摘要
目的:研究微小RNA-153(miR-153)对脂多糖(LPS)诱导的胚胎大鼠心肌H9C2细胞的炎症因子、细胞活力及凋亡的影响,并探讨其机制。方法:运用LPS建立H9C2细胞损伤模型;将细胞分为anti-miR-Con组(转染anti-miR-Con)、anti-miR-153组(转染anti-miR-153)、pcDNA组(转染pcDNA)、pcDNA-SORBS2组(转染pcDNA-SORBS2)、anti-miR-153+si-Con组(共转染anti-miR-153和si-Con)和anti-miR-153+si-SORBS2组(共转染anti-miR-153和si-SORBS2),转染后用LPS处理。RT-qPCR法检测细胞中miR-153和SORBS2 mRNA的表达;MTT法检测细胞活力;Western blot检测细胞中SORBS2的蛋白表达;ELISA实验检测细胞中肿瘤坏死因子α(TNF-α)和白细胞介素6(IL-6)的含量;流式细胞术检测细胞凋亡;双萤光素酶报告基因实验验证miR-153与SORBS2的靶向关系。结果:成功构建LPS诱导的H9C2细胞损伤模型;与对照的PBS组相比,LPS处理后H9C2细胞中miR-153的表达明显升高,SORBS2的表达明显降低;敲减miR-153表达或过表达SORBS2均可减少LPS诱导的TNF-α和IL-6释放及细胞凋亡,提高细胞活力;miR-153可抑制含野生型SORBS2的H9C2细胞的萤光素酶活性;敲减SORBS2的表达可逆转抑制miR-153对LPS诱导H9C2细胞的抗炎、抗凋亡及提高细胞活力的作用。结论:miR-153可促进LPS诱导的H9C2细胞炎症因子分泌和凋亡,抑制细胞的活力,发挥促进损伤的作用,其作用机制与靶向SORBS2有关。SORBS2可能成为心肌损伤治疗的新靶点。
AIM: To study the effects of microRNA-153(miR-153) on inflammatory factors, cell viability and apoptosis of embryonic rat H9 C2 cardiomyocytes induced by lipopolysaccharide(LPS), and to explore its mechanism. METHODS: The injury model of H9 C2 cells was established by LPS stimulation. The H9 C2 cells were divided into anti-miR-Con group(transfected with anti-miR-Con), anti-miR-153 group(transfected with anti-miR-153), pcDNA group(transfected with pcDNA), pcDNA-SORBS2 group(transfected with pcDNA-SORBS2), anti-miR-153+si-Con group(co-transfected with anti-miR-153 and si-Con) and anti-miR-153+si-SORBS2 group(co-transfected with anti-miR-153 and si-SORBS2), and treated with LPS after transfection. The expression of miR-153 and SORBS2 mRNA in the cells was detected by RT-qPCR. The viability of H9 C2 cells was measured by MTT assay. The protein expression of SORBS2 in the H9 C2 cells was determined by Western blot. The contents of tumor necrosis factor-α(TNF-α) and interleukin-6(IL-6) were detected by ELISA. The apoptosis of the H9 C2 cells was analyzed by flow cytometry. The targeting relationship between miR-153 and SORBS2 was verified by dual-luciferase reporter assay. RESULTS: The LPS-induced H9 C2 cell injury model was successfully constructed. Compared with PBS group, the expression of miR-153 was significantly increased and the expression of SORBS2 was significantly decreased in the H9 C2 cells treated with LPS. The inhibition of miR-153 and over-expression of SORBS2 decreased the contents of TNF-α and IL-6 and the level of apoptosis, but increased the cell viability. miR-153 inhibited the luciferase activity of the H9 C2 cells containing wild-type SORBS2. Inhibition of SORBS2 reversibly inhi-bited the anti-inflammatory effects of miR-153 on LPS-induced H9 C2 cells and increased the viability of the cells. CONCLUSION: miR-153 promotes the secretion of inflammatory factors, induces apoptosis, and inhibits the viability of H9 C2 cells induced by LPS, thus enhancing the damage. Its mechanism may be related to t
作者
吴瑞霞
黄粮
白玉鹏
刘晓刚
胡立群
WU Rui-xia;HUANG Liang;BAI Yu-peng;LIU Xiao-gang;HU Li-qun(Department of Cardiovascular Medicine,The Fourth Hospital of Wuhan,Wuhan 430000,China)
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2020年第1期22-28,共7页
Chinese Journal of Pathophysiology
基金
武汉市卫生和计划生育委员会科研项目(No.WX16B12)
