Control by IFN-gamma and PGE2 of TNF alpha and IL-1 production by human monocytes.

There have been suggestions that the production of pro-inflammatory mediators by human monocytes in response to interferon-gamma (IFN-gamma) may be controlled by changes in prostaglandins. Therefore we investigated tumour necrosis factor alpha (TNF alpha) and interleukin-1 (IL-1) activities and prostaglandin E2 (PGE2) levels in the supernatants of highly purified human monocytes cultured for 18 hr with recombinant human IFN-gamma. IFN-gamma (100 U/ml) did not stimulate monocytes isolated by counter-current centrifugal elutriation for detectable TNF alpha or IL-1 activities, or PGE2 production. However, IFN-gamma synergistically enhanced lipopolysaccharide (LPS)-induced TNF alpha and IL-1 activities. In contrast, there was no consistent change in PGE2 levels upon addition of IFN-gamma to LPS-treated monocyte cultures. The TNF alpha and IL-1 activities induced by LPS and by LPS with IFN-gamma were reduced by PGE2, and stimulated by indomethacin. As reported previously for IL-1 activities, the regulation by cyclo-oxygenase products of TNF alpha activities reflected predominantly a control of the production of immunoreactive TNF alpha, rather than the measurement of TNF alpha bio-activity. However, the addition of indomethacin or PGE2 to monocyte cultures did not change the extent of IFN-gamma synergy with LPS for increased TNF alpha and IL-1 activities. The results of this study suggest that, despite control by cyclo-oxygenase products of TNF alpha and IL-1 production in human monocytes, IFN-gamma may enhance TNF alpha and IL-1 activities independently of this regulatory mechanism. These findings are contrary to those suggested for the regulation by prostanoids of IL-1 production by murine macrophages.     

Interleukin-2 augmentation of interleukin-1 and prostaglandin E2 production

Some of the major side effects of interleukin-2 (IL-2) therapy in the treatment of malignancies may be related to increased interleukin-1 (IL-1) and/or prostaglandin E2 (PGE2) production. We examined the effect of recombinant (rIL-2) on the in vitro production of IL-1 beta and PGE2 by unstimulated and LPS-activated human blood mononuclear cells (PBMC). We also compared the effect of rIL-2 on IL-1 beta production by adherent and nonadherent blood mononuclear cell populations. Cultures of PBMC (5 x 10(6)/ml) were incubated for 24 hr in media only (control, 1,000 U/ml rIL-2, 2 micrograms/ml LPS, or both LPS and rIL-2. Supernatants obtained from these cultures were analyzed for levels of IL-1 beta and PGE2 by radioimmunoassays. The addition of rIL-2 caused an increase in IL-1 beta production in 13 of 13 control PBMC cultures and in 11 of 13 LPS-stimulated cultures, which were significant increases as determined by paired t tests. When PBMC were fractionated into plastic adherent and nonadherent populations, the rIL-2 induced increases in IL-1 beta production were more consistent in control (six of seven cases) and LPS (seven of seven cases) cultures of plastic nonadherent cells than in control (three of seven cases) and LPS (four of seven cases) cultures of plastic adherent cells. Recombinant IL-2 did not increase PGE2 production in control PBMC cultures (none of four cases), but did so in LPS-stimulated PBMC cultures (three of four cases]. These results suggest that rIL-2 may increase IL-1 production in vivo and thus possibly account for some of the side effects of this therapy.     

