Latest Articles Include:
- Perfecting peer review?
- Nat Med 17(1):1-2 (2011)
Nature Medicine | Editorial Perfecting peer review? Journal name:Nature MedicineVolume: 17,Pages:1–2Year published:(2011)DOI:doi:10.1038/nm0111-1Published online07 January 2011 Online science blogs are a valuable forum for commenting on published research, but their present importance lies in complementing rather than replacing the current system of peer review. View full text Additional data - Clinical drive prompts pharma and academia to partner up
- Nat Med 17(1):3 (2011)
Nature Medicine | News Clinical drive prompts pharma and academia to partner up * Megan Scudellari Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:3Year published:(2011)DOI:doi:10.1038/nm0111-3Published online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Pharmaceutical companies have sponsored research at academic centers for decades. But in the past few years, these collaborative agreements have escalated from small one-off contracts with individual labs to broad, big-money alliances that offer a hefty supply of perks but also a fair share of conflicts. "Over the years, things have changed," says Inder Verma, a Sanofi-Aventis–funded gene therapy researcher at the Salk Institute in La Jolla, California. "There's greater and greater interest in seeing whether our discoveries have the potential to be used in translational research. And, really, who is better at that than industry?" View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - A registry of registries? The US backs the idea for patients
- Nat Med 17(1):4 (2011)
Nature Medicine | News A registry of registries? The US backs the idea for patients * Monica Heger Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:4Year published:(2011)DOI:doi:10.1038/nm0111-4aPublished online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Building on the success of ClinicalTrials.gov, a registry of nearly 100,000 federally and privately funded clinical trials around the world, the US government is now planning to build a registry of patient registries. The ultimate goal of the effort is to create a one-stop shop where physicians, patients and researchers can find these lists of individuals who have made themselves available for observational medical studies. The 'metaregistry' will be searchable, and each entry will contain contact information for the person running the registry. At least initially, the catalog will not contain patient-specific information, but researchers could contact a given registry owner to obtain that data. The goal is that the database would serve patients and physicians looking for specific disease registries, researchers investigating a particular disease and drug developers. View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - New York Academy of Sciences launches angel investment network
- Nat Med 17(1):4 (2011)
Nature Medicine | News New York Academy of Sciences launches angel investment network * Cassandra Willyard Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:4Year published:(2011)DOI:doi:10.1038/nm0111-4bPublished online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. NEW YORK—New York City's burgeoning bioscience industry got a boost on 18 November when the New York Academy of Sciences launched a new network to connect private investors with fledgling life science companies looking for funding. The network, which is comprised of angel investors—wealthy individuals who risk their own money—will finance companies working to commercialize drugs, medical devices and other healthcare products. The goal is to bridge the funding gap between academic technology transfer offices and late-stage investors such as big corporations and venture capital firms. To be eligible for funding, inventors must submit an application and business plan to the Life Science Angel Network's (LSAN) screening committee, a panel of scientists, physicians, venture capitalists and other technology development experts tasked with vetting proposals. The most promising applicants will have the opportunity to present their idea to the entire network. View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Panel backs new NIH center devoted to translational medicine
- Nat Med 17(1):5 (2011)
Nature Medicine | News Panel backs new NIH center devoted to translational medicine * Brian Vastag Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:5Year published:(2011)DOI:doi:10.1038/nm0111-5aPublished online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. BETHESDA, MARYLAND—Seven years after the US National Institutes of Health (NIH) first launched its Roadmap for Medical Research, a broad-sweeping initiative designed to bridge the so-called 'valley of death' of drug development, the agency is on the verge of creating a new center devoted specifically to the goal of accelerating the transition of therapies from the lab to the clinic. On 7 December, the NIH's Scientific Management Review Board (SMRB) voted 12 to 1 in favor of forming a center for translational medicine and therapeutics. At the meeting, NIH director Francis Collins called the decision "a momentous occasion" but stopped short of immediately endorsing the center. (In NIH parlance, a 'center' is one step down the structural hierarchy from an 'institute'.) Agency watchers, however, widely expect Collins to move ahead with the idea. View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Amid legal uncertainties, NIH approves more embryonic stem cells
- Nat Med 17(1):5 (2011)
Nature Medicine | News Amid legal uncertainties, NIH approves more embryonic stem cells * Elie Dolgin Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:5Year published:(2011)DOI:doi:10.1038/nm0111-5bPublished online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. The legal battle over whether taxpayer dollars can go toward human embryonic stem cells research continues to drag on, but the US National Institutes of Health (NIH) is not waiting for a final court decision before adding new cell lines to its list of those eligible for financial backing. In August, a federal district judge issued a preliminary injunction against federally funded studies using such cells. But the nine-member working group tasked with determining whether embryonic stem cell lines are scientifically and ethically appropriate for federal backing has been "moving forward with our regular reviews" ever since an appeals court suspended the injunction in September, says the panel's chair Jeffrey Botkin, a medical ethicist from the University of Utah School of Medicine in Salt Lake City. View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Researchers sound alarm on 'silent' drug interactions
- Nat Med 17(1):6 (2011)
Nature Medicine | News Researchers sound alarm on 'silent' drug interactions * Stu Hutson Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:6Year published:(2011)DOI:doi:10.1038/nm0111-6Published online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. The number of people concurrently taking multiple prescription drugs is on the rise. According to numbers released last fall by the US Centers for Disease Control and Prevention, one in ten Americans is now on five or more medications—nearly twice as many people as in 2000—and the number of people taking at least two drugs has risen from a quarter to a third of the population over the same time period. Similar statistics are also reported throughout Europe. And as drug combinations grow more common, so does the risk that these medicines may interact in unpredictable ways that may go undetected until it's too late. Indeed, researchers across the world say that such 'silent' drug interactions pose a growing and widely misunderstood threat. View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Cancer drugs should add months, not weeks, say experts
- Nat Med 17(1):7 (2011)
Nature Medicine | News Cancer drugs should add months, not weeks, say experts * Hannah Hoag Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:7Year published:(2011)DOI:doi:10.1038/nm0111-7aPublished online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. In the last decade, the world's drug regulatory agencies have approved dozens of new anticancer therapies for everything from lung carcinoma to skin melanoma. Some of these new drugs add months to a patient's life. But others may offer only an extra week or two, on average, often with considerable toxicity and at a cost of thousands of dollars. Now experts are questioning whether these outcomes provide meaningful benefit to people's quality of life and are urging regulatory agencies to toughen the criteria for drug approval. Such a measure would push pharmaceutical companies to modify the design of clinical trials—a move that some drug makers and doctors worry could shrink the drug market. View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Funds go toward biomedical business incubators in Mexico
- Nat Med 17(1):7 (2011)
Nature Medicine | News Funds go toward biomedical business incubators in Mexico * Laura Vargas-Parada Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:7Year published:(2011)DOI:doi:10.1038/nm0111-7bPublished online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. MEXICO CITY—For decades, Mexican biomedical scientists have focused on basic research, looking with suspicion on those few colleagues who spoke of applying discoveries in the clinic or generating profits. However, this attitude is changing, according to Juan Pedro Laclette, general coordinator of the Scientific and Technological Consultative Forum, the main research advisory agency to the Mexican government: "Scientists are showing more interest in innovation and its potential to generate revenues," Laclette says. In the past two years, a group of high-ranking scientists from the National Autonomous University of Mexico (UNAM), the largest federally funded university in the country, have poured 15 million pesos ($1.2 million) of their own money into getting startups off the ground. "This is unprecedented," says Carlos Arias, director of the UNAM's Biotechnology Institute in Cuernavaca, in the state of Morelos, where most of the entrepreneur-scientists also work. Arias notes that the establishment of a technology transfer office at the institute has contributed to the entrepreneurial environment. View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Nomination bungle leaves CIRM leadership in limbo
- Nat Med 17(1):8 (2011)
Nature Medicine | News Nomination bungle leaves CIRM leadership in limbo * Elie Dolgin Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:8Year published:(2011)DOI:doi:10.1038/nm0111-8aPublished online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. The architect of California's $3 billion stem cell agency is set to keep his job for a little longer. Bob Klein—the man who co-wrote the ballot initiative that created the California Institute for Regenerative Medicine (CIRM) and then served as the agency's only chairman for the past six years—was reelected by its board of directors in a near-unanimous vote last month, meaning he will stay in his post for another six months while the organization regroups in its search for a suitable successor. State officials tasked with nominating his replacement had originally tapped current vice-chairman Art Torres as well as Alan Bernstein, executive director of the New York–based Global HIV Vaccine Enterprise as possible successors. But Bernstein, a Canadian national, was forced to pull out of the race at the beginning of December because of a state law requiring the head of a public agency to hold US citizenship, and Torres, a former state senator, withdrew his nomination a week later. As a result, Klein effectively became a shoo-in for the post. View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Conference brings Asia-Pacific research funding scheme closer
- Nat Med 17(1):8 (2011)
Nature Medicine | News Conference brings Asia-Pacific research funding scheme closer * Branwen Morgan Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:8Year published:(2011)DOI:doi:10.1038/nm0111-8bPublished online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. For several years, the idea of a multinational funding scheme to support research in the Asia-Pacific region has been brewing. Finally, at the biannual Australian Health and Medical Research Congress late last year, policymakers publicly discussed the merits and challenges of such a venture for the first time. The congress in Melbourne, convened by the Australian Society for Medical Research (ASMR), attracted about 1,500 delegates from 18 countries. Dignitaries at the November event included Australia's minister for mental health and aging and the acting consul-general of Japan. View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Drug developers explore vitamin D benefits without the vitamin
- Nat Med 17(1):9 (2011)
Nature Medicine | News Drug developers explore vitamin D benefits without the vitamin * Cassandra Willyard Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:9Year published:(2011)DOI:doi:10.1038/nm0111-9Published online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Late last year, the US Institute of Medicine released a report stating that research claiming any benefits from vitamin D beyond building strong bones is "inconsistent and inconclusive." Although inert vitamin D supplements have yet to prove their worth, new research is increasingly showing that active analogs of the molecule can help fight a number of diseases, including cancer and chronic kidney failure. Physicians already prescribe one such drug—calcitriol, an active form of vitamin D first synthesized in the 1970s—to treat rickets, psoriasis and vitamin D deficiencies sometimes seen with kidney disease. View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - 'Game changer' antibiotic and others in works for superbug
- Nat Med 17(1):10 (2011)
Nature Medicine | News 'Game changer' antibiotic and others in works for superbug * Elie Dolgin Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:10Year published:(2011)DOI:doi:10.1038/nm0111-10Published online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Last summer, researchers reported that the Clostridium difficile superbug had overtaken methicillin-resistant Staphylococcus aureus as the most common hospital-acquired infection in the US, where around half a million people fall ill from the diarrhea-inducing bacterium each year. For most people, the decades-old antibiotics metronidazole and vancomycin can clear the infection. But in around a quarter of all cases, the symptoms come roaring back, often with life-threatening consequences. Given the potential for relapse to occur, researchers have been desperately seeking new, more effective drug options. The leading candidate to reach the market within the year is fidaxomicin, an antibiotic pill that inhibits the enzyme RNA polymerase in bacteria. In October, the drug's manufacturer, San Diego–based Optimer Pharmaceuticals, reported data at the Infectious Diseases Society of America annual meeting in Vancouver, British Columbia from two phase 3 trials showing that two daily doses of fidaxomicin were as effective at clearing C. difficile infections as four pills per day of vancomycin. What really stuck out, though, was that fidaxomicin cut the recurrence rate in half. A month later, Optimer announced that it had filed a new drug application with the US Food and Drug Administration (FDA) and asked the drug regulator for a faster-than-usual review. View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Straight talk with...Phil Willis
- Nat Med 17(1):11 (2011)
Nature Medicine | News Straight talk with...Phil Willis * Asher Mullard Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:11Year published:(2011)DOI:doi:10.1038/nm0111-11Published online07 January 2011 Abstract Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg When Phil Willis, a former school headmaster turned politician, landed a seat on the UK government's Science and Technology Committee in 2005, he found a way to make his mark as a nonscientist by obsessively asking for evidence from researchers. Now in the House of Lords, he continues to advise the government on biomedicine, among other topics. Willis recently found a new cause to champion as well, the Association of Medical Research Charities (AMRC). The AMRC represents 124 UK nonprofits that collectively spend £1 billion ($1.6 billion) a year on biomedical research, around one third of the total amount of money put toward health research in the country. As Willis stepped into his role as chairman of the association in November, he spoke with about his plans to drive the sector forward through tough economic times. View full text Additional data - News in brief
- Nat Med 17(1):12-13 (2011)
Nature Medicine | News News in brief Journal name:Nature MedicineVolume: 17,Pages:12–13Year published:(2011)DOI:doi:10.1038/nm0111-12Published online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Policy Playback Responding to recommendations from a group of outside advisers, Francis Collins, director of the US National Institutes of Health, announced that he was forming a task force to see how the country's National Institute on Drug Abuse and National Institute on Alcohol Abuse and Alcoholism could be merged into a . View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Putting sleeping sickness to bed
- Nat Med 17(1):14-17 (2011)
Nature Medicine | News Putting sleeping sickness to bed * Cassandra Willyard1 Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Pages:14–17Year published:(2011)DOI:doi:10.1038/nm0111-14Published online07 January 2011 For most people, a single bite from a parasite-infected tsetse fly can trigger a slow, agonizing and sometimes fatal disease known as African sleeping sickness. But new research shows that some people, as well as baboons and other great apes, are naturally resistant to infection. awakens to the possibility of using existing immunity to engineer new therapies and transgenic livestock. View full text Additional data Affiliations * Cassandra Willyard is a science writer based in Brooklyn, New York - Scarred by disease
- Nat Med 17(1):18-20 (2011)
Nature Medicine | News Scarred by disease * Thomas Hayden1 Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Pages:18–20Year published:(2011)DOI:doi:10.1038/nm0111-18Published online07 January 2011 Collagen may be the darling of the beauty world for its purported antiwrinkle function, but too much of the protein can have ugly—even deadly—consequences. Measuring and treating this overabundance, known as fibrosis, presents a seemingly impossible challenge. visits a company in California offering a weighty solution. View full text Additional data Affiliations * Thomas Hayden is a San Francisco–based science writer. - To improve science literacy, researchers should run for school board
- Nat Med 17(1):21 (2011)
Nature Medicine | News To improve science literacy, researchers should run for school board * Jon D. Miller1 Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:21Year published:(2011)DOI:doi:10.1038/nm0111-21Published online07 January 2011 This century will bring exciting biomedical advances thanks to stem cells and genetic engineering. If scientists want the public to grasp the meaning of these developments, they need to start getting personally involved in improving the education system. View full text Additional data Affiliations * Jon D. Miller is director of the International Center for the Advancement of Scientific Literacy at the University of Michigan, Ann Arbor, USA. - Searching for the source
- Nat Med 17(1):23 (2011)
Nature Medicine | Book Review Searching for the source * Anne-Emanuelle Birn1 Contact Anne-Emanuelle Birn Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:23Year published:(2011)DOI:doi:10.1038/nm0111-23Published online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Inside the Outbreaks: The Elite Medical Detectives of the Epidemic Intelligence Service Mark Pendergrast Houghton Mifflin Harcourt, 2010 432 pp., hardcover, $28.00 ISBN: 0151011206 View full text Author information Affiliations * Anne-Emanuelle Birn is Professor and Canada Research Chair in International Health at the University of Toronto, Toronto, Ontario, Canada. ae.birn@utoronto.ca Competing financial interests The author declares no competing financial interests. Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data * Journal home * Current issue * For authors * Subscribe * E-alert sign up * RSS feed Open innovation challenges * Low-Volume Liquid Dispersion Mechanism Deadline:Feb 20 2011Reward:$25,000 USD The Seeker is looking for mechanisms to enable dispersion of a fluid sample of 40 nanoliters (nl) ov… * Chordoma Cancer Cell Lines Needed to Save Lives! Deadline:Mar 13 2011Reward:$10,000 USD The Chordoma Foundation requests cell lines or animal models that can be used for research into chor… * Powered by: * More challenges Top content Emailed * Neurodegeneration and the neuroimmune system Nature Medicine 06 Dec 2010 * Newer antidepressants go beyond serotonin—and the synapse Nature Medicine 06 Dec 2010 * Nicotine fix Nature Medicine 01 Jul 2004 * Universities evolve, looking to Darwin for new medical insights Nature Medicine 06 Dec 2010 * Fluoroquinolone-modifying enzyme: a new adaptation of a common aminoglycoside acetyltransferase Nature Medicine 20 Dec 2005 View all Downloaded * Receptor-mediated activation of ceramidase activity initiates the pleiotropic actions of adiponectin Nature Medicine 26 Dec 2010 * Conversion of vascular endothelial cells into multipotent stem-like cells Nature Medicine 21 Nov 2010 * Retractions of the year Nature Medicine 06 Dec 2010 * miR-499 regulates mitochondrial dynamics by targeting calcineurin and dynamin-related protein-1 Nature Medicine 26 Dec 2010 * Notable advances 2010 Nature Medicine 06 Dec 2010 View all Blogged * Severe pandemic 2009 H1N1 influenza disease due to pathogenic immune complexes Nature Medicine 05 Dec 2010 View all - Lack of HIF-2α in limb bud mesenchyme causes a modest and transient delay of endochondral bone development
- Nat Med 17(1):25-26 (2011)
Nature Medicine | Correspondence Lack of HIF-2α in limb bud mesenchyme causes a modest and transient delay of endochondral bone development * Elisa Araldi1 Search for this author in: * NPG journals * PubMed * Google Scholar * Richa Khatri1 Search for this author in: * NPG journals * PubMed * Google Scholar * Amato J Giaccia2 Search for this author in: * NPG journals * PubMed * Google Scholar * M Celeste Simon3 Search for this author in: * NPG journals * PubMed * Google Scholar * Ernestina Schipani1 Contact Ernestina Schipani Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:25–26Year published:(2011)DOI:doi:10.1038/nm0111-25Published online07 January 2011 To the Editor: Two papers recently published in Nature Medicine provide evidence that hypoxia-inducible factor-2α (HIF-2α) is crucial for articular surface homeostasis through mechanisms that involve, at least in part, regulation of genes such as Col10a1 (encoding the α1 chain of type X collagen, Col10A1), Mmp13 (encoding matrix metalloproteinase-13, MMP-13) and Vegfa (encoding vascular endothelial growth factor-A, VEGF)1, 2. Interestingly, Saito et al.1 also report a mild delay of chondrocyte hypertrophy in fetal growth plates of mice heterozygous for genetic knockout of HIF-2α (encoded by Epas1). The key implication of this finding is that homozygous loss of HIF-2α would markedly delay chondrocyte hypertrophy and replacement of cartilage by bone. However, as shown below, we found that this was not the case. View full text Author information * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Affiliations * Endocrine Unit, Department of Medicine, Massachusetts General Hospital–Harvard Medical School, Boston, Massachusetts, USA. * Elisa Araldi, * Richa Khatri & * Ernestina Schipani * Division of Cancer and Radiation Biology, Department of Radiation Oncology, Stanford University School of Medicine, Stanford, California, USA. * Amato J Giaccia * Abramson Family Cancer Research Institute, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, USA. * M Celeste Simon Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Ernestina Schipani Supplementary information * Author information * Supplementary information PDF files * Supplementary Text and Figures (75K) Supplementary Fig. 1 and Supplementary Methods Additional data - Replication studies in various ethnic populations do not support the association of the HIF-2α SNP rs17039192 with knee osteoarthritis
- Nat Med 17(1):26-27 (2011)
Nature Medicine | Correspondence Replication studies in various ethnic populations do not support the association of the HIF-2α SNP rs17039192 with knee osteoarthritis * Masahiro Nakajima1, 7 Search for this author in: * NPG journals * PubMed * Google Scholar * Dongquan Shi2, 3, 7 Search for this author in: * NPG journals * PubMed * Google Scholar * Jin Dai1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Aspasia Tsezou4 Search for this author in: * NPG journals * PubMed * Google Scholar * Minghao Zheng5 Search for this author in: * NPG journals * PubMed * Google Scholar * Paul E Norman5 Search for this author in: * NPG journals * PubMed * Google Scholar * Atsushi Takahashi6 Search for this author in: * NPG journals * PubMed * Google Scholar * Shiro Ikegawa1, 3, 5 Contact Shiro Ikegawa Search for this author in: * NPG journals * PubMed * Google Scholar * Qing Jiang2, 3 Contact Qing Jiang Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Corresponding authorsJournal name:Nature MedicineVolume: 17,Pages:26–27Year published:(2011)DOI:doi:10.1038/nm0111-26Published online07 January 2011 To the Editor: We read with great interest the recent article in Nature Medicine by Saito et al.1 describing the role of hypoxia-inducible factor-2α (HIF-2α) in skeletal growth and osteoarthritis development. The authors report that HIF-2α is a central transactivator targeting several crucial genes for endochondral ossification in mice1. The role of HIF-2α in human osteoarthritis is buttressed by their Japanese association study suggesting that rs17039192, a functional single nucleotide polymorphism (SNP) in the human EPAS1 gene encoding HIF-2α, is associated with knee osteoarthritis1. Although the study clearly contains interesting and compelling data on the effect of HIF-2α in endochondral ossification during skeletal growth in mice, the human evidence, supporting the role of HIF-2α in osteoarthritis, is limited by its small sample size and a marginal P value. Given that lack of replication and false positive findings are common pitfalls of association studies2, we sought to furth! er clarify the association of the HIF-2α SNP rs17039192 with osteoarthritis. View full text Author information * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Primary authors * These authors contributed equally to this work. * Masahiro Nakajima & * Dongquan Shi Affiliations * Laboratory for Bone and Joint Diseases, Center for Genomic Medicine, RIKEN, Tokyo, Japan. * Masahiro Nakajima, * Jin Dai & * Shiro Ikegawa * The Center of Diagnosis and Treatment for Joint Disease, Drum Tower Hospital Affiliated to Medical School of Nanjing University, Nanjing, China. * Dongquan Shi, * Jin Dai & * Qing Jiang * Laboratory for Bone and Joint Diseases, Model Animal Research Center, Nanjing University, Nanjing, China. * Dongquan Shi, * Shiro Ikegawa & * Qing Jiang * Laboratory of Cytogenetics and Molecular Genetics, Medical School, University of Thessaly, Larissa, Greece. * Aspasia Tsezou * School of Surgery, University of Western Australia, Perth, Australia. * Minghao Zheng, * Paul E Norman & * Shiro Ikegawa * Laboratory for Statistical Analysis, Center for Genomic Medicine, RIKEN, Tokyo, Japan. * Atsushi Takahashi Competing financial interests The authors declare no competing financial interests. Corresponding authors Correspondence to: * Shiro Ikegawa or * Qing Jiang Supplementary information * Author information * Supplementary information PDF files * Supplementary Text and Figures (108K) Supplementary Fig. 1 and Supplementary Table 1 Additional data - Reply to: "Lack of HIF-2α in limb bud mesenchyme causes a modest and transient delay of endochondral bone development" and "Replication studies in various ethnic populations do not support the association of the HIF-2α SNP rs17039192 with knee osteoarthritis"
- Nat Med 17(1):27-29 (2011)
Nature Medicine | Correspondence Reply to: "Lack of HIF-2α in limb bud mesenchyme causes a modest and transient delay of endochondral bone development" and "Replication studies in various ethnic populations do not support the association of the HIF-2α SNP rs17039192 with knee osteoarthritis" * Taku Saito1 Search for this author in: * NPG journals * PubMed * Google Scholar * Akihiko Mabuchi2 Search for this author in: * NPG journals * PubMed * Google Scholar * Toru Akune1 Search for this author in: * NPG journals * PubMed * Google Scholar * Ung-il Chung1 Search for this author in: * NPG journals * PubMed * Google Scholar * Katsushi Tokunaga2 Search for this author in: * NPG journals * PubMed * Google Scholar * Hiroshi Kawaguchi1 Contact Hiroshi Kawaguchi Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:27–29Year published:(2011)DOI:doi:10.1038/nm0111-27Published online07 January 2011 Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Saito et al. reply: As Araldi et al.1 mention in their correspondence, our previous article describes that mice heterozygous for knockout of the gene encoding hypoxia-inducible factor-2α (Epas1+/− mice) show mild and transient impairment in endochondral ossification during skeletal growth mainly in the embryonic stage2. We disagree with their hypothesis that there would be marked impairment due to homozygous deficiency of the gene, as the amount of HIF-2α essential for the maintenance of normal skeletal growth is unknown. Therefore, we are interested in their findings in more sophisticated mouse mutants with homozygous deficiency of Epas1 in limb bud mesenchyme, achieved by tissue-selective knockout. View full text Author information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Affiliations * Sensory & Motor System Medicine, Hongo, Bunkyo-ku, Tokyo, Japan. * Taku Saito, * Toru Akune, * Ung-il Chung & * Hiroshi Kawaguchi * Human Genetics, University of Tokyo, Hongo, Bunkyo-ku, Tokyo, Japan. * Akihiko Mabuchi & * Katsushi Tokunaga Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Hiroshi Kawaguchi Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Does thrombocytopenia contribute to patent ductus arteriosus?
- Nat Med 17(1):29-30 (2011)
Nature Medicine | Correspondence Does thrombocytopenia contribute to patent ductus arteriosus? * Kazumichi Fujioka1 Search for this author in: * NPG journals * PubMed * Google Scholar * Ichiro Morioka1 Contact Ichiro Morioka Search for this author in: * NPG journals * PubMed * Google Scholar * Akihiro Miwa1 Search for this author in: * NPG journals * PubMed * Google Scholar * Satoru Morikawa1 Search for this author in: * NPG journals * PubMed * Google Scholar * Akio Shibata1 Search for this author in: * NPG journals * PubMed * Google Scholar * Naoki Yokoyama1 Search for this author in: * NPG journals * PubMed * Google Scholar * Masafumi Matsuo1 Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:29–30Year published:(2011)DOI:doi:10.1038/nm0111-29Published online07 January 2011 Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. To the Editor: The persistence of a patent ductus arteriosus (PDA) in premature newborns causes serious hemodynamic changes that sometimes result in their death1. A better understanding of the mechanism of ductus arteriosus (DA) closure may lead to new strategies for the management of PDA in premature newborns. The article by Echtler et al.2 described an essential role of platelets in postnatal DA closure in a mouse model. In addition, they showed in a retrospective clinical study that thrombocytopenia increases the risk for PDA in German premature newborns2. View full text Author information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Affiliations * Department of Pediatrics, Kobe University Graduate School of Medicine, Kobe, Japan. * Kazumichi Fujioka, * Ichiro Morioka, * Akihiro Miwa, * Satoru Morikawa, * Akio Shibata, * Naoki Yokoyama & * Masafumi Matsuo Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Ichiro Morioka Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Reply to: "Does thrombocytopenia contribute to patent ductus arteriosus?"
