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目的:探讨靶向抑制ADAM17基因对雄激素非信赖性前列腺癌PC-3细胞增殖的影响。方法:应用ADAM17 siRNA转染PC-3细胞后,通过RT-PCR、Western印迹方法分别检测ADAM17 mRNA和蛋白表达变化;MTT、BrdU掺入法检测下调ADAM17对PC-3细胞的增殖和DNA合成能力的影响;流式细胞术检测ADAM17 siR-NA对PC-3细胞细胞周期的影响;Western印迹检测下调ADAM17对PC-3细胞增殖相关基因表达的影响。结果:两对ADAM17 siRNAs均可有效地降低PC-3细胞ADAM17 mRNA和蛋白的表达;MTT结果显示与对照组(0.80±0.51)相比,两对ADAM17 siRNAs均可显著抑制细胞的生长(0.43±0.57、0.44±0.64,P均<0.05);Br-dU掺入实验显示与对照组(0.79±0.72)相比,ADAM17 siRNAs均能显著下调DNA的合成能力(0.48±0.43、0.54±0.59,P<0.05);流式细胞术结果显示,与对照组(41.38±1.53)%相比,ADAM17 siRNAs可显著增加G1期细胞数量[(61.83±2.41)%、(59.78±1.92)%,P均<0.05]、降低S期细胞数量[从(33.51±1.47)%减少到(23.64±2.56)%、(25.24±1.86)%,P均<0.05],同时伴随着cyclin D1蛋白的表达下降而p21蛋白的表达升高。结论:ADAM17 siRNA可以通过下调cyclin D1、上调p21的表达而抑制前列腺癌PC-3细胞增殖,ADAM17可能成为前列腺癌基因治疗的靶点。
Objective: To investigate the effect of targeted inhibition of ADAM17 gene on the proliferation of androgen-independent prostate cancer PC-3 cells. Methods: ADAM17 siRNA was transfected into PC-3 cells. The mRNA and protein expression of ADAM17 were detected by RT-PCR and Western blotting respectively. MTT and BrdU incorporation assay were used to detect the proliferation and DNA synthesis ability of ADAM17 on PC-3 cells The effect of ADAM17 siR-NA on the cell cycle of PC-3 cells was detected by flow cytometry. The effect of downregulation of ADAM17 on the expression of proliferation-related genes in PC-3 cells was detected by Western blotting. Results: Two pairs of ADAM17 siRNAs could effectively decrease the expression of ADAM17 mRNA and protein in PC-3 cells. MTT assay showed that both ADAM17 siRNAs significantly inhibited the cell growth (0.43 ± 0.51) compared with the control group (0.80 ± 0.51) 0.57,0.44 ± 0.64, P <0.05). The Br-dU incorporation assay showed that compared with the control group (0.79 ± 0.72), ADAM17 siRNAs significantly decreased the DNA synthesis ability (0.48 ± 0.43,0.54 ± 0.59, P (P <0.05). The results of flow cytometry showed that ADAM17 siRNAs significantly increased the number of cells in G1 phase [(61.83 ± 2.41)% and (59.78 ± 1.92)%, respectively, compared with the control group (41.38 ± 1.53% 0.05]. The number of cells in S phase decreased from (33.51 ± 1.47)% to (23.64 ± 2.56)%, (25.24 ± 1.86)%, P <0.05, respectively) Increased expression. Conclusion: ADAM17 siRNA can inhibit the proliferation of PC-3 cells by down-regulating the expression of cyclin D1 and up-regulating the expression of p21. ADAM17 may be the target of prostate cancer gene therapy.