Hepatitis: Tang H

Zhang YY, Chen EQ, Yang J, Duan YR, Tang H. · Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu 610041, Sichuan Province, PR China. · Virol J. · Pubmed #19467157 links to  free full text

Abstract: BACKGROUND: Currently, there is no evidence on the combination of lamivudine and thymosin alpha-1 on chronic hepatitis B patients. The aim of this study was to compare the effect of lamivudine monotherapy with that of lamivudine and thymosin alpha-1 combination therapy for the treatment of hepatitis B e antigen (HBeAg)-positive hepatitis B patients. RESULTS: We searched PUBMED (from 1966 onwards), EMBASE (from 1966), CBMdisk (Chinese Biomedical Database, from 1978), CNKI (National Knowledge Infrastructure, from 1980), the Cochrane Central Register of Controlled Trials and the Cochrane Database of Systematic Reviews. Eight trials (583 patients in total) were identified. The lamivudine and thymosin alpha-1 combination treatment was significantly superior to lamivudine treatment in terms of ALT normalization rate (80.2% vs. 68.8%, P = 0.01), virological response rate (84.7% vs. 74.9%, P = 0.002), and HBeAg seroconversion rate (45.1% vs. 15.2%, P < 0.00001). CONCLUSION: Among HBeAg-positive patients, thymosin alpha-1 and lamivudine combination therapy may be more effective than lamivudine monotherapy, providing superior rates of biochemical response, virological response, and HBeAg seroconversion.

Tang H, Oishi N, Kaneko S, Murakami S. · Division of Biotherapy of Infectious Diseases, Key Laboratory of Biotherapy of Human Diseases, Ministry of Education, Chengdu, Sichuan, China. · Cancer Sci. · Pubmed #16984372 No free full text.

Abstract: Chronic infection of hepatitis B virus (HBV) is one of the major causes of hepatocellular carcinoma (HCC) in the world. Hepatitis B virus X protein (HBx) has been long suspected to be involved in hepatocarcinogenesis, although its oncogenic role remains controversial. HBx is a multifunctional regulator that modulates transcription, signal transduction, cell cycle progress, protein degradation pathways, apoptosis, and genetic stability by directly or indirectly interacting with host factors. This review focuses on the biological roles of HBx in HBV replication and cellular transformation in terms of the molecular functions of HBx. Using the transient HBV replication assay, ectopically expressed HBx could stimulate HBV transcription and replication with the X-defective replicon to the level of those with the wild one. The transcription coactivation is mainly contributing to the stimulatory role of HBx on HBV replication although the other functions may affect HBV replication. Effect of HBx on cellular transformation remains controversial and was never addressed with human primary or immortal cells. Using the human immortalized primary cells, HBx was found to retain the ability to overcome active oncogene RAS-induced senescence that requires full-length HBx. At least two functions of HBx, the coactivation function and the ability to overcome oncogene-induced senescence, may be cooperatively involved in HBV-related hepatocarcinogenesis.

Tang H, Peng T, Wong-Staal F. · Immusol Inc., San Diego, CA 92121, USA. · Curr Opin Pharmacol. · Pubmed #12324256 No free full text.

Abstract: A new paradigm for antiviral therapy focuses on targeting cellular genes that are critical for viral replication and pathogenesis. Several new technologies have contributed to the identification of such genes for human immunodeficiency virus and hepatitis C virus. These include proteomic approaches that identify cellular proteins that physically interact with viral proteins, genomic approaches that identify and functionally validate essential cellular co-factor genes, and novel cell biology systems that facilitate studies of virus-host interactions.

He Q, Chen SC, Wang S, Jiang XL, Xu C, Zhang B, Li LX, Tang H, Yang Y, Wang WY, Zhao LS. · East Lake Hospital of Shenzhen, Shenzhen 518020, China. · Zhonghua Gan Zang Bing Za Zhi. · Pubmed #12716519 No free full text.

