Cell Transplantation and “Stem Cell Therapy” in the Treatment of Myopathies: Many Promises in Mice, Few Realities in Humans
Figure 6
One of the techniques developed by the author for the intramuscular transplantation of myoblasts in nonhuman primates is illustrated. In this case, the protocol implies matrices of parallel close intramuscular injections, which is done to deliver myogenic cells the most homogeneously as possible throughout muscle regions that do not receive another treatment than the cell injections. For small to medium-sized regions, this is easily done with a Hamilton syringe attached to a repeating dispenser (a). The pattern of cell injections is controlled with a sterile transparent dressing with a 5 mm grid, adhered on the cell-grafted region (the biceps brachii of a macaque in the picture). The black arrow indicates the repetitive movements of penetration and subsequent removal of the needle, during which cells are injected by pressing the button on the device. After one month, the fusion of the grafted cells with the recipient’s myofibers is analyzed in cross sections of muscle biopsies, using histological techniques to detect the transgenic proteins that labeled the grafted cells ((b) and (c)). In this case, a cross section of a monkey muscle is grafted with transgenic myoblasts labeled with a microdystrophin coupled with a V5 peptide tag (μDys-V5). Hybrid myofibers expressing proteins coded by the myonuclei of the grafted cells are observed by fluorescent immunodetection of the V5 peptide tag. The distribution of the hybrid myofibers in “bands of engraftment” (b) reproduces the pattern of the original cell-injection trajectories (indicated by the yellow arrow). At a higher magnification (c), the labeling is essentially sarcoplasmic according to the normal location of dystrophin, although some intracellular staining is also observed. A serial section of (c) stained with hematoxylin and eosin is shown (d) to illustrate the structure of the muscle. Some of the V5-positive myofibers exhibit internal nuclei, indicating recent regeneration. Scale bars: 1 mm (b) and 100 μm ((c) and (d)).
