Roles of PKLR in NAFLD using loss- and gain-of-function mouse models
The etiology of non-alcoholic fatty liver disease (NAFLD) is poorly understood, with males and Mexican population exhibiting markedly increased susceptibility. Using a systems genetics approach, involving multi-omic analysis of ~100 inbred strains, we recently identified several candidate genes driving NAFLD. We investigated the role of one of these candidates, liver pyruvate kinase (L-PK or Pklr) in NAFLD using patient samples and mouse models. We examined L-PK expression in mice of both sexes and in a cohort of bariatric surgery patients. We used liver-specific loss- and gain-of-function strategies in independent animal models of diet-induced steatosis and fibrosis, involving both sexes. Following treatment, we measured several metabolic phenotypes associated with obesity, insulin resistance, dyslipidemia, liver steatosis and fibrosis. Furthermore, liver tissues were used for gene expression and western blots, and liver mitochondria for bioenergetics. Our results demonstrate that L-PK acts in a male-specific manner in the development of liver steatosis and fibrosis. Given that NAFLD/NASH exhibit sexual dimorphism, our results have important implications for the development of personalized therapeutics. Overall design: Eight-weeks old male C57BL/6J mice were randomly injected intravenously with adeno-associated viral 8 (AAV8) vectors containing either the cDNA or the shRNA sequences against Pyruvate kinase, liver isoform (PKLR) under a thyroxine binding globulin promoter (1012 titer). Control mice were injected with either AAV8 expressing GFP gene (for overexpression models) or AAV8 expressing shRNA against firefly luciferase gene (for knockdown models). The animals were then maintained on a HF/HS diet (Research Diets D12266B) until 25 weeks of age. Liver samples (n = 5 per group) were taken for RNASeq expression profiling.