Gammadelta T cell function varies with the expressed WC1 coreceptor

WC1 molecules are transmembrane glycoproteins belonging to the scavenger receptor cysteine-rich family and uniquely expressed on gammadelta T cells. Although participation of WC1+ gammadelta T cells in immune responses is well established, very little is understood regarding the significance of expressing different forms of the WC1 molecule. Two forms previously identified by mAbs, i.e., WC1.1 and WC1.2, are expressed by largely nonoverlapping subpopulations of gammadelta T cells. In this study it was shown that expression of the WC1.1 coreceptor was the main indicator of proliferation and IFN-gamma production in response to autologous and bacterial Ags as well as for IFN-gamma production without proliferation in Th1-polarizing, IL-12-containing cultures. Nevertheless, after culture in either Th1-polarizing or neutral conditions, mRNA was present for both T-bet and GATA-3 as well as for IL-12Rbeta2 in WC1.1+ and WC1.2+ subpopulations, and neither produced IL-4 under any conditions. Although the steady decrease in the proportion of WC1.1+ cells, but not WC1.2+ cells, within PBMC with animal aging suggested that the two subpopulations may have different roles in immune regulation, cells bearing either WC1.1 or WC1.2 expressed mRNA for regulatory cytokines IL-10 and TGF-beta, with TGF-beta being constitutively expressed by ex vivo cells. Overall, the results demonstrate that the form of the WC1 coreceptor expressed on gammadelta T cells divides them into functional subsets according to IFN-gamma production and proliferative capacity to specific stimuli as well as with regard to representation within PBMC. Finally, evidence is provided for minor differences in the intracytoplasmic tail sequences of WC1.1 and WC1.2 that may affect signaling.     

Gammadelta T cells promote a Th1 response during coxsackievirus B3 infection in vivo: role of Fas and Fas ligand

Fas/Fas ligand (FasL) interactions regulate disease outcome in coxsackievirus B3 (CVB3)-induced myocarditis. MRL(+/+) mice infected with CVB3 develop severe myocarditis, a dominant CD4(+) Th1 (gamma interferon [IFN-gamma(+)]) response to the virus, and a predominance of gammadelta T cells in the myocardial infiltrates. MRL lpr/lpr and MRL gld/gld mice, which lack normal expression of Fas and express a mutated FasL, respectively, have minimal myocarditis and show a dominant CD4(+) Th2 (interleukin-4 [IL-4(+)]) phenotype to CVB3. Spleen cells from virus-infected wild-type, lpr, and gld animals proliferate equally to virus in vitro. Adoptive transfer of gammadelta T cells from hearts of CVB3-infected MRL(+/+) mice (FasL(+)) into infected MRL gld/gld recipients (FasL(-)/Fas(+)) restores both disease susceptibility and Th1 cell phenotype. However, transfer of these cells into MRL lpr/lpr recipients (FasL(+)/Fas(-)) did not promote myocarditis and the viral response remained Th2 biased. This paralleled the expression of very high surface levels of FasL by myocardial gammadelta T cells, as well as their propensity to selectively lyse Th2 virus-specific CD4(+) T cells. These results demonstrate that Fas/FasL interactions conferred by gammadelta T cells on lymphocyte subpopulations may regulate the cytokine response to CVB3 infection and pathogenicity.     

Both gamma delta T cells and NK cells inhibit the engraftment of xenogeneic rat bone marrow cells and the induction of xenograft tolerance in mice

