Tick saliva inhibits dendritic cell migration, maturation, and function while promoting development of Th2 responses

Similarly to other blood-feeding arthropods, ticks have evolved immunosuppressive mechanisms enabling them to overcome the host immune system. Although the immunomodulatory effect of tick saliva on several cell populations of the immune system has been extensively studied, little is known about its impact on dendritic cells (DCs). We have examined the effect of Ixodes ricinus tick saliva on DC function in vitro and in vivo. Exposure of DCs to tick saliva in vitro resulted in impaired maturation, upon CD40 or TLR9, TLR3 and TLR7 ligation, as well as reduced Ag presentation capacity. Administration of tick saliva in vivo significantly inhibited maturation and early migration of DCs from inflamed skin to draining lymph nodes, and decreased the capacity of lymph node DCs to present soluble Ag to specific T cells. Moreover, saliva-exposed DCs failed to induce efficient Th1 and Th17 polarization and promoted development of Th2 responses. Our data reveal a complex inhibitory effect exerted by tick saliva on DC function. Given the role of DCs as the key instigators of adaptive immune responses, alteration of their function might represent a major mechanism of tick-mediated immune evasion.     

Mycobacterium abscessus MAB2560 induces maturation of dendritic cells via Toll-like receptor 4 and drives Th1 immune response

In this study, we showed that Mycobacterium abscessus MAB2560 induces the maturation of dendritic cells (DCs), which are representative antigen-presenting cells (APCs). M. abscessus MAB2560 stimulate the production of pro-inflammatory cytokines [interleukin (IL)-6, tumor necrosis factor (TNF)-α, IL-1β, and IL-12p70] and reduce the endocytic capacity and maturation of DCs. Using TLR4^-/- DCs, we found that MAB2560 mediated DC maturation via Toll-like receptor 4 (TLR4). MAB2560 also activated the MAPK signaling pathway, which was essential for DC maturation. Furthermore, MAB2560-treated DCs induced the transformation of naïve T cells to polarized CD4⁺ and CD8⁺ T cells, which would be crucial for Th1 polarization of the immune response. Taken together, our results indicate that MAB2560 could potentially regulate the host immune response to M. abscessus and may have critical implications for the manipulation of DC functions for developing DC-based immunotherapy. [BMB Reports 2014;47(9): 512-517]     

Silibinin polarizes Th1/Th2 immune responses through the inhibition of immunostimulatory function of dendritic cells

Silibinin is the primary active compound in silymarin. It has been demonstrated to exert anti-carcinogenic effects and hepato-protective effects. However, the effects of silibinin on the maturation and immunostimulatory activities exhibited by dendritic cells (DCs) remain, for the most part, unknown. In this study, we have attempted to determine whether silibinin can influence surface molecule expression, dextran uptake, cytokine production, capacity to induce T-cell differentiation, and the signaling pathways underlying these phenomena in murine bone marrow-derived DCs. Silibinin was shown to significantly suppress the expression of CD80, CD86, MHC class I, and MHC class II in the DCs, and was also associated with impairments of LPS-induced IL-12 expression in the DCs. Silibinin-treated DCs proved highly efficient with regard to Ag capture via mannose receptor-mediated endocytosis. Silibinin also inhibited the LPS-induced activation of MAPKs and the nuclear translocation of the NF-kappaB p65 subunit. Additionally, silibinin-treated DCs evidenced an impaired induction of Th1 response, and a normal cell-mediated immune response. These findings provide new insight into the immunopharmacological functions of silibinin, especially with regard to their impact on the DCs. These findings expand our current understanding of the immunopharmacological functions of silibinin, and may prove useful in the development of therapeutic adjuvants for acute and chronic DC-associated diseases.     

