RSRT Advisor Makes Significant Discovery

Glia in Rett Syndrome: New Findings

Gail Mandel’s new work reveals not only the important information that MECP2 is expressed in glia as well as in neurons, but the discovery that MECP2-deficient astrocytes (a subset of glia) seem to effectively stunt the development of neurons.

Neuroglia, Nursemaids to the Neurons

Parents of children with Rett Syndrome have become accustomed to hearing about MECP2 in neurons, but the very large world of glia may be new to them. Glial cells, which are found throughout the nervous system, comprise the vast majority of cells in the brain, where they outnumber neurons ten to one. They support and interact with neurons in innumerable ways, ranging from structural underpinnings and guidance of the neurons’ processes, which are the axons and dendrites that carry information, to creating protective insulation (myelin) for these processes, to bringing in the groceries and taking out the garbage for the neurons. There is constant “cross-talk” between neurons and surrounding glial cells. The health of the neuron requires healthy responses and support from the glia. There are several different types of glial cells, including astrocytes, oligodendrocytes, and microglia. We will be focusing in this interview on the astrocytes, so named because of their star-like shape.

Ruining the Neighborhood

In the majority of Rett cases, X inactivation results in neurons functioning with a “good” copy of MECP2 scattered among a fairly equal number of neurons with faulty, mutated MECP2. Until now, any special influence the glia might exert on these mixtures of good and faulty neurons, living side by side, was unknown. We now have a very new perspective: even the neurons that aren’t crippled by mutated MECP2 may be, instead, poisoned by malfunctioning astrocytes carrying a faulty copy of MECP2.

Restoration

In vitro experiments show that damaged neurons can recover normal growth when surrounded by healthy astrocytes.

An interview with Gail Mandel, Ph.D.

Gail Mandel, an advisor to RSRT, is a Howard Hughes Medical Institute investigator at the Vollum Institute . She discusses this new work and sudden turn in Rett research in a conversation with Monica Coenraads, Executive Director of the Rett Syndrome Research Trust.

MC: Congratulations on this significant step forward in understanding the far-reaching influence of MECP2, and of the pathology of Rett Syndrome. Until now, it was thought that MECP2 was not expressed in glia. What motivated you to explore glial function in Rett Syndrome?

GM: I got involved in this research from a different perspective all together. My previous work was dealing with neuronal repressors during neuronal differentiation. I’ve worked for a long time on a repressor protein called REST. This is a repressor which is a master regulator of the neuronal phenotype. It ensures that only neurons express neuronal genes, by keeping neuronal genes turned off outside of the nervous system. REST is present for a brief time during the earliest stages of development, including at the embryonic stem cell (ESC) stage. As the ESC differentiates into neurons, REST is lost, and other regulating genes take over.
In stages, our involvement with REST led us toward MECP2 as an important regulator in neuronal maturation. We began to study MECP2, and through the work of Adrian Bird, Huda Zoghbi, Rudolf Jaenisch and others, were introduced to Rett Syndrome.
What didn’t make a lot of sense to us is that MECP2 is found everywhere in the body and yet when you lose it via mutations the deficits are mostly neurological. So we started thinking what else does the brain have that other tissues don’t have, and one thing is glia. My research assistant, Dr. Nurit Ballas and I, decided to look more carefully at MECP2 in glia, and to do this we developed a new and very sensitive antibody which can detect even small amounts of MECP2 protein in glial cells. When I first started telling fellow scientists that we saw MECP2 in glia they said, “So what?” My response: If defective MECP2 severely compromises glial function, it’s a big “So what!”

MC: Your work suggests that a neuron with normal MECP2 will nevertheless be crippled if the surrounding glial cells cannot support normal growth. Can you elaborate?

GM: We found that MECP2 is present in all kinds of glia. There is more known about the affects of astrocytes on neurons than other types of glia. Astrocytes help neuronal synapses function properly. We’ve known for a long time that astrocytic conditioned medium keeps neurons healthy. (note: “Astrocytic conditioned medium” is comprised of the substances secreted by astrocytes.)
We saw that wildtype (healthy) neurons co-cultured with MECP2 mutant astrocytes exhibited very stunted growth.

