- Email: [email protected]
815 IDENIX NS5A HCV REPLICATION INHIBITORS WITH LOW PICOMOLAR, PAN-GENOTYPIC IN VITRO ANTIVIRAL ACTIVITY
POSTERS performed using a non-targeted multiple platform methodology combining ultrahigh performance liquid chromatography/tandem mass spectrometry (UHPLC/MS) and gas chromatography/mass spectrometry (GC/MS). Metabolites were identified by automated comparison of ion features to a reference of chemical standard entries. Welch’s two-sample t-tests were used to identify biochemicals that differed significantly between HCV infected and un-infected cells. Results: A total of 250 metabolites were detected and quantified, of which 73 were differentially regulated. At the 24-hour time point, there was a significant increase in a number of metabolites involved in nucleotide synthesis and RNA replication. NAD levels were also significantly increased along with several amino acids. A number of lipid metabolic pathways were disrupted by HCV infection, resulting in an increase in cholesterol and sphingolipid levels, altered phospholipid metabolism and a possible disruption in mitochondrial fatty acid transport. Fluctuations in methylthioadenosine (MTA) levels were also noted, along with alterations in the glutathione synthesis pathway. Conclusions: These results suggest that elevated metabolism and altered energy status are associated with early HCV infection. These findings also provide new information on the effect HCV infection has on the disruption of lipid metabolism. 813 HEPATITIS C VIRUS-SPECIFIC CELLULAR IMMUNITY DOES NOT PROTECT AGAINST FUTURE HCV INFECTION IN ANTI-HCV NEGATIVE INJECTING DRUG USERS R. Sacks-Davis1,2 , C. Aitken1,2 , P. Higgs1,2,3 , S. Moneer4 , J. Flynn5,6 , V. Suppiah7,8 , L. Tracy9 , R. Ffrench5,6 , D. Bowden9 , H. Drummer4,10,11 , J. George7 , M. Bharadwaj4 , M. Hellard1,2 . 1 Centre for Population Health, Burnet Institute, 2 Department of Epidemiology and Preventative Medicine, Monash University, Melbourne, VIC, 3 National Centre for HIV Epidemiology & Clinical Research, University of New South Wales, Darlinghurst, NSW, 4 Department of Microbiology and Immunology, University of Melbourne, Parkville, 5 Centre for Immunology, Burnet Institute, 6 Department of Immunology, Monash University, Melbourne, VIC, 7 Storr Liver Unit, Westmead Millennium Institute, 8 Institute for Immunology and Allergy Research, Westmead Millennium Institute, University of Sydney, Sydney, NSW, 9 Victorian Infectious Diseases Reference Laboratory, North Melbourne, 10 Centre for Virology, Burnet Institute, Melbourne, 11 Department of Microbiology, Monash University, Clayton, VIC, Australia E-mail: [email protected] Background and Aims: Findings of large proportions of hepatitis C virus (HCV) anti-HCV negative injecting drug users (IDUs) with detectable HCV-specific cellular immune responses raise the question of whether some people have immunity to HCV infection. We conducted a longitudinal study to determine whether antiHCV negative IDUs with detectable HCV-specific cellular immune responses were more or less likely than other anti-HCV negative participants to develop persistent HCV infection. Methods: Cohort study participants who were anti-HCV antibody and HCV RNA negative at baseline and had at least one further HCV test were eligible for participation. HCV-specific cellular immune responses were measured at baseline and one subsequent time point in an ex vivo interferon-g enzymelinked immunospot (ELISpot) assay with pooled HCV peptides. Participants with responses of at least 25 spot forming cells per million were classified as ELISpot positive. Potential behavioral, social, environmental and genetic variation (single nucleotide polymorphism in the IL28 region) that might correlate with ELISpot positivity were examined. Results: Of 53 anti-HCV negative participants, 20 were HCV ELISpot positive at baseline. Age, gender and HCV risk behaviors did not differ by HCV ELISpot status. Similar rates of HCV incidence were S326
