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The CD8+ and CD4+ T Cell Immunogen Atlas of Zika Virus Reveals E, NS1 and NS4 Proteins as the Vaccine Targets. Viruses 2022; 14:v14112332. [PMID: 36366430 PMCID: PMC9696057 DOI: 10.3390/v14112332] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/21/2022] [Revised: 10/14/2022] [Accepted: 10/17/2022] [Indexed: 02/01/2023] Open
Abstract
Zika virus (ZIKV)-specific T cells are activated by different peptides derived from virus structural and nonstructural proteins, and contributed to the viral clearance or protective immunity. Herein, we have depicted the profile of CD8+ and CD4+ T cell immunogenicity of ZIKV proteins in C57BL/6 (H-2b) and BALB/c (H-2d) mice, and found that featured cellular immunity antigens were variant among different murine alleles. In H-2b mice, the proteins E, NS2, NS3 and NS5 are recognized as immunodominant antigens by CD8+ T cells, while NS4 is dominantly recognized by CD4+ T cells. In contrast, in H-2d mice, NS1 and NS4 are the dominant CD8+ T cell antigen and NS4 as the dominant CD4+ T cell antigen, respectively. Among the synthesized 364 overlapping polypeptides spanning the whole proteome of ZIKV, we mapped 91 and 39 polypeptides which can induce ZIKV-specific T cell responses in H-2b and H-2d mice, respectively. Through the identification of CD8+ T cell epitopes, we found that immunodominant regions E294-302 and NS42351-2360 are hotspots epitopes with a distinct immunodominance hierarchy present in H-2b and H-2d mice, respectively. Our data characterized an overall landscape of the immunogenic spectrum of the ZIKV polyprotein, and provide useful insight into the vaccine development.
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Saraceni C, Birk J. A Review of Hepatitis B Virus and Hepatitis C Virus Immunopathogenesis. J Clin Transl Hepatol 2021; 9:409-418. [PMID: 34221927 PMCID: PMC8237136 DOI: 10.14218/jcth.2020.00095] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/20/2020] [Revised: 02/21/2021] [Accepted: 04/22/2021] [Indexed: 12/13/2022] Open
Abstract
Despite the advances in therapy, hepatitis B virus (HBV) and hepatitis C virus (HCV) still represent a significant global health burden, both as major causes of cirrhosis, hepatocellular carcinoma, and death worldwide. HBV is capable of incorporating its covalently closed circular DNA into the host cell's hepatocyte genome, making it rather difficult to eradicate its chronic stage. Successful viral clearance depends on the complex interactions between the virus and host's innate and adaptive immune response. One encouraging fact on hepatitis B is the development and effective distribution of the HBV vaccine. This has significantly reduced the spread of this virus. HCV is a RNA virus with high mutagenic capacity, thus enabling it to evade the immune system and have a high rate of chronic progression. High levels of HCV heterogeneity and its mutagenic capacity have made it difficult to create an effective vaccine. The recent advent of direct acting antivirals has ushered in a new era in hepatitis C therapy. Sustained virologic response is achieved with DAAs in 85-99% of cases. However, this still leads to a large population of treatment failures, so further advances in therapy are still needed. This article reviews the immunopathogenesis of HBV and HCV, their properties contributing to host immune system avoidance, chronic disease progression, vaccine efficacy and limitations, as well as treatment options and common pitfalls of said therapy.
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Affiliation(s)
- Corey Saraceni
- Correspondence to: Corey Saraceni, University of Connecticut School of Medicine, Department of Medicine, Division of Gastroenterology and Hepatology, 263 Farmington Avenue, Farmington, CT 06030-8074, USA. Tel: +1-203-733-7408, Fax: +1-860-679-3159, E-mail:
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Gomez-Perosanz M, Sanchez-Trincado JL, Fernandez-Arquero M, Sidney J, Sette A, Lafuente EM, Reche PA. Human rhinovirus-specific CD8 T cell responses target conserved and unusual epitopes. FASEB J 2020; 35:e21208. [PMID: 33230881 PMCID: PMC7753581 DOI: 10.1096/fj.202002165r] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/22/2020] [Revised: 10/30/2020] [Accepted: 11/04/2020] [Indexed: 12/14/2022]
Abstract
Human Rhinovirus (HRV) is a major cause of common cold, bronchiolitis, and exacerbations of chronic pulmonary diseases such as asthma. CD8 T cell responses likely play an important role in the control of HRV infection but, surprisingly, HRV‐specific CD8 T cell epitopes remain yet to be identified. Here, we approached the discovery and characterization of conserved HRV‐specific CD8 T cell epitopes from species A (HRV A) and C (HRV C), the most frequent subtypes in the clinics of various pulmonary diseases. We found IFNγ‐ELISPOT positive responses to 23 conserved HRV‐specific peptides on peripheral blood mononuclear cells (PBMCs) from 14 HLA I typed subjects. Peptide‐specific IFNγ production by CD8 T cells and binding to the relevant HLA I were confirmed for six HRV A‐specific and three HRV C‐specific CD8 T cell epitopes. In addition, we validated A*02:01‐restricted epitopes by DimerX staining and found out that these peptides mediated cytotoxicity. All these A*02:01‐restricted epitopes were 9‐mers but, interestingly, we also identified and validated an unusually long 16‐mer epitope peptide restricted by A*02:01, HRVC1791‐1806 (GLEPLDLNTSAGFPYV). HRV‐specific CD8 T cell epitopes describe here are expected to elicit CD8 T cell responses in up to 87% of the population and could be key for developing an HRV vaccine.
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Affiliation(s)
- Marta Gomez-Perosanz
- Department of Immunology, School of Medicine, Complutense University of Madrid, Madrid, Spain
| | - Jose L Sanchez-Trincado
- Department of Immunology, School of Medicine, Complutense University of Madrid, Madrid, Spain
| | | | - John Sidney
- Division of Vaccine Discovery, La Jolla Institute for Immunology, La Jolla, CA, USA
| | - Alessandro Sette
- Division of Vaccine Discovery, La Jolla Institute for Immunology, La Jolla, CA, USA
| | - Esther M Lafuente
- Department of Immunology, School of Medicine, Complutense University of Madrid, Madrid, Spain
| | - Pedro A Reche
- Department of Immunology, School of Medicine, Complutense University of Madrid, Madrid, Spain
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Antigenic cooperation among intrahost HCV variants organized into a complex network of cross-immunoreactivity. Proc Natl Acad Sci U S A 2015; 112:6653-8. [PMID: 25941392 DOI: 10.1073/pnas.1422942112] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/29/2023] Open
Abstract
Hepatitis C virus (HCV) has the propensity to cause chronic infection. Continuous immune escape has been proposed as a mechanism of intrahost viral evolution contributing to HCV persistence. Although the pronounced genetic diversity of intrahost HCV populations supports this hypothesis, recent observations of long-term persistence of individual HCV variants, negative selection increase, and complex dynamics of viral subpopulations during infection as well as broad cross-immunoreactivity (CR) among variants are inconsistent with the immune-escape hypothesis. Here, we present a mathematical model of intrahost viral population dynamics under the condition of a complex CR network (CRN) of viral variants and examine the contribution of CR to establishing persistent HCV infection. The model suggests a mechanism of viral adaptation by antigenic cooperation (AC), with immune responses against one variant protecting other variants. AC reduces the capacity of the host's immune system to neutralize certain viral variants. CRN structure determines specific roles for each viral variant in host adaptation, with variants eliciting broad-CR antibodies facilitating persistence of other variants immunoreacting with these antibodies. The proposed mechanism is supported by empirical observations of intrahost HCV evolution. Interference with AC is a potential strategy for interruption and prevention of chronic HCV infection.
