Cancer-associated fibroblasts: Vital suppressors of the immune response in the tumor microenvironment

Since the "seed and soil" hypothesis was proposed, the biological functions of the tumor microenvironment (TME), especially its stromal components, have received increasing attention. Cancer-associated fibroblasts (CAFs) are the major components of the stromal region, providing material support for tumor cell proliferation, migration, and invasion. Furthermore, CAFs are important mediators of suppressing immune responses by attracting the accumulation of immunosuppressive cells through cytokine/chemokine secretion. In this review, we summarized the major cytokines, chemokines and metabolites, including transforming growth factor-β (TGF-β), interleukin-6 (IL-6), C-X-C chemokine ligand (CXCL)12, C–C chemokine ligand (CCL) 2, prostaglandin E2 (PGE2), and other factors, by which CAFs suppress the immune systems in a variety of cancers. More importantly, we highlight potential therapeutic strategies to alleviate the immunosuppression produced by CAFs, thereby inhibiting tumor progression.     

Human intestinal mast cells are a potent source of multiple chemokines

Mast cells are key effector cells of immediate type allergic reactions. Upon activation they release a broad array of pre-stored and de novo synthesized mediators including immunoregulatory cytokines and chemokines. Here, we analyzed the chemokine profile expressed by mature human mast cells. Human mast cells were isolated from intestinal tissue and cultured with stem cell factor (SCF) in the presence or absence of IL-4 for 10d. Cells were stimulated by cross-linking of the high affinity IgE receptor (FcεRI) and/or by SCF. Chemokine and chemokine receptor mRNA expression was determined by real-time RT-PCR and chemokine release was measured by multiplex bead immunoassay. Out of 43 chemokines and 19 chemokine receptors human intestinal mast cells express 27 chemokines and nine chemokine receptors. Twelve chemokines (CCL1, CCL2, CCL3, CCL4, CCL5, CCL7, CCL18, CCL20, CXCL2, CXCL3, CXCL8, and XCL1) were more than four-fold up-regulated in response to FcεRI cross-linking. Combination of pre-culture with IL-4 and/or stimulation with SCF in addition to FcεRI cross-linking further increased the antigen-dependent expression of mRNA for most chemokines. In contrast, the expression of CCL20, CXCL2, and CXCL3 was strongly inhibited by IL-4 treatment. In conclusion, human intestinal mast cells express a broad spectrum of different chemokines underlining their important role as immunoregulatory cells. Furthermore, combined treatment with IL-4 and SCF increases the antigen-mediated expression and release of multiple chemokines, but IL-4 priming inhibits the expression of CCL20, CXCL2, and CXCL3.     

Variability in chemokine-induced adhesion of human mesenchymal stromal cells

BACKGROUND AIMS. Intravenously applied mesenchymal stromal cells (MSC) are under investigation for numerous clinical indications. However, their capacity to activate shear stress-dependent adhesion to endothelial ligands is incompletely characterized. METHODS. Parallel-plate flow chambers were used to induce firm adhesion of MSC to integrin ligand vascular cell adhesion molecule (VCAM)-1. Human MSC were stimulated by chemokine (C-C motif) ligand (CCL15)/macrophage inflammatory protein (MIP-5), CCL19/MIP-3β chemokine (C-X-C motif) ligand (CXCL8)/interleukin (IL)-8, CXCL12/ stromal derived factor (SDF-1) or CXCL13/B lymphocyte chemoattractant (BLC). RESULTS. Two MSC isolates responded to three chemokines (either to CCL15, CCL19 and CXCL13, or to CCL19, CXCL12 and CXCL13), two isolates responded to two chemokines (to CCL15 and CCL19, or to CCL19 and CXCL13), and one isolate responded to CCL19 only. In contrast, all tested MSC isolates responded to selectins (P-selectin and E-selectin) or integrin ligand VCAM-1, as visualized by a velocity reduction under flow. CONCLUSIONS. Inter-individual variability of chemokine-induced integrin activation should be considered when evaluating human MSC as cellular therapies.     

Interleukin-1beta induces interleukin-6 production through the production of prostaglandin E(2) in human osteoblasts, MG-63 cells.

