Analysis of soluble and cell surface factors regulating anti-DNA topoisomerase I autoantibody production demonstrates synergy between Th1 and Th2 autoreactive T cells

The cellular and subcellular events governing Ab production with specificity for self Ags are poorly understood. In this study we examined the role of cellular interactions and cytokines in regulating the production of anti-DNA topoisomerase I (topo I) Ab, a major autoantibody in patients with systemic sclerosis (SSc). Topo I-specific T cell clones derived from SSc subjects and healthy donors were cultured with autologous peripheral blood B cells. Anti-topo I Ab production was induced by five of seven topo I-specific T cell clones derived from SSc subjects, but by none of eight T cell clones generated from healthy controls. However, two of the T cell clones from healthy controls provided help to HLA-DR-matched SSc B cells to produce anti-topo I Ab. The analysis of cytokine mRNA expression revealed that the ability to promote anti-topo I autoantibody production was strictly correlated with IL-2 and IL-6 expression by the T cell clones. Kinetic studies showed that IL-2 was required throughout the culture period for maximal autoantibody production and that both MHC-TCR and CD40-CD40L interactions were essential during the early phase of the culture. IL-6 was important in the late phase. Th1 clones (producing IL-2, but no IL-6) and Th2 clones (producing IL-6, but no IL-2) synergically activated autologous B cells to produce anti-topo I Ab. These results indicate that T cell-dependent B cell activation resulting in anti-topo I autoantibody production requires a series of temporally defined cell contact and soluble stimuli.     

Differential T cell-mediated regulation of CD23 (Fc epsilonRII) in B cells and follicular dendritic cells

Differences in murine follicular dendritic cells (FDC)-CD23 expression under Th1 vs Th2 conditions prompted the hypothesis that T cells help regulate the phenotype of FDCs. FDCs express CD40, suggesting that T cell-CD40L and lymphokines may be involved in regulating FDC-CD23. To test this, highly enriched FDCs were incubated with CD40L trimer or anti-CD40 to mimic T cell signaling in the presence of IFN-gamma or IL-4. Surface expression of CD23 was determined by flow cytometry, whereas mRNA levels of CD23 and its isoforms CD23a and CD23b were independently measured by quantitative PCR. When FDCs were incubated with either CD40L trimer or agonistic anti-CD40 Ab, the expression of FDC-CD23 was increased both at the mRNA and protein levels. Moreover, engagement of FDC-CD40 enhanced mRNA levels for both CD23a and CD23b isoforms. In addition, IFN-gamma substantially enhanced CD23a and CD23b mRNA levels in CD40-stimulated FDCs. Curiously, IL-4 could also up-regulate FDC-CD23a but not -CD23b. Anti-IFN-gamma dramatically inhibited FDC-CD23 in mice immunized with CFA, whereas anti-IL-4 had only a modest inhibitory effect. In contrast with FDCs, IFN-gamma inhibited surface expression of murine B cell-CD23 as well as mRNA for B cell CD23a and -CD23b, whereas IL-4 dramatically enhanced message for both isoforms as well as protein expression. In short, CD23 was regulated very differently in FDCs and B cells. Previous studies suggest that high levels of FDC-CD23 inhibit IgE production, and this IFN-gamma and CD40L-mediated up-regulation of FDC-CD23 may explain, at least in part, why Th1 responses are associated with low IgE responses in vivo.     

Paradoxical effect of reduced costimulation in T cell-mediated colitis

B7-1 and B7-2 play different roles in the pathogenesis of autoimmunity, but this is controversial. We analyzed colitis induced by transfer of CD45RB(high)CD4(+) T cells to RAG(-/-) recipients lacking B7-1 and/or B7-2. Surprisingly, disease was greatly accelerated in RAG(-/-) recipients deficient for either B7-1 or B7-2, especially in the B7-2(-/-) recipients. This accelerated colitis induction correlated with increased T cell division in vivo and production of Th1 cytokines. Although colitis pathogenesis following T cell transfer was inhibited in the absence of CD40L expression, CD40-CD40L interactions were not required in the B7-2(-/-) RAG(-/-) recipients. In vitro priming by APCs lacking either B7-1 or B7-2 caused decreased IL-2 production, which led to decreased CTLA-4 expression, although T cells primed in this way could respond vigorously upon restimulation by producing increased IL-2 and proinflammatory cytokines. Consistent with this mechanism, we demonstrate that blocking IL-2 early after T cell transfer accelerated colitis. Our data therefore outline a mechanism whereby synergistic costimulation by B7-1 and B7-2 molecules during priming is required for optimal IL-2 production. The consequent inhibitory effect of full CTLA-4 expression, induced by IL-2, may slow colitis, even in the absence of regulatory T cells.     