- Nat Med 17(1):30-31 (2011)
Nature Medicine | Correspondence Reply to: "Does thrombocytopenia contribute to patent ductus arteriosus?" * Katrin Echtler1 Search for this author in: * NPG journals * PubMed * Google Scholar * Konstantin Stark1 Search for this author in: * NPG journals * PubMed * Google Scholar * Orsolya Genzel-Borovicz2 Search for this author in: * NPG journals * PubMed * Google Scholar * Adnan Kastrati1 Search for this author in: * NPG journals * PubMed * Google Scholar * Steffen Massberg1 Contact Steffen Massberg Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:30–31Year published:(2011)DOI:doi:10.1038/nm0111-30Published online07 January 2011 Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Echtler et al. reply: We have previously shown in a retrospective study including 123 infants born prematurely at 24–30 weeks' gestation that thrombocytopenia is associated with a higher incidence of patent ductus arteriosus (PDA)1. This finding is consistent with a prospective study published previously revealing that lower platelet counts are a major predictor of PDA following a single course of indomethacin in Malaysian preterm infants2. But, not in line with the abovementioned findings, Fujioka et al.3 in their correspondence do not find an association between low platelet counts and PDA in a cohort of Japanese preterm newborns. View full text Author information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Affiliations * Deutsches Herzzentrum, Klinik für Herz- und Kreislauferkrankungen, Technische Universität, Munich, Germany. * Katrin Echtler, * Konstantin Stark, * Adnan Kastrati & * Steffen Massberg * Klinik und Poliklinik für Frauenheilkunde und Geburtshilfe, Perinatalzentrum, Ludwig-Maximilians Universität, Munich, Germany. * Orsolya Genzel-Borovicz Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Steffen Massberg Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - NOX-4 is expressed in thickened pulmonary arteries in idiopathic pulmonary fibrosis
- Nat Med 17(1):31-32 (2011)
Nature Medicine | Correspondence NOX-4 is expressed in thickened pulmonary arteries in idiopathic pulmonary fibrosis * Jean-Claude Pache1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Stephanie Carnesecchi1, 3 Search for this author in: * NPG journals * PubMed * Google Scholar * Christine Deffert1, 2, 3 Search for this author in: * NPG journals * PubMed * Google Scholar * Yves Donati1, 3 Search for this author in: * NPG journals * PubMed * Google Scholar * François R Herrmann4 Search for this author in: * NPG journals * PubMed * Google Scholar * Constance Barazzone-Argiroffo1, 3 Search for this author in: * NPG journals * PubMed * Google Scholar * Karl-Heinz Krause1, 2 Contact Karl-Heinz Krause Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:31–32Year published:(2011)DOI:doi:10.1038/nm0111-31Published online07 January 2011 Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. To the Editor: Idiopathic pulmonary fibrosis (IPF) is a fatal disease, and there is a need for an improved understanding of its pathophysiology and for new therapeutic concepts. Recent work by Hecker et al.1 suggests that the generation of reactive oxygen species by the NADPH oxidase NOX-4 is involved in the pathophysiology of IPF. More specifically, the work suggests that NOX-4–dependent generation of reactive oxygen species is required for transforming growth factor-β1 (TGF-β1)-induced myofibroblast differentiation, extracellular matrix production and contractility. The authors provide the first elements to support the concept that NOX-4 might be a target for the treatment of pulmonary fibrosis. View full text Author information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Affiliations * Department of Pathology and Immunology, Medical School, University of Geneva, Geneva, Switzerland. * Jean-Claude Pache, * Stephanie Carnesecchi, * Christine Deffert, * Yves Donati, * Constance Barazzone-Argiroffo & * Karl-Heinz Krause * Department of Genetic and Laboratory Medicine, Geneva University Hospitals, Geneva, Switzerland. * Jean-Claude Pache, * Christine Deffert & * Karl-Heinz Krause * Department of Pediatrics, Medical School, University of Geneva, Geneva, Switzerland. * Stephanie Carnesecchi, * Christine Deffert, * Yves Donati & * Constance Barazzone-Argiroffo * Department of Rehabilitation and Geriatrics, Hospital of Trois Chêne, University Hospitals of Geneva, Geneva, Switzerland. * François R Herrmann Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Karl-Heinz Krause Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Reply to: "NOX-4 is expressed in thickened pulmonary arteries in idiopathic pulmonary fibrosis"
- Nat Med 17(1):32-33 (2011)
Nature Medicine | Correspondence Reply to: "NOX-4 is expressed in thickened pulmonary arteries in idiopathic pulmonary fibrosis" * Louise Hecker1 Search for this author in: * NPG journals * PubMed * Google Scholar * David R Crowe2 Search for this author in: * NPG journals * PubMed * Google Scholar * Victor J Thannickal1 Contact Victor J Thannickal Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:32–33Year published:(2011)DOI:doi:10.1038/nm0111-32Published online07 January 2011 Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Hecker, Crowe and Thannickal reply: In their correspondence, Pache et al.1 raise the intriguing idea that NOX-4 is expressed in the remodeled vascular structures resulting from chronic hypoxia in idiopathic pulmonary fibrosis (IPF). Their immunohistochemical analyses on lung tissue sections from control subjects and subjects with IPF show α-smooth muscle actin (α-SMA)-expressing cells in pulmonary vasculature1. In their morphometric analysis of pulmonary arteries, these investigators report a statistically significant increase in the thickness of the vessel walls in IPF1, thus supporting the concept of vascular remodeling in IPF, which may or may not be secondary to chronic hypoxia2. Although the authors do not provide NOX-4 staining data, they refer to our immunohistochemical studies of IPF tissue sections published earlier in Nature Medicine3 as evidence that the NADPH oxidase NOX-4 is expressed in remodeled blood vessels of the IPF lung, and they suggest that "NOX-4 might also be involved in the pathoph! ysiology of lung artery hypertrophy in human IPF."1. View full text Author information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Affiliations * Division of Pulmonary, Allergy and Critical Care Medicine, Department of Medicine, University of Alabama at Birmingham, Birmingham, Alabama, USA. * Louise Hecker & * Victor J Thannickal * Department of Pathology, University of Alabama at Birmingham, Birmingham, Alabama, USA. * David R Crowe Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Victor J Thannickal Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Triclosan is minimally effective in rodent malaria models
- Nat Med 17(1):33-34 (2011)
Nature Medicine | Correspondence Triclosan is minimally effective in rodent malaria models * Werner Baschong1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Sergio Wittlin3 Search for this author in: * NPG journals * PubMed * Google Scholar * Kirstine A Inglis4 Search for this author in: * NPG journals * PubMed * Google Scholar * Alan H Fairlamb4 Search for this author in: * NPG journals * PubMed * Google Scholar * Simon L Croft5 Search for this author in: * NPG journals * PubMed * Google Scholar * T R Santha Kumar6 Search for this author in: * NPG journals * PubMed * Google Scholar * David A Fidock6, 7 Search for this author in: * NPG journals * PubMed * Google Scholar * Reto Brun3 Contact Reto Brun Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:33–34Year published:(2011)DOI:doi:10.1038/nm0111-33Published online07 January 2011 To the Editor: The discovery several years ago that Plasmodium parasites use a fatty acid synthesis type II pathway (FAS-II), shared by plants and bacteria, raised hopes for the discovery of new antimalarial targets1 and stimulated the search for FAS-II inhibitors. This pathway is harbored in the Plasmodium apicoplast, a nonphotosynthetic plastid of cyanobacterial origin, and is composed of four enzymes. The rate-limiting step is mediated by enoyl acyl carrier protein reductase (FabI)2. In mammals, all four enzymatic steps reside in the single multifunctional fatty acid synthesis type I (FAS-I) protein. View full text Author information * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Affiliations * Department of Oral Surgery, Oral Radiology and Oral Medicine, University of Basel, Basel, Switzerland. * Werner Baschong * BASF Schweiz AG, Basel, Switzerland. * Werner Baschong * Department of Medical Parasitology and Infection Biology, Swiss Tropical and Public Health Institute, Basel, Switzerland. * Sergio Wittlin & * Reto Brun * Division of Biological Chemistry & Drug Discovery, University of Dundee, Scotland, UK. * Kirstine A Inglis & * Alan H Fairlamb * Department of Infectious and Tropical Diseases, London School of Hygiene & Tropical Medicine, England, UK. * Simon L Croft * Department of Microbiology & Immunology, Columbia University Medical Center, New York, New York, USA. * T R Santha Kumar & * David A Fidock * Division of Infectious Diseases in the Department of Medicine, Columbia University Medical Center, New York, New York, USA. * David A Fidock Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Reto Brun Supplementary information * Author information * Supplementary information PDF files * Supplementary Text and Figures (78K) Supplementary Table 1 and Supplementary Methods Additional data - Reply to: "Triclosan is minimally effective in rodent malaria models"
- Nat Med 17(1):34-35 (2011)
Nature Medicine | Correspondence Reply to: "Triclosan is minimally effective in rodent malaria models" * Namita Surolia1 Search for this author in: * NPG journals * PubMed * Google Scholar * Avadhesha Surolia2 Contact Avadhesha Surolia Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:34–35Year published:(2011)DOI:doi:10.1038/nm0111-34Published online07 January 2011 Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Surolia and Surolia reply: We previously showed that Plasmodium falciparum has the type II fatty acid synthesis (FAS) pathway and outlined the uniqueness of this pathway in terminating at C14 acyl chain. That the pathway shows termination predominantly at C14 acyl chain is an exclusive feature of Plasmodium falciparum, and no other organism shows this property. We found that triclosan inhibits FAS with a half-maximal inhibitory concentration of ~800 nM, impeding blood-stage parasitic growth1. These findings were validated by several independent groups2, 3. Targeting of enoyl–acyl carrier protein (ACP) reductase (FabI) by triclosan in malaria was subsequently claimed to be of great therapeutic promise4. View full text Author information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Affiliations * Molecular Biology and Genetics Unit, Jawaharlal Nehru Centre for Advanced Scientific Research, Jakkur, Bangalore, India. * Namita Surolia * Molecular Biophysics Unit, Indian Institute of Science, Bangalore, India. * Avadhesha Surolia Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Avadhesha Surolia Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Adiponectin sphings into action
- Nat Med 17(1):37-38 (2011)
Nature Medicine | Article Receptor-mediated activation of ceramidase activity initiates the pleiotropic actions of adiponectin * William L Holland1 Search for this author in: * NPG journals * PubMed * Google Scholar * Russell A Miller2 Search for this author in: * NPG journals * PubMed * Google Scholar * Zhao V Wang1 Search for this author in: * NPG journals * PubMed * Google Scholar * Kai Sun1 Search for this author in: * NPG journals * PubMed * Google Scholar * Brian M Barth3 Search for this author in: * NPG journals * PubMed * Google Scholar * Hai H Bui4 Search for this author in: * NPG journals * PubMed * Google Scholar * Kathryn E Davis1 Search for this author in: * NPG journals * PubMed * Google Scholar * Benjamin T Bikman5 Search for this author in: * NPG journals * PubMed * Google Scholar * Nils Halberg1, 6 Search for this author in: * NPG journals * PubMed * Google Scholar * Joseph M Rutkowski1 Search for this author in: * NPG journals * PubMed * Google Scholar * Mark R Wade4 Search for this author in: * NPG journals * PubMed * Google Scholar * Vincent M Tenorio1 Search for this author in: * NPG journals * PubMed * Google Scholar * Ming-Shang Kuo4 Search for this author in: * NPG journals * PubMed * Google Scholar * Joseph T Brozinick4 Search for this author in: * NPG journals * PubMed * Google Scholar * Bei B Zhang7 Search for this author in: * NPG journals * PubMed * Google Scholar * Morris J Birnbaum2 Search for this author in: * NPG journals * PubMed * Google Scholar * Scott A Summers3, 5, 8 Search for this author in: * NPG journals * PubMed * Google Scholar * Philipp E Scherer1, 9 Contact Philipp E Scherer Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:55–63Year published:(2011)DOI:doi:10.1038/nm.2277Received15 September 2010Accepted20 October 2010Published online26 December 2010 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Abstract * Abstract * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg The adipocyte-derived secretory factor adiponectin promotes insulin sensitivity, decreases inflammation and promotes cell survival. No unifying mechanism has yet explained how adiponectin can exert such a variety of beneficial systemic effects. Here, we show that adiponectin potently stimulates a ceramidase activity associated with its two receptors, AdipoR1 and AdipoR2, and enhances ceramide catabolism and formation of its antiapoptotic metabolite—sphingosine-1-phosphate (S1P)—independently of AMP-dependent kinase (AMPK). Using models of inducible apoptosis in pancreatic beta cells and cardiomyocytes, we show that transgenic overproduction of adiponectin decreases caspase-8-mediated death, whereas genetic ablation of adiponectin enhances apoptosis in vivo through a sphingolipid-mediated pathway. Ceramidase activity is impaired in cells lacking both adiponectin receptor isoforms, leading to elevated ceramide levels and enhanced susceptibility to palmitate-induced cell de! ath. Combined, our observations suggest a unifying mechanism of action for the beneficial systemic effects exerted by adiponectin, with sphingolipid metabolism as its core upstream signaling component. View full text Figures at a glance * Figure 1: Adiponectin rapidly lowers hepatic ceramide content and improves glucose homeostasis. () Total ceramide levels from liver of leptin-deficient (ob/ob) mice after 60-min intravenous (i.v.) treatments with full-length adiponectin (Adn, 2 mg kg−1) or PBS (n = 6 per group). () Glucose infusion rates during hyperinsulinemic-euglycemic clamps performed on conscious unrestrained ob/ob mice before and after a bolus of Adn (2 mg kg−1, i.v.) or PBS (n = 5 per group). () Total ceramide levels from livers of mice with diet-induced obesity after 60-min treatments with full-length Adn (2 mg kg−1, i.v.) or PBS (n = 9 per group). (,) Insulin tolerance () and hepatic ceramide content () of adiponectin-deficient (Adipoq−/−), wild-type (+/+) or overexpressing (Tg/+) mice maintained on HFD (solid lines) or normal chow (dashed line) for 8 weeks (n = 7 per group). (–) Results from Lkb1fl/fl mice infected with adenovirus (Ad) encoding either GFP or Cre recombinase 16 d before experiments (n = 8 per group). () Triplicate western blots of liver proteins, probed for LKB1, T! hr172-phosphorylated AMPK (pAMPK), AMPK, pSer79-ACC and ACC. () Whole blood glucose monitored for 6 h after injection of PBS (solid lines) or Adn (34 mg kg−1, i.v., dashed line). () Quantification of total hepatic ceramide, dihydroceramide (dhCeramide), glucosylceramide and GM3 ganglioside. *P < 0.01 effect of adiponectin. †P < 0.05 compared to lean wild-type controls. Error bars, s.e.m. * Figure 2: Adiponectin promotes cardiomyocyte and HEART-ATTAC survival. () Survival Kaplan-Meier plot of female HEART-ATTAC transgenic mice crossed into indicated adiponectin backgrounds and challenged with AP20187 (0.010 μg kg−1, i.p.) (n = 12 per group). () Ceramide quantified from left ventricle or serum and normalized to the average content from adiponectin wild-type (WT) mice (63.9 pmol mg−1 in left ventricle, 9.5 pmol μl−1 in serum) (n = 12 per group). () Dihydro-S1P (dhS1P), S1P, dihydrosphingosine (dhSph) and sphingosine (Sph) quantified in left ventricle of WT (Adipoq+/+), adiponectin heterozygote (Adipoq−/+) and null (Adipoq−/−) mice (n = 6 per group). () Survival of male AdipoqTg/+ mice treated with PBS or male Adipoq+/+ HEART-ATTAC mice treated with S1P (1 mg kg−1, i.p.), FTY720 (FTY; 1 mg kg−1, i.p.), myriocin (Myr; 0.3 mg kg−1, i.p.) or PBS immediately before injection with AP20187 (0.05 mg kg−1, i.p.) (n = 10 per group). () Cell death, determined with or without AP20187-induced apoptosis in HEART-ATTAC transg! enic primary cardiomyocytes after treatment with BSA alone, C2-ceramide or palmitate. Cells were co-treated with PBS, adiponectin (Adn), S1P or myriocin (Myr) (n = 6 per group from three separate experiments). *P < 0.05 difference from wild-type control. †P < 0.01 effect of lipid treatment. Error bars, s.e.m. * Figure 3: Adiponectin targets the endocrine pancreas and maintains beta cell mass. () Adiponectin (Adn), insulin and nuclei (DAPI) visualized by immunofluorescence after injection of PBS (top) or Adn (bottom) into adiponectin null mice. Scale bar, 100 μm. (,) Random-fed blood glucose (n = 12 per group) () and total pancreatic insulin content (n = 6 per group) (), assessed in male adiponectin transgenic (AdipoqTg/+) versus wild-type (Adipoq+/+) mice 10 d after treatment with vehicle or AP20187 (0.2 mg kg−1, i.p., single injection). (,) Random-fed blood glucose (n = 12 per group) () and total pancreatic insulin content (), determined in female wild-type (Adipoq+/+) and adiponectin null (Adipoq−/−) PANIC-ATTAC mice 10 d after initiating treatment with AP20187 (0.2 mg kg−1, i.p., twice daily for 3 d) (n = 6 per group). () Islet size, calculated by mean cross-sectional area of multicelled islets from pancreata of male adiponectin-overexpressing (AdipoqTg/+), wild-type (Adipoq+/+) or adiponectin-lacking (Adipoq−/−) mice, 10 d after treatment with AP! 20187 or vehicle (n = 6 per condition). *P < 0.02 difference between adiponectin transgenic (or adiponectin null) and WT mouse of the same treatment. †P < 0.02 effect of AP20187 treatment. Error bars, s.e.m. * Figure 4: Adiponectin alters sensitivity to ceramide-induced apoptosis in INS-1 beta cells. () Cell viability, assessed in INS-1 cells challenged with BSA, palmitate (Pal) or C2-ceramide (C2) in the presence or absence of adiponectin (Adn) (n = 6 per group from three separate experiments). () Cell viability determined on INS-1 cells that were pretreated with sphingosine kinase inhibitor (SKI) or DMSO before delivery of adiponectin or PBS and challenged with BSA or palmitate. () Ceramidase activity, determined in lysates from cultured INS-1 cells under a range of pH conditions (n = 4 from separate experiments) in the presence or absence of Adn (in vitro). Baseline is BSA treatment without Adn. () Live cells imaged by staining with carboxyfluorescein diacetate (cFDA), and dead cells stained for annexin V, after treatment with C2-ceramide in the presence or absence of S1P. Representative of three separate experiments; scale bar, 50 μm. () Apoptosis of INS-1 cells determined by FACS analysis of annexin V and propidium iodide staining after treatment with BSA, palmitat! e or coadministered palmitate and S1P (representative of three independent experiments). *P < 0.01 effect of Adn. †P < 0.01 effect of proapoptotic insult. Error bars, s.e.m. * Figure 5: Adiponectin receptors 1 and 2 confer ceramidase activity in vivo. (,) Ceramidase activity after in vitro treatment with adiponectin (Adn) or PBS () and protein expression () assessed 48 h after transient transfection of HEK-293T cells with constructs encoding GFP alone or cotransfection with GFP and mouse AdipoR1, mouse AdipoR2, or indicated point mutants for conserved histidine residues in AdipoR1 (H141R or H191R) or AdipoR2 (H152R or H202R) (n = 5 from separate experiments). WT, wild type; R1, AdipoR1; R2, AdipoR2. (–) Results from hepatic overexpression of human AdipoR1, human AdipoR2 or GFP by adenovirus in vivo. () Hepatic ceramidase activity determined from fresh lysates 5 d after infection of 9-week-old WT mice C57/Bl6J mice (n = 5 per group). () Hepatic ceramides, measured after 6 h infusion of lard oil emulsions or fat-free glycerol control emulsions (n = 6 per group). (,) Insulin tolerance () and hepatic ceramide content (), determined 8 d after infection of high fat–fed mice or chow-fed controls (n = 6–8 per group). *P < 0! .05 effect of Adn or lipid administration. †P < 0.02 effect of genetic overexpression. Error bars, s.e.m. * Figure 6: Ablating adiponectin receptors 1 and 2 impairs ceramidase activity, S1P generation and cell survival. () Ceramidase activity from wild-type (WT) or AdipoR1 AdipoR2 DKO MEFs in the presence or absence of adiponectin (Adn) (n = 4). () S1P and dihydro-S1P quantified from WT or DKO MEFs after 12 h incubation in palmitate (750 μM) or BSA (n = 6). () Ceramide quantified from WT, AdipoR1 AdipoR2 (R1/R2) DKO or LKB1 knockout (KO) MEFs after 12-h incubations with palmitate (Pal) or BSA supplemented with Adn (5 μg ml−1) or PBS (n = 6 from three separate experiments). () Cell viability assessed in MEFs treated as in after 16 h of palmitate treatment (n = 5). () Total and phosphorylated AMPK, probed by western blot after pretreatment with D-e-MAPP or DMSO and treatment with full-length Adn, globular adiponectin (gAdn), S1P, AICAR or C2-ceramide (representative of four independent experiments). () Ceramide promotes apoptosis by aiding death receptor clustering, apoptosome formation and Bcl-2-associated X protein (Bax) translocation. Ceramide impairs activation of Akt, also known as p! rotein kinase B (PKB), via activation of protein kinase C (PKC)-ζ or protein phosphatase 2A (PP2A). Adiponectin promotes the deacylation of ceramide by activating adiponectin receptors. The resulting sphingosine and S1P increase intracellular calcium and activate AMPK through stimulation of CAMKK. These actions promote survival, nutrient uptake, nutrient utilization and mitochondrial proliferation. CAMKK, calmodulin-dependent protein kinase kinase; ER, endoplasmic reticulum; SM, sphingomyelin; SphK, sphingosine kinase; TNF; tumor necrosis factor-α. *P < 0.05 effect of Adn. †P < 0.05 compared to WT cells. Error bars, s.e.m. Author information * Abstract * Author information * Supplementary information Affiliations * Touchstone Diabetes Center, The University of Texas Southwestern Medical Center, Dallas, Texas, USA. * William L Holland, * Zhao V Wang, * Kai Sun, * Kathryn E Davis, * Nils Halberg, * Joseph M Rutkowski, * Vincent M Tenorio & * Philipp E Scherer * Institute for Diabetes, Obesity and Metabolism, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, USA. * Russell A Miller & * Morris J Birnbaum * Department of Biochemistry and Molecular Biology, Colorado State University, Fort Collins, Colorado, USA. * Brian M Barth & * Scott A Summers * Eli Lilly and Company, Lilly Corporate Center, Indianapolis, Indiana, USA. * Hai H Bui, * Mark R Wade, * Ming-Shang Kuo & * Joseph T Brozinick * Program in Cardiovascular and Metabolic Diseases, Duke-National University of Singapore Graduate Medical School, Singapore. * Benjamin T Bikman & * Scott A Summers * Department of Biomedical Sciences, Faculty of Health Science, University of Copenhagen, Copenhagen, Denmark. * Nils Halberg * Department of Metabolic Disorders, Merck Research Laboratories, Rahway, New Jersey, USA. * Bei B Zhang * Stedman Center for Nutrition and Metabolism Research, Duke University Medical Center, Durham, North Carolina, USA. * Scott A Summers * Department of Cell Biology, The University of Texas Southwestern Medical Center, Dallas, Texas, USA. * Philipp E Scherer Contributions W.L.H. conducted all experiments, except the portions indicated below, and contributed to the writing of the manuscript. R.A.M. conducted in vivo experiments with liver-specific Lkb1−/− mice. Z.V.W. generated all of the ATTAC mouse models used here. K.S. was responsible for the mutagenesis studies of AdipoR1 and AdipoR2. B.M.B. was involved in the studies with INS-1 cells. H.H.B., M.R.W., M.-S.K. and J.T.B. were involved in liquid chromatography–tandem mass spectrometry analysis for determination of sphingolipid content of the samples. K.E.D. assisted in the generation of the Adipor1−/−Adipor2−/− MEFs and high-fat feeding studies using adiponectin transgenic mice. B.T.B. helped in data analysis and RT-PCR of sphingolipid metabolism genes. N.H. performed the experiments with in vivo injections of adiponectin and detection of the protein in beta cells. J.M.R. was involved in designing experiments and protein production. V.M.T. performed ceramidase assays and geno! typing. B.B.Z., M.J.B., S.A.S. and P.E.S. were involved in experimental design, data analysis and in the writing of the manuscript. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Philipp E Scherer Supplementary information * Abstract * Author information * Supplementary information PDF files * Supplementary Text and Figures (2M) Supplementary Figures 1–6, Supplementary Table 1 and Supplementary Methods Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Adenosine receptor crossroads in sickle cell disease
- Nat Med 17(1):38-40 (2011)
Nature Medicine | Article Detrimental effects of adenosine signaling in sickle cell disease * Yujin Zhang1 Search for this author in: * NPG journals * PubMed * Google Scholar * Yingbo Dai1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Jiaming Wen1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Weiru Zhang1, 3 Search for this author in: * NPG journals * PubMed * Google Scholar * Almut Grenz4 Search for this author in: * NPG journals * PubMed * Google Scholar * Hong Sun5 Search for this author in: * NPG journals * PubMed * Google Scholar * Lijian Tao3 Search for this author in: * NPG journals * PubMed * Google Scholar * Guangxiu Lu6 Search for this author in: * NPG journals * PubMed * Google Scholar * Danny C Alexander7 Search for this author in: * NPG journals * PubMed * Google Scholar * Michael V Milburn7 Search for this author in: * NPG journals * PubMed * Google Scholar * Louvenia Carter-Dawson8 Search for this author in: * NPG journals * PubMed * Google Scholar * Dorothy E Lewis9 Search for this author in: * NPG journals * PubMed * Google Scholar * Wenzheng Zhang9 Search for this author in: * NPG journals * PubMed * Google Scholar * Holger K Eltzschig4 Search for this author in: * NPG journals * PubMed * Google Scholar * Rodney E Kellems1 Search for this author in: * NPG journals * PubMed * Google Scholar * Michael R Blackburn1 Search for this author in: * NPG journals * PubMed * Google Scholar * Harinder S Juneja9 Search for this author in: * NPG journals * PubMed * Google Scholar * Yang Xia1, 6 Contact Yang Xia Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:79–86Year published:(2011)DOI:doi:10.1038/nm.2280Received11 May 2010Accepted19 November 2010Published online19 December 2010 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Abstract * Abstract * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Hypoxia can act as an initial trigger to induce erythrocyte sickling and eventual end organ damage in sickle cell disease (SCD). Many factors and metabolites are altered in response to hypoxia and may contribute to the pathogenesis of the disease. Using metabolomic profiling, we found that the steady-state concentration of adenosine in the blood was elevated in a transgenic mouse model of SCD. Adenosine concentrations were similarly elevated in the blood of humans with SCD. Increased adenosine levels promoted sickling, hemolysis and damage to multiple tissues in SCD transgenic mice and promoted sickling of human erythrocytes. Using biochemical, genetic and pharmacological approaches, we showed that adenosine A2B receptor (A2BR)-mediated induction of 2,3-diphosphoglycerate, an erythrocyte-specific metabolite that decreases the oxygen binding affinity of hemoglobin, underlies the induction of erythrocyte sickling by excess adenosine both in cultured human red blood cells and i! n SCD transgenic mice. Thus, excessive adenosine signaling through the A2BR has a pathological role in SCD. These findings may provide new therapeutic possibilities for this disease. View full text Figures at a glance * Figure 1: Increased adenosine levels contribute to sickling and hemolysis in SCD transgenic mice. () Representative HPLC profile showing adenosine concentrations in the plasma of wild-type (WT) and SCD transgenic (Tg) mice at steady state. () The effect of chronic PEG-ADA treatment on adenosine concentrations in the plasma of wild-type and SCD transgenic mice. () Blood smears of SCD transgenic mice without or with PEG-ADA enzyme therapy. (–) Effects of PEG-ADA treatment on plasma hemoglobin (), plasma haptoglobin () and plasma total bilirubin () concentrations in wild-type and SCD transgenic mice. () Lifespan of RBCs in SCD transgenic mice treated with or without PEG-ADA. Mean ± s.e.m; n = 4–8 mice per group; *P < 0.05 versus wild type; **P < 0.05 versus untreated SCD transgenic mice. * Figure 2: In vivo effects of PEG-ADA treatment on multiple organ damage and renal dysfunction in SCD transgenic mice. () H&E staining of lung, liver and spleen of wild-type and SCD transgenic mice after 8 weeks with or without PEG-ADA treatment. Substantial vascular congestion, vascular damage and necrosis in the lungs, livers and spleens observed in SCD transgenic mice was reduced by PEG-ADA treatment. For lung and spleen, scale bar, 20 μm; for liver, scale bar, 10 μm. (–) Semiquantitative analysis of histological changes in lung, liver and spleen of wild-type and SCD transgenic mice using Image-Pro Plus software (Media Cybernetics). () The effect of PEG-ADA enzyme therapy on heme content in lungs, livers and spleens of wild-type and SCD transgenic mice. () The effect of PEG-ADA enzyme therapy on microinfarction and cysts in renal cortex and on congestion in renal medulla of wild-type and SCD transgenic mice, as analyzed by H&E staining. Scale bar, 10 μm. (,) Semiquantitative analysis of histological changes in renal cortex and renal medulla of wild-type and SCD transgenic mice using ! Image-Pro Plus. (,) The effect of PEG-ADA enzyme therapy on proteinuria () and urine osmolarity () in wild-type and SCD transgenic mice. Mean ± s.e.m.; n = 4–8 mice per group; *P < 0.05 versus wild type; **P < 0.05 versus untreated SCD transgenic mice. ND, not detected. * Figure 3: PEG-ADA treatment attenuates hypoxia-reoxygenation–induced acute sickle crisis in SCD transgenic mice. (–) The effects of hypoxia-reoxygenation, with or without PEG-ADA, on plasma adenosine concentration (), percentage of sickled cells (), plasma hemoglobin concentration () and plasma bilirubin concentration () in SCD transgenic mice. () Lung sections from SCD transgenic mice were stained with an antibody that recognizes Ly-6B.2 (a polymorphic 40-kDa antigen expressed by neutrophils) to visualize infiltrated tissue neutrophils (brown). Scale bar, 10 μm. () Quantification of neutrophil infiltration in the lungs of SCD transgenic mice by Image-Pro