Subpopulations of mature murine thymocytes: properties of CD4-CD8+ and CD4+CD8- thymocytes lacking the heat-stable antigen

The heat-stable antigen (HSA), recognized by the monoclonal antibodies M1/69, B2A2, and J11d, is low or absent on the surface of most murine peripheral T cells but present on all but 3% of thymocytes. The CD4-CD8+ and CD4+CD8- or "single positive" thymic populations may be divided into further subgroups based on surface HSA expression. One group, CD4-CD8+ and expressing very high levels of HSA (HSA++), is an immature, T cell antigen receptor (TcR) negative, outer cortical blast cell. However, a further subdivision of CD4-CD8+ and CD4+CD8- single positives may be made, into those negative to low for HSA (HSA-) and those expressing moderate amounts of HSA (HSA+). The proportion of HSA- single positives is low in the thymus of young mice, whereas the proportion of HSA+ single positives is similar to that of the adult. Both the HSA- and the HSA+ subsets of single positive thymocytes from adult mice are CD3+ and express the normal peripheral T cell incidence of V beta 8 determinants on the TcR. On stimulation with concanavalin A in limit-dilution culture both HSA- and HSA+ subsets of single positive thymocytes give a high frequency of proliferating clones, and the clones from both HSA- and HSA+ subsets of CD4-CD8+ thymocytes are cytotoxic. Thus both HSA- and HSA+ single positive thymocytes are functionally mature. The HSA- subsets of single positive thymocytes differ from the HSA+ subsets in being slightly larger in size, in expressing higher levels of MEL-14, in binding more peanut agglutinin, and in including a proportion of cells expressing high levels of the Pgp-1 glycoprotein. It is suggested that HSA- CD4-CD8+ and HSA- CD4+CD8- thymocytes are more mature than their HSA+ counterparts, and might represent a previously activated or "memory" thymic subpopulation.     

Human thymocytes do not respond to interleukin-2 after removal of mature "bright" CD5 positive cells

Interleukin-2 receptors (IL-2R) are expressed on minor populations of immature and mature human thymocytes. These studies were designed to determine if immature T cells could respond to the mitogen phytohemagglutinin (PHA-P) plus IL-2 in vitro by increasing the expression of IL-2R and by proliferation. Using monoclonal antibodies to CD5 and magnetic immunobeads we were able to remove all mature, "bright" CD5+ cells from nylon wool-purified thymocytes and to obtain less mature cells which consisted almost completely of cells with the CD4+CD8+ phenotype. These immature cells were mostly "dim" CD5+ and less than 5% CD5- and a small percentage expressed the IL-2R. After culture in serum-free medium with PHA-P, these cells showed only a slight increase in the percentage of IL-2R+ cells and the addition of IL-2 did not increase the percentage of IL-2R+ cells and no proliferation was observed. Unseparated, nylon wool-purified thymocytes contained 14% bright CD5+ cells. These bright CD5+ cells had a mature phenotype of CD4+CD8- (52%) and CD4-CD8+ (27%) cells. A small percentage of these cells were IL-2R+. These bright CD5+IL-2R+ cells were predominantly mature CD4+CD8- cells as measured by three-color flow cytometry. After culture with PHA-P and IL-2, the percentage of IL-2R+ cells increased and they were now found not only on CD4+CD8- but also on CD4-CD8+ and on CD4+CD8+ cells. IL-2 plus PHA-P increased proliferation of these cells as compared to those cultured in medium with PHA-P without IL-2. Thus, we show that human immature thymocytes in contrast to mature thymocytes are not responsive to IL-2 as measured by a lack of IL-2R expression and proliferation. These data indicate that mature thymocytes can express a functional high affinity receptor for IL-2 and suggest that immature thymocytes may not possess a (functional) p75 chain of the IL-2R.     

B7 costimulates proliferation of CD4-8+ T lymphocytes but is not required for the deletion of immature CD4+8+ thymocytes.

