Stem Cell Clinic

Scientists give star treatment to lesser-known cells crucial for brain development – Seacoastonline.com

After decades of relative neglect, star-shaped brain cells called astrocytes are getting their due. To gather insight into a critical aspect of brain development, a team of scientists examined the maturation of astrocytes in 3-D structures grown in culture dishes to resemble human brain tissue. The study, which confirms the lab-grown cells develop at the same rate as those found in human brains, was published in Neuron and funded in part by the National Institutes of Healths National Institute of Neurological Disorders and Stroke.

This work addresses a significant gap in human brain research by providing an invaluable technique to investigate the role of astrocytes in both normal development and disease, said NINDS program director Jill Morris, Ph.D.

In 2015, a team directed by Dr. Sergiu Pasca, an assistant professor of psychiatry and behavioral science at Stanford University in California, and Dr. Ben Barres, Ph.D., a Stanford professor of neurobiology, published a method for taking adult skin cells, converting them to induced pluripotent stem cells, and then growing them as 3-D clusters of brain cells called human cortical spheroids (hCSs). These hCSs, which closely resemble miniature versions of a particular brain region, can be grown for many months. The cells in the cluster eventually develop into neurons, astrocytes, and other cells found in the human brain.

One of the challenges of studying the human brain is the difficulty of examining it at different stages of development, Dr. Pasca said. This is a system that tries to simulate brain development step by step.

In the new study, Steven Sloan, a student in Stanfords M.D./Ph.D. program, led a series of experiments comparing astrocytes from hCSs to those found in tissue from the developing and adult human brain. The team grew the hCSs for 20 months, one of the longest-ever studies of lab-grown human brain cells.

The results verified that the lab-grown cells change over time in a similar manner to cells taken directly from brain tissue during very early life, a critical time for brain growth. This process is considered critical for normal brain development and deviations are thought to cause a variety of neurological and mental health disorders, such as schizophrenia and autism. Creating hCSs using cells from patients could allow scientists to uncover the underlying developmental biology at the core of these disorders.

The hCS system makes it possible to replay astrocyte development from any patient, Dr. Barres said. Thats huge. Theres no other way one could ever do that without this method.

The current study showed that hCS-grown astrocytes develop at the same rate as those found in human brains, in terms of their gene activity, their shapes, and their functions. For example, astrocytes taken from hCSs that were less than six months old multiplied rapidly and were highly engaged in eliminating unnecessary connections between neurons, just like astrocytes in babies growing in the womb. But astrocytes grown in hCSs for more than nine months could not reproduce and removed significantly fewer of those connections, mirroring astrocytes in infants 6 to 12 months old. On the other hand, just like astrocytes from developing and adult brains, the early- and late-stage astrocytes from hCSs were equally effective at encouraging new connections to form between neurons.

Astrocytes are not just bystanders in the brain, Dr. Pasca said. Theyre not just there to keep neurons warm; they actually participate actively in neurological function.

Since astrocytes make up a greater proportion of brain cells in humans than in other species, it may reflect a greater need for astrocytes in normal human brain function, with more significant consequences when they dont work correctly, added David Panchision, Ph.D., program director at the National Institute of Mental Health (NIMH), which also helped fund the study.

The researchers caution that hCSs are only a model and lack many features of real brains. Moreover, certain genes that are active in fully mature astrocytes never switched on in the hCS-grown astrocytes, which they could conceivably do if the cells had more time to develop. To address this question, the researchers now hope to identify ways to produce mature brain cells more quickly. hCSs could also be used to scrutinize precisely what causes astrocytes to change over time and to screen drugs that might correct any differences that occur in brain disease.

These are questions that are going to be very exciting to explore, Dr. Barres said.

The study was funded by NINDS, the National Institute of Mental Health, the National Institute of General Medical Sciences, the National Center for Advancing Translational Sciences, the California Institute of Regenerative Medicine, the MQ Fellow Award, and Stanford University.

The NINDS is the nations leading funder of research on the brain and nervous system. The mission of NINDS is to seek fundamental knowledge about the brain and nervous system and to use that knowledge to reduce the burden of neurological disease.

The mission of the NIMH is to transform the understanding and treatment of mental illnesses through basic and clinical research, paving the way for prevention, recovery and cure. For information, visit the NIMH website.

The National Institute of General Medical Sciences supports basic research that increases understanding of biological processes and lays the foundation for advances in disease diagnosis, treatment and prevention. For information, visit the NIGMS website.

The National Center for Advancing Translational Sciences (NCATS) was established to transform the translational process so that new treatments and cures for disease can be delivered to patients faster. For information, visit the NCATS website.

The National Institutes of Health, the nation's medical research agency, encompasses 27 Institutes and Centers and is a component of the U.S. Department of Health and Human Services. NIH is the primary federal agency conducting and supporting basic, clinical, and translational medical research, and is investigating the causes, treatments, and cures for both common and rare diseases. For information about NIH and its programs, visit http://www.nih.gov.

