Abstract
Background/aimsLiver lipid accumulation induced by high-fat diet (HFD) is an early onset process of non-alcoholic fatty liver diseases (NAFLD). Protein kinase A (PKA) is known to be involved in hepatic lipid metabolism. However, the role of PKA in NAFLD has not been well tested in vivo due to the lack of optimal PKA deficient mouse model.MethodsA novel PKA-specific inhibitor gene was conditionally overexpressed in mouse (PKAi mouse) liver using LoxP/Cre system. PKA activity in the liver extract was measured with a commercial assay kit. The PKAi and control mice of 8-week age, were subjected to HFD or chow diet (CD) for 2 months. Body weight, liver index, and triglyceride in the liver were measured. RNA sequencing was performed for the liver tissues and analyzed with Gene Ontology (GO) and pathway enrichment.ResultsPKAi-GFP protein was overexpressed in the liver and the PKA activation was significantly inhibited in the liver of PKAi mouse. When fed with CD, RNA sequencing revealed 56 up-regulated and 51 down-regulated genes in PKAi mice compared with control mice, which were mainly involved in lipid metabolism though no significant differences in the body weight, liver index, triglyceride accumulation were observed between PKAi and control mice. However, when fed with HFD for 2 months, the liver was enlarged more, and the accumulation of triglyceride in the liver was more severe in PKAi mice. When comparing the transcriptomes of CD-fed and HFD-fed control mice, GO enrichment showed that the genes down-regulated by HFD were mainly enriched in immune-related GO terms, and up-regulated genes were enriched in metabolism. When comparing the transcriptomes of CD-fed and HFD-fed PKAi mice, GO analysis showed that the down-regulated genes were enriched in metabolism, while the up-regulated genes were clustered in ER stress-related pathways. When comparing HFD-fed PKAi and HFD-fed control mice, the genes with lower expression level in PKAi mice were enriched in the lipoprotein synthesis, which might explain that more TG is accumulated in PKAi liver after HFD feeding.ConclusionsReduced PKA activity could be a factor promoting the TG accumulation in the liver and the development of NAFLD.
Highlights
Nonalcoholic fatty liver disease (NAFLD) consists a wide pathological spectrum ranging from steatosis, nonalcoholic steatohepatitis (NASH), fibrosis, and cirrhosis to hepatocellular carcinoma; and has become the fastest growing liver disease worldwide with a prevalence as high as a 25% of the population
PKAi-GFP is expressed in the mouse liver to suppress Protein kinase A (PKA) activity Hepatic-specific PKA inhibition mice and corresponding control mice were obtained by crossing the heterozygotes CAG-LoxP-CAT-LoxP-PKAi-GFP transgenic and Alb-cre transgenic mice (Fig. 1a)
When crude liver tissue extract was stimulated by 1 μmol/L cyclic adenosine monophosphate (cAMP), PKA activity was significantly increased in control mouse liver samples but not as much in PKAi transgenic samples, indicating that overexpressed PKAi-GFP in PKAi mouse liver was sufficient to inhibit most of cAMP-induced PKA activity (Fig. 1c)
Summary
Nonalcoholic fatty liver disease (NAFLD) consists a wide pathological spectrum ranging from steatosis, nonalcoholic steatohepatitis (NASH), fibrosis, and cirrhosis to hepatocellular carcinoma; and has become the fastest growing liver disease worldwide with a prevalence as high as a 25% of the population. The underlying mechanism for the development and progression of NAFLD is complex and multifactorial, among which lipid accumulation in liver is an early step [3, 4]. There are multiple mechanisms leading to lipid accumulation in hepatocytes, including increased free fatty acids supply and de novo hepatic lipogenesis; and decreased free fatty β-oxidation in the liver as well as decreased hepatic lipid export [5]. Lipid accumulation may result in the early “inflammatory” hit, leading to the whole spectrum of NAFLD pathologies [3]
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