This study was conducted to investigate the mechanism of interleukin-1beta (IL-1beta)-induced IL-6 production in human osteoblasts (MG-63 cells). Stimulation with IL-1beta resulted in the production of IL-6 and prostaglandin E(2) (PGE(2)). IL-6 production gradually increased and peaked 96 h after stimulation. IL-6 mRNA was detected between 4 and 72 h after IL-1beta stimulation. The patterns of PGE(2) production and the expression of cyclooxygenase-2 (COX-2) mRNA were biphasic after stimulation. Actinomycin D, cycloheximide, indomethacin, and NS-398 (COX-2 inhibitor) suppressed the production of IL-6 and PGE(2). Anti-PGE(2) antibody markedly reduced the production of IL-6. In addition, stimulation with 17-phenyl-PGE(2), a PGE receptor-1 (EP-1 receptor) agonist, led to the expression of IL-6 mRNA after pretreatment with IL-1beta. These findings indicate that IL-1beta-induced IL-6 production in MG-63 cells involves the following sequence of steps: IL-1beta-induced COX-2 activation, PGE(2) production, and EP-1 receptor signaling prior to IL-6 production.     

Chemokines regulate small leucine-rich proteoglycans in the extracellular matrix of the pressure-overloaded right ventricle

Chemokines have been suggested to play a role during development of left ventricular failure, but little is known about their role during right ventricular (RV) remodeling and dysfunction. We have previously shown that the chemokine (C-X-C motif) ligand 13 (CXCL13) regulates small leucine-rich proteoglycans (SLRPs). We hypothesized that chemokines are upregulated in the pressure-overloaded RV, and that they regulate SLRPs. Mice with RV pressure overload following pulmonary banding (PB) had a significant increase in RV weight and an increase in liver weight after 1 wk. Microarray analysis (Affymetrix) of RV tissue from mice with PB revealed that CXCL10, CXCL6, chemokine (C-X3-C motif) ligand 1 (CX3CL1), chemokine (C-C motif) ligand 5 (CCL5), CXCL16, and CCL2 were the most upregulated chemokines. Stimulation of cardiac fibroblasts with these same chemokines showed that CXCL16 increased the expression of the four SLRPs: decorin, lumican, biglycan, and fibromodulin. CCL5 increased the same SLRPs, except decorin, whereas CX3CL1 increased the expression of decorin and lumican. CXCL16, CX3CL1, and CCL5 were also shown to increase the levels of glycosylated decorin and lumican in the medium after stimulation of fibroblasts. In the pressure-overloaded RV tissue, Western blotting revealed an increase in the total protein level of lumican and a glycosylated form of decorin with a higher molecular weight compared with control mice. Both mice with PB and patients with pulmonary stenosis had significantly increased circulating levels of CXCL16 compared with healthy controls measured by enzyme immunoassay. In conclusion, we have found that chemokines are upregulated in the pressure-overloaded RV and that CXCL16, CX3CL1, and CCL5 regulate expression and posttranslational modifications of SLRPs in cardiac fibroblasts. In the pressure-overloaded RV, protein levels of lumican were increased, and a glycosylated form of decorin with a high molecular weight appeared.     

Differing activities of homeostatic chemokines CCL19, CCL21, and CXCL12 in lymphocyte and dendritic cell recruitment and lymphoid neogenesis

Despite their widespread expression, the in vivo recruitment activities of CCL19 (EBV-induced molecule 1 ligand chemokine) and CXCL12 (stromal cell-derived factor 1) have not been established. Furthermore, although CXCL13 (B lymphocyte chemoattractant) has been shown to induce lymphoid neogenesis through induction of lymphotoxin (LT)alpha1beta2, it is unclear whether other homeostatic chemokines have this property. In this work we show that ectopic expression in pancreatic islets of CCL19 leads to small infiltrates composed of lymphocytes and dendritic cells and containing high endothelial venules and stromal cells. Ectopic CXCL12 induced small infiltrates containing few T cells but enriched in dendritic cells, B cells, and plasma cells. Comparison of CCL19 transgenic mice with mice expressing CCL21 (secondary lymphoid tissue chemokine) revealed that CCL21 induced larger and more organized infiltrates. A more significant role for CCL21 is also suggested in lymphoid tissues, as CCL21 protein was found to be present in lymph nodes and spleen at much higher concentrations than CCL19. CCL19 and CCL21 but not CXCL12 induced LTalpha1beta2 expression on naive CD4 T cells, and treatment of CCL21 transgenic mice with LTbetaR-Fc antagonized development of organized lymphoid structures. LTalpha1beta2 was also induced on naive T cells by the cytokines IL-4 and IL-7. These studies establish that CCL19 and CXCL12 are sufficient to mediate cell recruitment in vivo and they indicate that LTalpha1beta2 may function downstream of CCL21, CCL19, and IL-2 family cytokines in normal and pathological lymphoid tissue development.     

