干扰FSCN1基因表达对前列腺癌细胞凋亡、活性氧水平影响的研究

目的 探讨干扰FSCN1基因表达对前列腺癌细胞凋亡、活性氧(ROS)水平影响及机制.方法 以正常前列腺上皮细胞RWPE-1为对照细胞,通过RT-PCR及Western blot检测前列腺癌LNCaP、DU145和PC-3细胞中FSCN1 mRNA及蛋白表达;以LipofectamineTM 2000为载体,DU145细...

Effect of interfering FSCN1 gene expression on apoptosis and ROS content in prostate cancer cells

Objective To investigate the effect and mechanism of interfering FSCN1 gene expression on apoptosis and reactive oxygen species(ROS)level in prostate cancer cells.Methods The normal prostate epithelial cell RWPE-1 was used as the control,and the expression of FSCN1 mRNA and protein in the LNCaP,DU145 and PC-3 cells of prostate cancer were detected by RT-PCR and Western blot,respectively.LipofectamineTM 2000 was used as the carrier,DU145 cells were divided into si-FSCN1 group(small interfering RNAs targeting FSCN1 transfected DU145 cells),negative control group(negative control siRNA transfected DU145 cells)and blank control group(untransfected cells),siRNA was transfected for 48 h,and Western blot was used to detect the protein expression of FSCN1,PCNA,Cleaved caspase3,NF-κB p65,p-NF-κB p65,IKK and p-IKKα.Cell viability was detected by CCK8,and apoptosis rate and ROS level were detected by flow cytometry.One-way analysis of variance was used for comparison between groups,and SNK-q test was used for pairwise comparisons between groups.Results Compared with RWPE-1 cells,the expressions of FSCN1 mRNA in LNCaP,DU145 and PC-3cells(1 vs 2.561±0.189,7.183±0.882,4.796±0.567,4.796±0.567)and protein(0.053±0.007 vs 0.217±0.013,0.654±0.058,0.316±0.035)were increased(all P<0.05).Compared with the negative control group,the expression of FSCN1 in the si-FSCN1 group were decreased(0.473±0.052 vs 0.086±0.010,P<0.05).Compared with the si-FSCN1 group,the cell viability of the blank control group and the negative control group(0.302±0.033 vs 0.787±0.069,0.764±0.063)were increased,while the apoptotic rate(24.54±1.47﹪vs 3.04﹪±0.36﹪,3.28﹪±0.40﹪)and ROS relative fluorescence intensity(90.04±5.73 vs 47.88±3.62,49.62±4.11)were decreased(all P<0.05).Compared with si-FSCN1 group,the expression of PCNA(0.255±0.032 vs 0.654±0.062,0.631±0.058),NF-κB p65(0.092±0.011 vs 0.296±0.032,0.318±0.037),p-NF-κB p65(0.042±0.008 vs 0.155±0.018,0.151±0.016),IKKα(0.112±0.01 vs 0.172±0.020,0.192±0.023)and p-IKKαprotein(0.051±0.005 vs 0.102±0.011,0.091±0.009)were increased in blank control group and negative control group,while the expression of Cleaved caspase3 protein(0.206±0.018 vs 0.074±0.009,0.085±0.010)decreased(all P<0.05).There was no significant difference in cell viability,apoptosis rate,ROS levels,and protein expression of FSCN1,PCNA,Cleaved caspase3,NF-κB p65,p-NF-κB p65,IKKα,and p-IKKαin the negative control group and the blank control group(P>0.05).Conclusion Interfering with FSCN1 gene expression could reduce the activity and induce apoptosis of prostate cancer cells.The mechanism may be related to the increase of ROS level and the decrease of NF-κB signal.