miR-873通过靶向Beclin1基因调控细胞自噬促进支气管上皮细胞炎症与凋亡

MiR-873 regulates cell autophagy by targeting Beclin1 to promot inflammation and apoptosis of bronchial epithelial cells

目的:探究miR-873对支气管上皮细胞凋亡、自噬的影响,以及对Beclin1的调控作用。方法:使用miR-873 mimic转染16HBE细胞48 h,qRT-PCR检测miR-873的mRNA水平,并使用CCK-8实验评估细胞活力。使用ELISA试剂盒检测细胞上清液中IL-2、IL-6、IL-10和TNF-α的含量,并采用TUNEL染色法检测细胞凋亡情况。利用免疫荧光技术检测LC-3蛋白表达情况,以及通过qRT-PCR和Western blot技术检测Beclin1基因的mRNA和蛋白表达水平。同时,采用双荧光素酶报告基因技术检测miR-873与Beclin1的结合位点。结果:miR-873 mimic转染显著上调了16HBE细胞中miR-873的mRNA水平,抑制了16HBE细胞的增殖,并促进了其分泌IL-2、IL-6和TNF-α,同时抑制了IL-10的分泌。此外,miR-873还促进了16HBE细胞的凋亡并抑制了LC-3的表达。双荧光素酶报告基因证实miR-873与Beclin1基因存在结合位点,并可以靶向抑制Beclin1的mRNA和蛋白表达。结论:miR-873可能通过靶向Beclin1基因调控细胞自噬,从而促进支气管上皮细胞炎症与凋亡。

Objective:Investigating the effects of miR-873 on apoptosis and autophagy in bronchial epithelial cells,as well as its regulatory role on Beclin1.Methods:Following transfection of miR-873 mimic into 16HBE cells for 48 hours,the mRNA level of miR-873 was quantified by qRT-PCR,and cell viability was evaluated by CCK-8 assay.The levels of IL-2,IL-6,IL-10,and TNF-αin the cell supernatant were determined using ELISA assay,while cell apoptosis was detected by TUNEL staining.LC-3 protein expression was examined by immunofluorescence,and mRNA and protein expression levels of Beclin1 were ana-lyzed by qRT-PCR and Western blot,respectively.Moreover,dual-luciferase reporter gene technology was employed to investi-gate the binding site between miR-873 and Beclin1.Results:Transfection of miR-873 mimic into 16HBE cells significantly upreg-ulated the mRNA level of miR-873,which led to the inhibition of cell proliferation and the promotion of secretion of pro-inflammatory cytokines IL-2,IL-6,and TNF-α,while suppressing the secretion of anti-inflammatory cytokine IL-10.More-over,miR-873 induced cell apoptosis and inhibited the expression of LC-3.Dual-luciferase reporter gene assay further confirmed the presence of binding sites between miR-873 and Beclin1 gene.Besides,miR-873 could target and suppress the mRNA and pro-tein expression levels of Beclin1.Conclusion:miR-873 might modulate cell autophagy by targeting the Beclin1 gene,which can po-tentially promote inflammation and apoptosis in bronchial epithelial cells.