CD4+ T cell responses in mice lacking MHC class II molecules specifically on B cells

The role of B lymphocytes in initiating and maintaining a CD4+ T cell response has been examined using a variety of strategies, but remains controversial because of weaknesses inherent to each of the approaches. Here, we address this issue by measuring CD4+ T cell priming both in mutant mice devoid of B cells and in chimeric animals lacking major histocompatibility complex class II molecules specifically on B cells. We find that peptide and some protein antigens do not require B cells expressing class II molecules, nor B cells themselves, to efficiently prime. This could be demonstrated by the usual lymph node proliferation assay, a rather indirect in vitro measure of priming, and by a direct ex vivo assay of population expansion and activation marker expression. Interestingly, one protein antigen, conalbumin, could not prime in the absence of B cells, but could in the presence of B cells devoid of class II molecules. This finding constrains the possible mechanisms whereby B lymphocytes contribute to the initiation of a CD4+ T cell response, arguing against the importance of surface immunoglobulin-mediated antigen presentation by B cells.     

Effects of cytokines and glucocorticoids on endogenous class II MHC antigen expression by activated rat CD4+ T cells. Mature CD4+ CD8- thymocytes are phenotypically heterogeneous on activation

By the use of mixed leukocyte cultures it was shown that a population of allogeneically activated rat T cells synthesize and express class II MHC antigens, in confirmation of other studies. Compatible with the finding that the MHC molecules detected on these cells were of T cell origin rather than passively acquired, it was found that mRNA for class II transactivator could readily be detected in the T cells stimulated in these cultures. In contrast there was no evidence that mouse T cells synthesized class II MHC antigens. The size of the population of activated rat T cells expressing class II MHC antigens was affected by the presence of IL-4 and glucocorticoids in the activating cultures. However, whereas IL-4 increased the frequency of thymocytes and peripheral T cells expressing class II antigens in culture, glucocorticoids diminished this frequency. The expression of class II MHC antigens by allogeneically activated thymocytes demonstrated a novel heterogeneity amongst mature CD4+ CD8- thymocytes that could not readily be accounted for in terms of differences in maturity of the cells, in the affinity of the TCR for the stimulating ligands or in the stage in the cell cycle. The data suggest that CD4+ single-positive thymocytes do not constitute a homogeneous population differing only in TCR clonotypes.     

Adoptively transferred CD4+ lymphocytes from CD8 -/- mice are sufficient to mediate the rejection of MHC class II or class I disparate skin grafts

Recent studies revealed that CD4+ cells initiate allograft rejection through direct recognition of allogeneic MHC class II Ags and indirect recognition of MHC peptides processed by self APCs. Both pathways were shown to help CD8+ cells that eventually lysed allogeneic MHC class I-presenting targets. There was little evidence, however, that CD4+ cells are sufficient for graft rejection. We studied skin graft rejection by CD8-deficient (CD8 -/-) mice. We showed that BALB/cJ(H-2d) CD8 -/- mice could reject allogeneic skin from C57BL/6J(H-2b) mice deficient in MHC class I or in MHC class II Ags. To understand the role of CD4+ cells in this process, we isolated them from CD8 -/- mice and transferred them to BALB/cJ nude mice that had been grafted with allogeneic skin (H-2b) from animals deficient in MHC class I or MHC class II. Nude mice injected with CD4+ cells rejected MHC class II and, albeit more slowly, MHC class I disparate skins. We showed in vitro evidence that CD4+ cells were not cytotoxic toward MHC class I or MHC class II disparate targets and that they recognized MHC class I allogeneic targets through indirect recognition. CD4+ cells produced Th1 cytokines, but not IL-4, following stimulation with allogeneic cells. Furthermore, intragraft TNF-alpha was elevated in skin grafted onto nude mice reconstituted with CD4+ cells compared with nonreconstituted mice. This suggests that MHC class II- or MHC class I-guided CD4+ cells alone are sufficient to induce rejection by the generation of cytokine-induced lesions.     

