The immune response to Epstein-Barr virus

Epstein-Barr virus is a gammaherpes virus that establishes persistent infection in the majority of humans. Despite the potent oncogenic potential of the virus it is relatively rarely associated with malignant disease. This review focuses on the cellular immune responses that successfully control Epstein-Barr virus infection in most individuals.     

T-lymphocyte subset interactions in the cell-mediated immune response to Epstein-Barr virus

The T-lymphocyte subset interactions in the cell-mediated response to Epstein-Barr virus (EBV) were studied in an in vitro system in which the ability of T lymphocytes to inhibit outgrowth of autologous EBV-transformed B lymphocytes is assessed. Inhibition could be demonstrated within lymphocytes of both the T4 and T8 surface phenotypes. Outgrowth inhibition was observed more frequently when the effector T-cell population contained cells of both surface phenotypes. In blocking experiments the OKT3 antibody completely prevents development of outgrowth inhibitory activity; the OKT4 and OKT8 antibodies were less effective in interfering with outgrowth inhibitory function. Maximal blocking activity occurred when antibody addition occurred in the early phase of generation of suppressor function. Pharmacologically achievable concentrations of interferon-alpha restored outgrowth inhibitory activity after blocking with monoclonal antibody. EBV reactivation was easily demonstrable in a group of 10 renal allograft recipients who received OKT3 antibody for treatment of acute rejection. These studies suggest that the immunoregulatory control of proliferation of EBV-transformed B lymphocytes is complex, and involves a collaborative interaction of lymphocytes of both the T4 and T8 surface phenotypes.     

CD4+ T cell responses in the immune control against latent infection by Epstein-Barr virus

The human gamma-herpesvirus Epstein-Barr virus establishes latent, life-long infection in more than 95% of the human adult population. Despite its growth transforming capacity, most carriers control EBV associated malignancies efficiently and remain free of EBV+ tumors. It is commonly accepted that lymphoblastoid cells, expressing all EBV latent antigens, are targeted by the immune system and cause tumors only in immune-suppressed individuals. However, immune control of EBV associated malignancies which express only three or one EBV latent antigen is less obvious. Recent studies have addressed the pattern of EBV latent infection in healthy EBV carriers and the identity of EBV derived target antigens for CD4+ T cells. The results suggest that immune surveillance also extends to tumors, which have down-regulated most EBV latent antigens and therefore escape EBV specific immune recognition at least in part. EBV specific immunity that targets these tumors in healthy EBV carriers seems to fail specifically during the development of Hodgkin's disease, nasopharyngeal carcinoma and Burkitt's lymphoma. These three EBV+ tumors appear to subdue EBV immunity against the remaining EBV latent antigens in different ways or profit from the effect of other pathogens on EBV specific immune responses, when they develop in otherwise immune competent individuals. While immune control and immune escape of these so-called spontaneously arising EBV associated malignancies is just beginning to be understood, immune control of persisting EBV infection can serve as a model for tumor immune surveillance in general.     

The immune response to herpes simplex virus

Several host immune mechanisms are activated in the course of a herpes simplex virus infection. These include natural resistance mechanisms (natural killer cells and interferon), antiviral antibodies and effector CD4 and CD8 T lymphocytes. An important mechanism in the control of viral replication in epidermal cells involves the recruitment and activation of macrophages by CD4 T cells. In some instances, the action of CD4 T cells can lead to immune pathology following infection of the eye (stromal ketatitis) or central nervous system (demyelination). Despite the efficiency of the immune response in countering infection, the virus has evolved strategies to subvert the action of antibodies and complement and the detection of infected cells by cytotoxic T lymphocytes.     

The cell-mediated immune response to ectromelia virus infection. I. Kinetics and characteristics of the primary effector T cell response in vivo.

