长链非编码RNA核富集转录本1通过靶向微小RNA-761对缺氧/复氧大鼠星形胶质细胞损伤的影响

Effect of long non-coding RNA nuclear-enriched abundant transcript on hypoxia-reoxygenation rat astrocyte injury by targeting microRNA-761

目的探讨长链非编码RNA(lncRNA)核富集转录本1(NEAT1)对缺氧/复氧(H/R)胶质细胞损伤的影响,及其机制是否与调控微小RNA-761(miR-761)有关。方法构建大鼠皮质星形胶质细胞H/R损伤模型。分为对照组、H/R组、H/R+小干扰RNA阴性对照(si-NC)组、H/R+si-NEAT1组、H/R+miR-NC组、H/R+miR-761组、H/R+si-NEAT1+miRNA抑制物(anti-miR-NC)组、H/R+si-NEAT1+anti-miR-761组。Real-time PCR分析NEAT1和miR-761的mRNA表达。流式细胞术分析细胞凋亡。试剂盒检测细胞中丙二醛(MDA)含量以及超氧化物歧化酶(SOD)和过氧化氢酶(CAT)的活性。双荧光素酶报告基因实验和Real-time PCR分析NEAT1和miR-761靶向关系。实验重复3次。结果与对照组比较,H/R组细胞凋亡率和MDA含量显著升高,SOD和CAT活性显著降低,NEAT1表达显著升高,miR-761表达显著降低(P<0.05)。与H/R+si-NC组比较,H/R+si-NEAT1组细胞凋亡率和MDA含量显著降低,SOD和CAT活性显著升高(P<0.05)。与H/R+miR-NC组比较,H/R+miR-761组细胞凋亡率和MDA含量显著降低,SOD和CAT活性显著升高(P<0.05)。MiR-761是NEAT1的靶基因,NEAT1负调控miR-761表达。与H/R+si-NEAT1+anti-miR-NC组比较,H/R+si-NEAT1+anti-miR-761组细胞凋亡率、MDA含量显著升高,SOD和CAT活性显著降低(P<0.05)。结论干扰NEAT1表达通过上调miR-761对H/R损伤的星形胶质细胞具有保护作用。

Objective To investigate the effect of long non-coding RNA(lncRNA)nuclear-enriched abundant transcript 1(NEAT1)on hypoxia-reoxygenation(H/R)glial astrocyte injury,and to explore whether the mechanism was related to the regulation of micro RNA(miR)-761.Methods Rat cortical astrocytes were cultured to construct a H/R injury model.Astrocytes were divided into control group,model group,model+small interfering RNA negative control(si-NC)group,model+si-NEAT1 group,model+miR-NC group,model+miR-761 group,model+si-NEAT1+anti-miR-NC group,model+si-NEAT1+anti-miR-761 group.Expression of NEAT1 and miR-761 were detected by Real-time PCR.The experiment was repeated 3 times.The content of malonaldehyde(MDA),and the activity of superoxide dismutase(SOD)and catalase(CAT)were detected by kits.Dual luciferase reporter experiment and Real-time PCR were used to analyze the targeting relationship between NEAT1 and miR-761.The experiment was repeated 3 times.Results Compared with the control group,the cell apoptosis rate and MDA content of the model group increased significantly,SOD and CAT activities decreased significantly,NEAT1 expression increased significantly,and miR-761 expression decreased significantly(P<0.05).Compared with the model+si-NC group,the apoptosis rate and MDA content of the model+si-NEAT1 group reduced significantly,and SOD and CAT activities increased significantly(P<0.05).Compared with the model+miR-NC group,the apoptosis rate and MDA content of the model+miR-761 group reduced significantly,and SOD and CAT activities increased significantly(P<0.05).MiR-761 was the target gene of NEAT1,and NEAT1 negatively regulated miR-761 expression.Compared with the model+si-NEAT1+anti-miR-NC group,the apoptosis rate and MDA content of the model+si-NEAT1+anti-miR-761 group increased significantly,and SOD and CAT activities decreased significantly(P<0.05).Conclusion Interference with NEAT1 expression can protect astrocytes from H/R injury by up-regulating miR-761.