CD8alpha-11b+ dendritic cells but not CD8alpha+ dendritic cells mediate cross-tolerance toward intestinal antigens

Cross-presentation is a critical process by which antigen is displayed to CD8 T cells to induce tolerance. It is believed that CD8alpha+ dendritic cells (DCs) are responsible for cross-presentation, suggesting that the CD8alpha+ DC population is capable of inducing both cross-priming and cross-tolerance to antigen. We found that cross-tolerance against intestinal soluble antigen was abrogated in C57BL/6 mice lacking mesenteric lymph nodes (MLNs) and Peyer patches (PPs), whereas mice lacking PPs alone were capable of developing CD8 T-cell tolerance. CD8alpha-CD11b+ DCs but not CD8alpha+ DCs in the MLNs present intestinal antigens to relevant CD8 T cells, while CD8alpha+ DCs but not CD8alpha-CD11b+ DCs in the spleen exclusively cross-present intravenous soluble antigen. Thus, CD8alpha-CD11b+ DCs in the MLNs play a critical role for induction of cross-tolerance to dietary proteins.     

CD8alpha+ dendritic cells originate from the CD8alpha- dendritic cell subset by a maturation process involving CD8alpha, DEC-205, and CD24 up-regulation.

CD8alpha+ and CD8alpha- dendritic cells (DCs) have been considered as independent DC subpopulations both ontogenetically and functionally during recent years. However, it has been demonstrated that both DC subsets can be generated from a single precursor population, supporting the concept that they do not represent separate DC lineages. By using highly purified splenic CD8alpha- DCs, which were injected intravenously and traced by means of an Ly5.1/Ly5.2 transfer system, this study shows that CD8alpha- DCs acquired the phenotypic characteristics of CD8alpha+ DCs, by a differentiation process involving CD8alpha, DEC-205, and CD24 up-regulation, paralleled by the down-regulation of CD11b, F4/80, and CD4. These data demonstrate that CD8alpha+ DCs derive from CD8alpha- DCs, and strongly support that CD8alpha- and CD8alpha+ DCs represent different maturation or differentiation stages of the same DC population. Therefore, CD8alpha+ DCs would represent the last stage of DC differentiation, playing an essential role in the induction of T-cell responses, due to their antigen-presenting potential, cross-priming ability, and capacity to secrete large amounts of key cytokines such as interferon gamma and interleukin-12.     

CD8+ T-cell-mediated killing of donor dendritic cells prevents alloreactive T helper type-2 responses in vivo

Accumulating evidence indicates that, in absence of CD8+ T-cell activation, CD4+ T-cell-mediated allograft rejection is associated with a dominant Th2-cell response and eosinophil infiltrates. In this study, we analyzed the mechanisms by which CD8+ T cells regulate alloreactive CD4+ T-cell priming and differentiation into interleukin 4 (IL-4)-producing cells. We showed that interferon gamma (IFN-gamma) production by CD8+ T cells was dispensable for the inhibition of Th2-cell development, as well as tissue eosinophilia and type 2 cytokine production in the rejected grafts. Since we noticed that CD8+ T cells not only suppressed Th2 differentiation, but also down-modulated the overall priming of alloreactive CD4+ T cells, we evaluated whether CD8+ T cells act by limiting the accumulation of donor-derived dendritic cells (DCs) in lymph nodes. We found that indeed, alloreactive CD8+ T cells rapidly eliminated allogeneic DCs from T-cell areas of draining lymph nodes, through a perforin-dependent mechanism. Thus, our data demonstrate that cytotoxic T lymphocyte (CTL)-mediated clearance of allogeneic DCs is a negative feedback mechanism that limits the duration of alloantigen presentation in draining lymph nodes, thereby modulating the amplitude and polarization of the primary alloreactive CD4+ T-cell responses.     