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Campo DS, Dimitrova Z, Yamasaki L, Skums P, Lau DTY, Vaughan G, Forbi JC, Teo CG, Khudyakov Y. Next-generation sequencing reveals large connected networks of intra-host HCV variants. BMC Genomics 2014; 15 Suppl 5:S4. [PMID: 25081811 PMCID: PMC4120142 DOI: 10.1186/1471-2164-15-s5-s4] [Citation(s) in RCA: 43] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022] Open
Abstract
BACKGROUND Next-generation sequencing (NGS) allows for sampling numerous viral variants from infected patients. This provides a novel opportunity to represent and study the mutational landscape of Hepatitis C Virus (HCV) within a single host. RESULTS Intra-host variants of the HCV E1/E2 region were extensively sampled from 58 chronically infected patients. After NGS error correction, the average number of reads and variants obtained from each sample were 3202 and 464, respectively. The distance between each pair of variants was calculated and networks were created for each patient, where each node is a variant and two nodes are connected by a link if the nucleotide distance between them is 1. The work focused on large components having > 5% of all reads, which in average account for 93.7% of all reads found in a patient. CONCLUSIONS Most intra-host variants are organized into distinct single-mutation components that are: well separated from each other, represent genetic distances between viral variants, robust to sampling, reproducible and likely seeded during transmission events. Facilitated by NGS, large components offer a novel evolutionary framework for genetic analysis of intra-host viral populations and understanding transmission, immune escape and drug resistance.
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Affiliation(s)
- David S Campo
- Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
| | - Zoya Dimitrova
- Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
| | - Lilian Yamasaki
- Laboratory of Genomic Studies, Department of Biology, UNESP - São Paulo State University, Brazil
| | - Pavel Skums
- Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
| | - Daryl TY Lau
- Liver Center, Division of Gastroenterology, Beth Israel Deaconess Medical Center, Harvard Medical School, Massachusetts, USA
| | - Gilberto Vaughan
- Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
| | - Joseph C Forbi
- Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
| | - Chong-Gee Teo
- Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
| | - Yury Khudyakov
- Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
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Cusick MF, Libbey JE, Cox Gill J, Fujinami RS, Eckels DD. CD4 + T-cell engagement by both wild-type and variant HCV peptides modulates the conversion of viral clearing helper T cells to Tregs. Future Virol 2013; 8. [PMID: 24421862 DOI: 10.2217/fvl.13.49] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/11/2023]
Abstract
AIM To determine whether modulation of T-cell responses by naturally occurring viral variants caused an increase in numbers of Tregs in HCV-infected patients. PATIENTS MATERIALS & METHODS Human peripheral blood mononuclear cells, having proliferative responses to a wild-type HCV-specific CD4+ T-cell epitope, were used to quantify, via proliferative assays, flow cytometry and class II tetramers, the effects of naturally occurring viral variants arising in the immunodominant epitope. RESULTS In combination, the wild-type and variant peptides led to enhanced suppression of an anti-HCV T-cell response. The variant had a lower avidity for the wild-type-specific CD4+ T cell. Variant-stimulated CD4+ T cells had increased Foxp3, compared with wild-type-stimulated cells. CONCLUSION A stable viral variant from a chronic HCV subject was able to induce Tregs in multiple individuals that responded to the wild-type HCV-specific CD4+ T-cell epitope.
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Affiliation(s)
- Matthew F Cusick
- Department of Pathology, University of Utah School of Medicine, 30 North 1900 East, 3R330 SOM, Salt Lake City, UT 84132, USA
| | - Jane E Libbey
- Department of Pathology, University of Utah School of Medicine, 30 North 1900 East, 3R330 SOM, Salt Lake City, UT 84132, USA
| | - Joan Cox Gill
- Blood Research Institute, Blood Center of Wisconsin, Milwaukee, WI 53226, USA
| | - Robert S Fujinami
- Department of Pathology, University of Utah School of Medicine, 30 North 1900 East, 3R330 SOM, Salt Lake City, UT 84132, USA
| | - David D Eckels
- Department of Pathology, University of Utah School of Medicine, 30 North 1900 East, 3R330 SOM, Salt Lake City, UT 84132, USA
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Hepatitis C virus adaptation to T-cell immune pressure. ScientificWorldJournal 2013; 2013:673240. [PMID: 23554569 PMCID: PMC3608127 DOI: 10.1155/2013/673240] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/22/2013] [Accepted: 02/17/2013] [Indexed: 01/07/2023] Open
Abstract
Replication of the hepatitis C virus (HCV) is an error-prone process. This high error rate results in the emergence of viral populations (quasispecies) within hosts and contributes to interhost variability. Numerous studies have demonstrated that both viral and host factors contribute to this viral diversity, which can ultimately affect disease outcome. As the host's immune response is an important correlate of infection outcome for HCV, many of these viral variations are strongly influenced by T-cell immune pressure and accordingly constitute an efficient strategy to subvert such pressures (viral adaptations). This paper will review the data on viral diversity observed between and within hosts infected with HCV from the acute to the chronic stage of infection and will focus on viral adaptation to the host's T-cell immune response.
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Cusick MF, Libbey JE, Fujinami RS, Eckels DD. In vitro antigen-specific induction of IL-22 in human subjects that resolved HCV infection. Future Virol 2012. [PMID: 23185211 DOI: 10.2217/fvl.12.58] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022]
Abstract
AIMS: To determine if in vitro production of IL-22 and IL-17 correlated with resolution of HCV infection. MATERIALS #ENTITYSTARTX00026; METHODS: Human peripheral blood cells isolated from a well-defined cohort of resolved and chronic HCV-infected subjects were used to measure HCV-, influenza- and mitogen-activated T-cell proliferation. In addition, IL-22 and IL-17 production was measured via ELISAs and flow cytometry. RESULTS: Resolved HCV subjects had a significantly higher T-cell proliferative response to recombinant NS3 protein compared with chronic HCV subjects. Resolved subjects had a dose-dependent IL-22 response to recombinant NS3 compared with chronic HCV subjects. CONCLUSION: IL-22 production is associated with antigen-specific induction of CD4 (+) T cells in individuals that resolved HCV infection, suggesting a potential role for IL-22 in HCV clearance.
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Affiliation(s)
- Matthew F Cusick
- Department of Pathology, University of Utah School of Medicine, 30 North 1900 East, 3R330 SOM, Salt Lake City, UT 84132, USA
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Skums P, Dimitrova Z, Campo DS, Vaughan G, Rossi L, Forbi JC, Yokosawa J, Zelikovsky A, Khudyakov Y. Efficient error correction for next-generation sequencing of viral amplicons. BMC Bioinformatics 2012; 13 Suppl 10:S6. [PMID: 22759430 PMCID: PMC3382444 DOI: 10.1186/1471-2105-13-s10-s6] [Citation(s) in RCA: 79] [Impact Index Per Article: 6.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/05/2023] Open
Abstract
BACKGROUND Next-generation sequencing allows the analysis of an unprecedented number of viral sequence variants from infected patients, presenting a novel opportunity for understanding virus evolution, drug resistance and immune escape. However, sequencing in bulk is error prone. Thus, the generated data require error identification and correction. Most error-correction methods to date are not optimized for amplicon analysis and assume that the error rate is randomly distributed. Recent quality assessment of amplicon sequences obtained using 454-sequencing showed that the error rate is strongly linked to the presence and size of homopolymers, position in the sequence and length of the amplicon. All these parameters are strongly sequence specific and should be incorporated into the calibration of error-correction algorithms designed for amplicon sequencing. RESULTS In this paper, we present two new efficient error correction algorithms optimized for viral amplicons: (i) k-mer-based error correction (KEC) and (ii) empirical frequency threshold (ET). Both were compared to a previously published clustering algorithm (SHORAH), in order to evaluate their relative performance on 24 experimental datasets obtained by 454-sequencing of amplicons with known sequences. All three algorithms show similar accuracy in finding true haplotypes. However, KEC and ET were significantly more efficient than SHORAH in removing false haplotypes and estimating the frequency of true ones. CONCLUSIONS Both algorithms, KEC and ET, are highly suitable for rapid recovery of error-free haplotypes obtained by 454-sequencing of amplicons from heterogeneous viruses.The implementations of the algorithms and data sets used for their testing are available at: http://alan.cs.gsu.edu/NGS/?q=content/pyrosequencing-error-correction-algorithm.
