摘要
目的 探讨长链非编码RNA(lncRNA)SNHG15对脂多糖(LPS)诱导的肺泡上皮细胞损伤的影响及其分子机制.方法 LPS处理A549细胞,构建新生儿呼吸窘迫综合征(NRDS)细胞损伤模型;将A549细胞分为对照(Con-trol)组、模型(LPS)组、转染 si-NC 和 LPS 处理(LPS+si-NC)组、转染 si-lncRNA SNHG15 和 LPS 处理(LPS+si-ln-cRNA SNHG15)组、转染 miR-NC 和 LPS 处理(LPS+miR-NC)组、转染 miR-942-5p 和 LPS 处理(LPS+miR-942-5p)组、转染 si-lncRNA SNHG15、anti-miR-NC 和 LPS 处理(LPS+si-lncRNA SNHG15+anti-miR-NC)组和转染 si-lncRNA SNHG15、anti-miR-942-5p 和 LPS 处理(LPS+si-lncRNA SNHG15+anti-miR-942-5p)组;实时荧光定量 PCR(qRT-PCR)检测分子表达;流式细胞术和Western blot检测细胞凋亡;ELISA检测白介素-6(IL-6)、白介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)的表达水平.结果 与Control组比较,LPS组lncRNA SNHG15表达、细胞凋亡率、Bax、IL-6、IL-1β、TNF-α水平上调(均P＜0.05),而miR-942-5p表达、Bcl-2水平下调(均P＜0.05).沉默lncRNA SNHG15或过表达miR-942-5p降低了 LPS作用下的A549细胞凋亡率、Bax、IL-6、IL-1β、TNF-α水平,而升高了 Bcl-2水平(均P＜0.05).lncRNA SNHG15靶向miR-942-5p,且下调miR-942-5p逆转了沉默lncRNA SNHG15对LPS诱导A549细胞损伤的影响(均P＜0.05).结论 沉默lncRNA SNHG15通过靶向上调miR-942-5p减轻LPS诱导的A549细胞损伤.
Abstract
Objective To investigate the effects of long non-coding RNA(lncRNA)SNHG15 on lipopolysaccharide(LPS)-induced injury of human alveolar epithelial cells and the underlying molecular mechanism.Methods A549 cells were treated with LPS to construct a neonatal acute respiratory distress syndrome(NRDS)cell model.A549 cells were divided into Control group,LPS group,LPS+si-NC group,LPS+si-lncRNA SNHG15 group,LPS+miR-NC group,LPS+miR-942-5p group,LPS+si-lncRNA SNHG15+anti-miR-NC group and LPS+si-lncRNA SNHG15+anti-miR-942-5p group.Real-time fluorescence quantitative PCR(qRT-PCR)was used to detect the expression levels of lncRNA SNHG15 and miR-942-5p.Flow cytometry and Western blot were conducted to detect cell apoptosis.ELISA was used to detect the expression levels of interleukin-6(IL-6),in-terleukin-1β(IL-1β),and tumor necrosis factor-α(TNF-α).Dual-luciferase reporter assay was used to identify the targeting rela-tionship between lncRNA SNHG15 and miR-942-5p.Results Compared with the Control group,lncRNA SNHG15 expression,apoptosis rate,and levels of Bax,IL-6,IL-1β and TNF-α were increased in the LPS group,while miR-942-5p expression and Bcl-2 protein level were decreased(all P＜0.05).After knockdown of lncRNA SNHG15 or overexpression of miR-942-5p,cell apop-tosis rate and levels of Bax,IL-6,IL-1β and TNF-α were decreased,while Bcl-2 level was increased(all P＜0.05).lncRNA SNHG15 targeted miR-942-5p,and downregulation of miR-942-5p reversed the effect of lncRNA SNHG15 knockdown on LPS-induced A549 cell injury(all P＜0.05).Conclusion Silencing of lncRNA SNHG15 alleviates LPS-induced A549 cell injury by upregulation of miR-942-5p.
基金项目
上海市2021年度"科技创新行动计划"医学创新研究专项项目(21Y21900802)
万方数据