Preferential expansion of V gamma 9 V delta 2 T cells following stimulation of peripheral blood lymphocytes with extracts of Plasmodium falciparum.

A Plasmodium falciparum schizont lysate has been previously described as being a powerful inducer of proliferation for human peripheral T lymphocytes. In this report we study the phenotype of cycling T cells from unexposed donors and examine how the P. falciparum lysate compares with the conventional T cell mitogen phytohemagglutinin (PHA), a known superantigen staphylococcal enterotoxin B (SEB), and a classical antigen pure protein derivative (PPD). We show that for this lymphoproliferative activity interaction with the MHC class II molecule is required and that in the presence of P. falciparum the great majority of the cycling cells at day 6 are gamma delta T cells, all of them bearing V gamma 9 V delta 2. Our results suggest that P. falciparum induces a T cell proliferative response that resembles a response of human peripheral blood gamma delta T cells to superantigen. This observation is in agreement with the elevated level of peripheral gamma delta lymphocytes observed during and after malaria acute infection.     

Mechanism of murine Vgamma1+ gamma delta T cell-mediated innate immune response against Listeria monocytogenes infection

Murine gamma delta T cells participate in innate immune response against infection of the intracellular bacterium Listeria monocytogenes. In the present report, we analyzed the mechanism of the gamma delta T cell-mediated response against L. monocytogenes infection. gamma delta T cell-enriched spleen cells of L. monocytogenes-infected mice produced IFN-gamma in vitro in response to L. monocytogenes-infected spleen cells. The IFN-gamma production was abrogated by depletion of Vgamma1+ gamma delta T cells. IFN-gamma production of the Vgamma1+ gamma delta T cells in response to L. monocytogenes-infected spleen cells required IL-12. However, addition of Fab fragment of anti-TCR gamma delta monoclonal antibodies (mAb) failed to block the response, suggesting that the response requires no TCR-mediated antigen recognition. Interestingly, Vgamma1+ gamma delta T cells of naive mice also produced IFN-gamma in response to L. monocytogenes-infected spleen cells in an IL-12-dependent manner. Furthermore, the IL-12 receptor (IL-12R) gene was expressed on the Vgamma1+ gamma delta T cells of naive mice as well as those of L. monocytogenes-infected mice although naive alpha beta T cells lack IL-12R expression. All the results suggest that the Vgamma1+ gamma delta T cells participate in immune surveillance against intracellular bacterial infection through quick production of IFN-gamma in response to infection-induced IL-12 without antigen-driven clonal expansion and maturation.     

Increase of gamma/delta T cells in hospital workers who are in close contact with tuberculosis patients.

gamma/delta T cells are likely to participate in the immune response to tuberculous infection in humans. In this study, we carried out an investigation to characterize the responsiveness of gamma/delta T cells from tuberculous patients and healthy individuals to mycobacterial stimulation in vitro. Healthy subjects were assigned to the following two groups: those who had been exposed to tuberculosis (contacts) and those who had not been exposed (noncontacts). The percent gamma/delta T cells in fresh peripheral blood obtained from health care workers who were tuberculin skin test positive and who had constant contact with patients with active tuberculosis (healthy contacts) was significantly higher, whereas healthy noncontacts showed the normal range of gamma/delta T cells. Patients with active pulmonary tuberculosis also had low levels of gamma/delta T cells. HLA-DR antigen-bearing activated gamma/delta T cells were observed in higher percentages among healthy contacts than among healthy noncontacts or patients with pulmonary tuberculosis. In healthy contacts, gamma/delta T cells increased as a percentage of peripheral blood mononuclear cells after in vitro stimulation with purified protein derivative (PPD) tuberculin compared with the percentage of fresh peripheral blood mononuclear cells that they made up, whereas no such increase was observed in patients with tuberculosis or in healthy noncontacts. Phenotypic analysis of the gamma/delta T cells in healthy contacts, which increased in number in vitro in response to PPD, revealed the preferential outgrowth of CD4+ V gamma 2+ gamma/delta T cells. This expansion of gamma/delta T cells by PPD required accessory cells, and it was inhibited by the addition of an antibody against HLA-DR in culture. Proteolytic digestion of PPD showed that gamma/delta T cells increased in number in response to peptide, but not nonpeptide, components of PPD. These findings suggest that gamma/delta T cells, especially CD4+ V gamma 2+ gamma/delta T cells, may participate in the immune surveillance of tuberculous infections in humans.     

