Expression of intrahepatic hepatitis D viral antigen in chronic hepatitis D virus infection.

To elucidate the biological importance of intrahepatic hepatitis D virus antigen, its expression was correlated with biochemical and histological inflammatory activity in 98 biopsy specimens from 68 patients seropositive for total antibody to the virus. Seventy five specimens were positive for intrahepatic nuclear antigen for HDV antigen accompanied by cytoplasmic HDV antigen in only one biopsy specimen. This group had significantly higher serum transaminase activities and inflammatory activity than the remaining cases that were negative for HDV antigen. Among the group positive for HDV antigen, there was no correlation between the proportion of hepatocytes containing HDV antigen and either serum transaminase activity or histological inflammatory indices. In 22 HDV antigen positive patients who had follow up biopsy specimens taken at a median of two years, the proportion with cirrhosis increased from 36% to 73%. Serum transaminase activities remained the same during this period, but the proportion of HDV antigen positive cells dropped. Follow up of 51 patients showed that 21 died or underwent liver transplantation within three years. The absence of an association between intrahepatic HDV antigen expression and progression of histological liver damage does not support the view that HDV is directly cytopathic to hepatocytes. Immune mediated mechanisms may have a role in the pathogenesis of chronic liver disease related to HDV infection.     

Mouse hepatitis virus infection suppresses modulation of mouse spleen T-cell activation.

Natural infection by mouse hepatitis virus (MHV) can affect interpretation of immunological studies in mice. MHV, a collective term describing a group of corona viruses, is found in natural infections in over 70% of laboratory mouse populations in the U.S.A. and Canada. Natural outbreaks of MHV in our animal colony afforded us the opportunity to study MHV-induced immunosuppression as well as the effects of MHV infection on neurotransmitter immunomodulation. Concanavalin A (Con A)-stimulated DNA synthesis by spleen T lymphocytes from MHV-infected mice was 20-50% that of non-infected mice. The MHV infection also altered neurotransmitter modulation of spleen T-lymphocyte activation. In contrast to noradrenaline ablation of Con A-activated DNA synthesis by spleen lymphocytes from non-infected mice, DNA synthesis by the infected group was not inhibited by noradrenaline or dibutyryl-cAMP. These effects of MHV infection were specific for spleen T lymphocytes since MHV infection did not alter Con A stimulation of thymocytes, lipopolysaccharide stimulation of spleen B lymphocytes, or noradrenaline inhibition of thymocyte and B-cell DNA synthesis. MHV infection also did not alter spleen T-lymphocyte subset proportions. Thus, MHV infection inhibits spleen T-lymphocyte activation and blocks in vitro catecholamine and cAMP regulation of spleen T-cell activation but does not affect activation of thymic cells or spleen B cells.     

Differential effect of chronic hepatitis D virus infection on intrahepatic expression of hepatitis B viral antigen.

AIMS: To determine how chronic hepatitis D virus (HDV) infection affects intrahepatic hepatitis B virus (HBV) antigen expression. METHODS: Ninety eight liver biopsy specimens from 68 patients seropositive for total antibody to HDV were studied by immunohistochemistry, and the amount of HBV antigens was also quantified by radioimmunoassay in 12 patients and compared with 30 patients with chronic HBV infection. RESULTS: Forty nine of the 68 patients were positive for intrahepatic HDV antigen and only five were positive for HBV core antigen (HBcAg). HBV surface antigen (HBsAg) was present in 55 (80.9%) patients and was always cytoplasmic in distribution. Hepatic pre-S1 and pre-S2 expressions paralleled that of HBsAg, and were detected in 53 (77.9%) and 54 (79.4%) patients, respectively. There was no relation between the intrahepatic expression of HDV antigen and HBsAg/pre-S1/pre-S2. Follow up biopsy specimens in 25 patients showed either static or deteriorating histology while intrahepatic HDV antigen remained the same or fell. The patients with intrahepatic expression of HBcAg had either absent or noticeably decreased expression of HBcAg in their follow up biopsy specimens (median two years). In contrast, HBsAg/pre-S1/pre-S2 were the same or increased (p less than 0.001). Quantification of intrahepatic HBsAg in patients with chronic HDV infection (0.61 pg/hepatocyte, range: 0.05-1.08, n = 12) showed no difference with patients with chronic HBV infection alone (0.64 pg/hepatocyte, range: 0.02-1.02, n = 30, p = NS). CONCLUSION: These data indicate that chronic HDV infection suppresses intrahepatic expression of HBcAg but not HbsAg and pre-S antigens, suggesting a differential effect of chronic HDV infection on HBV gene expression.     

