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Fan H.,CAS Institute of Microbiology | Yan X.,CAS Institute of Microbiology | Zhang Y.,CAS Institute of Microbiology | Zhang X.,CAS Institute of Microbiology | And 4 more authors.
PLoS ONE | Year: 2013

Elevated expression of heat shock protein gp96 in hepatitis B virus (HBV)-infected patients is positively correlated with the progress of HBV-induced diseases, but little is known regarding the molecular mechanism of virus-induced gp96 expression and its impact on HBV infection. In this study, up-regulation of gp96 by HBV replication was confirmed both in vitro and in vivo. Among HBV components, HBV x protein (HBx) was found to increase gp96 promoter activity and enhance gp96 expression by using a luciferase reporter system, and western blot analysis. Further, we found that HBx-mediated regulation of gp96 expression requires a NF-κB cis-regulatory element on the gp96 promoter, and chromatin immunoprecipitation results demonstrated that HBx promotes the binding of NF-κB to the gp96 promoter. Significantly, both gain- and loss-of-function studies showed that gp96 enhances HBV production in HBV-transfected cells and a mouse model based on hydrodynamic transfection. Moreover, up-regulated gp96 expression was observed in HBV-infected patients, and gp96 levels were correlated with serum viral loads. Thus, our work demonstrates a positive feedback regulatory pathway involving gp96 and HBV, which may contribute to persistent HBV infection. Our data also indicate that modulation of gp96 function may represent a novel strategy for the intervention of HBV infection. © 2013 Fan et al. Source


Wang S.,CAS Institute of Microbiology | Qiu L.,CAS Institute of Microbiology | Yan X.,CAS Institute of Microbiology | Jin W.,CAS Institute of Microbiology | And 9 more authors.
Hepatology | Year: 2012

Hepatitis B virus (HBV) causes chronic infection in about 350 million people worldwide. Given the important role of the most abundant liver-specific microRNA, miR-122, in hepatic function and liver pathology, here we investigated the potential role and mechanism of miR-122 in regulating HBV replication. We found that miR-122 expression in liver was significantly down-regulated in patients with HBV infection compared with healthy controls, and the miR-122 levels were negatively correlated with intrahepatic viral load and hepatic necroinflammation. The depletion of endogenous miR-122 by its antisense inhibitor led to enhanced HBV replication, whereas overexpression of miR-122 by transfection of mimic or its expression vector inhibited viral production. We next identified cyclin G 1 as an miR-122 target from multiple candidate target genes that are involved in the regulation of HBV replication. Overexpression and knockdown studies both showed that cyclin G 1 regulated viral replication in HBV transfected cells. We also observed that cyclin G 1 expression was up-regulated in HBV-infected patients, and cyclin G 1 levels were inversely associated with miR-122 expression in liver tissues. Using coimmunoprecipitation, a luciferase reporter system, and electrophoretic mobility shift assay, we further demonstrated that cyclin G 1 specifically interacted with p53, and this interaction blocked the specific binding of p53 to HBV enhancer elements and simultaneously abrogated p53-mediated inhibition of HBV transcription. Finally, we show that miR-122 suppressed HBV replication in p53 wildtype cells but not in null isogenic cells. Conclusion: miR-122 down-regulates its target cyclin G 1, and thus interrupts the interaction between cyclin G 1 and p53 and abrogates p53-mediated inhibition of HBV replication. Our work shows that miR-122 down-regulation induced by HBV infection can impact HBV replication and possibly contribute to viral persistence and carcinogenesis. (HEPATOLOGY 2012;) © 2011 American Association for the Study of Liver Diseases. Source


Zhao J.,Beijing 302 Hospital | Zhou G.,Beijing 302 Hospital | Li M.,Beijing 302 Hospital | Li W.,Beijing 302 Hospital | And 6 more authors.
Scandinavian Journal of Gastroenterology | Year: 2010

