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Devi P, Punga T, Bergqvist A. Activation of the Ca2+/NFAT Pathway by Assembly of Hepatitis C Virus Core Protein into Nucleocapsid-like Particles. Viruses 2022; 14:v14040761. [PMID: 35458491 PMCID: PMC9031069 DOI: 10.3390/v14040761] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/15/2022] [Revised: 03/31/2022] [Accepted: 04/03/2022] [Indexed: 02/05/2023] Open
Abstract
Hepatitis C virus (HCV) is the primary pathogen responsible for liver cirrhosis and hepatocellular carcinoma. The main virion component, the core (C) protein, has been linked to several aspects of HCV pathology, including oncogenesis, immune evasion and stress responses. We and others have previously shown that C expression in various cell lines activates Ca2+ signaling and alters Ca2+ homeostasis. In this study, we identified two distinct C protein regions that are required for the activation of Ca2+/NFAT signaling. In the basic N-terminal domain, which has been implicated in self-association of C, amino acids 1–68 were critical for NFAT activation. Sedimentation analysis of four mutants in this domain revealed that association of the C protein into nucleocapsid-like particles correlated with NFAT-activated transcription. The internal, lipid droplet-targeting domain was not required for NFAT-activated transcription. Finally, the C-terminal ER-targeting domain was required in extenso for the C protein to function. Our results indicate that targeting of HCV C to the ER is necessary but not sufficient for inducing Ca2+/NFAT signaling. Taken together, our data are consistent with a model whereby proteolytic intermediates of C with an intact transmembrane ER-anchor assemble into pore-like structures in the ER membrane.
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Affiliation(s)
- Priya Devi
- Department of Medical Sciences, Uppsala University, SE 75185 Uppsala, Sweden;
| | - Tanel Punga
- Department of Medical Biochemistry and Microbiology, Uppsala University, SE 75123 Uppsala, Sweden;
| | - Anders Bergqvist
- Department of Medical Sciences, Uppsala University, SE 75185 Uppsala, Sweden;
- Clinical Microbiology and Hospital Infection Control, Uppsala University Hospital, SE 75185 Uppsala, Sweden
- Correspondence: ; Tel.: +46-186113937
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Dutkiewicz M, Ciesiołka J. Form confers function: Case of the 3’X region of the hepatitis C virus genome. World J Gastroenterol 2018; 24:3374-3383. [PMID: 30122877 PMCID: PMC6092582 DOI: 10.3748/wjg.v24.i30.3374] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/27/2018] [Revised: 06/25/2018] [Accepted: 06/30/2018] [Indexed: 02/06/2023] Open
Abstract
At the 3’ end of genomic hepatitis C virus (HCV) RNA there is a highly conserved untranslated region, the 3’X-tail, which forms part of the 3’UTR. This region plays key functions in regulation of critical processes of the viral life cycle. The 3’X region is essential for viral replication and infectivity. It is also responsible for regulation of switching between translation and transcription of the viral RNA. There is some evidence indicating the contribution of the 3’X region to the translation efficiency of the viral polyprotein and to the encapsidation process. Several different secondary structure models of the 3’X region, based on computer predictions and experimental structure probing, have been proposed. It is likely that the 3’X region adopts more than one structural form in infected cells and that a specific equilibrium between the various forms regulates several aspects of the viral life cycle. The most intriguing explanations of the structural heterogeneity problem of the 3’X region came with the discovery of its involvement in long-range RNA-RNA interactions and the potential for homodimer formation. This article summarizes current knowledge on the structure and function of the 3’X region of hepatitis C genomic RNA, reviews previous opinions, presents new hypotheses and summarizes the questions that still remain unanswered.
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Affiliation(s)
- Mariola Dutkiewicz
- Institute of Bioorganic Chemistry, Polish Academy of Sciences, Poznan 61-704, Poland
| | - Jerzy Ciesiołka
- Institute of Bioorganic Chemistry, Polish Academy of Sciences, Poznan 61-704, Poland
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3
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Abd Alla MDA, El Awady MK, Dawood RM, Elhawary MA, Al-Azhari SS, Galal ASGM. Hepatitis C virus serologic relapse after treatment with direct-acting antivirals is dependent on viral RNA levels in peripheral blood mononuclear cells and the grade of liver cirrhosis. Arch Virol 2018; 163:2765-2774. [PMID: 29971486 DOI: 10.1007/s00705-018-3922-7] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/21/2018] [Accepted: 06/13/2018] [Indexed: 12/12/2022]
Abstract
The disappearance of hepatitis C virus (HCV) from serum and tissues for 12 weeks after the end of treatment (EOT) with direct-acting antivirals (DAAs) is known as a "sustained virologic response" (SVR) and occurs more frequently in non-cirrhotic patients than in cirrhotic patients. In this study, we evaluated the outcome of HCV treatment with sofosbuvir (SOF) plus ledipasvir (LDV) at both EOT and 12 weeks after EOT in patients with and without hepatic cirrhosis to address the relationship of serologic relapse to persistent infection of PBMCs and the frequency of hepatic encephalopathy and hepatocellular carcinoma (HCC) after treatment. Seventy-five patients with post-HCV liver cirrhosis were assigned to one of three groups (A, B, and C), each of which included 25 patients and corresponded to the patients' Child-Turcotte-Pugh (CTP) classification. All of the patients received a daily dose of SOF (400 mg) plus LDV (90 mg) for 24 weeks and were tested using HCV single-strand reverse transcription (SRT) and PCR analysis of PBMCs at both EOT and 12 weeks after EOT. Fourteen (18.7%) out of 75 patients (all study populations) had intra-PBMC HCV RNA, but only nine of them (64.3%) developed HCV RNA serum relapse (seroconversion) 12 weeks after EOT (P < 0.001). Encephalopathy was significantly higher in group C at EOT and 12 weeks after EOT (P < 0.05). Development of HCC was observed in decompensated patients of group C (2 out of 5 = 40.0%) 12 weeks post-EOT (P = 0.03). In conclusion, detection of HCV RNA within PBMCs at the EOT provides an indication of potential relapse after 12 weeks. Moreover, development of encephalopathy and HCC after HCV eradication by SOF plus LDV therapy is perhaps a future warning for post-treatment hepatic decompensation in cirrhotic patients.
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Affiliation(s)
- Mohamed Darwish Ahmed Abd Alla
- Tropical Medicine Department, Faculty of Medicine, El-Hussein University Hospital, Al-Azhar University, Gouhar Al-Kaed Street, Al-Darasah, Cairo, 11675, Egypt.
| | | | - Reham M Dawood
- Micrbial Biotechnology Department, National Research Center, Dokki, Cairo, 12622, Egypt
| | - Mostafa Abdelaziz Elhawary
- Tropical Medicine Department, Faculty of Medicine, El-Hussein University Hospital, Al-Azhar University, Gouhar Al-Kaed Street, Al-Darasah, Cairo, 11675, Egypt
| | - Shabaan Salah Al-Azhari
- Tropical Medicine Department, Faculty of Medicine, El-Hussein University Hospital, Al-Azhar University, Gouhar Al-Kaed Street, Al-Darasah, Cairo, 11675, Egypt
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4
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Blackard JT, Kong L, Lombardi A, Homann D, Hammerstad SS, Tomer Y. A preliminary analysis of hepatitis C virus in pancreatic islet cells. Virol J 2017; 14:237. [PMID: 29258547 PMCID: PMC5738208 DOI: 10.1186/s12985-017-0905-3] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/01/2017] [Accepted: 12/01/2017] [Indexed: 02/08/2023] Open
Abstract
BACKGROUND An association between hepatitis C virus (HCV) and type 2 diabetes (T2D) is supported by numerous epidemiologic studies. We hypothesized that HCV could infect human pancreatic islet cells in vitro. METHODS Measures of HCV RNA synthesis and protein production were used to evaluate HCV infection of pancreatic islets recovered from human donors. RESULTS Significant co-staining of insulin and the HCV entry factor CD81 was observed in pancreatic islets. Positive- and negative-sense HCV RNA were detected in HCV-exposed islets at days 1, 3, 7, and 14 post-infection. The HCV core and NS3 proteins were expressed and increased with time providing further evidence of viral replication. Interferon and an HCV polymerase inhibitor reduced viral replication in islet cells. In HCV-infected islets, TNFα levels were elevated at days 1, 3, and 7 post-infection, while IL-6 levels were elevated at day 1 but not days 3 or 7. Overall, the expression of miR-122 was low in islets compared to the Huh7.5 hepatocyte-derived cell line, although the relative expression of miR-122 increased in islet cells after viral infection (1, 6.63, and 5.83 at days 1, 3, and 7, respectively). CONCLUSIONS In this pilot study, viral infection was demonstrated in pancreatic islet cells from multiple donors using complementary measures of viral replication, thus providing evidence of in vitro infection. Altered cytokine expression may contribute to the development of insulin deficiency, and understanding the etiology of diabetes in individuals with HCV infection may facilitate the development of novel treatment modalities and prevention strategies. This in vitro system provides an important model for mechanistic studies of HCV-pancreas interactions and facilitates future studies of the potential impact of viral infection on islet cell function.
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Affiliation(s)
- Jason T Blackard
- Division of Digestive Diseases, Department of Internal Medicine, University of Cincinnati College of Medicine, ML 0595, 231 Albert Sabin Way, Cincinnati, OH, 45267, USA.
| | - Ling Kong
- Division of Digestive Diseases, Department of Internal Medicine, University of Cincinnati College of Medicine, ML 0595, 231 Albert Sabin Way, Cincinnati, OH, 45267, USA
| | - Angela Lombardi
- Department of Medicine, Albert Einstein College of Medicine and Montefiore Medical Center, Bronx, NY, USA
| | - Dirk Homann
- Diabetes Obesity and Metabolism Institute, Mount Sinai Medical Center, New York, NY, USA
| | | | - Yaron Tomer
- Department of Medicine, Albert Einstein College of Medicine and Montefiore Medical Center, Bronx, NY, USA
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Abd Alla MDA, El Awady MK. Hepatitis C Virus RNA Strands Detection in Peripheral Blood Mononuclear Cells Legitimizes Virus Eradication in Negative Serum PCR Naïve and Post-treatment Patients. J Clin Transl Hepatol 2017; 5:1-8. [PMID: 28507919 PMCID: PMC5415493 DOI: 10.14218/jcth.2016.00054] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/24/2016] [Revised: 01/14/2017] [Accepted: 02/03/2017] [Indexed: 12/11/2022] Open
Abstract
Background and Aims: Hepatitis C virus (HCV) hepatotropism is associated with intra-peripheral blood mononuclear cell (PBMC) infection that causes post-treatment relapse in RNA seronegative patients. Our understanding of the association of non-viremic hepatic fibrosis with positive anti-HCV IgG antibodies and active hepatocellular damage might be increased by PBMCs screening for intracellular infection. Thus, the goals of this study included evaluation of PBMCs PCR for diagnosing HCV infection, addressing PBMCs plus serum real-time (SRT) PCR benefits over SRT-PCR alone, studying intra-PBMCs distribution of RNA sense and antisense strands, and identifying treatment feasibility in solitary intracellular infection. Methods: Enzyme-linked immunosorbent assay, SRT-PCR and PBMCs PCR were used to evaluate HCV infection in 401 subjects. The patients were classified into groups of negative controls (n = 30), positive controls (n = 63), non-viremia post-treatment (experienced; n = 166) and naïve (n = 49) cases, and non-viremia positive PBMCs PCR naïve (n = 35) and experienced (n = 58) patients. Results: The diagnosis of true positive and negative by PBMCs PCR and SRT-PCR had 100% and 96.7% compatibility respectively. PBMCs PCR detected intracellular HCV infection in 49 out of 215 non-viremia patients; among them, naïve cirrhotics had significantly higher number of intracellular infection than the naïve non-cirrhotic (p < 0.001) and experienced patients (p < 0.0001). Antisense and sense strands were respectively recognized in naïve and experienced cases (p = 0.01218). Intracellular HCV strands were detected in 18.02% of experienced patients. Recognition of intracellular RNA strands showed significant decline in experienced compared to naïve patients (p < 0.05). Conclusion: PBMCs PCR is a valid diagnostic test that can diagnose intracellular HCV when SRT-PCR is negative. Antisense and sense strands are respectively recognized more often in naïve and experienced patients. The expected overall relapsing rate in our cohort was 18.02%. Intra-PBMC infections are associated with liver cirrhosis in naïve non-viremia patients. Eradication of intracellular strands is recommended to avoid RNA seroconversion. Ethical approval certificate: Registration number 10231.
