抗SMO多克隆抗体对DU145前列腺癌细胞生长及侵袭力的影响

目的 探讨抗SMO多克隆抗体对DU145前列腺癌细胞生长及侵袭能力的影响。方法 预测SMO蛋白抗原表位并制备SMO抗体,评估其效价和特异性；实验分组包括空白对照组、兔IgG抗体处理组、抗SMO抗体处理组、shRNA空载对照组、TRPC-6蛋白shRNA稳转细胞处理组以及合并正常IgG抗体和抗SMO抗体处理组。MTT法和细胞凋亡实验检测抗SMO抗体对DU145细胞增殖和细胞凋亡的影响；Transwell实验检测抗SMO抗体对正常以及稳转TRPC-6shRNA蛋白的DU145细胞侵袭能力的影响。结果 制备的抗体效价为1∶51200,且抗体特异性良好；随着抗SMO抗体浓度的升高,细胞存活百分比逐渐下降(P<0.05),随着时间的发展,48h和72h之间的细胞存活百分比存在差异(P<0.05)。凋亡实验显示,早期凋亡细胞(Annexin V+/PI-)凋亡百分比在抗SMO抗体处理组较空白对照组中明显升高(P<0.05),随着抗体浓度的升高,凋亡百分比逐渐升高。Transwell实验显示,随着抗SMO抗体的处理,细胞的细胞侵袭抑制率逐渐升高(P<0.05),TRPC-6蛋白shRNA稳转DU145细胞处理组中,随着抗体浓度的升高,细胞的细胞侵袭抑制率逐渐升高(P<0.05)。结论 抗SMO多克隆抗体对正常以及TRPC-6蛋白shRNA稳转的DU145细胞生长以及侵袭能力均有明显的抑制作用,为临床治疗前列腺癌提供一个潜在的方向。
Objective To investigate the effects of SMO polyclonal antibody on proliferation and invasive ability of human prostate cancer DU145 cells. Methods SMO protein epitope was predicted and used for preparation of polyclonal antibody. Titer and specificity of antibody was evaluated. DU145 cells were divided into blank control group, normal rabbit IgG group, SMO antibody group, shRNA vector control group, TRPC-6 shRNA group and combined normal IgG and SMO antibody group. Cell proliferation and apoptosis was detected by MTT assay and flow cytometry, respectively; and cell invasive ability was examined by Transwell assay. Results Titer of antibody in this study was 1∶51200. The antibody was verified to be specific. MTT assay indicated that the proliferation of DU145 cells was inhibited with the increasing of SMO antibody concentrations (P<0.05). Flow cytometry showed that the ratio of apoptotic cells increased in SMO antibody group compared to normal IgG group (P<0.05); Transwell assay showed that invasive ability was decreased by SMO antibody. In TRPC-6shRNA stable transfected DU145 cells, invasive inhibition rate was increased (P<0.05). Conclusion SMO polyclonal antibody can inhibit proliferation and enhance apoptosis of prostate cancer DU145 cells, and it may have a synergistic effect with TRPC-6shRNA