Plus software. () The effect of hypoxia-reoxygenation with and without PEG-ADA treatment on proinflammatory cytokine abundance (IFN-γ, IL-6, IL-1β and GM-CSF) in lung homogenates from SCD transgenic mice. Mean ± s.e.m. from 5–7 mice per group; *P < 0.05 relative to SCD transgenic mice under normoxic conditions and **P < 0.05 relative to untreated SCD transgenic mice under hypoxia-reoxygenation conditions. * Figure 4: Excess adenosine acts through A2BRs to induce 2,3-DPG and subsequent sickling in SCD transgenic mice. () Concentration of 2,3-DPG in RBCs of wild-type and SCD transgenic mice with or without chronic PEG-ADA enzyme therapy. () In vivo measurement of oxygen saturation of hemoglobin in wild-type and SCD transgenic mice with or without PEG-ADA treatment for 8 weeks. () 2,3-DPG concentrations in isolated RBCs treated with NECA in the presence or absence of theophylline. () 2,3-DPG concentrations in RBCs isolated from wild-type or mice deficient in each of the four types of adenosine receptors cultured in the presence or absence of NECA. () Immunostaining of A2BRs in RBCs from wild-type and A2BR-deficient mice. () 2,3-DPG concentrations in RBCs isolated from wild-type and adenosine receptor–deficient mice under hypoxic conditions. () cAMP concentrations in RBCs isolated from wild-type and A2BR-deficient mice treated with or without NECA. () 2,3-DPG concentrations in RBCs isolated from wild-type mice treated with NECA in the presence or absence of the PKA-specific inhibitor H-89.! () 2,3-DPG concentrations in RBCs of wild-type and SCD transgenic mice treated with or without PSB1115. () Lifespan of RBCs in SCD transgenic mice treated with or without PSB1115. Mean ± s.e.m.; *P < 0.05 relative to untreated controls, **P < 0.05 relative to treated or positive samples; Each experiment was repeated five to seven times. n = 5–8 mice per group. * Figure 5: Adenosine levels are elevated in individuals with SCD and A2BR-mediated elevation of 2,3-DPG concentrations is required for hypoxia-induced human erythrocyte sickling. () Average adenosine concentrations in plasma from individuals with SCD (SCD, n = 12) and healthy volunteers (control, n = 11). () 2,3-DPG concentration in RBCs isolated from controls (n = 11) and individuals with SCD (n = 12). In and , *P < 0.05. () Changes in 2,3-DPG concentrations in isolated SCD RBCs after incubation under hypoxic conditions in the absence or presence of PEG-ADA, MRS1754 (A2BR antagonist), H-89 (PKA-specific inhibitor) or glycolate (GA; promotes degradation of 2,3-DPG). *P < 0.05 versus normoxic condition; **P < 0.05 versus untreated samples under hypoxic condition; ***P < 0.05 versus NECA-treated samples under hypoxic condition. () Changes in the percentage of sickled cells in isolated SCD RBCs after incubation under varying oxygen concentrations in the absence or presence of PEG-ADA, MRS1754, H-89 or glycolic acid. *P < 0.05 versus untreated cells. In and , each experiment was repeated four to six times. Data in – are mean ± s.e.m. () Working model ! of excessive adenosine signaling in erythrocyte sickling and potential mechanism-based therapies in SCD. Under hypoxic conditions, increased adenosine-mediated 2,3-DPG production induced by activation of A2BRs decreases the O2 binding affinity of HbS, resulting in increased amounts of deoxy-HbS and increased sickling, hemolysis and multiple tissue damage and dysfunction. Without interference, hemolysis and multiple tissue injury lead to increased release of ATP, which is converted to adenosine by the combined action of the ectonucleotidases CD39 and CD73. The use of PEG-ADA to lower adenosine concentrations or an A2BR antagonist to block receptor activation would reduce the production of erythrocyte 2,3-DPG and reduce sickling. Author information * Abstract * Author information * Supplementary information Affiliations * Department of Biochemistry and Molecular Biology, The University of Texas Health Science Center at Houston, Houston, Texas, USA. * Yujin Zhang, * Yingbo Dai, * Jiaming Wen, * Weiru Zhang, * Rodney E Kellems, * Michael R Blackburn & * Yang Xia * Department of Urology, The Third Xiangya Hospital, Changsha, Hunan, China. * Yingbo Dai & * Jiaming Wen * Department of Nephrology, The First Xiangya Hospital, Changsha, Hunan, China. * Weiru Zhang & * Lijian Tao * Department of Anesthesiology, The University of Colorado, Denver, USA. * Almut Grenz & * Holger K Eltzschig * Department of Otorhinolaryngology, The Third Xiangya Hospital, Changsha, Hunan, China. * Hong Sun * Institute of Reproductive & Genetic Medicine, Central South University, Changsha, Hunan, China. * Guangxiu Lu & * Yang Xia * Metabolon, Inc., Durham, North Carolina, USA. * Danny C Alexander & * Michael V Milburn * Department of Ophthalmology and Visual Science, The University of Texas Health Science Center at Houston, Houston, Texas, USA. * Louvenia Carter-Dawson * Department of Internal Medicine, The University of Texas Health Science Center at Houston, Houston, Texas, USA. * Dorothy E Lewis, * Wenzheng Zhang & * Harinder S Juneja Contributions Y.Z. carried out the measurement of adenosine and 2,3-DPG in humans and mice, analysis of sickling, hemolysis and lifespan of mouse RBCs, histological analysis of multiple tissues and image quantification, ELISA analysis of inflammatory cytokines in the lung homogenates and immunostaining of lung tissues with neutrophil markers, human erythrocyte culture and analysis of sickling under hypoxic conditions, and immunostaining of A2BRs on human and mouse RBCs, and contributed to generation of figures. Y.D. conducted PEG-ADA purification, treatment of mice with PEG-ADA or PSB1115 and proteinuria measurement, and isolation of multiple organs and blood from mice. J.W. was involved in the purification of PEG-ADA and treatment of mice with PEG-ADA or PSB1115, and performed heme content measurement and histological analysis of kidneys. W.Z. was involved in the purification of PEG-ADA; treated mice with PEG-ADA or PSB1115 and contributed to immunostaining of lung tissues with neutrophi! l markers. A.G. treated normal human erythrocyte cultures with A2BR or A2AR agonists. D.C.A. and M.V.M. conducted metabolomic screens in blood of wild-type and SCD transgenic mice. L.C.-D. provided expertise in confocal analysis of A2BR expression on RBCs. D.E.L. provided expertise in flow cytometry to measure the lifespan of RBCs. W.Z. assisted with urine osmolality analysis. H.S., L.T. and G.L. provided expertise in hemolytic disorders and kidney dysfunction. H.K.E. assisted A.G. with experiments on the effects of A2AR and A2BR agonists on 2,3-DPG induction in normal RBCs. R.E.K. provided expertise in adenosine signaling and helped edit the manuscript; M.R.B. provided mice deficient in each of the four types of adenosine receptors; H.S.J. provided expertise in hemolytic disorders, procured human subjects' approval and maintained the database of de-identified human subject information. Y.X. was the principal investigator, oversaw the design of experiments and interpretatio! n of results, wrote and organized the manuscript, including th! e text and figures, and edited the manuscript. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Yang Xia Supplementary information * Abstract * Author information * Supplementary information PDF files * Supplementary Text and Figures (944K) Supplementary Methods, Supplementary Tables 1 and 2 and Supplementary Figures 1–5 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Catapulting clopidogrel pharmacogenomics forward
- Nat Med 17(1):40-41 (2011)
Nature Medicine | Letter Paraoxonase-1 is a major determinant of clopidogrel efficacy * Heleen J Bouman1, 2, 3 Search for this author in: * NPG journals * PubMed * Google Scholar * Edgar Schömig4 Search for this author in: * NPG journals * PubMed * Google Scholar * Jochem W van Werkum1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Janna Velder5 Search for this author in: * NPG journals * PubMed * Google Scholar * Christian M Hackeng1, 6 Search for this author in: * NPG journals * PubMed * Google Scholar * Christoph Hirschhäuser5 Search for this author in: * NPG journals * PubMed * Google Scholar * Christopher Waldmann7 Search for this author in: * NPG journals * PubMed * Google Scholar * Hans-Günther Schmalz5 Search for this author in: * NPG journals * PubMed * Google Scholar * Jurriën M ten Berg1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Dirk Taubert4 Contact Dirk Taubert Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:110–116Year published:(2011)DOI:doi:10.1038/nm.2281Received14 July 2010Accepted22 November 2010Published online19 December 2010Corrected online21 December 2010 Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Clinical efficacy of the antiplatelet drug clopidogrel is hampered by its variable biotransformation into the active metabolite1, 2. The variability in the clinical response to clopidogrel treatment has been attributed to genetic factors, but the specific genes and mechanisms underlying clopidogrel bioactivation remain unclear. Using in vitro metabolomic profiling techniques, we identified paraoxonase-1 (PON1) as the crucial enzyme for clopidogrel bioactivation, with its common Q192R polymorphism determining the rate of active metabolite formation. We tested the clinical relevance of the PON1 Q192R genotype in a population of individuals with coronary artery disease who underwent stent implantation and received clopidogrel therapy. PON1 QQ192 homozygous individuals showed a considerably higher risk than RR192 homozygous individuals of stent thrombosis, lower PON1 plasma activity, lower plasma concentrations of active metabolite and lower platelet inhibition. Thus, we identif! ied PON1 as a key factor for the bioactivation and clinical activity of clopidogrel. These findings have therapeutic implications and may be exploited to prospectively assess the clinical efficacy of clopidogrel. View full text Figures at a glance * Figure 1: Kinetics of clopidogrel-metabolizing enzymes. (,) Substrate saturation kinetics for microsomal preparations of human cytochrome P450 isozymes () and esterases () involved in clopidogrel metabolism. Microsomes were obtained from stably transfected human embryonic kidney cells (HEK 293). Conversion of clopidogrel to 2-oxo-clopidogrel, of 2-oxo-clopidogrel to the active thiol metabolite and of clopidogrel to the thiol metabolite was assayed by incubation with increasing concentrations of substrate for 5 min at 37 °C. For esterases, the conversion of clopidogrel to clopidogrel-carboxylate, of 2-oxo-clopidogrel to 2-oxo-clopidogrel-carboxylate, and of the thiol metabolite to thiol metabolite-carboxylate was also assayed. Kinetics of the PON1 allozymes with the L55M and Q192R polymorphisms were determined. For each enzyme a specific probe reaction (positive control) was performed (black circles). Symbols and error bars represent means ± s.e.m. of three independent incubation experiments each. Supplementary Figure 1 shows si! milar kinetic measurements for enzymes not involved in clopidogrel metabolism. BChE, butyrylcholinesterase; CES1, carboxylesterase-1; CES2, carboxylesterase-2. DTNB, 5,5′-dithiobis-(2-nitrobenzoic acid). * Figure 2: Kaplan-Meier curves for individuals with coronary stent implantation and their pharmacokinetic and pharmacodynamic responses to clopidogrel. (,) Kaplan-Meier survival curves showing cumulative probabilities of case-cohort subjects without incident nonfatal definite stent thrombosis over 18 months of follow-up according to PON1 Q192R gene polymorphism () and according to tertiles (T1, T2 and T3) of paraoxonase plasma activity, with T1 < 104.9, T2 < 180.4–104.9 and T3 ≥ 180.4 nmol min−1 ml−1 (). Genotype distribution and tertile limits were extrapolated to the total cohort, and subcohort noncases were weighted with the inverse of the sampling fraction according to a previously described method33. Numbers of individuals at risk at the indicated time points are shown. P values for the total model coefficients were calculated by univariate Cox regression. () Box-and-whisker plots showing median response values with twenty-fifth and seventy-fifth percentiles (box) and tenth and ninetieth percentiles (whisker). After the 18-month follow-up, the clopidogrel-free cases with nonfatal stent thrombosis (ST, n = 41) a! nd the subcohort noncases without stent thrombosis (No ST, n = 71) were given a single 600-mg clopidogrel dose, and pharmacokinetic and platelet responses were compared by univariate Cox regression. Pharmacokinetic responses are indicated as maximum plasma concentrations (cmax, ng ml−1) of active thiol metabolite, 2-oxo-clopidogrel, parent clopidogrel and carboxylic acid metabolite and as ratio of the maximum plasma concentrations of 2-oxo-clopidogrel to thiol metabolite. Platelet response is indicated as percentage of maximal predose versus maximal 6-h postdose aggregation (Δ% induced by 20 μM ADP), Paraoxonase and arylesterase plasma activities are indicated as the velocity of transformation (nmol min−1 ml−1) of paraoxon to p-nitrophenol and of phenylacetate to phenol, respectively. P < 0.05 was considered a statistically significant difference. Change history * Change history * Author information * Supplementary informationErratum 21 December 2010 In the version of this article originally published online, the affiliations for Hans-Günther Schmalz and Dirk Taubert appeared incorrectly. Hans-Günther Schmalz is in the Department für Chemie, Universität zu Köln, Cologne, Germany, and Dirk Taubert is in the Department of Pharmacology, University Hospital of Cologne, Cologne, Germany. These errors have been corrected for the print, PDF and HTML versions of the article. Author information * Change history * Author information * Supplementary information Affiliations * Department of Cardiology, St. Antonius Hospital Nieuwegein, Nieuwegein, The Netherlands. * Heleen J Bouman, * Jochem W van Werkum, * Christian M Hackeng & * Jurriën M ten Berg * St. Antonius Center for Platelet Function Research, St. Antonius Hospital Nieuwegein, Nieuwegein, The Netherlands. * Heleen J Bouman, * Jochem W van Werkum & * Jurriën M ten Berg * Department of Biochemistry, Cardiovascular Research Institute Maastricht, University Maastricht, Maastricht, The Netherlands. * Heleen J Bouman * Department of Pharmacology, University Hospital of Cologne, Cologne, Germany. * Edgar Schömig & * Dirk Taubert * Department für Chemie, Universität zu Köln, Cologne, Germany. * Janna Velder, * Christoph Hirschhäuser & * Hans-Günther Schmalz * Department of Clinical Chemistry, St. Antonius Hospital Nieuwegein, Nieuwegein, The Netherlands. * Christian M Hackeng * Klinik und Poliklinik für Nuklearmedizin, Universität Münster, Münster, Germany. * Christopher Waldmann Contributions J.W.v.W. and D.T. conceived the project. E.S., J.M.t.B. and D.T. supervised the project. H.J.B., J.W.v.W., J.V., C.M.H., C.H., C.W., H.-G.S. and D.T. conducted or directed bioanalytics of clopidogrel and its metabolites, metabolomic profiling, blood collection and sample preparation, aggregometry, genotyping and PON1 phenotyping. H.J.B., E.S., J.W.v.W., C.M.H., J.M.t.B. and D.T. conducted or directed recruitment of subjects, disease assessment and follow-up assessments. H.J.B., J.W.v.W. and D.T. did the computational and statistical data analyses. H.J.B. and D.T. wrote the manuscript. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Dirk Taubert Supplementary information * Change history * Author information * Supplementary information PDF files * Supplementary Text and Figures (968K) Supplementary Tables 1–10, Supplementary Figures 1–9 and Supplementary Methods Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Behind the paper: Stepping up to the antiplatelet
- Nat Med 17(1):42 (2011)
Nature Medicine | News and Views Behind the paper: Stepping up to the antiplatelet * Elie Dolgin Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:42Year published:(2011)DOI:doi:10.1038/nm0111-42Published online07 January 2011 Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. In March 2002, a middle-aged man was admitted to the German Heart Centre in Munich to receive treatment for a blocked coronary artery. After undergoing balloon angioplasty and having a stent implanted, the man needed some sort of blood-thinning therapy, yet aspirin, the workhorse treatment for preventing arterial clots, gave him asthma attacks. His cardiologist, Nicolas von Beckerath, turned to clopidogrel, an antiplatelet drug that had been approved in the EU four years earlier under the brand name Plavix. The man tolerated the drug well, but clopidogrel did not seem to inhibit his platelet activity, even at very high doses. "Surprisingly, we saw that this patient was absorbing the drug very well, but he wasn't metabolizing it," says von Beckerath, now at the Viersen General Hospital in Germany. View full text Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Histamine restricts cancer: nothing to sneeze at
- Nat Med 17(1):43-44 (2011)
Nature Medicine | Article Histamine deficiency promotes inflammation-associated carcinogenesis through reduced myeloid maturation and accumulation of CD11b+Ly6G+ immature myeloid cells * Xiang Dong Yang1 Search for this author in: * NPG journals * PubMed * Google Scholar * Walden Ai1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Samuel Asfaha1 Search for this author in: * NPG journals * PubMed * Google Scholar * Govind Bhagat3 Search for this author in: * NPG journals * PubMed * Google Scholar * Richard A Friedman4 Search for this author in: * NPG journals * PubMed * Google Scholar * Guangchun Jin1 Search for this author in: * NPG journals * PubMed * Google Scholar * Heuijoon Park1 Search for this author in: * NPG journals * PubMed * Google Scholar * Benjamin Shykind5 Search for this author in: * NPG journals * PubMed * Google Scholar * Thomas G Diacovo6 Search for this author in: * NPG journals * PubMed * Google Scholar * Andras Falus7 Search for this author in: * NPG journals * PubMed * Google Scholar * Timothy C Wang1 Contact Timothy C Wang Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:87–95Year published:(2011)DOI:doi:10.1038/nm.2278Received18 August 2010Accepted16 November 2010Published online19 December 2010 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Abstract * Abstract * Accession codes * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Histidine decarboxylase (HDC), the unique enzyme responsible for histamine generation, is highly expressed in myeloid cells, but its function in these cells is poorly understood. Here we show that Hdc-knockout mice show a high rate of colon and skin carcinogenesis. Using Hdc-EGFP bacterial artificial chromosome (BAC) transgenic mice in which EGFP expression is controlled by the Hdc promoter, we show that Hdc is expressed primarily in CD11b+Ly6G+ immature myeloid cells (IMCs) that are recruited early on in chemical carcinogenesis. Transplant of Hdc-deficient bone marrow to wild-type recipients results in increased CD11b+Ly6G+ cell mobilization and reproduces the cancer susceptibility phenotype of Hdc-knockout mice. In addition, Hdc-deficient IMCs promote the growth of tumor allografts, whereas mouse CT26 colon cancer cells downregulate Hdc expression through promoter hypermethylation and inhibit myeloid cell maturation. Exogenous histamine induces the differentiation of IMCs ! and suppresses their ability to support the growth of tumor allografts. These data indicate key roles for Hdc and histamine in myeloid cell differentiation and CD11b+Ly6G+ IMCs in early cancer development. View full text Figures at a glance * Figure 1: Histamine-deficient Hdc−/− mice are highly susceptible to colorectal and skin carcinogenesis. () Macroscopic appearance of colorectal tumors from wild-type (WT) and Hdc−/− mice. Hdc−/− mice and wild-type mice (both male BALB/c background) were injected with a single dose of AOM followed by DSS in the drinking water for 10 d. () The number of large (diameter >3 mm) and small (diameter <3 mm) colorectal tumors in Hdc−/− mice versus wild-type mice (*P < 0.05; means ± s.d.; n = 8, each group). () Representative H&E-stained colonic tumor sections from wild-type and Hdc−/− mice. Scale bar, 50 μm. () Macroscopic appearance of the skin tumors from wild-type and Hdc−/− mice. Wild-type and Hdc−/− mice were treated with the tumor initiator DMBA and then treated twice weekly with the tumor promoter TPA. () The number of skin papillomas in Hdc−/− mice versus wild-type mice, including both large (diameter >2.5 mm) and small (diameter <2.5 mm) papillomas (*P < 0.05; means ± s.d.; n = 5, each group). () Representative H&E-stained skin tumor sections fr! om wild-type and Hdc−/− mice. Scale bar, 50 μm. * Figure 2: CD11b+Ly6G+ IMCs are the predominant source of Hdc-EGFP expression in the bone marrow. () Left, the percentage of EGFP+ cells within the bone marrow, spleen and peripheral blood of Hdc-EGFP mice. Right, the relative proportion of CD11b+ and Gr-1+ cells gated from the EGFP+ subset. FSC, forward scatter. Gr-1, granulocyte-differentiation antigen-1. () Expression of CD11b+Ly6G+ cells. CD11b+Gr-1+ myeloid cells were gated from the bone marrow of Hdc-EGFP mice. This FACS analysis indicates the proportion of EGFP+ cells that were CD11b+Ly6Ghigh, CD11b+Ly6Gmedium and CD11b+Ly6G− cells. This panel shows representative data from five experiments. Ly6G, lymphocyte antigen 6G. () FACS analysis of peritoneal exudative cells (PECs) of Hdc-EGFP mice with cell surface markers for myeloid cells and mast cells. c-kit, mouse CD117. FcεR, high affinity receptor. () Differentiation of splenic Hdc-EGFP+ IMCs. Immunofluorescence of DAPI-stained EGFP+CD11b+Ly6G+ IMCs treated with or without G-CSF for 48 h, showing suppression of EGFP expression with granulocyte differentiation (t! he images are representative of three independent experiments). * Figure 3: Hdc deficiency upregulates CD11b+Gr-1+ and CD11b+Ly6G+ IMCs. () FACS analysis of the percentage of CD11b+Gr-1+ IMCs in the bone marrow, spleen and peripheral blood in wild-type and Hdc−/− mice (*P < 0.05; means ± s.d.; n = 10, each group). () The relative proportion of CD11b+Ly6G+ cells in the peripheral blood and spleen of wild-type and Hdc−/− mice, with and without AOM plus DSS–induced colon tumors, as measured by FACS analysis (*P < 0.05; **P < 0.01; means ± s.d.; n = 5, each group). () FACS analysis showing the abundance of the Ly6G and Ly6C subsets in CD11b+ cells in the peripheral blood and spleen of wild-type and Hdc−/− mice. The panel is representative of data from five mice of each group. () Differentiation of IMCs. In vitro incubation of bone marrow–derived EGFP+ CD11b+Gr-1+ IMCs with GM-CSF and histamine followed by FACS analysis for Ly6G and Ly6C surface expression. The figure is representative of data from three independent experiments. () FACS analysis of the proportion of CD11b+Ly6C+ monocytes among CD! 11b+Gr-1+ IMCs sorted from the bone marrow of Hdc−/− mice and incubated with histamine; GM-CSF; histamine and GM-CSF; histamine, GM-CSF and H1R antagonist; histamine, GM-CSF and H2R antagonist; or histamine, GM-CSF, H1R and H2R antagonists (*P < 0.05; **P < 0.01; Student's t test. Data represent means ± s.d. from three independent experiments). () Effect of exogenous histamine on circulating CD11b+Gr-1+ IMCs in Hdc−/− mice. The percentage of CD11b+Gr-1+ IMCs by FACS in the peripheral blood of wild-type, Hdc−/−, and Hdc−/− mice treated with exogenous histamine (*P < 0.05; means ± s.d.; n = 5, each group). * Figure 4: EGFP+ IMCs are recruited to inflamed tissue and stroma of colon tumor. () Immunofluorescence and H&E sections from AOM plus DSS–treated (20 weeks) and control Hdc-EGFP mice (n = 5). Top, rare submucosal EGFP+ cells in the colon of control mice and a large number of EGFP+ cells in the colonic adenomas of AOM plus DSS–treated mice. H&E staining of colonic tumors confirmed abundant inflammatory cells in the stroma. Bottom, immunostaining of serial sections showing that EGFP+ cells are Hdc+, and approximately 50% of CD11b+ and Gr-1+ cells are EGFP+. () Left, immunofluorescence image showing EGFP+ cells in the DAPI-stained colon of Hdc-EGFP mice treated with DSS (10 d). Right, immunofluorescence image showing a greater number of EGFP+ cells in the DAPI-stained colon of Hdc-EGFP mice 8 weeks after AOM plus DSS treatment (compared to , before tumor development). () Immunofluorescence image of DAPI-stained colon from bone marrow–transplanted (BMT) mice 10 d after DSS, showing that infiltrating EGFP+ cells are bone marrow–derived (donor: Hdc-EGF! P mice; recipient: wild-type B6 mice). () Immunofluorescence stained human colonic inflammatory bowel disease (IBD) and colitis-associated carcinoma tissue sections showing HDC (green), CD11b (red) or tryptase (red). White arrows show HDC+ cells (which all colocalize with CD11b+ cells in IBD tissue); red arrows show cells that express CD11b or tryptase alone. The figure is representative of analyses of five IBD and five cancer samples. () Immunofluorescence and H&E-stained sections showing the presence of inflammatory cells and EGFP+ cells in the skin after a single TPA treatment. Top, H&E-stained skin sections. Bottom, immunofluorescence showing a large number of EGFP+ inflammatory cells in the dermis 24 h after TPA treatment, compared to 6 h after TPA treatment or to acetone-treated controls (n = 5). * Figure 5: Bone marrow derived IMCs from Hdc−/− mice accelerate tumor growth. () Colonic tumor numbers in bone marrow–transplanted mice at 20 weeks after AOM plus DSS treatment. Wild-type and Hdc−/− mice were lethally irradiated and reconstituted with wild-type or Hdc−/− bone marrow (*P < 0.05 compared to indicated control; means ± s.d. n = 6, each group). () Allograft tumor weight. CT26 colon cancer cells were implanted in NOD-SCID mice alone (control) or with CD11b+Ly6G+ bone marrow–derived IMCs from wild-type mice or Hdc−/− mice, with or without exogenous histamine (800 μg per kg body weight per day, intraperitoneal injection) (*P < 0.05; **P < 0.01; means ± s.d.; n = 7, each group). () Cytokine mRNA expression in bone marrow–derived IMCs. mRNA expression of IL-1α, IL-1β, IL-6, TGF-β and tumor necrosis factor-α (TNF-α) in CD11b+Ly6G+ IMCs from Hdc−/− mice compared to IMCs from wild-type mice were analyzed by qRT-PCR. () Effect of Il6 deficiency on promotion of tumor allograft growth by IMCs. CT26 cancer cells were impl! anted alone or concurrently with CD11b+Ly6G+ bone marrow IMCs isolated from wild-type or Il6−/− mice into NOD-SCID mice. Tumor weight was measured (*P < 0.05; means ± s.d.; n = 7). (,) The number of vessel branch points in allograft tumors derived from CT26 cancer cells injected with no CD11b+Ly6G+ IMCs (control) or IMCs from wild-type mice, Hdc−/− mice, Hdc−/− mice plus histamine or Il6−/− mice (*P < 0.05; means ± s.d.; n = 7, each group). * Figure 6: Migration of circulating CD11b+Gr-1+ IMCs is RAGE dependent, and suppression of Hdc occurs through a methylation-dependent mechanism. () Immunofluorescence staining of allograft tumor sections with or without concurrently implanted EGFP+ CD11b+Gr-1+ IMCs. Antibodies to proliferation marker Ki-67 and to myofibroblast α-SMA were applied. () Immunohistochemistry (IHC) staining show α-SMA+ myofibroblasts in the stroma of colonic adenomas from Hdc−/− mice treated with AOM and DSS compared to wild-type mice. The images in and show representative data from five tumors in each group. () qRT-PCR analysis of RAGE ligand S100A8 and monocyte chemoattractant protein-1 (MCP-1) mRNA expression in DSS-induced colitis tissue (*P < 0.05; means ± s.d.; n = 3, each group). () qRT-PCR analysis of RAGE ligands S100A8 and S100A9 mRNA expression in colonic tumors of Hdc−/− mice compared to wild-type mice treated with AOM and DSS (*P < 0.05; means ± s.d.; n = 3, each group). () FACS analysis of CD11b+Gr-1+ IMCs in the peripheral blood of Rage−/− mice after AOM plus DSS treatment compared to wild-type mice (*P < 0.0! 5; means ± s.d.; n = 5, each group). () Bone marrow–derived Hdc-EGFP–expressing CD11b+Ly6G+ IMCs were cultured with IEC-6 rat intestinal epithelial cells and CT26 cancer cells, respectively for 48 h; FACS analysis showing the percentage of EGFP−CD11b+ myeloid cells after co-culture with CT26 cancer cells compared to co-culture with IEC-6 cells and control IMCs (*P < 0.05; **P < 0.01; means ± s.d.; n = 3, each group). () Hdc promoter methylation analysis by sodium bisulfate DNA sequencing. Bone marrow–derived Hdc-EGFP+ CD11b+Ly6G+ IMCs were cultured with CT26 cancer cells for 48 h, and DNA of IMCs was extracted for detection of DNA CpG methylation sites. Top, bent arrow shows the transcription start site and boxes represent the GC box and exon 1. Short vertical lines across the horizontal line indicate CpG sites (−800 and +200 bp from the transcript start site). Bottom, filled and open circles represent methylated and unmethylated cytosine residues respectively. Accession codes * Abstract * Accession codes * Author information * Supplementary information Referenced accessions Gene Expression Omnibus * GSE23502 Author information * Abstract * Accession codes * Author information * Supplementary information Affiliations * Division of Digestive and Liver Diseases, Department of Medicine and Irving Cancer Center, Columbia University, New York, New York, USA. * Xiang Dong Yang, * Walden Ai, * Samuel Asfaha, * Guangchun Jin, * Heuijoon Park & * Timothy C Wang * Department of Pathology, Microbiology and Immunology, University of South Carolina School of Medicine, Columbia, South Carolina, USA. * Walden Ai * Department of Pathology and Cell Biology, Columbia University, New York, New York, USA. * Govind Bhagat * Department of Biomedical Informatics, Columbia University, New York, New York, USA. * Richard A Friedman * Department of Neuroscience, Columbia University, New York, New York, USA. * Benjamin Shykind * Department of Pediatrics, Columbia University, New York, New York, USA. * Thomas G Diacovo * Department of Genetics, Cell and Immunobiology, Semmelweis University, Budapest, Hungary. * Andras Falus Contributions X.D.Y. was involved in the study design, completion of experiments, data analysis and interpretation and manuscript preparation. W.A. constructed the Hdc-EGFP transgenic mouse and helped with examination of Hdc-EGFP expression. S.A. helped with the data interpretation and contributed to the manuscript preparation and revision. G.B. provided human colon specimen collection and did pathology assessments. R.A.F. carried out the microarray data analysis. G.J. helped with the colon cancer experiments and data analysis. H.P. helped with the skin carcinogenesis experiments and data analysis. B.S. performed histological analysis of the brains of Hdc-EGFP mice. T.G.D. carried out intravital microscopy studies. A.F. constructed the Hdc-knockout mice and provided helpful suggestions for our study. T.C.W. designed the study and contributed to the data analysis and writing of the manuscript. All authors discussed the results and commented on the manuscript. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Timothy C Wang Supplementary information * Abstract * Accession codes * Author information * Supplementary information Movies * Supplementary Video 1 (2M) Circulating EGFP-expressing blood cells in the ears of Hdc-EGFP mice with TPA-induced skin inflammation. An acute inflammatory response was induced in the ears of Hdc-EGFP mice by the application of a single dose of TPA (8.5 nmol, 20 μl per mouse). The video records 10 seconds of EGFP-expressing blood cells in the vessels using intravital microscopy 6 h after TPA treatment. PDF files * Supplementary Text and Figures (2M) Supplementary Figures 1–17, Supplementary Tables 1–4 and Supplementary Methods Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Filtering new facts about kidney disease