CD8alpha+ mouse spleen dendritic cells do not originate from the CD8alpha- dendritic cell subset

Although previous studies had indicated that the CD8alpha- and CD8alpha+ subtypes of murine dendritic cells (DCs) differ in immediate origin, a recent study found that intravenous transfer of CD8alpha- DCs led to CD8alpha+ DCs in the spleen several days later, suggesting a direct precursor-product relationship. We have repeated these experiments with a balance sheet approach. We find that though a few CD8alpha+ DCs can be generated in such experiments, this is a rare event and could be the result of a contaminant precursor. Most of the immediate precursors of CD8alpha+ DCs are cells that lack the phenotype of a recognizable DC. CD8alpha- DCs and CD8alpha+ DCs are not precursor-product related, though these sublineages may be connected further upstream.     

Concept of lymphoid versus myeloid dendritic cell lineages revisited: both CD8alpha(-) and CD8alpha(+) dendritic cells are generated from CD4(low) lymphoid-committed precursors

Two dendritic cell (DC) subsets have been identified in the murine system on the basis of their differential CD8alpha expression. CD8alpha(+) DCs and CD8alpha(-) DCs are considered as lymphoid- and myeloid-derived, respectively, because CD8alpha(+) but not CD8alpha(-) splenic DCs were generated from lymphoid CD4(low) precursors, devoid of myeloid reconstitution potential. Although CD8alpha(-) DCs were first described as negative for CD4, our results demonstrate that approximately 70% of them are CD4(+). Besides CD4(-) CD8alpha(-) and CD4(+) CD8alpha(-) DCs displayed a similar phenotype and T-cell stimulatory potential in mixed lymphocyte reaction (MLR), although among CD8alpha(-) DCs, the CD4(+) subset appears to have a higher endocytic capacity. Finally, experiments of DC reconstitution after irradiation in which, in contrast to previous studies, donor-type DCs were analyzed without depleting CD4(+) cells, revealed that both CD8alpha(+) DCs and CD8alpha(-) DCs were generated after transfer of CD4(low) precursors. These data suggest that both CD8alpha(+) and CD8alpha(-) DCs derive from a common precursor and, hence, do not support the concept of the CD8alpha(+) lymphoid-derived and CD8alpha(-) myeloid-derived DC lineages. However, because this hypothesis has to be confirmed at the clonal level, it remains possible that CD8alpha(-) DCs arise from a myeloid precursor within the CD4(low) precursor population or, alternatively, that both CD8alpha(+) and CD8alpha(-) DCs derive from an independent nonlymphoid, nonmyeloid DC precursor. In conclusion, although we favor the hypothesis that both CD8alpha(+) and CD8alpha(-) DCs derive from a lymphoid-committed precursor, a precise study of the differentiation process of CD8alpha(+) and CD8alpha(-) DCs is required to define conclusively their origin.     

Dendritic cell differentiation potential of mouse monocytes: monocytes represent immediate precursors of CD8- and CD8+ splenic dendritic cells

The monocyte capacity to differentiate into dendritic cells (DCs) was originally demonstrated by human in vitro DC differentiation assays that have subsequently become the essential methodologic approach for the production of DCs to be used in DC-mediated cancer immunotherapy protocols. In addition, in vitro DC generation from monocytes is a powerful tool to study DC differentiation and maturation. However, whether DC differentiation from monocytes occurs in vivo remains controversial, and the physiologic counterparts of in vitro monocyte-derived DCs are unknown. In addition, information on murine monocytes and monocyte-derived DCs is scarce. Here we show that mouse bone marrow monocytes can be differentiated in vitro into DCs using similar conditions as those defined in humans, including in vitro cultures with granulocyte-macrophage colony-stimulating factor and interleukin 4 and reverse transendothelial migration assays. Importantly, we demonstrate that after in vivo transfer monocytes generate CD8- and CD8+ DCs in the spleen, but differentiate into macrophages on migration to the thoracic cavity. In conclusion, we support the hypothesis that monocytes generate DCs not only on entry into the lymph and migration to the lymph nodes as proposed, but also on extravasation from blood and homing to the spleen, suggesting that monocytes represent immediate precursors of lymphoid organ DCs.     

