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PubMed1995

Antibody to hepatitis C virus second envelope (HCV-E2) glycoprotein: a new marker of HCV infection closely associated with viremia.

The second envelope protein (E2) of the hepatitis C virus (HCV) was cloned and expressed in Chinese hamster ovary (CHO) cells. This E2 glycoprotein was purified using ion exchange and lectin chromatography and used to construct an enzyme immunoassay for HCV E2 antibodies. The assay was shown to have good specificity, and detection of E2 antibodies was positively correlated (97.3%) to the presence of HCV RNA in serum and plasma. A high concordance between HCV 2.0 and E2 EIA reactivities was also observed. E2 antibody was the first serological marker to appear in 3/5 HCV seroconversion panels. This work demonstrated that 42.4% of core and 15.4% of NS3 indeterminate specimens also contained antibodies to E2, suggesting that HCV infection had occurred in these individuals. The E2 antibody assay was used to evaluate HCV 2.0 EIA-positive, HCV 3.0 EIA-negative plasma donors with indeterminate reactivity on RIBA HCV 2.0 or MATRIX HCV 1.0. Several HCV 3.0-negative specimens were shown to contain E2 antibodies in addition to an original indeterminate serological marker, primarily core. It is concluded that anti-E2 is a useful marker for determining HCV infection, and that the presence of antibodies to two nonoverlapping viral gene products suggests true HCV exposure. New HCV 3.0 blood screening tests should detect HCV 2.0-positive donors who present with an indeterminate pattern by RIBA or MATRIX and who also carry E2 antibodies.

Animals↗

PubMed1995

HCV replication in mononuclear cells stimulates anti-HCV-secreting B cells and reflects nonresponsiveness to interferon-alpha.

Recently, it was demonstrated in chronic hepatitis C that the release of IgG and IgM anti-HCV antibodies by mononuclear cells (PBMCs) correlated with inflammatory activity, HCV persistence in serum, and negative outcome from antiviral therapy. Thus, persistent antigenic stimulation of the antibody-secreting B cells has been suggested. In this study, PBMCs were derived from 13 patients with chronic hepatitis C. Nucleic acids were extracted by the guanidine-thiocyanate-method, and plus- and minus-stranded HCV-RNAs were determined using primers from the 5'-untranslated region of HCV. Simultaneously, unstimulated PBMCs were cultured for 8 days and anti-HCV antibodies were detected in the supernatants by EIA and RIBA. Seven patients (53.8%) had both plus- and minus-stranded HCV-RNA in PBMCs, while anti-HCV antibodies were secreted in vitro. One of 2 patients with plus- but not minus-stranded HCV-RNA in PBMCs was anti-HCV positive in vitro, whereas 4 patients without HCV-infected PBMCs were anti-HCV negative in vitro. Eight patients received antiviral therapy with interferon-alpha 2b. Four nonresponders and 1 partial responder had plus- and minus-stranded HCV-RNA in PBMCs and anti-HCV secretion in vitro. On the other hand, 2 complete responders and another partial responder showed neither HCV infection of PBMCs nor anti-HCV secretion in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed2006

Virus-specific T-cell responses associated with hepatitis C virus (HCV) persistence in the liver after apparent recovery from HCV infection.

Hepatitis C virus (HCV) RNA persistence in the liver has been described even after apparent resolution of HCV infection. Because T-cell reactivity plays a role in recovery from HCV infection, virus-specific T-cell responses were investigated in apparently recovered individuals in whom hepatic HCV RNA persistence was documented: 15 sustained virological responders to interferon (IFN)-treatment and 9 asymptomatic aviremic anti-HCV carriers. HCV-specific CD4(+) T-cell proliferative responses were detected significantly more often in apparently recovered individuals (sustained virological responders: 60%; asymptomatic anti-HCV carriers: 66%) compared with 50 chronic hepatitis C patients (28%; P < 0.05). However, T-cell frequencies and numbers tended to decline over time and the number of HCV proteins targeted by CD4(+) T-cell proliferative responses was limited. Interestingly, liver viral load correlated inversely with virus-specific immune responses. Thus, CD4(+) T-cell responders showed significantly lower hepatic HCV RNA levels (P < 0.05). HCV-specific IFN-gamma-secreting CD4(+) T-cells were not detected in all the apparently recovered patients although they were found significantly more often compared with chronic hepatitis C patients (P < 0.05). Also, HCV NS3-specific CD8(+) T-cells were detected in 11 HLA-A2-positive apparently recovered individuals (8 sustained virological responders and 3 asymptomatic anti-HCV carriers); T-cell frequencies tended to be greater in those patients who had lower hepatic viral levels. In conclusion, HCV-specific T-cells are detectable in apparently recovered individuals in whom HCV RNA can persist in the liver indicating that HCV replication may be prolonged in the face of an insufficient or inadequate virus-specific CD4(+) and CD8(+) T-cell response.

