Suppression Of IFN-gamma Research Articles

Cellular and transcriptional impacts of Janus kinase and/or IFN-gamma inhibition in a mouse model of primary hemophagocytic lymphohistiocytosis.

Primary hemophagocytic lymphohistiocytosis (pHLH) is an inherited inflammatory syndrome driven by the exuberant activation of interferon-gamma (IFNg)-producing CD8 T cells. Towards this end, ruxolitinib treatment or IFNg neutralization (aIFNg) lessens immunopathology in a model of pHLH in which perforin-deficient mice (Prf1-/-) are infected with Lymphocytic Choriomeningitis virus (LCMV). However, neither agent completely eradicates inflammation. Two studies combining ruxolitinib with aIFNg report conflicting results with one demonstrating improvement and the other worsening of disease manifestations. As these studies used differing doses of drugs and varying LCMV strains, it remained unclear whether combination therapy is safe and effective. We previously showed that a ruxolitinib dose of 90 mg/kg lessens inflammation in Prf1-/- mice infected with LCMV-Armstrong. To determine whether this dose controls inflammation induced by a different LCMV strain, we administered ruxolitinib at 90mg/kg to Prf1-/- mice infected with LCMV-WE. To elucidate the impacts of single agent versus combination therapy, Prf1-/- animals were infected with LCMV, treated or not with ruxolitinib, aIFNg or both agents, and analyzed for disease features and the transcriptional impacts of therapy within purified CD8 T cells. Ruxolitinib is well-tolerated and controls disease regardless of the viral strain used. aIFNg, administered alone or with ruxolitinib, is most effective at reversing anemia and reducing serum IFNg levels. In contrast, ruxolitinib appears better than aIFNg, and equally or more effective than combination therapy, at lessening immune cell expansion and cytokine production. Each treatment targets distinct gene expression pathways with aIFNg downregulating IFNg, IFNa, and IL-6-STAT3 pathways, and ruxolitinib downregulating IL-6-STAT3, glycolysis, and reactive oxygen species pathways. Unexpectedly, combination therapy is associated with upregulation of genes driving cell survival and proliferation. Ruxolitinib is tolerated and curtails inflammation regardless of the inciting viral strain and whether it is given alone or in combination with aIFNg. When administered at the doses used in this study, the combination of ruxolitinb and aIFNg appears no better than treatment with either drug alone in lessening inflammation. Further studies are warranted to elucidate the optimal doses, schedules, and combinations of these agents for the treatment of patients with pHLH.

Augmentation of IFN-γ by bone marrow derived immune cells in the presence of severe suppression of IFN-γ in gingivae induced by zoledronic acid and denosumab in Hu-BLT mice model of ONJ.

The potential mechanisms governing drug induced osteonecrosis of the jaw (ONJ) is not well understood, and is one of the objectives of this study. Thus, we tested the release of IFN-γ within different immune compartments including bone marrow and gingivae upon treatment with zoledronic acid (ZOL) and denosumab which are known to induce ONJ in susceptible individuals. We used humanized-BLT mouse model for the in-vivo studies reported in this paper. To determine the effects of zoledronic acid and denosumab on IFN-γ secretion and NK cell-mediated cytotoxicity; peripheral blood, bone marrow, spleen and gingiva were obtained after the injection of ZOL and denosumab in mice. Percentages of B cells are much higher in wild-type mice whereas the proportions of immune subsets in humans and reconstituted hu-BLT peripheral-blood are similar. Therefore, hu-BLT mice are preferable model to study human disease, in particular, immune-pathologies induced by ZOL and denosumab. Both agents resulted in a severe suppression of IFN-γ in the gingiva, whereas they heightened the release of IFN-γ and NK cell-mediated cytotoxicity by the BM-derived immune cells. ZOL increased the IFN-γ secretion by the spleen and peripheral blood immune cells, whereas denosumab decreased the release IFN-γ by these cells significantly. ZOL and denosumab may likely suppress IFN-γ secretion in gingiva through different mechanisms. In addition, to the suppression of IFN-γ secretion, denosumab mediated effect could in part be due to the decrease in the bone resorptive function of osteoclasts due to the induction of antibody dependent cellular cytotoxicity and lysis of osteoclasts, whereas ZOL is able to mediate cell death of osteoclasts directly. Suppression of IFN-gamma in gingiva is largely responsible for the inhibition of immune cell function, leading to dysregulated osteoblastic and osteoclastic activities. Restoration of IFN-gamma in the local microenvironment may result in establishment of homeostatic balance in the gingiva and prevention of osteonecrosis of jaw.

