Posts tagged stem cells
Articles and news from the latest research reports.
Faulty stem cell regulation may contribute to cognitive deficits associated with Down syndrome
The learning and physical disabilities that affect people with Down syndrome may be due at least in part to defective stem cell regulation throughout the body, according to researchers at the Stanford University School of Medicine. The defects in stem cell growth and self-renewal observed by the researchers can be alleviated by reducing the expression of just one gene on chromosome 21, they found.
The finding marks the first time Down syndrome has been linked to stem cells, and addresses some long-standing mysteries about the disorder. Although the gene, called Usp16, is unlikely to be the only contributor to the disease, the finding raises the possibility of an eventual therapy based on reducing its expression.
“There appear to be defects in the stem cells in all the tissues that we tested, including the brain,” said Michael Clarke, MD, Stanford’s Karel H. and Avice N. Beekhuis Professor in Cancer Biology. The researchers conducted their studies in both mouse and human cells. “We believe Usp16 overexpression is a major contributor to the neurological deficits seen in Down syndrome.”
Clarke is the senior author of the research, published Sept. 11 in Nature. Postdoctoral scholar Maddalena Adorno, PhD, is the lead author.
“Conceptually, this study suggests that drug-based strategies to slow the rate of stem cell use could have profound effects on cognitive function, aging and risk for Alzheimer’s disease in people with Down syndrome,” said co-author Craig Garner, PhD, who is the co-director of Stanford’s Center for Research and Treatment of Down Syndrome and a professor of psychiatry and behavioral sciences.
Down syndrome, which is caused by an extra copy of chromosome 21, affects about 400,000 people in the United States and 6 million worldwide. It causes both physical and cognitive problems. While many of the physical issues, such as vulnerability to heart problems, can now be treated, no treatments exist for poor cognitive function.
The new study’s findings suggest answers to many long-standing mysteries about the condition, including why people with Down syndrome appear to age faster and exhibit early Alzheimer’s disease.
“This study is the first to provide a possible explanation for these tendencies,” said Garner. The fact that people with Down syndrome have three copies of chromosome 21 and the Usp16 gene “accelerates the rate at which stem cells are used during early development, which likely exhausts stem cell pools and impairs tissue regeneration in adults with Down syndrome. As a result, their brains age faster and are susceptible to early onset neurodegenerative disorders.”
The researchers didn’t confine their studies to laboratory mice. They also investigated the effect of Usp16 overexpression in human cells. Adorno and colleagues in the laboratory of co-author Samuel Cheshier, MD, assistant professor of neurosurgery, found that the presence of excess Usp16 caused skin cells from unaffected people to grow more slowly. Furthermore, neural progenitor cells (those self-renewing cellular factories responsible for the development and maintenance of many of the cell types in the brain) were less able to form balls of cells called neurospheres — a laboratory test that reflects the number and robustness of nerve stem cells in a culture. Conversely, reducing Usp16 expression in skin and nerve-progenitor cells from people with Down syndrome allowed the cells, which usually proliferate slowly, to assume normal growth patterns.
“This gene is clearly regulating processes that are central to aging in mice and humans,” said Clarke, “and stem cells are severely compromised. Reducing Usp16 expression gives an unambiguous rescue at the stem cell level. The fact that it’s also involved in this human disorder highlights how critical stem cells are to our well-being.”
Adorno and Clarke didn’t set out to study Down syndrome. Clarke’s past research has focused on how normal stem cells and cancer stem cells regenerate themselves, and Adorno was searching for genes that could inhibit a specific molecular pathway involved in the self-renewal of these cells. Understanding how normal stem cells regenerate themselves could help to repair tissue and organ damage from disease, and understanding how cancer stem cells maintain themselves could help explain why they are unusually resistant to chemotherapy or radiation therapy — often resulting in a patient’s relapse after seemingly successful treatment. Usp16 seemed to fit the bill; it plays a critical role in a self-renewal pathway previously identified by Clarke and his colleagues.
But Adorno and Clarke soon realized that Usp16 had another interesting property: in humans, it is found on chromosome 21.
They turned to Garner and Cheshier to help them evaluate a possible link to Down syndrome. Garner supplied two strains of mice commonly used to study the condition. One, Ts65Dn, has three copies of 132 genes found on human chromosome 21 — including Usp16. The second, Ts1Cje, has three copies of 79 genes from the chromosome, but only two copies of Usp16. Although both mice display some symptoms of the disorder, Ts65Dn more closely mimics the craniofacial structure and learning and memory disabilities seen in affected humans.
Colleagues in the Cheshier laboratory found that neural stem cells from the more-severely affected Ts65Dn mice were less able to self-renew and grow normally than were cells from the Ts1Cje mice. Reducing the expression of Usp16 in the cells from the Ts65Dn mice to more normal levels largely corrected these functional defects.
“We demonstrated that central nervous system stem cells in Down syndrome mice were defective in their ability to self-renew — the process by which stem cells regenerate themselves upon cell division. Blocking Usp16 expression in these cells restored this ability,” said Cheshier. “We hope in the future that correcting this Usp16 defect can lead to therapeutics that will ameliorate the central nervous system defects seen in patients with Down syndrome.”
