IgG subclass composition of antibodies to HBsAg in circulating immune complexes from patients with hepatitis B virus infections

The IgG subclass of antibody associated with hepatitis B surface antigen (HBsAg) in circulating immune complexes (CIC) from patients with either acute or chronic hepatitis B virus (HBV) infections was measured using an isotype and antigen-specific ELISA. All patients were HBsAg positive but were negative for free anti-HBs antibody. The subclass of antibody associated with HBsAg in CIC in both groups was predominantly IgG1 and IgG4. This is in contrast to free anti-HBs in convalescent sera from patients recovering from HBV infection, which are highly restricted to IgG1 and IgG3. The finding of high levels of IgG4 antibodies in CIC suggest that CIC containing this subclass may be cleared less efficiently than CIC containing antibodies of other subclasses. Formation of these CIC may be an important factor in the progression of infection to chronicity and may also be involved in the antigen-specific immunosuppression seen in early acute and chronic HBV infections.     

Study of IgG subclass profiles of anti-HBs in populations with different HBV infection status

To study IgG-specific subclasses of hepatitis B virus (HBV) surface antigen (anti-HBs), in different populations in Taiwan, a comparison was made between 104 chronic carriers (60 male and 44 female) and 439 recovered individuals (247 male and 192 female). Biochemical analyses of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were also performed. Among the 104 chronic carriers, 21 patients had abnormal ALT and AST levels (> 25 IU/ml). When comparing the patients with abnormal ALT and AST levels to chronic carriers with normal ALT and AST levels, no statistical difference was observed for anti-HBs levels (p > 0.05). The IgG subclass pattern of the relative anti-HBs IgG subclass titers was IgG1 > IgG3 = IgG4 in both chronic carriers and recovered individuals (p < 0.05). IgG1 is the predominant anti-HBs antibody after HBV infection, in either chronic carriers or in HBV-cured individuals. This finding is partly inconsistent with data reported from other group who suggested in individuals naturally infected, the anti-HBs IgG consists mainly of IgG3 and IgG1. In contrast to that of our previous studies of anti-HBe and anti-HBc, the mean OD values of anti-HBs total IgG, and all IgG subclasses except for IgG2, of either males or females, were significantly higher in recovered individuals than in chronic carriers, while the mean OD values of anti-HBe and anti-HBc were significantly higher in chronic carriers than in recovered individuals (P < 0.05). The IgG subclass profile of anti-HBs in chronic carriers was not changed with liver inflammation and was independent of sex and age, except in individuals with abnormal ALT and AST for whom anti-HBs IgG1 was not significantly higher than IgG3 (p > 0.05), in spite of that whose mean O.D. value is higher.     

Anti-hepatitis B surface antigen IgG1 subclass is predominant in individuals who have recovered from hepatitis B virus infection, chronic carriers and vaccinees

Patterns of each IgG-specific subclass for hepatitis B virus (HBV) core antigen (anti-HBc) are remarkably different among individuals with different infection status, i.e., completely recovered or chronic carrier. Each of the IgG-specific subclasses of HBV surface antigen (anti-HBs) was tested for ELISA sensitivity using four commercially available hepatitis B surface antigen (HBsAg) kits and one self-prepared plate. The specificity in 18 serum samples obtained from chronic HBV carriers, recovered individuals, vaccinees and non-infected individuals was investigated. Differences in absorbance values were obtained by comparing results from these different plates. Data on the absorbance values of anti-HBs IgG subclasses obtained indicated that one to four subjects had a false-negative or false-positive result using the four commercial plates. Only the self-prepared plate demonstrated 100% specificity and sensitivity for anti-HBs subclasses. Moreover, the results indicate that anti-HBs subclass IgG1 was predominant in cured patients, chronic carriers and vaccinees. The samples from both chronic carriers and vaccinees exhibited a significantly higher concentration of total IgG and IgG1 than samples in recovered individuals (P<0.05).     

