Two distinct T cell subsets, CD4+ and CD8+CD60+, and their cytokines are required for in vitro induction of human ragweed-specific memory IgE responses

CD8(+)CD60(+) T cells (80-98% CD45RO(+); 20% CD23(+)) are significantly increased in the blood of serum IgE(+) ragweed-sensitized (RS) compared with serum IgE-nonatopic humans (p = 0.001). CD8(+)CD60(+) T cells of the RS patients produced IL-2, IL-4, IL-10, IL-12, IFN-alpha. and IFN-gamma, but not IL-6 or IL-13. When their PBMC were cultured with ragweed Ag (RA), peak IgE responses occurred on day 10; none was induced with non-cross-reacting or without Ag; nonatopic PBMC did not respond to any stimulant. When either CD4(+) or CD8(+)CD60(+) T cells were depleted from RS PBMC before culture with RA, no IgE responses were induced. If purified CD4(+) T cells or low numbers of CD8(+)CD60(+) T cells were added back to the depleted PBMC, IgE responses were restored. However, higher numbers of CD8(+)CD60(+) T cells totally suppressed IgE responses. Total suppression also was obtained when RS PBMC were cultured with RA and either anti-IL-2, IL-4, IL-10, IL-12, IFN-gamma (all concentrations), or IFN-alpha (low concentrations), but not anti-IL-6 or IL-13. Higher concentrations of anti-IFN-alpha potentiated IgE responses.     

Measles virus infection in adults induces production of IL-10 and is associated with increased CD4+ CD25+ regulatory T cells

Despite steady progress in elimination of measles virus globally, measles infection still causes 500,000 annual deaths, mostly in developing countries where endemic measles strains still circulate. Many adults are infected every year in China, with symptoms more severe than those observed in children. In this study, we have used blood samples from adult measles patients in Shanghai and age-matched healthy controls to gain an understanding of the immune status of adult measles patients. IFN-alpha mRNA was reduced in patient PBMC compared with healthy controls. In contrast, gene expression and plasma production of IL-2, IL-10, and IFN-gamma were elevated in patient blood. A similar cytokine profile was observed at early times when cultured PBMC were infected with a clinical isolate of measles virus. In contrast to previous studies in pediatric patients, we did not find a reduction in total CD4(+) and CD8(+) T cells in patient PBMC. Interestingly, we found that CD4(+)CD25(+)CD127(low) regulatory T cells were significantly increased in patient PBMC compared with controls. Using intracellular cytokine staining we also show that the measles virus induces IL-10-producing CD14(+) and CD4(+)CD25(+) cells in PBMC. Our results show that adult measles patients in the acute phase of the disease have a mixed Th1/Th2 type response, accompanied with severe immunosuppression of both innate and adaptive responses including suppression of type I IFN. Both regulatory T cells and plasma IL-10 may contribute to the immunosuppression.     

Suppression of T lymphocyte function by measles virus is due to cell cycle arrest in G1

Measles virus suppresses T lymphocyte functions in vitro. When measles virus-infected T lymphocytes are stimulated with PHA or 12-O-tetradecanoylphorbol-13-acetate, plus calcium ionophore, the cells secrete IL-2 and express the IL-2R or Tac Ag to a similar extent as uninfected cells, yet proliferation is reduced by 50 to 90%. Stimulated infected T cells also express the cell surface activation Ag 4F2, transferrin R, and HLA-DR. The secretion of IFN-gamma by infected T cells in response to PHA is not suppressed at 24 to 72 h after stimulation. Total RNA synthesis at 48 and 72 h after stimulation is reduced in infected T lymphocytes. Infectious measles virus progeny are produced during this interval. Thus infected T lymphocytes can become activated in response to mitogenic stimuli and the cells support efficient viral replication before the block in cell proliferation.     