Monophosphoryl lipid A (MPL) promotes allergen-induced immune deviation in favour of Th1 responses

BACKGROUND: Monophosphoryl lipid A (MPL) is a nontoxic derivative of the lipopolysaccharide (LPS) of Salmonella minnesota R595. MPL has been used as an adjuvant in grass and tree pollen vaccines for the treatment of seasonal allergic rhinitis. Little is known about the influence of MPL on cellular responses to allergens in man. We therefore studied the effects of MPL in vitro on peripheral blood mononuclear cells (PBMC) obtained from patients with grass pollen hay fever. METHODS: The PBMCs from 13 subjects were cultured with grass pollen Phleum pratense extract (0, 2 and 20 microg/ml) and MPL (0 and 10 microg/ml; defined as an optimal concentration in preliminary studies) and after 6 days proliferative responses were measured by thymidine incorporation and cytokine production by enzyme-linked immunosorbent assay (ELISA). RESULTS: Proliferative responses were unaffected by the presence of MPL whereas MPL induced a significant increase in allergen-induced interferon (IFN)-gamma production [allergen alone, 645 +/- 466 pg/ml (mean +/- SE) vs allergen + MPL, 3232 +/- 818 pg/ml; P < 0.001]. In addition, there was a significant decrease in interleukin (IL)-5 production (4307 +/- 1030 pg/ml vs 2997 +/- 826 pg/ml; P < 0.01). Although MPL alone could induce modest increases in IL-10 production, MPL did not influence the production of this cytokine in allergen-stimulated cultures. Addition of neutralizing antibody against IL-12 resulted in 95% inhibition of MPL-induced IFN-gamma production. Depletion of monocytes from the culture system abrogated the effects of MPL on elevated cytokine production. CONCLUSIONS: In summary, use of MPL with grass pollen extract results in immune deviation of allergen-induced peripheral Th2-cell responses in favour of 'protective' Th1 responses in an IL-12 and monocyte-dependent fashion.     

Effect of LACK and KMP11 on IFN-gamma production by peripheral blood mononuclear cells from cutaneous and mucosal leishmaniasis patients

The immune modulatory properties of recombinant antigens Kinetoplasmid membrane protein-11 (KMP11) and Leishmania homologue of receptors for activated C kinase (LACK) in cutaneous leishmaniasis (CL) and mucosal leishmaniasis (ML) patients were evaluated. The mean levels of interferon-gamma (IFN-gamma) in soluble leishmania antigen (SLA) stimulated peripheral blood mononuclear cells (PBMC) of ML and CL patients were 5625 +/- 2333 pg/ml and 4422 +/- 3665 pg/ml, respectively. IFN-gamma was not detected in cultures stimulated with KMP11 or LACK. Interleukin-10 (IL-10) concentration in SLA, KMP11 and LACK-stimulated PBMC of ML patients was 13 +/- 12 pg/ml, 285 +/- 388 pg/ml and 802 +/- 483 pg/ml, respectively. Addition of KMP11 or LACK to SLA-stimulated PBMC of CL and ML patients enhanced IL-10 production (P < 0.05). Addition of KMP11 decreased IFN-gamma levels by 52% in CL patients and by 19% in ML patients. Addition of LACK to SLA-stimulated cultures decreased IFN-gamma levels by 58% in CL patients and by 30% in ML patients. Neutralization of IL-10 abrogated the downregulatory effect of LACK and KMP11. The modulatory properties of LACK and KMP11 are due to induction of IL-10 production and may be helpful for attenuating chronic inflammatory diseases. However, in some clinical conditions, as demonstrated for ML, these molecules are not able to suppress the IFN-gamma response, even inducing IL-10 production.     

Endotoxin-Stimulated Human Monocyte Secretion of Interleukin 1, Tumour Necrosis Factor Alpha, and Prostaglandin E2 Shows Stable Interindividual Differences

