The surprising kinetics of the T cell response to live antigenic cells.

Cooperation between CD4(+) and CD8(+) T cells is required for the proper development of primary effector and memory CD8(+) T cells following immunization with noninflammatory immunogens. In this study, we characterized murine CD4(+) and CD8(+) T cell responses to male-specific minor histocompatibility (HY) Ags following injection of live male cells into females of the same strain. Male cells are rejected 10-12 days after transfer, coinciding with the expansion and effector function of CD8(+) CTLs to two H-2D(b)-restricted epitopes. Although anti-HY CD4(+) T cell responses are readily detectable day 5 posttransfer, CD8(+) responses are undetectable until day 10. The early CD4(+) response is not dependent on direct presentation of Ag by donor male cells, but depends on presentation of the male cells by recipient APC. The CD4(+) T cell response is required for the priming of CD8(+) T cell effector responses and rejection of HY-incompatible cells. Unexpectedly, HY-specific CD4(+) T cells are also capable of efficiently lysing target cells in vivo. The delay in the CD8(+) T cell response can be largely abrogated by depleting T cells from the male inoculum, and donor male CD8(+) T cells in particular suppress host anti-HY CD8(+) responses. These data demonstrate dramatic differences in host T cell responses to noninflammatory Ags compared with responses to pathogens. We explain the delayed CD8(+) response by proposing that there is a balance between cross-presentation of Ag by helper cell-licensed dendritic cells, on the one hand, and veto suppression by live male lymphocytes on the other.

Functional implication of cellular prion protein in antigen-driven interactions between T cells and dendritic cells.

The cellular prion protein (PrPC) is a host-encoded, GPI-anchored cell surface protein, expressed on a wide range of tissues including neuronal and lymphoreticular cells. PrPC may undergo posttranslational conversion, giving rise to scrapie PrP, the pathogenic conformer considered as responsible for prion diseases. Despite intensive studies, the normal function of PrPC is still enigmatic. Starting from microscope observations showing an accumulation of PrPC at the sites of contact between T cells and Ag-loaded dendritic cells (DC), we have studied the contribution of PrPC in alloantigen and peptide-MHC-driven T/DC interactions. Whereas the absence of PrPC on the DC results in a reduced allogeneic T cell response, its absence on the T cell partner has no apparent effect upon this response. Therefore, PrPC seems to fulfill different functions on the two cell partners forming the synapse. In contrast, PrPC mobilization by Ab reduces the stimulatory properties of DC and the proliferative potential of responding T cells. The contrasted consequences, regarding T cell function, between PrPC deletion and PrPC coating by Abs, suggests that the prion protein acts as a signaling molecule on T cells. Furthermore, our results show that the absence of PrPC has consequences in vivo also, upon the ability of APCs to stimulate proliferative T cell responses. Thus, independent of neurological considerations, some of the evolutionary constraints that may have contributed to the conservation of the Prnp gene in mammalians, could be of immunological origin.

The male minor transplantation antigen preferentially activates recipient CD4+ T cells through the indirect presentation pathway in vivo.

To evaluate the priming and trafficking of male Ag-reactive CD4(+) T cells in vivo, we developed an adoptive transfer model, using Marilyn (Mar) TCR transgenic T cells that are specific for the H-Y minor transplantation Ag plus I-A(b). By manipulating donor and recipient strain combinations, we permitted the Mar CD4(+) T cells to respond to the H-Y Ag after processing and presentation by recipient APCs (indirect pathway), or to the male Ag as expressed on donor APCs (direct pathway). Mar CD4(+) T cells responding through the indirect pathway specifically proliferated and expressed activation markers between days 2 and 4 posttransplant, migrated to the graft 2-3 days before cessation of graft heartbeat, and were detected in close proximity to transplant-infiltrating recipient APCs. Intriguingly, adoptively transferred Mar T cells did not respond to male heart or skin grafts placed onto syngeneic MHC class II-deficient female recipients, demonstrating that activation of Mar T cell preferentially occurs through cognate interactions with processed male Ag expressed on recipient APCs. The data highlight the potency of indirect processing and presentation pathways in vivo and underscore the importance of indirectly primed CD4(+) T cells as relevant participants in both the priming and effector phases of acute graft rejection.

