Brainpower applied to understanding of neural stem cells

New Strategy to Treat Multiple Sclerosis Shows Promise in Mice

Scientists at The Scripps Research Institute (TSRI) have identified a set of compounds that may be used to treat multiple sclerosis (MS) in a new way. Unlike existing MS therapies that suppress the immune system, the compounds boost a population of progenitor cells that can in turn repair MS-damaged nerve fibers.

One of the newly identified compounds, a Parkinson’s disease drug called benztropine, was highly effective in treating a standard model of MS in mice, both alone and in combination with existing MS therapies.

“We’re excited about these results, and are now considering how to design an initial clinical trial,” said Luke L. Lairson, an assistant professor of chemistry at TSRI and a senior author of the study, which is reported online in Nature on October 9, 2013.

Lairson cautioned that benztropine is a drug with dose-related adverse side effects, and has yet to be proven effective at a safe dose in human MS patients. “People shouldn’t start using it off-label for MS,” he said.

A New Approach

An autoimmune disease of the brain and spinal cord, MS currently affects more than half a million people in North America and Europe, and more than two million worldwide. Its precise triggers are unknown, but certain infections and a lack of vitamin D are thought to be risk factors. The disease is much more common among those of Northern European heritage, and occurs about twice as often in women as in men.

In MS, immune cells known as T cells infiltrate the upper spinal cord and brain, causing inflammation and ultimately the loss of an insulating coating called myelin on some nerve fibers. As nerve fibers lose this myelin coating, they lose their ability to transmit signals efficiently, and in time may begin to degenerate. The resulting symptoms, which commonly occur in a stop-start, “relapsing-remitting” pattern, may include limb weakness, numbness and tingling, fatigue, vision problems, slurred speech, memory difficulties and depression, among other problems.

Current therapies, such as interferon beta, aim to suppress the immune attack that de-myelinates nerve fibers. But they are only partially effective and are apt to have significant adverse side effects.

In the new study, Lairson and his colleagues decided to try a complementary approach, aimed at restoring a population of progenitor cells called oligodendrocytes. These cells normally keep the myelin sheaths of nerve fibers in good repair and in principle could fix these coatings after MS damages them. But oligodendrocyte numbers decline sharply in MS, due to a still-mysterious problem with the stem-like precursor cells that produce them. “Oligodendrocyte precursor cells (OPCs) are present during progressive phases of MS, but for unknown reasons don’t mature into functional oligodendrocytes,” Lairson said.

A 100,000-Molecule Screen

Using a sophisticated small-molecule screening laboratory that TSRI manages in conjunction with the California Institute of Regenerative Medicine and in collaboration with the California Institute for Biomedical Research (Calibr), Lairson and his team screened a library of about 100,000 diverse compounds for any that could potently induce OPCs to mature or “differentiate.”

Several compounds scored well as OPC differentiation-inducers. Most were compounds of unknown activity —but one, benztropine, had been well characterized and indeed was already FDA-approved for treating Parkinson’s disease. “That was a surprise, and it meant that we could move forward relatively quickly in testing it,” said graduate student Vishal A. Deshmukh, first author of the paper who performed most of these experiments.

With the help of Brian R. Lawson, a senior author of the paper and assistant professor of immunology at TSRI, and his colleague Research Associate Virginie Tardif, Deshmukh set up tests of benztropine in mice with an induced MS-like autoimmune disease—a model commonly used for testing prospective MS drugs.

In these tests, benztropine showed a powerful ability to prevent autoimmune disease and also was effective in treating it after symptoms had arisen—virtually eliminating the disease’s ability to relapse. Although benztropine on its own worked about as well as existing treatments, it also showed a remarkable ability to complement these existing treatments, in particular two first-line immune-suppressant therapies, interferon-beta and fingolimod.

“Adding even a suboptimal level of benztropine effectively allowed us, for example, to cut the dose of fingolimod by 90%—and achieve the same disease-modifying effect as a normal dose of fingolimod,” said Lawson. “In a clinical setting that dose-lowering could translate into a big reduction in fingolimod’s potentially serious side effects.”

