Decreased IFN-gamma Production Research Articles

Abstract 991: Platelets inhibit hepatocellular carcinoma growth in the context of nonalcoholic fatty liver disease

Abstract Purpose: Non-alcoholic fatty liver disease (NAFLD) is a major risk factor for hepatocellular carcinoma (HCC) in the US and globally. Platelets have been recognized as an important element of immune system and recently, platelets have been shown to be activated and aggerate in the livers of mice and humans with NAFLD. However, the role of platelets in HCC disease progression in NAFLD is still unknown. Methods: In this study, mice underwent an intrahepatic injection of an HCC cell line and were fed a methionine choline deficient (MCD) diet to generate an HCC NAFLD mouse model. The mouse model was then used to interrogate the interplay between platelets, immune responses and HCC growth within a liver with NAFLD. Anti-GP1bα or clopidogrel were used to deplete or inhibit platelet function and effects on tumor growth and immune cell activation were studied. Platelets express leukotriene receptors, and the possible involvement of leukotrienes was tested with zileuton, a leukotriene inhibitor. Finally, antibody dependent T cell depletion was used to identify the effector cells controlling HCC tumor development. Results: NAFLD mice with HCC treated with clopidogrel showed a close to 3-fold increase in tumor weight compared to vehicle control (50.6g vs 147.8g P<0.05) as well as tumor-to-liver weight ratio (0.05 vs 0.13 P<0.05). CD4+ and CD8+ T-cells from the livers of NAFLD mice showed a 2- to 3-fold increase in CD69+ T-cells compared to non-NAFLD mice (24.2% vs 9.1% P<0.05 in CD4+ and 23.7% vs 8.9% P<0.05 in CD8+, respectively). Interestingly, in NAFLD mice treated with clopidogrel, there was a decrease in CD69+ compared to NAFLD mice treated with control in both CD4+ (18.3% vs 24.2% P<0.05, respectively) and CD8+ (15.32% vs 23.7% P<0.05, respectively) T-cells. Consistently, when NAFLD mice were depleted of platelets with anti-GP1bα, there was a significant decreased in T cell activation compared to IgG control. Similarly, when NAFLD mice were treated with zileuton, there was a decrease in IFN-gamma production in liver isolated CD4+ (5.4% vs 3.1% P=0.12) and CD8+ (23.7% vs 16.7% P<0.05) T-cells compared to vehicle control treated mice. Finally, the clopidogrel treatment increased HCC tumor-to-liver weight ratio was abolished in mice treated with anti-CD8 (0.29 vs 0.23 P>0.05) but not in mice treated with anti-CD4 (0.11 vs 0.16 P<0.05). Conclusion: The results of this study show that in NAFLD livers, platelets increase immune cell activation and inhibit HCC in a CD8+ T-cell dependent manner. The results also suggest that leukotrienes may play a key role in modulating this response, but further studies are required to better characterize this observation. Citation Format: John C. McVey, Chi Ma, Laurence Diggs, Qiong Fu, Tim F. Greten. Platelets inhibit hepatocellular carcinoma growth in the context of nonalcoholic fatty liver disease [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 991.

Mechanism of interferon-gamma production by monocytes stimulated with myeloperoxidase and neutrophil extracellular traps

Neutrophil extracellular traps (NETs) have an important role in antimicrobial innate immunity and release substances that may modulate the immune response. We investigated the effects of soluble factors from NETs and neutrophil granule proteins on human monocyte function by using the Transwell system to prevent cell-cell contact. NET formation was induced by exposing human neutrophils to phorbol myristate acetate (PMA). When monocytes were incubated with PMA alone, expression of interleukin (IL)-4, IL-6, IL-8, and tumor necrosis factor (TNF)-alpha mRNA was upregulated, but IL-10, IL-12, and interferon (IFN)-gamma mRNA were not detected. Incubation of monocytes with NETs enhanced the expression of IL-10 and IFN-gamma mRNA, but not IL-12 mRNA. Myeloperoxidase stimulated IFN-gamma production by monocytes in a dose-dependent manner. Both a nuclear factor-kappaB inhibitor (PDTC) and an intracellular calcium antagonist (TMB-8) prevented upregulation of IFN-gamma production. Neither a combined p38alpha and p38beta inhibitor (SB203580) nor an extracellular signal-regulated kinase inhibitor (PD98059) suppressed IFN-gamma production. Interestingly, a combined p38gamma and p38delta inhibitor (BIRB796) significantly decreased IFN-gamma production. These findings suggest that myeloperoxidase induces IFN-gamma production by monocytes via p38gamma/delta mitogen-activated protein kinase.