Small vessel vasculitis caused by hepatitis B virus immune complexes: Small vessel vasculitis and HBsAg

In a comprehensive study of 80 patients with vasculitis, 4 had concurrent hepatitis B virus (HBV) infection. Polyarteritis nodosa was present in 2 and in the other 2, cutaneous vasculitis, presenting clinically as palpable or Henoch-Schönlein purpura. In one of these patients skin biopsies demonstrated granular deposits of IgM, C3, C4, and the hepatitis B surface antigen (HBsAg) and electron-dense deposits of aggregated 20-nm particles resembling HBsAg in postcapillary venules. Evidence for circulating HBsAg-immune complexes included increased serum C1q binding activity, decreased serum complement, and a cryoprecipitate containing both HBsAg and IgM anti-HBs. Aggregated 20-nm particles resembling intact HBsAg were also seen by negative staining electron microscopy of the serum cryoprecipitate. This patient fulfills all the criteria for a specific immune complex vasculitis caused by his immune response to a chronic HBV infection. These findings emphasize that HBV infection may be associated with small vessel vasculitis as well as polyarteritis nodosa, mixed cryoglobulinemia, and glomerulonephritis. A similar immune response to other viral infections may be expressed as palpable (Henoch-Schönlein) purpura also.     

The humoral immune response in acute and chronic hepatitis B virus infection

Conclusions Resolution of an AH-B infection and the associated liver injury are believed to be immune mediated. Therefore, the inability to resolve an HBV infection reflects a defect(s) in the HBV-specific immune response resulting in chronic infection. A number of studies suggest that the immunological defect may reside at the level of T cell responsiveness. Although any number of defects in the complex interactions between the host immune response and HBV may result in persistent infection, a likely candidate is an inefficient HBeAg-specific Th cell response. For example, we have proposed a model in which exposure to HBeAg in utero tolerizes the HBe/HBcAg-specific Th cell response and results in chronicity after neonatal or perinatal HBV infection [53]. Immunosuppression or genetic T cell nonresponsiveness to HBe/HBcAg may explain chronicity after adult HBV infection. However, the chronicity rate after symptomatic HBV infection in adults is relatively low (i.e., < 10%). Chronic or persistent HBV infection encompasses a heterogeneous patient population ranging from severe to limited liver disease to the asymptomatic carrier state. Presumably, chronic liver disease reflects ongoing HBV-specific immune responses sufficient to cause liver cell injury but insufficient to clear the infection completely. Nevertheless, the currently available serological assays suggest that the humoral immune responses to HBeAg and HBsAg during CH-B infection are universally negative regardless of clinical status or degree of liver injury. To resolve this apparent contradiction, we and others have designed more sensitive serological assays capable of detecting anti-HBe and anti-HBs antibodies in the presence of excess circulating antigens. Using sensitive experimental immunoassays it is clear that chronically infected HBV patients produce a variety of antibodies and that the quantity and quality of antibody production correlates with the degree of liver disease. In fact, HBV-specific humoral immune responses in chronic infection appear remarkably intact considering the degree to which Th cell function can be compromised and the excess production of subviral particles and proteins during chronic HBV infection. However, at least one study did suggest a B cell defect during HBV infection [78]. It was suggested that envelope-specific cytotoxic T lymphocytes may lyse HBsAg-specific B cells that present envelope peptides in the context of the MHC class I pathway. Such a mechanism would be expected to suppress anti-HBs production.In summary, the serological responses of CH-B patients serve as efficient markers of the degree and specificity of underlying T cell responsiveness. This information is relevant to the diagnosis and prognosis of CH-B patients and may be useful in determining appropriate treatment modalities. For example, the use of the experimental immunoassays can distinguish between asymptomatic and symptomatic CH-B patients and between AH-B patients and CH-B patients undergoing an AE of liver disease. Furthermore, a survey of a large number of CH-B patients using the more sensitive immunoassays suggested that the balance between HBV-specific Th₁ and Th₂-type cells may be important in preventing and/or resolving chronic HBV infection. Lastly, use of the sensitive experimental immunoassays may allow the selection of CH-B patients most likely to benefit from immunomodulatory therapy. For example, asymptomatic as well as symptomatic CH-B patients with evidence of pre-existing HBV-specific immune responses may be more responsive to immune-enhancing therapies (i.e., IFN-).     