In addition to TCR-derived signals, costimulatory signals derived from stimulation of the CD28 molecule by its natural ligand, B7, have been shown to be required for CD4+8- T cell activation. We investigate the ability of B7 to provide costimulatory signals necessary to drive proliferation and differentiation of virgin CD4-8+ T-cells that express a transgenic TCR specific for the male (H-Y) Ag presented by H-2Db class I MHC molecules. Virgin male-specific CD4-8+ T cells can be activated either with B7 transfected chinese hamster ovary (CHO) cells and T3.70, a mAb specific for the transgenic TCR-alpha chain that is associated with male-reactivity, or by male dendritic cells (DC). Activated CD4-8+ T cells proliferated in the absence of exogenously added IL-2. IL-2 activity was detected in supernatants of CD4-8+T3.70+ cells that were stimulated with T3.70 and B7+CHO cells. The response of CD4-8+T3.70+ cells to T3.70/B7+CHO or to male DC stimulation were inhibited by CTLA4Ig, a fusion protein comprising the extracellular portion of CTLA4 and human IgG C gamma 1. It has been previously shown that CTLA4Ig binds B7 with high affinity. Staining with CTLA4Ig revealed that DC express about 50 times more B7 than CD4-8+ T cells. CTLA4Ig also specifically blocked the proliferation of male-reactive cells in vivo. We have also used an in vitro deletion assay whereby immature CD4+8+ thymocytes expressing the transgenic male-specific TCR are deleted by overnight incubation with either immobilized T3.70 or male DC to investigate the participation of the CD28/B7 pathway in the negative selection of immature thymocytes. Staining with B7Ig established that both immature murine CD4+8+ and mature CD4-8+ thymocytes express a high level of CD28. However, despite the high expression of CD28 on CD4+8+ thymocytes, it was found that deletion of CD4+8+ thymocytes expressing the male-specific TCR by the T3.70 mAb was not inhibited by B7+CHO cells. Furthermore, the deletion of these thymocytes by DC also was not inhibited by CTLA4Ig. These findings provide evidence that although signaling through CD28 can costimulate a primary anti-male response in mature CD4-8+ T cells, the CD28/B7 pathway does not appear to participate in the negative selection of immature CD4+8+ thymocytes.     

Kinetics and efficacy of positive selection in the thymus of normal and T cell receptor transgenic mice

DNA-labeling studies in alpha beta T cell receptor (TCR) transgenic mice show that the lifespan of immature CD4+8+ thymocytes is 3.5 days irrespective of whether they are selected for maturation or not. While nonselected cells die, the binding of the TCR to thymic major histocompatibility complex molecules rescues CD4+8+ cells from programmed cell death and induces first upregulation of the TCR level and then differentiation into CD4+8- or CD4-8+ cells in the absence of any cell division. When most CD4+8+ thymocytes express a selectable transgenic TCR the formation of mature cells with high TCR levels is 10-20 times as efficient as observed in normal mice, yet still only 20% of the CD4+8+ cells become mature. This is due to the limited availability of selecting 'niches': most CD4+8+ thymocytes with a selectable transgenic TCR will undergo maturation when they represent only 5% or less of all CD4+8+ cells.     

Regulated costimulation in the thymus is critical for T cell development: dysregulated CD28 costimulation can bypass the pre-TCR checkpoint

Expression of CD28 is highly regulated during thymic development, with CD28 levels extremely low on immature thymocytes but increasing dramatically as CD4- CD8- cells initiate expression of TCRbeta. B7-1 and B7-2, the ligands for CD28, have a restricted distribution in the thymic cortex where immature thymocytes reside and are more highly expressed in the medulla where the most mature thymocytes are located. To determine the importance of this regulated CD28/B7 expression for T cell development, we examined the effect of induced CD28 signaling of immature thymocytes in CD28/B7-2 double-transgenic mice. Strikingly, we found that differentiation to the CD4+ CD8+ stage in CD28/B7-2 transgenics proceeds independent of the requirement for TCRbeta expression manifest in wild-type thymocytes, occurring even in Rag- or CD3epsilon- knockouts. These findings indicate that signaling of immature thymocytes through CD28 in the absence of TCR- or pre-TCR-derived signals can promote an aberrant pathway of T cell differentiation and highlight the importance of finely regulated physiologic expression of CD28 and B7 in maintaining integrity of the "beta" checkpoint for pre-TCR/TCR-dependent thymic differentiation.     