Finally, the researchers created a new, Ts65Dn-derived mouse strain in which one of the three copies of Usp16 was mutated. This normalized the level of expression of that gene, without affecting the overexpression of the other 131 triplicated genes in these mice. Nerve progenitor cells from these mice were equally able as normal cells to form neurospheres. The researchers are now continuing their studies of these mice.
“We are really interested in learning how other genes in this chromosomal region may be affecting stem cell renewal,” said Clarke. “We also want to understand how much we’re able to rescue the neurological defect by normalizing the expression of Usp16 in this mouse model. How does this compare to what is happening in humans? We’re sure it plays some significant role.”
(Source: med.stanford.edu)
Cell transplants may be a novel treatment for schizophrenia
Rodent research suggests feasibility of restoring neuron function
Research from the School of Medicine at The University of Texas Health Science Center at San Antonio suggests the exciting possibility of using cell transplants to treat schizophrenia.
Cells called “interneurons” inhibit activity within brain regions, but this braking or governing function is impaired in schizophrenia. Consequently, a group of nerve cells called the dopamine system go into overdrive. Different branches of the dopamine system are involved in cognition, movement and emotions.
“Since these cells are not functioning properly, our idea is to replace them,” said study senior author Daniel Lodge, Ph.D., assistant professor of pharmacology in the School of Medicine.
Transplant restored normal function
Dr. Lodge and lead author Stephanie Perez, graduate student in his laboratory, biopsied tissue from rat fetuses, isolated cells from the tissue and injected the cells into a brain center called the hippocampus. This center regulates the dopamine system and plays a role in learning, memory and executive functions such as decision making. Rats treated with the transplanted cells have restored hippocampal and dopamine function.
Stem cells are able to become different types of cells, and in this case interneurons were selected. “We put in a lot of cells and not all survived, but a significant portion did and restored hippocampal and dopamine function back to normal,” Dr. Lodge said.
‘You can essentially fix the problem’
Unlike traditional approaches to treating schizophrenia, such as medications and deep-brain stimulation, transplantation of interneurons potentially can produce a permanent solution. “You can essentially fix the problem,” Dr. Lodge said. “Ultimately, if this is translated to humans, we want to reprogram a patient’s own cells and use them.”
After meeting with other students, Perez brought the research idea to Dr. Lodge. “The students have journal club, and somebody had done a similar experiment to restore motor deficits and had good results,” Perez said. “We thought, why can’t we use it for schizophrenia and have good results, and so far we have.”
The study is in Molecular Psychiatry.
(Source: uthscsa.edu)
Scientists use latest stem cell and gene-editing techniques to generate neurons in a dish, and reveal new clues behind deadly diseases of the brain
There is no easy way to study diseases of the brain. Extracting neurons from a living patient is both difficult and risky, while examining a patient’s brain post-mortem usually only reveals the disease’s final stages. And animal models, while incredibly informative, have frequently fallen short during the crucial drug-development stage of research. The result: we are woefully unprepared to fight—and win—the war against this class of diseases.
But scientists at the Gladstone Institutes and the University of California, San Francisco (UCSF) are taking a potentially more powerful approach: an advanced stem-cell technique that creates a human model of degenerative disease in a dish.
Using this model, the team uncovered a molecular process that causes neurons to degenerate, a hallmark sign of conditions such as Alzheimer’s disease and frontotemporal dementia (FTD). The results, published in the latest issue of Stem Cell Reports, offer fresh ammunition in the continued battle against these and other deadly neurodegenerative disorders.
The research team, led by Gladstone Investigator Yadong Huang, MD, PhD, identified an important mechanism behind tauopathies. A group of disorders that includes both Alzheimer’s and FTD, tauopathies are characterized by the abnormal accumulation of the protein Tau in neurons. This buildup is thought to contribute to the degeneration of these neurons over time, leading to debilitating symptoms such as dementia and memory loss. But while this notion has been around for a long time, the underlying processes have largely remained unclear.
“So much about the mechanisms that cause tauopathies is a mystery, in part because traditional approaches—such as post-mortem brain analysis and animal models—give an incomplete picture,” explained Dr. Huang. “But by using the latest stem-cell technology, we generated human neurons in a dish that exhibited the same pattern of cell degeneration and death that occurs inside a patient’s brain. Studying these models allowed us to see for the first time how a specific genetic mutation may kick start the tauopathy process.”
Other scientists recently discovered that the Tau mutation in question could increase a person’s risk of developing different tauopathies, including Alzheimer’s or FTD. So the research team, in collaboration with Bruce Miller, MD, who directs the UCSF Memory and Aging Center and who provided skin cells from a patient with this mutation, transformed these cells into induced pluripotent stem cells, or iPS cells. This technique, pioneered by Gladstone Investigator and 2012 Nobel Laureate Shinya Yamanaka, MD, PhD, allows scientists to reprogram adult skin cells into cells that are virtually identical to stem cells. These stem cells can then develop into almost any cell in the body.
