반복적인 근 수축으로 인한 ROS 방어 체계의 향상이 Palmitate로 인한 ER stress 유발 및 미토콘드리아 칼슘에 미치는 효과

Abstract

PURPOSE: This study, it was intended to induce repetitive muscular dystrophy in skeletal muscle cells (C2C12) with continuous electro stimulation, check the improvement of the antioxidant system, and determine whether the ER stress by the palmitate can be controlled by iterative muscle contraction.<BR> METHODS: To examine the expression of catalase on palmitate-induced ER stress and mitochondria calcium level, 100uM palmitate was treated with C2C12 cell. Palmitate-induced reactive oxygen species(ROS) formation was measured by DCFDA fluorescence dye. ER Ca<SUP>2+</SUP> level was measured by using thapsigargin-induced ER Ca<SUP>2+</SUP> release method which was detected by Flou-4 am, Ca<SUP>2+</SUP> dye. palmitateinduced mitochondria calcium was measured by TMRM dye. ER stress protein PERK expression, phosphorylation, and ER Ca<SUP>2+</SUP> releasing channel related protein, RYR1 and p-RYR were analyzed by western blotting. All parameters were compared between control cell, palmitate - treated cell and palmitate and electro stimulation - treated cell. In addition, the catalase inhibitor, 3-Amino-1,2,4-triazole (3-AT), was treated in cells where the expression of the catalase was increased through electrical stimulation, and an experiment was conducted to establish the relationship between the catalase suppression and the ER stress.<BR> RESULTS: The one-week electro stimilation significantly increased the expression of the catalase in skeletal muscle cells and was shown to significantly inhibit the level of ER Ca<SUP>2+</SUP> by Palmitate. In addition, muscle contraction reduced of mitochondria Ca<SUP>2+</SUP> levels by palmitate was also shown to be inhibited, and the expression and phosphorylation of PERK / RYR1, an indicator were reduced. The result of processing 3-AT on cells that performed electro stimulation shows that the increase in ER stress indicators due to palmitate is significantly higher than that of electro stimulating cells. Phosphorylation of PERK / RYR also tends to increase.<BR> CONCLUSIONS: Repeated muscle contraction improves anti-oxidization capability in skeletal muscle, and activates the protective system of skeletal muscle related to metabolic disease. Palmitate increase the ROS of skeletal muscle cells and increase the ER stress. This study suggests that the incidence of catalase increases through skeletal muscle contraction an important role in defending the ER stress from palmitate processing, and suggests that the ROS defense system due to the Catalase is an important mechanism for suppressing ER stress and maintaining the mitochondria Ca<SUP>2+</SUP> level.