Regulation Of IFN-γ Production Research Articles

CREATINE KINASE EXPRESSION IS PROTECTIVE IN A MOUSE COLITIS MODEL

Abstract BACKGROUND We have previously identified a loss of expression of creatine pathway proteins in mucosal tissue samples from inflammatory bowel disease patients. This is important as creatine is vital to ATP buffering in energetically active cells. The goal of this study was to determine the impact of decreased creatine kinase (CK) expression on mucosal inflammation by utilizing a whole body CK Brain and CK Mitochondrial knockout mouse (CKB/MitKO) in the dextran sulfate sodium (DSS) model of acute colitis. METHODS CKB/MitKO and matched controls were administered 1.5% DSS in drinking water for 5 days and assessed by body weight loss, disease activity index, mucosal permeability, histology and inflammatory cytokine production by Mesoscale analysis. In subsequent experiments, interferon-γ (IFN-γ) was given by intraperitoneal injection during the DSS experiment. We also analyzed transcriptional differences between CKB/MitKO and controls using a transcription factor array and western blot. Finally, we evaluated the impact of CK on the regulation of IFN-γ and transcriptional regulation of IFN-γ production in immune cells by flow cytometry. RESULTS Loss of CKB/Mit did not result in detectable baseline inflammation or barrier dysfunction in CKB/MitKO mice as compared to controls. However, in the setting of the chemical irritant DSS, CKB/MitKO mice had significantly increased weight loss, disease activity, intestinal permeability, histologic inflammation and decreased colon length. While control mice treated with DSS had significant increases in mucosal IFN-γ, the CKB/MitKO mice had strikingly absent mucosal protein concentrations of IFN-γ. Although IFN-γ is commonly considered a pro-inflammatory cytokine, it has also been shown to help promote inflammatory resolution. Therefore, we proceeded to add back IFN-γ in DSS experiments and we identified significant protection from weight loss and histologic damage with the addition of IFN-γ. We went on to identify transcription factor differences between CKB/MitKO and control splenocytes that revealed a >40-fold increase in the hypoxia responsive factor, HIF-1a, in CKB/MitKO splenocytes. We confirmed a causative relationship between HIF-1 expression and IFN-γ by treating mouse splenocytes with a HIF stabilizer and identified that HIF stabilization results in a reduction of IFN-γ production by CD4+ and CD8+ T cells. CONCLUSIONS CK is an essential energetic regulator. Using CK knockout mice we have identified an important role for CK in regulating IFN-γ production, mediated at least in part by HIF stabilization. These findings support a novel role for CK in tissue protection by immune regulation during acute colitis. Better understanding the role of energetics and the creatine pathway in particular may provide novel therapeutic options in inflammatory bowel disease.

Klf10 favors Mycobacterium tuberculosis survival by impairing IFN-γ production and preventing macrophages reprograming to macropinocytosis.

Mycobacterium tuberculosis has developed diverse mechanisms to survive inside phagocytic cells, such as macrophages. Phagocytosis is a key process in eliminating invading pathogens; thus, M. tuberculosis efficiently disrupts phagosome maturation to ensure infection. However, inflammatory cytokines produced by macrophages in response to early M. tuberculosis infection are key to promoting bacterial clarification. IFN-γ enhances M. tuberculosis engulfment and destruction by reprogramming macrophages from phagocytosis to macropinocytosis. Here, we show that the transcription factor Krüppel-like factor 10 (Klf10) plays a positive role in M. tuberculosis survival and infection by negatively modulating IFN-γ levels. Naïve Klf10-deficient macrophages produce more IFN-γ upon stimulation than wild-type macrophages, thus enhancing bacterial uptake and bactericidal activity achieved by macropinocytosis. Moreover, Klf10⁻/ ⁻ macrophages showed cytoplasmic distribution of coronin 1 correlated with increased pseudopod count and length. In agreement with these observations, Klf10⁻/ ⁻ mice showed improved bacterial clearance from the lungs and increased viability. Altogether, our data indicate that Klf10 plays a critical role in M. tuberculosis survival by preventing macrophage reprogramming from phagocytosis to macropinocytosis by negatively regulating IFN-γ production upon macrophage infection.