MHC class II-transfected tumor cells directly present antigen to tumor-specific CD4+ T lymphocytes

We have developed and shown to be efficacious an immunotherapeutic strategy to enhance the generation of tumor-specific CD4+ T helper lymphocytes. The approach uses autologous tumor cells genetically modified to express syngeneic MHC class II genes as cell-based immunogens and is based on the hypothesis that tumor cells directly present tumor Ags to CD4+ T cells. Since the conventional pathway for CD4+ T cell activation is indirect via professional APC, induction of immunity following immunization with class II-transfected tumor cells was examined in bone marrow chimeric mice. Both tumor and host-derived cells are APC for tumor Ags, suggesting that the efficacy of tumor cell vaccines can be significantly improved by genetic modifications that enhance tumor cell Ag presentation.     

Cellular mechanisms involved in protection against influenza virus infection in transgenic mice expressing a TCR receptor specific for class II hemagglutinin peptide in CD4+ and CD8+ T cells

Mice transgenic for a TCR that recognizes peptide110-120 of hemagglutinin of PR8 influenza virus in the context of MHC class II I-Ed molecules express the transgenes in both CD4+ and CD8+ T cells. We have found that these TCR-hemagglutinin (TCR-HA) transgenic mice display a significantly increased resistance to the primary infection with PR8 virus compared with the wild-type mice. The TCR-HA transgenic mice mounted significant MHC type II and enhanced MHC type I-restricted cytotoxicity as well as increased cytokine responses in both spleen and lungs after infection with PR8 virus. In contrast, the primary humoral response against PR8 virus was not significantly different from that of the wild-type mice. In vivo depletion and adoptive cell transfer experiments demonstrated that both CD4+ and CD8+ TCR-HA+ T cell subsets were required for the complete clearance of pulmonary virus following infection with a dose that is 100% lethal in wild-type mice. Whereas CD4+ TCR-HA+ T cells were necessary for effective activation and local recruitment of CD8+ T cells, CD8+ TCR-HA+ T cells showed a Th1-biased pattern and MHC type II-restricted cytotoxicity. However, in the absence of in vivo expression of MHC type I molecules on the infected cells, the protection conferred by the TCR-HA+ T cells was impaired, indicating that the enhanced MHC class I-restricted cytotoxicity due to TCR-HA+ CD4+ Th cells was a critical element for clearance of the pulmonary virus by the transgenic mice.     

Coreceptor-independent T cell activation in mice expressing MHC class II molecules mutated in the CD4 binding domain

We have previously reported that efficient selection of the mature CD4+ T cell repertoire requires a functional interaction between the CD4 coreceptor on the developing thymocyte and the MHC class II molecule on the thymic epithelium. Mice expressing a class II protein carrying the EA137/VA142 double mutation in the CD4 binding domain develop fewer than one-third the number of CD4+ T cells found in wild-type mice. In this report we describe the functional characteristics of this population of CD4+ T cells. CD4+ T cells that develop under these conditions are predicted to be a CD4-independent subset of T cells, bearing TCRs of sufficient affinity for the class II ligand to undergo selection despite the absence of accessory class II-CD4 interactions. We show that CD4+ T cells from the class II mutant mice are indeed CD4 independent in their peripheral activation requirements. Surprisingly, we find that CD4+ T cells from the class II mutant mice, having been selected in the absence of a productive class II-CD4 interaction, fail to functionally engage CD4 even when subsequently provided with a wild-type class II ligand. Nevertheless, CD4+ T cells from EA137/VA142 class II mutant mice can respond to T-dependent Ags and support Ig isotype switching.     

Cytotoxic CD4+ T cells associated with the expression of major histocompatibility complex class II antigen of mouse myeloma cells secreting interferon-gamma are cytolytic in vitro and tumoricidal in vivo

The kidney is responsible for regulation of water and electrolyte balance, filtration and absorption of plasma proteins, and control of blood volume and pressure. Homeostasis achieved by the kidney is controlled in large part by the action of hormones or proteins on specific transmembrane receptors. Conversely, many renal diseases, including that resulting from atherosclerosis, are characterised by scarring and abnormal proliferation of cellular components of the kidney, and these processes are mediated in large part by these same receptors. The G protein-coupled receptors constitute a large and diverse class of proteins, characterised by the possession of seven transmembrane-spanning domains. These receptors bind polypeptide growth factors, which function to transmit a variety of signals from the extracellular to the intracellular milieu. The receptor-associated G proteins utilised by the kidney derive their specificity not only by activating or inhibiting various second-messenger molecules, but also by their location on particular cell types. In this review, several G protein-coupled receptors will be discussed from the perspective of their importance to kidney function and to the pathogenesis of renal disease, atherosclerosis, and hypertension.     