Target specific hyaluronic acid-interferon alpha conjugate for the treatment of hepatitis C virus infection

Interferon alpha (IFNα) conjugated with polyethylene glycol (PEG) has been widely used for the treatment of hepatitis C virus (HCV) infection as a once-a-week injection formulation. However, the PEGylated IFNα has a low efficacy of ca. 39% and a side effect after repeated injections possibly due to the non-specific delivery with PEGylation. In this work, target specific long-acting hyaluronic acid-interferon alpha (HA-IFNα) conjugate was successfully developed for the treatment of HCV infection. HA-IFNα conjugate was synthesized by coupling reaction between aldehyde modified HA and the N-terminal group of IFNα. The IFNα content could be controlled in the range of 2-9 molecules per single HA chain with a bioconjugation efficiency higher than 95%. According to in vitro anti-proliferation assay using Daudi cells, HA-IFNα conjugate showed a comparable biological activity to PEG-Intron. In vivo real-time bioimaging confirmed the target specific delivery of near-infrared fluorescence (NIRF) dye labeled HA-IFNα conjugate to the liver in mice. In addition, pharmacokinetic analysis revealed the enhanced residence time longer than 4 days. After tail-vein injection, HA-IFNα conjugate induced ca. 60% higher expression of 2',5'-oligoadenylate synthetase 1 (OAS 1) for innate immune responses to viral infection in the murine liver tissues than IFNα and PEG-Intron.     

Serological markers of posttransfusion hepatitis C viral infection.

Serological markers for hepatitis C virus (HCV) infection were measured in serial samples from 14 posttransfusion chronic non-A, non-B hepatitis patients by a semiquantitative dot blot immunoassay. The assay detected antibodies to HCV by use of recombinant proteins that represent putative HCV capsid (core), nonstructural protein 3 (NS3) (33c), and NS4 (c100) epitopes. Seroconversion to anti-HCV antibodies (anti-HCV) was detected in all patients. The average time to active antibody production detected by any of the recombinant proteins was 13.8 (range, 3.6 to 22.0) weeks posttransfusion or 4.6 (range, -4.5 to 13.4) weeks after the first biochemical marker of illness. Anti-HCV were detected earliest by the core antigen in most cases; however, the patterns of anti-HCV responses varied significantly among individuals. Overall, the addition of the core and NS3 antigens to the assay enabled the detection of the antibody response 4 to 5 weeks earlier than did the addition of the c100 antigen, the sole antigen used in current screening tests in the United States. Passively transferred antibodies were detected by at least one antigen in early posttransfusion samples from 12 patients and decayed below detectable levels for all antigens in only 2 patients. Antibodies to all three gene products were evident in the last sample from all five patients monitored for greater than 3 years from transfusion indicating the persistence of antibodies in patients with chronic illness. Our data show that the period following the onset of hepatitis during which anti-HCV are not detected by current screening assays can be greatly shortened by the detection of anti-HCV responses by a combination of core, NS3, and c100 antigens.     