Interleukin-1 regulates FGF-2 mRNA and localization of FGF-2 protein in human osteoblasts

Interleukin-1 (IL-1) and basic fibroblast growth factor (FGF-2) are potent stimulators of osteoclast formation. However, the role of FGF-2 in the responses to IL-1 in bone has not been reported. We examined the effect of IL-1 on FGF-2 mRNA and protein expression in human osteosarcoma MG-63 osteoblasts, normal human osteoblasts (NHOB), and osteoblasts from osteoarthritic patients (F2 and F13). IL-1 increased FGF-2 mRNA expression in osteoblasts within 1.5 to 3 h. Multiple FGF-2 protein isoforms were expressed in human osteoblasts. Twenty-four hours of treatment of MG-63 and NHOB cells with IL-1 increased the high-molecular-weight(HMW, 22/24 kDa) and low-molecular-weight (LMW, 18 kDa) FGF-2 proteins intracellularly. In contrast, IL-1 preferentially increased the LMW protein signal intracellularly as well as on the cell surface of F2 and F13 osteoblasts. We conclude that IL-1 is a major stimulator of FGF-2 expression in human osteoblasts. Furthermore, selective increases in the exportable LMW protein in osteoblasts from osteoarthritic patients may be of clinical relevance.     

Monocyte-astrocyte networks and the regulation of chemokine secretion in neurocysticercosis

Neurocysticercosis, caused by infection with larval Taenia solium, is a major cause of epilepsy worldwide. Larval degeneration, which is symptomatic, results in inflammatory cell influx. Astrocytes, the most abundant cell type and major cytokine-producing cell within the CNS, may be important in orchestrating inflammatory responses after larval degeneration. We investigated the effects of direct stimulation and of conditioned medium from T. solium larval Ag (TsAg)-stimulated monocytes (CoMTsAg) on neutrophil and astrocyte chemokine release. CoMTsAg, but not control conditioned medium, stimulated astrocyte CCL2/MCP-1 (161.5 +/- 16 ng/ml), CXCL8/IL-8 (416 +/- 6.2 ng/ml), and CXCL10/IFN-gamma-inducible protein (9.07 +/- 0.6 ng/ml) secretion after 24 h, whereas direct astrocyte or neutrophil stimulation with TsAg had no effect. There was rapid accumulation of CCL2 and CXCL8 mRNA within 1 h, with somewhat delayed expression of CXCL10 mRNA initially detected 8 h poststimulation. Neutralizing anti-TNF-alpha inhibited CoMTsAg-induced CCL2 mRNA accumulation by up to 99%, causing total abolition of CXCL10 and up to 77% reduction in CXCL8 mRNA. CoMTsAg induced maximal nuclear binding of NF-kappaB p65 and p50 by 1 h, with IkappaBalpha and IkappaBbeta decay within 15 min. In addition, CoMTsAg induced transient nuclear binding of AP-1, which peaked 4 h poststimulation. In NF-kappaB blocking experiments using pyrrolidine dithiocarbamate, CoMTsAg-induced CCL2 secretion was reduced by up to 80% (p = 0.0006), whereas CXCL8 was inhibited by up to 75% (p = 0.0003). In summary, the data show that astrocytes are an important source of chemokines following larval Ag stimulation. Such chemokine secretion is NF-kappaB dependent, likely to involve AP-1, and is regulated in a paracrine loop by monocyte-derived TNF-alpha.     

Stimulation of prostaglandin E2 (PGE2) production by arachidonic acid, oestrogen and parathyroid hormone in MG-63 and MC3T3-E1 osteoblast-like cells