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Affiliation(s)
- Pavel Skums
- Laboratory of Molecular Epidemiology and Bioinformatics, Division of Viral Hepatitis, Centers for Disease Control and Prevention, 1600 Clifton Road NE, 30333 Atlanta, GA, USA
| | - Zoya Dimitrova
- Laboratory of Molecular Epidemiology and Bioinformatics, Division of Viral Hepatitis, Centers for Disease Control and Prevention, 1600 Clifton Road NE, 30333 Atlanta, GA, USA
| | - David S Campo
- Laboratory of Molecular Epidemiology and Bioinformatics, Division of Viral Hepatitis, Centers for Disease Control and Prevention, 1600 Clifton Road NE, 30333 Atlanta, GA, USA
| | - Gilberto Vaughan
- Laboratory of Molecular Epidemiology and Bioinformatics, Division of Viral Hepatitis, Centers for Disease Control and Prevention, 1600 Clifton Road NE, 30333 Atlanta, GA, USA
| | - Livia Rossi
- Laboratory of Molecular Epidemiology and Bioinformatics, Division of Viral Hepatitis, Centers for Disease Control and Prevention, 1600 Clifton Road NE, 30333 Atlanta, GA, USA
| | - Joseph C Forbi
- Laboratory of Molecular Epidemiology and Bioinformatics, Division of Viral Hepatitis, Centers for Disease Control and Prevention, 1600 Clifton Road NE, 30333 Atlanta, GA, USA
| | - Jonny Yokosawa
- Laboratório de Virologia, Instituto de Ciências Biomédicas, Universidade Federal de Uberlândia, Uberlândia 38400-902, Brazil
| | - Alex Zelikovsky
- Department of Computer Science, Georgia State University, 34 Peachtree str., 30303, Atlanta, GA, USA
| | - Yury Khudyakov
- Laboratory of Molecular Epidemiology and Bioinformatics, Division of Viral Hepatitis, Centers for Disease Control and Prevention, 1600 Clifton Road NE, 30333 Atlanta, GA, USA
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Campo DS, Dimitrova Z, Yokosawa J, Hoang D, Perez NO, Ramachandran S, Khudyakov Y. Hepatitis C virus antigenic convergence. Sci Rep 2012; 2:267. [PMID: 22355779 PMCID: PMC3279735 DOI: 10.1038/srep00267] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/12/2012] [Accepted: 01/20/2012] [Indexed: 12/13/2022] Open
Abstract
Vaccine development against hepatitis C virus (HCV) is hindered by poor understanding of factors defining cross-immunoreactivity among heterogeneous epitopes. Using synthetic peptides and mouse immunization as a model, we conducted a quantitative analysis of cross-immunoreactivity among variants of the HCV hypervariable region 1 (HVR1). Analysis of 26,883 immunological reactions among pairs of peptides showed that the distribution of cross-immunoreactivity among HVR1 variants was skewed, with antibodies against a few variants reacting with all tested peptides. The HVR1 cross-immunoreactivity was accurately modeled based on amino acid sequence alone. The tested peptides were mapped in the HVR1 sequence space, which was visualized as a network of 11,319 sequences. The HVR1 variants with a greater network centrality showed a broader cross-immunoreactivity. The entire sequence space is explored by each HCV genotype and subtype. These findings indicate that HVR1 antigenic diversity is extensively convergent and effectively limited, suggesting significant implications for vaccine development.
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Affiliation(s)
- David S. Campo
- Molecular Epidemiology & Bioinformatics Laboratory, Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, USA, 30329
| | - Zoya Dimitrova
- Molecular Epidemiology & Bioinformatics Laboratory, Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, USA, 30329
| | - Jonny Yokosawa
- Molecular Epidemiology & Bioinformatics Laboratory, Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, USA, 30329
- Laboratório de Virologia, Instituto de Ciências Biomédicas, Universidade Federal de Uberlândia, Uberlândia, Brazil
| | - Duc Hoang
- Molecular Epidemiology & Bioinformatics Laboratory, Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, USA, 30329
- National Institute of Hygiene and Epidemiology, Hanoi, Vietnam
| | - Nestor O. Perez
- Molecular Epidemiology & Bioinformatics Laboratory, Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, USA, 30329
- Probiomed S.A., Tenancingo, Mexico
| | - Sumathi Ramachandran
- Molecular Epidemiology & Bioinformatics Laboratory, Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, USA, 30329
| | - Yury Khudyakov
- Molecular Epidemiology & Bioinformatics Laboratory, Division of Viral Hepatitis, Centers for Disease Control and Prevention, Atlanta, USA, 30329
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Human T cell expansion and experimental autoimmune encephalomyelitis inhibited by Lenaldekar, a small molecule discovered in a zebrafish screen. J Neuroimmunol 2012; 244:35-44. [PMID: 22245285 DOI: 10.1016/j.jneuroim.2011.12.024] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2011] [Revised: 12/15/2011] [Accepted: 12/16/2011] [Indexed: 11/22/2022]
Abstract
Immune-mediated diseases [multiple sclerosis (MS), experimental autoimmune encephalomyelitis (EAE)] are driven by proliferating, highly activated autoreactive T-cells that are unresponsive to in vivo immunoregulatory mechanisms. The compound Lenaldekar (LDK) was identified in a zebrafish screen by inhibiting T-cell expansion. By monitoring mitogen- and antigen-driven proliferation, we found that LDK inhibited human and murine T-cell expansion in a non-cytolytic manner. This suppressive activity directly correlated with the degree of activation/proliferation of the T-cells. In testing LDK in an EAE model of MS, exacerbations were suppressed in treated animals. Therefore, LDK represents a novel therapeutic approach to T-cell-mediated autoimmune diseases.
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Wang JH, Pianko MJ, Ke X, Herskovic A, Hershow R, Cotler SJ, Chen W, Chen ZW, Rong L. Characterization of antigenic variants of hepatitis C virus in immune evasion. Virol J 2011; 8:377. [PMID: 21801418 PMCID: PMC3158126 DOI: 10.1186/1743-422x-8-377] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/21/2011] [Accepted: 07/29/2011] [Indexed: 12/11/2022] Open
Abstract
BACKGROUND Antigenic variation is an effective way by which viruses evade host immune defense leading to viral persistence. Little is known about the inhibitory mechanisms of viral variants on CD4 T cell functions. RESULTS Using sythetic peptides of a HLA-DRB1*15-restricted CD4 epitope derived from the non-structural (NS) 3 protein of hepatitis C virus (HCV) and its antigenic variants and the peripheral blood mononuclear cells (PBMC) from six HLA-DRB1*15-positive patients chronically infected with HCV and 3 healthy subjects, the in vitro immune responses and the phenotypes of CD4+CD25+ cells of chronic HCV infection were investigated. The variants resulting from single or double amino acid substitutions at the center of the core region of the Th1 peptide not only induce failed T cell activation but also simultaneously up-regulate inhibitory IL-10, CD25-TGF-β+ Th3 and CD4+IL-10+ Tr1 cells. In contrast, other variants promote differentiation of CD25+TGF-β+ Th3 suppressors that attenuate T cell proliferation. CONCLUSIONS Naturally occuring HCV antigenic mutants of a CD4 epitope can shift a protective peripheral Th1 immune response into an inhibitory Th3 and/or Tr1 response. The modulation of antigenic variants on CD4 response is efficient and extensive, and is likely critical in viral persistence in HCV infection.