The FOXP3+ subset of human CD4+CD8+ thymocytes is immature and subject to intrathymic selection

FOXP3, believed to be the regulatory T (Treg)-cell determining factor, is already expressed at the CD4+CD8+ thymocyte stage, but there is disagreement whether these cells are the precursors of mature CD4+CD8(-) Treg cells. Here, we provide a quantitative analysis of FOXP3 expression in the human thymus. We show that a subset of CD4+CD8+ cells already expressed as much FOXP3 as the FOXP3+ CD4+CD8(-) cells, and like mature Treg cells were CD127 low. In contrast to earlier data, CD8+CD4(-) thymocytes expressed significantly lower levels of FOXP3 than either the CD4+CD8+ or CD4+CD8(-) subsets. The CD4+CD8+ double-positive cells also expressed recombination-activating gene-2, suggesting that they were still immature. Although the FOXP3+ double-positive cells are thus putatively the precursors of the mature CD4+CD8(-)FOXP3+ subset, their frequency did not predict the frequency of more mature Treg cells, and analysis of T-cell antigen receptor repertoire showed clear differences between the two subsets. Although these data do not rule out an independent CD4+CD8+ Treg cell subset, they are consistent with a model of human Treg cell development in which the upregulation of FOXP3 is an early event, but the first FOXP3+ population is still immature and subject to further selection. The upregulation of FOXP3 may thus not be the final determining factor in the commitment of human thymocytes to the Treg cell lineage.     

Clonogenic potential of murine CD4+8+ thymocytes. Direct demonstration using a V beta 6-specific proliferative stimulus in Mlsa mice

Although considerable indirect evidence supports the hypothesis that CD4+8+ thymocytes are developmental intermediates in the generation of mature (CD4+8- or CD4-8+) T cells, the ability of these cells to proliferate in vitro has been highly controversial. We demonstrate here that a fraction of purified murine CD4+8+ thymocytes can be induced to proliferate in response to immobilized anti-TCR mAb. To exclude possible proliferation by trace mature T cell contaminants, we have exploited our recent finding that in Mlsa mice mature V beta 6-bearing thymic T cells are virtually absent (less than or equal to 0.5%) due to clonal deletion, whereas V beta 6 +CD4+8+ thymocytes are present in much higher numbers (approximately 3%). Proliferation of sorted CD4+8+ thymocytes from Mlsa mice was therefore induced at limiting dilution with immobilized anti-V beta 6 mAb to select against any contaminating mature T cells. Under optimal culture conditions, the frequency of CD4+8+ thymocytes proliferating specifically to anti-V beta 6 mAb (1/1000) was higher than those obtained for purified CD4-8+ (1/2000) or CD4+8- (1/5000) subsets, thus demonstrating directly that a proportion (in this case 3%) of CD4+8+ thymocytes are potentially clonable. During culture, V beta 6 +CD4+8+ thymocytes gave rise to a mixture of phenotypically "immature" (CD4-8-) and "mature" (CD4-8+) T cells. This system should be valuable for further analysis of the elusive CD4+8+ thymocyte subset.     

The majority of CD4+8- thymocytes are functionally immature.