The team combined this method with a cutting-edge gene-editing technique that essentially eliminated the Tau mutation in some of the iPS cells. The result was a system that allowed the team to compare neurons that had the mutation to those that did not.
“Our approach allowed us to grow human neurons in a dish that contained the exact same mutation as the neurons in the brain of the patient,” explained first author Helen Fong, PhD, who is also a California Institute for Regenerative Medicine postdoctoral scholar. “By comparing these diseased neurons with the ‘genetically corrected’ healthy neurons, we could see—cell by cell—how the Tau mutation leads to the abnormal build up of Tau and, over time, neuronal degeneration and death.”
“Tau’s main functions include keeping the skeletal structure of individual neurons intact and regulating neuronal activity,” said Dr. Huang. “But our research showed that the Tau produced by neurons from people with the Tau mutation is different; so it is red-flagged by the cell and targeted for destruction. However, instead of being flushed out, Tau gets chopped into pieces. These potentially toxic fragments accumulate over time and may in fact cause the neuron to degenerate and die.”
But by correcting the Tau mutation, the team effectively removed Tau’s red flag. The protein remained in one piece, the abnormal buildup ceased and the neurons remained healthy. Ongoing studies aim to determine whether the abnormal fragmentation and buildup of mutant tau is really the main cause of the neuronal death and, if so, how to block it.
Finding a way to block this toxic buildup of tau fragments has been a key focus of drug development—but has thus far been unsuccessful. But Dr. Huang and his colleagues are optimistic that their approach could be exactly what researchers need to fight back against deadly tauopathies.
“These findings not only offer a glimpse into how these powerful new models can shed light on mechanisms of disease” said Dr. Miller, “They may also prove invaluable for screening potential drugs that could be developed into better treatments for Alzheimer’s disease, FTD and related conditions.”
Scientists Succeed in Growing Human Brain Tissue in “Test Tubes”
Complex human brain tissue has been successfully developed in a three-dimensional culture system established in an Austrian laboratory. The method described in the current issue of NATURE allows pluripotent stem cells to develop into cerebral organoids – or “mini brains” – that consist of several discrete brain regions. Instead of using so-called patterning growth factors to achieve this, scientists at the renowned Institute of Molecular Biotechnology (IMBA) of the Austrian Academy of Sciences (OeAW) fine-tuned growth conditions and provided a conducive environment. As a result, intrinsic cues from the stem cells guided the development towards different interdependent brain tissues. Using the “mini brains”, the scientists were also able to model the development of a human neuronal disorder and identify its origin – opening up routes to long hoped-for model systems of the human brain.
The development of the human brain remains one of the greatest mysteries in biology. Derived from a simple tissue, it develops into the most complex natural structure known to man. Studies of the human brain’s development and associated human disorders are extremely difficult, as no scientist has thus far successfully established a three-dimensional culture model of the developing brain as a whole. Now, a research group lead by Dr. Jürgen Knoblich at the Institute of Molecular Biotechnology of the Austrian Academy of Sciences (IMBA) has changed just that.
Brain Size Matters
Starting with established human embryonic stem cell lines and induced pluripotent stem (iPS) cells, the group identified growth conditions that aided the differentiation of the stem cells into several brain tissues. While using media for neuronal induction and differentiation, the group was able to avoid the use of patterning growth factor conditions, which are usually applied in order to generate specific cell identities from stem cells. Dr. Knoblich explains the new method: “We modified an established approach to generate so-called neuroectoderm, a cell layer from which the nervous system derives. Fragments of this tissue were then maintained in a 3D-culture and embedded in droplets of a specific gel that provided a scaffold for complex tissue growth. In order to enhance nutrient absorption, we later transferred the gel droplets to a spinning bioreactor. Within three to four weeks defined brain regions were formed.”
Already after 15 – 20 days, so-called “cerebral organoids” formed which consisted of continuous tissue (neuroepithelia) surrounding a fluid-filled cavity that was reminiscent of a cerebral ventricle. After 20 – 30 days, defined brain regions, including a cerebral cortex, retina, meninges as well as choroid plexus, developed. After two months, the mini brains reached a maximum size, but they could survive indefinitely (currently up to 10 months) in the spinning bioreactor. Further growth, however, was not achieved, most likely due to the lack of a circulation system and hence a lack of nutrients and oxygen at the core of the mini brains.
Microcephaly in Mini Brains
The new method also offers great potential for establishing model systems for human brain disorders. Such models are urgently needed, as the commonly used animal models are of considerably lower complexity, and often do not adequately recapitulate the human disease. Knoblich’s group has now demonstrated that the mini brains offer great potential as a human model system by analysing the onset of microcephaly, a human genetic disorder in which brain size is significantly reduced. By generating iPS cells from skin tissue of a microcephaly patient, the scientists were able to grow mini brains affected by this disorder. As expected, the patient derived organoids grew to a lesser size. Further analysis led to a surprising finding: while the neuroepithilial tissue was smaller than in mini brains unaffected by the disorder, increased neuronal outgrowth could be observed. This lead to the hypothesis that, during brain development of patients with microcephaly, the neural differentiation happens prematurely at the expense of stem and progenitor cells which would otherwise contribute to a more pronounced growth in brain size. Further experiments also revealed that a change in the direction in which the stem cells divide might be causal for the disorder.
