She and her team are exploring the causes of muscle diseases. Evidence shows that satellite cells are active in people with severe muscle diseases such as Duchenne muscular dystrophy, a severe genetic disease leading to muscle degeneration. "But at some point," she adds, "the reservoir is depleted of muscle stem cells and muscle wasting cannot be stopped." All attempts to rebuild muscle tissue by transplanting satellite cells in patients with this condition have failed in the past. The transplanted cells were not viable. Furthermore, the use of other cells with potential to regenerate muscle cells has shown little success. But how would it be possible to use the body's own self-renewal potential?, the researchers asked themselves.
After due approval and informed consent, Spuler and Marg obtained specimens of fresh thigh muscle tissue from patients between 20 and 80 years of age from neurosurgeons of Helios Klinikum Berlin-Buch, located close to the ECRC. From these biopsy specimens, the researchers dissected more than 1000 muscle fiber fragments, each about 2-3 millimeters long. Remarkably, the researchers found the number of stem cells in the individual tissue specimens to be independent of the age of the donor, and thousands of myoblasts developed from a small number of satellite cells. After further developmental steps, these fuse into muscle fibers.
Prof. Spuler and her co-workers cultivated the muscle fiber fragments with the satellite cells, initially for up to three weeks. During this time, the satellite cells increased by 20- to 50-fold, but numerous connective tissue cells also developed in these cultures. To prevent this, the researchers concurrently subjected the muscle fragments to oxygen deprivation (hypoxia) and to cooling (hypothermia) at 4 degrees Celsius. Under these conditions, only satellite cells are able to survive in their stem cell niche, in contrast to the connective tissue cells. "Apparently, the satellite cells receive the proper nutrients in their own 'local milieu'," Dr. Marg explains.
Next, the ECRC researchers have succeeded for the first time in showing how human satellite cells can be cultivated and proliferated while retaining their regeneration potential for several weeks. This is an important step for using the patient's own cells for therapeutic purposes.
The ECRC researchers also tried their approach in mice whose muscle regeneration had been inhibited by irradiation. The researchers grafted the muscle fragments containing satellite cells into the tibalis anterior muscle. They found that the muscles of animals that had been treated with these fiber fragments regenerated particularly well.
However, a genetic muscle disease cannot be successfully treated alone by transplanting muscle fragments. Prof. Spuler: "The idea is to equip the satellite cells additionally with a healthy gene that repairs the defective gene and then to transfect it with the aid of a non-viral 'gene taxi' into the muscles to be treated." In a first experiment with a 'reporter gene' in the Petri dish, Spuler and her team proved that this is basically possible. The reporter gene fluoresces green when it is transfected into the satellite cell. As gene taxi the researchers use the Sleeping Beauty transposon – a jumping gene that can change its position in the genome. This transposon technique was developed several years ago by Dr. Zsuzsanna Izsvák at the MDC and Dr. Zoltán Ivics from the Paul Ehrlich Institute in Frankfurt, and is considered to be a very promising delivery vehicle, or vector, for gene therapy.
Before the method developed by Prof. Spuler and her group can be used to benefit patients, some hurdles remain to be taken. So far, the transplantation has succeeded in small mice muscles. In clinical trials, the scientists and physicians want to determine whether this technique can be used in large human thigh muscles that may be severely altered due to a muscular disease. (© Max Delbrück Center for Molecular Medicine MDC, AcademiaNet)