Nitric Oxide Sustains IL-1β Expression in Human Dendritic Cells Enhancing Their Capacity to Induce IL-17–Producing T-Cells

The role played by lung dendritic cells (DCs) which are influenced by external antigens and by their redox state in controlling inflammation is unclear. We studied the role played by nitric oxide (NO) in DC maturation and function. Human DCs were stimulated with a long-acting NO donor, DPTA NONOate, prior to exposure to lipopolysaccharide (LPS). Dose-and time-dependent experiments were performed with DCs with the aim of measuring the release and gene expression of inflammatory cytokines capable of modifying T-cell differentiation, towardsTh1, Th2 and Th17 cells. NO changed the pattern of cytokine release by LPS-matured DCs, dependent on the concentration of NO, as well as on the timing of its addition to the cells during maturation. Addition of NO before LPS-induced maturation strongly inhibited the release of IL-12, while increasing the expression and release of IL-23, IL-1β and IL-6, which are all involved in Th17 polarization. Indeed, DCs treated with NO efficiently induced the release of IL-17 by T-cells through IL-1β. Our work highlights the important role that NO may play in sustaining inflammation during an infection through the preferential differentiation of the Th17 lineage.     

Novel and detrimental effects of lipopolysaccharide on in vitro generation of immature dendritic cells: involvement of mitogen-activated protein kinase p38

Dendritic cells (DCs) are recognized as major players in the regulation of immune responses to a variety of Ags, including bacterial agents. LPS, a Gram-negative bacterial cell wall component, has been shown to fully activate DCs both in vitro and in vivo. LPS-induced DC maturation involves activation of p38, extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinases, and NF-kappaB. Blocking p38 inhibits LPS-induced maturation of DCs. In this study we investigated the role of LPS in the in vitro generation of immature DCs. We report here that in contrast to the observed beneficial effects on DCs, the presence of LPS in monocyte culture retarded the generation of immature DCs. LPS not only impaired the morphology and reduced the yields of the cultured cells, but also inhibited the up-regulation of surface expression of CD1a, costimulatory and adhesion molecules. Furthermore, LPS up-regulated the secretion of IL-1beta, IL-6, IL-8, IL-10, and TNF-alpha; reduced Ag presentation capacity; and inhibited phosphorylation of ERK, but activated p38, leading to a reduced NF-kappaB activity in treated cells. Neutralizing Ab against IL-10, but not other cytokines, partially blocked the effects of LPS. Inhibiting p38 (by inhibitor SB203580) restored the morphology, phenotype, and Ag presentation capacity of LPS-treated cells. SB203580 also inhibited LPS-induced production of IL-1beta, IL-10, and TNF-alpha; enhanced IL-12 production; and recovered the activity of ERK and NF-kappaB. Thus, our study reveals that LPS has dual effects on DCs that are biologically important: activating existing DCs to initiate an immune response, and inhibiting the generation of new DCs to limit such a response.     

Loss of Jak2 Selectively Suppresses DC-Mediated Innate Immune Response and Protects Mice from Lethal Dose of LPS-Induced Septic Shock

Given the importance of Jak2 in cell signaling, a critical role for Jak2 in immune cells especially dendritic cells (DCs) has long been proposed. The exact function for Jak2 in DCs, however, remained poorly understood as Jak2 deficiency leads to embryonic lethality. Here we established Jak2 deficiency in adult Cre^+/+Jak2^fl/fl mice by tamoxifen induction. Loss of Jak2 significantly impaired DC development as manifested by reduced BMDC yield, smaller spleen size and reduced percentage of DCs in total splenocytes. Jak2 was also crucial for the capacity of DCs to mediate innate immune response. Jak2^−/− DCs were less potent in response to inflammatory stimuli and showed reduced capacity to secrete proinflammatory cytokines such as TNFα and IL-12. As a result, Jak2^−/− mice were defective for the early clearance of Listeria after infection. However, their potency to mediate adaptive immune response was not affected. Unlike DCs, Jak2^−/− macrophages showed similar capacity secretion of proinflammatory cytokines, suggesting that Jak2 selectively modulates innate immune response in a DC-dependent manner. Consistent with these results, Jak2^−/− mice were remarkably resistant to lethal dose of LPS-induced septic shock, a deadly sepsis characterized by the excessive innate immune response, and adoptive transfer of normal DCs restored their susceptibility to LPS-induced septic shock. Mechanistic studies revealed that Jak2/SATA5 signaling is pivotal for DC development and maturation, while the capacity for DCs secretion of proinflammatory cytokines is regulated by both Jak2/STAT5 and Jak2/STAT6 signaling.     