IFNAR signaling augments T cell and plasmacytoid dendritic cell activation during costimulation blockade resistant rejection

Abstract Costimulation blockade (CoB) is a promising new transplant immunosuppression strategy offering improved long-term allograft survival without the nephrotoxicity of calcineurin inhibitors. However, increased risks of T cell mediated acute rejection have impeded the widespread adoption of CoB. Type I interferon (IFN) produced predominantly by plasmacytoid dendritic cells (pDCs) in response to innate immune signals, induce systemic inflammation that may prime the adaptive immune system for acute rejection. Here we investigate how signaling through the type I interferon receptor (IFNAR) contributes to CoB-resistant allograft rejection. We performed fully MHC-mismatched skin grafts from Balb/cJ donors to C57BL/6 recipients. Mice were treated with CoB alone (250 μg CTLA4-Ig + 250 μg anti-CD154 given by IP injection day 0, 2, 4, 6) or in conjunction with anti-IFNAR (250 μg given day 0, 2, 4, 6). CoB treatment improved graft survival but resulted in CoB-resistant rejection, while CoB+anti-IFNAR significantly improved graft survival. We found that CoB+anti-IFNAR led to a reduction in the activation and effector function of CD8+ T cells compared to CoB alone, demonstrated by reduced frequencies of CD44+CD8+ T cells and an inhibition of IFN-gamma and TNF-alpha production by CD8+ T cells. In addition, we observed reduced CD80 and MHC I expression on conventional DCs (cDCs) and pDCs treated with CoB+anti-IFNAR compared to treatment with CoB alone. These data suggest that signaling through IFNAR augments cDC and pDC activation and is associated with CD8+ T cell activation and effector functions in order to promote CoB-resistant allograft rejection.

Low Dose IL-2 and Rapamycin Leads to Prolongation of Human Islet Allograft Survival by Inhibition of IFN-gamma+ T cells and Expansion of FOXP3+CD25+ Tregs in a Humanized Islet Transplant Mouse Model

Low Dose IL-2 and Rapamycin Leads to Prolongation of Human Islet Allograft Survival by Inhibition of IFN-gamma+ T cells and Expansion of FOXP3+CD25+ Tregs in a Humanized Islet Transplant Mouse Model

Effect of azithromycin on natural killer cell function

Azithromycin (AZM), a macrolide antibiotic for treating mycoplasma infections, may exhibit anti-inflammatory activity aside from its antimicrobial effect, providing additional therapeutic benefit. Natural killer (NK) cells, a first-line innate immune defense against microbial invasions, paradoxically exert a detrimental effect in protecting mycoplasma infection. Little was known regarding the effect of AZM on NK cells. In the present study, we investigated the ability of azithromycin to influence natural killer (NK) cell function with regard to activation, apoptosis and cytotoxic function. AZM had little effect on NK receptor expression and cytotoxic function of NK-92 cells. However, AZM did show a dose-dependent suppression on IL-15-induced CD69 expression of primary NK cells. AZM inhibited the cytotoxicity against K562 cells of resting and IL-15 activated primary NK cells possibly through down-regulation of perforin expression, especially on CD16+CD56+ NK subsets. AZM exerted a dose-dependent inhibition of IFN-gamma and TNF-alpha production from NK-92 cells, but did not affect the cytokine production of IL-15 activated primary NK cells. Taken together, AZM down-regulates NK cytotoxicity and cytokine production and may provide therapeutic benefits aside from its antimicrobial activity.

