Evidence for a functional interaction between the beta chain of major histocompatibility complex class II and the T cell receptor alpha chain during recognition of a bacterial superantigen.

Several studies have suggested that there is a direct interaction between the T cell receptor (TCR) and the major histocompatibility complex (MHC) molecule during T cell recognition of superantigen. To further investigate this possibility, we have analyzed T cell recognition of a bacterial superantigen, Staphylococcal enterotoxin B (SEB), presented by a series of mutant murine I-Ek molecules in which residues of either the alpha or beta chain predicted to interact with the TCR have been substituted. Individual T cell hybridomas gave distinct patterns of responsiveness to SEB presented by the I-E beta k mutants that could not be attributed to differences in the binding of SEB to the mutants. This effect appeared to be dependent on the TCR-alpha chain because some of these hybridomas expressed identical TCR transgenic beta chains. In contrast, none of the hybridomas gave distinct patterns of responsiveness to SEB presented by the I-E alpha k mutants. Taken together, these observations support the idea that there is a functional interaction between the alpha chain of the TCR and the beta chain of the MHC class II molecule. The data also support the idea that this interaction might enhance superantigen recognition in some cases.

Major histocompatibility complex-restricted recognition of retroviral superantigens by V beta 17+ T cells.

It has been established that at least some V beta 17+ T cells interact with an endogenous superantigen encoded by the murine retrovirus, Mtv-9. To analyze the role of major histocompatibility complex (MHC) class II molecules in presenting the Mtv-9 encoded superantigen, vSAG-9 to V beta 17+ hybridomas, a panel of nine hybridomas was tested for their ability to respond to A20/2J (H-2d) and LBK (H-2a) cells which had been transfected with the vSAG-9 gene. Whereas some of the hybridomas recognized vSAG-9 exclusively in the context of H-2a, other hybridomas recognized vSAG-9 exclusively in the context of H-2d or in the context of both H-2d and H-2a. These results suggest that: (a) the class II MHC molecule plays a direct role in the recognition of retroviral superantigen by T cells, rather than serving simply as a platform for presentation; and, (b) it is likely that components of the TCR other than V beta are involved in the vSAG-9/TCR/class II interaction.

Major histocompatibility complex class II-associated peptides control the presentation of bacterial superantigens to T cells.

Recent studies have shown that only a subset of major histocompatibility complex (MHC) class II molecules are able to present bacterial superantigens to T cells, leading to the suggestion that class-II associated peptides may influence superantigen presentation. Here, we have assessed the potential role of peptides on superantigen presentation by (a) analyzing the ability of superantigens to block peptide-specific T cell responses and (b) analyzing the ability of individual peptides to promote superantigen presentation on I-Ab-expressing T2 cells that have a quantitative defect in antigen processing. A series of peptides is described that specifically promote either toxic shock syndrome toxin (TSST) 1 or staphylococcal enterotoxin A (SEA) presentation. Whereas some peptides promoted the presentation of TSST-1 (almost 5,000-fold in the case of one peptide), other peptides promoted the presentation of SEA. These data demonstrate that MHC class II-associated peptides differentially influence the presentation of bacterial superantigens to T cells.

Major histocompatibility complex-specific recognition of Mls-1 is mediated by multiple elements of the T cell receptor.

