Laboratory of Liver Research, Graduate School of Medical Science and Engineering, Korea Advanced Institute of Science and Technology, Daejeon, Republic of Korea.
Laboratory of Liver Diseases, National Institute on Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD.
Hepatology. 2020 Aug;72(2):609-625. doi: 10.1002/hep.31041. Epub 2020 May 8.
Mitochondrial double-stranded RNA (mtdsRNA) and its innate immune responses have been reported previously; however, mtdsRNA generation and its effects on alcohol-associated liver disease (ALD) remain unclear. Here, we report that hepatic mtdsRNA stimulates toll-like receptor 3 (TLR3) in Kupffer cells through the exosome (Exo) to enhance interleukin (IL)-17A (IL-17A) production in ALD.
Following binge ethanol (EtOH) drinking, IL-17A production primarily increased in γδ T cells of wild-type (WT) mice, whereas the production of IL-17A was mainly facilitated by CD4 T cells in acute-on-chronic EtOH consumption. These were not observed in TLR3 knockout (KO) or Kupffer cell-depleted WT mice. The expression of polynucleotide phosphorylase, an mtdsRNA-restricting enzyme, was significantly decreased in EtOH-exposed livers and hepatocytes of WT mice. Immunostaining revealed that mtdsRNA colocalized with the mitochondria in EtOH-treated hepatocytes from WT mice and healthy humans. Bioanalyzer analysis revealed that small-sized RNAs were enriched in EtOH-treated Exos (EtOH-Exos) rather than EtOH-treated microvesicles in hepatocytes of WT mice and humans. Quantitative real-time PCR and RNA sequencing analyses indicated that mRNA expression of mitochondrial genes encoded by heavy and light strands was robustly increased in EtOH-Exos from mice and humans. After direct treatment with EtOH-Exos, IL-1β expression was significantly increased in WT Kupffer cells but not in TLR3 KO Kupffer cells, augmenting IL-17A production of γδ T cells in mice and humans.
EtOH-mediated generation of mtdsRNA contributes to TLR3 activation in Kupffer cells through exosomal delivery. Consequently, increased IL-1β expression in Kupffer cells triggers IL-17A production in γδ T cells at the early stage that may accelerate IL-17A expression in CD4 T cells in the later stage of ALD. Therefore, mtdsRNA and TLR3 may function as therapeutic targets in ALD.
先前已有报道称线粒体双链 RNA（mtdsRNA）及其固有免疫反应；然而，mtdsRNA 的产生及其对酒精相关肝病（ALD）的影响尚不清楚。在这里，我们报告肝 mtdsRNA 通过外泌体（Exo）刺激枯否细胞中的 toll 样受体 3（TLR3），以增强 ALD 中白细胞介素（IL）-17A（IL-17A）的产生。
在 binge 乙醇（EtOH）饮用后，IL-17A 的产生主要增加野生型（WT）小鼠的γδ T 细胞，而在急性慢性 EtOH 消耗中，IL-17A 的产生主要由 CD4 T 细胞促进。在 TLR3 敲除（KO）或枯否细胞耗尽的 WT 小鼠中未观察到这些现象。多核苷酸磷酸化酶的表达，一种限制 mtdsRNA 的酶，在 WT 小鼠的 EtOH 暴露肝脏和肝细胞中显著降低。免疫染色显示，mtdsRNA 与 WT 小鼠和健康人乙醇处理的肝细胞中的线粒体共定位。生物分析仪分析显示，在 WT 小鼠和人肝细胞中，乙醇处理的 Exos（EtOH-Exos）中富含小 RNA，而不是乙醇处理的微泡。定量实时 PCR 和 RNA 测序分析表明，来自小鼠和人的 EtOH-Exos 中重链和轻链编码的线粒体基因的 mRNA 表达显著增加。直接用 EtOH-Exos 处理后，WT 枯否细胞中 IL-1β 的表达显著增加，但 TLR3 KO 枯否细胞中没有增加，从而增加了小鼠和人 γδ T 细胞中 IL-17A 的产生。
乙醇介导的 mtdsRNA 的产生通过外泌体递送至枯否细胞中 TLR3 的激活。因此，枯否细胞中 IL-1β 表达的增加触发了 γδ T 细胞中 IL-17A 的产生，这可能在 ALD 的后期阶段加速 CD4 T 细胞中 IL-17A 的表达。因此，mtdsRNA 和 TLR3 可能是 ALD 的治疗靶点。
