CD38 expression on mouse T cells: CD38 defines functionally distinct subsets of {alpha}{beta} TCR+CD4 CD8 thymocytes

We have examined CD38 expression on mouse lymphocytes usingthe rat mAb NIM-R5 and demonstrate that CD38 expression is restrictedto {small tilde}8% of thymocytes. Although CD38 is absent fromthe majority of CD4+ CD8^– and CD4^–CD8⁺ T cells,we detected a strong correlation between CD36 expression andß⁺CD4^–CD8^– T cells in the thymus, withnearly 80% of ß TCR⁺CD4^–CD8^– thymocytesbeing CD38⁺. Using heat stable antigen (HSA) and CD38, we dividedß⁺CD4⁺CD8^– thymocytes into four subsets: HSA⁺CD38^–,HSA^– CD38^hi, HSA^–CD3810^low and HSA^– CD38^–.Two established characteristics of ß TCR⁺CD4CD8^–cells, bias towards Vß 8.2 TCR expression and highlevels of IL-4 production, were used to establish a possiblerelationship between the above thymocyte subsets. Our presentdata show that the HSA⁺CD38^– subset is not biased towardsVß8.2 TCR expression whereas the HSA^– CD38^–subset does show this bias (–47%). Neither of these subsetsmake IL-4 upon CD3 mediated stimulation. In contrast, the CD38⁺subsets are heavily biased toward Vß8.2 expressionand produce large amounts of IL-4 upon stimulation, particularlythe CD38^low cells. Taken together, these data suggest that thesefour subsets represent various stages of a possible differentiationpathway for ß TCR⁺ CD4^–CD8^– cells, withthe HSA⁺CD38^– subset being the most Immature while theHSA^–CD38^low subset is the most functionally mature. Thesecharacteristics support the view that ap TCR⁺CD4^–CD8^–T cells represent an independent lineage with a distinct, butas yet obscure, role in immunity     

IL-4 producing CD4+ TCR{alpha} {beta}int liver lymphocytes: influence of thymus, {beta}2-microglobulin and NK1.1 expression

The present report describes developmental, phenotypic and functionalfeatures of unconventional CD4⁺ TCRß lymphocytes.In C57BL/6 mice, the majority of liver lymphocytes expressingintermediate intensity of TCRß (TCRß^int)are CD4⁺NK1.1⁺ and express a highly restricted TCR V_ßrepertoire, dominated by V_ß8 with some contributionby V^ß7 and V^ß2. Although these cells expressthe CD4 co-receptor, they are present in H2-l Aß (Aß)^+/–gene disruption mutants but are markedly reduced in ß₂-microglobulin(ß₂m)^–/– mutant mice and hence are ß₂mdependent. Thymocytes expressing the CD4⁺NK1.1⁺ TCR ßphenotype are also (ß₂m) contingent, suggesting thatthese two T lymphocyte populations are related. The CD4⁺NK1.1⁺TCRßlymphocytes in liver and thymus share several markers such asLFA-1⁺, CD44⁺, CD5⁺, LECAM-1^– and IL-2Rßa. TheCD4⁺NK1.1 ⁺ TCRß^int liver lymphocytes were not detectedin athymic nuinu mice. We conclude that ß₂m expressionis crucial for development of the CD4⁺NK1.1⁺ TCRß^intliver lymphocytes and that thymus plays a major role. CD4⁺ TCRß^intliver lymphocytes were also identified in NK1.1⁺ mouse strains,there lacking the NK1.1 marker. We assume that the NK1.1 moleculeis a characteristic marker of the CD4⁺TCR"^int liver lymphocytesin NK1.1⁺ mouse strains, although its expression is not obligatoryfor their development. The liver lymphocytes from +₂m^+/–,but not from +₂m^–/–mice are potent IL-4 producersin response to CD3 or TCRß engagement and the IL-4production by liver lymphocytes was markedly reduced by treatmentwith anti-NK1.1 mAb. We conclude that the CD4+NK1.1⁺ TCRß^intliver lymphocytes are capable of producing IL-4 in responseto TCR stimulation.     