Alterations in CD4-binding regions of the MHC class II molecule I-Ek do not impede CD4+ T cell development

The T cell coreceptors CD4 and CD8 enhance T cell responses to TCR signals by participating in complexes containing TCR, coreceptor, and MHC molecules. These ternary complexes are also hypothesized to play a seminal role during T cell development, although the precise timing, frequency, and consequences of TCR-coreceptor-MHC interactions during positive selection and lineage commitment remain unclear. To address these issues, we designed transgenic mice expressing mutant I-Ek molecules with reduced CD4-binding capability. These transgenic lines were crossed to three different lines of I-Ek-specific TCR transgenic mice, and the efficiency of production of CD4+ lineage cells in the doubly transgenic progeny was assessed. Surprisingly, replacing wild-type I-Ek molecules with these mutant molecules did not affect the production of CD4+CD8- thymocytes or CD4+ peripheral T cells expressing any of the three TCRs examined. These data, when considered together with other experiments addressing the role of coreceptor during development, suggest that not all MHC class II-specific thymocytes require optimal and simultaneous TCR-CD4-MHC interactions to mature. Alternatively, it is possible that these particular alterations of I-Ek do not disrupt the CD4-MHC interaction adequately, potentially indicating functional differences between I-A and I-E MHC class II molecules.     

Protective antibodies develop, and murine Lyme arthritis regresses, in the absence of MHC class II and CD4+ T cells

Murine Lyme borreliosis is characterized by arthritis and carditis that are most severe at 2 to 3 wk, then regress during the course of persistent infection. Borrelia burgdorferi-specific Abs and CD4+ T cells have been implicated in the resolution phase of arthritis. Therefore, MHC class II transactivator (CIITA)-deficient mice that do not express conventional class II molecules and lack the normal CD4 repertoire were used to investigate the role of MHC class II-mediated responses in Lyme disease. The development of arthritis and carditis, and the resolution of arthritis, were similar in CIITA-deficient and control C57/BL6 mice. In contrast, the resolution of carditis was delayed in CIITA-deficient animals compared with controls. Moreover, CIITA-deficient mice developed B. burgdorferi-specific IgG2b Abs, and sera from these animals passively protected naive C3H/HeN mice from challenge inoculation and cleared B. burgdorferi from 2 day-infected C.B.17 SCID mice. These data suggest that CD4+ T cells and MHC class II-mediated responses are not required for the generation of protective Abs or the regression of arthritis, but may be important in the resolution of Lyme carditis in mice.     

In the normal repertoire of CD4+ T cells, a single class II MHC/peptide complex positively selects TCRs with various antigen specificities

In the thymus, immature T cells are positively and negatively selected by multiple interactions between their Ag receptors (TCRs) and self MHC/peptide complexes expressed on thymic stromal cells. Here we show that in the milieu of negative selection on physiological self class II MHC/peptide complexes (Abwt), a single class II/peptide complex AbEp52-68 positively selects a number of TCRs with various Ag specificities. This TCR repertoire is semidiverse and not biased toward Ep-like Ags. Our finding implies that the degeneracy of positive selection for peptide ligands exceeds peptide-specific negative selection and is essential to increase the efficiency and diversity of the repertoire so that T cells with the same Ag specificity can be selected by different self MHC/ peptide complexes.     

The response of gamma delta T cells to Plasmodium falciparum is dependent on activated CD4+ T cells and the recognition of MHC class I molecules

Peripheral blood gamma delta T cells from non-exposed individuals respond to antigens of the malaria parasite, Plasmodium falciparum, in vitro. This response, largely caused by T cells bearing the V gamma 9+ chain of the T-cell receptor, is stimulated by components of the parasite expressed on the schizont stage and released at schizont rupture. The response of V gamma 9+ T cells to parasite components is inhibited by antibodies to major histocompatibility complex (MHC) class I and class II. However, the inhibition by anti-MHC class II antibodies can be overcome by the addition of interleukin-2 (IL-2) to the cultures, suggesting that gamma delta T cells themselves do not recognize MHC class II molecules but require an MHC class II-dependent response taking place in the culture. In contrast, the inhibition by anti-class I antibodies cannot be reversed by addition of IL-2. Since an accompanying CD4+ T-cell response occurred in peripheral blood mononuclear cells cultured with P falciparum antigens, it was considered that these cells provide the cytokines necessary for the subsequent activation and expansion of V gamma 9+ T cells recognizing components of the parasite and MHC class I molecules. This was confirmed by reconstituting the response of enriched gamma delta T cells to P falciparum schizont extract by addition of purified CD4+ T cells.     

