靶向MIF的siRNA对大肠癌细胞侵袭的影响

Small interfering RNA-mediated MIF knockdown reduces cell invasion in murine colorectal cancer cell line CT-26

目的:研究化学合成的siRNA干扰巨噬细胞移动抑制因子(macrophage migration-inhibitory factor,MIF)后,对大肠癌CT-26细胞侵袭的影响并探讨其可能的机制.方法:实验组用靶向MIF的siRNA处理CT-26细胞,对照组用非特异性的siRNA处理CT-26细胞,空白组不添加任何干扰剂.Transwell细胞侵袭试验检测细胞侵袭情况,ELISA检测培养上清中MIF蛋白的含量,逆转录聚合酶链反应(RT-PCR)检测MIF、CD74、T淋巴瘤侵袭转移因子1(tiam1)、E-Cadherin mRNA的表达,Western blot检测细胞内MIF及CD74蛋白表达.结果:实验组经化学合成的MIFsiRNA处理大肠癌CT-26细胞24h后,与对照组和空白组相比,其侵袭受到明显抑制(P=0.012);实验组培养上清中MIF蛋白的含量与对照组和空白组比较有显著下降(P=0.02);实验组中MIF、CD74、tiam1的mRNA的表达较对照组和空白组相比显著下降(PMIF=0.001;PCD74=0.001;Ptiam1=0.004);实验组中E-Cadherin的表达较对照组和空白组升高(PE-Cadherin=0.001);实验组细胞内的MIF及CD74蛋白较对照组和空白组表达明显下降(PMIF=0.006;PCD74=0.016).结论:化学合成的MIFsiRNA抑制了CT-26细胞的侵袭,其可能机制是靶向MIF的siRNA抑制了MIF的表达,降低了CD74与tiam1的表达,升高了E-Cadherin的表达.

AIM：To investigate the effect of small interfering RNA（siRNA）-mediated knockdown of the mac-rophage migration-inhibitory factor（MIF） geneon cell invasion in murine colorectal cancer cell line CT-26 and to explore possible mechanisms involved.METHODS：CT-26 cells were divided into three groups：experimental group,control group,and blank group.The experimental group and con-trol group were treated with a siRNA specific for the MIF gene（MIF siRNA） and a nonspecif ic siRNA,respectively,while the blank group was not treated with any agent.Transwell assay was used to determine cell invasion.ELISA was used to determine the level of MIF protein in cell supernatants,and the expression of MIF,CD74,tiam1 and E-cadherin mRNAs was detected by RT-PCR.RESULTS：Twenty-four hours after treatment,cell invasion was signifi cantly inhibited and the level of MIF protein in supernatants signif icantly declined in the experimental group compared to the control and blank groups（P = 0.012,0.020）.Compared to the control and blank groups,the expression of MIF,CD74 and tiam1 mRNAs decreased significantly and that of E-cadherin mRNA increased signif icantly（PE-Cadherin = 0.001） in the experimental group.In addition,the levels of MIF and CD74 proteins signifi cantly declined in the experimental group compared to the con-trol and the blank groups（PMIF = 0.006;PCD74 = 0.016）.CONCLUSION：MIF siRNA inhibits the inva-sion of CT-26 cells possibly by down-regulating the expression of MIF,CD74 and tiam1 and up-regulating the expression of E-cadherin.