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Affiliation(s)
- Jane H Wang
- Section of Hepatology, Department of Medicine, University of Illinois at Chicago, Illinois, USA
- Division of Gastroenterology, Hepatology and Nutrition, Department of Pediatrics, the Feinberg School of Medicine, Northwestern University, Chicago, Illinois, USA
- Department of Microbiology and Immunology, University of Illinois at Chicago, Illinois, USA
| | - Matthew J Pianko
- Division of Gastroenterology, Hepatology and Nutrition, Department of Pediatrics, the Feinberg School of Medicine, Northwestern University, Chicago, Illinois, USA
| | - Xiaogang Ke
- Section of Hepatology, Department of Medicine, University of Illinois at Chicago, Illinois, USA
| | - Alex Herskovic
- Division of Gastroenterology, Hepatology and Nutrition, Department of Pediatrics, the Feinberg School of Medicine, Northwestern University, Chicago, Illinois, USA
| | - Ronald Hershow
- Division of Epidemiology and Biostatistics, School of Public Health, University of Illinois at Chicago, Illinois, USA
| | - Scott J Cotler
- Section of Hepatology, Department of Medicine, University of Illinois at Chicago, Illinois, USA
| | - Weijin Chen
- Changchun Institute of Biological Products, China National Biotec Group Int. Changchun, China
| | - Zheng W Chen
- Department of Microbiology and Immunology, University of Illinois at Chicago, Illinois, USA
| | - Lijun Rong
- Department of Microbiology and Immunology, University of Illinois at Chicago, Illinois, USA
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Abstract
It is generally believed that the role of CD4(+) T cells is to coordinate the different arms of the adaptive immune system to shape an effective response against a pathogen and regulate nonessential or deleterious activities. However, a growing body of evidence suggests that effector CD4(+) T cells can directly display potent antiviral activity themselves. The presence of cytolytic CD4(+) T cells has been demonstrated in the immune response to numerous viral infections in both humans and in animal models and it is likely that they play a critical role in the control of viral replication in vivo. This article describes the current research on virus-specific cytolytic CD4(+) T cells, with a focus on HIV-1 infection and the implications that this immune response has for vaccine design.
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Affiliation(s)
- Damien Z Soghoian
- Ragon Institute of MGH, MIT and Harvard Massachusetts General Hospital, Harvard Medical School Building 149, 13th Street, 5th floor, #5217, Charlestown, Boston, MA 02129, USA
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Hepatitis C virus induces regulatory T cells by naturally occurring viral variants to suppress T cell responses. Clin Dev Immunol 2010; 2011:806061. [PMID: 21197453 PMCID: PMC3004422 DOI: 10.1155/2011/806061] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/26/2010] [Accepted: 10/24/2010] [Indexed: 02/06/2023]
Abstract
Regulatory T cell markers are increased in chronically infected individuals with the hepatitis C virus (HCV), but to date, the induction and maintenance of Tregs in HCV infection has not been clearly defined. In this paper, we demonstrate that naturally occurring viral variants suppress T cell responses to cognate NS3358-375 in an antigen-specific manner. Of four archetypal variants, S370P induced regulatory T cell markers in comparison to NS3358-375-stimulated CD4 T cells. Further, the addition of variant-specific CD4 T cells back into a polyclonal culture in a dose-dependent manner inhibited the T cell response. These results suggest that HCV is able to induce antigen-specific regulatory T cells to suppress the antiviral T cell response in an antigen-specific manner, thus contributing to a niche within the host that could be conducive to HCV persistence.
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Mueller M, Spangenberg HC, Kersting N, Altay T, Blum HE, Klenerman P, Thimme R, Semmo N. Virus-specific CD4+ T cell responses in chronic HCV infection in blood and liver identified by antigen-specific upregulation of CD154. J Hepatol 2010; 52:800-11. [PMID: 20416967 DOI: 10.1016/j.jhep.2009.12.038] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/05/2009] [Revised: 11/30/2009] [Accepted: 12/19/2009] [Indexed: 01/27/2023]
Abstract
BACKGROUND & AIMS Virus-specific CD4+ T cells play a major role in hepatitis C virus (HCV) infection. Viral clearance is associated with vigorous and multispecific CD4+ T cell responses, while chronic infection has been shown to be associated with weak or absent T cell responses. Most of these studies, however, have used functional assays to analyse virus-specific CD4+ T cell responses. Therefore, the important question, of whether virus-specific CD4+ T cells are completely absent or primarily impaired in specific effector functions during chronic infection, has yet to be analysed in detail. METHODS To address this issue, a novel assay, where CD4+ T cell frequencies can be determined by de novo CD154 (CD40 ligand) expression in response to HCV antigens, was used in a cohort of chronically infected HCV patients and patients who spontaneously resolved HCV infection. These responses were compared to functional assays, such as the IFN-gamma ELISpot and flow cytometry-based proliferative assays. RESULTS Our results reveal that using the CD154 assay, virus-specific CD4+ T cells are readily detectable during chronic HCV infection albeit at a lower frequency when compared to patients who spontaneously resolved the infection. Importantly, no CD4+ T cell responses were detectable from these patients when using functional assays. Finally, these cell populations were enriched in the intrahepatic compartment. CONCLUSIONS Our findings suggest that HCV-specific CD4+ T cell responses are readily detectable in chronic HCV infection and enriched in the infected liver.
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Affiliation(s)
- Mareike Mueller
- Department of Medicine II, University Hospital Freiburg, Freiburg, Germany
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16
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Wang S, Buchli R, Schiller J, Gao J, VanGundy RS, Hildebrand WH, Eckels DD. Natural epitope variants of the hepatitis C virus impair cytotoxic T lymphocyte activity. World J Gastroenterol 2010; 16:1953-69. [PMID: 20419832 PMCID: PMC2860072 DOI: 10.3748/wjg.v16.i16.1953] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/07/2023] Open
Abstract
AIM: To understand how interactions between hepatitis C virus (HCV) and the host’s immune system might lead to viral persistence or effective elimination of HCV.
METHODS: Nucleotides 3519-3935 of the non-structural 3 (NS3) region were amplified by using reverse transcription polymerase chain reaction (PCR). PCR products of the HCV NS3 regions were integrated into a PCR® T7TOPO® TA vector and then sequenced in both directions using an automated DNA sequencer. Relative major histocompatibility complex binding levels of wild-type and variant peptides were performed by fluorescence polarization-based peptide competition assays. Peptides with wild type and variant sequences of NS3 were synthesized locally using F-moc chemistry and purified by high-performance liquid chromatography. Specific cytotoxic T lymphocytes (CTLs) clones toward HCV NS3 wild-type peptides were generated through limiting dilution cloning. The CTL clones specifically recognizing HCV NS3 wild-type peptides were tested by tetramer staining and flow cytometry. Cytolytic activity of CTL clones was measured using target cells labeled with the fluorescence enhancing ligand, DELFIA EuTDA.
RESULTS: The pattern of natural variants within three human leukocyte antigen (HLA)-A2-restricted NS3 epitopes has been examined in one patient with chronic HCV infection at 12, 28 and 63 mo post-infection. Results obtained may provide convincing evidence of immune selection pressure for all epitopes investigated. Statistical analysis of the extensive sequence variation found within these NS3 epitopes favors a Darwinian selection model of variant viruses. Mutations within the epitopes coincided with the decline of CTL responses, and peptide-binding studies suggested a significant impact of the mutation on T cell recognition rather than peptide presentation by HLA molecules. While most variants were either not recognized or elicited low responses, such could antagonize CTL responses to target cells pulsed with wild-type peptides.
CONCLUSION: Cross-recognition of CTL epitopes from wild-type and naturally-occurring HCV variants may lead to impaired immune responses and ultimately contribute to viral persistence.
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17
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Fleming VM, Harcourt G, Barnes E, Klenerman P. Virological footprint of CD4+ T-cell responses during chronic hepatitis C virus infection. J Gen Virol 2010; 91:1396-406. [PMID: 20107020 PMCID: PMC3052717 DOI: 10.1099/vir.0.017699-0] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/28/2022] Open
Abstract
Human and animal model evidence suggests that CD4+ T cells play a critical role in the control of chronic hepatitis C virus (HCV) infection. However, despite their importance, the mechanism behind the failure of such populations in chronic disease is not understood and the contribution of viral mutation is not known. To address this, this study defined the specificity and virological footprint of CD4+ T cells in chronic infection. CD8+ T-cell-depleted peripheral blood mononuclear cells from 61 HCV genotype 1-infected patients were analysed against a panel of peptides covering the HCV genotype 1 core – a region where CD4+ T-cell responses may be reproducibly obtained. In parallel, the core region and E2 protein were sequenced. Gamma interferon-secreting CD4+ T-cell responses directed against multiple epitopes were detected in 53 % of individuals, targeting between one and four peptides in the HCV core. Viral sequence evaluation revealed that these CD4+ T-cell responses were associated with mutants in 2/21 individuals. In these two cases, the circulating sequence variant was poorly recognized by host CD4+ T cells. Bioinformatics analyses revealed no overall evidence of selection in the target epitopes and no differences between the groups with and without detectable CD4+ T-cell responses. It was concluded that sustained core peptide-specific CD4+ T-cell responses may be reproducibly measured during chronic HCV infection and that immune escape may occur in specific instances. However, overall the virological impact of such responses is limited and other causes for CD4+ T-cell failure in HCV must be sought.