The serpin secreted by Brugia malayi microfilariae, Bm-SPN-2, elicits strong, but short-lived, immune responses in mice and humans

Understanding the basic immunology of an infectious disease requires insight into the pattern of T cell reactivity and specificity. Although lymphatic filariasis is a major tropical disease, the predominant T cell Ags of filarial species such as Brugia malayi are still undefined. We have now identified a prominent T cell Ag from B. malayi microfilariae (Mf) as Bm-SPN-2, a serpin secreted exclusively by this stage. Mf-infected mice mounted strong, but short-lived, Bm-SPN-2-specific Th1 responses, measured by in vitro production of IFN-gamma, but not IL-4 or IL-5, 14 days postinfection. By day 35, responsiveness to Bm-SPN-2 was lost despite enhanced reactivity to whole Mf extract. Single immunization with Mf extract also stimulated typical Th1 reactions to Bm-SPN-2, but IgG1 Ab responses dominated after repeated immunizations. Human patients displayed potent humoral responses to Bm-SPN-2 in both IgG1 and IgG4 subclasses. Thus, 100% (20 of 20) of the microfilaremic (MF(+)) patients bore IgG4 responses to Bm-SPN-2, while only 30% of endemic normal subjects were similarly positive. Following chemotherapy, Bm-SPN-2-specific Abs disappeared in 12 of 13 MF(+) patients, although the majority remained seropositive for whole parasite extract. PBMC from most, but not all, endemic subjects were induced to secrete IFN-gamma when stimulated with Bm-SPN-2. These findings demonstrate that Bm-SPN-2 is recognized by both murine and human T and B cells and indicate that their responses are under relatively stringent temporal control. This study also provides the first example of a stage-specific secreted molecule that acts as a major T cell Ag from filarial parasites and is a prime candidate for a serodiagnostic probe.     

IL-18-induced expression of intercellular adhesion molecule-1 in human monocytes: involvement in IL-12 and IFN-gamma production in PBMC

IL-18 time- and concentration-dependently upregulated the expression of intercellular adhesion molecule-1 (ICAM-1) in a monocyte population in human PBMC as determined by FACS analysis while the expression of CD11a, CD18, CD29, CD44, and CD62L in monocytes and that of ICAM-1, CD11a, CD18, CD29, CD44, and CD62L in T cells was not influenced by IL-18. IL-18 in the same concentration range stimulated the production of IL-12, TNF-alpha, and IFN-gamma in culture of PBMC; however, IL-18-induced expression of ICAM-1 in monocytes was not inhibited by anti-IL-12, anti-TNF-alpha, or anti-IFN-gamma Ab, suggesting the independence of the upregulating effect of IL-18 on endogenous IL-12, TNF-alpha, and IFN-gamma production. IL-18 also induced the aggregation of PBMC, which was prevented by anti-ICAM-1 and anti-LFA-1 Abs. On the other hand, anti-ICAM-1 and anti-LFA-1 Abs inhibited IL-18-induced production of three cytokines, IL-12, IFN-gamma, and TNF-alpha, by 60 and 40%, respectively. These results strongly suggested that the IL-18-induced upregulation of ICAM-1 and the subsequent adhesive interaction through ICAM-1 on monocytes and LFA-1 on T/NK cells generate an additional stimulatory signaling as well as an efficient paracrine environment for the IL-18-initiated cytokine cascade.     

Successful induction of CD8 T cell-dependent protection against malaria by sequential immunization with DNA and recombinant poxvirus of neonatal mice born to immune mothers

In some parts of Africa, 50% of deaths attributed to malaria occur in infants less than 8 mo. Thus, immunization against malaria may have to begin in the neonatal period, when neonates have maternally acquired Abs against malaria parasite proteins. Many malaria vaccines in development rely upon CD8 cells as immune effectors. Some studies indicate that neonates do not mount optimal CD8 cell responses. We report that BALB/c mice first immunized as neonates (7 days) with a Plasmodium yoelii circumsporozoite protein (PyCSP) DNA vaccine mixed with a plasmid expressing murine GM-CSF (DG) and boosted at 28 days with poxvirus expressing PyCSP were protected (93%) as well as mice immunized entirely as adults (70%). Protection was dependent on CD8 cells, and mice had excellent anti-PyCSP IFN-gamma and cytotoxic T lymphocyte responses. Mice born of mothers previously exposed to P. yoelii parasites or immunized with the vaccine were protected and had excellent T cell responses. These data support assessment of this immunization strategy in neonates/young infants in areas in which malaria exacts its greatest toll.     

