Dai Zhenzhao, Hu Jiansheng, Luo Zhiying, Xiao Jianhua
Department of Pediatrics, Affiliated Hospital of Jinggangshan University, Ji'an, Jiangxi, China.
Jinggangshan University, 28 Xueyuan Road, Qingyuan District, Ji'an, 343000, Jiangxi, China.
Biochem Genet. 2024 Apr;62(2):915-930. doi: 10.1007/s10528-023-10455-0. Epub 2023 Jul 27.
Circular RNAs (circRNAs) have been confirmed to mediate infantile pneumonia development. In this, we investigated the role and new mechanism of circ_0035292 regulating infantile pneumonia progression. Lipopolysaccharide (LPS)-treated WI-38 cells were used to mimic infantile pneumonia cell injury models. Quantitative real-time PCR was used to measure circ_0035292, microRNA (miR)-494-3p and toll-like receptor 4 (TLR4). Cell proliferation and apoptosis were assessed by MTT assay, EdU assay, and flow cytometry. Protein expression was tested using western blot analysis. Inflammation and oxidative stress were evaluated by measuring IL-6, IL-1β, MDA and SOD levels using ELISA assay and corresponding kits. RNA interaction was confirmed by dual-luciferase reporter assay and RIP assay. Circ_0035292 had elevated expression in infantile pneumonia patients and LPS-induced WI-38 cells. Silenced circ_0035292 could enhance WI-38 cell proliferation, while suppress apoptosis, inflammation and oxidative stress under LPS treatment. Mechanically, circ_0035292 targeted miR-494-3p to positively regulate TLR4. The rescue experiments indicated that miR-494-3p inhibitor abolished the function of circ_0035292 knockdown, and TLR4 overexpression reversed the inhibitory effect of miR-494-3p on LPS-induced WI-38 cell injury. Circ_0035292 might be a potential target for infantile pneumonia treatment, which knockdown could relieve LPS-induced cell injury via the regulation of miR-494-3p/TLR4 axis.
环状RNA（circRNAs）已被证实可介导婴幼儿肺炎的发展。在此研究中，我们探究了circ_0035292调控婴幼儿肺炎进展的作用及新机制。使用脂多糖（LPS）处理的WI-38细胞来模拟婴幼儿肺炎细胞损伤模型。采用定量实时PCR检测circ_0035292、微小RNA（miR）-494-3p和Toll样受体4（TLR4）。通过MTT法、EdU法和流式细胞术评估细胞增殖和凋亡。使用蛋白质印迹分析检测蛋白质表达。通过ELISA检测试剂盒检测IL-6、IL-1β、丙二醛（MDA）和超氧化物歧化酶（SOD）水平来评估炎症和氧化应激。通过双荧光素酶报告基因检测和RNA免疫沉淀（RIP）检测来确认RNA相互作用。circ_0035292在婴幼儿肺炎患者和LPS诱导的WI-38细胞中表达升高。沉默circ_0035292可增强WI-38细胞增殖，同时抑制LPS处理下的细胞凋亡、炎症和氧化应激。机制上，circ_0035292靶向miR-494-3p以正向调节TLR4。挽救实验表明，miR-494-3p抑制剂消除了circ_0035292敲低的作用，而TLR4过表达逆转了miR-494-3p对LPS诱导的WI-38细胞损伤的抑制作用。circ_0035292可能是婴幼儿肺炎治疗的潜在靶点，敲低该靶点可通过调节miR-494-3p/TLR4轴减轻LPS诱导的细胞损伤。
