Robust intrinsic differences in mitochondrial respiration and H₂O₂ emission between L6 and C2C12 cells

Abstract

Rat L6 and mouse C2C12 cell lines are commonly used to investigate myocellular metabolism. Mitochondrial characteristics of these cell lines remain poorly understood despite mitochondria being implicated in the development of various metabolic diseases. To address this need, we performed high-resolution respirometry to determine rates of oxygen consumption and H2O2 emission in suspended myoblasts during multiple substrate-uncoupler-inhibitor titration protocols. The capacity for oxidative phosphorylation supported by glutamate and malate, with and without succinate, or supported by palmitoyl-l-carnitine was lower in L6 compared with C2C12 myoblasts (all P < 0.01 for L6 vs. C2C12). Conversely, H2O2 emission during oxidative phosphorylation was greater in L6 than C2C12 myoblasts ( P < 0.01 for L6 vs. C2C12). Induction of noncoupled respiration revealed a significantly greater electron transfer capacity in C2C12 compared with L6 myoblasts, regardless of the substrate(s) provided. Mitochondrial metabolism was also investigated in differentiated L6 and C2C12 myotubes. Basal rates of oxygen consumption were not different between intact, adherent L6, and C2C12 myotubes; however, noncoupled respiration was significantly lower in L6 compared with C2C12 myotubes ( P = 0.01). In summary, L6 myoblasts had lower respiration rates than C2C12 myoblasts, including lesser capacity for fatty acid oxidation and greater electron leak toward H2O2. L6 cells also retain a lower capacity for electron transfer compared with C2C12 following differentiation to form fused myotubes. Intrinsic differences in mitochondrial metabolism between these cell lines should be considered when modeling and investigating myocellular metabolism.

Effects of PCB126 on Adipose-to-Muscle Communication in an in Vitro Model

“…Thus, C2C12 myotubes may have a more oxidative metabolism than L6 myotubes. This hypothesis has been confirmed in a recent study in which it was demonstrated that L6 cells had lower mitochondrial and fatty acid oxidation capacities than C2C12 cells while having higher emission of ROS ( Robinson et al. 2019 ).…”

“…Thus, C2C12 myotubes may have a more oxidative metabolism than L6 myotubes. This hypothesis has been confirmed in a recent study in which it was demonstrated that L6 cells had lower mitochondrial and fatty acid oxidation capacities than C2C12 cells while having higher emission of ROS (Robinson et al 2019). Future studies should therefore determine whether the higher glycolytic capacity and higher ROS emission in L6 cells exposed to PCB126 may explain the fact that this muscle cell model was more sensitive to PCB126 than C2C12.…”

PCB126-mediated effects on adipocyte energy metabolism and adipokine secretion may result in abnormal glucose uptake in muscle cells

“…Thus, C2C12 myotubes may have a more oxidative metabolism than L6 myotubes. This hypothesis has been confirmed in a recent study in which it was demonstrated that L6 cells have lower mitochondrial and fatty acid oxidation capacities than C2C12 cells, while having higher emission of ROS (Robinson et al, 2019). Future studies should therefore determine whether the higher glycolytic capacity and higher ROS emission in L6 cells may explain the fact that this muscle cell model is more sensitive to PCB126 than C2C12 when exposed to PCB126.…”

C2C12 cell model: its role in understanding of insulin resistance at the molecular level and pharmaceutical development at the preclinical stage

“…One of the studies revealed that rat L6 cells had lower rate of oxygen consumption and respiration when compared to mouse C2C12 cells during the oxidative phosphorylation and noncoupled respiration. [ 27 ] The mechanisms that are associated with the intrinsic differences in mitochondrial metabolism between mouse C2C12 and rat L6 cell will require further investigation. [ 27 ] Overall, insulin‐responsive mouse C2C12 cell line is a useful model in investigation of insulin resistance.…”