普鲁卡因调控CXCR7并影响AKT和STAT3信号通路从而抑制膀胱癌细胞活力、迁移和侵袭

Procaine regulates CXCR7 and affects AKT/STAT3 signaling pathway to inhibit viability,migration and invasion of bladder cancer cells

目的:探讨普鲁卡因(PCA)及CXC趋化因子受体7(CXCR7)对膀胱癌细胞活力、迁移和侵袭的影响及潜在的作用机制。方法:用不同浓度PCA处理人膀胱癌RT4细胞,并将细胞分为PBS组(无PCA处理)、PCA组(4 mmol/L PCA处理)、siRNA阴性对照(si-Con)组(转染si-Con)、CXCR7 siRNA(si-CXCR7)组(转染si-CXCR7)、PCA+pcDNA组(4 mmol/L PCA处理并转染pcDNA)和PCA+pcDNA-CXCR7组(4 mmol/L PCA处理并转染pcDNACXCR7),siRNA和pcDNA均用脂质体法转染至RT4细胞。运用RT-qPCR检测RT4细胞中CXCR7的mRNA表达水平;采用CCK-8法检测各组细胞的活力;Transwell实验检测各组细胞的侵袭和迁移能力;Western blot检测各组细胞中CXCR7、AKT、STAT3、p-AKT和p-STAT3的蛋白水平。结果:相较于PBS组,不同浓度PCA处理后RT4细胞的活力、迁移能力和侵袭能力显著降低(P<0.05),PCA组RT4细胞中CXCR7的mRNA和蛋白表达水平显著降低(P<0.05)。相较于si-Con组,si-CXCR7组RT4细胞中CXCR7的mRNA和蛋白表达水平显著降低,且细胞的活力、迁移能力和侵袭能力显著降低(P<0.05);相较于PCA+pcDNA组,PCA+pcDNA-CXCR7组RT4细胞中CXCR7的mRNA和蛋白表达水平显著升高,且细胞的活力、迁移能力和侵袭能力显著提高(P<0.05)。相较于PBS组,PCA组的p-AKT和p-STAT3蛋白水平显著降低;相较于PCA+pcDNA组,PCA+pcDNA-CXCR7组的p-AKT和p-STAT3蛋白水平显著升高(P<0.05)。AKT和STAT3的蛋白水平在各组之间的差异无统计学显著性。结论:PCA可能通过抑制CXCR7的表达抑制膀胱癌细胞的活力、迁移和侵袭。过表达CXCR7可逆转PCA的作用,其机制可能与AKT/STAT3信号通路有关。

AIM:To investigate the effects of procaine(PCA)and CXC chemokine receptor 7(CXCR7)on the viability,migration and invasion of bladder cancer cells and its potential mechanism.METHODS:Human bladder cancer RT4 cells were treated with PCA at different concentrations,and were divided into PBS group(without PCA treatment),PCA group(treated with 4 mmol/L PCA),siRNA negative control(si-Con)group(transfected with si-Con),CXCR7 siRNA(si-CXCR7)group(transfected with si-CXCR7),PCA+pcDNA group(treated with 4 mmol/L PCA and transfected with pcDNA)and PCA+pcDNA-CXCR7 group(treated with 4 mmol/L PCA and transfected with pcDNACXCR7).The siRNA and pcDNA were transfected into the RT4 cells by liposome method.The mRNA expression of CXCR7 in the RT4 cells was detected by RT-qPCR.The cell viability was measured by CCK-8 assay.The invasion and migration abilities of the cells were detected by Transwell assays.The protein levels of CXCR7,AKT,STAT3,p-AKT and p-STAT3 were determined by Western blot.RESULTS:Compared with PBS group,the viability,migration ability and invasion ability of the RT4 cells treated with PCA at different concentrations were significantly decreased(P<0.05),and the expression of CXCR7 at mRNA and protein levels in PCA group was also significantly decreased(P<0.05).Compared with si-Con group,the expression of CXCR7 at mRNA and protein levels in si-CXCR7 group was significantly decreased,and the viability,migration ability and invasion ability of the cells were also significantly decreased(P<0.05).Compared with PCA+pcDNA group,the expression of CXCR7 at mRNA and protein levels in PCA+pcDNA-CXCR7 group was significantly increased,and the viability,migration ability and invasion ability of the cells were also significantly increased(P<0.05).Compared with PBS group,the protein levels of p-AKT and p-STAT3 in PCA group were significantly decreased(P<0.05).Compared with PCA+pcDNA group,the protein levels of p-AKT and p-STAT3 in PCA+pcDNACXCR7 group were significantly increased(P<0.05).No significant difference in the protein levels of AKT and STAT3 among the groups was observed.CONCLUSION:Treatment with PCA inhibits the viability,migration and invasion of bladder cancer cells by inhibiting the expression of CXCR7.Over-expression of CXCR7 reverses this effect of PCA.Its mechanism may be related to AKT/STAT3 signaling pathway.