Glutamine depletion disrupts mitochondrial integrity and impairs C2C12 myoblast proliferation, differentiation, and the heat-shock response

Abstract

Glutamine and glucose are both oxidized in the mitochondria to supply the majority of usable energy for processes of cellular function. Low levels of plasma and skeletal muscle glutamine are associated with severe illness. We hypothesized that glutamine deficiency would disrupt mitochondrial integrity and impair cell function. C2C12 mouse myoblasts were cultured in control media supplemented with 5.6 mmol/L glucose and 2 mmol/L glutamine, glutamine depletion (Gln^-) or glucose depletion (Glc^-) media. We compared mitochondrial morphology and function, as well as cell proliferation, myogenic differentiation, and heat-shock response in these cells. Glc^- cells exhibited slightly elongated mitochondrial networks and increased mitochondrial mass, with normal membrane potential (ΔΨm). Mitochondria in Gln^- cells became hyperfused and swollen, which were accompanied by severe disruption of cristae and decreases in ΔΨm, mitochondrial mass, the inner mitochondrial membrane remodeling protein OPA1, electron transport chain complex IV protein expression, and markers of mitochondrial biogenesis and bioenergetics. In addition, Gln^- increased the autophagy marker LC3B-II on the mitochondrial membrane. Notably, basal mitochondrial respiration was increased in Glc^- cells as compared to control cells, whereas maximal respiration remained unchanged. In contrast, basal respiration, maximal respiration and reserve capacity were all decreased in Gln^- cells. Consistent with the aforementioned mitochondrial deficits, Gln^- cells had lower growth rates and myogenic differentiation, as well as a higher rate of cell death under heat stress conditions than Glc^- and control cells. We conclude that glutamine is essential for mitochondrial integrity and function; glutamine depletion impairs myoblast proliferation, differentiation, and the heat-shock response.

Keywords: mitochondrial bioenergetics; mitochondrial biogenesis; mitochondrial fission; mitochondrial fusion; mitophagy; nutrient depletion; oxidative phosphorylation.