observed amongst HCV ELISpot positive participants (28.7 per 100 py, 95% CI 12.9, 63.8) and those who were HCV ELISpot negative (19.4 per 100 py, 95% CI 8.1, 46.6). Genetic variations in the IL28B region previously identified as associated with viral clearance were no more prevalent amongst participants who were ELISpot positive and anti-HCV antibody negative, HCV RNA negative than those who were anti-HCV antibody positive or HCV RNA positive. Conclusions: HCV-specific cellular immune responses were observed in more than one third of anti-HCV negative IDUs but our data contain no evidence that this subgroup of IDUs is protected from future HCV infection. 814 INDUCTION OF EPIGENETIC CELL MODIFICATIONS MEDIATED BY HEPATITIS C VIRUS J. Sheldon1 , A. Madejon2 , F. Antonucci3 , C. Perales1 , J. Garc´ıa3 1 Samaniego2 , E. Domingo1 , A. Sanchez-Pacheco ´ . Department of Virology and Microbiology, Centro de Biologia Molecular Severo Ochoa, CSIC-Universidad Autonoma de Madrid, 2 Hepatology Unit, Hospital Carlos III. CIBERehd, 3 Department of Biochemistry, Universidad Autonoma de Madrid-Instituto de Investigaciones Biom´edicas A. Sols (UAM-CSIC), Madrid, Spain E-mail: [email protected] Background and Aims: Epigenetic regulation mechanisms are essential for the modulation of eukaryotic gene expression. These processes are regulated by nuclear enzymatic activities such as kinases, acetyltranferases or demethylases. There are not too much knowledges about if HCV, an infectious agent with a cytoplasmic replicative cycle can induce epigenetic modifications in the host cell. Material and Methods: Huh 7.5 cells were infected with Jc1 virus supernatant (HCV cell culture infectious clone) or DMEM media and passaged until >90% cells were infected. In these cells several epigenetic markers were analyzed. Phosphorylation of histone-3 in serine-10 position (H3Ser10ph) and methylation of histone 4 in lysine-20 residue (H4K20me3) was tested by western blot analysis using specific antibodies against the histone modifications. The expression levels of EGFR and IGBP3 genes were tested by reverse-transcription/quantitative-real time PCR. All the assays were performed in triplicate in independent experiments Results: Data obtained showed that the infection of the Huh 7.5 cells with the Jc1 is associated with the induction of several epigenetic changes. The levels of H3Ser10ph were higher in the mock compared to the infected cells. In addition H4K20me3 levels were significantly reduced (70%) in the infected cells when compared to the mock. The level of EGFR mRNA expressed was also 75% lower in the infected cells, while the IGBP3 expression didn’t change by Jc1 infection. Conclusions: Our data shows that HCV infection can induce epigenetic modifications within the host cell. We found both H3Ser10ph and H4K20me3 levels were reduced in the presence of HCV infection. These epigenetic modifications correlated with changes in the expression levels of EGFR gene. Since HCV replicates in the cytoplasm, this induction process should occur by indirect regulation of nuclear enzymatic activities implicated in the epigenetic regulation. 815 IDENIX NS5A HCV REPLICATION INHIBITORS WITH LOW PICOMOLAR, PAN-GENOTYPIC IN VITRO ANTIVIRAL ACTIVITY C.B. Dousson1 , C. Chapron2 , D. Standring2 , J.P. Bilello2 , J. McCarville2 , M. La Colla2 , M. Seifer2 , C. Parsy1 , D. Dukhan1 , C. Pierra1 , D. Surleraux1 . 1 Idenix Pharmaceuticals, Montpellier, France; 2 Idenix Pharmaceuticals, Cambridge, MA, USA E-mail: [email protected] Background: The NS5A protein is essential to the life cycle of the hepatitis C virus (HCV) and provides an important antiviral drug
Journal of Hepatology 2011 vol. 54 | S209–S361
POSTERS target. Here we describe the in vivo and in vitro characterization of NS5A-targeted compounds from different structural series that inhibit the replication of multiple HCV genotypes with picomolar (pM) potencies. Methods: Replicons were generated containing the NS5A protein sequence from HCV of genotypes 1a, 1b, 2a, 3a and 4a. Compounds with potent activity against the NS5A genotype 1 replicons were further evaluated against genotypes 2–4. Selected compounds were examined for cell permeability, protein-binding, and PK characteristics in rodents and monkeys. Results: Compounds potently active against HCV replicons of multiple genotypes were identified from several distinct molecular scaffolds. The in vitro activities against genotypes 1–4 of three such compounds from two scaffold classes are compared to BMS790052 in the table. EC50 (pM)
