(The) role of exosomal mitochondrial double-stranded RNA-mediated activation of toll-like receptor 3 in kupffer cells in alcoholic liver disease

Abstract

Alcohol consumption contributes ROS production by CYP2E1 in mitochondria and recent studies show that mitochondrial double-stranded RNA (mtdsRNA) produced by ROS affects antiviral signaling. Hepatic extracellular vesicles (EVs) can be an intercellular modulator when released into extracellular milieu. Here, we report alcohol increases mtdsRNA and stimulates toll-like receptor 3 (TLR3) in Kupffer cells (KCs), enhancing IL-17A production in $\gamma \delta$ T cells in alcoholic liver disease (ALD). ALD was induced by binge ethanol drinking in wild-type (WT) and TLR3 knock-out mice with or without Kupffer cell depletion by clodronate. In intracellular cytokine analysis, IL-17A producing cells were analyzed among liver mononuclear cells (MNCs). To investigate DMSE 20165395 이준희. 알코올성 간질환에서 엑소좀 내 미토콘드리아 이중 가닥 리보핵산 매개 톨-유사 수용체 3 활성화를 통한 쿠퍼세포의 역할 규명. 의과학대학원. 2020년. 57+ⅲ 쪽. 지도교수: 정원일. (영문 논문) Jun-Hee Lee. The Role of Exosomal Mitochondrial Double-Stranded RNA-Mediated Activation of Toll-Like Receptor 3 in Kupffer Cells in Alcoholic Liver Disease. Graduate School of Medical Science and Engineering. 2020. 57+iii pages. Advisor: Won-Il Jeong. (Text in English) the presence of mtdsRNA, immunofluorescent staining and RNA sequencing were performed in exosomes. After binge ethanol drinking, $\gamma \delta$ T cells were the most IL-17A-producing cells and the expression of IL-1$\beta$ increase in KCs. IL-17A production in $\gamma \delta$ T cells was significantly decreased when KCs were depleted by clodronate. EVs isolated from EtOH-treated hepatocytes increased the expression of IL-1$\beta$ in KCs. The expression of PNPase, an RNA restricting enzyme, was significantly decreased in ethanol-treated livers. We found accumulation of dsRNA in mitochondria of ethanol-treated hepatocytes by J2 immunostaining and entrapment of J2 foci in exosomes when treated into KCs. We identified enrichment of small-sized RNA in exosomes by Bioanalyzer analysis. When ablated TLR3 in KCs, IL-17A decreased in $\gamma \delta$ T cells after binge drinking. In RNA sequencing data, exosomes contain small-sized RNA contents from mitochondrial origin. MtdsRNA generated by ethanol-induced suppression of PNPase transffered into nearby KCs and TLR3 activation through exosomal delivery. Consequently, increased IL-1$\beta$ in KCs augments IL-17A production in $\gamma \delta$ T cells in ALD. Therefore, mtdsRNA and TLR3 may function as novel therapeutic targets in ALD.