In murine allogeneic bone marrow transplantation recipients, treatment of the hosts with a nonmyeloablative regimen, including depleting anti-CD4 and anti-CD8 mAbs, allows establishment of long-term mixed chimerism and donor-specific tolerance. However, in the xenogeneic rat-to-mouse combination, additional anti-Thy1.2 and anti-NK1.1 mAbs are required. We have now attempted to identify the xenoresistant mouse cell populations that are targeted by anti-NK1.1 and anti-Thy1.2 mAbs. C57BL/6 (B6) wild-type, B6 TCRbeta(-/-), and B6 TCRdelta(-/-) mice received anti-CD4 and anti-CD8 mAbs, followed by 3 Gy of whole body irradiation, 7 Gy of thymic irradiation, and transplantation of T cell-depleted rat bone marrow cells. Anti-NK1.1 and anti-Thy1.2 mAbs were additionally administered to some groups. Increased rat chimerism was observed in TCRdelta(-/-) mice treated with anti-CD4, anti-CD8, and anti-NK1.1 mAbs compared with similarly treated TCRbeta(-/-) mice. In TCRbeta(-/-) mice, but not in TCR delta(-/-) mice, donor chimerism was increased by treatment with anti-Thy1.2 mAb, indicating that CD4(-)CD8(-)TCRgammadelta(+)Thy1. 2(+)NK1.1(-) cells (gammadelta T cells) are involved in the rejection of rat marrow. In addition, chimerism was enhanced in both TCRbeta(-/-) and TCRdelta(-/-) mice treated with anti-CD4, anti-CD8, and anti-Thy1.2 mAbs by the addition of anti-NK1.1 mAb to the conditioning regimen. Donor-specific skin graft prolongation was enhanced by anti-Thy1.2 and anti-NK1.1 mAbs in TCRdelta(-/-) mice. Therefore, in addition to CD4 and CD8 T cells, gammadelta T cells and NK cells play a role in resisting engraftment of rat marrow and the induction of xenograft tolerance in mice.     

Bovine gamma delta T cell subsets express distinct patterns of chemokine responsiveness and adhesion molecules: a mechanism for tissue-specific gamma delta T cell subset accumulation

Subsets of gammadelta T cells localize to distinct tissue sites in the absence of exogenous Ag stimulation or development of effector/memory cells. Selective lymphocyte homing from the blood into tissues is controlled by a multistep process involving vascular and lymphocyte adhesion molecules, and G protein-linked chemokine receptors. The role of these mechanisms in the tissue tropism of gammadelta T cells is still poorly understood. In this study, we demonstrate that a subset of gammadelta T cells, most of which express an antigenically distinct TCR and are characterized by coexpression of CD8, selectively accumulated in tissues that expressed high levels of the mucosal vascular addressin, mucosal addressin cell adhesion molecule 1. These cells expressed higher levels of alpha(4)beta(7) integrins than other gammadelta T cell subsets and selectively migrated to the CCR7 ligand secondary lymphoid-tissue chemokine (CCL21). Integrin activation by CCL21 selectively increased CD8(+)gammadelta T cell binding to recombinant mucosal addressin cell adhesion molecule 1. These results suggest that the tropism of circulating CD8(+)gammadelta T cells for mucosal tissues is due, at least in part, to selective developmental expression of adhesion molecules and chemokine receptors.     

Homing and function of human skin gammadelta T cells and NK cells: relevance for tumor surveillance

Normal (noninflamed) human skin contains a network of lymphocytes, but little is known about the homing and function of these cells. The majority of alphabeta T cells in normal skin express CCR8 and produce proinflammatory cytokines. In this study we examined other subsets of cutaneous lymphocytes, focusing on those with potential function in purging healthy tissue of transformed and stressed cells. Human dermal cell suspensions contained significant populations of Vdelta1(+) gammadelta T cells and CD56(+)CD16(-) NK cells, but lacked the subsets of Vdelta2(+) gammadelta T cells and CD56(+)CD16(+) NK cells, which predominate in peripheral blood. The skin-homing receptors CCR8 and CLA were expressed by a large fraction of both cell types, whereas chemokine receptors associated with lymphocyte migration to inflamed skin were absent. Neither cell type expressed CCR7, although gammadelta T cells up-regulated this lymph node-homing receptor upon TCR triggering. Stimulation of cutaneous Vdelta1(+) gammadelta T cell lines induced secretion of large amounts of TNF-alpha, IFN-gamma, and the CCR8 ligand CCL1. In contrast to cutaneous alphabeta T cells, both cell types had the capacity to produce intracellular perforin and displayed strong cytotoxic activity against melanoma cells. We therefore propose that gammadelta T cells and NK cells are regular constituents of normal human skin with potential function in the clearance of tumor and otherwise stressed tissue cells.     

