Han Yu, Zhao Jun, Liao Xiuge, Wang Ruifeng, Dong Lixia
Department of Respiratory and Critical Care Medicine, Tianjin Medical University General Hospital Affiliated to Tianjin Medical University, No.154 Heping Road to Anshan, Tianjin City, 300052, People's Republic of China.
Department of Respiratory and Critical Care Medicine, Central Hospital of Ordos City, Ordos, Inner Mongolia Autonomous Region, China.
Appl Biochem Biotechnol. 2025 May;197(5):2927-2943. doi: 10.1007/s12010-024-05168-y. Epub 2025 Jan 14.
Dysregulated circular RNAs (circRNAs) has been revealed to be involved in pulmonary fibrosis progression. Herein, this study focused on exploring the function and mechanism of circRNA Zinc Finger MYM-Type Containing 2 (circZMYM2) on idiopathic pulmonary fibrosis (IPF) using transforming growth factor (TGF)-β1-stimulated fibroblasts. Human fibroblast cell lines IMR-90 and HFL1 were stimulated with TGF-β1 to mimic fibrosis condition in vitro. Levels of genes and proteins were detected by qRT-PCR and western blotting. Cell proliferation and migration were analyzed using cell counting kit-8 assay, 5-Ethynyl-2'-deoxyuridine (EdU) and wound healing assays. The fibrosis progression was determined by the change of E-cadherin, α-smooth muscle actin (α-SMA), collagen type I α 1 (COL1A1) and collagen type III α 1 (COL3A1). The interaction between miR-199b-5p and circZMYM2 or KLF13 (Kruppel Like Factor 13) was analyzed using dual-luciferase reporter, RIP and RNA-pull-down assays. CircZMYM2 was decreased in TGF-β1-induced IMR-90 and HFL1 fibroblasts. Functionally, re-expression of circZMYM2 in IMR-90 and HFL1 cells could attenuate TGF-β1-evoked proliferation, migration and fibrosis in cells. Mechanistically, the circZMYM2/miR-199b-5p/KLF13 constituted a competing endogenous RNA (ceRNA). TGF-β1 reduced KLF13 expression and increased miR-199b-5p expression in IMR-90 and HFL1 cells. Further rescue experiments suggested that miR-199b-5p up-regulation or KLF13 knockdown reversed the anti-fibrotic effects of circZMYM2; moreover, silencing of miR-199b-5p exhibited anti-fibrotic effects, which was counteracted by KLF13 knockdown. CircZMYM2 had an anti-fibrotic effect that could suppress fibroblast activation via miR-199b-5p/KLF13 axis, pointing a novel perspective into the potential action pattern of circ_0022383 in IPF.
失调的环状RNA（circRNAs）已被揭示参与肺纤维化的进展。在此，本研究聚焦于利用转化生长因子（TGF）-β1刺激的成纤维细胞探索环状RNA锌指MYM型包含2（circZMYM2）在特发性肺纤维化（IPF）中的功能及机制。用人成纤维细胞系IMR-90和HFL1用TGF-β1刺激以在体外模拟纤维化状态。通过qRT-PCR和蛋白质印迹法检测基因和蛋白质水平。使用细胞计数试剂盒-8检测、5-乙炔基-2'-脱氧尿苷（EdU）和伤口愈合检测分析细胞增殖和迁移。通过E-钙黏蛋白、α-平滑肌肌动蛋白（α-SMA）、I型胶原蛋白α1（COL1A1）和III型胶原蛋白α1（COL3A1）的变化确定纤维化进展。使用双荧光素酶报告基因、RIP和RNA下拉检测分析miR-199b-5p与circZMYM2或Kruppel样因子13（KLF13）之间的相互作用。在TGF-β1诱导的IMR-90和HFL1成纤维细胞中circZMYM2表达降低。在功能上，在IMR-90和HFL1细胞中重新表达circZMYM2可减弱TGF-β1诱导的细胞增殖、迁移和纤维化。机制上，circZMYM2/miR-199b-5p/KLF13构成竞争性内源RNA（ceRNA）。TGF-β1降低IMR-90和HFL1细胞中KLF13表达并增加miR-199b-5p表达。进一步的挽救实验表明，miR-199b-5p上调或KLF13敲低可逆转circZMYM2的抗纤维化作用；此外，沉默miR-199b-5p表现出抗纤维化作用，而KLF13敲低可抵消该作用。CircZMYM2具有抗纤维化作用，可通过miR-199b-5p/KLF13轴抑制成纤维细胞活化，为circ_0022383在IPF中的潜在作用模式指明了新的视角。