The secretions of interleukin 1 (IL-1), tumour necrosis factor alpha (TNF), and prostaglandin E2 (PGE2) of low-dose E. coli lipopolysaccharide (LPS)-stimulated human monocytes (M phi) were investigated in an endotoxin (ET)-free milieu (less than 1.6 pg LPS/ml). Human M phi cultures from nine healthy men were stimulated with 0, 12.5-500, and 250,000 pg LPS/ml as measured by a very sensitive Limulus test. The IL-1 activity was tested by the mouse costimulatory thymocyte (LAF) assay, which was thoroughly standardized and characterized (interassay variation 22-24%, intra-assay variation 3-7%). Spontaneous M phi secretions of IL-1, TNF, and PGE2 were negligible, but 12.5 pg LPS/ml significantly stimulated the secretions of these M phi products and the monokine responses to 500 and 250,000 pg LPS/ml were almost in the same range. It was demonstrated that the secretions of IL-1-TNF and TNF-PGE2 were strongly correlated. Pronounced interindividual differences in LPS responsiveness were demonstrated, and two low-responders, one of whom was HLA-DR1,2-positive, were identified. Three first-degree relatives of the DR1,2-positive low-responder had similar low responses. Furthermore, M phi cultures were prepared weekly for 4 weeks from four HLA-DR different men and the only DR2,2 homozygous individual had low monokine responses. In conclusion, stable interindividual differences in in vitro monokine and PGE2 secretions of LPS-stimulated M phi were demonstrated. It is suggested that HLA-DR2-positive individuals may be low responders.     

Expression of IL-18 by Mycobacterium avium-infected human monocytes; association with M. avium virulence

Disseminated Mycobacterium avium infection is the most frequent bacterial infection in patients with advanced AIDS and also associated with interferon-gamma (IFN-gamma) or IL-12 receptor deficiency. IFN-gamma is a key cytokine in host defence against M. avium infection. Expression of IL-18, a potent IFN-gamma inducer, and IFN-gamma by human monocytes after infection with M. avium was examined. Monocytes were co-cultured with isogenic smooth-transparent (SmT: virulent) or smooth-domed (SmD: avirulent) M. avium strains (10 organisms per monocyte). Infection with the SmD strain induced significantly higher concentration of IL-18 and IFN-gamma in culture supernatants than did the SmT strain. IFN-gamma production in response to M. avium was partially inhibited by anti-human IL-18 MoAb. Both recombinant human IL-12 (77 +/- 42 pg/ml, control versus 1492 +/- 141 pg/ml, cultures with IL-12 1 ng/ml) and IL-18 (126 +/- 37 pg/ml, control versus 2683 +/- 864 pg/ml, cultures with IL-18 10 ng/ml) augmented M. avium-induced IFN-gamma production. Freshly isolated uninfected monocytes expressed constitutive levels of IL-18. Following infection with M. avium, enhancement of IL-18 mRNA expression peaked at 3-6 h. IL-18 protein was detected in monocyte lysates as early as 1 h after infection with both SmT and SmD M. avium strains by Western blotting. Higher IL-18 expression by monocytes infected with the avirulent strain may result in more IFN-gamma production, thus modulating its pathogenicity. Local induction of IL-18 may be important both for M. avium pathogenicity and host defence and become a potential candidate for immunotherapy.     

The expression of functional IL-2 receptor on activated macrophages depends on the stimulus applied.

Human peripheral blood monocytes (Mo) synthesize prostaglandin E2 (PGE2) when activated with lipopolysaccharide (LPS). This production is strongly enhanced by the addition of supernatant from phytohaemagglutinin (PHA)-activated T cells. To evaluate the factor(s) responsible for this enhancement we studied the effect of several cytokines on the PGE2 metabolism. Recombinant interleukin-1 (IL-1) or recombinant IL-2 strongly enhanced PGE2 synthesis in LPS-stimulated Mo cultures, whereas recombinant interferon-gamma (IFN-gamma) partially inhibited its production. To see whether the effect of IL-2 on Mo was due to the presence of IL-2 receptor (IL-2R) on the cell surface, flow cytometric analysis and electron microscopy were used to investigate IL-2R expression in unstimulated and stimulated Mo. Stimulated, but not resting, Mo displayed the p55 IL-2R chain on their cellular surface and associated with the polyribosomes of the rough endoplasmic reticulum in the cytoplasm. This finding strongly suggested that the p55 chain of the IL-2R was synthesized by activated Mo. To confirm this result, 125I-labelled IL-2 was bound under high- and low-affinity conditions and cross-linked to Mo cultured in the presence of LPS, IFN-gamma or IL-1. The cross-linked 125I-IL-2/IL-2R complexes were analysed by SDS-PAGE. Mo cultured with LPS, IFN-gamma and IL-1 expressed the p55 protein detected by low-affinity cross-linking, whereas only LPS-stimulated Mo displayed a barely detectable band with an apparent MW of 70,000 under high-affinity binding conditions. In addition, stimulated Mo were found capable of producing the soluble form of IL-2R. Finally, LPS-activated Mo only responded to the addition of IL-2 by an increase in PGE2 production, suggesting that the function of IL-2R on activated Mo is linked to the stimulus applied.     