Chimeras of labile toxin one and cholera toxin retain mucosal adjuvanticity and direct Th cell subsets via their B subunit

Native cholera toxin (nCT) and the heat-labile toxin 1 (nLT) of enterotoxigenic Escherichia coli are AB5-type enterotoxins. Both nCT and nLT are effective adjuvants that promote mucosal and systemic immunity to protein Ags given by either oral or nasal routes. Previous studies have shown that nCT as mucosal adjuvant requires IL-4 and induces CD4-positive (CD4+) Th2-type responses, while nLT up-regulates Th1 cell production of IFN-gamma and IL-4-independent Th2-type responses. To address the relative importance of the A or B subunits in CD4+ Th cell subset responses, chimeras of CT-A/LT-B and LT-A/CT-B were constructed. Mice nasally immunized with CT-A/LT-B or LT-A/CT-B and the weak immunogen OVA developed OVA-specific, plasma IgG Abs titers similar to those induced by either nCT or nLT. Both CT-A/LT-B and LT-A/CT-B promoted secretory IgA anti-OVA Ab, which established their retention of mucosal adjuvant activity. The CT-A/LT-B chimera, like nLT, induced OVA-specific mucosal and peripheral CD4+ T cells secreting IFN-gamma and IL-4-independent Th2-type responses, with plasma IgG2a anti-OVA Abs. Further, LT-A/CT-B, like nCT, promoted plasma IgG1 more than IgG2a and IgE Abs with OVA-specific CD4+ Th2 cells secreting high levels of IL-4, but not IFN-gamma. The LT-A/CT-B chimera and nCT, but not the CT-A/LT-B chimera or nLT, suppressed IL-12R expression and IFN-gamma production by activated T cells. Our results show that the B subunits of enterotoxin adjuvants regulate IL-12R expression and subsequent Th cell subset responses.     

Diversity of the human allergen-specific T cell repertoire associated with distinct skin test reactions: delayed-type hypersensitivity-associated major epitopes induce Th1- and Th2-dominated responses.

Distinct immune responses in humans to the Trichophyton rubrum Ag, Tri r 2, are associated with different patterns of T cell epitope recognition based on in vitro proliferation to peptides derived from this 29-kDa protein. Specifically, the amino-terminal immunodominant epitope, peptide 5 (P5), stimulates strong T cell proliferative responses in subjects with delayed (DTH), but not immediate (IH) hypersensitivity skin tests. Evidence of a role for cytokines or changes in epitope recognition over time was examined in responses to Trichophyton using primary PBMC cultures established from seven IH and seven DTH subjects. Responses stimulated by Tri r 2 were dominated by the Th1 cytokine IFN-gamma (IFN-gamma:IL-5 > or = 4:1) in five DTH subjects, even in the presence of Th2-dominated responses (IFN-gamma:IL-5 < or = 3:1) to a subset of major epitopes. Paradoxically, P5 induced IL-5 and IL-10 production in DTH, but not IH subjects (p = 0.003 (IL-5), p = 0.024 (IL-10)), with no significant difference in IFN-gamma levels between the two groups. In cultures from IH responders, no IL-5 was measurable after stimulation with P6 and P7 (as well as P5); this region of the molecule was shown previously to stimulate markedly reduced T cell proliferation in these individuals. Repeat proliferation assays confirmed no change in the pattern of peptide recognition after > or =20 mo in IH or DTH subjects. We conclude that T cell repertoires associated with distinct immune responses to Tri r 2 can be distinguished based on Th2 cytokine induction by DTH-associated major epitopes localizing to the amino-terminal region of the molecule.     