Role of 4-1BB Receptor in the Control Played by CD8+ T Cells on IFN-γ Production by Mycobacterium tuberculosis Antigen-Specific CD4+ T Cells

BackgroundAntigen-specific IFN-γ producing CD4+ T cells are the main mediators of protection against Mycobacterium tuberculosis infection both under natural conditions and following vaccination. However these cells are responsible for lung damage and poor vaccine efficacy when not tightly controlled. Discovering new tools to control nonprotective antigen-specific IFN-γ production without affecting protective IFN-γ is a challenge in tuberculosis research.Methods and FindingsImmunization with DNA encoding Ag85B, a candidate vaccine antigen of Mycobacterium tuberculosis, elicited in mice a low but protective CD4+ T cell-mediated IFN-γ response, while in mice primed with DNA and boosted with Ag85B protein a massive increase in IFN-γ response was associated with loss of protection. Both protective and non-protective Ag85B-immunization generated antigen-specific CD8+ T cells which suppressed IFN-γ-secreting CD4+ T cells. However, ex vivo ligation of 4-1BB, a member of TNF-receptor super-family, reduced the massive, non-protective IFN-γ responses by CD4+ T cells in protein-boosted mice without affecting the low protective IFN-γ-secretion in mice immunized with DNA. This selective inhibition was due to the induction of 4-1BB exclusively on CD8+ T cells of DNA-primed and protein-boosted mice following Ag85B protein stimulation. The 4-1BB-mediated IFN-γ inhibition did not require soluble IL-10, TGF-β, XCL-1 and MIP-1β. In vivo Ag85B stimulation induced 4-1BB expression on CD8+ T cells and in vivo 4-1BB ligation reduced the activation, IFN-γ production and expansion of Ag85B-specific CD4+ T cells of DNA-primed and protein-boosted mice.Conclusion/SignificanceAntigen-specific suppressor CD8+ T cells are elicited through immunization with the mycobacterial antigen Ag85B. Ligation of 4-1BB receptor further enhanced their suppressive activity on IFN-γ-secreting CD4+ T cells. The selective expression of 4-1BB only on CD8+ T cells in mice developing a massive, non-protective IFN-γ response opens novel strategies for intervention in tuberculosis pathology and vaccination through T-cell co-stimulatory-based molecular targeting.

Differential dose effects of recombinant IL-25 on the development of dextran sulfate sodium-induced colitis

ObjectiveWe evaluated different dose effects of rIL-25 on acute ulcerative colitis.Materials and methodsMice were fed 2.5% dextran sulfate sodium (DSS) for 5 days while infused i.p. with repeated doses of rIL-25 (0.2, 0.4 and 0.8 μg) in PBS or PBS only after every 24 h at the same time as the start of the DSS exposure. Clinical, macroscopical and microscopical assessment of colitis severity with survival study was performed. Colonic IL-25 expression and production of IFN-γ, IL-10 and IL-4 was also analyzed.ResultsAt a dose of 0.2 μg, colitis was aggravated with high mortality, better improvements were observed at a dose of 0.4 μg, and colitis-induced diarrhea was reversed at a dose of 0.8 μg. The expression of IL-25 was found to decrease in severe colitis. Moreover, IL-25 inhibited production of mucosal IFN-γ, induced increase in IL-10 but not IL-4.ConclusionImprovements in DSS-induced colitis in response to IL-25 suggest IL-25's protective role by mechanisms including inhibition of IFN-γ with enhancement of anti-inflammatory release.

Cellular and molecular basis for the regulation of inflammation by TGF-

Transforming growth factor-beta (TGF-beta) has been shown to play an essential role in the suppression of inflammation, yet recent studies have revealed the positive roles of TGF-beta in inflammatory responses. For example, TGF-beta induces Foxp3-positive regulatory T cells (iTregs) in the presence of interleukin-2 (IL-2), while in the presence of IL-6, it induces pathogenic IL-17 producing Th17 cells. TGF-beta inhibits the proliferation of immune cells as well as cytokine production via Foxp3-dependent and -independent mechanisms. Little is known about molecular mechanisms involved in immune suppression via TGF-beta; however, Smad2/3 have been shown to play essential roles in Foxp3 induction as well as in IL-2 and IFN-gamma suppression, whereas Th17 differentiation is promoted via the Smad-independent pathway. Interaction between TGF-beta and other cytokine signaling is important in establishing the balance of immunity and tolerance.