Gut-associated lymphoid tissue-primed CD4+ T cells display CCR9-dependent and -independent homing to the small intestine

CD4(+) T-cell entry to the intestinal mucosa is central to the generation of mucosal immunity as well as chronic intestinal inflammation, yet the mechanisms regulating this process remain poorly defined. Here we show that murine small intestinal CD4(+) lamina propria lymphocytes express a heterogeneous but restricted array of chemokine receptors including CCR5, CCR6, CCR9, CXCR3, and CXCR6. CD4(+) T-cell receptor transgenic OT-II cells activated in mesenteric lymph nodes acquired a distinct chemokine receptor profile, including expression of CCR6, CCR9, and CXCR3 that was only partially reproduced in vitro after priming with mesenteric lymph node dendritic cells. A subset of these effector CD4(+) T cells, expressing CD69 and alpha(4)beta(7), entered the intestinal lamina propria and the majority of these cells expressed CCR9. CCR9(-/-) OT-II cells were disadvantaged in their ability to localize to the intestinal lamina propria; however, they were readily detected at this site and expressed alpha(4)beta(7), but little CCR2, CCR5, CCR6, CCR8, CCR10, CXCR3, or CXCR6. Thus, whereas CD4(+) T cells activated in gut-associated lymphoid tissue express a restricted chemokine receptor profile, including CCR9, targeting both CCR9-dependent and CCR9-independent entry mechanisms is likely to be important to maximally inhibit accumulation of these cells within the small intestinal mucosa.     

Anatomic location and T-cell stimulatory functions of mouse dendritic cell subsets defined by CD4 and CD8 expression

Mouse spleen contains CD4+, CD8alpha+, and CD4-/CD8alpha- dendritic cells (DCs) in a 2:1:1 ratio. An analysis of 70 surface and cytoplasmic antigens revealed several differences in antigen expression between the 3 subsets. Notably, the Birbeck granule-associated Langerin antigen, as well as CD103 (the mouse homologue of the rat DC marker OX62), were specifically expressed by the CD8alpha+ DC subset. All DC types were apparent in the T-cell areas as well as in the splenic marginal zones and showed similar migratory capacity in collagen lattices. The 3 DC subtypes stimulated allogeneic CD4+ T cells comparably. However, CD8alpha+ DCs were very weak stimulators of resting or activated allogeneic CD8+ T cells, even at high stimulator-to-responder ratios, although this defect could be overcome under optimal DC/T cell ratios and peptide concentrations using CD8+ F5 T-cell receptor (TCR)-transgenic T cells. CD8alpha- or CD8alpha+ DCs presented alloantigens with the same efficiency for lysis by cytotoxic T lymphocytes (CTLs), and their turnover rate of class I-peptide complexes was similar, thus neither an inability to present, nor rapid loss of antigenic complexes from CD8alpha DCs was responsible for the low allostimulatory capacity of CD8alpha+ DCs in vitro. Surprisingly, both CD8alpha+ DCs and CD4-/CD8- DCs efficiently primed minor histocompatibility (H-Y male antigen) cytotoxicity following intravenous injection, whereas CD4+ DCs were weak inducers of CTLs. Thus, the inability of CD8alpha+ DCs to stimulate CD8+ T cells is limited to certain in vitro assays that must lack certain enhancing signals present during in vivo interaction between CD8alpha+ DCs and CD8+ T cells.     