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Affiliation(s)
- Mohamed Darwish Ahmed Abd Alla
- Tropical Medicine Department, Faculty of Medicine, Al-Azhar University, Cairo, Egypt
- *Correspondence to: Mohamed Darwish Ahmed Abd Alla, El-Hussein University Hospital, Al-Azhar University, Gouhar Al-Kaed Street, Al-Darasah, Cairo 11675, Egypt. Tel: +20-109-417-5209, Fax: +20-2512-3091, E-mail:
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6
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Abstract
HIV/hepatitis C virus (HCV) coinfection is estimated to affect 2 million individuals globally. The acceleration of HCV-associated complications, particularly hepatic fibrosis, because of HIV coinfection has been well established, whereas the impact of HCV on HIV progression remains unclear. In this review, we summarize the current evidence on the impact of coinfection on the transmission and clinical progression of each infection. We focus on the virological and immunological alterations that contribute to HIV and HCV pathogenesis in coinfection and also review the disease-modifying effects of antiretroviral therapy as they pertain to HCV.
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Nakai M, Seya T, Matsumoto M, Shimotohno K, Sakamoto N, Aly HH. The J6JFH1 Strain of Hepatitis C Virus Infects Human B-Cells with Low Replication Efficacy. Viral Immunol 2014; 27:285-94. [DOI: 10.1089/vim.2013.0140] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/19/2023] Open
Affiliation(s)
- Masato Nakai
- Department of Microbiology and Immunology, Hokkaido University Graduate School of Medicine, Kita-ku, Japan
- Department of Gastroenterology, Hokkaido University Graduate School of Medicine, Kita-ku, Japan
| | - Tsukasa Seya
- Department of Microbiology and Immunology, Hokkaido University Graduate School of Medicine, Kita-ku, Japan
| | - Misako Matsumoto
- Department of Microbiology and Immunology, Hokkaido University Graduate School of Medicine, Kita-ku, Japan
| | - Kunitada Shimotohno
- Research Center for Hepatitis and Immunology, National Center for Global Health and Medicine, Ichikawa, Japan
| | - Naoya Sakamoto
- Department of Gastroenterology, Hokkaido University Graduate School of Medicine, Kita-ku, Japan
| | - Hussein H. Aly
- Department of Microbiology and Immunology, Hokkaido University Graduate School of Medicine, Kita-ku, Japan
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Kisiel E, Radkowski M, Pawelczyk A, Horban A, Stanczak J, Bukowska-Ośko I, Caraballo Cortes K, Kaźmierczak J, Popiel M, Laskus T. Seronegative hepatitis C virus infection in patients with lymphoproliferative disorders. J Viral Hepat 2014; 21:424-9. [PMID: 24138606 DOI: 10.1111/jvh.12181] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/24/2022]
Abstract
It has been reported that hepatitis C virus (HCV) RNA may be present in serum and/or lymphoid cells in the absence of specific circulating antibodies. The current study analysed seronegative HCV infection in patients with lymphoproliferative disorders. We studied 77 anti-HCV-negative patients (45 male and 32 female, mean age 54.8 ± 14.2 years) with various lymphoproliferative disorders. HCV-RNA was detected by RT-PCR in plasma, peripheral blood mononuclear cells (PBMC) and bone marrow. Furthermore, the presence of viral nonstructural protein 3 (NS3) was determined in PBMC and bone marrow by immunostaining. HCV-RNA was detectable in at least one compartment in 27 (35.1%) patients. Viral RNA was found in bone marrow in 22 patients (28.6%), in PBMC in 13 (16.9%) and in plasma in 10 (13%) patients. In nine patients, evidence of infection was confined to the bone marrow compartment. Viral load in HCV-RNA-positive plasma ranged from 15 to 1.17 × 10(3) IU/mL. NS3 was detected in all but two HCV-RNA-positive bone marrow samples and in all but one HCV-RNA-positive PBMC samples. All 27 HCV-RNA-positive patients remained anti-HCV-negative when tested again after 6-12 months, but only four remained HCV-RNA positive. In conclusion, among patients with lymphoproliferative disorders, HCV can be present in plasma, PBMC and bone marrow despite the lack of circulating specific antibodies. Further studies are required to analyse the phenomenon of seronegative infection and to determine whether such patients are infectious.
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Affiliation(s)
- E Kisiel
- Department of Hematology, Institute of Hematology and Blood Transfusion, Warsaw, Poland
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Fernandez-Ponce C, Dominguez-Villar M, Aguado E, Garcia-Cozar F. CD4+ primary T cells expressing HCV-core protein upregulate Foxp3 and IL-10, suppressing CD4 and CD8 T cells. PLoS One 2014; 9:e85191. [PMID: 24465502 PMCID: PMC3896374 DOI: 10.1371/journal.pone.0085191] [Citation(s) in RCA: 25] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/23/2012] [Accepted: 11/30/2013] [Indexed: 12/11/2022] Open
Abstract
Adaptive T cell responses are critical for controlling HCV infection. While there is clinical evidence of a relevant role for regulatory T cells in chronic HCV-infected patients, based on their increased number and function; mechanisms underlying such a phenomena are still poorly understood. Accumulating evidence suggests that proteins from Hepatitis C virus can suppress host immune responses. We and others have shown that HCV is present in CD4+ lymphocytes from chronically infected patients and that HCV-core protein induces a state of unresponsiveness in the CD4+ tumor cell line Jurkat. Here we show that CD4+ primary T cells lentivirally transduced with HCV-core, not only acquire an anergic phenotype but also inhibit IL-2 production and proliferation of bystander CD4+ or CD8+ T cells in response to anti-CD3 plus anti-CD28 stimulation. Core-transduced CD4+ T cells show a phenotype characterized by an increased basal secretion of the regulatory cytokine IL-10, a decreased IFN-γ production upon stimulation, as well as expression of regulatory T cell markers, CTLA-4, and Foxp3. A significant induction of CD4+CD25+CD127(low)PD-1(high)TIM-3(high) regulatory T cells with an exhausted phenotype was also observed. Moreover, CCR7 expression decreased in HCV-core expressing CD4+ T cells explaining their sequestration in inflamed tissues such as the infected liver. This work provides a new perspective on de novo generation of regulatory CD4+ T cells in the periphery, induced by the expression of a single viral protein.
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Affiliation(s)
- Cecilia Fernandez-Ponce
- Department of Biomedicine, Biotechnology and Public Health (Immunology), University of Cadiz and Puerto Real University Hospital Research Unit, School of Medicine, Cadiz, Spain
| | - Margarita Dominguez-Villar
- Department of Biomedicine, Biotechnology and Public Health (Immunology), University of Cadiz and Puerto Real University Hospital Research Unit, School of Medicine, Cadiz, Spain
| | - Enrique Aguado
- Department of Biomedicine, Biotechnology and Public Health (Immunology), University of Cadiz and Puerto Real University Hospital Research Unit, School of Medicine, Cadiz, Spain
| | - Francisco Garcia-Cozar
- Department of Biomedicine, Biotechnology and Public Health (Immunology), University of Cadiz and Puerto Real University Hospital Research Unit, School of Medicine, Cadiz, Spain
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Kondo Y, Shimosegawa T. Direct effects of hepatitis C virus on the lymphoid cells. World J Gastroenterol 2013; 19:7889-7895. [PMID: 24307783 PMCID: PMC3848137 DOI: 10.3748/wjg.v19.i44.7889] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/11/2013] [Revised: 10/01/2013] [Accepted: 11/13/2013] [Indexed: 02/06/2023] Open
Abstract
It has been reported that the direct binding of hepatitis C virus (HCV) and/or the replication of HCV in the extrahepatic organs and, especially, lymphoid cells, might affect the pathogenesis of extrahepatic diseases with HCV infection. More than one decade ago, several reports described the existence of HCV-RNA in peripheral blood mononuclear cells. Moreover, many reports describing the existence of HCV in B lymphocytes and B cell lymphoma have been published. In addition to B lymphocytes, it was reported that HCV replication could be detected in T lymphocytes and T cell lines. Among the extrahepatic diseases with HCV infection, mixed cryoglobulinemia-related diseases and autoimmune-related diseases are important for understanding the immunopathogensis of HCV persistent infection. Moreover, HCV persistent infection can cause malignant lymphoma. The biological significance of lymphotropic HCV has not yet become clear. However, several candidates have been considered for a long time. One is that lymphotropic HCV is an HCV reservoir that might contribute to the recurrence of HCV infection and difficult-to-treat disease status. The other important issue is the carcinogenesis of the lymphoid cells and disturbances of the immune responses. Therefore, the extrahepatic diseases might be induced by direct interaction between HCV and lymphoid cells. In this article, we summarize various studies showing the direct effect of HCV on lymphoid cells and discuss the biological significance of lymphotropic HCV.
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11
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Pawełczyk A, Kubisa N, Jabłońska J, Bukowska-Ośko I, Caraballo Cortes K, Fic M, Laskus T, Radkowski M. Detection of hepatitis C virus (HCV) negative strand RNA and NS3 protein in peripheral blood mononuclear cells (PBMC): CD3+, CD14+ and CD19+. Virol J 2013; 10:346. [PMID: 24279719 PMCID: PMC4222874 DOI: 10.1186/1743-422x-10-346] [Citation(s) in RCA: 36] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/29/2013] [Accepted: 11/12/2013] [Indexed: 12/19/2022] Open
Abstract
Background Although hepatitis C virus (HCV) is primarily hepatotropic, markers of HCV replication were detected in peripheral blood mononuclear cells (PBMC) as well as in ex vivo collected tissues and organs. Specific strains of HCV were found to be capable to infect cells of the immune system: T and B cells and monocytes/macrophages as well as cell lines in vitro. The direct invasion of cells of the immune system by the virus may be responsible for extrahepatic consequences of HCV infection: cryoglobulinemia and non-Hodgkin’s lymphoma. The aim of the present study was to determine the prevalence of markers of HCV infection: negative strand HCV RNA and non-structural NS3 protein in PBMC subpopulations: CD3+, CD14+ and CD19+. The presence of virus and the proportion of affected cells within a particular PBMC fraction could indicate a principal target cell susceptible for HCV. Methods PBMC samples were collected from 26 treatment-free patients chronically infected with HCV. PBMC subpopulations: CD3+, CD14+, CD19+ were obtained using positive magnetic separation. The presence of negative strand RNA HCV and viral NS3 protein were analyzed by strand-specific RT-PCR and NS3 immunocytochemistry staining. Results Negative strand HCV RNA was detectable in 7/26 (27%), whereas NS3 protein in 15/26 (57.6%) of PBMC samples. At least one replication marker was found in 13/26 (50%) of CD3+ cells then in 8/26 (30.8%) of CD14+ and CD19+ cells. The highest percentage of cells harboring viral markers in single specimen was also observed in CD3+ (2.4%), then in CD19+ (1.2%), and much lower in CD14+ (0.4%) cells. Conclusions Our results indicate that CD3+ cells are a dominant site for extrahepatic HCV replication, although other PBMC subpopulations may also support virus replication.
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Affiliation(s)
- Agnieszka Pawełczyk
- Department of Immunopathology of Infectious and Parasitic Diseases, Warsaw Medical University, Warsaw, Poland.