- Nat Med 17(1):44-45 (2011)
Nature Medicine | Letter Podocyte-secreted angiopoietin-like-4 mediates proteinuria in glucocorticoid-sensitive nephrotic syndrome * Lionel C Clement1 Search for this author in: * NPG journals * PubMed * Google Scholar * Carmen Avila-Casado2, 5 Search for this author in: * NPG journals * PubMed * Google Scholar * Camille Macé1, 5 Search for this author in: * NPG journals * PubMed * Google Scholar * Elizabeth Soria2 Search for this author in: * NPG journals * PubMed * Google Scholar * Winston W Bakker3 Search for this author in: * NPG journals * PubMed * Google Scholar * Sander Kersten4 Search for this author in: * NPG journals * PubMed * Google Scholar * Sumant S Chugh1 Contact Sumant S Chugh Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:117–122Year published:(2011)DOI:doi:10.1038/nm.2261Received11 August 2010Accepted18 October 2010Published online12 December 2010 Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg The main manifestations of nephrotic syndrome include proteinuria, hypoalbuminemia, edema, hyperlipidemia and lipiduria. Common causes of nephrotic syndrome are diabetic nephropathy, minimal change disease (MCD), focal and segmental glomerulosclerosis (FSGS) and membranous nephropathy. Among the primary glomerular diseases, MCD is usually sensitive to glucocorticoid treatment, whereas the other diseases show variable responses1. Despite the identification of key structural proteins in the glomerular capillary loop which may contribute to defects in ultrafiltration, many of the disease mechanisms of nephrotic syndrome remain unresolved. In this study, we show that the glomerular expression of angiopoietin-like-4 (Angptl4), a secreted glycoprotein, is glucocorticoid sensitive and is highly upregulated in the serum and in podocytes in experimental models of MCD and in the human disease. Podocyte-specific transgenic overexpression of Angptl4 (NPHS2-Angptl4) in rats induced nephr! otic-range, and selective, proteinuria (over 500-fold increase in albuminuria), loss of glomerular basement membrane (GBM) charge and foot process effacement, whereas transgenic expression specifically in the adipose tissue (aP2-Angptl4) resulted in increased circulating Angptl4, but no proteinuria. Angptl4−/− mice that were injected with lipopolysaccharide (LPS) or nephritogenic antisera developed markedly less proteinuria than did control mice. Angptl4 secreted from podocytes in some forms of nephrotic syndrome lacks normal sialylation. When we fed the sialic acid precursor N-acetyl-D-mannosamine (ManNAc) to NPHS2-Angptl4 transgenic rats it increased the sialylation of Angptl4 and decreased albuminuria by more than 40%. These results suggest that podocyte-secreted Angptl4 has a key role in nephrotic syndrome. View full text Figures at a glance * Figure 1: Angptl4 mRNA and protein expression in experimental glomerular disease. () Induction of proteinuria in rats 24 h after injection of γ2-NTS. () Upregulation of glomerular Angptl4 mRNA in rats injected with γ2-NTS. () Proteinuria in Angptl4−/− and Angptl4+/+ mice after injection of PBS or LPS. () Confocal expression of Angptl4 in rat glomeruli and colocalization with podocyte protein CD2AP. Controls to show the specificity of staining by the Angptl4-specific antibody involved the use of post-absorbed antibodies ('post abs.'); details are in the Online Methods. () Assessment of changes in glomerular Angptl4 expression in rat models of MCD (puromycin nephrosis, PAN), membranous nephropathy (passive Heymann nephritis, PHN), mesangial injury (anti-Thy1.1 nephritis) and severe focal and segmental glomerulosclerosis (non-HIV collapsing glomerulopathy, CG). Threshold for significance was threefold change. Red arrows, onset of proteinuria. D1, D3, D6 and D10 represent days after induction. Numbers above each bar are mean values. () Confocal assessme! nt of Angptl4 expression (red) in control and PAN day 6 glomeruli, and colocalization with GBM heparan sulfate proteoglycan (white; top and middle images) and podocyte protein nephrin (green, overlap yellow; bottom image). () Immunogold electron microscopy of PAN day 6 rat glomeruli to demonstrate Angptl4 expression (gold particles) in the podocytes (yellow arrows) and GBM (black arrows). Scale bars: 7.5 μm (), 10 μm () and 0.33 μm (). LCM, laser capture microdissection; EFP, effaced foot processes; endo, endothelium. * Figure 2: Characterization of male Angptl4 transgenic mice and rats. () Light microscopy (left) and confocal assessment (middle) of Angptl4 and confocal assessment of ZO-1 (right) expression in Angptl4 transgenic (TG) and wild-type (WT) mice. () Electron microscopy of 3-month-old transgenic mouse glomeruli, showing intact (FP) and effaced podocyte foot processes (EFP). () Immunogold electron microscopy for Angptl4 showing gold particles in podocytes and GBM (arrows) in Angptl4 transgenic mice. () Proteinuria in 3-month-old Angptl4 transgenic mice. () Rat Angptl4 transgenic constructs for the targeted expression of Angptl4 in podocytes (NPHS2-Angptl4, left) and adipose tissue (aP2-Angptl4, right) in rats. () Multi-organ mRNA expression profile of Angptl4 in podocyte-specific (left) and adipose tissue–specific (right) transgenic rats. Sk. muscle, skeletal muscle; BAT, brown adipose tissue; WAT, white adipose tissue. () Periodic acid Schiff–stained sections from 3-month-old wild-type and heterozygous transgenic rats. Arrows, prominent podocy! tes in NPHS2-Angptl4 transgenic rats. () Confocal expression of Angptl4 (red) in NPHS2-Angptl4 transgenic rat glomeruli and colocalization with podocyte protein nephrin (green, overlap yellow) and GBM heparan sulfate proteoglycan (blue, overlap fuchsia). () Electron microscopy of a glomerular capillary loop from a 5-month-old homozygous NPHS2-Angptl4 transgenic rat, showing diffuse foot process effacement (arrows). () Immunogold electron microscopy for Angptl4 in NPHS2-Angptl4 transgenic rats of increasing age (left to right), with transition from intact foot processes to foot process effacement (first noted around age 3 months, middle image), and clustering of gold particles in the GBM noted prominently in areas opposite to effaced foot processes (middle and right panels). Scale bars: 10 μm (), 1 μm (), 0.25 μm (), 10 μm (), 8 μm (), 1 μm (), 0.2 μm (). Endo, endothelium. **P < 0.01; ***P < 0.001. * Figure 3: Relationship between Angptl4 overexpression and proteinuria. () Albuminuria in female NPHS2-Angptl4 transgenic rats. () Albuminuria in male heterozygous NPHS2-Angptl4 transgenic rats. () Albuminuria in male homozygous NPHS2-Angptl4 transgenic rats. () GelCode blue–stained SDS PAGE of urinary protein from transgenic rats, rats with PAN and individuals with MCD and membranous nephropathy (MN). Arrow indicates prominent 70-kDa intact albumin band. Mean percentage densitometry of intact albumin is shown for each lane (see Supplementary Fig. 3d). MW, molecular weight; NA, not applicable. () Proteinuria after induction of low-dose PAN in heterozygous male NPHS2-Angptl4 transgenic and wild-type littermates. () Proteinuria in Wistar rats treated with glucocorticoids (PAN-S) or PBS (PAN) on alternate days starting 1 d after induction of PAN. () Glomerular Angptl4 mRNA expression in PAN rats described in . *P < 0.05; **P < 0.01; ***P < 0.001. * Figure 4: Relationship between Angptl4 sialylation and proteinuria. () Two-dimensional gel electrophoresis and western blot of protein from perfused glomeruli show neutral and high pI, low-order Angptl4 oligomers (pink and orange arrows) in control, PAN day 6 and glucocorticoid-treated PAN day 6 rats (from experiment in Fig. 3f). Reactivity of sialic acid binding lectin MAA to these oligomers is also assessed (exemplified for PAN, excerpts from independent blots). () Densitometry of total, neutral- and high-pI oligomers shown in . () Two-dimensional gel electrophoresis and western blot of concentrated supernatant from Angptl4-HEK293 stable cell line incubated with ManNAc or control, and analyzed for Angptl4 expression and binding with sialic acid–binding lectin MAA. Green arrow and line highlight high-pI protein in the control treated group; blue arrow in the ManNAc-treated group shows neutral-pI protein. () Same study as , except done with supernatant from the GEC stable cell line. () Two-dimensional gel electrophoresis and western blot s! tudies of glomerular protein from NPHS2-Angptl4 transgenic rats given tap water or tap water with ManNAc for 12 d. Blots were analyzed for neutral pI (enclosed in red ovals) and high pI (enclosed in green ovals) Angptl4 using anti-Angptl4 antibody and sialic acid–binding lectin from Sambucus nigra (SNA I). () Percentage of neutral- and high-pI Angptl4 in each group assessed by densitometry. () Albuminuria in NPHS2-Angptl4 transgenic rats given tap water (Control group) or tap water with ManNAc (Treatment group) for 12 d (Treatment group, ManNAc phase), followed by plain tap water for 24 d (Treatment group, Washout phase). Values are expressed as a percentage of the baseline albuminuria (designated as 100%). Individual tracings are shown in Supplementary Figure 8. In , significance is for difference from control values. In , significance is for difference from baseline values. Loading controls shown in Supplementary Figure 6. **P < 0.01; ***P < 0.001. Accession codes * Accession codes * Author information * Supplementary information Referenced accessions Entrez Nucleotide * AF487463.1 Author information * Accession codes * Author information * Supplementary information Primary authors * These authors contributed equally to this work. * Carmen Avila-Casado & * Camille Macé Affiliations * Glomerular Disease Therapeutics Laboratory, and Nephrology Research and Training Center, University of Alabama at Birmingham, Birmingham, Alabama, USA. * Lionel C Clement, * Camille Macé & * Sumant S Chugh * Department of Pathology, Instituto Nacional de Cardiologia, Mexico City, Mexico. * Carmen Avila-Casado & * Elizabeth Soria * Department of Pathology and Medical Biology, University Medical Center of Groningen, Groningen, The Netherlands. * Winston W Bakker * Division of Human Nutrition, Wageningen University, Wageningen, The Netherlands. * Sander Kersten Contributions L.C.C. was lead postdoctoral fellow, conducted most of the animal studies, generated stable cell lines and purified recombinant protein, maintained transgenic rats and conducted all studies on transgenic rats, confocal imaging and in situ hybridization, most gene expression studies and selected two-dimensional gel studies. C.A.-C. interpreted and analyzed light microscopy, electron microscopy and immunogold electron microscopy sections for the study and conducted studies on induction of collapsing glomerulopathy in rats. C.M. conducted most of the two-dimensional gel electrophoresis and proteomic work, and most of the albumin ELISA assays. E.S. acted as electron microscopist and morphometrics expert, prepared tissue for electron microscopy, conducted and imaged conventional and most immunogold electron microscopy studies, and conducted alcian blue charge studies. W.W.B. provided human sera and assisted in study design and preparation of the manuscript. S.K. conducted studies! on Angptl4 transgenic mice, provided tissue for histological and gene expression analysis, conducted Angptl4−/− mouse studies with NTS and made substantial contributions to the preparation of the manuscript. S.S.C. acted as senior investigator, planned and supervised the study, generated constructs for transgenic rats, conducted molecular biology and gene expression studies, conducted early animal studies, and wrote and revised the manuscript with input from the other authors. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Sumant S Chugh Supplementary information * Accession codes * Author information * Supplementary information PDF files * Supplementary Text and Figures (2M) Supplementary Figures 1–11, Supplementary Tables 1 and 2 and Supplementary Methods Read the full article * Instant access to this article: US$18Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - The ART of preventing HIV
- Nat Med 17(1):46-47 (2011)
Nature Medicine | Community Corner The ART of preventing HIV Journal name:Nature MedicineVolume: 17,Pages:46–47Year published:(2011)DOI:doi:10.1038/nm0111-46Published online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Whereas antiretroviral therapy of HIV infection has enormously increased the life expectancy of infected individuals, efforts to prevent HIV have lagged behind—despite the recognized importance of prophylaxis for this disease. Now Robert Grant et al.1 show how taking antiretroviral agents given daily in a single pill reduced the risk of HIV acquisition in 44% of 2,499 uninfected men who had sex with men, indicating that a preexposure prophylaxis (PrEP) regimen with antiretroviral drugs can protect against HIV infection in this group of people at risk. We asked the experts about the implications for the implementation of HIV prevention programs with antiretroviral drugs, the impact on public health and the next step in HIV therapeutics. Diane Havlir This past year, investigators showed how tenofovir-containing microbocide gel reduced the risk of HIV acquisition by 39% among HIV-uninfected women2 and that antiretroviral treatment of HIV-infected adults was associated with a 92% reduction in HIV transmission to their uninfected partners3. On the heels of these two studies, HIV PrEP with tenofovir and emtricitabine worked, reducing HIV acquisition by 44% in the first randomized efficacy study (iPrEx) conducted in HIV-uninfected men who have sex with men1. 2010 was undoubtedly a banner year for HIV prevention research. View full text Author information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Competing financial interests S.G.D. has received research support from Gilead Sciences. J.W.M. is a consultant to Gilead Sciences, Merck and RFS Pharma. He also holds share options in RFS Pharma, an early-stage biotech company that is developing new therapeutics for HIV and hepatitis C virus. Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Defying malaria: Fathoming severe Plasmodium vivax disease
- Nat Med 17(1):48-49 (2011)
Nature Medicine | Between Bedside and Bench Defying malaria: Fathoming severe Plasmodium vivax disease * Quique Bassat1 Contact Quique Bassat Search for this author in: * NPG journals * PubMed * Google Scholar * Pedro L Alonso1 Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:48–49Year published:(2011)DOI:doi:10.1038/nm0111-48Published online07 January 2011 The pathogen causing malaria, Plasmodium, is a perfect escapist that causes millions of infections and deaths—mostly in endemic areas plagued with poverty and lack of resources. Efforts in developing vaccines against the parasite focus on several immunological strategies, but they still fail to control it. In 'Bedside to Bench', Pedro Alonso and Quique Bassat examine recent observational studies where Plasmodium vivax was associated with severe malaria—usually linked to Plasmodium falciparum—in non-African endemic areas. Understanding what factors add to this morbidity and how this species severely sickens children and adults may help pave the way to eradicate malaria worldwide. In 'Bench to Bedside', Michael Good and Christian Engwerda discuss how a CD8+ T cell–mediated strategy may be useful in a vaccine to tackle the blood-stage parasite. Stimulation of these immune cells with the correct vaccination approach could open new doors to prevent disease in people infec! ted with malaria. View full text Figures at a glance * Figure 1: P. vivax can cause a wide spectrum of clinical symptoms, ranging from single-organ affectation to multiple life-threatening organ failure. * Figure 2: The transition from P. vivax asymptomatic infection to clinical disease depends on the interaction of various known or potential parasite, host and external factors. Parasite factors include antimalarial drug sensitivity, multiplicative potential and relapsing patterns, erythrocyte invasion capabilities, induction of endothelial adhesion and the expression of variant genes important for host immune evasion. Host factors include age and gender, immunity, pregnancy, ethnicity and genetic polymorphisms, coexisting infections or chronic conditions. Other external factors include access to health care and effective antimalarial treatment, vectorial capacity in the area and other sociocultural factors. Author information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Affiliations * Quique Bassat and Pedro Alonso are at the Barcelona Centre for International Health Research, Hospital Clínic-Universitat de Barcelona, Barcelona, Spain, and at the Centro de Investigação em Saúde de Manhiça, Maputo, Mozambique, Africa. * Pedro L Alonso Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Quique Bassat Additional data - Defying malaria: Arming T cells to halt malaria
- Nat Med 17(1):49-51 (2011)
Nature Medicine | Between Bedside and Bench Defying malaria: Arming T cells to halt malaria * Michael F Good1 Contact Michael F Good Search for this author in: * NPG journals * PubMed * Google Scholar * Christian Engwerda2 Contact Christian Engwerda Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Corresponding authorsJournal name:Nature MedicineVolume: 17,Pages:49–51Year published:(2011)DOI:doi:10.1038/nm0111-49Published online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Bench to Bedside Like most organisms that have jumped from animals to humans, the parasite responsible for the most serious cases of malaria infection, Plasmodium falciparum, is a cunning evader and suppressor of host immunity, which makes vaccine development especially difficult. A thorough understanding of all potential antiparasitic immune mechanisms is therefore required to design an effective vaccine. Until recently, the immune components able to kill malaria parasites resident within red blood cells were thought to be limited to antibodies, CD4+ T cells and γδ T cells (found mainly in peripheral tissues)1, 2, 3. In contrast, CD8+ T cells, a T cell subset known to kill tumor cells and virus-infected cells, were not thought to have a role, even though they were shown to kill the malaria parasite during its pre-erythrocytic stage within liver hepatocytes4, 5. View full text Author information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Affiliations * Michael F. Good is at the Glycomics Institute, Griffith University, Gold Coast, Australia * Christian Engwerda is at the Queensland Institute of Medical Research, Brisbane, Australia. Competing financial interests The authors declare no competing financial interests. Corresponding authors Correspondence to: * Michael F Good or * Christian Engwerda Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Research Highlights
- Nat Med 17(1):52-53 (2011)
Nature Medicine | Research Highlights Research Highlights Journal name:Nature MedicineVolume: 17,Pages:52–53Year published:(2011)DOI:doi:10.1038/nm0111-52Published online07 January 2011 Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Immunology: Taming IL-17 inflammation Interleukin-17 (IL-17) signaling is crucial in driving inflammatory autoimmune disease, and researchers are investigating ways of inhibiting this pathway in the clinic. But little is known about endogenous mechanisms aimed at dampening IL-17 signaling. Youcun Qian and colleagues (J. Exp. Med., 2647–2662, 2010) found that tumor necrosis factor receptor–associated factor-3 (TRAF3) is recruited to the IL-17 receptor upon binding of IL-17. This inhibits formation of a key signaling complex comprised of the adaptor Act1 and TRAF6, suppressing downstream signaling and cytokine production. In a mouse model of multiple sclerosis, overexpression of TRAF3 suppresses disease development and expression of inflammatory cytokines, whereas silencing of TRAF3 in the brain worsens disease. View full text Read the full article * Instant access to this article: US$32Buy now * Subscribe to Nature Medicine for full access: SubscribeLogin for existing subscribers Additional access options: * Use a document delivery service * Rent this article from DeepDyve * Login via Athens * Purchase a site license * Institutional access * British Library Document Supply Centre * Infotrieve * Thompson ISI Document Delivery * You can also request this document from your local library through inter-library loan services. Additional data - Receptor-mediated activation of ceramidase activity initiates the pleiotropic actions of adiponectin
- Nat Med 17(1):55-63 (2011)
Nature Medicine | Article Receptor-mediated activation of ceramidase activity initiates the pleiotropic actions of adiponectin * William L Holland1 Search for this author in: * NPG journals * PubMed * Google Scholar * Russell A Miller2 Search for this author in: * NPG journals * PubMed * Google Scholar * Zhao V Wang1 Search for this author in: * NPG journals * PubMed * Google Scholar * Kai Sun1 Search for this author in: * NPG journals * PubMed * Google Scholar * Brian M Barth3 Search for this author in: * NPG journals * PubMed * Google Scholar * Hai H Bui4 Search for this author in: * NPG journals * PubMed * Google Scholar * Kathryn E Davis1 Search for this author in: * NPG journals * PubMed * Google Scholar * Benjamin T Bikman5 Search for this author in: * NPG journals * PubMed * Google Scholar * Nils Halberg1, 6 Search for this author in: * NPG journals * PubMed * Google Scholar * Joseph M Rutkowski1 Search for this author in: * NPG journals * PubMed * Google Scholar * Mark R Wade4 Search for this author in: * NPG journals * PubMed * Google Scholar * Vincent M Tenorio1 Search for this author in: * NPG journals * PubMed * Google Scholar * Ming-Shang Kuo4 Search for this author in: * NPG journals * PubMed * Google Scholar * Joseph T Brozinick4 Search for this author in: * NPG journals * PubMed * Google Scholar * Bei B Zhang7 Search for this author in: * NPG journals * PubMed * Google Scholar * Morris J Birnbaum2 Search for this author in: * NPG journals * PubMed * Google Scholar * Scott A Summers3, 5, 8 Search for this author in: * NPG journals * PubMed * Google Scholar * Philipp E Scherer1, 9 Contact Philipp E Scherer Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:55–63Year published:(2011)DOI:doi:10.1038/nm.2277Received15 September 2010Accepted20 October 2010Published online26 December 2010 Abstract * Abstract * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg The adipocyte-derived secretory factor adiponectin promotes insulin sensitivity, decreases inflammation and promotes cell survival. No unifying mechanism has yet explained how adiponectin can exert such a variety of beneficial systemic effects. Here, we show that adiponectin potently stimulates a ceramidase activity associated with its two receptors, AdipoR1 and AdipoR2, and enhances ceramide catabolism and formation of its antiapoptotic metabolite—sphingosine-1-phosphate (S1P)—independently of AMP-dependent kinase (AMPK). Using models of inducible apoptosis in pancreatic beta cells and cardiomyocytes, we show that transgenic overproduction of adiponectin decreases caspase-8-mediated death, whereas genetic ablation of adiponectin enhances apoptosis in vivo through a sphingolipid-mediated pathway. Ceramidase activity is impaired in cells lacking both adiponectin receptor isoforms, leading to elevated ceramide levels and enhanced susceptibility to palmitate-induced cell de! ath. Combined, our observations suggest a unifying mechanism of action for the beneficial systemic effects exerted by adiponectin, with sphingolipid metabolism as its core upstream signaling component. View full text Figures at a glance * Figure 1: Adiponectin rapidly lowers hepatic ceramide content and improves glucose homeostasis. () Total ceramide levels from liver of leptin-deficient (ob/ob) mice after 60-min intravenous (i.v.) treatments with full-length adiponectin (Adn, 2 mg kg−1) or PBS (n = 6 per group). () Glucose infusion rates during hyperinsulinemic-euglycemic clamps performed on conscious unrestrained ob/ob mice before and after a bolus of Adn (2 mg kg−1, i.v.) or PBS (n = 5 per group). () Total ceramide levels from livers of mice with diet-induced obesity after 60-min treatments with full-length Adn (2 mg kg−1, i.v.) or PBS (n = 9 per group). (,) Insulin tolerance () and hepatic ceramide content () of adiponectin-deficient (Adipoq−/−), wild-type (+/+) or overexpressing (Tg/+) mice maintained on HFD (solid lines) or normal chow (dashed line) for 8 weeks (n = 7 per group). (–) Results from Lkb1fl/fl mice infected with adenovirus (Ad) encoding either GFP or Cre recombinase 16 d before experiments (n = 8 per group). () Triplicate western blots of liver proteins, probed for LKB1, T! hr172-phosphorylated AMPK (pAMPK), AMPK, pSer79-ACC and ACC. () Whole blood glucose monitored for 6 h after injection of PBS (solid lines) or Adn (34 mg kg−1, i.v., dashed line). () Quantification of total hepatic ceramide, dihydroceramide (dhCeramide), glucosylceramide and GM3 ganglioside. *P < 0.01 effect of adiponectin. †P < 0.05 compared to lean wild-type controls. Error bars, s.e.m. * Figure 2: Adiponectin promotes cardiomyocyte and HEART-ATTAC survival. () Survival Kaplan-Meier plot of female HEART-ATTAC transgenic mice crossed into indicated adiponectin backgrounds and challenged with AP20187 (0.010 μg kg−1, i.p.) (n = 12 per group). () Ceramide quantified from left ventricle or serum and normalized to the average content from adiponectin wild-type (WT) mice (63.9 pmol mg−1 in left ventricle, 9.5 pmol μl−1 in serum) (n = 12 per group). () Dihydro-S1P (dhS1P), S1P, dihydrosphingosine (dhSph) and sphingosine (Sph) quantified in left ventricle of WT (Adipoq+/+), adiponectin heterozygote (Adipoq−/+) and null (Adipoq−/−) mice (n = 6 per group). () Survival of male AdipoqTg/+ mice treated with PBS or male Adipoq+/+ HEART-ATTAC mice treated with S1P (1 mg kg−1, i.p.), FTY720 (FTY; 1 mg kg−1, i.p.), myriocin (Myr; 0.3 mg kg−1, i.p.) or PBS immediately before injection with AP20187 (0.05 mg kg−1, i.p.) (n = 10 per group). () Cell death, determined with or without AP20187-induced apoptosis in HEART-ATTAC transg! enic primary cardiomyocytes after treatment with BSA alone, C2-ceramide or palmitate. Cells were co-treated with PBS, adiponectin (Adn), S1P or myriocin (Myr) (n = 6 per group from three separate experiments). *P < 0.05 difference from wild-type control. †P < 0.01 effect of lipid treatment. Error bars, s.e.m. * Figure 3: Adiponectin targets the endocrine pancreas and maintains beta cell mass. () Adiponectin (Adn), insulin and nuclei (DAPI) visualized by immunofluorescence after injection of PBS (top) or Adn (bottom) into adiponectin null mice. Scale bar, 100 μm. (,) Random-fed blood glucose (n = 12 per group) () and total pancreatic insulin content (n = 6 per group) (), assessed in male adiponectin transgenic (AdipoqTg/+) versus wild-type (Adipoq+/+) mice 10 d after treatment with vehicle or AP20187 (0.2 mg kg−1, i.p., single injection). (,) Random-fed blood glucose (n = 12 per group) () and total pancreatic insulin content (), determined in female wild-type (Adipoq+/+) and adiponectin null (Adipoq−/−) PANIC-ATTAC mice 10 d after initiating treatment with AP20187 (0.2 mg kg−1, i.p., twice daily for 3 d) (n = 6 per group). () Islet size, calculated by mean cross-sectional area of multicelled islets from pancreata of male adiponectin-overexpressing (AdipoqTg/+), wild-type (Adipoq+/+) or adiponectin-lacking (Adipoq−/−) mice, 10 d after treatment with AP! 20187 or vehicle (n = 6 per condition). *P < 0.02 difference between adiponectin transgenic (or adiponectin null) and WT mouse of the same treatment. †P < 0.02 effect of AP20187 treatment. Error bars, s.e.m. * Figure 4: Adiponectin alters sensitivity to ceramide-induced apoptosis in INS-1 beta cells. () Cell viability, assessed in INS-1 cells challenged with BSA, palmitate (Pal) or C2-ceramide (C2) in the presence or absence of adiponectin (Adn) (n = 6 per group from three separate experiments). () Cell viability determined on INS-1 cells that were pretreated with sphingosine kinase inhibitor (SKI) or DMSO before delivery of adiponectin or PBS and challenged with BSA or palmitate. () Ceramidase activity, determined in lysates from cultured INS-1 cells under a range of pH conditions (n = 4 from separate experiments) in the presence or absence of Adn (in vitro). Baseline is BSA treatment without Adn. () Live cells imaged by staining with carboxyfluorescein diacetate (cFDA), and dead cells stained for annexin V, after treatment with C2-ceramide in the presence or absence of S1P. Representative of three separate experiments; scale bar, 50 μm. () Apoptosis of INS-1 cells determined by FACS analysis of annexin V and propidium iodide staining after treatment with BSA, palmitat! e or coadministered palmitate and S1P (representative of three independent experiments). *P < 0.01 effect of Adn. †P < 0.01 effect of proapoptotic insult. Error bars, s.e.m. * Figure 5: Adiponectin receptors 1 and 2 confer ceramidase activity in vivo. (,) Ceramidase activity after in vitro treatment with adiponectin (Adn) or PBS () and protein expression () assessed 48 h after transient transfection of HEK-293T cells with constructs encoding GFP alone or cotransfection with GFP and mouse AdipoR1, mouse AdipoR2, or indicated point mutants for conserved histidine residues in AdipoR1 (H141R or H191R) or AdipoR2 (H152R or H202R) (n = 5 from separate experiments). WT, wild type; R1, AdipoR1; R2, AdipoR2. (–) Results from hepatic overexpression of human AdipoR1, human AdipoR2 or GFP by adenovirus in vivo. () Hepatic ceramidase activity determined from fresh lysates 5 d after infection of 9-week-old WT mice C57/Bl6J mice (n = 5 per group). () Hepatic ceramides, measured after 6 h infusion of lard oil emulsions or fat-free glycerol control emulsions (n = 6 per group). (,) Insulin tolerance () and hepatic ceramide content (), determined 8 d after infection of high fat–fed mice or chow-fed controls (n = 6–8 per group). *P < 0! .05 effect of Adn or lipid administration. †P < 0.02 effect of genetic overexpression. Error bars, s.e.m. * Figure 6: Ablating adiponectin receptors 1 and 2 impairs ceramidase activity, S1P generation and cell survival. () Ceramidase activity from wild-type (WT) or AdipoR1 AdipoR2 DKO MEFs in the presence or absence of adiponectin (Adn) (n = 4). () S1P and dihydro-S1P quantified from WT or DKO MEFs after 12 h incubation in palmitate (750 μM) or BSA (n = 6). () Ceramide quantified from WT, AdipoR1 AdipoR2 (R1/R2) DKO or LKB1 knockout (KO) MEFs after 12-h incubations with palmitate (Pal) or BSA supplemented with Adn (5 μg ml−1) or PBS (n = 6 from three separate experiments). () Cell viability assessed in MEFs treated as in after 16 h of palmitate treatment (n = 5). () Total and phosphorylated AMPK, probed by western blot after pretreatment with D-e-MAPP or DMSO and treatment with full-length Adn, globular adiponectin (gAdn), S1P, AICAR or C2-ceramide (representative of four independent experiments). () Ceramide promotes apoptosis by aiding death receptor clustering, apoptosome formation and Bcl-2-associated X protein (Bax) translocation. Ceramide impairs activation of Akt, also known as p! rotein kinase B (PKB), via activation of protein kinase C (PKC)-ζ or protein phosphatase 2A (PP2A). Adiponectin promotes the deacylation of ceramide by activating adiponectin receptors. The resulting sphingosine and S1P increase intracellular calcium and activate AMPK through stimulation of CAMKK. These actions promote survival, nutrient uptake, nutrient utilization and mitochondrial proliferation. CAMKK, calmodulin-dependent protein kinase kinase; ER, endoplasmic reticulum; SM, sphingomyelin; SphK, sphingosine kinase; TNF; tumor necrosis factor-α. *P < 0.05 effect of Adn. †P < 0.05 compared to WT cells. Error bars, s.e.m. Author information * Abstract * Author information * Supplementary information Affiliations * Touchstone Diabetes Center, The University of Texas Southwestern Medical Center, Dallas, Texas, USA. * William L Holland, * Zhao V Wang, * Kai Sun, * Kathryn E Davis, * Nils Halberg, * Joseph M Rutkowski, * Vincent M Tenorio & * Philipp E Scherer * Institute for Diabetes, Obesity and Metabolism, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, USA. * Russell A Miller & * Morris J Birnbaum * Department of Biochemistry and Molecular Biology, Colorado State University, Fort Collins, Colorado, USA. * Brian M Barth & * Scott A Summers * Eli Lilly and Company, Lilly Corporate Center, Indianapolis, Indiana, USA. * Hai H Bui, * Mark R Wade, * Ming-Shang Kuo & * Joseph T Brozinick * Program in Cardiovascular and Metabolic Diseases, Duke-National University of Singapore Graduate Medical School, Singapore. * Benjamin T Bikman & * Scott A Summers * Department of Biomedical Sciences, Faculty of Health Science, University of Copenhagen, Copenhagen, Denmark. * Nils Halberg * Department of Metabolic Disorders, Merck Research Laboratories, Rahway, New Jersey, USA. * Bei B Zhang * Stedman Center for Nutrition and Metabolism Research, Duke University Medical Center, Durham, North Carolina, USA. * Scott A Summers * Department of Cell Biology, The University of Texas Southwestern Medical Center, Dallas, Texas, USA. * Philipp E Scherer Contributions W.L.H. conducted all experiments, except the portions indicated below, and contributed to the writing of the manuscript. R.A.M. conducted in vivo experiments with liver-specific Lkb1−/− mice. Z.V.W. generated all of the ATTAC mouse models used here. K.S. was responsible for the mutagenesis studies of AdipoR1 and AdipoR2. B.M.B. was involved in the studies with INS-1 cells. H.H.B., M.R.W., M.-S.K. and J.T.B. were involved in liquid chromatography–tandem mass spectrometry analysis for determination of sphingolipid content of the samples. K.E.D. assisted in the generation of the Adipor1−/−Adipor2−/− MEFs and high-fat feeding studies using adiponectin transgenic mice. B.T.B. helped in data analysis and RT-PCR of sphingolipid metabolism genes. N.H. performed the experiments with in vivo injections of adiponectin and detection of the protein in beta cells. J.M.R. was involved in designing experiments and protein production. V.M.T. performed ceramidase assays and geno! typing. B.B.Z., M.J.B., S.A.S. and P.E.S. were involved in experimental design, data analysis and in the writing of the manuscript. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Philipp E Scherer Supplementary information * Abstract * Author information * Supplementary information PDF files * Supplementary Text and Figures (2M) Supplementary Figures 1–6, Supplementary Table 1 and Supplementary Methods Additional data - MicroRNA-124 promotes microglia quiescence and suppresses EAE by deactivating macrophages via the C/EBP-α–PU.1 pathway
