CB1 knockout alleviates hepatic steatosis via lipophagy and lipolysis in HBs transgenic mice

The endocannabinoid (EC) system has been implicated in the pathogenesis of several metabolic diseases, including nonalcoholic fatty liver diseases (NAFLD). The present study aims to describe the effect of global CB1 knockout on the hepatic lipid metabolism in a HBs transgenic mouse model.

Hybrids of HBs transgenic mice and CB1 knockout mice were bred on genetic backgrounds of B6. The intracellular lipids were assessed using Oil-Red-O staining, biochemical triglyceride (TGs) assays and thin layer chromatography (TLC). The expression of key proteins regulating autophagy and lipid metabolism were analyzed by immunohistochemistry (IHC), immune fluorescence (IF) and Western blotting (WB).

CB1 knockout mice showed a reduction in body weight and lipid droplets (LDs) sizes as well as TGs in HBs/CB1-/- mice compared to HBs transgenic mice. CB1-/- mediated metabolic disturbances as shown by increased GLUT1 expression on the surface of hepatocytes and activation of the transcription factors CREB and FOXO1. Metabolic disturbances induced autophagy flux progression as shown by decreased LC3B-II and LAMP1 in WB and IF of HBs/CB1-/- compared to HBs. LAMP1 and lipid droplets binding protein PLIN2 were co-localized in HBs/CB1-/- indicating autophagy of LDs i.e. lipophagy. Autophagy mediated decrease of PLIN2 protein level was also associated with elevated expression of lysosomal acid lipase (LAL), cytoplasmic lipases, hepatic lipase (LIPC) and adipose triglyceride lipase (ATGL). ATGL and PLIN2 co-localization further indicated enhanced ATGL interaction with LDs promoting lipophagy.

In conclusion, the present study shows that CB1 receptor knockout decreases hepatic steatosis via lipophagy and cytoplasmic lipolysis in HBs transgenic mice. Our results might have an impact on other liver diseases with enhanced steatosis.