The affinity/avidity and length of exposure to the deleting ligand determine dependence on CD28 for the efficient deletion of self-specific CD4+CD8+ thymocytes.

Whether the CD28/B7 signaling pathway is essential for the negative selection of immature CD4+CD8+ (DP) thymocytes expressing self-specific alphabeta TCRs is a controversial issue. In this study we examined the role of CD28 in the deletion of thymocytes that express either the H-Y or the 2C transgenic TCR. In H-2(b) male mice that expressed the H-Y TCR, negative selection of DP H-Y TCR+ thymocytes occurred very efficiently and this deletion was unaffected by the CD28(-/-) mutation. In H-2(b) 2C mice, where the deletion of DP 2C TCR+ thymocytes occurred less efficiently, the CD28(-/-) mutation led to a higher recovery of DP thymocytes. Using an in vitro deletion assay, a requirement for the CD28 signaling pathway in the deletion of DP H-Y TCR+ thymocytes was evident at low, but not high, densities of the antigenic ligand. Similar results were also observed in an in vivo assay for the deletion of these thymocytes. Intraperitoneal administration of an anti-CD3epsilon mAb led to the intrathymic deletion of DP H-Y TCR+ thymocytes in a CD28-dependent manner at the 24-h time point. However, the CD28 dependence was less evident at the 40-h time point. These results indicate that the dependence on CD28 for the efficient deletion of self-specific thymocytes is determined by the concentration, affinity/avidity, and length of exposure to the deleting ligand.

The regulation and function of the Id proteins in lymphocyte development.

Helix-loop-helix (HLH) proteins are essential factors for lymphocyte development and function. One class of HLH proteins, the E-proteins, regulate many aspects of lymphocyte maturation, survival, proliferation, and differentiation. E-proteins are negatively regulated by another class of HLH proteins known as the Id proteins. The Id proteins function as dominant negative inhibitors of E-proteins by inhibiting their ability to bind DNA. Here we discuss the function and regulation of the Id proteins in lymphocyte development.

TNF receptor-deficient mice reveal striking differences between several models of thymocyte negative selection.

Central tolerance depends upon Ag-mediated cell death in developing thymocytes. However, the mechanism of induced death is poorly understood. Among the known death-inducing proteins, TNF was previously found to be constitutively expressed in the thymus. The role of TNF in thymocyte negative selection was therefore investigated using TNF receptor (TNFR)-deficient mice containing a TCR transgene. TNFR-deficient mice displayed aberrant negative selection in two models: an in vitro system in which APC are cultured with thymocytes, and a popular in vivo system in which mice are treated with anti-CD3 Abs. In contrast, TNFR-deficient mice displayed normal thymocyte deletion in two Ag-induced in vivo models of negative selection. Current models of negative selection and the role of TNFR family members in this process are discussed in light of these results.

Minor histocompatibility antigens HA-1-, -2-, and -4-, and HY-specific cytotoxic T-cell clones inhibit human hematopoietic progenitor cell growth by a mechanism that is dependent on direct cell-cell contact.