Hormonal regulation of CD4(+) T-cell responses in coxsackievirus B3-induced myocarditis in mice

Coxsackievirus B3 infection causes significant cardiac inflammation in male, but not female, B1.Tg.Ealpha mice. This gender difference in disease susceptibility correlates with selective induction of CD4(+) Th1 (gamma interferon-positive) cell responses in animals with testosterone, whereas estradiol promotes preferential CD4(+) Th2 (interleukin-4 positive [IL-4(+)]) cell responses. Differences in immune deviation of CD4(+) T cells cannot be explained by variation in B7-1 or B7-2 expression. Infection significantly upregulated both molecules, but no differences were detected between estradiol- and testosterone-treated groups. Significantly increased numbers of activated (CD69(+)) T cells expressing the gammadelta T-cell receptor were found in male and testosterone-treated male and female mice. In vivo depletion of gammadelta+ cells by using monoclonal antibodies inhibited myocarditis and resulted in a shift from a Th1 to Th2 response phenotype. Taken together, our results indicate that testosterone promotes a CD4(+) Th1 cell response and myocarditis by promoting increased gammadelta+ cell activation.     

Infiltration of canonical Vgamma4/Vdelta1 gammadelta T cells in an adriamycin-induced progressive renal failure model

We have previously reported an infiltration of renal interstitial gammadelta T cells in Adriamycin-induced progressive glomerulosclerosis in the rat kidney. The TCR repertoire and sequences used by these gammadelta T cells have now been studied. Two injections of Adriamycin 14 days apart caused segmental glomerulosclerosis, massive interstitial infiltration of mononuclear cells, and end-stage renal failure. Flow cytometry of lymphocyte subpopulations with Abs to CD3, the gammadelta TCR, and the alphabeta TCR showed that gammadelta T cells as a proportion of CD3(+) cells were increased in Adriamycin-treated kidneys (8.5 +/- 5.4%), but not in lymph nodes (1.3 +/- 0.4%). A semiquantitative score of glomerular damage (r = 0.65; p < 0.01) and creatinine (r = 0.62; p < 0.01) correlated significantly with the presence of gammadelta T cells. TCR Vgamma repertoire analysis by RT-PCR and Southern blotting showed that Vgamma2 was the dominant subfamily in lymph nodes, whereas Vgamma4 became the predominant subfamily in advanced stages of the rat Adriamycin-treated kidney. Sequencing of the Vgamma4-Jgamma junctional region showed an invariant sequence. The amino acid sequence of the junctional region of the Vgamma4 TCR was the same as the reported mouse canonical Vgamma4 TCR sequence. Analysis of the kidney Vdelta repertoire showed dominant expression of Vdelta1, and sequencing again revealed the selective expression of a canonical Vdelta1 gene. Semiquantitative RT-PCR for cytokine gene expression showed that gammadelta T cells from the kidneys expressed TGF-beta, but not IL-4, IL-10, or IFN-gamma. These results suggest that the predominant gammadelta T cells in the Adriamycin kidney use an invariant Vgamma4/Vdelta1 receptor.     

Depletion of a gamma delta T cell subset can increase host resistance to a bacterial infection

Gammadelta T lymphocytes have been shown to regulate immune responses in diverse experimental systems. Because distinct gammadelta T cell subsets, as defined by the usage of certain TCR V genes, preferentially respond in various diseases and disease models, we have hypothesized that the various gammadelta T cell subsets carry out different functions. To test this, we compared one particular gammadelta T cell subset, the Vgamma1(+) subset, which represents a major gammadelta T cell type in the lymphoid organs and blood of mice, to other subsets and to gammadelta T cells as a whole. Using LISTERIA: monocytogenes infection as an infectious disease model, we found that bacterial containment improves in mice depleted of Vgamma1(+) gammadelta T cells, albeit mice lacking all gammadelta T cells are instead impaired in their ability to control LISTERIA: expansion. Our findings indicate that Vgamma1(+) gammadelta T cells reduce the ability of the innate immune system to destroy LISTERIA:, even though other gammadelta T cells as a whole promote clearance of this pathogen.     