"In addition to the potential for new insights into the development of human brain disorders, mini brains will also be of great interest to the pharmaceutical and chemical industry," explains Dr. Madeline A. Lancaster, team member and first author of the publication. "They allow for the testing of therapies against brain defects and other neuronal disorders. Furthermore, they will enable the analysis of the effects that specific chemicals have on brain development."
UC Davis team “spikes” stem cells to generate myelin
Findings hold promise for developing regenerative therapies for spinal cord injuries and diseases such as multiple sclerosis
Stem cell technology has long offered the hope of regenerating tissue to repair broken or damaged neural tissue. Findings from a team of UC Davis investigators have brought this dream a step closer by developing a method to generate functioning brain cells that produce myelin — a fatty, insulating sheath essential to normal neural conduction.
“Our findings represent an important conceptual advance in stem cell research,” said Wenbin Deng, principal investigator of the study and associate professor at the UC Davis Department of Biochemistry and Molecular Medicine. “We have bioengineered the first generation of myelin-producing cells with superior regenerative capacity.”
The brain is made up predominantly of two cell types: neurons and glial cells. Neurons are regarded as responsible for thought and sensation. Glial cells surround, support and communicate with neurons, helping neurons process and transmit information using electrical and chemical signals. One type of glial cell — the oligodendrocyte — produces a sheath called myelin that provides support and insulation to neurons. Myelin, which has been compared to insulation around electrical wires that helps to prevent short circuits, is essential for normal neural conduction and brain function; well-recognized conditions involving defective myelin development or myelin loss include multiple sclerosis and leukodystrophies.
In this study, the UC Davis team first developed a novel protocol to efficiently induce embryonic stem cells (ESCs) to differentiate into oligodendroglial progenitor cells (OPCs), early cells that normally develop into oligodendrocytes. Although this has been successfully done by other researchers, the UC Davis method results in a purer population of OPCs, according to Deng, with fewer other cell types arising from their technique.
They next compared electrophysiological properties of the derived OPCs to naturally occurring OPCs. They found that unlike natural OPCs, the ESC-derived OPCs lacked sodium ion channels in their cell membranes, making them unable to generate spikes when electrically stimulated. Using a technique called viral transduction, they then introduced DNA that codes for sodium channels into the ESC-derived OPCs. These OPCs then expressed ion channels in their cells and developed the ability to generate spikes.
According to Deng, this is the first time that scientists have successfully generated OPCs with so-called spiking properties. This achievement allowed them to compare the capabilities of spiking cells to non-spiking cells.
In cell culture, they found that only spiking OPCs received electrical input from neurons, and they showed superior capability to mature into oligodendrocytes.
They also transplanted spiking and non-spiking OPCs into the spinal cord and brains of mice that are genetically unable to produce myelin. Both types of OPCs had the capability to mature into oligo-dendrocytes and produce myelin, but those from spiking OPCs produced longer and thicker myelin sheaths around axons.
“We actually developed ‘super cells’ with an even greater capacity to spike than natural cells,” Deng said. “This appears to give them an edge for maturing into oligodendrocytes and producing better myelin.”
It is well known that adult human neural tissue has a poor capacity to regenerate naturally. Although early cells such as OPCs are present, they do not regenerate tissue very effectively when disease or injury strikes.
Deng believes that replacing glial cells with the enhanced spiking OPCs to treat neural injuries and diseases has the potential to be a better strategy than replacing neurons, which tend to be more problematic to work with. Providing the proper structure and environment for neurons to live may be the best approach to regenerate healthy neural tissue. He also notes that many diverse conditions that have not traditionally been considered to be myelin-related diseases – including schizophrenia, epilepsy and amyotrophic lateral sclerosis (ALS) – actually are now recognized to involve defective myelin.
There’s Life After Radiation for Brain Cells
Johns Hopkins researchers suggest neural stem cells may regenerate after anti-cancer treatment
Scientists have long believed that healthy brain cells, once damaged by radiation designed to kill brain tumors, cannot regenerate. But new Johns Hopkins research in mice suggests that neural stem cells, the body’s source of new brain cells, are resistant to radiation, and can be roused from a hibernation-like state to reproduce and generate new cells able to migrate, replace injured cells and potentially restore lost function.
“Despite being hit hard by radiation, it turns out that neural stem cells are like the special forces, on standby waiting to be activated,” says Alfredo Quiñones-Hinojosa, M.D., a professor of neurosurgery at the Johns Hopkins University School of Medicine and leader of a study described online today in the journal Stem Cells. “Now we might figure out how to unleash the potential of these stem cells to repair human brain damage.”
The findings, Quiñones-Hinojosa adds, may have implications not only for brain cancer patients, but also for people with progressive neurological diseases such as multiple sclerosis (MS) and Parkinson’s disease (PD), in which cognitive functions worsen as the brain suffers permanent damage over time.