The cell-mediated immune (CMI) response to ectromelia virus infection in mice was studied. Virus doses from 4 × 10² up to 5 × 10⁴ PFU of an attenuated strain inoculated intravenously (iv) all induced cytotoxic T cell responses in the spleen as measured in a ⁵¹Cr release assay using virus-infected target cells. Higher virus doses gave larger responses. There was little variation between individual animals, and mice ranging in age from 4–22 weeks gave similar responses. Following iv infection, virus grew logarithmically in spleen for 2 days, then titers declined to undetectable levels by day 5. The peak of the virus-specific cytotoxic T cell response occurred at 5–6 days post-infection, as determined by calculation of effector units based on a linear log-log relationship between killer cells added and targets lysed. T cells responsible for virus clearance in vivo gave similar kinetics, suggesting the possibility that both functions are mediated by the same T cell subset. Two other categories of cytotoxic activity were also generated at low levels in the spleen during ectromelia infection or during infection with a bacterium, Listeria monocytogenes. These activities were significantly sensitive to anti-δ and complement treatment, suggesting T cell dependence, but participation of other mechanisms has not been rigorously excluded. One category lysed allogenic target cells and reached a peak at 4 days post-infection. The other lysed H-2-compatible cells, syngeneic embryo cells, and some syngeneic tumor cells but not syngeneic macrophages, and was present at similar low levels through days 1–4. These different kinetics and evidence from “cold” target competition experiments suggested that the total cytotoxic activity of immune spleen cell populations was a composite of the activities of separate cellular subsets (probably mainly T cells), killing of any one target cell type being the responsibility of a subset with receptors at least partly specific for antigens on that target cell.     

Rectification of age-associated deficiency in cytotoxic T cell response to influenza A virus by immunization with immune complexes

Decline in cellular immunity in aging compromises protection against infectious diseases and leads to the increased susceptibility of the elderly to infection. In particular, Ag-specific cytotoxic T lymphocyte (CTL) response against virus is markedly reduced in an aged immune system. It is of great importance to explore novel strategy in eliciting effective antiviral CTL activity in the elderly. In this study, the efficacy and mechanisms of immunization with immune complexes in overcoming age-associated deficiency in cellular immunity were investigated. In this study, we show that the severely depressed CTL response to influenza A in aged mice can be significantly restored by immunization with immune complexes consisting of influenza A virus and mAb to influenza A nucleoprotein. The main mechanisms underlying this recovery of CTL response induced by immune complex immunization in aged mice are enhanced dendritic cell function and elevated production of IFN-gamma in both CD4(+) Th1 and CD8(+) CTLs. Thus, these results demonstrate that immune complex immunization may represent a novel strategy to elicit effective virus-specific cytotoxic response in an aged immune system, and possibly, to overcome age-related immune deficiency in general.     

Specific immune serum to the Epstein-Barr virus DNA polymerase.

Epstein-Barr virus (EBV) DNA polymerase was released from phorbol ester-treated tamarin (Saguinus oedipus) cells (B95-8) and prepared for use as an antigen by sequential column chromatography with DEAE-Sephadex A-25, DEAE-cellulose, phosphocellulose, and single-stranded DNA cellulose. Proteins from single-stranded DNA cellulose with DNA polymerase activity in 100 mM ammonium sulfate were mixed with complete Freund adjuvant and injected intradermally into rats and rabbits. Immune sera that were screened for specific antibody by indirect immunofluorescence procedures reacted with approximately 3% of the cells in EBV-producer cultures (B95-8 and P3HR-1) but not with EBV genome-negative cells (BJAB). In functional enzyme assays, immune sera or the immunoglobulin fraction inhibited the activity of purified EBV DNA polymerase 90%. Inhibition of enzyme activity was not affected by absorption of immune sera with insoluble matrices of proteins prepared with tamarin and human cells which lacked the EBV genome. Cellular DNA polymerase alpha was not inhibited by immune sera to the EBV enzyme.     

