血红素加氧酶-1表达对HBV复制的反向调控作用  

Heme oxygenase-1 reversely regulates HBV replication in vitro and in vivo

作　　者：李磊[1] 韩华[1] 金坤[1] 刘磊[1] 吴晓玲[1] 高人焘[1] 

机构地区：[1]安徽医科大学附属省立医院感染病科,合肥230001

出　　处：《肝脏》2013年第10期667-670,679,共5页Chinese Hepatology

基　　金：安徽省教育厅自然基金资助项目(KJ2012A174);安徽省科技厅自然科学基金资助项目(1208085QH147);安医大校基金资助项目(2010xkj084)

摘　　要：目的评价血红素加氧酶-1(HO-1)对HBV复制的调控作用。方法采用分子克隆的方法分别构建人HO-1真核表达载体——pHO和HO-1 RNA干扰质粒pHi,同HBV可复制性克隆pHBV1.3体外共转染人肝癌细胞系Huh-7,检测HBV抗原分泌情况和HBV相关mRNA含量。利用HBV急性感染小鼠模型,腹腔注射钴原卟啉(CoPP)IX和锌原卟啉(ZnPP)IX分别诱导和抑制HO-1表达,检测血清HO-1水平、HBV效价,免疫组织化学染色观察HBcAg在肝细胞内表达情况。分别在细胞水平和动物体内水平,评价HO-1表达对HBV复制的调控作用。结果同空载体共转染细胞栩比,pHO共转染后HO-1的分泌水平明显上调(P<0.01).HBsAg/HBeAg的分泌受到抑制(均P<0.01);pHi共转染后HO-1的分泌水平下降(P<0.01).而HBsAg/HBeAg分泌增加(P均<0.01);但各组HBV相关RNA水平无明显差异。CoPPIX注射后诱导小鼠血清HO-1高表达(P<0.01),ZnPPIX则有效抑制了HO-1表达(P<0.01)。同对照组相比,CoPPIX组小鼠血清HBV DNA含量降低,肝脏内HBcAg阳性染色信号也随之明显减弱(P均<0.01);而ZnPPIX组小鼠血清HBV DNA含量增加,肝脏内HBcAg表达明显增强(均P<0.01)。结论上调HO-1表达可有效抑制HBV复制,而下调其表达有利于HBV复制,且其可能是在转录后环节上发挥抗HBV作用。Objective To evaluate the regulation of heme oxygenase-1 on HBV replication. Methods Human HO-1 eukaryotic expression vector -pHO and siRNA expression plasmid-pHi were constructed by molecular cloning, respectively. Then they were co-transfected into Huh-7 cell line with HBV replication competent clone-pHBV1. 3 in vitro. Levels of HBsAg and HBeAg in supernatant were detected by ELISA. HBV related mRNA was quantified by RT-PCR. Acute HBV infection mice model were intraperitoneal injected with CoPPIX and ZnPPIX to induce and suppress HO-1 expression in vivo, respectively. Level of HO-1 and expression of HBV from serum, and expression of HBcAg from liver by immunohis- tochemical staining were detected. It was evaluated that the expression of HO-1 regulated HBV replication in vitro and in vivo. Results Compared to control, pHO plasmid co-transfected cells secreted more level of HO-1 （P〈0. 01）, and secretion of HBsAg/HBeAg was inhibited （P〈0.01）. On the contrary, secretion of HO-1 was suppressed significantly （P〈0.01） in phi co-transfected cells, while secretion of HBsAg/HBeAg was increased （P〈0. 01 ）. No significant difference on HBV related RNA level was observed in different transfected cells. HO-1 expression was induced by CoPPIX injection in mice （P〈0.01）, while its expression was effectively inhibited by ZnPPIX （P〈0.01）. Compared to control, serum HBV DNA level was lower and positive staining signal of HBcAg in the liver was also decreased obviously in mice treated with CoPPIX （P〈0. 01）. Meanwhile, serum HBV DNA and HBeAg expression increased in mice treated with ZnPPIX （P〈0.01）. Conclusion HBV replication can be effectively suppressed by up-regulating HO-1 expression, On the contrary, HBV replication can be promoted by down-regulating HO-1 expression. The mechanism of HO-l＇s anti-HBV function probably acts in the post-transcriptional phase in HBV replication life cycle.

关 键 词：血红素加氧酶1 乙型肝炎病毒 复制 高压水注射 RNA干扰 

分 类 号：R512.62[医药卫生—内科学]