Predominant expression of invariant V{alpha}14+ TCR {alpha} chain in NK1.1+ T cell populations

A novel T cell subset characterized by cell surface NK1.1⁺ TCRß⁺expression was investigated for its TCR usage, particularlythat of invariant V14 TCR, which was found to be preferentiallyused in peripheral CD4^–CD8^–T cells developed atextrathymic sites. We found that NK₊ ß T cell subsetsaccount for 0.4% in thymocytes, 5% in the splenic T cells and40.5% in the bone marrow T cells. Among these NK⁺ ßT cells, two distinct subsets were detected; cell surface TCRV14⁺and V14^– subpopulations. Almost all of NK⁺ ßthymocytes express V14 mRNA; however, only<20% were positive,while >80% were negative or undetectable for V14 TCR expressionon the cell surface in the thymus. Similarly,50% of NK⁺ ßT cells in spleen and bone marrow are V14⁺; as revealed by FACS.TCR repertoire analysis by nucleotide sequences on inverse PCRproducts demonstrated that most NK⁺ ß T cells expressan invariant TCR encoded by the V14J281 gene with a 1 base N-regionin all tissues. Thus, invariant V14 TCR is uniquely expressedon NK T cells, and can be a marker to distinguish NK, NK T andT cells.     

Developmentally regulated expression of the PD-1 protein on the surface of double-negative(CD4 CD8 ) thymocytes

PD-1, a member of the Ig superfamily, was previously isolatedfrom an apoptosis-induced T cell hybridoma 2B4.11 by subtractivehybridization. Expression of the PD-1 mRNA is restricted tothymus in adult mice. Using an anti-PD-1 mAb (J43), we examinedexpression of the PD-1 protein during differentiation of thymocytesin normal adult, fetal and RAG-2^-/- mice with or without anti-CD3mAb stimulation. While PD-1 was expressed only on 3–5%of total normal thymocytes, –34% of the CD4^-CD8^- double-negative(DN) fraction are PD-1⁺ cells with two distinct expression levels(low and high). PD-1^high thymocytes belonged to TCR lineagecells. In the DN compartment of the TCR ß lineage,PD-1 expression started at the low level from the CD44⁺CD25⁺stage and the majority of thymocytes expressed PD-1 at the CD44^-CD25^-stage in which thymocytes express TCR ß chains. Theanti-CD3 antibody administration augmented the PD-1 expressionas well as the differentiation of the CD44^-CD25⁺ DN cells intothe CD44^-CD25^- DN stage, not only in normal mice but also inRAG-2-deficient mice. The fraction of the PD-1^low cells in theCD4⁺CD8⁺ double-positive (DP) compartment was very small (>5%)but increased by stimulation with the anti-CD3 antibody, althoughthe total number of DP cells was drastically reduced. The resultsshow that PD-1 expression is specifically induced at the stagespreceding clonal selection.     

Entry of CD4 CD8 immature thymocytes into the CD4/CD8 developmental pathway is controlled by tyrosine kinase signals that can be provided through TCR components

Entry of thymus-migrated precursor cells into the CD4/CD8 developmentalpathway was analyzed by using the short-term organ culturesof day 14 fetal mouse thymus lobes. Organ cultures of CD4^–CD8^–day 14 fetal thymocytes for 1-2 days resulted in the generationof CD4^–CD8⁺ cells, which were mostly immediate precursorcells for CD4⁺CD8⁺ thymocytes. This differentiation of CD4^–CD8^–thymocytes into CD4^–CD8⁺ cells was strongly enhanced byanti-CD3 antibodies. The anti-CD3-induced generation of CD4^–CD8⁺cells was even found in the immunodeficient scid fetal thymuscultures, and the cell surface CD3 expression on the scid fetalthymocytes could be directly visualized, indicating that functionalCD3 could be expressed on CD4^–CD8^– immature thymocyteswithout being associated with rearranged TCR components. Theanti-CD3-lnduced generation of CD4^–CD8⁺ cells from scidand normal fetal thymus cultures was inhibited by tyrosine kinaseinhibitors Herblmycin A and Tyrphostin. The generation of CD4^–CD8⁺cells in unstimulated normal fetal thymus cultures was alsomarkedly inhibited by the tyrosine kinase inhibitors but notby Cyclosporin A, suggesting that tyrosine klnase-dependentbut calclneurin-lndependent signals were essential for the differentiationof CD4^–CD8^– thymocytes. Interestingly, the generationof CD4^–CD8⁺ cells from the normal fetal thymus cultureswas modestly but consistently enhanced by anti-TCRßantibody, suggesting that functional TCRß in additionto CD3 was expressed on normal CD4^–CDS⁺ immature thymocytes.On the other hand, anti-TCR antibody did not affect this differentiationin the normal fetal thymus cultures and the generation of CD4^–CD8⁺cells from the normal fetal thymus cultures of TCR-deficientmice was still enhanced by anti-TCRß or anti-CD3 antibodies,indicating that either TCR chains or TCR⁺ cells were not involvedin the control of the differentiation into CD4^–CD8⁺ cells.These results indicate that the entry of CD4^–CD8^–immature thymocytes into the CD4/CD8 developmental pathway iscontrolled by tyrosine kinase signals and that these signalscan be provided through the engagement of TCR-CD3 complexeswith or without TCRß chains expressed on the CD4^–CD8^–immature thymocytes.     