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Affiliation(s)
- Vicki M Fleming
- Peter Medawar Building for Pathogen Research, Nuffield Department of Clinical Medicine, University of Oxford, South Parks Road, Oxford, UK.
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18
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Kumar D, Malik A, Asim M, Chakravarti A, Das RH, Kar P. Influence of quasispecies on virological responses and disease severity in patients with chronic hepatitis C. World J Gastroenterol 2008; 14:701-8. [PMID: 18205258 PMCID: PMC2683995 DOI: 10.3748/wjg.14.701] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To elucidate the influence of quasispecies on virological response and disease severity in patients with chronic hepatitis C.
METHODS: Forty seven patients with hepatitis C [32 with chronic active hepatitis (CAH), 9 with cirrhosis, and 6 with hepatocellular carcinoma (HCC)] were screened for the presence of quasispecies by single stranded conformational polymorphism (SSCP) analysis in the hypervariable region (HVR) and non-structural 5B (NS5B) viral genes of hepatitis C virus. The 41 patients excluding those with HCC were on therapy and followed up for a year with the determination of virological response and disease severity. Virus isolated from twenty three randomly selected patients (11 non-responders and 12 showing a sustained virological response) was sequenced for the assessment of mutations.
RESULTS: The occurrence of quasispecies was proportionately higher in patients with HCC and cirrhosis than in those with CAH, revealing a significant correlation between the molecular evolution of quasispecies and the severity of disease in patients with hepatitis C. The occurrence of complex quasispecies has a significant association (P < 0.05) with the non-responders, and leads to persistence of infection. Significant differences (P < 0.05) in viral load (log10 IU/mL) were observed among patients infected with complex quasispecies (CQS), those infected with simple quasispecies (SQS) and those with no quasispecies (NQS), after 12 wk (CQS-5.2 ± 2.3, SQS-3.2 ± 1.9, NQS-2.8 ± 2.4) and 24 wk (CQS-3.9 ± 2.2, SQS-3.0 ± 2.2, NQS-2.1 ± 2.3) in the HVR region. However, a statistically significant difference (P < 0.05) was observed between the viral loads of patients infected with CQS and those infected with NQS in NS5B viral gene after 24 wk (CQS-3.9 ± 2.2, SQS-3.0 ± 2.2, and NQS-2.1 ± 2.3) and 48 wk (CQS-3.1 ± 2.7, SQS-2.3 ± 2.4, NQS-2.0 ± 2.3) of therapy. Disease severity was significantly associated with viral load during therapy. The strains isolated from non-responders showed close pairing on phylogeny based on the NS5B gene, but dissimilar HVR regions. This revealed the possibility of the selection of resistant strains during the evolution of quasispecies in NS5B.
CONCLUSION: Viral quasispecies may be an important predictor of virological responses to combination therapy in patients with chronic hepatitis C. Complex quasispecies and resistant strains may lead to high viral loads during therapy, with a concerted effect on disease severity.
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19
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Tarr AW, Owsianka AM, Jayaraj D, Brown RJP, Hickling TP, Irving WL, Patel AH, Ball JK. Determination of the human antibody response to the epitope defined by the hepatitis C virus-neutralizing monoclonal antibody AP33. J Gen Virol 2007; 88:2991-3001. [PMID: 17947521 DOI: 10.1099/vir.0.83065-0] [Citation(s) in RCA: 57] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022] Open
Abstract
Hepatitis C virus (HCV) is a major cause of liver disease worldwide and there is a pressing need for the development of a preventative vaccine as well as new treatments. It was recently demonstrated that the mouse monoclonal antibody (mAb) AP33 potently neutralizes infectivity of HCV pseudoparticles (HCVpp) carrying E1E2 envelopes representative of all of the major genotypes of HCV. This study determined the prevalence of human serum antibodies reactive to the region of HCV E2 recognized by AP33. Antibodies recognizing this region were present in less than 2.5 % of sera obtained from individuals with chronic HCV infection. A similar prevalence was found in a smaller cohort of individuals who had experienced an acute infection, suggesting that AP33-like antibodies do not play a major role in natural clearance of HCV infection. Sera exhibited different patterns of reactivity to a panel of peptides representing circulating variants, highlighting the presence of distinct epitopes in this region. Only two sera contained antibodies that could recognize a specific AP33-reactive peptide mimotope. AP33-like antibodies made a measurable contribution to the ability of these sera to inhibit E2-CD81 interaction, but not to the overall neutralization of cell entry. Together, these data show that antibodies to the AP33 epitope are not commonly generated during natural infection and that generation of such antibodies via vaccination may require modified immunogens to focus the generation of specific antibodies. Importantly, individuals harbouring AP33-like antibodies are an important potential source of human mAbs for future therapeutic development.
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Affiliation(s)
- Alexander W Tarr
- The Institute of Infection, Immunity and Inflammation and Division of Microbiology, The University of Nottingham, Queen's Medical Centre, Nottingham NG7 2UH, UK
| | - Ania M Owsianka
- MRC Virology Unit, Institute of Virology, University of Glasgow, Church Street, Glasgow G11 5JR, UK
| | - Dhanya Jayaraj
- The Institute of Infection, Immunity and Inflammation and Division of Microbiology, The University of Nottingham, Queen's Medical Centre, Nottingham NG7 2UH, UK
| | - Richard J P Brown
- The Institute of Infection, Immunity and Inflammation and Division of Microbiology, The University of Nottingham, Queen's Medical Centre, Nottingham NG7 2UH, UK
| | - Timothy P Hickling
- The Institute of Infection, Immunity and Inflammation and Division of Microbiology, The University of Nottingham, Queen's Medical Centre, Nottingham NG7 2UH, UK
| | - William L Irving
- The Institute of Infection, Immunity and Inflammation and Division of Microbiology, The University of Nottingham, Queen's Medical Centre, Nottingham NG7 2UH, UK
| | - Arvind H Patel
- MRC Virology Unit, Institute of Virology, University of Glasgow, Church Street, Glasgow G11 5JR, UK
| | - Jonathan K Ball
- The Institute of Infection, Immunity and Inflammation and Division of Microbiology, The University of Nottingham, Queen's Medical Centre, Nottingham NG7 2UH, UK
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20
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Timm J, Roggendorf M. Sequence diversity of hepatitis C virus: implications for immune control and therapy. World J Gastroenterol 2007; 13:4808-17. [PMID: 17828811 PMCID: PMC4611758 DOI: 10.3748/wjg.v13.i36.4808] [Citation(s) in RCA: 67] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/26/2007] [Revised: 07/02/2007] [Accepted: 07/09/2007] [Indexed: 02/06/2023] Open
Abstract
With approximately 3% of the world's population (170 million people) infected with the hepatitis C virus (HCV), the WHO has declared HCV a global health problem. Upon acute infection about 50%-80% of subjects develop chronic hepatitis with viral persistence being at risk to develop liver cirrhosis and hepatocellular carcinoma. One characteristic of HCV is its enormous sequence diversity, which represents a significant hurdle to the development of both effective vaccines as well as to novel therapeutic interventions. Due to a polymerase that lacks a proofreading function HCV presents with a high rate of evolution, which enables rapid adaptation to a new environment including an activated immune system upon acute infection. Similarly, novel drugs designed to specifically inhibit viral proteins will face the potential problem of rapid selection of drug resistance mutations. This review focuses on the sequence diversity of HCV, the driving forces of evolution and the impact on immune control and treatment response. An important feature of any therapeutic or prophylactic intervention will be an efficient attack of a structurally or functionally important region in the viral protein. The understanding of the driving forces, but also the limits of viral evolution, will be fundamental for the design of novel therapies.