In further analyses, the researchers confirmed that benztropine works against disease in this mouse model by boosting the population of mature oligodendrocytes, which in turn restore the myelin sheaths of damaged nerves—even as the immune attack continues. “The benztropine-treated mice showed no change in the usual signs of inflammation, yet their myelin was mostly intact, suggesting that it was probably being repaired as rapidly as it was being destroyed,” said Lawson.

Benztropine is known to have multiple specific effects on brain cells, including the blocking of activity at acetylcholine and histamine receptors and a boosting of activity at dopamine receptors. But Lairson and his colleagues found evidence that the drug stimulates OPCs to differentiate mainly by blocking M1 or M3 acetylcholine receptors on these cells.

In addition to setting up initial clinical trials, Lairson and his team hope to learn more about how benztropine induces OPC maturation, and how its molecular structure might be optimized for this purpose. “We’re also looking at some of the other, relatively unknown molecules that we identified in our initial screen, to see if any of those has better clinical potential than benztropine,” he said.

“This work, like our previous studies with hematopoietic and mesenchymal stem cells, illustrates the power of small molecules to control stem and precursor cells in ways that may ultimately lead to a new generation of drugs for regenerative medicine,” said Peter G. Schultz, the Scripps Family Chair Professor in the Department of Chemistry at TSRI and one of the study’s senior authors.

Same musicians: brand new tune

A small ensemble of musicians can produce an infinite number of melodies, harmonies and rhythms. So too, do a handful of workhorse signaling pathways that interact to construct multiple structures that comprise the vertebrate body. In fact, crosstalk between two of those pathways—those governed by proteins known as Notch and BMP (for Bone Morphogenetic Protein) receptors—occurs over and over in processes as diverse as forming a tooth, sculpting a heart valve and building a brain.

A new study by Stowers Institute for Medical Research Investigator Ting Xie, Ph.D., reveals yet another duet played by Notch and BMP signals, this time with Notch calling the tune. That work, published in this week’s online issue of PNAS, uses mouse genetics to demonstrate how one Notch family protein, Notch2, shapes an eye structure known as the ciliary body (CB), most likely by ensuring that BMP signals remain loud and clear.

In vertebrates, the CB encircles the lens and performs two tasks essential for normal vision. First, it contains a tiny muscle that reshapes the lens when you change focus, or “accommodate”. And it also secretes liquid aqueous humor into the front compartment of the eye where it likely maintains correct eye pressure. Understanding CB construction is critical, as excessive pressure is one risk factor for glaucoma.

Eye development is a relatively new field for Xie, a recognized leader in the study of adult stem cells in the fruit fly: only recently did he branch out into mouse studies. “A few years ago I was asked to participate in a think tank-type meeting to discuss the potential application of cell therapy to treat glaucoma,” he says.
“I became interested in using retinal progenitor cells to treat diseases like glaucoma or macular degeneration. But I realized that first we needed to understand eye disease at the molecular level.” The new study is an important step in that direction.

Previously, investigators knew that once cells that form the CB are established in an embryo, the BMP pathway drives their “morphogenesis”, the term used by developmental biologists to describe the process of expanding and then sculpting a committed population of cells into a unique structure. “The Notch2 receptor was previously shown to be expressed in the developing mouse eye,” explains Chris Tanzie, M.D., Ph.D., a former graduate student in the Xie lab and the study’s co-first author. “But its function was unknown, and no one connected how various signaling pathways direct CB morphogenesis.”

To determine what Notch2 was doing in the developing eye, the Stowers team constructed a conditional knockout mouse, meaning that the Notch2 gene is deleted from the genome only in eye cells that give rise to the CB. In normal newborn mice a series of cellular “folds” that characterize the CB emerges over the first 7 days of life. But the mutant knockout mice showed a complete absence of folds, dramatic evidence that Notch2 is required to elaborate a CB.