Generation of T cells from cytokine-mobilized peripheral blood and adult bone marrow CD34+ cells

The present study compared the T-cell progenitor content of CD34+ lineage (Lin)- cells isolated from normal adult bone marrow (ABM) and mobilized peripheral blood (MPB). Both cell populations were found to differentiate into T cells when injected into human fetal thymi implanted into severe combined immunodeficient mice. Cytokine-MPB cells were less efficient than ABM cells in engrafting in the fetal human thymus, although both gave rise to thymocytes with identical phenotypes based on the analysis of CD1a, CD3, CD4, and CD8 expression. Thymocytes derived from adult CD34+ Lin- cells were capable of fully differentiating into mature CD3+ T cells expressing either the T-cell receptor (TCR) gamma delta or the TCR alpha beta (the later associated with CD4 or CD8), showing that the T-cell progenies of adult CD34+ cells were polyclonal and functional. Our data indicate that human MPB CD34+ cells are qualitatively identical to their BM counterparts, and demonstrate the existence of T-lymphoid progenitor cell activity in MPB.     

CD8alpha alpha memory effector T cells descend directly from clonally expanded CD8alpha +beta high TCRalpha beta T cells in vivo

Whereas most peripheral CD8(+) alphabeta T cells highly express CD8alphabeta heterodimer in healthy individuals, there is an increase of CD8alpha(+)beta(low) or CD8alphaalpha alphabeta T cells in HIV infection or Wiskott-Aldrich syndrome and after bone marrow transplantation. The significance of these uncommon cell populations is not well understood. There has been some question as to whether these subsets and CD8alpha(+)beta(high) cells belong to different ontogenic lineages or whether a fraction of CD8alpha(+)beta(high) cells have down-regulated CD8beta chain. Here we assessed clonality of CD8alphaalpha and CD8alpha(+)beta(low) alphabeta T cells as well as their phenotypic and functional characteristics. Deduced from surface antigens, cytotoxic granule constituents, and cytokine production, CD8alpha(+)beta(low) cells are exclusively composed of effector memory cells. CD8alphaalpha cells comprise effector memory cells and terminally differentiated CD45RO(-)CCR7(-) memory cells. T-cell receptor (TCR) Vbeta complementarity-determining region 3 (CDR3) spectratyping analysis and subsequent sequencing of CDR3 cDNA clones revealed polyclonality of CD8alpha(+)beta(high) cells and oligoclonality of CD8alpha(+)beta(low) and CD8alphaalpha cells. Importantly, some expanded clones within CD8alphaalpha cells were also identified within CD8alpha(+)beta(high) and CD8alpha(+)beta(low) subpopulations. Furthermore, signal-joint TCR rearrangement excision circles concentration was reduced with the loss of CD8beta expression. These results indicated that some specific CD8alpha(+)beta(high) alphabeta T cells expand clonally, differentiate, and simultaneously down-regulate CD8beta chain possibly by an antigen-driven mechanism. Provided that antigenic stimulation directly influences the emergence of CD8alphaalpha alphabeta T cells, these cells, which have been previously regarded as of extrathymic origin, may present new insights into the mechanisms of autoimmune diseases and immunodeficiencies, and also serve as a useful biomarker to evaluate the disease activities.     

Characterization of a new subpopulation of mouse CD8alpha+ B220+ dendritic cells endowed with type 1 interferon production capacity and tolerogenic potential

We describe a new B220+ subpopulation of immaturelike dendritic cells (B220+ DCs) with low levels of expression of major histocompatibility complex (MHC) and costimulatory molecules and markedly reduced T-cell stimulatory potential, located in the thymus, bone marrow, spleen, and lymph nodes. B220+ DCs display ultrastructural characteristics resembling those of human plasmacytoid cells and accordingly produce interferon-alpha after virus stimulation. B220+ DCs acquired a strong antigen-presenting cell capacity on incubation with CpG oligodeoxynucleotides, concomitant with a remarkable up-regulation of MHC and costimulatory molecules and the production of interleukin-12 (IL-12) and IL-10. Importantly, our data suggest that nonstimulated B220+ DCs represent a subset of physiological tolerogenic DCs endowed with the capacity to induce a nonanergic state of T-cell unresponsiveness, involving the differentiation of T regulatory cells capable of suppressing antigen-specific T-cell proliferation. In conclusion, our data support the hypothesis that B220+ DCs represent a lymphoid organ subset of immature DCs with a dual role in the immune system-exerting a tolerogenic function in steady state but differentiating on microbial stimulation into potent antigen-presenting cells with type 1 interferon production capacity.     