Polyunsaturated fatty acids (PUFAs) as well as oestrogen (E2) and parathyroid hormone (PTH) affect bone cells. The aim of the study was to determine whether arachidonic acid (AA), E2, and PTH increase prostaglandin E₂ (PGE₂) synthesis in MG-63 and MC3T3-E1 osteoblastic cells and the level of mediation by COX-1 and COX-2. PGE₂ levels were determined in the conditioned culture media of MG-63 and MC3T3-E1 osteoblasts after exposure to AA, PTH and E2. Cells were pre-incubated in some experiments with the unselective COX inhibitor indomethacin or the COX-2 specific blocker NS-398. Indirect immunofluorescence was performed on MG-63 cells to detect the presence and location of the two enzymes involved. AA increased PGE₂ secretion in both cell lines; production by MC3T3-E1 cells, however, was significantly higher than that of MG-63 cells. This could be due to autoamplification via the EP₁ subtype of PGE receptors in mouse MC3T3-E1 osteoblasts. Both COX-1 and COX-2 affected the regulation of PGE₂ synthesis in MG-63 cells. E2 had no effect on PGE₂ secretion in both cell lines, while PTH caused a slight increase in PGE₂ synthesis in the MG-63 cell line.     

Modulatory effects of luteolin on osteoblastic function and inflammatory mediators in osteoblastic MC3T3-E1 cells

The effects of luteolin on the function of osteoblastic MC3T3-E1 cells and the production of local factors in osteoblasts were investigated. Luteolin (1microM) caused a significant elevation of collagen content, alkaline phosphatase (ALP) activity, and osteocalcin secretion in the cells (P<0.05). The effect of luteolin in increasing collagen content and ALP activity was completely prevented by the presence of 10(-6)M cycloheximide and 10(-6)M tamoxifen, suggesting that luteolin's effect results from a newly synthesized protein component and might be partly involved in estrogen action. We then examined the effect of luteolin on the 3-morpholinosydnonimine (SIN-1)-induced production of oxidative stress markers [nitric oxide (NO) and prostaglan E(2) (PGE(2))] and cytokines [tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6)] in osteoblasts. Luteolin (1 and 10microM) decreased the SIN-1-induced production of NO, PGE(2), TNF-alpha, and IL-6 in osteoblasts. These results suggest that inflammatory mediators can be regulated by luteolin stimulating osteoblastic function.     

Modulation by soluble factors of gelatinase activities released by osteoblastic cells

This study investigated the ability of normal human osteoblasts (hOb) and osteogenic sarcoma cells (MG-63 and SaOS2) to produce gelatinases and undergo modulation by interleukin 1beta (IL-1beta), interleukin 6 (IL-6), oncostatin M (OSM), leukaemia inhibitory factor (LIF), growth hormone (GH) and insulin-like growth factor-I (IGF-I). Gelatinase activities were determined by zymogaphy, and a quantitative analysis was performed by ELISA. The MMP-2 activities of the three cell lines were significantly increased in the presence of IL-1beta and IL-6, but no modulation of MMP-2 activities was observed in the presence of OSM, LIF and GH. IGF-I increased the activity released by SaOS2 and hOb, but no modulation was detectable in MG-63 cell conditioned medium. An upmodulation of pro-MMP-2 secretion by SaOS2 and hOb was observed for all soluble factors used, whereas an upmodulation of pro-MMP-2 secretion by MG-63 was observed only in the presence of IL-1beta, IL-6 and IGF-I. Thus, osteoblastic cells modulated by cytokines can be involved in bone resorption as a result of the protease activities released.     

Azithromycin treatment modulates the extracellular signal-regulated kinase mediated pathway and inhibits inflammatory cytokines and chemokines in epithelial cells from infertile women with recurrent Chlamydia trachomatis infection

Epidemiological and animal model studies suggest that sequelae of genital Chlamydia trachomatis infection are more often associated with second or subsequent infections than with initial infection. Further, in order to establish an acute or long-term persistent infection, C. trachomatis develops several strategies to circumvent host immune responses. Hence, resolution of the C. trachomatis infection may require modulation of host factors especially during persistent or chronic infection. Moreover, azithromycin treatment has been reported to possess anti-inflammatory properties but its mechanism of action is still not elucidated. Therefore, in order to better understand the effect of azithromycin in chronic conditions, our aim was to study changes in expression of key genes associated with inflammatory cytokines and receptors, mitogen-activated protein kinase (MAPK) signaling pathway, and apoptosis pathway before and after therapy with azithromycin in infertile women with recurrent C. trachomatis infection. Real-time polymerase chain reaction was performed to study inflammatory cytokines and receptors, MAPK signaling pathway, and apoptosis pathway before and after therapy with azithromycin in infertile women with recurrent C. trachomatis infection. Further, effect of azithromycin on activation of extracellular signal-regulated kinase was studied in epithelial cells by western blotting. Chemokine (C-C motif) ligand 2 (CCL2), CCL5, chemokine (C-X-C motif) ligand 1 (CXCL1), CXCL5, CXCL9, interleukin-1B (IL-1B), IL-8, baculoviral IAP repeat-containing 3 (BIRC3), myeloid cell leukemia sequence 1 (MCL1), and MAPK1 were downregualted after azithromycin treatment. In addition, phosphorylation of extracellular signal-regulated kinase was inhibited after azithromycin treatment in epithelial cells obtained from women with recurrent infection. Hence, our data suggest that azithromycin with its properties apart from antibacterial activity may contribute to its therapeutic potential in treatment of chronic recurrent infection in infertile women.     