- Nat Med 17(1):64-70 (2011)
Nature Medicine | Article MicroRNA-124 promotes microglia quiescence and suppresses EAE by deactivating macrophages via the C/EBP-α–PU.1 pathway * Eugene D Ponomarev1 Search for this author in: * NPG journals * PubMed * Google Scholar * Tatyana Veremeyko1 Search for this author in: * NPG journals * PubMed * Google Scholar * Natasha Barteneva2 Search for this author in: * NPG journals * PubMed * Google Scholar * Anna M Krichevsky1, 3 Contact Anna M Krichevsky Search for this author in: * NPG journals * PubMed * Google Scholar * Howard L Weiner1, 3 Contact Howard L Weiner Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorsJournal name:Nature MedicineVolume: 17,Pages:64–70Year published:(2011)DOI:doi:10.1038/nm.2266Received11 March 2010Accepted22 October 2010Published online05 December 2010 Abstract * Abstract * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg MicroRNAs are a family of regulatory molecules involved in many physiological processes, including differentiation and activation of cells of the immune system. We found that brain-specific miR-124 is expressed in microglia but not in peripheral monocytes or macrophages. When overexpressed in macrophages, miR-124 directly inhibited the transcription factor CCAAT/enhancer-binding protein-α (C/EBP-α) and its downstream target PU.1, resulting in transformation of these cells from an activated phenotype into a quiescent CD45low, major histocompatibility complex (MHC) class IIlow phenotype resembling resting microglia. During experimental autoimmune encephalomyelitis (EAE), miR-124 was downregulated in activated microglia. Peripheral administration of miR-124 in EAE caused systemic deactivation of macrophages, reduced activation of myelin-specific T cells and marked suppression of disease. Conversely, knockdown of miR-124 in microglia and macrophages resulted in activation of t! hese cells in vitro and in vivo. These findings identify miR-124 both as a key regulator of microglia quiescence in the central nervous system and as a previously unknown modulator of monocyte and macrophage activation. View full text Figures at a glance * Figure 1: Analysis of expression of miR-124 in CNS-resident microglia compared to peripheral macrophages. (,) Real-time qRT-PCR analysis of miR-124 () and miR-223 () expression in F4/80+CD11b+ mononuclear cells isolated from different tissues of healthy adult C57BL/6 mice. Mean ± s.d. of triplicate wells is shown. The data are representative of three different experiments. BM, bone marrow; Perit., peritoneal cavity; TGM, thioglycolate-elicited inflammatory macrophages. (,) Analysis of expression of miR-124 in populations of microglia and peripheral macrophages isolated from the CNS of chimeric mice during EAE. () GFP fluorescence (x axis) and staining for CD11b (y axis) are shown for mononuclear cells isolated from the CNS of chimeric mice at the peak of EAE (day 21). In , miR-124 expression in populations of F4/80+CD11b+GFP− microglia (, left rectangle) and F4/80+CD11b+GFP+ peripheral macrophages (, right rectangle) sorted from CNS of healthy chimeric mice (no disease) or chimeras with EAE at the disease onset (day 14), peak (day 21) and recovery phase (day 40). One represen! tative of four experiments is shown. * Figure 2: Quantitative analysis of expression of miR-124 in activated microglia. () Analysis of expression of activation markers CD45 and MHC class II in populations of gated F4/80+CD11b+GFP− microglia (upper row) and F4/80+CD11b+GFP+ peripheral macrophages (lower row) isolated from CNS of healthy chimeric mice (no disease) or chimeric mice with EAE at the disease onset (day 14), peak (day 21) or recovery (day 40). Four to five mice per group were used. The isotype controls are shown in upper left quadrants of contour plots. Percentages of CD45+ cells (right quadrants) and MHC class II+ cells (upper quadrants) are shown. () Comparison of miR-124 expression in populations of resting CD45lowMHC class II−GFP− and of activated CD45int–hiMHC class II+GFP− microglia, as well as populations of activated CD45hiMHC class II+GFP+ and deactivated CD45int–hiMHC class II−GFP+ peripheral macrophages. The cells were sorted by FACS from the CNS of chimeric mice with EAE at day 14, and miR-124 expression was assessed by real-time qRT-PCR. Representative res! ults from two independent experiments are shown, with mean ± s.d. of quadruplicate wells plotted; **P < 0.01. () Analysis of expression of miR-124 in microglia activated in vitro by GM-CSF and IFN-γ or LPS and IFN-γ. Microglia cells were isolated from healthy mice and incubated in medium alone, with GM-CSF and IFN-γ, or with LPS and IFN-γ for 6 h, and miR-124 expression was assessed by real-time qRT-PCR. Mean ± s.d. of triplicate experiments is shown; *P < 0.05 compared to ex vivo–isolated microglia. * Figure 3: Analysis of expression of activation markers, pro- and anti-inflammatory cytokines and markers for M2 macrophages in BMDMs with ectopic overexpression of miR-124. (,) Flow cytometry analysis of the expression of CD45 (x axes) and either CD11b, F4/80, MHC class II or CD86 (y axes) in BMDMs transfected with miR-124 or control miRNA. Percentages of CD45hi cells (right quadrants) and activation marker–positive cells (upper quadrants) are shown. A representative experiment is shown in , and mean ± s.e.m. of four independent experiments is shown in . MFI, mean fluorescence intensity. *P < 0.05. () Flow cytometry analysis of TNF-α production by BMDMs transfected with miR-124 or control miRNA. () Real-time qRT-pPCR analysis of the mRNA expression of C/EBP-α, inducible form of nitric oxide synthase (iNOS), IL-4, IL-10, TGF-β1, arginase I and FIZZ1 in BMDMs transfected with miR-124 or control miRNA. () Analysis of TGF-β1 protein expression. The cells were stained for intracellular TGF-β1 or for isotype control and analyzed by FACS. Open histograms show TGF-β1 staining; filled histograms show staining for isotype control. Percentages of! TGF-β1+ cells are shown. * Figure 4: Validation of downstream target genes for miR-124. () Alignment of three predicted miR-124 binding sites to C/EBP-α 3′ UTR is shown for different species (Mus musculus, Homo sapiens, Pan troglodytes, Macaca mulatta, Rattus norvegicus and Oryctolagus cuniculus). () Western blot analysis of C/EBP-α expression in BMDMs transfected with miR-124 or control miRNA () Flow cytometry analysis of the expression levels of C/EBP-α and CD45, or PU.1 and CD45 in BMDMs transfected with miR-124 or control miRNA. Percentages of CD45hi C/EBP-α+ and CD45hiPU.1+ cells are shown in upper right quadrants. Populations of miR-124–transfected CD45low cells were negative for C/EBP-α and PU.1 expression, as shown in double staining for cell-surface CD45 and intracellular C/EBP-α or PU.1 (lower left quadrants). Staining for CD45 (x axes) and either C/EBP-α, PU.1 or corresponding isotype controls (y axes) are shown. () The mean ± s.e.m. of percentages of CD45hi C/EBP-α+ and CD45hiPU.1+ cells from four independent experiments. **P < 0.01. ()! Luciferase activity in NIE115 cells transfected with reporter constructs containing either intact or mutated C/EBP-α 3′ UTR. The NIE115 cell line was co-transfected with the indicated constructs and either miR-124 or control miRNA, and normalized levels of luciferase activity are shown. * Figure 5: Flow cytometry and histology analysis of extent of inflammation and demyelination in the CNS of mice treated with miR-124 or control miRNA. (,) EAE disease course (scored as described in Online Methods) in mice treated with miR-124. Mice with EAE were injected intravenously (i.v.) with miR-124 or control miRNA on days 7, 11, 15 and 18 () or days 13, 16, 18, 20 and 22 () after EAE induction, as indicated by arrows. The data represent average disease scores from three experiments with four or five mice per group. () Flow cytometry analysis of immune cell infiltrate in the CNS of mice with EAE treated with miR-124 or control miRNA. Mononuclear cells were isolated from the CNS on day 21 after induction of EAE, stained for CD11b and CD45 and analyzed by FACS. Percentages of resting CD11b+CD45low microglia (region R1), CD11b+CD45hi activated microglia and peripheral macrophages (R2) and CD11b−CD45hi lymphocytes (R3) are shown. () Quantification of absolute number of activated microglia and macrophages (CD11b+CD45hi), lymphocytes and CD4 T cells in the CNS of mice treated with either miR-124 or control miRNA. Mean ±! s.e.m. of three independent experiments is shown. () Histology analysis of extent of inflammation and demyelination in the spinal cords of mice with EAE treated with miR-124 or control miRNA. Spinal cords were harvested on day 21 after induction of EAE, and sections of spinal cord were stained for myelin or CD11b. Each panel shows a representative histopathology image (scale bar, 200 μm); three mice were analyzed. Myelin sheath is stained light blue and nucleated cells are stained dark blue (left images). The cells colored dark gray are positive for CD11b (right images). * Figure 6: Effect of miR-124 inhibitor on phenotype of macrophages cocultured with neural and astroglial cells. () Flow cytometry analysis of the expression of CD45 (x axis) and MHC class II (y axis) in populations of CD11b+GFP+ gated BMDMs cultured alone (left contour plot) or cocultured with an astroglial (middle contour plot) or neuronal (right contour plot) cell line. Percentages of CD45hiMHC class II+ cells are shown in upper right quadrants. BMDMs were isolated from ACTB-GFP transgenic mice that ubiquitously express GFP under the actin promoter. The cells were then analyzed for the expression of GFP, CD11b, MHC class II and CD45 using four-color flow cytometry. () qRT-PCR analysis of miR-124 expression in microglia, astroglial and neuronal lines, in BMDMs cultured alone or in CD11b+GFP+ BMDMs sorted from the cocultures. (–) BMDMs isolated from ACTB-GFP transgenic mice were cocultured with either an astroglial () or neuronal () cell line in the presence of anti–miR-124 or a control antagomir. (,) CD11b+GFP+ gated cells were analyzed for expression of CD45 (x axis) and MHC cla! ss II (y axis). Percentages of CD45hiMHC class II+ cells are shown in upper right quadrants. () Data from three independent experiments are summarized, which show mean ± s.e.m. of percentages of CD45hiMHC class II+ macrophages. *P < 0.05; **P < 0.01. Author information * Abstract * Author information * Supplementary information Primary authors * These authors contributed equally to this work. * Anna M Krichevsky & * Howard L Weiner Affiliations * Center for Neurologic Diseases, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA. * Eugene D Ponomarev, * Tatyana Veremeyko, * Anna M Krichevsky & * Howard L Weiner * The Immune Disease Institute, Harvard Medical School, Boston, Massachusetts, USA. * Natasha Barteneva Contributions E.D.P. performed all flow-cytometry assays, EAE experiments, experiments with chimeric, knockout and transgenic mice, in vivo injections of oligonucleotides, cell isolations, cell cultures, coculture assays and immunohistochemistry; collected and analyzed the data; and wrote the manuscript. T.V. performed in vitro transfections, miRNA and mRNA expression assays and data analysis, western blots, luciferase target validation assay, immunohistochemistry and in silico target prediction analysis and helped to write the manuscript. N.B. performed imaging cytometry. E.D.P. and A.M.K. conceived the project. E.D.P., T.V. and A.M.K. developed the hypothesis and designed the experiments. A.M.K. and H.L.W. discussed the hypothesis, helped with data interpretation, coordinated and directed the project and wrote the manuscript. Competing financial interests The authors declare no competing financial interests. Corresponding authors Correspondence to: * Howard L Weiner or * Anna M Krichevsky Supplementary information * Abstract * Author information * Supplementary information PDF files * Supplementary Text and Figures (3M) Supplementary Methods, Supplementary Data, Supplementary Discussion, Supplementary Figures 1–16 and Supplementary Tables 1–3 Additional data - miR-499 regulates mitochondrial dynamics by targeting calcineurin and dynamin-related protein-1
- Nat Med 17(1):71-78 (2011)
Nature Medicine | Article miR-499 regulates mitochondrial dynamics by targeting calcineurin and dynamin-related protein-1 * Jian-Xun Wang1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Jian-Qin Jiao1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Qian Li1 Search for this author in: * NPG journals * PubMed * Google Scholar * Bo Long1 Search for this author in: * NPG journals * PubMed * Google Scholar * Kun Wang1 Search for this author in: * NPG journals * PubMed * Google Scholar * Jin-Ping Liu1 Search for this author in: * NPG journals * PubMed * Google Scholar * Yan-Rui Li1 Search for this author in: * NPG journals * PubMed * Google Scholar * Pei-Feng Li1 Contact Pei-Feng Li Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:71–78Year published:(2011)DOI:doi:10.1038/nm.2282Received11 August 2010Accepted22 November 2010Published online26 December 2010 Abstract * Abstract * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Myocardial infarction is a leading cause of mortality worldwide. Here we report that modulation of microRNA-499 (miR-499) levels affects apoptosis and the severity of myocardial infarction and cardiac dysfunction induced by ischemia-reperfusion. We found that both the α- and β-isoforms of the calcineurin catalytic subunit are direct targets of miR-499 and that miR-499 inhibits cardiomyocyte apoptosis through its suppression of calcineurin-mediated dephosphorylation of dynamin-related protein-1 (Drp1), thereby decreasing Drp1 accumulation in mitochondria and Drp1-mediated activation of the mitochondrial fission program. We also found that p53 transcriptionally downregulates miR-499 expression. Our data reveal a role for miR-499 in regulating the mitochondrial fission machinery and we suggest that modulation of miR-499 levels may provide a therapeutic approach for treating myocardial infarction. View full text Figures at a glance * Figure 1: miR-499 prevents apoptosis and myocardial infarction. () miR-499 levels during myocardial ischemia in rats. n = 5 or 6 rats per time point; *P < 0.05 compared with 0 min or sham operation. () miR-499 levels in neonatal rat cardiomyocytes exposed to anoxia. *P < 0.05 compared with control. Data represent three separate experiments. (,) Cell death () and miR-499 levels () of cardiomyocytes infected with adenoviral miR-499 or its mutated form (miR-499-mut), and then exposed to anoxia. *P < 0.05. Data represent three separate experiments. () Representative images of ventricular myocardium sections from wild-type (WT) and miR-499 transgenic (Tg) mice exposed to sham operation or ischemia-reperfusion (I/R). Green, TUNEL-positive myocyte nuclei; blue, DAPI-stained nuclei; red, cardiomyocytes labeled with antibody to α-actinin; scale bar, 50 μm. Quantitative analysis of apoptosis is shown at right. n = 5 or 6 mice per group. () Infarct sizes (bottom) and representative images of midventricular myocardial slices (top) from Tg and WT m! ice exposed to I/R or sham operation. AAR, area at risk; LV, left ventricular area, INF, infarct area. The ratios of AAR to LV, INF to AAR and INF to LV are shown. n = 5 mice per group; *P < 0.05 compared with WT. Scale bar, 2 mm. (,) I/R-induced apoptosis analyzed by TUNEL () and myocardial infarction () in mice treated with miR-499 antagomir, control antagomir or buffer only (PBS). n = 5 or 6 mice per group. For , *P < 0.05 compared with antagomir control. For , *P < 0.05 compared with antagomir control plus I/R. Data are expressed as the mean ± s.e.m. * Figure 2: miR-499 transgenic mice are resistant to left ventricular remodeling after ischemia-reperfusion. (–) Heart/body weight ratio, cross-sectional areas and collagen areas of wild-type (WT) and miR-499 transgenic (Tg) mice subjected to ischemia-reperfusion (I/R) or sham operation. Representative photomicrographs show FITC-conjugated wheat germ agglutinin staining for cross-sectional analysis () and Masson trichrome staining for collagen (). n = 5 or 6 mice per group. *P < 0.05; scale bars, 20 μm. () Echocardiographic analysis of left ventricular dimensions and cardiac function in mice exposed to I/R. Pre-I/R, 1 d before I/R surgery; post-I/R, 2 weeks after surgery; LVIDs, systolic left ventricular internal diameters; LVIDd, diastolic left ventricular internal diameters; FS, fractional shortening of left ventricular diameter, calculated as [(LVIDd – LVIDs)/LVIDd] × 100. *P < 0.05; n = 5 or 6 mice per group. (–) Quantitative reverse-transcription PCR results showing mRNA levels of α-MHC, β-MHC, troponin I type 2 (Tnni2) and troponin T type 3 (Tnnt3) in hearts of WT o! r Tg mice exposed to sham operation or I/R. n = 5 or 6 mice per group; *P < 0.05 compared with WT subjected to I/R. Data are expressed as the mean ± s.e.m. * Figure 3: CnAα and CnAβ are targets of miR-499. () Putative miR-499 binding sites (potential complementary residues shown in red). () Calcineurin (Cn) activity (bottom) and immunoblots of CnAα and CnAβ proteins (top) in the hearts of rats during ischemia. *P < 0.05 compared with 0 min or sham operation; n = 5 rats per time point. Results are representative of three independent experiments. Numbers above immunoblots in – show the ratio of the band intensity of CnA to that of actin. (–) Immunoblots of CnAα and CnAβ proteins from cardiomyocytes exposed to anoxia (), transfected with miR-499 antagomir or the antagomir control (the number of hours after transfection is indicated) (), or infected with adenoviral miR-499 or its mutated form and then exposed to anoxia (). Results are representative of three independent experiments. () Luciferase activity measured from HEK293 cells transfected with luciferase constructs that report translation driven by the CnAα or CnAβ 3′ UTR, along with plasmids expressing miR-499 or! its mutated form. Cells transfected with pGL3 served as a control. Data represent three separate experiments. *P < 0.05 compared with transfection of CnAα 3′ UTR construct alone; #P < 0.05 compared with transfection of CnAβ 3′ UTR construct alone. () Luciferase activity measured from cardiomyocytes infected with adenoviral miR-499 or its mutated form, and transfected with a luciferase construct containing the CnAα 3′ UTR, the miR-499 antagomir and/or the negative-control antagomir (NC). Data represent three separate experiments. () Immunoblots of CnAα and CnAβ proteins from cardiomyocytes infected with adenoviral constructs expressing CnAα- or CnAβ-targeting siRNAs or scrambled forms of these siRNAs (sc) and treated or not with anoxia. Results are representative of three independent experiments. () The percentage of cells undergoing cell death after treatment or not with anoxia and after knockdown of CnAα or CnAβ with siRNA constructs in . Data represent thr! ee separate experiments, *P < 0.05 compared with anoxia alone.! Data are expressed as the mean ± s.e.m. * Figure 4: Drp1 participates in the regulation of mitochondrial fission and apoptosis in cardiomyocytes. () Immunoblot for Drp1 from cardiomyocytes 48 h after infection with adenoviral constructs encoding Drp1 siRNA or its scrambled form (Drp1-sc). () Mitochondrial fission and cell death in cardiomyocytes infected as in and then exposed to anoxia. *P < 0.05 compared with anoxia alone. Data represent three separate experiments. () Myocardial infarct sizes (as in Fig. 1f) in hearts of rats infected with adenoviral Drp1 siRNA or Drp1-sc and exposed to ischemia-reperfusion (I/R). n = 5 or 6 rats per group; *P < 0.05. () Immunoblots showing distribution of unphosphorylated Drp1 and phosphorylated Drp1 (p-Drp1) in mitochondria-enriched heavy membranes (HM) or cytosol of cardiomyocytes exposed to anoxia. Results are representative of three independent experiments. Data are expressed as the mean ± s.e.m. * Figure 5: miR-499 regulates mitochondrial fission through calcineurin and Drp1. () Left, mitochondrial morphology in cardiomyocytes infected with adenoviral miR-499 or its mutated form. Scale bar, 10 μm. Right, quantification of mitochondrial fission. Data represent three separate experiments. *P < 0.05 compared with control. () Immunoblots showing subcellular distribution of unphosphorylated Drp1 and phosphorylated Drp1 (p-Drp1) in mitochondria-enriched heavy membranes (HM) or cytosol of cardiomyocytes infected with adenoviral miR-499. Numbers indicate the ratio of the band intensity of Drp1 or p-Drp1 to that of Cox IV or actin. Results are representative of three independent experiments. () Left, representative EM images of mitochondria from miR-499 transgenic (Tg) mice or wild-type (WT) mice subjected to ischemia-reperfusion (I/R). Scale bar, 1 μm; arrows indicate fission mitochondria. Right, percentage of mitochondrial fission. n = 6 mice per group; *P < 0.05 compared with WT mice subjected to I/R. () Immunoblots showing Drp1 levels in mitochondri! a of Tg or WT mice subjected to I/R. Numbers indicate the ratio of Drp1 or p-Drp1 to Cox IV or actin. n = 5 mice per group. () Top, immunoblots showing CnAα in whole-cell lysates and Drp1 in HM of cardiomyocytes infected with adenoviral constructs expressing miR-499, along with adenoviral constructs encoding CnAα with a wild-type or mutated 3′ UTR (CnAα-WT-3′ UTR or CnAα-M-3′ UTR, respectively). Numbers indicate the ratio of the band intensity of CnA to that of actin, or the band intensity of Drp1 to that of Cox IV. Bottom, the percentages of cells with mitochondrial fission or undergoing cell death. Data represent three separate experiments. *P < 0.05. () Cardiomyocytes were infected with adenoviral constructs expressing miR-499 or miR-499-mut, and the percentage of cells with mitochondrial fission was determined. Data are expressed as the mean ± s.e.m. * Figure 6: p53 transcriptionally downregulates miR-499. () p53 binding sites in the sequence flanking rat miR-499 on the 5′ side. BS1, binding site 1; BS2, binding site 2. () ChIP analysis of p53 binding to the miR-499 promoter in cardiomyocytes treated or not with anoxia. An actin-specific antibody was used as a negative control in the immunoprecipitation step. () Luciferase activity measured from cardiomyocytes infected with adenovirus expressing p53 or β-galactosidase (β-gal) and transfected with empty vector (pGL4) or with constructs containing the wild-type (WT) miR-499 promoter or the miR-499 promoter mutated at either of the putative p53 binding sites (m-BS1 and m-BS2). Data represent three separate experiments. *P < 0.05 compared with WT construct alone; #P < 0.05 compared with m-BS2 construct alone. () miR-499 levels (bottom) and immunoblot showing p53 (top) in cardiomyocytes infected with adenoviral p53 or β-gal. Numbers above blot indicate ratio of p53 to actin. () miR-499 levels (bottom) and immunoblot for p53 (t! op) in cardiomyocytes infected with adenoviral p53 siRNA or its scrambled form (p53-sc) as indicated. Data represent three separate experiments. *P < 0.05 compared with anoxia alone. Numbers above the blot indicate the ratio of the band intensity of p53 to that of actin. () Cardiomyocytes were infected with adenoviral p53 or β-gal and the percentage of cells with mitochondrial fission or undergoing cell death was determined. The number of hours after infection is indicated. Data represent three separate experiments. *P < 0.05 compared with control. () Cardiomyocytes treated with anoxia or not were infected with adenoviral p53 siRNA or its control siRNA p53-sc and the percentage of cells with mitochondrial fission or undergoing cell death was determined. Data represent three separate experiments. *P < 0.05 compared with anoxia alone. () Immunoblots for CnA and showing the subcellular distribution of unphosphorylated Drp1 and phosphorylated Drp1 (p-Drp1) in cardiomyocytes tr! eated with anoxia or not and infected with adenoviral p53 siRN! A or p53-sc. Results are representative of three independent experiments. HM, heavy membranes. Numbers show the ratio of the band intensity of CnA to that of actin, or the band intensity of Drp1 or p-Drp1 to that of Cox IV or actin. () miR-499 levels and myocardial infarct sizes (as in Fig. 1f) in the hearts of rats subjected to ischemia-reperfusion (I/R) or not and infected with adenoviral p53 siRNA or p53-sc. n = 5 or 6 rats per group. Top graph, *P < 0.05 compared with ischemia-reperfusion alone. Bottom graph, *P < 0.05 compared with control (Con) subjected to I/R. Scale bar, 2 mm. () Schematic model of miR-499's role in regulating mitochondrial fission and cell survival. Data are expressed as the mean ± s.e.m. Author information * Abstract * Author information * Supplementary information Primary authors * These authors contributed equally to this work. * Jian-Xun Wang & * Jian-Qin Jiao Affiliations * Division of Cardiovascular Research, National Key Laboratory of Biomembrane and Membrane Biotechnology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China. * Jian-Xun Wang, * Jian-Qin Jiao, * Qian Li, * Bo Long, * Kun Wang, * Jin-Ping Liu, * Yan-Rui Li & * Pei-Feng Li Contributions P.-F.L. and J.-X.W. designed research. J.-X.W. performed cellular experiments. J.-Q.J. conducted animal experiments. Y.-R.L. created 3′ UTR constructs. Q.L. and B.L. constructed adenoviruses and generated miR-499 transgenic mice. J.-P.L. measured calcium. K.W. analyzed hypertrophy. P.-F.L. and J.-X.W. wrote the manuscript. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Pei-Feng Li Supplementary information * Abstract * Author information * Supplementary information PDF files * Supplementary Text and Figures (2M) Supplementary Figures 1–16 and Supplementary Methods Additional data - Detrimental effects of adenosine signaling in sickle cell disease
- Nat Med 17(1):79-86 (2011)