P0404B CELLS FROM PATIENTS WITH GRANULOMATOSIS WITH POLYANGIITIS FAIL TO REGULATE T CELL INTERFERON GAMMA RELEASE BUT DO REGULATE OTHER ASPECTS OF T CELL ACTIVATION

Abstract Background and Aims Regulatory features of B-cells have gained increased attention in patients with systemic vasculitis, including granulomatosis with polyangiitis (GPA). The aim with this study was to investigate the frequencies and phenotypes of two putative regulatory B (Breg) cell populations and assess the capacity of B-cells from GPA patients to regulate T-cell activation. Method 37 GPA patients (22 remission and 15 active) and 31 healthy controls (HC) were included for measurement of CD19+CD24highCD27+ (memory) and CD19+CD24highCD38high (transitional) Breg cells, and 11 GPA patients in remission and 12 HC for the functional studies of B-cells. HC were matched for age and gender. Results Memory Breg cells were reduced in GPA both during active disease and remission compared with HC, whereas transitional Breg cells did not differ between the groups. Memory Breg cells consisted of less CD25+ cells during both active disease and remission but expressed more PD-L1 and CD86 during remission compared with active disease and HC. B-cells from GPA patients regulated T-cell proliferation and IL-17a production but failed to regulate IFN-γ production (Fig 1). The levels of IFN-γ and the IFN-γ regulated cytokine CXCL10 were elevated in patient plasma. Conclusion Despite reduced levels and altered phenotypes of memory Breg cells, the capacity of GPA patient B-cells to regulate T-cell proliferation and IL-17a production was unaffected. However, we found a profound inability of patient B-cells to regulate T-cell IFN-γ production. This alteration could be important for the persistent inflammation that continues during remission and results in the chronic relapsing nature of the disease

MicroRNA-29b modulates the innate immune response by suppressing IFNγs production in orange-spotted grouper (Epinephelus coioides)

Interferon-γ (IFNγ), a type II interferon, is essential to host resistance against various infections. Unlike other vertebrates, fish have two types of IFNγs, IFNγ1 (also named IFNγ-rel) and IFNγ2. MicroRNAs (miRNAs) regulate multiple biological processes by suppressing mRNA translation or inducing mRNA degradation. Among them, miR-29 can directly target IFNγ and affact innate and adaptive immune responses in mice. There are five members of the miR-29 family in orange-spotted grouper (Epinephelus coioides), which share the same miRNA seed region. However, whether miR-29 directly targets E. coioides IFNγs and regulate IFNγ production is still unknown. In the present study, the negative correlation between miR-29b and both IFNγs in immune tissues of healthy E. coioides and grouper spleen cells (GS cells) stimulated with LPS or poly I:C was demonstrated. Moreover, dual-luciferase reporter assays and western blotting were performed to demonstrate that miR-29b suppressed E. coioides IFNγ production. Studies of NO production in GS cells after miR-29b transfection revealed that miR-29b overexpression affected NO production through the downregulation of IFNγ expression. Taken together, our results suggest that miR-29b may directly target E. coioides IFNγs and modulate IFNγ-mediated innate immune responses by suppressing E. coioides IFNγs production.

CD56−CD16+ NK cells from HIV-infected individuals negatively regulate IFN-γ production by autologous CD8+ T cells

The percentage of human CD56- CD16+ NK cells increases during chronic infection with human HIV; however, the biologic role of CD56- CD16+ NK cells in HIV infection is unclear. Our results demonstrate that the percentage of CD56- CD16+ NK cells producing IL-10 and TGF-β was higher than CD56dim CD16+ NK cells. CD56- CD16+ NK cells could inhibit IFN-γ production by autologous CD8+ T cells, and this inhibition could be partially reversed by anti-IL-10, anti-TGF-β, or anti-PD-L1 mAbs. CD56- CD16+ NK cells are potential targets for the development of novel immune therapies against HIV infection.