BMS790052 IDX380 IDX719 IDX210
1a
1b
2a
3a
4a
87 28 9 175
15 8 4 11
156 41 21 14
429 62 27 24
33 10 5 6
The following pharmacologic characteristics, generally comparable to BMS790052, were observed: high selectivity indices (CC50 s ~1 million-fold >EC50 s); low protein binding (<10-fold shift in EC50 in 40%-45% human serum); low to moderate permeability in a Caco-2 assay with no P-glycoprotein efflux, and low to moderate clearance in monkey and human primary hepatocytes. IDX210 showed similar bioavailability in cynomolgus monkeys compared to BMS790052. Plasma drug levels remained far above the EC50 values 24 h after administration of a single 10 mg/kg oral dose in the rat and monkey. Conclusions: Novel NS5A compounds were identified with low pM potency against HCV genotypes 1a, 1b, 2a, 3a and 4a, high selectivity indices in vitro and favorable pharmacokinetic properties. Extensive in vitro and in vivo evaluations of these new NS5A compounds are in progress to identify the preferred clinical drug candidate. 816 DYSREGULATION OF HEPATOCYTE SIGNAL TRANSDUCTION, CIAP1 AND APOPTOSIS BY HEPATITIS C VIRUS K. El Khobar1 , R. Chin1,2 , L. Earnest-Silveira1,2 , U. Nachbur3 , J. Silke3 , C.-T. Bock4 , J. Torresi1,2,5 . 1 Medicine, Austin Hospital, The University of Melbourne, 2 Austin Centre for Infection Research, Austin Hospital, Heidelberg, 3 Biochemistry, LaTrobe University, Bundoora, VIC, Australia; 4 Virology, Robert Koch Institute, Berlin, Germany; 5 Infectious Diseases, Austin Hospital, Heidelberg, VIC, Australia E-mail: [email protected] Introduction: Hepatitis C virus dysregulates JAK/STAT/SOCS, ERK, AKT/mTOR and apoptotic pathways. The NS5A protein blocks STAT3 activation and interacts with PI3K to activate the PI3K-AKT pathway and block hepatocyte apoptosis. The core and E2 envelope proteins activate MAPK/ERK signaling resulting in cellular proliferation. These changes are associated with up-regulation of TGFb and promotion of liver fibrosis. The effect of hepatitis C on the cellular inhibitor of apoptosis protein 1 (cIAP1) has not been previously reported. In this study, we determined the effects of HCV core-E1E2 and NS3-NS5b protein expression on JAK/STAT, ERK, AKT/mTOR and cIAP1 in hepatocytes. Methods: Recombinant adenoviruses expressing HCV genotype 1a CoreE1E2 and NS3 to 5b were used to infect Huh7 cells and primary mouse hepatocytes. Control infections were performed with rAdGFP. Western immunoblots were performed to determine the effects on total and phosphorylated STAT1, STAT3, AKT, mTOR and ERK and total SOCS3. We also investigated the effects on
activation of caspase 3, cleaved PARP, cIAP1 expression and oxidative stress. Results: Infection with rAdHCV-NS3–5b and rAdHCV-CE1E2/ rAdHCV-NS3–5b co-infection produced a 7-fold decline in pSTAT3 levels between 24 h and 72 h pi. A sustained 13fold increase in SOCS3 expression was observed in coinfected hepatocytes. Phospho-ERK expression was increased 15- and 30-fold in rAdHCVCoreE1E2 and rAdHCVNS35b-infected cells respectively, compared to >100-fold increase in co-infected hepatocytes. Phospho-AKT levels were increased 7 to 12-fold in rAdHCV-CoreE1E2, rAdHCVNS35b- and co-infected hepatocytes. Correspondingly, mTOR levels were also increased. Levels of cleaved caspase 3 and PARP were significantly increased and infected hepatocytes were sensitized to the cytotoxic effects of TNF-alpha. This was accompanied by a decline in levels of cIAP1. Co-infected cells also showed evidence of oxidative stress with increased levels of the GRP94 protein. Discussion: The expression of HCV structural and non-structural proteins in primary hepatocytes results in dysregulation of ERK, and AKT signalling, down regulation of pSTAT3 and increased SOCS3. HCV infection decreased levels of cIAP1 with a concomitant increase in cleaved caspase 3 and PARP consistent with increased apoptosis. These changes are likely to contribute to persistent inflammation, activation of stellate cells and fibrogenesis. 817 CD4+ CD25+ CD127LOW REGULATORY T CELLS: A POTENTIAL RESERVOIR OF HEPATITIS C VIRUS REPLICATION? T. Vausselin1 , C. Miroux1 , C. Wychowski2 , J. Dubuisson2 , F. Conti3 , V. Pancre´ 1 , N. Delhem1 . 