STIM1基因在人下咽癌细胞系FaDu中的表达及对细胞凋亡的影响

Expression of STIM1 gene in human hypopharyngeal carcinoma cell line FaDu and its effect on FaDu cell apoptosis

目的探索STIM1基因在人下咽癌细胞系Fa Du中的表达情况及其表达沉默后对细胞凋亡的影响。方法培养人下咽癌Fa Du细胞,根据不同慢病毒感染分2组。STIM1-siRNA组为经STIM1-siRNA慢病毒感染的下咽癌细胞;对照组为经阴性对照慢病毒感染的下咽癌细胞。Real-Time PCR法检测STIM1在人下咽癌Fa Du细胞中的表达以及siRNA慢病毒感染后STIM1mRNA水平的表达;Western blot检测siRNA慢病毒感染后STIM1在蛋白水平的表达;流式细胞仪检测STIM1基因沉默后的人下咽癌Fa Du细胞的凋亡情况。采用SPSS 17.0软件对数据进行统计学分析。结果以GAPDH为内参(Ct=12.08±0.05),STIM1在下咽癌细胞系Fa Du中表达显著(Ct=22.21±0.05,P<0.01);Real-time PCR结果显示对照组和STIM1-siRNA组人下咽癌Fa Du细胞的表达值分别为(1.00±0.08)和(0.12±0.01),2组差异具有统计学意义(P<0.01);Western blot的结果显示,STIM1-siRNA组Fa Du细胞中STIM1蛋白明显受抑制,与Real-time PCR结果一致,慢病毒感染成功;流式细胞仪检测结果显示对照组凋亡率为(4.36±1.32)%;实验组凋亡率为(9.81±0.56)%,差异具有统计学意义(P<0.05)。结论 STIM1基因与人下咽癌Fa Du细胞凋亡显著相关,人下咽癌Fa Du细胞中,STIM1可能抑制凋亡,可能成为下咽癌诊治的全新切入点。

Objective To study the expression of STIM1 gene in human hypopharyngeal carcinoma cell line Fa Du and its effect on Fa Du cell apoptosis. Methods Lentivirus infection was used to knock STIM1 down in Fa Du cells. Group STIM1-siRNA： the expression of STIM1 in Fa Du cell was inhibited by STIM1-siRNA lentivirus. Group control： Fa Du cells were infected by negative control siRNA lentivirus.Real-Time PCR was applied to identify the efficacy of lenticirus infection and the expression of STIM1 in Fa Du cells. Western blot was used to identify the expression of STIM1 protein after lenticirus infection. Flow cytometry assay was performed to detect the apoptosis of Fa Du cells in the two groups. The data were statistically analyzed with SPSS 17. 0 software. Results Compared with GAPDH（ Ct = 12. 08 ± 0. 05）,the expression of STIM1 in Fa Du cells was significant expressed（ Ct = 22. 21 ± 0. 05,P〈0. 001）. Real-Time PCR analysis the relative mRNA expression of STIM1 in Fa Du cells of control group and STIM1-siRNA group were（ 1. 00 ± 0. 08） and（ 0. 12 ± 0. 01） respectively（ P〈0. 001）.Western blot showed that the expression of STIM1 gene and protein in Fa Du cells were inhibited significantly after STIM1-siRNA lentiviral infection,which was in accordance with the results of Real-Time PCR analysis. Flow cytometry assay showed that the siRNA-mRNA group had a higher apoptosis percentage（ 9. 81 ± 0. 56） % compared to the control group（ 4. 36 ± 1. 32） %,with statistically significant difference（ P〈0. 05）. Conclusion STIM1 gene correlated significantly with Fa Du cell apoptosis. It inhibits apoptosis of Fa Du cells,and it may be a potential diagnostic and therapeutic target for the hypopharyngeal carcinoma.