Abstract: OBJECTIVE: To summarize the clinical changing characters of the clinical markers after interferon treatment in chronic hepatitis B (CHB) and make out practical indexes to predict the effect. METHODS: 150 CHB patients were randomly divided into two groups: therapeutic group (90) and control group (60) in the prospective controlled trial. The levels of endogenous interferon before treatment, interferon antibody at the end of the second month and fourth month after treatment, alanine aminotransferase (ALT) and HBV DNA in the serum were detected. Then the data was analysed to find out indexes for predicting the effect. RESULTS: (1) The clearance rate of HBeAg had no significant difference in age except for 20 - 30 and 30 - 40 (t > 2.331 2, P < 0.01). (2) It was more effective if ALT level was higher than 400 U/L before treatment and it decreased more than 50% two months after treatment. (3) The patients whose HBV DNA was negative (dot hybridization) or less than 10(6) copies/ml before treatment had higher rate of HBeAg clearance. (4) There was no effect on patients whose interferon antibody turned positive at the end of the second month. (5)A predictive method of comprehensive factors was made out, whose sensitivity, specificity, and accuracy were 80%, 100% and 90%, respectively. CONCLUSION: The clinical characters of these Chinese patients are different from those of the westerners and the effects of interferon have close relation to the levels of ALT, HBV DNA and interferon antibody.

Mutimer D, Dowling D, Cane P, Ratcliffe D, Tang H, O'Donnell K, Shaw J, Elias E, Pillay D. · Liver and Hepatobiliary Unit, Queen Elizabeth Hospital, Birmingham, UK. · Antivir Ther. · Pubmed #11142622 No free full text.

Abstract: alpha-Interferon has limited efficacy against chronic hepatitis B virus (HBV) infection. Nucleoside analogues may confer greater benefits, however, it is likely that combination therapies will be required for effective control of this infection. We investigated the antiviral effect of lamivudine and interferon therapy in eight patients with high HBV-DNA levels. Six patients received lamivudine/interferon combination therapy followed, after a 6-month drug-free period, with lamivudine monotherapy. Mean HBV viral load (copies/ml) reduction was significantly greater after 4 months of combination therapy (4.3 x 10(3)) compared to an equivalent period of lamivudine monotherapy (2.9 x 10(2)) (P=0.03). Two patients were given 6 months of lamivudine/interferon combination therapy followed immediately by lamivudine monotherapy. Cessation of interferon in these patients led to a rapid 1-2 log10 increase in HBV viral load. These findings suggest that alpha-interferon has a direct antiviral effect on chronic HBV infection, which may be additive to, or synergistic with lamivudine.

Mutimer D, Pillay D, Dragon E, Tang H, Ahmed M, O'Donnell K, Shaw J, Burroughs N, Rand D, Cane P, Martin B, Buchan S, Boxall E, Barmat S, Gutekunst K, McMaster P, Elias E. · Liver and Hepatobiliary Unit, Queen Elizabeth Hospital, Birmingham, UK. · J Hepatol. · Pubmed #10207815 No free full text.

Abstract: BACKGROUND/AIMS: Orthotopic liver transplantation has an established role for the treatment of patients with chronic liver failure secondary to hepatitis B virus (HBV) infection. Unfortunately, recurrent infection of the graft can be associated with aggressive disease, and with diminished graft and patient survival. Currently, the role of nucleoside analogues for prevention of graft re-infection is being evaluated. Preliminary results are encouraging, but treatment failure has been associated with emergence of drug-resistant virus. METHODS: We have studied ten consecutive patients who received lamivudine prophylaxis for prevention of HBV graft reinfection. Sequential sera, collected prelamivudine then during treatment before and after liver transplantation, were examined. Conventional serological markers were measured, as were serum viral DNA levels with a sensitive quantitative polymerase chain reaction assay. RESULTS: Lamivudine treatment effected a reduction in serum HBV levels, but six patients still had measurable viral DNA at the time of transplantation. Five patients developed graft re-infection with lamivudine-resistant virus. Resistant virus emerged 8 to 15 months post-transplant. The likelihood of emergence of resistant virus was related to the pre-treatment serum HBV titre. Persistent serum viral DNA positivity and evidence of graft re-infection during the early post-transplant period did not predict the subsequent emergence of resistant virus. CONCLUSIONS: Our observations suggest that the resistant species may be present in the viral quasispecies in the serum and liver of patients with high-level replication prior to lamivudine exposure. The resistant species can persist during lamivudine treatment prior to transplantation, and emerge following transplantation. These observations suggest strategies which might prevent the emergence of drug-resistant species, and imply that graft re-infection may be a preventable phenomenon.

Tang H, Chen EQ. · Center for Infectious Diseases, West China Hospital, Sichuan University, Chengdu, China. · Zhonghua Gan Zang Bing Za Zhi. · Pubmed #19554777 No free full text.

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Tang H, Grisé H. · Department of Biological Science, Florida State University, Tallahassee, FL 32306-4295, USA. · Clin Sci (Lond). · Pubmed #19515018 No free full text.