Targeting Tumors with IL-10 Prevents Dendritic Cell-Mediated CD8+ T Cell Apoptosis

Increasing evidence demonstrates that interleukin-10 (IL-10), known as an immunosuppressive cytokine, induces antitumor effects depending on CD8+ Tcells. However, it remains elusive how immunosuppressive effects of IL-10 contribute to CD8+ Tcell-mediated antitumor immunity. We generated Cetuximab-based IL-10 fusion protein (CmAb-(IL10)2) to prolong its half-life and allow tumor-targeted delivery of IL-10. Our results demonstrated potent antitumor effects of CmAb-(IL10)2 with reduced toxicity. Moreover, we revealed a mechanism of CmAb-(IL10)2 preventing dendritic cell (DC)-mediated CD8+ tumor-infiltrating lymphocyte apoptosis through regulating IFN-γ production. When combined with immune checkpoint blockade, CmAb-(IL10)2 significantly improves antitumor effects in mice with advanced tumors. Our findings reveal a DC-regulating role of IL-10 to potentiate CD8+ Tcell-mediated antitumor immunity and provide a potential strategy to improve cancer immunotherapy.

CFTR, which not only serves as a TCR signaling molecule but also function as an anion channel, dual-negatively regulates IFN-γ production and tumor immunity in γδ T cells

Abstract γδ T cells have been demonstrated to have the MHC independent-broad anti-tumor activities with high IFN-γ production in the infiltration of the tumor tissue, therefore, they become the strongest interest for cancer immunotherapy. Ion channels play critical roles in the cell functions, especially in the regulation of immune cells functions. Cystic fibrosis transmembrane conductance regulator (CFTR) is an important chloride channel and regulator in the apical membrane of epithelial cells. Recently, CFTR was reported to participate in the immune regulation and likely account for the risk of developing cancer. However, little is known about the effect of CFTR on γδ T cells tumor immunity. In this study, we would like to elucidate the regulatory mechanisms and indispensable functions of CFTR on γδ T cells in tumor immunity. We found that genetic deletion of CFTR displayed the increased TCR signaling and elevates IFN-γ release in peripheral γδ T cells, but not in CD4+ T cells. The absence of CFTR in γδ T cells strengthened the TCR signaling. The underlying mechanism of higher TCR signaling was likely that CFTR may participate in TCR signaling via interacting with the PLCγ-1-LCK-ZAP70 signaling complex in γδ T cells. Consistent with the data in mice, CFTR inhibition, which increased intracellular Ca2+ concentration, significantly increased the IFN-γ production in γδ T cells. Finally, CFTR−/−γδ T cells displayed higher cytolytic ability against the B16 melanoma and protected against B16 tumor growth. This study systematically defined the roles of CFTR in the activation of γδ T cells and its potential contribution to tumor immunity, which will enrich the γδ T cells biology and expand the sight for the application of CFTR in γδ T cells cancer immunotherapy.

Inducible Costimulator Expressing T Cells Promote Parasitic Growth During Blood Stage Plasmodium berghei ANKA Infection.

The lethality of blood stage Plasmodium berghei ANKA (PbA) infection is associated with the expression of T-bet and production of cytokine IFN-γ. Expression of inducible costimulator (ICOS) and its downstream signaling has been shown to play a critical role in the T-bet expression and IFN-γ production. Although earlier studies have examined the role of ICOS in the control of acute blood-stage infection of Plasmodium chabaudi chabaudi AS (a non-lethal model of malaria infection), its significance in the lethal blood-stage of PbA infection remains unclear. Thus, to address the seminal role of ICOS in lethal blood-stage of PbA infection, we treated PbA-infected mice with anti-ICOS antibody and observed that these mice survived longer than their infected counterparts with significantly lower parasitemia. Anti-ICOS treatment notably depleted ICOS expressing CD4+ and CD8+ T cells with a concurrent reduction in plasma IFN-γ, which strongly indicated that ICOS expressing T cells are major IFN-γ producers. Interestingly, we observed that while ICOS expressing CD4+ and CD8+ T cells produced IFN-γ, ICOS−CD8+ T cells were also found to be producers of IFN-γ. However, we report that ICOS+CD8+ T cells were higher producers of IFN-γ than ICOS−CD8+ T cells. Moreover, correlation of ICOS expression with IFN-γ production in ICOS+IFN-γ+ T cell population (CD4+ and CD8+ T cells) suggested that ICOS and IFN-γ could positively regulate each other. Further, master transcription factor T-bet importantly involved in regulating IFN-γ production was also found to be expressed by ICOS expressing CD4+ and CD8+ T cells during PbA infection. As noted above with IFN-γ and ICOS, a positive correlation of expression of ICOS with the transcription factor T-bet suggested that both of them could regulate each other. Taken together, our results depicted the importance of ICOS expressing CD4+ and CD8+ T cells in malaria parasite growth and lethality through IFN-γ production and T-bet expression.