Utilizing self-prepared ELISA plates for a cross-population study of different anti-HBe IgG subclass profiles

Fourteen serum samples obtained from hepatitis B virus (HBV) chronic carriers and patients recovered from hepatitis B infection were used with four sodium dodecyl sulfate-treated enzyme-linked immunosorbent assay (ELISA) plates available commercially, and one self-prepared HBcAg analog for evaluation of anti-HBe subclass pattern absorbance. The self-prepared plates had the best performance and were thus used for samples obtained from 104 (60 male and 44 female) HBV chronic carriers and 439 (247 male and 192 female) recovered individuals. Tests for aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were also carried out in 21 of the subjects (>25 IU/ml). Statistical comparison of these patients with elevated ALT/AST levels with other ALT/AST-normal chronic carriers revealed no significant differences in the anti-HBe OD, although the mean optical density (OD) of patients with elevated ALT/AST levels was higher. The results suggest that the anti-HBe IgG subclass profiles in the chronic carriers did not change with inflammation of the liver, and were independent of sex and age. In contrast to previous anti-HBc findings, the distribution pattern of anti-HBe subclasses in HBV chronic carriers was IgG1 > IgG4 > IgG3 while in the recovered individuals it was IgG1 > IgG3 > IgG4, for both males and females. Subclasses IgG1 and IgG2 were the most and least prevalent isotypes, respectively, in both study groups. The results of the study suggest that induction of IgG1 and/or IgG3 antibodies is important for effective virus neutralization, while IgG2 antibodies are of limited importance. Significantly higher OD values for anti-HBe IgG4 were observed when comparing samples from the chronic carriers and recovered individuals, which may reflect the effects of persistence. Further, in contrast to previous anti-HBs results, the concentrations of total IgG and IgG1 were higher in the samples from chronic carriers relative to those from recovered individuals.     

Passively acquired antibody to hepatitis B surface antigen. Pitfall in evaluating immunity to hepatitis B viral infections

Antibody to hepatitis B surface antigen (anti-HBs) has been used clinically to indicate an immune response to hepatitis B virus (HBV) and a protection against reinfection with the virus. We describe a child with hemophilia who had high-titer IgG anti-HBs in his serum and who subsequently developed viral B hepatitis. The child had received a unit of fresh frozen plasma 17 days prior to the determination of anti-HBs. The fresh frozen plasma donor was later found to be anti-HBs positive. The patient's anti-HBs was most likely passively acquired and therefore did not signify immunity to HBV. Various tests, including hepatitis B surface antigen group-specific and subtype determinants, ratio units of anti-HBs, and antibody class, have been used to determine whether or not anti-HBs will confer immunity. Although these tests have been thought to accurately predict immune status against infection with HBV, our case shows this may not be true, especially in patients who have been recently transfused. Anti-HBs testing may be predictive of immunity to HBV in the absence of a source of passively acquired anti-HBs.     

Regulation of the Immune Response to Hepatitis B Virus and Human Serum Albumin

The circulatory pool of B cells, from donors immune lo hepatitis B (HB) through natural infection, contained sensitized B cells with the capacity to secrete antibodies with specificity for human serum albumin (USA) when stimulated with purified hepatitis B surface antigen (HBsAg) in vitro The immunoglobulin secretion was dependent upon and regulated by T cells and specifically induced, since it was not obtained in cell cultures from HB-susceptible donors. Culture supernatants with anti-USA reactivity also contained specific antibodies to HBsAg (anti-HBs). indicating that the outer coat of HBV normally provokes an immune response to both the viral antigen and a self component. Perturbation in the regulation of the immune response triggered by USA in association with HBV/HBsAg particles may involve a putative risk for development of chronic HBsAg carriership.     

IL-17A in hepatitis B infection: friend or foe?

Hepatitis B virus (HBV) is one of the most prevalent and infectious agents that leads to liver disease in humans. Five clinical forms of HBV infection exist, including fulminant, acute, chronic, asymptomatic and occult. The chronic, asymptomatic and occult forms are long-term infections that can lead to hepatocellular carcinoma (HCC) and liver cirrhosis. The mechanisms responsible for progression of these forms of the infection to HCC and liver cirrhosis are not yet clearly understood or characterised. However, genetic and immunological parameters may play important roles in the disease. IL-17A is an important cytokine involved in early immune responses against fungal and bacterial infections, but its role in the response against viral infections is yet to be fully clarified. The crucial roles of IL-17A in the pathogenesis of autoimmune and destructive immune-related diseases have been documented and may provide insights into its functions during hepatitis infection. Therefore, the aim of this review was to address the recent information regarding the status and association of IL-17A during hepatitis B infection and its related disorders, including HCC and liver cirrhosis.     