Prevalence of CD8(+)alpha beta T cells in Trypanosoma cruzi-elicited myocarditis is associated with acquisition of CD62L(Low)LFA-1(High)VLA-4(High) activation phenotype and expression of IFN-gamma-inducible adhesion and chemoattractant molecules

The determinants of the prevalence of CD8(+) T cells in the inflamed myocardium of Trypanosoma cruzi-infected patients and experimental animals are undefined. Using C3H/He mice infected with the Colombiana strain of T. cruzi, we found that the distribution of CD4(+)/CD8(-) and CD4(-)/CD8(+) T cells in the myocardium mirrors the frequency of cells expressing the CD62L(Low)LFA-1(High)VLA-4(High) activation phenotype among CD4(+)/CD8(-) and CD4(-)/CD8(+ )peripheral blood T cells. Consistently, vascular cell adhesion molecule-1-positive endothelial cells and a fine fibronectin network surrounding VLA-4(+) mononuclear cells were found in the inflamed myocardium. Further, interferon gamma (IFN-gamma) and IFN-gamma-induced chemokines (RANTES, MIG and CRG-2/IP-10), as well as JE/MCP-1 and MIP1-alpha, were found to be the dominant cytokines expressed in situ during acute and chronic myocarditis elicited by T. cruzi. In contrast, interleukin 4 mRNA was only detected during the chronic phase. Altogether, the results indicate that the distribution of T-cell subsets in the myocardium of T. cruzi-infected mice reflects the particular profile of adhesion molecules acquired by most peripheral CD8(+) T lymphocytes and point to the possibility that multiple IFN-gamma-inducible molecules present in the inflamed tissue contribute to the establishment and maintenance of T. cruzi-induced myocarditis.     

Gammadelta T cells enhance the expression of experimental autoimmune encephalomyelitis by promoting antigen presentation and IL-12 production

Using an adoptive transfer model of experimental autoimmune encephalomyelitis (EAE) induced by myelin basic protein (MBP)-reactive lymph node cells (LNC), we have shown that depletion of gammadelta T cells from LNC resulted in diminished severity of EAE in recipient mice, both clinically and histopathologically. The reduced potency of gammadelta T cell-depleted LNC to induce EAE correlated with decreased cell proliferation in response to MBP. The gammadelta T cell effect upon the threshold of MBP-induced LNC proliferation and EAE transfer was restored by reconstitution of gammadelta T cells derived from either MBP-immunized or naive mice, indicating that this effect was not Ag specific. The enhancing effect of gammadelta T cells on MBP-induced proliferation and EAE transfer required direct cell-to-cell contact with LNC. The gammadelta T cell effect upon the LNC response to MBP did not involve a change in expression of the costimulatory molecules CD28, CD40L, and CTLA-4 on TCRalphabeta(+) cells, and CD40, CD80, and CD86 on CD19(+) and CD11b(+) cells. However, depletion of gammadelta T cells resulted in significant reduction in IL-12 production by LNC. That gammadelta T cells enhanced the MBP response and severity of adoptive EAE by stimulating IL-12 production was supported by experiments showing that reconstitution of the gammadelta T cell population restored IL-12 production, and that gammadelta T cell depletion-induced effects were reversed by the addition of IL-12. These results suggest a role for gammadelta T cells in the early effector phase of the immune response in EAE.     

Selective expansion and partial activation of human NK cells and NK receptor-positive T cells by IL-2 and IL-15.

IL-2 and IL-15 are lymphocyte growth factors produced by different cell types with overlapping functions in immune responses. Both cytokines costimulate lymphocyte proliferation and activation, while IL-15 additionally promotes the development and survival of NK cells, NKT cells, and intraepithelial lymphocytes. We have investigated the effects of IL-2 and IL-15 on proliferation, cytotoxicity, and cytokine secretion by human PBMC subpopulations in vitro. Both cytokines selectively induced the proliferation of NK cells and CD56(+) T cells, but not CD56(-) lymphocytes. All NK and CD56(+) T cell subpopulations tested (CD4(+), CD8(+), CD4(-)CD8(-), alphabetaTCR(+), gammadeltaTCR(+), CD16(+), CD161(+), CD158a(+), CD158b(+), KIR3DL1(+), and CD94(+)) expanded in response to both cytokines, whereas all CD56(-) cell subpopulations did not. Therefore, previously reported IL-15-induced gammadelta and CD8(+) T cell expansions reflect proliferations of NK and CD56(+) T cells that most frequently express these phenotypes. IL-15 also expanded CD8alpha(+)beta(-) and Valpha24Vbeta11 TCR(+) T cells. Both cytokines stimulated cytotoxicity by NK and CD56(+) T cells against K562 targets, but not the production of IFN-gamma, TNF-alpha, IL-2, or IL-4. However, they augmented cytokine production in response to phorbol ester stimulation or CD3 cross-linking by inducing the proliferation of NK cells and CD56(+) T cells that produce these cytokines at greater frequencies than other T cells. These results indicate that IL-2 and IL-15 act at different stages of the immune response by expanding and partially activating NK receptor-positive lymphocytes, but, on their own, do not influence the Th1/Th2 balance of adaptive immune responses.     