We have recently shown that recognition of the mouse mammary tumor virus 9-associated superantigen (vSAG-9) by murine V beta 17+ T cells is strongly influenced by the major histocompatibility complex (MHC) class II haplotype of the presenting cells, resulting in a form of MHC-restricted recognition. This finding was unexpected, because T cell recognition of another well-characterized retroviral superantigen, minor lymphocyte-stimulating antigen 1 (Mls-1), had been shown to be independent of the MHC haplotype of the presenting cell. To determine whether recognition of vSAG-9 and Mls-1 is fundamentally different, we undertook an extensive analysis of MHC haplotype influences on vSAG-9 and Mls-1 recognition by panels of T cell hybridomas. Our results show that, although most hybridomas recognized Mls-1 regardless of the MHC haplotype of the presenting cells, as previously described by others, some hybridomas exhibited unique patterns of MHC fine specificity. Thus, T cell recognition of vSAG-9 and Mls-1 is not fundamentally different, but the apparent differences can be explained in terms of frequency. The MHC fine specificity of individual Mls-1-reactive hybridomas was influenced by both V beta and non-V beta T cell receptor (TCR) elements. First, the influence of the V beta element was apparent from the observation that V beta 8.2+ hybridomas were significantly more MHC specific in their recognition of Mls-1 than V beta 8.1 hybridomas. Second, a role for the TCR alpha chain was implicated from the distinct patterns of fine specificity of Mls-1 reactivity among a panel of transgenic hybridomas that expressed an identical beta chain (V beta 8.1D beta 2J beta 2.3C beta 2). Sequence analysis revealed that junctional residues of the TCR alpha chain and/or V alpha/J alpha combinations influenced the MHC haplotype fine specificity for Mls-1. Third, D beta J beta influences were implicated, in that the transgenic hybridomas expressed distinctive patterns of Mls-1 fine specificity not represented among V beta 8.1+ nontransgenic hybridomas. The findings that T cell recognition of endogenous superantigen is MHC specific, and that this specificity correlates with non-V beta elements of the TCR, support the hypothesis that there is a direct interaction between the TCR and either polymorphic residues of the MHC class II molecule or haplotype-specific dominant peptides presented by class II.

Endogenous superantigen expression is controlled by mouse mammary tumor proviral loci.

Superantigens are defined by their ability to stimulate T cells based predominantly on their V beta expression and ability to delete T cells in the thymus when expressed endogenously. We show here that the expression of one endogenous superantigen, Etc-1, is controlled by the expression of the open reading frame region of the 3' long terminal repeat of the mouse mammary tumor proviral gene, Mtv-9. We show that Mtv-8 controls a superantigen with similar specificity, and that both Mtv-8 and Mtv-9 stimulate some V beta 17+ T cells. A third provirus, Mtv-6, controls a superantigen with specificity for V beta 3. Data presented raise the possibility that endogenous superantigens may compete for class II molecules in a single B cell.

Pathogenesis of an infectious mononucleosis-like disease induced by a murine gamma-herpesvirus: role for a viral superantigen?

The murine gamma-herpesvirus 68 has many similarities to EBV, and induces a syndrome comparable to infectious mononucleosis (IM). The frequency of activated CD8+ T cells (CD62L(lo)) in the peripheral blood increased greater than fourfold by 21 d after infection of C57BL/6J (H-2(b)) mice, and remained high for at least a further month. The spectrum of T cell receptor usage was greatly skewed, with as many as 75% of the CD8+ T cells in the blood expressing a Vbeta4+ phenotype. Interestingly, the Vbeta4 dominance was also seen, to varying extents, in H-2(k), H-2(d), H-2(u), and H-2(q) strains of mice. In addition, although CD4 depletion from day 11 had no effect on the Vbeta4 bias of the T cells, the Vbeta4+CD8+ expansion was absent in H-2IA(b)-deficient congenic mice. However, the numbers of cycling cells in the CD4 antibody-depleted mice and mice that are CD4 deficient as a consequence of the deletion of MHC class II, were generally lower. The findings suggest that the IM-like disease is driven both by cytokines provided by CD4+ T cells and by a viral superantigen presented by MHC class II glycoproteins to Vbeta4+CD8+ T cells.

Control of gammaherpesvirus latency by latent antigen-specific CD8(+) T cells.

