Antibodies to different virus antigens in patients with chronic hepatitis C

A total of 176 hospital patients with chronic hepatitis C (CHC), among them 110 males and 66 females, were examined. The spectrum of antibodies to four hepatitis C virus (HCV) proteins (core, NS3, NS4, NS5) and in 142 patients --IgM antibodies to HCV (anti-HCV IgM) were determined. In 92% of the CHC patients antibodies to core, NS3 and NS4 proteins were simultaneously detected. Differences in the detection of antibodies to HCV in males and females were not statistically reliable. In CHC patients aged up to 20 years anti-NS4 and anti-NS5 were less frequently detected. Among males of different age groups reliable differences in the detection rate of anti-NS5 were registered, while among females of different age groups no such differences were observed. With the increase of age these antibodies were detected somewhat more often. In females over 60 years anti-HCV IgM occurred more often than in males of the same age. The levels of alanine aminotransferase (ALT) were higher in persons with the presence of anti-NS5 and anti-HCV IgM than in persons with their absence. In all groups of CHC patients with biochemical activity and liver cirrhosis the detection rate of anti-HCV IgM was significantly higher than in patients with normal ALT activity. The antibody spectrum with the simultaneous absence of HCV IgM and anti-NS5, while found to contain antibodies to other HCV antigens, was registered significantly less frequently in patients with moderate and high CHC activity and the liver cirrhosis induced by HCV infection.     

Co-infections with hepatitis G and TT virus in patients with chronic hepatitis C in Hungary

The significance of co-infections with novel hepatitis viruses Hepatitis G (GBV-C, HGV) and TT virus (TTV) in chronic hepatitis C is not clear. We determined the prevalence of HGV RNA and TTV DNA in chronic hepatitis C patients and in asymptomatic hepatitis C virus (HCV) carriers, and assessed the influence of these agents on the course of HCV infection. Seventy-seven patients with chronic hepatitis C--50 of them treated with interferon (IFN)--and 33 HCV carriers with normal alanine aminotransferase have been investigated. Previous HBV infection was detected by testing serum HBsAg and aHBc. HGV RNA and TTV DNA were detected by PCR. In the healthy population, the prevalence of anti-HCV was 0.3%, HGV RNA 8.0% and TTV DNA 18.5%. In chronic hepatitis C HGV RNA occurred in 9.09% and TTV DNA in 40.25% of cases. In IFN-treated patients with sustained remission, the frequency of TTV was 20% vs. 45.7% found in non-responders. Among asymptomatic HCV-carriers, the prevalence of HGV RNA was 9.09% and TTV DNA 75.7%. Neither HGV RNA nor TTV DNA had apparent effect on the HCV infection. TTV was detected with the lowest frequency in persons with sustained remission due to IFN, suggesting antiviral effect of IFN on TTV.     

Therapy-induced antibodies against the antiviral and antiproliferative effects of interferons in patients with chronic hepatitis C virus infection

Sera from 86 patients with chronic hepatitis C virus (HCV) infection treated with recombinant interferons-alpha (rIFN-alpha) were screened for IFN-binding and antiviral effect-neutralizing antibodies. Out of the 61 patients treated with rIFN-alpha2b, 46% had binding and 28% had neutralizing antibodies. 44% of the 25 patients treated with rIFN-alpha2a developed binding antibodies and 24% had neutralizing antibodies. Contradictory data were observed concerning the appearance of anti-IFN antibodies and the outcome of IFN therapy. A significantly higher number of the patients with a sustained response to rIFN-alpha2b therapy formed antibodies than the number among the non-responder patients. At the same time, in the patients treated with rIFN-alpha2a, opposite data were found. The activity of the antibodies in some sera was studied against the antiproliferative effect of IFNs on Daudi cells by measuring the [3H]thymidine incorporation. The binding antibodies without neutralization of the antiviral effect of the IFNs inhibited the antiproliferative activity of the rIFNs, similarly to antibodies having both IFN-binding and antiviral effect-neutralizing capacities. At the same time, the antiproliferative effect of the natural IFN was less affected. It is suggested that the antiproliferative assay is more sensitive than the antiviral method for demonstration of the presence of antibodies exerting an inhibitory effect on the biological activities of IFN.     

