Gstm3在何首乌苷延缓SH-SY5Y细胞衰老中的作用

Effect of Gstm3 on Tetrahydroxy stilbene glycoside delaying senescence of SH-SY5Y cells

目的初步探讨Gstm3在何首乌苷延缓SH-SY5Y细胞衰老中的作用及可能机制。方法从针对Gstm3基因的4组含不同shRNA序列的干扰载体中筛选沉默效率最高的一组,作为Gstm3沉默组(shGstm3-P/D),同时设空白对照组(NC-P/D,未转染)及阴性对照组(shNC-P/D,转染无意义的阴性对照病毒载体shRNA序列),对照组(NC组,SH-SY5Y细胞)、D-Gal处理组(D-Gal组,SH-SY5Y细胞用D-Gal处理)、何首乌苷和D-Gal共同处理组(P/D组,SH-SY5Y细胞用何首乌苷和D-Gal共同处理);采用qPCR检测SH-SY5Y细胞中Gstm3 mRNA水平,细胞端粒长短,CCK8法检测细胞活力;Western blotting检测转染后各组细胞Gstm3、Bax、Bcl-2蛋白的表达,流式细胞术检测转染后各组细胞凋亡率。结果qPCR和WB检测结果显示,与NC组和shNC组比较shGstm3-P/D组SH-SY5Y细胞中Gstm3 mRNA和蛋白的表达明显降低(P<0.01);与NC组比较,D-Gal组端粒长度明显缩短,细胞活力下降;与D-Gal组比较,P/D组端粒长度明显增加,细胞活力升高;而与P/D组比较,shGstm3-P/D组端粒长度明显缩短,细胞活力明显下降(P<0.05);与NC-P/D组和shNC-P/D组比较,shGstm3-P/D组Bcl-2蛋白水平降低、Bax蛋白水平增加(P<0.05),且细胞凋亡率分别增加11.22%和9.07%(P<0.001)。结论Gstm3通过调控细胞凋亡相关蛋白Bcl-2、Bax的表达及细胞凋亡率参与何首乌苷延缓SH-SY5Y细胞衰老。

Objective To explore the role and possible mechanism of GSTM3 in(Tetrahydroxy stilbene glycoside,TSG)delaying the aging of SH-SY5Y cells.Methods The group with the highest silencing efficiency was selected from the four groups of interference vectors containing different shRNA sequences for the Gstm3 gene as the Gstm3 silencing group(shGstm3-P/D).At the same time,the blank control group(NC-P/D,not transfected)and the negative control group(shNC-P/D,transfected with meaningless negative control virus vector shRNA sequence)were set.Control group(NC group,SH-SY5Y cells),D-Gal treatment group(D-Gal group,SH-SY5Y cells treated with D-Gal),TSG and D-Gal Co-treatment group were set(P/D group,SH-SY5Y cells treated with TSG and D-Gal);qPCR was used to detect the level of Gstm3 mRNA in SH-SY5Y cells and the changes of telomere length;The cell viability was detected by CCK8;Western blotting was used to detect the expression level of Gstm3,Bax and Bcl-2 protein,and flow cytometry was used to detect the apoptosis rate.Results The results of qPCR and WB showed that the expression levels of Gstm3 mRNA and protein in SH-SY5Y cells in shGstm3-P/D group were significantly lower than those in NC-P/D group and shNC-P/D group(P<0.01);compared with NC group,the telomere length in D-Gal group was significantly shorter and cell viability decreased;compared with D-Gal group,the telomere length in P/D group was significantly increased and cell viability increased;compared with P/D group,the telomere length in shGstm3-P/D group was shorter and cell viability decreased significantly(P<0.05).Compared with NC-P/D group and shNC-P/D group,the level of Bcl-2 protein decreased,the level of Bax protein increased(P<0.05),and the apoptosis rate increased by 11.22%and 9.07%respectively(P<0.01).Conclusion Gstm3 participates in TSG delaying the aging of SH-SY5Y cells by regulating the expression of apoptosis related proteins Bcl-2 and Bax and the rate of apoptosis.