Figure 4.

Putative donor-derived satellite cells in humans after myoblast transplantation. The images correspond to a cross-section of the cell-grafted muscle of a Duchenne muscular dystrophy patient, 1 month after normal myoblast transplantation. We codetected dystrophin (red fluorescence), X chromosomes by FISH (green-fluorescent dots), and nuclei (DAPI: blue fluorescence). (a) A region with several dystrophin⁺ myofibers is shown. This patient had 18.6% dystrophin⁺ myofibers in the myoblast-transplanted muscle and <0.5% dystrophin⁺ myofibers in the contralateral nontransplanted muscle. The area in the rectangle is enlarged in b. (c) The most evident pairs of FISH dots indicating female (donor-derived) nuclei are pointed: arrowheads indicate pairs of FISH dots inside dystrophin⁺ myofibers, and the arrow indicates a pair of FISH dots adjacent to the exterior side of the dystrophin⁺ sarcolemma. The Cy3 fluorescence of dystrophin appears with the filter used here for FITC, and the picture was taken in this form to allow confirming the position of the FISH dots in reference to the sarcolemma. The region containing the pair of FISH dots, circumscribed with white corners in b and c, is enlarged in the insets (d,e). d is a merge of the FISH dots with the dystrophin fluorescence, to better evidence the position of this donor-derived nucleus in a satellite cell position. There is a concavity in the sarcolemma under the FISH dots, which may correspond to the indentation of the myofiber profile produced by satellite cells. (f) In spite of some diffusion, DAPI is shown to indicate the nuclei corresponding to the pairs of FISH dots pointed in c. Bar = 25 µm. DAPI, 4′-6-Diamidino-2-phenylindole; FISH, fluorescent in situ hybridization; FITC, fluorescein isothiocyanate.