The contribution of the latent antigen-specific CD8(+) T cell response to the control of gammaherpesvirus latency is currently obscure. Some latent antigens induce potent T cell responses, but little is known about their induction or the role they play during the establishment of latency. Here we used the murine gammaherpesvirus system to examine the expression of the latency-associated M2 gene during latency and the induction of the CD8(+) T cell response to this protein. M2, in contrast to the M3 latency-associated antigen, was expressed at day 14 after infection but was undetectable during long-term latency. The induction of the M2(91-99)/K(d) CD8(+) T cell response was B cell dependent, transient, and apparently induced by the rapid increase in latently infected cells around day 14 after intranasal infection. These kinetics were consistent with a role in controlling the initial "burst" of latently infected cells. In support of this hypothesis, adoptive transfer of an M2-specific CD8(+) T cell line reduced the initial load of latently infected cells, although not the long-term load. These data represent the first description of a latent antigen-specific immune response in this model, and suggest that vaccination with latent antigens such as M2 may be capable of modulating latent gammaherpesvirus infection.

Influence of the major histocompatibility complex on positive thymic selection of V beta 17a+ T cells.

A monoclonal antibody was used to show directly positive thymic selection of the T cell repertoire in mouse strains expressing the 17a beta-chain variable domain (V beta 17a) of the T cell receptor. In the absence of the potent tolerizing class II major histocompatibility complex (MHC) molecule, I-E, peripheral expression of V beta 17a+ T cell receptors varied with the MHC haplotype of the mouse strain. In the most extreme case, H-2q mice expressed high peripheral levels of CD4+ V beta 17a+ T cells (14 to 19 percent), whereas H-2b mice expressed low levels (3 to 4 percent). Analysis of (b x q)F1 mice and chimeric mice showed that these differences were determined by positive thymic selection and implicated the thymic epithelium as the controlling cell type.

Natural killing of tumor cells by human peripheral blood cells. Suppression of killing in vitro by tumor-promoting phorbol diesters.

Tumor-promoting phorbol diesters were shown to suppress natural killing in vitro by human peripheral blood mononuclear cells. The inhibitory effect of different phorbol diesters and their analogues correlated with their potency as tumor promoters, the most effective agent being 12-O-tetradecanoylphorbol-13-acetate (TPA). Both peripheral blood cells and targets specifically bound TPA, and natural killing could be inhibited by pretreatment of either cell population with TPA, though this was less effective than direct addition of TPA to the assay. Cells that had been pretreated with TPA released TPA and metabolites of tPA during subsequent incubation in fresh medium. This release of tPA was evidently responsible for the inhibition of natural killing by pretreated target cells; in experiments where labeled and unlabeled target cells were mixed, pretreatment of unlabeled targets with TPA inhibited killing of labeled targets. Suppression of natural killing by TPA was greatly reduced when adherent cells were removed from the peripheral blood cells, suggesting that monocytes mediate suppression. Inhibition of natural killing by TPA provides a model for examining the regulation of natural killing. Suppression of natural killing by phorbol diesters may contribute to their activity as tumor promoters.

Suppression of natural killing in vitro by monocytes and polymorphonuclear leukocytes: requirement for reactive metabolites of oxygen.