Cutting Edge: A possible role for CD4+ thymic macrophages as professional scavengers of apoptotic thymocytes

A vast majority of thymocytes are eliminated during T cell development by apoptosis. However, apoptotic thymocytes are not usually found in the thymus, indicating that apoptotic thymocytes must be eliminated rapidly by scavengers. Although macrophages and dendritic cells are believed to play such role, little is known about scavengers in the thymus. We found that CD4(+)/CD11b(+)/CD11c(-) cells were present in the thymus and that they expressed costimulatory molecules for T cell selection and possessed Ag-presenting activity. Moreover, these CD4(+)/CD11b(+) cells phagocytosed apoptotic thymocytes much more efficiently than thymic CD4(-)/CD11b(+) cells as well as activated peritoneal macrophages. CD4(+)/CD11b(+) cells became larger along with thymus development, while no such change was observed in CD4(-)/CD11b(+) cells. Finally, engulfed nuclei were frequently found in CD4(+)/CD11b(+) cells. These results strongly suggest that thymic CD4(+)/CD11b(+) cells are major scavengers of apoptotic thymocytes.     

Murine T cell determination of pregnancy outcome.

At the fetomaternal interface, maternal effector cells come in intimate contact with fetal trophoblast cells which express paternal antigens. Failure of fetal trophoblast cells to activate maternal Th1 immune responses has been attributed in part to the absence of classical Class I and Class II major histocompatibilty complex (MHC) antigen expression and elaboration of factors which reduce TcR expression and shift any immune responses which may occur to Th2. Classical TcR alphabeta(+) T cells have not been found to be able to respond to trophoblasts. Recently, TcR gammadelta(+) T cells have been characterized in the low-abortion-rate pregnant C57Bl/10 mouse decidua, and the Vgamma1(+) subset may be able to respond to trophoblasts in a non-MHC-dependent manner. Trophoblast-recognizing T cells with Vgamma1 receptors are also present in the decidua of CBA/J mice pregnant by DBA/2, an abortion-prone mating combination. To test the role of the Vgamma1 subset of decidual gammadelta T cells in abortion-prone pregnancies, we altered this subset by injecting monoclonal anti-Vgamma1.1 antibody on gestation day 5.5, 1 day after implantation. This reduced detectability of a Vgammadelta subset producing TNF-alpha and reduced the abortion rate. Anti-Vgamma2, which reacts with a similar proportion of decidual gammadelta T cells as anti-Vgamma1.1, failed to prevent abortions. Vdelta6.3(+) cells are prominent at the fetomaternal interface, and anti-Vdelta6 antibody injected on day 5.5 prevented abortions. TGF-beta2(+) gammadelta cells first appear on day 8.5 of pregnancy; anti-Vgamma1.1 antibody injection on day 8.5 depleted these cells and boosted abortions; anti-Vdelta6.3 given on day 8.5 boosted abortions to the same level. These results suggest that two populations of Vgamma1.1(+)delta6.3(+) T cells may arise in the decidua: an early population that is Th1, abortogenic, and present during the time of implantation, and a Th2/3 cell subset that is present in the decidua later during pregnancy and which is pregnancy-protective.     