Adult↗

PubMed1998

Evidence that both HIV and HIV-induced immunodeficiency enhance HCV replication among HCV seroconverters.

The objective of this retrospective cohort study is to assess the mechanism by which human immunodeficiency virus type 1 (HIV) influences hepatitis C virus (HCV) replication in injecting drug users. Virological (HCV and HIV RNA levels) and immunological (CD4+, CD8+ cell counts, and anti-CD3 reactivity) parameters were determined in 19 HCV seroconverters in sequential samples over a period of 1 to 9 years. Among these subjects, 10 were HIV-seronegative (HIVneg), 4 were HIV-seropositive (HIVpos), and 5 seroconverted for HIV (HIVsc) during the observation period. HCV RNA levels were higher in HIVpos subjects than in HIVneg subjects. In subjects seroconverting for HIV, HCV, RNA levels increased significantly immediately after HIV seroconversion (P < 0.0001), while they remained stable over time in HIVpos and HIVneg subjects. HCV RNA correlated inversely with CD4+ cell counts in both the HIVpos population (R = -0.22, P < 0.05) and the HIVneg population (R = -0.45, P < 0.0001). In addition, when subjects were stratified according to CD4+ cell counts a significant difference was found in HCV RNA levels between HIVpos and HIVneg subjects with CD4+ cell counts > 500 cells/microliter (P = 0.001), but not in the population with CD4+ cell counts < 500 cells/microliter. In no population was a correlation found between HCV RNA levels and CD8+ cell counts or anti-CD3 reactivity. Both HIV infection and CD4+ cell counts are apparently associated with HCV RNA levels. The direct association, independent of CD4+ cell counts, between HIV infection and HCV replication appears to be stronger than the association between HIV-induced CD4+ cell decline and HCV replication. We conclude that (i) HCV replication is in some way directly influenced by the presence of HIV; (ii) HCV-specific host immunity controls, in part, HCV replication; and (iii) HCV replication increases when the immune system is impaired by HIV.

CD4 Lymphocyte Count↗

PubMed2004

Improved detection of HCV Infection in hemodialysis patients using a new HCV RNA qualitative assay: experience of a transplant center.

BACKGROUND: Hepatitis C virus (HCV) is frequently a silent infection in hemodialysis (HD) patients with a prevalence of 8-10%. Improving HCV detection in this population prior to transplantation is critical both for infection control and optimal patient care. OBJECTIVES: To assess the current HCV testing practice of the National Institute for Transplantation (PCR testing of enzyme immunoassay (EIA) positive HD patients) by evaluating a subset of EIA positive and EIA negative samples with the VERSANT HCV RNA Qualitative Assay based on transcription mediated amplification (HCV Qual (TMA)) (sensitivity < or = 9.6 IU/ml) and in-house PCR (HCV Qual (PCR)) (sensitivity approximately 149 IU/ml). STUDY DESIGN: 2321 HD patients were screened by Abbott HCV EIA 2.0. A subset of 80/169 E IA positive samples and 100/2152 EIA negative samples were tested by both assays. TMA/PCR discordant samples were genotyped. RESULTS: PCR and TMA gave concordant results in 67/80 (83.8%) of EIA positive samples. 11/80 (14.7%) were reactive by HCV Qual (TMA), but not by HCV Qual (PCR); 2/80 (2.7%) were reactive by HCV Qual (PCR), but not by HCV Qual (TMA). 2/100 (2%) EIA negative samples were reactive and 95/100 (95%) were non-reactive by both assays. Three (3%) were only HCV Qual (TMA) reactive. 11/14 TMA+/PCR-samples with sufficient volume were genotyped. CONCLUSIONS: HCV Qual (TMA) identified active HCV infection in more EIA positive and EIA negative patients than HCV Qual (PCR) and should be part of our testing algorithm.