Differential requirement for the CD40-CD154 costimulatory pathway during Th cell priming by CD8 alpha+ and CD8 alpha- murine dendritic cell subsets

Dendritic cells (DCs) regulate the development of distinct Th populations and thereby provoke appropriate immune responses to various kinds of Ags. In the present work, we investigated the role CD40-CD154 interactions play during the process of Th cell priming by CD8 alpha(+) and CD8 alpha(-) murine DC subsets, which have been reported to differently regulate the Th response. Adoptive transfer of Ag-pulsed CD8 alpha(+) DCs induced a Th1 response and the production of IgG2a Abs, whereas transfer of CD8 alpha(-) DCs induced Th2 cells and IgE Abs in vivo. Induction of distinct Th populations by each DC subset was also confirmed in vitro. Although interruption of CD80/CD86-CD28 interactions inhibited Th cell priming by both DC subsets, disruption of CD40-CD154 interactions only inhibited the induction of the Th1 response by CD8 alpha(+) DCs in vivo. CD40-CD154 interactions were not required for the proliferation of Ag-specific naive Th cells stimulated by either DC subset, but were indispensable in the production of IL-12 from CD8 alpha(+) DCs and their induction of Th1 cells in vitro. Taken together, in our immunization model of Ag-pulsed DC transfer, CD40-CD154 interactions play an important role in the development of CD8 alpha(+) DC-driven Th1 responses but not CD8 alpha(-) DC-driven Th2 responses to protein Ags.     

Th2-dependent B cell responses in the absence of CD40-CD40 ligand interactions

CD40 is thought to play a central role in T cell-dependent humoral responses through two distinct mechanisms. CD4+ T helper cells are activated via CD40-dependent Ag presentation in which CD80/CD86 provides costimulation through CD28. In addition, engagement of CD40 on B cells provides a direct pathway for activation of humoral responses. We used a model of adenovirus-mediated gene transfer of beta-galactosidase (lacZ) into murine lung to evaluate the specific CD40-dependent pathways required for humoral immunity at mucosal surfaces of the lung. Animals deficient in CD40L failed to develop T and B cell responses to vector. Activation of Th2 cells, which normally requires CD40-dependent stimulation of APCs, was selectively reconstituted in CD40 ligand-deficient mice by systemic administration of an Ab that is agonistic to CD28. Surprisingly, this resulted in the development of a functional humoral response to vector as evidenced by formation of germinal centers and production of antiadenovirus IgG1 and IgA that neutralized and prevented effective readministration of vector. The CD28-dependent B cell response required CD4+ T cells and was mediated via IL-4. These studies indicate that CD40 signals to the B cells are not necessary for CD4+ Th2 cell-dependent humoral responses to be generated.     

Enhanced IgA class switching in marginal zone and B1 B cells relative to follicular/B2 B cells

Mouse splenic marginal zone (MZ) B cells and B1 B cells enriched in the peritoneal cavity respond preferentially to T cell-independent Ags compared with follicular (FO)/B2 B cells. Despite the differential responses of B cell subsets to various stimuli, and despite the need for multiple stimuli to induce IgA class switching, the relative contribution of B cell subpopulations to IgA production is unknown. By culturing purified B cell populations, we find that MZ and peritoneal B1 cells switch more readily to IgA than do splenic FO or peritoneal B2 cells in BLyS/LPS/TGF-beta. Addition of IL-4, IL-5, and anti-IgD dextran to the cultures enhances IgA switching in FO/B2 and MZ B cells to a similar frequency, but this treatment suppresses IgA class switching in B1 cells. Thus, IgA switching differs among purified B cell subsets, suggesting that individual B cell populations could contribute differentially to IgA expression in vivo, depending on available stimuli.     

CD40-CD40 ligand costimulation is required for generating antiviral CD4 T cell responses but is dispensable for CD8 T cell responses.