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Affiliation(s)
- Joerg Timm
- University of Essen, Institute of Virology, Hufelandst 55, Essen 45122, Germany.
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21
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Abstract
Hepatitis C virus (HCV) infection is a major cause of liver damage, with virus-induced end-stage disease such as liver cirrhosis and hepatocellular carcinoma resulting in a high rate of morbidity and mortality worldwide. Evidence that CD4+ T cell responses to HCV play an important role in the outcome of acute infection has been shown in several studies. However, the mechanisms behind viral persistence and the failure of CD4+ T cell responses to contain virus are poorly understood. During chronic HCV infection, HCV-specific CD4+ T cell responses are relatively weak or absent whereas in resolved infection these responses are vigorous and multispecific. Persons with a T-helper type I profile, which promotes cellular effector mechanisms are thought to be more likely to experience viral clearance, but the overall role of these cells in the immunopathogenesis of chronic liver disease is not known. To define this, much more data is required on the function and specificity of virus-specific CD4+ T cells, especially in the early phases of acute disease and in the liver during chronic infection. The role and possible mechanisms of action of CD4+ T cell responses in determining the outcome of acute and chronic HCV infection will be discussed in this review.
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Affiliation(s)
- Nasser Semmo
- Nuffield Department of Clinical Medicine, University of Oxford, The Peter Medawar Building for Pathogen Research, South Parks Road, Oxford OX1 3SY, United Kingdom
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22
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Perrella A, Atripaldi L, Bellopede P, Patarino T, Sbreglia C, Tarantino G, Sorrentino P, Conca P, Ruggiero L, Perrella O. Flow cytometry assay of myeloid dendritic cells (mDCs) in peripheral blood during acute hepatitis C: possible pathogenetic mechanisms. World J Gastroenterol 2006; 12:1105-1109. [PMID: 16534853 PMCID: PMC4087904 DOI: 10.3748/wjg.v12.i7.1105] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/01/2005] [Revised: 10/06/2005] [Accepted: 10/26/2005] [Indexed: 02/06/2023] Open
Abstract
AIM To asses the expression of myeloid dendritic cells (CD11c+) subset during acute HCV hepatitis and its possible involvement in natural history of the infection. METHODS We enrolled 11 patients with acute hepatitis C (AHC) (Group A), 10 patients with acute hepatitis A (AHA) (as infective control-Group B) and 10 healthy donors (group C) in this study. All patients underwent selective flow cytometry gating strategies to assess the peripheral number of the myeloid dendritic cells (mDCs) to understand the possible role and differences during acute hepatitis. RESULTS Eight of 11 patients with acute HCV hepatitis did not show any increase of mDCs compared to healthy individuals, while a significant decrease of mDCs was found in absolute cell count (z = -2.37; P<0.05) and percentage (z = -2.30; P<0.05) as compared with AHA. On the contrary, The remaining three patients of the group A had a higher mDCs number and percentage as occur in group B. Interestingly, after six months, those patients did not show any increase of mDCs subset were chronically infected. while the three subjects with an increase of peripheral mDCs, as in HAV acute infection, resolved the illness. CONCLUSION The lack of increase of mDCs during acute hepatitis C might be an important factor involved in chronicization of the infection.
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Affiliation(s)
- Alessandro Perrella
- Department Infectious Disease and Immunology, Hospital D.Cotugno, Via E.A. Mario 35 , Zip code 80131, Naples, Italy.
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23
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Gerlach JT, Ulsenheimer A, Grüner NH, Jung MC, Schraut W, Schirren CA, Heeg M, Scholz S, Witter K, Zahn R, Vogler A, Zachoval R, Pape GR, Diepolder HM. Minimal T-cell-stimulatory sequences and spectrum of HLA restriction of immunodominant CD4+ T-cell epitopes within hepatitis C virus NS3 and NS4 proteins. J Virol 2005; 79:12425-33. [PMID: 16160170 PMCID: PMC1211510 DOI: 10.1128/jvi.79.19.12425-12433.2005] [Citation(s) in RCA: 52] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/30/2023] Open
Abstract
The hepatitis C virus (HCV)-specific CD4+ T-cell response against nonstructural proteins is strongly associated with successful viral clearance during acute hepatitis C. To further develop these observations into peptide-based vaccines and clinical immunomonitoring tools like HLA class II tetramers, a detailed characterization of immunodominant CD4+ T-cell epitopes is required. We studied peripheral blood mononuclear cells from 20 patients with acute hepatitis C using 83 overlapping 20-mer peptides covering the NS3 helicase and NS4. Eight peptides were recognized by > or = 40% of patients, and specific CD4+ T-cell clones were obtained for seven of these and three additional, subdominant epitopes. Mapping of minimal stimulatory sequences defined epitopes of 8 to 13 amino acids in length, but optimal T-cell stimulation was observed with 10- to 15-mers. While some epitopes were presented by different HLA molecules, others were presented by only a single HLA class II molecule, which has implications for patient selection in clinical trials of peptide-based immunotherapies. In conclusion, using two different approaches we identified and characterized a set of CD4+ T-cell epitopes in the HCV NS3-NS4 region which are immunodominant in patients achieving transient or persistent viral control. This information allows the construction of a valuable panel of HCV-specific HLA class II tetramers for further study of CD4+ T-cell responses in chronic hepatitis C. The finding of immunodominant epitopes with very constrained HLA restriction has implications for patient selection in clinical trials of peptide-based immunotherapies.
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Affiliation(s)
- J T Gerlach
- Department of Gastroenterology and Hepatology, University Hospital Zurich, Rämistrasse 100, 8091 Zurich, Switzerland.
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24
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Affiliation(s)
- Stefan F Wieland
- Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, CA 92037, USA
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25
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Zhu F, Yang M, Eckels DD. Interactions between helper T-cell epitopes of hepatitis C virus. Vaccine 2005; 23:3572-80. [PMID: 15855016 DOI: 10.1016/j.vaccine.2005.01.157] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/04/2004] [Revised: 01/24/2005] [Accepted: 01/24/2005] [Indexed: 12/09/2022]
Abstract
The premise of this work is that within a given hepatitis C virus (HCV) protein there exists an array of Th1 and Th2 epitopes, each of which can provide synergistic (positive or negative) effects upon other epitopes by intramolecular, cytokine-mediated immunoregulation of helper T-cell responses. To address this question, we constructed minigene plasmids pHCVTh1, pHCVTh1X3 and pHCVThR, and HCV NS3 full-length plasmid pHCVNS3. 293T cells were transfected with these plasmids and cell lysates from the transfected cells were used to stimulate PBMC from a patient with chronic HCV infection. IL-2 and IFN-gamma in the supernatant of the cultured PBMC were tested and proliferation of the PBMC was measured. The results demonstrate that interactions exist among helper T-cell epitopes; the synergistic effects of suppressive Th2 epitopes upon Th1 epitopes will inhibit the responses induced by Th1 epitopes, which may contribute to chronic infection by HCV; synergistic effects among Th1 epitopes induce higher levels of IFN-gamma, which may suggest a new strategy for HCV vaccine development. Further, stimulation of an HCV NS3 specific clone with cell lysates from 293T cells transfected with different constructs shows that the HCV NS3 clone could respond to all suggesting that the epitope-specific suppression may be due to an imbalance of Type 1 and Type 2 cytokines or regulatory T-cells.