Comparison of media and serum supplementation for generation of monophosphoryl lipid A/interferon-γ–matured type I dendritic cells for immunotherapy

Background aimsEx vivo–generated monocyte-derived dendritic cells (DCs) matured with monophosphoryl lipid A (MPLA) and interferon-γ (IFN-γ) can be used as cancer immunotherapy. MPLA/IFN-γ DCs induce Th1 T cell responses and have migratory capacity. Different culture regimens have been used for generation of immunotherapeutic DCs, with varying results. In the present study, culture conditions for MPLA/IFN-γ–matured type I DCs were optimized for clinical application.MethodsDCs were generated from monocytes in the clinical grade culture media CellGro DC, AIM V or X-VIVO 15 in the absence or presence of 2% human serum (HS) and matured with the use of MPLA/IFN-γ. DC yield and DC functionality were assessed. DC functionality was determined by means of analysis of cytokines in culture supernatant, migratory capacity, expression of co-stimulatory molecules, T cell stimulatory capacity of DCs and T helper cell (Th) polarization by the DCs.ResultsDCs generated in the presence of 2% HS produced low amounts of pro-inflammatory cytokines and could not migrate irrespective of the medium used. In the absence of HS, MPLA/IFN-γ DCs generated in X-VIVO did not migrate either. MPLA/IFN-γ DCs generated in AIM V have slightly lower capacity to induce Th1 cells than do DCs generated in CellGro or X-VIVO.ConclusionsAddition of HS to different GMP culture media is detrimental for pro-inflammatory DC maturation and migration. In the absence of serum, CellGro is the most optimal medium tested for generation of migratory and Th1-inducing MPLA/IFN-γ DCs for cancer immunotherapy.     

Impaired pro-inflammatory cytokine production and increased Th2-biasing ability of dendritic cells exposed to Taenia excreted/secreted antigens: A critical role for carbohydrates but not for STAT6 signaling

In cysticercosis, a parasitic disease caused by cestodes, the details of early interactions between parasite antigens and innate cells from the host are not well understood. In this study, the role of cestode-conditioned dendritic cells (DCs) in priming Th1 versus Th2 responses to bystander antigen was examined by using CD11c⁺ DCs as antigen-presenting cells and naive CD4⁺ DO11.10 lymphocytes specific to ovalbumin (OVA) as responding cells. No conventional maturation was induced in DCs exposed to Taenia crassiceps excreted/secreted antigens (TcES). The ability of TcES to affect Toll-like receptor (TLR)-mediated maturation and the pro-inflammatory response was analyzed by co-pulsing DCs with TcES and TLR ligands. DCs exposed to TcES blocked TLR4, TLR9 and Toxoplasma soluble antigen-induced phenotypic maturation. TcES-exposed DCs also blocked secretion of pro-inflammatory cytokines and alloreactive T cell proliferation, while preserving IL-10 production. DCs pulsed with TcES + OVA suppressed IFN-γ, whereas they induced greater IL-4 production by CD4⁺ DO11.10 cells. TcES with chemically-altered glycans failed to modulate TLR-mediated activation of DCs and their Th1-inhibitng ability, which was STAT6-independent. Our results reflect the capacity of TcES glyco-antigens to modulate Th1-type and inflammatory responses mediated through DC activation.     

Inhibition of myeloid dendritic cell accessory cell function and induction of T cell anergy by alcohol correlates with decreased IL-12 production

