Parasitic infection may have spoiled zebrafish experiments

A common parasite that infects laboratory zebrafish may have been confounding the results of years of behavioural experiments, researchers say – but critics say the case isn’t proven.

Like the rat, the zebrafish (Danio rerio) is used in labs worldwide to study everything from the effects of drugs, to genetic diseases and disorders such as schizophrenia and autism. Since both zebrafish and people are highly social, researchers think that zebrafish may be a better lab model for some human behaviours than rodents.

Zebrafish demonstrate their preference for each other by clustering into shoals – a social behaviour that researchers measure when they want to test how drugs affect zebrafish stress and anxiety levels, as a proxy for potential human responses. But this behaviour can change when fish are infected with a neural parasite called Pseudoloma neurophilia, scientists from Oregon State University in Corvallis report in a paper published on 11 July in the Journal of Fish Diseases.

The team say that individual fish infected with P. neurophilia swim closer to each other than do non-infected fish, a behaviour that is also associated with increased stress and anxiety. The finding casts doubt on results from previous experiments, says lead study author Sean Spagnoli, a veterinary surgeon – since the infection may have scrambled researchers’ interpretations of shoaling behaviour.

Spagnoli first heard that a parasite was infecting many laboratory zebrafish when he was working at the Zebrafish International Resource Center (ZIRC) in Eugene, Oregon – a central repository which sends out zebrafish strains to researchers and also tests zebrafish health. P. neurophilia settles in the brain, spinal cord and nerves of zebrafish.

“The paper is great, as it raises some doubts about the way behaviour may be used to study brain function in zebrafish,” says Robert Gerlai, a behavioural geneticist from the University of Toronto Mississauga in Canada. But he advises not jumping to conclusions on the basis of one study. Gerlai has concerns about the work; in particular, he says, Spagnoli’s team relied on a low-tech method to measure their fish shoals, taking screen snapshots and measuring the distance between each fish rather than more precise continuous tracking. And the researchers didn’t check what else might have been affecting the zebrafish, he adds.

Elena Dreosti, a geneticist at University College London, says that the paper’s data are weak and the effects it shows are small. “Considerable additional work is needed to know if this is likely to have a significant impact on the type of behaviour research that is done by the community working with zebrafish,” she says.

But Spagnoli says that his low-tech method is all that’s needed to raise the red flag that infection can influence behaviours such as shoaling. He agrees that he hasn’t proven that the P. neurophilia is directly responsible for the changed behaviour – but says that his study suggests that shoaling changes when the parasite is present.

As many as half of all laboratory facilities may be using some infected zebrafish, according to ZIRC data from 2015 – although only 28 facilities submitted their zebrafish to the centre for health checks that year. Within a facility, infection rates hover around 7-10%; some tanks may have no infected zebrafish, but others have many, Spagnoli says.

Nuno Pereira, a zebrafish veterinarian at the Gulbenkian Science Institute in Oeiras, Portugal, says that most researchers are already aware of the importance of testing for the parasite – and Spagnoli agrees that labs have drastically improved their screening protocols.

But Spagnoli thinks that many labs may still have a significant number of fish that are infected. “I haven’t seen a single paper that stated that ‘fish used were certified pathogen-free for P. neurophilia’,” he says. The team will continue to study the parasite’s effects, he says, and is also looking at the potential influence of another common contaminant, Mycobacterium chelonae, on shoaling behaviour.

Nature doi:10.1038/nature.2016.20308  Nature  Original web page at Nature 


* New technique helps link complex mouse behaviors to genes that influence them

Mice are one of the most commonly used laboratory organisms, widely used to study everything from autism to infectious diseases. Yet genomic studies in mice have lagged behind those in humans.

“Genome-wide association studies — matching genes to diseases and other traits — have been a big deal in human genetics for the past decade,” said Abraham Palmer, PhD, professor of psychiatry at University of California San Diego School of Medicine. “But progress hasn’t been so great in animal genetics. That’s because researchers have mostly been using crosses between inbred strains, making it impossible to pinpoint specific genomic regions or individual genes associated with a trait. In addition, we didn’t previously have good ways of genotyping animals in a cost-efficient way.”

Now, in a study published July 4, 2016 in Nature Genetics, Palmer’s team used 1,200 outbred mice, which are more similar to a natural population, to test a new cost-effective technique to search for specific genes linked with 66 different physical and behavioral traits.

“This is a system that could be used to discover genes associated with any complex trait a researcher is interested in, in any animal model,” Palmer said. “We can look at any trait and rapidly develop hypotheses about specific genes. It’s like genome-wide association studies in humans, but less expensive. And we can look at certain traits that we can’t in humans.”

Previously, only large regions of a chromosome could be associated with a particular mouse trait or behavior. Palmer’s method takes advantage of the superior mixing that is present in an outbred population to help drill down to specific genes using two steps: genotype-by-sequencing, which sequences about one percent of the mouse genome; and RNA sequencing, which identifies only genes turned “on” in a particular tissue, such as the brain.

With this approach, the researchers found numerous associations between genes and the traits they are associated with. For example, they report that the mouse gene Azi2 is associated with the effects methamphetamines have on body movements, and that mouse gene Zmynd11 is associated with anxiety-like behavior. The findings may be clinically relevant, as humans have analogous genes, Palmer said.

Next, the team will engineer mice that specifically lack these genes to determine if the associations are truly causal and to better understand the underlying mechanisms.

“This study has been extremely gratifying since this is the first time these two genes have been identified as playing roles in psychological conditions,” Palmer said. “And now we can think about targeting these genes or the proteins they encode with novel therapeutics.” Science Daily  Original web page at Science Daily


US government issues historic $3.5-million fine over animal welfare

Antibody provider Santa Cruz Biotechnology settles with government after complaints about treatment of goats. Santa Cruz Biotech has used goats to produce antibodies.

The US government has fined Santa Cruz Biotechnology, a major antibody provider, US$3.5 million over alleged violations of the US Animal Welfare Act. The penalty from the US Department of Agriculture is the largest in the agency’s history.

The company, which is headquartered in Dallas, Texas, will pay the fine as part of a settlement with the US Department of Agriculture (USDA). The agency had lodged three animal-welfare complaints against Santa Cruz Biotech, after USDA inspectors found evidence that the company mistreated goats at its facility in California.

Santa Cruz Biotech contested the government complaints, and the 19 May settlement agreement says that the company “neither admits nor denies” that it violated US animal-welfare regulations.

The settlement also permanently revokes Santa Cruz Biotech’s government licence to sell, buy, trade or import animals. And it requires the company to cancel its registration to operate as a research facility that uses animals. The company had extracted antibodies for research from animals such as goats and rabbits after injecting the animals with proteins to stimulate antibody production.

Neither Santa Cruz Biotech nor representatives of Covington & Burling, a Washington DC law firm that represents the company, have responded to Nature’s request for comment on the settlement.

Cathy Liss, president of the Animal Welfare Institute, an advocacy group in Washington DC, says that she is shocked by the unprecedented size of the fine on Santa Cruz Biotech. The largest previous fine that the USDA had imposed for animal-welfare complaints was a $270,000 penalty levied in 2011 against Feld Entertainment, which operates the Ringling Brothers and Barnum & Bailey Circus.

The settlement with Santa Cruz Biotech marks the end of a long-running investigation of the company’s animal-welfare practices. The USDA has lodged three animal-welfare complaints against Santa Cruz Biotech since 2007, after agency inspectors reported finding problems such as goats with untreated coyote bites and massive tumours, and rabbits being housed in cruel conditions. USDA inspectors also discovered that Santa Cruz was keeping 841 goats in a hidden facility.

In February, Nature reported that more than 5,000 goats and rabbits had disappeared from Santa Cruz’s facilities before a scheduled hearing on the USDA complaints. Santa Cruz would not confirm whether the animals were killed or sold.

After news of the animals’ disappearance became public, some scientists took to social media to call for a boycott of Santa Cruz’s products. Among them was Stephen Floor, a biologist at the University of California, Berkeley, who says that his lab has since sought out other antibody providers.

Floor says that losing Santa Cruz as an antibody provider will create extra work for researchers. Because the quality and type of antibodies varies widely, individual labs often stick with products from a single company to ensure that their experiments are replicable. “That said, I think any scientist will happily do that work in order to ensure that animal rights are a priority,” he says.

Nature doi:10.1038/nature.2016.19958  Nature  Original web page at Nature


* Scientists still fail to record age and sex of lab mice

The largest-ever analysis of the quality of mouse studies reveals that as recently as 2014, only around 50% of research papers recorded both the sex and age of the animals used — key details needed for others to assess and reproduce the research.

The analysis, which used software to trawl through the text of more than 15,000 open-access papers published between 1994 and 2014, also reveals the preferences of different research fields. Cardiovascular research tends to use male mice, whereas research on infectious diseases such as HIV and tuberculosis favours female mice, for example.

The study is “the strongest evidence about sex and age bias through biomedical research to date”, say its authors.

Many researchers have pointed out that male and female mice — like men and women — can have different responses to drugs or different behaviours in laboratory experiments. One study last year, for instance, found that although inhibiting the function of immune cells called microglia helps to relieve pain in male mice, it doesn’t do so in female mice. The difference might explain why some clinical trials of pain drugs have failed.

Recognizing the problem, the US National Institutes of Health said in 2014 that it would require researchers to report their plans for the balance of male and female animals in preclinical studies. But even before that, scientists had been urged to report in more detail the strains, sexes and ages of the mice they experimented on.

“It’s useful to see what’s happening in a large-scale study — which has thrown up some things we weren’t expecting,” says Andrew Brass, a bioinformatician at the University of Manchester, UK, and a co-author of the study, which is published in eLife.

One surprise was that although recording of animal studies improved through the 1990s and 2000s, standards seem to have plateaued after 2010 — despite the introduction that year of a voluntary checklist to improve reporting, called the ARRIVE guideline.

The study shows the power of being able to mine the full text of research papers, says Malcolm Macleod, a stroke researcher and specialist in trial design at the University of Edinburgh, UK. But he notes that the fully open papers that can be mined may not be representative of the entire scientific literature: scientists who want to make their papers open may be “among the more enlightened”, he suggests.

Previous research has consistently suggested that researchers tend to use male mice in studies — but the eLife paper reports that, where the sex of animals is recorded, the mice are more likely to be female. That surprises Caroline Zeiss, a veterinary neuropathologist at Yale University in New Haven, Connecticut. The discrepancy might result from biases in the specific fields of research that the eLife paper examined, she says.

The sex bias varied according to the type of biomedical study, and even between studies of the same disease. Diabetes research, for example, was found to be male-biased overall — but studies on the immunology of the condition tended to use female mice. There was no correlation between reporting standards and the impact factor of the journal in which the studies were published, the researchers found.

Nature doi:10.1038/nature.2016.19500  Nature  Original web page at Nature


Thousands of goats and rabbits vanish from major biotech lab

Santa Cruz Biotechnology has used goats to make antibodies for research. In July 2015, the major antibody provider Santa Cruz Biotechnology owned 2,471 rabbits and 3,202 goats. Now the animals have vanished, according to a recent federal inspection report from the US Department of Agriculture (USDA).

The company, which is headquartered in Dallas, Texas, is one of the world’s largest providers of antibodies — extracting them from animals such as goats and rabbits by injecting the animals with proteins to stimulate antibody production. Biomedical researchers can then use these antibodies to detect and label the same protein in cell or tissue samples.

But Santa Cruz Biotech is also the subject of three animal-welfare complaints filed by the USDA after its inspectors found evidence that the firm mistreated goats at its facility in California. Discovery of goat facility adds to antibody provider’s woes.

Santa Cruz Biotech has contested the complaints, prompting a hearing in August before a USDA administrative law judge in Washington DC. Four days into the hearing, both parties asked to suspend the proceedings in order to negotiate a settlement. But those negotiations fell through in September.