Maturation of medullary thymic epithelium requires thymocytes expressing fully assembled CD3-TCR complexes

Unlike meduilary thymic epithelial cells (TEC) of normal mice,meduilary TEC of TCR^– SCID mice are immature and disorganized.In order to assess directly the role of TCR⁺ cells in the developmentof medullary TEC, we bred mice which co-expressed the SCID geneticdefect and transgenes encoding clonotypic TCR chains. Immunohistologicexamination revealed that meduilary thymic epithelial cellsfrom TCRß transgenic SCID mice, whose thymocytes onlyexpress TCRß chains that inefficiently associate withCD3 and , components, remained immature and disorganized. Incontrast, meduilary TEC from TCRß transgenic SCIDmice, whose thymocytes express fully assembled CD3--TCRßcomplexes were mature and organized. Interestingly, the abilityof TCRß⁺-⁺-CD3³ thymocytes to induce maturation ofmeduilary TEC appeared not to be related to the antigen specificityof the TCR as thyml from positively selecting, negatively selectingand non-selecting TCRß transgenic SCID mice all possessedinduced meduilary thymic epithelial cells. In addition, we foundthat induction of meduilary TEC cells was associated with thepresence of meduilary thymocytes, including those of the CD4-CD8-TCRß⁺phenotype. The present findings demonstrate that fully assembledCD3--TCR complexes are required to induce maturation of meduilarythymic epithelial cells and indicate that thymocyte inductionof meduilary thymic epithelial cells may result from signalingindependently of their clonotyplc chains.     

Migration pathways of CD4 T cell subsets in vivo: the CD45RC- subset enters the thymus via {alpha}4 integrin-VCAM-1 interaction

The present investigation examines the localization and migrationof purified T cell subsets in comparison with B cells, CD8 Tcells and CD4⁺CD8^– single-positive thymocytes. CD4 T cellsubsets in the rat are defined by mAb MRC OX22 ( anti-CD45RC),which distinguishes resting CD4 T cells (CD45RC⁺) from those(CD45RC^–) which have encountered antigen in the recentpast– subpopulations often referred to as ‘naive’and ‘memory’. Purified, ⁵¹Cr-labelled CD45RC⁺ CD4T cells broadly reflected the migration pattern of CD8 T cellsand B cells. Early localization to the spleen was followed bya redistribution to mesenteric lymph nodes (MLN) and cervicallymph nodes ( CLN) , B cells migrating at a slightly slowertempo. There was almost no localization of these subpopulationsto the small or large intestine [Peyer's patches (PP) excluded].In contrast, CD45RC^– CD4 T cells (indistinguishable insize from the CD45RC⁺ subset) localized in large numbers tothe intestine; they were present here at the earliest time point(0.5 h) , persisted for at least 48 h but did not accumulate,indicating a rapid exit. Numerically, localization of CD45RC^–CD4 T cells in the MLN could be accounted for entirely by afferentdrainage from the intestine. Unexpectedly, CD45RC^– CD4T cells (but not other subsets) localized and accumulated inthe thymus. In vivo treatment with mAb HP2/1 against the integrin₄ subunit inhibited almost entirely CD45RCT^– CD4 T cellmigration into the PP (98.1%), intestine (87.1%) , MLN (89.1%)and thymus (93.5%) migration into the CLN was only reduced byhalf. To distinguish between recognition of MAdCAM-1 and VCAM-1by ^4–containing integrins, recipients were treated withmAb 5F10 against rat VCAM-1. Except for the thymus and a smallreduction in CLN, localization of CD45RC^– CD4 T cellswas unaffected; entry to the thymus was almost completely blocked(92.3%) by anti-VCAM-1. The results indicated (i) that CD45RC^–CD4 T cells alone showed enhanced localization to the gut andPP, probably via ₄ß₇-MAdCAM-1 interaction; ( II) thatmany CD45RC^– cells entered nonmucosal LN independentlyof ₄ integrin or VCAM-1; and (III) that entry of mature recirculatlngCD45RC^– CD4 T cells into the thymus across thymic endothellumwas apparently regulated by ₄ integrln-VCAM-1 interaction.     