Alcohol consumption inhibits accessory cell function and Ag-specific T cell responses. Myeloid dendritic cells (DCs) coordinate innate immune responses and T cell activation. In this report, we found that in vivo moderate alcohol intake (0.8 g/kg of body weight) in normal volunteers inhibited DC allostimulatory capacity. Furthermore, in vitro alcohol treatment during DC differentiation significantly reduced allostimulatory activity in a MLR using naive CD4(+) T cells, and inhibited tetanus toxoid Ag presentation by DCs. Alcohol-treated DCs showed reduced IL-12, increased IL-10 production, and a decrease in expression of the costimulatory molecules CD80 and CD86. Addition of exogenous IL-12 and IL-2, but not neutralization of IL-10, during MLR ameliorated the reduced allostimulatory capacity of alcohol-treated DCs. Naive CD4(+) T cells primed with alcohol-treated DCs showed decreased IFN-gamma production that was restored by exogenous IL-12, indicating inhibition of Th1 responses. Furthermore, CD4(+) T cells primed with alcohol-treated DCs were hyporesponsive to subsequent stimulation with the same donor-derived normal DCs, suggesting the ability of alcohol-treated DCs to induce T cell anergy. LPS-induced maturation of alcohol-treated immature DCs partially restored the reduced allostimulatory activity, whereas alcohol given only during DC maturation failed to inhibit DC functions, suggesting that alcohol primarily impairs DC differentiation rather than maturation. NFkappaB activation, a marker of DC maturation was not affected by alcohol. Taken together, alcohol both in vitro and in vivo can impair generation of Th1 immune responses via inhibition of DC differentiation and accessory cell function through mechanisms that involve decreased IL-12 induction.     

IL-10 alters DC function via modulation of cell surface molecules resulting in impaired T-cell responses

IL-10 is a potent inhibitor of T-cell activation and has tolerizing effects on these cells. These effects are primarily mediated via modulation of antigen presenting cell function. Here, it is demonstrated that IL-10 completely inhibits LPS-induced DC maturation, resulting in altered DC-T-cell interactions and reduced T-cell responses. IL-10 inhibited LPS-induced upregulation of costimulatory molecules, MHC Class II, and the secretion of IL-12, TNF-alpha, IL-6, and IL-1beta by DCs, although it upregulated the SLAM (CD150) expression at both the mRNA and protein levels. IL-10 pre-treated DC did not respond to subsequent LPS activation and its stimulatory ability for allogeneic and antigen-specific T-cells was severely impaired. Importantly, T-cells derived from co-cultures with Ag-pulsed, IL-10-treated DC were impaired in their responses to subsequent Ag-specific restimulation. Transwell and DC-derived plasma membrane experiments indicated that the capacity of IL-10-treated DC to induce T-cell unresponsiveness results from alterations in the cell surface molecules rather than modulation of cytokine secretion.     

Neonatal B cells suppress innate toll-like receptor immune responses and modulate alloimmunity

It has been known for decades that neonates are susceptible to transplant tolerance, but the immunological mechanisms involved remain to be fully elucidated. Recent evidence indicates that the maturation state of DCs responding to an allograft may have a profound impact on whether immunity or tolerance ensues. Given that TLR activation is a key process leading to DC maturation, we hypothesized that DCs from neonates have defective TLR immune responses. Contrary to our hypothesis, we found that murine neonatal DCs demonstrated enhanced TLR responses in comparison to adult counterparts in vitro. However, we found that neonatal B cells possess unique immunoregulatory functions as they impaired DC responses to TLR activation in an IL-10-dependent fashion. Functionally, we demonstrated that TLR-activated neonatal, but not adult, B cells impaired Th1, but not Th2, T cell alloimmune responses in vitro and in vivo, in models of alloimmune priming and allotransplantation. We conclude that neonatal B cells possess unique immunoregulatory properties that inhibit DC function and modulate alloimmunity in our murine experimental systems.     

Oncostatin M induces dendritic cell maturation and Th1 polarization

Oncostatin M (OSM) is a pleiotropic cytokine and a member of the gp130/IL-6 cytokine family that has been found to be involved in both pro- and anti-inflammatory responses in cell-mediated immunity. Maturation of dendritic cells (DCs) is crucial for initiation of primary immune responses and is regulated by several stimuli. In this study, the role of OSM in the phenotypic and functional maturation of DCs was evaluated in vitro. Stimulation with OSM upregulated the expression of CD80, CD86, MHC class I and MHC class II and reduced the endocytic capacity of immature DCs. Moreover, OSM induced the allogeneic immunostimulatory capacity of DCs by stimulating the production of the Th1-promoting cytokine IL-12. OSM also increased the production of IFN-γ by T cells in mixed-lymphocyte reactions, which would be expected to contribute to the Th1 polarization of the immune response. The expression of surface markers and cytokine production in DCs was mediated by both the MAPK and NF-κB pathways. Taken together, these results indicate that OSM may play a role in innate immunity and in acquired immunity by enhancing DCs maturation and promoting Th1 immune responses.     