HLA-identical bone marrow transplantation (BMT) may be complicated by graft-versus-host disease or graft rejection. Both complications are thought to be initiated by recognition of minor histocompatibility (mH) antigens by HLA-restricted mH-antigen-specific T lymphocytes. Using HLA-A2-restricted mH antigens HA-1-, -2-, and -4-, and HY-specific cytotoxic T lymphocyte (CTL) clones, we studied the recognition by these CTL clones of interleukin-2 (IL-2)-stimulated T cells (IL-2 blasts), BM mononuclear cells (BMMNCs), and hematopoietic progenitor cells (HPCs). We showed that, when IL-2 blasts from the BM donors who were investigated were recognized by the HA-1-, -2-, and -4-, and HY-specific CTL clones, their BMMNCs and HPCs were recognized as well by these CTL clones, resulting in antigen-specific growth inhibition of erythrocyte burst-forming units (BFU-E), colony-forming units-granulocyte (CFU-G), and CFU-macrophage (CFU-M). the HA-2-specific CTL clone, however, inhibited BFU-E and CFU-G growth from four donors to a lesser extent than from two other donors. We further investigated whether inhibitory cytokines released into the culture medium by the antigen-specific stimulated CTLs or by stimulated BMMNCs were responsible for suppression of HPC growth or whether this effect was caused by direct cell-cell contact between CTLs and HPCs. HPC growth inhibition was only observed after preincubation of BMMNCs and CTLs together for 4 hours before plating the cells in semisolid HPC culture medium. When no cell-cell contact was permitted before plating, neither antigen-stimulated CTL nor antigen-nonstimulated CTLs provoked HPC growth inhibition. Culturing BMMNCs in the presence of supernatants harvested after incubation of BMMNCs and CTL clones together for 4 or 72 hours did also not result in HPC growth inhibition. Both suppression of HPC growth and lysis of IL-2 blasts and BMMNCs in the 51Cr-release assay appeared to be dependent on direct cell-cell contact between target cells and CTLs and were not caused by the release of inhibitory cytokines into the culture medium by antigen-specific stimulated CTLs or by stimulated BMMNCs. Our results show that mH-antigen-specific CTLs can inhibit HPC growth by a direct cytolytic effect and may therefore be responsible for BM graft rejection after HLA-identical BMT.

[The human Y chromosome]. / El cromosoma Y humano.

Recent findings concerning the human Y chromosome are reviewed. Normally the presence of the Y chromosome is related to male differentiation; however until recently nothing was known about chromosome Y and the development of the testis at the molecular level. Initially the most plausible assumption in this field was the "H-Y hypothesis" which suggested that the presence of a male specific antigen called H-Y, controlled by a gene(s) on the Y chromosome, was responsible for the differentiation of the primitive gonad into a testis. An alternative was a series of DNA sequences termed Bkm which were thought to play an important role in sex determination. These two hypothesis were refuted. More recently, molecular studies in XX males have shown the presence of Y-DNA in their genome suggesting that a small fragment of the Y (probably a single gene) is responsible for testis development in the majority of these patients. This gene has been termed TDF or ZFY and encodes a protein that could act as a regulating factor of the male pathway. Also reviewed are the actions of various genes recently assigned to the Y chromosome.

Serological H-Y antigen and gonadal status in XYDOM sex-reversed mice.

Y chromosomes of feral mice (Mus musculus domesticus) from various localities, when introduced into the C57BL/6 laboratory strain, give rise to phenotypic females and true hermaphrodites both with the sex chromosome constellation XY. Sex-reversed animals of each type were examined macroscopically or histologically for gonadal status and H-Y antigenic activity by serological assay methods. Most XY females with histologically confirmed bilateral ovaries did not differ from XX female controls with respect to serological H-Y antigen, i.e. they were H-Y negative. The true hermaphrodites were H-Y positive, though H-Y antigenic activity was intermediate to male and female controls in the majority of cases. The findings support a relationship between the presence of serological H-Y antigen and gonadal status.

Serological H-Y antigen in the female chicken occurs during gonadal differentiation.