This study documents a striking dichotomy between CD4 and CD8 T cells in terms of their requirements for CD40-CD40 ligand (CD40L) costimulation. CD40L-deficient (-/-) mice made potent virus-specific CD8 T cell responses to dominant as well as subdominant epitopes following infection with lymphocytic choriomeningitis virus. In contrast, in the very same mice, virus-specific CD4 T cell responses were severely compromised. There were 10-fold fewer virus-specific CD4 T cells in CD40L-/- mice compared with those in CD40L+/+ mice, and this inhibition was seen for both Th1 (IFN-gamma, IL-2) and Th2 (IL-4) responses. An in vivo functional consequence of this Th cell defect was the inability of CD40L-/- mice to control a chronic lymphocytic choriomeningitis virus infection. This study highlights the importance of CD40-CD40L interactions in generating virus-specific CD4 T cell responses and in resolving chronic viral infection.     

SAP enables T cells to help B cells by a mechanism distinct from Th cell programming or CD40 ligand regulation

Genetic mutations disrupting the function of signaling lymphocytic activation molecule-associated protein (SAP) lead to T cell intrinsic defects in T cell-dependent Ab responses. To better understand how SAP enables Th cells to help B cells, we first assessed whether molecules important for B cell help are dysregulated in SAP-deficient (SAP knockout (KO)) mice. CD40 ligand (CD40L) expression was enhanced on unpolarized SAP KO T cells; however, Th2 polarization returned their CD40L expression to wild-type levels without rescuing their ability to help B cells. CD40L also localized normally to the site of contact between SAP KO T cells and Ag-bearing B cells. Finally, CD40L-deficient Th cells and SAP KO Th cells differed in their abilities to help B cells in vitro. These data argue that Ab defects caused by SAP deficiency do not result from a loss of CD40L regulation or CD40L function on CD4 T cells. SAP KO Th cells additionally displayed normal patterns of migration and expression of ICOS and CXCR5. Global gene expression was remarkably similar in activated SAP KO vs wild-type T cells, prompting us to investigate whether SAP is necessary for "programming" T cells to become B cell helpers. By restricting SAP expression during differentiation, we determined that SAP is not required during the first 5 days of T cell activation/differentiation to generate Th cells capable of helping B cells. Instead, SAP is necessary for very late stages of differentiation or, most likely, for allowing Th cells to communicate during cognate T:B interactions.     

Isotype switching by human B cells is division-associated and regulated by cytokines

Isotype switching by murine B cells follows a pattern whereby the proportion of cells undergoing switching increases with division number and is regulated by cytokines. Here we explored whether human B cells behaved in a similar manner. The effect of IL-4, IL-10, and IL-13, alone or in combination, on Ig isotype switching by highly purified naive human CD40 ligand (CD40L)-activated B cells was measured against division number over various harvest times. Switching to IgG was induced by IL-4 and, to a lesser extent, IL-13 and IL-10. The combination of IL-10 with IL-4, but not IL-13, induced a higher percentage of cells to undergo switching. Isotype switching to IgG by human CD40L-activated naive B cells was found to be linked to the division history of the cells: IgG(+) cells appeared in cultures of B cells stimulated with CD40L and IL-4 after approximately the third cell division, with the majority expressing IgG1, thus revealing a predictable pattern of IgG isotype switching. These results reveal a useful quantitative framework for monitoring the effects of cytokines on proliferation and isotype switching that should prove valuable for screening Ig immunodeficiencies and polymorphisms in the population for a better understanding of the regulation of human humoral immune responses.     

Differential requirements for expression of CD80/86 and CD40 on B cells for T-dependent antibody responses in vivo

The CD80/86-CD28 and CD40-CD40 ligand costimulatory pathways are essential for Th cell-dependent B cell responses that generate high-affinity, class-switched Ab in vivo. Disruption of either costimulatory pathway results in defective in vivo humoral immune responses, but it remains unclear to what extent this is due to deficient activation of Th cells and/or of B cells. To address this issue, we generated mixed chimeras in which CD80/86- or CD40-deficient bone marrow-derived cells coexist with wild-type (WT) cells, thereby providing the functional T cell help and accessory cell functions required for fully competent B cell responses. We were then able to assess the requirement for CD80/86 or CD40 expression on B cells producing class-switched Ig in response to a T-dependent Ag. In CD80/86 WT plus CD80/86 double-knockout mixed chimeras, both WT- and CD80/86-deficient B cells produced IgG1 and IgE responses, indicating that direct signaling by CD80/86 is not essential for efficient B cell activation. In marked contrast, only WT IgG1 and IgE responses were detected in the chimeras containing CD40-deficient cells, demonstrating that CD40 expression on B cells is essential for class switching by those B cells. Thus, while disrupting either the CD80/86-CD28 or the CD40-CD40 ligand costimulatory pathway abrogates T-dependent B cell immune responses, the two pathways are nonredundant and mediated by distinct mechanisms.     