In Quiñones-Hinojosa’s laboratory, the researchers examined the impact of radiation on mouse neural stem cells by testing the rodents’ responses to a subsequent brain injury. To do the experiment, the researchers used a device invented and used only at Johns Hopkins that accurately simulates localized radiation used in human cancer therapy. Other techniques, the researchers say, use too much radiation to precisely mimic the clinical experience of brain cancer patients.
In the weeks after radiation, the researchers injected the mice with lysolecithin, a substance that caused brain damage by inducing a demyelinating brain lesion, much like that present in MS. They found that neural stem cells within the irradiated subventricular zone of the brain generated new cells, which rushed to the damaged site to rescue newly injured cells. A month later, the new cells had incorporated into the demyelinated area where new myelin, the protein insulation that protects nerves, was being produced.
“These mice have brain damage, but that doesn’t mean it’s irreparable,” Quiñones-Hinojosa says. “This research is like detective work. We’re putting a lot of different clues together. This is another tiny piece of the puzzle. The brain has some innate capabilities to regenerate and we hope there is a way to take advantage of them. If we can let loose this potential in humans, we may be able to help them recover from radiation therapy, strokes, brain trauma, you name it.”
His findings may not be all good news, however. Neural stem cells have been linked to brain tumor development, Quiñones-Hinojosa cautions. The radiation resistance his experiments uncovered, he says, could explain why glioblastoma, the deadliest and most aggressive form of brain cancer, is so hard to treat with radiation.
(Source: hopkinsmedicine.org)
Study Reveals Genes That Drive Brain Cancer
About 15 percent of glioblastoma patients could receive personalized treatment with drugs currently used in other cancers
A team of researchers at the Herbert Irving Comprehensive Cancer Center at Columbia University Medical Center has identified 18 new genes responsible for driving glioblastoma multiforme, the most common—and most aggressive—form of brain cancer in adults. The study was published August 5, 2013, in Nature Genetics.
“Cancers rely on driver genes to remain cancers, and driver genes are the best targets for therapy,” said Antonio Iavarone, MD, professor of pathology and neurology at Columbia University Medical Center and a principal author of the study.
“Once you know the driver in a particular tumor and you hit it, the cancer collapses. We think our study has identified the vast majority of drivers in glioblastoma, and therefore a list of the most important targets for glioblastoma drug development and the basis for personalized treatment of brain cancer.”
Personalized treatment could be a reality soon for about 15 percent of glioblastoma patients, said Anna Lasorella, MD, associate professor of pediatrics and of pathology & cell biology at CUMC.
“This study—together with our study from last year, Research May Lead to New Treatment for Type of Brain Cancer—shows that about 15 percent of glioblastomas are driven by genes that could be targeted with currently available FDA-approved drugs,” she said. “There is no reason why these patients couldn’t receive these drugs now in clinical trials.”
New Bioinformatics Technique Distinguishes Driver Genes from Other Mutations
In any single tumor, hundreds of genes may be mutated, but distinguishing the mutations that drive cancer from mutations that have no effect has been a longstanding problem for researchers.
An analysis of all gene mutations in nearly 140 brain tumors has uncovered most of the genes responsible for driving glioblastoma. The analysis found 18 new driver genes (labeled red), never before implicated in glioblastoma and correctly identified the 15 previously known driver genes (labeled blue). The graphs show mutated genes that are commonly found in varying numbers in glioblastoma (left), that frequently contain insertions (middle), and that frequently contain deletions (right). Genes represented by blue dots in the graphs were statistically most likely to be driver genes. Image: Raul Rabadan/Columbia University Medical Center.
The Columbia team used a combination of high throughput DNA sequencing and a new method of statistical analysis to generate a short list of driver candidates. The massive study of nearly 140 brain tumors sequenced the DNA and RNA of every gene in the tumors to identify all the mutations in each tumor. A statistical algorithm designed by co-author Raul Rabadan, PhD, assistant professor of biomedical informatics and systems biology, was then used to identify the mutations most likely to be driver mutations. The algorithm differs from other techniques to distinguish drivers from other mutations in that it considers not only how often the gene is mutated in different tumors, but also the manner in which it is mutated.
“If one copy of the gene in a tumor is mutated at a single point and the second copy is mutated in a different way, there’s a higher probability that the gene is a driver,” Dr. Iavarone said.
The analysis identified 15 driver genes that had been previously identified in other studies—confirming the accuracy of the technique—and 18 new driver genes that had never been implicated in glioblastoma.
Significantly, some of the most important candidates among the 18 new genes, such as LZTR1 and delta catenin, were confirmed to be driver genes in laboratory studies involving cancer stem cells taken from human tumors and examined in culture, as well as after they had been implanted into mice.
A New Model for Personalized Cancer Treatment
Because patients’ tumors are powered by different driver genes, the researchers say that a complicated analysis will be needed for personalized glioblastoma treatment to become a reality. First, all the genes in a patient’s tumor must be sequenced and analyzed to identify its driver gene.
“In some tumors it’s obvious what the driver is; but in others, it’s harder to figure out,” said Dr.Iavarone.
Once the candidate driver is identified, it must be confirmed in laboratory tests with cancer stem cells isolated from the patient’s tumor.