Glycans from Fasciola hepatica Modulate the Host Immune Response and TLR-Induced Maturation of Dendritic Cells

Helminths express various carbohydrate-containing glycoconjugates on their surface, and they release glycan-rich excretion/secretion products that can be very important in their life cycles, infection and pathology. Recent evidence suggests that parasite glycoconjugates could play a role in the evasion of the immune response, leading to a modified Th2-polarized immune response that favors parasite survival in the host. Nevertheless, there is limited information about the nature or function of glycans produced by the trematode Fasciola hepatica, the causative agent of fasciolosis. In this paper, we investigate whether glycosylated molecules from F. hepatica participate in the modulation of host immunity. We also focus on dendritic cells, since they are an important target of immune-modulation by helminths, affecting their activity or function. Our results indicate that glycans from F. hepatica promote the production of IL-4 and IL-10, suppressing IFNγ production. During infection, this parasite is able to induce a semi-mature phenotype of DCs expressing low levels of MHCII and secrete IL-10. Furthermore, we show that parasite glycoconjugates mediate the modulation of LPS-induced maturation of DCs since their oxidation restores the capacity of LPS-treated DCs to secrete high levels of the pro-inflammatory cytokines IL-6 and IL-12/23p40 and low levels of the anti-inflammatory cytokine IL-10. Inhibition assays using carbohydrates suggest that the immune-modulation is mediated, at least in part, by the recognition of a mannose specific-CLR that signals by recruiting the phosphatase Php2. The results presented here contribute to the understanding of the role of parasite glycosylated molecules in the modulation of the host immunity and might be useful in the design of vaccines against fasciolosis.     

Folate deficiency affects dendritic cell function and subsequent T helper cell differentiation

Insufficient folate status may be related to the increasing prevalence of immune- or inflammation-related chronic diseases. To investigate the effects of folate on immune regulation, we examined the impact of folate deficiency (FD) on dendritic cell (DC) maturation and function and, thus, T helper (Th) cells differentiation. First, bone marrow-derived DCs (BMDCs) were generated from BALB/c mice bone marrow cells cultured in folate-containing (F-BMDCs) or folate-deficient (FD-BMDCs) medium. FD-BMDC displayed more immature phenotype including reduced levels of major histocompatibility complex class II (MHC II), co-stimulatory molecules and characteristic of higher endocytic activity. FD-BMDC produced less IL-12p70 and proinflammatory cytokines in response to lipopolysaccharide. This aberrant DC maturation due to FD resulted in reduced BMDC-induced Th cell activity and lower IL-2, IFNγ, IL-13 and IL-10 productions. Further in vivo study confirmed significantly lower IFNγ and IL-10 productions by T cells and showed higher splenic naïve Th and lower memory T, effector T and regulatory T cell (Treg) percentages in mice fed with the FD diet for 13 weeks. To investigate the role of DCs on T cell activity, splenic DCs (spDC) from FD mice were cocultured with Th cells. The FD spDC had lower MHC II and CD80 expressions and subsequently impaired DC-induced Th differentiation, shown as decreased cytokine productions. This study demonstrated that folate deficiency impaired DC functions and, thus, Th differentiation and responses, suggesting that folate plays a crucial role in maintaining Th cells homeostasis.     

CD86 and IL-12p70 Are Key Players for T Helper 1 Polarization and Natural Killer Cell Activation by Toll-Like Receptor-Induced Dendritic Cells

Background

Dendritic cells (DCs) determine the activation and polarization of T cells via expression of costimulatory molecules and secretion of cytokines. The function of DCs derived from monocytes ex vivo strongly depends on the composition of the maturation cocktail used.