Abstract: HCV (hepatitis C virus) infects nearly 3% of the population worldwide and has emerged as a major causative agent of liver disease, resulting in acute and chronic infections that can lead to fibrosis, cirrhosis and hepatocellular carcinoma. Hepatitis C represents the leading cause of liver transplantation in the United States and Europe. A positive-strand RNA virus of the Flaviviridae family, HCV contains a single-stranded RNA genome of approx. 9600 nucleotides. The genome RNA serves as both mRNA for translation of viral proteins and the template for RNA replication. Cis-acting RNA elements within the genome regulate RNA replication by forming secondary structures that interact with each other and trans-acting factors. Although structural proteins are clearly dispensable for RNA replication, recent evidence points to an important role of several non-structural proteins in particle assembly and release, turning their designation on its head. HCV enters host cells through receptor-mediated endocytosis, and the process requires the co-ordination of multiple cellular receptors and co-receptors. RNA replication takes place at specialized intracellular membrane structures called 'membranous webs' or 'membrane-associated foci', whereas viral assembly probably occurs on lipid droplets and endoplasmic reticulum. Liver inflammation plays a central role in the liver damage seen in hepatitis C, but many HCV proteins also directly contribute to HCV pathogenesis. In the present review, the molecular and cellular aspects of the HCV life cycle and the role of viral proteins in pathological liver conditions caused by HCV infection are described.

Tang H, Liu L, Liu FJ, Chen EQ, Murakami S, Lin Y, He F, Zhou TY, Huang FJ. · Center of Infectious Diseases, West China Hospital of Sichuan University, Chengdu, People's Republic of China. · Arch Virol. · Pubmed #19495936 No free full text.

Abstract: The hepatitis B virus (HBV) X gene plays an important role in HBV-associated pathogenesis, especially hepatocarcinogenesis. Establishment of a stable and regulable HBx expression system will allow study of the function of this gene. Here, we describe the development of a doxycycline-inducible recombinant plasmid (pBPSTR3-FlagX) with the full-length HBV X gene and all components of the tetracycline-on ("Tet-on") gene expression system. This vector exhibited dose-dependent doxycycline-dependent induction of the Flag-HBx protein in HepG2 and Hep3B cells. We also observed dose-dependent doxycycline transactivation of HBx in HepG2 cells. After transfecting HepG2 cells with the pBPSTR3-FlagX plasmid, we isolated five puromycin-resistant cell clones with stable HBx expression, two of which exhibited stable and tight control of HBx expression by doxycycline. This new system has great potential for functional studies of the HBV X gene.

Liu Z, Robida JM, Chinnaswamy S, Yi G, Robotham JM, Nelson HB, Irsigler A, Kao CC, Tang H. · Department of Biological Science, Florida State University, Tallahassee, FL 32306-4295, USA. · Hepatology. · Pubmed #19489073 No free full text.

Abstract: Hepatitis C virus (HCV) infection leads to acute and chronic liver diseases, and new classes of anti-HCV therapeutics are needed. Cyclosporine A (CsA) inhibits HCV replication and CsA derivatives that lack the immunosuppressive function are currently in clinical trials as candidate anti-HCV drugs. Here we characterize several independently derived HCV replicons with varying levels of CsA resistance due to mutations in nonstructural protein 5B (NS5B), the HCV-encoded polymerase. Mutant HCV replicons engineered with these mutations showed resistance to CsA. The mutations reside in two distinct patches in the polymerase: the template channel and one face of a concave surface behind the template channel. Mutant NS5B made by cells expressing the HCV replicon had increased ability to bind to RNA in the presence of CsA. Purified recombinant NS5B proteins containing the mutations were better at de novo initiated RNA synthesis than the wild-type control. Furthermore, the mutant proteins were able to bind RNA with approximately 8-fold higher affinity. Last, mutation near the template channel alleviated the lethal phenotype of a mutation in the concave patch, P540A. This intramolecular compensation for the HCV replicase function by amino acid changes in different domains was further confirmed in an infectious cell culture-derived virus system. Conclusion: An increased level of CsA resistance is associated with distinct mutations in the NS5B gene that increase RNA binding in the presence of CsA, and the intramolecular communications between residues of the thumb and the C-terminal domains are important for HCV replicase function.

Liu Z, Yang F, Robotham JM, Tang H. · Department of Biological Science, Florida State University, Tallahassee, FL 32306-4295, USA. · J Virol. · Pubmed #19386705 No free full text.

