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Gray seals may be becoming the great white sharks of Dutch beaches

After 10 years of criminal scene investigation–style work, researchers have pinpointed the gray seal as the culprit behind mutilated, stranded harbor porpoises on Dutch beaches. After 10 years of criminal scene investigation–style work, researchers have pinpointed the gray seal as the culprit behind mutilated, stranded harbor porpoises on Dutch beaches. Gray seals may be becoming the great white sharks of Dutch beaches. The bodies kept washing ashore—dozens of mutilated harbor porpoises stranded on the Dutch beaches every year, their bloody remains discovered by screaming vacationers. Now, after 10 years of crime scene investigation–style work—complete with autopsies and DNA testing—biologists and veterinary pathologists have finally cracked the identity of the murderers: big-eyed, chubby-faced gray seals. The finding could lead conservationists to rethink gray seal rehabilitation, and it even raises the specter of a new “great white” in the North Sea. When harbor porpoises with missing bellies first appeared on Dutch coastlines in 2006, local biologists thought someone was deliberately hurting the animals. But the numbers soon rose to dozens a year—impossible to attribute to a single person. So the scientists looked elsewhere: Perhaps it was ducted propellers that sucked the porpoises in? Or fishermen cutting up unintentionally trapped porpoises? Then in 2012, a group of Belgian researchers noticed that some of the wounds on dead porpoises found on Belgium beaches bore the canine teeth marks of gray seals. “We thought, ‘Of course, how silly,’ ” says biologist Mardik Leopold of the Wageningen University and Research Centre in the Netherlands. “You think seals are nice, cuddly animals—they are not. They are predators.” With a towering height of 2.5 meters and the weight of two linebackers, gray seals are the largest predators in the southern North Sea. Though they are known as fish hunters, an adult male seal could easily overpower a 30-kilogram juvenile harbor porpoise. So the researchers examined photographs and autopsy results of more than 1000 stranded harbor porpoises collected from 2003 to 2013. The wounds further implicated gray seals. There was the torn blubber, the fatty nourishment that seals seek; the rows of canine teeth imprints on the tailstock, the thin part connecting the body and the tail; and the telltale scratch marks—four parallel lines left by seal claws grabbing onto the porpoises. Analysis indicates that close to a fifth of the stranded porpoises had a lethal encounter with gray seals—mostly naive juveniles who probably saw seals for the first time in their lives as they migrated north in the spring, Leopold says.

But finding the smoking gun proved to be a challenge. Short of analyzing the stomach contents of living seals, the only way to ascertain the identity of the predator was to find saliva DNA in the inflicted wounds: a seemingly impossible task, as seawater should quickly wash away any traces of DNA. Even human forensics rarely employs such analysis on drowned corpses, Leopold says. “Everyone thought we were crazy in even trying.” Indeed, nothing turned up from the obvious tear wounds. But at the bottom of deep, narrow bite marks on three porpoise bodies—where the flesh veered back after the seals pulled out their teeth and formed sealed pockets—the biologists found the iron-clad DNA evidence, they report online today in the Proceedings of the Royal Society B. That solves the “who” question, but it still doesn’t answer the “why”: What caused the gray seals to suddenly eye harbor porpoises for dinner in 2006? Humans may be to blame, the researchers suggest. As gas prices went up in recent years, Dutch fishermen switched from trolling to using cheaper set nets anchored to the seabed, which trapped harbor porpoises as by-catch. The team speculates that the gray seal, known to steal fish from the set nets, may have stumbled on a much larger, fattier “fish” and went on to actively hunt porpoises. The situation poses a dilemma for conservationists, as both the gray seals and the harbor porpoises are protected species. The Netherlands currently operates three rehabilitation centers for seals: The marine mammals disappeared from the region in the Middle Ages due to excessive hunting and only reemerged in the 1980s. But conservationists may need to reconsider the strategy, says biologist Jan Haelters of the Royal Belgian Institute of Natural Sciences in Ostend, who was not involved with the study. “In Africa, if you take care of all the lions and release them to the wild, it would affect the natural balance.” Still, he says, the study “gives us a very good framework for coordinated, coherent monitoring of harbor porpoises.” The seal predation could have long-term impacts on the local porpoise population, Leopold says. Researchers studying bottlenose dolphin attacks on porpoises in Scotland have found that the porpoises adapt by becoming leaner, faster swimmers. But speed comes at a cost. Compared with larger marine mammals such as whales, porpoises have more surface area relative to their body volume, which means they need to feed every hour to compensate for heat loss through the skin. Without food, a porpoise would die of starvation after 3 days, whereas a whale can fast for a month. “They are between a rock and a hard place really,” Leopold says. That seals would hunt down such large prey hints at a bigger problem, Leopold and Haelters warn. The Dutch shores are frequented by human bathers and surfers, raising the specter of a new “great white” terrorizing the North Sea beaches. “Statistically,” Haelters says, “an accident will happen.”

http://www.sciencemag.org/  Science Magazine

http://news.sciencemag.org/biology/2014/11/gray-seals-may-be-becoming-great-white-sharks-dutch-beaches?rss=1  Original web page at Science Magazine

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‘Safe’ levels of environmental pollution may have long-term health consequences

If you’re eating better and exercising regularly, but still aren’t seeing improvements in your health, there might be a reason: pollution. According to a new research report published in the September issue of The FASEB Journal, what you are eating and doing may not be the problem, but what’s in what you are eating could be the culprit. “This study adds evidences for rethinking the way of addressing risk assessment especially when considering that the human population is widely exposed to low levels of thousands of chemicals, and that the health impact of realistic mixtures of pollutants will have to be tested as well,” said Brigitte Le Magueresse-Battistoni, a researcher involved in the work from the French National Institute of Health and Medical Research (INSERM). “Indeed, one pollutant could have a different effect when in mixture with other pollutants. Thus, our study may have strong implications in terms of recommendations for food security. Our data also bring new light to the understanding of the impact of environmental food contaminants in the development of metabolic diseases.”

To make this discovery, scientists used two groups of obese mice. Both were fed a high-fat, high-sucrose enriched diet, with one group receiving a cocktail of pollutants added to its diet at a very low dosage. These pollutants were given to the mice throughout — from pre-conception to adulthood. Although the researchers did not observe toxicity or excess of weight gain in the group having received the cocktail of pollutants, they did see a deterioration of glucose tolerance in females, suggesting a defect in insulin signaling. Study results suggest that the mixture of pollutants reduced estrogen activity in the liver through enhancing an enzyme in charge of estrogen elimination. In contrast to females, glucose tolerance was not impacted in males exposed to the cocktail of pollutants. However, males did show some changes in liver related to cholesterol synthesis and transport. This study fuels the concept that pollutants may contribute to the current prevalence of chronic diseases including metabolic diseases and diabetes. “This report that confirms something we’ve known for a long time: pollution is bad for us,” said Gerald Weissmann, M.D., Editor-in-Chief of The FASEB Journal. “But, what’s equally important, it shows that evaluating food contaminants and pollutants on an individual basis may be too simplistic. We can see that when “safe” levels of contaminants and pollutants act together, they have significant impact on public health.”

Science Daily
September 17, 2013

Original web page at Science Daily

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Flexible tail of the prion protein poisons brain cells

Prion proteins are the infectious pathogens that cause Mad Cow disease and Creutzfeldt-Jakob disease. They occur when a normal prion protein becomes deformed and clumped. The naturally occurring prion protein is harmless and can be found in most organisms. In humans, it is found in our brain cell membrane. By contrast, the abnormally deformed prion protein is poisonous for the brain cells. Adriano Aguzzi, Professor of Neuropathology at the University of Zurich and University Hospital Zurich, has spent many years exploring why this deformation is poisonous. Aguzzi’s team has now discovered that the prion protein has a kind of “switch” that controls its toxicity. This switch covers a tiny area on the surface of the protein. If another molecule, for example an antibody, touches this switch, a lethal mechanism is triggered that can lead to very fast cell death. In the current edition of Nature, the scientists demonstrate that the prion protein molecule comprises two functionally distinct parts: a globular domain, which is tethered to the cell membrane, and a long and unstructured tail. Under normal conditions, this tail is very important in order to maintain the functioning of nerve cells. By contrast, in the case of a prion infection the pathogenic prion protein interacts with the globular part and the tail causes cell death — this is the hypothesis put forward by the researchers.