Furthermore, in normal mice a protein called Jagged-1, which activates Notch2, was expressed in cells adjacent to Notch2-expressing CB cells during the same developmental period. Strikingly, the team’s collaborators in Richard Libby’s laboratory at the University of Rochester Medical Center, were able to demonstrate that just like the Notch2 mutants, Jagged-1 conditional knockout mice showed almost total loss of CB fold structures, a major hint that Notch2 was switched on by Jagged1 to drive CB formation.

Biochemical and microarray analysis provided further explanation for defects observed after Notch2 loss. Comparison of normal and Notch2-mutant eye cells revealed that not only did cells of mutant mice lose BMP signaling but that expression of two proteins known to interfere with BMP increased in those cells.

“Up-regulation of BMP antagonists following Notch2 loss is an important observation,” says Xie. “In other systems people often observe that Notch and BMP cooperatively regulate common targets by transcription factor collaboration at the transcriptional level, but this is a unique mechanism. We find that Notch2 keeps BMP signaling active by inhibiting its inhibitors.”

The study’s second co-first author is Yi Zhou, a University of Kansas Medical Center graduate student earning his Ph.D. in Xie’s lab. “Our work reveals a novel link between Notch and BMP pathways potentially involved in the pathogenesis of glaucoma,” says Zhou, noting one more tantalizing implication of the paper. “In addition, mutations in Jagged-1 and Notch2 are thought to underlie the human genetic disease known as Alagille Syndrome. Our work may lead to a better understanding of both.”

Alagille Syndrome is an inherited childhood disorder causing defects in organ systems including liver, heart and the skeleton. Xie is equally intrigued by potential connections between his group’s observations in the mouse eye and Alagille outcomes in humans. Nonetheless, he remains focused on nailing down how perturbation of the Jagged1-Notch2-BMP axis might cause eye disease.

“We now know how to build better mouse mutants to study CB development. In this work we show that Notch regulates BMP signaling but have not yet determined whether alterations in CB structure actually change interocular pressure,” he says. “Answering that question is our future goal.”

Serotonin Mediates Exercise-Induced Generation of New Neurons

Mice that exercise in running wheels exhibit increased neurogenesis in the brain. Crucial to this process is serotonin signaling. These are the findings of a study by Dr. Friederike Klempin, Daniel Beis and Dr. Natalia Alenina from the research group led by Professor Michael Bader at the Max Delbrück Center (MDC) Berlin-Buch. Surprisingly, mice lacking brain serotonin due to a genetic mutation exhibited normal baseline neurogenesis. However, in these serotonin-deficient mice, activity-induced proliferation was impaired, and wheel running did not induce increased generation of new neurons. (Journal of Neuroscience)

Scientists have known for some time that exercise induces neurogenesis in a specific brain region, the hippocampus. However, until this study, the underlying mechanism was not fully understood. The hippocampus plays an important role in learning and in memory and is one of the brain regions where new neurons are generated throughout life.

Serotonin facilitates precursor cell maturation

The researchers demonstrated that mice with the ability to produce serotonin are likely to release more of this hormone during exercise, which in turn increases cell proliferation of precursor cells in the hippocampus. Furthermore, serotonin seems to facilitate the transition of stem to progenitor cells that become neurons in the adult mouse brain.

For Dr. Klempin and Dr. Alenina it was surprising that normal baseline neurogenesis occurs in mice that, due to a genetic mutation, cannot produce serotonin in the brain. However, they noted that some of the stem cells in serotonin-deficient mice either die or fail to become neurons.

Yet, these animals seem to have a mechanism that allows compensation for the deficit, in that progenitor cells, an intermediate stage in the development from a stem cell to a neuron, divide more frequently. According to the researchers, this is to maintain the pool of these cells.

However, the group of wheel-running mice that do not produce serotonin did not exhibit an exercise-induced increase in neurogenesis. The compensatory mechanism failed following running. The researchers concluded: “Serotonin is not necessarily required for baseline generation of new neurons in the adult brain, but is essential for exercise-induced hippocampal neurogenesis.”

Hope for new approaches to treat depression and memory loss in the elderly

Deficiency in serotonin, popularly known as the “molecule of happiness”, has been considered in the context of theories linking major depression to declining neurogenesis in the adult brain. “Our findings could potentially help to develop new approaches to prevent and treat depression as well as age-related decline in learning and memory,” said Dr. Klempin and Dr. Alenina.