Abstract: Replication of hepatitis C virus (HCV) RNA occurs on intracellular membranes, and the replication complex (RC) contains viral RNA, nonstructural proteins, and cellular cofactors. We previously demonstrated that cyclophilin A (CyPA) is an essential cofactor for HCV infection and the intracellular target of cyclosporine's anti-HCV effect. Here we investigate the mechanism by which CyPA facilitates HCV replication. Cyclosporine treatment specifically blocked the incorporation of NS5B into the RC without affecting either the total protein level or the membrane association of the protein. Other nonstructural proteins or viral RNAs in the RC were not affected. NS5B from the cyclosporine-resistant replicon was resistant to this disruption of RC incorporation. We also isolated membrane fractions from both naïve and HCV-positive cells and found that CyPA is recruited into membrane fractions in HCV-replicating cells via an interaction with RC-associated NS5B, which is sensitive to cyclosporine treatment. Finally, we introduced point mutations in the prolyl-peptidyl isomerase (PPIase) motif of CyPA and demonstrated a critical role of this motif in HCV replication in cDNA rescue experiments. We propose a model in which the incorporation of the HCV polymerase into the RC depends on its interaction with a cellular chaperone protein and in which cyclosporine inhibits HCV replication by blocking this critical interaction and the PPIase activity of CyPA. Our results provide a mechanism of action for the cyclosporine-mediated inhibition of HCV and identify a critical role of CyPA's PPIase activity in the proper assembly and function of the HCV RC.

Chatterji U, Bobardt M, Selvarajah S, Yang F, Tang H, Sakamoto N, Vuagniaux G, Parkinson T, Gallay P. · From the Scripps Research Institute, La Jolla, California 92037. · J Biol Chem. · Pubmed #19380579 No free full text.

Abstract: Cyclosporine A and nonimmunosuppressive cyclophilin (Cyp) inhibitors such as Debio 025, NIM811, and SCY-635 block hepatitis C virus (HCV) replication in vitro. This effect was recently confirmed in HCV-infected patients where Debio 025 treatment dramatically decreased HCV viral load, suggesting that Cyps inhibitors represent a novel class of anti-HCV agents. However, it remains unclear how these compounds control HCV replication. Recent studies suggest that Cyps are important for HCV replication. However, a profound disagreement currently exists as to the respective roles of Cyp members in HCV replication. In this study, we analyzed the respective contribution of Cyp members to HCV replication by specifically knocking down their expression by both transient and stable small RNA interference. Only the CypA knockdown drastically decreased HCV replication. The re-expression of an exogenous CypA escape protein, which contains escape mutations at the small RNA interference recognition site, restored HCV replication, demonstrating the specificity for the CypA requirement. We then mutated residues that reside in the hydrophobic pocket of CypA where proline-containing peptide substrates and cyclosporine A bind and that are vital for the enzymatic or the hydrophobic pocket binding activity of CypA. Remarkably, these CypA mutants fail to restore HCV replication, suggesting for the first time that HCV exploits either the isomerase or the chaperone activity of CypA to replicate in hepatocytes and that CypA is the principal mediator of the Cyp inhibitor anti-HCV activity. Moreover, we demonstrated that the HCV NS5B polymerase associates with CypA via its enzymatic pocket. The study of the roles of Cyps in HCV replication should lead to the identification of new targets for the development of alternate anti-HCV therapies.

Li Y, Huang T, Zhang X, Wan T, Hu J, Huang A, Tang H. · Key Laboratory of Molecular Infectious Diseases, Ministry of Education, Chongqing Medical University, 1 Yi Xue Yuan Road, 400016, Chongqing, China. · Arch Virol. · Pubmed #19225715 No free full text.

Abstract: The hepatitis B virus (HBV) posttranscriptional regulatory element (HPRE) has been demonstrated to facilitate the cytoplasmic localization of unspliced transcripts. One cellular protein, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), is known to combine with this element. However, its function on HPRE remains unclear. Here, we show that recombinant GAPDH protein binds to HPRE RNA in vitro in streptavidin pull-down assays. Functional analysis demonstrated that GAPDH inhibited HPRE function in a pDM138-HPRE chloramphenicol acetyltransferase reporter assay system. Overexpression of GAPDH depressed the expression of HBs antigen, as detected both in cells transiently expressing HBs-HPRE and in HepG2.2.15 cells. These data indicate that GAPDH may be involved in the posttranscriptional regulation of HBV, which is critical for the life cycle of HBV.

Feng Y, He F, Zhang P, Wu Q, Huang N, Tang H, Kong X, Li Y, Lu J, Chen Q, Wang B. · State Key Laboratory of Oral Diseases, Sichuan University, Chengdu 610041, China. · Antiviral Res. · Pubmed #19150374 No free full text.

