CircRNA97编码的截短型SMOX在肝纤维化逆转中的作用机制研究

Oxidative stress and inflammation induced by chronic liver injury are two important factors for the occurrence of liver fibrosis. Using high-throughput sequencing screening approach, we previously showed a significant downregulation of circRNA97 in the fibrotic liver, but the mechanism is still unclear. Recent studies showed that circRNA97 encodes a truncated peptide SMOX (circSMOX). Furthermore, it has been reported that SMOX protein is upregulated by inflammation and significantly elevated in the fibrotic liver, and that SMOX can promote tumorigenesis by inducing oxidative stress. Previous experiments of the applicant also confirmed the high expression of SMOX protein in liver fibrosis tissue, and the up-regulation of circSMOX can inhibit the activity of TGF-β and SMOX.Accordingly, a hypothesis was proposed that circRNA97 could inhibit the oxidative damage of SMOX protein by encoding SMOX peptide, thereby affecting the development of liver fibrosis. To examine this hypothesis, this application is aimed to investigate (1) the potential binding of circSMOX with SMOX and its impacts on the formation of active SMOX dimer in vitro; (2) the functions of circSMOX and circRNA97 silencing in regulating the development of liver fibrosis in vivo liver fibrosis model; (3) finally their regulatory roles in patients with liver fibrosis. These experiments proposed here will provide better understanding for the development of liver fibrosis and novel evidence for the discovery of new therapeutic targets.

慢性肝损伤导致的氧化应激和炎症反应是肝纤维化发生发展的重要因素。前期通过高通量测序显示肝纤维化组织中circRNA97显著降低，但作用机制尚不明确，进一步研究发现circRNA97能够编码一个截短型SMOX肽段(circSMOX)。既往文献报道SMOX蛋白由炎症诱导并通过氧化应激可导致肿瘤的发生。申请者在预实验也证实SMOX蛋白在肝纤维化组织中高表达，上调circSMOX能够抑制TGF-β和SMOX的活性。据此提出假说circRNA97通过编码SMOX肽段拮抗SMOX蛋白活性抑制氧化损伤，从而影响肝纤维化发展的进程。为验证假说，本申请拟在体外实验研究circSMOX结合SMOX蛋白的区域和位点，以及对活性SMOX二聚体形成的影响；通过构建肝纤维化大鼠模型，探讨circRNA97编码circSMOX逆转肝纤维化的作用机制；并在临床水平进行验证。为阐明肝纤维化的发病机制和治疗靶点提供新思路。

慢性肝损伤导致的氧化应激和炎症反应是肝纤维化发生发展的重要因素。SMOX蛋白具有催化精胺产生活性氧的作用，且研究表明SMOX的表达水平与肝细胞癌临床症状的严重性呈正相关关系，提示SMOX可能促进肝细胞癌的发生。近年来发现circRNA与许多疾病的发生和发展密切相关。前期通过高通量测序显示肝纤维化组织中circRNA97显著降低，但作用机制尚不明确，进一步研究发现circRNA97能够编码一个截短型SMOX肽段(circSMOX)。在此基础上，本项目组通过动物实验和体外实验发现：（1）上调circSMOX能够抑制肝纤维化相关蛋白、TGF-β和SMOX的表达。（2）在肝纤维化组织中检测到SMOX上调和miR-139-5p下调。（3）过表达的miR-139-5p可显著抑制LPS诱导的HSC-T6细胞的活化，降低活性氧水平，同时下调SMOX、α-SMA 和 TGF-β等肝纤维化相关因子。（4）SMOX是miR-139-5p的直接靶点，miR-139-5p通过靶向和下调SMOX抑制肝纤维化的进展。本项目组研究提示circRNA97可编码circSMOX并通过介导miR-139-5p靶向作用于SMOX从而抑制肝纤维化的进展，为阐明肝纤维化的发病机制和治疗靶点提供新思路。