Manufacturing autologous myoblast for regenerative medicine applications

Abstract

Background: Autologous myoblasts have been tested in the treatment of muscle-related diseases. However, the standardization of manufacturing myoblasts is still not established. Here we report a flask and animal-free medium-based method of manufacturing clinical-grade myoblast together with establishing releasing criteria for myoblast products under Good Manufacturing Practice (GMP). Methods: Quadriceps muscle biopsy samples were donated from three patients with myogenic ptosis. After biopsy samples were digested through enzymatic dissociation, the cells were grown in T175 flasks (passage 0) and hyperflasks (passage 1) in the animal-free SkGM TM -2 skeletal muscle cell growth medium containing 5% human platelet lysate for 15-17 days. The harvested cells were released based on cell morphology, cell dose, viability, sterility, endotoxin, mycoplasma and immunophenotype. Myotube differentiation was also evaluated. Results: 400 to 500 million myoblast cells were manufactured within 15 to 17 days by the end of passage 1, which met predetermined releasing criteria. The manufactured myoblast cells could differentiate and fuse into myotubes in vitro, with the possible trend that the Electronic supplementary material The online version of this article (

Myogenic differentiation of human myoblasts and Mesenchymal stromal cells under GDF11 on Poly-ɛ-caprolactone-collagen I-Polyethylene-nanofibers

“…The purchased myoblasts were all from the quadriceps muscle and human subjects were all male to limit heterogeneity. Other factors like age are known to influence the degree of myotube formation [ 37 , 38 ]. Furthermore, maintenance and myogenic differentiation of primary cells isolated from adult muscle tissue is a larger challenge than culturing immortal cell lines like C2C12 with a rapid proliferative and differentiation capacity [ 39 ].…”

Myogenic Differentiation of Human Myoblasts and Mesenchymal Stromal Stem Cells under GDF11 on Poly-ɛ-Caprolactone-Collagen I-Polyethylene-Nanofibers

“…The purchased myoblasts were all from the quadriceps muscle and human subjects were all male to limit heterogeneity. Other factors like age are known to in uence the degree of myotube formation 31,32 . Furthermore, maintaining and myogenic differentiation of primary cells isolated from adult muscle tissue is a larger challenge than culturing immortal cell lines like C2C12 without contact inhibition.…”