Pathogenesis of hepatitis B virus infection

The adaptive immune response is thought to be responsible for viral clearance and disease pathogenesis during hepatitis B virus infection. It is generally acknowledged that the humoral antibody response contributes to the clearance of circulating virus particles and the prevention of viral spread within the host while the cellular immune response eliminates infected cells. The T cell response to the hepatitis B virus (HBV) is vigorous, polyclonal and multispecific in acutely infected patients who successfully clear the virus and relatively weak and narrowly focussed in chronically infected patients, suggesting that clearance of HBV is T cell dependent. The pathogenetic and antiviral potential of the cytotoxic T lymphocyte (CTL) response to HBV has been proven by the induction of a severe necroinflammatory liver disease following the adoptive transfer of HBsAg specific CTL into HBV transgenic mice. Remarkably, the CTLs also purge HBV replicative intermediates from the liver by secreting type 1 inflammatory cytokines thereby limiting virus spread to uninfected cells and reducing the degree of immunopathology required to terminate the infection. Persistent HBV infection is characterized by a weak adaptive immune response, thought to be due to inefficient CD4+ T cell priming early in the infection and subsequent development of a quantitatively and qualitatively ineffective CD8+ T cell response. Other factors that could contribute to viral persistence are immunological tolerance, mutational epitope inactivation, T cell receptor antagonism, incomplete down-regulation of viral replication and infection of immunologically privileged tissues. However, these pathways become apparent only in the setting of an ineffective immune response, which is, therefore, the fundamental underlying cause. Persistent infection is characterized by chronic liver cell injury, regeneration, inflammation, widespread DNA damage and insertional deregulation of cellular growth control genes, which, collectively, lead to cirrhosis of the liver and hepatocellular carcinoma.     

Immunoglobulin G subclass antibodies to measles virus in patients with subacute sclerosing panencephalitis or multiple sclerosis

Mouse monoclonal antibodies specific for human immunoglobulin G (IgG) subclasses and a sensitive immunoassay were used to evaluate the IgG subclass antibody response to measles virus antigens in cerebrospinal fluid and serum samples from 20 patients with subacute sclerosing panencephalitis (SSPE), 12 patients with multiple sclerosis (MS), and 11 controls with high measles virus antibody titers in serum. In patients with SSPE, measles virus-specific antibodies were found mainly in the IgG1 subclass and the IgG subclass distribution remained unchanged, irrespective of the clinical stage or duration of the disease. In patients with MS and in controls, measles virus activity was also associated mainly with IgG1. However, the activity was significantly lower than that found in patients with SSPE. The results suggest that there is no primary abnormality in humoral immune response to measles virus in patients with MS. The disproportionately high levels of the measles virus-specific IgG1 subclass found in patients with SSPE may be due to persistent antigenic stimulation or reflect a defect in immunoregulatory mechanisms in response to viral infection.     

IgG subclass response to Helicobacter pylori and CagA antigens in children

Specific serum IgG subclass antibodies against Helicobacter pylori antigens and recombinant CagA were analysed in 75 symptomatic children with histologically confirmed H. pylori infection. H. pylori stimulated an IgG1 predominant response, and IgG3 titres showed a positive association with peptic ulcer disease, chronicity of antral inflammation and density of H. pylori colonization. Two methods used for assessing serum IgG CagA antibody status, i.e. Western blotting and enzyme-linked immunosorbent assay (ELISA), were concordant. CagA stimulated an IgG1 and IgG3 predominant humoral response. Total CagA IgG titres were higher in children with active and more severe chronic antral inflammation. These findings suggest that in children the systemic humoral immune response to H. pylori infection may reflect gastroduodenal pathology.     

