engleski

Human myoblast cell culture-model for adult stem cell activation after muscle injury and disease

Introduction Skeletal muscle is composed of muscle cells (myoblasts) which unite into myofibrils and later in the muscle fibers. Activation of myoblasts after muscle injury or muscle disease is an important step in the process of muscle regeneration. During muscle regeneration myoblasts enter the proliferative process at the end of which the majority of myoblasts are differentiated in myotube. Materials and Methods Myoblasts were isolated by enzymatic digestion of skeletal muscle biopsy. After the cells reached a density about 70-80%, muscle stem cells were prepared for the separation of myoblasts from fibroblasts. Myoblasts were first characterized by specific monoclonal antibody (NCAM/CD56) and IgG microspheres boding. Then the Magnetic-activated cell sorting (Miltenyl Biotec, GmbH) separates from fibroblasts. Differentiation of myoblasts in the myotubes can be traced by the phase-contrast microscope. Results We have established a cultures of stem cells, myoblasts and fibroblasts from a very small piece of muscle biopsy. Using a magnetic cell separator, myoblasts were separated from fibroblasts and as such stored in liquid nitrogen. Purity of myoblasts was confirmed with markers CD 56+ and CD 45- by flow cytometry. It was confirmed that 81.4% of the cells carry a myoblast marker. Conclusion Cell culture of human myoblasts opens up new possibilities in the field of research of adult stem cells and rare diseases, especially mitochondrial myopathy. This biological material allows the study of cellular and molecular mechanisms that may contribute to the understanding of the pathophysiology and new therapeutic approaches by using stem cells.

myoblast cell culture; stem cells; muscle regeneration

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