Modulation of immune responses to Mycobacterium bovis in cattle depleted of WC1(+) gamma delta T cells

It is accepted that cell-mediated immune responses predominate in mycobacterial infections. Many studies have shown that CD4(+) T cells produce Th1 cytokines, such as gamma interferon (IFN-gamma), in response to mycobacterial antigens and that the cytolytic activity of CD8(+) cells toward infected macrophages is important. However, the extent and manner in which gamma delta T cells participate in this response remain unclear. In ruminants, gamma delta T cells comprise a major proportion of the peripheral blood mononuclear cell population. We have previously shown that WC1(+) gamma delta T cells are involved early in Mycobacterium bovis infection of cattle, but their specific functions are not well understood. Here we describe an in vivo model of bovine tuberculosis in which the WC1(+) gamma delta T cells were depleted from the peripheral circulation and respiratory tract, by infusion of WC1(+)-specific monoclonal antibody, prior to infection. While no effects on disease pathology were observed in this experiment, results indicate that WC1(+) gamma delta T cells, which become significantly activated (CD25(+)) in the circulation of control calves from 21 days postinfection, may play a role in modulating the developing immune response to M. bovis. WC1(+)-depleted animals exhibited decreased antigen-specific lymphocyte proliferative response, an increased antigen-specific production of interleukin-4, and a lack of specific immunoglobulin G2 antibody. This suggests that WC1(+) gamma delta TCR(+) cells contribute, either directly or indirectly, toward the Th1 bias of the immune response in bovine tuberculosis--a hypothesis supported by the decreased innate production of IFN-gamma, which was observed in WC1(+)-depleted calves.     

Quantitative analysis of the response of human T cell receptor V gamma 9+ cells to Plasmodium falciparum.

Plasmodium falciparum stimulates peripheral blood gamma delta + T cells from unexposed donors. The responding cells bear V gamma 9+ chains of the T cell receptor, the majority of which, but not all, are associated with V delta 2 chains. We have analyzed whether the precursor frequency of these V gamma 9+ cells approaches that expected of superantigens or mitogens and whether, like a superantigen, the response is major histocompatibility complex (MHC) unrestricted. Limiting dilution analyses of the response of enriched gamma delta + T cells to P.falciparum using autologous and heterologous antigen-presenting cells suggest that the response is more characteristic of an antigen-specific MHC-restricted response. The frequencies are lower than would be expected if all V gamma 9+ cells respond, and there is a dramatically reduced response when allogeneic antigen-presenting cells are used.     

Perturbation and proinflammatory type activation of V delta 1(+) gamma delta T cells in African children with Plasmodium falciparum malaria

gamma delta T cells have variously been implicated in the protection against, and the pathogenesis of, malaria, but few studies have examined the gamma delta T-cell response to malaria in African children, who suffer the large majority of malaria-associated morbidity and mortality. This is unfortunate, since available data suggest that simple extrapolation of conclusions drawn from studies of nonimmune adults ex vivo and in vitro is not always possible. Here we show that both the frequencies and the absolute numbers of gamma delta T cells are transiently increased following treatment of Plasmodium falciparum malaria in Ghanaian children and they can constitute 30 to 50% of all T cells shortly after initiation of antimalarial chemotherapy. The bulk of the gamma delta T cells involved in this perturbation expressed V delta 1 and had a highly activated phenotype. Analysis of the T-cell receptors (TCR) of the V delta 1(+) cell population at the peak of their increase showed that all expressed V gamma chains were used, and CDR3 length polymorphism indicated that the expanded V delta 1 population was highly polyclonal. A very high proportion of the V delta 1(+) T cells produced gamma interferon, while fewer V delta 1(+) cells than the average proportion of all CD3(+) cells produced tumor necrosis factor alpha. No interleukin 10 production was detected among TCR-gamma delta(+) cells in general or V delta 1(+) cells in particular. Taken together, our data point to an immunoregulatory role of the expanded V delta 1(+) T-cell population in this group of semi-immune P. falciparum malaria patients.     

Human TcR gamma delta+ lymphocyte response on primary exposure to Plasmodium falciparum.