ICOS signaling in CD8+ CTL contributes to the antitumor efficacy (P2122)

Abstract The expression of Inducible costimulator (ICOS) on CD8+ CTL and its role in antitumor immunity remains largely unknown. To address this issue, we crossed ICOS-deficient mice with pmel-1 transgenic mice to generate pmel1/ICOS-/- mice. The loss of ICOS in pmel-1CD8+ CTL cells did not affect the expression of other T cell activation markers (e.g. CD25 and CD69) but led to a significant decrease in IFN-gamma production and killing of target tumor cells. Similarly, adoptive transfer of pmel-1 cells into ICOS-ligand knockout mice showed impaired cell proliferation and decreased expression of IFN-gamma and CD107a. Most importantly, when activated ICOS- / - pmel-1 cells were adoptively tranfered into tumor-bearing mice they displayed poor proliferation and diminished antitumor activity as compared to ICOS-intact pmel-1 cells. These results provide direct evidence for the important role of ICOS in CD8+ CTL mediated antitumor immunity and have implications in the development of novel immunotherapy for cancer by manupilation of ICOS pathway.

Dendritic/pancreatic carcinoma fusions for clinical use: Comparative functional analysis of healthy- versus patient-derived fusions

Fetal calf serum (FCS)-independent pancreatic cancer cells were established in plasma protein fraction (PPF)-supplemented medium that is an agent of good manufacturing practice (GMP) grade. Dendritic cells (DCs) were activated with the Toll-like receptor agonist, penicillin-inactivated Streptococcus pyogenes (OK-432) that is also a GMP grade agent. Therefore, sufficient amounts of FCS-independent fusions were successfully generated with decreased potential hazards of FCS. The FCS-independent fusions expressed tumor-associated antigens, HLA-DR, costimulatory molecules, IL-12, and IL-10. Stimulation of T cells with fusions from healthy donors resulted in proliferation of T cells with high expression levels of perforin/granzyme B and IFN-gamma and efficient induction of antigen-specific cytotoxic T lymphocytes (CTLs). Selection and expansion of T-cell clones were confirmed by TCR Vbeta analysis. However, fusions from patients with metastatic pancreatic cancer induced increased expression levels of TGF-beta1 in CD4+ CD25high T cells and low levels of CTLs with decreased IFN-gamma production.

Fas Ligand Expression on T Cells Is Sufficient to Prevent Prolonged Airway Inflammation in a Murine Model of Asthma

Our previous studies revealed that, in a murine model of asthma, mice that received Fas-deficient T cells developed a prolonged phase of airway inflammation, mucus production, and airway hyperreactivity that failed to resolve even 6 weeks after the last challenge. To investigate how Fas-Fas ligand (FasL) interaction occurs between T cells and other cells in vivo, Gld mice with abnormalities of the FasL signaling pathway were used. The reconstituted mice were made by transferring T cells from B6 or Gld mice to Rag(-/-) or FasL-deficient Rag(-/-) mice. We found that Rag(-/-) mice that received B6 T cells resolved the airway inflammation, whereas FasL-deficient Rag(-/-) mice that received Gld T cells developed a prolonged airway inflammation at Day 28, with decreased IFN-gamma production. Both FasL-deficient Rag(-/-) mice that received B6 T cells and Rag(-/-) mice that received Gld T cells also had completely resolved their airway inflammation by Day 28 after challenge. Interestingly, FasL-deficient Rag(-/-) mice that received Gld T cells eventually resolved airway inflammation at Day 42, with a similar level of IFN-gamma production to that of control group. These results demonstrate that FasL expression on either T cells only or non-T cells only was sufficient for the eventual resolution of airway inflammation, and the prolonged airway inflammation in FasL-deficient Rag(-/-) mice that received Gld T cells was correlated with decreased IFN-gamma production by Gld T cells.