Innate immunity: Fibrinolytic and innate systems collide.

Nuclear factor of activated T cells (NFAT) regulates IFNγ production in natural killer cells and controls fibrinolytic processes required to clear skin infections.

Naive T cells are intrinsically biased in helper T cell functions

Abstract CD5, a negative regulator of T cell receptor (TCR) signals, is a cell surface protein whose expression reflects the self-reactivity of the TCR for self-peptide. Recent studies have shown that CD5lo versus CD5hi T cells differentially respond to foreign antigen in primary and recall responses and may also have biased differentiation potential. Within the CD4+ T cell lineage, in particular, evidence points to the preferential recruitment of CD5hi CD4+ cells into the memory compartment and biased regulatory T cell development. However, the extent to which CD5 levels influence T cell differentiation and effector cytokine production is unknown. We have expanded upon these findings to show that ex vivo activated murine CD5lo CD4+ T cells produce relatively greater amounts of the Th1 cytokine IFNγ compared to their CD5hi counterparts. Interestingly, this difference is specific to IFNγ production and is not attributable to differential expression of the Th1 cell-specific transcription factor T-bet. Given the distinct metabolic programs associated with these T cell subsets and recent evidence that glycolysis regulates IFNγ production through an epigenetic mechanism, we propose that CD5 levels shape T cell function by direct or indirect regulation of cellular metabolism. Consistent with this, our preliminary data suggests that modulation of cellular acetate levels can restore IFNγ production in CD5hi cells similar to that of CD5lo cells. Our ongoing studies are focused on epigenetic and metabolic profiling of CD5lo and CD5hi cells in order to better understand the observed lineage bias.

NFAT2 Regulates Generation of Innate-Like CD8+ T Lymphocytes and CD8+ T Lymphocytes Responses

Nuclear factor of activated T cells (NFAT) 2 null mutant mice die in utero of cardiac failure, precluding analysis of the role of NFAT2 in lymphocyte responses. Only the NFAT2−/−/Rag-1−/− chimeric mice model gave insight into the role of NFAT2 transcription factor in T lymphocyte development, activation, and differentiation. As reports are mainly focused on the role of NFAT2 in CD4+ T lymphocytes activation and differentiation, we decided to investigate NFAT2’s impact on CD8+ T lymphocyte responses. We report that NFAT2 is phosphorylated and inactive in the cytoplasm of naive CD8+ T cells, and upon TCR stimulation, it is dephosphorylated and translocated into the nucleus. To study the role of NFAT2 in CD8+ T responses, we employed NFAT2fl/flCD4-Cre mice with NFAT2 deletion specifically in T cells. Interestingly, the absence of NFAT2 in T cells resulted in increased percentage of non-conventional innate-like CD8+ T cells. These cells were CD122+, rapid producer of interferon gamma (IFN-γ) and had characteristics of conventional memory CD8+ T cells. We also observed an expansion of PLZF+ expressing CD3+ thymocyte population in the absence of NFAT2 and increased IL-4 production. Furthermore, we found that CD8+ T lymphocytes deficient in NFAT2 had reduced activation, proliferation, and IFN-γ and IL-2 production at suboptimal TCR strength. NFAT2 absence did not significantly influence differentiation of CD8+ T cells into cytotoxic effector cells but reduced their IFN-γ production. This work documents NFAT2 as a negative regulator of innate-like CD8+ T cells development. NFAT2 is required for complete CD8+ T cell responses at suboptimal TCR stimulation and regulates IFN-γ production by cytotoxic CD8+ T cells in vitro.