Type-specific antibodies to hepatitis C virus in sera from patients with various hematologic diseases

The reactivity of 100 sera taken from patients with different blood diseases and donors with respect to synthesized peptides in the variable area of protein NS4 of hepatitis C virus was studied. The presence of type-specific antibodies in the blood sera of patients with hepatitis C was shown. Two antigenic determinant corresponding to 1683-1705 and 1711-1732 amino acid residues in the protein area under study were detected. In hematological patients undergoing frequent blood transfusions mixed infection with different types of hepatitis C virus was registered; these types could be reliably determined with the use of synthetic peptides. The serotype determined with the use of peptides corresponded to the type of the circulating virus.     

Serum levels of Th17 associated cytokines in chronic hepatitis C virus infection

The Th17-mediated immune response was investigated in patients chronically infected with hepatitis C virus (HCV) by determining the serum levels of the cytokines involved in the induction of the Th17 response (TGF-β and IL-6), the cytokines produced by Th17 cells (IL-17A, IL-17F and IL-22) and the cytokines whose production is stimulated by Th17 lymphocytes (IL-8 and GM-CSF). We investigated the relationships among the levels of these cytokines by assessing clinical findings, liver histology and viremia. Sixty untreated patients and 28 healthy individuals were included in the study. Cytokine levels were determined using ELISA. Differences between HCV and control groups were identified in the median levels of IL-17F (controls=172.4pg/mL; HCV=96.8pg/mL, p<0.001) and IL-8 (controls=30.1pg/mL; HCV=18.1pg/mL, p<0.05). IL-6 levels were higher in patients presenting moderate liver necroinflammation than in patients with mild or no liver necroinflammation (p<0.05). IL-17F levels were increased in patients that had increased ALT levels. Additionally, a strong positive correlation was observed between IL-17F and IL-22 levels in the two groups investigated, and the IL-17F/IL-22 ratio was lower in the patients infected with HCV (p<0.0001). Patients with low HCV viral loads had higher median levels of IL-8 (32.5pg/mL) than did patients with high HCV loads (16.7pg/mL, p<0.05). These results suggest that in chronic hepatitis C infection, IL-17F and IL-8 could be associated with the control of liver injury and infection, respectively.     

Circulating Th1 and Th2 cytokines in patients with hepatitis C virus infection.

The imbalance of T-helper (Th) lymphocyte cytokine production may play an important role in immunopathogenesis of persistent hepatitis C virus (HCV) infection. To know whether an imbalance between Th1 and Th2 cytokines is present in chronic HCV infection, serum levels of Th1 cytokines, interferon gamma (IFN-gamma) and interleukin (IL)-2, and Th2 cytokines, IL-4 and IL-10, were measured using enzyme-linked immunosorbent assay in this study. Eighteen individuals with chronic HCV infection, 11 healthy subjects as normal controls and 10 chronic HBV infected patients as disease controls were observed. The results showed that the levels of Th2 cytokines (IL-4 and IL-10) were significantly increased in chronic HCV infected patients compared with normal controls (IL-4: 30.49+/-17.55 vs. 14.94+/-13.73, pg/ml, P<0.025; IL-10: 50.30+/-19.59 vs. 17.87+/-9.49, pg/ml, P<0.001). Similarly, the levels of Th1 cytokine, IL-2, was also elevated in individuals with chronic HCV infection when compared with normal controls (IL-2: 118.53+/-95.23 vs. 61.57+/-28.70, pg/ml, P<0.05). However, Th1 cytokine IFN-gamma level was not significantly changed during HCV infection (IFN-gamma: 28.09+/-15.65 vs. 24.10+/-15.61, pg/ml, P>0.05). Furthermore, the elevated levels of Th2 cytokines are greater than Th1 cytokines in HCV infection. Thus, the study indicates that an enhanced Th2 responses are present during chronic HCV infection, which may partly be responsible for the persistence of HCV infection.     

Increased serum oxysterol concentrations in patients with chronic hepatitis C virus infection

Oxidative stress and dysregulated cholesterol metabolism are characteristic features of chronic hepatitis C virus infection (CHC). Therefore, we analyzed serum oxysterol profiles in CHC patients and examined the significance of oxysterols in CHC. The concentrations of 7α-hydroxycholesterol, 4β-hydroxycholesterol and 25-hydroxycholesterol as determined by LC–ESI–MS/MS were significantly elevated by +236%, +29% and +44%, respectively, in CHC patients compared with controls. Moreover, the elevated levels were significantly decreased by anti-viral therapy using PEGylated-interferon and ribavirin for 3 months. In contrast, 24S-hydroxycholesterol, 27-hydroxycholesterol and 7α-hydroxy-4-cholesten-3-one concentrations were not affected by CHC or anti-viral treatment. These results suggest that some oxysterols that are elevated in CHC are produced by cholesterol autoxidation due to oxidative stress or inflammation in the liver. Oxysterols may represent novel targets for the inhibition of disease progression and the prevention of hepatocarcinogenesis in CHC patients.     