Phenotypically distinct subsets of CD4+ T cells induce or protect from chronic intestinal inflammation in C. B-17 scid mice

CD4⁺ T cells in the mouse can be subdivided into two fractionsbased on the level of expression of the CD45RB determinant.Previous studies have shown that these subsets are functionallydistinct. We have further characterized the properties of thesesubpopulations in vivo by injecting them into C. B-17 scid mice.The animals restored with the CD45RB^highCD4⁺ T cell populationdeveloped a lethal wasting disease with severe mononuclear cellinfiltrates into the colon and elevated levels of IFN- mRNA.In contrast, animals restored with the reciprocal CD45RB^lowsubset or with unfractionated CD4⁺ T cells did not develop thewasting or colitis. Importantly, the co-transfer of the CD45RB^lowpopulation with the CD45RB^high population prevented the wastingdisease and colitis. These data indicate that important regulatoryinteractions occur between the CD45RB^high and CD45RB^lowCD4⁺T cell subsets and that disruption of this mechanism has fatalconsequences.     

T cell receptor-triggered activation of intraepithelial lymphocytes in vitro

Intraepithelial lymphocytes (IEL) of the mouse small intestinewere examined for their potential to respond to TCR signallingin vitro. Purified IEL subsets were activated using mAbs specificfor CD3, TCRßor TCR&. Thy-1⁺IEL, regardless ofTCR type, proliferated equally well in response to anti-TCRmAb with or without exogenous IL-2. In contrast, Thy-1^–TCR, CD8^– IEL required exogenous IL-2 for proliferation.No such requirement was observed for Thy-1^– TCR& IELproliferation. IEL proliferation in the absence of added IL-2was due to an IL-2 secretion/IL-2 receptor (IL-2R) autocrinepathway, since mAbs specific for IL-2 and IL-2R inhibited IELproliferation. Thy-1⁺ CD8ß^– CD4⁺CD8⁺ IEL wereunresponsive to TCR-induced proliferation but exhibited highlevels of cytolytic activity upon TCR-triggerlng. Thy-1^–non-cytolytic IEL were induced to express Thy-1 and cytolytlcactivity following activation in vitro. In addition, the involvementof the co-stimulatory molecule CD28 in IEL activation was tested.CD28 was weakly expressed by fresh IEL and anti-CD28 mAb hadno effect on TCR-triggered proliferation. However, anti-TCRstimulation increased CD28 expression on a subset of TCRßIEL and the addition of anti-CD28 mAb resulted in increasedIL-2 production, but not in increased proliferation. Our resultsindicate that IEL, including the purported extrathymlc CD8ß^–subset, can respond to TCR-driven signals via proliferationand/or cytolytlc activity.     

CD2 expression on murine intestinal intraepithelial lymphocytes is bimodal and defines proliferative capacity

The CD2 molecule is normally expressed on nearly all murinelymphocytes, and is co-stimulatory in T cell activation viathe antigen receptor (TCR). A naturally occurring T lymphocytepopulation that is bimodal for CD2 expression was found in theintestinal intraepithelial lymphocytes (IEL). TCRß⁺IEL contain CD2^– and CD2⁺ cells of approximately equalproportion, while TCR⁺ IEL are predominantly CD2^–. Theproliferative response of IEL to stimulation with an anti-CD3mAb or with PMA plus ionomycin co-segregated with CD2 expression;the CD2⁺ subset proliferated vigorously under these conditionswhile the CD2^– subset was much less responsive. The respondingCD2⁺ IEL contained both TCRß⁺ and TCR⁺ cells. However,activation of the CD2^– IEL with anti-CD3 mAb resultedin only the expansion of TCR⁺ IEL, while activation with PMAplus ionomycin did not promote expansion of either the TCRß⁺or the TCR⁺ IEL. These findings parallel observations in theautoimmune lpr mouse, where massive numbers of peripheral TCRß⁺CD4^–CD8^–T cells that lack CD2 expression are also hyporesponsive tomltogenic stimulation. The apparent energy of CD2^–TCRß⁺IEL, as well as CD2^– T cells from lpr mice, demonstratesthat the absence of CD2 on TCRß⁺ T lymphocytes co-segregateswith nonresponsiveness.     