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12
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Sengupta S, Powell E, Kong L, Blackard JT. Effects of HCV on basal and tat-induced HIV LTR activation. PLoS One 2013; 8:e64956. [PMID: 23762271 PMCID: PMC3677892 DOI: 10.1371/journal.pone.0064956] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/19/2012] [Accepted: 04/23/2013] [Indexed: 01/19/2023] Open
Abstract
Hepatitis C virus (HCV) co-infection occurs in ∼30–40% of the HIV-infected population in the US. While a significant body of research suggests an adverse effect of HIV on HCV replication and disease progression, the impact of HCV on HIV infection has not been well studied. Increasing data suggest that hepatocytes and other liver cell populations can serve as reservoirs for HIV replication. Therefore, to gain insight into the impact of HCV on HIV, the effects of the HCV Core protein and infectious hepatitis C virions were evaluated on basal and Tat-induced activation of the HIV long terminal repeat (LTR) in hepatocytes. The HIV LTR was highly induced by the HIV transactivator protein Tat in hepatocytes. Activation varied according to the number of NF-kB binding sites present in the LTRs from different HIV subtypes. Involvement of the NF-kB binding pathway in LTR activation was demonstrated using an NF-kB inhibitor and deletion of the NF-kB binding sites. TNFα, a pro-inflammatory cytokine that plays an important role in HIV pathogenesis, also induced LTR activity in hepatocytes. However, HIV LTR activity was suppressed in hepatocytes in the presence of HCV Core protein, and the suppressive effect persisted in the presence of TNFα. In contrast, infectious hepatitis C virions upregulated HIV LTR activation and gene transcription. Core-mediated suppression remained unaltered in the presence of HCV NS3/4A protein, suggesting the involvement of other viral/cellular factors. These findings have significant clinical implications as they imply that HCV could accelerate HIV disease progression in HIV/HCV co-infected patients. Such analyses are important to elucidate the mechanisms by which these viruses interact and could facilitate the development of more effective therapies to treat HIV/HCV co-infection.
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Affiliation(s)
- Satarupa Sengupta
- Division of Digestive Diseases, Department of Internal Medicine, University of Cincinnati College of Medicine, Cincinnati, Ohio, United States of America
| | - Eleanor Powell
- Division of Digestive Diseases, Department of Internal Medicine, University of Cincinnati College of Medicine, Cincinnati, Ohio, United States of America
| | - Ling Kong
- Division of Digestive Diseases, Department of Internal Medicine, University of Cincinnati College of Medicine, Cincinnati, Ohio, United States of America
| | - Jason T. Blackard
- Division of Digestive Diseases, Department of Internal Medicine, University of Cincinnati College of Medicine, Cincinnati, Ohio, United States of America
- * E-mail:
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Dominguez-Villar M, Fernandez-Ponce C, Munoz-Suano A, Gomez E, Rodríguez-Iglesias M, Garcia-Cozar F. Up-regulation of FOXP3 and induction of suppressive function in CD4+ Jurkat T-cells expressing hepatitis C virus core protein. Clin Sci (Lond) 2012; 123:15-27. [PMID: 22214248 DOI: 10.1042/cs20110631] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/04/2025]
Abstract
HCV (hepatitis C virus) infection is a serious health care problem that affects more than 170 million people worldwide. Viral clearance depends on the development of a successful cellular immune response against the virus. Interestingly, such a response is altered in chronically infected patients, leading to chronic hepatitis that can result in liver fibrosis, cirrhosis and hepatocellular carcinoma. Among the mechanisms that have been described as being responsible for the immune suppression caused by the virus, Treg-cells (regulatory T-cells) are emerging as an essential component. In the present work we aim to study the effect of HCV-core protein in the development of T-cells with regulatory-like function. Using a third-generation lentiviral system to express HCV-core in CD4+ Jurkat T-cells, we describe that HCV-core-expressing Jurkat cells show an up-regulation of FOXP3 (forkhead box P3) and CTLA-4 (cytotoxic T-lymphocyte antigen-4). Moreover, we show that HCV-core-transduced Jurkat cells are able to suppress CD4+ and CD8+ T-cell responses to anti-CD3 plus anti-CD28 stimulation.
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Affiliation(s)
- Margarita Dominguez-Villar
- Puerto Real University Hospital Research Unit, School of Medicine, Department of Biomedicine, Biotechnology (Immunology), University of Cadiz, Cadiz, Spain
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Chary A, Winters MA, Eisen R, Knight TH, Asmuth DM, Holodniy M. Quantitation of hepatitis C virus RNA in peripheral blood mononuclear cells in HCV-monoinfection and HIV/HCV-coinfection. J Med Virol 2012; 84:431-7. [PMID: 22246828 DOI: 10.1002/jmv.23210] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/28/2023]
Abstract
Peripheral blood mononuclear cells (PBMCs) represent an extrahepatic hepatitis C virus (HCV) reservoir, the significance of which is unclear due to limited studies and varying test methodologies. In this study, a commercial viral load assay for measuring cell-associated PBMC HCV RNA was evaluated. HCV RNA was extracted from PBMCs, sorted CD14+, and CD19+ cells and corresponding plasma samples using the Abbott m2000 and Real-Time HCV assay. Test performance and influence of HIV seropositivity on plasma and PBMC HCV RNA were studied. Among 51 patients, 67 and 62 unique patient samples had detectable plasma and PBMC HCV viral load, respectively. The median PBMC viral load was 535 IU/1 M cells (range 29-5,190). CD19+ cells had significantly higher viral load than CD14+ cells (median log(10) HCV viral load 2.63 vs. 1.50 IU/ml; P< 0.001). Stability of PBMC viral load over time was demonstrated in untreated patients; all patients with an undetectable plasma HCV viral load after HCV treatment also demonstrated undetectable PBMC viral load. Repeated testing in nine samples yielded consistent PBMC viral load, differing by only 1.3-fold (range 1.0-1.7-fold). Among samples with detectable plasma HCV RNA, the correlation between PBMC and plasma viral load was moderate (r = 0.66) and was greater among HCV mono-infected compared to HIV/HCV co-infected subjects (r = 0.80 vs. 0.52). Measurement of cell-associated PBMC HCV RNA using a commercial assay demonstrated promising test characteristics. Differences in PBMC HCV viral load based on HIV-coinfection status and the significance of greater copy number in B-cells requires further study.
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Affiliation(s)
- Aarthi Chary
- AIDS Research Center, VA Palo Alto Health Care System, Palo Alto, California, USA.
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15
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Biological significance of HCV in various kinds of lymphoid cells. Int J Microbiol 2012; 2012:647581. [PMID: 22518147 PMCID: PMC3299277 DOI: 10.1155/2012/647581] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/16/2011] [Accepted: 12/12/2011] [Indexed: 12/21/2022] Open
Abstract
It has been reported that HCV can infect not only hepatocytes but also various kinds of lymphoid cells. Although many reports have described the biological significance of lymphotropic HCV, the issue remains controversial since the target lymphoid cells might have various kinds of functions in the immune system. One of the important roles of lymphoid cells in HCV replication is being a reservoir of HCV. Several groups described the detection of HCV-RNA in lymphoid cells after HCV eradication in plasma. Another important role of lymphotropic HCV is that it acts as a carcinogenic agent and induces immune dysfunction. In this paper, we summarize the reports regarding the biological significance of lymphotropic HCV in representative lymphoid cells.
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16
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Ippagunta SK, Naik S, Jameel S, KN SR, Aggarwal R. Viral RNA but no evidence of replication can be detected in the peripheral blood mononuclear cells of hepatitis E virus-infected patients. J Viral Hepat 2011; 18:668-72. [PMID: 20659304 PMCID: PMC3075346 DOI: 10.1111/j.1365-2893.2010.01351.x] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/24/2022]
Abstract
Hepatitis E virus (HEV) infection is an important cause of acute viral hepatitis in several developing countries but has recently been shown to cause chronic hepatitis in immunosuppressed persons. Other hepatotropic viruses that cause chronic infection have been shown to infect peripheral blood mononuclear cells (PBMCs) and to persist in those cells. We therefore decided to look for evidence of replication of HEV in PBMCs obtained from patients with acute hepatitis E, using strand-specific assays for positive and negative HEV RNA. Of the 44 patients with acute hepatitis E during an outbreak in India, including 27 with detectable IgM anti-HEV and 19 with detectable serum HEV RNA, 11 had detectable HEV RNA in their PBMCs. However, of the six PBMC specimens with strong HEV RNA signal, none had detectable negative-strand HEV RNA, a marker of viral replication. These findings indicate the presence of HEV RNA but the absence of its replication in PBMCs from patients with acute hepatitis E.
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Affiliation(s)
- Sirish Kumar Ippagunta
- Department of Gastroenterology, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow 226014, India
| | - Sita Naik
- Department of Immunology, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow 226014, India
| | - Shahid Jameel
- Virology Group, International Centre for Genetic Engineering and Biotechnology, New Delhi 110067, India
| | - Sudha Ramana KN
- Sir Ronald Ross Institute of Tropical and Communicable Diseases, Government Fever Hospital, Hyderabad 530 044, India
| | - Rakesh Aggarwal
- Department of Gastroenterology, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow 226014, India
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Barrett L, Gallant M, Howley C, Ian Bowmer M, Hirsch G, Peltekian K, Grant M. Stronger hepatitis C virus-specific CD8+ T-cell responses in HIV coinfection. J Viral Hepat 2011; 18:170-80. [PMID: 20497309 DOI: 10.1111/j.1365-2893.2010.01293.x] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/09/2022]
Abstract
Hepatitis C virus (HCV) is a widespread chronic infection that shares routes of transmission with human immunodeficiency virus (HIV). Thus, coinfection with these viruses is a relatively common and growing problem. In general, liver disease develops over years with HIV coinfection, when compared to decades in HCV monoinfection. The role of the immune system in the accelerated pathogenesis of liver disease in HIV/HCV coinfection is not clear. In this study, we compared the frequency, magnitude, breadth and specificity of peripheral blood CD4+ and CD8+ T-cell responses between HCV-monoinfected and HCV/HIV-coinfected individuals and between HIV/HCV-coinfected subgroups distinguished by anti-HCV antibody and HCV RNA status. While HIV coinfection tended to reduce the frequency and breadth of anti-HCV CD8+ T-cell responses in general, responses that were present were substantially stronger than in monoinfection. In all groups, HCV-specific CD4+ T-cell responses were rare and weak, independent of either nadir or concurrent CD4+ T-cell counts of HIV-infected individuals. Subgroup analysis demonstrated restricted breadth of CD8+ HCV-specific T-cell responses and lower B-cell counts in HIV/HCV-coinfected individuals without anti-HCV antibodies. The greatest difference between HIV/HCV-coinfected and HCV-monoinfected groups was substantially stronger HCV-specific CD8+ T-cell responses in the HIV-coinfected group, which may relate to accelerated liver disease in this setting.
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Affiliation(s)
- L Barrett
- Immunology Program, Division of BioMedical Sciences, Faculty of Medicine, Memorial University of Newfoundland, St. John's, NL, Canada.