Role of Th1 and Th2 cytokines in immune response to uncomplicated Plasmodium falciparum malaria

The relative balance between Th1 and Th2 cytokines appears crucial, since the role of cytokines has been evaluated in several studies by comparison of clinically heterogeneous groups of patients. The aim of this study is to determine the role of proinflammatory Th1 cytokines, interleukin-12 (IL-12) and gamma interferon (IFN-gamma), and anti-inflammatory Th2 cytokines, IL-4 and IL-10, in a homogeneous group of patients with uncomplicated Plasmodium falciparum malaria. Levels of IL-12, IFN-gamma, Il-4, and IL-10 in serum for 20 adult patients and 15 healthy control subjects were determined by an immunoenzymatic assay. Serum levels of Th1 cytokines, IL-12 (8.6 +/- 2.8 pg/ml; controls, 3.2 +/- 0.7 pg/ml) and IFN-gamma (39.2 +/- 67.6 pg/ml; controls, 8.4 +/- 6.3 pg/ml), were significantly increased at admission; 3 days later, levels of IL-12 in serum remained significantly high (8.8 +/- 2.6 pg/ml), whereas IFN-gamma levels returned to control values. The anti-inflammatory response of Th2 cytokines (IL-10 and IL-4) was distinct. Levels of IL-10 in serum were not significantly increased at day 0 and day 3 (306.6 +/- 200.4 pg/ml and 56.6 +/- 38.4 pg/ml, respectively; controls, 17.4 +/- 9.0 pg/ml). In contrast, levels of IL-4 in serum were not increased on admission (3.4 +/- 1.2 pg/ml; controls, 2.4 +/- 0.8 pg/ml), but at day 3 a moderate and significant increase of IL-4 levels was observed (4.5 +/- 1.7 pg/ml). In conclusion, the increase of Th1 cytokine IL-12 and IFN-gamma levels during the acute phase of uncomplicated P. falciparum malaria may reflect an early and effective immune response regulated by proinflammatory Th1 cytokines, and in particular IFN-gamma may play a role in limiting progression from uncomplicated malaria to severe and life-threatening complications.     

Staphylococcal glycocalyx activates macrophage prostaglandin E2 and interleukin 1 production and modulates tumor necrosis factor alpha and nitric oxide production.

We have examined the effect of staphylococcal glycocalyces on the ability of murine peritoneal macrophages to produce prostaglandin E2 (PGE2) and the inflammatory cytokines interleukin 1 (IL-1) and tumor necrosis factor alpha (TNF-alpha) and to generate nitric oxide. Glycocalyx partially purified under endotoxin-free conditions from defined liquid medium cultures of Staphylococcus lugdunensis or Staphylococcus epidermidis was a strong stimulator of PGE2 and IL-1 production. The addition of 10 to 100 micrograms of glycocalyx per ml induced levels of IL-1 and PGE2 production similar to that induced by 0.1 to 1 micrograms of Escherichia coli lipopolysaccharide (LPS) per ml. In contrast, glycocalyx induced ninefold less TNF-alpha and three- to fourfold less nitrite than LPS. A modulatory effect was suggested by the observation that the amount of TNF-alpha and nitrite generated remained constant whether the macrophages were stimulated with 10 or 100 micrograms of glycocalyx per ml. A selective modulation of macrophage activation was confirmed by the demonstration that costimulation of macrophages with both glycocalyx and LPS resulted in a reduction in TNF-alpha and nitrite generation relative to stimulation with LPS alone even though costimulation had no effect on PGE2 production and increased IL-1 production. Involvement of PGE2 in this modulatory effect was suggested by the ability of indomethacin to augment glycocalyx-stimulated TNF-alpha production and to reverse the inhibitory effect of glycocalyx on LPS induction of TNF-alpha production. However, the inability of indomethacin to reverse the inhibitory effect of glycocalyx on LPS-induced nitric oxide generation suggests that the selective modulation of macrophage function by glycocalyx may be more complex than increased sensitivity to PGE2 feedback inhibition.     

