The fusion index (FI) was calculated as the percentage of nuclei fused within myotubes/total nuclei. Cell transplantation in vivo and engraftment analyses 100,000 MCAM+CD82+ or MCAM+ cells were injected IM in opposite TA muscles of immune-deficient dystrophic Dansylamide recipients. dystrophy patients, together suggesting that CD82 function may be important for muscle stem cell function in muscular disorders. Graphical Abstract Introduction The muscular dystrophies are progressive disorders affecting children and adults (Chelly and Desguerre, 2013; Mercuri and Muntoni, 2013). One example of this class of diseases is usually Duchenne muscular dystrophy (DMD), which is caused by mutations in dystrophin (Monaco et al., 1986), a large cytoplasmic protein located at the sub-sarcolemma of myofibers (Hoffman et al., 1987). Dystrophin functions in muscle by interacting with a group of proteins known collectively as Dansylamide the Dystrophin-Associated Glycoprotein Complex (DAPC) (Yoshida and Ozawa, 1990; Ibraghimov-Beskrovnaya et al., 1992). In the absence of dystrophin, the cellular levels of many DAPC proteins are severely reduced (Ervasti et al., 1990), thus when dystrophin is usually mutated in DMD the function of other proteins is compromised. A second protein complex located at the sarcolemma of myofibers is the 71 integrin. This protein complex is thought to provide membrane stabilization by linking the cytoskeleton Dansylamide to the extracellular matrix (Burkin and Kaufman, 1999). Mutations 7 integrin (7-ITG) cause muscle disease in humans (Mayer et Dansylamide al., 1997; Hayashi et al., 1998). Overexpression of 7-ITG in dystrophic mice, a mouse model for DMD (Bulfield et al., 1984), significantly ameliorates the dystrophic pathology via increased stability of the link between 7-ITG and laminin (Burkin et al., 2005). The tetraspanin sarcospan, an associated member of the DAPC, interacts with the 71 integrin (Marshall et al., 2015), however whether other proteins are also associated with this complex or link the DAPC and 71 integrin protein complexes is not entirely known. In the present study, we demonstrate that this tetraspanin KAI/CD82 is an excellent prospective marker for purification of stem cells from human fetal and adult skeletal muscles. CD82+ human muscle cells successfully engraft in an immune-deficient mouse model of muscular dystrophy. CD82 interacts with 71-ITG in human myogenic cells and it is Erg linked to the DAPC complex via conversation with -sarcoglycan. Expression of CD82 is usually decreased in muscle tissue and myoblasts from DMD patients, suggesting that CD82 function may be linked to muscular dystrophies. Results To uncover regulators of human myogenesis, we sought to identify markers that label myogenic cells in developing human muscle. Melanoma Cell-Adhesion Molecule (MCAM) enriches for myogenic cells in human fetal muscle (Lapan and Gussoni, 2012), however MCAM is expressed in both myogenic Pax7+ satellite cells, mature myofibers and a subfraction (~25%) of Pax7? cells (Figure S1A, B). To further refine the myogenic from Dansylamide non-myogenic cells within the MCAM-positive fraction, comparison of the transcriptome of MCAM+ versus MCAM? cells identified CD82 as one candidate preferentially expressed in MCAM+ cells (Table S1). CD82 showed partial myofiber staining and it outlined cells that co-stained with Pax7 (Figure 1A). By western blot, CD82 protein was detected as a band of ~30Kd in uncultured, proliferating and differentiating human fetal myogenic cells (Figure 1B). FACS analysis of freshly dissociated human fetal muscle cells using CD82 and MCAM confirmed that CD82 marked a subpopulation of MCAM+ cells (Figure 1C). Sorted cell populations were induced to differentiate and myotube forming activity was restricted to the CD82+ subpopulation of MCAM+ cells, while neither double negative, nor MCAM+CD82? retained myotube-forming potential (Figure 1D). To confirm enrichment in myogenic activity when CD82 and MCAM were used in conjunction, the myotube formation ability of MCAM+CD82+ cells was compared to MCAM+ (total) cells. The fusion index (Figure 1E) and myotube size (Figure 1F) were significantly higher for MCAM+CD82+ compared to MCAM+ cells. By immunofluorescence, co-staining of CD82, dystrophin.