摘要
目的探讨siRNA沉默葡萄糖调节蛋白78(GRP78)增强顺铂对乳腺癌细胞凋亡的敏感性,以期为乳腺癌的临床治疗提供新的靶点。方法 MTT法检测不同浓度(0、1、2、4、8、16μmol/L)顺铂对乳腺癌细胞MDA-MB-231的增殖抑制作用;PI/AnnexinV双染检测顺铂(8μmol/L)对siRNA转染沉默GRP78乳腺癌细胞MDA-MB-231凋亡的影响,同时将未转染和转染空质粒乳腺癌细胞MDA-MB-231作为空白和阴性对照;Western blot检测顺铂(8μmol/L)处理乳腺癌细胞MDA-MB-231不同时间(0、6、16、24 h)GRP78的表达以及siRNA转染沉默GRP78乳腺癌细胞MDA-MB-231中GRP78的表达。结果 8μmol/L顺铂作用于乳腺癌细胞MDA-MB-231 24、48、72 h细胞存活率分别为83.13%、54.22%、35.79%,但24 h细胞凋亡率仅为10.8%。将GRP78沉默后,细胞的凋亡率为24.6%,沉默GRP78并合用顺铂进行刺激,细胞的凋亡率为48.9%,明显高于单用顺铂组。用顺铂刺激乳腺癌细胞MDA-MB-231 GRP78表达先上调后有减弱趋势,而siRNA转染沉默GRP78后,GRP78表达明显下降。结论抑制GRP78的表达可以增加乳腺癌细胞的凋亡率,为乳腺癌的临床治疗提供了新的靶点。
Objective To investigate the effect of small interfering RNA-mediated glucose-regulated protein 78 （GRP78） knockdown on the chemosensitivity of breast cancer cells to cisplatin. Methods Human breast cancer cell line MDA-MB-231 was exposed to different doses of cisplatin （0, 1, 2, 4, 8, and 16 pmol/L）, and the changes in cell viability were detected using MTT assay. PI/Annexin V staining was used to observe the apoptosis of the cells in response to transfection with a small interfering RNA targeting GRP78 （pSHISi-GRP78）. Western blotting was employed to detect GRP78 expression in pSHISi- GRP78-transfected cells after exposure to 8 pmol/L cisplatin for 24, 48 and 72 h. Results Exposure of the cells to 8 pmol/L cisplatin for 24, 48 and 72 h resulted in a cell survival rate of 83.13%, 54.22% and 35.79%, respectively, but the cell apoptosis rate was only 10.8% at 24 h. Transfection of MDA-MB-231 cells with pSHISi-GRP78 caused a cell apoptosis rate of 24.6%, which increased to 48.9% in cells with both pSH1Si-GRP78 transfection and cisplatin exposure. Cisplatin exposure caused an initial up-regulation followed then by a down-regulation of GRP78 expression in MDA-MB-231 cells, while pSHISi-GRP78 transfection produced an obvious down-regulation of GRP78 expression. Conclusions Inhibition of GRP78 expression increases the apoptosis and enhance cisplatin chemosensitivity of breast cancer cells in vitro, suggesting the value of GRP78 as a potential therapeutic target in the clinical treatment of breast cancer.
出处
《南方医科大学学报》
CAS
CSCD
北大核心
2013年第1期44-47,共4页
Journal of Southern Medical University
基金
国家自然科学基金(81000992
81072207)
安徽省教育厅自然科学研究重点项目(KJ2012A201)
安徽省自然科学基金(1208085MH136)~~
关键词
乳腺癌
葡萄糖调节蛋白78
小干扰RNA
顺铂
凋亡
breast cancer
glucose-regulated protein 78
small interfering RNA
cisplatin
apoptosis