Transcriptional regulator Bhlhe40 works as a cofactor of T-bet in the regulation of IFN-γ production in iNKT cells

Invariant natural killer T (iNKT) cells are a subset of innate-like T cells that act as important mediators of immune responses. In particular, iNKT cells have the ability to immediately produce large amounts of IFN-γ upon activation and thus initiate immune responses in various pathological conditions. However, molecular mechanisms that control IFN-γ production in iNKT cells are not fully understood. Here, we report that basic helix-loop-helix transcription factor family, member e40 (Bhlhe40), is an important regulator for IFN-γ production in iNKT cells. Bhlhe40 is highly expressed in stage 3 thymic iNKT cells and iNKT1 subsets, and the level of Bhlhe40 mRNA expression is correlated with Ifng mRNA expression in the resting state. Although Bhlhe40-deficient mice show normal iNKT cell development, Bhlhe40-deficient iNKT cells show significant impairment of IFN-γ production and antitumor effects. Bhlhe40 alone shows no significant effects on Ifng promoter activities but contributes to enhance T-box transcription factor Tbx21 (T-bet)-mediated Ifng promoter activation. Chromatin immunoprecipitation analysis revealed that Bhlhe40 accumulates in the T-box region of the Ifng locus and contributes to histone H3-lysine 9 acetylation of the Ifng locus, which is impaired without T-bet conditions. These results indicate that Bhlhe40 works as a cofactor of T-bet for enhancing IFN-γ production in iNKT cells.

T Cell Intrinsic USP15 Deficiency Promotes Excessive IFN-γ Production and an Immunosuppressive Tumor Microenvironment in MCA-Induced Fibrosarcoma.

USP15 is a deubiquitinase that negatively regulates activation of naive CD4(+) T cells and generation of IFN-γ-producing T helper 1 (Th1) cells. USP15 deficiency in mice promotes antitumor T cell responses in a transplantable cancer model; however, it has remained unclear how deregulated T cell activation impacts primary tumor development during the prolonged interplay between tumors and the immune system. Here, we find that the USP15-deficient mice are hypersensitive to methylcholantrene (MCA)-induced fibrosarcomas. Excessive IFN-γ production in USP15-deficient mice promotes expression of the immunosuppressive molecule PD-L1 and the chemokine CXCL12, causing accumulation of T-bet(+) regulatory T cells and CD11b(+)Gr-1(+) myeloid-derived suppressor cells at tumor site. Mixed bone marrow adoptive transfer studies further reveals a T cell-intrinsic role for USP15 in regulating IFN-γ production and tumor development. These findings suggest that T cell intrinsic USP15 deficiency causes excessive production of IFN-γ, which promotes an immunosuppressive tumor microenvironment during MCA-induced primary tumorigenesis.

ID: 226: DCIR maintains bone homeostasis by regulating IFN-γ production in T cells

Dendritic cell immunoreceptor (DCIR) is a C-type lectin receptor with a canonical signal motif for initiating inhibitory signals in the cytoplasmic region. In accordance with the structural insights, Dcir−/− mice spontaneously developed autoimmune-like diseases like sialadenitis and swelling and redness of joints. Thus, DCIR is a pivotal regulator for maintaining the homeostasis of the immune system. Furthermore, Dcir−/− mice not only developed ankylosis, characterized by aberrant cartilage and bone formation, but also increased bone volume, accompanied with the higher number of osteoclasts and osteoblasts (OBs). To understand the role of DCIR in bone metabolism, we dissected the molecular mechanism affecting the bone volume in Dcir−/− mice. Flow cytometry analyses revealed that Dcir−/− mice increased IFN- γ -positive T cells in peripheral blood and lymph nodes. Dcir−/− dendritic cells had higher potency to shape IFN- γ -positive T cells in the co-culture with wild-type T cells. X-ray histomorphometric analysis revealed that both Rag2−/− Dcir−/− and Ifng−/− Dcir−/− negated the increase of bone volume, suggesting that T cell-secreting IFN- γ be a pathogenic in the increased bone mass. In in vitro OB culture system, we identified IFN- γ as an osteogenic factor capable of facilitating mineralization of OBs, indicating that IFN- γ released from T cells acts on OBs. These findings demonstrate that Dcir−/− mice had increased IFN- γ -positive T cells, which accelerates mineralization of OBs, leading to the increased bone volume. Collectively, DCIR plays a key role in maintaining bone homeostasis and IFN- γ functions as an enhancing factor in bone formation.