Peroxisome proliferator-activated receptor α agonists modulate Th1 and Th2 chemokine secretion in normal thyrocytes and Graves' disease

Until now, no data are present about the effect of peroxisome proliferator-activated receptor (PPAR)α activation on the prototype Th1 [chemokine (C–X–C motif) ligand (CXCL)10] (CXCL10) and Th2 [chemokine (C–C motif) ligand 2] (CCL2) chemokines secretion in thyroid cells.The role of PPARα and PPARγ activation on CXCL10 and CCL2 secretion was tested in Graves' disease (GD) and control primary thyrocytes stimulated with interferon (IFN)γ and tumor necrosis factor (TNF)α.IFNγ stimulated both CXCL10 and CCL2 secretion in primary GD and control thyrocytes. TNFα alone stimulated CCL2 secretion, while had no effect on CXCL10. The combination of IFNγ and TNFα had a synergistic effect both on CXCL10 and CCL2 chemokines in GD thyrocytes at levels comparable to those of controls. PPARα activators inhibited the secretion of both chemokines (stimulated with IFNγ and TNFα) at a level higher (for CXCL10, about 60–72%) than PPARγ agonists (about 25–35%), which were confirmed to inhibit CXCL10, but not CCL2.Our data show that CCL2 is modulated by IFNγ and TNFα in GD and normal thyrocytes. Furthermore we first show that PPARα activators inhibit the secretion of CXCL10 and CCL2 in thyrocytes, suggesting that PPARα may be involved in the modulation of the immune response in the thyroid.     

Stimulation of oral fibroblast chemokine receptors identifies CCR3 and CCR4 as potential wound healing targets

The focus of this study was to determine which chemokine receptors are present on oral fibroblasts and whether these receptors influence proliferation, migration, and/or the release of wound healing mediators. This information may provide insight into the superior wound healing characteristics of the oral mucosa. The gingiva fibroblasts expressed 12 different chemokine receptors (CCR3, CCR4, CCR6, CCR9, CCR10, CXCR1, CXCR2, CXCR4, CXCR5, CXCR7, CX3CR1, and XCR1), as analyzed by flow cytometry. Fourteen corresponding chemokines (CCL5, CCL15, CCL20, CCL22, CCL25, CCL27, CCL28, CXCL1, CXCL8, CXCL11, CXCL12, CXCL13, CX3CL1, and XCL1) were used to study the activation of these receptors on gingiva fibroblasts. Twelve of these fourteen chemokines stimulated gingiva fibroblast migration (all except for CXCL8 and CXCL12). Five of the chemokines stimulated proliferation (CCL5/CCR3, CCL15/CCR3, CCL22/CCR4, CCL28/CCR3/CCR10, and XCL1/XCR1). Furthermore, CCL28/CCR3/CCR10 and CCL22/CCR4 stimulation increased IL‐6 secretion and CCL28/CCR3/CCR10 together with CCL27/CCR10 upregulated HGF secretion. Moreover, TIMP‐1 secretion was reduced by CCL15/CCR3. In conclusion, this in‐vitro study identifies chemokine receptor‐ligand pairs which may be used in future targeted wound healing strategies. In particular, we identified the chemokine receptors CCR3 and CCR4, and the mucosa specific chemokine CCL28, as having an predominant role in oral wound healing by increasing human gingiva fibroblast proliferation, migration, and the secretion of IL‐6 and HGF and reducing the secretion of TIMP‐1.     