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18
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Bernini F, Ebranati E, De Maddalena C, Shkjezi R, Milazzo L, Lo Presti A, Ciccozzi M, Galli M, Zehender G. Within-host dynamics of the hepatitis C virus quasispecies population in HIV-1/HCV coinfected patients. PLoS One 2011; 6:e16551. [PMID: 21304985 PMCID: PMC3031583 DOI: 10.1371/journal.pone.0016551] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/28/2010] [Accepted: 12/29/2010] [Indexed: 12/16/2022] Open
Abstract
HIV/HCV coinfected individuals under highly active antiretroviral therapy (HAART) represent an interesting model for the investigation of the role played by the immune system in driving the evolution of the HCV quasispecies. We prospectively studied the intra-host evolution of the HCV heterogeneity in 8 coinfected subjects, selected from a cohort of 32 patients initiating HAART: 5 immunological responders (group A) and 3 immunological non-responders (group B), and in two HCV singly infected controls not assuming drugs (group C). For all these subjects at least two serial samples obtained at the first observation (before HAART) and more than 1 year later, underwent clonal sequence analysis of partial E1/E2 sequences, encompassing the whole HVR1. Evolutionary rates, dated phylogenies and population dynamics were co-estimated by using a Bayesian Markov Chain Monte Carlo approach, and site specific selection pressures were estimated by maximum likelihood-based methods. The intra-host evolutionary rates of HCV quasispecies was 10 times higher in subjects treated with HAART than in controls without immunodeficiency (1.9 and 2.3×10−3 sub/site/month in group A and B and 0.29×10−3 sub/site/month in group C individuals). The within-host Bayesian Skyline plot analysis showed an exponential growth of the quasispecies populations in immunological responders, coinciding with a peak in CD4 cell counts. On the contrary, quasispecies population remained constant in group B and in group C controls. A significant positive selection pressure was detected in a half of the patients under HAART and in none of the group C controls. Several sites under significant positive selection were described, mainly included in the HVR1. Our data indicate that different forces, in addition to the selection pressure, drive an exceptionally fast evolution of HCV during HAART immune restoration. We hypothesize that an important role is played by the enlargement of the viral replicative space.
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Affiliation(s)
- Flavia Bernini
- Department of Clinical Sciences “L. Sacco”, University of Milan, Milan, Italy
| | - Erika Ebranati
- Department of Clinical Sciences “L. Sacco”, University of Milan, Milan, Italy
| | - Chiara De Maddalena
- Department of Clinical Sciences “L. Sacco”, University of Milan, Milan, Italy
| | - Renata Shkjezi
- Department of Clinical Sciences “L. Sacco”, University of Milan, Milan, Italy
| | - Laura Milazzo
- Department of Clinical Sciences “L. Sacco”, University of Milan, Milan, Italy
| | - Alessandra Lo Presti
- Department of Infectious, Parasitic and Immune-Mediated Diseases, Istituto Superiore di Sanità, Rome, Italy
| | - Massimo Ciccozzi
- Department of Infectious, Parasitic and Immune-Mediated Diseases, Istituto Superiore di Sanità, Rome, Italy
| | - Massimo Galli
- Department of Clinical Sciences “L. Sacco”, University of Milan, Milan, Italy
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19
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[Tropism of hepatitis C virus for leukocytes-- importance of the analysis of viral E1 and E2 envelope glycoprotein genes by sequencing]. ACTA ACUST UNITED AC 2009; 58:170-4. [PMID: 19892492 DOI: 10.1016/j.patbio.2009.06.010] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2009] [Accepted: 06/26/2009] [Indexed: 11/23/2022]
Abstract
The ability of hepatitis C virus (HCV) to infect leukocytes could favour HCV pathogenesis. Although viral infection of these immunocompetent cells is poorly (or not) productive, the impact on their immunomodulatory functions could be important. Viral envelope glycoproteins E1 and E2, because of their crucial role in the recognition of viral receptors on permissive cells, could contribute to viral leukocytic tropism and, as a consequence, to the pathophysiology of HCV chronic infection.
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20
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Ye L, Wang X, Wang S, Wang Y, Song L, Hou W, Zhou L, Li H, Ho W. CD56+ T cells inhibit hepatitis C virus replication in human hepatocytes. Hepatology 2009; 49:753-62. [PMID: 19085952 PMCID: PMC2677088 DOI: 10.1002/hep.22715] [Citation(s) in RCA: 46] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Abstract
UNLABELLED CD56(+) T cells are abundant in liver and play an important role in defense against viral infections. However, the role of CD56(+) T cells in control of hepatitis C virus (HCV) infection remains to be determined. We investigated the noncytolytic anti-HCV activity of primary CD56(+) T cells in human hepatocytes. When HCV Japanese fulminant hepatitis-1 (JFH-1)-infected hepatocytes were co-cultured with CD56(+) T cells or incubated in media conditioned with CD56(+) T cell culture supernatants (SN), HCV infectivity and replication were significantly inhibited. The antibodies to interferon (IFN)-gamma or IFN-gamma receptor could largely block CD56(+) T cell-mediated anti-HCV activity. Investigation of mechanism(s) responsible for CD56(+) T cell-mediated noncytolytic anti-HCV activity showed that CD56(+) T SN activated the multiple elements of janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway and enhanced the expression of IFN regulatory factors (IRFs) 1, 3, 7, 8, and 9, resulting in the induction of endogenous IFN-alpha/beta expression in hepatocytes. Moreover, CD56(+) T SN treatment inhibited the expression of HCV-supportive micro RNA (miRNA)-122 and enhanced the levels of anti-HCV miRNA-196a in human hepatocytes. CONCLUSION These findings provide direct in vitro evidence at cellular and molecular levels that CD56(+) T cells may have an essential role in innate immune cell-mediated defense against HCV infection. (HEPATOLOGY 2009.).
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Affiliation(s)
- Li Ye
- From Division of Allergy & Immunology, Joseph Stokes, Jr. Research Institute at The Children’s Hospital of Philadelphia, Department of Pediatrics, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA
| | - Xu Wang
- From Division of Allergy & Immunology, Joseph Stokes, Jr. Research Institute at The Children’s Hospital of Philadelphia, Department of Pediatrics, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA
| | - Shihong Wang
- From Division of Allergy & Immunology, Joseph Stokes, Jr. Research Institute at The Children’s Hospital of Philadelphia, Department of Pediatrics, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA
| | - Yanjian Wang
- From Division of Allergy & Immunology, Joseph Stokes, Jr. Research Institute at The Children’s Hospital of Philadelphia, Department of Pediatrics, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA
| | - Li Song
- From Division of Allergy & Immunology, Joseph Stokes, Jr. Research Institute at The Children’s Hospital of Philadelphia, Department of Pediatrics, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA
| | - Wei Hou
- From Division of Allergy & Immunology, Joseph Stokes, Jr. Research Institute at The Children’s Hospital of Philadelphia, Department of Pediatrics, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA
| | - Lin Zhou
- From Division of Allergy & Immunology, Joseph Stokes, Jr. Research Institute at The Children’s Hospital of Philadelphia, Department of Pediatrics, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA.
,Division of Histology and Embryology, Department of Anatomy, Tongji Medical College, Huazhong University of Science and Technology, Hubei 430030, China
| | - He Li
- Division of Histology and Embryology, Department of Anatomy, Tongji Medical College, Huazhong University of Science and Technology, Hubei 430030, China
| | - Wenzhe Ho
- From Division of Allergy & Immunology, Joseph Stokes, Jr. Research Institute at The Children’s Hospital of Philadelphia, Department of Pediatrics, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA
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21
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Gazzola L, Tincati C, Bellistre GM, d'Arminio Monforte A, Marchetti G. The Absence of CD4+ T Cell Count Recovery Despite Receipt of Virologically Suppressive Highly Active Antiretroviral Therapy: Clinical Risk, Immunological Gaps, and Therapeutic Options. Clin Infect Dis 2009. [DOI: 10.1086/695852] [Citation(s) in RCA: 92] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/03/2022] Open
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22
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Gazzola L, Tincati C, Bellistrì GM, Monforte AD, Marchetti G. The absence of CD4+ T cell count recovery despite receipt of virologically suppressive highly active antiretroviral therapy: clinical risk, immunological gaps, and therapeutic options. Clin Infect Dis 2009; 48:328-37. [PMID: 19123868 DOI: 10.1086/595851] [Citation(s) in RCA: 113] [Impact Index Per Article: 7.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/03/2022] Open
Abstract
Up to 30% of human immunodeficiency virus (HIV)-infected patients who are receiving long-term highly active antiretroviral therapy do not exhibit a marked increase in the CD4(+) T cell count, despite achieving complete suppression of the HIV load. These patients are referred to as "immunological nonresponders." When treating immunological nonresponders, the practicing clinician has several questions, including questions about the clinical risk associated with persistent immunodeficiency and about possible approaches to treatment that would provide clinical and immunological benefits. However, tentative answers to these questions require investigations of the mechanisms that underlie the lack of immune recovery, because only the deepest comprehension of the immunological gaps underlying functional defects will allow administration of highly targeted and efficacious treatment strategies. The aim of our review is to provide a thorough assessment of the clinical implications of a lack of increase in the CD4(+) T cell count in immunological nonresponders, to examine the immunological gaps limiting recovery of the CD4(+) T cell count, and to note possible therapeutic avenues, which may offer clinicians guidance regarding how to most efficaciously treat these critical patients.
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Affiliation(s)
- Lidia Gazzola
- Department of Medicine, Surgery, and Dentistry, Clinic of Infectious Diseases, San Paolo Hospital, University of Milan, Milan, Italy
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23
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Schramm F, Moenne-Loccoz R, Fafi-Kremer S, Soulier E, Royer C, Weitten T, Brignon N, Ellero B, Woehl-Jaegle ML, Meyer C, Wolf P, Doffoel M, Baumert TF, Gut JP, Stoll-Keller F, Schvoerer E. [Study of hepatitis C virus leukotropism by characterization of viral quasispecies in the liver transplantation setting]. ACTA ACUST UNITED AC 2008; 56:487-91. [PMID: 18842359 DOI: 10.1016/j.patbio.2008.07.007] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/28/2008] [Accepted: 07/03/2008] [Indexed: 01/28/2023]
Abstract
Besides hepatocytes, representing the main replication site of hepatitis C virus, peripheral blood mononuclear cells also represent a crucial target for viral infection. Hepatitis C virus compartmentalization (i.e., non-random distribution) of viral variants between plasma and peripheral blood mononuclear cells, more frequently observed in liver transplant patients compared to non-transplanted patients, makes liver transplantation an interesting model for the analysis of hepatitis C leukotropism. This article aims to present, firstly, in clinical and biological features arguing favour of hepatitis C virus infection leukotropism and, secondly, to review current knowledge about compartmentalization between plasma and peripheral blood mononuclear cells, especially in the liver transplantation setting.
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Affiliation(s)
- F Schramm
- Unité Inserm 748, 3, rue Koeberlé, 67000 Strasbourg, France.
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24
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Abstract
Because of major advances in the treatment of HIV/AIDS, HIV-positive persons now live longer, healthier lives; however, hepatitis C virus (HCV) is increasingly recognized as a major cause of morbidity and mortality in this population. Among HCV-infected persons, HIV co-infection is associated with increased HCV RNA levels, increased hepatic inflammation and fibrosis, and more rapid progression to cirrhosis and end-stage liver disease. Compounding this problem are reduced HCV treatment response rates among HCV/HIV co-infected persons. Moreover, antiretroviral therapy used to suppress HIV replication is often associated with a paradoxical increase in HCV RNA levels, as well as hepatotoxicity. Despite the adverse clinical consequences of HCV/HIV co-infection, the mechanisms by which these two viruses interact at the cellular level remain largely unexplored. This review focuses on the evidence demonstrating direct infection of hepatocytes by HIV, as well as the indirect mechanisms by which HIV may regulate HCV replication at the cellular level. A comprehensive understanding of virus-virus and virus-cell interactions is critical to the development of novel treatment strategies to combat HCV/HIV co-infection.
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Affiliation(s)
- J T Blackard
- Division of Digestive Diseases, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA.