Aberrant CD8+ T-cell responses and memory differentiation upon viral infection of an ataxia-telangiectasia mouse model driven by hyper-activated Akt and mTORC1 signaling

Immune system-related pathology is common in ataxia-telangiectasia (A-T) patients and mice that lack the protein kinase, A-T mutated (ATM). However, it has not been studied how ATM influences immune responses to a viral infection. Using the lymphocytic choriomeningitis virus (LCMV) infection model, we show that ATM^−/− mice, despite having fewer naïve CD8⁺ T cells, effectively clear the virus. However, aberrant CD8⁺ T-cell responses are observed, including defective expansion and contraction, effector-to-memory differentiation, and a switch in viral-epitope immunodominance. T-cell receptor-activated, but not naïve, ATM^−/− splenic CD8⁺ T cells have increased ribosomal protein S6 and Akt phosphorylation and do not proliferate well in response to IL-15, a cytokine important for memory T-cell development. Accordingly, pharmacological Akt or mammalian target of rapamycin complex 1 (mTORC1) inhibition during T-cell receptor activation alone rescues the IL-15 proliferation defect. Finally, rapamycin treatment during LCMV infection in vivo increases the number of memory T cells in ATM^−/− mice. Altogether, these results show that CD8⁺ T cells lacking ATM have hyperactive Akt and mTORC1 signaling in response to T-cell receptor activation, which results in aberrant cytokine responses and memory T-cell development. We speculate that similar signaling defects contribute to the immune system pathology of A-T, and that inhibition of Akt and/or mTORC1 may be of therapeutic value.Ataxia-telangiectasia (A-T) is a human disease caused by mutations in the gene encoding the PI3-kinase-like protein kinase A-T mutated (ATM).¹ A-T is a multifaceted disease with complex pathology. Cerebellar degeneration underlies the hallmark ataxia symptoms, but another prominent issue is immune system-related pathology, including immunodeficiency and lymphoid cancers.² A-T patients commonly acquire hematological malignancies (eg, leukemia and lymphoma) that together with recurrent bronchial infections account for most of the mortality from the disease.³ ATM gene knock-out mouse models of A-T exhibit many features of the human disease,^4–6 including sexual immaturity, immune system defects, hematopoietic stem cell defects, and thymic lymphoma, the latter of which is the most common cause of death in these animals.^4,7Immunodeficiency associated with decreased production of immunoglobulins A, E, and G2, and thymic hypoplasia has been documented in A-T patients.^8,9 The latter involves decreased peripheral CD4⁺ and CD8⁺ T-lymphocyte pools resulting from developmental defects in the thymic microenvironment.⁹ Because ATM is recruited to double-strand breaks, it is likely that defects in the V(D)J recombination process, which results in a block in differentiation at the CD4⁺/CD8⁺ double-positive stage in the thymus, cause lower thymic output of mature CD4⁺ and CD8⁺ cells. This is corroborated by the ability of a functional T-cell receptor (TCR)-αβ transgene to rescue the deficit in peripheral T cells in ATM^−/− mice.¹⁰ Despite the defective thymic development of T cells in A-T patients, the immune function of mature T cells has been reported to be essentially normal.¹¹ However, to date, there have been no studies of how deficiency of ATM affects the response to an infection in A-T patients or in the mouse models of the disease.The best-defined role for ATM is in the nuclear DNA damage response^2,12; however, other functions for ATM have been described.^12,13 For example, ATM is important for mitochondrial homeostasis,^14,15 insulin signaling,^13,16 phosphorylation of 5''-AMP-activated protein kinase (AMPK),^17–19 and activation of Akt.^16,20 In addition, ATM signals to TSC2 in response to reactive oxygen species,²¹ to inhibit mammalian target of rapamycin complex 1 (mTORC1) that itself is regulated by AMPK and Akt.^22–24 Finally, treatment of mice with the mTORC1 inhibitor rapamycin significantly increases the life span of ATM^−/− mice by delaying development of thymic lymphoma.²⁵ Altogether, these results highlight how the loss of ATM might disrupt the integration of signals that feed into the nutrient-sensing mTORC1 pathway.The CD8⁺ T-cell response is a crucial arm of the adaptive immune system. In response to an infection, these cells are activated through the TCR, proliferate, and differentiate into cytotoxic effector cells that kill infected cells. Most of these cells die after clearance of the pathogen, but a subpopulation survives, loses effector cell properties, and become memory T cells.^26,27 Memory T cells are important for fighting recurrent infections, as they are programmed to respond faster and more effectively to the pathogen. The CD8⁺ T-cell response to infection involves differentiation into short-lived effector cells and memory-precursor cells^26,28 that can be monitored based on surface expression of KLRG1 and CD127 markers. Effector CD8⁺ T cells are preferentially represented in the KLRG1^hi population,^29,30 whereas cells that are CD127^hi, which is the receptor for IL-7, and KLRG1^lo preferentially become long-lived memory T cells.^31–33The response of CD8⁺ T cells to TCR activation and the pathways involved in effector and memory cell differentiation are well-documented.³⁴ These include roles of the AMPK and mTORC1 pathways at several levels. For example, TCR activation leads to rapid activation of AMPK in response to Ca²⁺ signaling, presumably in anticipation of the enormous energy demand required for T-cell expansion.³⁵ In addition, we have shown that AMPK/mTORC1 signaling dynamically regulates mitochondrial biogenesis during TCR activation.³⁶ Finally, treatment of mice with the AMPK activator metformin or the mTORC1 inhibitor rapamycin enhances memory T-cell differentiation by boosting fatty acid oxidation.^33,37 Similarly, mTORC1 regulates differentially effector and memory T-cell commitment,³⁸ and it is a negative regulator of memory T-cell differentiation in mice.^33,37,39The goal of this study was to determine how loss of ATM affects normal CD8⁺ T-cell activation and differentiation upon viral infection and to understand how alterations in mTORC1 and related pathways due to lack of ATM might contribute to the immune-related pathology of A-T using ATM^−/− mice as a model of the disease and the well-characterized murine lymphocytic choriomeningitis virus (LCMV) infection paradigm.     