The USDA hearing is set to resume on 5 April. If Santa Cruz Biotech is found to have violated the US Animal Welfare Act, it could be fined or lose its licence to keep animals for commercial use. The USDA says that the company could face a maximum fine of US$10,000 per violation for each day that a given violation persists. The agency has reported 31 alleged violations by the company.

In the meantime, the company seems to have done away with its entire animal inventory. When the USDA inspected the firm’s California facility on 12 January, it found no animal-welfare violations — and no animals.

The fate of the goats and rabbits is unclear. The company did not respond to questions about the matter, and David Schaefer, director of public relations for the law firm Covington & Burling in Washington DC, which is representing Santa Cruz Biotechnology, declined to comment on the animals’ fate.

As research animals, the goats and rabbits could not be sold for meat, although they could be sold to another business or research entity. Cathy Liss, president of the Animal Welfare Institute, an advocacy group in Washington DC, suspects that the animals were killed. She says that it is unlikely that such a large number of animals bred for such a specific purpose would find a buyer.

David Favre, an expert in animal-welfare law at Michigan State University in East Lansing, argues that the disappearance of the animals should not sway the legal proceedings over whether the company violated the Animal Welfare Act, because the USDA complaints refer to past events. But he says that the court could still take this latest development into account.

Favre is also unhappy with the USDA’s decision to delay action on the complaints. The department’s “whole attitude is ‘we’ll give them time to fix it rather than impose punishment’,” Favre says. “There’s no excuse for a company that size not to be able to comply with the Animal Welfare Act.”

The USDA’s complaints about Santa Cruz Biotech’s operations detail alleged violations of the law since 2007 — among them accounts of goats with untreated coyote bites and massive tumours, and rabbits being housed in cruel conditions. A sick goat died in front of a USDA inspector during one 2012 inspection.

The company also angered the USDA by keeping 841 goats in an unreported facility, which concealed the animals and their living conditions from regulatory scrutiny until 2012. During the August 2015 USDA hearing, former Santa Cruz Biotech veterinarian Robin Parker testified that company president John Stephenson had decided not to tell the USDA inspectors about the second site. She said that she had been told that the regulators tended to “nitpick” the company’s operations.

After leaving Santa Cruz Biotech in 2012, Parker notified the USDA about the unreported facility. When inspectors visited the site, they found animals housed in poor living conditions.

“So much of what was cited involves great animal suffering,” says Liss, whose advocacy group has campaigned for the USDA to take action against the company. “We are hoping that USDA holds out and if there is going to be a settlement, that it includes them permanently losing their licence as a dealer.”

Favre says that the USDA and Santa Cruz Biotech may have already reached some sort of agreement that requires the company to cease keeping animals for commercial use in advance of the 5 April court hearing; that could explain why the company no longer keeps goats or rabbits. But the USDA administrative-law court says that the company has not submitted any exhibits related to animal removal for the upcoming hearing, although its attorneys could petition to add more evidence before the hearing begins.

Were Santa Cruz Biotech to stop producing antibodies, there would still be hundreds of other providers supplying the materials to researchers, says David Rimm, a pathologist at Yale University in New Haven, Connecticut. “I think people might notice, but I don’t think it would impact science at all,” says Rimm.

He speculates that the company may be moving towards making ‘recombinant’ antibodies. These are produced in cells cultured in the lab that are genetically engineered to make specific antibodies. Many companies have switched to this method instead of producing antibodies using animals, Rimm says.

Alice Ra’anan, director of government relations and science policy at the American Physiological Society in Bethesda, Maryland, hopes that the allegations against Santa Cruz Biotechnology will prompt a cultural change among those involved in animal research. The ideal outcome, she says, is “that people will give animal welfare the level of consideration it deserves”.

Nature doi:10.1038/nature.2016.19411  Nature  Original web page at Nature


Proposal to ban imported monkeys catches scientists off guard

Australian bill provokes rush of protests ahead of parliamentary deadline.

Nicholas Price works to understand the brain’s fundamental functions, with a view towards developing a bionic eye. The neuroscientist uses marmosets and macaques in his experiments at Monash University’s Biomedicine Discovery Institute in Melbourne. In late January, he was shocked to discover a bill before the Australian Parliament that calls for a ban on the import of non-human primates for medical research.

Australia’s three main breeding colonies of research primates consist of several hundred macaques, marmosets and baboons. Regular imports of the animals are vital to maintain the genetic diversity of these colonies, says Price.

Senator Lee Rhiannon, a member of the Greens party, introduced the bill on 17 September last year as an amendment to Australia’s federal Environment Protection and Biodiversity Conservation Act. But because the Senate committee that deals with this piece of legislation is not usually of interest to those in the medical research community, the amendment almost slipped under the community’s radar, says Price. By the time he heard about the proposed ban, from another researcher, the window for public comment was days away from closing, though it was later extended.

As soon as they found out, Price and his Monash colleagues James Bourne and Marcello Rosa began e-mailing researchers around the world. Several institutions rushed to submit statements opposing the bill, including the Federation of European Neuroscience Societies (FENS), the Society for Neuroscience, headquartered in Washington DC, and the Basel Declaration Society, which promotes the open, transparent and ethical use of animals in research. Australia’s National Health and Medical Research Council and the Australasian Neuroscience Society also sent statements of opposition to the Senate committee.

Animal research, including that on non-human primates, “continues to be the basis for medical advances that have extended our life expectancy”, says the FENS statement, submitted on 4 February. The Society for Neuroscience’s statement, dated the same day, says that the proposed legislation would lead to “the loss of critical research resources that will be devastating for Australian science”. The committee is now in the process of considering the bill, and will report on it on 1 March, as a prelude to an eventual Senate vote.

Rhiannon, who trained as a zoologist, told Nature that the Greens are not calling for a ban on non-human primates being used in research. “There’s certainly a live debate but it’s not the party position,” she says. She calls the bill a “modest” way to improve the welfare of research animals, and, when she launched the bill last year, spoke at length about the “the cruelty during their capture, confinement and transportation”.

Price and Bourne say that cutting off access to the genetic diversity required to maintain the monkey colonies would eventually spell the end of Australian research on non-human primates. “After a while you’ll get genetic inbreeding,” says Price, “and that can lead to exacerbation of health problems that you may not be able to see until you’re a few generations down the track.” He regards non-human primate research as one of the country’s major sources of scientific innovation.

In introducing the bill, Rhiannon also said that it would ensure that Australia “does not participate in the unethical trade of wild-caught primates for use in experimentation for the research industry”. But Australian regulations already prohibit the use of wild-caught primates in medical research, says Bourne, who is chairman of the National Non-human Primate Breeding and Research Facility Board. “They have to be certified from a registered and accredited breeding facility in another country.”

Rhiannon introduced a similar bill in 2012 that never made it to a Senate vote before Parliament was dissolved ahead of a federal election. The latest bill may face a similar challenge with an election required by January 2017 at the latest. But Rhiannon says that even if it doesn’t get passed into legislation anytime soon, the bill is a way to highlight that there is a problem with the use of non-human primates, without going so far as to call for banning such research entirely.

Price says that the events have convinced him of a need to be more public about the importance of his work. “If we can demonstrate the value and outcomes, especially in medical research, then we feel that the majority of the public would be very supportive.”

Nature doi:10.1038/nature.2016.19419  Nature  Original web page at Nature


Zika researchers release real-time data on viral infection study in monkeys

Researchers in the United States who have infected monkeys with Zika virus made their first data public last week.  But instead  of publishing them in a journal, they have released them online for anyone to view- and are updating their results day by day.

The team is posting raw data on the amount of virus detected in the blood, saliva and urine of three Indian rhesus macaques, which they injected with Zika on 15 February. “This is the first time Researchers in the United States who have infected monkeys with Zika virus made their first data public last week. But instead of publishing them in a journal, they have released them online for anyone to view — and are updating their results day by day.

that our group has made data available in real time,” says David O’Connor, a virologist at the University of Wisconsin–Madison and a leader of the project, whose scientists have dubbed themselves ZEST (the Zika experimental-science team). He hopes that releasing the data will help to speed up research into the nature of the virus that has spread across the Americas.

Although a few teams have begun to share genomic data online during disease outbreaks, instant open-data release remains the exception rather than the rule, particularly in clinical research. O’Connor says that he was inspired by researchers during the Ebola epidemic who rapidly published genomic-sequencing data online and encouraged others to re-analyse them. At the time, O’Connor’s group downloaded raw data shared by a team led by Pardis Sabeti, a computational geneticist at the Broad Institute and Harvard University in Cambridge, Massachusetts; it immediately helped to advance their own Ebola research, he says, and led to a collaboration with Sabeti’s group.

“O’Connor’s team is to be lauded for their efforts to make their Zika virus data publicly available as soon as possible,” says Nathan Yozwiak, a senior scientist in Sabeti’s laboratory. “Distributing up-to-date information — in this case, animal model data — as widely and openly as possible is critical during emergencies such as Zika, where relatively little is known about its pathogenesis, yet public concerns and attention are so high.”

“This is exemplary for research,” agrees Koen Van Rompay, a specialist in non-human primate models of HIV infection at the California National Primate Research Center at the University of California, Davis. Van Rompay is part of a consortium that plans to inject pregnant macaques with Zika. He says that his team will also share data openly in real time.

David O’Connor, whose ‘ZEST’ team at the University of Wisconsin-Madison are releasing Zika data in real time. O’Connor’s team seems to be the first to have detailed information from macaques infected with Zika virus, so the rapid data release will enable other researchers planning similar experiments to take the work into account, saving time and resources, Van Rompay adds. “This is such an urgent public-health emergency that this should not be a race of scientists competing against each other. We’re in a race against the Zika virus, a race against time,” he says.

Like other researchers, the ZEST team wants to understand when a developing fetus might be at risk of birth defects from Zika. Typically, the virus gives rise to no or mild symptoms — but scientists are urgently working to estimate the strength of any association between Zika infection and an apparent rise in the number of babies born with microcephaly (abnormally small heads and brains) in northeastern Brazil.

If the virus behaves the same way in macaques that it does in humans, O’Connor says, researchers will be able to glean information by infecting monkeys with varying doses of Zika — data that would be impossible to gain rapidly or ethically from people. Scientists could repeatedly sample amniotic fluid in pregnant macaques, for example, to determine whether, and how quickly, the virus can infect a fetus.

The team is starting with male monkeys to get information on how the virus behaves in macaques and determine which dose would be most suitable for later experiments. They have already shown that Zika can infect macaques and that it is detectable not only in blood, but also in cerebrospinal fluid and urine. They will follow up their work with experiments in macaques at different stages of pregnancy, checking for the virus’s presence in a wide range of tissues and organs.

Even if a link to birth defects is proven, it may still be that very few Zika infections during pregnancy lead to microcephaly, O’Connor says. But he thinks that even with a small number of animals, the team can assess important questions such as whether fetuses become infected with Zika virus and whether they develop abnormalities as a result.

Pregnant rhesus macaques have been used in past to study congenital birth defects, O’Connor adds. Research on the effects of cytomegalovirus or Listeria, for example, have revealed that the diseases produce similar effects in macaques and in humans. The team also hopes to carry out Zika studies in marmosets, which are native to northeastern Brazil and smaller than macaques, making them easier to work with in the lab. If it is possible for the virus to infect marmosets, this might also suggest that the animals are involved in Zika transmission in Brazil, O’Connor says.

It was easy for the ZEST members to make their online lab notebook open to all, O’Connor says. The team uses the biomedical-research collaboration system LabKey Server, as does the Wisconsin National Primate Research Center in Madison, which is where many of the ZEST collaborators work and which (along with the US National Institutes of Health) is supporting the research. Researchers created a study to store and update their data, and simply had to switch permissions to allow anyone to view it. Meanwhile, regulatory agencies at the University of Wisconsin–Madison understood that the work was time-sensitive and expedited approvals for animal care and biosafety (without reducing scrutiny, O’Connor adds).