To Make Mice Smarter, Add A Few Human Brain Cells

For more than a century, neurons have been the superstars of the brain. Their less glamorous partners, glial cells, can’t send electric signals, and so they’ve been mostly ignored.

Now scientists have injected some human glial cells into the brains of newborn mice. When the mice grew up, they were faster learners. The study, published Thursday in Cell Stem Cell, not only introduces a new tool to study the mechanisms of the human brain, it supports the hypothesis that glial cells — and not just neurons — play an important role in learning.

The scientific obsession with neurons really began at the end of the 19th century. Spanish anatomy professor Santiago Ramon y Cajal used a special dye to stain brain tissue. Under the microscope, neurons were revealed in exquisite detail. “A dense forest,” Ramón y Cajal called it — a field of little branching cells that would soon be named neurons.

With beautiful ink drawings, Ramón y Cajal painstakingly mapped neural networks and slowly developed the theory that neurons are the telegraph lines of thought (an idea later embraced by Schoolhouse Rock). Every idea and memory — every aspect of learning — could be traced back to the electric signals sent between neurons. Ramón y Cajal won the Nobel Prize for his work, and scientists focused on neurons for the next century.

But neurons aren’t the only cells in the brain.

"We’ve overlooked half the brain," says Douglas Fields, a neuroscientist at the National Institutes of Health. "We’ve only been studying one kind of cell in the brain." The other kind of cell — glial cells — are at least as abundant as neurons. But early scientists thought they were so boring they didn’t even merit a singular noun. "Glia is plural — there is no singular," Fields says. "We have ‘neuron’ but we don’t have ‘glion.’ "

Glial cells lacked the ability to send electric signals, and most scientists thought they were housekeeping cells, helping provide nutrients and insulation.

It was only in the last decade or so that scientists realized glial cells were more than that. Special types of glial cells, called astrocytes, which are named for the star-like patterns of their cellular structure, have their own form of chemical signaling. They have the potential to coordinate whole groups of neurons. “Glia are in a position to regulate the flow of information through the brain,” Fields says. “This is all missing from our models.”

And there’s something else. This type of glial cell, these astrocytes, have changed a lot as humans have evolved, while neurons have pretty much stayed the same. A mouse neuron and a human neuron look so much alike, even experienced neuroscientists can’t tell them apart.

"I can’t tell the differences between a neuron from a bird or a mouse or a primate or a human," says Steve Goldman, a neuroscientist at the University of Rochester who has studied brain cells for decades. But Goldman says glial cells are easy to tell apart.

"Human glial cells — human astrocytes — are much larger than those of lower species," he says. "They have more fibers and they send those fibers out over greater distances."

The thought is maybe these glial cells have played a role in making humans smarter. So Goldman teamed up with this wife, Maiken Nedergaard, to test this idea.

They injected some human glial cells into the brains of newborn mice. The mice grew up, and so did the human glial cells. The cells spread through the mouse brain, integrating perfectly with mouse neurons and, in some areas, outnumbering their mouse counterparts. All the while Goldman says the glial cells maintained their human characteristics.

"They very much thought that they were in the human brain, in terms of how they developed and integrated," he says.

So what are these mice like, the ones with brains full of functioning human cells? Their neural circuitry is still just the same, so they act completely normal. They still socialize with other mice and still seem interested in mousey things.

But the researchers say these mice are measurably smarter. In classic maze tests, they learn faster. “They make many fewer errors, and it takes them less time to come to the appropriate answer,” Goldman says.

It might take a normal mouse four or five attempts to learn the correct route, for example. But a mouse with human brain cells could get it on the second try. Glial cells — those boring glial cells — somehow enhance learning.

In fact, they could be changing what it means to be a mouse, and that raises ethical questions for this kind of research.

"Maybe bioethicists have been a little bit too cavalier assuming that a mouse with some human brain cells in it is just your normal old mouse," says Robert Streiffer, a bioethicist from the University of Wisconsin-Madison. "Well, it’s not going to be human, but that doesn’t mean it’s a normal old mouse either."