Recombinant Human Granulocyte Colony-Stimulating Factor Significantly Decreases the Expression of CXCR3 and CCR6 on T Cells and Preferentially Induces T helper Cells to a T helper 17 Phenotype in Peripheral Blood Harvests

The aim of this study was to investigate the expression of chemokine receptors on T cells and functional changes of T helper (Th) cells in peripheral blood stem cell (PBSC) harvests after treating healthy donors with recombinant human granulocyte colony-stimulating factor (rhG-CSF). Using multiparameter flow cytometry, we analyzed the expression of CXCR3 and CCR6 on T cells and the production of interferon-gamma (IFN-gamma), interleukin-4 (IL-4), and IL-17 by CD4(+) Th cells in PBSC grafts of healthy donors after in vivo rhG-CSF application. Alterations in the relative expression levels of T cell receptor beta variable (TCRBV) family members were determined using real-time polymerase chain reaction (PCR). rhG-CSF mobilization significantly decreased the expression of CXCR3 and CCR6 on T cells. Treating donors with rhG-CSF resulted in decreased IFN-gamma production and dramatically increased IL-4 and IL-17 secretion by CD4(+) Th cells, leading to T cell polarization from the Th1 to the Th2 phenotype and a preferential increase in IL-17-producing CD4(+) Th cells. We did not observe any differences in the relative expression levels of TCRBV family members before and after in vivo rhG-CSF application. Our results suggest that the expression of CXCR3 and CCR6 on donor T cells was dramatically downregulated and an IL-17 phenotype of CD4(+) Th cells was preferentially induced in PBSC grafts after treating healthy donors with rhG-CSF. The observed effects of rhG-CSF on T cells may be independent of the relative expression levels of TCRBV family members.

Anti-inflammatory effects of cannabinoid receptor-2-selective agonists in immune cells (98.12)

Abstract A few studies have been published showing effects of cannabinoids in modulating immune response from T helper cell type 1 (Th1) to T helper cell type 2 (Th2) in a model of Legionella pneumophila infection. Cannabinoids act through two cloned receptors, CB1 and CB2. The CB1 receptor is expressed primarily in the CNS, whereas the CB2 receptor is highly expressed in immune cells in the periphery. Previous results from our laboratories indicated that the CB2 agonist had an anti-inflammatory effect on dendritic cells (DCs) and T cells. DCs differentiated in the presence of CB2 agonist produced less IL-12 upon LPS stimulation and T cells produced less IFN-gamma when co-cultured with these DCs in syngeneic culture in the presence of specific antigen. Here, we report that CB2 agonist treatment of DCs resulted in reduced STAT-1 phosphorylation. Also, treatment of polyclonally activated CD4+ T cells resulted in reduced expression of IL-12 receptor beta 2 chain, which correlated with decreased IFN-gamma production. Our data lead us to believe that CB2 agonists can modulate immune responses by acting directly on CD4+ T cells or through their effects on DCs. This work is supported by NIH/NIAID R01AI05230, and training grant T32 DA07237 (SA)

Adenosine A2Areceptor activation inhibits T helper 1 and T helper 2 cell development and effector function

Adenosine is an immunosuppressive nucleoside, and adenosine A(2A) receptors inhibit T-cell activation. We investigated the role of A(2A) receptors in regulating T helper (Th)1- and Th2-cell development and effector function. A(2A)-receptor stimulation suppressed the development of T-cell receptor (TCR) -stimulated naive T cells into both Th1 and Th2 cells, as indicated by decreased IFN-gamma production by cells developed under Th1-skewing conditions and decreased interleukin (IL) -4, IL-5, and IL-10 production by cells developed under Th2-skewing conditions. Using A(2A) receptor-deficient mice, we demonstrate that A(2A) receptor activation inhibits Th1- and Th2-cell development by decreasing the proliferation and IL-2 production of naive T cells, irrespective of whether the cells are expanded under Th1- or Th2-skewing environment. Using in vivo established Th1 and Th2 cells, we further demonstrate the nonselective nature of A(2A) receptor-mediated immunosuppressive effects, because A(2A) receptor activation decreased IFN-gamma and IL-4 secretion and mRNA level of TCR-stimulated effector Th1 and Th2 cells, respectively. A(2A) receptor mRNA expression in both Th1 and Th2 effector cells increased following TCR stimulation. In summary, these data demonstrate that A(2A) receptor activation has strong inhibitory actions during early developmental, as well as late effector, stages of Th1- and Th2-cell responses.

High frequency of spontaneous interferon-gamma-producing cells in human tonsils: role of local accessory cells and soluble factors.