Abnormal T-cell activation in chronic hepatitis B viral infection: a consequence of monocyte dysfunction?

The process of T-cell activation in chronic hepatitis B virus (HBV) carriers has been investigated by measurement of membrane expression of lymphocyte-activation markers in response to a variety of mitogenic stimuli in order to delineate further the abnormality of T-cell-mediated immunity present in such patients. A substantial proportion of unstimulated T cells from the peripheral blood of patients but not controls expressed HLA-DR; in contrast the IL-2 and transferrin receptors were rarely expressed spontaneously in either group and there was no difference in spontaneous lymphocyte transformation. After stimulation with monocyte-dependent T-cell mitogens, phytohaemagglutinin (PHA) or anti-T3, patients had significantly reduced expression of the IL-2 and transferrin receptors and of HLA-DR in association with impaired lymphocyte transformations compared to controls. In contrast, lymphocyte activation was normal in response to the monocyte-independent T-cell mitogen phorbol-myristate-acetate (PMA). These data confirm that the process of T-cell activation is abnormal in chronic HBV carriers but suggest that the T cell is intrinsically normal. In allogeneic co-cultures, monocytes from patients inhibited the transformation of normal and patients' lymphocytes in response to PHA, suggesting that defects of T-cell-mediated immunity in chronic HBV carriers may be a consequence of monocyte dysfunction.     

Prospective study of viral clearance and CD4(+) T-cell response in acute hepatitis C primary infection and reinfection.