About 15 percent of glioblastoma driver genes can be targeted with currently available drugs, suggesting that personalized treatment for some patients may be possible in the near future. Personalized therapy for glioblastoma patients could be achieved by isolating the most aggressive cells from the patient’s tumor and identifying the driver gene responsible for the tumor’s growth (different tumors will be driven by different genes). Drugs can then be tested on the isolated cells to find the most promising candidate. In this image, the gene mutation driving the malignant tumor has been replaced with the normal gene, transforming malignant cells back into normal brain cells. Image: Anna Lasorella.
“Cancer stem cells are the tumor’s most aggressive cells and the critical cellular targets for cancer therapies,” said Dr. Lasorella. “Drugs that prove successful in hitting driver genes in cancer stem cells and slowing cancer growth in cell culture and animal models would then be tried in the patient.”
Personalized Treatment Already Possible for Some Patients
For 85 percent of the known glioblastoma drivers, no drugs that target them have yet been approved.
But the Columbia team has found that about 15 percent of patients whose tumors are driven by certain gene fusions, FDA-approved drugs that target those drivers are available.
The study found that half of these patients have tumors driven by a fusion between the gene EGFR and one of several other genes. The fusion makes EGFR—a growth factor already implicated in cancer—hyperactive; hyperactive EGFR drives tumor growth in these glioblastomas.
“When this gene fusion is present, tumors become addicted to it—they can’t live without it,” Dr. Iavarone said. “We think patients with this fusion might benefit from EGFR inhibitors that are already on the market. In our study, when we gave the inhibitors to mice with these human glioblastomas, tumor growth was strongly inhibited.”
Other patients have tumors that harbor a fusion of the genes FGFR (fibroblast growth factor receptor) and TACC (transforming acidic coiled-coil), first reported by the Columbia team last year. These patients may benefit from FGFR kinase inhibitors. Preliminary trials of these drugs (for treatment of other forms of cancer) have shown that they have a good safety profile, which should accelerate testing in patients with glioblastoma.
Epilepsy in a dish: Stem cell research reveals clues to disease’s origins and possible treatment
A new stem cell-based approach to studying epilepsy has yielded a surprising discovery about what causes one form of the disease, and may help in the search for better medicines to treat all kinds of seizure disorders.
The findings, reported by a team of scientists from the University of Michigan Medical School and colleagues, use a technique that could be called “epilepsy in a dish”.
By turning skin cells of epilepsy patients into stem cells, and then turning those stem cells into neurons, or brain nerve cells, the team created a miniature testing ground for epilepsy. They could even measure the signals that the cells were sending to one another, through tiny portals called sodium channels.
In neurons derived from the cells of children who have a severe, rare genetic form of epilepsy called Dravet syndrome, the researchers report abnormally high levels of sodium current activity. They saw spontaneous bursts of communication and “hyperexcitability” that could potentially set off seizures. Neurons made from the skin cells of people without epilepsy showed none of this abnormal activity.
They report their results online in the Annals of Neurology, and have further work in progress to create induced pluripotent stem cell lines from the cells of patients with other genetic forms of epilepsy. The work is funded by the National Institutes of Health, the American Epilepsy Society, the Epilepsy Foundation and U-M.
The new findings differs from what other scientists have seen in mice — demonstrating the importance of studying cells made from human epilepsy patients. Because the cells came from patients, they contained the hallmark seen in most patients with Dravet syndrome: a new mutation in SCN1A, the gene that encodes the crucial sodium channel protein called Nav1.1. That mutation reduces the number of channels to half the normal number in patients’ brains.
"With this technique, we can study cells that closely resemble the patient’s own brain cells, without doing a brain biopsy," says senior author and team leader Jack M. Parent, M.D., professor of neurology at U-M and a researcher at the VA Ann Arbor Healthcare System. "It appears that the cells are overcompensating for the loss of channels due to the mutation. These patient-specific induced neurons hold great promise for modeling seizure disorders, and potentially screening medications."
With the new paper, Parent, postdoctoral fellow Yu Liu, Ph.D. and their collaborators Lori Isom, Ph.D., professor of Pharmacology and of Molecular and Integrative Physiology at U-M, and Miriam Meisler, Ph.D., Distinguished University Professor of Human Genetics at U-M, report striking discoveries about what is happening at the cell level in the neurons of Dravet syndrome patients with a mutated SCN1A gene.
They also demonstrated that the effect is rooted in something that happens after function of the gene is reduced due to the mutation, though they don’t yet know how or why the nerve cells overcompensate for partial loss of this channel.
And, they found that the neurons didn’t show the telltale signs of hyperexcitability in the first few weeks after they were made — consistent with the fact that children with Dravet syndrome often don’t suffer their first seizures until they are several months old.
"In addition, reproduction of the hyperactivity of epileptic neurons in these cell cultures demonstrates that there is an intrinsic change in the neurons that does not depend on input from circuits in the brain," says co-author Meisler.
A platform for testing medications
Many Dravet patients don’t respond to current epilepsy medications, making the search for new options urgent. Their lives are constantly under threat by the risk of SUDEP, sudden unexplained death in epilepsy – and they never outgrow their condition, which delays their development and often requires round-the-clock care.