The frequency of mononuclear cells (MNC) spontaneously secreting interferon-gamma (IFN-gamma) has been examined in freshly isolated cell suspensions from human palatine tonsils. Two-site reverse enzyme-linked immunospot (ELISPOT) analyses, involving short term (20 h) incubation of MNC in the absence of any added exogenous stimulus, revealed that tonsillar MNC suspensions contain exceptionally large numbers of cells secreting IFN-gamma. No significant differences were observed when comparing the frequency of IFN-gamma-producing cells between cell suspensions obtained from hyperplastic and tonsillitis specimens. Cell-sorting experiments disclosed that spontaneous tonsillar IFN-gamma production was essentially contributed by CD4+ T cells, and required the presence of accessory cells and/or soluble factors to be detected. Thus, depletion of plastic adherent cells or monocytes from the tonsillar MNC suspensions resulted in reduced numbers of detectable IFN-gamma-secreting cells. Addition of very small numbers of autologous monocytes restored spontaneous IFN-gamma production in tonsillar MNC cultures depleted of monocytes. Neutralization of endogenous IL-1 beta and IL-2, as well as blocking of the IL-2 receptor, also decreased IFN-gamma production from unfractionated tonsillar cells. Addition of exogenous IL-1 beta restored IFN-gamma production in cultures of tonsillar MNC depleted of plastic adherent cells. Furthermore, IL-1 beta synergized with IL-2 by tonsillar MNC depleted of plastic adherent cells. Furthermore, IL-1 beta synergized with IL-2 by increasing intracellular as well as cell-free levels of IFN-gamma in cultures of unfractionated tonsillar MNC. This study further establishes that the tonsils are highly active immunological organs containing large numbers of T cells spontaneously producing IFN-gamma whose detection is contingent upon the presence of functional accessory cells. It also demonstrates that concomitant production of IL-1 beta and IL-2 occurs in tonsils and is necessary to maintain ongoing synthesis and extracellular accumulation of IFN-gamma in these organs.

Staphylococcus aureusskin colonization in atopic dermatitis children is associated with decreased IFN‐γproduction by peripheral blood CD4+and CD8+T cells

Atopic dermatitis (AD) is a chronic inflammatory skin disorder, which is associated with an increased expression of Th2 cytokines with concomitant decrease in IFN-gamma production by circulating CD4+ and CD8+ T cells. The skin of patients with AD is often colonized by Staphylococcus aureus, which may reflect in changes in immunological parameters. The aim of the study was flow cytometric measurement of some peripheral blood lymphocyte subsets expressing naive/memory marker (RA/RO) and activation marker (CD25) as well as intracellular production of IFN-gamma by peripheral blood CD4+ and CD8+ T cells from varied severity AD children and determine the impact of S. aureus skin colonization on cytokines profiles. There was a significant increase in the percentage of CD4+ and CD8+ T cells producing IL-4 and IL-13 and decrease in the percentage of CD4+ and CD8+ T cells producing IFN-gamma upon in vitro stimulation with phorbol 12-myristate 13-acetate and ionomycin in children with AD compared to healthy ones. The absolute number of CD4+ and CD8+ T cells expressing memory marker CD45RO was elevated as compared with controls. The severity of AD was positively correlated with the percentage of lymphocyte subsets: CD45RO+, CD4+CD45RO+, and the percentage of CD3+ and CD4+ expressing CD25 as well as the number of S. aureus on the skin. In conclusion, both CD4+ and CD8+ memory T cells are involved in the immunopathogenesis of AD. S. aureus skin colonization is related with disease severity and changes in expression of CD45RO and CD25 on T cells. A decrease in the percentage of CD4+ and CD8+ T cells producing IFN-gamma in AD children may explain propensity for skin infection.

