The Effect Of Lactate On Proliferation And Differentiation Behaviour Of C2c12 And Primary Human Myoblasts

Abstract

Lactate (La) has long been considered as a waste product of energy metabolism and the cause of decrease in muscle pH and hence muscle fatigue. This idea has changed massively in the past. Now it is known that La is rather an intermediate of glucose metabolism as well as it has been termed Lactormon for it has signalling properties inducing gene expression for skeletal muscle adaptation. It has been shown to increase MCT content as well as to increase mitochondrial biogenesis. Such skeletal muscle adaptation is one of the major foci in sports medicine. Numerous adaptational mechanisms and signalling pathways have been described in the literature, investigating among other mechanical, metabolical, nutritional, and hormonal stimuli and their effects for the improvement of physiological functioning of skeletal muscle tissue. La, as a metabolic stimulus, has not yet been investigated extensively in this context. PURPOSE: We investigated the effects of different lactate concentrations on the proliferation status and differentiation behaviour of C2C12 mouse immortal and primary human myoblasts. METHODS: C2C12 mouse and primary human myoblasts were constantly incubated with differentiation medium containing different lactate concentrations (10 mM, 20 mM) at 37°C and 5% CO2 over a time period of 15 days. Cells were fixed using 4% PFA in PBS for immunocytochemical staining for Ki67, activated Caspase-3, F5D (myogenin) and Mf-20 (myosin heavy chain). Additionally, cell lysates were made for Western Blot analysis using the same markers as for immunostaining. Furthermore, we repeated the experiments with an intermittent incubation of 2 h every day to simulate a more realistic training situation. RESULTS: Densitometrical analysis of cells stained for Ki67 showed a significant decrease if treated with La in a dose-dependent manner. Controversially, apoptotic induction by activated caspase-3 was increased if treated with La. Differentiation was shown to be delayed, but not decreased if cells received the La intervention. CONCLUSION: Lactate seems to exhibit an inhibitory effect on the proliferation and apoptotic behaviour of C2C12 and primary human myoblasts. Furthermore, differentiation seems to be delayed but not reduced in a dose-dependent manner.