Aguzzi and his team tested this by generating mimetic antibodies in tissue sections from the cerebellum of mice which have a similar toxicity to that of a prion infection. The researchers found that these antibodies tripped the switch of the prion protein. “Prion proteins with a trimmed version of the flexible tail can, however, no longer damage the brain cells, even if their switch has been recognized by antibodies,” explains Adriano Aguzzi. “This flexible tail is responsible for causing cell death.” If the tail is bound and made inaccessible using a further antibody, activation of the switch can likewise no longer trigger cell death. “Our discovery has far-reaching consequences for understanding prion diseases,” says Aguzzi. The findings reveal that only those antibodies that target the prion protein tail are suitable for use as potential drugs. By contrast, antibodies that trip the switch of the prion are very harmful and dangerous.

Science Daily
August 20, 2013

Original web page at Science Daily

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New model for staph bone infections outlined

Osteomyelitis, a debilitating bone infection most frequently caused by Staphylococcus aureus (“staph”) bacteria, is particularly challenging to treat. Now, Vanderbilt microbiologist Eric Skaar, Ph.D., MPH, and colleagues have identified a staph-killing compound that may be an effective treatment for osteomyelitis, and they have developed a new mouse model that will be useful for testing this compound and for generating additional therapeutic strategies. James Cassat, M.D., Ph.D., a fellow in Pediatric Infectious Diseases who is interested in improving treatments for children with bone infections, led the mouse model studies. Working with colleagues in the Vanderbilt Center for Bone Biology and the Vanderbilt University Institute of Imaging Science, Cassat developed micro-computed tomography (micro-CT) imaging technologies to visualize a surgically introduced bone infection in progress. “The micro-CT gives excellent resolution images of the damage that’s being done to the bone,” said Skaar, the Ernest W. Goodpasture Professor of Pathology. “We found that staph is not only destroying bone, but it’s also promoting new bone growth. Staph is causing profound changes in bone remodeling.” Cassat also established methods for recovering — and counting — bacteria from the infected bone.

“We’re not aware of any other bone infection models where you can pull the bacteria out of a bone and count them in a highly reproducible manner,” Skaar said. “From a therapeutic development standpoint, we think this model is going to allow investigators to test new compounds for efficacy against bone infections caused by staph or any other bacteria that cause osteomyelitis.” Several pharmaceutical companies have already approached Skaar and his team about testing compounds in the new bone infection model, which the investigators describe in the June 12 issue of Cell Host & Microbe. Using the model, the team demonstrated that a certain protein secreted by staph plays a critical role in the pathogenesis of osteomyelitis. Understanding the specific bacterial factors — and the bone cell signals — that promote bone destruction and formation during infection could lead to new strategies for restoring bone balance, Skaar said. “Even if it’s not possible to kill the bacteria, compounds that manipulate bone growth or destruction might have some therapeutic benefit.” Still, Skaar is interested in treatments that will eliminate the infection.

The staph bacteria involved in osteomyelitis and in other persistent infections (such as lung infections in cystic fibrosis) are often a sub-class of staph known as “small colony variants.” These staph variants grow slowly and are resistant to entire classes of antibiotics commonly used to treat bone and lung infections, Skaar said. One way that staph bacteria become antibiotic-resistant small colony variants is by changing the way they generate energy. Instead of using respiration, they switch to fermentation, which blocks antibiotic entry and slows bacterial growth. In a high-throughput screen for compounds that activate a heme-sensing bacterial pathway, graduate student Laura Mike identified a compound that kills fermenting staph. The findings are reported in a recent issue of the Proceedings of the National Academy of Sciences. “This is a completely new molecular activity,” Skaar said. “We don’t know of other molecules that are toxic against fermenting bacteria.” The compound — and derivatives synthesized by Gary Sulikowski, Ph.D., and his team — might be useful in treating staph small colony variants, or in preventing their emergence.

The investigators demonstrated in culture that treating staph with the antibiotic gentamicin forced it to become a small colony variant and ferment, and that co-treatment with the new compound prevented resistance and killed all of the bacteria. “We think a really interesting therapeutic strategy for this compound is that it might augment the antimicrobial activity of existing classes of antibiotics by preventing resistance to them — it might extend the lifetime of these classes of antibiotics,” Skaar said. This would be similar to the drug Augmentin, which combines a traditional penicillin-type antibiotic and a compound that blocks bacterial resistance. The investigators are excited to test the new compound in the mouse model of osteomyelitis. First, they will treat the mice with gentamicin and assess whether staph small colony variants form. If so, they will co-administer the new compound to test if it prevents resistance, and they will also assess it as a single treatment for the persistent infection. Skaar stressed that Vanderbilt’s collaborative environment made these studies possible. Daniel Perrien, Ph.D., and Florent Elefteriou, Ph.D., in the Vanderbilt Center for Bone Biology and colleagues in the Vanderbilt University Institute of Imaging Science were critical in facilitating development of the bone infection model. Sulikowski and other colleagues in the Vanderbilt Institute of Chemical Biology (VICB) enabled the compound development. “This is exactly the kind of work the VICB is promoting — getting biologists like me together with chemists, to make new therapeutics,” Skaar said.

Science Daily
August 20, 2013

Original web page at Science Daily

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Physicians call for change in cancer tissue handling

Genetic sequencing technology is altering the way cancer is diagnosed and treated, but traditional specimen handling methods threaten to slow that progress. That’s the message delivered this week in a column appearing in the Journal of the American Medical Association (JAMA) by Scripps Clinic physicians Eric Topol, Kelly Bethel and Laura Goetz. Dr. Topol is a cardiologist who serves as chief academic officer of Scripps Health and director of the Scripps Translational Science Institute (STSI), leading Scripps’ genomic medicine research efforts. Dr. Bethel is a pathologist, and Dr. Goetz is a general surgeon and a researcher at STSI. “Deciding how best to obtain (tumor) samples and how best to process them for whole genome or exome sequencing is a pivotal yet unresolved issue with several layers of complexity,” the doctors wrote. “As the new clinical applicability of genomics emerges at a fairly rapid rate, the field of pathology will arrive at a tipping point for a fundamental change in how cancer specimens are handled.” Currently, tumor tissue obtained through a biopsy is fixed in formalin, a mixture of formaldehyde and water, and embedded in paraffin for microscopic viewing. However, because the chemical mixture damages DNA, sequencing tissue processed in this way can be difficult, if not impossible.

A better alternative is to also routinely freeze a portion of the specimen, which retains the tissue’s genetic coding while preserving it for future analysis. In order to have enough tissue to freeze, larger or additional biopsy samples may be required, especially when using minimally invasive needle biopsy procedures. “We need to completely rethink the way we have collected and stored cancer tissue samples for decades,” said Dr. Topol, “It’s becoming increasingly clear that obtaining an accurate map of a tumor’s DNA can be the key to determining the specific mutations that are driving a person’s cancer, how best to treat it and how likely it is to recur.” Even though complete genetic evaluations of tumors might require higher sample-storage costs and a more invasive biopsy procedure, most patients would likely agree to that option if it translates into a better diagnosis and possible treatment, the authors wrote. Evidence of such benefit must come from randomized clinical trials that compare detailed genetic evaluation of tumor tissue with the current standard of care for cancer patients, they said. “Drs. Goetz, Bethel and Topol’s editorial acknowledges kindly the critical role pathologists play in patient care,” said Dr. Stanley Robboy, president of the College of American Pathologists. “This type of change will require discussion about new operative standards, which will need the cooperation of surgeons, pathologists, ethicists and, of course, appropriate patient consents. It’s these types of implications we will need to consider and incorporate as a progressive healthcare agenda is moved forward.” Patients and their advocates have a direct role to play in this transformation and should raise questions about tumor specimen storage and genetic testing with their doctor, said Dr. Topol, who outlines a consumer-led digital health revolution in his book “The Creative Destruction of Medicine.”