"Working with patient families, and translating our sodium channel research to a pediatric disease, has made our basic science work much more immediate and critical," says Isom, who serves on the scientific advisory board of the Dravet Syndrome Foundation along with Meisler. Parent, who co-directs U-M’s Comprehensive Epilepsy Program, was recently honored by the foundation.
The team is now working toward screening specific compounds for seizure-calming potential in Dravet syndrome, by testing their impact on the cells in the “epilepsy in a dish” model. The National Institutes of Health has made a library of drugs that have been approved by the U.S. Food and Drug Administration available for researchers to use — potentially allowing older drugs to have a second life treating an entirely different disease from what they were initially intended.
Parent and his colleagues hope to identify drugs that affect certain aspects of sodium channels, to see if they can dampen the sodium currents and calm hyperexcitability. The team is exploring new techniques that can make this process faster, using microelectrodes and calcium-sensitive dyes. They also hope to use the model to study potential drugs for non-genetic forms of epilepsy.
Having a U-M team that includes experts in induced pluripotent stem cell biology, sodium channel physiology and epilepsy genetics expertise helps the research progress, Parent notes. “Epilepsy is a complicated brain network disease,” he says. “It takes team-based science to address it.”
Patients as part of the research team
The U-M team’s research wouldn’t be possible without the participation of patients with Dravet syndrome and other genetic forms of epilepsy, and their parents.
More than 100 of them have joined the International Ion Channel Epilepsy Patient Registry, which is based at U-M and Miami Children’s Hospital and co-funded by the Dravet Syndrome Foundation and the ICE Epilepsy Alliance. The researchers hope to be able to conduct clinical trials of potential drugs with participation by these patients and others.
Meanwhile, patients with other genetically based neurological diseases can also help U-M scientists discover more about their conditions, by taking part in other efforts to create induced neurons from skin cells. Parent and his team have worked with several other U-M faculty to create stem cell lines from skin cells provided by patients with other diseases including forms of ataxia and lysosmal storage disease.
UC Davis stem cell study uncovers the brain-protective powers of astrocytes
One of regenerative medicine’s greatest goals is to develop new treatments for stroke. So far, stem cell research for the disease has focused on developing therapeutic neurons — the primary movers of electrical impulses in the brain — to repair tissue damaged when oxygen to the brain is limited by a blood clot or break in a vessel. New UC Davis research, however, shows that other cells may be better suited for the task.
Published today in the journal Nature Communications, the large, collaborative study found that astrocytes — neural cells that transport key nutrients and form the blood-brain barrier — can protect brain tissue and reduce disability due to stroke and other ischemic brain disorders.
“Astrocytes are often considered just ‘housekeeping’ cells because of their supportive roles to neurons, but they’re actually much more sophisticated,” said Wenbin Deng, associate professor of biochemistry and molecular medicine at UC Davis and senior author of the study. “They are critical to several brain functions and are believed to protect neurons from injury and death. They are not excitable cells like neurons and are easier to harness. We wanted to explore their potential in treating neurological disorders, beginning with stroke.”
Deng added that the therapeutic potential of astrocytes has not been investigated in this context, since making them at the purity levels necessary for stem cell therapies is challenging. In addition, the specific types of astrocytes linked with protecting and repairing brain injuries were not well understood.
The team began by using a transcription factor (a protein that turns on genes) known as Olig2 to differentiate human embryonic stem cells into astrocytes. This approach generated a previously undiscovered type of astrocyte called Olig2PC-Astros. More importantly, it produced those astrocytes at almost 100 percent purity.
The researchers then compared the effects of Olig2PC-Astros, another type of astrocyte called NPC-Astros and no treatment whatsoever on three groups of rats with ischemic brain injuries. The rats transplanted with Olig2PC-Astros experienced superior neuroprotection together with higher levels of brain-derived neurotrophic factor (BDNF), a protein associated with nerve growth and survival. The rats transplanted with NPC-Astros or that received no treatment showed much higher levels of neuronal loss.
To determine whether the astrocytes impacted behavior, the researchers used a water maze to measure the rats’ learning and memory. In the maze, the rats were required to use memory rather than vision to reach a destination. When tested 14 days after transplantation, the rats receiving Olig2PC-Astros navigated the maze in significantly less time than the rats that received NPC-Astros or no treatment.
The investigators used cell culture experiments to determine whether the astrocytes could protect neurons from oxidative stress, which plays a significant role in brain injury following stroke. They exposed neurons co-cultured with both types of astrocytes to hydrogen peroxide to replicate oxidative stress. They found that, while both types of astrocytes provided protection, the Olig2PC-Astros had greater antioxidant effects. Further investigation showed that the Olig2PC-Astros had higher levels of the protein Nrf2, which increased antioxidant activity in the mouse neurons.
“We were surprised and delighted to find that the Olig2PC-Astros protected neurons from oxidative stress in addition to rebuilding the neural circuits that improved learning and memory,” said Deng.
