Stem Cells from Muscle Can Repair Nerve Damage After Injury


Researchers from the University of Pittsburgh School of Medicine have discovered that stem cells derived from human muscle tissue can repair nerve damage and restore function in an animal model of sciatic nerve injury. These data have been recently published online in the Journal of Clinical Investigation, but more importantly, this work demonstrates the feasibility of cell therapy for certain nerve diseases, such as multiple sclerosis.

Presently there are few treatments for peripheral nerve damage. Peripheral nerve damage can leave patients with chronic pain, impaired muscle control and decreased sensation.

The senior author of this work, Henry J. Mankin, serves as the Chair in Orthopedic Surgery Research, Pitt School of Medicine, and deputy director for cellular therapy, McGowan Institute for Regenerative Medicine, and said, “This study indicates that placing adult, human muscle-derived stem cells at the site of peripheral nerve injury can help heal the lesion. The stem cells were able to make non-neuronal support cells to promote regeneration of the damaged nerve fiber.”

Muscle-derived stem cells

Workers in Mankin’s laboratory, in collaboration with Dr. Mitra Lavasani, assistant professor of orthopedic surgery, Pitt School of Medicine, grew human muscle-derived stem/progenitor cells in culture by using a culture medium suitable for nerve cells. In culture, Lavasani, Mankin and their colleagues found that when these muscle-derived stem cells were grown in the presence of specific nerve-growth factors, they differentiated into neurons and glial cells. Glial cells act as support cells from neurons. One type of glial cell that these muscle-derived stem cells could differentiate into was Schwann cells, which are the cells that form the myelin sheath around the axons of neurons to accelerate the speed at which nerve impulses are conducted.

Schwann Cell

Mankin and his colleagues then injected these human muscle-derived stem/progenitor cells into mice that had a quarter-inch injury in their right sciatic nerve. The sciatic nerve controls right leg movement. Six weeks later, the nerve had fully regenerated in stem-cell treated mice, but the untreated group showed only limited nerve regrowth and functionality. In other tests, 12 weeks after treatments, the stem cell-treated mice were able to keep their treated and untreated legs balanced at the same level while being held vertically by their tails. When the treated mice ran through a special maze, analyses of their paw prints showed that their gait, which had been abnormal, was now completely normal. Finally, treated and untreated mice experienced loss of muscle mass after nerve damage, but only the stem cell-treated mice regained normal muscle mass by 72 weeks after nerve damage.

sciatic-nerve

“Even 12 weeks after the injury, the regenerated sciatic nerve looked and behaved like a normal nerve,” Dr. Lavasani said. “This approach has great potential for not only acute nerve injury, but also conditions of chronic damage, such as diabetic neuropathy and multiple sclerosis.”

Drs. Huard and Lavasani and the team are now trying to understand how the human muscle-derived stem/progenitor cells triggered injury repair. They are also developing delivery systems, such as gels, that could hold the cells in place at larger injury sites.

The co-authors of this paper included Seth D. Thompson, Jonathan B. Pollett, Arvydas Usas, Aiping Lu, Donna B. Stolz, Katherine A. Clark, Bin Sun, and Bruno Péault, all of whom are from the University of Pittsburgh.

Teaching Old Neural Stem Cells New Tricks


In our brains, cells called neurons produce nerve impulses and are responsible for thinking, learning memory, reasoning, and so on. Neurons do not exist in isolation, but in combination with cells called glial cells that support the neurons, nourish them, and protects them from stress damage. Neurons and glial cells are replenished by brain-specific neural stem cell populations in the brain.

Unfortunately, the neural stem cell population in our brains tends to produce far fewer neurons as they age. This deficit of new neurons can play a role in the onset of neurodegenerative diseases such as Alzheimer’s disease and Parkinson’s disease. Also, our own “senior moments” when we forget where we placed our iPod or car keys comes from a loss of neurons as we age.

Fortunately, some recent research might change this trend. A team from Japan’s Keio University, and the Riken national research institute, has reported the discovery of a small RNA molecule (micro-RNA) that controls neuron production in young mice. When this micro-RNA was manipulated in older mice, their neural stem cells started to make neurons again. The Japanese team also has reasons to believe that the same mechanism is at work in human brains as well. This research was reported in the journal Proceedings of the National Academies of Science. The mechanism is believed to exist in humans as well.

Senior Author Hideyuki Okano said, “We observed the neurogenic-to-gliogenic switching in developing NSCs.” Translation: Okano and his team examined embryonic mouse brains and their neural stem cell (NSC) populations. They found what many other groups have previously observed: that the developing embryonic brain NSCs create neurons first, then switch over to making glial cells later. Okano’s team also discovered the microRNA-17/106-p38 axis that is responsible for this initial neuron-to-glial cell switch during embryonic development.

When they manipulated this embryonic microRNA-17/106-p38 pathway in older, post-natal NSCs in culture, these older post-natal NSCs switched from making glial cells to producing neurons.

In culture, NSCs are difficult to control, since getting large supplies of neurons from cell cultures that various research groups call NSCs is very difficult.