Modulation of T-effector function by imatinib at the level of cytokine secretion

Recently, evidence was provided, that the selective tyrosine kinase inhibitor imatinib mesylate (imatinib) has immunomodulatory or suppressive effects. However, the discussion about imatinib's influence on immune cells is still controversial. The aim of this study was to clarify the effect of imatinib on CD8+ and CD4+ T-cell effector functions. For analyzing T-cell effector functions T-cell receptor-transgenic ovalbumin-specific CD8+ T cells and in vivo primed CD4+ Th1 cells were used. T-cell effector functions were analyzed on the level of antigen responsiveness by intracellular cytokine staining, by measuring cytokine secretion in an interferon-gamma (IFN-gamma) enzyme-linked immunosorbent assay and by detecting cytotoxicity using the fluorescein-activated cell sorting-based fluorometric assessment of T-lymphocyte antigen-specific lysis assay. It was demonstrated that imatinib inhibits antigen-specific IFN-gamma secretion of both CD4+ and CD8+ T-effector cells at therapeutically relevant concentrations, while T cells remain responsive. The decrease of IFN-gamma production was not due to the loss of T-cell viability. Further, it was shown that the effector T cells are modulated rather than suppressed, because the cytolytic functions of CD8+ cytotoxic T cells were not altered. Residual cytolytic activity in the presence of imatinib was not due to FasL interaction. These experiments provide evidence for a therapeutically relevant modulation of T-cell effector functions by imatinib. This might open a possible applicability of imatinib in various autoimmune and inflammatory diseases, including multiple sclerosis and rheumatoid arthritis.

Influence of interleukin-15 on CD8+ natural killer cells in human immunodeficiency virus type 1-infected chimpanzees

Chimpanzees are susceptible to human immunodeficiency virus type-1 (HIV-1) and develop persistent infection but generally do not progress to full-blown AIDS. Several host and immunological factors have been implicated in mediating resistance to disease progression. Chimpanzees have a higher prevalence of circulating natural killer (NK) cells than humans; however, their role in mediating resistance to disease progression is not well understood. Furthermore, NK cell survival and activity have been shown to be dependent on interleukin-15 (IL-15). Accordingly, the influence of IL-15 on NK cell activity and gamma interferon (IFN-gamma) production was evaluated in naive and HIV-1-infected chimpanzees. In vitro stimulation of whole-blood cultures with recombinant gp120 (rgp120) resulted in enhanced IFN-gamma production predominantly by the CD3(-) CD8(+) subset of NK cells, and addition of anti-IL-15 to the system decreased IFN-gamma production. Moreover, in vitro stimulation with recombinant IL-15 (rIL-15) augmented IFN-gamma production from this subset of NK cells and increased NK cell cytotoxic activity. Stimulation with rgp120 also resulted in a 2- to 7-fold increase in IL-15 production. These findings suggest that chimpanzee CD3(-) CD8(+) NK cells play a vital role in controlling HIV-1 infection by producing high levels of IFN-gamma, and that IL-15 elicits IFN-gamma production in this subpopulation of NK cells in HIV-1-infected chimpanzees.

Recurrent Headache as the Main Symptom of Acquired Cerebral Toxoplasmosis in Nonhuman Immunodeficiency Virus-infected Subjects With no Lymphadenopathy