Neutralization of hepatitis B virus infectivity by a murine monoclonal antibody: an experimental study in the chimpanzee

Two study chimpanzees were inoculated intravenously with approximately 1,000 chimpanzee infectious doses of hepatitis B virus (HBV), one with subtype adr and one with subtype ayw, each previously incubated with 0.1 ml of a murine monoclonal antibody (IgG 1(K) class) directed against a single epitope on hepatitis B surface antigen common to most or all HBV. Two control chimpanzees received identical doses of HBV not incubated with the murine anti-HBs. Neither study chimpanzee developed HBV infection during 12 months of follow-up as judged by normal serum aminotransferase activity, normal liver biopsies, and negative serological tests for HBV-associated antigens and antibodies. In contrast, both control chimpanzees became infected by HBV as evidenced by elevated serum aminotransferase activity, liver biopsy changes characteristic of viral hepatitis, and the appearance of hepatitis B surface antigen (HBsAg) in their sera. Both study chimpanzees were shown to be fully susceptible to infection with these same HBV inocula when challenged 15 months after the initial inoculations at a time when passively administered anti-HBs was no longer detectable. Prior to challenge with HBV, one of the two study chimpanzees received a second injection of the same volume of the murine monoclonal anti-HBs. The survival of this anti-HBs in serum was reduced from six weeks (after the initial injection) to approximately two weeks.(ABSTRACT TRUNCATED AT 250 WORDS)     

Hepatitis B precore protein: pathogenic potential and therapeutic promise

Hepatitis B virus (HBV), a small and economically packaged double-stranded DNA virus, represents an enormous global health care burden. In spite of an effective vaccine, HBV is endemic in many countries. Chronic hepatitis B (CHB) results in the development of significant clinical outcomes such as liver disease and hepatocellular carcinoma (HCC), which are associated with high mortality rates. HBV is a non-cytopathic virus, with the host's immune response responsible for the associated liver damage. Indeed, HBV appears to be a master of manipulating and modulating the immune response to achieve persistent and chronic infection. The HBV precore protein or hepatitis B e antigen (HBeAg) is a key viral protein involved in these processes, for instance though the down-regulation of the innate immune response. The development of new therapies that target viral proteins, such as HBeAg, which regulates of the immune system, may offer a new wave of potential therapeutics to circumvent progression to CHB and liver disease.     

The role of innate immunity in HBV infection

Hepatitis B virus (HBV) infection is one of the main causes of chronic liver diseases. Whether HBV infection is cleared or persists is determined by both viral factors and host immune responses. It becomes clear that innate immunity is of importance in protecting the host from HBV infection and persistence. However, HBV develops strategies to suppress the antiviral immune responses. A combined therapeutic strategy with both viral suppression and enhancement of antiviral immune responses is needed for effective long-term clearance and cure for chronic HBV infection. We and others confirmed that bifunctional siRNAs with both gene silencing and innate immune activation properties are beneficial for inhibition of HBV and represent a potential approach for treatment of viral infection. Understanding the nature of liver innate immunity and their roles in chronic HBV progression and HBV clearance may aid in the design of novel therapeutic strategies for chronic HBV infection.     

Vaccination with a fusion DNA vaccine encoding hepatitis B surface antigen fused to the extracellular domain of CTLA4 enhances HBV-specific immune responses in mice: Implication of its potential use as a therapeutic vaccine

Fusion of specific antigens to extracellular domain of cytotoxic-T-lymphocyte-associated antigen 4 (CTLA4) represents a promising approach to increase the immunogenicity of DNA vaccines. We evaluated this interesting approach for its enhancement on HBV-specific immune responses and its antiviral effects in HBV transgenic mice. A fusion plasmid encoding the extracellular domain of CTLA4 linked with HBsAg was constructed. Mice were immunized by this fusion plasmid. Vaccination with the CTLA4-fused DNA not only induced much higher level of anti-HBs antibody, but also increased HBsAg-specific CD8+ response as well as CTL response in BALB/c mice. Furthermore, both Th1 and Th2 responses were augmented. In HBV transgenic mice, the levels of circulating HBsAg and HBV DNA replication were down-regulated by induction of higher anti-HBs antibody and HBsAg-specific CD8+ response after vaccination with the fusion plasmid. Thus, the CTLA4-fused DNA vaccine led to breakdown of immune tolerance to viral infection in HBV transgenic mice, which might be used as a therapeutic vaccine in HBV infection.     

Lipopolysaccharide is present in immune complexes isolated from sputum in patients with cystic fibrosis and chronic Pseudomonas aeruginosa lung infection.