The thymus is the major site of T cell development and repertoire selection. During these processes, T cells segregate into two subsets that express either CD4 or CD8 accessory molecules, the phenotype of peripheral T cells. Analysis of CD4+8- thymocytes revealed that the majority of these cells express the heat-stable Ag (HSA) but not the nonclassical class I Ag, Qa-2. This HSA+, Qa-2- phenotype is similar to that of the less mature, CD4+8+ thymocytes. The remaining CD4+8- thymocytes possess the HSA-, Qa-2+ phenotype of peripheral T cells. To determine whether the Qa-2-, CD4+8- thymic subset is fully mature, we have analyzed the functional status of these CD4+8- subpopulations. The results indicate that only those thymocytes which express Qa-2 are fully responsive to anti-TCR stimulation in a manner analogous to peripheral T cells. The Qa-2- subset is nonresponsive to stimulation by anti-TCR antibodies that have been immobilized to plastic, even in the presence of lymphokines or syngeneic APC. This subset is, however, capable of proliferating to allogeneic cells or to anti-TCR on the surface of syngeneic APC, although not to the levels achieved by Qa-2+ thymocytes. Thus, the Qa-2- subset appears to require additional interactions which are not necessary for peripheral T cells or Qa-2+ thymocytes. Relevant to this issue, the Qa-2+ thymocyte subset does not appear until relatively late in development, and does not reach adult frequencies until several weeks after birth. These results would suggest that there is a progression from HSA+, Qa-2- to HSA-, Qa-2+ which parallels the maturation of functional responsiveness. These findings are important to understanding T cell selection since thymocytes with such a decreased responsiveness may have a differential capacity for tolerance induction. The results presented suggest that the bulk of CD4+8- thymocytes are not fully mature and that Qa-2 may serve as a marker for T cells with a more complete functional competence.     

Inverse correlation between steady-state RNA and cell surface T cell receptor levels.

The relationship between steady-state RNA and cell surface levels of T cell receptor (TCR) was examined in mature T cells and immature CD4+CD8+ double positive thymocytes. TCR is expressed at high levels on the surface of mature T cells and at much lower levels on double positive thymocytes. We demonstrate that in direct contrast to surface expression, TCR-alpha, -beta, CD3-delta, -epsilon, -gamma, and sigma RNA levels are much higher in the immature double positive thymocyte population than in mature T cells. These results demonstrate that quantitative differences in TCR surface expression in immature and mature T cells are not due to increases in TCR RNA levels.     

Identification and distribution of the costimulatory receptor CD28 in the mouse.

T cell activation requires Ag-specific stimulation mediated by the TCR as well as an additional stimulus provided by Ag presenting cells. On human T cells, it has been shown that antibodies to the Ag CD28 can provide a potent amplification signal for cytokine production and proliferation. Here we describe the production of a mAb to the murine homologue of CD28, and the use of this antibody to examine the function and distribution of CD28 in the mouse. Anti-murine CD28 synergizes with TCR-mediated signals to greatly enhance lymphokine production and proliferation of T cells, and the CD28 signal is not blocked by cyclosporin A. In the peripheral lymphoid organs and in the blood of the mouse, all CD4+ and CD8+ T cells express CD28. In the thymus, CD28 expression is highest on immature CD3-, CD8+ and CD4+8+ cells, and on CD4-8- cells that express alpha beta and tau delta TCR. The level of CD28 on mature CD4+ and CD8+ alpha beta TCR+ thymocytes is two- to fourfold lower than on the immature cells. The potent costimulatory function of CD28 on mature T cells, together with the high level of expression on CD4+8+ thymocytes, suggest that this costimulatory receptor might play an important role in T cell development and activation.     

Selective stimulation of thymocyte precursors mediated by specific cytokines. Different CD3+ subsets are generated by IL-1 versus IL-2.

