钙释放激活钙通道调节分子1促进结肠癌细胞系SW480的增殖

Calcium release-activated calcium channel modulator 1 promotes the proliferation of SW480 colon cancer cell line

目的探讨钙释放激活钙通道调节分子1(ORAI1)对结肠癌细胞系SW480增殖的影响及其机制。方法以ORAI1干扰慢病毒感染SW480细胞,分别用实时荧光定量PCR和Western blot检测细胞中ORAI1mRNA和蛋白的表达,通过MTT法、集落形成实验和倍增时间检测细胞的增殖,流式细胞仪检测其周期,激光共聚焦显微镜检测细胞钙内流(SOCE),Western blot法检测细胞中ERK1/2、p-ERK1/2和Cyclin D1蛋白表达。结果 SW480转染ORAI1干扰慢病毒72 h后,可见明显的荧光表达;干扰组较空病毒组和对照组,ORAI1的表达降低(P<0.01)、细胞生长减慢(P<0.05)、集落形成能力减弱(P<0.01)、倍增时间延长(P<0.01)、G1期细胞比例增高(P<0.05)、SOCE内流峰值降低(P<0.05)、p-ERK1/2和Cyclin D1的蛋白表达量降低(P<0.01)。结论 ORAI1可能通过SOCE调节胞内Ca2+浓度,并进一步通过MAPK信号通路促进SW480细胞的增殖。

Objective To explore the effect of calcium release-activated calcium channel modulator 1 （ ORAI1 ） on the proliferation of colon cancer cell line SW480 and its mechanism. Methods SW480 cells were infected with ORAI1 interference lentivirus. The expressions of ORAI1 mRNA and protein were confirmed by fluorescence quantitative PCR（FQ-PCR） and Western blot MTT, colony-forming test and doubling time were used to examined the proliferation of SW480. Flow cytometry was used to detect the periodic variation of SW480. Confocal microscopy was used to detect the difference of SOCE （store-operated Ca^2＋ entry, SOCE） in each group. Meanwhile,Western blot was used to detect the expressions of ERK1/2, p-ERK1/2 and CyclinD1 protein. Results After infecting SW480 with ORAI1 interference lentivirns for 72 h, significant fluorescence expression can be seen. Compared with the empty vector group and the control group, the expression of ORAI1 was lower （P 〈 0. 01 ）, the cells grew slower（P 〈 0. 05 ）, the colony-forming ability was decreased （P 〈0. 01 ）, the doubling time was delay（P 〈0. 01 ） ,the proportion of cells in G1 phase was higher（ P 〈0.05 ）, the Internal flow peak of SOCE was lower（ P 〈0.05 ）, and the expressions of p-ERK1/2 and CyclinD1 proteins deseased（P 〈0. 01 ）. Conclusions ORAI1 may regulate the intracellular calcium concentration by SOCE, and promote the proliferation of SW480 through MAPK signaling pathways.