Streiffer says it’s not just that these mice can get through a maze more quickly — they’re better at recognizing things that scare them. And perception of fear is one of the things bioethicists must weigh when they decide the types of experiments you can do on an animal.

"So you have to sort of step back and do some hardcore philosophy," he says. Like, will these types of human-animal hybrids eventually get close enough to humanity that we would feel uncomfortable performing experiments on them?

The researchers in this study say we’re really, really far from that point. And if you want to investigate the role of glial cells, these hybrid mice are the best tools available.

Microglia controls neuron production as brain develops

In a surprise breakthrough, researchers at the UC Davis MIND Institute and their colleagues have found that microglia remove healthy neural progenitor cells (NPCs) through phagocytosis to control neuron production during brain development. This newly discovered mechanism keeps neuron numbers in check, preventing brain overgrowth.

The discovery could open up new avenues for brain research and lead to therapies for a variety of neurological conditions.

The study was published online in the The Journal of Neuroscience.

Microglia are the immune component cell of the central nervous system. Similar to macrophages, microglia provide the brain’s primary defense against pathogens and foreign bodies, clear away dying cells and help repair neural damage. When inactive, they act as sentinels. When a problem is located, they activate and eliminate it. However, until recently, no one had realized the important roles they play in brain development.

"We have known for some time that neurons can undergo apoptosis, a form of cell death, and ultimately be removed by microglia," said Stephen Noctor, assistant professor in the Department of Psychiatry and Behavioral Sciences and the study’s lead author. "But this is new. Microglia are actually eating healthy progenitor cells, thereby regulating the number of neurons produced in the developing brain."

During development, NPCs produce neurons in the brain’s proliferative zones. However, creating too many or too few neurons can have dire consequences.

"If you have too many cells, there’s only so much trophic support (brain infrastructure for cell growth and survival) to keep neurons alive," Noctor said. "All these cells competing for resources could easily throw off connectional properties, altering the way surviving neurons interact. Likewise, having too few cortical cells would have profoundly negative consequences."

(Image: Antoine Triller, Alain Bessis & Serge Marty - Département de Biologie, ENS)

Now hear this: Researchers identify forerunners of inner-ear cells that enable hearing

Researchers at the Stanford University School of Medicine have identified a group of progenitor cells in the inner ear that can become the sensory hair cells and adjacent supporting cells that enable hearing. Studying these progenitor cells could someday lead to discoveries that help millions of Americans suffering from hearing loss due to damaged or impaired sensory hair cells.

“It’s well known that, in mammals, these specialized sensory cells don’t regenerate after damage,” said Alan Cheng, MD, assistant professor of otolaryngology. (In contrast, birds and fish are much better equipped: They can regain their sensory cells after trauma caused by noise or certain drugs.) “Identifying the progenitor cells, and the cues that trigger them to become sensory cells, will allow us to better understand not just how the inner ear develops, but also how to devise new ways to treat hearing loss and deafness.”

The research was published online Feb. 26 in Development. Cheng is the senior author. Former medical student Taha Jan, MD, and postdoctoral scholar Renjie Chai, PhD, share lead authorship of the study. Roel Nusse, PhD, a professor of developmental biology, is a co-senior author of the research.

The inner ear is a highly specialized structure for gathering and transmitting vibrations in the air. The auditory compartment, called the cochlea, is a snail-shaped cavity that houses specialized cells with hair-like projections that sense vibration, much like seaweed waving in the ocean current. These hair cells are responsible for both hearing and balance, and are surrounded by supporting cells that are also critical for hearing.

Twenty percent of all Americans, and up to 33 percent of those ages 65-74, suffer from hearing loss. Hearing aids and, in severe cases, cochlear implants can be helpful for many people, but neither address the underlying cause: the loss of hair cells in the inner ear. Cheng and his colleagues identified a class of cells called tympanic border cells that can give rise to hair cells and the cells that support them during a phase of cochlear maturation right after birth.