In 29 patients experiencing their first P. falciparum malarial attack, blood levels of TcR gamma delta+ lymphocytes were studied from the onset of infection to up to 6-9 months later. Blood TcR gamma delta+ lymphocytes, revealed using the TcR delta 1 monoclonal antibody (MoAb), were increased both in absolute and relative numbers. Alterations lasted for up to 3-4 months following the attack. A Ti gamma A/BB3 reactive V gamma 9 subset was preferentially amplified. In vitro, TcR gamma delta+ lymphocytes from both malaria-sensitized and unprimed donors responded to P. falciparum schizont extract (PFSE). PFSE-stimulated polyclonal T cell lines consisted principally in TcR gamma delta+ cells with a Ti gamma A+/BB3+ phenotype. Several TcR gamma delta+ T cell clones obtained from patients recovering from acute malarial attack were maintained in the presence of PFSE and autologous irradiated PBL. They belong to the V gamma 9 subset. In long-term cultures, TcR gamma delta+ clones progressively lost their capacity to react to PFSE antigen while they were able to proliferate and to exert cytotoxic activity in response to autologous TcR alpha beta+, PFSE-specific T lymphocyte clones. This suggests that regulatory interactions occur between activated TcR gamma delta+ and TcR alpha beta+ cells generated by P. falciparum. Sequential variations in blood TcR gamma delta+ and TcR alpha beta+ lymphocyte levels after primary exposure to P. falciparum suggest that such regulatory interactions may occur in vivo.     

Paracoccidioides brasiliensis-stimulated human gamma/delta T cells support antibody production by B cells.

Paracoccidioidomycosis patients show hyperactive humoral immune responses. Consequently, we investigated whether cytokines in supernatants from Paracoccidioides brasiliensis-stimulated gamma/delta T cells support B-cell activation. We detected proliferation of B cells and increased immunoglobulin M (IgM) and IgG production. Thus, gamma/delta T cells may participate in polyclonal B-cell activation during paracoccidioidomycosis.     

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

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

Recognition of heat shock proteins and gamma delta cell function

Recently evidence has accumulated suggesting that gamma delta cells may participate in the immune response to mycobacteria and other infectious organisms. Many mouse gamma delta cells are stimulated by the 65 kDa heat shock protein of M. bovis and human gamma delta cell lines reactive with this mycobacterial protein have also been isolated. Indirect evidence further suggests that gamma delta cells can recognize autologous heat shock proteins. In this article, Willi Born and colleagues focus on these and other recent findings and speculate on their importance to gamma delta cell function in vivo.     

Escherichia coli infection induces only fetal thymus-derived gamma delta T cells at the infected site

Intraperitoneal infection of mice with Escherichia coli induced activated TCR gamma delta T cells in the peritoneal cavity. We provide evidence that the E. coli-induced gamma delta T cells are derived only from the fetal thymus on the following grounds. The gamma delta T cells were not induced in athymic nude mice and irradiated bone marrow-transferred mice which lack fetal thymus-derived T cells. However, E. coli infection of fetal thymus-grafted nude mice did induce fetal thymus-derived gamma delta T cells. These results suggest that the fetal thymus-derived gamma delta T cells colonize the periphery during early ontogeny, and are maintained until adult age. The E. coli-induced gamma delta T cells express only the Vdelta1 gene. Vgamma6 was predominantly expressed whereas anti-Vgamma1 and anti-Vgamma4 monoclonal antibodies stained less than 3 % of the cells. Direct sequencing of PCR products revealed that Vgamma6 and Vdelta1 genes expressed by the E. coli-induced gamma delta T cells were invariant sequences identical to those expressed in the fetal thymus. The antigen (Ag) specificity of a T cell hybridoma expressing the fetal type Vgamma6 / Vdelta1(+) TCR could not be identified as the cells failed to respond to lipopolysaccharide, E. coli Ag, mycobacterial heat shock protein 65, or isopentenyl pyrophosphate. These results suggest that the Vgamma6 / Vdelta1(+) gamma delta T cells derived from fetal thymus can participate in immune responses against bacterial infection through recognition of a novel class of Ag which is not yet identified.     

Gamma delta T cells inhibit in vitro growth of the asexual blood stages of Plasmodium falciparum by a granule exocytosis-dependent cytotoxic pathway that requires granulysin

Several reports have stated the ability of gamma delta T cells to inhibit the growth of the asexual blood stages of Plasmodium falciparum in vitro. However, little information is available about the mechanisms involved. In this study, in vitro systems were used to study the role of the granule exocytosis-dependent cytotoxic pathway in the growth inhibition/killing of P. falciparum by human gamma delta T cells. Our results show that the inhibition requires cell-to-cell contact and that gamma delta T cells kill the asexual blood stages of P. falciparum through a granule exocytosis-dependent cytotoxic pathway after recognition of certain ligands or molecules expressed on the surface of infected erythrocytes or merozoites. The in vitro inhibitory capacity of gamma delta T cells was strongly correlated with the expression of granulysin in the cytotoxic granules, while non-inhibitory CD4+ and CD8+ T cells expressed very little, implicating a role for granulysin in parasite inhibition. This was further suggested by the addition of neutralizing anti-granulysin antibodies, which abrogated the parasite inhibitory capacity of the gamma delta T cells. Taken together, our results suggest that the capacity of gamma delta T cells for inhibition/killing of P. falciparum is based on the granule exocytosis-dependent cytotoxic pathway and that the presence of granulysin is essential to maintain efficient killing.     