On 10 February, dozens of major funders, government agencies and journals released a statement supporting open-data sharing — even before publication — during public-health emergencies such as the Zika and Ebola epidemic. “In the context of a public-health emergency of international concern, there is an imperative on all parties to make any information available that might have value in combatting the crisis,” it concluded.

Jeremy Farrar, director of the Wellcome Trust in London – one of the research funders that signed the statement – says he welcomes the “increasing commitment” of scientists to sharing information during public health emergencies. “The world is changing and all of us involved need to encourage, facilitate and give thanks and credit to these teams and the approach they are taking,” he says.

“I hope that even those who disagree in principle with animals in research realize that making data available publicly works towards a common goal,” O’Connor adds. “Fewer animals will be used in research if groups know what others are doing, and the information that is gained from each animal is maximized.”

Nature doi:10.1038/nature.2016.19438  Nature  Original web page at Nature


Proposal to ban imported monkeys catches scientists off guard

Nicholas Price works to understand the brain’s fundamental functions, with a view towards developing a bionic eye. The neuroscientist uses marmosets and macaques in his experiments at Monash University’s Biomedicine Discovery Institute in Melbourne. In late January, he was shocked to discover a bill before the Australian Parliament that calls for a ban on the import of non-human primates for medical research.

Australia’s three main breeding colonies of research primates consist of several hundred macaques, marmosets and baboons. Regular imports of the animals are vital to maintain the genetic diversity of these colonies, says Price.

Senator Lee Rhiannon, a member of the Greens party, introduced the bill on 17 September last year as an amendment to Australia’s federal Environment Protection and Biodiversity Conservation Act. But because the Senate committee that deals with this piece of legislation is not usually of interest to those in the medical research community, the amendment almost slipped under the community’s radar, says Price. By the time he heard about the proposed ban, from another researcher, the window for public comment was days away from closing, though it was later extended.

As soon as they found out, Price and his Monash colleagues James Bourne and Marcello Rosa began e-mailing researchers around the world. Several institutions rushed to submit statements opposing the bill, including the Federation of European Neuroscience Societies (FENS), the Society for Neuroscience, headquartered in Washington DC, and the Basel Declaration Society, which promotes the open, transparent and ethical use of animals in research. Australia’s National Health and Medical Research Council and the Australasian Neuroscience Society also sent statements of opposition to the Senate committee.

Animal research, including that on non-human primates, “continues to be the basis for medical advances that have extended our life expectancy”, says the FENS statement, submitted on 4 February. The Society for Neuroscience’s statement, dated the same day, says that the proposed legislation would lead to “the loss of critical research resources that will be devastating for Australian science”. The committee is now in the process of considering the bill, and will report on it on 1 March, as a prelude to an eventual Senate vote.

Rhiannon, who trained as a zoologist, told Nature that the Greens are not calling for a ban on non-human primates being used in research. “There’s certainly a live debate but it’s not the party position,” she says. She calls the bill a “modest” way to improve the welfare of research animals, and, when she launched the bill last year, spoke at length about the “the cruelty during their capture, confinement and transportation”.

Price and Bourne say that cutting off access to the genetic diversity required to maintain the monkey colonies would eventually spell the end of Australian research on non-human primates. “After a while you’ll get genetic inbreeding,” says Price, “and that can lead to exacerbation of health problems that you may not be able to see until you’re a few generations down the track.” He regards non-human primate research as one of the country’s major sources of scientific innovation.

In introducing the bill, Rhiannon also said that it would ensure that Australia “does not participate in the unethical trade of wild-caught primates for use in experimentation for the research industry”. But Australian regulations already prohibit the use of wild-caught primates in medical research, says Bourne, who is chairman of the National Non-human Primate Breeding and Research Facility Board. “They have to be certified from a registered and accredited breeding facility in another country.”

Rhiannon introduced a similar bill in 2012 that never made it to a Senate vote before Parliament was dissolved ahead of a federal election. The latest bill may face a similar challenge with an election required by January 2017 at the latest. But Rhiannon says that even if it doesn’t get passed into legislation anytime soon, the bill is a way to highlight that there is a problem with the use of non-human primates, without going so far as to call for banning such research entirely.

Price says that the events have convinced him of a need to be more public about the importance of his work. “If we can demonstrate the value and outcomes, especially in medical research, then we feel that the majority of the public would be very supportive.”

Nature 530, 394 (25 February 2016) doi:10.1038/nature.2016.19419  Nature Original web page at Nature


Team suppresses oxidative stress, neuronal death associated with Alzheimer’s disease

The brain is an enormous network of communication, containing over 100 billion nerve cells, or neurons, with branches that connect at more than 100 trillion points. They are constantly sending signals through a vast neuron forest that forms memories, thoughts and feelings; these patterns of activity form the essence of each person. Alzheimer’s disease (AD) disrupts both the way electrical charges travel within cells and the activity of neurotransmitters. An AD brain has fewer nerve cells and synapses than a healthy brain; plaques and abnormal clusters of protein fragments accumulate between nerve cells. The major pathological indicators of AD are the accumulation of amyloid beta plaques and neurofibrillary tangles in the brain. The pathways in our neuron forest are systemically attacked and destroyed by amyloid beta (Aβ): a solitary molecule that evolves into plaque clusters, which block cell-to-cell signalling at synapses. They may also activate immune system cells that result in inflammation and destroy damaged cells.

In a healthy brain, orderly parallel strands, akin to railroad tracks, permit nutrients and essential proteins to move between cells. The protein tau helps these tracks remain intact and functioning. In an AD affected brain tau breaks down, collapses and forms tangles that prevent transmission along the tracks. The tracks fall apart and disintegrate. Essential proteins, including nutrients, can no longer reach brain cells, which eventually die. The plaques and tangles described above are currently the leading working theory explaining the cell death and tissue loss found in an AD brain, though the theory is yet to be irrefutably confirmed. The effects of AD on the brain, however, are well known: brain cells slowly disintegrate, the disease progressively invades different parts of the brain, creating unique changes that signal the various stages of Alzheimer’s. Short term memory loss, logical thoughts and emotions are all obstructed, fundamentally altering and ultimately eradicating an affected individual’s personality. Over time, Alzheimer’s leads to nerve cell death and dramatic shrinking of the brain, which affects nearly all of its functions.

The scientific team from the Center for Nanoparticle Research within the Institute for Basic Science (IBS) has developed a novel mitochondria-targeting ceria nanoparticle that can effectively impede the process of neuronal cell death, in collaboration with the research group of Seoul National University led by professor Inhee Mook. Cells in our brain are powered by mitochondria; tiny power plants within cells that produce a body’s essential energy, which is required for each cell to function. Reactive oxygen species (ROS) are formed as a natural by-product of normal metabolism of oxygen. Abnormal generations of ROS, resulting from mitochondrial dysfunction, can lead to neuronal cell death. Additionally, Aβ-induced mitochondrial dysfunction also has been known to be a possible cause of AD through abnormal production of ROS. Ceria nanoparticles function as known to function as strong and recyclable ROS scavengers, eliminating abnormal ROS, by shuttling between Ce3+ and Ce4+ oxidation states.

The research team, under the direction of the IBS Center’s director Taeghwan Hyeon, synthesized a ceria nanoparticle, mitochondria-specific antioxidant and investigated the effect of the new therapeutic agent in suppressing the pathogenesis of AD using an in vivo mouse model. The team introduced the powerful ceria nanoparticles (CeO2 NP) to mitochondria by using small, mitochondria-targeting materials (triphenylphosphonium-conjugating) and recorded quite remarkable results in a transgenic AD mouse model. Two months after the mouse was injected, positive cells were quantified. According to the results, published online in ACS Nano on February 11, the CeO2 NPs localised to mitochondria had effectively suppressed neuronal death in the mouse model, demonstrating that the administration of mitochondria-targeting ceria NPs significantly restored neuronal viability of the AD-affected mouse. Since the accumulation of Aβ did not differ significantly between the brains of the affected and non-treated mouse, it is concluded that the mitochondria-targeting ceria NPs ameliorate the neuronal damage of the test subject in an indirect way, independent of the Aβ accumulation. The team’s paper stressed that the data “indicated that the mitochondria-targeting ceria NPs are a potential therapeutic candidate for treating mitochondrial oxidative-stress-induced damage in AD.” Director Hyeon said, “This study is quite remarkable in that the collaborative research between nano science and biomedical science has led to a potent therapeutic agent against reactive oxygen species in the mitochondria, which is deemed to be one of major culprits in a number of diseases.   Science Daily Original web page at Science Daily


Brain boost: Research to improve memory through electricity?

In a breakthrough study that could improve how people learn and retain information, researchers at the Catholic University Medical School in Rome significantly boosted the memory and mental performance of laboratory mice through electrical stimulation.

The study, sponsored by the Office of Naval Research (ONR) Global, involved the use of Transcranial Direct Current Stimulation, or tDCS, on the mice. A noninvasive technique for brain stimulation, tDCS is applied using two small electrodes placed on the scalp, delivering short bursts of extremely low-intensity electrical currents.

“In addition to potentially enhancing task performance for Sailors and Marines,” said ONR Global Commanding Officer Capt. Clark Troyer, “understanding how this technique works biochemically may lead to advances in the treatment of conditions like post-traumatic stress disorder, depression and anxiety–which affect learning and memory in otherwise healthy individuals.”

The implications of this research also have great potential to strengthen learning and memory in both healthy people and those with cognitive deficits such as Alzheimer’s.

“We already have promising results in animal models of Alzheimer’s disease,” said Dr. Claudio Grassi, who leads the research team. “In the near future, we will continue this research and extend analyses of tDCS to other brain areas and functions.”

After exposing the mice to single 20-minute tDCS sessions, the researchers saw signs of improved memory and brain plasticity (the ability to form new connections between neurons when learning new information), which lasted at least a week. This intellectual boost was demonstrated by the enhanced performance of the mice during tests requiring them to navigate a water maze and distinguish between known and unknown objects.

Using data gathered from the sessions, Grassi’s team discovered increased synaptic plasticity in the hippocampus, a region of the brain critical to memory processing and storage.

Although tDCS has been used for years to treat patients suffering from conditions such as stroke, depression and bipolar disorder, there are few studies supporting a direct link between tDCS and improved plasticity–making Grassi’s work unique.

More important, the researchers identified the actual molecular trigger behind the bolstered memory and plasticity–increased production of BDNF, a protein essential to brain growth. BDNF, which stands for “brain-derived neurotrophic factor,” is synthesized naturally by neurons and is crucial to neuronal development and specialization.

“While the technique and behavioral effects of tDCS are not new,” said ONR Global Associate Director Dr. Monique Beaudoin, “Dr. Grassi’s work is the first to describe BDNF as a mechanism for the behavioral changes that occur after tDCS treatment. This is an exciting and growing research area of great interest to ONR.”

Beaudoin said tDCS treatment could one day benefit Sailors and Marines, from helping them learn faster and more effectively to easing the effects of post-traumatic stress disorder.

“Our warfighters face tremendous challenges that are both physically and cognitively taxing,” she said. “They perform their duties in stressful environments where there are often suddenly and randomly varying levels of environmental stimulation, disrupted sleep cycles and a constant need to stay alert and vigilant.

“We want to explore all interventions that could help them stay healthy and perform optimally in these environments–especially when treatments are potentially noninvasive, effective and lead to long-lasting changes.”  Science Daily Original web page at Science Daily


Web tool aims to reduce flaws in animal studies

A free online tool that visualizes the design of animal experiments and gives critical feedback could save scientists from embarking on poorly designed research, the software’s developers hope.