Objective. There is no animal model that displays the features of non-alcoholic steatohepatitis (NASH) characterized by insulin resistance (IR) and fibrosing steatohepatitis. This study aimed to develop a novel IR-associated rat model of NASH. Material and methods. Male SpragueDawley rats were fed with the high-fat diet (HFD) supplemented with 0.25% propylthiouracil for 2, 4, 6, 8 and 12 weeks. The IR-associated metabolic parameters, histological assessment and the expression of key insulin signaling molecules were determined. The circulating and tissue pro-inflammatory factors and adipocytokines were examined. Results. In the HFD-fed rats, the systemic and multiple-organ IR was developed after 4 weeks, whereas the histological changes characterized by steatohepatitis, inflammatory response in the visceral adipose tissue and proliferative pancreatic islet β-cells appeared after 6 weeks, concomitant with altered expression of key insulin signaling molecules. In addition, the elevated levels of serum tumor necrosis factor α (TNF-α), soluble TNF receptor2, interleukin (IL)-6, CC-chemokine ligand (CCL)2 and resistin were parallel with the severity of hepatic inflammation, while the levels of serum adiponectin, leptin and TNF-α, but not resistin, were correlated with IR. Conclusions . We have developed a systemic IR-associated NASH model of rats, with impaired insulin signaling, systemic inflammation and appropriate pathology characterized by human NASH, and provided a realistic experimental model for elucidating the association between IR and the pathogenesis of NASH. © 2010 Informa Healthcare. Source


Zhao J.,Beijing 302 Hospital | Zhao S.,Beijing 302 Hospital | Zhou G.,Beijing 302 Hospital | Liang L.,Beijing 302 Hospital | And 7 more authors.
Scandinavian Journal of Gastroenterology | Year: 2011

Objective: Lipopolysaccharide (LPS) is suspected to trigger primary biliary cirrhosis (PBC) in susceptible individuals, yet the precise mechanism of its effect in PBC remains largely unknown. The aim of this study was to investigate altered responses to LPS ligand for Toll-like receptors (TLRs) in pathogenesis of PBC in vivo and in vitro. Material and methods. In vivo, we investigated levels of LPS and pro-inflammatory cytokines in sera and expression of LPS receptors in liver tissues from 162 patients with PBC, 325 patients with other liver diseases and 80 healthy controls. In vitro, altered responses to LPS on monocytes and cultured human biliary epithelial cells (BECs) from patients with PBC were determined. Results. Significantly higher levels of LPS in patients with PBC were detected, compared with patients with other liver diseases and healthy controls. Immunohistochemically, expression of TLR4, CD14, CD68 and NF-κB was significantly enhanced in liver tissues from patients with PBC. Before LPS stimulation, we found significantly higher serum levels of tumor necrosis factor-α, interleukin (IL)-1β, IL-6 and IL-8 in patients with PBC than those in healthy controls. After LPS stimulation, TLR4 expression and pro-inflammatory cytokine production in CD14-positive monocytes and cultured BEC from patients with PBC increased significantly. Conclusions. These results indicated that patients with PBC were prone to exhibit higher serum LPS level, hypersensitivity of monocytes and BEC to LPS, and enhanced production of pro-inflammatory cytokines. LPS altered expression of TLR4, CD14 and NF-κB on monocytes and BEC, which may be implicated in the pathogenesis and progression of PBC. © 2011 Informa Healthcare. Source


Zheng L.,CAS Institute of Biophysics | Zheng L.,University of Chinese Academy of Sciences | Li D.,CAS Institute of Biophysics | Wang F.,CAS Institute of Biophysics | And 9 more authors.
Journal of Clinical Immunology | Year: 2010

Background PD-1, encoded by PDCD1, is highly expressed on virus-specific T cells and plays critical roles in modulating anti-virus immune responses in chronic viral infection. It is unknown, however, whether polymorphisms of the PDCD1 are associated with viral clearance during chronic viral infections. Methodology and principal findings Here, we used the polymerase chain reaction-restriction fragment length polymorphism method to genotype two single nucleotide polymorphisms (SNPs) of PDCD1 in 502 patients with chronic hepatitis B virus (HBV) infection and 359 healthy controls to determine the association between PDCD1 genotypes and serum viral load as well as the risk of chronic infection. Our results showed that although neither the P7209C/T SNP site nor the P8737A/G site was associated with the risk of chronic HBV infection, the P7209T allele in intron 4 is significantly associated with lower viral burden in the blood. Using a luciferase reporter assay, we demonstrated that the P7209T allele creates a negative cis-element for gene transcription. Conclusions and significance Our data provide the first evidence that PDCD1 polymorphisms is a genetic factor in pathogenesis of chronic viral infection and reveal the functional significance of the P7209 SNP of the PDCD1. © Springer Science+Business Media, LLC 2010. Source

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