Nature Medicine | Article Detrimental effects of adenosine signaling in sickle cell disease * Yujin Zhang1 Search for this author in: * NPG journals * PubMed * Google Scholar * Yingbo Dai1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Jiaming Wen1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Weiru Zhang1, 3 Search for this author in: * NPG journals * PubMed * Google Scholar * Almut Grenz4 Search for this author in: * NPG journals * PubMed * Google Scholar * Hong Sun5 Search for this author in: * NPG journals * PubMed * Google Scholar * Lijian Tao3 Search for this author in: * NPG journals * PubMed * Google Scholar * Guangxiu Lu6 Search for this author in: * NPG journals * PubMed * Google Scholar * Danny C Alexander7 Search for this author in: * NPG journals * PubMed * Google Scholar * Michael V Milburn7 Search for this author in: * NPG journals * PubMed * Google Scholar * Louvenia Carter-Dawson8 Search for this author in: * NPG journals * PubMed * Google Scholar * Dorothy E Lewis9 Search for this author in: * NPG journals * PubMed * Google Scholar * Wenzheng Zhang9 Search for this author in: * NPG journals * PubMed * Google Scholar * Holger K Eltzschig4 Search for this author in: * NPG journals * PubMed * Google Scholar * Rodney E Kellems1 Search for this author in: * NPG journals * PubMed * Google Scholar * Michael R Blackburn1 Search for this author in: * NPG journals * PubMed * Google Scholar * Harinder S Juneja9 Search for this author in: * NPG journals * PubMed * Google Scholar * Yang Xia1, 6 Contact Yang Xia Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:79–86Year published:(2011)DOI:doi:10.1038/nm.2280Received11 May 2010Accepted19 November 2010Published online19 December 2010 Abstract * Abstract * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Hypoxia can act as an initial trigger to induce erythrocyte sickling and eventual end organ damage in sickle cell disease (SCD). Many factors and metabolites are altered in response to hypoxia and may contribute to the pathogenesis of the disease. Using metabolomic profiling, we found that the steady-state concentration of adenosine in the blood was elevated in a transgenic mouse model of SCD. Adenosine concentrations were similarly elevated in the blood of humans with SCD. Increased adenosine levels promoted sickling, hemolysis and damage to multiple tissues in SCD transgenic mice and promoted sickling of human erythrocytes. Using biochemical, genetic and pharmacological approaches, we showed that adenosine A2B receptor (A2BR)-mediated induction of 2,3-diphosphoglycerate, an erythrocyte-specific metabolite that decreases the oxygen binding affinity of hemoglobin, underlies the induction of erythrocyte sickling by excess adenosine both in cultured human red blood cells and i! n SCD transgenic mice. Thus, excessive adenosine signaling through the A2BR has a pathological role in SCD. These findings may provide new therapeutic possibilities for this disease. View full text Figures at a glance * Figure 1: Increased adenosine levels contribute to sickling and hemolysis in SCD transgenic mice. () Representative HPLC profile showing adenosine concentrations in the plasma of wild-type (WT) and SCD transgenic (Tg) mice at steady state. () The effect of chronic PEG-ADA treatment on adenosine concentrations in the plasma of wild-type and SCD transgenic mice. () Blood smears of SCD transgenic mice without or with PEG-ADA enzyme therapy. (–) Effects of PEG-ADA treatment on plasma hemoglobin (), plasma haptoglobin () and plasma total bilirubin () concentrations in wild-type and SCD transgenic mice. () Lifespan of RBCs in SCD transgenic mice treated with or without PEG-ADA. Mean ± s.e.m; n = 4–8 mice per group; *P < 0.05 versus wild type; **P < 0.05 versus untreated SCD transgenic mice. * Figure 2: In vivo effects of PEG-ADA treatment on multiple organ damage and renal dysfunction in SCD transgenic mice. () H&E staining of lung, liver and spleen of wild-type and SCD transgenic mice after 8 weeks with or without PEG-ADA treatment. Substantial vascular congestion, vascular damage and necrosis in the lungs, livers and spleens observed in SCD transgenic mice was reduced by PEG-ADA treatment. For lung and spleen, scale bar, 20 μm; for liver, scale bar, 10 μm. (–) Semiquantitative analysis of histological changes in lung, liver and spleen of wild-type and SCD transgenic mice using Image-Pro Plus software (Media Cybernetics). () The effect of PEG-ADA enzyme therapy on heme content in lungs, livers and spleens of wild-type and SCD transgenic mice. () The effect of PEG-ADA enzyme therapy on microinfarction and cysts in renal cortex and on congestion in renal medulla of wild-type and SCD transgenic mice, as analyzed by H&E staining. Scale bar, 10 μm. (,) Semiquantitative analysis of histological changes in renal cortex and renal medulla of wild-type and SCD transgenic mice using ! Image-Pro Plus. (,) The effect of PEG-ADA enzyme therapy on proteinuria () and urine osmolarity () in wild-type and SCD transgenic mice. Mean ± s.e.m.; n = 4–8 mice per group; *P < 0.05 versus wild type; **P < 0.05 versus untreated SCD transgenic mice. ND, not detected. * Figure 3: PEG-ADA treatment attenuates hypoxia-reoxygenation–induced acute sickle crisis in SCD transgenic mice. (–) The effects of hypoxia-reoxygenation, with or without PEG-ADA, on plasma adenosine concentration (), percentage of sickled cells (), plasma hemoglobin concentration () and plasma bilirubin concentration () in SCD transgenic mice. () Lung sections from SCD transgenic mice were stained with an antibody that recognizes Ly-6B.2 (a polymorphic 40-kDa antigen expressed by neutrophils) to visualize infiltrated tissue neutrophils (brown). Scale bar, 10 μm. () Quantification of neutrophil infiltration in the lungs of SCD transgenic mice by Image-Pro Plus software. () The effect of hypoxia-reoxygenation with and without PEG-ADA treatment on proinflammatory cytokine abundance (IFN-γ, IL-6, IL-1β and GM-CSF) in lung homogenates from SCD transgenic mice. Mean ± s.e.m. from 5–7 mice per group; *P < 0.05 relative to SCD transgenic mice under normoxic conditions and **P < 0.05 relative to untreated SCD transgenic mice under hypoxia-reoxygenation conditions. * Figure 4: Excess adenosine acts through A2BRs to induce 2,3-DPG and subsequent sickling in SCD transgenic mice. () Concentration of 2,3-DPG in RBCs of wild-type and SCD transgenic mice with or without chronic PEG-ADA enzyme therapy. () In vivo measurement of oxygen saturation of hemoglobin in wild-type and SCD transgenic mice with or without PEG-ADA treatment for 8 weeks. () 2,3-DPG concentrations in isolated RBCs treated with NECA in the presence or absence of theophylline. () 2,3-DPG concentrations in RBCs isolated from wild-type or mice deficient in each of the four types of adenosine receptors cultured in the presence or absence of NECA. () Immunostaining of A2BRs in RBCs from wild-type and A2BR-deficient mice. () 2,3-DPG concentrations in RBCs isolated from wild-type and adenosine receptor–deficient mice under hypoxic conditions. () cAMP concentrations in RBCs isolated from wild-type and A2BR-deficient mice treated with or without NECA. () 2,3-DPG concentrations in RBCs isolated from wild-type mice treated with NECA in the presence or absence of the PKA-specific inhibitor H-89.! () 2,3-DPG concentrations in RBCs of wild-type and SCD transgenic mice treated with or without PSB1115. () Lifespan of RBCs in SCD transgenic mice treated with or without PSB1115. Mean ± s.e.m.; *P < 0.05 relative to untreated controls, **P < 0.05 relative to treated or positive samples; Each experiment was repeated five to seven times. n = 5–8 mice per group. * Figure 5: Adenosine levels are elevated in individuals with SCD and A2BR-mediated elevation of 2,3-DPG concentrations is required for hypoxia-induced human erythrocyte sickling. () Average adenosine concentrations in plasma from individuals with SCD (SCD, n = 12) and healthy volunteers (control, n = 11). () 2,3-DPG concentration in RBCs isolated from controls (n = 11) and individuals with SCD (n = 12). In and , *P < 0.05. () Changes in 2,3-DPG concentrations in isolated SCD RBCs after incubation under hypoxic conditions in the absence or presence of PEG-ADA, MRS1754 (A2BR antagonist), H-89 (PKA-specific inhibitor) or glycolate (GA; promotes degradation of 2,3-DPG). *P < 0.05 versus normoxic condition; **P < 0.05 versus untreated samples under hypoxic condition; ***P < 0.05 versus NECA-treated samples under hypoxic condition. () Changes in the percentage of sickled cells in isolated SCD RBCs after incubation under varying oxygen concentrations in the absence or presence of PEG-ADA, MRS1754, H-89 or glycolic acid. *P < 0.05 versus untreated cells. In and , each experiment was repeated four to six times. Data in – are mean ± s.e.m. () Working model ! of excessive adenosine signaling in erythrocyte sickling and potential mechanism-based therapies in SCD. Under hypoxic conditions, increased adenosine-mediated 2,3-DPG production induced by activation of A2BRs decreases the O2 binding affinity of HbS, resulting in increased amounts of deoxy-HbS and increased sickling, hemolysis and multiple tissue damage and dysfunction. Without interference, hemolysis and multiple tissue injury lead to increased release of ATP, which is converted to adenosine by the combined action of the ectonucleotidases CD39 and CD73. The use of PEG-ADA to lower adenosine concentrations or an A2BR antagonist to block receptor activation would reduce the production of erythrocyte 2,3-DPG and reduce sickling. Author information * Abstract * Author information * Supplementary information Affiliations * Department of Biochemistry and Molecular Biology, The University of Texas Health Science Center at Houston, Houston, Texas, USA. * Yujin Zhang, * Yingbo Dai, * Jiaming Wen, * Weiru Zhang, * Rodney E Kellems, * Michael R Blackburn & * Yang Xia * Department of Urology, The Third Xiangya Hospital, Changsha, Hunan, China. * Yingbo Dai & * Jiaming Wen * Department of Nephrology, The First Xiangya Hospital, Changsha, Hunan, China. * Weiru Zhang & * Lijian Tao * Department of Anesthesiology, The University of Colorado, Denver, USA. * Almut Grenz & * Holger K Eltzschig * Department of Otorhinolaryngology, The Third Xiangya Hospital, Changsha, Hunan, China. * Hong Sun * Institute of Reproductive & Genetic Medicine, Central South University, Changsha, Hunan, China. * Guangxiu Lu & * Yang Xia * Metabolon, Inc., Durham, North Carolina, USA. * Danny C Alexander & * Michael V Milburn * Department of Ophthalmology and Visual Science, The University of Texas Health Science Center at Houston, Houston, Texas, USA. * Louvenia Carter-Dawson * Department of Internal Medicine, The University of Texas Health Science Center at Houston, Houston, Texas, USA. * Dorothy E Lewis, * Wenzheng Zhang & * Harinder S Juneja Contributions Y.Z. carried out the measurement of adenosine and 2,3-DPG in humans and mice, analysis of sickling, hemolysis and lifespan of mouse RBCs, histological analysis of multiple tissues and image quantification, ELISA analysis of inflammatory cytokines in the lung homogenates and immunostaining of lung tissues with neutrophil markers, human erythrocyte culture and analysis of sickling under hypoxic conditions, and immunostaining of A2BRs on human and mouse RBCs, and contributed to generation of figures. Y.D. conducted PEG-ADA purification, treatment of mice with PEG-ADA or PSB1115 and proteinuria measurement, and isolation of multiple organs and blood from mice. J.W. was involved in the purification of PEG-ADA and treatment of mice with PEG-ADA or PSB1115, and performed heme content measurement and histological analysis of kidneys. W.Z. was involved in the purification of PEG-ADA; treated mice with PEG-ADA or PSB1115 and contributed to immunostaining of lung tissues with neutrophi! l markers. A.G. treated normal human erythrocyte cultures with A2BR or A2AR agonists. D.C.A. and M.V.M. conducted metabolomic screens in blood of wild-type and SCD transgenic mice. L.C.-D. provided expertise in confocal analysis of A2BR expression on RBCs. D.E.L. provided expertise in flow cytometry to measure the lifespan of RBCs. W.Z. assisted with urine osmolality analysis. H.S., L.T. and G.L. provided expertise in hemolytic disorders and kidney dysfunction. H.K.E. assisted A.G. with experiments on the effects of A2AR and A2BR agonists on 2,3-DPG induction in normal RBCs. R.E.K. provided expertise in adenosine signaling and helped edit the manuscript; M.R.B. provided mice deficient in each of the four types of adenosine receptors; H.S.J. provided expertise in hemolytic disorders, procured human subjects' approval and maintained the database of de-identified human subject information. Y.X. was the principal investigator, oversaw the design of experiments and interpretatio! n of results, wrote and organized the manuscript, including th! e text and figures, and edited the manuscript. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Yang Xia Supplementary information * Abstract * Author information * Supplementary information PDF files * Supplementary Text and Figures (944K) Supplementary Methods, Supplementary Tables 1 and 2 and Supplementary Figures 1–5 Additional data - Histamine deficiency promotes inflammation-associated carcinogenesis through reduced myeloid maturation and accumulation of CD11b+Ly6G+ immature myeloid cells
- Nat Med 17(1):87-95 (2011)
Nature Medicine | Article Histamine deficiency promotes inflammation-associated carcinogenesis through reduced myeloid maturation and accumulation of CD11b+Ly6G+ immature myeloid cells * Xiang Dong Yang1 Search for this author in: * NPG journals * PubMed * Google Scholar * Walden Ai1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Samuel Asfaha1 Search for this author in: * NPG journals * PubMed * Google Scholar * Govind Bhagat3 Search for this author in: * NPG journals * PubMed * Google Scholar * Richard A Friedman4 Search for this author in: * NPG journals * PubMed * Google Scholar * Guangchun Jin1 Search for this author in: * NPG journals * PubMed * Google Scholar * Heuijoon Park1 Search for this author in: * NPG journals * PubMed * Google Scholar * Benjamin Shykind5 Search for this author in: * NPG journals * PubMed * Google Scholar * Thomas G Diacovo6 Search for this author in: * NPG journals * PubMed * Google Scholar * Andras Falus7 Search for this author in: * NPG journals * PubMed * Google Scholar * Timothy C Wang1 Contact Timothy C Wang Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:87–95Year published:(2011)DOI:doi:10.1038/nm.2278Received18 August 2010Accepted16 November 2010Published online19 December 2010 Abstract * Abstract * Accession codes * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Histidine decarboxylase (HDC), the unique enzyme responsible for histamine generation, is highly expressed in myeloid cells, but its function in these cells is poorly understood. Here we show that Hdc-knockout mice show a high rate of colon and skin carcinogenesis. Using Hdc-EGFP bacterial artificial chromosome (BAC) transgenic mice in which EGFP expression is controlled by the Hdc promoter, we show that Hdc is expressed primarily in CD11b+Ly6G+ immature myeloid cells (IMCs) that are recruited early on in chemical carcinogenesis. Transplant of Hdc-deficient bone marrow to wild-type recipients results in increased CD11b+Ly6G+ cell mobilization and reproduces the cancer susceptibility phenotype of Hdc-knockout mice. In addition, Hdc-deficient IMCs promote the growth of tumor allografts, whereas mouse CT26 colon cancer cells downregulate Hdc expression through promoter hypermethylation and inhibit myeloid cell maturation. Exogenous histamine induces the differentiation of IMCs ! and suppresses their ability to support the growth of tumor allografts. These data indicate key roles for Hdc and histamine in myeloid cell differentiation and CD11b+Ly6G+ IMCs in early cancer development. View full text Figures at a glance * Figure 1: Histamine-deficient Hdc−/− mice are highly susceptible to colorectal and skin carcinogenesis. () Macroscopic appearance of colorectal tumors from wild-type (WT) and Hdc−/− mice. Hdc−/− mice and wild-type mice (both male BALB/c background) were injected with a single dose of AOM followed by DSS in the drinking water for 10 d. () The number of large (diameter >3 mm) and small (diameter <3 mm) colorectal tumors in Hdc−/− mice versus wild-type mice (*P < 0.05; means ± s.d.; n = 8, each group). () Representative H&E-stained colonic tumor sections from wild-type and Hdc−/− mice. Scale bar, 50 μm. () Macroscopic appearance of the skin tumors from wild-type and Hdc−/− mice. Wild-type and Hdc−/− mice were treated with the tumor initiator DMBA and then treated twice weekly with the tumor promoter TPA. () The number of skin papillomas in Hdc−/− mice versus wild-type mice, including both large (diameter >2.5 mm) and small (diameter <2.5 mm) papillomas (*P < 0.05; means ± s.d.; n = 5, each group). () Representative H&E-stained skin tumor sections fr! om wild-type and Hdc−/− mice. Scale bar, 50 μm. * Figure 2: CD11b+Ly6G+ IMCs are the predominant source of Hdc-EGFP expression in the bone marrow. () Left, the percentage of EGFP+ cells within the bone marrow, spleen and peripheral blood of Hdc-EGFP mice. Right, the relative proportion of CD11b+ and Gr-1+ cells gated from the EGFP+ subset. FSC, forward scatter. Gr-1, granulocyte-differentiation antigen-1. () Expression of CD11b+Ly6G+ cells. CD11b+Gr-1+ myeloid cells were gated from the bone marrow of Hdc-EGFP mice. This FACS analysis indicates the proportion of EGFP+ cells that were CD11b+Ly6Ghigh, CD11b+Ly6Gmedium and CD11b+Ly6G− cells. This panel shows representative data from five experiments. Ly6G, lymphocyte antigen 6G. () FACS analysis of peritoneal exudative cells (PECs) of Hdc-EGFP mice with cell surface markers for myeloid cells and mast cells. c-kit, mouse CD117. FcεR, high affinity receptor. () Differentiation of splenic Hdc-EGFP+ IMCs. Immunofluorescence of DAPI-stained EGFP+CD11b+Ly6G+ IMCs treated with or without G-CSF for 48 h, showing suppression of EGFP expression with granulocyte differentiation (t! he images are representative of three independent experiments). * Figure 3: Hdc deficiency upregulates CD11b+Gr-1+ and CD11b+Ly6G+ IMCs. () FACS analysis of the percentage of CD11b+Gr-1+ IMCs in the bone marrow, spleen and peripheral blood in wild-type and Hdc−/− mice (*P < 0.05; means ± s.d.; n = 10, each group). () The relative proportion of CD11b+Ly6G+ cells in the peripheral blood and spleen of wild-type and Hdc−/− mice, with and without AOM plus DSS–induced colon tumors, as measured by FACS analysis (*P < 0.05; **P < 0.01; means ± s.d.; n = 5, each group). () FACS analysis showing the abundance of the Ly6G and Ly6C subsets in CD11b+ cells in the peripheral blood and spleen of wild-type and Hdc−/− mice. The panel is representative of data from five mice of each group. () Differentiation of IMCs. In vitro incubation of bone marrow–derived EGFP+ CD11b+Gr-1+ IMCs with GM-CSF and histamine followed by FACS analysis for Ly6G and Ly6C surface expression. The figure is representative of data from three independent experiments. () FACS analysis of the proportion of CD11b+Ly6C+ monocytes among CD! 11b+Gr-1+ IMCs sorted from the bone marrow of Hdc−/− mice and incubated with histamine; GM-CSF; histamine and GM-CSF; histamine, GM-CSF and H1R antagonist; histamine, GM-CSF and H2R antagonist; or histamine, GM-CSF, H1R and H2R antagonists (*P < 0.05; **P < 0.01; Student's t test. Data represent means ± s.d. from three independent experiments). () Effect of exogenous histamine on circulating CD11b+Gr-1+ IMCs in Hdc−/− mice. The percentage of CD11b+Gr-1+ IMCs by FACS in the peripheral blood of wild-type, Hdc−/−, and Hdc−/− mice treated with exogenous histamine (*P < 0.05; means ± s.d.; n = 5, each group). * Figure 4: EGFP+ IMCs are recruited to inflamed tissue and stroma of colon tumor. () Immunofluorescence and H&E sections from AOM plus DSS–treated (20 weeks) and control Hdc-EGFP mice (n = 5). Top, rare submucosal EGFP+ cells in the colon of control mice and a large number of EGFP+ cells in the colonic adenomas of AOM plus DSS–treated mice. H&E staining of colonic tumors confirmed abundant inflammatory cells in the stroma. Bottom, immunostaining of serial sections showing that EGFP+ cells are Hdc+, and approximately 50% of CD11b+ and Gr-1+ cells are EGFP+. () Left, immunofluorescence image showing EGFP+ cells in the DAPI-stained colon of Hdc-EGFP mice treated with DSS (10 d). Right, immunofluorescence image showing a greater number of EGFP+ cells in the DAPI-stained colon of Hdc-EGFP mice 8 weeks after AOM plus DSS treatment (compared to , before tumor development). () Immunofluorescence image of DAPI-stained colon from bone marrow–transplanted (BMT) mice 10 d after DSS, showing that infiltrating EGFP+ cells are bone marrow–derived (donor: Hdc-EGF! P mice; recipient: wild-type B6 mice). () Immunofluorescence stained human colonic inflammatory bowel disease (IBD) and colitis-associated carcinoma tissue sections showing HDC (green), CD11b (red) or tryptase (red). White arrows show HDC+ cells (which all colocalize with CD11b+ cells in IBD tissue); red arrows show cells that express CD11b or tryptase alone. The figure is representative of analyses of five IBD and five cancer samples. () Immunofluorescence and H&E-stained sections showing the presence of inflammatory cells and EGFP+ cells in the skin after a single TPA treatment. Top, H&E-stained skin sections. Bottom, immunofluorescence showing a large number of EGFP+ inflammatory cells in the dermis 24 h after TPA treatment, compared to 6 h after TPA treatment or to acetone-treated controls (n = 5). * Figure 5: Bone marrow derived IMCs from Hdc−/− mice accelerate tumor growth. () Colonic tumor numbers in bone marrow–transplanted mice at 20 weeks after AOM plus DSS treatment. Wild-type and Hdc−/− mice were lethally irradiated and reconstituted with wild-type or Hdc−/− bone marrow (*P < 0.05 compared to indicated control; means ± s.d. n = 6, each group). () Allograft tumor weight. CT26 colon cancer cells were implanted in NOD-SCID mice alone (control) or with CD11b+Ly6G+ bone marrow–derived IMCs from wild-type mice or Hdc−/− mice, with or without exogenous histamine (800 μg per kg body weight per day, intraperitoneal injection) (*P < 0.05; **P < 0.01; means ± s.d.; n = 7, each group). () Cytokine mRNA expression in bone marrow–derived IMCs. mRNA expression of IL-1α, IL-1β, IL-6, TGF-β and tumor necrosis factor-α (TNF-α) in CD11b+Ly6G+ IMCs from Hdc−/− mice compared to IMCs from wild-type mice were analyzed by qRT-PCR. () Effect of Il6 deficiency on promotion of tumor allograft growth by IMCs. CT26 cancer cells were impl! anted alone or concurrently with CD11b+Ly6G+ bone marrow IMCs isolated from wild-type or Il6−/− mice into NOD-SCID mice. Tumor weight was measured (*P < 0.05; means ± s.d.; n = 7). (,) The number of vessel branch points in allograft tumors derived from CT26 cancer cells injected with no CD11b+Ly6G+ IMCs (control) or IMCs from wild-type mice, Hdc−/− mice, Hdc−/− mice plus histamine or Il6−/− mice (*P < 0.05; means ± s.d.; n = 7, each group). * Figure 6: Migration of circulating CD11b+Gr-1+ IMCs is RAGE dependent, and suppression of Hdc occurs through a methylation-dependent mechanism. () Immunofluorescence staining of allograft tumor sections with or without concurrently implanted EGFP+ CD11b+Gr-1+ IMCs. Antibodies to proliferation marker Ki-67 and to myofibroblast α-SMA were applied. () Immunohistochemistry (IHC) staining show α-SMA+ myofibroblasts in the stroma of colonic adenomas from Hdc−/− mice treated with AOM and DSS compared to wild-type mice. The images in and show representative data from five tumors in each group. () qRT-PCR analysis of RAGE ligand S100A8 and monocyte chemoattractant protein-1 (MCP-1) mRNA expression in DSS-induced colitis tissue (*P < 0.05; means ± s.d.; n = 3, each group). () qRT-PCR analysis of RAGE ligands S100A8 and S100A9 mRNA expression in colonic tumors of Hdc−/− mice compared to wild-type mice treated with AOM and DSS (*P < 0.05; means ± s.d.; n = 3, each group). () FACS analysis of CD11b+Gr-1+ IMCs in the peripheral blood of Rage−/− mice after AOM plus DSS treatment compared to wild-type mice (*P < 0.0! 5; means ± s.d.; n = 5, each group). () Bone marrow–derived Hdc-EGFP–expressing CD11b+Ly6G+ IMCs were cultured with IEC-6 rat intestinal epithelial cells and CT26 cancer cells, respectively for 48 h; FACS analysis showing the percentage of EGFP−CD11b+ myeloid cells after co-culture with CT26 cancer cells compared to co-culture with IEC-6 cells and control IMCs (*P < 0.05; **P < 0.01; means ± s.d.; n = 3, each group). () Hdc promoter methylation analysis by sodium bisulfate DNA sequencing. Bone marrow–derived Hdc-EGFP+ CD11b+Ly6G+ IMCs were cultured with CT26 cancer cells for 48 h, and DNA of IMCs was extracted for detection of DNA CpG methylation sites. Top, bent arrow shows the transcription start site and boxes represent the GC box and exon 1. Short vertical lines across the horizontal line indicate CpG sites (−800 and +200 bp from the transcript start site). Bottom, filled and open circles represent methylated and unmethylated cytosine residues respectively. Accession codes * Abstract * Accession codes * Author information * Supplementary information Referenced accessions Gene Expression Omnibus * GSE23502 Author information * Abstract * Accession codes * Author information * Supplementary information Affiliations * Division of Digestive and Liver Diseases, Department of Medicine and Irving Cancer Center, Columbia University, New York, New York, USA. * Xiang Dong Yang, * Walden Ai, * Samuel Asfaha, * Guangchun Jin, * Heuijoon Park & * Timothy C Wang * Department of Pathology, Microbiology and Immunology, University of South Carolina School of Medicine, Columbia, South Carolina, USA. * Walden Ai * Department of Pathology and Cell Biology, Columbia University, New York, New York, USA. * Govind Bhagat * Department of Biomedical Informatics, Columbia University, New York, New York, USA. * Richard A Friedman * Department of Neuroscience, Columbia University, New York, New York, USA. * Benjamin Shykind * Department of Pediatrics, Columbia University, New York, New York, USA. * Thomas G Diacovo * Department of Genetics, Cell and Immunobiology, Semmelweis University, Budapest, Hungary. * Andras Falus Contributions X.D.Y. was involved in the study design, completion of experiments, data analysis and interpretation and manuscript preparation. W.A. constructed the Hdc-EGFP transgenic mouse and helped with examination of Hdc-EGFP expression. S.A. helped with the data interpretation and contributed to the manuscript preparation and revision. G.B. provided human colon specimen collection and did pathology assessments. R.A.F. carried out the microarray data analysis. G.J. helped with the colon cancer experiments and data analysis. H.P. helped with the skin carcinogenesis experiments and data analysis. B.S. performed histological analysis of the brains of Hdc-EGFP mice. T.G.D. carried out intravital microscopy studies. A.F. constructed the Hdc-knockout mice and provided helpful suggestions for our study. T.C.W. designed the study and contributed to the data analysis and writing of the manuscript. All authors discussed the results and commented on the manuscript. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Timothy C Wang Supplementary information * Abstract * Accession codes * Author information * Supplementary information Movies * Supplementary Video 1 (2M) Circulating EGFP-expressing blood cells in the ears of Hdc-EGFP mice with TPA-induced skin inflammation. An acute inflammatory response was induced in the ears of Hdc-EGFP mice by the application of a single dose of TPA (8.5 nmol, 20 μl per mouse). The video records 10 seconds of EGFP-expressing blood cells in the vessels using intravital microscopy 6 h after TPA treatment. PDF files * Supplementary Text and Figures (2M) Supplementary Figures 1–17, Supplementary Tables 1–4 and Supplementary Methods Additional data - Desmoglein 2 is a receptor for adenovirus serotypes 3, 7, 11 and 14
- Nat Med 17(1):96-104 (2011)
Nature Medicine | Article Desmoglein 2 is a receptor for adenovirus serotypes 3, 7, 11 and 14 * Hongjie Wang1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Zong-Yi Li1 Search for this author in: * NPG journals * PubMed * Google Scholar * Ying Liu1 Search for this author in: * NPG journals * PubMed * Google Scholar * Jonas Persson1 Search for this author in: * NPG journals * PubMed * Google Scholar * Ines Beyer1 Search for this author in: * NPG journals * PubMed * Google Scholar * Thomas Möller3 Search for this author in: * NPG journals * PubMed * Google Scholar * Dilara Koyuncu4 Search for this author in: * NPG journals * PubMed * Google Scholar * Max R Drescher1 Search for this author in: * NPG journals * PubMed * Google Scholar * Robert Strauss1 Search for this author in: * NPG journals * PubMed * Google Scholar * Xiao-Bing Zhang5 Search for this author in: * NPG journals * PubMed * Google Scholar * James K Wahl III6 Search for this author in: * NPG journals * PubMed * Google Scholar * Nicole Urban7 Search for this author in: * NPG journals * PubMed * Google Scholar * Charles Drescher7 Search for this author in: * NPG journals * PubMed * Google Scholar * Akseli Hemminki2 Search for this author in: * NPG journals * PubMed * Google Scholar * Pascal Fender8 Search for this author in: * NPG journals * PubMed * Google Scholar * André Lieber1 Contact André Lieber Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:96–104Year published:(2011)DOI:doi:10.1038/nm.2270Received28 September 2010Accepted08 November 2010Published online12 December 2010 Abstract * Abstract * Accession codes * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg We have identified desmoglein-2 (DSG-2) as the primary high-affinity receptor used by adenoviruses Ad3, Ad7, Ad11 and Ad14. These serotypes represent key human pathogens causing respiratory and urinary tract infections. In epithelial cells, adenovirus binding of DSG-2 triggers events reminiscent of epithelial-to-mesenchymal transition, leading to transient opening of intercellular junctions. This opening improves access to receptors, for example, CD46 and Her2/neu, that are trapped in intercellular junctions. In addition to complete virions, dodecahedral particles (PtDds), formed by excess amounts of viral capsid proteins, penton base and fiber during viral replication, can trigger DSG-2–mediated opening of intercellular junctions as shown by studies with recombinant Ad3 PtDds. Our findings shed light on adenovirus biology and pathogenesis and may have implications for cancer therapy. View full text Figures at a glance * Figure 1: Identification of receptor X using Ad3 virions and Ad3 PtDds. () Competition of 3H-labeled Ad3 and Ad5 virus attachment to HeLa cells after preincubation with Ad3 BsDds, PtDds or adenovirus fiber knobs. Attachment in PBS-treated cells was taken as 100%. n = 5. Data are represented as means ± s.e.m. Ad3-PtDds versus Ad3 knob: P = 0.0033. () Competition of Ad3-GFP and Ad35-GFP virus infection. HeLa cells were pretreated with Ad3 fiber knob or PtDds at increasing concentrations and then exposed to Ad3-GFP (left) or Ad35-GFP (right) virus at an MOI of 100 PFU per cell. GFP expression was measured 18 h later by flow cytometry. Data are represented as means. s.d. was less than 10% for all data points. () Attachment of 3H-labeled Ad3 and Ad35 viruses to human and nonhuman cell lines. Y79 and Ramos are human retinoblastoma and lymphoma cells, respectively. CHO cells are Chinese hamster ovary cells. MMC and TC1 cells are mouse mammary carcinoma and lung carcinoma cells, respectively. TC1-CD46 cells express human CD46. Shown are the average num! ber of viral particles (VP) attached per cell. n = 5. (,) Identification of receptor X by affinity capture and MS-MS. Membrane protein fractions were prepared from HeLa and Ramos cells. Protein blots were hybridized with Ad5/35++ virions () and Ad3 virions or Ad3 PtDd (). Binding was visualized with polyclonal antibodies against Ad35++ knob () or Ad3 knob () (see also Supplementary Fig. 1f,g). Solubilized HeLa cell membrane lysates were also immunoprecipitated with DSG-2–specific mAb 6D8 crosslinked with protein A/G plus agarose. Western blotting of immunoprecipitates was performed with DSG-2–specific monoclonal antibody AH12.2 (anti-DSG-2-IP). () MS-MS analysis of the 160-kDa band. Top, structure of DSG-2. EC, extracellular domain; EA, juxtamembrane extracellular anchor domain; TM, transmembrane domain. Bottom, amino acid sequence of DSG-2. Highlighted are the peptide sequences captured by MS-MS analysis of the 160-kDa band. The triangles in the DSG-2 schematic (top) i! ndicate the localization of the identified peptides with regar! d to the different domains. (–) Biacore plasmon surface resonance studies with recombinant human DSG-2 immobilized on sensor chips. Ad2, Ad3 and Ad5 at 5 × 109 viral particles per ml (), various concentrations of PtDds (1, 3, 10, 30 or 100 μg ml−1) or 100 μg ml−1 BsDs () or PtDds and Ad3 fiber knob () were injected over the activated surface, and response signals were collected over the indicated time periods. * Figure 2: Validation of DSG-2 as adenovirus receptor by loss-of-function studies. ) Competition of 3H-labeled adenovirus binding by recombinant DSG-2. 