Differentiation of myeloid dendritic cells into CD8alpha-positive dendritic cells in vivo

Bone marrow-derived dendritic cells (DC) represent a family of antigen-presenting cells (APC) with varying phenotypes. For example, in mice, CD8alpha(+) and CD8alpha(-) DC are thought to represent cells of lymphoid and myeloid origin, respectively. Langerhans cells (LC) of the epidermis are typical myeloid DC; they do not express CD8alpha, but they do express high levels of myeloid antigens such as CD11b and FcgammaR. By contrast, thymic DC, which derive from a lymphoid-related progenitor, express CD8alpha but only low levels of myeloid antigens. CD8alpha(+) DC are also found in the spleen and lymph nodes (LN), but the origin of these cells has not been determined. By activating and labeling CD8alpha(-) epidermal LC in vivo, it was found that these cells expressed CD8alpha on migration to the draining LN. Similarly, CD8alpha(-) LC generated in vitro from a CD8 wild-type mouse and injected into the skin of a CD8alphaKO mouse expressed CD8alpha when they reached the draining LN. The results also show that CD8alpha(+) LC are potent APC. After migration from skin, they localized in the T-cell areas of LN, secreted high levels of interleukin-12, interferon-gamma, and chemokine-attracting T cells, and they induced antigen-specific T-cell activation. These results demonstrate that myeloid DC in the periphery can express CD8alpha when they migrate to the draining LN. CD8alpha expression on these DC appears to reflect a state of activation, mobilization, or both, rather than lineage. (Blood. 2000;96:1865-1872)     

CD4+CD25+ T regulatory cells control anti-islet CD8+ T cells through TGF-beta-TGF-beta receptor interactions in type 1 diabetes

Pancreatic lymph node-derived CD4+CD25+ T regulatory (Treg) cells inhibit in situ differentiation of islet-reactive CD8+ T cells into cytotoxic T lymphocytes, thereby preventing diabetes progression. The mechanism by which these Treg cells suppress anti-islet CD8+ T cells is unknown. Here, we show by using a CD8+ T cell-mediated model of type 1 diabetes that transforming growth factor (TGF)-beta-TGF-beta receptor signals are critical for CD4+CD25+ Treg cell regulation of autoreactive islet-specific cytotoxic T lymphocytes. Transgenic expression of tumor necrosis factor alpha from birth to 25 days of age in the islets of B6 mice that constitutively express CD80 on their beta cells results in accumulation of CD4+CD25+TGF-beta+ cells exclusively in the islets and pancreatic lymph nodes, which delays diabetes progression. In contrast, expression of tumor necrosis factor alpha until 28 days of age prevents islet accumulation of CD4+CD25+TGF-beta+ Treg cells, resulting in acceleration to diabetes. Furthermore, adoptive transfer experiments demonstrated that CD4+CD25+ Treg cells could not control na?ve or activated islet-reactive CD8+ T cells bearing a dominant negative TGF-beta receptor type II. Our data demonstrate that, in vivo, TGF-beta signaling in CD8+ T cells is critical for CD4+CD25+ Treg cell suppression of islet-reactive CD8+ T cells in type 1 diabetes.     

