Na^(+)/K^(+) ATPase β1亚基对乙型肝炎病毒复制的影响及其作用机制  

Effect of Na^(+)/K^(+) ATPase β1 subunit on hepatitis B virus replication and relevant mechanism

作　　者：曹扬[1] 王莹[1] CAO Yang;WANG Ying(First Hospital of Jilin University t Changchun 130021,Jilin Province,China)

机构地区：[1]吉林大学第一医院,吉林长春130021

出　　处：《中国生物制品学杂志》2021年第8期910-916,共7页Chinese Journal of Biologicals

基　　金：国家自然科学基金(81802050)。

摘　　要：目的探讨Na^(+)/K^(+) ATPaseβ1亚基(ATP1B1)对乙型肝炎病毒(hepatitis B virus,HBV)复制的影响及其作用机制。方法将稳定表达HBV的质粒pHBV1.3分别与质粒VR1012-ATP1B1及siRNA ATP1B1序列共转染Hep G2细胞,ELISA法检测细胞培养上清中HBs Ag的含量,qRT-PCR法检测细胞中HBV DNA及RNA含量。Western blot法检测过表达ATP1B1的Hep G2细胞中维甲酸诱导基因蛋白-Ⅰ(retinoic acid-inducible gene-Ⅰ,RIG-Ⅰ)和黑色素瘤分化相关抗原5(melanoma differentiation-associated 5,MDA5)蛋白的表达,qRT-PCR法检测细胞因子(IFNα、IFNβ、IFNγ、IL-28a、IL-29)及IFN诱导基因(interferon-stimulated gene,ISG)mRNA含量,以转染载体VR012为对照。结果过表达ATP1B1可显著降低HepG2细胞培养上清中HBs Ag及HBV DNA、RNA含量(P <0.01);沉默ATP1B1可显著升高HepG2细胞培养上清中HBsAg及HBV DNA、RNA含量(P <0.01)。与VR012组比较,过表达ATP1B1的Hep G2细胞中RIG-Ⅰ和MDA5蛋白表达升高;IFNα、IFNβ和IL-29 mRNA含量明显升高(P <0.05),IFNγ和IL-28A mRNA含量差异无统计学意义(P> 0.05):ISG15、ISG56、OAS2和BST-2 mRNA含量显著升高(P <0.01),GBP-1 mRNA含量差异无统计学意义(P> 0.05)。结论 ATP1B1可抑制HBV的复制,可能是通过上调细胞内RIG-Ⅰ及MDA5蛋白,诱导Ⅰ型IFN表达,激活下游抗病毒因子ISG发挥作用。Objective To investigate the effect of Na^(+)/K^(+) ATPase β1 subunit(ATP1B1)on hepatitis B virus(HBV)replication as well as the relevant mechanism. Methods HepG2 cells were co-transfected with plasmid pHBV1.3 stably expressing HBV,plasmid VR1012-ATP1B1 and siRNA ATP1B1. The HBsAg content in cell culture supernatant was determined by ELISA,while the HBV DNA and RNA contents in cells by qRT-PCR. The expression levels of retinoic acidinducible gene-Ⅰ(RIG-Ⅰ)and melanoma differentiation-associated gene 5(MDA5)in HepG2 cells overexpressing ATP1B1 were determined by Western blot,while the mRNA contents of cytokines(IFNα,IFNβ,IFNγ,IL-28 a,IL-29)and interferonstimulated gene(ISG)by qRT-PCR,using those in the cells transfected with vector VR1012 as control. Results Overexpression of ATP1B1 significantly decreased the contents of HBsAg and HBV DNA and RNA in cultured supernatant of HepG2 cells(P < 0. 01),while silencing ATP1B1 significantly increased the above-mentioned contents(P < 0. 01). Compared with those in the cells transfected with vector VR1012, the expressions of RIG-Ⅰ and MDA5 in HepG2 cells overexpressing ATP1B1 increased,while the IFNα,IFNβ and IL-29 mRNA contents increased significantly(P < 0. 05),the IFNγ and IL-28 A mRNA contents showed no significant difference(P > 0. 05),the ISG15,ISG56,OAS2 and BST-2 mRNA contents increased significantly(P < 0. 01),and GBP-1 mRNA content showed no significant difference(P > 0. 05).Conclusion ATP1B1 inhibited HBV replication by a possible mechanism of upregulating intracellular RIG-Ⅰand MDA5,inducing the expression of type Ⅰ IFN and activating downstream antiviral factor ISG.

关 键 词：Na^(+)/K^(+)ATPaseβ1亚基 乙型肝炎病毒 维甲酸诱导基因蛋白-Ⅰ 黑色素瘤分化相关抗原5 干扰素诱导基因 

分 类 号：R373.9[医药卫生—病原生物学]