Headache and/or migraine, a common problem in pediatrics and internal medicine, affect about 5% to 10% children and adolescents, and nearly 30% of middle-aged women. Headache is also one of the most common clinical manifestations of acquired Toxoplasma gondii infection of the central nervous system (CNS) in immunosuppressed subjects. We present 11 apparently nonhuman immunodeficiency virus-infected children aged 7 to 17 years (8 girls, 3 boys) and 1 adult woman with recurrent severe headaches in whom latent chronic CNS T. gondii infection not manifested by enlarged peripheral lymph nodes typical for toxoplasmosis, was found. In 7 patients, the mean serum IgG Toxoplasma antibodies concentration was 189 +/- 85 (SD) IU/mL (range 89 to 300 IU/mL), and in 5 other subjects, the indirect fluorescent antibody test titer ranged from 1:40 to 1:5120 IU/mL (n= <1:10 IU/mL). Some of the patients suffered also from atopic dermatitis (AD) and were exposed to cat and/or other pet allergens, associated with an increased IL-4 and decreased IFN-gamma production. These cytokine irregularities caused limited control of cerebral toxoplasmosis probably because IL-4 down-regulated both the production of IFN-gamma and its activity, and stimulated production of a low NO-producing population of monocytes, which allowed cysts rupture, increased parasite multiplication and finally reactivation of T. gondii infection. The immune studies performed in 4 subjects showed a decreased percentage of T lymphocytes, increased total number of lymphocytes B and serum IgM concentration, and impaired phagocytosis. In addition, few of them had also urinary tract diseases known to produce IL-6 that can mediate immunosuppressive functions, involving induction of the anti-inflammatory cytokine IL-10. These disturbances probably resulted from the host protective immune reactions associated with the chronic latent CNS T. gondii infection/inflammation. This is consistent with significantly lower enzyme indoleamine 2,3-dioxygenase (IDO) activity reported in atopic than in nonatopic individuals, and an important role that IDO and tryptophan degradation pathways plays in both, the host resistance to T. gondii infection and its reactivation. Analysis of literature information on the subjects with different types of headaches caused by foods, medications, and other substances, may suggest that their clinical symptoms and changes in laboratory data result at least in part from interference of these factors with dietary tryptophan biotransformation pathways. Several of these agents caused headache attacks through enhancing NO production via the conversion of arginine to citrulline and NO by the inducible nitric oxide synthase enzyme, which results in the high-output pathway of NO synthesis. This increased production of NO is, however, quickly down-regulated by NO itself because this biomolecule can directly inactivate NOS, may inhibit Ia expression on IFN-gamma-activated macrophages, which would limit antigen-presenting capability, and block T-cell proliferation, thus decreasing the antitoxoplasmatic activity. Moreover, NO inhibits IDO activity, thereby suppressing kynurenine formation, and at least one member of the kynurenine pathway, 3-hydroxyanthranilic acid, has been shown to inhibit NOS enzyme activity, the expression of NOS mRNA, and activation of the inflammatory transcription factor, nuclear factor-kB. In addition, the anti-inflammatory cytokines IL-4 and IL-10, TGF-beta, and a cytokine known as macrophage deactivating factor, have been shown to directly modulate NO production, sometimes expressing synergistic activity. On the other hand, IL-4 and TGF-beta can suppress IDO activity in some cells, for example human monocytes and fibroblasts, which is consistent with metabolic pathways controlled by IDO being a significant contributor to the proinflammatory system. Also, it seems that idiopathic intracranial hypertension, pseudotumor cerebri, and aseptic meningitis, induced by various factors, may result from their interference with IDO and inducible nitric oxide synthase activities, endogenous NO level, and cytokine irregularities which finally affect former T. gondii status 2mo in the brain. All these biochemical disturbances caused by the CNS T. gondii infection/inflammation may also be responsible for the relationship found between neurologic symptoms, such as headache, vertigo, and syncope observed in apparently immunocompetent children and adolescents, and physical and psychiatric symptoms in adulthood. We therefore believe that tests for T. gondii should be performed obligatorily in apparently immunocompetent patients with different types of headaches, even if they have no enlarged peripheral lymph nodes. This may help to avoid overlooking this treatable cause of the CNS disease, markedly reduce costs of hospitalization, diagnosis and treatment, and eventually prevent developing serious neurologic and psychiatric disorders.

Comparison of interferon-γ response between tuberculosis and non-tubercular Pneumonia

Macrophages aided by interferon-gamma (IFN-gamma) are vital to controlling Mycobacterium tuberculosis (M. tuberculosis) infection. Although numerous studies have compared IFN-gamma response between tubercular patients and healthy controls, no studies have investigated IFN-gamma response in patients with pulmonary tuberculosis and non-tubercular pneumonia. The aim of this work was to examine the difference in IFN-gamma response between patients with tuberculosis and non-tubercular pneumonia. IFN-gamma production was detected based on the difference in supernatants between non-stimulated and stimulated peripheral blood mononuclear cells by phytohemagglutinin in 83 tubercular patients and 47 patients with pneumonia. Presence of a cavity on chest radiography and co-morbidities of pneumoconiosis, bronchiectasis, liver cirrhosis, renal failure on hemodialysis, diabetes mellitus (DM) and lung cancer were recorded for analysis. Interferon-gamma response, DM and a cavity on chest radiography were independent factors for predicting active pulmonary tuberculosis. Interferon-gamma response was decreased in patients with pulmonary tuberculosis compared with that in patients with non-tubercular pneumonia. Notably, M. tuberculosis infection was the principal factor correlated with IFN-gamma response. The IFN-gamma response was principally affected by M. tuberculosis infection and not by other co-morbidities. Further study is required to identify the mechanism of decreased IFN-gamma production.