TGF-β activity protects against inflammatory aortic aneurysm progression and complications in angiotensin II–infused mice

Complicated abdominal aortic aneurysm (AAA) is a major cause of mortality in elderly men. Ang II-dependent TGF-beta activity promotes aortic aneurysm progression in experimental Marfan syndrome. However, the role of TGF-beta in experimental models of AAA has not been comprehensively assessed. Here, we show that systemic neutralization of TGF-beta activity breaks the resistance of normocholesterolemic C57BL/6 mice to Ang II-induced AAA formation and markedly increases their susceptibility to the disease. These aneurysms displayed a large spectrum of complications on echography, including fissuration, double channel formation, and rupture, leading to death from aneurysm complications. The disease was refractory to inhibition of IFN-gamma, IL-4, IL-6, or TNF-alpha signaling. Genetic deletion of T and B cells or inhibition of the CX3CR1 pathway resulted in partial protection. Interestingly, neutralization of TGF-beta activity enhanced monocyte invasiveness, and monocyte depletion markedly inhibited aneurysm progression and complications. Finally, TGF-beta neutralization increased MMP-12 activity, and MMP-12 deficiency prevented aneurysm rupture. These results clearly identify a critical role for TGF-beta in the taming of the innate immune response and the preservation of vessel integrity in C57BL/6 mice, which contrasts with its reported pathogenic role in Marfan syndrome.

Human Guanylate Binding Proteins Potentiate the Anti-Chlamydia Effects of Interferon-γ

Chlamydiae are obligate intracellular pathogens that are sensitive to pro-inflammatory cytokine interferon-γ. IFN-γ-inducible murine p47 GTPases have been demonstrated to function in resistance to chlamydia infection in vivo and in vitro. Because the human genome does not encode IFN-γ-inducible homologues of these proteins, the significance of the p47 GTPase findings to chlamydia pathogenesis in humans is unclear. Here we report a pair of IFN-γ-inducible proteins, the human guanylate binding proteins (hGBPs) 1 and 2 that potentiate the anti-chlamydial properties of IFN-γ. hGBP1 and 2 localize to the inclusion membrane, and their anti-chlamydial functions required the GTPase domain. Alone, hGBP1 or 2 have mild, but statistically significant and reproducible negative effects on the growth of Chlamydia trachomatis, whilst having potent anti-chlamydial activity in conjunction with treatment with a sub-inhibitory concentration of IFN-γ. Thus, hGBPs appear to potentiate the anti-chlamydial effects of IFN-γ. Indeed, depletion of hGBP1 and 2 in cells treated with IFN-γ led to an increase in inclusion size, indicative of better growth. Interestingly, chlamydia species/strains harboring the full-length version of the putative cytotoxin gene, which has been suggested to confer resistance to IFN-γ was not affected by hGBP overexpression. These findings identify the guanylate binding proteins as potentiators of IFN-γ inhibition of C. trachomatis growth, and may be the targets of the chlamydial cytotoxin.

Suppression of IFNγ + mycobacterial lipoarabinomannan-induced NO by IL-4 is due to decreased IRF-1 expression

In mice, and possibly in humans, nitric oxide (NO) is an important host-defense molecule against Mycobacterium tuberculosis. Inducible nitric oxide synthase (iNOS) and NO are upregulated in murine macrophages stimulated with interferon-gamma (IFNgamma) and mannose-capped lipoarabinomannan (ManLAM), a major lipoglycan in the cell wall of M. tuberculosis. Interleukin-4 (IL-4) can inhibit NO expression and may impair host immune response to M. tuberculosis. Therefore, we sought to determine the mechanism by which IL-4 inhibits IFNgamma+ManLAM-induced NO production. Since l-arginine is the substrate for both iNOS and arginase, and IL-4 increases arginase activity by inducing its production, a plausible mechanism of IL-4 inhibition of NO expression is via depletion of l-arginine through increased arginase activity. Herein, we show that IL-4 inhibited iNOS gene expression at the transcriptional level, suggesting an inhibitory mechanism that is independent of the competition for l-arginine between iNOS and arginase. Furthermore, pharmacologic inhibition of IL-4-induced arginase activity did not abrogate IL-4 inhibition of IFNgamma+ManLAM-induced NO expression. Instead, inhibition by IL-4 was mediated principally by the ability of IL-4 to inhibit the production of IFNgamma-induced interferon-gamma response factor-1 (IRF-1) protein, a critically important transcriptional element that enhances expression of IFNgamma-inducible genes such as iNOS.