Ferritin and liver fibrosis among patients with chronic hepatitis C virus infection

IntroductionIn chronic hepatitis C virus (HCV) infection there is increased iron absorption leading to iron overload, a fact that may promote ferritin synthesis. Theoretically, increased ferritin should promote ongoing liver fibrosis but disparate results have been described.ObjectiveWe analyze the behavior of iron metabolism- related variables, comparing them with fibrosis and inflammatory activity in liver biopsy in HCV infected patients.Patients and MethodsWe analyzed among 90 HCV patients subjected to liver biopsy prior to antiviral treatment the relationships of serum levels of iron, ferritin, transferrin, transferrin saturation index (TSI) and total iron binding capacity (TIBC) with liver fibrosis and histological severity, assessed by Metavir-f, Metavir-a and Knodell indices, as well as with liver function, and also compared the aforementioned iron metabolism- related variables with 34 controls.ResultsPatients showed higher values of sideremia (T = 2.04; p = 0.044) and transferrin (T = 2.29; p = 0.004) compared with controls; but not ferritin, that was significantly higher among the 33 patients who also consumed alcohol (Z = 2.05; p = 0.041). Most patients showed a well preserved liver function (86 cases, Child A). Patients with Child B or C showed higher ferritin levels (Z = 2.68; p = 0.007) and TSI (Z = 2.41; p = 0.016), but lower transferrin and TIBC (Z = 3.25; p = 0.001) than Child A patients. Transferrin and TIBC were directly related to albumin (ρ = 0.24; p = 0.026), whereas bilirubin showed direct relationships with iron (ρ = 0.25; p = 0.016), TSI (ρ = 0.39; p < 0.001) and ferritin (ρ = 0.36; p < 0.001). Both ferritin (ρ = −0.22; p = 0.04) and TSI (ρ = −0.25; p = 0.016) were related to platelet count. No relationships were observed between iron variables and Knodell index, but serum iron, serum transferrin, and TSI were directly related to Metavir-f score (ρ = 0.28; p = 0.009, ρ = 0.22; p = 0.044, and ρ = 0.22; p = 0.044, in this order).ConclusionAlterations of iron related variables are relatively subtle in our series of 90 well compensated HCV patients. Serum ferritin was not related to liver fibrosis and increases only when alcoholism co-exists with HCV infection.     

Levels of hepatitis B virus replicative intermediate in serum samples of chronic hepatitis B patients

Hepatitis B virus (HBV) cccDNA levels is an absolute marker of HBV replication in the liver of HBV infected patients. This study aimed to quantify the HBV cccDNA levels in sera and liver tissue samples of treatment naïve patients with chronic hepatitis B. Eighty one chronic hepatitis B (CHB) treatment naïve patients were enrolled from January 2009 to June 2011. Total HBV DNA and HBV cccDNA levels were quantified using sensitive real time PCR assay. The mean age of recruited patients was 34 ± 11.5 years. Fifty four (66.7 %) patients were HBeAg negative. Liver tissue samples were available from 2 HBeAg positive and 21 HBeAg negative CHB patients. The amount of total intrahepatic HBV DNA ranged from 0.09 to 1508.92 copies/cell. The median intrahepatic HBV cccDNA was 0.31 and 0.20 copies/cell in HBeAg positive and HBeAg negative cases, respectively. Serum HBV cccDNA was detectable in 85.2 % HBeAg positive and 48.1 % HBeAg negative CHB patients. Median serum HBV cccDNA was 46,000 and 26,350 copies/mL in HBeAg positive and HBeAg negative subjects, respectively. There was a significant positive correlation between the levels of intrahepatic total HBV DNA and intrahepatic HBV cccDNA (r = 0.533, p = 0.009). A positive correlation was also seen between serum HBV cccDNA levels and serum HBV DNA levels (r = 0.871, p < 0.001). It was concluded that serum HBV cccDNA could be detectable in higher proportion of HBeAg positive patients compared to HBeAg negative patients. Moreover, the median level of serum HBV cccDNA was significantly higher in HBeAg positive patients in contrast to HBeAg negative subjects.     