Endothelial stroma programs hematopoietic stem cells to differentiate into regulatory dendritic cells through IL-10

Regulatory dendritic cells (DCs) have been reported recently, but their origin is poorly understood. Our previous study demonstrated that splenic stroma can drive mature DCs to proliferate and differentiate into regulatory DCs, and their natural counterpart with similar regulatory function in normal spleens has been identified. Considering that the spleen microenvironment supports hematopoiesis and that hematopoietic stem cells (HSCs) are found in spleens of adult mice, we wondered whether splenic microenvironment could differentiate HSCs into regulatory DCs. In this report, we demonstrate that endothelial splenic stroma induce HSCs to differentiate into a distinct regulatory DC subset with high expression of CD11b but low expression of Ia. CD11b(hi)Ia(lo) DCs secreting high levels of TGF-beta, IL-10, and NO can suppress T-cell proliferation both in vitro and in vivo. Furthermore, CD11b(hi)Ia(lo) DCs have the ability to potently suppress allo-DTH in vivo, indicating their preventive or therapeutic perspectives for some immunologic disorders. The inhibitory function of CD11b(hi)Ia(lo) DCs is mediated through NO but not through induction of regulatory T (Treg) cells or T-cell anergy. IL-10, which is secreted by endothelial splenic stroma, plays a critical role in the differentiation of the regulatory CD11b(hi)Ia(lo) DCs from HSCs. These results suggest that splenic microenvironment may physiologically induce regulatory DC differentiation in situ.     

Mouse CD11c(+) B220(+) Gr1(+) plasmacytoid dendritic cells develop independently of the T-cell lineage

The developmental origin of dendritic cells (DCs) is controversial. In the mouse CD8alpha(+) and CD8alpha(-) DC subsets are often considered to be of lymphoid and myeloid origin respectively, although evidence on this point is conflicting. Very recently a novel CD11c(+) B220(+) DC subset has been identified that appears to be the murine counterpart to interferon alpha (IFNalpha)-producing human plasmacytoid DCs (PDCs). We show here that CD11c(+) B220(+) mouse PDCs, like human PDCs, are present in the thymus and express T lineage markers such as CD8alpha and CD4. However, the intrathymic development of PDCs can be completely dissociated from immature T lineage cells in mixed chimeras established with bone marrow cells from mice deficient for either Notch-1 or T-cell factor 1, two independent mutations that severely block early T-cell development. Our data indicate that thymic PDCs do not arise from a bipotential T/DC precursor.     

Priming CD8+ T cells with dendritic cells matured using TLR4 and TLR7/8 ligands together enhances generation of CD8+ T cells retaining CD28

TLRs expressed on dendritic cells (DCs) differentially activate DCs when activated alone or in combination, inducing distinct cytokines and costimulatory molecules that influence T-cell responses. Defining the requirements of DCs to program T cells during priming to become memory rather than effector cells could enhance vaccine development. We used an in vitro system to assess the influence of DC maturation signals on priming naive human CD8+ T cells. Maturation of DCs with lipopolysaccharide (LPS; TLR4) concurrently with R848 (TLR7/8) induced a heterogeneous population of DCs that produced high levels of IL12 p70. Compared with DCs matured with LPS or R848 alone, the DC population matured with both adjuvants primed CD8+ T-cell responses containing an increased proportion of antigen-specific T cells retaining CD28 expression. Priming with a homogenous subpopulation of LPS/R848-matured DCs that were CD83(Hi)/CD80+/CD86+ reduced this CD28+ subpopulation and induced T cells with an effector cytokine signature, whereas priming with the less mature subpopulations of DCs resulted in minimal T-cell expansion. These results suggest that TLR4 and TLR7/8 signals together induce DCs with fully mature and less mature phenotypes that are both required to more efficiently prime CD8+ T cells with qualities associated with memory T cells.     

Engagement of the external domains of CD45 tyrosine phosphatase can regulate the differentiation of immature CD4+CD8+ thymocytes into mature T cells.