Cutting edge: mouse IgG1 antibodies comprise two functionally distinct types that are differentially regulated by IL-4 and IL-12.

IL-4-dependent and -independent IgG1 Abs differ in their ability to induce mast cell degranulation as measured by passive cutaneous anaphylaxis (PCA). Mice immunized with OVA or PIII (fraction of Ascaris suum) produced high titers of IgG1 as shown by ELISA and PCA. In contrast, another A. suum fraction, PI, elicited IgG1 Abs with no PCA activity. IgG1 with anaphylactic activity required IL-4, as IgG1 responses to OVA and PIII in IL-4-/- mice gave no PCA. PI-specific IgG1 was IL-4-independent, because no difference was found between the responses of IL-4-/- and IL-4+/+ mice. Significant PCA reactions were elicited, however, with PI-specific IgG1 from IL-12-/- or anti-IFN-gamma Ab-treated mice, although less Ab was measured by ELISA. These results indicate that one type of IgG1 has anaphylactic activity and its synthesis is IL-4-dependent, being inhibited by IL-12 or IFN-gamma; the other lacks this activity and its synthesis is stimulated by IL-12 or IFN-gamma.     

Enhancement of T helper2 response in the absence of interleukin (IL-)6; an inhibition of IL-4-mediated T helper2 cell differentiation by IL-6

Functional roles of interleukin (IL-)6 in T cell response were investigated. Mice deficient in IL-6 and wild mice were immunized with antigens (myelin oligodendrocyte glycoprotein or methylated BSA) and production of IL-4 and interferon (IFN)-gamma by regional lymph nodes was measured. IL-6 deficiency led to an enhancement of IL-4 and an inhibition of IFN-gamma production. Moreover, polyclonal stimulation of spleen T cells from unimmunized IL-6-deficient mice with anti-CD3 plus anti-CD28 antibodies (Abs) demonstrated an enhancement of T helper (Th)(2)responses. The presence of IL-6, however, augmented IL-4 production but it inhibited IFN-gamma expression by spleen T cells in response to polyclonal stimulation and by antigen-primed spleen T cells in response to re-challenge with the antigen. In contrast, the induction of spleen CD4-positive T cells into Th(2)cells in vitro by the anti-CD3 plus IL-4 was completely suppressed by exogenously added IL-6, whereas Th(1)differentiation of T cells by the anti-CD3 plus IL-12 was not inhibited by the presence of IL-6. Thus, these results indicate that IL-6 physiologically could modulate qualitative T cell response and suggest that it augments Th(1)responses partly through its inhibitory capability of IL-4-induced Th(2)differentiation of naive T cells.     

Transmission intensity determines lymphocyte responsiveness and cytokine bias in human lymphatic filariasis

Humans living in areas where filariasis is endemic vary greatly in their exposure to mosquito-borne infective third-stage larvae (L3) of these parasitic helminths. Because the intensity of exposure to Ags affects T cell differentiation and susceptibility to parasitic infections in murine models, we compared T cell and cytokine responses in 97 residents of two villages in Papua New Guinea, where transmission intensity of Wuchereria bancrofti differed by 63-fold (37 vs 2355 L3 per person per year). Residents of the high transmission village had 4- to 11-fold lower proliferation and IFN-gamma responses to filarial Ags, nonparasite Ag, and PHA by PBMC compared with the low transmission village (p < 0.01) even when subjects were matched for intensity of infection. In contrast, filarial Ag-driven IL-5 production was 5.5-fold greater (p < 0.001), and plasma IL-4 and TGF-beta levels were 4-fold and 34% higher, respectively, in residents of the high transmission village. IL-4 and IL-10 responses by PBMC differed little according to village, and increased production of the counterregulatory cytokines IL-10 or TGF-beta by PBMC did not correlate with weak proliferation and IFN-gamma responses. Plasma IL-5, IFN-gamma, and IL-10 levels were similar in the two villages. These data demonstrate that the intensity of exposure to L3 affects lymphocyte responsiveness and cytokine bias possibly by a mechanism that alters APC function.     