The sequence of activation signals that stimulate proliferation, differentiation, and selection of mature T cell subsets from immature, dull-CD5+/CD4-, CD8- double negative (bCD5), (dCD5/DN) thymocytes are still unclear. However, it is likely that cytokines play integral roles in these events. Here we report that IL-1, in the presence of Con A, supports the proliferation and differentiation of highly purified dCD5/DN precursors into bright-CD5+ DN, CD2- lymphocytes with an apparently mature phenotype. These cells express CD3 and preferentially express the products of two TCR gene families, V beta 8 and V beta 6, whose expression is dependent on the allelic expression of the Mls-1 locus. Experiments, using DN thymocytes mixed with purified dCD5 subset of DN cells from a congenic strain of mice (i.e., expressing two different alleles of CD5) have shown that the cells that are stimulated by IL-1 and comitogen are derived from the immature dCD5 subset and not from the mature bCD5 cells contained within the DN subset. In contrast, IL-2 with the co-mitogen stimulates three- to fourfold higher levels of proliferation, from the same purified immature precursor population, and nearly a twofold increase in cell yield. However, the cells that were generated from precursor thymic cells stimulated with IL-2 represent a completely different T cell subset compared to IL-1-generated cells; these IL-2-stimulated cells express comparable levels of CD3, but also express substantial levels of CD2 and the TCR-gamma/delta, and a subset expresses CD8. These data suggest that these two TCR-alpha/beta and TCR-gamma/delta subsets of mature thymocytes use different cytokines and therefore possibly different stromal interactions to initiate differentiation.     

MHC non-restricted, CD95-independent apoptosis of immature thymocytes induced by thymic epithelial cells

The interaction of thymocytes with thymic epithelial cells in the absence of an exogenous antigen was studied in vitro. Thymic, but not splenic epithelial cells induced apoptosis of thymocytes. A thymic epithelial cell line (TEC) induced apoptosis of thymocytes but not of splenic T-cells. The target population for TEC-induced death were immature CD4(+)8(+) (double positive), but not mature single positive thymocytes. TEC also induced DNA fragmentation in day 18 foetal thymocytes, most of which are CD4(+)8(+) cells. Radiation leukemia virus (RadLV)-transformed thymic lymphoma clones expressing various phenotypes reflected this sensitivity, in that a CD4(+)8(+)3(+) clone apoptosed by thymic epithelial cells or TEC. Other, single positive or double negative clones were resistant. Thymocytes from C3H (H-2(k)), C57BL/6 (H-2(b)) and Balb/C (H-2(d)) mice apoptosed equally in response to either C57BL/6 thymic epithelial cells or TEC (H-2(b) x H-2(d)). Likewise, thymocytes from MRLIpr((-/-)) and B6Ipr((-/-)) mice, which do not express CD95 were also apoptosed by TEC.The data suggest that thymic epithelial cells induce MHC non-restricted, Fas-independent apoptosis of immature thymocytes. This response may reflect a mechanism through which thymocytes expressing TcR with no affinity to self MHC/peptide complexes are eliminated.     

T cell development in mice lacking the CD3-zeta/eta gene.

The CD3-zeta and CD3-eta polypeptides are two of the components of the T cell antigen receptor (TCR) which contribute to its efficient cell surface expression and account for part of its transducing capability. CD3-zeta and CD3-eta result from the alternative splicing of a single gene designated CD3-zeta/eta. To evaluate the role of these subunits during T cell development, we have produced mice with a disrupted CD3-zeta/eta gene. The analysis of thymocyte populations from the CD3-zeta/eta-/- homozygous mutant mice revealed that they have a profound reduction in the surface levels of TCR complexes and that the products of the CD3-zeta/eta gene appear to be needed for the efficient generation and/or survival of CD4+CD8+ thymocytes. Despite the almost total absence of mature single positive thymocytes, the lymph nodes from zeta/eta-/- mice were found to contain unusual CD4+CD8- and CD4-CD8+ single positive cells which were CD3-. In contrast to the situation observed in the thymus, the thymus-independent gut intraepithelial lymphocytes present in zeta/eta-/- mice do express TCR complexes on their surface and these are associated with Fc epsilon RI gamma homodimers. These results establish an essential role for the CD3-zeta/eta gene products during intrathymic T cell differentiation and further emphasize the difference between conventional T cells and thymus-independent gut intraepithelial lymphocytes.     