Major histocompatibility complex (MHC) class I KbDb -/- deficient mice possess functional CD8+ T cells and natural killer cells

We obtained mice deficient for major histocompatibility complex (MHC) molecules encoded by the H-2K and H-2D genes. H-2 KbDb -/- mice express no detectable classical MHC class I-region associated (Ia) heavy chains, although beta2-microglobulin and the nonclassical class Ib proteins examined are expressed normally. KbDb -/- mice have greatly reduced numbers of mature CD8+ T cells, indicating that selection of the vast majority (>90%) of CD8+ T cells cannot be compensated for by beta2-microglobulin-associated molecules other than classical H-2K and D locus products. In accord with the greatly reduced number of CD8+ T cells, spleen cells from KbDb -/- mice do not generate cytotoxic responses in primary mixed-lymphocyte cultures against MHC-disparate (allogeneic) cells. However, in vivo priming of KbDb -/- mice with allogeneic cells resulted in strong CD8+ MHC class Ia-specific allogeneic responses. Thus, a minor population of functionally competent peripheral CD8+ T cells capable of strong cytotoxic activity arises in the complete absence of classical MHC class Ia molecules. KbDb -/- animals also have natural killer cells that retain their cytotoxic potential.     

Killing of rat adenocarcinoma 13762 in situ by adoptive transfer of CD4+ anti-tumor T cells requires tumor expression of cell surface MHC class II molecules

Cerebrovascular disease exemplifies the poor regenerative capacity of the CNS. While there are methods to prevent cerebral infarction, there is no effective therapy available to ameliorate the anatomical, neurochemical and behavioral deficits which follow cerebral ischemia. Focal and transient occlusion of the middle cerebral artery (MCA) in rodents has been reported to result in neuropathology similar to that seen in clinical cerebral ischemia. Using specific techniques, this MCA occlusion can result in a well-localized infarct of the striatum. This review article will provide data accumulated from animal studies using the MCA occlusion technique in rodents to examine whether neural transplantation can ameliorate behavioral and morphological deficits associated with cerebral infarction. Recent advances in neural transplantation as a treatment modality for neurodegenerative disorders such as Parkinson's disease, have revealed that fetal tissue transplantation may produce neurobehavioral recovery. Accordingly, fetal tissue transplantation may provide a potential therapy for cerebral infarction. Preliminary findings in rodents subjected to unilateral MCA occlusion, and subsequently transplanted with fetal striatal tissue into the infarcted striatum have produced encouraging results. Transplanted fetal tissue, assessed immunohistochemically, has been demonstrated to survive and integrate with the host tissue, and, more importantly, ameliorate the ischemia-related behavioral deficits, at least in the short term. Although, this review will focus primarily on cerebral ischemia, characterized by a localized CNS lesion within the striatum, it is envisioned that this baseline data may be extrapolated and applied to cerebral infarction in other brain areas.     

Regulatory CD4(+) T cells expressing endogenous T cell receptor chains protect myelin basic protein-specific transgenic mice from spontaneous autoimmune encephalomyelitis

The development of T cell-mediated autoimmune diseases hinges on the balance between effector and regulatory mechanisms. Using two transgenic mouse lines expressing identical myelin basic protein (MBP)-specific T cell receptor (TCR) genes, we have previously shown that mice bearing exclusively MBP-specific T cells (designated T/R-) spontaneously develop experimental autoimmune encephalomyelitis (EAE), whereas mice bearing MBP-specific T cells as well as other lymphocytes (designated T/R+) did not. Here we demonstrate that T/R- mice can be protected from EAE by the early transfer of total splenocytes or purified CD4(+) T cells from normal donors. Moreover, whereas T/R+ mice crossed with B cell-deficient, gamma/delta T cell-deficient, or major histocompatibility complex class I-deficient mice did not develop EAE spontaneously, T/R+ mice crossed with TCR-alpha and -beta knockout mice developed EAE with the same incidence and severity as T/R- mice. In addition, MBP-specific transgenic mice that lack only endogenous TCR-alpha chains developed EAE with high incidence but reduced severity. Surprisingly, two-thirds of MBP-specific transgenic mice lacking only endogenous TCR-beta chains also developed EAE, suggesting that in T/R+ mice, cells with high protective activity escape TCR-beta chain allelic exclusion. Our study identifies CD4(+) T cells bearing endogenous alpha and beta TCR chains as the lymphocytes that prevent spontaneous EAE in T/R+ mice.     

