Objective: To investigate the effects of siRNAs targeting CD97 immune epitopes on proliferation, infiltration, apoptosis and cell cycle of breast cancer cells.
Methods: siRNA sequences targeting CD97EGF and CD97Stalk immune epitopes were designed according to Gene Bank NM_001025160.2 with smart siCatchTM siRNA design software. CD97siRNAs were transfected into MDA-MB231 cells in which CD97 was highly expressed. Highest sensitive CD97EGF and CD97Stalk siRNA were screened by Western blotting. Inverted microscope was used to observe the growth of CD97siRNAs-transfected MDA-MB231 cells; the proliferation activity of MDA-MB231 cells was detected by MTT method; the wound healing assay and Transwell migration test were performed to examine the migration and infiltration ability of CD97EGF and CD97Stalk siRNA-transfected MDA-MB231 cells; the effects of CD97EGF siRNA and CD97Stalk siRNA on cell apoptosis and cell cycle of MDA-MB231 cells were detected by TUNEL and flow cytometry.
Results: The growth and proliferation activity of CD97siRNAs-transfected MDA-MB231 cells were significantly lower than those in the control groups, and such differences were more significant in CD97Stalk siRNA-transfected group (all P<0.05); scratch test showed that the wound healing rate was lower in CD97siRNAs-transfected groups, especially in CD97Stalk siRNA-transfected group (all P<0.05); Transwell migration showed that the number of MDA-MB231 cells crossing through chambers were less in CD97siRNAs-transfected groups, especially in CD97Stalk siRNA-transfected group (all P<0.05); no significant difference in cell apoptosis was observed between CD97siRNAs-transfected groups and control groups; cell cycle detection showed that CD97siRNAs-transfected groups had less cells in G0/G1 phase and more cells in S phase compared with the control groups, and such effect on cell cycle was more marked in CD97Stalk siRNA-transfected group (all P<0.05).
Conclusions: CD97 plays an important role in the cell growth, proliferation, migration and invasion of breast cancer MDA-MB231 cells, and compared with CD97EGF, CD97Stalk may have more effective inhibitory effects on cellular malignant behaviors.
目的: 明确CD97免疫表位CD97 EGF和CD97 Stalk对乳腺癌细胞生物学行为的影响。
方法: 采用siCatch TM siRNA design软件设计CD97 EGF小干扰RNA（siRNA）和CD97 Stalk siRNA并转染CD97高表达乳腺癌细胞株MDA-MB231；蛋白质印迹法筛选高敏感CD97 EGFsiRNA和CD97 Stalk siRNA。倒置显微镜观察和计数siRNA转染后乳腺癌细胞的生长；四唑盐（MTT）法检测siRNA转染后乳腺癌细胞的增殖活性；划痕试验和Transwell试验观察siRNA转染对乳腺癌细胞迁移和侵袭力的影响；TUNEL法检测siRNA转染对乳腺癌细胞凋亡的影响；流式细胞术检测siRNA转染对乳腺癌细胞周期的影响。
结果: siRNA转染后，乳腺癌细胞的生长数量和增殖活性明显低于对照组，且CD97 Stalk siRNA组的变化较CD97 EGFsiRNA组更为显著（均 P < 0.05）；划痕试验结果显示，siRNA转染组乳腺癌细胞的移动速率明显慢于对照组（均 P < 0.01），且CD97 Stalk siRNA对乳腺癌细胞移动速率的影响明显大于CD97 EGFsiRNA（ P < 0.05）；Transwell试验结果显示，siRNA转染组迁移过膜的细胞数量明显少于对照组，且以CD97 Stalk siRNA组减少最为显著（均 P < 0.05）。siRNA转染对乳腺癌细胞凋亡率的影响不明显，但siRNA转染组中G 0/G 1期细胞所占比例较对照组明显减小，而S期细胞所占的比例明显增加，且CD97 Stalk siRNA对细胞周期的影响较CD97 EGFsiRNA更为显著（均 P < 0.05）。
结论: CD97蛋白分子可能参与乳腺癌细胞株的生长、增殖、迁移和侵袭等生物学行为，CD97 Stalk免疫表位对于乳腺癌细胞株生物学行为影响较CD97 EGF免疫表位更为显著。