Natural killer cells spontaneously lyse certain tumor cells and may defend against malignancy. We have previously shown that natural killing (NK) by human peripheral blood mononuclear cells (PBMC) is suppressed in vitro by phorbol diester tumor promoters, including 12-O-tetradecanoylphorbol-13-acetate (TPA). We here demonstrate that suppression of NK is mediated by monocytes or polymorphonuclear leukocytes (PMN) and that suppression is dependent on the generation of reactive forms of molecular oxygen (RO), particularly hydrogen peroxide (H2O2). NK was suppressed not only by TPA but also by opsonized zymosan (yeast cell walls), which, like TPA, was not toxic to PBMC. Both TPA and zymosan stimulated the production of superoxide anion (O2-) and H2O2 by PBMC. Production of RO correlated with suppression of NK. When PBMC were depleted of monocytes, the production of RO and the suppression of NK were both markedly reduced. Suppression could be restored by monocytes or PMN, both of which produced RO in response to TPA or zymosan. Suppression of NK was dependent on RO. Monocytes or PMN from a patient with chronic granulomatous disease, whose cells cannot generate RO, did not mediate suppression of NK. Suppression was also reduced in glucose-free medium, which did not support the generation of RO. Suppression of NK by TPA was inhibited by catalase. Bovine superoxide dismutase had a limited effect on suppression, even in high concentration, and tyrosine-copper (II) complex, which also enhances dismutation of O2- to H2O2, had almost no effect on suppression. When H2O2 was directly generated enzymatically from glucose oxidase and glucose, NK was suppressed and suppression was reversed by catalase. NK was also suppressed by the enzymatic generation of O2- from xanthine oxidase and xanthine, but suppression under these conditions was again inhibited by catalase and not by superoxide dismutase, indicating that suppression was due to the secondary formation of H2O2 from O2-. These results indicate that H2O2 is important in suppression of NK. Myeloperoxidase did not appear to play a role in suppression because inhibition of this enzyme by sodium azide, cyanide, or aminotriazole did not prevent suppression of NK. Suppression of NK was reversible; after exposure to zymosan, NK could be partially restored by the addition of catalase and superoxide dismutase or by the removal of zymosan. These studies demonstrate cellular regulation of NK by monocytes or polymorphonuclear leukocytes and indicate a role for RO in immunoregulation.

Tuning into immunological dissonance: an experimental model for infectious mononucleosis.

Virus infections cause a much more profound perturbation of the lymphoid tissue than can be accounted for by the exigencies of the antigen-specific response. The extent of this 'immunological dissonance' is seen most dramatically in mice infected with a persistent gamma-herpesvirus, MHV-68. A profile of massive, continuing proliferation of both T and B cells in the lymph nodes and spleen leads to a dramatic increase in the prevalence of a CD62Llow CD8+ T cell subset in the blood, a pattern first detected two to three weeks after intranasal exposure to the inducing virus. This syndrome, which seems identical to human infectious mononucleosis (IM), persists for a further month or more. Part of the IM-like phase of MHV-68 infection reflects the selective expansion of Vbeta4+ CD8+ T cells, with the Vbeta4 effect being apparent for several different MHC class I H-2 types but not in mice that are deficient in MHC class II glycoprotein expression. Depleting CD4(+) T helper cells in MHV-68-infected mice leads to the decreased proliferation of the CD8+ T cells in the spleen and fewer CD62Llow CD8+ T lymphocytes than would be expected in peripheral blood, but fails to diminish the prominence of the V4beta+ CD8+ population. The results so far of this unique experimental mouse model of IM suggest that both cytokine-mediated effects and a viral superantigen are operating to promote the dramatic expansion and persistence of activated CD8+ T cells in the vascular compartment.

Role of the T cell receptor alpha-chain in superantigen recognition.

Superantigens bind to antigen-presenting cells on the outside of the major histocompatibility complex (MHC) class II molecule and to T cells via the external face of the T cell receptor (TCR) V beta element. As a consequence, superantigens stimulate populations of T cells in a V beta-specific, non-MHC-restricted manner. However, accumulating evidence has shown an additional contribution of the TCR alpha-chain and polymorphic residues of the MHC molecule to superantigen recognition by some T cells. These data suggest that the TCR and MHC come into contact during superantigen engagement and indirectly modulate the superantigen reactivity. Thus, additional interactions between non-V beta elements of the TCR and MHC play a role in the overall stability of the superantigen/MHC/TCR complex, explaining the influence of the TCR alpha-chain. It is likely that this additional interaction is of greater consequence for weakly reactive T cells. This modulation of superantigen reactivity in individual T cells may have physiological consequences, for example, in the induction of autoimmunity.

Limiting the available T cell receptor repertoire modifies acute lymphocytic choriomeningitis virus-induced immunopathology.