BACKGROUND: The outcome of acute hepatitis C is determined by early host-virus interactions, particularly involving the antiviral T-cell response. OBJECTIVES: To identify early prognostic markers of spontaneous resolution of acute hepatitis C by performing a comprehensive analysis of viral and immunological factors during the natural course of acute HCV infection and reinfection. STUDY DESIGN: 20 patients were investigated prospectively during acute HC or confirmed reinfection and 18 of them during follow up after spontaneous or treatment-induced elimination of the virus and resolution of the disease. Multiparameter flow cytometry was used to functionally characterize virus-specific CD4(+) T-cell responses relative to the virologic outcome. RESULTS: Parallel immunologic and virologic monitoring of patients with acute HC identified distinct patterns of host-virus interaction related to HCV persistence or clearance. The highest frequency of antiviral Th1 cytokine-producing CD4(+) T-cells was observed in patients with HCV reinfection, preceding rapid viral clearance within 3 weeks after disease onset. In all patients who subsequently cleared viremia, CD4(+) T-cells produced Th1 cytokines following stimulation with non-structural HCV antigens (NS3 and NS4). In contrast, a chronic course of disease was associated with the absence of antiviral Th1 cytokine producing cells from the first weeks after onset of disease (acute persistent HC), or with fluctuating RNA levels (yo-yo pattern) and gradual waning of antiviral Th1 cells. CONCLUSIONS: The results highlight the variability of immune response pattern in acute hepatitis C. Most importantly, "acute persistent hepatitis C" and a lack of TH1 effector cells within the first months of acute hepatitis C represent efficacious predictors of viral persistence and could thus be used as criteria in selecting candidates for early antiviral treatment.     

Induction of EAE by T cells specific for alpha B-crystallin depends on prior viral infection in the CNS

While myelin-reactive T cells are widely believed to play a pathogenic role in multiple sclerosis (MS), no substantial differences appear to exist in T-cell responses to myelin antigens between MS patients and healthy subjects. As an example, indistinguishable peripheral T-cell responses and serum antibody levels have been found in MS patients and healthy controls to alpha B-crystallin, a dominant antigen in MS-affected brain myelin. This suggests that additional factors are relevant in allowing myelin-reactive T cells to become pathogenic. In this study, we examined whether the inflammatory state of the CNS is relevant to the pathogenicity of alpha B-crystallin-specific T cells in mice. In normal mice, T-cell responses against alpha B-crystallin are limited by robust immunological tolerance. Reactive T cells were therefore generated in alpha B-crystallin-deficient mice, and these T cells were transferred into C57BL/6 recipients. While such a transfer in itself never induced any clinical signs of experimental autoimmune encephalomyelitis (EAE) in healthy recipient mice, acute EAE could be induced in animals that had been infected 7 days before with the avirulent A7(74) strain of Semliki Forest virus (SFV). SFV infection alone did not induce clinical disease, nor did it alter the expression levels of the target antigen. Our findings indicate that at least in mice, alpha B-crystallin-specific T cells can trigger EAE but only when prior viral infection has induced an inflammatory state in the CNS that helps recruit and activate T cells.     

Quantitative signal of anti-HCV by an automated assay predicts viremia in a population at high prevalence of hepatitis C virus infection.

BACKGROUND AND AIMS: The diagnosis of ongoing hepatitis C virus (HCV) infection involves the detection of specific antibodies and of HCV-RNA. We aimed to assess the relationship between these two parameters in a representative sample of a population at high risk for HCV infection. METHODS: Plasma and serum samples were respectively tested for HCV-RNA by a qualitative PCR (Cobas Amplicor HCV, Roche) and for HCV antibodies by a MEIA screening assay (AxSYM HCV 3.0, Abbott) and an immunoblot (Inno-LIA-III, Innogenetics). RESULTS AND CONCLUSIONS: Out of 888 samples assayed, 579 (65.2%) were positive for HCV-RNA, while anti-HCV antibodies were detected respectively in 802 sera by AxSYM (90.3%) and in 783 by LIA (706 positive and 77 indeterminate, 88.2%). The anti-core antibodies displayed the best correlation with viremia, since they were present in 97.1% of the PCR+ samples, followed by anti-NS3 (90.2%) and anti-NS4 (89.6%). Only one HCV-RNA positive sample was negative by LIA and MEIA (early seroconversion). The AxSYM sample/cutoff (S/CO) values were directly correlated with the presence of HCV-RNA: a PCR positivity was found in 4.9% of samples with a S/CO < or =10, in 60.8% of samples with a S/CO between 11 and 50 and in 93.6% of cases with a S/CO >50, (P < 0.005). The immunoblot adds little, on a single specimen, to the information yielded by the AxSYM screening test. A suitable diagnostic algorithm for HCV in high-risk settings could be the anti-HCV screening by MEIA and a qualitative assay for HCV-RNA on samples with low reactivity.     

