环状RNA608-微小RNA222-PINK1轴调控非酒精性脂肪性肝炎相关纤维化中肝星状细胞的线粒体自噬

CircRNA608-microRNA222-PINK1 axis regulates the mitophagy of hepatic stellate cells in NASH related fibrosis.

作者信息

Xu Zi-Xin, Li Jing-Ze, Li Qin, Xu Ming-Yi, Li Hui-Yi

机构信息

Department of Gastroenterology, Shanghai East Hospital, School of Medicine, Tongji University, Shanghai, 200092, China; Department of Gastroenterology, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200080, China.

Department of Gastroenterology, Shanghai East Hospital, School of Medicine, Tongji University, Shanghai, 200092, China.

出版信息

Biochem Biophys Res Commun. 2022 Jun 25;610:35-42. doi: 10.1016/j.bbrc.2022.04.008. Epub 2022 Apr 5.

DOI:10.1016/j.bbrc.2022.04.008

PMID:35436629

Abstract

BACKGROUND

Increasing evidences have confirmed the relationship between mitophagy and nonalcoholic steatohepatitis (NASH). The exact mechanism of upstream circular RNAs (circRNAs) regulating PTEN-induced putative kinase 1 (PINK1) mediated mitophagy and its contribution to NASH-related liver fibrosis was explored in our study.

METHODS

Primary hepatic stellate cells (PHSCs) from C57BL/6 mice transfected with small interfering RNAs against PINK1 (si-PINK1) and negative control (si-NC) were prepared to perform circRNA sequence. Differentially expressed circRNAs, bioinformatic analysis and predicting software were performed to select axis of circ608/miR-222/PINK1. The expressions of circ608/miR-222/PINK1 were verified by RT-qPCR. The mitochondrial function was evaluated by immunofluorescence staining of COX4 and LC3B.

RESULTS

PINK1-mediated mitophagy was inhibited in NASH-related liver fibrosis mice. CircRNA sequence revealed there were 37 DE-circRNAs between si-PINK1 PHSCs and si-NC PHSCs. Bioinformatic analysis showed these DE-circRNAs were related to enriched signaling pathways (such as Wnt, Rap1, mTOR, Hippo) regulating liver fibrosis and mitophagy. Circ608 was significantly down-regulated in lipotoxic HSCs and in livers of NASH-related liver fibrosis mice. MiR222 was identified to be the target miRNA of circ608 and was negatively regulated by circ608 in lipotoxic HSCs. MiR222 also had a binding site with PINK1 and could negatively regulate PINK1. So, the axis of circ608-miR222-PINK1 was proved to participate in NASH-related liver fibrosis by regulating mitophagy. These results illustrated that circ608 might promote PINK1-mediated mitophagy though inhibiting miR222 in lipotoxic HSCs.

CONCLUSION

Circ608 could promote PINK1-mediated mitophagy of HSCs though inhibiting miR222 in NASH-related liver fibrosis.

摘要

背景

越来越多的证据证实了线粒体自噬与非酒精性脂肪性肝炎（NASH）之间的关系。本研究探讨了上游环状RNA（circRNA）调控PTEN诱导的假定激酶1（PINK1）介导的线粒体自噬的确切机制及其对NASH相关肝纤维化的作用。

方法

制备用针对PINK1的小干扰RNA（si-PINK1）和阴性对照（si-NC）转染的C57BL/6小鼠原代肝星状细胞（PHSC），以进行circRNA测序。进行差异表达circRNA、生物信息学分析和预测软件以选择circ608/miR-222/PINK1轴。通过RT-qPCR验证circ608/miR-222/PINK1的表达。通过COX4和LC3B的免疫荧光染色评估线粒体功能。

结果

在NASH相关肝纤维化小鼠中，PINK1介导的线粒体自噬受到抑制。circRNA测序显示，si-PINK1 PHSC与si-NC PHSC之间有37种差异表达的circRNA（DE-circRNA）。生物信息学分析表明，这些DE-circRNA与调节肝纤维化和线粒体自噬的富集信号通路（如Wnt、Rap1、mTOR、Hippo）相关。Circ608在脂毒性肝星状细胞和NASH相关肝纤维化小鼠肝脏中显著下调。MiR222被鉴定为circ608的靶标miRNA，在脂毒性肝星状细胞中受circ608负调控。MiR222也与PINK1有结合位点，并可负调控PINK1。因此，circ608-miR222-PINK1轴被证明通过调节线粒体自噬参与NASH相关肝纤维化。这些结果表明，circ608可能通过在脂毒性肝星状细胞中抑制miR222来促进PINK1介导的线粒体自噬。