Production of interferon γ in asthmatic patients with small bacille Calmette-Guerin scars: A pilot study

Some studies suggest a decrease in interferon (IFN) gamma production among atopic individuals, and others refer to low IFN-gamma levels as a characteristic of asthma, regardless of the atopic state. Recent research has indicated a relation between asthma and a tendency toward a bacille Calmette-Guerin (BCG) vaccine scar with a small diameter, suggesting an association of this illness with the low production of IFN-gamma. The aim of this study was to check the hypothesis that asthmatic patients with a small BCG scar present low IFN-gamma production. This work was a quasi-experimental study. The capacity to produce IFN-gamma among 12 asthmatic patients with an average BCG vaccine scar diameter <5 mm was compared with 7 nonasthmatic individuals with scars > or =5 mm using whole blood cells stimulated by purified protein derivative (PPD), phorbol myristate acetatelionomycin (PMA/Iono) and medium (no stimulus). The study also analyzed the relation between IFN-gamma production after PPD stimulation and eosinophil count, total IgE, and dust-mite-specific IgE levels. The asthmatic patients presented a decrease in IFN-gamma production in comparison with a control group after PPD stimulation (p < 0.02). However, the same was not observed when PMA/Iono stimulation was used or when no stimulation was used. Asthmatic patients with small BCG scars showed a decrease in IFN-gamma production, suggesting that there may be an intrinsic characteristic of these patients.

A Synergistic Effect Between PG490-88 and Tacrolimus Prolongs Renal Allograft Survival in Monkeys

This study was undertaken to determine if PG490-88 and tacrolimus (Tac) act synergistically to prevent renal allograft rejection in monkeys and to explore possible mechanisms of synergy between these agents. MHC-mismatched renal allografts were transplanted into cynomolgus monkeys after bilateral nephrectomy. Recipients were divided into the following groups: (i) no treatment; (ii) PG490-88 (0.03 mg/kg); (iii) Tac (1 mg/kg); (iv) PG490-88 (0.01 mg/kg) + Tac (1 mg/kg) and (v) PG490-88 (0.03 mg/kg) + Tac (1 mg/kg). Through synergy PG490-88 and Tac inhibited anti-CD3/PMA-induced T-cell proliferation and IFN-gamma expression in vitro. Tac monotherapy only marginally prolonged survival (27 +/- 3.2 days), while the combination of PG490-88 and Tac significantly prolonged graft survival to a median of 99 days (PG490-88 at 0.03 mg) and 38.5 days (PG490-88 at 0.01 mg/kg). Prolonged survival correlated with inhibited IgM production as well as reduced T-cell infiltration, IL-2 protein expression and NF-AT/NF-kappaB activity. We conclude that PG490-88 and a subtherapeutic dose of Tac significantly prolong renal allograft survival in monkeys through the synergistic inhibition of T-cell activation and a decrease in IFN-gamma production and NF-AT/NF-kappaB activity.

The presence of IFNG 3/3 genotype in the recipient associates with increased risk for Epstein–Barr virus reactivation after allogeneic haematopoietic stem cell transplantation

Recent studies have shown that interferon-gamma gene (IFNG) polymorphism constitutes a risk factor for acute and chronic graft-versus-host disease (GvHD) after allogeneic haematopoietic stem cell transplantation (HSCT). Patients with IFNG 3/3 have been found to be more prone to GvHD. This rather puzzling result, as 3/3 genotype is associated with a decreased IFN-gamma production, was investigated in the present study in the context of Epstein-Barr virus (EBV) reactivation. Microsatellite polymorphism (CA)n within the first intron of IFNG gene was assessed in 83 HSCT recipients and related to EBV load. Quantification of EBV copies was performed by a real-time polymerase chain reaction in peripheral blood cells taken from the patients 2-3 months after HSCT. It was found, that patients having IFNG 3/3 genotype presented with a high number of EBV copies (over 10/10(5) blood cells) when compared with the recipients with other IFNG genotypes (10/14 vs. 17/69, P < 0.001). This association was independent of recipient's age, underlying disease, conditioning regimen, type of donor, source of stem cells or pretransplant donor and recipient EBV serological status. Thus IFNG 3/3 genotype, known to be associated with a decreased IFN-gamma production, appeared as a factor significantly contributing to the risk of EBV reactivation after allogeneic HSCT.

