趋化因子C-C基序配体6对心肌细胞糖氧剥夺损伤的保护作用和机制研究

摘要

目的 研究趋化因子C-C基序配体6(chemokine C-C motif ligand 6,CCL6)对急性心肌细胞糖氧剥夺损伤的保护作用及其可能的分子机制.方法 在公共基因芯片数据库(GEO)下载,并分析假手术组(Sham组)和缺血-再灌注组(IR组)心肌组织基因表达差异.体外培养大鼠心肌细胞H9C2,通过氧葡萄糖剥夺(oxygen glucose deprivation,OGD)方法建立心肌细胞损伤模型.采用MTT法检测细胞活力;采用Annex V/PI双染检测细胞凋亡;采用荧光定量PCR和Western blot检测相关基因表达情况.结果 和Sham组相比,基因芯片分析和荧光定量PCR结果显示缺血-再灌注大鼠心肌组织中CCL6表达水平显著降低(P＜0.01).氧葡萄糖剥夺可诱导H9C2心肌细胞中CCL6表达水平降低,且呈现时间依赖性.此外,氧葡萄糖剥夺导致H9C2心肌细胞活力降低,凋亡增加;而加入CCL6则促进细胞活力并减少细胞凋亡率.lncRNA芯片和荧光定量PCR显示,CCL6处理心肌细胞后导致lncRNA IGF2-AS表达水平显著降低,并进一步导致Akt和GSK-3β磷酸化水平增强.结论 CCL6在一定程度上可以抑制糖氧剥夺诱导的心肌细胞损伤,其分子机制可能与抑制IGF2-AS以及增强Akt/GSK-3β信号通路相关.%Objective To study the protective effect of chemokine C-C motif ligand 6 (CCL6) on glucose-oxygen deprivation induced injury in cardiomyocytes and its possible molecular mechanism.Methods Gene expression was analyzed in the public database Gene Expression Omnibus (GEO) database and gene expression of analyzed for myocardial tissue was analyzed gene expression in the sham group and the ischemia-reperfusion group (IR group).Rat H9C2 cardiomyocytes were cultured in vitro,and myocardial cell injury model was established by oxygen glucose deprivation (OGD).Cell viability was detected by MTT assay;apoptosis was determined by Annex V/PI double staining;the expression of related genes was detected by real-time PCR and Western blot.Results Compared with the sham group,transcriptome analysis and real-time PCR showed that the expression of CCL6 in the myocardial tissue of the IR group was significantly decreased (P＜0.01).Oxygen glucose deprivation induced a decrease in CCL6 expression levels in H9C2 cardiomyocytes in a time-dependent manner.In addition,oxygen glucose deprivation leads to decreased cell viability and increased apoptosis;while addition of CCL6 promotes cell viability and reduces apoptosis.The IncRNA microarray and real-time PCR showed that CCL6 treatment of cardiomyocytes resulted in a significant decrease in the expression of hicRNA IGF2-AS and further increased the phosphorylation of Akt and GSK-3β.Conclusion CCL6 can inhibit cardiomyocyte injury induced by glucose deprivation,and its molecular mechanism may be related to inhibition of IGF2-AS and enhancement of Akt/GSK-3β signaling pathway.