학술논문
STIM1对乳腺癌细胞存活与增殖的影响及初步机制分析 / Effect of STIM1 on survival and proliferation of breast cancer cells and its pre-liminary mechanism analysis
Document Type
Academic Journal
Author
Source
中国免疫学杂志 / Chinese Journal of Immunology. 34(5):681-687
Subject
Language
Chinese
ISSN
1000-484X
Abstract
目的:探讨STIM1对乳腺癌细胞存活与增殖的影响及初步机制分析.方法:以正常乳腺上皮细胞MCF-10A作为对照,Western blot检测乳腺癌 MCF7、HCC1569、MDA-MB-231、BT549细胞中STIM1的蛋白表达;将STIM1的靶向siRNA序列(STIM1-siRNA组)转染MDA-MB-231细胞,并设置阴性对照组和空白对照组,转染48 h后检测各组细胞STIM1的蛋白表达;MTT法检测转染24、48和72 h的细胞活力;流式细胞术检测转染48 h的细胞凋亡率;RT-PCR检测IL-6和 TNF-α 的mRNA表达;Western blot检测增殖相关蛋白细胞增殖核抗原(PCNA)、凋亡蛋白B细胞淋巴瘤/白血病-2(Bcl-2)、Bcl-2相关X蛋白(Bax)、含半胱氨酸的天冬氨酸蛋白水解酶3(Caspase3)及STAT3和磷酸化的信号转导与转录因子3(p-STAT3)的蛋白表达.结果:乳腺癌细胞中STIM1的蛋白表达均显著高于在MCF-10A细胞表达(P<0.05);转染STIM1的siRNA后MDA-MB-231细胞中STIM1的蛋白表达显著低于空白对照组(P<0.05);与空白对照组比较,STIM1-siRNA组细胞在48 h和72 h的细胞活力显著降低,在48 h的细胞凋亡率显著升高,IL-6和TNF-α的mRNA表达显著降低,PCNA、Bcl-2和p-STAT3的蛋白表达显著降低,Bax和Caspase3蛋白表达显著升高.结论:STIM1基因在乳腺癌细胞高表达,通过RNA干扰抑制其表达可降低癌细胞活力,诱导细胞凋亡、提高免疫及下调STAT3信号.
Objective:To investigate the effect of STIM1 on the survival and proliferation of breast cancer cells and its preliminary mechanism analysis.Methods:Normal mammary epithelial cells MCF-10A as control,the expression of STIM1 in MCF7, HCC1569,MDA-MB-231 and BT549 breast cancer cells were detected by Western blot;STIM1 siRNA sequence(STIM1-siRNA group) were transfected into MDA-MB-231 cells,and the negative control group and the blank control group were set up,the protein expression of STIM1 in each group were detected after cells were transfected for 48 h;MTT method was used to detect cell activity cells were transfected for 24 h,48 h and 72 h;cell apoptosis was detected after cells were transfected for 48 h by flow cytometry;the mRNA expression of IL-6 and TNF-α were detected by RT-PCR;the expression of PCNA,Bcl-2,Bax,Caspase3,STAT3 and p-STAT3 protein were detected by Western blot.Results:The expression of STIM1 protein in breast cancer cells was significantly higher than that in MCF-10A cells (P<0.05);the expression of STIM1 protein in MDA-MB-231 cells transfected STIM1-siRNA was significantly lower than the control group(P<0.05);compared with the control group,cell viability in STIM1-siRNA group decreased significantly in cells were transfected for 48 h and 72 h,the apoptosis rate in 48 h was significantly increased,the expression of IL-6 and TNF-α mRNA sig-nificantly decreased,the expression of PCNA,Bcl-2 and p-STAT3 protein were significantly decreased,the expression of Bax and Caspase3 protein increased significantly.Conclusion:STIM1 gene is highly expressed in breast cancer cells.Inhibition of STIM1 expression by RNA interference can down regulate the activity of cancer cells,induce apoptosis,enhance immunity and by down regulation STAT3 signal.
Objective:To investigate the effect of STIM1 on the survival and proliferation of breast cancer cells and its preliminary mechanism analysis.Methods:Normal mammary epithelial cells MCF-10A as control,the expression of STIM1 in MCF7, HCC1569,MDA-MB-231 and BT549 breast cancer cells were detected by Western blot;STIM1 siRNA sequence(STIM1-siRNA group) were transfected into MDA-MB-231 cells,and the negative control group and the blank control group were set up,the protein expression of STIM1 in each group were detected after cells were transfected for 48 h;MTT method was used to detect cell activity cells were transfected for 24 h,48 h and 72 h;cell apoptosis was detected after cells were transfected for 48 h by flow cytometry;the mRNA expression of IL-6 and TNF-α were detected by RT-PCR;the expression of PCNA,Bcl-2,Bax,Caspase3,STAT3 and p-STAT3 protein were detected by Western blot.Results:The expression of STIM1 protein in breast cancer cells was significantly higher than that in MCF-10A cells (P<0.05);the expression of STIM1 protein in MDA-MB-231 cells transfected STIM1-siRNA was significantly lower than the control group(P<0.05);compared with the control group,cell viability in STIM1-siRNA group decreased significantly in cells were transfected for 48 h and 72 h,the apoptosis rate in 48 h was significantly increased,the expression of IL-6 and TNF-α mRNA sig-nificantly decreased,the expression of PCNA,Bcl-2 and p-STAT3 protein were significantly decreased,the expression of Bax and Caspase3 protein increased significantly.Conclusion:STIM1 gene is highly expressed in breast cancer cells.Inhibition of STIM1 expression by RNA interference can down regulate the activity of cancer cells,induce apoptosis,enhance immunity and by down regulation STAT3 signal.