Science Daily
January 22, 2013

Original web page at Science Daily

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Surgery has a more profound effect than anesthesia on brain pathology and cognition in Alzheimer’s animal model

A syndrome called “post-operative cognitive decline” has been coined to refer to the commonly reported loss of cognitive abilities, usually in older adults, in the days to weeks after surgery. In fact, some patients time the onset of their Alzheimer’s disease symptoms from a surgical procedure. Exactly how the trio of anesthesia, surgery, and dementia interact is clinically inconclusive, yet of great concern to patients, their families and physicians. A year ago, researchers at the Perelman School of Medicine at the University of Pennsylvania reported that Alzheimer’s pathology, as reflected by cerebral spinal fluid biomarkers, might be increased in patients after surgery and anesthesia. However, it is not clear whether the anesthetic drugs or the surgical procedure itself was responsible. To separate these possibilities, the group turned to a mouse model of Alzheimer’s disease. The results, published online this month in the Annals of Surgery, shows that surgery itself, rather than anesthesia, has the more profound impact on a dementia-vulnerable brain.

The team, led by Roderic Eckenhoff, MD, Austin Lamont Professor of Anesthesia, exposed mice with human Alzheimer disease genes, to either anesthesia alone, or anesthesia and an abdominal surgery. The surgery was similar to appendectomy or colectomy, very common procedures in humans. They found that surgery causes a lasting increase in Alzheimer’s pathology, primarily through a transient activation of brain inflammation. Also, a significant cognitive impairment persisted for at least 14 weeks after surgery compared to controls receiving anesthesia alone. Neither surgery nor anesthesia produced changes in normal non-transgenic animals. “In the mice, there was a clear and persistent decrement in learning and memory caused by surgery as compared with inhalational anesthesia — but only in the context of a brain made vulnerable by human Alzheimer-associated transgenes,” notes Eckenhoff. He also notes that at the time of surgery, the AD mice showed no outward symptoms of AD, despite having subtle evidence of ongoing neuropathology. “This timeline is analogous to both the age range and cognitive status of many of our patients presenting for a surgical procedure and suggests the window of vulnerability to surgery of the Alzheimer’s brain extends into this pre-symptomatic period,” says Eckenhoff. This period might be analogous to what is now called prodromal AD. “On the other hand,” cautions Maryellen Eckenhoff, PhD, a neuroscientist on the team, “the brain vulnerability seen in the AD mice may not translate well to people.” The AD mice used, like all current mouse models of Alzheimer disease, more closely resemble the situation in familial Alzheimer disease, which constitutes only a small minority of patients. She points out that it is not yet clear whether results from AD mouse models will represent patients who eventually get late-onset, or “sporadic” Alzheimer disease. These mice are, however, the current standard of choice for screening new drugs and have yielded considerable insight into Alzheimer pathogenesis.

The mechanism linking surgery and the cognitive effects seems to be inflammation. An inflammatory process is well known to occur as a result of surgery, at least outside the central nervous system. How this inflammatory process gains access to the brain, and accelerates AD pathology in a persistent way is still unclear. Postoperative cognitive decline has not been convincingly demonstrated to persist after three months in most people, and whether it predicts later dementia is still unclear. This study suggests that in the setting of a vulnerable brain, the cognitive deficits after surgery might be irreversible. However, the finding that inflammation is the underlying mechanism, immediately suggests a strategy for mitigating injury. “Human studies will be needed to first confirm these findings and then begin to deploy anti-inflammatory strategies to minimize injury,” adds Eckenhoff. “As a profession, doctors need to understand the long-term implications of our care, both positive and negative, and do all we can to delay the onset of dementia.”

Science Daily
October 2, 2012

Original web page at Science Daily

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Study finds ‘mad cow disease’ in cattle can spread widely in ANS before detectable in CNS

Bovine spongiform encephalopathy (BSE, or “mad cow disease”) is a fatal disease in cattle that causes portions of the brain to turn sponge-like. This transmissible disease is caused by the propagation of a misfolded form of protein known as a prion, rather than by a bacterium or virus. The average time from infection to signs of illness is about 60 months. Little is known about the pathogenesis of BSE in the early incubation period. Previous research has reported that the autonomic nervous system (ANS) becomes affected by the disease only after the central nervous system (CNS) has been infected. In a new study published online in the August issue of The American Journal of Pathology, researchers found that the ANS can show signs of infection prior to involvement of the CNS. “Our results clearly indicate that both pathways are involved in the early pathogenesis of BSE, but not necessarily simultaneously,” reports lead investigator Martin H. Groschup, PhD, Institute for Novel and Emerging Infectious Diseases at the Friedrich-Loeffler-Institut, Riems, Germany.

To understand the pathogenesis of BSE, fifty-six calves between four and six months of age were infected orally with BSE from infected cattle. Eighteen calves were inoculated orally with BSE-negative material from calf brainstem as controls. The study also included samples collected from a calf that had died naturally of BSE. Tissue samples from the gut, the CNS, and the ANS were collected from animals every four months from 16 to 44 months after infection. The samples were examined for the presence of prions by immunohistochemistry. Samples were also used to infect experimental mice that are highly sensitive to a BSE infection. A distinct accumulation of the pathological prion protein was observed in the gut in almost all samples. BSE prions were found in the sympathetic ANS system, located in the thoracic and lumbar spinal cord, starting at 16 months after infection; and in the parasympathetic ANS, located in the sacral region of the spinal cord and the medulla, from 20 months post infection. There was little or no sign of infection in the CNS in these samples. The sympathetic part of the ANS was more widely involved in the early pathogenesis than its parasympathetic counterpart. More bovines showing clinical symptoms revealed signs of infection in the sympathetic nervous system structures at a higher degree than in the parasympathetic tissue samples. The earliest detection of BSE prions in the brainstem was at 24 months post infection. However, infection detected in the spinal cord of one animal at 16 months post infection suggests the existence of an additional pathway to the brain.

“The clear involvement of the sympathetic nervous system illustrates that it plays an important role in the pathogenesis of BSE in cattle,” notes Dr. Groschup. “Nevertheless, our results also support earlier research that postulated an early parasympathetic route for BSE.” The results, Dr. Groschup says, indicate three possible neuronal routes for the ascension of BSE prions to the brain: sympathetic, parasympathetic, and spinal cord projections, in order of importance. “Our study sheds light on the pathogenesis of BSE in cattle during the early incubation period, with implications for diagnostic strategies and food-safety measures.”

