Ezetimibe ameliorates steatohepatitis via AMP activated protein kinase-TFEB-mediated activation of autophagy and NLRP3 inflammasome inhibition

Abstract

Impairment in macroautophagy/autophagy flux and inflammasome activation are common characteristics of nonalcoholic steatohepatitis (NASH). Considering the lack of approved agents for treating NASH, drugs that can enhance autophagy and modulate inflammasome pathways may be beneficial. Here, we investigated the novel mechanism of ezetimibe, a widely prescribed drug for hypercholesterolemia, as a therapeutic option for ameliorating NASH. Human liver samples with steatosis and NASH were analyzed. For in vitro studies of autophagy and inflammasomes, primary mouse hepatocytes, human hepatoma cells, mouse embryonic fibroblasts with Ampk or Tsc2 knockout, and human or primary mouse macrophages were treated with ezetimibe and palmitate. Steatohepatitis and fibrosis were induced by feeding Atg7 wild-type, haploinsufficient, and knockout mice a methionine- and choline-deficient diet with ezetimibe (10 mg/kg) for 4 wk. Human livers with steatosis or NASH presented impaired autophagy with decreased nuclear TFEB and increased SQSTM1, MAP1LC3-II, and NLRP3 expression. Ezetimibe increased autophagy flux and concomitantly ameliorated lipid accumulation and apoptosis in palmitate-exposed hepatocytes. Ezetimibe induced AMPK phosphorylation and subsequent TFEB nuclear translocation, related to MAPK/ERK. In macrophages, ezetimibe blocked the NLRP3 inflammasome-IL1B pathway in an autophagy-dependent manner and modulated hepatocyte-macrophage interaction via extracellular vesicles. Ezetimibe attenuated lipid accumulation, inflammation, and fibrosis in liver-specific Atg7 wild-type and haploinsufficient mice, but not in knockout mice. Ezetimibe ameliorates steatohepatitis by autophagy induction through AMPK activation and TFEB nuclear translocation, related to an independent MTOR ameliorative effect and the MAPK/ERK pathway. Ezetimibe dampens NLRP3 inflammasome activation in macrophages by modulating autophagy and a hepatocyte-driven exosome pathway.

MPB, a novel berberine derivative, enhances lysosomal and bactericidal properties via TGF‐β–activated kinase 1‐dependent activation of the transcription factor EB

“…Pharmaceutical or genetic inhibition of AMPK activity abolished MPB-induced TFEB nuclear translocation. Our results are in agreement with recent reports (21,52) showing that AMPK activation promotes TFEB nuclear translocation. Thus, MPB-induced AMPK activation and subsequent inhibition of mTOR activity reduces TFEB phosphorylation, allowing it to translocate.…”

Autophagy limits activation of the inflammasomes

“…147 Alternatively, chemical compounds such as rapamycin, AICAR, resveratrol, celastrol, ezetimibe, and polyunsaturated fatty acids, are known to induce autophagy. 105,[148][149][150][151][152] Although a hypocaloric diet and these compounds regulate not only autophagy but also various cellular responses, they are reported to ameliorate inflammation. [150][151][152][153][154][155] Thus, pharmacological modulation of autophagy may provide a potential therapy for inflammasome-related diseases.…”

Ezetimibe ameliorates lipid accumulation during adipogenesis by regulating the AMPK–mTORC1 pathway

“…20,35 Furthermore, recent studies showed that ezetimibe has antioxidant functions through the activation of AMPK, and therefore, it can ameliorate hepatic lipid accumulation and protect the liver from oxidative damage caused by diet-induced NASH in a mouse model. 21,26 However, its effects on adipogenesis have not been previously determined; thus, in this study, we demonstrate that ezetimibe has anti-adipogenic effects on 3T3-L1 preadipocytes.…”

“…[32][33][34] Several studies have reported that ezetimibe can improve the symptoms of metabolic disorder accompanied by obesity. 21,26 However, its effects on adipogenesis F I G U R E 7 AMPK is required for ezetimibe-mediated inhibition of adipogenesis. 20,35 Furthermore, recent studies showed that ezetimibe has antioxidant functions through the activation of AMPK, and therefore, it can ameliorate hepatic lipid accumulation and protect the liver from oxidative damage caused by diet-induced NASH in a mouse model.…”

“…20,21 To investigate whether it can ameliorate lipid accumulation in adipocytes, we incubated 3T3-L1 preadipocytes in differentiation medium with increasing concentration of ezetimibe, added every 2 days for up to 8 days (Supplemental Figure S1A). 20,21 To investigate whether it can ameliorate lipid accumulation in adipocytes, we incubated 3T3-L1 preadipocytes in differentiation medium with increasing concentration of ezetimibe, added every 2 days for up to 8 days (Supplemental Figure S1A).…”