Over the past few years, researchers have picked out numerous flaws in the design and reporting of published animal experiments, which, they warn, could lead to bias. In response, hundreds of journals have agreed to voluntary guidelines for reporting animal studies: checklists of best practice, such as what statistical calculations to use to ward off error.

But these lists kick in after scientists submit a paper, says Nathalie Percie du Sert, who specializes in experimental design at the National Centre for the Replacement, Refinement and Reduction of Animals in Research (NC3Rs) in London. “When you get to the reporting stage, that’s a bit too late,” she says. “We want researchers to think about these issues at the design stage.”

Percie du Sert’s solution is a programme called the Experimental Design Assistant (EDA), which launched in October 2015. She hopes that it will help to improve the quality of animal research and perhaps even become an integral part of the conduct of animal studies.

The EDA allows scientists to create a visual representation of an experiment by laying out its key elements — hypothesis, experimental method and planned analysis — in logically connected, coloured boxes. The software then uses a built-in set of rules to spot potential problems, and suggests refinements. These may be simple — the researcher hasn’t specified how to randomize animals to the control or treatment arm — or more complex: there are potential confounding variables in the control and trial arms. The tool can also assist scientists with calculating the sample size needed to ensure a statistically robust result, or with randomization.

There’s nothing fundamentally new in the EDA, says Percie du Sert. It builds on existing knowledge of good experimental design. But it can aid scientists who have little training in the area, she says, and teach them design choices.

Since the EDA’s launch, around 400 accounts have been created to use it, producing between 50 and 100 experimental diagrams in total each month, says Percie du Sert. She does not have access to detailed information about its users; the sensitivities around animal research and the need to protect researchers mean that data on who is using it, and how, are secured.

The Wellcome Trust’s Sanger Institute, a genome-research centre in Cambridge, UK, is rolling out an internal training programme that includes lessons on design and use of the EDA; the agency is encouraging staff to use the software to present experiments to ethical-review committees, says Natasha Karp, a biostatistician at the institute. Karp took part in the working group that oversaw the tool’s development, and says that she uses it to visualize the experiments of the biologists whom she supports.

The EDA is not the only software that aims to improve research quality and reproducibility. Other tools check manuscripts before publication for issues such as errors in formatting or omission of P values. These include Penelope, a paid-for service aimed at journal publishers; another tool called WebCONSORT (which is not yet freely available) is being tested as a way to improve reporting of clinical trials. Protocol Navigator, a free web application created by scientists at Cardiff University, UK, also produces visual experiment maps that can be shared.

But the EDA specifically targets animal research, and as such, is unique in its ability to give a rapid overview of the design and analysis of animal experiments, says Karp. “There isn’t anything else quite like this system.”

Percie du Sert hopes that a visual representation of experiments could become common practice, used in research papers or lab meeting presentations. Eventually, the EDA might even produce time-stamped versions to prove that an experiment was conducted and analysed as designed, she adds, rather than being the product of a scientist searching for meaning in data after the fact — a frowned-upon practice sometimes called HARKing (‘hypothesizing after the results are known’).

The online tool can seem a little complicated, says Jeffrey Mogil, who studies pain at McGill University in Montreal, Canada. “But I actually think that people might get a big kick out of using this,” he says. “It looks like a cool way to break in new grad students or teach the scientific method to undergrads.”

Nature 531, 128 (03 March 2016) doi:10.1038/531128a  Nature  Original web page at Nature


Legal tussle delays launch of huge toxicity database

A database of the toxicity of nearly 10,000 chemicals might reduce the need for animal safety-testing. A giant database on the health risks of nearly 10,000 chemicals will make it easier to predict the toxicity of tens of thousands of consumer products for which no data exist, say researchers. But a legal disagreement means they haven’t yet been able to make the database public, as they had hoped to do.

“This has the potential to save millions of animals and reduce testing costs by hundreds of millions,” says Thomas Hartung, a toxicologist at Johns Hopkins University in Baltimore, Maryland, who led the team that created the database. He describes his work in papers published on 11 February in Alternatives to Animal Experiments.

The index is built from a mountain of safety data collected over the past decade by the European Chemicals Agency (ECHA) in Helsinki, under a 2006 law known as REACH (registration, evaluation, authorization and restriction of chemicals). The information is already public, but not held in an easy-to-analyse format — so Hartung’s team developed software that extracted it and converted it into a searchable database.

But the ECHA says that it has exclusive rights to the information, and that Hartung did not gain the specific permission he needed from the agency in order to duplicate it. For the moment, Hartung has agreed to hold off making his team’s database public.

With the database, Hartung hopes that companies and regulators will find it easier to infer the toxic effects of untested substances by comparing them with structurally or biologically similar chemicals with known effects — a method called read-across.

Thomas Hartung has long promoted alternatives to animal safety-tests. That concept is already popular among chemical companies that seek alternatives to safety tests on animals, says Markus Wahl, a regulatory toxicologist at chemicals producer BASF in Ludwigshafen, Germany. The database will provide “helpful supporting evidence”, he says, but he adds that European regulators accept the results of read-across (in place of animal-safety tests) only occasionally. Hartung, a long-time promoter of alternatives to animal-based safety testing, hopes to change that.

A spokesperson for the ECHA says that read-across is a “good approach” for checking relatively simple concerns such as harmful effects to skin and eyes — but that it “proves to be challenging” for complex issues such as the effects of repeated exposure to chemicals. “Companies quite often fail to substantiate why the read-across is scientifically justified,” the agency says.

Hartung argues that the database will strengthen the scientific case for read-across, because the certainty of the comparison “increases with the extent and quality of data,” he says. The data might also prove very useful to US regulators, as the country’s lawmakers attempt to tighten legislation governing the safe use of chemicals. Currently, chemicals can go on the US market with little regulatory scrutiny.

Analysis of the database has shown that some animal tests are woefully irreproducible, Hartung adds. The Draize eye test, for example, in which chemicals are applied to rabbits’ eyes to check for harmful effects, is a “big lottery”, he says. The test, in use for decades, has been widely criticized for producing inconsistent results and raising animal-welfare concerns, but the Johns Hopkins team analysed 9,700 Draize results from the ECHA data to quantify the problem. The results, published alongside the description of the database, suggest that substances found to cause serious irreversible eye damage in one test have a 10% chance of being found harmless in a subsequent test. By contrast, substances found to cause reversible irritation to rabbits’ eyes have nearly a 60% chance of being found harmless in a later test.

Hartung says that numerous researchers have expressed interest in mining the database, including the US Environmental Protection Agency, the US National Institutes of Health and some academic and industry groups.

But the database has already run into legal trouble with the ECHA. Information on the ECHA’s website is “proprietary”, the agency says, and “may be subject to intellectual property rights or copyright” belonging to the chemical companies. “This is not a parochial or bureaucratic requirement,” a spokesperson added: the ECHA is “keen” to see the data being used, but says that it also needs to protect the rights of the companies that own the data.

Hartung counters that the research “makes use only of the publicly available data”, and that the agency shouldn’t prohibit their use for academic enquiry. He has agreed to delay making the database public until he gets the specific permission he needs from the ECHA, but it is not clear how long that will take.

Nature doi:10.1038/nature.2016.19365  Nature  Original web page at Nature


A mouse’s house may ruin experiments

Mice are sensitive to minor changes in food, bedding and light exposure.

It’s no secret that therapies that look promising in mice rarely work in people. But too often, experimental treatments that succeed in one mouse population do not even work in other mice, suggesting that many rodent studies may be flawed from the start.

“We say mice are simpler, but I think the problem is deeper than that,” says Caroline Zeiss, a veterinary neuropathologist at Yale University in New Haven, Connecticut. Researchers rarely report on subtle environmental factors such as their mice’s food, bedding or exposure to light; as a result, conditions vary widely across labs despite an enormous body of research showing that these factors can significantly affect the animals’ biology.

“It’s sort of surprising how many people are surprised by the extent of the variation” between mice that receive different care, says Cory Brayton, a pathologist at Johns Hopkins University in Baltimore, Maryland. At a meeting on mouse models at the Wellcome Genome Campus in Hinxton, UK, on 9–11 February, she and others explored the many biological factors that prevent mouse studies from being reproduced.

Christopher Colwell, a neuroscientist at the University of California, Los Angeles, has first-hand experience with these issues. He and a colleague studied autism in the same genetically modified mouse line, but obtained different results on the same behaviour tests. Eventually they worked out why: Colwell, who studies circadian rhythms, keeps his mice dark in the daytime to trick their body clocks into thinking day is night, so that the nocturnal animals are more alert when tested during the day. His colleague does not.

Colwell notes that disregarding mouse circadian rhythms could bias many behaviour experiments. Most humans would not perform well on social and cognitive tests either if made to do them in the middle of the night, he adds.

Nutrition can also determine whether a mouse study succeeds or fails, yet Brayton says that many researchers cannot even say where their animals’ feed comes from. Some mouse foods contain oestrogens and endocrine-disrupting chemicals that can affect research on cancer, among other diseases. And the high-fat, high-sugar food used in obesity studies goes rancid quickly; when it does, mice may stop eating and lose weight without researchers realizing why.

Food choices can also alter a mouse’s gut microbiome. Catherine Hagan Gillespie, a veterinary pathologist at the Jackson Laboratory in Sacramento, California, has found that species of bacteria in the gut vary widely between mice from different vendors. In another, unpublished, study, she found that mice with different assortments of gut bacteria showed different anxiety levels in behavioural tests.

But few behavioural scientists think about running microbiology assessments, says Hagan Gillespie. Even when they do, the extra work involved can increase the complexity and cost of the study. Yet the mouse microbiome is sensitive to a wide array of factors, such as air quality, maternal stress and immune function.

Differences in the gut microbiome may explain why mice with the same genetic mutation can have different characteristics, or phenotypes, says George Weinstock, associate director for microbial genomics at the Jackson Lab’s site in Farmington, Connecticut. Jackson Lab, which breeds and supplies mice for use in studies around the world, tightly controls factors such as the type and quantity of food and the pH of water that animals receive. Even so, it finds differences between the mice at its three sites. Weinstock says that the company has begun researching ways to standardize its customers’ experiments by providing special food and care instructions for the mice that it provides.

But even when improved mice and food are available, some researchers resist using them out of concern that it will affect their results, says Graham Tobin, former technical director of the mouse-diet vendor Teklad in Alconbury, UK. Yet he argues that standardizing results across labs is worth this inconvenience to individual scientists. Tobin also notes that researchers rarely resist adopting other new technologies — such as improved DNA sequencers — that can throw older data into question.

Zeiss says that the competitive nature of science might increase researchers’ resistance to changing how they consider animals in research design. If scientists have to treat their animals at the right point in the experiment, analyse both clinical and biomarker changes, include old mice and both sexes to ensure that results are representative of broad populations, and control environmental variables, each experiment will take much longer and they’re probably not going to be able to publish as much, she says.

The US National Institutes of Health (NIH) has taken steps to address some of these problems. Some of its institutes require certain animal trials to be replicated before a therapy can move into clinical trials, but the NIH says that it has no plans to require this agency-wide. And in 2014, the NIH began requiring researchers to include female animals in studies, and started to give out supplementary grants to researchers who complained about the cost. But the agency has not issued any specific grants or supplements to study other confounding factors.

That is disappointing to those who would like to see researchers control — or at least report — factors such as the strain of mice used and the type of environment they are raised in. This would allow researchers to perform metanalyses of published literature that could identify any confounding factors. “The information and the wisdom is out there,” says Zeiss, “but studies get funded without necessarily a lot of attention to that.”