EurekAlert! Medicine
July 24, 2012

Original web page at EurekAlert! Medicine

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Schmallenberg virus in calf born at term with porencephaly

From the end of August through the end of October 2011, a clinical syndrome involving adult cattle and the fetuses of pregnant cows emerged in the border area between the Netherlands and North Rhine-Westphalia, Germany. The syndrome was characterized by nonspecific clinical signs (fever, decreased milk production), severe diarrhea, and some abortions. A metagenomic analysis was conducted on pooled samples from cattle with acute signs on a farm in the city of Schmallenberg, Germany. The analysis detected nucleotide sequences homologous to arthropod-borne Akabane, Aino, and Shamonda viruses, all belonging to the family Bunyaviridae, genus Orthobunyavirus, and Simbu serogroup. Real-time PCR detected the genomic RNA of the new and emerging virus, tentatively designated Schmallenberg virus (SBV), in the blood of adult cattle, abdominal fluid of a stillborn calf, and brains of lambs born with birth defects on dozens of farms in the Netherlands, Germany, and Belgium. No data are yet available to predict how the emerging virus might affect the cattle industry. We report the case of a 1-week old calf with severe central nervous system (CNS) lesions probably caused by in utero infection with the new virus.

In Belgium in January 2012, a Belgian Blue multiparous cow gave birth to a 45-kg female calf that was morphologically normal but hypertonic and hyperreflexic. Pregnancy had proceeded uneventfully and lasted 9 months and 4 days. Spontaneous reflexes such as sucking, swallowing, micturition, defecation, and crying were completely preserved, but the calf was unable to stand, and its consciousness alternated from mild to severe depression. It was obviously blind and showed ventrolateral strabismus, but the pupils functioned normally. Muscle tone was permanently increased, as indicated by tetanus-like erection of the ears and by a violent but brief startle response to the slightest acoustic or tactile stimulation. When the calf was placed upright, loss of conscious proprioception was obvious; it maintained its position only a few seconds before collapsing. Altogether, the clinical signs suggested severe dysfunctions of the cerebral cortex, basal ganglia, and mesencephalon. The calf drank from a bottle twice a day for a week, but then was euthanized for humane reasons (infected decubital ulcers).

However, the cerebral hemispheres were replaced by 2 thin-walled, fluid-filled cysts with some floating islets and peninsulae corresponding to preserved cortex. There was variable preservation of the cerebrum, total liquefaction of occipital lobes, and irregular preservation of the outer layers of some parts of the temporal and frontal lobes. Altogether, the picture was compatible with severe porencephaly or hydranencephaly. The spine showed no sign of scoliosis, and movement of the limb joints was not restricted (i.e., no arthrogryposis). Samples were removed from the remnants of the cerebrum, diencephalon, and organs (thymus, lung, myocardium, jejunum, ileum, mesenteric lymph node, liver, spleen, kidney, and striated muscle), and 3 independent real-time PCR protocols were conducted to detect genomes of bovine viral diarrhea/mucosal disease virus, bluetongue virus serotype 8, and the novel SBV. Initial retrotranscription of the RNA genomes was followed by quantitative (real-time) PCR. The process was conducted by using our procedures and, for SBV, by following the protocol and using recently developed control reagents as described. The SBV genome was detected in only CNS samples (quantification cycle value 28.8); bovine viral diarrhea/mucosal disease virus and BTV-8 genomes were not detected. The new virus genome load was 1.61 × 104 copies per gram of cerebrum sample.

Emerging Infectious Diseases
May 1, 2012

Original web page at Emerging Infectious Diseases

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How cells limit inflammation in lung injury

Researchers at the University of Illinois at Chicago College of Medicine have found in an animal model of acute lung injury a molecular mechanism that allows cells of the immune system to reduce tissue damage from inflammation. Inflammation is part of the normal response to infection. One aspect of inflammation is the production of negatively charged oxygen-rich molecules by specialized white blood cells called phagocytes. The molecules, called reactive oxygen species (ROS), help to break up bacteria, allowing the phagocytes to “mop up” the broken pieces and clear out the infection. Unfortunately, ROS can also cause damage to normal tissue. The UIC researchers found that a channel through the cell membrane of phagocytes is able to modulate this destructive phase of inflammation. “Although the channel, called TRPM2, is found in many cell types in the immune system, including phagocytes, it’s function in these cells has been unknown,” said Anke Di, UIC research assistant professor in pharmacology and first author of the study.

The researchers were able to show that TRPM2 had a protective anti-inflammatory role in the animal model of ALI, and, further, it played a previously unknown role in protecting against inflammation and tissue injury generally. TRPM2’s protective effect was a result of its ability to dampen the production of the negatively charged ROS by modulating the electrochemical gradient — the difference in charge between molecules within the cell and outside the plasma membrane of the cell. ALI and its more severe form, acute respiratory distress syndrome (ARDS) result from pulmonary edema (leaky blood vessels) and inflammation. Both direct lung injury from infection and indirect lung injury from trauma, sepsis, pancreatitis, transfusions, radiation exposure and drug overdose can trigger ALI. It is fatal in almost 40 percent of cases. Inflammation plays an important role in ALI and a number of other human diseases, said Dr. Asrar Malik, UIC Schweppe Family Distinguished Professor and head of pharmacology and principal investigator of the study. Understanding how inflammatory damage to tissues is controlled normally may help develop therapies in the future, he said.

Science Daily
January 10, 2012

Original web page at Science Daily

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A new target to inhibit malaria and toxoplasmosis infection

Maryse Lebrun, Research Director at Inserm, and her fellow researchers at the Laboratoire Dynamique des Interactions Membranaires Normales et Pathologiques (Laboratory of Dynamics of Membrane Interactions in Normal and Pathological Cells — CNRS/ Montpellier Universities 1 and 2) in France, have characterised a protein complex that allows the agents that cause malaria and toxoplasmosis to infect host cells. This is a highly original mechanism, since the parasite supplies both the receptor which it inserts into the host cell membrane and the ligand it exposes at its surface. The three-dimensional structure of this complex has been now solved in collaboration with Dr. Martin Boulanger (Department of Biochemistry and Microbiology, University of Victoria, in Victoria, British Columbia, Canada). The new data is published in Science on 22 July 2011. It paves the way for new drugs designed to inhibit the formation of the protein complex in question and block invasion by Plasmodium falciparum in red blood cells.

Apicomplexa form a huge family of parasites that cause many different illnesses in humans and animals, and which includes Plasmodium, the parasite that causes malaria and Toxoplasma gondii, the agent that causes toxoplasmosis. Over a million people die of malaria every year. Over a third of the world’s population is at risk and the parasite has developed resistance to most of the anti-malaria drugs currently available. Toxoplasmosis is one of the most widespread congenital infections. Intracellular parasites such as these penetrate inside the cells of the infected organism. The research laboratory in Montpellier is investigating the mechanisms related to invasion of the host cell, a crucial step in the development of infection, the specific nature of which they want to identify so that new treatment targets can be designed. Researchers at Inserm and the CNRS recently deciphered the mechanism used by Apicomplexa to penetrate the cell, at cellular and molecular level. This involves a protein complex that assembles at the interface between the host cell membrane and the parasite membrane, forming a structure known as a moving junction (MJ). This is a highly original mechanism, since the parasite supplies both the receptor (RON2) which it inserts into the host cell membrane and the ligand (AMA1) which it exposes at its surface. These proteins are not present in the host, so this mechanism is specific to Apicomplexa parasites. The researchers have identified a short-peptide region in RON2 capable of binding AMA1 which has very strong affinity and inhibits invasion by the parasite.

In collaboration with a Canadian team (University of Victoria, Vancouver), the researchers crystallised the AMA1- RON2 peptide complex in Toxoplasma and mapped out the amino acids necessary for AMA1 and RON2 to interact and form the MJ in vivo. The RON2 peptide is inserted into a hydrophobic groove in AMA1, allowing the parasite to overcome the mechanical constraints it encounters as it invades the host cell. “By analysing the relation between structure and function and modelling the equivalent Plasmodium falciparum complex, we were able to identify an area on RON2 that is strongly implicated in the interaction specific to the various Apicomplexa parasites and thus understand the mechanisms used by antibodies targeting AMA1 to inhibit invasion,” explained Maryse Lebrun. Together, all this data can be used as the basis for developing drugs able to inhibit assembly of the AMA1-RON2 complex and the invasion of red blood cells by P. falciparum. Furthermore, since the AMA1-RON2 complex is also found for Apicomplexa that cause extremely costly veterinary infections such as avian coccidiosis, piroplasmosis, neosporosis and ovine toxoplasmosis, this strategy can also be applied in treating other parasitic infections. The results of the project may therefore have direct applications in human and veterinary medicine.