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Scientists give star treatment to lesser-known cells crucial for brain development - Seacoastonline.com

Breakthrough in Gene Editing Comes as Scientists Correct Disease-Causing Mutation in Human Embryo – TrendinTech

Its an exciting time for gene editing currently as scientists achieve the first ever safe repair of a single-gene mutation in a human embryo. Using the CRISPR-Cas9 system, medical professionals used the technique to correct the mutation for a heart condition. The procedure was carried out early enough in the embryonic development so that the defect wouldnt be passed down to further generations.

The study is an important breakthrough and could pave the way for improved in vitro fertilization (IVF) outcomes further down the line as well as finding cures for even a few of the thousands of diseases caused by single gene mutations. Juan Carlos Izpisua Belmonte, a professor in Salks Gene Expression Laboratory and corresponding author of the paper commented that, Gene editing is still in its infancy so even though this preliminary effort was found to be safe and effective, it is crucial that we continue to proceed with the utmost caution, paying the highest attention to ethical considerations. Although gene editing has become relatively easy to carry out for scientists these days, they do still proceed with much caution. This is partly to avoid introducing any mutations into the germ line.

*Hypertrophic cardiomyopathy (HCM) affects around 1 in every 500 people and is the most common cause of sudden death in otherwise healthy, young athletes. A dominant mutation in the MYBPC3 gene is the cause of it and theres a 50 percent chance of a carrier passing it to their offspring. If the mutation in the embryo was corrected it would prevent the disease in children as well as their descendants. During the study, researchers produced induced pluripotent stem cells from a sample taken from a male with HCM. They also developed a CRISPR-Cas9 gene editing strategy that was able to target specifically the mutated copy of the MYBPC3 gene for repair. The MYBPC3 gene was cut by the Cas9 enzyme allowing the mutation to be fixed in the next round of cell division. Researchers then used IVF techniques to inject top gene-editing components into healthy donor eggs which had been newly fertilized. They were then able to monitor just how well the mutation was repaired.

This method proved to be both extremely safe and efficient, much to the surprise of the researchers. No strange mutations were induced and a high number of embryonic cells were repaired. They also developed a way in which they could ensure the repair happened consistently in all of the embryonic cells which is a bonus as spotty repairs have been known to cause mutations. Even though the success rate in patient cells cultured in a dish was low, we saw that the gene correction seems to be very robust in embryos of which one copy of the MYBPC3 gene is mutated, states Salk staff scientists and one of the papers first authors, Jun Wu. Our technology successfully repairs the disease-causing gene mutation by taking advantage of a DNA repair response unique to early embryos. However, it is still early days and both Izpisua Belmonte and Wu agree further research is required to ensure no unintended effects occur.

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Fertile offspring produced from sterile mice using iPS cells – Kyodo News Plus

Researchers at Kyoto University have succeeded in producing fertile offspring from sterile mice with chromosome abnormality by using iPS cells, the university said Friday.

The research outcome by the international team was published online by the U.S. magazine Science with the headline "Fertile offspring from sterile sex chromosome trisomic mice."

The team intentionally produced sterile trisomic mice and showed that fibroblasts from the abnormal mice lose the extra sex chromosome during reprogramming to induced pluripotent stem cells, or iPS cells.

The team termed the phenomenon trisomy-biased chromosome loss.

It is vital to have the correct number of chromosomes for normal development and health.

The researchers have successfully produced fertile offspring with a usual pair of sex chromosome by injecting functional sperm originated from the euploid iPS cells into eggs.

"(The finding) could lead to the development of treatment for infertility caused by chromosome or other genetic abnormalities," said Michinori Saito, a professor of the Graduate School of Medicine at Kyoto University and a research team member.

The team also involves James Turner of the Francis Crick Institute in Britain.

Sex chromosome trisomy, associated with infertility, affects 0.1 percent of the human population, according to the research team.

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Fertile offspring produced from sterile mice using iPS cells - Kyodo News Plus

Brain Spheroids Hatch Mature Astrocytes | ALZFORUM – Alzforum

18 Aug 2017

Astrocytes are more than bystanders in neurotransmissionthey take an active role in synaptic activity. However, their functions are hard to study because the cells are difficult to grow in vitro and its hard to coax them to mature from progenitors. Now, researchers from the labs of Sergiu Paca and Ben Barres, both at Stanford University School of Medicine, California, report that astrocytes come of age in spherical balls of human brain cells cultured in a dish for almost two years. As reported in the August 16 Neuron, these astrocytes develop much like those from real brains, undergoing similar transcriptomic, morphologic, and functional changes. Studying the processes involved in this astrocyte maturation will help researchers understand neurodevelopmental disorders such as autism and schizophrenia, researchers say, and might even shed light on problems in adultbrains.

That these 3D cultures can be maintained for such a long time allows us to capture an interesting transition in astrocytes, said Paca. We are starting to appreciate aspects of human brain development to which we would not otherwise haveaccess.