Nature doi:10.1038/nature.2016.19335  Nature  Original web page at Nature


* Researchers’ preclinical trial upends conventional wisdom about responses to fear

In a new study researchers found that female rats often respond to fear by ‘darting.’ For more than a century scientists have recognized ‘freezing’ as the natural fear response. But in a new study found that female rats often respond to fear by ‘darting.’ The findings not only raise questions about the veracity of previous studies that rely on freezing to indicate fear, but could also lead to better treatments for post-traumatic stress disorder.

Fear. You’ve been there: Your heart races, even jumps to your throat. Your hands grow clammy and your stomach churns. Your mind goes blank. Rats have been there, too. We don’t know their feelings, of course, but we do know their response: They freeze in their tracks. Or at least that’s been the consensus among scientists since 1899, when experimental psychologist Willard Stanton Small first noted the behavior.

But now new research led by Rebecca Shansky, assistant professor of psychology at Northeastern University, upends that conventional wisdom.

In a study recently published in the online journal eLife, Shansky’s team found that female rats often respond to fear by “darting.” “They start running around like crazy,” Shansky says. “It looks like they’re trying to escape.”

In addition, the darting rats were more successful at integrating a process that suppressed the fear response, says Shansky, exhibiting a “cognitive flexibility” that the freezers lacked.

The findings not only raise questions about the veracity of previous studies that rely on freezing to indicate fear. They could also lead to better treatments for post-traumatic stress disorder, a condition that, in the U.S. alone, affects about 8 mil-lion adults during a given year, according to the National Center for PTSD of the U.S. Department of Veterans Affairs.

“If we can harness whatever is going on when an animal becomes a darter,” says Shansky, “we could try to apply that to treatments.”

Shansky had not set out to challenge a century-old assumption. Rather, she stumbled across the findings while performing a common behavioral test called “fear conditioning” in an effort to see how individual males and females differed in their fear responses, and to explore what brain changes related to those differences.

The test involved teaching the animals to associate a tone with a foot shock, and then–with a video camera connected to a computer–measuring the duration of their reaction as the training proceeded. “Animals who exhibit low levels of freezing would traditionally be interpreted as either not learning or naturally fear-less,” says Shansky.

Because computers may mistake sleeping for freezing, graduate student Tina Gruene, PhD, watched the videos afterward as a backup check. What she saw shocked her: Scores of the female rats not only didn’t freeze at the sound of the tone; they darted hither and yon, as if looking for an exit.

What did that mean? The study had a large number of rats–120 as opposed to the standard 20–so Shansky set out to quantify the behavior. “We wanted to see if this was something real,” she says.

The researchers fed the videos into a behavioral analysis program that tracks motion to monitor the velocity of the animals’ movement. Their plotted graphs confirmed their hunch: Darting was not a sign of fearlessness or an inability to learn. It was just as much a learned response as freezing.

“The learning curve for darting was the same as the learning curve for freezing,” says Shansky, pointing to graphics in the paper. “But we saw it almost exclusively in the females–more than 40 percent of them.”

The findings go beyond clarifying differences in fear behavior among male and female rats. They also point to possibilities for better treatments for people with PTSD.

Following the fear conditioning, the researchers used a process called “extinction” to suppress the rats’ fear response: By playing the tone repeatedly without the shock, a “good” memory may come to replace the bad one. Extinction is akin to exposure therapy for people with PTSD. Exposure therapy works, but not for everyone: it’s effective in only about 50 percent of cases, according to numerous studies, and it has a very high dropout rate.

The darters, it turned out, were more successful at extinction than the freezers, suggesting that the neurobiological processes of the males and females differed; the females, it appeared, had an edge. “Females may have developed adaptive strategies to fearful events,” says Shansky.

The results raise the question of whether PTSD treatments for women–who develop the disorder twice as frequently as men–should be different from those for men. Even more radically: Might it be possible to develop a therapy that alters the neural circuits of freezers to more closely resemble those of darters?  Science Daily  Original web page at Science Daily



* Surge in support for animal-research guidelines

Journals throw their weight behind checklist for rigorous animal experiments.

More than 600 research journals have now signed up to voluntary guidelines that are designed to improve the reporting of animal experiments.

Scientists have repeatedly pointed out that many published papers on animal studies suffer from poor study design and sloppy reporting — leaving the research at a substantial risk of bias.

So in 2010, the ARRIVE guidelines (Animals in Research: Reporting In Vivo Experiments) were introduced by a team led by the UK National Centre for the Replacement, Refinement and Reduction of Animals in Research (NC3Rs). They provide a detailed checklist of elements that should be included in any reporting of animal research, such as information about animal strain and sex, appropriate statistical calculations and disclosure of adverse events.

More than 150 journals endorsed the ARRIVE guidelines in 2015 alone, according to NC3Rs data — the highest number of signatories in a single year since the checklist’s release in 2010. By the end of January 2016, the total number had passed 600.

Still, endorsement does not mean enforcement: a 2014 study suggested that researchers largely ignore the voluntary guidelines. By comparing some papers published before and after the guidelines were issued, it found that there was little difference in the quality of reporting.

Nature doi:10.1038/nature.2016.  Nature  Original web page at Nature


Poorly-designed animal experiments in the spotlight

Preclinical research to test drugs in animals suffers from a “substantial” risk of bias because of poor study design, even when it is published in the most-acclaimed journals or done at top-tier institutions, an analysis of thousands of papers suggests. “You can’t rely on where the work was done or where it was published.”

Scientists can take basic steps to avoid possible biases in such experiments, says Malcolm Macleod, a stroke researcher and trial-design expert at the University of Edinburgh, UK. These include randomizing the assignment of animals to the trial’s treatment or control arm; calculating how large the sample needs to be to produce a statistically robust result; ‘blinding’ investigators as to which animals were assigned to which treatment until the end of the study; and producing a conflict-of-interest (COI) statement.

But many published papers make no mention of these methods, according to an analysis that Macleod conducted with Emily Sena, at the University of Edinburgh, and other colleagues. Looking at 2,671 papers from 1992 to 2011 that reported trials in animals, the team found randomization reported in 25%, blinding in 30%, sample-size calculations in fewer than 1% and COI statements in 12%. The papers were not selected at random; they had been included in meta-analyses of experimental disease treatments. Later studies reported randomization, blinding and COI statements at higher rates than did earlier ones, but rates never reached above 45%. “We could clearly be doing a lot better,” Macleod says.

The most-cited scientific journals don’t necessarily publish papers with more robust methods, Macleod adds. In fact, in 2011, the median journal impact factor was generally lower for studies that reported randomization than for publications that didn’t.

The researchers also looked at papers submitted by leading UK institutions to a national research-quality audit. They found that work done at the University of Oxford, the University of Cambridge, University College London, Imperial College London and the University of Edinburgh reported randomization only 14% of the time, and blinding only 17% of the time where it would have been appropriate. Of more than 1,000 publications, only one reported all four bias-reducing measures.

“Although sobering, the findings of this paper are not a surprise, as they add to the existing body of evidence on the need for more rigorous assessments of the experimental design and methodology used in animal research. This is another wake-up call for the scientific community,” said Vicky Robinson, chief executive of the London-based National Centre for the Replacement, Refinement and Reduction of Animals in Research, in a statement distributed by the UK Science Media Centre.

A separate analysis, also published today, shows that animal-based research on the cancer drug sunitinib is plagued with poor study design. A team at McGill University in Montreal, Canada, analysed the design of 158 published preclinical experiments, finding that none reported blinding or sample-size calculations and only 58 reported randomization. The researchers reported that publications were skewed towards those that reported positive effects — so much so that the team believes that published studies overestimate the effect of sunitinib on cancer by 45%.

Jonathan Kimmelman, the biomedical ethicist who led the work, says that journal editors, referees, institutions and researchers must all take responsibility for the poor quality of reporting and the consequent risk of bias. “There’s plenty of blame to go round,” he says.

But journals have in the last few years made efforts to address the problem. In 2010, researchers published ARRIVE guidelines for reporting animal research, which many journals have now endorsed, including Cell, Nature and Science Translational Medicine. And Philip Campbell, editor-in-chief of Nature, notes that since 2013, the journal has asked authors of life-sciences articles to include details about experimental and analytical design in their papers, and, during peer review, to complete a checklist focusing on often poorly-reported methods such as sample size, randomization and blinding.

Nature doi:10.1038/nature.2015.18559   Nature  Original web page at Nature


Self-propelled powder designed to stop severe bleeding

UBC researchers have created the first self-propelled particles capable of delivering coagulants against the flow of blood to treat severe bleeding, a potentially huge advancement in trauma care.

“Bleeding is the number one killer of young people, and maternal death from postpartum hemorrhage can be as high as one in 50 births in low resource settings so these are extreme problems,” explains Christian Kastrup, an assistant professor in the Department of Biochemistry and Molecular Biology and the Michael Smith Laboratories at the University of British Columbia.

Traditional methods of halting severe bleeding are not very effective when the blood loss originates inside the body like the uterus, sinus or abdomen.

“People have developed hundreds of agents that can clot blood but the issue is that it’s hard to push these therapies against severe blood flow, especially far enough upstream to reach the leaking vessels. Here, for the first time, we’ve come up with an agent that can do that,” Kastrup said

Kastrup teamed up with a group of researchers, biochemical engineers and emergency physicians to develop simple, gas-generating calcium carbonate micro-particles that can be applied in powder form to stop critical bleeding. The particles work by releasing carbon dioxide gas, like antacid tablets, to propel them toward the source of bleeding. The carbonate forms porous micro particles that can bind with a clotting agent known as tranexamic acid, and transport it through wounds and deep into the damaged tissue

After studying and modeling the movement of the particles in vitro, the researchers confirmed their results using two animal models. Even in a scenario that mimicked a catastrophic event like a gunshot wound to a femoral artery, the particles proved highly effective in stopping the bleeding

While much more rigorous testing and development is needed to bring the agent to market, the particles could have a wide range of uses, from sinus operations to treating combat wounds.

“The area we’re really focusing on is postpartum hemorrhage: in the uterus, after childbirth where you can’t see the damaged vessels but you can put the powder into that area and the particles can propel and find those damaged vessels,” said Kastrup.  Science Daily  Original web page at Science Daily


* An inside look at the first pig biobank

Boar 1339 was genetically engineered to have diabetes; its body parts, now in the Munich MIDY-PIG Biobank in Germany, are freely available to researchers. First out is a kidney: its dark red fades to beige as it is washed of its blood. The pancreas, harder to find amid the tangle of inner organs, is rushed on to dry ice. Speed is essential because tissues degrade after death — and each detail counts in this autopsy.

The precious organs belong to Boar 1339, which, for 3.5 years, had lived a normal pig’s life on the farm of a German university, despite the diabetes it was born with. Earlier this month, the animal was killed, and the body parts placed in the service of science, as part of a growing movement to maximize the scientific benefits of every animal used in research. Thousands of tiny tissue and fluid samples from the boar’s 226-kilogram body now sit in the newly constructed Munich MIDY-PIG Biobank in Germany — the world’s first systematic repository of tissue from a large, genetically engineered, non-human animal.

The biobank, part of Munich’s Ludwig Maximilian University (LMU), houses tissue taken from throughout the body and from pigs of different ages. It is designed to help diabetes researchers to discover the molecules and mechanisms involved in the long-term complications of the disease, including the degeneration of small blood vessels and nerves, heart and kidney disease, and blindness. These develop over a lifetime, and are poorly understood. “We are more similar to pigs than we like to think, so this resource will be very valuable,” says diabetes researcher Patrik Rorsman at the University of Oxford, UK. The samples are freely available to researchers anywhere in the world, who need only pay for the postage. As pressure grows to reduce the number of animals used in research, biobanks are becoming attractive because they allow teams working on different organs and aspects of the disease to use the same animal. “Biobanking means that no part of the animal is wasted,” says LMU geneticist and veterinary surgeon Eckhard Wolf, who launched the pig biobank.