“Until now, these cells have had no clear function,” said Cheng. “We used several techniques to define their behavior in cell culture dishes, as well as in mice. I hope these findings will lead to new areas of research to better understand how our ears develop and perhaps new ways to stimulate the regeneration of sensory cells in the cochlea.”

Monell scientists identify elusive taste stem cells

Scientists at the Monell Center have identified the location and certain genetic characteristics of taste stem cells on the tongue. The findings will facilitate techniques to grow and manipulate new functional taste cells for both clinical and research purposes.

"Cancer patients who have taste loss following radiation to the head and neck and elderly individuals with diminished taste function are just two populations who could benefit from the ability to activate adult taste stem cells," said Robert Margolskee, M.D., Ph.D., a molecular neurobiologist at Monell who is one of the study’s authors.

Taste cells are located in clusters called taste buds, which in turn are found in papillae, the raised bumps visible on the tongue’s surface.

Two types of taste cells contain chemical receptors that initiate perception of sweet, bitter, umami, salty, and sour taste qualities. A third type appears to serve as a supporting cell.

A remarkable characteristic of these sensory cells is that they regularly regenerate. All three taste cell types undergo frequent turnover, with an average lifespan of 10-16 days. As such, new taste cells must constantly be regenerated to replace cells that have died.

For decades, taste scientists have attempted to identify the stem or progenitor cells that spawn the different taste receptor cells. The elusive challenge also sought to establish whether one or several progenitors are involved and where they are located, whether in or near the taste bud.

Drawing on the strong physiological relationship between oral taste cells and endocrine (hormone producing) cells in the intestine, the Monell team used a marker for intestinal stem cells to probe for stem cells in taste tissue on the tongue.

Stains for the stem cell marker, known as Lgr5 (leucine-rich repeat-containing G-protein-coupled receptor 5), showed two patterns of expression in taste tissue. The first was a strong signal underlying taste papillae at the back of the tongue and the second was a weaker signal immediately underneath taste buds in those papillae.

The Monell scientists hypothesize that the two levels of expression could indicate two different populations of cells. The cells that more strongly express Lgr5 could be true taste stem cells, whereas those with weaker expression could represent those stem cells that have begun the transformation into functional taste cells.

Additional studies revealed that the Lgr5-expressing cells were capable of becoming any one of the three major taste cell types.

The findings are published online in the journal Stem Cells.

"This is just the tip of the iceberg," said senior author Peihua Jiang, Ph.D., also a Monell molecular neurobiologist. "Identification of these cells opens up a whole new area for studying taste cell renewal, and contributes to stem cell biology in general."

Future studies will focus on identifying the factors that program the Lgr5-expressing cells to differentiate into the different taste cell types, and explore how to grow these cells in culture, thus providing a renewable source of taste receptor cells for research and perhaps even clinical use.

(Image: Getty)

For brain tumors, origins matter

Cancers arise when a normal cell acquires a mutation in a gene that regulates cellular growth or survival. But the particular cell this mutation happens in—the cell of origin—can have an enormous impact on the behavior of the tumor, and on the strategies used to treat it.

Robert Wechsler-Reya, Ph.D., professor and director of the Tumor Development Program in Sanford-Burnham’s NCI-designated Cancer Center, and his team study medulloblastoma, the most common malignant brain cancer in children. A few years ago, they made an important discovery: medulloblastoma can originate from one of two cell types: 1) stem cells, which can make all the different cell types in the brain or 2) neuronal progenitor cells, which can only make neurons.

Stem cells and progenitor cells are regulated by different growth factors. So, Wechsler-Reya thought, maybe the tumors arising from these cells respond differently to different therapies…

In a study published recently in the journal Oncogene, he and his team show that this is indeed the case. They looked at one growth factor in particular—basic fibroblast growth factor (bFGF)—and found that while it induces stem cell growth, it also inhibits neuronal progenitor cell growth.

What’s more, the researchers discovered that bFGF also blocks the growth of tumors that originate from progenitors. When they treated a mouse model of medulloblastoma with bFGF, it dramatically inhibited tumor growth.