Abstract: Natural killer (NK) cells and cytolytic T lymphocytes (CTL) have been implicated as important effectors of antiviral defense. We previously isolated a novel antibacterial polypeptide, which was identified as high mobility group nucleosomal-binding domain 2 (HMGN2), from human mononuclear leukocytes. This study examined the antiviral activity of HMGN2 against human hepatitis virus B. HMGN2 was isolated and purified from the acid soluble proteins of the human THP-1 cell line, and identified by mass spectrum, Western blot and antibacterial assay. The hepatitis B virus (HBV)-transfected HepG2.2.15 cell line was used in the in vitro assay system. In the range of 1-100 microg/ml HMGN2, no cytotoxicity for HepG2.2.15 cells was detected by MTT assay. When incubated with HMGN2 at 1-100 microg/ml for 72 or 144 h, there was a significant reduction in hepatitis B e antigen (HBeAg) and hepatitis B surface antigen (HBsAg) expression, which were detected by ELISA, and a significant reduction in HBV DNA copies, which was determined by the real time quantitative PCR, in the supernatant of HepG2.2.15 cells. Northern and Southern blot analysis also showed that the levels of the HBV 3.5 kb and the 2.4/2.1 kb mRNA species and HBV replicative intermediate DNA were significantly reduced in the HMGN2-treated HepG2.2.15 cells. These results indicated that HMGN2 protein could markedly inhibit HBV protein expression and replication in vitro.

Tong A, Gou L, Lau QC, Chen B, Zhao X, Li J, Tang H, Chen L, Tang M, Huang C, Wei YQ. · West China Hospital, Sichuan University, Chengdu, 610041, People's Republic of China. · J Proteome Res. · Pubmed #19117405 No free full text.

Abstract: The hepatitis B virus-encoded X (HBx) protein coactivates transcription of a variety of viral and cellular genes and it is believed to play essential roles in viral replication and hepatocarcinogenesis. To examine the pleiotropic effects of HBx protein on host cell protein expression, we utilized 2-DE and MS analysis to compare and identify differentially expressed proteins between a stable HBx-transfected cell line (HepG2-HBx), constitutively expressing HBx, and vector control cells. Of the 60 spots identified as differentially expressed (+/- over 2-fold, p < 0.05) between the two cell lines, 54 spots were positively identified by MS/MS analysis. Several recent studies suggested that HBx was involved in regional hypermethylation of tumor suppressor genes and global hypomethylation of satellite 2 repeats during hepatocarcinogenesis; however, no specific gene has been reported as hypomethylated by HBx. Promoter methylation analysis was examined for those protein spots showing significant alterations, and our results revealed that specific genes, such as aldehyde dehydrogenase 1 (ALDH1), can be hypomethylated by HBx, and two calcium ion-binding proteins, S100A6 and S100A4, were hypermethylated by HBx and could be re-expressed by AZA (DNA methylase inhibitor) treatment. Moreover, via cluster and pathway analysis, we proposed a hypothetical model for the HBx regulatory circuit involving aberrant methylation of retinol metabolism-related genes and calcium homeostasis-related genes. In summary, we profiled proteome alterations between HepG2-HBx and control cells, and found that HBx not only induces regional hypermethylation but also specific hypomethylation of host cell genes.

Tang H, Da L, Mao Y, Li Y, Li D, Xu Z, Li F, Wang Y, Tiollais P, Li T, Zhao M. · State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, PR China. · Hepatology. · Pubmed #19065679 No free full text.

Abstract: Human beclin 1 is the first identified mammalian gene to induce autophagy. It is commonly expressed at reduced levels in breast tumors; however, it is overexpressed in hepatitis B virus (HBV)-infected cancerous liver tissues. To expose the possible mechanism and biological significance of this up-regulation of beclin 1, we investigated the regulation of beclin 1 expression by HBV x protein (HBx) in hepatic or hepatoma cell lines. Here, we showed that enforced expression of HBx by transfection technology results in the up-regulation of the endogenous messenger RNA (mRNA) and protein levels of Beclin 1 in the tested cells. Using a luciferase- reporter assay, we demonstrated that HBx transactivates beclin 1 promoter activity in a dose-dependent manner. The promoter region of the beclin 1 gene identified in this study is located at nt -277/+197 and has the maximum transcriptional activity. HBx-mediated up-regulation of beclin 1 expression might be direct, that is, via its promoter. Furthermore, the cells that transiently or stably expressed HBx showed an enhanced accumulation of vacuoles carrying the autophagy marker LC3 as compared with the control cells, which was induced by nutrient starvation, indicating HBx-enhanced autophagy. Moreover, this enhanced autophagy occurred in HepG2.2.15 cells that replicate HBV and in cells transfected with HBV genomic DNA, suggesting that HBV infection also causes increased levels of autophagy under starvation conditions. Treatment of cells with beclin 1 small interfering RNA (siRNA) blocked HBx-enhanced autophagy, demonstrating that the function of HBx in influencing autophagy is Beclin 1 dependent. Conclusion: Our findings suggest a novel function of HBx in increasing autophagy through the up-regulation of beclin1 expression, and this may provide an important mechanism in HBV-infected hepatocytes growing under nutrient-deficient conditions.

