MMP-2基因对前列腺癌细胞恶性表型的影响及其机制研究

Effect of MMP-2 gene on malignant phenotype of prostate cancer cells and its mechanism

目的探讨基质金属蛋白酶2(MMP-2)基因对前列腺癌细胞恶性表型的影响及其机制。方法 Western blot和RT-PCR检测人正常前列腺上皮RWPE-1细胞和前列腺癌PC3细胞中MMP-2的表达情况;以脂质体Lipfectamine 2000将MMP-2 siRNA和MMP-2 control质粒分别转染至siRNA MMP-2组和siRNA control组细胞中,以不做处理的细胞为对照组。Western blot和RT-PCR检测其转染效果;CCK-8法检测细胞增殖;Transwell法检测细胞的侵袭和迁移;FCM检测细胞凋亡;Western blot检测细胞中血管内皮生长因子(VEGF)和Cleaved Caspase-8蛋白的表达。结果与RWPE-1细胞相比,PC3细胞中MMP-2蛋白和mRNA的表达水平显著升高(P<0.05)。MMP-2在siRNA MMP-2组细胞中的蛋白和mRNA表达显著低于对照组(P<0.05),而siRNA control组与对照组间MMP-2的蛋白和mRNA表达水平差异无显著性(P>0.05)。siRNA MMP-2组细胞的OD值、侵袭细胞数、迁移细胞数和VEGF蛋白的表达水平显著下降,而细胞凋亡率和Cleaved Caspase-8蛋白水平显著升高,差异均显著性(P<0.05);对照组和siRNA control组间OD值、侵袭细胞数、迁移细胞数、细胞凋亡率、VEGF和Cleaved Caspase-8蛋白表达均未见显著差异(P>0.05)。结论 MMP-2在前列腺癌PC3细胞中高表达,干扰其表达可抑制细胞的增殖、侵袭和迁移,并促进细胞凋亡,其作用机制可能与下调VEGF蛋白和上调Cleaved Caspase-8蛋白的表达有关。

Objective To investigate the effect of matrix metalloproteinase-2（ MMP-2） gene on the malignant phenotype of prostate cancer cells and its mechanism. Methods The expression of MMP-2 in human prostate epithelial cells RWPE-1 and prostate cancer PC3 cells were detected by Western blot and RT-PCR. The MMP-2 siRNA and MMP-2 control plasmids were transfected into siRNA MMP-2 group and siRNA control group by liposome Lipfectamine 2000,and the cells without treatment were served as control group. Transfection effects were detected by Western blot and RT-PCR. Cell proliferation was detected by CCK-8 assay. Cell invasion and migration were detected by Transwell assay and apoptosis was detected by FCM. The expression of VEGF and Cleaved Caspase-8 protein in cells was detected by Western blot. Results Compared with RWPE-1 cells,the expression levels of MMP-2 protein and mRNA in PC3 cells were significantly increased（ P 〈 0. 05）.The protein and mRNA expression of MMP-2 in siRNA MMP-2 group was significantly lower than that in control group（ P 〈 0. 05）. There was no significant difference in the expression of MMP-2 and mRNA between siRNA control group and control group（ P 〈 0. 05）. The OD value of the cells,the number of invasion and migrated cells,and the expression of VEGF protein were significantly decreased,but the rate of apoptosis and Cleaved Caspase-8 protein expression levels were significantly increased in siRNA MMP-2 group,and the differences were statistically significant（ P 〈 0. 05）. OD value,the number of invasion and migrated cells,apoptosis rate,VEGF and Cleaved Caspase-8 protein expression levels had no significant difference between control group and siRNA control group（ P 〈 0. 05）. Conclusion MMP-2 is highly expressed in prostate cancer PC3 cells,interfering with its expression can inhibit cell proliferation,invasion and migration,and promote apoptosis,and its mechanism may be related to down regulation of VEGF protein and up regulation of Cleaved Caspase-8 protein expression.