Genotype↗

PubMed2003

Mucosal humoral immune response to hepatitis C virus E1/E2 surface glycoproteins and HCV shedding in saliva and cervicovaginal fluids from chronically HCV-infected patients.

BACKGROUND/AIMS: We herein focused on identifying biological factors possibly involved in non-parenteral transmission of hepatitis C virus (HCV), such as HCV excretion patterns and antibody-based immunity to the virus in saliva and/or cervicovaginal secretions (CVS). METHODS: Paired blood, saliva and cervicovaginal lavage samples were obtained from HCV-RNA plasma-positive hemoglobin (Hb) antigen and HIV-seronegative, HCV-seropositive males (n=13) and females (n=21). HCV-specific antibodies were detected by ELISA in paired samples, and HCV-RNA was detected in cell-free and cell-associated body fluids. RESULTS: Antibodies to E1 HCV surface glycoprotein of the IgG and IgA isotypes showed similar, but less pronounced, profiles as IgG and IgA to E2. HCV-specific IgG and IgA in mucosal fluids likely originated predominantly from the systemic compartment, because HCV-specific mucosal immunoglobulins involved primarily monomeric antibodies, including monomeric IgA, and because their specific activities for HCV antigens in corporeal fluids were similar to those in paired serum (Se). Viral shedding in saliva or CVS was restricted to cell-associated, non-replicating strand((+)) HCV-RNA in 42% (12 out of 28) of saliva and in 19% (four out of 21) of cervicovaginal fluids. CONCLUSIONS: The association in body fluids of HCV-specific IgG, and to a lesser extent IgA, directed to E1/E2 surface glycoproteins (which may block critical steps of virus-cell interactions), of undetectable free viral RNA, and of occasional non-replicating cell-associated HCV, suggests a resulting poor infectivity of saliva or cervicovaginal fluid in chronically HCV-infected individuals. Taken together, these observations provide the basis for the low risk of non-parenteral transmission of HCV infection.

Adult↗

PubMed1999

HCV seroreactivity and detection of HCV RNA in cirrhotics.

This study was carried out to determine the presence of HCV infection in cirrhotic patients and to compare their serologic profiles with detection of HCV RNA by RT-PCR. Liver function profiles were assessed and correlated with infection. Coinfection of HCV with HBV was studied in the Indian context Sera from 62 patients of biopsy confirmed cirrhosis and an equal number of asymptomatic controls were tested for HCV by two ELISA (third generation) kits and nested reverse transcription PCR using primers from the 5'NCR. Other tests included HBV serology (ELISA for HBsAg and anti-HBc) and liver function tests. Twenty-four (38.7%) cases were HCV infected, of which 17 (70.8%) had past exposure to HBV or were coinfected with HBV (either being chronically infected or carriers of HBV). There was no significant difference in the clinical and liver function profiles of HCV infected and uninfected cirrhotics. Similarly, no difference was observed in cases coinfected with both HCV and HBV compared with those infected with HCV alone. Although the difference between positivity of HCV RNA and of anti-HCV was not significant, HCV RNA and anti HCV were present together in only 7/62 (11.3%) cases. Thus testing for both antibody and HCV RNA would be more appropriate than either test alone. HBV infection was seen in about one-fourth of HCV infected cirrhotics. About one-fourth (25.8%) patients had non-B, non-C cirrhosis in whom testing for HCV variants and other recently characterized hepatitis viruses could be performed.

Adult↗

PubMed1997

Genetic analysis of the hepatitis C virus (HCV) genome from HCV-infected human T cells.

We recently established a cell culture system for the replication of hepatitis C virus (HCV) by using a human T cell leukaemia virus type I-infected cell line, MT-2, and showed that the quasi-species of the hypervariable region 1 observed in the original inoculum became homogeneous in MT-2 cells 10 days after inoculation of HCV. In this study, we obtained HCV cDNA clones covering the whole viral genome by RT-nested PCR using RNA from HCV-infected cloned MT-2C cells, which support viral replication more efficiently, at 12 days after inoculation. A total of 41 distinct HCV cDNA clones covering almost the whole viral genome were also isolated from a cDNA library derived from the original inoculum. Molecular evolutional analyses comparing the sequences of the HCV clones obtained from both sources revealed that the HCV populations became homogeneous in more than half of the compared regions. This finding suggests that limited HCV populations are able to replicate in MT-2C cells. In addition, we isolated cDNA clones containing a 3' X-tail sequence, which was recently identified as a bona fide 3' terminus of the HCV genome, in the HCV-infected MT-2C cells and confirmed that the nucleotide sequence of the 3' X-tail was highly conserved, suggesting its implication in HCV replication. Finally, on the basis of the sequences of HCV cDNA clones obtained from HCV-infected MT-2C cells, we determined the entire nucleotide sequence of the HCV genome containing the 3' X-tail as a candidate for an infectious HCV molecular clone.