A role for BP-3/BST-1 antigen in early T cell development

In the mouse thymus, pre-T cells are defined by their CD3^–CD4^–CD8^–triple-negative, CD44^10/– CD25⁺ phenotype. We made a ratmAb IF-7, that, among all Tcell subsets analyzed, reacted exclusivelywith pre-T cells. Molecular cloning revealed that the antigenrecognized by IF-7 was identical to BP-3/BST-1, a glycosyl-phosphatidylinositol-linked,CD38-related molecule previously described asa possible co-activationmolecule of pre-B cells. We found that IF-7 cross-linking enhancesthe proliferative response ofsorted pre-T cells to anti-CD3stimulation. In addition, IF-7 enhances and accelerates thedevelopment of fetal thymic organ culture (FTOC), although the lineage is unaffected by the treatment. In addition, sortedIF-7⁺ pre-T cells give preferentially rise to ß TCR⁺thymocytes in FTOC. Our observations strongly suggest that BP-3/BST-1is implicated in both early B and T cell growth and development,and is an early marker for the ß lineage.     

CD4+Thy1- thymocytes with a Th-type 2 cytokine response

We have identified a small subset of CD3⁺, CD4⁺, CD8^–thymocytes that do not express Thy1 (CD90). This Thy1^–subset represents 1–3.7% of the total number of thymocytesin a naive mouse. CD4⁺Thy1^– thymocytes express high levelsof CD3, intermediate to high levels of heat-stable antigen (HSA),and low levels of CD25, CD45RB, CD69, CD44 and CD62L. They producehigh titers of IL-4 and no IFN- upon stimulation in vitro, aresponse characteristic of T_h2 cells. In the thymi of mice infectedneonatally with a high dose of the retrovirus Cas-Br-E MuLV,the frequency of CD4⁺Thy1^– cells increased ~10-fold. High-dosevirus infection resulted in decreased HSA and increased CD44expression on CD4⁺Thy1^– cells relative to cells from naivemice. CD4⁺Thy1^– cells from high-dose infected mice alsosecreted IL-4 and not IFN- upon in vitro stimulation. We previouslyreported that infection of newborn mice with a high dose ofmurine retrovirus results in the induction of a non-protectiveanti-viral T_h2 T cell response; CD4⁺Thy1^– thymocytes witha T_h2-like cytokine profile may play a role in determining thecytokine bias of this anti-viral response.     

In vivo and in vitro repertoire of CD3+CD4 CD8 thymocytes

CD4-CD8- thymocytes contain a cell subset which expresses thecomplete CD3-TCR complex (/ or /) of which the ontogeny andfiliation are unknown. One of the questions is whether thispopulation can be intrathymically selected, an obligatory stepfor the mature CD4⁺ and CD8⁺ cell differentiation pathway, orif the absence of CD4 and CD8 allows them to escape thymic selection.The repertoire of CD3 ⁺ CD4 - CD8 - (CD3 ⁺ DN) thymocytes wasanalyzed in different strains of mice with different combinationsof H-2 and Mls expression. The expression of V_ß8.1in freshly isolated CD3⁺ DN cells is the highest in Mls-l^b miceand the lowest in MIS-l^a and F₁ mice, suggesting that selectionagainst this specificity can be achieved in vivo. By contrast,a low percentage of V_ß6⁺ cells is found in all thestrains, with no correlation according to MIS expression. Invitro culture of DN thymocytes with IL-1 and IL-2 leads to theproliferation of CD3⁺ DN cells. In vitro culture amplifies thein vivo pattern of V_ß8.1 expression, while V_ß6⁺cells only expand in DN cells of 66 and 61002 Mls-l^b mice withthe same genetic background. These results show: (i) CD3⁺ DNthymic cells can be intrathymically selected but the repertoireis different from that of mature T cells; (ii) V_ß6and V_ß8.1 selection do not follow the same pattern;(iii) this repertoire can be modified by In vitro culture, towarda more mature type; and (iv) comparison of DN cell repertoireof BALBlc, BALB.D2 (congenic for MLs), and other strains ofmice suggests a multigenic control for this selection, and aninvolvement of background genes.     