CXCL8 and CCL20 Enhance Osteoclastogenesis via Modulation of Cytokine Production by Human Primary Osteoblasts

Generalized osteoporosis is common in patients with inflammatory diseases, possibly because of circulating inflammatory factors that affect osteoblast and osteoclast formation and activity. Serum levels of the inflammatory factors CXCL8 and CCL20 are elevated in rheumatoid arthritis, but whether these factors affect bone metabolism is unknown. We hypothesized that CXCL8 and CCL20 decrease osteoblast proliferation and differentiation, and enhance osteoblast-mediated osteoclast formation and activity. Human primary osteoblasts were cultured with or without CXCL8 (2–200 pg/ml) or CCL20 (5–500 pg/ml) for 14 days. Osteoblast proliferation and gene expression of matrix proteins and cytokines were analyzed. Osteoclast precursors were cultured with CXCL8 (200 pg/ml) and CCL20 (500 pg/ml), or with conditioned medium (CM) from CXCL8 and CCL20-treated osteoblasts with or without IL-6 inhibitor. After 3 weeks osteoclast formation and activity were determined. CXCL8 (200 pg/ml) and CCL20 (500 pg/ml) enhanced mRNA expression of KI67 (2.5–2.7-fold), ALP (1.6–1.7-fold), and IL-6 protein production (1.3–1.6-fold) by osteoblasts. CXCL8-CM enhanced the number of osteoclasts with 3–5 nuclei (1.7-fold), and with >5 nuclei (3-fold). CCL20-CM enhanced the number of osteoclasts with 3–5 nuclei (1.3-fold), and with >5 nuclei (2.8-fold). IL-6 inhibition reduced the stimulatory effect of CXCL8-CM and CCL20-CM on formation of osteoclasts. In conclusion, CXCL8 and CCL20 did not decrease osteoblast proliferation or gene expression of matrix proteins. CXCL8 and CCL20 did not directly affect osteoclastogenesis. However, CXCL8 and CCL20 enhanced osteoblast-mediated osteoclastogenesis, partly via IL-6 production, suggesting that CXCL8 and CCL20 may contribute to osteoporosis in rheumatoid arthritis by affecting bone cell communication.     

Cutting edge: TLR2-mediated proinflammatory cytokine and chemokine production by microglial cells in response to herpes simplex virus

Recent studies indicate that TLRs are critical in generating innate immune responses during infection with HSV-1. In this study, we investigated the role of TLR2 signaling in regulating the production of neuroimmune mediators by examining cytokine and chemokine expression using primary microglial cells obtained from TLR2-/- as well as wild-type mice. Data presented here demonstrate that TLR2 signaling is required for the production of proinflammatory cytokines and chemokines: TNF-alpha, IL-1beta, IL-6, IL-12, CCL7, CCL8, CCL9, CXCL1, CXCL2, CXCL4, and CXCL5. CXCL9 and CXCL10 were also induced by HSV, but their production was not dependent upon TLR2 signaling. Because TLR2-/- mice display significantly reduced mortality and diminished neuroinflammation in response to brain infection with HSV, the TLR2-dependent cytokines identified here might function as key players influencing viral neuropathogenesis.     

Metabolism of nitric oxide by Neisseria meningitidis modifies release of NO-regulated cytokines and chemokines by human macrophages

Macrophages produce nitric oxide (NO) via the inducible nitric oxide synthase as part of a successful response to infection. The gene norB of Neisseria meningitidis encodes a NO reductase which enables utilization and consumption of NO during microaerobic respiration and confers resistance to nitrosative stress-related killing by human monocyte-derived macrophages (MDM). In this study we confirmed that NO regulates cytokine and chemokine release by resting MDM: accumulation of TNF-alpha, IL-12, IL-10, CCL5 (RANTES) and CXCL8 (IL-8) in MDM supernatants was significantly modified by the NO-donor S-nitroso-N-penicillamine (SNAP). Using a protein array, infection of MDM with N. meningitidis was shown to be associated with secretion of a wide range of cytokines and chemokines. To test whether NO metabolism by N. meningitidis modifies release of NO-regulated cytokines, we infected MDM with wild-type organisms and an isogenic norB strain. Resulting expression of the cytokines TNF-alpha and IL-12, and the chemokine CXCL8 was increased and production of the cytokine IL-10 and the chemokine CCL5 was decreased in norB-infected MDM, in comparison to wild-type. Addition of SNAP to cultures infected with wild-type mimicked the effect observed in cultures infected with the norB mutant. In conclusion, NorB-catalysed removal of NO modifies cellular release of NO-regulated cytokines and chemokines.