The investigators also investigated the genetic qualities of the newly identified astrocytes. Global microarray studies showed they were genetically similar to the standard NPC-Astros. The Olig2PC-Astros, however, expressed more genes (such as BDNF and vasoactive endothelial growth factor, or VEGF) associated with neuroprotection. Many of these genes help regulate the formation and function of synapses, which carry signals between neurons.
Additional experiments showed that both the Olig2PC-Astros and NPC-Astros accelerated synapse development in mouse neurons. The Olig2PC-Astros, however, had significantly greater protective effects over the NPC-Astros.
In addition to being therapeutically helpful, the Olig2PC-Astros showed no tumor formation, remained in brain areas where they were transplanted and did not differentiate into other cell types, such as neurons.
“Dr. Deng’s team has shown that this new method for deriving astrocytes from embryonic stem cells creates a cell population that is more pure and functionally superior to the standard method for astrocyte derivation,” said Jan Nolta, director of the UC Davis Institute for Regenerative Cures. “The functional improvement seen in the brain injury models is impressive, as are the higher levels of BDNF. I will be excited to see this work extended to other brain disease models such as Huntington’s disease and others, where it is known that BDNF has a positive effect.”
Deng added that the results could lead to stem cell treatments for many neurodegenerative diseases.
“By creating a highly purified population of astrocytes and showing both their therapeutic benefits and safety, we open up the possibility of using these cells to restore brain function for conditions such as Alzheimer’s disease, epilepsy, traumatic brain disorder, cerebral palsy and spinal cord injury,” said Deng.
Stem cells reprogrammed using chemicals alone
Scientists have demonstrated a new way to reprogram adult tissue to become cells as versatile as embryonic stem cells — without the addition of extra genes that could increase the risk of dangerous mutations or cancer.
Researchers have been striving to achieve this since 2006, when the creation of so-called induced pluripotent (iPS) cells was first reported. Previously, they had managed to reduce the number of genes needed using small-molecule chemical compounds (1, 2), but those attempts always required at least one gene, Oct4.
Now, writing in Science, researchers report success in creating iPS cells using chemical compounds only — what they call CiPS cells.
Hongkui Deng, a stem-cell biologist at Peking University in Beijing, and his team screened 10,000 small molecules to find chemical substitutes for the gene. Whereas other groups looked for compounds that would directly stand in for Oct4, Deng’s team took an indirect approach: searching for small-molecule compounds that could reprogram the cells in the presence of all the usual genes except Oct4.
Then came the most difficult part. When the group teamed the Oct4 replacements with replacements for the other three genes, the adult cells did not become pluripotent, or able to turn into any cell type, says Deng.
Fine-tuning
The researchers tinkered with the combinations of chemicals for more than a year, until they finally found one that produced some cells that were in an early stage of reprogramming. But the cells still lacked the hallmark genes indicating pluripotency. By adding DZNep, a compound known to catalyse late reprogramming stages, they finally got fully reprogrammed cells, but in only very small numbers. One further chemical increased efficiency by 40 times. Finally, using a cocktail of seven compounds, the group was able to get 0.2% of cells to convert — results comparable to those from standard iPS production techniques.
The team proved that the cells were pluripotent by introducing them into developing mouse embryos. In the resulting animals, the CiPS cells had contributed to all major cell types, including liver, heart, brain, skin and muscle.
“People have always wondered whether all factors can be replaced by small molecules. The paper shows they can,” says Rudolf Jaenisch, a cell biologist at the Whitehead Institute for Biomedical Research in Cambridge, Massachusetts, who was among the first researchers to produce iPS cells. Studies of CiPS cells could give insight into the mechanisms of reprogramming, says Jaenisch.
The frog’s secret
The achievement could even help regenerative biologists to work out how amphibians grow new limbs. Deng’s group found that one gene indicative of pluripotency, Sall4, was expressed much earlier in the CiPS-cell reprogramming process than in iPS-cell reprogramming. The same Sall4 involvement is seen in frogs that regenerate a lost a limb: before the regeneration, cells in the limb de-differentiate, a process akin to reprogramming, and Sall4 is active early in that process.
The discovery “provides an important framework to decipher the signalling pathways leading to Sall4 expression” in regulating limb regeneration, says Anton Neff, who studies organ regeneration at Indiana University in Bloomington.
Sheng Ding, a reprogramming researcher at the Gladstone Institutes in San Francisco, California, says that the study marks “significant progress” in the field, but notes that chemical reprogramming is unlikely to be used widely until the team can show that it can work for human cells, not just mouse ones. Other strategies, including one that uses RNA, can complete reprogramming with less risk of disturbing the genes than the original iPS-generation method, and are already in use in humans. Indeed, clinical trials with iPS cells derived through such means are already being planned.
Deng has made some progress towards using his method in human cells, but it will require tweaks. ”Maybe some additional small molecules are needed,” he says.
If it the technique is found to be safe and effective in humans, it could be useful for the clinic. It does not risk causing mutations, and the compounds themselves seem to be safe — four of them are in fact already in clinical use. The small molecules can easily pass through cell membranes, so they can be washed away after they have initiated the reprogramming.