Only a few animal biobanks have so far been built, and most are for mice. Pigs, although more expensive to house and breed, could be more useful because of their larger size and the greater similarity of their physiology and metabolism to those of humans. To create the pig biobank, scientists used genetic engineering and cloning techniques to create animals with a damaged gene called MIDY, which means that they need a daily insulin injection. The animals were then bred with healthy pigs so that, on average, half of the second-generation litters had diabetes and the other half were healthy, and thus able to serve as experimental controls. The 12th addition to the bank from this process, Boar 1339 was already anaesthetized when it arrived in the cavernous autopsy room of the 101-year-old veterinary school in a leafy suburb of Munich. Waiting for the pig was a team of 25 veterinary surgeons and technicians, gowned up, masked and alert at parallel dissection tables.

A hoist raised the pig by its hind legs and pathologists injected the animal with a lethal dose of anaesthetic. Then, in swift, precision choreography, the team moved in with their knives, cleavers, hammers and pincers. They removed organs, muscles, nerves and more, and transferred the tissues to the tables at which the waiting dissectors set about their complicated sampling process, strictly following an 80-page protocol. “We don’t want to complete a super-sophisticated sampling and then realize we forgot to weigh the liver,” says Andreas Blutke, one of the chief pathologists at the LMU. The chopping, hacking and puncturing are loud; the concentration of the workers keeps the background noise to a murmur. The whole process takes a mere 2 hours 15 minutes. Because the cells in any organ are of different sizes, shapes and orientations, and are unevenly distributed, the team takes numerous samples using different methods to ensure that the whole organ is appropriately represented. The system is so sophisticated that it gives researchers a three-dimensional anatomical reconstruction of the exact cell types.

Crucially, the researchers divide each sample and preserve the portions in different ways, each optimized for either structural or molecular analysis. This allows both types of analysis to be done on the same sample. “I think this is the only service which allows you to do molecular profiling and cellular anatomy from the same sample,” says Wolf. As soon as Rorsman heard about the biobank, he saw its benefit. He suspects that the long-term complications of diabetes are caused by changes in a particular molecule in the cells of several different tissues. Material from the biobank will allow him to confirm that he is on the right track, he says, before he obtains samples from human biobanks, which takes a long time because of ethical constraints. Herbert Tempfer, a diabetes researcher at the University of Salzburg in Austria, is already analysing samples of tendons — notoriously fragile in people with diabetes — from the biobank.

Others want to know how similar pig diabetes is to the human disease. Immunologist Åsa Hidmark at the University of Heidelberg in Germany made the three-hour journey to Boar 1339’s dissection to cut samples of skin from its trotter. She hopes to discover that the nerve endings in the outer layer have been lost, as happens in people with diabetes. The ultimate value of the bank will depend on how much it is used — and there are no guarantees. Despite its collection of 42 tissues taken from 940 mouse lines, a mouse biobank at the Wellcome Trust Sanger Institute near Cambridge, UK, has so far received only 50 or so requests for material. “There is a lack of awareness of its value,” says Jacqui White, who leads this Sanger mouse-autopsy project. Wolf plans to extend his biobank to other genetic pig models as they are developed. Next in line are probably pigs engineered to have Duchenne muscular dystrophy. A sow implanted with a cloned genetically modified embryo is now pregnant.

Nature 519, 397–398 (26 March 2015) doi:10.1038/519397a  Nature  Original web page at Nature


New compound prevents type 1 diabetes in animal models, before it begins

Scientists from the Florida campus of The Scripps Research Institute (TSRI) have successfully tested a potent synthetic compound that prevents type 1 diabetes in animal models of the disease. “The animals in our study never developed high blood sugar indicative of diabetes, and beta cell damage was significantly reduced compared to animals that hadn’t been treated with our compound,” said Laura Solt, PhD, a TSRI biologist who was the lead author of the study. Type 1 diabetes is a consequence of the autoimmune destruction of insulin-producing beta cells in the pancreas. While standard treatment for the disease aims to replace lost insulin, the new study focuses instead on the possibility of preventing the initial devastation caused by the immune system — stopping the disease before it even gets started.

In the study, published in the March 2015 issue of the journal Endocrinology, the scientists tested an experimental compound known as SR1001 in non-obese diabetic animal models. The compound targets a pair of nuclear receptors (RORα and RORγ) that play critical roles in the development of a specific population (Th17) of immune cells associated with the disease. “Because Th17 cells have been linked to a number of autoimmune diseases, including multiple sclerosis, we thought our compound might inhibit Th17 cells in type 1 diabetes and possibly interfere with disease progression,” said Solt. “We were right.” The researchers found SR1001 eliminated the incidence of diabetes and minimized insulitis, which is the inflammation associated with, and destroyer of, insulin-producing cells, in the treated animals.

The compound suppressed the immune response, including the production of Th17 cells, while maintaining normal insulin levels; it also increased the frequency of the expression of Foxp3 in T cells, which controls the development and function of a type of immune cell known as T regulatory cells. Solt notes that the study strongly suggests that Th17 cells have a pathological role in the development of type 1 diabetes and use of ROR-specific synthetic compounds targeting this cell type may have potential as a preventative therapy for type 1 diabetes. “It certainly opens the door for other areas to be looked at,” she said.  Science Daily  Original web page at Science Daily


How much does the public support animal research? Depends on the question

Embattled U.K. biomedical researchers are drawing some comfort from a new survey showing that a sizable majority of the public continues to support the use of animals in research. But there’s another twist that should interest social scientists as well: The government’s decision this year to field two almost identical surveys on the topic offers fresh evidence that the way you ask a question affects how people answer it. Since 1999, the U.K. Department for Business, Innovation & Skills (BIS) has been funding a survey of 1000 adults about their attitudes toward animal experimentation. But this year the government asked the London-based pollsters, Ipsos MORI, to carry out a new survey, changing the wording of several questions. The company also collected additional information, including public attitudes toward different animal species and current rules regarding their use. For example, the phrase “animal experimentation” was replaced by “animal research” because the latter is “less inflammatory,” notes Ipsos MORI Research Manager Jerry Latter. In addition, says Emma Brown, a BIS spokeswoman, the word research “more accurately reflects the range of procedures that animals may be involved in, including the breeding of genetically modified animals.” But government officials also value the information about long-term trends in public attitudes that can be gleaned from the current survey. So they told the company to conduct one last round—the 10th in the series—at the same time they deployed the new survey. Each survey went to a representative, but different, sample of U.K. adults. The changes in wording seem to have given animal researchers a bit of a ratings bump. In the new survey, some 68% agreed that “I can accept the use of animals in scientific research as long as it is for medical research purposes and there is no alternative.” By comparison, only 64% of respondents to the ongoing (trends) survey said yes when asked if they “can accept animal experimentation so long as it is for medical research purposes.” Although still strongly positive, that number was down from the roughly 75% recorded throughout the previous decade (the figure had dipped to 66% in 2012). One confounding factor in the new survey is the addition of a reference to “no alternatives.” When that phrase is added to a comparable question in the trends survey, the positive response drops from 64% to 60%.

The use of the word “medical” seems to have an even larger impact on public attitudes. When the trends survey asked a question that didn’t use the word “medical,” i.e., “I agree with animal experimentation for all types of research where there is no alternative,” only 47% of respondents say yes. And the positive response to the comparable question in the new survey is even lower. Only 37% say “it is acceptable to use animals for all types of research where there is no alternative.” Despite these subtleties, U.K. scientists believe the surveys show that the public will stand behind them if they speak out. “In the past, many scientists were understandably afraid of talking about their use of animals, but the climate has very much changed,” says Frances Rawle, head of policy at the Medical Research Council. “We encourage our researchers to be open about this work because it’s important that the public … are aware that research using animals is still an important and very necessary part of medical science.” At the same time, the new survey reveals that many people don’t understand what animal research is now being done and how it is regulated. For example, nearly one in three say that scientists in the United Kingdom can use animals to test cosmetics even though the practice has been banned for more than 15 years. Only 30% feel they are “well informed” about the use of animals in research, and some 55% would like to know more about efforts to find alternatives and improve animal welfare. That level of ignorance worries government officials. “It is of concern that most people feel uninformed about the use of animals in research,” BIS’s Brown says. “The lack of knowledge could be impacting people’s view of the need for animal research and their perception of the regulatory system around it.”  Science Magazine  Original web page at Science Magazine


UK institutions sign up to animal-research openness

British research organizations hope that more disclosure on how and why they conduct animal studies will shore up public support of their work. A who’s who of the United Kingdom’s most respected life-sciences organizations has today signed a document committing them to be open about animal research. In the so-called ‘concordat’, 72 organizations including universities, charities, industry and government funders say that they will provide more information about their work, engage more readily with media, and encourage more public discussion of animal research. Dominic Wells, who studies neuromuscular diseases at the Royal Veterinary College in London and was heavily involved in drawing up the concordat, says that today’s announcement probably makes the United Kingdom “the most open place in the world” regarding animal research. “I do feel we’re leading the way on this,” he says. Support for more transparency in animal research has gained ground in recent years, with many scientists hoping that better information about their work with animals and its goals will bring more public support. In 2010 researchers in Germany and Switzerland produced the Basel Declaration, pledging to be more open about their work. And the UK government earlier this month proposed to remove rules that prevent the Home Office from releasing much of the information it holds on animal research in the country. But despite these moves towards openness, there has still been resistance from some institutions. Wells says that universities sometimes block their researchers from talking about their animal research in press conferences, and job advertisements avoid mentioning animal research. A key aim of today’s concordat is to change such practices. The concordat comprises four commitments. It aims to put into practice a pledge made in 2012 by a number of UK life-sciences institutions, with detailed steps such as being realistic about the harms and limitations of animal research, releasing public statements on the use of animals in research and responding to “reasonable inquiries” about work undertaken or supported.

The document did not satisfy all anti-animal-research activists. The British Union for the Abolition of Vivisection in London said in a statement that the concordat was “transparency on their terms” and that it would continue its undercover work, in which activists have infiltrated laboratories at Imperial College London and elsewhere. Those supporting the concordat insist that it is a major advance. Wendy Jarrett, chief executive of the London-based group Understanding Animal Research, which campaigns in support of animal research and has worked on the concordat, says that there will be genuine change as a result. “We have a broad spectrum of signatories here,” she told reporters in London. Some of them “have been open for many years and doing practically everything that’s in the concordat already. In some cases there are organizations who have signed up who know they have quite a way to go.”

Nature doi:10.1038/nature.2014.15222  Nature

June 10, 2014  Original web page at Nature


Misleading mouse studies waste medical resources

A running joke among health researchers is that everything has been cured — in mice. However that may not be always true. The failure of experimental drugs that had once looked promising could have been prevented with better animal studies, according to a re-examination of past clinical trials. “I hear too many stories about patients who have used their one shot at getting into a trial on a drug that didn’t have enough legs to begin with, and that’s a tragedy,” says Steve Perrin, an amyotrophic lateral sclerosis (ALS) researcher who led the work. Perrin, chief scientific officer of the ALS Therapy Development Institute in Cambridge, Massachusetts, used mice with symptoms similar to ALS to test more than 100 compounds that had previously been identified as candidate drugs. Most — including eight that had shown promise in previous mouse work but ultimately failed in humans trials — failed to slow the progressive, fatal degenerative disease, also called Lou Gehrig’s disease or motor neuron disease. Perrin argues that positive results seen in previous mouse trials were spurious, probably resulting from poorly conducted studies. In a Comment published today in Nature, he is urging researchers to boost the quality of animal studies by better characterizing and understanding how mouse models correspond to human disease, minimizing result-muddling variation between animals, and using statistical models to guide study design. “The recommendations could have a profound positive impact on translational research, strongly improving the quality of preclinical studies,” says Adriano Chiò, a neurologist at the University of Turin in Italy. Perrin found that one type of mouse model for ALS, in which animals express a mutant version of the protein TDP43, differs in key ways from the human disease. For example, TDP43 mice usually died of bowel obstructions, whereas humans with the disease tend to succumb to muscle wasting, which often results in the inability to breathe. He further found that although the first generation of TDP43 mice had been reported to die within 200 days, later generations bred from those originals lived for up to 400 days without signs of disease.