Hepatitis C virus genotypes in patients with chronic hepatitis C infection in southern Iran from 2016 to 2019

Hepatitis C is a liver disease caused by the hepatitis C virus (HCV). The treatment of HCV infection has become more complicated due to various genotypes and subtypes of HCV. The treatment of HCV has made significant advances with direct-acting antivirals. However, for the choice of medicine or the combination of drugs for hepatitis C, it is imperative to detect and discriminate the crucial HCV genotypes. The main objective of this study was to determine the pattern of circulating HCV genotypes in southern Iran, from 2016 until 2019. The other aim of the study was to determine possible associations of patients’ risk factors with HCV genotypes. A total of 803 serum samples were collected in 4 years (2016–2019) from patients with HCV antibody positive results. A total of 728 serum samples were HCV-RNA positive. The prevalence of HCV genotypes was detected using the genotype-specific RT-PCR test for serum samples obtained from 615 patients. The HCV genotype 1 (G1) was the most prevalent (48.8%) genotype in the area, with G1a, G1b, and mixed G1a/b representing 38.4%, 10.1%, and 0.3%, respectively. Genotype 3a was the next most prevalent (47.2%). Mixed genotypes 1a/3a were detected in 22 (3.6%) and finally G4 was found in 3 (0.5%) patients. The other HCV genotypes were not detected in any patient. Genotype 1 (1a and 1b alone, 1a/1b and 1a/3a coinfections) is the most prevalent HCV genotype in southern Iran. HCV G1 shows a significantly higher rate in people under 40 years old.     

Strain-specific T-cell suppression and protective immunity in patients with chronic hepatitis C virus infection

Hepatitis C virus (HCV) frequently persists with an apparently ineffective antiviral T-cell response. We hypothesized that some patients may be exposed to multiple HCV subtypes and that strain-specific T cells could contribute to the viral dynamics in this setting. To test this hypothesis, CD4 T-cell responses to three genotype 1a-derived HCV antigens and HCV antibody serotype were examined in chronically HCV infected (genotypes 1a, 1b, 2, 3, and 4) and spontaneously HCV recovered subjects. Consistent with multiple HCV exposure, 63% of patients infected with genotypes 2 to 4 (genotypes 2-4) and 36% of those infected with genotype 1b displayed CD4 T-cell responses to 1a-derived HCV antigens, while 29% of genotype 2-4-infected patients showed serotype responses to genotype 1. Detection of 1a-specific T cells in patients without active 1a infection suggested prior self-limited 1a infection with T-cell-mediated protection from 1a but not from non-1a viruses. Remarkably, CD4 T-cell responses to 1a-derived HCV antigens were weakest in patients with homologous 1a infection and greater in non-1a-infected patients: proportions of patients responding were 19% (1a), 36% (1b), and 63% (2-4) (P = 0.0006). Increased 1a-specific CD4 T-cell responsiveness in non-1a-infected patients was not due to increased immunogenicity or cross-reactivity of non-1a viruses but directly related to sequence divergence. We conclude that the T-cell response to the circulating virus is either suppressed or not induced in a strain-specific manner in chronically HCV infected patients and that, despite their ability to clear one HCV strain, patients may be reinfected with a heterologous strain that can then persist. These findings provide new insights into host-virus interactions in HCV infection that have implications for vaccine development.     

Intrahepatic hepatitis C virus replication correlates with chronic hepatitis C disease severity in vivo

The role of viral factors in the pathogenesis of chronic hepatitis C is unknown. The objective of the present study was to characterize markers of hepatitis C virus (HCV) infection and replication in liver biopsy specimens obtained from 65 genotype 1-infected subjects, including 31 who were coinfected with human immunodeficiency virus (HIV), and to analyze associations between intrahepatic viral markers and hepatitis C disease severity. The percentages of liver cells harboring HCV genomes (%G) and replicative-intermediate RNAs (%RI) were evaluated using strand-specific in situ hybridization, while HCV core and NS3 antigens were assessed by immunocytochemistry. HIV-positive and HIV-negative subjects had similar mean grades and stages of liver disease and had similar indices of HCV infection and replication in liver, even though coinfected subjects had significantly shorter mean disease duration (P = 0.0003). Multivariate analysis showed that %G was not associated with grade or stage of liver disease (P = 0.5 and 0.4, respectively), while %RI was strongly associated with liver inflammation (P < 0.001), liver fibrosis (P < 0.001), and serum alanine aminotransferase levels (P = 0.01). NS3 antigen (but not core) was more frequently detected in HCV RI-positive versus RI-negative specimens (P = 0.028). These findings demonstrate a link between HCV proliferation and hepatitis C disease severity and suggest similar pathogenic mechanisms in HIV-positive and HIV-negative individuals.     