Science Daily
August 9, 2011

Original web page at Science daily

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Leaking blood vessels may be cause of infection-spreading condition

Researchers at St. Michael’s Hospital have put forward a new theory as to what causes sepsis, an often fatal condition that occurs when infection spreads throughout the body. Leaking blood vessels may actually be a cause of sepsis, rather than a symptom as previously thought, said Dr. Warren Lee. Dr. Lee’s hypothesis and a review of recent research on sepsis were published in the journal Science Translational Medicine. Sepsis kills about 1,400 people a day worldwide and is the second-leading cause of death in intensive care units in Canada. Despite intense research, the death rate remains high, between 30 and 50 per cent. Treatments largely consist of surgical removal or drainage of the infected site, antibiotics and supportive care. “The lack of therapies is not for want of effort. On the basis of promising in vitro and animal studies, many drugs have entered clinical trials only to fail, resulting in the description of sepsis research as a ‘graveyard’ of discovery,” said Dr. Lee, a critical care physician and researcher in the hospital’s L Ka Shing Knowledge Institute.

“The repeated failure of clinical trials suggests that some fundamental knowledge is lacking in our current understanding of the pathogenesis of human sepsis.” Dr. Lee said one reason previous trials may have failed is that they haven’t taken into account a key feature of sepsis in humans — diffuse microvascular leakage (widespread leakage from small blood vessels) leading to swelling in tissues and organs (edema). This causes organs throughout the body to malfunction. There have been no clinical trials of drugs designed to prevent or treat leaky blood vessels. “This is a major change in our thinking about sepsis that could lead to novel treatments to save lives,” Dr. Lee said. “Sepsis is a devastating illness. If we could develop drugs to stop blood vessels from leaking, we could save lives.”

Science Daily
July 12, 2011

Original web page at Science Daily

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New study upends thinking about how liver disease develops

In the latest of a series of related papers, researchers at the University of California, San Diego School of Medicine, with colleagues in Austria and elsewhere, present a new and more definitive explanation of how fibrotic cells form, multiply and eventually destroy the human liver, resulting in cirrhosis. In doing so, the findings upend the standing of a long-presumed marker for multiple fibrotic diseases and reveal the existence of a previously unknown kind of inflammatory white blood cell. The results are published in the early online edition of the Proceedings of the National Academy of Sciences. In all types of chronic diseases, healthy, functioning tissues are progressively replaced by fibrous scarring, which renders the tissues or larger organ increasingly dysfunctional until, eventually, it fails. The process is called fibrosis. In the human liver, the end result is cirrhosis, the 12th leading cause of death by disease in the United States with roughly 27,000 deaths annually. Fibrosis occurs in other organs as well, such as the heart, kidneys and lungs, with comparable deadly effect.

Scientists do not fully understand the process of fibrosis, particularly how problematic fibroblast cells are created. For years, conventional wisdom has posited that fibroblasts are likely to be transformed epithelial cells, a conversion called “epithelial to mesenchymal transition” or EMT. A protein called fibroblast-specific protein 1 (FSP1) has long been considered to be a reliable indicator of fibroblasts in injured organs undergoing tissue remodeling and has been broadly used to identify the presence of fibrotic disease. The new research undermines the validity of prevailing assumptions about EMT and FSP1, but also opens the door to new avenues of investigation that could ultimately lead to improved detection and treatment of cirrhosis and similar conditions. “This work, along with earlier papers, puts into question a whole area of research — at least in terms of the liver” said David Brenner, MD, Vice Chancellor for Health Sciences, dean of the UC San Diego School of Medicine and co-author of the paper. “The old evidence and assumptions about the source of fibroblasts and the role of FSP1 as a marker are not valid.”

Specifically, in experiments using cell cultures, human liver samples and mouse models, the researchers found no evidence of EMT — that transformed epithelial cells became liver fibroblasts. Rather, endogenous stellate cells appear to be the culprit, though the scientists note many types of cells seem to contribute, directly or indirectly, to liver fibrosis. Likewise, experiments proved FSP1 to be an unreliable marker for fibrosis. Cells containing FSP1 increased in human and experimental liver disease and in liver cancer, but researchers found that liver fibroblasts do not express the protein, nor do hepatic stellate cells — a major cell type involved in liver fibrosis. Similarly, FSP1 was determined not to be a marker for myofibroblasts (a fibroblast with some properties of a smooth muscle cell) or any precursors of myofibroblasts. “There have been hundreds of papers based on FSP1 as a marker,” said Brenner. “That thinking now seems to have been a mistake. One of the take-home messages of this paper is that FSP1 clearly can’t be reliably used as a marker.” On the other hand, the scientists discovered that FSP1 is a consistent marker for a previously unknown subset of inflammatory white blood cells or macrophages found in injured livers. The protein appears to also perform biological functions in the macrophages, though these remain to be determined.

Science Daily
January 11, 2011

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How key drug kills worms in tropical diseases

In a major breakthrough that comes after decades of research and nearly half a billion treatments in humans, scientists have finally unlocked how a key anti-parasitic drug kills the worms brought on by the filarial diseases river blindness and elephantitis. Onderstanding how the drug ivermectin works has the potential to lead to new treatments for the diseases, in which the body is infected with parasitic worms, said Charles Mackenzie, a professor of veterinary pathology in the College of Veterinary Medicine and researcher on the project. The diseases afflict about 140 million people worldwide, doing much of their damage in equatorial Africa. “Ivermectin is one of the most important veterinary and human anti-parasitic agents ever,” Mackenzie said. “Knowing specifically how it interacts with the body’s own immune system and kills parasitic worms opens up whole new treatment avenues.” The research appears in the current edition of the Proceedings of the National Academy of Sciences.

Elephantiasis (lymphatic filariasis) is caused by tiny worms spread via mosquitoes and results in severe swelling of the legs, arms and torso. River blindness (onchocerciasis) is spread by black flies, and after the worms die in a person’s eye, they can cause blindness and debilitating skin disease. Ivermectin works by killing the first stage of the worm in the human body, and also appears to paralyze the reproductive tract of the adult female worms, stopping reproduction of new parasites. What the researchers discovered is that the drug does this by preventing the worm from secreting proteins through a pore in its mid-body; ivermectin binds to receptors at the pore and blocks the secretions. It is the secretions that normally block a person’s ability to attack and kill the worm; after the drug prevents them, the host’s own immune system is able to attack and kill the parasites.

“Understanding how the worms were avoiding the host’s immune responses will greatly enhance our ability to manipulate the immune system to the advantage of the host, and perhaps develop vaccines,” Mackenzie said. “Also, one of the most important challenges in the overall effort against filarial infections relates to the development of resistance and the loss of efficacy of the drugs we use; this new knowledge provides an important key to understanding and perhaps preventing resistance.” Ivermectin was developed by pharmaceutical firm Merck & Co. in the 1970s. It was donated in 1987 for use to treat river blindness, as existing drugs were in fact inducing blindness. Ivermectin was able to be used safely in mass drug administration programs in many developing countries, shifting the paradigm for how public health programs delivered medicines in rural areas. The drug then was used in other parasitic disease programs, such as the one for elephantiasis, treating more than 100 million people for that disease. Mackenzie has worked for more than 20 years on tropical filarial diseases, much of that time partnering with Tim Geary at McGill University in Montreal. Geary’s lab was critical in the ivermectin findings, as was McGill graduate student Yovany Moreno.