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Affiliation(s)
- Fenlu Zhu
- Department of Pediatrics, Medical College of Wisconsin, 8701 Watertown Plank Road, P.O. Box 26509, Milwaukee, WI 53226-0509, USA
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26
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Giuggio VM, Bonkovsky HL, Rothman AL. Evolution of the Intrahepatic T Cell Repertoire during Chronic Hepatitis C Virus Infection. Viral Immunol 2005; 18:179-89. [PMID: 15802962 DOI: 10.1089/vim.2005.18.179] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022] Open
Abstract
Chronic hepatitis C virus (HCV) infection is characterized by extensive infiltration of inflammatory cells in the liver, where there is a compartmentalization of HCV-reactive T lymphocytes. Previous studies have demonstrated a broad intrahepatic TCR repertoire; however, there is little information regarding the stability of this intrahepatic T cell population. We studied the T cell repertoires in sequential liver biopsy samples from five individuals with chronic HCV infection using TCR spectratype analysis; four subjects had been treated with IFN-alpha during the interval between biopsies. Transcripts from most TCRBV families were detectable in the liver tissues, and 25-85% of these had skewed spectratype profiles indicative of T cell clonal expansions. Most of the intrahepatic T cell expansions were not evident in an analysis of peripheral blood T cells collected at the same time as the liver biopsy. Detailed analysis using TCRBJ-primed run-off reactions revealed that the intrahepatic TCR repertoires were not stable within an individual, although some TCR clonotypes were maintained for at least 45 months.
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Affiliation(s)
- Vicki M Giuggio
- Center for Infectious Disease and Vaccine Research, University of Massachusetts Medical School, 55 Lake Ave. North, Worcester, MA 01655, USA
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27
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Affiliation(s)
- David G Bowen
- Center for Vaccines and Immunity, Columbus Children's Research Institute, 700 Childrens Dr, Columbus, OH 43205, USA
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28
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Perrella A, Borgia G, Perrella O. Immune system homeostasis during acute hepatitis C: viral escape or T-cell regulation? Hepatology 2004; 40:1229-1230. [PMID: 15486981 DOI: 10.1002/hep.20470] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/05/2023]
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29
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Abstract
The hepatitis C virus (HCV) infects approximately three percent of the world's population. Some individuals resolve the infection spontaneously, but the majority develop persistent viremia that often causes progressive liver disease. There is an emerging consensus that cellular immune responses are essential for spontaneous resolution of acute hepatitis C and long-term protection from persistent infection. This review focuses on the recent advances in understanding mechanisms of protective immunity and why they fail in most infected individuals. The distinct yet complementary role of CD4+ and CD8+ T lymphocytes in this process is highlighted.
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Affiliation(s)
- Naglaa H Shoukry
- Center for Vaccines and Immunity, Columbus Children's Research Institute, Columbus, Ohio 43205, USA.
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30
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Wang JH, Layden TJ, Eckels DD. Modulation of the peripheral T-Cell response by CD4 mutants of hepatitis C virus: transition from a Th1 to a Th2 response. Hum Immunol 2003; 64:662-73. [PMID: 12826368 DOI: 10.1016/s0198-8859(03)00070-3] [Citation(s) in RCA: 43] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
A disturbing feature of hepatitis C virus (HCV) is its long-term persistence in roughly 85% of those infected. Escape mutants may play a major role in HCV persistence. Our previous studies have identified a human leukocyte antigen DRB1*15 (HLA-DRB1*15) restricted Th1 epitope in the HCV NS3 protein, NS3(358-375), and escape variants of this epitope that may emerge under immune selection. Such variants attenuate or fail to stimulate T-cell proliferation. Here we provide data from peripheral blood mononuclear cells derived from four HLA-DRB1*15 patients chronically infected with HCV, and report that naturally occurring single amino acid substitutions in the Th1 epitope NS3(358-375) fail to stimulate proliferation, which is accompanied by a shift in cytokine secretion patterns from one characteristic of a Th1 antiviral responses to a Th2 form. Further, in one patient, we demonstrate that HCV variant peptides can effectively inhibit host polyclonal peripheral T-cell proliferation. We speculate that this phenomenon may be a factor in chronic HCV infection.
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Affiliation(s)
- Jane H Wang
- Section of Liver Diseases, University of Illinois at Chicago, Chicago, IL, USA
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31
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Abstract
After a virus infects an animal, antiviral responses are generated that attempt to prevent dissemination. Interferons, antibody, complement, T and natural killer cells all contribute to the control and eradication of viral infections. Most flaviviruses, with the exception of some of the encephalitic viruses, cause acute disease and do not establish persistent infection. The outcome of flavivirus infection in an animal is determined by a balance between the speed of viral replication and spread, and the immune system response. Although many of the mechanistic details require further elucidation, flaviviruses have evolved specific tactics to evade the innate and adaptive immune response. A more thorough understanding of these principles could lead to improved models for viral pathogenesis and to strategies for the development of novel antiviral agents.
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Affiliation(s)
- Michael S Diamond
- Department of Medicine, Washington University School of Medicine, Saint Louis, Missouri 63110, United States of America.
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Sreenarasimhaiah J, Jaramillo A, Crippin J, Lisker-Melman M, Chapman WC, Mohanakumar T. Lack of optimal T-cell reactivity against the hepatitis C virus is associated with the development of fibrosis/cirrhosis during chronic hepatitis. Hum Immunol 2003; 64:224-30. [PMID: 12559624 DOI: 10.1016/s0198-8859(02)00781-4] [Citation(s) in RCA: 20] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
Chronic hepatitis C virus (HCV) infection develops in 85% of exposed individuals and 20% develop cirrhosis. However, the pathogenesis of this process is not well-understood. The objective of this study was to determine whether HCV-reactive T cells play a role in the process of development of cirrhosis during chronic HCV infection. We analyzed 21 human leukocyte antigen (HLA)-A2 patients with chronic HCV infection (9 with histology of inflammation and 12 with histology of fibrosis/cirrhosis). The frequency of CD8(+) T cells reactive to 12 HCV-derived epitopes was determined by an interferon-gamma enzyme-linked immunospot (ELISPOT) assay. The frequency of CD4(+) Th1 and Th2 cells reactive to the HCV core antigen was determined by interferon-gamma and interleukin-5 ELISPOT assays, respectively. Patients with histology of inflammation showed a significantly higher CD8(+) T-cell response to five HCV-derived epitopes (YLLPRRGPRL [core], CINGVCWTV [NS3], LLCPAGHAV [NS3], ILAGYGAGV [NS4B], and GLQDCTMLV [NS5B]) as compared with patients with histology of fibrosis/cirrhosis. Also, patients with histology of inflammation showed a significantly higher CD4(+) Th1 response to the HCV core antigen as compared to patients with histology of fibrosis/cirrhosis. These results indicate that a lack of an optimal T-cell response to HCV is associated with the development of cirrhosis during chronic HCV infection.
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McNeil LK, Evavold BD. TCR reserve: a novel principle of CD4 T cell activation by weak ligands. JOURNAL OF IMMUNOLOGY (BALTIMORE, MD. : 1950) 2003; 170:1224-30. [PMID: 12538680 DOI: 10.4049/jimmunol.170.3.1224] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Abstract
Some ligand-receptor systems have a receptor reserve where a maximal response can be achieved by occupation of a fraction of available receptors. An implication of a receptor reserve is the expansion of the number of ligands for response. To determine whether T cells follow receptor reserve, we have characterized the effect of reducing TCR levels on CD4 T cell responses elicited by altered peptide ligands that vary in potency. Agonist peptide is unaffected by a 90% reduction in TCR level while proliferation to weak agonists is significantly inhibited when TCR expression is reduced by 40%. Thymocyte-negative selection similarly demonstrates a differential requirement of TCR for response to agonist, weak agonist, and partial agonist. Therefore, our data demonstrate receptor reserve as a novel principle of T cell activation in which excess TCRs expand the antigenic repertoire to include less potent ligands.
