Skeletal myoblast cells have a high potential for cell therapy based repair of the injured myocardium. Stem cell differentiation markers are important for finding an appropriate cellular state optimal for engraftment. We followed differentiation-induced changes in cellular size, the distribution of mitochondria, adenine and pyridine nucleotide content, NAD(P)H fluorescence in skeletal myogenic stem cell populations derived from rabbit skeletal muscle. The fraction of larger size cells increased during differentiation. Mitochondrial distribution changed from perinuclear in undifferentiated cells to even in the cytoplasm of differentiated cells. The most remarkable feature of undifferentiated myoblasts was a very low NAD(P)H fluorescence which augmented after initiating cell differentiation, due to metabolic and physiological changes, but that did not correlate with an increase in cellular size. A significant increase of NAD(P)H concentration was chromatographically detected after 8 days of differentiation, although the NAD(P)H fluorescence increased by 42% as soon as after three days and 2.7-fold after five days of differentiation as compared with the control. We show that the early increase in fluorescence is determined by mitochondrial NADH, but upon a longer differentiation NADPH amount also increases. The total amount of adenine nucleotides (ATP, ADP, AMP) increased, but the ATP / ADP ratio decreased during differentiation. We suggest that an increase in NAD(P)H fluorescence may be useful for a non-invasive detection of the onset and course of myoblast differentiation.

Keywords: myogenic stem cells, differentiation, NAD(P)H fluorescence