Inhibition of HBV replication by VPS4B and its dominant negative mutant VPS4B-K180Q in vivo.
10.1007/s11596-012-0054-2
- Author:
Jianbo XIA
1
;
Weipeng WANG
;
Lei LI
;
Zhi LIU
;
Min LIU
;
Dongliang YANG
Author Information
1. Division of Clinical Immunology, Huazhong University of Science and Technology, Wuhan, China. xjb915@126.com
- Publication Type:Journal Article
- MeSH:
ATPases Associated with Diverse Cellular Activities;
Adenosine Triphosphatases;
physiology;
Animals;
Endosomal Sorting Complexes Required for Transport;
physiology;
Female;
Genes, Dominant;
genetics;
Hepatitis B;
metabolism;
virology;
Hepatitis B virus;
physiology;
Liver;
virology;
Mice;
Mice, Inbred BALB C;
Mutation;
genetics;
Virus Inactivation
- From:
Journal of Huazhong University of Science and Technology (Medical Sciences)
2012;32(3):311-316
- CountryChina
- Language:English
-
Abstract:
This study examined the anti-hepatitis B virus (HBV) effect of wild-type (WT) vacuolar protein sorting 4B (VPS4B) and its dominant negative (DN) mutant VPS4B-K180Q in vivo in order to further explore the relationship between HBV and the host cellular factor VPS4. VPS4B gene was amplified from Huh7 cells by RT-PCR and cloned into the eukaryotic expression vector pXF3H. Then, the VPS4B plasmid and the VPS4B-K180Q mutation plasmid were constructed by using the overlap extension PCR site-directed mutagenesis technique. VPS4B and HBV vectors were co-delivered into mice by the hydrodynamic tail-vein injection to establish HBV vector-based models. Quantities of HBsAg and HBeAg in the mouse sera were determined by ElectroChemiLuminescence (ECL). HBV DNA in sera was measured by real-time quantitative PCR. Southern blot analysis was used to assay the intracellular HBV nuclear capsid-related DNA, real-time quantitative PCR to detect the HBV-related mRNA and immunohistochemical staining to observe the HBcAg expression in the mouse liver tissues. Our results showed that VPS4B and its mutant VPS4B-K180Q could decrease the levels of serum HBsAg, HBeAg and HBV-DNA. In addition, the HBV DNA replication and the mRNA level of HBV in the liver tissues of treated mice could be suppressed by VPS4B and VPS4B-K180Q. It was also found that VPS4B and VPS4B-K180Q had an ability to inhibit core antigen expression in the infected mouse liver. Furthermore, the anti-HBV effect of mutant VPS4B-K180Q was more potent than that of wild-type VPS4B. Taken together, it was concluded that VPS4B and its DN mutant VPS4B-K180Q have anti-HBV effect in vivo, which helps develop molecular therapeutic strategies for HBV infection.