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25
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Villacres MC, Literat O, DeGiacomo M, Du W, Frederick T, Kovacs A. Defective response to Toll-like receptor 3 and 4 ligands by activated monocytes in chronic hepatitis C virus infection. J Viral Hepat 2008; 15:137-44. [PMID: 18184197 PMCID: PMC3118839 DOI: 10.1111/j.1365-2893.2007.00904.x] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Toll-like receptors (TLR) have a critical role in innate immunity against pathogens. We investigated the cytokine response to TLR stimulation in peripheral blood cells of subjects infected with hepatitis C virus (HCV) and/or human immunodeficiency virus (HIV) in the Women Interagency HIV Study (WIHS) cohort. Interleukin (IL)-6 in response to TLR3 and TLR4 ligands such as polyinosinic-polycytidylic acid and lipopolysaccharide was significantly compromised in HCV-infected women. High spontaneous secretion of IL-6 suggested pre-existing cell activation as a factor mediating reduced responses to TLR3 and TLR4 stimulation. To a lesser extent, tumour necrosis factor-alpha and IL-1beta responses to TLR stimulation were also compromised. Monocytes, but not B cells or NK cells, were identified as the cell population spontaneously secreting cytokines and also as the cells responding to TLR stimulation. These results highlight a functional defect in antigen-presenting cells of women with HCV infection or co-infection. In women with existing HIV co-infection, decreased cytokine function of antigen-presenting cells suggests another mechanism contributing to immune dysfunction in addition to the HIV-associated CD4 defect.
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Affiliation(s)
- M C Villacres
- Maternal, Child and Adolescent Center for Infectious Diseases and Virology, Keck School of Medicine, University of Southern California, Los Angeles, CA 90033, USA.
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26
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Impact of hepatitis C virus coinfection on immune restoration during successful antiretroviral therapy in chronic human immunodeficiency virus type 1 disease. Eur J Clin Microbiol Infect Dis 2007; 27:65-73. [DOI: 10.1007/s10096-007-0384-3] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/27/2022]
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27
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Rodríguez-Torres M, Rodríguez-Orengo JF, Ríos-Bedoya CF, Fernández-Carbia A, González-Lassalle E, Salgado-Mercado R, Marxuach-Cuétara AM. Efficacy and safety of peg-IFN alfa-2a with ribavirin for the treatment of HCV/HIV coinfected patients who failed previous IFN based therapy. J Clin Virol 2007; 38:32-8. [PMID: 17064957 DOI: 10.1016/j.jcv.2006.09.009] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/30/2005] [Revised: 06/20/2006] [Accepted: 09/19/2006] [Indexed: 02/05/2023]
Abstract
BACKGROUND Interferon (IFN) regimens for HCV treatment are less effective in HCV/HIV-coinfected patients. There are no effective treatments for patients who fail IFN therapies. We examined the safety and efficacy of peginterferon alfa-2a (peg-IFNalpha-2a) plus ribavirin (RBV) in 41HCV/HIV-coinfected patients non-responsive to prior IFN treatment. METHODS Patients received peg-IFNalpha-2a (180mg/week) plus RBV (800mg/day) for 24 weeks (n=41). At week 24, patients with non-detectable HCV RNA or > or =2-log decrease from baseline, received peg-IFNalpha-2a (180mg/week) plus RBV (800mg/day) for 24 weeks further. Patients not responding to treatment at week 24 were discontinued. RESULTS Intent to treat (ITT) sustained viral response (SVR) was 21.9%. Patients who received at least 24 weeks of peg-IFNalpha-2a plus RBV treatment (n=35), SVR rates were 25.7%. SVR was associated with significant improvements in liver histology grade (p=0.02), stage (p=0.02), and fibrosis progression rate (FPR) (p=0.03). Patients that failed to achieve SVR had statistically significant decreases in grade (p=0.09) and FPR (p=0.01). CONCLUSION peg-IFNalpha-2a plus RBV is effective and safe to achieve SVR in HCV/HIV coinfected patients non-responsive to prior IFN treatment. Patients that achieve SVR have significant improvements in liver histology parameters. In patients that do not achieve SVR there are histological benefits beyond virological response that suggest that peg-IFNalpha-2a+RBV therapy may decrease risk of progression to end stage liver disease.
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28
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MacParland SA, Pham TNQ, Gujar SA, Michalak TI. De novo infection and propagation of wild-type Hepatitis C virus in human T lymphocytes in vitro. J Gen Virol 2006; 87:3577-3586. [PMID: 17098973 DOI: 10.1099/vir.0.81868-0] [Citation(s) in RCA: 39] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
While exploring previous findings that ex vivo treatment of lymphoid cells from Hepatitis C virus (HCV)-infected individuals with T cell-stimulating mitogens augments detection of the residing virus, an in vitro HCV replication system was established, in which mitogen-induced T cell-enriched cultures served as HCV targets and the derived T cells multiplied virus during repeated serial passage. HCV replication was ascertained by detecting HCV RNA positive and negative strands, HCV NS5a and E2 proteins, release of HCV virions and nucleocapsids (confirmed by immunoelectron microscopy) and de novo infection of mitogen-induced T cells prepared from healthy donors. Further, affinity-purified normal human T lymphocytes were also susceptible to HCV infection in vitro and HCV replication was detected in pure T cells isolated from a patient with chronic hepatitis C. These results document that T cells can support propagation of HCV both in vivo and in vitro. The infection system established offers a valuable tool for in vitro studies on the entire cycle of HCV replication, virus cytopathogenicity and evaluation of antiviral agents against wild-type HCV in the natural host-cell milieu.
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Affiliation(s)
- Sonya A MacParland
- Molecular Virology and Hepatology Research, Division of Basic Medical Sciences, Faculty of Medicine, Health Sciences Centre, Memorial University, St John's, NL A1B 3V6, Canada
| | - Tram N Q Pham
- Molecular Virology and Hepatology Research, Division of Basic Medical Sciences, Faculty of Medicine, Health Sciences Centre, Memorial University, St John's, NL A1B 3V6, Canada
| | - Shashi A Gujar
- Molecular Virology and Hepatology Research, Division of Basic Medical Sciences, Faculty of Medicine, Health Sciences Centre, Memorial University, St John's, NL A1B 3V6, Canada
| | - Tomasz I Michalak
- Discipline of Laboratory Medicine, Faculty of Medicine, Health Sciences Centre, Memorial University, St John's, NL A1B 3V6, Canada
- Molecular Virology and Hepatology Research, Division of Basic Medical Sciences, Faculty of Medicine, Health Sciences Centre, Memorial University, St John's, NL A1B 3V6, Canada
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29
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Laskus T, Operskalski EA, Radkowski M, Wilkinson J, Mack WJ, deGiacomo M, Al-Harthi L, Chen Z, Xu J, Kovacs A. Negative-strand hepatitis C virus (HCV) RNA in peripheral blood mononuclear cells from anti-HCV-positive/HIV-infected women. J Infect Dis 2006; 195:124-33. [PMID: 17152016 PMCID: PMC3319123 DOI: 10.1086/509897] [Citation(s) in RCA: 57] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2006] [Accepted: 08/31/2006] [Indexed: 01/08/2023] Open
Abstract
BACKGROUND Hepatitis C virus (HCV) has been reported to replicate in peripheral blood mononuclear cells (PBMCs), particularly in patients coinfected with HCV and human immunodeficiency virus (HIV). However, there are limited data regarding the prevalence of and the factors associated with extrahepatic replication. METHODS The presence of negative-strand HCV RNA in PBMCs was evaluated by a strand-specific assay for 144 anti-HCV-positive/HIV-infected women enrolled in the Women's Interagency HIV Study. One to 5 PBMC samples obtained from each woman were tested. Multivariate analyses were used to assess for associations with the clinical and demographic characteristics of the women. RESULTS Negative-strand HCV RNA was detected in 78 (25%) of 315 specimens, and, for 61 women (42%), > or = 1 specimen was found to have positive results. The presence of negative-strand HCV RNA in PBMCs was significantly positively associated with an HCV RNA plasma level of > or = 6.75 log copies/mL (P=.04) and consumption of > or = 7 alcoholic drinks per week (P=.02). It was also negatively associated with injection drug use occurring in the past 6 months (P=.03). A negative association with a CD4+ CD38+ DR+ cell percentage of > 10% and a positive association with acquired immunodeficiency syndrome were borderline significant (P=.05). CONCLUSIONS HCV replication in PBMCs is common among HIV-coinfected women and appears to be a dynamic process related to lifestyle, virologic, and immunologic factors.
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Affiliation(s)
- Tomasz Laskus
- Department of Medicine, St. Joseph's Hospital and Medical Center, Phoenix, AZ 85013, USA.
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Nguyen H, Sankaran S, Dandekar S. Hepatitis C virus core protein induces expression of genes regulating immune evasion and anti-apoptosis in hepatocytes. Virology 2006; 354:58-68. [PMID: 16876223 DOI: 10.1016/j.virol.2006.04.028] [Citation(s) in RCA: 54] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/14/2006] [Revised: 03/17/2006] [Accepted: 04/24/2006] [Indexed: 12/12/2022]
Abstract
Hepatitis C virus (HCV) Core protein is implicated in the development of hepatocellular carcinoma (HCC). We utilized a HepG2 human hepatocyte cell line with inducible expression of HCV Core protein (HCV-1b) to investigate the early effects of Core protein on hepatocyte gene expression and to identify molecular processes modulated by the Core protein. A significant change was observed in the expression of 407 genes, which included genes regulating apoptosis, immune response, and cell cycle. Some of these genes were previously known to be tumor markers. The decreased expression of chemo-attractants such as TNFSF10, CCL20, and osteopontin was observed, which suggested that HCV Core expression could lead to suppression of inflammatory response as well as trafficking of macrophages and neutrophils to the site of HCV infection. An increased expression of anti-apoptosis factors including PAK2, API5, BH1, Tax1BP1, DAXX, and TNFAIP3/A20 was observed. Some of these genes were also linked to the regulation of NFKB activation and that the alteration of their expression levels, by HCV Core, might lead to the suppression NFKB activation of inflammatory responses. Our data suggested that Core expression may contribute to the viral persistence by protecting infected hepatocytes from cell death by the suppressing apoptosis and inflammatory reaction to HCV viral infection.
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Affiliation(s)
- Hau Nguyen
- Department of Medical Microbiology and Immunology, School of Medicine, Topper Hall, Room 3146, University of California, Davis, CA 95616, USA
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Di Liberto G, Roque-Afonso AM, Kara R, Ducoulombier D, Fallot G, Samuel D, Feray C. Clinical and therapeutic implications of hepatitis C virus compartmentalization. Gastroenterology 2006; 131:76-84. [PMID: 16831592 DOI: 10.1053/j.gastro.2006.04.016] [Citation(s) in RCA: 69] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/23/2005] [Accepted: 04/07/2006] [Indexed: 12/17/2022]
Abstract
BACKGROUND & AIMS Blood mononuclear cells (BMCs) frequently are infected by hepatitis C virus (HCV) variants that are not found in plasma. The influence of this compartmentalization on the natural and therapeutic outcome of hepatitis C is unknown. METHODS We studied 119 patients with previously untreated chronic HCV infection. Sixty-five of these patients started first-line treatment with pegylated interferon-alfa and ribavirin after enrollment in the study. The internal ribosomal entry site (IRES) of HCV RNA was amplified and compared between plasma and BMCs by means of single-strand conformational polymorphism (SSCP) analysis, line-probe assay, and cloning sequencing. RESULTS The IRES SSCP patterns differed between plasma and BMCs in 54 (48%) of 113 assessable patients. Twenty-seven (24%) of these patients were co-infected by 2 HCV types or subtypes, only 1 of which was detectable in BMCs (n = 25) or in plasma (n = 2). SSCP-defined compartmentalization was more frequent in former drug users than in others (35/56 [60%] vs 19/56 [34%]; P < .01), and less frequent in patients with genotype 1 HCV in plasma (26/73 [24%] vs 28/40 [65%]; P < .01). The only variables that were independently predictive of a sustained virologic response were SSCP-defined compartmentalization (25/31 vs 10/32; P = .0001) and genotype 2 or 3 infection of BMCs (22/31 vs 8/34; P = .002). CONCLUSIONS A significant proportion of patients with hepatitis C are co-infected by 2 or more HCV variants with distinct IRES sequences and distinct cellular tropism. This compartmentalization is a strong independent predictor of treatment efficacy.