Other researchers have highlighted problems with reproducibility of animal studies in cancer. Neurobiologist Caterina Bendotti of the Mario Negri Institute for Pharmacological Research in Milan, Italy, agrees that the issues Perrin describes are not unique to his field: “The poor reproducibility of preclinical results, particularly in animal models, goes beyond ALS,” she says. Irreproducible preclinical results can lead to a massive waste of time and money in clinical trials. Chiò points to the example of a 2008 study in mice and 44 humans with ALS, which suggested that therapeutic lithium might slow the disease. The metal is already used as a drug to treat mental disorders including schizophrenia, so it is available cheaply, and people with ALS began administering it to themselves. Perrin’s and Bendotti’s groups both tried to replicate the original results, and found that they could not — but not before clinical trials began to test the effect. Ultimately, five trials involving more than 1,000 people with ALS in five countries found no benefit, Chiò says, and the initial mouse results were never duplicated. Untreated animals in the original study survived for 20 days less than those in other mouse studies, suggesting that there may have been some anomaly with these animals. “It is our stringent responsibility to avoid a new lithium saga, and Perrin’s recommendations go exactly in this direction,” Chiò says. Other researchers say they agree broadly with Perrin, but that they would also like to see the data from his group’s experiments, and that it may not be necessary to find a positive animal result to progress to a human trial. “Many ALS researchers would say that if there is other preclinical evidence that a drug might work, it may be sufficient to go into human trials,” says neurologist Leonard van den Berg, director of the NetherlandsALSCenter at University Medical Center Utrecht. But work to discover how relevant animal models are to human disease — such as Perrin’s studies on TDP43 mice — is expensive and unrewarding for researchers and their teams. Perrin argues that they must be funded specifically, perhaps by private–public collaborations. “Somebody has to do this, or else we’re wasting precious resources,” says Perrin.

Nature doi:10.1038/nature.2014.14938 Nature

April 15, 2014  Original web page at Nature


First glimpse of brain circuit that helps experience to shape perception

In work published today in Nature Neuroscience, scientists from Cold Spring Harbor Laboratory (CSHL) demonstrate for the first time a way to observe this process in awake animals. The team, led by Assistant Professor Stephen Shea, was able to measure the activity of a group of inhibitory neurons that links the odor-sensing area of the brain with brain areas responsible for thought and cognition. This connection provides feedback so that memories and experiences can alter the way smells are interpreted. The inhibitory neurons that forge the link are known as granule cells. They are found in the core of the olfactory bulb, the area of the mouse brain responsible for receiving odor information from the nose. Granule cells in the olfactory bulb receive inputs from areas deep within the brain involved in memory formation and cognition. Despite their importance, it has been almost impossible to collect information about how granule cells function. They are extremely small and, in the past, scientists have only been able to measure their activity in anesthetized animals. But the animal must be awake and conscious in order to for experiences to alter sensory interpretation. Shea worked with lead authors on the study: Brittany Cazakoff, graduate student in CSHL’s Watson School of Biological Sciences, and Billy Lau, PhD, a postdoctoral fellow. They engineered a system to observe granule cells for the first time in awake animals.

Granule cells relay the information they receive from neurons involved in memory and cognition back to the olfactory bulb. There, the granule cells inhibit the neurons that receive sensory inputs. In this way, “the granule cells provide a way for the brain to ‘talk’ to the sensory information as it comes in,” explains Shea. “You can think of these cells as conduits which allow experiences to shape incoming data.” Why might an animal want to inhibit or block out specific parts of a stimulus, like an odor? Every scent is made up of hundreds of different chemicals, and “granule cells might help animals to emphasize the important components of complex mixtures,” says Shea. For example, an animal might have learned through experience to associate a particular scent, such as a predator’s urine, with danger. But each encounter with the smell is likely to be different. Maybe it is mixed with the smell of pine on one occasion and seawater on another. Granule cells provide the brain with an opportunity to filter away the less important odors and to focus sensory neurons only on the salient part of the stimulus. Now that it is possible to measure the activity of granule cells in awake animals, Shea and his team are eager to look at how sensory information changes when the expectations and memories associated with an odor change. “The interplay between a stimulus and our expectations is truly the merger of ourselves with the world. It exciting to see just how the brain mediates that interaction,” says Shea.  Science Daily

March18, 2014  Original web page at Science Daily


Discovery of a ‘conductor’ in muscle development could impact on the treatment of muscular diseases

A team led by Jean-François Côté, researcher at the IRCM, identified a ”conductor” in the development of muscle tissue. The discovery, published online by the scientific journal Proceedings of the National Academy of Sciences (PNAS), could have an important impact on the treatment of muscular diseases such as myopathies and muscular dystrophies. For several years, we have been studying myogenesis, a process by which muscles are formed during embryonic development,” says Jean-François Côté, PhD, Director of the Cytoskeletal Organization and Cell Migration research unit at the IRCM. “During the last step of this process, muscle cells called myoblasts align and fuse together to form muscle fibers.” The fusion of myoblasts is a critical step in the formation of embryonic muscle fibers as it determines muscle size, among other things. This process is also important in adult life because muscle stem cells fuse with existing fibers to achieve muscle growth and help regenerate damaged muscles. However, until now, fusion remained a poorly understood step within the scientific community. “We were able to identify the receptor BAI3, a protein at the surface of myoblasts, as one of the crucial missing links in the fusion of muscle cells,” adds Dr. Côté. “In fact, this receptor acts much like an orchestra conductor by activating a signalling pathway required for this important process.”

In 2008, Dr. Côté’s team explained the role of the DOCK1 and DOCK5 genes in the development of muscle tissue by showing that these two genes were critical regulators of the fusion process in mice. In their most recent study, the researchers confirmed receptor BAI3’s essential role by blocking its interaction with the DOCK signalling pathway. They discovered that, as a result, myoblast fusion was also blocked. “Our scientific breakthrough will undoubtedly have a translational research application on the regeneration of tissue from stem cells, given that a better understanding of the molecular mechanisms of fusion are required for the development of such therapies,” concludes Dr. Côté. “This could therefore have an impact on the treatment of muscular diseases, including myopathies and muscular dystrophies.”  Science Daily  Original web page at Science Daily


UK ‘absolutely committed’ to reducing animals used in research

Government stands by pledge but shies away from hard target as number of experiments rises. The UK government says that it will intensify steps to reduce the use of animals in laboratory research. British ministers insisted today that they are still committed to reducing the number of animals used in research, but warned that this might not mean a reduction in the overall number of scientific procedures involving animals. Science minister David Willetts told reporters in London that the government was “absolutely committed” to the so-called 3Rs of reducing, replacing and refining the use of animals. “This is about the scientific community doing its best whenever possible to reduce and replace the use of animals,” he added. “This isn’t about a numerical target.” The number of scientific procedures involving animals in the United Kingdom reached a peak of around 5.5 million in the 1970s before dropping to just over 2.5 million in 2000. Since then, however, it has increased to more than 4 million in 2012, and despite the government’s promise in 2010 to “work to reduce the use of animals in scientific research”. Today’s action plan pledges support for the London-based National Centre for the 3Rs (NC3Rs); to encourage data sharing between animal researchers to minimise duplication; and to increase the role of government inspectors of animal research in promoting the 3Rs. For example, inspectors will give more guidance to researchers on alternative lab technique that do not require lab animals.

Norman Baker, the Home Office minister responsible for animal research, insisted that there was no other country doing as much as the UK to reduce the use of lab animals. He said that the government had already backed work — such as developing non-animal tests for detection of toxins in commercial shellfish — that had led to reductions. Had such work not been done, he added, “we would have a higher number than we’ve currently got”. Echoing Willetts, Baker said it would be “artificial” for the UK to try and set an overall target for the number of animal experiments, given the global nature of science. UK animal-rights groups criticised today’s announcement. The Nottingham-based Fund for the Replacement of Animals In Medical Experiments said it was disappointed by the lack of targets, while the London-based British Union for the Abolition of Vivisection said it showed that the government was abandoning its 2010 pledge. Mark Walport, the government’s chief scientific adviser, cautioned that the increase in animal-research procedures seen in official statistics was mainly down to an increase in the breeding of genetically-modified animals — whose births are counted as procedures — and not to what might more generally be considered ‘experiments’, which have remained roughly stable at 2 million per year in the past decade. Walport said that scientists were increasingly transparent about their use of animals, and increasingly sophisticated in how they used them. Vicky Robinson, chief executive of the NC3Rs, welcomed the report and said progress was being made. “Most people are starting to get it [3Rs] isn’t a regulatory tick-box. It’s about how we do the best science,” she told Nature. Nature March 4, 2014  Original web page at Nature


Scientists reprogram skin cells into insulin-producing pancreas cells

A cure for type 1 diabetes has long eluded even the top experts. Not because they do not know what must be done — but because the tools did not exist to do it. But now scientists at the Gladstone Institutes, harnessing the power of regenerative medicine, have developed a technique in animal models that could replenish the very cells destroyed by the disease. The team’s findings, published online in the journal Cell Stem Cell, are an important step towards freeing an entire generation of patients from the life-long injections that characterize this devastating disease. Type 1 diabetes, which usually manifests during childhood, is caused by the destruction of ß-cells, a type of cell that normally resides in the pancreas and produces a hormone called insulin. Without insulin, the body’s organs have difficulty absorbing sugars, such as glucose, from the blood. Once a death sentence, the disease can now be managed with regular glucose monitoring and insulin injections. A more permanent solution, however, would be to replace the missing ß-cells. But these cells are hard to come by, so researchers have looked towards stem cell technology as a way to make them. “The power of regenerative medicine is that it can potentially provide an unlimited source of functional, insulin-producing ß-cells that can then be transplanted into the patient,” said Dr. Ding, who is also a professor at the University of California, San Francisco (UCSF), with which Gladstone is affiliated. “But previous attempts to produce large quantities of healthy ß-cells — and to develop a workable delivery system — have not been entirely successful. So we took a somewhat different approach.”

One of the major challenges to generating large quantities of ß-cells is that these cells have limited regenerative ability; once they mature it’s difficult to make more. So the team decided to go one step backwards in the life cycle of the cell. The team first collected skin cells, called fibroblasts, from laboratory mice. Then, by treating the fibroblasts with a unique ‘cocktail’ of molecules and reprogramming factors, they transformed the cells into endoderm-like cells. Endoderm cells are a type of cell found in the early embryo, and which eventually mature into the body’s major organs — including the pancreas. “Using another chemical cocktail, we then transformed these endoderm-like cells into cells that mimicked early pancreas-like cells, which we called PPLC’s,” said Gladstone Postdoctoral Scholar Ke Li, PhD, the paper’s lead author. “Our initial goal was to see whether we could coax these PPLC’s to mature into cells that, like ß-cells, respond to the correct chemical signals and — most importantly — secrete insulin. And our initial experiments, performed in a petri dish, revealed that they did.” The research team then wanted to see whether the same would occur in live animal models. So they transplanted PPLC’s into mice modified to have hyperglycemia (high glucose levels), a key indicator of diabetes. 