The breakthrough is that they can develop human astrocytes very close to maturity in their 3D culture models, said Doo Yeon Kim, Massachusetts General Hospital, Charlestown, who uses 3D culture models to study pathological process that occur in Alzheimers disease. Some researchers are using 3D cultures to model other neurodegenerative disorders, such as ALS, and still others are planning to use cultured astrocytes for cell therapy. If astrocytes are not mature enough in culture, patterns [we see] may not be the same as in the diseased brain, saidKim.

This developing human astrocyte (red), which comes from a 350-day-old cortical spheroid, is taking shape as a mature cell. [Image courtesy of Sloan et al.Neuron]

A few years back, Pacas group developed a method for differentiating human induced pluripotent stem cells (hiPSCs) into a 3D culture of brain cells. They used special dishes that the cells could not easily attach to, coaxing them to stick to each other instead. Under these conditions the iPSCs balled up into neural spheroids that grew to about 4 mm in diameter. A cocktail of growth factors early on encouraged them to form excitatory pyramidal cells like those in the cortex, and the cells spontaneously organized into layers. These cortical spheroids survived a year or more and spontaneously grew astrocytes in addition to neurons (Paca et al., 2015). Not long after, the Barres lab reported that astrocytes in the adult human brain look different from those isolated from fetuses. They called the latter astrocyte progenitor cells (APCs). Each had their own transcriptional patterns and functions (Jan 2016 news). Together, Barres and Paca wondered if it was possible to see the APCs morph into mature astrocytes in these long-lived corticalspheroids.

To find out, first author Steven Sloan and colleagues examined spheroids generated from iPSCs derived from healthy human fibroblasts. Sloan grew the spheroids for about 20 months. Along the way, he took samples, isolated the astrocytes, and compared them to those isolated from fetal and postnatal humanbrain.

At about 100 days in culture, astrocytes began to sprout spontaneously from within the mostly neuronal milieu of the cortical spheroids. At first, these cells were simple, adorned by few branches and expressing genes akin to those active in APCs. But as the spheroids reached about 250 days, the astrocytes therein looked more mature, having numerous processes. After this point, APC gene expression tapered off and the astrocytes started producing proteins typical of matureastrocytes.

Astrocytes also underwent functional changes as they matured. Early versions divided in fast and furious fashion, much like their counterparts from the fetal tissue. That division slowed as the spheroids aged. Dividing APCs dropped from 35 percent of all astrocytes at day 167 to 3 percent at day 590. Taken from the spheroids at day 150 and cultured in a 2D layer, immature astrocytes also harbored a voracious appetite for added synaptosomes, much like immature astrocytes recently characterized in mice (see image below; Dec 2013 conference news on Chung et al., 2013). However, that hunger waned as astrocytes approached the 590-daymark.

At the older end of the spectrum, mature astrocytes seemed to take on a supportive role, strengthening calcium signaling in nearbyneurons.

Studying the neurons and astrocytes in these cortical spheroids could be useful for addressing certain unanswered questions about human biology, said other researchers. This could be a very strong opportunity to understand what goes wrong in human genetic disorders that affect astrocyte function, said M. Kerry OBanion, University of Rochester Medical Center, New York. Its also possible that such cultures could reveal as yet unknown facets of familial mutations that cause Alzheimers disease, he suggested. However, given that these cultures take a long time to grow and develop, they are unlikely to completely supplant other types of cultures or faster-maturing animal models, hesaid.

Kim agreed, saying, The results are very exciting, but not practical yet for disease modeling." However, Kim hopes that researchers will make progress on accelerating the maturationprocess.

The Barres and Paca labs are trying just that with the spheroid. They will also analyze what they secrete to support neuronal signaling. In addition, they are exploring how to make the astrocytes reactive, as they often are in neurodegenerative diseases, such as Alzheimers. Doing so might reveal how such astrocytes interact withneurons.

An immature astrocyte taken from a 150-day-old spheroid gobbles up added synaptosomes (red). [Neuron, Sloan et al.2017]

To Pacas knowledge, these cortical spheroids are some of the longest human cell cultures ever reported. His group has continued to cultivate these clumps, with the oldest still going strong at day 850. Granted, these systems are missing many cell types: endothelial cells, oligodendrocytes, and microglia to name a few, he said. However, his lab has introduced new ways to add in other cells. Earlier this year, he reported 3D cultures of cortical glutamatergic neurons and GABAergic interneurons that fused together when they were placed side-by-side (Birey et al., 2017).

Clive Svendsen, Cedars-Sinai Medical Center in Los Angeles, California, saw clinical implications for this paper. It shows iPSC derived astrocytes can mature to an adult phenotype, he said. This further supports their use in clinical transplantation, as we are planning to do. His group has begun a Phase 1 clinical trial that implants human fetal astrocytes into the spinal cords of ALS patients.Gwyneth DickeyZakaib

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Brain Spheroids Hatch Mature Astrocytes | ALZFORUM - Alzforum