Both LFA-1-positive and -deficient T cell clones require the CD2/LFA-3 interaction for specific cytolytic activation.

We investigated the capacity of T lymphocytes from a leukocyte adhesion-deficient (LAD) patient to respond to alloantigen. Leukocytes of this patient completely lacked LFA-1 surface expression due to the absence of mRNA coding for the LFA-1 beta chain. Despite the absence of LFA-1, T lymphocytes obtained from this patient, cultured with allogeneic stimulator cells (lymphoblastoid B cells JY), were capable of lysing JY cells. Furthermore, two T cell clones (one CD4+ and one CD8+), generated from this lymphocyte culture, specifically lysed the allogeneic lymphoblastoid JY cells. The cytolytic capacity of LFA-1-negative T lymphocytes and T cell clones was comparable to that of control LFA-1-positive T cells with allospecificity against JY. Detailed analysis of the CD4 positive and LFA-1-negative T cell clone demonstrated that it specifically recognized HLA-DQ. Antibody inhibition studies showed that the CTL/target cell interaction was mediated through the CD2/LFA-3 adhesion pathway. LFA-1 expressed by the target cells did not participate in the CTL/target cell conjugate formation and contributed only minimally to the cytotoxic activity. Moreover, when allogeneic LFA-1-deficient B cells, bearing the appropriate HLA-DQ alloantigen, were used as target cells, significant levels of specific cytotoxicity were measured, further excluding a role for LFA-1 in this interaction. The adhesion molecules, VLA-4, CD44 and L-selectin (LECAM1) were not involved. These results demonstrate that LFA-1-negative T lymphocytes can exert allospecific cytotoxicity and that CTL/target cell contact is mediated through the CD2/LFA-3 route. This observation may explain in part why in LAD patients viral infections, cleared largely by T cells, are less frequently observed than bacterial infections, in which phagocytic cells play a major role.     

Age-related accumulation of LFA-1high cells in a CD8+CD45RAhigh T cell population

The differential expression of CD45 isoforms has been suggested as a marker for stages of post-thymic T cell development, that is, CD45RA+CD45R0^? T cells and CD45RA^?CD45R0⁺ T cells are supposed to be virgin and memory cells respectively. Recently, several adhesion molecules have been shown to be up-regulated on the cell surface of memory T cells, and have been suggested to serve as a memory marker. In this study, we investigated the levels of LFA-1 expression on T cells in various subpopulations defined by CD45 isoform expression in donors of various ages. In CD4⁺ T cells, the proportion of LFA-1^high cells among CD45RA^highCD45R0-T cells remained low in all age groups and did not show significant accumulation with age. CD4⁺CD45RA^?CD45R0^highTcells expressed LFA-1 at a higher level than CD4⁺CD45RA^highCD45R0^? T cells. Thus, the currently prevailing view that CD45RA and CD45R0 can be markers for virgin and primed cells was consistent with LFA-1 expression in CD4⁺ T cell population. In CD8⁺ T cells, however, CD45RA^highCD45R0^? T cells consisted of two distinct subpopulations, LFA-1^low and LFA-1^high cells, whereas CD45RA^?CD45R0^high T cells were almost exclusively LFA-1^high When CD29 expression was examined in place of LFA-1 expression, similar results were obtained; CD45RA^high CD45R0^? T cells consisted of two distinct subpopulations, CD29-^{to low} and CD^29high cells, while CD45RA-CD45R0^high T cells were mostly CD29^high. The proportion of LFA-1^high cells in the CD8+CD45RA^high T cell subpopulation increased significantly as a function of age (r = 0.9, p < 0.001). It ranged from 8% in a 14-year-old donor to 94% in a 79-year-old donor. Furthermore, the proportion of CD8⁺CD45RA^highLFA-1^high cells in the CD8⁺ T cell population increased significantly as a function of age (r = 0.85, p < 0.001). On the other hand, the proportion of LEA-1^high cells in CD8⁺CD45RA^? T cell subpopulation exceeded 90% in most donors irrespective of age. These results indicate that the CD8⁺CD45RA^highCD45R0^? T cell subpopulation contains a considerable number of LFA-1^high cells and CD29^high cells, phenotypically similar to previously activated cells. Thus, in terms of LFA-1 and CD29 expressions, the simple scheme that CD45RA is a marker of virgin cells is not applicable to the CD8⁺ T cell population.     