Skewing of cytotoxic activity and chemokine production,but not of chemokine receptor expression,in human type-1/-2 gamma delta T lymphocytes

Human Vgamma9/Vdelta2(+) T lymphocytes participate in the immune response against intracellular pathogens through the secretion of type-1 cytokines and chemokines and by killing of infected cells. Little is known of the effects by type-2 differentiation of gamma delta cells on these functions. Here, we report that bona fide naive cord blood-derived gamma delta lymphocytes expanded in vitro with the mycobacterial antigen isopentenyl pyrophosphate (IPP) can be differentiated as either type-1 or type-2 cells, in the presence of an appropriate cytokine milieu. Instead, peripheral gamma delta cells from PPD-negative healthy adults displayed a type-1 cytokine profile, i.e. IPP-stimulated secretion of IFN-gamma, but not of IL-4 and IL-10. Moreover, they released the macrophage inflammatory protein (MIP)-1beta, but not IL-8 nor the Th2 chemoattractants I-309 and TARC (thymus and activation-regulated chemokine). This cytokine profile was not significantly affected by in vitro culture in Th2 polarizing conditions. Only in one case out of seven were peripheral gamma delta cells fully differentiated to type-2 lymphocytes, characterized by sustained IL-4 and IL-10 production, along with secretion of substantial amounts of IL-8, I-309 and TARC. Type-2 gamma delta T lymphocytes preferentially expressed the co-stimulatory molecule CD30; conversely, no skewing in chemokine receptor expression was observed. Both polarized populations displayed high levels of CXCR3 in the absence of CCR3, CCR4 and CCR5. Finally, type-1, but not type-2, gamma delta T lymphocytes killed IPP-pulsed U937 cells and displayed elevated perforin content. Overall, our data suggest that type-2 differentiation of gamma delta T lymphocytes profoundly affects both their effector functions and their potential to recruit the appropriate leukocyte subsets to the sites of inflammation.     

Exaggerated Proinflammatory and Th1 Responses in the Absence of γ/δ T Cells after Infection with Listeria monocytogenes

While gamma/delta T cells are involved in host defense and immunopathology in a variety of infectious diseases, their precise role is not yet clearly defined. In the absence of gamma/delta T cells, mice die after infection with a dose of Listeria monocytogenes that is not lethal in immunologically intact animals. Morbidity might result from insufficient levels of cytokines normally produced by gamma/delta T cells or conversely from an excess of cytokines due to a lack of down-regulation of the inflammatory response in the absence of gamma/delta T cells. Consistent with a regulatory role, we found that systemic levels of proinflammatory cytokines (interleukin-6 [IL-6], IL-12, and gamma interferon [IFN-gamma]) were significantly higher in the absence of gamma/delta T cells during the innate phase of the response. Using combinations of genetically altered and immunodepleted mice, we found evidence for gamma/delta T-cell-mediated regulation of IFN-gamma production by multiple cell types of both lymphoid and myeloid lineages. The antigen-specific alpha/beta T-cell response that followed the exaggerated innate response was also increased in gamma/delta T-cell-deficient mice. These findings are consistent with an emerging picture from a variety of immune response models of a critical role for gamma/delta T cells in down-modulation of the immune response.     

Cross-talk between gammadelta T lymphocytes and immune cells in humoral response.

The role of gamma delta T cells in immunoregulation is largely unknown. In the current study we noted that gamma delta T cells play a positive role in the humoral response. These positively acting gamma delta T cells are required for the successful adoptive cell transfer of the humoral response, as well as for in vitro generation of plaque-forming cells (PFC). The presented results show that gammadelta T cells cause an increase in interleukin-10 (IL-10) production, which partly elucidates the mechanism of action of these cells. However, experiments with cell culture inserts strongly suggest that direct cell-cell contact between immune and gamma delta H-2-compatible regulatory T cells is critical to the exertion of the positive immunoregulatory function of gamma delta cells. The mechanism of cross-talk between these two cell populations is still not clear but we regard as most likely that the positively acting gamma delta T cells may interact with a complex of heat-shock protein-non-polymorphic MHC (IB) on the surface of T helper type 2 and/or B cells. This could provide, by direct cell-cell contact, the cognate recognition between gamma delta T-cell receptors and heat-shock protein-MHC that leads to positive internal signalling in the immune cells.     

