hTERT启动子介导的PRPS2基因表达下调对胶质瘤U251细胞增殖和凋亡的影响

The Influence of hTERT Promoter Mediated Down-regulation of PRPS2 on the Proliferation and Apoptosis of Glioma U251 Cells

目的探讨人端粒酶逆转录酶(hTERT)启动子介导的磷酸核糖焦磷酸合成酶2(PRPS2)表达下调对胶质瘤U251细胞生长增殖、细胞活力以及细胞周期和凋亡的影响。方法构建pG GN-hTERT-PRPS2干扰载体及阴性对照载体,转染U251细胞后;采用RT-PCR检测空白对照组、阴性对照组及实验组中PRPS2基因表达情况;采用Cell Counting Kit8细胞增殖试剂盒(CCK-8)检测细胞活力;采用Hoechst 33342染色检测细胞凋亡与坏死情况;进一步通过流式细胞术检测PRPS2对细胞周期和凋亡的影响。结果阴性对照组(转染阴性对照载体)与空白对照组相比,U251细胞中PRPS2基因表达及各项生物学特性均差异无统计学意义(P>0.05);实验组(转染pG GN-hTERT-PRPS2干扰载体)与阴性对照组相比,PRPS2表达下调60%,CCK-8结果显示细胞活力明显下降(P <0.05),Hoechst 33342染色后发现典型凋亡形态细胞,流式细胞术结果发现凋亡率显著上升(P <0.05),细胞阻滞于G0/G1期(P <0.05)。结论特异性下调U251细胞中PRPS2的表达,可降低细胞活力、抑制细胞增殖、促进细胞凋亡。

Objective To investigate the effect of human telomerase reverse transcriptase （hTERT） promoter mediated down- regulation of phosphoribosyl -pyrophosphate synthase 2 （PRPS2） on proliferation,cell viability,cell cycle,and apoptosis of U251 glioma cells.Methods The interference vector and negative control vector of PRPS2 gene were constructed.plasma was transfected into U251cells.The expression level of PRPS2 was detected by RT -PCR.The ability of cell proliferation was detected by CCK 8.The cell cycleand apoptosis were detected by flow cytometry （FCM） and Hoechst33342 dyeing.Results There were no significantly difference be -tween PRPS2 gene expression and the biological characteristics of U251 cells （ P 〉0.05） in the negative and blank control groups.The expression of PRPS2 decreased by 60% in the experimental group.And the PRPS2- knockdown significantly inhibited the viabili-ty and proliferation of U251 cells （P 〈0.05）.Hoechst33342 dyeing showed typical apoptotic cell.FCM result showed higher apotosisrate,and PRPS2- knockdown contributed to G0/G1 arrest （P 〈0.05）.Conclusion Specific down- regulation of PRPS2 expression inU251 cells could decrease cell viability and proliferation as well as promote cell apoptosis.