Rong Yan1, Chun-Yan Niu2, Yu Tian3. 1. Department of Gastroenterology, First Affiliated Hospital of Xi'an Medical University, Xi'an, Shaanxi Province, China. Electronic address: ashleyyan0117@126.com. 2. Department of Gastroenterology, Xiang'an Hospital of Xiamen University, Xiamen, Fujian Province, China. 3. Clinical Laboratory, First Affiliated Hospital of Xi'an Medical University, Xi'an, Shaanxi Province, China.
Abstract
OBJECTIVE: To investigate interaction between autophagy and PKC-ε in lipid metabolism of NAFLD cell models. METHODS: HL-7702 cells and SK-HEP-1 cells were cultured in vitro as NAFLD cell models and treated with RAPA to induce autophagy. 3-MA was used to inhibit cell autophagy. And HL-7702 and SK-HEP-1 cell were ordinary cultured as control groups. Cell viability was determined by MTT colorimetric assay. The levels of TG, TC and PKC-ε were detected by ELISA. PKC-ε was detected by IF. LC3-II/LC3-I, P62, IRS-1, IRS-2, PI3Kp85, mTOR were detected by Western-blot. SPSS 20 software was used for statistical analysis. RESULTS: The values of PKC-ε were the highest in the steatosis groups (HL-7702 cells were 91.10%, SK-HEP-1 cells were 98.20%). Compared with the steatosis groups, the LC3-II/LC3-I ratio in the induced autophagy groups increased obviously (p <0.05). P62/β-actin grayscale ratio of the induced autophagy groups decreased significantly compared with the steatosis group (p <0.05). MTOR/β-actin grayscale ratio in the induced autophagy groups were significantly lower than those in the steatosis groups (p <0.05). PI3Kp85, IRS-1 and IRS-2/β-actin grayscale ratio of the induced autophagy groups increased significantly compared with the steatosis group (p <0.05). CONCLUSION: Up-regulation of autophagy can promote the elimination of liver fat; while down-regulation can promote lipid accumulation. The expression of PKC-ε is positively related to the degree of hepatic steatosis. PI3K was involved in both autophagy and IR induced by PKC-ε. PKC-ε might participate in hepatocyte autophagy by regulating PI3K.
OBJECTIVE: To investigate interaction between autophagy and PKC-ε in lipid metabolism of NAFLD cell models. METHODS: HL-7702 cells and SK-HEP-1 cells were cultured in vitro as NAFLD cell models and treated with RAPA to induce autophagy. 3-MA was used to inhibit cell autophagy. And HL-7702 and SK-HEP-1 cell were ordinary cultured as control groups. Cell viability was determined by MTT colorimetric assay. The levels of TG, TC and PKC-ε were detected by ELISA. PKC-ε was detected by IF. LC3-II/LC3-I, P62, IRS-1, IRS-2, PI3Kp85, mTOR were detected by Western-blot. SPSS 20 software was used for statistical analysis. RESULTS: The values of PKC-ε were the highest in the steatosis groups (HL-7702 cells were 91.10%, SK-HEP-1 cells were 98.20%). Compared with the steatosis groups, the LC3-II/LC3-I ratio in the induced autophagy groups increased obviously (p <0.05). P62/β-actin grayscale ratio of the induced autophagy groups decreased significantly compared with the steatosis group (p <0.05). MTOR/β-actin grayscale ratio in the induced autophagy groups were significantly lower than those in the steatosis groups (p <0.05). PI3Kp85, IRS-1 and IRS-2/β-actin grayscale ratio of the induced autophagy groups increased significantly compared with the steatosis group (p <0.05). CONCLUSION: Up-regulation of autophagy can promote the elimination of liver fat; while down-regulation can promote lipid accumulation. The expression of PKC-ε is positively related to the degree of hepatic steatosis. PI3K was involved in both autophagy and IR induced by PKC-ε. PKC-ε might participate in hepatocyte autophagy by regulating PI3K.
Authors: Jiayao Yang; Ying Zhang; Hongfeng Yi; Yan Liao; Lei Shu; Shu Zhang; Chenyu Li; Liu An; Nianlong Du; Zhaohong Shi; Wei Ma Journal: Evid Based Complement Alternat Med Date: 2022-08-26 Impact factor: 2.650