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Affiliation(s)
- Gaëtana Di Liberto
- Institut National de la Santé et de la Recherche Médicale, INSERM, Centre de Recherche Biomedicale Beaujon-Bichat, Faculté de Médecine Xavier Bichat, Paris, France
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Blackard JT, Kemmer N, Sherman KE. Extrahepatic replication of HCV: insights into clinical manifestations and biological consequences. Hepatology 2006; 44:15-22. [PMID: 16799966 DOI: 10.1002/hep.21283] [Citation(s) in RCA: 134] [Impact Index Per Article: 7.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
An estimated 170 million persons are infected with the hepatitis C virus (HCV) worldwide. While hepatocytes are the major site of infection, a broad clinical spectrum of extrahepatic complications and diseases are associated with chronic HCV infection, highlighting the involvement of HCV in a variety of non-hepatic pathogenic processes. There is a growing body of evidence to suggest that HCV can replicate efficiently in extrahepatic tissues and cell types, including peripheral blood mononuclear cells. Nonetheless, laboratory confirmation of HCV replication in extrahepatic sites is fraught with technical challenges, and in vitro systems to investigate extrahepatic replication of HCV are severely limited. Thus, future studies of extrahepatic replication should combine innovative in vitro assays with a prospective cohort design to maximize our understanding of this important phenomenon to the pathogenesis and treatment response rates of HCV.
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Affiliation(s)
- Jason T Blackard
- Division of Digestive Diseases, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA.
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Powers KA, Ribeiro RM, Patel K, Pianko S, Nyberg L, Pockros P, Conrad AJ, McHutchison J, Perelson AS. Kinetics of hepatitis C virus reinfection after liver transplantation. Liver Transpl 2006; 12:207-16. [PMID: 16447184 DOI: 10.1002/lt.20572] [Citation(s) in RCA: 64] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
Improved understanding of hepatitis C virus (HCV) dynamics during and after liver transplantation can be useful in optimizing antiviral therapy in transplant recipients. We analyzed serum HCV ribonucleic acid (RNA) levels during and after cadaveric liver transplantation in 6 HCV patients. After removal of the liver and before the new liver started producing virions, HCV RNA levels dropped with an average half-life (t(1/2)) of 0.8 hours. Viral loads then continued to drop up to 23 hours postimplantation (t(1/2) = 3.4 hours), and began to rise (doubling-time = 2.0 days) as soon as 15 hours after the anhepatic phase. In 3 patients the viral load reached a plateau before rising, suggesting that a nonhepatic source supplied virions and balanced their intrinsic clearance. However, from the decline in viral load over the first 24 hours of the postanhepatic phase, we estimate that nonhepatic sources can at most correspond to 4% of total viral production, 96% of which occurs in the liver, even after we corrected for fluid exchanges during surgery. As the new liver was reinfected, production increased and viral load rose to a new steady state. Using nonlinear regression, we were able to fit the patients' HCV RNA data to a viral dynamic model and estimate the de novo infection rate (mean 1.5 x 10(-6) mL/virion/day), as well as the average percentage of hepatocytes infected at the posttransplantation steady state (19%). In conclusion, we have quantified liver reinfection dynamics in the absence of posttransplantation antiviral therapy. Our findings support the notion that early antiviral therapy may delay or prevent reinfection.
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Affiliation(s)
- Kimberly A Powers
- Theoretical Biology & Biophysics Group, Los Alamos National Laboratory, Los Alamos, NM 87545, USA
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Georgopoulou U, Tsitoura P, Kalamvoki M, Mavromara P. The protein phosphatase 2A represents a novel cellular target for hepatitis C virus NS5A protein. Biochimie 2006; 88:651-62. [PMID: 16460864 DOI: 10.1016/j.biochi.2005.12.003] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/04/2005] [Accepted: 12/16/2005] [Indexed: 01/15/2023]
Abstract
It is well established that HCV NS5A protein when expressed in mammalian cells perturbs the extracellular signal regulated kinase (ERK) pathway. The protein serine/threonine phosphatase 2A controls the phosphorylation of numerous proteins involved in cell signaling and one characterized function is the regulation of Ras-Raf mitogen activated protein (MAP) kinase signaling pathways. Our results showed that expression of HCV NS5A protein stimulates phosphatase 2A (PP2A) activity in cells, indicating the relevance of NS5A as a regulator of PP2A in vivo. We found that transient expression of the full length NS5A protein in different cell lines leads to a significant increase of the PP2A activity and this activity is specifically inhibited by the addition of okadaic acid, a PP2A inhibitor, in living cells. Further investigation showed that NS5A protein interacts in vivo and in vitro with the scaffolding A and the catalytic C subunits of PP2A. We propose that HCV NS5A represents a viral PP2A regulatory protein. This is a novel function for the NS5A protein which may have a key role in the ability of the virus to deregulate cell growth and survival.
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Affiliation(s)
- Urania Georgopoulou
- Molecular Virology Laboratory, Hellenic Pasteur Institute, 127, Vas. Sofias Avenue, Athens 11521, Greece.
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Abstract
Chronic hepatitis C virus (HCV) infection is frequently associated with a number of extrahepatic complications. In the majority of cases the underlying pathogenetic mechanisms are immune mediated, as evidenced by the presence of circulating autoantibodies (mixed cryoglobulinemia), whereas for others a localized host cellular immune response is implicated (e.g. sialadenitis, lichen planus). In this review, the latest data on the pathogenesis and clinical manifestations of the most common autoimmune extrahepatic manifestations of chronic HCV infection are presented.
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Affiliation(s)
- Dimitrios Vassilopoulos
- Academic Department of Medicine, Athens University School of Medicine, Hippokration General Hospital, Athens, Greece
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Villacres MC, Literat O, Degiacomo M, Du W, La Rosa C, Diamond DJ, Kovacs A. Reduced type 1 and type 2 cytokines in antiviral memory T helper function among women coinfected with HIV and HCV. J Clin Immunol 2005; 25:134-41. [PMID: 15821890 PMCID: PMC3127261 DOI: 10.1007/s10875-005-2819-x] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 12/17/2004] [Indexed: 10/25/2022]
Abstract
Bias in cytokine responses has been proposed as a contributing mechanism to pathogenesis in persistent HIV or hepatitis C virus (HCV) infections. We investigated whether coinfection with HCV modifies the profile of antigen-specific cytokine secretion in women persistently infected with HIV compared to women with single HIV or HCV infection. The T helper response to HIV, HCV and cytomegalovirus (CMV) as a positive viral control was dominated by type 1 cytokines (interleukin- [IL] 2, interferon- [IFN] gamma and tumor necrosis factor- [TNF] alpha), with IFN-gamma as the most abundantly secreted. IL-4, IL-5 and IL-10 were low in healthy controls and patients. Robust CMV-specific responses contrasted with curtailed HCV-specific responses in HCV-infected women. The overall anti-viral profile was dominated by Th1 cytokines even in coinfected women but both type 1 and type 2 responses were reduced in HIV-infected women and more extensively in women with HCV/HIV coinfection.
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Affiliation(s)
- Maria C Villacres
- Maternal, Child and Adolescent Center for Infectious Diseases and Virology, Keck School of Medicine, University of Southern California, Los Angeles, CA 99033, USA.
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Zehender G, De Maddalena C, Bernini F, Ebranati E, Monti G, Pioltelli P, Galli M. Compartmentalization of hepatitis C virus quasispecies in blood mononuclear cells of patients with mixed cryoglobulinemic syndrome. J Virol 2005; 79:9145-56. [PMID: 15994809 PMCID: PMC1168762 DOI: 10.1128/jvi.79.14.9145-9156.2005] [Citation(s) in RCA: 32] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022] Open
Abstract
The aim of this study was to investigate the quasispecies heterogeneity of hepatitis C virus (HCV) in the plasma, cryoprecipitate, and peripheral lymphocytes of chronically infected HCV patients with mixed cryoglobulinemia (MC). We studied 360 clones from 10 HCV-positive patients with MC and 8 age-, gender- and HCV genotype-matched subjects with chronic HCV infection but without MC. A partial nucleotide sequence encompassing the E1/E2 region, including hypervariable region 1 (HVR1), was amplified and cloned from plasma, cryoprecipitates, and peripheral blood mononuclear cells (PBMC), and the genetic diversity and complexity and synonymous and nonsynonymous substitution rates were determined. Heterogeneous selection pressure at codon sites was evaluated. Compartmentalization was estimated by phylogenetic and phenetic (Mantel's test) approaches. The patients with MC had 3.3 times lower nonsynonymous substitution rates (1.7 versus 5.7 substitutions/100 sites). Among the subjects with HCV genotype 1, the MC patients had significantly less complexity than the controls, whereas the diversity and complexity were similar in the genotype 2 patients and controls. Site-specific selection analysis confirmed the low frequency of MC patients showing positive selection. There was a significant correlation between positive selection and the infecting HCV genotype. The quasispecies were less heterogeneous in PBMC than in plasma. Significant compartmentalization of HCV quasispecies was observed in the PBMC of four of nine subjects (three with MC) and seven of nine cryoprecipitates. In one subject with MC, we detected a 5-amino-acid insertion at codons 385 to 389 of HVR1. Our results suggest reduced quasispecies heterogeneity in MC patients that is related to a low selection pressure which is probably due to an impaired immune response, the HCV genotype, and/or the duration of the infection. The frequent HCV quasispecies compartmentalization in patients' PBMC suggests a possible pathogenetic significance.
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Affiliation(s)
- Gianguglielmo Zehender
- Istituto di Malattie Infettive e Tropicali, Università di Milano c/o Ospedale L. Sacco, Via G.B. Grassi 74, 20157 Milan, Italy.
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Roque-Afonso AM, Ducoulombier D, Di Liberto G, Kara R, Gigou M, Dussaix E, Samuel D, Féray C. Compartmentalization of hepatitis C virus genotypes between plasma and peripheral blood mononuclear cells. J Virol 2005; 79:6349-57. [PMID: 15858018 PMCID: PMC1091708 DOI: 10.1128/jvi.79.10.6349-6357.2005] [Citation(s) in RCA: 119] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023] Open
Abstract
Differences in hepatitis C virus (HCV) variants of the highly conserved 5' untranslated region (UTR) have been observed between plasma and peripheral blood mononuclear cells (PBMC). The prevalence and the mechanisms of this compartmentalization are unknown. Plasma and PBMC HCV variants were compared by single-strand conformation polymorphism (SSCP) and by cloning or by genotyping with a line probe assay (LiPA) in 116 chronically infected patients, including 44 liver transplant recipients. SSCP patterns differed between compartments in 43/109 analyzable patients (39%). Differences were significantly more frequent in patients with transplants (21/38 [55%] versus 22/71 [31%]; P < 0.01) and in those who acquired HCV through multiple transfusions before 1991 (15/20; 75%) or through drug injection (16/31; 52%) than in those infected through an unknown route (7/29; 24%) or through a single transfusion (5/29; 17%; P < 0.001). Cloning of the 5' UTR, LiPA analysis, and nonstructural region 5B sequencing revealed different genotypes in the two compartments from 10 patients (9%). In nine patients, the genotype detected in PBMC was not detected in plasma and was weak or undetectable in the liver in three cases. This genotypic compartmentalization persisted for years in three patients and after liver transplantation in two. The present study shows that a significant proportion of HCV-infected subjects harbor in their PBMC highly divergent variants which were likely acquired through superinfections.