1 Institut de Biologie de Lille UMR8161, 2 Centre d’Immunologie et d’Infection de Lille, 3 Hˆ opital Saint Antoine, Lille, France E-mail: [email protected] Background and Aim: Although hepatocytes are commonly considered to be the main reservoir of Hepatitis C Virus (HCV) replication, detection of viral RNA in some immune cell introduced the new notion of HCV lymphotropism. Moreover, the inevitable hepatitis C recurrence after orthotopic liver transplantation (LT) supports this hypothesis. Previously, we have showed that enhanced regulatory T cells (Treg) frequency was correlated with the liver fibrosis progression and the severity of the recurrence after LT. In this context, we have investigated the possibility for Treg cells to directly interact with HCV and to potentially support the virus replication. Methods: Human CD4+ CD25+ CD127Low cells were isolated from healthy donors and the presence of HCV receptors (CD81, SRB1, LDL-R and Claudin-1) were analyzed by Q-PCR, FACS and immunofluorescence. HCV internalization was assessed in detection of the viral glycoproteins E1 and E2 and the non-structural protein NS5A by Immunofluorescence assays, after Treg infection with HCVcc (JFH-1). Finally, the viral replication was measured by the presence of RNA negative-strand (sensitive and specific RT-PCR). Results: Freshly isolated Treg cells express all of the HCV specific receptors tested, both at RNA or protein level. Interestingly, it appears that HCV could invade Treg cells, since E1, E2 and NS5A have been observed within the cytoplasm. Moreover, HCV-RNA negative-strand has been detected in Treg samples, suggesting a potential viral replication in these cells. Conclusions: We showed for the first time that Treg cells express HCV receptors allowing viral entry and internalization. Treg cells seem also able to support HCV replication, which could explain the viral persistence during hepatitis C infection and the recurrence after liver transplantation.
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observed amongst HCV ELISpot positive participants (28.7 per 100 py, 95% CI 12.9, 63.8) and those who were HCV ELISpot negative (19.4 per 100 py, 95% CI 8.1, 46.6). Genetic variations in the IL28B region previously identified as associated with viral clearance were no more prevalent amongst participants who were ELISpot positive and anti-HCV antibody negative, HCV RNA negative than those who were anti-HCV antibody positive or HCV RNA positive. Conclusions: HCV-specific cellular immune responses were observed in more than one third of anti-HCV negative IDUs but our data contain no evidence that this subgroup of IDUs is protected from future HCV infection. 814 INDUCTION OF EPIGENETIC CELL MODIFICATIONS MEDIATED BY HEPATITIS C VIRUS J. Sheldon1 , A. Madejon2 , F. Antonucci3 , C. Perales1 , J. Garc´ıa3 1 Samaniego2 , E. Domingo1 , A. Sanchez-Pacheco ´ . Department of Virology and Microbiology, Centro de Biologia Molecular Severo Ochoa, CSIC-Universidad Autonoma de Madrid, 2 Hepatology Unit, Hospital Carlos III. CIBERehd, 3 Department of Biochemistry, Universidad Autonoma de Madrid-Instituto de Investigaciones Biom´edicas A. Sols (UAM-CSIC), Madrid, Spain E-mail: [email protected] Background and Aims: Epigenetic regulation mechanisms are essential for the modulation of eukaryotic gene expression. These processes are regulated by nuclear enzymatic activities such as kinases, acetyltranferases or demethylases. There are not too much knowledges about if HCV, an infectious agent with a cytoplasmic replicative cycle can induce epigenetic modifications in the host cell. Material and Methods: Huh 7.5 cells were infected with Jc1 virus supernatant (HCV cell culture infectious clone) or DMEM media and passaged until >90% cells were infected. In these cells several epigenetic markers were analyzed. Phosphorylation of histone-3 in serine-10 position (H3Ser10ph) and methylation of histone 4 in lysine-20 residue (H4K20me3) was tested by western blot analysis using specific antibodies against the histone modifications. The expression levels of EGFR and IGBP3 genes were tested by reverse-transcription/quantitative-real time PCR. All the assays were performed in triplicate in independent experiments Results: Data obtained showed that the infection