The Role And Mechanism Of PP2A Inhibitor LB100 In Nonalcoholic Fatty Liver Disease

Backgrounds and Aims:Nonalcoholic fatty liver disease(NAFLD)is one of the most common chronic liver diseases in the world nowadays,with rapidly increasing prevalence in the globe and unclear mechanism,and the efficiency of present treatments of NAFLD is limited.Emerging evidence suggests that overactivation of Serine/threonine protein phosphatase 2A(PP2A)is closely related to elevated insulin resistance(IR)in the liver,while IR is one of the most important pathogenesis of NAFLD.LB 100 is a small molecule inhibitor of PP2A,however,whether LB 100 plays an important role in NAFLD and its specific mechanism remains unknown.This study aimed to elucidate the effect and underlying mechanism of LB 100 in NAFLD.Materials and Methods:Human normal liver cell line,L02 was stimulated by free fatty acid(FFA)for 24 hours to induce NAFLD cell model,with or without LB 100 stimulation.Six to eightweek-old C57BL/6 mice were fed with high fat diet(HFD)for 16 weeks to establish in vivo model of NAFLD,after 10 weeks of HFD feeding,mice were injected intraperitoneally with vehicle or LB 100 1.5mg/kg for an additional 6 weeks(three times a week).Hematoxylin-Eosin(H&E)staining,oil red O staining and triglyceride determination were performed to estimate liver steatosis.The level of hepatic insulin resistance was assessed by glucose and insulin tolerance test and hepatic insulin pathway.The cellular and hepatic PP2A phosphatase activity was determined by enzyme linked immunosorbent assay(ELISA).Western blot analysis was used to detect the protein expression of Sirtuinl(Sirtl),total and phosphorylated AMP-activated protein kinase α(AMPKα),and the proteins involved in lipogenesis and fatty acid oxidation.The mRNA levels were determined by Quantitative PCR(qPCR).Pharmacological inhibition of AMPK was performed to further examine the exact mechanism of LB 100 in NAFLD.Results:(1)This study successfully established a cellular model of NAFLD,in this model,LB 100 significantly reduced cellular PP2A activity and activated the AMPK/Sirtl pathway.LB100 treatment significantly downregulated the levels of proteins involved in fatty acid synthesis while increased the expression of proteins involved in fatty acid β-oxidation.(2)This study successfully established an animal model of NAFLD.In the animal model,LB 100 significantly ameliorated HFD-induced obesity,IR,hepatic lipid accumulation and hepatic injury in mice.Further mechanistic studies have found that LB 100 significantly reduced liver PP2A activity and activated the AMPK/Sirtl pathway.LB 100 treatment significantly downregulated the levels of proteins involved in fatty acid synthesis while increased the expression of proteins involved in fatty acid β-oxidation.(3)In the NAFLD cell model,after pharmacological inhibition of AMPK with Compound C,the improvement of NAFLD by LB 100 was reversed,indicating that the role of LB 100 in NAFLD was mediated by AMPK/Sirtl pathway.Conclusions:PP2A inhibition by LB 100 significantly ameliorates hepatic steatosis by regulating hepatic lipogenesis and fatty acid oxidation via the AMPK/Sirt1 pathway.LB 100 may be a potential therapeutic agent for NAFLD.