Identification and characterization of subpopulations in undifferentiated ES cell culture

Embryonic stem (ES) cells are pluripotent cells derived from the inner cell mass (ICM) and the epiblast, and have been suggested to be a homogeneous population with characteristics intermediate between them. These cells express Oct3/4 and Rex1 genes, which have been used as markers to indicate the undifferentiated state of ES cells. Whereas Oct3/4 is expressed in totipotent and pluripotent cells in the mouse life cycle, Rex1 expression is restricted to the ICM, and is downregulated in pluripotent cell populations in the later stages, i.e. the epiblast and primitive ectoderm (PrE). To address whether ES cells comprise a homogeneous population equivalent to a certain developmental stage of pluripotent cells or a heterogeneous population composed of cells corresponding to various stages of differentiation, we established knock-in ES cell lines in which genes for fluorescent proteins were inserted into the Rex1 and Oct3/4 gene loci to visualize the expression of these genes. We found that undifferentiated ES cells included at least two different populations, Rex1(+)/Oct3/4(+) cells and Rex1(-)/Oct3/4(+) cells. The Rex1(-)/Oct3/4(+) and Rex1(+)/Oct3/4(+) populations could convert into each other in the presence of LIF. In accordance with our assumption that Rex1(+)/Oct3/4(+) cells and Rex1(-)/Oct3/4(+) cells have characteristics similar to those of ICM and early-PrE cells, Rex1(+)/Oct3/4(+) cells predominantly differentiated into primitive ectoderm and contributed to chimera formation, whereas Rex1(-)/Oct3/4(+) cells differentiated into cells of the somatic lineage more efficiently than non-fractionated ES cells in vitro and showed poor ability to contribute to chimera formation. These results confirmed that undifferentiated ES cell culture contains subpopulations corresponding to ICM, epiblast and PrE.     

The restricted expression of granzyme M in human lymphocytes

We have analyzed the expression of human granzyme M (Gzm M) in various human leukocyte subsets using the specific mAb 4H10. Using FACS and Western blotting analysis we compared the expression of Gzm M with that of other granzymes (Gzm A and Gzm B) and the lytic protein perforin. Human Gzm M was constitutively highly expressed in NK cells as was perforin and Gzm A. Surprisingly, freshly isolated NK cells had very low (sometimes undetectable) levels of Gzm B. In contrast to Gzm B and perforin, Gzm M was not detected in highly purified CD4(+) and CD8(+) T cells either constitutively or after short term activation in vitro. However, low levels of Gzm M were observed in some T cell clones on prolonged passage in vitro. Gzm M was not detected in highly purified neutrophils, monocytes, or tumor cells of the myelomonocytic lineage. Examination of minor T cell subsets from human peripheral blood showed detectable Gzm M in CD3(+), CD56(+) T cells and gammadelta T cells. A histological staining procedure was developed that demonstrated a granular staining pattern for Gzm M and a cellular distribution similar to that observed by Western blotting. These data indicate that the expression of Gzm M does not always correlate with the lytic activity of cytotoxic cells. However, expression of Gzm M in NK cells, CD3(+), CD56(+) T cells, and gammadelta T cells suggests that this enzyme may play some role in innate immune responses.     

Primary hyperparathyroidism is associated with increased circulating bone marrow-derived progenitor cells

Recently, parathyroid hormone (PTH) was shown to support survival of progenitor cells in bone marrow. The release of progenitor cells occurs in physiological and pathological conditions and was shown to contribute to neovascularization in tumors and ischemic tissues. In the present study we sought to investigate prospectively the effect of primary hyperparathyroidism (PHPT) on mobilization of bone marrow-derived progenitor cells. In 22 patients with PHPT and 10 controls, defined subpopulations of circulating bone marrow-derived progenitor cells (BMCs) were analyzed by flow cytometry (CD45(+)/CD34(+)/CD31(+) cells indicating endothelial progenitor cells, CD45(+)/CD34(+)/c-kit(+) cells indicating hematopoietic stem cells, and CD45(+)/CD34(+)/CXCR4(+) cells indicating progenitor cells with the homing receptor CXCR4). Cytokine serum levels (SCF, SDF-1, VEGF, EPO, and G-CSF) were assessed using ELISA. Levels of PTH and thyroid hormone as well as serum electrolytes, renal and liver parameters, and blood count were analyzed. Our data show for the first time a significant increase of circulating BMCs and an upregulation of SDF-1 and VEGF serum levels in patients with PHPT. The number of circulating BMCs returned to control levels measured 16.7 +/- 2.3 mo after surgery. There was a positive correlation of PTH levels with the number of CD45(+)/CD34(+)/CD31(+), CD45(+)/CD34(+)/c-kit(+), and CD45(+)/CD34(+)/CXCR4(+) cells. However, there was no correlation between cytokine serum concentrations (SDF-1, VEGF) and circulating BMCs. Serum levels of G-CSF, EPO, and SCF known to mobilize BMCs were even decreased or remained unchanged, suggesting a direct effect of PTH on stem cell mobilization. Our data suggest a new function of PTH mobilizing BMCs into peripheral blood.     

