Author(s): WU, J., ZHANG, Y., CUI, R., ZHENG, H., LUO, Y., SUN, W., XIANG, M., Institution: 南京体育学院, Country: CHINA, Abstract-ID: 1013

INTRODUCTION:
Impaired autophagy in hepatocytes may not only result in steatosis, but also lead to steatohepatitis. Mitophagy is an important part of autophagy and plays an important role in regulating liver homeostasis. This study aim to explore the effect of exercise on hepatocyte lipid deposition and autophagy in NAFLD mice, which to provide theoretical basis for NAFLD preventive treatment.
METHODS:
30 8-week male C57BL/6J mice were adaptively fed for one week, then randomly divided into normal diet group (NC, n=10) and high-fat, fructose, and cholesterol diet group (HFC, n=20). After 10-week feeding, 10 mice from HFC mice were randomly selected for 8 weeks of voluntary wheel running (HFE, n=10). Animal serum and fresh liver tissue were collected. Serum lipids level were detected. Liver mitochondria were isolated by gradient centrifugation.The supernate from the mitochondrial isolation step was removed and centrifuged at 35,000 g for obtaining the cytoplasm. HE and oil red straining were used to observe the histopathological changes and hepatic lipid deposition. Immunofluorescence co-localization of p62+LC3 and LAMP-2+LC3 was used to detect the formation and degradation of autophagosomes, and P62,LC3, mitochondrial PINK1, Parkin and cytoplasmic Parkin expressions were detected by western blot.
RESULTS:
1)The body weight(+30.56%)、liver weight(+93.04%)、and TG(+31.33%) content in HFC mice were remarkably higher than NC(P<0.05).Compare to HFC, mice in HFE group showed decreased body weight (-14.07%),liver weight (-27.48%) and TG (-46.79%) (P<0.05) levels.
2)Oil red staining indicated that HFFC diet significantly increased the liver lipid area (XXfold,P<0.01), and the liver lipid area was significantly decreased after exercise intervention(-50%, P<0.01).
3)Compared to NC mice, HFC mice exhibited significantly reduced the level of co-localization in P62+LC3(4.00±0.32vs2.4±0.24) and LAMP-2+LC3(7.40±0.24vs3.6±0.51), as well as increased P62 expression (0.71±0.09vs1.47±0.13)(P<0.01). Exercise markedly decreased the expression of P62 by 55.78%, accompanied by higher LC3II/LC3I ratio(0.74±0.13vs1.29±0.11, P < 0.05) and increased P62+LC3 (2.4±0.24vs6.4±0.51), LAMP-2+LC3 (3.6±0.51vs12.00±1.05) co-localization.
4)Mice in HFC demonstrated reduced mitochondrial PINK1(-33.53%) and Parkin (-39.60%)expression(P<0.05), but increased the cytosolic Parkin content (+157.5%,P<0.01). Compared with HFC, mitochondrial PINK1 and Parkin levels in HFE mice were elevated by 42.48% and 80.33%, respectively.
CONCLUSION:
1)HFFC diet lead to hepatocyte structure damage and lipid deposition. Exercise intervention can reduce the body weight, liver index, and alleviate the liver lipid deposition of HFFC diet mice.
2)HFFC diet resulted in p62 protein accumulation and impaired autophagy flux in hepatocytes. Exercise significantly improved the autophagy flux via increasing p62+LC3 and LAMP2+LC3 co-localization, and enhanced mitophagy by up-regulating PINK1/Parkin signaling pathway.