Downregulation of pro‐inflammatory cytokines by lupeol measured using cytometric bead array immunoassay

The objective of the study was to investigate the activity of Lupeol (LUP) on proinflammatory and anti-inflammatory cytokines in the pleural exudate from male swiss albino mice. We applied Cytometric bead array technology for simultaneously measurement of these cytokines in pleurisy induced mice treated with lupeol in graded oral doses. Cytometric bead array uses the sensitivity of amplified fluorescence detection by flowcytometer to measure soluble analytes in a particle based immune assay. This assay can accurately quantitate 5 cytokines in a 50 microlitre sample volume. Oral administration of LUP at doses of 25, 50, 100 and 200 mg/kg p.o. produced dose related inhibition of IL-2, IFN-gamma and TNF-alpha in the pleural exudate with the most significant effect at 100 mg/kg oral dose. LUP had a non significant inhibitory effect on the levels of IL-4 and IL-5.

Phenotypic and Functional Effects of Novel Akt Inhibitor Perifosine on Immune System.

Perifosine is a synthetic novel alkylphospholipid, a new class of antitumor agent which targets cell membranes and inhibits Akt activation. Perifosine inhibits multiple myeloma (MM) cell growth in vitro and in vivo. Currently perifosine is under phase II clinical evaluation in MM. Although perifosine has shown significant direct antitumor effects, its effect on immune system has not yet been clarified. The objective of this study was to investigate the effects of perifosine on immune system including the activity of antigen presenting cells (APCs), T cells and NK cells. Perifosine was used at a clinically relevant concentrations of 2.5 μM, 5 μM and 10 μM. Monocyte-derived dendritic cells (DCs) from healthy donors were used as the APCs. We observed that the perifosine up to 48 hours had no effect on viability (>90%), as assessed by annexin V and PI staining. Alteration of DC phenotype by perifosine was further examined by flow cytometry. Our results demonstrated that perifosine treatment led to a dose-dependent downregulation of surface antigen expression, associated with costimulation (CD40, CD80 and CD86), antigen presentation (HLA-ABC, HLA-DQ) and maturation (CD83) on immature DCs at 24 and 48 hours. The significant downregulation of CD40, CD83, HLA-ABC and HLA-DQ was also observed on mature DCs. Perifosine also inhibited immature DC uptake of antigens (40-kDa Dextran-FITC, 45-kDa protein A-Alexa Fluor 488 and 20-kDa protein G-Alexa Fluor 488) in a dose-dependent manner. Since DCs play a crucial regulatory role via cytokine production, we next determined IL-12p70 and IL-10 secretion by LPS-induced DCs with and without perifosine treatment. Compared to controls, perifosine treatment at 24 hours significantly inhibited LPS-induced-IL-12p70 production by DCs (trt vs. untrt = 166 pg/ml (2.5μM), 111 pg/ml (5μM) and 45 pg/ml (10μM) vs. 192 pg/ml), as well as inhibited IL-10 production (trt vs. untrt = 371 pg/ml (2.5μM), 306 pg/ml (5μM) and 179 pg/ml (10μM) vs. 472 pg/ml). These data suggest that perifosine is able to affect both immature and mature DCs and could contribute to inhibition of DC-mediated immune responses. Furthermore, we evaluated effect of perifosine on T cells obtained from healthy donor peripheral blood mononuclear cells (PBMCs) by negative selection. Although perifosine treatment, up to 10μM, had no effect on T cell survival, it inhibited IFN-gamma production by T cells stimulated with PMA and ionomycin compared to that of the control (intracellular flow cytometry, trt vs. untrt= 11.7% vs. 21.3%). The IFN-gamma inhibition by perifosine treatment (10μM) was further confirmed by ELISA upon stimulation with anti- CD2/CD3/CD28 activation beads (trt vs. untrt =11340 pg/ml vs. 18150 pg/mL). Similarly, viability of NK cells obtained from PBMCs by negative selection was not significantly affected by perifosine; however, decreased cytotoxicities were observed by NK cells treated with perifosine (10μM) against the U266 myeloma cell line (trt vs. untrt=30% vs. 51% (E:T=10:1) and 17% vs 30% (E:T=5:1) respectively). In addition, the IFN-gamma production by IL-2 and IL-12- induced NK cells was significantly inhibited following the perifosine treatment (trt vs. untrt=286 pg/ml (2.5μM), 210 pg/ml (10μM) vs. 439 pg/ml). These studies demonstrate that perifosine treatment significantly affects phenotype and function of human DCs, T cells and NK cells. Our pre-clinical data therefore indicates the need to monitor immune functions in patients under the Akt inhibitor treatment.