Quantities of interleukin-12p40 in mature CD8alpha negative dendritic cells correlate with strength of TCR signal and determine Th cell development

Strength of T cell antigen receptor (TCR) signaling drives the development of Th1 and Th2 subsets from naive T helper precursors. The quantity of interleukin-12 (IL-12) from antigen presenting cells (APC) is also profoundly involved in Th development. TCR signal strength and IL-12 production from dendritic cells (DCs) are linked by CD40 ligand (CD40L) expression on activated T cells. CD40L on the activated T cells interacts with CD40 on DC, resulting in induction of IL-12 from DCs. However, the subsets of DC in spleen that produce the IL-12 have not been clearly identified. Purification of DC subsets itself may provide maturation signals to immature DCs. Thus, we used non-purified mouse spleen cells to analyze IL-12 producing cells, near to steady states, during the interaction of naive T cells either with or without agonist. Mature CD86highCD8alpha- DCs in spleen mainly produced IL-12p40 after stimulation of high dose agonist. The ratio of CD40L positive T cells and IL-12p40 secreting CD86high DCs correlated with the concentration of agonist and Th1 development. However, anti-IL-12 did not completely inhibit the Th1 development. Altogether, strength of TCR signaling directs Th cell development by regulating CD40L expression on T cells which determines production of IL-12p40 from CD86high CD8alpha- DC via CD40.     

Disruption of CD40-CD40 ligand pathway inhibits the development of intestinal muscle hypercontractility and protective immunity in nematode infection

In our previous studies, we demonstrated that during Trichinella spiralis infection, T helper (Th) 2 cells contribute to the development of intestinal muscle hypercontractility and worm expulsion from the gut via STAT6. In addition, we have linked the altered muscle contractility to the eviction of the parasite and thereby to the host defense. However, the initial events linking infection to the development of muscle hypercontractility are poorly understood. In this study, we examined the contribution of CD40-CD40 ligand (CD40L) interaction in the development of intestinal muscle hypercontractility, in monocyte chemoattractant protein-1 (MCP-1) production, and in the Th2 response in CD40 ligand-deficient (CD40L -/-) mice infected with T. spiralis. Expulsion of intestinal worms was substantially delayed in CD40L -/- mice compared with the wild-type mice after T. spiralis infection. Consistent with delayed worm expulsion, there was a significant attenuation of intestinal muscle contractility in CD40L -/- mice. Infected CD40L -/- mice also exhibited marked impairment in the production of MCP-1, IL-4, IL-13, IgG1, IgE, and mouse mucosal MCP 1 (MMCP-1), and in goblet cell response. These results demonstrate that CD40-CD40 ligand interaction plays an important role in MCP-1 production, Th2 response, intestinal muscle hypercontractility, and worm expulsion in nematode infection. The present data suggest that the early events leading to the generation of Th2 response include CD40-CD40 ligand interaction, which subsequently influences the production of Th2 cytokines, most likely via upregulation of MCP-1.     

B cell antigen presentation promotes Th2 responses and immunopathology during chronic allergic lung disease

Background

The role of B cells in allergic asthma remains undefined. One mechanism by which B cells clearly contribute to allergic disease is via the production of specific immunoglobulin, and especially IgE. Cognate interactions with specific T cells result in T cell help for B cells, resulting in differentiation and immunoglobulin secretion. Proximal to (and required for) T cell-dependent immunoglobulin production, however, is antigen presentation by B cells. While interaction with T cells clearly has implications for B cell function and differentiation, this study investigated the role that B cells have in shaping the T cell response during chronic allergic lung disease.