Science Daily
November 23, 2010

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Real-world proof of hand washing’s effectiveness

Scientists are reporting dramatic new real-world evidence supporting the idea that hand washing can prevent the spread of water-borne disease. It appears in a new study showing a connection between fecal bacteria contamination on hands, fecal contamination of stored drinking water, and health in households in a developing country in Africa. The study is in ACS’ Environmental Science & Technology, a semi-monthly journal. Alexandria Boehm, Jenna Davis, and their students note that almost half of the world’s population — over 3 billion people — have no access to municipal drinking water supply systems. They obtain drinking water wells, springs, and other sources, and store it in jugs and other containers in their homes. Past research showed that this stored water can have higher levels of bacterial contamination than its source. But nobody knew why. The scientists found a strong link between fecal contamination on the hands of household residents and bacterial contamination in stored water in Dar es Salaam, Tanzania. Stored water contained nearly 100 times more fecal bacteria than the source where it was collected. “The results suggest that reducing fecal contamination on hands should be investigated as a strategy for improving stored drinking water quality and health among households using non-networked water supplies,” the report notes.

Science Daily
June 8, 2010

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Utility of immunohistochemical markers in differentiating benign from malignant follicular-derived thyroid nodules

Thyroid nodules are common among adults though only a small percentage is malignant, which can histologically mimic benign nodules. Accurate diagnosis of these thyroid nodules is critical for the proper clinical management. We investigated immunoexpression in 98 surgically removed benign thyroid nodules including 52 hyperplastic nodules (HN) and 46 follicular/ Hurthle cell adenomas (FA), and 54 malignant tumors including 22 follicular carcinoma (FC), 20 classic papillary carcinoma (PTC), and 12 follicular variant papillary carcinoma (FVPC). The staining results showed that malignant tumors express galectin-3, HBME-1, CK19 and Ret oncoprotein significantly more than benign nodules. The sensitivity of these markers for the distinction between benign and malignant lesions ranged from 83.3% to 87%. The sensitivity of two-marker panels was not significantly different. Immunoexpression was usually diffuse and strong in malignant tumors, and focal and weak in the benign lesions. Our findings indicate that these immunomarkers are significantly more expressed in malignant tumors compared to benign lesions and may be of additional diagnostic value when combined with routine histology.

BioMed Central
February 23, 2010

Original web page at Diagnostic Pathology

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Nuclear magnetic resonance technology could reduce time spent under the knife

Chemical fingerprints of tissue samples taken during operations could soon help surgeons to decide where to make their incisions. Two groups — one based in the United Kingdom, the other in France — are leading efforts to use nuclear magnetic resonance (NMR) spectroscopy to analyse the metabolites in biopsies. The analyses should reveal whether cells in the sample are healthy and — for the first time — relay that information back to the operating theatre within minutes. In February, a team led by Jeremy Nicholson, head of the department of surgery and cancer at Imperial College London, is planning to install a £300,000 (US$490,000) NMR machine that can study solid samples at a surgical unit at St Mary’s Hospital. It is the first step in a programme that could later see the instruments rolled out to intensive-care units and other wards in hospitals affiliated with Imperial. The machine will be able to rapidly load samples and disposable inserts, allowing the needles used for biopsies to go directly into the spectrometer.

Nature
January 12, 2010

Original web page at Nature

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West Nile virus infection may persist in kidneys years after initial infection

A new study shows that people who have been infected with West Nile virus may have persistent virus in their kidneys for years after initial infection, potentially leading to kidney problems. The research appeared in the January 1 issue of The Journal of Infectious Diseases. Spread by infected mosquitoes, West Nile virus was first detected in the United States in 1999. Since then, approximately 25,000 human cases have been reported, causing more than 1,000 deaths. Many more have become infected without showing symptoms. Previous animal studies raised the possibility that patients may still be infected with the virus several years after recovering from their initial illness. Prior to this latest research, however, humans were thought to remain infected with West Nile virus only for the first few days of illness. The study, led by Kristy Murray, DVM, PhD at the University of Texas School of Public Health in Houston, demonstrates that not all individuals clear the virus from their system within the first few days — and that it can remain in the kidneys for years, potentially leading to kidney failure.

Dr. Murray and her colleagues followed more than 100 patients in Houston with severe initial West Nile virus infections for seven years. Individuals were evaluated and blood samples collected every six months. More than half continued to have infection-related symptoms years after their initial illness, although symptoms began to plateau around two years after infection. The deaths of five participants due to kidney failure led researchers to consider whether the kidney could be a preferred replication site for the virus. To test this hypothesis, Dr. Murray and her team collected urine samples from 25 patients from their original cohort and tested them for presence of West Nile virus. In this group, five patients (20 percent) tested positive for the virus. Viral RNA could be detected in the urine for at least six years following infection. Four of the five patients who tested positive for virus also experienced chronic symptoms. Of these five, one patient developed kidney failure. These results show that West Nile virus is capable of long term persistence in patients, particularly when chronic symptoms are present.

In an accompanying editorial, Ernest Gould, PhD, of the Centre for Ecology and Hydrology in Oxford, England, points out that this study raises the additional concern that West Nile virus and other flaviviruses may be transmitted to mosquitoes by apparently healthy humans or animals. This possibility has the potential to start epidemics in new regions of the world. According to Dr. Murray, patients who have been infected with West Nile virus should “have their kidneys monitored by their physician for any evidence of disease and be aware that persistent infection of the kidneys can happen.” Dr. Murray also reminds the public to take proper precautions to protect themselves from mosquito bites during transmission seasons, typically the summer and fall, to avoid infection. More research is needed to “understand the underlying mechanisms related to the shedding of virus particles in urine, whether shedding of the virus is constant or intermittent, and whether or not this represents true infection resulting in kidney disease,” the investigators say. They continue to evaluate all study participants, particularly in regard to kidney function. In addition, they are focusing on developing treatment options for those who remain infected with the virus.

Science Daily
January 12, 2010

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How diarrheal bacteria cause some colon cancers revealed in mouse studies

Johns Hopkins scientists say they have figured out how bacteria that cause diarrhea may also be the culprit in some colon cancers. The investigators say that strains of the common Bacteroides fragilis (ETBF) dupe immune system cells into permitting runaway colon tissue inflammation, a precursor for malignant growth. This could be the H. pylori of colon cancer,” says Johns Hopkins infectious disease specialist, Cynthia Sears, M.D., referring to the bacteria long known to cause stomach ulcers and suspected of causing the majority of stomach cancers. Her studies suggest that ETBF uses tissue inflammation to cause colon cancer in a similar way that H. pylori causes stomach tumors. A so-called enterotoxigenic bacterium, the germ is widely known to cause diarrhea in children and adults in the developing and developed world, and a previous study in Turkey has linked it to colon cancer.

The bacteria, which colonize in the gut, cause no symptoms in some individuals, but others develop diarrhea and colon inflammation, which has been linked to cancer growth. Unlike the case with H. pylori, it is unknown whether standard antibiotics can eradicate the microbe, experts say. To track the link between ETBF and colon cancer, the Johns Hopkins researchers conducted a series of tests in mice bred to carry mutations in a colon cancer-causing gene called APC. Their results, published in the August 23 issue of Nature Medicine, show that mice infected with ETBF developed diarrhea which resolved quickly, but within a week, developed inflammation and small tumors in the colon. One month later, the colons were pockmarked with tumors. Mice infected with a non-toxin producing strain of the bacteria were free of diarrhea, inflammation and tumors.