Activated T lymphocytes regulate hyaluronan binding to monocyte CD44 via production of IL-2 and IFN-gamma

Interactions of the cell surface proteoglycan CD44 with the extracellular matrix glycosaminoglycan hyaluronan (HA) are important during inflammatory immune responses. Our previous studies indicated that monocyte HA binding could be induced by TNF-alpha. Moreover, monocyte HA binding could be markedly up-regulated by culturing PBMC with anti-CD3 (TCR complex) mAbs. The present study was undertaken to identify soluble factors and/or cell surface molecules of activated T lymphocytes that might regulate HA binding to monocytes. Abs to IL-1 alpha, IL-1 beta, IL-2, IL-3, IL-10, IL-15, GM-CSF, IFN-gamma, and TNF-alpha were tested for their effects on anti-CD3 mAb-, Con A-, and PMA/ionomycin-mediated monocyte HA binding in PBMC cultures. Anti-TNF-alpha, anti-IL-2, and anti-IFN-gamma Abs, when added together to PBMC cultures, completely blocked Con A- and partially blocked anti-CD3- and PMA/ionomycin-induced monocyte HA binding. Furthermore, when added together to PBMC cultures, IL-2 and TNF-alpha induced high levels of monocyte HA binding. Likewise, IFN-gamma augmented TNF-alpha-induced monocyte HA binding. To investigate the role of T cell-monocyte direct contact in induction of monocyte HA binding, we studied PMA/ionomycin-activated, paraformaldehyde-fixed CD4(+) T cells in these assays. Fixed, PMA/ionomycin-activated CD4(+) T lymphocytes induced monocyte HA binding, but direct T cell-monocyte contact was not required. Moreover, anti-IFN-gamma and anti-TNF-alpha Abs blocked fixed PMA/ionomycin-activated CD4(+) T cell-induced monocyte HA binding. Taken together, these studies indicate roles for soluble T lymphocyte-derived factor(s), such as IL-2 and IFN-gamma, and a role for monocyte-derived TNF-alpha in Con A-, TCR complex-, and PMA/ionomycin-induced HA binding to monocyte CD44.     

Adjuvant costimulation during secondary antigen challenge directs qualitative aspects of oral tolerance induction, particularly during the neonatal period

In this report we demonstrate that although passive feeding of specific Ag to mice as neonates or adults can induce oral tolerance in both the cellular and humoral arms of the immune response, quantitative and, in particular, qualitative aspects of the tolerance process are determined by the nature of the inflammatory costimuli provided at the time of secondary Ag challenge. Moreover, this dependency upon nonspecific costimulation is more profound in Ag-fed neonates than in their adult counterparts. Thus, administration of Ag in the Th1-selective adjuvant CFA to prefed animals resulted in significant inhibition of IgG2a, IL-2, and IFN-gamma responses, whereas IL-5 responses were increased. In contrast, rechallenge with Ag in the Th2-selective adjuvant aluminum hydroxide resulted in significant inhibition of IgG1, IgE, IL-2, and IL-5 responses, whereas IFN-gamma responses were increased. Additionally, although soluble Ag challenge of prefed adults revealed marginal tolerogenic effects, the same challenge protocol in animals prefed as neonates elicited enhanced Th2-dependent IgG1 production. These results suggest that inflammatory stimulation at the time of Ag challenge is obligatory to trigger oral tolerance mechanisms, particularly in animals fed as neonates and also that the type of adjuvant used at the time of challenge selects for the type of Th cell population to be inhibited.     