Methodology/Principal Findings

We analyzed the effect of costimulatory molecule expression and cytokine secretion by DCs on T and natural killer (NK) cell activation by conducting a head-to-head comparison of a Toll-like receptor (TLR) agonist-based cocktail with the standard combination of proinflammatory cytokines or IL-10 alone. We could show that TLR-induced DCs are characterized by a predominance of costimulatory over coinhibitory molecules and by high secretion of IL-12p70, but not IL-10. Functionally, these signals translated into an increase in IFN-γ secreting Th1 cells and a decrease in regulatory T cells. T cell activation and polarization were dependent on IL-12p70 and CD86, but remarkably not on CD80 signaling. By means of IL-12p70 secretion, only TLR-induced DCs activated NK cells.

Conclusions/Significance

TLR-matured DCs are highly suitable for application in immunotherapeutic strategies that rely on strong type 1 polarization and NK cell activation. Their effects particularly depend on high CD86 expression and IL-12p70 secretion.     

The effects of 1,25-dihydroxyvitamin D3 on markers related to the differentiation and maturation of bone marrow-derived dendritic cells from control and obese mice

Vitamin D has been reported to regulate the maturation and function of dendritic cells (DCs). Obesity was shown to be associated with the dysregulation of vitamin D metabolism and malfunction of DCs. We investigated the effects of in vitro 1,25(OH)₂D₃ treatment (0, 1, or 10 nM) on phenotype and expression of genes related to function of bone marrow-derived DCs (BMDCs) from control and obese mice. C57BL/6 N mice were fed a control or high-fat (10% or 45% kcal fat: CON or HFD) diets for 15 weeks. Differentiation toward DCs was induced with GM-CSF (20 ng/ml) and maturation was induced by LPS (50 ng/ml); 10 nM 1,25(OH)₂D₃ treatment inhibited BMDC differentiation (CD11c⁺) and decreased the percentage of mature DCs (MHCII^highCD11c⁺ and CD86^highCD11c⁺) in both CON and HFD groups. The Il10 expression in stimulated BMDCs from the CON group increased with the 10 nM 1,25(OH)₂D₃ treatment, but not in those from the HFD group. The Il12b mRNA levels in stimulated BMDCs were lower in the HFD group than in the CON group. In conclusion, lower levels of Cd 40, Cd83 and Il12 mRNA in LPS-stimulated BMDCs from obese mice suggest malfunction of DCs as antigen presenting cells. 1,25(OH)₂D₃ treatment inhibited the differentiation and maturation of BMDCs in both control and obese mice. Differential effects of 1,25(OH)₂D₃ on the expression of Il10 between control and obese mice suggest that regulation of immune response by vitamin D could be influenced by obesity.     

Stimulation of dopamine receptor D5 expressed on dendritic cells potentiates Th17-mediated immunity

Dendritic cells (DCs) are responsible for priming T cells and for promoting their differentiation from naive T cells into appropriate effector cells. Emerging evidence suggests that neurotransmitters can modulate T cell-mediated immunity. However, the involvement of specific neurotransmitters or receptors remains poorly understood. In this study, we analyzed the role of dopamine in the regulation of DC function. We found that DCs express dopamine receptors as well as the machinery necessary to synthesize, store, and degrade dopamine. Notably, the expression of D5R decreased upon LPS-induced DC maturation. Deficiency of D5R on the surface of DCs impaired LPS-induced IL-23 and IL-12 production and consequently attenuated the activation and proliferation of Ag-specific CD4(+) T cells. To determine the relevance of D5R expressed on DCs in vivo, we studied the role of this receptor in the modulation of a CD4(+) T cell-driven autoimmunity model. Importantly, D5R-deficient DCs prophylactically transferred into wild-type recipients were able to reduce the severity of experimental autoimmune encephalomyelitis. Furthermore, mice transferred with D5R-deficient DCs displayed a significant reduction in the percentage of Th17 cells infiltrating the CNS without differences in the percentage of Th1 cells compared with animals transferred with wild-type DCs. Our findings demonstrate that by contributing to CD4(+) T cell activation and differentiation to Th17 phenotype, D5R expressed on DCs is able to modulate the development of an autoimmune response in vivo.     