Base Sequence↗

PubMed2003

The regulation of hepatitis C virus (HCV) internal ribosome-entry site-mediated translation by HCV replicons and nonstructural proteins.

Hepatitis C virus (HCV), the global leading cause of chronic liver disease, has a positive-sense, ssRNA genome that encodes a large polyprotein. HCV polyprotein translation is initiated by an internal ribosome-entry site (IRES) located at the 5' end of the viral genome, in a cap-independent manner, but the regulatory mechanism of this process remains poorly understood. In this study, we characterized the effect of HCV nonstructural proteins on HCV IRES-directed translation in both HCV replicon cells and transiently transfected human liver cells expressing HCV nonstructural proteins. Using bicistronic reporter gene constructs carrying either HCV or other viral IRES sequences, we found that the HCV IRES-mediated translation was specifically upregulated in HCV replicon cells. This enhancement of HCV IRES-mediated translation by the replicon cells was inhibited by treatment with either type I interferon or ribavirin, drugs that perturb HCV genome replication, suggesting that the enhancement is probably due to HCV-encoded protein function(s). Reduced phosphorylation levels of both eIF2alpha and eIF4E were observed in the replicon cells, which is consistent with our previous findings and indicates that the NS5A nonstructural protein may be involved in the regulatory mechanism(s). Indeed, transient expression of NS5A or NS4B in human liver cells stimulated HCV IRES activity. Interestingly, mutation in the ISDR of NS5A perturbed this stimulation of HCV IRES activity. All these results suggest, for the first time, that HCV nonstructural proteins preferentially stimulate the viral cap-independent, IRES-mediated translation.

Antiviral Agents↗

PubMed2005

HCV viremia is associated with drug use in young HIV-1 and HCV coinfected pregnant and non-pregnant women.

AIMS: Vertical transmission of HCV is increased among HIV-1/HCV coinfected women and is related to HCV viral load. In this study we assessed clinical and demographic factors associated with HCV viremia in a cohort of young pregnant and non-pregnant mothers coinfected with HIV-1. DESIGN: A cross-sectional clinic-based study nested within a prospective cohort study. METHODS: From 1988 to 2000, HIV-1 + pregnant and non-pregnant women with children followed in a large maternal, child and adolescent HIV-1 clinic were evaluated for HCV infection using EIA 3.0. HCV RNA levels were determined for HCV antibody + women using polymerase chain reaction. Demographic and clinical characteristics between HCV-RNA(+) and HCV-RNA(-) women and between pregnant and non-pregnant HIV-1/HCV coinfected women were compared using univariate and multivariate analyses. FINDINGS: Among 359 HIV-1(+) women, 84 (23%) were HCV-ab + and 49/84 (58%) had detectable HCV-RNA in plasma. Median age was 31. CD4 counts, HIV-1 RNA levels and demographic characteristics were similar for viremic and non-viremic women and pregnant and non-pregnant women. However, viremic women were more likely to report a history of (88% versus 43%; P < 0.001) or active injection drug use (AIDU) (83% versus 29%; P < 0.001). Logistic regression analysis showed that HCV viremia was associated significantly with AIDU (adjusted OR: 15.17; 95% CI: 3.56, 64.56) after adjusting for age, race, number of sexual partners, pregnancy status, CD4 counts and HIV-1 viral load. CONCLUSION: In this cohort of young HIV-1 and HCV coinfected women, HCV viremia was associated strongly with active injection drug use, perhaps due to reinfection or reactivation of HCV. Thus, careful evaluation for HCV infection and counseling related to drug use may be necessary.

Adult↗

PubMed2007

Induction of hepatitis C virus (HCV)-specific T cells by needle stick injury in the absence of HCV-viraemia.