In the chicken, serological H-Y antigen is specific for the female sex. Male gonad differentiation can be experimentally influenced by estrogens, resulting in the transient formation of an ovotestis. The sex-inverted gonad becomes positive for H-Y antigen. Therefore, the question arises whether, in normal gonadogenesis also, the female gonad at the indifferent stage, before estrogens are produced, is negative for H-Y antigen. Here we show that this is indeed the case. The female gonad becomes positive for H-Y antigen when the ovary starts its organotypic differentiation at about day 6 1/2 of embryonal development. It is assumed that estrogens are responsible for the occurrence of H-Y antigen. This finding supports the view that H-Y antigen plays a role in primary ovogenesis in the chicken.

An in vitro model of gonad differentiation in the chicken. Estradiol-induced sex-inversion results in the occurrence of serological H-Y antigen.

Dissociated cells from the gonads and mesonephros of 8-day-old chicken embryos were reorganized in rotation culture. The aggregates obtained from gonadal cells exhibited specific morphologic and histologic sex differences. In the presence of estradiol, aggregates from testicular cells showed characteristics similar to control ovarian aggregates, while in ovarian aggregates under estradiol treatment the female organization became more pronounced. Determination of serological H-Y antigen revealed that male aggregates of gonads and mesonephros were negative for H-Y and those of female embryos were positive for H-Y. Administration of estradiol did not change the H-Y findings in female aggregates. In contrast, in the male, gonadal cultures became H-Y positive while mesonephros cultures remained negative. It is assumed that estradiol induces the occurrence of H-Y antigen in the gonads.

Sexological theory, H-Y antigen, chromosomes, gonads, and cyclicity: two syndromes compared.

The androgen-insensitivity syndrome (AIS) and Rokitansky syndrome (RS) are clinically similar except for H-Y antigen, chromosomal sex, gonadal sex, and the cyclicity of sex hormonal functioning. Nine AIS women and nine RS women could not be distinguished systematically on any of the following variables: romantic and genitoerotic imagery/ideation; sensory channels of erotic arousal; gender orientation; satisfaction with feminine status; self-rating of sexual frequency and interest; masturbation experience; orgasm experience; attitude toward marriage; and attitude toward infant and child care. These findings indicate that neither H-Y antigen status, chromosomal sex, nor hormonal cyclicity directly determines the differentiation of gender-identity/role (G-I/R) as male, female, or ambiguous. By implication, these findings are relevant to heterosexual/homosexual theory.

Spermatogenic failure in male mice lacking H-Y antigen.

The mammalian Y chromosome carries a factor that initiates male sexual development by directing the fetal gonads to form testes. Wachtel and his colleagues proposed that this testis-determining function of the Y is mediated by the male-specific cell-surface antigen H-Y, originally defined by skin grafting. This attractive hypothesis, which has been widely accepted, was based on the assumption that serological tests using antisera raised against male cells were recognizing H-Y antigen. Although disputed this assumption is supported by some recent studies. However, mice have been described which develop testes but lack the cell-surface H-Y antigen as defined by T-cell-mediated transplantation tests. Thus, although it remains possible that a serologically detected male-specific antigen is responsible for testis determination, it seems that H-Y, as originally defined, is not. We show here that H-Y negative male mice, in losing the genetic information that encodes H-Y, have also lost genetic information required for spermatogenesis. This result identifies a gene on the mouse Y, distinct from the testis-determining gene, which is necessary for spermatogenesis, and raises the intriguing possibility that the product of this 'spermatogenesis gene' is H-Y antigen.

The H-Y antigen and its functions: a review and a hypothesis.

Having reviewed the status of H-Y as the sex-determining antigen concerned with the differentiation of the dominant gonad, we consider some of the problems deriving from the tests for this antigen, and from their application to the study of natural experiments. To reconcile the results of these studies with the alleged influence of H-Y on gonadal development, we propose and discuss a hypothesis on the genetic control of the synthesis of this antigen. This states that an autosomally-coded, positively cross-reacting precursor is rendered biologically active by a Y-chromosomal gene, and transformed (in a dose-dependent manner) into a biologically inactive, antigenically negative substance under the influence of an X-chromosomal gene.