Study of activation of murine T cells with bacterial superantigens. In vitro induction of enhanced responses in CD4+ T cells and of anergy in CD8+ T cells

Primary and secondary responses of murine CD4+ T cells and CD8+ T cells upon stimulation with staphylococcal enterotoxin E (SEE) bearing superantigenic properties were examined. Both isolated C57BL/6 splenic CD4+ T cells and CD8+ T cells proliferated and produced IL-2 and IFN-gamma upon stimulation with SEE in substantial levels. The amounts of IL-2 were greater in CD4+ T cells and those of IFN-gamma were somewhat greater in CD8+ T cells. SEE-induced CD4+ T lymphoblasts, larger parts of which bore the V beta 11 element in their TCR, proliferated, produced IL-2 and IFN-gamma, and showed toxin-dependent cytotoxicity in substantial levels upon restimulation with SEE. By contrast, SEE-induced CD8+ T lymphoblasts, the larger part of which bore the V beta 11 element, did not show the first two of the three responses at all upon restimulation with SEE, whereas these cells showed greater cytotoxicity. The CD8+ T lymphoblasts did not suppress the reactivity of the CD4+ T lymphoblasts. Both SEE-induced CD4+ T lymphoblasts and CD8+ T lymphoblasts proliferated and produced IL-2 and IFN-gamma in comparable levels upon stimulation with rIL-2 or mAb to CD3 or V beta 11.     

The interaction between CD4 and MHC class II molecules and its effect on T cell function

During T cell activation, CD4 and CD8 form a 'bridge' between the T cell receptor (TCR) and major histocompatibility complex (MHC) class II and class I molecules, respectively. Due to this intimate association, CD4 and CD8 are now termed co-receptors and considered an integral part of this multimolecular complex. In addition, interest in CD4 has been heightened by the discovery that it is, in part, the receptor for HIV. Although CD4 and CD8 appear to perform similar immune functions, they are structurally diverse suggesting that their mode of interaction with the TCR and MHC molecules may differ. This review will focus primarily on a series of studies which have attempted to map the residues which mediate CD4:MHC class II interaction. These data will be evaluated in light of our current understanding of CD8:MHC class I, and CD4:TCR interactions. In addition, a model to explain the structural and functional differences between CD4 and CD8 will be presented. Finally, the potential effect of these multiple interactions on T cell function will be discussed.     

Direct evidence that functionally impaired CD4+ T cells persist in vivo following induction of peripheral tolerance

A small population of CD4+ OVA-specific TCR transgenic T cells was tracked following the induction of peripheral tolerance by soluble Ag to address whether functionally unresponsive, or anergic T cells, persist in vivo for extended periods of time. Although injection of OVA peptide in the absence of adjuvant caused a transient expansion and deletion of the Ag-specific T cells, a population that showed signs of prior activation persisted in the lymphoid tissues for several months. These surviving OVA-specific T cells had long-lasting, but reversible defects in their ability to proliferate in lymph nodes and secrete IL-2 and TNF-alpha in vivo following an antigenic challenge. These defects were not associated with the production of Th2-type cytokines or the capacity to suppress the clonal expansion of a bystander population of T cells present in the same lymph nodes. Therefore, our results provide direct evidence that a long-lived population of functionally impaired Ag-specific CD4+ T cells is generated in vivo after exposure to soluble Ag.     