The invariably fatal immunopathological disease that follows intracerebral injection of CBA/Ca (H-2k) mice with 1000 PFU of lymphocytic choriomeningitis virus (LCMV) generally fails to develop in congenic mice transgenic for a V beta 8.1D beta 2J beta 2.3C beta 2 T cell receptor (TCR) gene. The majority of these LCMV-infected TCR-transgenic mice show a substantial meningitis of delayed onset, that resolves without causing any obvious clinical impairment. This inflammatory process depends on the involvement of V beta 8+ T cells, but does not require the participation of the CD4+ subset. The cytotoxic effectors that develop in both the transgenic mice and the CBA/Ca controls are lytic for target cells infected with a vaccinia construct expressing genes encoding the putative polymerase protein of LCMV. Limiting the available TCR repertoire to lymphocytes with a single V beta phenotype (not required for the generation of potent effectors in wild-type mice) thus modifies the development of the lethal neuropathology characteristic of LCMV infection, although the CD8+ cytotoxic T lymphocyte response is not greatly compromised.

Vaccination against murine gamma-herpesvirus infection.

The gamma-herpesviruses establish life-long latency in the host and are important human pathogens. T cells play a major role in controlling the initial acute infection and subsequently maintaining the virus in a quiescent state. However, the nature of the T-cell response to gamma-herpesvirus infection and the requirements for effective vaccination are poorly understood. The recent development of a murine gamma-herpesvirus (murine herpesvirus-68 [MHV-68]) has made it possible to analyze T-cell responses and test vaccination strategies in a small animal model. Intranasal infection with MHV-68 induces an acute infection in the lung and the subsequent establishment of long-term latency, which is associated with splenomegaly and an infectious mononucleosis-like syndrome. Here we review the T-cell response to different phases of the infection and the impact of vaccination against either lytic-cycle, or latency-associated T-cell epitopes.

Factors controlling levels of CD8+ T-cell lymphocytosis associated with murine gamma-herpesvirus infection.

Intranasal infection of mice with murine gamma-herpesvirus 68 (MHV-68) elicits a striking CD8+ T-cell lymphocytosis following the establishment of latency, which includes a marked increased frequency of Vbeta4+ CD8+ T cells. The Vbeta4+ CD8+ T cells do not recognize a conventional viral peptide, but are stimulated by an uncharacterized ligand expressed on latently infected, activated B cells. The selective expansion of Vbeta4+ CD8+ T cells after MHV-68 infection is observed in all mouse strains examined, although the fold-increase varies widely, ranging from less than twofold to greater than 10-fold. The factors controlling the variation are currently undefined. In the current study, CD8+ T cell activation and Vbeta4+ CD8+ T-cell frequencies were analyzed in 18 inbred strains of mice. The data show that the magnitude of the Vbeta4+ CD8+ T-cell response correlates with the degree of CD8+ T cell-activation, and that both major histocompatibility complex (MHC) and non-MHC genes contribute to the magnitude of the activation. Furthermore, the magnitude of the response does not reflect major differences in susceptibility to viral infection and/or corresponding differences in the acute response. Rather the degree of Vbeta4+ CD8+ T cell activation may be determined by differences in levels of expression of the stimulatory ligand at the peak of latency.

Retroviral super-antigens and T cells.

For many years immunologists have been intrigued by a series of potent antigens encoded in the murine genome. These antigens, originally termed minor lymphocyte stimulating (Mls) antigens, are capable of inducing extremely strong T cell proliferative responses when presented in the context of MHC class II molecules. Recently, Mls antigens have been shown to stimulate T cells bearing particular T cell receptor V beta elements, leading to the designation of super-antigens. The endogenous expression of these super-antigens in mice results in the clonal elimination of large numbers of T cells in order to maintain self-tolerance. In this review we discuss the recent identification of endogenous super-antigens as retroviral gene products. In addition, we analyze the role of class II MHC molecules in the presentation of endogenous super-antigens to T cells. Finally, we discuss the dramatic effect of retroviral super-antigens on the T cell repertoire.