AT1R blockade reduces IFN-γ production in lymphocytes in vivo and in vitro

Type 1 angiotensin II (Ang II) receptor (AT(1)R) signaling induces proinflammatory responses. Recent studies suggest that T lymphocytes express AT(1)R; yet the effects of Ang II binding to AT(1)R on T cells are poorly understood. We examined the effect of AT(1)R blockade on release of the proinflammatory cytokine, interferon-gamma (IFN-gamma) by human lymphocytes in vivo and in vitro. We used an AT(1)R blocker losartan in a randomized clinical trial in kidney transplant recipients over a 12-month period [AT(1)R blocker (N= 11) and control (N= 10)]. Peripheral blood lymphocytes, isolated from both cohorts, were analyzed by enzyme-linked immunosorbent spot assays (ELISPOT) analyses and real-time reverse transcription-polymerase chain reaction (RT-PCR) to enumerate IFN-gamma producing T cells and IFN-gamma mRNA levels. The effects of AT(1)R blockade in vitro were assessed using human alloreactive T cells and an IFN-gamma producing human cytotoxic T-lymphocyte line. Alloreactive T cells were treated with losartan or candesartan and enzyme-linked immunosorbant assay (ELISA) was used to measure IFN-gamma protein release. The cytotoxic T-lymphocyte line also was AT(1)R blocker-treated prior to determining IFN-gamma producing cells by intracellular cytokine staining. The AT(1)R blocker cohort had a significant decrease in IFN-gamma producing peripheral blood lymphocytes (P< or = 0.05 for each time point) and IFN-gamma mRNA levels (P= 0.01 vs. control patients). Losartan also decreased IFN-gamma production (P < 0.001) in purified alloreactive T cells in vitro as did candesartan. Moreover, Ang II amplified IFN-gamma generation (P < 0.05) in alloreactive T cells while AT(1)R blocker treatment inhibited Ang II's effect (P < 0.04). AT(1)R blocker treatment furthermore also inhibited IFN-gamma production in the cytotoxic T-lymphocyte line. AT(1)R blockers may have a clinically relevant immunomodulatory role by blocking IFN-gamma production in T cells.

Effects of polyphenols on human Th1 and Th2 cytokine production

Numerous phenolic compounds are consumed in the diet in a range of foods. There are very few studies of the effects of these compounds on the production of lymphocyte-derived cytokines. To investigate the effects of five phenolic compounds on cytokine production by cultured human lymphocytes. Human whole blood cultures were stimulated with the T cell stimulant concanavalin A for 48 h in the presence of phenolic compounds (vanillic acid, syringic acid, kaempferol, oleuropein and tyrosol) at concentrations up to 10(-4) M. Interleukin (IL)-2, IL-4 and interferon-gamma (IFN-gamma) concentrations were measured in the culture supernatants by ELISA. IFN-gamma concentration was significantly lower in cultures containing 10(-4) M kaempferol than in cultures with kaempferol at 10(-7), 10(-6)and 10(-5) M or without kaempferol. The other phenolic compounds did not affect IFN-gamma concentration and none of the phenolics tested affected IL-2 or IL-4 concentrations. Some, but not all, phenolic compounds can decrease IFN-gamma production by stimulated human whole blood cultures.

Impaired Th1 responses in mice deficient in Epstein‐Barr virus‐induced gene 3 and challenged with physiological doses of Leishmania major

Protection against Leishmania major is dependent on IL-12 release from L. major-infected dendritic cells (DC) that induce IFN-gamma-producing Th1/Tc1 cells. IL-27, a novel member of the IL-12 family, is a heterodimer composed of p28 and IL-12p40-related Epstein-Barr virus-induced gene 3 (EBI3), and was shown to be produced by DC. In this study, we utilized EBI3-deficient mice to investigate the role of IL-27 in leishmaniasis using physiological low-dose infections that mimic natural transmissions. Lesions in EBI3(-/-) mice were significantly larger between weeks 3 and 10 post infection, reaching up to approximately threefold increased lesion volumes compared to wild types. In parallel, dermal lesions of EBI3(-/-) mice contained greater parasite numbers, reaching a peak load that was 2-log higher than in C57BL/6 mice. However, lesions in EBI3(-/-) and wild-type mice resolved after 12 weeks. At early time points, the antigen-specific cytokine response in EBI3(-/-) lymph nodes showed increased levels of IL-4, IL-10 and IL-13 and decreased IFN-gamma production. IL-27 production was restricted to the DC population, since the majority of EBI3 expression in lymph nodes of infected mice was found in CD11c(+) cells. In conclusion, our data show that DC-derived IL-27 is critical for the timely initiation of efficient anti-parasite Th1 immunity early in infections.