表没食子儿茶素没食子酸酯对缺血再灌注损伤诱导的大鼠神经细胞凋亡的影响

目的探讨表没食子儿茶素没食子酸酯(EGCG)对缺血再灌注损伤诱导的大鼠神经细胞凋亡、氧化应激的影响及其对长链非编码RNA MALAT1(lncRNA MALAT1)/微小RNA-2115-3p(miR-2115-3p)分子轴的调控作用。方法体外培养大鼠神经元细胞HT22,建立氧糖剥夺/复氧损伤(OGD/R)神经细胞模型,随机分为Con组、OGD/R组、EGCG-L组、EGCG-M组、EGCG-H组、EGCG-H+pcDNA组、EGCG-H+pcDNA-MALAT1组。检测氧化应激指标超氧化物歧化酶(SOD)、丙二醛(MDA)的含量;流式细胞术检测细胞凋亡率;实时荧光定量聚合酶链反应(qRT-PCR)检测MALAT1、miR-2115-3p的表达量;双荧光素酶报告实验验证MALAT1、miR-2115-3p的靶向关系;蛋白免疫印迹法(Western blot)检测B淋巴细胞瘤-2相关蛋白(Bax)、B淋巴细胞瘤-2(Bcl-2)的表达量。结果与Con组比较,OGD/R组SOD含量降低(P<0.05),MDA含量升高(P<0.05),细胞凋亡率升高(P<0.05),Bax蛋白水平升高(P<0.05),Bcl-2蛋白水平降低(P<0.05),MALAT1表达水平升高(P<0.05),miR-2115-3p表达水平降低(P<0.05);与OGD/R组比较,EGCG-L组、EGCG-M组、EGCG-H组SOD含量升高(P<0.05),MDA含量降低(P<0.05),细胞凋亡率降低(P<0.05),Bax蛋白水平降低(P<0.05),Bcl-2蛋白水平升高(P<0.05),MALAT1表达水平降低(P<0.05),miR-2115-3p表达水平升高(P<0.05),且EGCG-L组、EGCG-M组、EGCG-H组各指标比较差异有统计学意义(P<0.05)。双荧光素酶报告实验证实MALAT1可靶向结合miR-2115-3p;MALAT1过表达可减弱EGCG对OGD/R诱导的神经细胞氧化应激及凋亡的作用。结论EGCG可通过调控MALAT1/miR-2115-3p分子轴,从而抑制缺血再灌注损伤诱导的大鼠神经细胞凋亡及减轻其氧化损伤。

Effect of epigalocatechin gallate on neuronal apoptosis induced by ischemia-reperfusion injury in rats

Objective To explore the effect of epigalocatechin galllate(EGCG)on neuronal apoptosis and oxidative stress induced by ischemia-reperfusion injury and its regulation on the molecular axis of lncRNA MALAT1/miR-2115-3 p.Methods The neuron cell of rats,HT22,was cultured in vitro to establish OGD/R injured nerve cell model,which was then randomly divided into Con group,OGD/R group,EGCG-L group,EGCG-M group,EGCG-H group,EGCG-H+pcDNA group,and EGCG-H+pcDNA-MALAT1 group.The content of oxidative stress indexes SOD and MDA was detected;flow cytometry was used to detect the apoptosis rate;qRT-PCR was used to detect the expression of MALAT1 and miR-2115-3 p;the dual luciferase report experiment was used to verify the targeting relationship between MALAT1 and miR-2115-3 p;Western blot was used to detect the expression of Bax and Bcl-2.Results Compared with Con group,the content of SOD in OGD/R group was reduced(P<0.05),the MDA content was increased(P<0.05),the apoptosis rate was increased(P<0.05),the level of Bax protein was increased(P<0.05),the level of Bcl-2 protein was reduced(P<0.05),the expression level of MALAT1 was increased(P<0.05),and the expression level of miR-2115-3 p was reduced(P<0.05).Compared with OGD/R group,the content of SOD in EGCG-L group,EGCG-M group and EGCG-H group was increased(P<0.05),the content of MDA was reduced(P<0.05),and the apoptosis rate was reduced(P<0.05),the level of Bax protein was reduced(P<0.05),the level of Bcl-2 protein was increased(P<0.05),the expression level of MALAT1 was reduced(P<0.05),the expression level of miR-2115-3 p was increased(P<0.05),and there were significant differences in the indexes among EGCG-L group,EGCG-M group and EGCG-H group(P<0.05).The dual luciferase report experiment confirmed that MALAT1 could targetedly bind to miR-2115-3 p.Overexpression of MALAT1 could reduce the effect of EGCG on OGD/R-induced neuronal oxidative stress and apoptosis.Conclusion EGCG can regulate the molecular axis of MALAT1/miR-2115-3 p to inhibit ischemia-reperfusion injury-induced neuronal cell apoptosis and reduce its oxidative damage.