小干扰RNA沉默巨噬细胞移动抑制因子基因对肝癌细胞p27表达的影响

目的 观察巨噬细胞移动抑制因子(MIF)和细胞周期调控因子p27在肝细胞癌中表达的相互关系,探讨小干扰RNA(siRNA)沉默MIF基因对肝癌细胞p27表达的影响.方法 免疫组织化学法和荧光定量PCR法检测MIF、p27的蛋白和mRNA在肝癌及其癌旁组织中的表达情况.化学合成MIF siRNA和对照siRNA,脂质体法转染肝癌细胞PLC和Hep3B,荧光定量PCR法检测MIF和p27 mRNA在实验组及对照组中的表达情况.根据不同资料分别采用X2检验、Logistic回归分析或单因素方差分析.结果 MIF蛋白及其mRNA在肝癌组织中过表达,在癌旁组织中低表达;p27蛋白及其mRNA在癌组织中低表达,在癌旁组织中高表达.Logistic回归分析提示MIF为肝癌发生的危险因素,p27为保护因素.MIF mRNA在肝癌细胞株中过表达(F=61.036,P＜0.01),p27 mRNA在正常肝细胞L02中高表达(F=529.853,P＜0.01).经MIFsiRNA转染后,MIF mRNA在PLC及Hep3B中的表达水平降低,并且呈剂量依赖关系(F值分别为320.1和201.2,P值均＜0.01);p27 mRNA伴随MIF mRNA的降低而增加(F值分别为419.4和459.9,P值均＜0.01).结论 MIF在肝细胞癌中过表达,MIF siRNA能特异性抑制其在肝癌细胞中的表达;MIF可能参与了p27基因表达的调控.

Decreased macrophage migration inhibitor factor expression up-regulats p27 in hepatocellular carcinoma

Objective To observe the expression of macrophage migration inhibition factor(MIF)and p27 in hepatocellular carcinoma tissue,and to investigate the effect of MIF on the expression of p27 in hepatocellular carcinoma(HCC)cells.Methods Immunohistochemistry and quantitative RT-PCR were performed to detect the expression of MIF and p27 in HCC tissues and peri-tumor tissues.Specific small interfering RNA(siRNA)targeting MIF gene was chemically synthesized and then transfected at the concentration of 50 nmol/L and 100 nmol/L into PLC cells and Hep3B cells.The mRNA levels of MIF and p27 after MIF siRNA treatment were quantified by real-time RT-PCR.Results MIF protein and mRNA were overexpressed in the HCC tumor tissues compared to these in the peri-tumor tissues (P＜0.01).The expression of p27 protein and mRNA was significantly lower in the HCC tumor tissues compared to these in the peri-tumor tissues(P＜0.01).Compared to nodal liver cell line L-02,HCC cell lines expressed higher level of MIF (F=61.036,P＜0.01)and lower level of p27(F=529.853,P＜0.01).In MIF siRNA treated PLC and Hep3B cells,the MIF mRNA was decreased in a dose-dependent manner(F=320.1,P＜0.01;F=201.2,P＜0.01).The p27 mRNA was significantly up-regulated in MIF siRNA treated PLC and Hep3B cells compared to control siRNA transfected cells(F=419.4,P＜0.01;F=459.9,P＜0.01).Conelusions MIF iS overexpressed in HCC tumor tissues,and the expression of p27 is repressed by MIF.