3H-Ad3, 3H-Ad7, 3H-Ad14, 3H-Ad14a, 3H-Ad11, 3H-Ad5 and 3H-Ad35 viruses were preincubated with 6 μg ml−1 recombinant human DSG-2 protein. Attachment of virus particles incubated with PBS was taken as 100%. For analysis of Ad11 attachment, cells were also incubated with 50 μg ml−1 of Ad35 fiber knob domain (Ad35K) before addition of Ad11 virus to block CD46. () Competition of adenovirus transduction by recombinant DSG-1, DSG-2 or desmocollin-2 (DSC-2) proteins. () Competition of 3H-Ad3 binding by DSG-2–specific antibodies. n = 5. PBS versus 6D8: P = 0.013; PBS versus 8E5: P = 0.0014. The specificity of mAbs to different DSG-2 domains is as follows (for a schematic of DSG-2, see Fig. 1f): 20G1 (propeptide region), 7H9 (propeptide region (Pro)/extracellular domain 1(EC1)), 13B11 (EC1/EC2), 10D2 (EC1/EC2), 8E5 (EC3) and 6D8 (EC3/EC4). (,) Effect of siRNA-mediated DSG-2 downregulation on adenovirus attach! ment () and transduction (). Shown are mean fluorescence intensity values. n = 5. () Cytolysis of Ad3-GFP–infected BT474 cells at day 7 after infection. siRNA-transfected cells were infected at adjusted MOIs that allow for comparable initial transduction rates, that is, MOI 1.0 PFU per cell and 0.5 PFU per cell for DSG-2 siRNA– and control siRNA–treated cells, respectively. Seven days later, viable cells were stained with crystal violet. () Cytolysis of Ad3-GFP–infected cells at day 7 after infection. siRNA-transfected small airway epithelial cells were infected at adjusted MOIs. Seven days later, cell viability was measured by WST-1 assay. Viability of PBS treated cells was taken as 100%. n = 3. Ad3-GFP or control siRNA versus Ad3-GFP DSG-2 siRNA: P < 0.001. * Figure 3: Validation of DSG-2 as adenovirus receptor by gain-of-function studies. () Transduction of human cell lines that express DSG-2 at various levels. Human erythromyeloblastoid leukemia K562 cells and Burkitt's B cell lymphoma BJAB and Ramos cells were infected with Ad3-GFP and Ad5/35-GFP at increasing MOIs, and GFP expression was measured 18 h later. n = 3. s.d. was less than 10% for all data points. () Ectopic DSG-2 expression. Human histiocytic lymphoma U937 cells were infected with a lentivirus vector carrying the DSG-2 cDNA under the control of the elongation factor 1α (EF1α) promoter. () Attachment of 3H-Ad3 and 3H-Ad35 to Raji, U937 and DSG-2-expressing U937 (U937–DSG-2) cells. () GFP expression after transduction of U937 and U937–DSG-2 cells with Ad3-GFP and Ad5/35-GFP. n = 3. * Figure 4: DSG-2 localization in human epithelial cells and interaction with Ad3. () Immunohistochemistry studies on human colon, foreskin and ovarian cancer paraffin sections with DSG-2–specific antibodies. Positive staining appears in brown. () Confocal microscopy immunofluorescence analysis of polarized human colon cancer T84 cells for DSG-2 (green) and the intercellular junction protein claudin-7 (red). Nuclei are blue. XY and XZ planes are shown. () Ad3 binding to DSG-2. T84 cells were incubated with Cy3-labeled Ad3 particles (red) for 15 min, washed and subjected to confocal microscopy. The top XZ image is a higher magnification. At least two (green) DSG-2 signals are associated with one (red) Cy3-Ad3 signal. () Confocal microscopy of normal human small airway epithelial cells (not grown in transwell chambers). Cells were incubated with Cy5-labeled PtDd for 15 min and then washed with PBS. The top XZ panel shows colocalization of DSG-2 (red) and E-cadherin (green). The bottom XZ panel is the same image showing purple Cy5-PtDd signals colocalized w! ith green E-cadherin signals. The XY panel shows purple (PtDd) and green E-cadherin channels. Thin arrows mark membrane-localized PtDds, thick arrows label cytoplasmic DSG-2. () Confocal microscopy immunofluorescence analysis of human cervical carcinoma HeLa cells. (The top XZ and XY images show staining of HeLa cells for DSG-2 (red) and claudin-7 (green). The bottom XZ image shows Cy3-Ad3 (red) and DSG2 (green) signals of HeLa cells incubated for 15 min with Cy3-Ad3). Scale bars, 20 μm. * Figure 5: EMT signaling induced by Ad3 virions and PtDds in epithelial cells. –) Phenotypic changes triggered by PtDds in breast cancer epithelial cells. 1 × 105 BT474 cells were incubated with 50 ng of PtDds or BsDds for the indicated time and subjected to staining with antibodies. The scale bar is 20 μm in all confocal images (,) and 40 μm in the standard immunofluorescence studies (,). () Graphic demonstration of array data for up- and downregulated genes (PtDd- versus PBS-treated cells). Each dot represents one gene. () Western blot analysis of ERK1/2 and PI3K phosphorylation analyzed 6 h after incubation of BT474 cells or BT474 cells transfected with siRNA with PtDds, BsDds, DSG-2-specific antibodies (10D2, 13B11 or 6D8) or control (Co) antibody (anti-GAPDH) at the indicated concentrations. For pathway inhibition, cells were treated overnight with the Erk1/2 inhibitor UO126 (5 μM) or the PI3K inhibitor wortmannin (2.5 μM) before PtDds were added (+ inhibitor). The efficacy of the drugs for inhibition of the specific pathway was validated i! n a previous study8. GAPDH is used to demonstrate equal loading. * Figure 6: Opening of intercellular junctions in epithelial breast cancer cells by interaction of Ad3 virions or PtDds with DSG-2. () FITC-dextran diffusion through monolayers of BT474 cells. BT474 cells cultured in a transwell chamber with 0.4 μm pore size were treated with 0.5 μg ml−1 BsDds or PtDds or 2 × 108 adenovirus particles per ml for 2 h, and then 4-kDa FITC-dextran was added to the apical (inner) compartment. Paracellular flux was assessed in aliquots from the apical and basal chambers. BsDds versus PtDds: P < 0.001. () Facilitation of 3H-Ad35 uptake by PtDds. Left, trapping of CD46 in intercellular junctions of T84 cells. Colocalization of CD46 and the intercellular junction protein claudin-7 results in yellow signals. Right, 3H-Ad35 attachment. BT474 cells were incubated with PtDds or BsDds and 3H-Ad35 for 2 h on ice, washed and then incubated at 37 °C for 60 min. Noninternalized adenovirus particles were removed by trypsin digestion, and cell-associated radioactivity was measured. () Mice carrying subcutaneous ovc316 tumors were injected intravenously with 50 μg PtDds or BsDds 8 h b! efore intravenous injection of 1 × 109 PFU of Ad5/35-βGal. Sections were stained with X-gal 72 h after injection. Scale bar, 40 μm. () Confocal microscopy for Her2/neu and claudin-7 in the Her2/neu-positive human breast cancer cell line BT474. () Confocal microscopy of BT474 cells 24 h after treatment with PBS or PtDds. () Viability of Her2/neu-positive breast cancer cells after treatment with PtDds, BsDds, UV-inactivated Ad3, UV-inactivated Ad5 and Herceptin. Viability of PBS-treated cells was taken as 100%. n = 5, *P < 0.05. () Effect of Ad3 and PtDd on Herceptin therapy in BT474 cells treated with inhibitors of ERK1/2 and PI3K pathways or DSG2-targeting siRNA. BT474 cells were transfected with control and DSG2 siRNA as described in Figure 2d and 48 h later treated with Ad3 or PtDds and Herceptin as described in . For inhibitor studies, BT474 cells were incubated with the indicated agents overnight. Cells were washed and treated with PtDd or UV-inactivated Ad3 and Herc! eptin as described above. n = 5, PBS versus wortmannin, UO126:! P < 0.05. () PtDd-mediated enhancement of Herceptin therapy in vivo. Shown is the tumor volume of individual mice at different days after BT474-M1 cell injection. The day of saline or Herceptin injection is indicated by an arrow. PtDds were injected 12 hours before Herceptin. Accession codes * Abstract * Accession codes * Author information * Supplementary information Referenced accessions Gene Expression Omnibus * GSE24138 Author information * Abstract * Accession codes * Author information * Supplementary information Affiliations * University of Washington, Division of Medical Genetics, Seattle, Washington, USA. * Hongjie Wang, * Zong-Yi Li, * Ying Liu, * Jonas Persson, * Ines Beyer, * Max R Drescher, * Robert Strauss & * André Lieber * Cancer Gene Therapy Group, University of Helsinki & Helsinki University Central Hospital, Helsinki, Finland. * Hongjie Wang & * Akseli Hemminki * University of Washington, Department of Neurology, Seattle, Washington, USA. * Thomas Möller * Bogazici University, Department of Molecular Biology and Genetics, Istanbul, Turkey. * Dilara Koyuncu * Loma Linda University, Department of Medicine, Division of Regenerative Medicine, Loma Linda, California, USA. * Xiao-Bing Zhang * University of Nebraska Medical Center, Omaha, Nebaska. * James K Wahl III * Fred Hutchinson Cancer Research Center, Seattle, Washington, USA. * Nicole Urban & * Charles Drescher * Unit of Virus Host Cell Interactions, Grenoble, France. * Pascal Fender Contributions H.W. conducted the studies for the identification and validation of DSG-2 as an adenovirus receptor; Z.-Y.L. performed the immunofluorescence studies; Y.L. performed the in vivo studies; J.P. contributed to adenovirus attachment assays; I.B. contributed to in vivo studies; T.M. helped with the expression array studies; D.K. and M.R.D. participated in in vitro studies with Herceptin; R.S. performed the western blot studies for kinase activation; X.-B.Z. produced the DSG-2–expressing lentivirus vector; J.K.W. III provided the DSG-2–specific antibodies; N.U. and C.D. provided tumor biopsies; A.H. helped with data interpretation; P.F. performed the Biacore studies and provided recombinant PtDds, and A.L. supervised the project and wrote the manuscript. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * André Lieber Supplementary information * Abstract * Accession codes * Author information * Supplementary information PDF files * Supplementary Text and Figures (1M) Supplementary Figures 1–7 and Supplementary Methods Additional data - The GD1a glycan is a cellular receptor for adenoviruses causing epidemic keratoconjunctivitis
- Nat Med 17(1):105-109 (2011)
Nature Medicine | Letter The GD1a glycan is a cellular receptor for adenoviruses causing epidemic keratoconjunctivitis * Emma C Nilsson1 Search for this author in: * NPG journals * PubMed * Google Scholar * Rickard J Storm1 Search for this author in: * NPG journals * PubMed * Google Scholar * Johannes Bauer2 Search for this author in: * NPG journals * PubMed * Google Scholar * Susanne M C Johansson1 Search for this author in: * NPG journals * PubMed * Google Scholar * Aivar Lookene3 Search for this author in: * NPG journals * PubMed * Google Scholar * Jonas Ångström4 Search for this author in: * NPG journals * PubMed * Google Scholar * Mattias Hedenström5 Search for this author in: * NPG journals * PubMed * Google Scholar * Therese L Eriksson1 Search for this author in: * NPG journals * PubMed * Google Scholar * Lars Frängsmyr1 Search for this author in: * NPG journals * PubMed * Google Scholar * Simon Rinaldi6 Search for this author in: * NPG journals * PubMed * Google Scholar * Hugh J Willison6 Search for this author in: * NPG journals * PubMed * Google Scholar * Fatima Pedrosa Domellöf7, 8 Search for this author in: * NPG journals * PubMed * Google Scholar * Thilo Stehle2, 9 Search for this author in: * NPG journals * PubMed * Google Scholar * Niklas Arnberg1, 10 Contact Niklas Arnberg Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:105–109Year published:(2011)DOI:doi:10.1038/nm.2267Received20 May 2010Accepted05 November 2010Published online12 December 2010 Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Adenovirus type 37 (Ad37) is a leading cause of epidemic keratoconjunctivitis (EKC)1, 2, a severe and highly contagious ocular disease. Whereas most other adenoviruses infect cells by engaging CD46 or the coxsackie and adenovirus receptor (CAR), Ad37 binds previously unknown sialic acid–containing cell surface molecules3, 4. By glycan array screening, we show here that the receptor-recognizing knob domain of the Ad37 fiber protein specifically binds a branched hexasaccharide that is present in the GD1a ganglioside and that features two terminal sialic acids. Soluble GD1a glycan and GD1a-binding antibodies efficiently prevented Ad37 virions from binding and infecting corneal cells. Unexpectedly, the receptor is constituted by one or more glycoproteins containing the GD1a glycan motif rather than the ganglioside itself, as shown by binding, infection and flow cytometry experiments. Molecular modeling, nuclear magnetic resonance and X-ray crystallography reveal that the two t! erminal sialic acids dock into two of three previously established sialic acid–binding sites in the trimeric Ad37 knob. Surface plasmon resonance analysis shows that the knob–GD1a glycan interaction has high affinity. Our findings therefore form a basis for the design and development of sialic acid–containing antiviral drugs for topical treatment of EKC. View full text Figures at a glance * Figure 1: Glycan array of Ad37 knob interactions with 260 different glycans. Elevated peaks indicate that the Ad37 knob bound with high efficiency to immobilized plasma glycoproteins (glycan nos. 1–6), sulfated glycans (nos. 26–40 and 42–47), monosialic acid–containing glycans (nos. 14–16), and one branched, disialic acid–containing glycan (no. 212) corresponding to the hexasaccharide that is found in ganglioside GD1a (see Supplementary Table 1 for details). RFU, relative fluorescent units. * Figure 2: Ad37 binding to the corneal epithelial cell surface depends on fiber knob binding to molecules containing the GD1a glycan. (,) Bars indicate the effect of glycans on Ad37 virion binding to () and infection of () HCE cells. (,) Bars show the effect of anti-glycan monoclonal antibodies on Ad37 virion binding to () and infection of () HCE cells. () Bars indicate the effect of glycans on Ad37 knob binding to HCE cells. Gal, galactose; SA, sialic acid. (–) Surface-staining of EM9 (), control sample without antibodies (), Ad37 knob (), and control without Ad37 () for all layers of human corneal epithelium. Scale bar, 20 μm. Controls were performed without glycans (, , ); without antibodies (, , ) and without Ad37 knob protein (, ). The control values were set to 100% on the y axes in –. Error bars in – show ± s.e.m. * Figure 3: X-ray crystallography and functional analysis of the Ad37 knob-GD1a glycan complex. () Overall structure of the Ad37 fiber knob in complex with GD1a. The three Ad37 chains A, B and C are shown as ribbon tracings and colored green, blue and gray. GD1a is bound on top of the Ad37 knob and shown in stick representation, with carbons in yellow, oxygens in red and nitrogens in blue. () Simulated annealing omit difference electron density map for GD1a, calculated at 1.65-Å resolution, contoured at 2.5 σ and shown with a radius of 4 Å around GD1a. Chains A, B and C are highlighted in green, blue and gray, respectively. One GD1a-molecule (yellow) is bound with its two terminal sialic acids (Neu5Ac residues) between chains A and B and between chains B and C. () Interactions between Ad37 fiber knob and Neu5AcB (yellow) of GD1a. Contacting residues are shown in stick representation. Residues involved in hydrogen bonds are highlighted blue (chain B) and gray (chain C), with oxygens in red and nitrogens in blue. Also shown: hydrogen bonds (broken lines), water molecu! les (red spheres) and residues only involved in van der Waals contacts (light gray). () Hydrogen bond network between Ad37 fiber knob and GD1a. Hydrogen bonds to the bridging sugar residues GalII and GlcI are shown as red broken lines. Previously introduced hydrogen bonds (shown in ) between Ad37 knob and Neu5AcB are indicated by black broken lines; water molecules are shown as red spheres. () Bars indicate flow cytometry analysis of knob binding to HCE cells treated with or without neuraminidase. WT, wild-type. Error bars show ± s.e.m. * Figure 4: Surface plasmon resonance analysis of GD1a glycan binding to immobilized Ad37 knob protein. Response values at equilibrium, ΔReq, are shown for each GD1a concentration. The curve was obtained by fitting the data to a two–binding site model. Author information * Author information * Supplementary information Affiliations * Department of Clinical Microbiology, Division of Virology, Umeå University, Umeå, Sweden. * Emma C Nilsson, * Rickard J Storm, * Susanne M C Johansson, * Therese L Eriksson, * Lars Frängsmyr & * Niklas Arnberg * Interfaculty Institute of Biochemistry, University of Tübingen, Tübingen, Germany. * Johannes Bauer & * Thilo Stehle * Department of Chemistry, Tallinn University of Technology, Tallinn, Estonia. * Aivar Lookene * Institute of Biomedicine, Department of Medical Biochemistry and Cell Biology, University of Göteborg, Göteborg, Sweden. * Jonas Ångström * Department of Chemistry, Computational Life Science Cluster (CLiC), Umeå University, Umeå, Sweden. * Mattias Hedenström * Institute of Infection, Immunity and Inflammation, College of Medical, Veterinary and Life Sciences, Glasgow Biomedical Research Centre, University of Glasgow, Glasgow, UK. * Simon Rinaldi & * Hugh J Willison * Department of Clinical Sciences, Ophthalmology, Umeå University, Umeå, Sweden. * Fatima Pedrosa Domellöf * Department of Integrative Medical Biology, Anatomy, Umeå University, Umeå, Sweden. * Fatima Pedrosa Domellöf * Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, Tennessee, USA. * Thilo Stehle * Laboratory for Molecular Infection Medicine in Sweden (MIMS), Umeå University, Umeå, Sweden. * Niklas Arnberg Contributions E.C.N. and R.J.S. contributed equally to design and conduction of binding, infection and flow cytometry experiments; S.M.C.J. produced GD1a glycan and performed NMR studies with M.H.; J.B. and T.S. carried out X-ray crystallography studies; S.R. and H.J.W. performed combinatorial glycolipid glycoarray; J.Å. conducted molecular modeling; F.P.D. carried out immunohistochemistry analysis; and A.L. performed SPR experiments. L.F. and T.L.E. conducted two-dimensional gel electrophoresis and blotting experiments, and L.F. did statistical calculations. T.S., M.H., F.P.D., A.L., J.Å., H.J.W., L.F., J.B. and N.A. discussed and wrote the manuscript, and T.S. and N.A. supervised the project. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Niklas Arnberg Supplementary information * Author information * Supplementary information Excel files * Supplementary Table 1 (120K) Summary of glycan array data PDF files * Supplementary Text and Figures (5M) Supplementary Figures 1–6 and Supplementary Tables 2–5 Additional data - Paraoxonase-1 is a major determinant of clopidogrel efficacy
- Nat Med 17(1):110-116 (2011)
Nature Medicine | Letter Paraoxonase-1 is a major determinant of clopidogrel efficacy * Heleen J Bouman1, 2, 3 Search for this author in: * NPG journals * PubMed * Google Scholar * Edgar Schömig4 Search for this author in: * NPG journals * PubMed * Google Scholar * Jochem W van Werkum1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Janna Velder5 Search for this author in: * NPG journals * PubMed * Google Scholar * Christian M Hackeng1, 6 Search for this author in: * NPG journals * PubMed * Google Scholar * Christoph Hirschhäuser5 Search for this author in: * NPG journals * PubMed * Google Scholar * Christopher Waldmann7 Search for this author in: * NPG journals * PubMed * Google Scholar * Hans-Günther Schmalz5 Search for this author in: * NPG journals * PubMed * Google Scholar * Jurriën M ten Berg1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Dirk Taubert4 Contact Dirk Taubert Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:110–116Year published:(2011)DOI:doi:10.1038/nm.2281Received14 July 2010Accepted22 November 2010Published online19 December 2010Corrected online21 December 2010 Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Clinical efficacy of the antiplatelet drug clopidogrel is hampered by its variable biotransformation into the active metabolite1, 2. The variability in the clinical response to clopidogrel treatment has been attributed to genetic factors, but the specific genes and mechanisms underlying clopidogrel bioactivation remain unclear. Using in vitro metabolomic profiling techniques, we identified paraoxonase-1 (PON1) as the crucial enzyme for clopidogrel bioactivation, with its common Q192R polymorphism determining the rate of active metabolite formation. We tested the clinical relevance of the PON1 Q192R genotype in a population of individuals with coronary artery disease who underwent stent implantation and received clopidogrel therapy. PON1 QQ192 homozygous individuals showed a considerably higher risk than RR192 homozygous individuals of stent thrombosis, lower PON1 plasma activity, lower plasma concentrations of active metabolite and lower platelet inhibition. Thus, we identif! ied PON1 as a key factor for the bioactivation and clinical activity of clopidogrel. These findings have therapeutic implications and may be exploited to prospectively assess the clinical efficacy of clopidogrel. View full text Figures at a glance * Figure 1: Kinetics of clopidogrel-metabolizing enzymes. (,) Substrate saturation kinetics for microsomal preparations of human cytochrome P450 isozymes () and esterases () involved in clopidogrel metabolism. Microsomes were obtained from stably transfected human embryonic kidney cells (HEK 293). Conversion of clopidogrel to 2-oxo-clopidogrel, of 2-oxo-clopidogrel to the active thiol metabolite and of clopidogrel to the thiol metabolite was assayed by incubation with increasing concentrations of substrate for 5 min at 37 °C. For esterases, the conversion of clopidogrel to clopidogrel-carboxylate, of 2-oxo-clopidogrel to 2-oxo-clopidogrel-carboxylate, and of the thiol metabolite to thiol metabolite-carboxylate was also assayed. Kinetics of the PON1 allozymes with the L55M and Q192R polymorphisms were determined. For each enzyme a specific probe reaction (positive control) was performed (black circles). Symbols and error bars represent means ± s.e.m. of three independent incubation experiments each. Supplementary Figure 1 shows si! milar kinetic measurements for enzymes not involved in clopidogrel metabolism. BChE, butyrylcholinesterase; CES1, carboxylesterase-1; CES2, carboxylesterase-2. DTNB, 5,5′-dithiobis-(2-nitrobenzoic acid). * Figure 2: Kaplan-Meier curves for individuals with coronary stent implantation and their pharmacokinetic and pharmacodynamic responses to clopidogrel. (,) Kaplan-Meier survival curves showing cumulative probabilities of case-cohort subjects without incident nonfatal definite stent thrombosis over 18 months of follow-up according to PON1 Q192R gene polymorphism () and according to tertiles (T1, T2 and T3) of paraoxonase plasma activity, with T1 < 104.9, T2 < 180.4–104.9 and T3 ≥ 180.4 nmol min−1 ml−1 (). Genotype distribution and tertile limits were extrapolated to the total cohort, and subcohort noncases were weighted with the inverse of the sampling fraction according to a previously described method33. Numbers of individuals at risk at the indicated time points are shown. P values for the total model coefficients were calculated by univariate Cox regression. () Box-and-whisker plots showing median response values with twenty-fifth and seventy-fifth percentiles (box) and tenth and ninetieth percentiles (whisker). After the 18-month follow-up, the clopidogrel-free cases with nonfatal stent thrombosis (ST, n = 41) a! nd the subcohort noncases without stent thrombosis (No ST, n = 71) were given a single 600-mg clopidogrel dose, and pharmacokinetic and platelet responses were compared by univariate Cox regression. Pharmacokinetic responses are indicated as maximum plasma concentrations (cmax, ng ml−1) of active thiol metabolite, 2-oxo-clopidogrel, parent clopidogrel and carboxylic acid metabolite and as ratio of the maximum plasma concentrations of 2-oxo-clopidogrel to thiol metabolite. Platelet response is indicated as percentage of maximal predose versus maximal 6-h postdose aggregation (Δ% induced by 20 μM ADP), Paraoxonase and arylesterase plasma activities are indicated as the velocity of transformation (nmol min−1 ml−1) of paraoxon to p-nitrophenol and of phenylacetate to phenol, respectively. P < 0.05 was considered a statistically significant difference. Change history * Change history * Author information * Supplementary informationErratum 21 December 2010 In the version of this article originally published online, the affiliations for Hans-Günther Schmalz and Dirk Taubert appeared incorrectly. Hans-Günther Schmalz is in the Department für Chemie, Universität zu Köln, Cologne, Germany, and Dirk Taubert is in the Department of Pharmacology, University Hospital of Cologne, Cologne, Germany. These errors have been corrected for the print, PDF and HTML versions of the article. Author information * Change history * Author information * Supplementary information Affiliations * Department of Cardiology, St. Antonius Hospital Nieuwegein, Nieuwegein, The Netherlands. * Heleen J Bouman, * Jochem W van Werkum, * Christian M Hackeng & * Jurriën M ten Berg * St. Antonius Center for Platelet Function Research, St. Antonius Hospital Nieuwegein, Nieuwegein, The Netherlands. * Heleen J Bouman, * Jochem W van Werkum & * Jurriën M ten Berg * Department of Biochemistry, Cardiovascular Research Institute Maastricht, University Maastricht, Maastricht, The Netherlands. * Heleen J Bouman * Department of Pharmacology, University Hospital of Cologne, Cologne, Germany. * Edgar Schömig & * Dirk Taubert * Department für Chemie, Universität zu Köln, Cologne, Germany. * Janna Velder, * Christoph Hirschhäuser & * Hans-Günther Schmalz * Department of Clinical Chemistry, St. Antonius Hospital Nieuwegein, Nieuwegein, The Netherlands. * Christian M Hackeng * Klinik und Poliklinik für Nuklearmedizin, Universität Münster, Münster, Germany. * Christopher Waldmann Contributions J.W.v.W. and D.T. conceived the project. E.S., J.M.t.B. and D.T. supervised the project. H.J.B., J.W.v.W., J.V., C.M.H., C.H., C.W., H.-G.S. and D.T. conducted or directed bioanalytics of clopidogrel and its metabolites, metabolomic profiling, blood collection and sample preparation, aggregometry, genotyping and PON1 phenotyping. H.J.B., E.S., J.W.v.W., C.M.H., J.M.t.B. and D.T. conducted or directed recruitment of subjects, disease assessment and follow-up assessments. H.J.B., J.W.v.W. and D.T. did the computational and statistical data analyses. H.J.B. and D.T. wrote the manuscript. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Dirk Taubert Supplementary information * Change history * Author information * Supplementary information PDF files * Supplementary Text and Figures (968K) Supplementary Tables 1–10, Supplementary Figures 1–9 and Supplementary Methods Additional data - Podocyte-secreted angiopoietin-like-4 mediates proteinuria in glucocorticoid-sensitive nephrotic syndrome
- Nat Med 17(1):117-122 (2011)