Hexabromocyclododecane and tetrabromobisphenol A alter secretion of interferon gamma (IFN-γ) from human immune cells.

Hexabromocyclododecane (HBCD) and tetrabromobisphenol A (TBBPA) are brominated flame-retardant compounds used in a variety of applications including insulation, upholstery, and epoxy resin circuit boards. Interferon gamma (IFN-γ) is an inflammatory cytokine produced by activated T and NK cells that regulates immune responsiveness. HBCD and TBBPA are found in human blood, and previous studies have shown that they alter the ability of human natural killer (NK) lymphocytes to destroy tumor cells. This study examines whether HBCD and TBBPA affect the secretion of IFN-γ from increasingly complex preparations of human immune cells-purified NK cells, monocyte-depleted (MD) peripheral blood mononuclear cells (PBMCs), and PBMCs. Both HBCD and TBBPA were tested at concentrations ranging from 0.05 to 5µM. HBCD generally caused increases in IFN-γ secretion after 24-h, 48-h, and 6-day exposures in each of the different cell preparations. The specific concentration of HBCD that caused increases as well as the magnitude of the increase varied from donor to donor. In contrast, TBBPA tended to decrease secretion of IFN-γ from NK cells, MD-PBMCs, and PBMCs. Thus, exposure to these compounds may potentially disrupt the immune regulation mediated by IFN-γ. Signaling pathways that have the capacity to regulate IFN-γ production (nuclear factor kappa B (NF-κB), p44/42, p38, JNK) were examined for their role in the HBCD-induced increases in IFN-γ. Results showed that the p44/42 (ERK1/2) MAPK pathway appears to be important in HBCD-induced increases in IFN-γ secretion from human immune cells.

DCIR maintains bone homeostasis by regulating IFN-γ production in T cells.

Dendritic cell immunoreceptor (DCIR) is a C-type lectin receptor mainly expressed in DCs. Dcir (-/-) mice spontaneously develop autoimmune enthesitis and ankylosis accompanied by fibrocartilage proliferation and ectopic ossification. However, the mechanisms of new bone/cartilage formation in Dcir (-/-) mice remain to be elucidated. In this study, we show that DCIR maintains bone homeostasis by regulating IFN-γ production under pathophysiological conditions. DCIR deficiency increased bone volume in femurs and caused aberrant ossification in joints, whereas these symptoms were abolished in Rag2(-/-)Dcir(-/-) mice. IFN-γ-producing T cells accumulated in lymph nodes and joints of Dcir(-/-) mice, and purified Dcir(-/-) DCs enhanced IFN-γ(+) T cell differentiation. The ankylotic changes and bone volume increase were suppressed in the absence of IFN-γ. Thus, IFN-γ is a positive chondrogenic and osteoblastogenic factor, and DCIR is a crucial regulator of bone metabolism; consequently, both factors are potential targets for therapies directed against bone metabolic diseases.

