Chronic-plus-binge alcohol intake induces production of proinflammatory mtDNA-enriched extracellular vesicles and steatohepatitis via ASK1/p38MAPKα-dependent mechanisms
Jing Ma, … , Jian Sun, Bin Gao
Jing Ma, … , Jian Sun, Bin Gao
Published June 16, 2020
Citation Information: JCI Insight. 2020;5(14):e136496. https://doi.org/10.1172/jci.insight.136496.
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Research Article Hepatology

Chronic-plus-binge alcohol intake induces production of proinflammatory mtDNA-enriched extracellular vesicles and steatohepatitis via ASK1/p38MAPKα-dependent mechanisms

Abstract

Alcohol-associated liver disease is a spectrum of liver disorders with histopathological changes ranging from simple steatosis to steatohepatitis, cirrhosis, and hepatocellular carcinoma. Recent data suggest that chronic-plus-binge ethanol intake induces steatohepatitis by promoting release by hepatocytes of proinflammatory mitochondrial DNA–enriched (mtDNA-enriched) extracellular vesicles (EVs). The aim of the present study was to investigate the role of the stress kinase apoptosis signal–regulating kinase 1 (ASK1) and p38 mitogen-activated protein kinase (p38) in chronic-plus-binge ethanol–induced steatohepatitis and mtDNA-enriched EV release. Microarray analysis revealed the greatest hepatic upregulation of metallothionein 1 and 2 (Mt1/2), which encode 2 of the most potent antioxidant proteins. Genetic deletion of the Mt1 and Mt2 genes aggravated ethanol-induced liver injury, as evidenced by elevation of serum ALT, neutrophil infiltration, oxidative stress, and ASK1/p38 activation in the liver. Inhibition or genetic deletion of Ask1 or p38 ameliorated ethanol-induced liver injury, inflammation, ROS levels, and expression of phagocytic oxidase and ER stress markers in the liver. In addition, inhibition of ASK1 or p38 also attenuated ethanol-induced mtDNA-enriched EV secretion from hepatocytes. Taken together, these findings indicate that induction of hepatic mtDNA-enriched EVs by ethanol is dependent on ASK1 and p38, thereby promoting alcoholic steatohepatitis.

Authors

Jing Ma, Haixia Cao, Robim M. Rodrigues, Mingjiang Xu, Tianyi Ren, Yong He, Seonghwan Hwang, Dechun Feng, Ruixue Ren, Peixin Yang, Suthat Liangpunsakul, Jian Sun, Bin Gao

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Figure 9

ASK1/p38 pathway activation contributes to elevated release of mtDNA enriched EVs in E10d+1B model.

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ASK1/p38 pathway activation contributes to elevated release of mtDNA enr...
(A) WT, Ask1–/–, and p38aHep–/– mice were subjected to E10+1B ethanol feeding and were euthanized 9 hours after gavage. Serum EVs were isolated and quantified by exosome quantitation assay. Cytochrome c oxidase (Cyto c ox) DNA levels in EVs were measured. (B) C57BL/6J mice were subjected to E10d+1B feeding and received i.p. injection of ASK1 or p38 inhibitors 30 minutes before gavage. Mice were euthanized 9 hours after gavage. Serum EVs were isolated and counted by exosome quantitation assay. Cyto c ox DNA levels in EVs were measured. (C) Hepatocytes were isolated from WT, Ask1–/–, or p38aHep–/– mice and treated with ethanol (100 mM) or PBS for 24 hours. EVs were isolated from culture medium and counted by exosome quantitation assay. (D and E) Hepatocytes were isolated from WT mice and treated with ethanol (100 mM) or ethanol plus ASK1 or p38 inhibitors for 24 hours. EVs were isolated from culture medium and counted by exosome quantitation assay. Cyto c ox DNA levels in the EVs were measured. Values represent mean ± SEM (each dot represents 1 mouse sample). Statistical evaluation was performed by Student’s t test or 1-way ANOVA with Tukey’s post hoc test for multiple comparisons. *P < 0.05, **P < 0.01, ***P < 0.001.