敲低同源盒A6（HOXA6）抑制HCT116细胞增殖、迁移和侵袭

Knockdown of homeobox A6 （HOXA6） inhibits proliferation，migration and invasion of HCT116 cells

摘要:

目的：探讨抑制同源盒A6（homeobox A6，HOXA6）的表达对HCT116结直肠癌细胞增殖、迁移和侵袭能力的影响。方法：构建重组干扰质粒（shRNA1，2，3，4-HOXA6）和阴性对照质粒（shRNA-NC），脂质体转染，荧光显微镜、RT-PCR和Western blot检测转染效率和表达情况。CCK-8法和克隆形成检测增殖，Transwell检测迁移和侵袭。Western blot检测E-cadherin、N-cadherin、Vimentin蛋白表达。结果：荧光显微镜显示各组转染效率均为30%左右；RT-PCR和Western blot结果显示各干扰组HOXA6 mRNA（P=0.003）和蛋白（P=0.000）的表达均低于shRNA-NC组，其中shRNA1-HOXA6组最低（P=0.000）。CCK-8结果显示shRNA1-HOXA6组的吸光度值明显低于shRNA-NC组（P=0.005）；克隆形成结果显示shRNA1-HOXA6组（132±45）克隆形成数较shRNA-NC组（353±45）明显减少（P=0.004）；Transwell迁移和侵袭实验显示shRNA1-HOXA6组[（134±28）、（60±4）]穿出细胞数较shRNA-NC组[（276±94）、（168±50）]明显减少（P=0.025、P=0.008）。Western blot实验显示shRNA1-HOXA6组E-cadherin（0.490±0.025）的表达较shRNA-NC组（0.326±0.032）明显升高（P=0.003），而N-cadherin（0.112±0.016）和Vimentin（0.886±0.033）较shRNA-NC组[（0.147±0.013）、（1.159±0.040）]明显降低（P=0.037、P=0.001）。结论：敲低HOXA6表达能显著抑制结直肠癌HCT116细胞的增殖、迁移和侵袭能力。

Abstract:

Objective：To investigate the effect of suppressed expression of homeobox A6（HOXA6） on the proliferation，migration and invasion of HCT116 colorectal cancer cells. Methods：Recombinant plasmids were constructed and transfected via Lipofectamine 2000. Transfection and interference efficiency was detected by fluorescence microscope，RT-PCR and Western blot. Moreover，cell proliferation，migration and invasion were measured by CCK-8，colony formation and Transwell assays. The protein expression of E-cadherin，N-cadherin and Vimentin was identified by Western blot. Results：The transfection efficiency was nearly 30%. The expres－sion of HOXA6 mRNA（P=0.003） and protein（P=0.000） was significantly lower in all interference groups than in shRNA-NC group，and the most interference efficient plasmid was shRNA1-HOXA6（P=0.000）. CCK-8 showed that the A450 values in shRNA1-HOXA6 group were lower than in shRNA-NC group（P=0.005）. Colony formation revealed that the colonies in shRNA1-HOXA6 group （132±45） were considerably fewer than in shRNA-NC group（353±45）（P=0.004）. Besides，Transwell assay showed that the num－ber of migration and invasion cells was significantly fewer in shRNA1-HOXA6 group[（134±28），（60±4）] than in shRNA-NC group [（276±94），（168±50）] （P=0.025，P=0.008）. Moreover，the protein expression of E-cadherin was higher in shRNA1-HOXA6 group （0.490±0.025） than in shRNA-NC group（0.326±0.032）（P=0.003），but N-cadherin（0.112±0.016） and Vimentin（0.886±0.033） expression was lower in shRNA1-HOXA6 group than in shRNA-NC group[（0.147±0.013），（1.159±0.040）]（P=0.037，P=0.001）. Conclusion：Knockdown of HOXA6 expression can remarkably suppress the proliferation，migration and invasion of HCT116 colorectal cancer cells.