Antigen expression during murine gamma-herpesvirus infection.

Gammaherpesviruses (gammaHV) establish a life-long latency in the host and are associated with a number of malignant human diseases. It is generally believed that T cells play a major role in controlling the initial acute infection and subsequently maintaining the virus in a quiescent state. However, the nature of the T cell response to gamma-herpesvirus infections is poorly understood. In the current report we took advantage of a mouse model of gammaHV infection (murine herpesvirus-68, MHV-68) to investigate the T cell response to different phases of the infection. Intranasal infection with MHV-68 induces an acute infection in lung epithelial cells and long-term latency in B cells. The kinetics of the CD8+ T cell response to different lytic cycle and latency-associated antigens was highly complex and distinct patterns of response could be identified. These responses were regulated by multiple factors including differences in temporal expression of the relevant antigens, differences in the presentation of antigen in different organs, and differential expression of antigen in different types of antigen presenting cells. For example, some antigens were expressed at distinct phases of the infection and in specific organs or subsets of antigen presenting cells. In addition, recent data suggest that in addition to B cells, both macrophages and dendritic cells harbor latent MHV-68 infection, adding further complexity to their role in controlling the T cell response to this infection.

In vivo effects of superantigens.

Superantigens are potent immunostimulatory molecules that activate both T cells and antigen presenting cells. The consequences of superantigen exposure range from induction of T cell proliferation, massive cytokine release and systemic shock to immunosuppression and tolerance. Superantigens have been directly implicated in a number of human conditions including food poisoning and toxic shock. In addition, there is evidence to suggest that superantigens are involved in the initiation of autoimmunity, and the immune dysfunction associated with HIV infection. Because of their possible role in human disease, and their potential use in immune therapy, it is important that we more completely understand the in vivo effects of superantigens.

Persistent transmission of mouse hepatitis virus by transgenic mice.

Variation in susceptibility to viral infection is well documented across mouse strains. Specific combinations of viral strains and murine hosts may favor viral infection and disease, and could potentially allow the unexpected development of chronic, persistent, or latent infections. In some genetically modified strains of mice, immune function and perhaps other physiologic or metabolic systems may be substantially or marginally impaired. In the case study reported here, we document the apparent persistent transmission of mouse hepatitis virus (MHV) over a two-year period by MHV-seropositive transgenic mice. Transmission occurred via direct contact with seropositive mice and exposure to contaminated bedding. However, MHV was not detected at diagnostic laboratories by use of viral isolation or reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of tissues from MHV-seropositive animals. Our observation, together with the constantly expanding varieties of immune-impaired or poorly characterized murine hosts and the burgeoning dissemination of these animals throughout the biomedical research community, indicate that unexpected pathophysiologic presentations of common murine viral diseases may present new challenges to the biomedical research community in the future.

The Yin and Yang of inflammation.

Inflammation is an essential protective part of the body's response to infection, yet many diseases are the product of inflammation. For example, inflammation can lead to autoimmune disease and tissue damage, and is a key element in chronic health conditions such as heart disease, diabetes, rheumatoid arthritis, and also drives changes associated with aging. Animal models of infectious and chronic disease are important tools with which to dissect the pathways whereby inflammatory responses are initiated and controlled. Animal models therefore provide a prism through which the role of inflammation in health and disease can be viewed, and are important means by which to dissect mechanisms and identify potential therapies to be tested in the clinic. A meeting, "The Yin and Yang of Inflammation" was organized by Trudeau Institute and was held between April 4-6, 2014. The main goal was to bring together experts from biotechnology and academic organizations to examine and describe critical pathways in inflammation and place these pathways within the context of human disease. A group of ~80 scientists met for three days of intense formal and informal exchanges. A key focus was to stimulate interactions between basic research and industry.