Robotham JM, Nelson HB, Tang H. · Department of Biological Science, Florida State University, Tallahassee, FL, USA. · Methods Mol Biol. · Pubmed #19009265 No free full text.

Abstract: Because HCV RNA-dependent RNA polymerase is error-prone and the viral RNA has a high turnover rate, the genetic diversity of HCV is very high both in vitro and in vivo. The mutation rate in long-term replicon cultures approaches 3.0 x 10(-3) base substitutions/site/year in this in vitro replication model. A direct consequence of the high mutation rate is the rapid emergence of drug-resistant variants, both in cell culture and in patients. Selectable replicons have been used extensively to isolate and characterize drug-resistant HCV genomes in vitro. Typically, replicon cells are plated at a low density and then subjected to a double selection by G418 and escalating dosages of a compound of choice. Here we describe an alternative screening assay that takes advantage of an HCV replicon that is amenable to live-cell sorting with a suitable flow cytometer. We also present a strategy for determining the relative contribution to the resistance by viral genome and host cells. We use selection and characterization of Cyclosporine A (CsA)-resistant replicons as a example to present the protocols, but this method can easily be adapted for the selection of replicon cells resistant to other chemical compounds as long as the compound does not fluoresce at the same wavelength as the fluorescent reporter protein in the replicon.

Zheng D, Chen G, Guo B, Cheng G, Tang H. · Center for Infection and Immunity, Institute of Biophysics, Chinese Academy of Sciences, 15 Da Tun Road, Chaoyang District, Beijing 100101, China. · Cell Res. · Pubmed #18957937 No free full text.

Abstract: Infections by coronaviruses such as severe acute respiratory syndrome (SARS) coronavirus (SCoV) and mouse hepatitis virus A59 (MHV-A59) result in very little type I interferon (IFN) production by host cells, which is potentially responsible for the rapid viral growth and severe immunopathology associated with SARS. However, the molecular mechanisms for the low IFN production in cells infected with coronaviruses remain unclear. Here, we provide evidence that Papain-like protease domain 2 (PLP2), a catalytic domain of the nonstructural protein 3 (nsp3) of MHV-A59, can bind to IRF3, cause its deubiquitination and prevent its nuclear translocation. As a consequence, co-expression of PLP2 strongly inhibits CARDIF-, TBK1- and IRF3-mediated IFNbeta reporter activities. In addition, we show that wild-type PLP2 but not the mutant PLP2 lacking the deubiquitinase (DUB) activity can reduce IFN induction and promote viral growth in cells infected with VSV. Thus, our study uncovered a viral DUB which coronaviruses may use to escape from the host innate antiviral responses.

He QS, Tang H, Zhang J, Truong K, Wong-Staal F, Zhou D. · iTherX Pharmaceutical Inc., San Diego, CA 92121, USA. · J Virol Methods. · Pubmed #18782589 No free full text.

Abstract: A major issue of current virology concerns the characterization of cellular proteins that operate as functional components of the viral multiplication process. RNAi is a powerful tool to elucidate gene functions. In this study three RNAi approaches (transient transfection, stable transduction and inducible RNAi) were assessed to validate human RNA helicase A (RHA) as an essential factor in hepatitis C virus (HCV) replication. It indicated that RHA transient knockdown by synthetic siRNA had no effect on HCV replication, while RHA stable knockdown via lentivector transduction caused cell lethality. The involvement of RHA in HCV replication was verified by an RNAi inducible system that, on the one hand, maintained long-term gene silencing, but on the other hand, alleviated siRNA toxicity during the essential gene silencing. A 21-day follow-up of the response of HCV replication to the presence and absence of RNAi indicated that RHA is a cellular factor involved in the HCV replication process.

Tang H, Huang Y, Chen J, Yu C, Huang AL. · Key Laboratory of Molecular Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing, China. · Intervirology. · Pubmed #18753794 No free full text.