The decline in antibodies to hepatitis C virus during antiviral therapy

The goal of the study was to analyze B-cell response to hepatitis C virus during antiviral therapy among responders and non-responders. The content of antibodies to individual structural and non-structural HCV proteins was investigated during two years in three groups of patients: initial responders, non-responders and a reference group (without therapy). Treated patients in all groups exhibited the decrease in antibodies to analyzed HCV proteins, but with different patterns. The first statistically significant differences in the decline of the virus-specific antibodies between initial responders and non-responders were observed within the first three months after the beginning of therapy. Some treated patients demonstrated the decrease in antibody levels to HCV proteins after the end of therapy.     

IgM anti-hepatitis C virus in patients with chronic non-A, non-B hepatitis and their relationship to viral replication

Patients with hepatitis C virus (HCV) infection may have different patterns of antibody response to various structural and non-structural viral antigens. We have correlated the serological patterns to the clinical features of chronic infection and to viral replication in 68 HCV-Ab-positive patients with chronic liver disease at different stages (19 with cirrhosis-hepatocellular carcinoma, 38 with chronic active hepatitis and 11 with chronic persistent hepatitis). Serum samples from each patient were assayed for HCV-IgM by enzyme immunoassay and for HCV-RNA by the polymerase chain reaction using primer sets derived from the 5'-non-coding region. The prevalence of HCV-IgM was high (54 patients (79.4%)) and the study showed a good correlation between high values of anti-HCV-IgM and the presence of HCV-RNA in serum, since HCV-RNA was detected in 35 of the 54 IgM-positive patients (64.8%) and notably in 19 of the 20 subjects with high levels of specific IgM. Conversely, all the 35 sera containing HCV-RNA were also reactive for HCV-IgM, while none of the HCV-IgM-negative sera was HCV-RNA reactive. Positivity rates for both HCV-RNA and IgM anti-HCV were higher in the more advanced stages of disease; thus, the clinical pattern of HCV chronic hepatitis seems to be strictly related to the serological pattern and the presence of HCV-RNA.     

Prevalence of antibodies to hepatitis C virus (HCV) in haemophiliacs

The prevalence of 1) hepatitis C virus (HCV), an agent likely to be responsible for parenterally transmitted hepatitis non-A, non-B, 2) hepatitis B virus (HBV) and 3) human immunodeficiency virus (HIV) infection was studied in 211 patients with clotting disorders (78% of the patients had residual factor activities of less than or equal to 2%). Of these patients 71% were positive for HBV markers and 44% for HIV markers. Using a new ELISA technique, 80% were anti-HCV-positive. The prevalence of anti-HCV was greater in patients with more severe clotting disorders and was related to the total amount of replacement therapy received; the prevalence was less in older patients. Seroconversion after a single exposure to dry heat-treated factor concentrates was documented in 3 patients 3-4 months after exposure.     

Inhibition of hepatitis C virus-like particle binding to target cells by antiviral antibodies in acute and chronic hepatitis C

Hepatitis C virus (HCV) is a leading cause of chronic viral hepatitis worldwide. The study of antibody-mediated virus neutralization has been hampered by the lack of an efficient and high-throughput cell culture system for the study of virus neutralization. The HCV structural proteins have been shown to assemble into noninfectious HCV-like particles (HCV-LPs). Similar to serum-derived virions, HCV-LPs bind and enter human hepatocytes and hepatoma cell lines. In this study, we developed an HCV-LP-based model system for a systematic functional analysis of antiviral antibodies from patients with acute or chronic hepatitis C. We demonstrate that cellular HCV-LP binding was specifically inhibited by antiviral antibodies from patients with acute or chronic hepatitis C in a dose-dependent manner. Using a library of homologous overlapping envelope peptides covering the entire HCV envelope, we identified an epitope in the N-terminal E2 region (SQKIQLVNTNGSWHI; amino acid positions 408 to 422) as one target of human antiviral antibodies inhibiting cellular particle binding. Using a large panel of serum samples from patients with acute and chronic hepatitis C, we demonstrated that the presence of antibodies with inhibition of binding activity was not associated with viral clearance. In conclusion, antibody-mediated inhibition of cellular HCV-LP binding represents a convenient system for the functional characterization of human anti-HCV antibodies, allowing the mapping of envelope neutralization epitopes targeted by naturally occurring antiviral antibodies.