In vitro evaluation of human muscle satellite cell migration prior to fusion into myotubes

We developed a short-time assay to evaluate muscle satellite cell migration , based on the fact that during myogenic differentiation, myoblasts migrate preferentially towards high cellular density areas where myotubes would fo rm. This assay consists of a computer-assisted count of cells within a rand omly chosen field, performed every hour for eight hours, and compared with the cell number at the start time of the experiment. Nine primary myoblast cultures were tested in triplicate. The method relies on several requisites . (1) Negligible cell proliferation: cell division was nearly absent in 8 h experiments. (2) Directional cell movement: a major flow of cells, either entering or exiting the fields, was constantly observed. 'Counter-flows', d etected by visual counting, involved minor percentages of cells. (3) Consta nt migration rate: a linear increase in cell count variations over 8 h and a very high degree of intra-assay homogeneity were observed. Individual pri mary cell culture characteristics (depending on characteristics of the diff erent donors) were the sole factor with a significant impact on migration r ate. Automatic cell counting conveniently assessed the inhibitory effect of GRGDTP, an inhibitor of integrin-mediated cell adhesion. The method descri bed here is rapid, does not require heavy equipment, and allows studies und er serum-free conditions required to test molecules interfering with cell m igration, in the course of the in vitro myogenic process. (C) Kluwer Academ ic Publishers.