丙型肝炎病毒非结构蛋白质5A抑制Hepcidin因表达并促进肝细胞内铁储留

HCV NS5A protein down-regulates hepcidin gene expression and increases hepatic intracellular iron storage

目的探讨丙型肝炎病毒（HCV）非结构蛋白质5A（NS5A）对Hepcidin基因表达的影响。方法用含HCVNS5A基因的表达质粒pcNS5A瞬时转染QSG7701细胞，采用逆转录-聚合酶链反应及Westemb10t试验观察HCVNS5A和Hepcidin的mRNA转录水平及蛋白质表达水平，铁染色后观察细胞内铁储留情况。检测数据的多组间比较行单因素方差分析，两两比较行LSD-t检验。结果转染pcNS5A质粒细胞内有HCVNS5A的mRNA和蛋白的表达，未转染质粒组和转染空白质粒pRc／CMV组细胞内无HCVNS5A的mRNA和蛋白的表达。未转染质粒组、转染pRc／CMV质粒组和转染pcNS5A质粒组细胞的HeIxzidinmRNA相对表达量分别为0．711±0．049、0．718±0．052和0．264±0．030，转染pcNS5A组HepcidinmRNA表达低于未转染质粒组和转染pRc／CMV质粒组（t值分别为-13．523和-13．045，P值均〈0．01）。转染pcNS5A质粒组细胞HeIxzidin蛋白表达较未转染质粒组及转染pRc／CMV质粒组下降，且随着转染pcNS5A质粒剂量的增加，Hepcidin蛋白表达下降更明显。铁染色结果显示，转染pcNS5A质粒细胞内铁较转染pRC／CMV质粒细胞和未转染质粒细胞高。结论HCVNS5A蛋白能抑制Hepcidin的mRNA和蛋白质表达，并引起细胞内铁的储留增加。

Objective To investigate whether the nonstructural protein 5A （NS5A） encoded by the hepatitis C virus RNA genome affects the expression of hepcidin gene. Methods HCV NS5A expression plasmid （pCN5A） and pRc/CMV were transfected into QSG7701 cells individually, RT-PCR was employed to detect the HCV NS5A and hepcidin mRNA transcription. Western blot was used for detection of HCV NS5A and hepcidin proteins. Iron was stained to evaluate the intracellular iron level. Results HCV NS5A plasmid was successfully transfected into QSG7701 cells, which was evidenced by HCV NS5A mRNA and protein from the transfected cells. The hepcidin mRNA relative quantification in untransfected cells, pRc/CMV transfected cells and pCNS5A transfected cells were 0.711 ± 0.049, 0.718 ± 0.052 and 0.264 ± 0.030 respectively. The transcription of hepcidin mRNA decreased remarkably in the cells transfected with pCNS5A plasmid as compared to the untransfected cells and pRc/CMV transfected cells（ P 〈 0.01 ）. The level of hepcidin protein expression was found also significantly lower in the pCN5A plasmid transfectedcells as compared to the untransfected cells and pRc/CMV transfected cells. The intracellular iron staining was remarkably higher in the pcNS5A transfected cells than untransfected or pRc/CMV transfected cells. Conclusions HCV NS5A inhibits the transcription of hepcidin mRNA and expression of hepcidin protein, inducing hepatic intracellular iron storage.