Epstein-Barr virus: exploiting the immune system

In vitro, Epstein-Barr virus (EBV) will infect any resting B cell, driving it out of the resting state to become an activated proliferating lymphoblast. Paradoxically, EBV persists in vivo in a quiescent state in resting memory B cells that circulate in the peripheral blood. How does the virus get there, and with such specificity for the memory compartment? An explanation comes from the idea that two genes encoded by the virus--LMP1 and LMP2A--allow EBV to exploit the normal pathways of B-cell differentiation so that the EBV-infected B blast can become a resting memory cell.     

Dominance and diversity in the primary human CD4 T cell response to replication-competent vaccinia virus

Vaccination with replication-competent vaccinia protects against heterologous orthopoxvirus challenge. CD4 T cells have essential roles helping functionally important Ab and CD8 antiviral responses, and contribute to the durability of vaccinia-specific memory. Little is known about the specificity, diversity, or dominance hierarchy of orthopoxvirus-specific CD4 T cell responses. We interrogated vaccinia-reactive CD4 in vitro T cell lines with vaccinia protein fragments expressed from an unbiased genomic library, and also with a panel of membrane proteins. CD4 T cells from three primary vaccinees reacted with 44 separate antigenic regions in 35 vaccinia proteins, recognizing 8 to 20 proteins per person. The integrated responses to the Ags that we defined accounted for 49 to 81% of the CD4 reactivity to whole vaccinia Ag. Individual dominant Ags drove up to 30% of the total response. The gene F11L-encoded protein was immunodominant in two of three subjects and is fragmented in a replication-incompetent vaccine candidate. The presence of protein in virions was strongly associated with CD4 antigenicity. These findings are consistent with models in which exogenous Ag drives CD4 immunodominance, and provides tools to investigate the relationship between Ab and CD4 T cell specificity for complex pathogens.     

T cell regulation of the immune response to infection in periodontal diseases

Although T cells have been implicated in the pathogenesis and are considered to be central both in progression and control of the chronic inflammatory periodontal diseases, the precise contribution of T cells to the regulation of tissue destruction has not been fully elucidated. Current dogma suggests that immunity to infection is controlled by distinct T helper 1 (Th1) and T helper 2 (Th2) subsets of T cells classified on the basis of their cytokine profile. Further, a subset of T cells with immunosuppressive function and cytokine profile distinct from Th1 or Th2 has been described and designated as regulatory T cells. Although these regulatory T cells have been considered to maintain self-tolerance resulting in the suppression of auto-immune responses, recent data suggest that these cells may also play a role in preventing infection-induced immunopathology. In this review, the role of functional and regulatory T cells in chronic inflammatory periodontal diseases will be summarized. This should not only provide an insight into the relationship between the immune response to periodontopathic bacteria and disease but should also highlight areas of development for potentially new therapeutic modalities.     

The human immune response to reconstituted bovine collagen

The human immune response to bovine dermal collagen was characterized through histologic, serologic, and immunoblotting methods. Collagen-sensitive patients were identified by hypersensitivity to intradermal exposure to ZYDERM Collagen Implant--a pepsin-solubilized, reconstituted, bovine dermal collagen. Biopsies of test sites in the forearm were obtained from several collagen-sensitive patients. Histologic examination revealed an implant-associated palisading foreign body granuloma. The lesion also contained a mixed cell infiltrate of histiocytes, lymphocytes, and eosinophils. Sera were collected from patients who developed erythema or induration at intradermal test or treatment sites, and were evaluated for antibodies to bovine dermal collagen by an enzyme-linked immunosorbent assay (ELISA). Sera with anti-collagen antibodies were further characterized in this study. The circulating antibodies were reactive with both native and heat-denatured bovine dermal collagen. By using purified alpha 1(I) and alpha 2(I) polypeptides, these sera were found to have antibodies reactive with both alpha-chains. Each alpha-chain was fragmented by using cyanogen bromide (CB). The CB peptides were electrophoretically separated, and these sera were evaluated for antibodies to the major fragments by using an immunoblotting technique. Of the sera evaluated by this method, 89% (23/26) had antibodies to alpha 1-CB6; 77% (20/26) had antibodies to alpha 2-CB4; and 65% (17/26) had antibodies reactive with both CB fragments. In addition, most sera (77%) contained antibodies reactive with two or more (up to five) of the major CB peptides. The least antigenic fragment was alpha 2-CB3,5 (8%). In addition, these sera had antibody activity against both native and heat-denaturated bovine types III and II collagens. Little or no interspecies (rat or guinea pig) cross-reactivity (types I and II) was detected. Furthermore, these sera did not have antibodies against human types I, II, and III collagens.     

