The first sign can appear all of a sudden: a movement that was previously natural such as the simple gesture of stretching out your hand to get a glass of water becomes difficult, as if the arm were locked. Over the course of months or years, the muscles increasingly begin to fail, to the point where they atrophy. In the end, the only movements that a person with amyotrophic lateral sclerosis can control are those of their eyes, as is the case of British physicist Stephen Hawking, who for more than 40 years has lived with this illness, which usually results in a person’s death within a decade. Although it manifests itself in the muscles, the actual problem lies in the degeneration of the motor neurons, which are responsible for muscular contractions and the body’s voluntary movements. Experiments carried out over the last few years with stem-cells have helped to understand the disease, but do not point to stem-cell treatment possibilities. “Can you imagine replacing a cell at the base of the spinal column that goes all the way down to your toes?” protests Brazilian biologist Maria Carolina Marchetto, a researcher at the Salk Institute in the United States. “This is a very complicated path and the cell is unable to find a way.” The December issue of the international journal Cell Stem Cell presents the researcher’s most recent contribution to understanding this disease, which it is estimated to claim 15 new victims daily in the United States.
Carol’s team, which includes another Brazilian, Alysson Muotri, along with the American Fred Gage, who is head of Salk’s Genetics Laboratory, came up with an innovative model for studying amyotrophic lateral sclerosis in human beings. “What was unprecedented in connection with our research,” explains Carol, “was the use for the very first time of a totally human in vitro model for studying amyotrophic lateral sclerosis”. On plastic sheets, she cultivated motor neurons based on human embryonic stem-cells and astrocytes, star-shaped brain cells that make up the glia – glue in Greek -, the tissue that nourishes and sustains the neurons and gives the brain its structure. This is a major advance because most studies are carried out on mice, but in general the drugs that work for mice do not have the same effect on humans.
This system makes it possible to study how mutations in specific genes give rise to different manifestations of amyotrophic lateral sclerosis. One of the mutations – the one studied by Carol – produces a deficient version of the protein SOD1, which fails to be metabolized and ends up accumulating in the motor neurons. Like a long, narrow corridor that is turned into a warehouse, the debris in the neurons blocks the flow of nutrients and signaling substances to the extremity that stimulates the muscle.
As a result, the neuron is unable to perform its function and it dies. And the muscle atrophies. The disease is not limited to neurons. In the last few years, the group of neuro-scientist Don Cleveland, from the University of California in San Diego, has shown that when the mutation occurs in the glia cells they produce substances that create an inflammatory reaction, which triggers the immune system. “The defense cells migrate to the nervous system and attack the neurons, killing them,” explains the biologist. This discovery was good news in terms of the outlook for treatment, since there are no drugs that can reestablish the health of the motor neuron. Recent studies carried out by other groups showed that if the support cells are healthy, the affected neurons live longer. The best alternative seems to be to keep the immune system from attacking the neuron terminals, by means of immune suppressor medication and antioxidants.
It is a therapeutic aboutface. The most effective medicine up until now against the disease, riluzole, eliminates the neurotransmitter glutamate that accumulates in the connections between the neurons in operation, a function usually performed by healthy astrocytes. But it does not do much to prolong the patient’s life. The San Diego based group tested five antioxidant compositions and managed to reduce the oxidizing activity of the cells. The most promising drug, apocynin, does indeed seem to improve the survival of the neurons in culture with mutation-bearing astrocytes.
The next step will be to build more complex in-vitro models, which should be three-dimensional and have a greater diversity of brain cells and even blood vessels. Therefore, the hope is to reproduce in a lab conditions that are as close as possible to those that assail people with the disease. These models should also make it possible to test the effect of the mutations responsible for other varieties of amyotrophic lateral sclerosis.
Studying SOD1 has been advantageous, although it only accounts for 2% of all cases. Another mutation was identified by the geneticist Mayana Zatz’s group from the University of São Paulo (USP) and described in 2004 in the American Journal of Human Genetics. It alters the protein VAP-B, which plays an essential role in transporting substances within the cells. Since then, a number of research groups have published papers setting out in detail the workings of the protein. It is already known that the mutation makes the protein insoluble; it ends up forming clusters within the cells. “The most important thing is that the identification of this protein, which seems to intervene in various types of amyotrophic lateral sclerosis, makes it clear that apparently rare genes can help unmask common pathological mechanisms,” explains the geneticist.
Mayana has put a lot of effort into understanding the disease better. Miguel Mitne-Neto, one of her doctoral students, is studying VAP-B’s interaction with other proteins. In an article published in 2007 in the journal Protein Expression and Purification, he showed that the mutation reduces VAP-B’s affinity with two other proteins at play in the brain – tubulin and GAPDH – which were seen to perform inadequately in other neurodegenerative diseases. The new model developed by Carol and Muotri comes in handy both for gaining a better understanding of how these proteins act in the brain and for testing possible treatments. “We are already in contact with them so that we can collaborate in this new phase of the study,” declares Mayana.
MARCHETTO, M. C. N. et al. Non-cell-autonomous effect of human SOD1 astrocytes on motor neurons derived from human embryonic stem cells. Cell Stem Cell, v. 3, no. 6, Dec. 2008.