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目的采用RNA干扰(RNAi)技术抑制前列腺癌PC3细胞系中Bloom解旋酶基因的表达,探讨Bloom解旋酶基因表达下调后对PC3细胞的抑制作用。方法使用实验室前期成功构建的两条针对于Bloom解旋酶基因的RNAi载体shRNA-1和shRNA-2转染前列腺癌PC3细胞,以未转染RNAi载体为对照组,分别在转染24、48、72h后通过MTT法检测细胞增殖情况,转染48h后通过Transwell小室法检验细胞侵袭、迁移能力,细胞划痕实验检测细胞迁移情况,Hoechst/PI双染法检测细胞凋亡情况。结果转染RNAi载体后,与未转染对照组相比,各实验时间点的转染组细胞增殖率降低(P<0.05),Transwell细胞侵袭和迁移实验中穿过室膜的细胞数与对照组比均减少(侵袭:119±24、118±30vs 227±38;迁移:122±13、121±47vs 277±32,P<0.05),划痕愈合率与划痕迁移距离均降低,48h时差异有统计学意义(P<0.05),而且细胞凋亡明显增加。结论以RNAi载体干扰Bloom解旋酶基因的表达可抑制前列腺癌PC3细胞的增殖、迁移和侵袭能力并促进其凋亡,为前列腺癌的靶向基因治疗提供了依据。
OBJECTIVE: To study the effect of RNA interference (RNAi) on the expression of implicase gene in PC3 cell line and to investigate the effect of down-regulation of the expression of implicase on PC3 cell line. Methods Two prostate cancer PC3 cells transfected with RNAi vector shRNA-1 and shRNA-2 targeting to the Bloom helicase gene were successfully constructed in the laboratory. The untransfected RNAi vector was used as the control group, Cell proliferation was detected by MTT assay 48 and 72 hours after transfection. Cell invasion and migration were assayed by Transwell chamber assay 48 hours after transfection. Migration was assessed by cell scratch assay. Hoechst / PI double staining was used to detect cell apoptosis. Results Compared with untransfected control group, the proliferation rate of transfected cells in each experimental time point decreased (P <0.05). The number of cells passing through the ventricular membrane in the process of Transwell invasion and migration was significantly lower than that of the control (Invasion: 119 ± 24,118 ± 30vs 227 ± 38; Migration: 122 ± 13,121 ± 47vs 277 ± 32, P <0.05). The healing rate and scratch migration distance of scratch scar decreased at 48h The difference was statistically significant (P <0.05), and the apoptosis was significantly increased. CONCLUSION: RNAi vector interferes with the expression of the Bloom helicase gene in prostate cancer PC3 cells can inhibit the proliferation, migration and invasion and promote apoptosis and provide the basis for targeted gene therapy of prostate cancer.