Analysis of Tpl2-deficient mice reveals functional redundancies in the regulation of IFNγ production by NK cells during MCMV infection (INM8P.401)

Abstract The serine-threonine kinase, Tpl2, promotes innate inflammatory responses and is considered as a therapeutic target for autoimmunity. We have demonstrated that Tpl2 is required for efficient IFNγ secretion by CD4+ T cells and Th1 effector functions, but little is known about how Tpl2 regulates IFNγ production by other cells, including natural killer (NK) cells. Herein, we addressed the contribution of Tpl2 to the development and function of NK cells. NK cell development and cytotoxic activity occurred independently of Tpl2. In contrast, IL-12-induced IFNγ secretion was nearly abrogated in Tpl2-/- NK cells but could be restored by concomitant IL-18 stimulation. Impaired IFNγ secretion was not attributed to differences in IL-12R expression or Stat4-dependent signals. Furthermore, only a modest reduction in IL-12-induced IFNγ mRNA expression was observed, suggesting that Tpl2 regulates IFNγ production primarily via translational control. Consistent with in vitro findings, Tpl2-/- mice mounted a diminished NK-derived systemic IFNγ response to murine cytomegalovirus (MCMV) infection, which could be compensated for in ex vivo restimulation assays by the presence of MCMV infection-induced IL-18 secretion. These data demonstrate the importance of Tpl2 in IL-12-induced NK cell IFNγ secretion. Furthermore, the synergism and functional redundancy of the IL-18 pathway with the IL-12-Tpl2 pathway underscores the criticality of ensuring appropriate NK-derived IFNγ to anti-viral immunity.

Mechanisms of Interferon-γ Production by Neutrophils and Its Function duringStreptococcus pneumoniaePneumonia

Bacterial pneumonia is a common public health problem associated with significant mortality, morbidity, and cost. Neutrophils are usually the earliest leukocytes to respond to bacteria in the lungs. Neutrophils rapidly sequester in the pulmonary microvasculature and migrate into the lung parenchyma and alveolar spaces, where they perform numerous effector functions for host defense. Previous studies showed that migrated neutrophils produce IFN-γ early during pneumonia induced by Streptococcus pneumoniae and that early production of IFN-γ regulates bacterial clearance. IFN-γ production by neutrophils requires Rac2, Hck/Lyn/Fgr Src family tyrosine kinases, and NADPH oxidase. Our current studies examined the mechanisms that regulate IFN-γ production by lung neutrophils during acute S. pneumoniae pneumonia in mice and its function. We demonstrate that IFN-γ production by neutrophils is a tightly regulated process that does not require IL-12. The adaptor molecule MyD88 is critical for IFN-γ production by neutrophils. The guanine nucleotide exchange factor CalDAG-GEFI modulates IFN-γ production. The CD11/CD18 complex, CD44, Toll-like receptors 2 and 4, TRIF, and Nrf2 are not required for IFN-γ production by neutrophils. The recently described neutrophil-dendritic cell hybrid cell, identified by its expression of Ly6G and CD11c, is present at low numbers in pneumonic lungs and is not a source of IFN-γ. IFN-γ produced by neutrophils early during acute S. pneumoniae pneumonia induces transcription of target genes in the lungs, which are critical for host defense. These studies underline the complexity of the neutrophil responses during pneumonia in the acute inflammatory response and in subsequent resolution or initiation of immune responses.

Epigenetic control of interferon-gamma expression in CD8 T cells.

Interferon- (IFN-) γ is an essential cytokine for immunity against intracellular pathogens and cancer. IFN-γ expression by CD4 T lymphocytes is observed only after T helper (Th) 1 differentiation and there are several studies about the molecular mechanisms that control Ifng expression in these cells. However, naïve CD8 T lymphocytes do not produce large amounts of IFN-γ, but after TCR stimulation there is a progressive acquisition of IFN-γ expression during differentiation into cytotoxic T lymphocytes (CTL) and memory cells, which are capable of producing high levels of this cytokine. Differential gene expression can be regulated from the selective action of transcriptional factors and also from epigenetic mechanisms, such as DNA CpG methylation or posttranslational histone modifications. Recently it has been recognized that epigenetic modification is an integral part of CD8 lymphocyte differentiation. This review will focus on the chromatin status of Ifng promoter in CD8 T cells and possible influences of epigenetic modifications in Ifng gene and conserved noncoding sequences (CNSs) in regulation of IFN-γ production by CD8 T lymphocytes.

Identification of genetic markers of susceptibility to Tuberculosis in signalling pathways that regulates IFN-γ production.

Identification of genetic markers of susceptibility to Tuberculosis in signalling pathways that regulates IFN-γ production.