Methodology/Principal Findings

In these studies, we used a clinically relevant mouse model of chronic allergic lung disease to study the role of B cells and B cell antigen presentation in this disease. In these studies we present several novel findings: 1) Lung B cells from chronically allergen challenged mice up-regulated MHC II and costimulatory molecules CD40, CD80 and CD86. 2) Using in vitro studies, B cells from the lungs of allergen challenged mice could present antigen to T cells, as assessed by T cell proliferation and the preferential production of Th2 cytokines. 3) Following chronic allergen challenge, the levels of Th2 cytokines IL-4 and IL-5 in the lungs and airways were significantly attenuated in B cell −/− mice, relative to controls. 4) B cell driven Th2 responses and mucus hyper secretion in the lungs were dependent upon MHC II expression by B cells.

Conclusions/Significance

Collectively, these results provide evidence for antigen presentation as a novel mechanism by which B cells contribute to chronic allergic disease. These findings give new insight into the mechanisms by which B cells promote asthma and other chronic diseases.     

Cytokine production by mature and immature CD4-CD8- T cells. Alpha beta-T cell receptor+ CD4-CD8- T cells produce IL-4.

This study follows our previous investigation describing the production of four cytokines (IL-2, IL-4, IFN-gamma, and TNF-alpha) by subsets of thymocytes defined by the expression of CD3, 4, 8, and 25. Here we investigate in greater detail subpopulations of CD4-CD8- double negative (DN) thymocytes. First we divided immature CD25-CD4-CD8-CD3- (CD25- triple negative) (TN) thymocytes into CD44+ and CD44- subsets. The CD44+ population includes very immature precursor T cells and produced high titers of IL-2, TNF-alpha, and IFN-gamma upon activation with calcium ionophore and phorbol ester. In contrast, the CD44- subset of CD25- TN thymocytes did not produce any of the cytokines studied under similar activation conditions. This observation indicates that the latter subset, which differentiates spontaneously in vitro into CD4+CD8+, already resembles CD4+CD8+ thymocytes (which do not produce any of the tested cytokines). We also subdivided the more mature CD3+ DN thymocytes into TCR-alpha beta- and TCR-gamma delta-bearing subsets. These cells produced cytokines upon activation with solid phase anti-CD3 mAb. gamma delta TCR+ DN thymocytes produced IL-2, IFN-gamma and TNF-alpha, whereas alpha beta TCR+ DN thymocytes produced IL-4, IFN-gamma, and TNF-alpha but not IL-2. We then studied alpha beta TCR+ DN T cells isolated from the spleen and found a similar cytokine production profile. Furthermore, splenic alpha beta TCR+ DN cells showed a TCR V beta gene expression profile reminiscent of alpha beta TCR+ DN thymocytes (predominant use of V beta 8.2). These observations suggest that at least some alpha beta TCR+ DN splenocytes are derived from alpha beta TCR+ DN thymocytes and also raises the possibility that these cells may play a role in the development of Th2 responses through their production of IL-4.     

Pertussis toxin-induced cytokine differentiation and clonal expansion of T cells is mediated predominantly via costimulation

Pertussis toxin (PTX) has potent immunologic adjuvant activity in vivo and concomitantly enhances both T helper type (Th1) and Th2 cytokine responses. The PTX-induced enhancement of Th1 and Th2 immunity is mediated via the activation of antigen presenting cells (APCs), but the underlying mechanism is not known. Here we asked whether the adjuvant activity of PTX on T cell immunity was mediated by cytokines and/or costimulatory signals. The results show that in vivo blockade of CD28-CD80/86 costimulation essentially abrogated PTX-mediated enhancement of antigen-specific Th1 and Th2 responses. Blockade of CD40L-CD40 interactions was less efficient in inhibiting PTX-mediated enhancement of Th1 and Th2 responses. In contrast, the adjuvant activity of PTX was not mediated via cytokines, because neither Th1 nor Th2 responses were substantially impaired in mice deficient for IL-12, IFN-gamma, IL-4, IL-5, or IL-6. Collectively, the data suggest that PTX mediates its adjuvant effects on T cell cytokine differentiation and clonal expansion via the modulation of costimulatory molecules on APCs. Understanding the costimulatory pathways targeted by PTX could lead to the design of novel adjuvants that selectively induce Th1 or Th2 immunity.     