Nature Medicine | Letter Podocyte-secreted angiopoietin-like-4 mediates proteinuria in glucocorticoid-sensitive nephrotic syndrome * Lionel C Clement1 Search for this author in: * NPG journals * PubMed * Google Scholar * Carmen Avila-Casado2, 5 Search for this author in: * NPG journals * PubMed * Google Scholar * Camille Macé1, 5 Search for this author in: * NPG journals * PubMed * Google Scholar * Elizabeth Soria2 Search for this author in: * NPG journals * PubMed * Google Scholar * Winston W Bakker3 Search for this author in: * NPG journals * PubMed * Google Scholar * Sander Kersten4 Search for this author in: * NPG journals * PubMed * Google Scholar * Sumant S Chugh1 Contact Sumant S Chugh Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:117–122Year published:(2011)DOI:doi:10.1038/nm.2261Received11 August 2010Accepted18 October 2010Published online12 December 2010 Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg The main manifestations of nephrotic syndrome include proteinuria, hypoalbuminemia, edema, hyperlipidemia and lipiduria. Common causes of nephrotic syndrome are diabetic nephropathy, minimal change disease (MCD), focal and segmental glomerulosclerosis (FSGS) and membranous nephropathy. Among the primary glomerular diseases, MCD is usually sensitive to glucocorticoid treatment, whereas the other diseases show variable responses1. Despite the identification of key structural proteins in the glomerular capillary loop which may contribute to defects in ultrafiltration, many of the disease mechanisms of nephrotic syndrome remain unresolved. In this study, we show that the glomerular expression of angiopoietin-like-4 (Angptl4), a secreted glycoprotein, is glucocorticoid sensitive and is highly upregulated in the serum and in podocytes in experimental models of MCD and in the human disease. Podocyte-specific transgenic overexpression of Angptl4 (NPHS2-Angptl4) in rats induced nephr! otic-range, and selective, proteinuria (over 500-fold increase in albuminuria), loss of glomerular basement membrane (GBM) charge and foot process effacement, whereas transgenic expression specifically in the adipose tissue (aP2-Angptl4) resulted in increased circulating Angptl4, but no proteinuria. Angptl4−/− mice that were injected with lipopolysaccharide (LPS) or nephritogenic antisera developed markedly less proteinuria than did control mice. Angptl4 secreted from podocytes in some forms of nephrotic syndrome lacks normal sialylation. When we fed the sialic acid precursor N-acetyl-D-mannosamine (ManNAc) to NPHS2-Angptl4 transgenic rats it increased the sialylation of Angptl4 and decreased albuminuria by more than 40%. These results suggest that podocyte-secreted Angptl4 has a key role in nephrotic syndrome. View full text Figures at a glance * Figure 1: Angptl4 mRNA and protein expression in experimental glomerular disease. () Induction of proteinuria in rats 24 h after injection of γ2-NTS. () Upregulation of glomerular Angptl4 mRNA in rats injected with γ2-NTS. () Proteinuria in Angptl4−/− and Angptl4+/+ mice after injection of PBS or LPS. () Confocal expression of Angptl4 in rat glomeruli and colocalization with podocyte protein CD2AP. Controls to show the specificity of staining by the Angptl4-specific antibody involved the use of post-absorbed antibodies ('post abs.'); details are in the Online Methods. () Assessment of changes in glomerular Angptl4 expression in rat models of MCD (puromycin nephrosis, PAN), membranous nephropathy (passive Heymann nephritis, PHN), mesangial injury (anti-Thy1.1 nephritis) and severe focal and segmental glomerulosclerosis (non-HIV collapsing glomerulopathy, CG). Threshold for significance was threefold change. Red arrows, onset of proteinuria. D1, D3, D6 and D10 represent days after induction. Numbers above each bar are mean values. () Confocal assessme! nt of Angptl4 expression (red) in control and PAN day 6 glomeruli, and colocalization with GBM heparan sulfate proteoglycan (white; top and middle images) and podocyte protein nephrin (green, overlap yellow; bottom image). () Immunogold electron microscopy of PAN day 6 rat glomeruli to demonstrate Angptl4 expression (gold particles) in the podocytes (yellow arrows) and GBM (black arrows). Scale bars: 7.5 μm (), 10 μm () and 0.33 μm (). LCM, laser capture microdissection; EFP, effaced foot processes; endo, endothelium. * Figure 2: Characterization of male Angptl4 transgenic mice and rats. () Light microscopy (left) and confocal assessment (middle) of Angptl4 and confocal assessment of ZO-1 (right) expression in Angptl4 transgenic (TG) and wild-type (WT) mice. () Electron microscopy of 3-month-old transgenic mouse glomeruli, showing intact (FP) and effaced podocyte foot processes (EFP). () Immunogold electron microscopy for Angptl4 showing gold particles in podocytes and GBM (arrows) in Angptl4 transgenic mice. () Proteinuria in 3-month-old Angptl4 transgenic mice. () Rat Angptl4 transgenic constructs for the targeted expression of Angptl4 in podocytes (NPHS2-Angptl4, left) and adipose tissue (aP2-Angptl4, right) in rats. () Multi-organ mRNA expression profile of Angptl4 in podocyte-specific (left) and adipose tissue–specific (right) transgenic rats. Sk. muscle, skeletal muscle; BAT, brown adipose tissue; WAT, white adipose tissue. () Periodic acid Schiff–stained sections from 3-month-old wild-type and heterozygous transgenic rats. Arrows, prominent podocy! tes in NPHS2-Angptl4 transgenic rats. () Confocal expression of Angptl4 (red) in NPHS2-Angptl4 transgenic rat glomeruli and colocalization with podocyte protein nephrin (green, overlap yellow) and GBM heparan sulfate proteoglycan (blue, overlap fuchsia). () Electron microscopy of a glomerular capillary loop from a 5-month-old homozygous NPHS2-Angptl4 transgenic rat, showing diffuse foot process effacement (arrows). () Immunogold electron microscopy for Angptl4 in NPHS2-Angptl4 transgenic rats of increasing age (left to right), with transition from intact foot processes to foot process effacement (first noted around age 3 months, middle image), and clustering of gold particles in the GBM noted prominently in areas opposite to effaced foot processes (middle and right panels). Scale bars: 10 μm (), 1 μm (), 0.25 μm (), 10 μm (), 8 μm (), 1 μm (), 0.2 μm (). Endo, endothelium. **P < 0.01; ***P < 0.001. * Figure 3: Relationship between Angptl4 overexpression and proteinuria. () Albuminuria in female NPHS2-Angptl4 transgenic rats. () Albuminuria in male heterozygous NPHS2-Angptl4 transgenic rats. () Albuminuria in male homozygous NPHS2-Angptl4 transgenic rats. () GelCode blue–stained SDS PAGE of urinary protein from transgenic rats, rats with PAN and individuals with MCD and membranous nephropathy (MN). Arrow indicates prominent 70-kDa intact albumin band. Mean percentage densitometry of intact albumin is shown for each lane (see Supplementary Fig. 3d). MW, molecular weight; NA, not applicable. () Proteinuria after induction of low-dose PAN in heterozygous male NPHS2-Angptl4 transgenic and wild-type littermates. () Proteinuria in Wistar rats treated with glucocorticoids (PAN-S) or PBS (PAN) on alternate days starting 1 d after induction of PAN. () Glomerular Angptl4 mRNA expression in PAN rats described in . *P < 0.05; **P < 0.01; ***P < 0.001. * Figure 4: Relationship between Angptl4 sialylation and proteinuria. () Two-dimensional gel electrophoresis and western blot of protein from perfused glomeruli show neutral and high pI, low-order Angptl4 oligomers (pink and orange arrows) in control, PAN day 6 and glucocorticoid-treated PAN day 6 rats (from experiment in Fig. 3f). Reactivity of sialic acid binding lectin MAA to these oligomers is also assessed (exemplified for PAN, excerpts from independent blots). () Densitometry of total, neutral- and high-pI oligomers shown in . () Two-dimensional gel electrophoresis and western blot of concentrated supernatant from Angptl4-HEK293 stable cell line incubated with ManNAc or control, and analyzed for Angptl4 expression and binding with sialic acid–binding lectin MAA. Green arrow and line highlight high-pI protein in the control treated group; blue arrow in the ManNAc-treated group shows neutral-pI protein. () Same study as , except done with supernatant from the GEC stable cell line. () Two-dimensional gel electrophoresis and western blot s! tudies of glomerular protein from NPHS2-Angptl4 transgenic rats given tap water or tap water with ManNAc for 12 d. Blots were analyzed for neutral pI (enclosed in red ovals) and high pI (enclosed in green ovals) Angptl4 using anti-Angptl4 antibody and sialic acid–binding lectin from Sambucus nigra (SNA I). () Percentage of neutral- and high-pI Angptl4 in each group assessed by densitometry. () Albuminuria in NPHS2-Angptl4 transgenic rats given tap water (Control group) or tap water with ManNAc (Treatment group) for 12 d (Treatment group, ManNAc phase), followed by plain tap water for 24 d (Treatment group, Washout phase). Values are expressed as a percentage of the baseline albuminuria (designated as 100%). Individual tracings are shown in Supplementary Figure 8. In , significance is for difference from control values. In , significance is for difference from baseline values. Loading controls shown in Supplementary Figure 6. **P < 0.01; ***P < 0.001. Accession codes * Accession codes * Author information * Supplementary information Referenced accessions Entrez Nucleotide * AF487463.1 Author information * Accession codes * Author information * Supplementary information Primary authors * These authors contributed equally to this work. * Carmen Avila-Casado & * Camille Macé Affiliations * Glomerular Disease Therapeutics Laboratory, and Nephrology Research and Training Center, University of Alabama at Birmingham, Birmingham, Alabama, USA. * Lionel C Clement, * Camille Macé & * Sumant S Chugh * Department of Pathology, Instituto Nacional de Cardiologia, Mexico City, Mexico. * Carmen Avila-Casado & * Elizabeth Soria * Department of Pathology and Medical Biology, University Medical Center of Groningen, Groningen, The Netherlands. * Winston W Bakker * Division of Human Nutrition, Wageningen University, Wageningen, The Netherlands. * Sander Kersten Contributions L.C.C. was lead postdoctoral fellow, conducted most of the animal studies, generated stable cell lines and purified recombinant protein, maintained transgenic rats and conducted all studies on transgenic rats, confocal imaging and in situ hybridization, most gene expression studies and selected two-dimensional gel studies. C.A.-C. interpreted and analyzed light microscopy, electron microscopy and immunogold electron microscopy sections for the study and conducted studies on induction of collapsing glomerulopathy in rats. C.M. conducted most of the two-dimensional gel electrophoresis and proteomic work, and most of the albumin ELISA assays. E.S. acted as electron microscopist and morphometrics expert, prepared tissue for electron microscopy, conducted and imaged conventional and most immunogold electron microscopy studies, and conducted alcian blue charge studies. W.W.B. provided human sera and assisted in study design and preparation of the manuscript. S.K. conducted studies! on Angptl4 transgenic mice, provided tissue for histological and gene expression analysis, conducted Angptl4−/− mouse studies with NTS and made substantial contributions to the preparation of the manuscript. S.S.C. acted as senior investigator, planned and supervised the study, generated constructs for transgenic rats, conducted molecular biology and gene expression studies, conducted early animal studies, and wrote and revised the manuscript with input from the other authors. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Sumant S Chugh Supplementary information * Accession codes * Author information * Supplementary information PDF files * Supplementary Text and Figures (2M) Supplementary Figures 1–11, Supplementary Tables 1 and 2 and Supplementary Methods Additional data - Tumor-specific imaging through progression elevated gene-3 promoter-driven gene expression
- Nat Med 17(1):123-129 (2011)
Nature Medicine | Technical Report Tumor-specific imaging through progression elevated gene-3 promoter-driven gene expression * Hyo-eun C Bhang1 Search for this author in: * NPG journals * PubMed * Google Scholar * Kathleen L Gabrielson2 Search for this author in: * NPG journals * PubMed * Google Scholar * John Laterra3, 4 Search for this author in: * NPG journals * PubMed * Google Scholar * Paul B Fisher5 Search for this author in: * NPG journals * PubMed * Google Scholar * Martin G Pomper1, 6 Contact Martin G Pomper Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:123–129Year published:(2011)DOI:doi:10.1038/nm.2269Received31 December 2009Accepted11 August 2010Published online12 December 2010 Abstract * Abstract * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Molecular-genetic imaging is advancing from a valuable preclinical tool to a guide for patient management. The strategy involves pairing an imaging reporter gene with a complementary imaging agent in a system that can be used to measure gene expression or protein interaction or track gene-tagged cells in vivo. Tissue-specific promoters can be used to delineate gene expression in certain tissues, particularly when coupled with an appropriate amplification mechanism. Here we show that the progression elevated gene-3 (PEG-3) promoter, derived from a rodent gene mediating tumor progression and metastatic phenotypes, can be used to drive imaging reporters selectively to enable detection of micrometastatic disease in mouse models of human melanoma and breast cancer using bioluminescence and radionuclide-based molecular imaging techniques. Because of its strong promoter activity, tumor specificity and capacity for clinical translation, PEG-3 promoter–driven gene expression may re! present a practical, new system for facilitating cancer imaging and therapy. View full text Figures at a glance * Figure 1: Cancer-specific PEG-3 promoter activity shown by bioluminescence imaging in experimental metastasis models of human melanoma (Mel) and breast cancer (BCa). () Bioluminescence imaging of a representative healthy control mouse (Ctrl-2). () Bioluminescence imaging showing firefly luciferase expression observed in a representative melanoma model (Mel-3). Each mouse was imaged from four directions (V, ventral; L, left side; R, right side; D, dorsal views) to cover the entire body. Pseudocolor images from the two groups were adjusted to the same threshold. () Quantification of bioluminescence imaging signal intensity in the control group (Ctrl) and melanoma group at 24 and 48 h after injection of pPEG-Luc–PEI polyplex. Quantified values are shown in total flux. ***P < 0.0001. (,) CT scans and gross anatomical views of lung from one representative mouse from the control group () and the melanoma group (). (,) Bioluminescence imaging of one representative mouse from the control group (, Ctrl-3) and the experimental breast cancer metastasis group (, BCa-1). The pseudocolor images were adjusted to the same threshold. () Quantification ! of bioluminescent signal intensity in the Ctrl and breast cancer groups at 24 and 48 h after injection of pPEG-Luc–PEI polyplex. **P = 0.0066. () A CT image and a macroscopic view of lung from a representative breast cancer mouse. Displayed bioluminescent images (,,,) were obtained at 48 h after the systemic delivery of pPEG-Luc–PEI polyplex. Black arrows (,) indicate metastatic nodules observed in the lung. Scale bars, 5 mm. * Figure 2: Correlation between PEG-3 promoter–driven Luc expression and microscopic metastatic sites shown by histopathological analysis in a mouse model of melanoma metastasis. () Bioluminescence imaging of a representative mouse Mel-2 from Supplementary Figure 2b. These images were acquired at 48 h after the delivery of pPEG-Luc–PEI polyplex. (–) H&E and firefly luciferase staining of the formalin-fixed, paraffin-embedded tissues collected from Mel-2. According to the imaging results (, white solid or dotted rectangles and circles), organs tentatively associated with luciferase expression were collected for histological analysis. H&E staining confirms metastatic foci of melanoma cells in the lung (, white solid rectangle in ), in the adrenal glands (, white dotted rectangle in ), inside the thoracic cage adjacent to the sternum (, white solid circle in ) and in abdominal inguinal adipose tissues adjacent to the urinary bladder (, white dotted circle in ). Immunostaining of the consecutive sections with rabbit luciferase-specific antibody (anti-Luc) shows precise correlation between the localization of microscopic metastasis and Luc expression ! (brown staining). Melanoma-associated melanin pigmentation was also observed in these organs (black arrows in 'macroscopic view', –). Scale bars, 100 μm (H&E) and 2 mm (macroscopic views). * Figure 3: Histological confirmation of the microscopic metastatic sites detected by the PEG-3 promoter–driven bioluminescence imaging system in a human breast cancer metastasis model. () Bioluminescence imaging of a representative mouse, BCa-3 from Supplementary Figure 3b, 24 h after the systemic delivery of pPEG-Luc–PEI polyplex. The organs associated with the expression of luciferase from (black or white circles and rectangles), were collected for histological correlation. () H&E and luciferase staining on cryosections of the lung from BCa-3, which correlates with bioluminescent light output shown in the white rectangle in . Stained lung cryosections of a control mouse (Ctrl-3) from Supplementary Figure 3a are shown for comparison. (–) H&E and luciferase staining of the formalin-fixed, paraffin-embedded tissue sections collected from BCa-3. In the peripancreatic area (, black circle in ), inside the thoracic wall adjacent to the sternum (, white rectangle in ) and in the lymph node located in the abdominal inguinal adipose tissues adjacent to the bladder (, white circle in ), H&E staining confirmed the presence of metastatic lesions. Thin layers of ! breast cancer cells were found inside the rib cage (, black rectangle in ): between the first and third ribs (, top) and fourth and seventh ribs (, bottom). Luciferase staining on the consecutive sections shows the colocalization of the metastatic sites and luciferase expression. Another consecutive section of peripancreatic tissues was stained for human pan-cytokeratin (Anti–PAN CK) to ensure the precise colocalization of luciferase expression and MDA-MB-231 cells (). Scale bars, 100 μm. * Figure 4: Cancer-specific expression of HSV1-TK driven by PEG-3 promoter shown by SPECT-CT imaging in an experimental model of human melanoma metastasis. (,) CT, SPECT and co-registered [125I]FIAU SPECT-CT images of lungs of the healthy control group (, n = 3; Ctrl-1–Ctrl-3) and of the metastasis model of melanoma (, n = 5; Mel-1–Mel-5). Images were acquired at 48 h after i.v. injection of [125I]FIAU, which was 94 h after i.v. administration of pPEG-HSV1tk–PEI polyplex. () Quantification of lung SPECT images in and . Regions of interest of the same size and shape were drawn in the right lobes of the lung of each mouse. **P = 0.0070. Error bars represent means ± s.e.m. * Figure 5: Detection and localization of metastatic masses by SPECT-CT imaging after the systemic administration of pPEG-HSV1tk. (–) Systemic metastatic sites were detected based on the whole body SPECT-CT images in three representative mice, Mel-2 (,), Mel-3 (–) and BCa-1 (,). (,,,) Transverse, coronal and sagittal views of co-registered SPECT-CT images of Mel-2 () and Mel-3 (,,) from Figure 4b. All images were obtained at 24 h after [125I]FIAU injection. (,,,) Gross anatomical details of the metastatic masses that were located on the basis of the SPECT-CT images (,,,). Multiple metastatic sites were detected by imaging in Mel-2 (, dotted circle). Necropsy of the corresponding area revealed melanoma masses under the brown adipose tissue in the upper dorsal area (, dotted circle). () Accumulated radioactivity was detected adjacent to the thoracic mid-spine (arrow), which corresponded to a tumor at this location (, arrow). Additional metastatic sites demonstrated by SPECT-CT imaging (,, arrow and dotted circle) correlated with melanoma masses uncovered immediately above the diaphragm (, dotted circ! le) and in the left inguinal lymph node (, arrow), respectively. (,) Cross-comparison of PEG-3 promoter–mediated imaging and FDG-PET in a breast cancer metastasis model, BCa-1. Two nodules (Tu-1 and Tu-2) were detected by [125I]FIAU-SPECT near the heart () and were confirmed by necropsy (). Although Tu-1 was also detected by [18F]FDG-PET, Tu-2, a smaller nodule attached to the heart, was not obvious in the PET image. SPECT images were acquired 48 h after injection of [125I]FIAU. The PET and SPECT images were acquired on the same day (). (Tu, tumor; Ht, heart; Bf, brown fat.) Scale bars, 10 mm. Author information * Abstract * Author information * Supplementary information Affiliations * Department of Pharmacology and Molecular Sciences, Johns Hopkins Medical Institutions, Baltimore, Maryland, USA. * Hyo-eun C Bhang & * Martin G Pomper * Department of Molecular and Comparative Pathobiology, Johns Hopkins Medical Institutions, Baltimore, Maryland, USA. * Kathleen L Gabrielson * Department of Neurology, Johns Hopkins Medical Institutions, Baltimore, Maryland, USA. * John Laterra * Kennedy Krieger Institute, Baltimore, Maryland, USA. * John Laterra * Department of Human and Molecular Genetics, Virginia Commonwealth University Institute of Molecular Medicine, Virginia Commonwealth University Massey Cancer Center, Virginia Commonwealth University School of Medicine, Richmond, Virginia, USA. * Paul B Fisher * Russell H. Morgan Department of Radiology and Radiological Science, Johns Hopkins Medical Institutions, Baltimore, Maryland, USA. * Martin G Pomper Contributions H.-e.C.B. designed and performed the experiments, analyzed data and prepared the manuscript. K.L.G. provided technical support in histopathology. J.L., K.L.G. and P.B.F. gave conceptual advice and edited the manuscript. M.G.P. and P.B.F. conceived of the project. M.G.P. supervised the project and prepared the manuscript. Competing financial interests The authors declare no competing financial interests. Corresponding author Correspondence to: * Martin G Pomper Supplementary information * Abstract * Author information * Supplementary information Movies * Supplementary Video 1 (3M) Detection of metastatic melanoma after systemic administration of pPEG-HSV1tk/PEI polyplex using SPECT-CT. Movie of a representative melanoma metastasis model, Mel-2 from Figure 4b and Figure 5a,b. The image was acquired at 24 h post injection of [125I]FIAU (1.4 mCi), which was 70 h after the pPEG-HSV1tk delivery. Multiple metastatic sites are predicted in the lung and upper dorsal area of the animal by [125I]FIAU SPECT. Melanoma masses were confirmed under the brown adipose tissue in the corresponding area (Fig. 5b) as well as in its lung by the gross pathological analysis. * Supplementary Video 2 (3M) Movie of a representative control animal, Ctrl-3 from Figure 4a. This whole body SPECT-CT image was acquired at 24 h post injection of 1.4 mCi of [125I]FIAU, which was 70 h after the IV injection of pPEG-HSV1tk/PEI polyplex. Accumulated radioactivity was only detected in the urinary bladder and intestines of the animal. The same pseudo color scale used for Mel-2 in Supplementary Video 1 was applied. PDF files * Supplementary Text and Figures (2M) Supplementary Methods and Supplementary Figures 1–8 Additional data - Differentiation between glioma and radiation necrosis using molecular magnetic resonance imaging of endogenous proteins and peptides
- Nat Med 17(1):130-134 (2011)
Nature Medicine | Technical Report Differentiation between glioma and radiation necrosis using molecular magnetic resonance imaging of endogenous proteins and peptides * Jinyuan Zhou1, 2 Contact Jinyuan Zhou Search for this author in: * NPG journals * PubMed * Google Scholar * Erik Tryggestad3 Search for this author in: * NPG journals * PubMed * Google Scholar * Zhibo Wen1, 4 Search for this author in: * NPG journals * PubMed * Google Scholar * Bachchu Lal5, 6 Search for this author in: * NPG journals * PubMed * Google Scholar * Tingting Zhou1 Search for this author in: * NPG journals * PubMed * Google Scholar * Rachel Grossman7 Search for this author in: * NPG journals * PubMed * Google Scholar * Silun Wang1 Search for this author in: * NPG journals * PubMed * Google Scholar * Kun Yan1 Search for this author in: * NPG journals * PubMed * Google Scholar * De-Xue Fu8 Search for this author in: * NPG journals * PubMed * Google Scholar * Eric Ford3 Search for this author in: * NPG journals * PubMed * Google Scholar * Betty Tyler7 Search for this author in: * NPG journals * PubMed * Google Scholar * Jaishri Blakeley5 Search for this author in: * NPG journals * PubMed * Google Scholar * John Laterra5, 6 Search for this author in: * NPG journals * PubMed * Google Scholar * Peter C M van Zijl1, 2 Search for this author in: * NPG journals * PubMed * Google Scholar * Affiliations * Contributions * Corresponding authorJournal name:Nature MedicineVolume: 17,Pages:130–134Year published:(2011)DOI:doi:10.1038/nm.2268Received29 January 2010Accepted01 September 2010Published online19 December 2010 Abstract * Abstract * Author information * Supplementary information Article tools * Full text * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg It remains difficult to distinguish tumor recurrence from radiation necrosis after brain tumor therapy. Here we show that these lesions can be distinguished using the amide proton transfer (APT) magnetic resonance imaging (MRI) signals of endogenous cellular proteins and peptides as an imaging biomarker. When comparing two models of orthotopic glioma (SF188/V+ glioma and 9L gliosarcoma) with a model of radiation necrosis in rats, we could clearly differentiate viable glioma (hyperintense) from radiation necrosis (hypointense to isointense) by APT MRI. When we irradiated rats with U87MG gliomas, the APT signals in the irradiated tumors had decreased substantially by 3 d and 6 d after radiation. The amide protons that can be detected by APT provide a unique and noninvasive MRI biomarker for distinguishing viable malignancy from radiation necrosis and predicting tumor response to therapy. View full text Figures at a glance * Figure 1: MRI characteristics of radiation necrosis (40 Gy, 178 d after radiation; black solid arrow) in a rat. () T2-weighted images. () Gd-enhanced T1-weighted images. Both axial (top) and coronal (bottom) planes were acquired, and three consecutive slices are shown. Radiation necrosis is heterogeneous on the T2-weighted images and shows large enhancement on the Gd-enhanced T1-weighted images. Scale bars, 2 mm. * Figure 2: Comparison of radiation necrosis and glioma by conventional MRI. () Radiation necrosis (40 Gy, 178 d after radiation; black solid arrow). () SF188/V+ human glioma (16 d after implantation; pink open arrow). () 9L gliosarcoma (12 d after implantation; red open arrow). All pathologies are heterogeneous on the T2-weighted images and show large enhancement on the Gd-enhanced T1-weighted images. The shifted midlines of the brain indicate possible mass effect. These MRI features are very similar and not predictive of the final pathology. Scale bars, 2 mm. * Figure 3: Comparison of radiation necrosis and glioma using APT MRI and histology. () Magnetic resonance images and H&E-stained histopathological sections of radiation necrosis (40 Gy, 178 d after radiation). Radiation necrosis (black solid arrow) is revealed by Gd enhancement. Compared with contralateral brain tissue, APT MRI is hypointense to isointense in the lesion, where the loss of normal brain tissue components with vacuolation changes of necrosis (black arrowhead), as well as dilated damaged vessels (white arrowhead), are clearly visible on the high-magnification histology images. () Magnetic resonance images and H&E staining of SF188/V+ human glioma (16 d after implantation). () Magnetic resonance images and H&E staining of 9L gliosarcoma (11 d after implantation). Both SF188/V+ (pink open arrow) and 9L (red open arrow) tumors are hyperintense on the APT images, corresponding to high cellularity on histology. Scale bars, 2 mm (Gd-enhanced, APT and H&E 1.25×) and 50 μm (H&E 20×). * Figure 4: Quantitative comparison of APT image intensities (in percentage change of bulk water signal intensity) for radiation necrosis and glioma. Radiation necrosis (40 Gy, 163−188 d after radiation, n = 9); SF188/V+ human glioma (9−35 d after implantation, n = 9); 9L gliosarcoma (10−12 d after implantation, n = 9). **P < 0.01; ***P < 0.001. The tumors and radiation necrosis have opposite APT signal intensities with respect to the contralateral brain tissue (Contral.). Data are means ± s.d. * Figure 5: Changes in APT signal intensity (in percentage change of bulk water signal intensity) for the radiated U87MG tumors as a function of time after radiation (40 Gy; before radiation, 3 d after radiation and 6 d after radiation). () T2-weighted and APT images and H&E-stained brain sections for a typical rat. Before radiation, tumors are hyperintense with minimal heterogeneity on the APT images. APT signals show a clear decrease after therapy. At 6 d after radiation, the image intensity of the mass is very heterogeneous with an area that is almost isointense to contralateral normal tissue (pink arrow). At this time point, the presence of severe radiation necrosis was confirmed on histology. Low density tumor cells were also found mainly on the edge of the lesion (white arrowhead). Scale bars, 2 mm (T2-weighted, APT and H&E 1.25×) and 50 μm (H&E 20×). () Quantitative comparisons of APT image intensities for the irradiated tumors and the contralateral normal-appearing brain tissue. n = 5; *P < 0.05; ***P < 0.001. The average APT signal intensities in the lesion significantly decreased at 3 d and 6 d after radiation (P < 0.05 and P < 0.001, respectively). Data are means ± s.d. Author information * Abstract * Author information * Supplementary information Affiliations * Department of Radiology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA. * Jinyuan Zhou, * Zhibo Wen, * Tingting Zhou, * Silun Wang, * Kun Yan & * Peter C M van Zijl * F.M. Kirby Research Center for Functional Brain Imaging, Kennedy Krieger Institute, Baltimore, Maryland, USA. * Jinyuan Zhou & * Peter C M van Zijl * Department of Radiation Oncology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA. * Erik Tryggestad & * Eric Ford * Department of Radiology, Zhujiang Hospital, Southern Medical University, Guangzhou, Guangdong, China. * Zhibo Wen * Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA. * Bachchu Lal, * Jaishri Blakeley & * John Laterra * Department of Neurology, Kennedy Krieger Institute, Baltimore, Maryland, USA. * Bachchu Lal & * John Laterra * Department of Neurosurgery, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA. * Rachel Grossman & * Betty Tyler * Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA. * De-Xue Fu Contributions J.Z. developed the APT methodology, designed and performed most of the MRI experiments, carried out data analysis, prepared figures, wrote the manuscript and supervised the project. E.T. and E.F. designed and performed the radiation experiments and contributed to manuscript preparation. Z.W. contributed to MRI experimental work and histological analysis, contributed to manuscript preparation and provided helpful discussions on related clinical issues. B.L. performed experimental work in the tumor models and performed pathology experiments. T.Z. contributed to MRI experimental work, statistical analysis and manuscript preparation. R.G. and B.T. contributed to experimental work, especially in the tumor models, and manuscript preparation. S.W. contributed to MRI experimental work, performed histological analysis and statistical analysis, prepared some figures and contributed to manuscript preparation. K.Y. & D.-X.F. contributed to experimental work and data analysis. J.B. and J! .L. contributed to experimental design and manuscript preparation and provided helpful discussions on related clinical issues. P.C.M.v.Z. developed the APT methodology, contributed to experimental design and edited the manuscript. Competing financial interests J.Z. and P.C.M.v.Z. are co-inventors on a patent at the US Patent and Trademark Office for the APT technology. This patent is owned and managed by Johns Hopkins University. Corresponding author Correspondence to: * Jinyuan Zhou Supplementary information * Abstract * Author information * Supplementary information PDF files * Supplementary Text and Figures (344K) Supplementary Figures 1–3, Supplementary Tables 1 and 2, Supplementary Discussion and Supplementary Methods Additional data - Retraction: Genomic signatures to guide the use of chemotherapeutics
- Nat Med 17(1):135 (2011)
Nature Medicine | Retraction Retraction: Genomic signatures to guide the use of chemotherapeutics * Anil Potti Search for this author in: * NPG journals * PubMed * Google Scholar * Holly K Dressman Search for this author in: * NPG journals * PubMed * Google Scholar * Andrea Bild Search for this author in: * NPG journals * PubMed * Google Scholar * Richard F Riedel Search for this author in: * NPG journals * PubMed * Google Scholar * Gina Chan Search for this author in: * NPG journals * PubMed * Google Scholar * Robyn Sayer Search for this author in: * NPG journals * PubMed * Google Scholar * Janiel Cragun Search for this author in: * NPG journals * PubMed * Google Scholar * Hope Cottrill Search for this author in: * NPG journals * PubMed * Google Scholar * Michael J Kelley Search for this author in: * NPG journals * PubMed * Google Scholar * Rebecca Petersen Search for this author in: * NPG journals * PubMed * Google Scholar * David Harpole Search for this author in: * NPG journals * PubMed * Google Scholar * Jeffrey Marks Search for this author in: * NPG journals * PubMed * Google Scholar * Andrew Berchuck Search for this author in: * NPG journals * PubMed * Google Scholar * Geoffrey S Ginsburg Search for this author in: * NPG journals * PubMed * Google Scholar * Phillip Febbo Search for this author in: * NPG journals * PubMed * Google Scholar * Johnathan Lancaster Search for this author in: * NPG journals * PubMed * Google Scholar * Joseph R Nevins Search for this author in: * NPG journals * PubMed * Google ScholarJournal name:Nature MedicineVolume: 17,Page:135Year published:(2011)DOI:doi:10.1038/nm0111-135Published online07 January 2011 Article tools * Print * Email * Download PDF * Download citation * Order reprints * Rights and permissions * Share/bookmark * Connotea * CiteULike * Facebook * Twitter * Delicious * Digg Nat. Med.12, 1294–1300 (2006); published online 22 October 2006; corrected online 27 October 2006, 10 May 2007 and 10 October 2007 and corrected after print 21 July 2008; retracted 7 January 2011 We wish to retract this article because we have been unable to reproduce certain crucial experiments showing validation of signatures for predicting response to chemotherapies, including docetaxel and topotecan. Although we believe that the underlying approach to developing predictive signatures is valid, a corruption of several validation data sets precludes conclusions regarding these signatures. As these results are fundamental to the conclusions of the paper, we formally retract the paper. We deeply regret the impact of this action on the work of other investigators. Nature Medicine would also like to note that several of the earlier correction dates were either omitted or incorrect. The corrigenda published online 10 May 2007, 10 October 2007 and 21 July 2008 mistakenly omitted the earlier correction date of 27 October 2006. The correction in July 2008 went online on 21 July 2008 but was incorrectly noted in the corrigendum as having gone online 18 July 2008. Additional data
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