The effects of physiological and pathophysiological concentrations of fatty acids on lipid droplet accumulation and metabolic function of LHCN-M2 human skeletal muscle cells

Introduction: Skeletal muscle (SkM) is a major site of carbohydrate and fatty acid (FA) utilisation, where efficient selection between these two substrates is paramount in maintaining metabolic homeostasis. The accumulation of FA in SkM has been suggested to contribute to metabolic diseases such as obesity and type 2 diabetes mellitus. Consequently, there is significant interest in SkM FA metabolism and importantly, how FAs contribute to the metabolic function of SkM.

Methods: LHCN-M2 human skeletal muscle cells were cultured and differentiated into multinucleated myotubes. After a total of 8 days differentiation, LHCN-M2 cells were cultured for a further 2 days with 0.25% fatty acid-free bovine serum albumin alone or conjugated to oleic, palmitic, linoleic, and α-linoleic acid (OPLA; physiological ratio 45:30:24:1%) at a concentration of 200µM and 800µM. Immunofluorescence confocal microscopy was used to determine the presence of lipid droplets in SkM and Immunoblotting to determine the expression of oxidative phosphorylation (OXPHOS) protein complexes (II-V). Cell respiration analysis was used to determine the effects of OPLA in mitochondrial function.

Results: Immunofluorescence based microscopy showed an increased in the presence of lipid droplets following exposure to increasing concentrations of OPLA. However, analysis of cellular respiration found no significant differences in basal or maximal respiration, coupling efficiency or spare capacity between conditions (P > 0.05). The expression OXPHOS protein complexes (II-V) was not different between conditions (P > 0.05).

Conclusions: Preliminary data from these experiments would suggest that although exposure to increasing concentrations of OPLA leads to an increase in the presence of lipids droplets, there are no differences in cell respiration or expression of OXPHOS protein complexes. Further experiments are needed to determine how changes in composition of lipids at these concentrations effect the metabolic function of human skeletal muscle in vitro.