The B Subunit ofEscherichia coliHeat-Labile Enterotoxin Inhibits Th1 but Not Th17 Cell Responses in Established Experimental Autoimmune Uveoretinitis

To investigate the efficacy of the B subunit of Escherichia coli heat-labile enterotoxin (EtxB) in the treatment of ocular autoimmune disease. Murine experimental autoimmune uveoretinitis (EAU) is an animal model of autoimmune posterior uveitis initiated by retinal antigen-specific Th1 and Th17 CD4(+) T cells, which activate myeloid cells, inducing retinal damage. EtxB is a potent immune modulator that ameliorates other Th1-mediated autoimmune diseases, enhancing regulatory T-cell activity. EAU was induced in B10.RIII mice by immunization with peptide hIRBP(161-180). Disease severity was measured by clinical and histologic assessment, and functional responses of macrophages (Mphis) and T cells were assessed, both in vivo and in cocultures in vitro. EtxB was administered intranasally daily for 4 days, starting either 3 days before or 3 days after EAU induction. Preimmunization treatment with EtxB protected mice from EAU, limiting both the number and the activation status of retinal infiltrating immune cells. Treatment after EAU induction did not alter the disease course, despite suppression of IFN-gamma. Although EtxB treatment of in vitro cocultures of T cells and Mphis increased IL-10 production, EtxB treatment in vivo increased the proportion and number of IL-17-producing CD4(+) cells infiltrating the eye. EtxB preimmunization protects mice from EAU induction by inhibiting Th1 responses, but the resultant reduction in IFN-gamma responses by EtxB does not effect infiltration or structural damage in established EAU, where Th17 responses predominate. These data highlight the critical importance of the dynamics of T-cell phenotype and infiltration during EAU when considering immunomodulatory therapy.

Children with egg allergy have evidence of reduced neonatal CD4 +CD25 +CD127 lo/− regulatory T cell function

The role of regulatory T (Treg) cells in allergic predisposition is not known. This study compared the frequency and function of cord blood Treg cells from nonallergic children (n = 18) with those from children who have egg allergy (n = 15) in the first year of life. CD4(+) effector T cells and autologous antigen-presenting cells isolated from cord blood mononuclear cells were cocultured with or without CD4(+)CD25(+)CD127(lo/-) Treg cells, and cytokine responses to staphylococcal endotoxin B were assessed after 48 hours. The addition of Treg cell populations to cord blood mononuclear cell cultures resulted in significant reduction in IL-10 (P = .002), IL-13 (P = .012), and IFN-gamma (P < .001) production. Consistent with other reports, effector CD4(+) T-cell responses (IFN-gamma and IL-13) tended to be lower in the allergic group. These neonates showed less significant Treg cell-associated suppression of IFN-gamma (P = .015) compared with that seen in the nonallergic group (P = .001). The allergic group was also less likely (44%) to show Treg cell-associated suppression of IFN-gamma effector responses compared with that seen in the nonallergic group (78%, P = .015). The magnitude of suppression (change in IFN-gamma level when CD4(+)CD25(+)CD127(lo/-) Treg cells were added to responding effector T-cell cultures) was significantly lower in the allergic group (P = .004). There were no between-group differences in the circulating CD4(+)CD25(+)CD127(lo/-) Treg cells (as a percentage of cord blood T cells) or in the FOXP3 expression of these cells. This study confirms the presence and activity of Treg cells in cord blood and provides preliminary evidence of differences in neonates who progress to allergic disease in the first year of life.