Immature precursor cells are induced in the thymus to express clonotypic T-cell antigen receptors (TCRs) and to differentiate into mature T cells. Perhaps the least understood event which occurs during intrathymic development is the positive selection of immature CD4+CD8+ thymocytes for differentiation into mature CD4+ and CD8+ T cells based on the TCR specificity individual thymocytes express. TCR expression by CD4+CD8+ thymocytes is quantitatively regulated by CD4-mediated activation of p56lck protein-tyrosine kinase whose activity can in turn be regulated by the membrane-bound protein-tyrosine-phosphatase CD45. Here we show that antibody engagement of CD45 external domains enhances Lck tyrosine kinase activity in CD4+CD8+ thymocytes, inhibits TCR expression, and inhibits differentiation of immature CD4+CD8+ thymocytes into mature T cells. Thus, engagement of the external domains of CD45 tyrosine phosphatase can regulate the ability of immature CD4+CD8+ thymocytes to undergo positive selection, suggesting an important regulatory role for intrathymic ligands that are capable of engaging CD45 within the thymus.     

Proliferation and differentiation potential of human CD8+ memory T-cell subsets in response to antigen or homeostatic cytokines

Four human CD8+ T-cell subsets, naive (CCR7+CD45RA+), central memory (TCM, CCR7+CD45RA-), effector memory (TEM, CCR7-CD45RA-), and CD45RA+ effector memory cells (TEMRA, CCR7-CD45RA+) were compared for their capacity to proliferate and differentiate in response to antigen or homeostatic cytokines. Cytokine responsiveness and interleukin-15 receptor expression were low in naive T cells and progressively increased from TCM to TEM and TEMRA. In contrast, the capacity to accumulate in response to T-cell receptor (TCR) or cytokine stimulation showed a reciprocal pattern and was associated with resistance to cell death and Bcl-2 expression. Whereas all TCR-stimulated cells acquired a CD45RA-CCR7- phenotype, cytokine-stimulated cells maintained their phenotype with the exception of TCM cells, which expressed CCR7, CD45RA, and perforin in various combinations. Single CD8+ TCM cells, but not TEM cells, could be expanded with cytokines, and the obtained clones displayed several distinct phenotypes, suggesting that TCM cells are heterogeneous. Consistently, CCR4 expression in the CD8+ TCM pool discriminated CCR4+ type 2 polarized cells (Tc2) and CCR4-CTL precursors. Finally, ex vivo bromodeoxyuridine (BrdU) incorporation experiments revealed that memory subsets have different in vivo proliferation rates, with CCR4-TCM having the highest turnover and TEMRA the lowest. These results show that human CD8+ memory T-cell subsets have different proliferation and differentiation potentials in vitro and in vivo. Furthermore, they suggest that TEMRA cells are generated from a TCM subset upon homeostatic proliferation in the absence of antigen.     

Precommitment of CD4+CD8+ thymocytes to either CD4 or CD8 lineages.

CD4+ and CD8+ mature T cells arise from CD4+CD8+ precursors in the thymus. During this process, cells expressing T-cell receptors (TCRs) reactive with self major histocompatibility complex (MHC) class I or II molecules are positively selected to the CD8 or CD4 lineage, respectively. It is controversial whether lineage commitment of CD4+CD8+ thymocytes is controlled directly by TCR specificity for MHC (instructional model) or, alternatively, by processes that operate independently of TCR specificity (stochastic model). We show here that CD4+CD8+ thymocytes bearing a MHC class I-restricted transgenic TCR can be subject to two alternative developmental fates. One population of CD4+CD8+ cells is positively selected by MHC class I molecules to the CD8 lineage as expected, whereas the other CD4+CD8+ population rearranges endogenous TCR genes and is positively selected by MHC class II molecules to the CD4 lineage. Blocking TCR-MHC class II interactions in vivo does not interfere with the generation of CD4+CD8+ cells expressing endogenous TCRs but does prevent their subsequent maturation to CD4+ cells. These data support a version of the stochastic model in which CD4+CD8+ thymocytes are precommitted to the CD4 or CD8 lineage independently of TCR specificity for MHC and prior to positive selection.