Conjugated linoleic acid suppresses dendritic cell activation and subsequent Th17 responses

PUFAs (polyunsaturated fatty acids) can modify immune responses, so they may have potential therapeutic effects in inflammatory disorders. We previously demonstrated that the cis-9, trans-11 isomer of the PUFA conjugated linoleic acid (CLA) can modulate dendritic cell (DC) cytokine production. Since DCs play a central role in initiating inflammation by directing T helper (Th) cell differentiation, here we examined the effects of CLA on DC maturation and migration and the subsequent generation of Th cell responses. We examined the effect of CLA in vitro on the function of lipopolysaccharide (LPS)-activated bone marrow-derived DCs and ex vivo using cells from mice with high levels of CLA in their diet. We report that CLA inhibits DC migration and modulates TLR-induced production of key cytokines involved in Th cell differentiation both in vitro and in vivo. These changes were accompanied by a significant decrease in expression of MHCII, CD80 and CD86 on the DC surface. Exposure of DCs to CLA suppressed their ability to promote differentiation of naïve T cells into Th1 and/or Th17 cells in vitro and following their adoptive transfer in vivo. Furthermore, in a murine model of endotoxic shock, treatment with CLA suppressed LPS-induced induction of circulating IFN-γ, IL-12p40 and IL-1β. This is the first study to demonstrate that exposure of antigen-presenting cells to CLA can modulate the subsequent Th cell response, and the findings may explain some of the beneficial effects of c9, t11-CLA in inflammatory diseases mediated by Th1 and Th17 cells.     

Dendritic cells engineered to express CD40L continuously produce IL12 and resist negative signals from Tr1/Th3 dominated tumors

TNFα-matured dendritic cells (DCs) pulsed with tumor antigens are being evaluated as cancer vaccines. It has been shown that DCs produce IL12 during a limited time span and subsequently enter a stage of IL12 exhaustion. If DCs are generated ex vivo, the patient could receive IL12-exhausted DCs which may be detrimental for stimulating anti-tumor Th1 responses. Furthermore, many cancer patients exhibit a cytokine profile skewed toward IL10 and TGFβ. This immunological profile, called the Tr1/Th3 response, is associated with the presence of regulatory T-cells. Tr1/Th3 responses potently inhibit DC maturation, thereby regulating Th1 responses. In the present study, we produced genetically engineered DCs that continuously express Th1-related cytokines such as IL12, and resist negative signals from Tr1/Th3-dominated bladder carcinoma cells. Human immature DCs were genetically engineered by adenoviral vectors to express CD40L, or were treated with TNFα as a positive control for maturation. The expression of different Th1/Th3 and inflammatory cytokines was monitored. IL12 and IFNγ were expressed by CD40L-engineered DCs, while TNFα-matured DCs lacked IFNγ and exhibited low IL12 expression. The addition of recombinant IL10 to genetically engineered DCs did not abolish their Th1 profile. Likewise, coculture with tumor cell lines expressing TGFβ with or without recombinant IL10 did not revert to the engineered DCs. We further demonstrate that the resistance of CD40L-expressing DCs to TGFβ and IL10 may be due to decreased levels of TGFβ and IL10 receptors. Thus, CD40L-engineered DCs are robust Th1-promoting ones that are resistant to Tr1/Th3-signaling via IL10 and TGFβ.     

Dendritic cell synthesis of C3 is required for full T cell activation and development of a Th1 phenotype

Previous studies have found that deficiency of complement component C3 is associated with reduced T cell responses in several disease models including viral infection, autoimmune disease, and transplantation. However, the underlying mechanism is unclear. In this study, we demonstrate that dendritic cells (DCs) are able to synthesize C3 and this synthesis is required for the capacity of DCs to stimulate alloreactive T cell responses in vitro and in vivo. Compared with C3-producing DCs, C3-nonproducing DCs exhibit reduced potency to stimulate an alloreactive T cell response, favor the polarization of CD4(+) T cells toward Th2 phenotype, and have regulatory T cell-driving capacity. In addition, priming mice with C3-deficient DCs compared with wild-type DCs led to delayed skin allograft rejection. Our findings that nonproduction of C3 by DCs significantly reduced T cell stimulation and impaired allograft rejection provide a potentially important explanation of how C3-deficient mice develop reduced T cell responses and of how C3-deficient donor kidney is protected from T cell-mediated graft rejection.