A dynamical model of human immune response to influenza A virus infection

We present a simplified dynamical model of immune response to uncomplicated influenza A virus (IAV) infection, which focuses on the control of the infection by the innate and adaptive immunity. Innate immunity is represented by interferon-induced resistance to infection of respiratory epithelial cells and by removal of infected cells by effector cells (cytotoxic T-cells and natural killer cells). Adaptive immunity is represented by virus-specific antibodies. Similar in spirit to the recent model of Bocharov and Romanyukha [1994. Mathematical model of antiviral immune response. III. Influenza A virus infection. J. Theor. Biol. 167, 323-360], the model is constructed as a system of 10 ordinary differential equations with 27 parameters characterizing the rates of various processes contributing to the course of disease. The parameters are derived from published experimental data or estimated so as to reproduce available data about the time course of IAV infection in a na?ve host. We explore the effect of initial viral load on the severity and duration of the disease, construct a phase diagram that sheds insight into the dynamics of the disease, and perform sensitivity analysis on the model parameters to explore which ones influence the most the onset, duration and severity of infection. To account for the variability and speed of adaptation of the adaptive response to a particular virus strain, we introduce a variable that quantifies the antigenic compatibility between the virus and the antibodies currently produced by the organism. We find that for small initial viral load the disease progresses through an asymptomatic course, for intermediate value it takes a typical course with constant duration and severity of infection but variable onset, and for large initial viral load the disease becomes severe. This behavior is robust to a wide range of parameter values. The absence of antibody response leads to recurrence of disease and appearance of a chronic state with nontrivial constant viral load.     

Regulation of T cell apoptosis during the immune response

Apoptosis of T-lymphocytes is a fundamental process regulating antigen receptor repertoire selection during T cell maturation and homeostasis of the immune system. It also plays a key role in elimination of autoreactive lymphocytes. Resting mature T cells are activated by antigen to elicit an appropriate immune response. In contrast, preactivated T cells undergo activation-induced cell death (AICD) in response to TCR triggering alone. Thus, death by apoptosis is essential for function, growth and differentiation of T-lymphocytes. This review focuses on apoptosis mechanisms involved in T cell development and during the course of an immune response.     

Use of fluoresceinated Epstein-Barr virus to study Epstein-Barr virus-lymphoid cell interactions.

As a direct approach to visualize Epstein-Barr virus (EBV) binding to its cellular receptors and to learn more on the nature of this binding, virus preparations were conjugated to fluorescein isothiocyanate and used to detect EBV receptors on lymphoid cells. Different enzymatic and chemical treatments were also applied either to the virus or to target cells or to both, and the effect of these treatments on virus binding was then examined. The results obtained show that: (i) EBV can be fluoresceinated without affecting its infectivity or cell binding ability, and the fluoresceinated virus represents an important tool to investigate the biology and nature of EBV interactions with its cellular receptors; (ii) the two virus strains (P3HR-1 and B95-8) share common receptors on Raji cells; (iii) protease treatment of EBV or target cells abrogates virus binding; (iv) EBV receptors regenerate after removal of the protease, and this regeneration is inhibited by cycloheximide or sucrose; (v) EBV particles bear concanavalin A receptors, and this lectin hinders the interaction of the virus with its cellular receptors; (vi) neuraminidase treatment, various monosaccharides, ovalbumin, and glycopeptides derived from EBV or cell surface do not inhibit virus binding. Taken together, the above data also demonstrate that some cellular and viral surface (glyco-) proteins are required for EBV binding to its targets.     