Sputum samples from seven patients with cystic fibrosis and chronic P. aeruginosa lung infection were investigated for immune complexes by PEG precipitation and in two different complement binding assays. All seven patients were immune complex positive. The components involved in immune complex formation were identified by SDS-PAGE and immunoblotting. We found P. aeruginosa lipopolysaccharide as a major antigen. Both core and O-specific saccharide antigens could be demonstrated. IgG and IgA were the immunoglobulins involved, with IgG2 as the dominating IgG subclass. Lipopolysaccharide has a number of biological activities and its presence in sputum may have consequences for the pathogenesis of lung disease in cystic fibrosis.     

Cellular and humoral immune reactions in chronic active liver disease. II. Lymphocyte subsets and viral antigens in liver biopsies of patients with acute and chronic hepatitis B

The characteristics and distribution of the inflammatory infiltrate in liver biopsies of 25 patients with hepatitis B viral (HBV) infection were studied in relation to the distribution and expression of HBV antigens. Mononuclear subsets were characterized with monoclonal (OKT, OKM, Leu) antibodies to surface antigens. For the demonstration of viral antigens directly conjugated antibodies to surface (HBsAg), core (HBcAg) and 'e' (HBeAg) antigen were used. For the study of mutual relations all methods were performed on serial cut tissue sections. In chronic active hepatitis B (CAH-B, n = 12) OKT8+ lymphocytes of T cell origin were the only cell type present in areas with liver cell degeneration and T cell cytotoxicity appears to be the only immune mechanism. In chronic persistent hepatitis B (CPH-B, n = 7) the only conspicuous feature was the presence of many Leu 3+ lymphocytes of the helper/inducer population in the portal tracts. In acute hepatitis B (AHB, n = 6) OKT8+ cells of non-T origin (OKT1-,3-) and Leu 7+ cells of presumed natural killer (NK) potential predominated in the areas with liver cell necrosis, and non-T cell cytotoxicity appears to be the predominant immune mechanism. In none of these disease entities a positive spatial relation could be established between the cytotoxic cells and the demonstrable expression of HBV antigens in hepatocytes. It is concluded that differences in immunological reaction pattern may explain the different course in the three forms of HBV infection studied.     

Lymphocyte proliferation and IgG production with herpesvirus antigens in solid phase

A technique for simultaneous specific lymphocyte proliferation and IgG production with herpesvirus antigens in solid phase was developed. The cell mediated immune response was highly specific. Stronger cellular responses were found after stimulation with nucleocapsid antigens than with membrane antigens. Varicella-zoster virus (VZV) and herpes simplex virus (HSV) specific antibody production in vitro were found in almost 100% of seropositive individuals, while cytomegalovirus (CMV) antibody production was detectable in 70%. A higher specific IgG production was found after stimulation with CMV and VZV membrane antigens than with CMV nucleocapsid and VZV cell antigens. No differences in IgG production were found with the two types of HSV antigens. A method for viral IgG subclass determination in vitro was also developed. These methods may be used for monitoring immunosuppressed patients for immunological responsiveness to herpesvirus infections.     

Different profiles of wheat antigens are recognised by patients suffering from coeliac disease and IgE-mediated food allergy

BACKGROUND: Dietary intake of wheat can cause two distinct immunologically mediated diseases with severe gastrointestinal manifestations, coeliac disease (CD) and IgE-mediated food allergy. The pathomechanisms underlying these diseases are different, but the profile of the target antigens in wheat has not been compared for the two diseases. METHODS: We compared IgA- and IgE-reactive antigens in wheat using sera from patients with coeliac disease (n = 35) and food allergy to wheat (n = 16) by one- and two-dimensional immunoblotting. Furthermore, the IgG subclass (IgG1-IgG4) reactivity to wheat antigens was studied by enzyme-linked immunosorbent assay. RESULTS: IgA antibodies from CD patients and IgE antibodies from allergic patients recognised distinct profiles of wheat antigens. Furthermore, the IgG subclass responses to wheat antigens were different in CD and wheat-allergic patients. CONCLUSION: This study thus demonstrates that wheat contains antigens/epitopes which are preferentially recognised by CD patients, whereas others elicit IgE-mediated food allergy. This finding suggests that the nature of a food antigen may influence the quality of the pathological immune response in the gut and has implications for the diagnosis and therapy of hypersensitivity to wheat.