Nevertheless, “there is general agreement that neurogenesis (make neurons) largely precedes gliogenesis (making glial cells) during CNS development in vertebrates,” Okano explained. And adult NSCs, according to Okano, clearly can produce neurons in the body, “whereas they exhibit strong gliogenic characteristics under culture conditions in vitro (that is, in the laboratory).”

Adult NSCs in two regions of the brain—the subventricular zone and hippocampus—also “make neurons, even though transplant studies have shown us that the adult CNS is a gliogenic environment.”

Subventricular Zone

So it seems clear that old NSCs can make neurons, at least under certain conditions. However, it is very difficult to determine the age at which NSCs begin making substantially more glial cells than neurons. According to Okano, “It is difficult to clearly explain the association between total glial cell number and changes in NSC abilities. Moreover, there is less evidence about gliogenic ability of aged NSCs because most of studies about NSCs have mainly focused on the neurogenic ability. “

Still, Okano says: “There are some reports about decline of neurogenesis ability of NSCs with age. These reports indicate that reduction in paracrine Wnt3 factors, and increase of (chemokine) CCL11 concentration in blood, impaired adult neurogenesis in the hippocampus, for example.”

Could the group’s microRNA approach improve memory in humans? Okano believes so, but says more work needs to be done.

“We observed the neurogenic effect by overexpression of miR-17 in primary cultured neurospheres” – spheres of a variety of cells, including NSCs—“derived from the SVZ at postnatal day 30. Similar phenomenon by overexpression of miR-106b-25 cluster has been reported by another group.”

Okano also warns that his approach has only been attempted in cultured cells. He cautioned, “There is no evidence using knock-out mice. Therefore, the functions of them in adult neurogenesis and learning/memory functions are still unclear.”

Next, Okano’s group will develop “a useful method for precise manipulation of cytogenesis from NSCs. “

However, he says, “we think that further understanding of basic molecular mechanisms underlying the neural development is also an important issue.” He will study the ways in which his microRNA system interacts with other glia-producing genes. He wants to fully understand the mechanisms underlying “the end of neurogenic competence and acquisition of gliogenic competence.”

Finally, the group will “examine the significance of miR-17/p38 pathway in various somatic stem cells other than NSCs,” he says.

How Neural Stem Cells Become Neurons and Glia


How do neural stem cells differentiate into neurons or glia? A new paper from researchers at the University of California, Los Angeles (UCLA) seeks to explain this very phenomenon.

Neurons serve as the conductive cells of the nervous system. They transmit electrochemical signals from one neuron to another and provide signals to muscles, glands, and so on. They are responsible for consciousness, thought, learning and memory, and personality.

Despite their immense utility, neurons are not the only cells in the nervous system. Glial cells or just glia support neurons, hold them in place, and supply neurons with oxygen and nutrients and protect them from pathogens.

Glial Cells

When mouse neural stem cells were grown in culture, Wange Lu, associate professor of biochemistry and molecular biology at the Keck School of Medicine, and his colleagues came upon a protein called SMEK1 that promotes the differentiation of neural stem and progenitor cells. SMEK1 also keeps neural stem cells in check by preventing them from dividing uncontrollably.

When Lu and others took a more detailed look at the role of SMEK1, they discovered that it does not work alone, but in concert with a protein called Protein Phosphatase 4 (PP4) to suppress the function of a third protein called PAR3. PAR3 discourages the birth of new neurons (neurogenesis), and PAR3 inhibition leads to the differentiation of neural stem progenitor cells into neurons and glia.

“These studies reveal the mechanisms of how the brain keeps the balance of stem cells and neurons when the brain is formed,” said Wange. “If this process goes wrong, it leads to cancer, or mental retardation or other neurological diseases.”

Neural stem and progenitor cells offer tremendous promise as a future treatment for neurodegenerative disorders, and understanding their differentiation is the first step towards co-opting the therapeutic potential of these cells. This could offer new treatments for patients who suffer from Alzheimer’s, Parkinson’s and many other currently incurable diseases.

This work is interesting. It was published in Cell Reports 5, 593–600, November 14, 2013. My only criticism of some of the thinking in this paper is that neural stem cell lines are usually made from aborted fetuses. I realize that some of these neural stem cell lines come from medical abortions in which the baby had already died, but many of them come from aborted babies. If we are going to use neural stem cells for therapeutic purposes, then we should make them from induced pluripotent stem cells and take them from aborted babies.

Studying Tough-to-Examine Disease by Using Brain Cells Made from Stem Cells


Diseases that are hard to study, such as Alzheimer’s, schizophrenia, and autism can be examined more safely and effectively thanks to an innovative new method for making mature brain cells from reprogrammed skin cells. Gong Chen, the Verne M. William Chair in Life Sciences and professor of biology at Penn State University and the leader of the research team that designed this method said this: “The most exciting part of this research is that it offers the promise of direct disease modeling, allowing for the creation, in a Petri dish, of mature human neurons that behave a lot like neurons that grow naturally in the human brain.”