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Antonucci G, Girardi E, Cozzi-Lepri A, Capobianchi MR, De Luca A, Puoti M, Petrelli E, Carnevale G, Rizzardini G, Grossi PA, Viganò P, Moioli MC, Carletti F, Solmone M, Ippolito G, Monforte AD. Role of hepatitis C virus (HCV) viremia and HCV genotype in the immune recovery from highly active antiretroviral therapy in a cohort of antiretroviral-naive HIV-infected individuals. Clin Infect Dis 2005; 40:e101-9. [PMID: 15909251 DOI: 10.1086/430445] [Citation(s) in RCA: 39] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/28/2004] [Accepted: 02/11/2005] [Indexed: 12/30/2022] Open
Abstract
BACKGROUND The roles of hepatitis C virus (HCV) viremia and HCV genotype in the immune response to highly active antiretroviral therapy (HAART) are poorly understood. Our aim was to assess the CD4+ cell count recovery after HAART in human immunodeficiency virus (HIV)-infected patients with HCV viremia and HIV-infected patients who tested negative for HCV antibody (HCV-Ab). We also aimed to assess whether the response to HAART in these patients varied according to HCV genotype. METHODS The analysis focused on 1219 HCV-Ab-negative patients and 284 HCV-viremic patients from a cohort of HIV-infected subjects that includes persons who were antiretroviral naive before initiating HAART after cohort enrollment. HCV RNA load and HCV genotype were determined in plasma specimens obtained and stored during the 6-month period preceding the initiation of HAART. RESULTS The chance of achieving a CD4+ cell count increase of > or = 100 cells/microL from the pre-HAART level tended to be poorer in HCV-viremic patients than in patients who tested negative for HCV-Ab (adjusted relative hazard [RH], 0.82; 95% confidence interval [CI], 0.66-1.01; P = .06). In contrast, a comparison of patients who had a HCV RNA load >1 x 10(6) IU/mL with patients who had a HCV RNA load of 5-1 x 10(6) IU/mL revealed no significant association between HCV RNA load and achievement of an increased CD4+ cell count (adjusted RH, 0.97; 95% CI, 0.75-1.27; P = .83). There was no clear association between HCV genotype and the probability of achieving a CD4+ cell count increase. CONCLUSIONS An association between the presence of HCV-Ab and immune reconstitution after HAART has been shown elsewhere. Results of our large, prospective study support a direct role of HCV viremia in the CD4+ cell count response to HAART. Moreover, our results underline the fact that, in individuals coinfected with HIV and HCV, the goal of treating HCV infection is to eradicate HCV, to both slow the rate of HCV progression and limit potential interference with the response to HAART.
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Affiliation(s)
- Giorgio Antonucci
- National Institute of Infectious Diseases, L. Spallanzani, Rome, Italy.
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Pham TNQ, MacParland SA, Coffin CS, Lee SS, Bursey FR, Michalak TI. Mitogen-induced upregulation of hepatitis C virus expression in human lymphoid cells. J Gen Virol 2005; 86:657-666. [PMID: 15722526 DOI: 10.1099/vir.0.80624-0] [Citation(s) in RCA: 58] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
Considering growing evidence indicating that hepatitis C virus (HCV) replicates in lymphoid cells, establishment of a reliable and sensitive method for detection of HCV in these cells may provide means for monitoring the infection and the efficacy of sterilizing antiviral therapy. In this study, conditions for ex vivo augmentation and detection of the HCV genome in peripheral blood mononuclear cells (PBMCs) from patients with chronic hepatitis C (CHC) or after a sustained virological response (SVR) to antiviral treatment were assessed. Following stimulation with combinations of mitogens and/or cytokines, PBMCs and, in certain cases, affinity-purified T and B cells were examined for HCV positive- and negative-strand RNA by using RT-PCR followed by nucleic acid hybridization, while the presence of viral NS3 protein was determined by flow cytometry. HCV RNA augmentation was assessed by quantification of Southern and dot-blot hybridization signals. The results showed that treatment of peripheral lymphoid cells with mitogens stimulating T- and B-cell proliferation and with cytokines supporting their growth significantly increased HCV RNA detection in patients with both CHC and SVR. This enhancement was up to 100-fold for the HCV genome and fivefold for the NS3 protein compared with untreated cells. In conclusion, HCV RNA can be readily detected in circulating lymphoid cells in progressing hepatitis C and following SVR after ex vivo cell stimulation. As such, this method offers a new investigative tool to study HCV lymphotropism and to monitor virus presence during the course of HCV infection.
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Affiliation(s)
- Tram N Q Pham
- Molecular Virology and Hepatology Research, Division of Basic Medical Science, Faculty of Medicine, Health Sciences Centre, Memorial University, St John's, Newfoundland, Canada A1B 3V6
| | - Sonya A MacParland
- Molecular Virology and Hepatology Research, Division of Basic Medical Science, Faculty of Medicine, Health Sciences Centre, Memorial University, St John's, Newfoundland, Canada A1B 3V6
| | - Carla S Coffin
- Liver Unit, Division of Gastroenterology, University of Calgary, Calgary, Alberta, Canada
- Molecular Virology and Hepatology Research, Division of Basic Medical Science, Faculty of Medicine, Health Sciences Centre, Memorial University, St John's, Newfoundland, Canada A1B 3V6
| | - Samuel S Lee
- Liver Unit, Division of Gastroenterology, University of Calgary, Calgary, Alberta, Canada
| | - Ford R Bursey
- Gastroenterology Unit, General Hospital, Faculty of Medicine, Memorial University, St John's, Newfoundland, Canada
| | - Tomasz I Michalak
- Discipline of Laboratory Medicine, Faculty of Medicine, Memorial University, St John's, Newfoundland, Canada
- Molecular Virology and Hepatology Research, Division of Basic Medical Science, Faculty of Medicine, Health Sciences Centre, Memorial University, St John's, Newfoundland, Canada A1B 3V6
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Carlos Martín J, Castilla J, López M, Arranz R, González-Lahoz J, Soriano V. Impact of chronic hepatitis C on HIV-1 disease progression. HIV CLINICAL TRIALS 2004; 5:125-31. [PMID: 15248136 DOI: 10.1310/yfv8-fe5k-5ln9-dq4c] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/09/2023]
Abstract
BACKGROUND Although there is clear evidence of an accelerated progression of liver fibrosis in HIV-positive patients with chronic hepatitis C virus (HCV) infection, it is unclear whether HCV infection may influence HIV-1 disease progression. We have analyzed the impact of HCV on CD4 counts and plasma HIV RNA in a large group of HIV-positive individuals. METHOD Epidemiological data, CD4 counts, and plasma HIV RNA values were recorded from 902 consecutive HIV-1-positive persons who attended our institution since 1998. RESULTS HCV infection was documented (antibodies and/or HCV RNA) in 72% of the total study population. The higher rates were seen among intravenous drug users (97%) compared to other groups (17% in homosexual men, 23% in patients who acquired HIV heterosexually). In a cross-sectional analysis performed at the first trimester of 2000, the mean CD4 count was lower among HCV-positive than among HCV-negative individuals (518 +/- 282 cells/microL vs. 620 +/- 302 cells/microL; p <.001). The mean plasma HIV RNA was 11,188 +/- 55,301 copies/mL in HCV-positive persons versus 6,352 +/- 32,152 copies/mL in HCV-negative persons (p =.03). Undetectable plasma HIV RNA (<50 copies/mL) was recognized in 54% of HCV-positive persons versus 64% of HCV negative persons (p =.04); a similar proportion of patients in each group was on antiretroviral therapy (90% vs. 93%) or HAART (86% vs. 89%). When comparing data from 1998 and 2000, the CD4 count increased an average of 53 cells/microL (11%) in HCV-positive persons versus 111 (19%) in HIV-negative persons during this 2-year interval (p <.05). Plasma HIV RNA on average declined 606 copies/mL (5%) in HCV-positive persons versus 5,788 copies/mL (54%) in HCV-negative persons (p <.05). A significant association between HCV infection and CD4 counts was recognized in the multivariate analysis, which was independent of gender, age, plasma HIV RNA, use of HAART, and adherence to therapy. In contrast, no significant effect of HCV on HIV RNA was found. CONCLUSION Hepatitis C may be associated with a poor immunologic outcome in HIV-infected persons. This worst influence is not explained by a lower rate of antiretroviral therapy among HCV-positive persons nor a much poorer drug adherence in this population. Therefore, hepatitis C may act as a direct cofactor for HIV disease progression. If so, treatment of chronic hepatitis C might indirectly benefit HIV disease.
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Affiliation(s)
- Juan Carlos Martín
- Service of Infectious Diseases, Hospital Carlos III, Carlos III Institute of Health, Madrid, Spain
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Laskus T, Radkowski M, Jablonska J, Kibler K, Wilkinson J, Adair D, Rakela J. Human immunodeficiency virus facilitates infection/replication of hepatitis C virus in native human macrophages. Blood 2004; 103:3854-9. [PMID: 14739225 DOI: 10.1182/blood-2003-08-2923] [Citation(s) in RCA: 67] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/25/2023] Open
Abstract
Hepatitis C virus (HCV) was found to replicate in monocytes/macrophages particularly in patients with human immunodeficiency virus type 1 (HIV-1) infection. This study was undertaken to determine whether HIV facilitates HCV infection of native human macrophages in vitro. Monocytes/macrophages were collected from healthy donors, infected with HIV M-tropic molecular clone, and then exposed to HCV-positive sera. Presence of positive and negative HCV RNA strands was determined with a novel strand-specific quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR). Preceding as well as near-simultaneous infection with HIV made the macrophages more susceptible to infection with HCV; in particular, an HCV RNA-negative strand was detectable almost exclusively in the setting of concomitant HIV infection. Furthermore, HCV RNA load correlated with HIV replication level in the early stage of infection. The ratio of positive to negative strand in macrophages was lower than in control liver samples. HIV infection was also found to facilitate HCV replication in a Daudi B-cell line with engineered CD4 expression. It seems that HIV infection can facilitate replication of HCV in monocytes/macrophages either by rendering cells more susceptible to HCV infection or by increasing HCV replication. This could explain the presence of extrahepatic HCV replication in HIV-coinfected individuals.
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Affiliation(s)
- Tomasz Laskus
- Department of Medicine, Mayo Clinic Scottsdale, AZ 85259, USA
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Baré P, Massud I, Belmonte L, Corti M, Villafañe M, Pérez Bianco R, de Tezanos-pinto M, de Bracco MME, Ruibal-Ares B. HCV recovery from peripheral blood mononuclear cell culture supernatants derived from HCV-HIV co-infected haemophilic patients with undetectable HCV viraemia. Haemophilia 2003; 9:598-604. [PMID: 14511301 DOI: 10.1046/j.1365-2516.2003.00808.x] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
Hepatitis C viraemia, in 38 human immunodeficiency virus positive (HIV+)/hepatitis C virus positive (HCV+) patients, was determined in haemophilic patients during the 4 years since initiation of highly active antiretroviral therapy (HAART). Six of 38 patients had persistently HCV-negative viraemia for more than 2 years. No correlation between HCV-negative viraemia and CD4+ T-cell counts, HIV viral load, age, type or severity of haemophilia could be established. Reduced levels of HIV viral load and the immune reconstitution that follows the initiation of HAART were not enough to explain the disappearance of HCV from plasma. Individuals who cleared plasma HCV had significantly higher CD8+ T-cell counts (P=0.0013) (mean +/- SE: 1153 +/- 117.8 cells microL(-1)) than those with HCV-positive viraemia (819.1 +/- 40.72 cells microL(-1)). Because HCV could maintain a low replication level in peripheral blood mononuclear cells (PBMC), we cultured PBMC of five of six patients with undetectable HCV viraemia. We found four of five HCV RNA-positive cultures. The presence of HCV RNA in our cultures proved that these cells may be an important viral reservoir that could contribute to HCV recurrence in plasma even after long periods of negative viraemia. In summary, our results indicate that in spite of prolonged HCV-negative plasma viraemia, HCV patients that are co-infected with HIV may harbour replication-competent HCV in their PBMC. Therefore, true clearance of HCV infection is difficult to achieve in these patients.
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Affiliation(s)
- P Baré
- Instituto de Investigaciones Hematológicas, Academia Nacional de Medicina, Pacheco de Melo 3081, Argentina.