Dendritic cells injected via different routes induce immunity in cancer patients

Dendritic cells (DC) represent potent APCs that are capable of generating tumor-specific immunity. We performed a pilot clinical trial using Ag-pulsed DC as a tumor vaccine. Twenty-one patients with metastatic prostate cancer received two monthly injections of DC enriched and activated from their PBMC. DC were cocultured ex vivo with recombinant mouse prostatic acid phosphatase as the target neoantigen. Following enrichment, DC developed an activated phenotype with up-regulation of CD80, CD86, and CD83 expression. During culture, the DC maintained their levels of various adhesion molecules, including CD44, LFA-1, cutaneous lymphocyte-associated Ag, and CD49d, up-regulated CCR7, but lost CD62 ligand and CCR5. In the absence of CD62 ligand, such cells would not be expected to prime T cells efficiently if administered i.v. due to their inability to access lymphoid tissue via high endothelial venules. To assess this possibility, three patient cohorts were immunized with Ag-pulsed DC by i.v., intradermal (i.d.), or intralymphatic (i.l.) injection. All patients developed Ag-specific T cell immune responses following immunization, regardless of route. Induction of IFN-gamma production, however, was seen only with i.d. and i.l. routes of administration, and no IL-4 responses were seen regardless of route, consistent with the induction of Th1-type immunity. Five of nine patients who were immunized by the i.v. route developed Ag-specific Abs compared with one of six for i.d. and two of six for i.l. routes. These results suggest that while activated DC can prime T cell immunity regardless of route, the quality of this response and induction of Ag-specific Abs may be affected by the route of administration.     

IL-2 receptor blockade inhibits late,but not early,IFN-gamma and CD40 ligand expression in human T cells: disruption of both IL-12-dependent and -independent pathways of IFN-gamma production

mAbs directed against the alpha-chain (Tac/CD25) of the IL-2R are an emerging therapy in both transplantation and autoimmune disease. However, the mechanisms underlying their therapeutic efficacy have not been fully elucidated. Therefore, we examined the effect of IL-2R blockade on Th1 and Th2 cytokine production from human PBMC. Addition of a humanized anti-Tac Ab (HAT) to activated PBMC cultures inhibited IFN-gamma production from CD4 and CD8 T cells by 80-90%. HAT partially inhibited production of TNF-alpha and completely inhibited production of IL-4, IL-5, and IL-10. Furthermore, IL-12, a central regulatory cytokine that induces IFN-gamma, was undetectable in treated cultures. As T cell-dependent induction of IL-12 is regulated via CD40/CD40 ligand (CD40L) interactions, we examined the effect of HAT on CD40L expression. We found CD40L expression to be biphasic with an early (6 h) peak that is CD28/IL-2-independent, but a later peak (48 h) being CD28/IL-2-dependent and inhibited by HAT. Similarly, IFN-gamma production at 6 h was CD28/IL-2-independent but CD28/IL-2-dependent and inhibited by HAT at 48 h. Nonetheless, addition of rCD40L or exogenous IL-12 to HAT-treated cultures could not restore IFN-gamma production. The IFN-gamma deficit in such cultures appears to be due to a direct inhibition by HAT of IL-12-independent IFN-gamma production from T cells rather than altered expression of either the IL-12Rbeta1 or IL-12Rbeta2 chains. These data demonstrate that IL-2 plays a critical role in the regulation of Th1 and Th2 responses and impacts both IL-12-dependent and -independent IFN-gamma production.     

Monophosphoryl lipid A and QS21 increase CD8 T lymphocyte cytotoxicity to herpes simplex virus-2 infected cell proteins 4 and 27 through IFN-gamma and IL-12 production