Next, Sears and the Johns Hopkins team evaluated the bacteria’s effect on immune responses that may contribute to cancer development. In ETBF-infected mice, they found high levels of a protein called pStat3, which, in its normal role, acts as a signal to trigger inflammation. One of those signals activates an immune cell called T-helper 17 (Th17). Th17 cells produce molecules that have been implicated in fostering inflammation of tissues. Th17 activity in the gut of germ-bearing mice was 100 times greater than normal, according to the investigators, and when they blocked the effects of Th17, they were able to reverse inflammation and tumor growth. Drew Pardoll, M.D., Ph.D., an immunologist and cancer researcher at Johns Hopkins, speculates that in humans, infection with ETBF “produces a low-level inflammation that persists for a long time.” “If what we are seeing in mice holds true in humans, the chronic inflammation damages genetic material in the colon cells, allowing them to grow uncontrollably and develop into tumors earlier and more progressively than if they were not infected with ETBF,” Pardoll says.

Sears first witnessed the impact of diarrheal pathogens two decades ago in a refugee camp in Thailand where children, especially, were vulnerable to infection where water sanitation is poor. Most diarrheal disease is short-lived but can be very severe, Sears says, and it is common worldwide. The ETBF microbe is found in the gut of up to 20 to 35 percent of children and adults and, according to the Turkish study, in as many as 40 percent of colon cancer patients. Sears and Pardoll believe that ETBF may collude with other types of normal bacteria in the gut to promote cancer. The microbe itself is difficult to culture from stool specimens, according to the investigators, so they are working on blood tests to detect antibodies to the pathogen’s toxin, which may show whether an individual has been exposed to it and perhaps determine who may be at risk for colon cancer. The investigators also envision vaccines and drug therapies that neutralize the pathogen’s toxin and its ability to inflame tissues.

Science Daily
September 8, 2009

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Drug shows cancer stem cells not invulnerable

A promising new cancer drug targets and kills the pernicious tumour cells responsible for metastasis and relapse. It is far too early to test the drug, called salinomycin, in humans, but the findings offer hope that the so-called cancer stem cells will eventually prove vulnerable to treatment. “It’s been thought that these cells are responsible both for metastasis and for recurrence following anti-cancer therapy,” says Piyush Gupta, a molecular biologist at the Broad Institute of MIT and Harvard in Cambridge, Massachusetts, who led the new study. “There’s been a lot of evidence to suggest that cancer stem cells are resistant to a variety of cell-death-inducing agents.” Gupta’s team found that salinomycin kills breast-cancer stem cells at least 100 times more effectively than another popular anti-cancer drug. And mice implanted with human breast-cancer cells and later treated with salinomycin showed fewer signs of metastases than mice given a standard cancer therapy. One reason cancer stem cells have been hard to target with drugs is their rarity in tumours and in laboratory cancer-cell cultures. This makes it difficult to hunt for drugs that selectively kill the cells.

To sidestep this problem, Gupta and his colleague Tamer Onder, formerly of the Whitehead Institute for Biomedical Research, also in Cambridge, reprogrammed normal skin cells into cells with some of the properties of cancer stem cells. Like normal cancer stem cells, the reprogrammed cells proved impervious to chemotherapy. Next, Gupta and Onder’s team tested about 16,000 different drugs against the reprogrammed cells: 32 of those compounds killed the reprogrammed stem cells, but not normal skin cells. Of the 32, salinomycin proved one of the most effective and the easiest to obtain in large quantities, Gupta says. Tests in laboratory-cultured human breast-cancer cells confirmed the drug’s potency. Mice injected with human breast-cancer cells – a common model for the disease – developed fewer aggressive tumours when treated with salinomycin, compared with a commonly used anti-cancer drug called paclitaxel. Further experiments in animals are needed before even thinking of trying salinomycin in humans, Gupta says. The researchers tested the drug on genetically homogenous laboratory cultures, so it also remains to be seen whether it will have the same effect on tumour cells collected from living people. Little is known about the drug’s safety, or whether it would find its way through the bloodstream to human tumours. Gupta says the significance of his team’s result is proof that cancer stem cells aren’t invincible to drugs. “They do have weaknesses and we’ve essentially shown an approach to find them systemically,” he says.
Source: Cell

New Scientist
August 25, 2009

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Gaining a better understanding of kidney diseases

By introducing a genetic switch in mice it is possible to increase or decrease the production of specific protein molecules in their kidneys. Thus, researchers can study the influence of specific proteins on disease development. Scientists of the German Cancer Research Center (Deutsches Krebsforschungszentrum, DKFZ), Heidelberg University Hospitals and other research institutes have published this model of investigating severe kidney diseases in the latest issue of Nature medicine. Cystic kidney disease, renal fibrosis, or renal cell carcinoma: Many diseases of the excretory organs are characterized by overproduction or – on the contrary – absence of characteristic proteins in the renal cells. An international research team under the leadership of scientists from DKFZ and Heidelberg University Hospitals has now developed an animal model to better investigate these conditions. The researchers introduced a genetic switch into the genome of mice. This switch allows to selectively turn on and off the production of disease-typical proteins in renal tissue. It is activated simply by adding the antibiotic tetracycline to the animal food.

To find out whether it is possible to study the development of kidney diseases in the genetically modified (transgenic) animals, the investigators stimulated the production of c-Myc in the renal tissue of the mice. Numerous tumors have been reported to be associated with elevated levels of this transcription factor. Shortly after activation of the c-Myc gene the animals started developing cysts that led to organ failure. Pathologists also discovered renal cell carcinomas in some of the mice. As a reaction to the overexpression of another signaling molecule, the mice developed renal fibrosis. Earlier attempts to study disease development using transgenic animals have often failed because the proteins to be studied are overproduced in the murine embryos already. As a result, the animals often develop severe malformations that make meaningful conclusions impossible. “A particular advantage of our model is that we can switch on and off disease-typical renal proteins at any given time,” explained Associate Professor (PD) Dr. Robert Kösters of the Institute of Human Genetics of the University of Heidelberg and Professor Dr. Hermann-Josef Gröne of the German Cancer Research Center. “Thus, we are able to simulate the natural course of disease development and also of healing processes.”

Science Daily
September 30, 2008

Original web page at Science Daily

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Biphasic onset of splenic apoptosis following hemorrhagic shock: critical implications for Bax, Bcl-2 and Mcl-1 proteins

The innate immune response to trauma-hemorrhage involves inflammatory mediators, thus promoting cellular dysfunction as well as cell death in diverse tissues. These effects ultimately bear the risk of post-traumatic complications, such as organ dysfunction, multiple organ failure or adult respiratory distress syndrome. In this study, a murine model of resuscitated hemorrhagic shock (HS) was used to determine the apoptosis in spleen as a marker of cellular injury and reduced immune functions. Male C57BL-6 mice were subjected to sham operation or resuscitated HS. At t = 0 hrs, t = 24 hrs and t = 72 hrs, mice were euthanized, the spleens were removed and evaluated for apoptotic changes via DNA fragmentation, caspase activities and activation of both extrinsic and intrinsic apoptotic pathways. Spleens from untreated mice were used as control samples.

HS was associated with distinct lymphocytopenia as early as t = 0 hrs after hemorrhage without regaining baseline levels within the consecutive 72 hrs when compared with sham and control group, respectively. A rapid activation of splenic apoptosis in HS mice was observed on t = 0 hrs and t = 72 hrs after hemorrhage and predominantly confirmed by increased DNA fragmentation, elevated caspase-3/7, -8, -9 activities and enhanced expression of intrinsic mitochondrial proteins, respectively. Accordingly, mitochondrial pro-apoptotic Bax and anti-apoptotic Bcl-2 proteins were inversely expressed within the 72 hrs observation period, thereby supporting significant pro-apoptotic changes. Solely on t = 24 hrs, expression of the anti-apoptotic Mcl-1 protein shows significant increase when compared with sham operated and control animals. Furthermore, expression of extrinsic death receptors was only slightly increased. Our data suggest that HS induces apoptotic changes in spleen through a biphasic caspase dependent mechanism and implies a detrimental imbalance of pro- and anti-apoptotic mitochondrial proteins Bax, Bcl-2 and Mcl-1, thereby promoting post-traumatic immunosuppression.