Cytokine dependence of V{gamma}3 thymocytes: mature but not immature V{gamma}3 cells require endogenous IL-2 and IL-7 to survive evidence for cytokine redundancy

It has previously been described that V3 cells can proliferateextensively in vitro in the presence of different cytokines.Here, the role of cytokines in the maintenance of V3 cells inthe thymus has been determined. Culture of fetal thymocytesin cell suspension for 24 h showed that, whereas immature TCR^lowHSA^highV3cells remained viable, all mature TCR^highHSA_lowV cells died.These cells died by apoptosis since protein synthesis was requiredand flow cytometric analysis as well as DNA gel electrophoresisshowed that the DNA was degraded to oligonucleosomal bands.Addition of IL-2, IL-4 or IL-7 to suspension cultures of fetalthymocytes rescued V3 cells from dying. Addition of IL-1, IL-3,IL-5, IL-6, IL-9, TNE- or IFN- was without effect. Phenotypicanalysis showed that the -chain of the IL-2 receptor (IL-2R)was expressed by part of the immature V3 thymocytes, all matureV3 cells expressed the p-chain of the IL-2 receptor (IL-2RP).Addition of anti-IL-2R mAb to fetal thymic organ culture (FTOC)resulted in a moderate reduction of the cell number of matureV3 thymocytes. Addition of anti-IL-2Ra, anti-IL-4 or anti-IL-7mAb had no effect. The cell number of mature V3 cells was highlyreduced when both anti-IL-2Rp and anti-IL-7 mAb were added toFTOC. These results show that IL-2 and IL-7 are actively involvedin the maintenance of mature V3 cells in the thymus. This cytokinedependence of mature Vthymocytes may explain their selectivelocalization in skin epithelium.     

Characterization of virus-primed CD8+ T cells with a type 1 cytokine profile

Infection with lymphocytic chorlomeningitis virus is associatedwith marked polyclonal activation of the CD8⁺ T cell subpopulation.In this report the cytokine production of virus-activated Tcells is analyzed and the producing cells subset is characterizedphenotypically. CoinciB. A. Askonasding with other parametersof cell-mediated immunity, splenic T cells appear which areable to release high amounts of IFN-, but not IL-5, IL-10 ortumor necrosis factor- upon short-term stimulation with anti-CD3in vitro. A similar profile is observed analyzing T cells takenfrom an inflammatory site. Phenotypically, the main cytokine-producingcell subset is found to be CD8⁺ cells targeted for homing toinflammatory sites (VLA-4^hiL-selectin^lo) of which 30–40%were positive by intracellular staining for IFN-. This subsetalso contains all T cells with a cytotoxic potential as measuredby redirected killing. An enhanced cytotoxic potential as wellas an increased capacity to produce IFN- is observed for atleast 2 months after infection and cell sorting analysis revealedthat this could be ascribed to a long-standing increase in thefrequency of CD8⁺Pgp-1^hi cells. Therefore, these results demonstratethat systemic virus infection may exert marked perturbationof the CD8⁺ T cell population resulting in generation of a long-livedsubset of primed cells with important effector potential.     

NK1.1+ T cells from IL-7-deficient mice have a normal distribution and selection but exhibit impaired cytokine production

Particular subsets of T cells expressing the NK1.1 antigen havebeen proposed to play an immune regulatory role by their fastand strong production of cytokines, in particular IL-4. We soughtto determine factors driving the functional differentiationof NK1.1⁺ T cells. Since NK1.1⁺ T cells are exquisitely sensitiveto IL-7 stimulation, we analyzed the development, selectionand IL-4 production of NK1.1⁺ T cells in IL-7 deficient mice(IL-7–/–mice). Besides a sharp reduction of allT cell subsets, NK1.1⁺ T cells develop at normal relative frequenciesin IL-7–/–;mice. They also undergo a normal selectionprocess, as revealed by the biased V_ß TCR repertoireidentical to the one in IL-7+/+ mice. However, NK1.1₊ T cellsfrom IL-7+/+ mice were found to be impaired in IL-4 and IFN-production in in vitro and in vivo models. In addition, IL-7was able to restore IL-4 production by NK1.1⁺ thymocytes fromIL-7–/– mice. Finally, IL-7 but not IL-4 was ableto maintain and increase IL-4 production by NK1.1⁺ thymocytesfrom normal mice. These data suggest that the functional maturationof NK1.1⁺ T cells requires a cytokine-driven differentiationprocess, in which IL-7 plays a major role.