epoxomicin可诱发前列腺癌细胞内质网应激蛋白GADD153促进细胞凋亡  

Epoxomicin induces endoplasmic reticulum stress protein GADD153 / CHOP to promote cell apoptosis in prostate cancer DU145 cells

作　　者：王利平[1,2] 罗荣城[1] 符鹏程[2] 黄朝刚[2] 

机构地区：[1]南方医科大学中西医结合医院肿瘤中心,广州510315 [2]郴州市第一人民医院肿瘤科,湖南郴州423000

出　　处：《第三军医大学学报》2014年第23期2367-2371,共5页Journal of Third Military Medical University

基　　金：郴州市第一人民医院青年基金(N2013-009)~~

摘　　要：目的探讨蛋白酶体抑制剂(epoxomicin,EPO)诱导前列腺癌DU145细胞凋亡及其与GADD153/CHOP的关系。方法不同浓度的EPO处理DU145不同时间,MTS检测细胞生长情况;流式细胞仪检测细胞凋亡率情况;Real-time PCR检测内质网应激的相关分子GADD153、GRP78 mRNA以及Western blot方法检测GADD153、GRP78蛋白表达情况;转染GADD153siRNA从而沉默基因GADD153,通过基因转录水平及蛋白水平检测GADD153证实转染是否成功;转染GADD153后,通过MTS法及流式细胞仪技术检测细胞增殖情况及凋亡情况。结果 EPO抑制DU145细胞生长,并呈现剂量、时间依赖。处理组细胞凋亡率高于未处理组,高剂量组凋亡率高于低剂量组。EPO处理后GADD153,GRP78基因转录水平随时间逐渐升高,72 h最为明显。GADD153、GRP78蛋白在EPO处理后不断增加,均在72 h增加到最高,且低剂量与高剂量组在72 h差异有统计学意义(P<0.05)。转染GADD153siRNA后,GADD153基因转录水平及蛋白表达水平明显降低,其中GADD153siRNA 50 nmol/L最为明显。沉默基因GADD153后,EPO处理DU145细胞,其细胞数目的减少部分被阻断,细胞凋亡部分被阻断。结论蛋白酶体抑制剂epoxomicin能抑制DU145细胞生长,诱导凋亡,其机制可能与激活内质网应激,诱发内质网的相关分子GADD153表达有关。Objective To investigate the proteasome inhibitor,epoxomicin（ EPO）,inducing the cell apoptosis and its relation with endoplasmic reticulum stress protein GADD153 / CHOP. Methods After DU145 cells were treated with EPO at different doses and / or for different times,the cell proliferation was detected by using MTS assay. Cell apoptosis was examined by flow cytometry. The mRNA expression of endoplasmic reticulum stress-related genes,GADD153 and GRP78 was tested by real-time PCR,while the protein levels were detected by Western blotting. GADD153 siRNA was transfected to silence the gene,which was verified by real-time PCR and Western blotting. Cell proliferation and apoptosis were tested again after the transfection with MTS assay and flow cytometry. Results Cell proliferation was inhibited after EPO treatment in a time- and dose-dependent fashion. The apoptotic rate was also in a dose-dependent manner. With the elapse of time,the mRNA levels of GADD153 and GRP78 were increased obviously when DU145 cells were treated with EPO,and reached the highest in 72 h after treatment. The same tendency was observed in the protein levels of GADD153 and GRP78,and the highest levels were also seen in 72 h（ P 〈 0. 05）. The mRNA and protein expression of GADD153 was decreased obviously after the transfection with GADD153 siRNA,with the dose of 50 nmol / L most significant. The inhibitory effects of EPO on the proliferation and apoptosis in DU145 cells were partly blocked by silencing the gene GADD153. Conclusion Proteasome inhibitor EPO inhibits the cell proliferation and apoptosis in DU145 cells,which might be due to activate the endoplasmic reticulum stress and up-regulate its associated molecule GADD153.

关 键 词：蛋白酶体抑制剂 GADD153/CHOP GRP78 内质网应激 

分 类 号：R737.25[医药卫生—肿瘤] R966[医药卫生—临床医学]