T-cell intrinsic effects of GITR and 4-1BB during viral infection and cancer immunotherapy

GITR [glucocorticoid inducible tumor necrosis factor receptor (TNFR)-related protein] and 4-1BB are costimulatory TNFR family members that are expressed on regulatory and effector T cells as well as on other cells of the immune system. Here we discuss the role of GITR and 4-1BB on T cells during viral infections and in cancer immunotherapy. Systemic treatment with agonistic anti-4-1BB antibody leads to a number of immune system abnormalities, and clinical trials of anti-4-1BB have been terminated. However, other modes of 4-1BB ligation may be less toxic. To date, similar toxicities have not been reported for anti-GITR treatment of mice, although anti-GITR antibodies can exacerbate mouse autoimmune models. Intrinsic effects of GITR and 4-1BB on effector T cells appear to predominate over their effects on other cell types in some models. Despite their similarities in enhancing T-cell survival, 4-1BB and GITR are clearly not redundant, and both pathways are required for maximal CD8(+) T-cell responses and mouse survival following severe respiratory influenza infection. GITR uses TNFR-associated factor (TRAF) 2 and TRAF5, whereas 4-1BB recruits TRAF1 and TRAF2 to mediate survival signaling in T cells. The differential use of signaling adapters combined with their differential expression may explain the non-redundant roles of GITR and 4-1BB in the immune system.     

Prevalence of GB virus-C/hepatitis G virus infection in an antenatal population

It is difficult to explain the high levels of infection seen with GBV-C/HGV if transmission relies on the parenteral route. A group of young women was investigated in order to establish the prevalence of infection in this age group of the general population and perhaps indicate other possible routes of infection, searching for both GBV-C/HGV RNA and HGV E2 antibodies. Evidence of infection was found in 11.8%. This is a higher prevalence than that found in blood donors but lower than in prostitutes. Evidence is accumulating from various groups that sexual/ close contact may result in transmission of this virus.     

Lessons from the study of T-cell differentiation in persistent human virus infection

Confusion surrounds the current classification of memory and effector T-cell subsets and there is a lack of consistency in the use of these terms between human and murine studies. The development of peptide-HLA tetrameric complexes ("tetramers") that accurately identify virus-specific T cells and can be used with a range of cell surface and intra-cellular markers has provided further insights in our understanding of the process of T-cell differentiation, or post-thymic development. We propose that T-cell differentiation subsets in human viral infection should be regarded as distinct from the current definitions of memory and effector cells; further work is needed to reveal the role of the differentiation process in anti-viral immunity.     

Mouse cytomegalovirus infection induces antibodies which cross-react with virus and cardiac myosin: a model for the study of molecular mimicry in the pathogenesis of viral myocarditis.