BACKGROUND: The risk of hepatitis C virus (HCV) infection after occupational exposure is low with seroconversion rates between 0 and 5%. However, factors associated with natural resistance against HCV after needle stick injury are poorly defined. HCV-specific T-cell responses have been described in cross-sectional studies of exposed HCV-seronegative individuals. MATERIALS AND METHODS: In this study, we prospectively followed 10 healthcare professionals who experienced an injury with an HCV-contaminated needle. Blood samples were taken on the day or the day after the event and at different time points during follow-up for up to 32 months. HCV-specific T-cell responses were investigated directly ex vivo and in T-cell lines. RESULTS: None of the individuals became positive for HCV-RNA in serum tested with the highly sensitive transcription-mediated amplification (TMA)-assay or in peripheral blood mononuclear cells (PBMC). All of them remained anti-HCV negative throughout follow-up. At the time of injury, HCV-specific CD4+ T-cell responses were already detectable in two individuals and became detectable thereafter in three additional persons. Transient HCV-specific CD8+ T-cell responses developed in two HLA-A2 positive patients, which became negative until the most recent follow-up after 5 and 17 months, respectively. CONCLUSION: We demonstrate the development of HCV-specific T cells in HCV-exposed individuals after needle stick injury indicating subinfectious exposure to HCV. T-cell immunity against HCV may contribute to the low prevalence of HCV in medical healthcare professionals in Western countries.

Adult↗

PubMed2001

Nested restriction site-specific PCR to detect and type hepatitis C virus (HCV): a rapid method to distinguish HCV subtype 1b from other genotypes.

Genotypic differentiation of hepatitis C virus (HCV) has become an integral part of clinical management and epidemiologic studies of hepatitis C infections. Thus, it is extremely important in areas such as the Czech Republic, where current instrumentation and kits for assessing HCV infection are too costly for widespread use. We describe a new and relatively inexpensive method called nested restriction site-specific PCR (RSS-PCR) that generates a "fingerprint" pattern to represent an HCV genotype without the use of restriction endonucleases and that specifically differentiates HCV genotype 1b from the other HCV genotypes. The RSS-PCR method was applied directly to serum samples from patients with hepatitis C from the Czech Republic and from patients with known HCV genotypes from the United States. The method was validated by comparison of the subtype determined by RSS-PCR to the subtype determined from analysis of the 5' noncoding region (NC) or the nonstructural protein gene (NS5b) nucleotide sequence of HCV in these clinical samples. From 75 Czech samples containing HCV RNA, three distinct RSS-PCR patterns were observed; 54 were predicted to contain subtype 1b, 19 were predicted to contain subtype 1a, and 2 were predicted to contain subtype 3a. Among 54 samples predicted to contain HCV genotype 1b, all were confirmed by their 5' NC or NS5b sequences to be subtype 1b. Thus, both the sensitivity and specificity of the RSS-PCR test for the differentiation of HCV subtype 1b from the others were 100%. While the assay described here was designed to specifically differentiate HCV subtype 1b from the other HCV genotypes, the RSS-PCR method can be modified to differentiate any HCV genotype or subtype of interest. Its simplicity and speed may provide new opportunities to study the epidemiology of HCV infections and the relationship between HCV genotypes and clinical outcome by more laboratories throughout the world.

Genotype↗

PubMed2004

[The difference in distribution of HCV genotypes between patients infected with HCV by transfusion and non-transfusion routes].

OBJECTIVE: To investigate the HCV genotypes distribution in northern and southern cities in China and the difference between patients infected with HCV by transfusion and non-transfusion routes. METHODS: The HCV genotypes of the patients with chronic hepatitis C from 9 cities belonging to different regions were genotyped by the PCR products of 5 prime untranslated region NTR digested with restriction endonucleases, and the HCV genotypes distribution among different cities or between the patients infected with HCV through transfusion and other routes was analyzed. RESULTS: The HCV genotypes of 214 in 219 cases were determined; 197 patients were infected with monogenotype HCV. The major epidemic genotypes of HCV isolates in China were 1b (76.64%) and 2a (18.22%), but 5.14% of patients were infected with HCV belonging to genotype 3b and this was the first report that there is genotype 4a in China. The HCV genotype distribution was not different in northern and southern areas, but was significantly different between patients infected with HCV through transfusion and non-transfusion routes (P=0.036). In patients infected trough transfusion, the rates of monogenotype HCV infection and genotype 1b were 93.88% and 76.87%, respectively, which were higher than those (86.57% and 58.21%) in the patients infected with HCV through non-transfusion routes. The rate of patient infected with mixed genotype HCV strains in non-transfusion group was 13.43%, which was higher than that (6.12%) of patients in transfusion group. CONCLUSION: The HCV genotype distribution in northern and southern regions were similar, but was significantly different between the patients infected through transfusion and other routes.