HLA-DR Alpha 2 Mediates Negative Signalling via Binding to Tirc7 Leading to Anti-Inflammatory and Apoptotic Effects in Lymphocytes In Vitro and In Vivo

Classically, HLA-DR expressed on antigen presenting cells (APC) initiates lymphocyte activation via presentation of peptides to TCR bearing CD4+ T-Cells. Here we demonstrate that HLA-DR alpha 2 domain (sHLA-DRα2) also induces negative signals by engaging TIRC7 on lymphocytes. This interaction inhibits proliferation and induces apoptosis in CD4+ and CD8+ T-cells via activation of the intrinsic pathway. Proliferation inhibition is associated with SHP-1 recruitment by TIRC7, decreased phosphorylation of STAT4, TCR-ζ chain & ZAP70, and inhibition of IFN-γ and FasL expression. HLA-DRα2 and TIRC7 co-localize at the APC-T cell interaction site. Triggering HLA-DR - TIRC7 pathway demonstrates that sHLA-DRα2 treatment inhibits proinflammatory-inflammatory cytokine expression in APC & T cells after lipopolysaccaride (LPS) stimulation in vitro and induces apoptosis in vivo. These results suggest a novel antiproliferative role for HLA-DR mediated via TIRC7, revise the notion of an exclusive stimulatory interaction of HLA-DR with CD4+ T cells and highlights a novel physiologically relevant regulatory pathway.

MDMA (“Ecstasy”) suppresses the innate IFN-γ response in vivo: A critical role for the anti-inflammatory cytokine IL-10

Here we demonstrate that the widely abused drug methylenedioxymethamphetamine (MDMA; "Ecstasy") suppresses innate interferon (IFN)-gamma production in mice following an in vivo lipopolysaccharide (LPS) challenge. IFN-gamma signalling was also impaired by MDMA, as indicated by reduced phosphorylation of signal transducer and activator of transcription-1 (STAT1) and reduced expression of interferon-gamma inducible protein 10 (IP-10/CXCL10); a chemokine induced by IFN-gamma. MDMA also suppressed production of interleukin (IL)-12 and IL-15; two cytokines that induce IFN-gamma production. Our results demonstrate that in vitro exposure to MDMA does not mimic the suppression of innate IFN-gamma observed in vivo, indicating that observed suppression is most likely due to the release of endogenous immunomodulatory substances following drug administration. In this regard, we previously demonstrated that MDMA increases production of the anti-inflammatory cytokine IL-10 in vivo, an event that is mediated by beta-adrenoceptor activation on immune cells. Considering that increased IL-10 production precedes suppression of IFN-gamma induced by MDMA, and also considering that IL-10 can inhibit IL-12 and IFN-gamma production, we examined the possibility that IL-10 was an essential mediator of the suppressive effect of MDMA on the IFN-gamma response. By pre-treating mice with an anti-IL-10 receptor antibody we demonstrate that IL-10 is a critical mediator of MDMA-induced suppression of IFN- gamma production and signalling. Consistent with a role for beta-adrenoceptor activation in the immunosuppressive actions of MDMA, pre-treatment with the beta-adrenoceptor antagonist nadolol blocked the MDMA-induced increase in IL-10, and also inhibited the suppressive action of MDMA on the innate IFN-gamma response. The potential clinical significance of these findings for MDMA users is discussed.

Exposure to bacterial DNA before hemorrhagic shock strongly aggravates systemic inflammation and gut barrier loss via an IFN-gamma-dependent route.

To investigate the role of bacterial DNA in development of an excessive inflammatory response and loss of gut barrier loss following systemic hypotension. Bacterial infection may contribute to development of inflammatory complications following major surgery; however, the pathogenesis is not clear. A common denominator of bacterial infection is bacterial DNA characterized by unmethylated CpG motifs. Recently, it has been shown that bacterial DNA or synthetic oligodeoxynucleotides containing unmethylated CpG motifs (CpG-ODN) are immunostimulatory leading to release of inflammatory mediators. Rats were exposed to CpG-ODN prior to a nonlethal hemorrhagic shock. The role of interferon-gamma (IFN-gamma) was investigated by administration of anti IFN-gamma antibodies. Exposure to CpG-ODN prior to hemorrhagic shock significantly augmented shock-induced release of IFN-gamma, tumor necrosis factor-alpha (TNF-alpha) (P < 0.05), interleukin (IL)-6 (P < 0.05), and nitrite levels (P < 0.05), while there was a defective IL-10 response (P < 0.05). Simultaneously, expression of Toll-like receptor (TLR) 4 in the liver was markedly enhanced. Furthermore, intestinal permeability for HRP significantly increased and bacterial translocation was enhanced in hemorrhagic shock rats pretreated with CpG-ODN. Interestingly, inhibition of IFN-gamma in CpG-treated animals reduced TNF-alpha (P < 0.05), IL-6 (P < 0.05), nitrite (P < 0.05), and intestinal permeability following hemorrhagic shock (P < 0.05) and down-regulated expression of TLR4. Exposure to bacterial DNA strongly aggravates the inflammatory response, disrupts the intestinal barrier, and up-regulates TLR4 expression in the liver following hemorrhagic shock. These effects are mediated via an IFN-gamma-dependent route. In the clinical setting, bacterial DNA may be important in development of inflammatory complications in surgical patients with bacterial infection.