Abstract: OBJECTIVE: To identify T-cell intracellular antigen 1 (TIA-1) interacting with the posttranscriptional regulatory element (PRE) of hepatitis B virus (HBV), and reveal TIA-1 function on HBV gene expression. METHODS: In order to demonstrate TIA-1 binding to PRE, electrophoretic mobility shift and RNA and protein pull-down assays were used. The pDM138-PRE reporter system, HepG2.2.15 cell line and HBs-PRE transient expression cells were adopted to identify TIA-1 function in the HBV life cycle. RESULTS: TIA-1 could directly bind to PRE RNA. The functional binding region of TIA-1 was identified. Functional analysis indicated that TIA-1 could dose-dependently inhibit PRE function in pDM138-PRE system. Down-expression of HBs antigen induced by TIA-1 was detected both in the HepG2.2.15 cell line and in HBs-PRE transient expression cells. CONCLUSION: These data demonstrated that TIA-1 inhibits HBs expression by interacting with PRE.

Tong A, Wu L, Lin Q, Lau QC, Zhao X, Li J, Chen P, Chen L, Tang H, Huang C, Wei YQ. · The State Key Laboratory of Biotherapy, West China Hospital, College of Life Science, Sichuan University, Chengdu, People's Republic of China. · Proteomics. · Pubmed #18491315 No free full text.

Abstract: Hepatitis B virus (HBV) is one of the major etiological factors responsible for acute and chronic liver disease and for the development of hepatocellular carcinoma (HCC). To determine the effects of HBV replication on host cell-protein expression, we utilized 2-DE and MS/MS analysis to compare and identify differentially expressed proteins between an HBV-producing cell line HepG2.2.15 and its parental cell line HepG2. Of the 66 spots identified as differentially expressed (+/- over twofold, p <0.05) between the two cell lines, 62 spots (corresponding to 61 unique proteins) were positively identified by MS/MS analysis. These proteins could be clearly divided into three major groups by cluster and metabolic/signaling pathway analysis: proteins involved in retinol metabolism pathway, calcium ion-binding proteins, and proteins associated with protein degradation pathways. Other proteins identified include those that function in diverse biological processes such as signal transduction, immune regulation, molecular chaperone, electron transport/redox regulation, cell proliferation/differentiation, and mRNA splicing. In summary, we profiled proteome alterations between HepG2.2.15 and HepG2 cells. The proteins identified in this study would be useful in revealing the mechanisms underlying HBV-host cell interactions and the development of HCC. This study can also provide some useful clues for antiviral research.

Tang H, Min G, Ge B, Li Y, Liu X, Jiang S. · Key Laboratory for Natural Medicine of Gansu Province, Lanzhou Institute of Chemical Physics, Chinese Academy of Sciences, Lanzhou 730000, PR China. · J Ethnopharmacol. · Pubmed #18403144 No free full text.

Abstract: AIM OF THE STUDY: Chi-Zhi-Huang decoction (PGR) is one of the traditional Chinese medicine (TCM) preparations with unique effect on withdrawing jaundice and has been used to treat icteric patients in China for many years. In this research, we aim at to evaluate the potential activity of PGR in restoring hepatic drug metabolism in a damaged liver. MATERIALS AND METHODS: A cocktail approach with caffeine (10mg/kg), dapsone (10mg/kg) and chlorzoxazone (20mg/kg) respectively as probe drug of cytochrome P450 (CYP) isoform of CYP 1A2, 3A4 and 2E1 was used to evaluate its possible effects on Phase I oxidative metabolism. Pretreated with three dosages of PGR water extract (0.75, 1.5 and 3g/kg, po) for 5 days, male Wistar rats (220-240 g) were intoxicated by phenylisothiocyanate (PITC, 100mg/kg, po) 24h before probes intravenous injection. The pharmacokinetics of the probes in the blood was determined simultaneously by HPLC, and their non-compartmental parameters were used to evaluate the metabolic difference among the groups. Moreover, the levels of liver enzymes (ALT, AST, ALP) and bilirubins were also measured for insight of liver function. RESULTS: The findings in this study suggest that PGR induces CYP 3A4, does not have much effect on CYP 2E1, and inhibits CYP 1A2 at high dosage. CONCLUSION: The current pharmacokinetic approach allowed the protective effects of PGR on oxidative drug metabolism in damaged liver to be systemically examined and will certainly help in the explanation of synergistic effect of the composites formula.