5' Untranslated Regions↗

PubMed1997

Prevalence and significance of hepatitis C virus (HCV) genotypes in anti-HCV positive patients in northwest Croatia.

Hepatitis C virus (HCV) transmission is related to blood transfusion and transmission in hemodialysis (HD) units. The aim of the study was to investigate the distribution of HCV genotypes and routes of HCV transmission in three groups of anti-HCV positive patients in north-west Croatia. A total of 111 patients were studied. Patients were classified into three groups: 45 HD patients with a low percentage of anti-HCV positivity (group 1), 60 HD patients treated at another HD unit and with high percentage of anti-HCV positivity (group 2), and six anti-HCV positive patients with chronic hepatitis (group 3). Most of the HD patients were treated during the sama shift, but with separate equipment. Serum HCV RNA was detected by reverse transcription-polymerase chain reaction (RT-PCR). Genotyping of HCV isolates was performed with a line-probe assay. Patients of groups 1 and 2 did not show significant differences with regard to the clinical profile. In group 1, all anti-HCV positive patients were RT-PCR positive and all of them were infected with the same genotype (genotype 3). In group 2, 89% of anti-HCV positive patients were RT-PCR positive and all infected with the same genotype (genotype 1 b). In group 3, 50% anti-HCV positive chronic hepatitis patients were RT-PCR positive. Two of them were infected with genotype 1 b and one with genotype 4. The homogeneity of HCV genotypes in the patients from both HD units (groups 1 and 2), seemed to indicate nosocomial transmission of HCV, whereas viremia was found to be related to blood transfusion in all group 3 patients. The exact mechanism involved in the transmission of HCV in HD units remains to be discovered.

Croatia↗

PubMed1992

Prevalence of anti-HCV and HCV viremia in hemodialysis patients in Taiwan.

Hepatitis C viral infection in 125 hemodialysis patients from Taiwan was studied using a second-generation anti-HCV immunoassay (EIA II) (Abbott HCV 2.0 EIA) and the polymerase chain reaction (PCR) to detect the HCV RNA in the serum. A total of 59 patients (47.2%) were positive by EIA II. In comparison, the conventional C100-3 anti-HCV assay was positive in 40 (32.0%). HCV RNA was found in 47 patients (37.6%). Patients with elevated serum transaminase level had a higher positive rate of anti-HCV and HCV RNA. The dialysis time was longer for those patients positive for anti-HCV than for those who were negative. A total of 57 of the 59 EIA II-positive cases had a history of blood transfusion. The HBsAg status did not influence the anti-HCV positivity. Among the 59 EIA II-positive patients, 66.1% were also positive for HCV RNA, and of the 47 HCV RNA-positive cases 83.0% were positive for EIA II. It is concluded that the high prevalence of specific HCV infection and HCV viremia was present in these patients. Prevention of cross-contamination during dialysis and blood screening before transfusion are important for the control of HCV infection in these patients.

Adult↗

PubMed1992

Serum hepatitis C virus (HCV)-RNA and response to alpha-interferon in anti-HCV positive chronic hepatitis.

Hepatitis C virus (HCV) replication was assessed before and during alpha-interferon (IFN) treatment in 22 anti-HCV positive patients with posttransfusion or sporadic chronic hepatitis (CH). Eleven patients were "responders" and 11 patients "non-responders" to IFN. Thirteen anti-HCV negative healthy subjects and five anti-HCV negative patients with autoimmune CH served as controls. Serum HCV-RNA was detected by the polymerase chain reaction (PCR) in all untreated anti-HCV positive patients but in none of the anti-HCV negative subjects. PCR primers from the 5'-noncoding (NC) region were more sensitive than primers from a non-structural (NS5) region in detecting HCV-RNA (21/22, 95% vs. 7/22, 32%, respectively). Positive strand HCV-RNA titre and positivity rate for the negative strand were similar in responders and non-responders before IFN treatment, as well as anti-c100-3 titre by enzyme-linked immunosorbent assay (ELISA), and anti-5-1-1, anti-c33c, anti-c22 positivity rate by immunoblot assay (RIBA). HCV-RNA positivity by both NC and NS primers was more frequent before IFN among responders. During IFN treatment, serum HCV-RNA was detectable, mostly at low titres, in 1 (NC positive) of the 11 responders and in 9 (4 NS positive and 5 NC positive) of the 11 non-responders. Among the four non-responders who were NS positive during IFN, three were NC positive before IFN. Serum HCV-RNA was always found in our post-transfusion or sporadic anti-HCV positive patients with CH. Viraemia generally decreased during IFN treatment, but no available HCV markers clearly distinguished responders from non-responders before IFN treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed2004

Polypyrimidine tract-binding protein (PTB) inhibits Hepatitis C virus internal ribosome entry site (HCV IRES)-mediated translation, but does not affect HCV replication.