We have shown previously that IFN-gamma pretreatment of human epidermal cells (ECs) cultured in vitro partially reverses down-regulation of surface MHC class I by HSV infection, allowing recognition by CD8 CTLs, and that HSV immediate early (IE)/early (E) proteins are the predominant targets for CD8 CTLs. In this study of 25 subjects, CD8 CTLs recognized the HSV-2 IE infected cell protein 27 (ICP27) (expressed in autologous IFN-gamma-pretreated, Vaccinia virus recombinant-infected ECs) in all subjects studied, ICP4 in 89%, and ICP0 in 11%. The main hierarchy of recognition was ICP27 > ICP4. ICP27 was the dominant target in 89% of subjects but showed great individual variability in the degree of cytotoxicity. CD8 cytotoxicity specific for HSV-2 IE proteins was enhanced by 48-67% when CD8 CTLs were coincubated with the combination of monophosphoryl lipid A and QS21 adjuvants at the time of Ag presentation. These adjuvants also significantly enhanced IL-12 and IFN-gamma production from nonadherent mononuclear cells stimulated by HSV-2-infected ECs. Addition of IL-12 and IFN-gamma at the time of initial Ag presentation enhanced CD8 cytotoxicity to levels comparable with those stimulated by the adjuvants. Addition of neutralizing Abs to IL-12 or IFN-gamma inhibited CD8 T cell cytotoxicity up to 95% when a combination of the Abs were added at the time of initial Ag presentation. Therefore, the mechanism for the enhancement of CD8 T cell cytotoxicity by adjuvants in this system appears to be via increased levels of IL-12 and IFN-gamma.     

Reduction in the number of UCHL-1+ cells and IL-2 production in the peripheral blood of patients with visceral leishmaniasis.

PBMC from patients with visceral leishmaniasis (VL), before and after successful antimony therapy, were analyzed for their phenotypes and for their ability to produce IL-2 and IFN-gamma and to proliferate against PHA and leishmanial Ag. In agreement with results of earlier studies, PBMC from active VL patients showed a markedly reduced proliferative response and IL-2 and IFN-gamma production, compared with those of healthy controls. The levels of CD4+ and CD8+ T cells were within the normal range, but there was a significant decrease in UCHL-1+ cells (helper-inducer), compared with healthy individuals. The inhibited cellular responses, and lymphokine secretion and decreased level of UCHL-1+ cells in the PBMC of the VL patients returned to the normal range after successful chemotherapy. PBMC from active VL patients were fractionated into adherent cells and nonadherent cells, and the non-adherent were further fractionated into UCHL-1+ and UCHL-1- subpopulations. Results from cell depletion and reconstitution experiments suggest that the IL-2 production by nonadherent cells stimulated with PHA was inhibited by adherent cells, but the IL-2 production by nonadherent cells in response to specific Ag was not. In contrast, UCHL-1- cells seem to mediate the inhibition of Ag-driven IL-2 production by nonadherent cells but not mitogen-stimulated IL-2 secretion by nonadherent cells. Ag-specific IL-2 production principally involves UCHL-1+ cells.     

Mechanisms of the natural reactivity of lymphocytes from noninfected individuals to membrane-associated Leishmania infantum antigens

Membrane-associated Leishmania Ags (MLA) or soluble Leishmania Ags were used in vitro to stimulate cord blood or PBMC from healthy donors noninfected by Leishmania parasites. MLA, but not soluble Leishmania Ags, constantly induce strong proliferation of cord blood mononuclear cells and PBMC from noninfected individuals. Responding cells are CD3+, CD4+, TCRalphabeta+, CD45RO+, and CD45RA+ and secrete IFN-gamma and IL-10, but not IL-4. MLA do not activate NK cells nor NKT cells. Membrane Ags also induce purified macrophages from noninfected individuals to secrete IL-10 and TNF-alpha, but have no effect on IL-1alpha or IL-12 secretion. The effects of MLA are proteinase K-sensitive and resistant to lipid extraction. The lymphoproliferative responses are inhibited by anti-HLA-DR Abs and require Ag processing by APCs, excluding that the biological effect of MLA could be attributed to a superantigen. Finally, TCR repertoire analysis shows that the T cell expansion induced by MLA uses TCR with various variable beta segment rearrangements and CDR3 lengths, features much more characteristic to those observed with a polyclonal activator than with a conventional Ag. These results suggest a particular mechanism developed during the host's natural response to Leishmania parasites that allows direct activation of naive CD4 lymphocytes by parasite membrane-associated Ags.