BioMed Central
February 5, 2008

Original web page at BioMed Central

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Apoptosis and pathogenesis of avian influenza A (H5N1) virus in humans

The pathogenesis of avian influenza A (H5N1) virus in humans has not been clearly elucidated. Apoptosis may also play an important role. We studied autopsy specimens from 2 patients who died of infection with this virus. Apoptosis was observed in alveolar epithelial cells, which is the major target cell type for the viral replication. Numerous apoptotic leukocytes were observed in the lung of a patient who died on day 6 of illness. Our data suggest that apoptosis may play a major role in the pathogenesis of influenza (H5N1) virus in humans by destroying alveolar epithelial cells. This pathogenesis causes pneumonia and destroys leukocytes, leading to leukopenia, which is a prominent clinical feature of influenza (H5N1) virus in humans. Whether observed apoptotic cells were a direct result of the viral replication or a consequence of an overactivation of the immune system requires further studies

Emerging Infectious Diseases
May 15, 2007

Original web page at Emerging Infectious Diseases

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New evidence questions the simple link between prion proteins and vCJD

While newly published research confirms that under laboratory circumstances prion-protein can be absorbed across the gut, it also shows that this is unlikely to occur in real life. In addition, the results show that the places in the gut that do take up these disease-associated proteins are different from the locations where infectivity is known to be amplified. The findings will be published in the Journal of Pathology. Since the outbreak of BSE in cattle and vCJD in humans, scientists have struggled to make sense of how an abnormal variation of a normal protein can trigger an infectious disease. Some are questioning whether this simple relationship exists at all. This paper adds new evidence that can inform the debate.

Firstly, it is known that individual people and animals have different levels of genetic susceptibility to this group of diseases, but no one knows how this resistance is achieved. One option is that resistant people do not absorb the disease-associated prion protein (PrP) from their guts. To test this, the researchers worked with 50 sheep, with different degrees of genetic resistance to scrapie – the sheep form of the disease. When they injected material containing abnormal prion protein (PrP) into the sheep’s gut, it was equally absorbed by all sheep. “This clearly shows that resistance is not achieved by blocking uptake of abnormal proteins from the gut – it must be achieved by some other mechanism,” says lead author Dr Martin Jeffrey.

Secondly, they looked in more detail at the route of absorption in the gut. Using surgically modified sheep, they loaded a small area of the gut with a fluid mixture containing 0.5 grams of scrapie infected brain containing a large amount of the disease specific variant of the PrP protein and watched how it was taken up. They saw the abnormal PrP was rapidly taken up by finger-like projections called villi and passed in to the lymph. It was not, however, taken up by structures called Peyer’s nodules, that are believed to be the places where animals amplify the infective agent. “The fact the PrP isn’t taken up by the Peyer’s nodules questions whether PrP is really infectious, or whether PrP is really just a secondary marker of the presence of the scrapie agent,” says Jeffrey.

His belief in this need to reappraise the fundamental understanding of prion diseases is enhanced by one more observation published in this same paper. The team pre-digested a mixture containing disease specific PrP with standard stomach contents, and then injected the resulting mixture into the gut. No PrP transferred into the villi. When they used a highly sensitive version of Western Blot analysis to examine the contents of this pre-digested mixture, they found only the faintest suggestion that some of the PrP had survived. This was despite the fact that the original mixture had a contained a high level of PrP. “Think about it – a sheep grazing in a field is not naturally exposed to highly infected brain and could only pick up a tiny amount of PrP from other tissues. This will then be exposed to 48 hours or more digestion before it arrives in the gut, and our experiments show that after this, the chance of there being more than an unmeasurably small amount of PrP left to absorb is very small,” says Jeffrey.

“As sheep can become infected, the theoretical probability of this being due to an invisible sub-fraction of digestion resistant PrP molecules is unlikely. The possibility of there being infectious molecules other than PrP must therefore be seriously considered,” says Jeffrey. “A lot of people are completely wedded to the prion hypothesis of diseases like vCJD, but the more you deal with whole animals as opposed to relying purely on in vitro studies, the more cautious you are about saying that prion proteins alone cause the disease,” says Martin Jeffrey. In a commentary published in the same edition of the journal Dr Nicole Sales of the Department of Infectology, at the Scripps Research Institute Jupiter, Florida, suggests that one possible explanation that keeps with the prion hypothesis is that infection occurs as PrPs are absorbed in the mouth, rather than in the gut. Dr Jeffrey, however, is not convinced by this argument. “Were infection to be acquired from the mouth then the first tissues to accumulate infectivity would be lymph nodes in the throat or the tonsils. But we don’t tend to see this in animals, and have no reason to believe it would be different in humans,” comments Jeffrey.

Source: John Wiley & Sons, Inc.

Bio com
April 11, 2006

Original web page at Bio com

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Liver C-reactive protein production linked to atherosclerosis

New research shows that levels of C-reactive protein (CRP), produced by the liver and not at the lesion site, correlate with the degree of atherosclerosis. The paper by Sun et al., “C-reactive protein in atherosclerotic lesions: its origin and pathophysiological significance,” appears in the October issue of The American Journal of Pathology and is accompanied by a commentary. C-reactive protein (CRP) is a hallmark of inflammation and tissue damage, as in arthritis or infection. It is also widely touted as a marker for cardiovascular disease, with doctors using patient CRP levels to improve risk assessment. However, whether CRP is merely a risk marker or is actually a contributing factor of cardiovascular disease has remained controversial.

To address these issues, Dr. Jianglin Fan’s group examined the role of CRP in two rabbit models of atherosclerosis: high cholesterol diet or LDL receptor deficiency. Rabbits represent highly suitable models as they quickly form atherosclerotic plaques in response to high serum cholesterol, and rabbit CRP shares 70% homology with human CRP. As expected, hypercholesterolemic rabbits developed atherosclerotic plaques. Upon further examination, serum CRP levels were found to positively correlate with plaque size. CRP was found in plaques of various stages, including early and advanced lesions, but it did not appear to associate with macrophages, as had been suggested. Similar results were seen in human aortic lesions.

To determine where CRP protein was being produced, CRP mRNA levels were measured by Northern blot and real-time RT-PCR. CRP mRNA was only detected in liver obtained from atherosclerotic rabbits but was undetectable in vascular cells or macrophages. Again, results were confirmed in human specimens: insignificant mRNA levels found in atherosclerotic aorta compared to high levels in liver.
Finally, in vitro analysis revealed that hepatocytes, but not macrophages, expressed CRP mRNA following stimulation by inflammatory molecules. These data highlight a powerful role for the liver in generating the CRP that is associated with atherosclerotic lesions. Nonetheless, we are still left wondering whether CRP is a cause, result, or both of heart disease, as presence at the scene of a crime is not necessarily evidence of guilt. Thus, CRP could be an innocent bystander, a victim, or possibly an atheroprotective force. Sun and colleagues acknowledge that “further studies will be required to clarify whether decreasing CRP alone without changing the plasma cholesterol level can be beneficial for the treatment of atherosclerosis.”

This study uncovers new approaches for the treatment of atherosclerosis. In the future, Dr. Fan hopes to “test whether any therapeutic inhibition of CRP levels can be beneficial for [coronary heart disease] patients or preventive from coronary artery syndrome or plaque rupture. Importantly, we will target the liver rather than the vascular wall (such as macrophages) for the inhibition of CRP.”

Science Daily
October 25, 2005

Original web page at Science Daily