Cx43对乳腺癌细胞侵袭和迁移及细胞间隙通讯的影响

Effect of Cx43 on the invasion and migration and Gap jjnctional intercellular communication in breast cancer cell

目的:探讨Cx43对乳腺癌侵袭、迁移以及细胞间隙通讯功能的影响。方法:用脂质体转染法将Cx43质粒、Cx43 siRNA及阴性对照转染入人乳腺癌细胞株MDA-MB-231。细胞黏附实验、Transwell试验检测细胞黏附、侵袭和迁移能力。分子生物学方法检测MMP-2、MMP-9、BRMS-1的表达。荧光染料传输实验检测不同Cx43表达的细胞间隙通讯的功能。结果:细胞黏附实验结果表明相较于野生型细胞,Cx43 siRNA组黏附能力(0.43±0.07)降低(t=20.801,P=0.002),Cx43质粒组黏附能力(1.63±0.26)增高(t=5.917,P=0.027)。Transwell实验表明相较野生组,Cx43质粒组侵出小室的细胞数增加(侵袭:1.25±0.04,t=16.339,P=0.004;迁移:1.33±0.02,t=22.770,P=0.002),而Cx43 siRNA组侵出小室的细胞数减少(迁移:0.84±0.04,t=11.139,P=0.008;侵袭:0.79±0.04,t=5.743,P=0.029)。分子生物学检测发现Cx43质粒组MMP-2、MMP-9表达增高,BRMS-1表达降低,而Cx43 siRNA组MMP-2、MMP-9表达降低,BRMS-1表达增高。染料传输实验证实相较野生组传输能力,Cx43质粒组传输作用增强;Cx43 siRNA组细胞间的传输作用减弱。结论:Cx43对乳腺癌细胞侵袭及迁移能力存在正调控。这一调控能力可能通过GJIC实现。

To explore the effects of Cx43 on the invasion,migration and gap junctional intercellular communication（GJIC）of breast cancer. Methods：The Cx43 plasmid,inhibitors and negative controls were transfected into human breast cancer cell line MDA - MB - 231 by liposome. The motility of cells was evaluated by adhesion and Transwell assay. MMP - 2,MMP - 9 and BRMS - 1 were detected by both qPCR and Western blot. A scrape - loading dye transfer assay was used to evaluate the function of GJIC. Results：Compared to the wild type,adhesion ratio of Cx43 siRNA group（0. 43 ± 0. 07）was significantly lower（t = 20. 801,P = 0. 002）,and that of Cx43 plasmid group （1. 63 ± 0. 26）was higher（t = 5. 917,P = 0. 027）by adhesion assay. Transwell tests showed the migration and inva-sion of Cx43 plasmid group was significantly increased（ invasion：1. 25 ± 0. 04,t = 16. 339,P = 0. 004;migration：1. 33 ± 0. 02,t = 22. 770,P = 0. 002）,but that of Cx43 siRNA group was significantly reduced（ migration：0. 84 ± 0. 04,t = 11. 139,P = 0. 008;invasion：0. 79 ± 0. 04,t = 5. 743,P = 0. 029）. The expression of MMP - 2 and MMP - 9 were enhanced and the expression of BRMS - 1 was reduced in Cx43 plasmid group detected by both qPCR and west-ern blot and vice versa in Cx43 siRNA group. Compared to the wild type,GJIC was enhanced in Cx43 plasmid group and reduced in Cx43 siRNA group by a scrape - loading dye transfer assay. Conclusion：Cx43 might have a positive control on invasion and migration of breast cancer cell by GJIC.