of the Huh 7.5 cells with the Jc1 is associated with the induction of several epigenetic changes. The levels of H3Ser10ph were higher in the mock compared to the infected cells. In addition H4K20me3 levels were significantly reduced (70%) in the infected cells when compared to the mock. The level of EGFR mRNA expressed was also 75% lower in the infected cells, while the IGBP3 expression didn’t change by Jc1 infection. Conclusions: Our data shows that HCV infection can induce epigenetic modifications within the host cell. We found both H3Ser10ph and H4K20me3 levels were reduced in the presence of HCV infection. These epigenetic modifications correlated with changes in the expression levels of EGFR gene. Since HCV replicates in the cytoplasm, this induction process should occur by indirect regulation of nuclear enzymatic activities implicated in the epigenetic regulation. 815 IDENIX NS5A HCV REPLICATION INHIBITORS WITH LOW PICOMOLAR, PAN-GENOTYPIC IN VITRO ANTIVIRAL ACTIVITY C.B. Dousson1 , C. Chapron2 , D. Standring2 , J.P. Bilello2 , J. McCarville2 , M. La Colla2 , M. Seifer2 , C. Parsy1 , D. Dukhan1 , C. Pierra1 , D. Surleraux1 . 1 Idenix Pharmaceuticals, Montpellier, France; 2 Idenix Pharmaceuticals, Cambridge, MA, USA E-mail: [email protected] Background: The NS5A protein is essential to the life cycle of the hepatitis C virus (HCV) and provides an important antiviral drug
Journal of Hepatology 2011 vol. 54 | S209–S361
POSTERS target. Here we describe the in vivo and in vitro characterization of NS5A-targeted compounds from different structural series that inhibit the replication of multiple HCV genotypes with picomolar (pM) potencies. Methods: Replicons were generated containing the NS5A protein sequence from HCV of genotypes 1a, 1b, 2a, 3a and 4a. Compounds with potent activity against the NS5A genotype 1 replicons were further evaluated against genotypes 2–4. Selected compounds were examined for cell permeability, protein-binding, and PK characteristics in rodents and monkeys. Results: Compounds potently active against HCV replicons of multiple genotypes were identified from several distinct molecular scaffolds. The in vitro activities against genotypes 1–4 of three such compounds from two scaffold classes are compared to BMS790052 in the table. EC50 (pM)
BMS790052 IDX380 IDX719 IDX210
1a
1b
2a
3a
4a
87 28 9 175
15 8 4 11
156 41 21 14
429 62 27 24
33 10 5 6
The following pharmacologic characteristics, generally comparable to BMS790052, were observed: high selectivity indices (CC50 s ~1 million-fold >EC50 s); low protein binding (<10-fold shift in EC50 in 40%-45% human serum); low to moderate permeability in a Caco-2 assay with no P-glycoprotein efflux, and low to moderate clearance in monkey and human primary hepatocytes. IDX210 showed similar bioavailability in cynomolgus monkeys compared to BMS790052. Plasma drug levels remained far above the EC50 values 24 h after administration of a single 10 mg/kg oral dose in the rat and monkey. Conclusions: Novel NS5A compounds were identified with low pM potency against HCV genotypes 1a, 1b, 2a, 3a and 4a, high selectivity indices in vitro and favorable pharmacokinetic properties. Extensive in vitro and in vivo evaluations of these new NS5A compounds are in progress to identify the preferred clinical drug candidate. 816 DYSREGULATION OF HEPATOCYTE SIGNAL TRANSDUCTION, CIAP1 AND APOPTOSIS BY HEPATITIS C VIRUS K. El Khobar1 , R. Chin1,2 , L. Earnest-Silveira1,2 , U. Nachbur3 , J. Silke3 , C.-T. Bock4 , J. Torresi1,2,5 . 1 Medicine, Austin Hospital, The University of Melbourne, 2 Austin Centre for Infection Research, Austin Hospital, Heidelberg, 3 Biochemistry, LaTrobe University, Bundoora, VIC, Australia; 4 Virology, Robert Koch Institute, Berlin, Germany; 5 Infectious Diseases, Austin Hospital, Heidelberg, VIC, Australia E-mail: [email protected] Introduction: Hepatitis C virus dysregulates JAK/STAT/SOCS, ERK, AKT/mTOR and apoptotic pathways. The NS5A protein blocks STAT3 activation and interacts with PI3K to activate the PI3K-AKT pathway and block hepatocyte apoptosis. The core and E2 envelope proteins activate MAPK/ERK signaling resulting in cellular proliferation. These changes are associated with up-regulation of TGFb and promotion of liver fibrosis. The effect of hepatitis C on the cellular inhibitor of apoptosis protein 1 (cIAP1) has not been previously reported. In this study, we determined the effects of HCV core-E1E2 and NS3-NS5b protein expression on JAK/STAT, ERK, AKT/mTOR and cIAP1 in hepatocytes. Methods: Recombinant adenoviruses expressing HCV genotype 1a CoreE1E2 and NS3 to 5b were used to infect Huh7 cells and primary mouse hepatocytes. Control infections were performed with rAdGFP. Western immunoblots were performed to determine the effects on total and phosphorylated STAT1, STAT3, AKT, mTOR and ERK and total SOCS3. We also investigated the effects on