Chen’s method could lead to customized treatment for individual patients that are based on their own genetic and cellular profile. Chen explained it this way: “Obviously we do not want to remove someone’s brain to experiment on, so recreating the patient’s brain cells in a Petri dish is the next best thing for research purposes and drug screening.”

In previous work, scientists at the University of Wisconsin in James Thomson’s laboratory and in Shinya Yamanaka’s laboratory at Kyoto University in Kyoto, Japan discovered a way to reprogram adult cells into pluripotent stem cells. Such stem cells are called induced pluripotent stem cells or iPSCs. To make iPSCs, scientists infect adult cells with genetically engineered viruses that introduce four specific genes (OCT4, SOX2, KLF4 and cMYC for those who are interested). These genes encode transcription factors, which are proteins that bind to DNA or to the machinery that directly regulates gene expression.  These transcription factors turn on those genes (e.g., OCT4, NANOG, REX1, DNMT3β and SALL4, and OCT4) that induce pluripotency, which means the ability to form any adult cell type.  Once in the pluripotent state, iPSCs can be cultured and grown life embryonic stem cells and can differentiate into adult cell types and tissues.

As Chen explained, “A pluripotent stem cell is a kind of blank slate.”  Chen continued, “During development, such stem cells differentiate into many diverse specialized cell types, such as a muscle cell, a brain cell, or a blood cell.  So, after generating iPSCs from skin cells, researchers then can culture them to become brain cells, or neurons, which can be studies safely in a Petri dish.”

Chen’s team invented a protocol to differentiate iPSCs into mature human neurons much more effectively than previous protocols.  This generates cells that behave neurons in our own brains and can be used to model the individualized disease of a single patient.

In the brain, neurons rarely work alone, but instead are usually in close proximity to star-shaped cells called astrocytes.  Astrocytes are very abundant cells and they assist neuron function and mediate neuronal survival.  “Because neurons are adjacent to astrocytes in the brain, we predicted that this direct physical contact might be an integral part of neuronal growth and health,” said Chen.  To test this hypothesis, Chen and his colleagues began by culturing iPSCs-derived neural stem cells, which are stem cells that have the potential to become neurons.  These cells were cultured on top of a one-cell-thick layer of astrocytes sop that the two cell types were physically touching each other.

Astrocytes
Astrocytes

“We found that these neural stem cells cultured on astrocytes differentiated into mature neurons much more effectively,” Chen said.  This contrasts Chen’s method with other neural stem cells that were cultured alone in a Petri dish.  As Chen put it, the astrocytes seems to be “cheering the stem cells on, telling them what to do, and helping them to fulfill their destiny to become neurons.”

While this sounds a little cheesy, it is undeniable that the astrocyte layer increases the efficiency of neuronal differentiation of iPSCs.  Personalized medicine is moving beyond the gene level, to the level of cellular organization and tissue physiology, and iPSCs are showing the way.

Induced Pluripotent Stem Cells do not Form Neural Stem Cells as well as Embryonic Stem Cells


Induced pluripotent stem cells show tremendous promise for regenerative medicine. However in a February 15th article in the Proceedings of the National Academy of Sciences, showed that induced pluripotent stem cells (iPSCs) were inefficient at forming the cells of the brain in comparison to their embryonic stem cell counterparts.

The senior author of the article , Su-Chun Zhang, (professor, University of Wisconsin-Madison School of Medicine and Public Health), said:  “Embryonic stem cells can pretty much be predicted,” and “Induced cells cannot. That means that at this point there is still some work to be done to generate ideal induced pluripotent stem cells for application.”

This study compared the ability of five different embryonic stem cell lines to 12 different iPSC lines to form nerve cell precursors.  Embryonic stem cells are considered the “gold standard” for all pluripotent stem cells, which are cells that can differentiate into all of the 220 cell types in the human body.  Zhang’s group found that the induced cells differentiated into progenitor neural cells and further into the different kinds of functional neurons that make up the brain, but they did not faithfully reproduce all the differentiation capabilities of embryonic stem cells.  This suggests that there are unknown factors at play that may limit the use of iPSCs when it comes to modeling diseases in the laboratory.  Such unknowns would also limit their use in clinical settings for such things as cell transplants.

Despite their unpredictability, Zhang notes that iPSCs can still be used to make pure populations of specific types of cells, which makes them useful for some applications like testing potential new drugs for efficacy and toxicity.  Zhang also noted that the limitations identified by his group are technical issues likely to be resolved relatively quickly.  “It appears to be a technical issue,” said Zhang.  “Technical things can usually be overcome,” he added.

This is very possibly a technical issue that is due to our inability to properly manipulate iPSCs to form nerve cells.  However, if the same protocols that drive embryonic stem cells to form nerve cells are used on iPSCs, they only form nerve cells poorly.  There are probably other protocols that can do just this.  We just haven’t found them yet.

Also, it is worth mentioning, that the ability of iPSCs to differentiate into neurons is probably a line-specific property.  Therefore if these lines to not form lines effectively, then perhaps other lines do.