Human T-cell leukemia virus type I-induced proliferation of human immature CD2+CD3- thymocytes.

The mitogenic activity of human T-cell leukemia virus type I (HTLV-I) is triggering the proliferation of human resting T lymphocytes through the induction of the interleukin-2 (IL-2)/IL-2 receptor autocrine loop. This HTLV-I-induced proliferation was found to be mainly mediated by the CD2 T-cell antigen, which is first expressed on double-negative lymphoid precursors after colonization of the thymus. Thus, immature thymocytes express the CD2 antigen before that of the CD3-TCR complex. We therefore investigated the responsiveness of these CD2+CD3- immature thymocytes and compared it with that of unseparated thymocytes, containing a majority of the CD2+CD3+ mature thymocytes, and that of the CD2-CD3- prothymocytes. Both immature and unseparated thymocytes were incorporating [3H]thymidine in response to the virus, provided that they were cultivated in the presence of submitogenic doses of phytohemagglutinin. In contrast, the prothymocytes did not proliferate. Downmodulation of the CD2 molecule by incubating unseparated and immature thymocytes with a single anti-CD2 monoclonal antibody inhibited the proliferative response to HTLV-I. These results clearly underline that the expression of the CD2 molecule is exclusively required in mediating the proliferative response to the synergistic effect of phytohemagglutinin and HTLV-I. Immature thymocytes treated with a pair of anti-CD2 monoclonal antibodies were shown to proliferate in response to HTLV-I, even in the absence of exogenous IL-2. We further verified that the proliferation of human thymocytes is consecutive to the expression of IL-2 receptors and the synthesis of IL-2. These observations provide evidence that the mitogenic stimulus delivered by HTLV-I is more efficient than that provided by other conventional mitogenic stimuli, which are unable to trigger the synthesis of endogenous IL-2. Collectively, these results show that the mitogenic activity of HTLV-I is able to trigger the proliferation of cells which are at an early stage of T-cell development. They might therefore represent target cells in which HTLV-I infection could favor the initiation of the multistep lymphoproliferative process leading to adult T-cell leukemia.     

Signal transduction via CD4, CD8, and CD28 in mature and immature thymocytes. Implications for thymic selection.

The positive and negative selection of immature thymocytes that shapes the mature T cell repertoire appears to occur at an intermediate stage of development when the cells express low levels of TCR/CD3. These cells are also CD4+CD8+ and CD28+ (dull), and signals delivered by these three accessory molecules have been implicated in the selection process. We have examined the regulatory function of these accessory molecules on responses of immature thymocytes stimulated through the TCR/CD3 complex. Cross-linking CD4 or CD8 with CD3 strongly enhanced signal transduction via CD3 as assessed by protein tyrosine phosphorylation and calcium mobilization. Subsequent cell proliferation could be induced by soluble anti-CD28 mAb, which was comitogenic for cells stimulated with CD3 x CD4 or CD3 x CD8 cross-linking, but was without effect on cells stimulated with CD3 x CD3 cross-linking. A potential role for CD28 signal transduction in thymic maturation is suggested by the demonstration that the BB-1 molecule, a natural ligand for CD28, is expressed on thymic stromal cells. Taken together, our data suggest a model of thymic development in which CD4 or CD8 may enhance TCR/CD3 signaling upon coligation by an MHC molecule. If the CD28 surface receptor is simultaneously stimulated by a BB-1 expressing stromal cell, this set of interactions could lead to proliferation and positive selection. In the absence of CD28 stimulation the enhanced TCR/CD3 signals might lead to apoptosis and negative selection.     