activation of caspase 3, cleaved PARP, cIAP1 expression and oxidative stress. Results: Infection with rAdHCV-NS3–5b and rAdHCV-CE1E2/ rAdHCV-NS3–5b co-infection produced a 7-fold decline in pSTAT3 levels between 24 h and 72 h pi. A sustained 13fold increase in SOCS3 expression was observed in coinfected hepatocytes. Phospho-ERK expression was increased 15- and 30-fold in rAdHCVCoreE1E2 and rAdHCVNS35b-infected cells respectively, compared to >100-fold increase in co-infected hepatocytes. Phospho-AKT levels were increased 7 to 12-fold in rAdHCV-CoreE1E2, rAdHCVNS35b- and co-infected hepatocytes. Correspondingly, mTOR levels were also increased. Levels of cleaved caspase 3 and PARP were significantly increased and infected hepatocytes were sensitized to the cytotoxic effects of TNF-alpha. This was accompanied by a decline in levels of cIAP1. Co-infected cells also showed evidence of oxidative stress with increased levels of the GRP94 protein. Discussion: The expression of HCV structural and non-structural proteins in primary hepatocytes results in dysregulation of ERK, and AKT signalling, down regulation of pSTAT3 and increased SOCS3. HCV infection decreased levels of cIAP1 with a concomitant increase in cleaved caspase 3 and PARP consistent with increased apoptosis. These changes are likely to contribute to persistent inflammation, activation of stellate cells and fibrogenesis. 817 CD4+ CD25+ CD127LOW REGULATORY T CELLS: A POTENTIAL RESERVOIR OF HEPATITIS C VIRUS REPLICATION? T. Vausselin1 , C. Miroux1 , C. Wychowski2 , J. Dubuisson2 , F. Conti3 , V. Pancre´ 1 , N. Delhem1 . 1 Institut de Biologie de Lille UMR8161, 2 Centre d’Immunologie et d’Infection de Lille, 3 Hˆ opital Saint Antoine, Lille, France E-mail: [email protected] Background and Aim: Although hepatocytes are commonly considered to be the main reservoir of Hepatitis C Virus (HCV) replication, detection of viral RNA in some immune cell introduced the new notion of HCV lymphotropism. Moreover, the inevitable hepatitis C recurrence after orthotopic liver transplantation (LT) supports this hypothesis. Previously, we have showed that enhanced regulatory T cells (Treg) frequency was correlated with the liver fibrosis progression and the severity of the recurrence after LT. In this context, we have investigated the possibility for Treg cells to directly interact with HCV and to potentially support the virus replication. Methods: Human CD4+ CD25+ CD127Low cells were isolated from healthy donors and the presence of HCV receptors (CD81, SRB1, LDL-R and Claudin-1) were analyzed by Q-PCR, FACS and immunofluorescence. HCV internalization was assessed in detection of the viral glycoproteins E1 and E2 and the non-structural protein NS5A by Immunofluorescence assays, after Treg infection with HCVcc (JFH-1). Finally, the viral replication was measured by the presence of RNA negative-strand (sensitive and specific RT-PCR). Results: Freshly isolated Treg cells express all of the HCV specific receptors tested, both at RNA or protein level. Interestingly, it appears that HCV could invade Treg cells, since E1, E2 and NS5A have been observed within the cytoplasm. Moreover, HCV-RNA negative-strand has been detected in Treg samples, suggesting a potential viral replication in these cells. Conclusions: We showed for the first time that Treg cells express HCV receptors allowing viral entry and internalization. Treg cells seem also able to support HCV replication, which could explain the viral persistence during hepatitis C infection and the recurrence after liver transplantation.
Journal of Hepatology 2011 vol. 54 | S209–S361
S327