Yang F, Robotham JM, Nelson HB, Irsigler A, Kenworthy R, Tang H. · Department of Biological Science, Florida State University, Tallahassee, Florida 32306-4370, USA. · J Virol. · Pubmed #18385230 links to  free full text

Abstract: Cyclosporine (CsA) and its derivatives potently suppress hepatitis C virus (HCV) replication. Recently, CsA-resistant HCV replicons have been identified in vitro. We examined the dependence of the wild-type and CsA-resistant replicons on various cyclophilins for replication. A strong correlation between CsA resistance and reduced dependency on cyclophilin A (CyPA) for replication was identified. Silencing of CyPB or CyPC expression had no significant effect on replication, whereas various forms of small interfering RNA (siRNA) directed at CyPA inhibited HCV replication of wild-type but not CsA-resistant replicons. The efficiency of a particular siRNA in suppressing CyPA expression was correlated with its potency in inhibiting HCV replication, and expression of an siRNA-resistant CyPA cDNA rescued replication. In addition, an anti-CyPA antibody blocked replication of the wild-type but not the resistant replicon in an in vitro replication assay. Depletion of CyPA alone in the CsA-resistant replicon cells eliminated CsA resistance, indicating that CyPA is the chief mediator of the observed CsA resistance. The dependency on CyPA for replication was observed for both genotype (GT) 1a and 1b replicons as well as a GT 2a infectious virus. An interaction between CyPA and HCV RNA as well as the viral polymerase that is sensitive to CsA treatment in wild-type but not in resistant replicons was detected. These findings reveal the molecular mechanism of CsA resistance and identify CyPA as a critical cellular cofactor for HCV replication and infection.

Liao Y, Lu X, Lu C, Li G, Jin Y, Tang H. · Department of Physiology, School of Basic Medicine, China Medical University, Shenyang, Liaoning, PR China. · Pharmacol Res. · Pubmed #18282716 No free full text.

Abstract: The objective of this study was to explore the optimal combination of agents used along with cisplatin for protection of hepatotoxicity. Animal experiment was carried out based on the orthogonal design L(8) (2(7)) setting seven factors with two different levels of each, and eight groups of mice were needed. The agents tested in this study were zinc, selenium, fosfomycin, sodium thiosulfate (STS), N-acetyl-cysteine (NAC), methionine and taurine. Mice were supplemented by gavage with various combinations of agents as designed in the orthogonal table once a day for nine days beginning two days before cisplatin administration. 3.5mg/kg body weight of cisplatin was given intraperitoneally once a day for five days simultaneously. After cessation of cisplatin administration, the agents were supplemented continuously for two days. Activities of alanine aminotransferase (ALT) in serum, levels of glutathione (GSH) and malondialdehyde (MDA) in liver were analyzed after cessation of supplementation. Results showed zinc, fosfomycin and methionine were the effective factors for protection of weight loss; fosfomycin and methionine were the effective factors for prevention of decreased liver ratio; selenium, fosfomycin and STS were the effective factors for prevention of increased ALT activities in serum. On the other hand, methionine was the only effective factor for prevention of decreased GSH levels in liver; zinc, selenium and fosfomycin were the effective factors for prevention of increased MDA levels in liver. Based on the data observed in this study, the optimum combinations of agents were selenium, fosfomycin, methionine and taurine, and zinc, selenium, STS and methionine. In conclusion, each agent used in this study could play a beneficial role for prevention of cisplatin hepatotoxicity, however, none could play the crucial role. The potentiated actions for prevention of cisplatin hepatotoxicity could be achieved via combined use of these agents.

Liu P, Xiang K, Tang H, Zhang W, Wang X, Tong X, Takebe Y, Yang R. · The State Key Laboratory of Virology, Wuhan Institution of Virology, Chinese Academy of Sciences, Wuhan, Hubei 430071, China. · AIDS Res Hum Retroviruses. · Pubmed #18275341 No free full text.

Abstract: The prevalence of hepatitis C virus (HCV) among human immunodeficiency virus (HIV-1)-positive former blood donors (FBDs) in Hubei province, central China, and the subtypes of these two viruses are identified. HIV-1-positive specimens were collected from FBDs, transfusion recipients, and their sexual partners in Hubei province, central China. The prevalence of HCV in HIV-1-positive FBDs was 78.6%. The dominant circulating HIV-1 subtype of FBDs was subtype B' (Thai-B); one belonged to U.S.-European subtype B. HCV genotypes 2a (78.6%) and 1b (21.4%) were detected. No recombinant form of HIV-1 was identified. Non-B' subtypes occurring among FBDs indicate the complexity of the HIV-1 prevalence in central China, where HIV is beginning to spread into the general population.