“Importantly, just one week post-transplant, the animals’ glucose levels started to decrease gradually approaching normal levels,” continued Dr. Li. “And when we removed the transplanted cells, we saw an immediate glucose spike, revealing a direct link between the transplantation of the PPLC’s and reduced hyperglycemia.” But it was when the team tested the mice eight weeks post-transplant that they saw more dramatic changes: the PPLC’s had given rise to fully functional, insulin-secreting ß-cells. “These results not only highlight the power of small molecules in cellular reprogramming, they are proof-of-principle that could one day be used as a personalized therapeutic approach in patients,” explained Dr. Ding. “I am particularly excited about the prospect of translating these findings to the human system,” said Matthias Hebrok, PhD, one of the study’s authors and director of the UCSF Diabetes Center. “Most immediately, this technology in human cells could significantly advance our understanding of how inherent defects in ß-cells result in diabetes, bringing us notably closer to a much-needed cure.”  Science Daily

March 4, 2014  Original web page at Science Daily


Report slams university’s animal research

Imperial College London accepts heavy criticism from independent reviewers after exposé by animal-rights group. The treatment of laboratory animals at one of the United Kingdom’s most prestigious universities came under severe criticism today from an independent review set up in the wake of allegations of malpractice. Imperial College London’s animal-research facilities are understaffed, under-resourced and operating without adequate systems for training, supervision, management and ethical review, according to the report released today. “Across a whole number of areas there needs to be significant improvement,” says Steve Brown, director of the Medical Research Council Mammalian Genetics Unit in Harwell, UK, and the man appointed to chair the review. The university has accepted all of the recommendations of the review, and said in a statement that “there is significant scope for improvement”. Imperial appointed Brown after an undercover investigation by London-based campaign group BUAV (British Union for the Abolition of Vivisection), which in April alleged that many animals at the university’s facilities were subjected to severe suffering. The BUAV’s exposé included graphic imagery and reported incidents such as a researcher leaving a clamp inside an animal after surgery and animals being left in distress in short-staffed laboratories over a weekend. The UK government’s Home Office launched an official investigation into these specific allegations, but Brown’s group was appointed in parallel to look at animal care and welfare across the whole university.

The report makes “substantive” recommendations for necessary improvement in four main areas: operations at the Central Biomedical Services facility in Hammersmith, west London; training; culture, leadership and management; and the university’s ethical-review body. Brown’s review found that Imperial’s Hammersmith facility was well equipped and that there was a high quality of animal husbandry, but there was still “considerable room for improvement” in working practices, operational standards and mechanisms for reporting concerns about animal welfare. The report also found that there were not enough staff at this facility, and there was too much reliance on temporary staff. Overall, the investigators highlighted the “ad hoc” nature of competency assessment, supervision and training, and found little provision for sharing information and best practice between staff members. In addition, the work culture was found to be flawed, with a focus on procedure to the detriment of improvements in the ‘three Rs’: approaches for replacing, reducing and refining the use of animals in experiments. “The committee found that there was a level of complacency at the College with little opportunity for challenge or bringing in new ideas and that there was scope for improvement in the culture, management and overall leadership,” says the report.

Finally, the university’s crucial Animal Welfare and Ethical Review Body — which is responsible for reviewing animal use — was found to be “not fit for purpose” and in need of “wholesale reform”, says the report. In its response, Imperial says it will invest in staffing and bring in a plan by February 2014 to respond to the committee’s report, and that it has already enhanced training for researchers who work on animals. “The College will now move quickly to implement the recommendations,” said a statement from the university. “The committee commended the high standards of husbandry and animal care at Imperial. However, the College accepts that there is significant scope for improvement in aspects of the operation, management and oversight in order to become a world leader in animal research.” Sarah Kite, director of special projects at the BUAV, says that the conclusions reached by the report are “damning”, and highlight that there are not enough Home Office inspectors to monitor the United Kingdom’s animal researchers. “It shouldn’t have been down to the BUAV to uncover these failings,” says Kite. “If this is happening in one of the UK’s leading universities, it raises the question of what could be going on in other facilities around the country.” Paul Flecknell, director of the research-animal facilities at Newcastle University, UK, and a member of the committee behind the report, stressed that the investigation was specific to Imperial, but he says that “every institution will pick up something we’ve said and think, ‘we should take more note of that’”. The Home Office investigation of the specific incidents uncovered by the BUAV was due to be produced by the end of the year, but a spokeswoman told Nature last week that there was currently no clear date for when it will be available.

January 7, 2014

Original web page at Nature


For first time, drug developed based on zebrafish studies passes Phase I clinical trial

Zebrafish research achieved a significant milestone when the first drug developed through studies utilizing the tiny animal and then put into clinical trials passed a Phase 1 trial aimed at establishing its safety. The drug, discovered in the laboratory of Leonard Zon, MD, at Boston Children’s Hospital, has already advanced to Phase II studies designed to determine its efficacy. Results of the safety trial were reported recently in the journal Blood. At only six years after Zon’s laboratory reported the discovery of the chemical from which the drug is derived, the Phase 1 data underscore the potential of zebrafish research to accelerate the journey from bench to bedside. “The zebrafish is a very good system for evaluating potential drugs,” Zon said. “When you discover a new treatment option and can see it go into patients, it’s quite a remarkable feeling.” The drug, which is being developed by Fate Therapeutics under the name ProHema®, is the result of the Zon laboratory’s search for a way to improve the success of hematopoietic stem cell (HSC) transplants using umbilical cord blood. ProHema is a chemical derivative of prostaglandin E2 (PGE2) that, according to preclinical and clinical data, might improve engraftment of transplanted umbilical cord blood cells by helping donated cells home in on the bone marrow.

Although umbilical cords are an effective transplant source in patients for whom a suitable donor cannot be found, a single umbilical cord rarely contains enough HSCs for a transplant for an adult patient. The current method is to use two cord blood units per transplant, raising the risk that the immune cells that arise from the two cords may start to attack each other. In addition, umbilical cords are expensive and in limited supply. This problem has led Zon, a co-author of the Blood study, and other researchers to search for molecules that could help expand cord blood stem cells or improve the efficiency of cord blood transplants and eliminate the need for cells from a second cord. Zon’s laboratory discovered PGE2’s properties after screening 2,500 chemicals for their effects on blood stem cell production in zebrafish, a popular and cost-effective research model for stem cell, genetic and developmental research. Not only are zebrafish genes surprisingly similar to human genes, but they can be inexpensively housed at high densities and female zebrafish lay 300 eggs per week, making them a promising vehicle for quickly and cheaply discovering new drugs. Zon and his colleagues reported their initial PGE2 findings in Nature in 2007.

“We think PGE2 acts as a kind of priming mechanism,” Zon said. “It gets the cell set so that it will function better once it is introduced into the recipient’s body.” Subsequent preclinical studies showed that PGE2 can trigger a four-fold increase in efficiency of stem cell engraftment, compared to untreated controls, by helping stem cells home more effectively to the bone marrow. The Phase I trial of ProHema, the drug derived from PGE2, was launched in 2009 at Dana-Farber Cancer Institute (DFCI) and Massachusetts General Hospital under the direction of DFCI’s Corey Cutler, MD, MPH. It showed that treatment of donated umbilical cord blood stem cells with the drug before transplant was safe. In addition, treated cells could engraft and rebuild a patient’s blood system more quickly than untreated ones. “These are very promising results,” Cutler said. “They suggest that by generating more effective stem cells, we might be able to lower the dose of stem cells needed for a successful transplant. And because this approach takes substantially less time than techniques that increase the number of stem cells prior to transplant, it can easily be performed by most stem cell-processing facilities.” Because the Food and Drug Administration has already approved PGE2 for other uses, researchers were able to move quickly into clinical trials. The Phase II study is underway at seven institutions nationwide.

EurekAlert! Medicine
November 12, 2013

Original web page at EurekAlert! Medicine


Knockout mouse grows larger, but weaker, muscles

Although muscle cells did not reduce in size or number in mice lacking a protective antioxidant protein, they were weaker than normal muscle cells, researchers from the Barshop Institute for Longevity and Aging Studies at The University of Texas Health Science Center San Antonio found. The scientists, who are faculty in the university’s School of Medicine, are studying how oxidative stress in cells impacts sarcopenia — a loss of muscle mass and strength that occurs in all humans as they age. The antioxidant protein is called SOD1. The researchers developed mice that did not have SOD1 in their muscles, though it was still present in other types of cells. Then they asked the question: Is lack of SOD1 at the muscle enough to cause atrophy? Surprisingly, the total muscle mass in this mouse was larger. “We think that lack of SOD1 could be priming the muscle to use all of its survival skills,” said Holly Van Remmen, Ph.D., professor of cellular and structural biology in the School of Medicine and associate director for basic research at the Barshop Institute. “The muscle knows things aren’t quite right. Its rescue mechanisms are pulled into play.” But even though the muscles were not atrophied, they were still weak.

Sarcopenia in people has two components: loss of muscle mass and loss of function (weakness). This study supports the idea that oxidative stress has a role in these detrimental effects. If a way can be found to curb the effects, then healthier, more productive aging could result, Dr. Van Remmen said. The oxidative stress theory of aging holds that oxidation from molecules called “free radicals” causes damage to cells over time, resulting in sarcopenia and other decline. The study is described in The FASEB Journal. Future research will assess whether limiting oxidative stress can effect muscle regeneration, Dr. Van Remmen said.

Science Daily
September 3, 2013

Original web page at Science Daily


New animal model may lead to treatments for common liver disease

Scientists at Texas Biomed have developed the laboratory opossum as a new animal model to study the most common liver disease in the nation — afflicting up to 15 million Americans — and for which there is no cure. The condition, nonalcoholic steatohepatitis (NASH), resembles alcoholic liver disease, but occurs in people who drink little or no alcohol. The major feature of NASH is accumulation of fat in the liver, along with inflammation and functional damage. Most people with NASH feel well and are not aware that they have a liver problem. Nevertheless, NASH can progress to cirrhosis, in which the liver is permanently damaged and no longer able to work properly. NASH-related cirrhosis is the fourth most common indication for liver transplantation in the U.S. NASH affects 2 to 5 percent of Americans — roughly six million to 15 million people. An additional 15 to 30 percent of Americans have excess fat in their livers, but no inflammation or liver damage, a condition called “fatty liver” or the non-progressive form of nonalcoholic fatty liver disease (NAFLD).

The study, published in the July issue of the American Journal of Physiology-Gastrointestinal and Liver Physiology, was supported by the National Institutes of Health and the Robert J. Kleberg, Jr., and Helen C. Kleberg Foundation. “This is the type of model in which to develop mechanism-based therapies,” writes Geoffrey C Farrell, M.D., of the Australian National University Medical School in Canberra, in a journal editorial. Both NASH and NAFLD are becoming more common, possibly because of the greater number of Americans with obesity and its important health complications, type 2 diabetes, high blood cholesterol levels, high blood pressure and other risk factors for heart attack and stroke. In the past 10 years, the prevalence of obesity has doubled in adults and tripled in children. It was previously reported by other scientists that the prevalence of NAFLD and NASH in a cohort of middle-aged patients in San Antonio is 46 percent and 12 percent, respectively. “It now seems likely that genetic factors, such as those important for diabetes and high cholesterol levels, are what determines why a small proportion of those with fatty liver develop NASH and its complications of cirrhosis and liver cancer,” said Farrell.

In the new study, high responding opossums developed elevated cholesterol and fatty liver disease when fed a high cholesterol and high fat diet, whereas low responding opossums did not. High responders carry a mutated ABCB4 gene, which affects their ability to secrete excess cholesterol from the liver into bile which, in turn, transports the cholesterol to the intestines for excretion from the body. As a consequence, opossums with the mutated gene accumulate cholesterol in the liver and ultimately in the blood. “We showed that the fatty livers of high responders contain a tremendous amount of cholesterol,” said first author Jeannie Chan, Ph.D., of Texas Biomed. “The opossum is a new animal model for investigating the mechanism by which cholesterol mediates liver injury, which will lead to a better understanding of the role of dietary cholesterol in the development of NASH.”

Science Daily
July 24, 2012

Original web page at Science Daily