Innate immune response to RNA virus infection

Viral RNA is recognized by RIG-I-like receptors and Toll-like receptors. RIG-I is a cytoplasmic viral RNA sensor. High Mobility Group Box (HMGB) proteins and DExD/H box RNA helicases, such as DDX3 and 60, associate with viral RNA. Those proteins promotes the RIG-I binding to viral RNA. RIG-I triggers the signal via IPS-1 adaptor molecule to induce type I IFN. RIG-I harbors Lys63-linked polyubiquitination by Riplet and TRIM25 ubiquitin ligases. The polyubiquitination is essential for RIG-I-mediated signaling. Toll-like receptors are located in endosome. TLR3 recognizes viral double-stranded RNA, and TLR7 and 8 recognize single-strand RNA. Virus has the ability to suppress these innate immune response. For example, to inhibit RIG-I-mediated signaling, HCV core protein suppresses the function of DDX3. In addition, HCV NS3-4A protein cleaves IPS-1 to inhibit the signal. Molecular mechanism of how viral RNA is recognized by innate immune system will make great progress on our understanding of how virus escapes from host immune system.     

Analysis of cellular immune response to EBV by using cloned T cell lines

Eight cloned T cell lines specific for Epstein Barr virus-transformed B lymphocytes were derived. In the presence of the autologous virus-infected B cells, the T cell lines show HLA-restricted cytotoxic activity and also secrete alpha-interferon in sufficient amounts to inhibit infection and transformation. Four of these clones showed restriction to a single HLA locus (two for A3, and two for B7) and three showed exquisite self-restriction lysing only autologous targets. These seven clones expressed the classical cell surface phenotype of cytotoxic T cells being T3, 8, 11, and la-positive and T4-negative. An eighth clone that lacked the T8 surface marker appeared to recognize both B7 and BW51. HLA restriction was confirmed: 1) by the ability of a monoclonal antibody against an HLA-A,B,C framework antigen (W6-32) to block the cytotoxicity; 2) the failure of the clones to lyse Daudi, an EBV-positive, HLA-A,B, C-negative cell line; and 3) successful competition of the cytotoxicity by autologous but not allogeneic cold targets. The cloned T cells do not kill EBV-negative targets such as autologous pokeweed mitogen blasts and cell lines including CEM and the natural killer cell target K562. The results suggest T cell clones may be generated against an EBV-associated membrane antigen on transformed B cells, perhaps equivalent to the lymphocyte-determined membrane antigen, and that the recognition is restricted by a single HLA determinant. We propose that single T cells can play multiple roles in controlling EBV infection in vitro and in vivo including the elimination of transformed cells by cytotoxicity and the prevention by secreted interferon of further re-infection and transformation.     

T cell cytokine profile during primary Epstein-Barr virus infection (infectious mononucleosis)

Cytokine profiles of CD4+ and CD8+ T-cell subsets were evaluated in 8 patients with infectious mononucleosis (IM). Intracellular detection of cytokines using flow cytometry revealed an expansion of IFN-gamma-expressing CD4+ T cells, and particularly CD8+ T cells, while IL-2 expressing cells were less frequently encountered when compared to healthy controls. Single TNF-alpha-expressing CD4+ and CD8+ T cells were likewise reduced and shifted towards IFN-gamma/TNF-alpha co-production. The predominant pro-inflammatory type 1-biased immune response during IM was emphasized by low frequencies of IL-10 expression in both T cell subsets, although some patients displayed elevated serum levels. Six months later, a decreased, but still elevated IFN-gamma expression within the CD8+ T cell subset, and an increased percentage of IL-2-expressing CD4+ and CD8+ T cells, reaching values shown for controls, were noted. Type 2-associated cytokines such as IL-4 and IL-13, as well as IL-6 and TNF-alpha were not significantly different when compared to controls at study entry and at follow-up. The striking expansion of IFN-gamma-producing CD8+ T cells with rather low expression of IL-10, appears to be a key factor for clinically overt disease, but is nevertheless compatible with successful control of the viral infection.