MC: Are you saying that, in effect, a healthy neuron may be poisoned by malfunctioning glial cells?

GM: There are two possibilities that could explain what we are seeing. 1) The astrocytes are secreting a toxic factor. This is the scenario that I’m hoping for because it could provide a therapeutic intervention – one could imagine finding a way to neutralize the factor. 2) The astrocytes are depleting a nutrient that the neurons need. This would be a more difficult scenario to deal with.
We did a classic experiment called the mixing experiment: we combined, half and half, conditioned medium from wildtype astrocytes with conditioned medium from the Rett mutant astrocytes. The interpretation is that if normal neurons in the half and half medium resemble neurons cultured in the mutant medium, it’s probably due to a toxic factor. If the astrocytes were depleting a nutrient you would expect that adding in more wildtype medium would help the problem. In our mixing experiment the neurons reacted as if they were in the straight mutant media. These results strengthen the argument that the astrocytes are secreting a toxic factor.

MC: One of the very fascinating aspects of your work is that an MECP2-deficient neuron seems to recover and produce good axonal and dendritic growth in vitro if supported by normal glia. What are the implications of this finding?

GM: It’s an encouraging finding. In theory it suggests that we could treat Rett, at least in part, by neutralizing the toxic factor or by over-expressing the normal factor that wildtype astrocytes secrete to nurture neurons. Given Adrian Bird’s 2007 reversal paper and our culture experiments we would predict that the abnormal phenotype of the neurons should be reversible.
It’s important to note that neutralizing the toxic factor would be a treatment strategy that is not directed at the MECP2 gene itself and would represent a novel approach.

MC: Daniel Lioy, a graduate student in your lab, shared a theory that resonated with me. A girl with Rett is born with roughly 50% wildtype neurons and 50% MECP2– mutated neurons. For a while (sometimes many months) she seems okay. Then the stagnation and regression kick in. Daniel theorizes that perhaps as the toxic factor from the mutated astrocytes poisons the whole neighborhood (wildtype cells as well as mutated cells) the child’s symptoms worsen.

GM: It’s a plausible theory but until we prove it…it’s just a theory. Our lab is currently undertaking several key experiments to further elucidate this proposition as well as to delve more deeply into the question of a toxic factor.

MC: There have been sporadic reports through the years of abnormal findings in the peripheral nervous system in Rett Syndrome. It’s conceivable these are related to some of Rett’s many symptoms. Is there any basis for examining the glial cells of the peripheral nervous system for MECP2 expression?

GM: Yes, I think it’s very important, and we are looking at this, as well.

MC: What are your impressions of the research progress in Rett during the last few years?

GM: I think it’s been really very good, especially when you consider the fact that Rett is a very complicated disease that has a fairly small, albeit growing, community of scientists. There are phenomenal people working in this field and they are making progress. And we don’t just have the Rett scientists in our court; we also have all the other basic scientists working on how genes are regulated in different cells. The body is linked by common solutions to cell functions. Science sometimes works by surprise discoveries that often are fundamental principles that govern how the nervous system works.

MC: Rett shares characteristics of so many neurological disorders, from autism to Parkinson’s. With this new understanding of glial involvement, are there even more disease models where research interests might overlap?

GM: Recent data suggests that toxic astrocytic secreted factors are at the heart of ALS and spinocerebellar ataxia type 7. In fact, it may turn out that the secreted factors may be similar. One could envision a scenario where the sick neurons (whether from an MECP2 mutation or other mutations that cause ALS or SCA7) cause, in turn, the astrocytes to secrete a factor in response to the original insult to the brain. This factor need not be specific to the disease.
I truly hope that my lab can contribute in a meaningful way to the development of treatments. I know the children are in dire need.

MC: On behalf of everyone who loves a child with Rett Syndrome I congratulate you on your discoveries. I look forward to your continued contributions as this work unfolds.