Polypyrimidine tract-binding protein (PTB) has previously been shown to affect Hepatitis C virus (HCV) IRES-mediated translation. In the present study we investigated the functional role of PTB for HCV translation, replication and chronic HCV infection. Bicistronic HCV IRES reporter plasmids and two different subgenomic replicons (bicistronic: pHCVrep1bBB7 (s1179I); monocistronic: pFK1-389/hyg-ubi/NS3-3'/5.1) were used to analyze the effects of PTB. Following transfection of plasmids expressing PTB RNA in sense or antisense orientation, translational activity and HCV RNA were analyzed by luciferase assay, quantitative real-time RT-PCR and northern blot analysis. Additionally, in liver tissue (n = 53) intrahepatic PTB RNA levels were determined by quantitative real-time RT-PCR. Significant inhibition of HCV IRES activity up to 42.6% was observed upon PTB sense RNA expression for HCV IRES reporter plasmids, while translational activity was enhanced up to 63.8% for PTB antisense RNA expression. In the HCV replicons PTB did not affect replication and no correlation was found between intrahepatic PTB mRNA levels and serum HCV RNA or histological changes in liver tissue of HCV infected patients. Although PTB inhibits HCV IRES-mediated translation from bicistronic reporter constructs, data obtained from two subgenomic HCV replicons and liver tissue do not indicate a significant role of PTB for HCV replication and chronic HCV infection.

5' Untranslated Regions↗

PubMed2006

Relationship between hepatitis C virus (HCV) and insulin resistance, endothelial perturbation, and platelet activation in HIV-HCV-coinfected patients under highly active antiretroviral treatment.

Insulin resistance is associated with highly active antiretroviral therapy in HIV-infected patients, and the risk of developing insulin resistance is increased in hepatitis C virus (HCV)-infected patients. The aim of the present study was to determine whether hepatitis C virus infection constitutes an additional risk factor for insulin resistance or other prothrombotic conditions in HIV-HCV coinfected patients under highly active antiretroviral therapy. One hundred eighteen HIV-infected patients were studied: 50 who had no history of anti-HIV treatment and 68 who were receiving therapy with highly active antiretroviral treatment. The treatment-naive group consisted of 35 HCV-negative subjects and 15 HCV-positive ones. Within the treated group, 50 patients were HCV negative and 18 were HCV positive. For each patient, the lipid profile was determined and the following values measured: glucose, soluble P-selectin (as a marker of platelet activation), soluble thrombomodulin, von Willebrand factor and soluble vascular cell adhesion molecule-1 (as endothelial markers), and insulin resistance. No significant difference (p>0.05) for any variable was found among subjects with or without HCV coinfection in the treatment-naïve group. Among patients under highly active antiretroviral therapy, however, those with HCV coinfection showed higher values (p<0.05) for insulin resistance (homeostasis model assessment value: 2.65 vs. 1.79), glucose (93 vs. 86 mg/dl), endothelial markers (von Willebrand factor, 204 vs. 123%; soluble vascular cell adhesion molecule-1, 650 vs. 482 ng/ml), and platelet activation marker (soluble P-selectin, 78 vs. 51 ng/ml) in parallel with lower CD4+ cells counts (289 vs. 402 cells/mm3) and higher HIV-1 viral loads (305 vs. 50 copies/ml) compared to patients without HCV coinfection. Glucose, soluble P-selectin, and von Willebrand factor were independently related to HCV infection. The presence of HCV coinfection during HIV treatment was closely related to higher values of insulin resistance, to activated platelets, and to endothelial perturbation in parallel with lower CD4+ cell counts and higher HIV-1 viral loads compared to patients without HCV coinfection. On the basis of these results, it may be preferable to treat HCV infection prior to initiating treatment for HIV infection in HIV-HCV-coinfected patients.

Adult↗