A subset of gamma delta lymphocytes is increased during HIV-1 infection.

The gamma delta T cell receptor (TcR) lymphocytes constitute 3-10% of human peripheral blood lymphocytes. Only a very small fraction of these cells is recognized by the delta TCS1 monoclonal antibody, directed against the V delta 1 chain of the receptor. We describe the immunological, virological and clinical data of a small group of seropositive subjects having high levels of gamma delta TcR T cells in the peripheral blood. Our flow cytometric studies show that most of these cells belong to the delta TCS1+ (V delta 1+), CD8 +/- (dim staining) subset. Patients with high gamma delta TcR T cell numbers were not characterized by the presence of an acute (IgM positive) or reactivated (as defined by high IgG titres against early antigen or IgA titres against viral capsidic antigen) Epstein-Barr virus infection. Cytomegalovirus infection was excluded by serological assays, and other herpes viral infections were not found after clinical examination. HIV p24 antigenaemia was present in two out of 11 subjects. AIDS patients had very high percentages of gamma delta TcR T cells. Altogether these data show that the selective expansion of delta TCS1+ cells in HIV1 seropositive subjects is not related to some exogenous antigen stimulation, but may be related to peculiar pathologic processes involving the immune system.     

Is there a role for gamma delta T cells in malaria?

Peripheral blood lymphocytes of the V gamma 9+ family of gamma delta T cells proliferate vigorously in response to Plasmodium falciparum. In this brief article, Jean Langhorne and colleagues discuss this response and assess the possible role of gamma delta T cells in the pathogenesis of malaria.     

The gamma/delta T-cell response to Plasmodium falciparum malaria in a population in which malaria is endemic.

Frequencies and absolute numbers of peripheral gamma/delta T cells have been reported to increase after episodes of Plasmodium falciparum malaria in adults with limited or no previous malaria exposure. In contrast, little is known about the gamma/delta T-cell response to malaria in children from areas where malaria is endemic, who bear the burden of malaria-related morbidity and mortality. We investigated the gamma/delta T-cell response in 19 Ghanaian children from an area of hyperendemic, seasonal malaria transmission. The children presented with cerebral malaria (n = 7), severe malarial anemia (n = 5), or uncomplicated malaria (n = 7) and were monitored from admission until 4 weeks later. We found no evidence of increased frequencies of gamma/delta T cells in any of the patient groups, whereas one adult expatriate studied in Ghana and three adults admitted to the hospital in Copenhagen, Denmark, all with uncomplicated, primary P. falciparum malaria, showed increased gamma/delta T-cell frequencies similar to those previously reported. All patients had lowered absolute numbers of peripheral gamma/delta T cells at admission, changing to increased numbers by days 7 to 14 and then returning to normal levels. The study raises questions regarding age and degree of previous exposure as determinants of malaria-induced gamma/delta T-cell responses.     

The role of gamma delta T lymphocytes in infection.

Many recent studies suggest an involvement of gamma delta T cells in the immune response to infectious pathogens including viruses, bacteria and eukaryotic parasites. However, it remains unclear whether the responses of gamma delta T cells are specifically directed against antigens derived from these pathogens or against infection-induced, host-derived ligands.     

Evidence that gamma delta T cells play a limited role in resistance to murine listeriosis.

A role for alpha beta and gamma delta T cells in protection against primary and secondary infection with Listeria monocytogenes was studied. The results show that mice depleted of either gamma delta T cells with 3A10 monoclonal antibody (mAb), or alpha beta T cells with anti-CD4 plus anti-CD8 mAb, or both types of T cells, remained capable of controlling Listeria multiplication during the first 4 days of primary sublethal infection. Moreover, mice depleted of either or both types of T cells also remained capable of resolving primary infection, although the absence of alpha beta T cells, but not gamma delta T cells, caused resolution to be slower. Likewise, Listeria-immune mice depleted of either alpha beta or gamma delta T cells remained capable of resolving secondary infection with a large inoculum of L. monocytogenes, although depletion of alpha beta T cells, and to a much lesser extent gamma delta T cells, resulted in early exacerbation of infection. However, immune mice depleted of both types of T cells lost their ability to resist a lethal Listeria challenge. Taken together, the results show that whereas neither type of T cell is needed for resistance to sublethal primary listeriosis, alpha beta T cells may act in concert with gamma delta T cells in protecting mice against lethal secondary infection. In addition, the results indicate that the role of gamma delta T cells in anti-Listeria resistance is much less important than the role of alpha beta T cells, and can be demonstrated mainly in the absence of alpha beta T cells.