Colitis induced by enteric bacterial antigen-specific CD4+ T cells requires CD40-CD40 ligand interactions for a sustained increase in mucosal IL-12

C3H/HeJBir is a mouse substrain that is highly susceptible to colitis. Their CD4+ T cells react to Ags of the commensal enteric bacteria, and the latter can mediate colitis when activated by these Ags and transferred to histocompatible scid recipients. In this study, multiple long-term C3H/HeJBir CD4+ T cell (Bir) lines reactive to commensal enteric bacterial Ags have been generated. All these were Ag specific, pauciclonal, and Th1 predominant; most induced colitis uniformly after transfer to scid recipients. Lesions were focal and marked by increased expression of IL-12p40 and IFN-gamma mRNA and protein. Pathogenic Bir T cell lines expressed CD40 ligand (CD40L) when cultured with Ag-pulsed APCs in vitro. Production of IL-12 was also increased in such cultures, an effect that was Ag- and T cell-dependent and required costimulation by CD40, but not by B7. The two Bir T cell lines that did not induce lesions after transfer failed to significantly express CD40L or increase IL-12 when cultured with Ag-pulsed APCs. Administration of anti-CD40L blocked disease expression induced by pathogenic T cells. We conclude that interactions in the colon mucosa between CD40L-expressing Bir Th1 cells with APCs endogenously loaded with commensal bacterial Ags are critical for sustained increases in local IL-12 production and progression to colitis.     

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.     

Vasoactive intestinal peptide and pituitary adenylate cyclase-activating polypeptide stimulate the induction of Th2 responses by up-regulating B7.2 expression.

Vasoactive intestinal peptide (VIP) and the pituitary adenylate cyclase-activating polypeptide (PACAP), two structurally related neuropeptides produced within the lymphoid microenvironment, modulate several immunologic functions. We have recently demonstrated that VIP and PACAP enhance the macrophage costimulatory activity for naive CD4+ T cells exposed to allogeneic or anti-CD3 stimuli through the differential regulation of the B7 costimulatory molecules. In this study, we report on the role of VIP and PACAP on macrophage B7 expression and costimulatory function for Ag-primed CD4+ T cells, and on the macrophage-induced regulation of Th1/Th2 differentiation in vitro and in vivo. VIP and PACAP up-regulate the costimulatory activity of macrophages for Ag-primed CD4+ T cells. VIP-/PACAP-treated macrophages gain the ability to induce Th2-type cytokines such as IL-4 and IL-5 and reduce Th1-type cytokines such as IFN-gamma and IL-2. In vivo administration of VIP or PACAP in Ag-immunized mice reduce the numbers of IFN-gamma-secreting cells and enhance the numbers of IL-4-secreting cells. One of the consequences of the VIP-/PACAP-induced shift in cytokine profile is a change in the Ag-specific Ig isotype, increasing IgG1 and decreasing IgG2a levels. Finally, the preferential differentiation into Th2 effector cells after Ag stimulation induced by VIP-/PACAP-treated macrophages is mediated through the up-regulation of B7.2 expression.     

A recombinant vector expressing transgenes for four T-cell costimulatory molecules (OX40L,B7-1, ICAM-1, LFA-3) induces sustained CD4+ and CD8+ T-cell activation,protection from apoptosis,and enhanced cytokine production

The role of OX40L on the activation of T cells was investigated using poxvirus vectors expressing OX40L alone or in combination with three other T-cell costimulatory molecules: B7-1, ICAM-1, and LFA-3. Poxvirus vector-infected cells were used to stimulate nai;ve or activated CD4(+) and CD8(+) T cells. These studies demonstrate that (a) OX40L plays a role in sustaining the long-term proliferation of CD8(+) T cells in addition to the known effect on CD4(+) T cells following activation, (b) OX40L enhances the production of Th1 cytokines (IL-2, IFN-gamma, and TNF-alpha) from both CD4(+) and CD8(+) while no change in IL-4 expression was observed, and (c) the anti-apoptotic effect of OX40L on T cells is likely the result of sustained expression of anti-apoptotic genes while genes involved in apoptosis are inhibited. In addition, these are the first studies to demonstrate that the combined use of a vector driving the expression of OX40L with three other costimulatory molecules (B7-1, ICAM-1, and LFA-3) both enhances initial activation and then further potentiates sustained activation of nai;ve and effector T cells.