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Abstract
With highly active antiretroviral therapy (HAART), HIV-infected patients can now live longer and healthier lives, and other comorbid diseases, such as chronic hepatitis C, have emerged as a significant health concern. Coinfection with the hepatitis C virus (HCV) may limit life expectancy because it can lead to serious liver disease including decompensated liver cirrhosis and hepatocellular carcinoma. HCV-induced fibrosis progresses faster in HIV/HCV-coinfected persons, although HAART may be able to decrease this disease acceleration. Combination therapy for HCV with interferon and ribavirin can achieve a sustained viral response, although at a lower rate than in HCV-monoinfected patients. Combination treatment with pegylated interferon and ribavirin will probably emerge as the next HCV therapy of choice for HIV/HCV-coinfected patients. HCV combination therapy is generally safe, but serious adverse reactions, like lactic acidosis, may occur. Cytopenia may present a problem leading to dose reductions, but the role of growth factors is under study. All HIV/HCV-coinfected patients should be evaluated for therapy against the hepatitis C virus. A sustained viral load will probably lead to regression of liver disease, and even interferon-based treatment without viral clearance may slow down progression of liver disease. HIV/HCV-coinfected patients who have progressed to end-stage liver disease have few therapeutic options other than palliative care, since liver transplants are generally unavailable. The mortality post-transplant may be higher than in HCV-monoinfected patients. We are entering an era where safe and effective HCV therapy is being defined for HIV/HCV-coinfected patients, and all eligible patients should be offered treatment.
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45
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Abstract
Hepatitis C virus (HCV) is the major cause for non-A, non-B hepatitis. Most HCV-infected individuals do not clear the virus resulting in a chronic infection that may potentially lead to liver cirrhosis and hepatocellular carcinoma. In addition to hepatic manifestations, HCV infection is associated with B cell lymphoproliferative disorders, including mixed cryoglobulinemia, usually a benign condition, and overt B cell lymphoma. A direct role of HCV infection in the genesis of these B cell lymphoproliferative disorders has been suggested initially by epidemiological studies and is supported by recent studies, which analyzed the monoclonal B cells that proliferate in these disorders. How HCV induces B cell lymphoproliferative disorders is still unclear, it is probably not due to direct change of phenotype in B cells after viral infection, but may be due to an HCV-antigen driven process. Support for this hypothesis comes from the analysis of monoclonal B cells found in these disorders, which use a restricted repertoire of immunoglobulin variable region genes that are similar to those used by B cells that secrete anti-HCV antibodies. The fact that monoclonal IgM is resolved in HCV-infected patients who responded to anti-viral treatment supports the linkage between antigen persistence and B cell proliferation. Finally, the linkage between benign B cell proliferation and overt lymphoma is supported by the identification of a pre-malignant B cell clone that subsequently converted to an overt B cell lymphoma. The molecular basis for viral induced B cell proliferation is still unknown. One possibility is that HCV stimulates the proliferation of monoclonal B cells via their HCV-specific B cell receptor (BCR) on the cell surface. Binding of the HCVenvelope proteins to a cellular ligand, CD81, may also enhance this antigen-driven process. A recent report on regression of splenic marginal zone lymphoma after anti-viral treatment with interferon and ribavirin has significantly strengthened the cause-effect relationship between HCV infection and lymphoma. Further studies should determine whether BCRs expressed on HCV-associated lymphomas, particularly those that regress in response to anti-viral therapy, bind HCV antigens that stimulate their proliferation.
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Affiliation(s)
- Wen-Kai Weng
- Department of Medicine, Division of Oncology, CCSR 1105a, Stanford University School of Medicine, Stanford, CA 94305, USA
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46
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Abstract
In summary, HCV-cell interactions include those directly involved with the HCV life cycle such as virus attachment, entry, and replication. Included within this broad area of research are the interactions of HCV proteins with the IFN system, cytokine and chemokine pathways such as IL-8, and various other cellular proteins and pathways. The plethora of contradictory and sometimes confusing accessory HCV-host interactions defies precise predictions of their role in HCV biology. It is clear that these virus-cell interactions affect HCV replication, antiviral resistance, persistence, and pathogenesis. Because HCV-host interactions are initiated immediately on infection, they are operative during acute HCV infection, whereby HCV interacts with innate cellular antiviral and immune systems. The magnitude and duration of these HCV-host interactions therefore may influence the development of acquired immunity. Because HCV exists as a quasispecies in all infected individuals, heterogeneity in biological responses to HCV-host interactions is predicted, revealing opportunities for the development of various genotypic and phenotypic prognostic indicators. With the model systems in place, these hypotheses can be tested. The challenge for the future is to determine if there is a hierarchical importance to these interactions, to delineate how these virus-cell interactions affect the patient infected with HCV, and to determine whether any of these interactions represents a target for therapeutic intervention.
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Affiliation(s)
- Stephen J Polyak
- Department of Laboratory Medicine, University of Washington, Box 359690, 325 9th Avenue, Seattle, WA 98104-2499, USA.
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Vargas HE, Laskus T, Radkowski M, Wilkinson J, Balan V, Douglas DD, Harrison ME, Mulligan DC, Olden K, Adair D, Rakela J. Detection of hepatitis C virus sequences in brain tissue obtained in recurrent hepatitis C after liver transplantation. Liver Transpl 2002; 8:1014-9. [PMID: 12424714 DOI: 10.1053/jlts.2002.36393] [Citation(s) in RCA: 49] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Patients with chronic hepatitis C frequently report tiredness, easy fatigability, and depression. The aim of this study is to determine whether hepatitis C virus (HCV) replication could be found in brain tissue in patients with hepatitis C and depression. We report two patients with recurrent hepatitis C after liver transplantation who also developed severe depression. One patient died of multiorgan failure and the other, septicemia caused by Staphylococcus aureussis. Both patients had evidence of severe hepatitis C recurrence with features of cholestatic fibrosing hepatitis. We were able to study samples of their central nervous system obtained at autopsy for evidence of HCV replication. The presence of HCV RNA-negative strand, which is the viral replicative form, was determined by strand-specific Tth-based reverse-transcriptase polymerase chain reaction. Viral sequences were compared by means of single-strand conformation polymorphism and direct sequencing. HCV RNA-negative strands were found in subcortical white matter from one patient and cerebral cortex from the other patient. HCV RNA-negative strands amplified from brain tissue differed by several nucleotide substitutions from serum consensus sequences in the 5' untranslated region. These findings support the concept of HCV neuroinvasion, and we speculate that it may provide a biological substrate to neuropsychiatric disorders observed in patients with chronic hepatitis C. The exact lineage of cells permissive for HCV replication and the possible interaction between viral replication and cerebral function that may lead to depression remain to be elucidated.
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Affiliation(s)
- Hugo E Vargas
- Division of Transplantation Medicine, Mayo Clinic Scottsdale, Scottsdale, AZ 85259, USA
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48
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Polyak SJ, Khabar KS, Paschal DM, Ezelle HJ, Duverlie G, Barber GN, Levy DE, Mukaida N, Gretch DR. Hepatitis C virus nonstructural 5A protein induces interleukin-8, leading to partial inhibition of the interferon-induced antiviral response. J Virol 2001; 75:6095-106. [PMID: 11390611 PMCID: PMC114325 DOI: 10.1128/jvi.75.13.6095-6106.2001] [Citation(s) in RCA: 239] [Impact Index Per Article: 10.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
Hepatitis C virus (HCV), a major cause of liver disease worldwide, is frequently resistant to the antiviral alpha interferon (IFN). The HCV nonstructural 5A (NS5A) protein has been implicated in HCV antiviral resistance in many studies. NS5A antagonizes the IFN antiviral response in vitro, and one mechanism is via inhibition of a key IFN-induced enzyme, the double-stranded-RNA-activated protein kinase (PKR). In the present study we determined if NS5A uses other strategies to subvert the IFN system. Expression of full-length NS5A proteins from patients who exhibited a complete response (FL-NS5A-CR) or were nonresponsive (FL-NS5A-NR) to IFN therapy in HeLa cells had no effect on IFN induction of IFN-stimulated gene factor 3 (ISGF-3). Expression of mutant NS5A proteins lacking 110 (NS5A-DeltaN110), 222 (NS5A-DeltaN222), and 334 amino-terminal amino acids and mutants lacking 117 and 230 carboxy-terminal amino acids also had no effect on ISGF-3 induction by IFN. Expression of FL-NS5A-CR and FL-NS5A-NR did not affect IFN-induced STAT-1 tyrosine phosphorylation or upregulation of PKR and major histocompatibility complex class I antigens. However, NS5A expression in human cells induced interleukin 8 (IL-8) mRNA and protein, and this effect correlated with inhibition of the antiviral effects of IFN in an in vitro bioassay. NS5A induced transcription of a reporter gene driven by the IL-8 promoter, and the first 133 bp of the IL-8 promoter made up the minimal domain required for NS5A transactivation. NS5A-DeltaN110 and NS5A-DeltaN222 stimulated the IL-8 promoter to higher levels than did the full-length NS5A protein, and this correlated with increased nuclear localization of the proteins. Additional mutagenesis of the IL-8 promoter suggested that NF-kappaB and AP-1 were important in NS5A-DeltaN222 transactivation in the presence of tumor necrosis factor alpha and that NF-IL-6 was inhibitory to this process. This study suggests that NS5A inhibits the antiviral actions of IFN by at least two mechanisms and provides the first evidence for a biological effect of the transcriptional activity of the NS5A protein. During HCV infection, viral proteins may induce chemokines that contribute to HCV antiviral resistance and pathogenesis.
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Affiliation(s)
- S J Polyak
- Department of Laboratory Medicine, Virology Division, University of Washington, 325 9th Ave., Seattle, WA 98104-2499, USA.
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49
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Charges virales VHC dans les cellules mononucléées sanguines chez les patients mono-infectés par le VHC et les patients co-infectés par le VIH et le VHC. Med Mal Infect 2000. [DOI: 10.1016/s0399-077x(01)80035-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022]
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50
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Greub G, Ledergerber B, Battegay M, Grob P, Perrin L, Furrer H, Burgisser P, Erb P, Boggian K, Piffaretti JC, Hirschel B, Janin P, Francioli P, Flepp M, Telenti A. Clinical progression, survival, and immune recovery during antiretroviral therapy in patients with HIV-1 and hepatitis C virus coinfection: the Swiss HIV Cohort Study. Lancet 2000; 356:1800-5. [PMID: 11117912 DOI: 10.1016/s0140-6736(00)03232-3] [Citation(s) in RCA: 627] [Impact Index Per Article: 25.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
BACKGROUND Hepatitis C virus (HCV) infection is highly prevalent among HIV-1-infected individuals, but its contribution to the morbidity and mortality of coinfected patients who receive potent antiretroviral therapy is controversial. We used data from the ongoing Swiss HIV Cohort Study to analyse clinical progression of HIV-1, and the virological and immunological response to potent antiretroviral therapy in HIV-1-infected patients with or without concurrent HCV infection. METHODS We analysed prospective data on survival, clinical disease progression, suppression of HIV-1 replication, CD4-cell recovery, and frequency of changes in antiretroviral therapy according to HCV status in 3111 patients starting potent antiretroviral therapy. RESULTS 1157 patients (37.2%) were coinfected with HCV, 1015 of whom (87.7%) had a history of intravenous drug use. In multivariate Cox's regression, the probability of progression to a new AIDS-defining clinical event or to death was independently associated with HCV seropositivity (hazard ratio 1.7 [95% CI 1.26-2.30]), and with active intravenous drug use (1.38 [1.02-1.88]). Virological response to antiretroviral therapy and the probability of treatment change were not associated with HCV serostatus. In contrast, HCV seropositivity was associated with a smaller CD4-cell recovery (hazard ratio for a CD4-cell count increase of at least 50 cells/microL=0.79 [0.72-0.87]). INTERPRETATION HCV and active intravenous drug use could be important factors in the morbidity and mortality among HIV-1-infected patients, possibly through impaired CD4-cell recovery in HCV seropositive patients receiving potent antiretroviral therapy. These findings are relevant for decisions about optimum timing for HCV treatment in the setting of HIV infection.
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Affiliation(s)
- G Greub
- Division of Infectious Diseases, University Hospital, Lausanne, Switzerland
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