lncIAPF介导的TGFβ1-lncIAPF-HuR-TGFβ1正反馈环路和lncIAPF-HuR-EZH2/STAT1自噬通路在特发性肺纤维化发生中的作用及其作用机制

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, and devastating interstitial lung disease with a median survival of 3–5 years. IPF remains a disease with an incompletely understood molecular mechanism, thereby leading to poor diagnosis/prognosis and limited clinical therapy. . Long non-coding RNA(lncRNA) that comprise more than 200 nucleotides has attracted considerable attention because of its wide range of biological regulatory functions, such as cell proliferation, migration, differentiation, and apoptosis, as well as immune responses, splicing, and protein localization. The influence of lncRNAs is achieved by transcriptional, post-transcriptional, and translational interferences. Emerging evidence has purported lncRNA as a new class of players involved in the development and progression of various diseases, such as cancer, steatohepatitis, and myocardial and liver fibrogenesis. Shen et al. demonstrated the roles of lncRNA Haunt in regulating the HOXA gene cluster during embryonic stem cell differentiation. Song et al. found the interaction of NF-kappa B with lncRNA is linked with breast cancer metastasis and patient prognosis. Although a set of human lncRNAs have been identified, their expression patterns, characteristics, and mechanism in IPF remain largely unexplored. . Our previous study has found that lncIAPF (Inhibited Autophagy in Pulmonary Fibrosis) can promote the development of fibrosis by inhibiting autophagy. The silence smart targeted lncIAPF can significantly upregulate the expression of autophagy-related genes ULK1, Atg5, LC3-II, Atg4, Atg12 and Atg16L1, and downregulate SQSTM1/p62. This project intends to use the gain- and loss-of-function of lncIAPF constructed by genome editing technology to obtain cell pulmonary fibrosis model, mouse pulmonary fibrosis model and IPF patient specimens. Our aims are to investigate whether TGFβ1, as a regulatory upstream molecule of lncIAPF, can enhance smad2/3 enrichment in the lncIAPF promoter to promote the transcription of lncIAPF, lncIAPF regulate its target gene HuR to facilitate the nucleosome shuttle of HuR protein and the lncIAPF- HuR target the autophagy-related gene EZH2/STAT1 leading to inhibiting the process of autophagy and promoting the process of fibrosis. The completion of our project will provide important experimental data for the clinical application of lncIAPF and identifying new drug and gene therapy targets.

特发性肺纤维化(IPF)是一种弥漫性肺间质疾病，成纤维细胞自噬平衡失调是其主要发病机制之一。项目组前期研究发现一种新的lncRNA-IAPF能够通过抑制自噬作用促进纤维化的发生，特异针对lncIAPF的silence smart能显著上调自噬相关基因ULK1、Atg5、LC3-II 、Atg4、Atg12、Atg16L1的表达，下调SQSTM1/p62的表达。本项目拟采用基因组编辑技术构建lncIAPF功能获得和缺失成纤维细胞、小鼠肺纤维化模型和IPF患者标本，探究lncIAPF介导的TGFβ1-lncIAPF-HuR-TGFβ1正反馈环路和lncIAPF-HuR-EZH2/STAT1自噬通路调控IPF发生的分子机制，为lncIAPF的临床应用和鉴定新的药物和基因治疗靶点提供重要的实验数据。

特发性肺纤维化（IPF）是一种弥漫性肺间质疾病，成纤维细胞自噬平衡失调是其主要发病机制之一。该项目鉴定了一个在IPF中上调表达的lncRNA，根据其功能将其命名为lncIAPF (Inhibited Autophagy in Pulmonary Fibrosis) 。功能实验发现lncIAPF可通过促进成纤维细胞向肌成纤维细胞分化、增殖和迁移并抑制自噬流促进肺纤维化进程。ATAC测序结果显示成纤维细胞分化中lncIAPF染色质可及性增强、开放区域组蛋白H3K27乙酰化水平显著增高，且转录因子ATF3在启动子区高度富集，促进lncIAPF基因转录。分段RNA pull-down、RNAase-RIP、半衰期和泛素化等实验表明，lncIAPF在1330 -1613 bp段与HuR结合形成RNA-蛋白复合体，并能延长各自半衰期增加稳定性。实时自噬流观测和自噬PCR Array等实验发现lncIAPF-HuR复合体通过促进靶基因EZH2、STAT1和FOXK1稳定性，阻断自噬小体与溶酶体融合诱导参与调控肌成纤维细胞表型。体内实验证实高表达lncIAPF可促进小鼠肺纤维化进程且被HuR干扰片段逆转，同时发现lncIAPF在IPF患者血液中显著高表达且与IPF疾病临床指标密切相关。该项目探究了lncIAPF介导的lncIAPF-HuR-EZH2/STAT1自噬通路调控IPF发生的分子机制，为lncIAPF的临床应用和鉴定新的药物和基因治疗靶点提供了重要的实验数据和设计思路。

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