TLR2 but not TLR4 Signalling is Critically Involved in the Inhibition of IFN‐γ‐induced Killing of Mycobacteria by Murine Macrophages

Gamma-interferon (IFN-gamma) plays a determinant role in activating macrophages that are critical to control Mycobacterium tuberculosis infection. However, M. tuberculosis can escape killing by attenuating the response of macrophages to IFN-gamma by blocking the transcription of a subset of IFN-gamma inducible genes. This inhibition occurs after signalling through Toll-like receptor 2 (TLR2). While most studies have investigated the inhibition of IFN-gamma responsive genes after TLR2 signalling, the present study focuses on the functional implications of inhibition of IFN-gamma signalling in macrophages with regard to mycobacteria killing. Here, we provide evidence that exposure of the murine macrophage cell line J774 to the TLR2 ligands; 19-kDa or zymosan, but not the TLR4 ligand LPS, inhibits IFN-gamma-induced killing of Mycobacterium bovis Bacillus Calmette-Guérin (BCG). Moreover, exposure of bone marrow-derived macrophages (BMM) from TLR4-deficient and wild-type (WT), but not from TLR2-deficient mice to 19-kDa lipoprotein (19-kDa) or zymosan, results in an impairment of IFN-gamma-mediated killing. We demonstrate that 19-kDa and zymosan inhibit the ability of IFN-gamma to activate murine macrophages to kill BCG without inhibiting nitric oxide (NO) or tumour necrosis factor (TNF) production. Finally, we demonstrate that the inhibitory effect of 19-kDa on IFN-gamma signalling is overcome with increasing amounts of IFN-gamma indicating that the refractoriness could be reversed at optimal IFN-gamma concentrations. The critical role of TLR2 but not TLR4 signalling in the inhibition of IFN-gamma promoted killing of mycobacteria is discussed.

Transient inhibition of Th1‐type cytokine production by CD4+ T cells in hepatitis B core antigen immunized mice is mediated by regulatory T cells

The non-cytopathic hepatitis B virus (HBV) can induce chronic infections characterized by weak and limited T cell responses against the virus. The factors contributing to the failure to clear HBV and subsequent development of chronic HBV infections are not clearly understood, but a strong interferon-gamma (IFN-gamma) response by CD4+ T cells against the nucleocapsid hepatitis B core antigen (HBcAg) of the virus appears to be important for viral clearance. The present study documents depressed numbers of CD4+ T cells secreting IFN-gamma and interleukin-2 (IL-2) in enzyme-linked immunospot assay (ELISPOT) assays restimulated for 24 hr with antigen following both primary and secondary immunizations of mice with recombinant hepatitis B core antigen (rHBcAg). The kinetics of these responses showed that the depression occurred following a peak response and lasted approximately 2 weeks before returning to the previous peak levels. The depression was abrogated by depletion of CD25+ cells prior to culture in the ELISPOT assay, suggesting inhibition by regulatory T cells. This inhibition of IFN-gamma and IL-2 production was also reversed by in vitro restimulation of the test cells for 48 hr rather than 24 hr in the assay. No such transient, reversible inhibition was detected in the production of IL-5, a Th2-type cytokine. The inhibition in cytokine production did not appear to correlate with the number of antibody-secreting cells or the isotypes produced. This delay by regulatory T cells of Th1-type cytokine production could contribute to viral persistence in chronic HBV infection by interfering with the critical role IFN-gamma plays in protection against viral infections.