CD69 expression discriminates MHC-dependent and -independent stages of thymocyte positive selection

In the thymus, phenotypically and functionally mature single positive cells are generated from immature CD4+8+ precursors by a process known as positive selection. Although this event is known to involve alphabetaTCR ligation by peptide/MHC complexes expressed on thymic stromal cells, it is clear that positive selection is a multistage process involving transition through an intermediate CD4+8+69+ phase as well as subsequent postselection phases. By analyzing the development of preselection CD4+8+69- and intermediate CD4+8+69+ thymocytes in the presence of MHC class I-deficient, MHC class II-deficient, and MHC double-deficient thymic stromal cells, we investigated the role of MHC molecules at three distinct points during positive selection. Although the initiation of positive selection is critically dependent upon MHC interactions, we find the that later stages of maturation, involving the differentiation of CD4+8- and CD4-8+ cells from CD4+8+69+ thymocytes, occur in the absence of MHC molecules. Moreover, an analysis of the postselection proliferation of newly generated CD4+8- and CD4-8+ thymocytes shows that this also occurs independently of MHC molecules. Thus, our data provide direct evidence that, although positive selection is a multistage process initiated by TCR-MHC interactions, continuation of this process and subsequent postselection events are independent of ongoing engagement of the TCR.     

Thymus and autoimmunity: production of CD25+CD4+ naturally anergic and suppressive T cells as a key function of the thymus in maintaining immunologic self-tolerance

This study shows that the normal thymus produces immunoregulatory CD25+4+8- thymocytes capable of controlling self-reactive T cells. Transfer of thymocyte suspensions depleted of CD25+4+8- thymocytes, which constitute approximately 5% of steroid-resistant mature CD4+8- thymocytes in normal naive mice, produces various autoimmune diseases in syngeneic athymic nude mice. These CD25+4+8- thymocytes are nonproliferative (anergic) to TCR stimulation in vitro, but potently suppress the proliferation of other CD4+8- or CD4-8+ thymocytes; breakage of their anergic state in vitro by high doses of IL-2 or anti-CD28 Ab simultaneously abrogates their suppressive activity; and transfer of such suppression-abrogated thymocyte suspensions produces autoimmune disease in nude mice. These immunoregulatory CD25+4+8- thymocytes/T cells are functionally distinct from activated CD25+4+ T cells derived from CD25-4+ thymocytes/T cells in that the latter scarcely exhibits suppressive activity in vitro, although both CD25+4+ populations express a similar profile of cell surface markers. Furthermore, the CD25+4+8- thymocytes appear to acquire their anergic and suppressive property through the thymic selection process, since TCR transgenic mice develop similar anergic/suppressive CD25+4+8- thymocytes and CD25+4+ T cells that predominantly express TCRs utilizing endogenous alpha-chains, but RAG-2-deficient TCR transgenic mice do not. These results taken together indicate that anergic/suppressive CD25+4+8- thymocytes and peripheral T cells in normal naive mice may constitute a common T cell lineage functionally and developmentally distinct from other T cells, and that production of this unique immunoregulatory T cell population can be another key function of the thymus in maintaining immunologic self-tolerance.     

Expression of CD1 and class I MHC antigens by human thymocytes

The acquisition of surface class I MHC molecules is associated with the maturation of thymocytes. Here, surface expression of class I MHC and CD1, which represents a family of MHC-related molecules, was analyzed on various human immature and mature thymocyte subpopulations. Class I expression was inversely related to the expression of CD1. The majority of CD4+ CD8+ cortical type thymocytes expressed low levels of class I MHC Ag, the previously described CD4+ CD8+ thymocyte subpopulation with low CD8 expression exhibited intermediate levels of class I MHC, whereas most of the single positive CD4 and CD8 thymocytes displayed high levels of class I MHC. Biochemical comparison of CD1 and class I showed that thymic class I molecules were post-translationally modified by phosphorylation, whereas CD1 was not phosphorylated. Furthermore, our studies suggested that in addition to CD1/CD8 complexes, thymocytes bear CD8/class I complexes. Chemical cross-linking and peptide mapping studies clearly identified the CD8-associated protein on thymic clones as the class I MHC molecule.