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MESH Headings
- Amino Acid Sequence
- Animals
- Antibodies, Blocking/pharmacology
- Antigens, CD/biosynthesis
- Antigens, Differentiation, T-Lymphocyte/biosynthesis
- CD4-Positive T-Lymphocytes/cytology
- CD4-Positive T-Lymphocytes/immunology
- CD4-Positive T-Lymphocytes/metabolism
- Cytochrome c Group/immunology
- Cytochrome c Group/pharmacology
- Dose-Response Relationship, Immunologic
- Down-Regulation/genetics
- Down-Regulation/immunology
- Growth Inhibitors/antagonists & inhibitors
- Growth Inhibitors/biosynthesis
- Growth Inhibitors/genetics
- Growth Inhibitors/immunology
- Immunoglobulin Fab Fragments/pharmacology
- Lectins, C-Type
- Ligands
- Lymphocyte Activation/genetics
- Mice
- Mice, Inbred A
- Mice, Inbred C57BL
- Mice, Transgenic
- Molecular Sequence Data
- Moths/enzymology
- Receptors, Antigen, T-Cell, alpha-beta/antagonists & inhibitors
- Receptors, Antigen, T-Cell, alpha-beta/biosynthesis
- Receptors, Antigen, T-Cell, alpha-beta/genetics
- Receptors, Antigen, T-Cell, alpha-beta/immunology
- T-Lymphocyte Subsets/cytology
- T-Lymphocyte Subsets/immunology
- T-Lymphocyte Subsets/metabolism
- Thymus Gland/cytology
- Thymus Gland/immunology
- Up-Regulation/immunology
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Affiliation(s)
- Lisa K McNeil
- Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, GA 30322, USA
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Fowler NL, Torresi J, Jackson DC, Brown LE, Gowans EJ. Immune responses in hepatitis C virus infection: the role of dendritic cells. Immunol Cell Biol 2003; 81:63-6. [PMID: 12534948 DOI: 10.1046/j.0818-9641.2002.01138.x] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023]
Abstract
Cellular immune responses are critical for the clearance of hepatitis C virus. Persistent infection results from a narrow and weak cellular immune response, in direct contrast to the broad, strong response associated with viral clearance in acute infection. The presence of dendritic cells in the liver facilitates presentation of viral antigens to both CD4+ and CD8+ T cell populations. Exploiting the potent antigen presentation capability of dendritic cells for immunotherapy of chronic hepatitis C is attractive; however, infection or transfection of segments or the entire hepatitis C virus genome appears to impair the allostimulation capacity of dendritic cells. If dendritic cell immunotherapy for hepatitis C virus infection is to become a reality, the mechanism behind the defective allostimulatory capacity needs to be deciphered.
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Affiliation(s)
- Nina L Fowler
- Macfarlane Burnet Institute for Medical Research and Public Health, Prahran, Victoria, Australia.
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Abstract
The chimpanzee (Pan troglodytes) is the only experimental animal susceptible to infection with hepatitis C virus (HCV). The chimpanzee model of HCV infection was instrumental in the initial studies on non-A, non-B hepatitis, including observations on the clinical course of infection, determination of the physical properties of the virus, and eventual cloning of the HCV nucleic acid. This review focuses on more recent aspects of the use of the chimpanzee in HCV research. The chimpanzee model has been critical for the analysis of early events in HCV infection because it represents a population for which samples are available from the time of exposure and all exposed animals are examined. For this reason, the chimpanzee represents a truly nonselected population. In contrast, human cohorts are often selected for disease status or antibody reactivity and typically include individuals that have been infected for decades. The chimpanzee model is essential to an improved understanding of the factors involved in viral clearance, analysis of the immune response to infection, and the development of vaccines. The development of infectious cDNA clones of HCV was dependent on the use of chimpanzees, and they will continue to be needed in the use of reverse genetics to evaluate critical sequences for viral replication. In addition, chimpanzees have been used in conjunction with DNA microarray technology to probe the entire spectrum of changes in liver gene expression during the course of HCV infection. The chimpanzee will continue to provide a critical aspect to the understanding of HCV disease and the development of therapeutic modalities.
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Affiliation(s)
- R E Lanford
- Department of Virology and Immunology, Southwest Regional Primate Research Center, Southwest Foundation for Biomedical Research, San Antonio, Texas, USA
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Abstract
Hepatitis C virus (HCV) is known for its ability to establish persistent infection and cause chronic hepatitis in most infected individuals. The pathogenesis of hepatic injury and the precise mechanisms underlying viral persistence are unknown. Accumulating evidence indicates that successful elimination of HCV is associated with the induction and maintaining of strong helper T-cell and cytotoxic T-cell responses against multiple viral epitopes. In contrast, patients who develop chronic HCV infection are characterized by the lack of strong viral-specific helper T-cell responses. The failure to mount and maintain strong HCV-specific T-cell responses may be determined by the genetics, especially the major histocompatibility complex background, of the host. However, it is likely that other host and viral factors are also involved in determining the outcome of HCV infection. Available data suggest that HCV is not cytopathic to hepatocytes and that liver injury associated with chronic HCV infection is likely to be mediated by immune responses against HCV-infected hepatocytes. In addition to hepatitis, HCV infection may also cause breaching of immune tolerance, leading to autoimmune disorders. Although the lack of a small animal model and a tissue culture system has impeded research on hepatitis C virus (HCV) infection, recent studies in humans and chimpanzees have significantly enhanced our understanding of the interaction between HCV and the host's immune system. This review focuses on the most recent advances in our understanding of the immunology of HCV infection. In particular, the possible mechanisms of how HCV establishes chronic infection are discussed. The pathogenesis of liver injury, the immunogenetics of HCV infection, and the effect of HCV infection on host's immune function are also reviewed.
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Affiliation(s)
- L Huang
- Division of Gastroenterology, Brigham and Women's Hospital, Beth Israel Deaconess Hospital, Boston, Massachusetts 02115, USA
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Brander C, Walker BD. Modulation of host immune responses by clinically relevant human DNA and RNA viruses. Curr Opin Microbiol 2000; 3:379-86. [PMID: 10972498 DOI: 10.1016/s1369-5274(00)00108-9] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
Numerous mechanisms allow viruses to evade host immune surveillance, and new evasion strategies continue to be identified. In addition to interference with antigen processing and presentation, direct viral modulation of host immune responses can also be achieved by altering the host cytokine milieu and the development of immunoregulatory cells. A better understanding of these viral evasion strategies will help to define critical host defense mechanisms and will lead to novel immune-based therapeutic strategies in the future.
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Affiliation(s)
- C Brander
- Partners AIDS Research Center, Massachusetts General Hospital and Harvard Medical School, Charlestown 02129, USA
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Pelletier SJ, Raymond DP, Crabtree TD, Iezzoni JC, Sawyer RG, Hahn YS, Pruett TL. Pretransplantation hepatitis C virus quasispecies may be predictive of outcome after liver transplantation. Hepatology 2000; 32:375-81. [PMID: 10915745 DOI: 10.1053/jhep.2000.9089] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/23/2022]
Abstract
The evolution of hepatitis C virus (HCV) envelope variation was studied using a liver-transplant model to evaluate the role of HCV quasispecies for hepatocyte infection. Twelve HCV polymerase chain reaction (PCR)-positive liver-transplant recipients (6 with posttransplantation biochemical hepatitis and 6 without hepatitis [controls]) were prospectively evaluated and underwent detailed quasispecies analysis of pre- and postoperative serum samples. HCV amino acid sequence diversity and complexity at the first hypervariable region (HVR1) of the second envelope protein (E2) was correlated with outcome. Recurrence of HCV-induced allograft injury was defined by persistently elevated serum alanine transaminase (ALT) levels. The major variant (sequences >10% of all clones) of recipients with hepatitis accounted for a significantly smaller percent of all preoperative clones compared with controls (41% +/- 6% vs. 69% +/- 8%; P <.015). Recipients with hepatitis had an increased number of pretransplantation major variants (2.5 +/- 0.3 vs. 1.1 +/- 0.2; P <.006). Eighty-three percent of controls had a predominant variant (accounting for >50% of clones) compared with 17% of those with recurrence (P <.05). These differences did not persist postoperatively. In addition, recipients without a pretransplantation predominant variant demonstrated an increased allograft fibrosis score (2.3 +/- 0.3 vs. 0.5 +/- 0.3; P <.015) at 181 to 360 days posttransplantation compared with those in whom a predominant variant was present. Increased HCV envelope complexity may act as a stronger antigenic stimulus or improve hepatocyte receptor binding and lead to allograft hepatitis and fibrosis. Although pretransplantation differences in HCV quasispecies did not persist postoperatively, pretransplantation quasispecies may be a predictor of HCV-induced hepatitis and graft fibrosis after liver transplantation.
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Affiliation(s)
- S J Pelletier
- Charles O. Strickler Transplant Center, University of Virginia Health Sciences Center, Charlottesville, VA, USA.
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