gammadelta T cells may dichotomously modulate infection with avirulent Salmonella choleraesuis via IFN-gamma and IL-13 in mice

To investigate the roles of gammadelta T cells in Salmonella infection, we examined the resolution of an intraperitoneal infection with avirulent Salmonella choleraesuis 31N-1 in mice lacking T-cell-receptor (TCR) alphabeta T cells by disruption of the TCRbeta chain gene (TCRbeta(-/-)). The bacteria in TCRbeta(-/-) mice decreased with kinetics similar to that seen in control mice (TCRbeta(+/+)) after infection. The number of natural killer (NK) cells in the peritoneal cavity increased on day 6 after infection and thereafter decreased in both TCRbeta(-/-) and TCRbeta(+/+) mice, whereas the number of gammadelta T cells, in place of alphabeta T cells, increased remarkably in the peritoneal cavity of TCRbeta(-/-) mice on day 6 after infection. The NK cells from Salmonella-infected TCRbeta(-/-) mice produced interferon-gamma (IFN-gamma) but neither interleukin-4 (IL-4) nor IL-13 in response to immobilized anti-NK1.1 monoclonal antibody (mAb). The gammadelta T cells produced IFN-gamma but neither IL-4 nor IL-13 in response to heat-killed Salmonella, whereas both IFN-gamma and IL-13 but no IL-4 was produced by the gammadelta T cells stimulated with immobilized anti-TCRgammadelta mAb. In vivo administration of anti-NK1.1 mAb inhibited the reduction of Salmonella, whereas anti-TCRgammadelta mAb treatment did not affect the bacterial growth in TCRbeta(-/-) mice after Salmonella infection. However, neutralization of endogenous IL-13 with anti-IL-13 mAb enhanced the bacterial clearance in TCRbeta(-/-) mice after infection. These results suggest that NK1.1(+) cells serve mainly to protect against avirulent Salmonella infection in the absence of alphabeta T cells, whereas gammadelta T cells may play dichotomous roles in Salmonella infection through IFN-gamma and IL-13 in TCRbeta(-/-) mice.     

Ligation of the WC1 receptor induces gamma delta T cell growth arrest through fumonisin B1-sensitive increases in cellular ceramide

Ceramide is a powerful regulator of cell fate, inducing either apoptosis or growth arrest. We have previously shown that an Ab to the gammadelta T cell-specific orphan receptor, WC1, is able to induce growth arrest in proliferating IL-2-dependent gammadelta T cells. We now show that this WC1-mediated growth arrest is associated with an increase in cellular ceramide, in the absence of any measurable changes in acidic/neutral sphingomyelinase activity. Moreover, cell-permeable analogues of ceramide also mimicked WC1-induced growth arrest along with an associated decrease in pocket protein expression and phosphorylation status. An important role for ceramide in WC1-induced growth arrest was confirmed by demonstrating that the specific ceramide synthase inhibitor fumonisin B1 blocked WC1-induced growth arrest and the associated molecular effects on the pocket proteins. Finally, we observed constitutive expression of both antiapoptotic factors bcl-2 and bcl-X, the former having increased expression upon WC1 stimulation. It is therefore proposed that ligation of WC1 leads to an accumulation in cellular ceramide through activation of ceramide synthase. This in turn results in a decreased overall expression of the pocket proteins pRb and p107, their hypophosphorylation, and an eventual growth arrest of the gammadelta T cell. To our knowledge, these results demonstrate for the first time that cell surface receptor-mediated ceramide synthase activation can affect cell fate through increases in cellular ceramide and provide further evidence that the orphan receptor WC1 regulates gammadelta T cell biology through a novel signaling pathway.