Mouse cytomegalovirus (MCMV) infection induces persisting myocarditis in the susceptible BALB/c strain. Autoantibodies to cardiac myosin are produced in both susceptible BALB/c and resistant C57BL/6 mice following MCMV infection. These affinity-purified anti-cardiac myosin antibodies cross-react with MCMV protein(s). The polypeptides of CMV which share immunological cross-reactivity with the 200,000 MW polypeptide, the heavy chain of myosin, were viral polypeptides of 83,000, 94,000 and 116,000 MW recognized by BALB/c post-infection sera and polypeptides of 66,000 and 94,000 MW recognized by C57BL/6 post-infection sera. Passive transfer of anti-cardiac myosin antibodies from Day 56 post-infection sera of the BALB/c strain induced inflammation and necrosis of the myocardium of uninfected BALB/c recipients. This late immune sera contains autoantibodies specific for the cardiac isoform of myosin. Furthermore, immunization with cardiac myosin induced myocarditis and high titres of cardiac myosin antibodies in uninfected mice of the susceptible BALB/c strain only. However, antibodies to myosin elicited in cardiac myosin-immunized BALB/c mice did not cross-react with MCMV by ELISA. We suggest that virus infection may modulate the immune recognition of the common-epitope(s) shared between MCMV protein(s) and the heavy chain of myosin. Of particular interest is the possibility that molecular mimicry of CMV with cardiac myosin may contribute to the pathogenesis of autoimmune myocarditis following virus infection.     

New tools to monitor the impact of viral infection on the alloreactive T-cell repertoire

Accumulating evidence suggests that alloreactive memory T-cells may be generated as a result of viral infection. So far, a suitable tool to define the individual human leukocyte antigen (HLA) cross-reactivity of virus-specific memory T-cells is not available. We therefore aimed to develop a novel system for the detection of cross-reactive alloresponses using single HLA antigen expressing cell lines (SALs) as stimulator. Herein, we generated Epstein-Barr Virus (EBV) EBNA3A specific CD8 memory T-cell clones (HLA-B*0801/FLRGRAYGL peptide restricted) and assayed for alloreactivity against a panel of SALs using interferon-γ Elispot as readout. Generation of the T-cell clones was performed by single cell sorting based on staining with viral peptide/major histocompatibility complex-specific tetramer. Monoclonality of the T-cell clones was confirmed by T-cell receptor (TCR) polymerase chain reaction analysis. First, we confirmed the previously described alloreactivity of the EBV EBNA3A-specific T-cell clones against SAL-expressing HLA-B*4402. Further screening against the entire panel of SALs also showed additional cross-reactivity against SAL-expressing HLA-B*5501. Functionality of the cross-reactive T-cell clones was confirmed by chromium release assay using phytohemagglutinin blasts as targets. SALs are an effective tool to detect cross-reactivity of viral-specific CD8 memory T-cell clones against individual class I HLA molecules. This technique may have important implications for donor selection and monitoring of transplant recipients.     

Association of hepatitis A virus infection with allergic sensitization in a population with high prevalence of hepatitis A virus exposure

BACKGROUND: An inverse association between allergic sensitization and markers of exposure to food-borne and orofecal infections (particularly hepatitis A virus, HAV) has been reported. The prevalence of HAV exposure and allergic sensitization vary widely in different areas, and vary along with age within a given area. AIM: To investigate the association between HAV exposure and allergic sensitization in adults from a mostly rural area of Spain. METHODS: An age-stratified random sample of 720 subjects was drawn from the population older than 18 years of A-Estrada, Spain. From 697 eligible subjects, 469 (67.2%, median age 54 years, range: 18-92) participated in the study. Positive skin prick tests to a panel of aeroallergens defined allergic sensitization. Positive serum HAV antibodies (assayed in 465 subjects) defined HAV exposure. RESULTS: The prevalence of HAV exposure was 83.6% (95% CI: 80.7-86.5). The prevalence of allergic sensitization was lower in subjects with HAV exposure than in patients without it (25.0%vs 40.0%, OR 0.44, 95% CI: 0.25-0.77, P=0.004), but this association became substantially altered after adjusting for age, which was closely linked to both allergic sensitization and HAV exposure (adjusted OR 1.15, 95% CI: 0.60-2.19, P=0.66). CONCLUSIONS: In a population with high prevalence of HAV exposure, no significant association between HAV exposure and allergic sensitization is observed after controlling for the confounding effect of age.