Endocrine Abstracts

Introduction: Skeletal muscle cells enable investigation of myogenesis and metabolic function in vitro. Exposure to human serum can provide insight into the impact of endocrine factors upon differentiation and mitochondrial function of skeletal muscle. The aim of these experiments was to optimise the culture conditions using human serum, which it was hypothesised would enhance myogenesis and mitochondrial function of LHCN-M2 human skeletal muscle cells.

Methods: LHCN-M2 skeletal muscle myoblasts were differentiated in serum-free media, 0.5% or 2% healthy human serum, for 0, 5 and 10 days. Skeletal muscle myotube formation was assessed morphologically by immunofluorescence (Myosin Heavy Chain (MHC)) and differentiation transcriptionally by mRNA expression (Myogenic differentation1 (MyoD1)) and (Myogenic regulatory factor 5 (Myf5)). At day 10, extracellular flux analysis was used to determine mitochondrial function following differentiation in different concentrations of serum.

Results: Differentiation with human serum increased the mRNA expression of MyoD1 (6.58 ± 1.33 fold at day 5 and 4.60 ± 1.43 fold at day 10 vs day 0) and reduced Myf5 (0.21 ± 0.11 fold at day 5 and 0.07 ± 0.03 fold at day 10 vs day 0) with time (P < 0.05). However, there was no difference in mRNA expression between conditions (P = NS). Extracellular flux analysis showed no differences in basal, maximal, ATP or spare capacity between conditions (P = NS). However, coupling efficiency was significantly lower following differentiation in 2% human serum compared to serum-free media (79.68 ± 3.62% vs. 92.08 ± 7.19%; P < 0.05).

Conclusions: These preliminary findings contradict the hypothesis and reveal that the concentration of human serum does not affect the mRNA expression of myogenic transcription factors during LHCN-M2 differentiation or mitochondrial respiratory capacity, with the exception of coupling efficiency, which is attenuated in 2% serum. These experiments have wider implications for investigating how human serum is used in skeletal muscle metabolism research.