Disturbed CD4+ T cell homeostasis and in vitro HIV-1 susceptibility in transgenic mice expressing T cell line-tropic HIV-1 receptors

T cell line-tropic (T-tropic) HIV type 1 strains enter cells by interacting with the cell-surface molecules CD4 and CXCR4. We have generated transgenic mice predominantly expressing human CD4 and CXCR4 on their CD4-positive T lymphocytes (CD4+ T cells). Their primary thymocytes are susceptible to T-tropic but not to macrophage-tropic HIV-1 infection in vitro, albeit with a viral antigen production less efficient than human peripheral blood mononuclear cells. Interestingly, even without HIV infection, transgenic mice display a CD4+ T cell depletion profile of peripheral blood reminiscent of that seen in AIDS patients. We demonstrate that CD4+ T cell trafficking in transgenic mice is biased toward bone marrow essentially due to CXCR4 overexpression, resulting in the severe loss of CD4+ T cells from circulating blood. Our data suggest that CXCR4 plays an important role in lymphocyte trafficking through tissues, especially between peripheral blood and bone marrow, participating in the regulation of lymphocyte homeostasis in these compartments. Based on these findings, we propose a hypothetical model in which the dual function of CXCR4 in HIV-1 infection and in lymphocyte trafficking may cooperatively induce progressive HIV-1 infection and CD4+ T cell decline in patients.     

Increased expression of CD80, CD86 and CD70 on T cells from HIV-infected individuals upon activation in vitro: regulation by CD4+ T cells

T cells express CD28 and CD27 which transduce co-stimulatory signals after interaction with their ligands on antigen-presenting cells (APC). These ligands, CD80, CD86 and CD70, are also expressed to some extent on activated T cells. Here, we show that in human immunodeficiency virus (HIV)-infected individuals, CD28 and CD27 expression is decreased on CD8+ T cells. On the other hand, T cell stimulation in vitro induced high CD80, CD86 and CD70 expression on T cells from HIV-infected individuals. It appeared that an inverted CD4:CD8 T cell ratio could explain this enhanced expression of co-stimulatory ligands. Indeed, high expression levels of CD80, CD86 and CD70 were found on activated CD8+ T cells from HIV- individuals cultured in the absence of CD4+ T cells. Addition of CD4+ T cells prevented this up-regulation. However, in HIV-infected individuals, addition of excess autologous or healthy control CD4+ T cells did not completely counteract up-regulation of co-stimulatory ligand expression on CD8+ T cells. Thus, to some extent, CD8+ T cells in HIV-infected individuals appeared to be refractory to CD4+ T cell-mediated regulation of ligand expression in vitro. Activated T cells from HIV-infected individuals and activated CD8+ T cells from healthy controls were able to act as accessory cells in CD3-induced T cell proliferation, which was dependent on cell-cell contact. Thus, we showed that T cells from HIV-infected individuals express enhanced levels of co-stimulatory ligands upon activation, which provides them with accessory cell properties. Enhanced stimulatory potential of these nonprofessional APC may contribute to persistently high levels of immune activation in HIV infection related to disease progression.     

MHC class II-dependent NK1.1+ gammadelta T cells are induced in mice by Salmonella infection

We observed the emergence of a novel population of gammadelta T cells expressing NK1.1 Ag in the peritoneal cavity of mice infected with Salmonella choleraesuis. The NK1.1+gammadelta T cells accounted for approximately 20% of all gammadelta T cells emerging in the peritoneal cavity of C57BL/6 mice and expressed preferentially rearranged Vgamma4-Jgamma1 and Vdelta6.3-Ddelta1-Ddelta2-Jdelta1 genes with N diversity. The gammadelta T cells proliferated vigorously in response to PHA-treated spleen cells and produced IFN-gamma in the culture supernatant. However, spleen cells from Abetab-deficient mice were unable to stimulate the gammadelta T cells. Furthermore, the NK1.1+gammadelta T cells were stimulated not only by Chinese hamster ovary (CHO) cells expressing wild-type IAb but also by those expressing IAb/Ealpha52-68 or IAb/pigeon cytochrome c-derived analogue peptide complex. These proliferation activities were inhibited by mAb specific for IAb chain. Consistent with these findings, the emergence of NK1.1+gammadelta T cells was reduced in the peritoneal cavity of Abetab-deficient mice after Salmonella infection, whereas NK1.1+gammadelta T cells were rather abundant in the peritoneal cavity of Salmonella-infected beta2m-deficient mice. Moreover, the NK1.1+gammadelta T cells were easily identified in the thymus of beta2m-deficient but not Abetab-deficient mice. Our results indicated that MHC class II expression is essential for development and activation of NK1. 1+gammadelta T cells in the thymus and the periphery.