RNA干扰BLIMP1表达对NCI-H929细胞增殖凋亡的影响及可能机制

Effects of BLIMP1 gene silencing by RNA interference on the proliferation of NCI-H929 cells and related mechanisms

目的 探讨RNA干扰B淋巴细胞诱导成熟蛋白（B lymphocyte induced maturationprotein,BLIMP1）表达对多发性骨髓瘤（multiple myeloma,MM）细胞株NCI-H929生物学功能的影响及其可能机制.方法 用慢病毒干扰载体及阴性对照载体转染NCI-H929细胞后采用RT-PCR及Western blot方法观察BLIMP1 mRNA及蛋白表达,CCK-8法检测细胞增殖作用,Annexin V/PI双染色流式细胞术检测细胞凋亡,RT-PCR及Western blot检测未折叠蛋白反应（unfolded protein response,UPR）信号通路中X盒结合蛋白1（X-box binding protein 1,XBP1）、肌醇必需酶（inositol-requiring enzyme,IRE）1α及凋亡蛋白C/EBP同源蛋白（C/EBP homologous protein,CHOP）的变化.结果 BLIMP1 shRNA转染NCI-H929细胞后,BLIMP1 mRNA、蛋白表达均下调（P〈0.05）;shRNA/BLIMP1组24、48及72 h细胞增殖率低于转染空载体（shRNA/con）组和对照组（P〈0.05）,shRNA/BLIMP1组细胞凋亡率〔（10.87±1.79）%〕高于shRNA/con组〔（5.57±0.87）%〕和对照组〔（4.90±0.57）%〕（P〈0.01）;凋亡蛋白CHOP表达上调,XBP1、IRE1α表达下调.结论 沉默BLIMP1可以下调XBP1的表达而抑制UPR反应,发挥其抑制NCI-H929细胞增殖及促进凋亡作用.

Objective To investigate the effects of BLIMP1 gene silencing by RNA interference on the proliferation and apoptosis of multiple myeloma （MM） cell line NCI-H929 and related mechanisms.Methods NCI-H929 cells were transfected with a lentiviral vector or a control vector.Then,the levels of BLIMP1 mRNA and protein were detected by RT-PCR and Western blotting.The proliferation of the above cells was detected by CCK-8 assay.Cell apoptosis was detected by Annexin V/PI double staining flow cytometry.The levels of XBP1,IRE1α and CHOP mRNA and protein were detected by RT-PCR and Western blotting.Results After transfection of BLIMP1 mRNA,the levels of BLIMP1 mRNA and protein were markedly suppressed （P〈0.05）.The shRNA/BLIMP1 group showed suppressed proliferation of NCI-H929 cells at 24,48 and 72 h,compared with the shRNA/con and control groups.The cell apoptotic rate was （10.87±1.79）% in the shRNA/BLIMP1 group,which was higer than （5.57±0.87）% for the shRNA/con group and （4.90±0.57）% for the control group （P〈0.01）.BLIMP1 shRNA up-regulated the expression of CHOP,but down-regulated the expression of XBP1 and IRE1α.Conclusions BLIMP1 gene silencing may inhibit the proliferation of NCI-H929 cells through suppressing unfolded protein response （UPR） by down-regulation of XBP1 expression.