Effects of natural human interferon-alpha, -beta and -gamma on interleukin 2 production in human peripheral lymphocytes

Effects of interferon (IFN) on PHA-induced interleukin 2 (IL-2) production by human peripheral mononuclear cells were studied comparatively with natural human IFN-alpha, IFN-beta and IFN-gamma, using an equivalent unit of their antiviral activity ranging from 10 to 1000 IU/ml. IL-2 activity was assessed in cultures with or without IFN by a standard bioassay using murine CTLL-2 cells. PHA-induced production of IL-2 in cultures of peripheral mononuclear cells was unaltered or slightly suppressed by the simultaneous presence of IFN-alpha and IFN-beta. The effect was the same, whether or not indomethacin was present in the cultures. In contrast, the addition of IFN-gamma to the PHA-stimulated cultures markedly enhanced IL-2 production, while IFN-gamma per se had no effect on IL-2 production in the absence of PHA. The enhancement of IL-2 production due to IFN-gamma was more marked in cultures which did not include indomethacin than in cultures which contained indomethacin (1 x 10(-6) M).     

Gram-positive bacteria are potent inducers of monocytic interleukin-12 (IL-12) while gram-negative bacteria preferentially stimulate IL-10 production

Interleukin-10 (IL-10) and IL-12 are two cytokines secreted by monocytes/macrophages in response to bacterial products which have largely opposite effects on the immune system. IL-12 activates cytotoxicity and gamma interferon (IFN-gamma) secretion by T cells and NK cells, whereas IL-10 inhibits these functions. In the present study, the capacities of gram-positive and gram-negative bacteria to induce IL-10 and IL-12 were compared. Monocytes from blood donors were stimulated with UV-killed bacteria from each of seven gram-positive and seven gram-negative bacterial species representing both aerobic and anaerobic commensals and pathogens. Gram-positive bacteria induced much more IL-12 than did gram-negative bacteria (median, 3,500 versus 120 pg/ml at an optimal dose of 25 bacteria/cell; P < 0.001), whereas gram-negative bacteria preferentially stimulated secretion of IL-10 (650 versus 200 pg/ml; P < 0.001). Gram-positive species also induced stronger major histocompatibility complex class II-restricted IFN-gamma production in unfractionated blood mononuclear cells than did gram-negative species (12,000 versus 3,600 pg/ml; P < 0.001). The poor IL-12-inducing capacity of gram-negative bacteria was not remediated by addition of blocking anti-IL-10 antibodies to the cultures. No isolated bacterial component could be identified that mimicked the potent induction of IL-12 by whole gram-positive bacteria, whereas purified LPS induced IL-10. The results suggest that gram-positive bacteria induce a cytokine pattern that promotes Th1 effector functions.     

Role of endogenous prostaglandin E2 in interleukin 1 production by peripheral blood monocytes from patients with rheumatoid arthritis

We studied the effect of endogenous prostaglandin E2 (PGE2) on interleukin 1 (IL-1) production by peripheral blood monocytes from patients with rheumatoid arthritis (RA). IL-1 production by RA monocytes was not different from that of monocytes from normal controls, when the cells were either unstimulated or stimulated with lipopolysaccharide (LPS, 20 micrograms/ml), as measured by two different bioassays (thymocyte or fibroblast proliferation assay) and enzyme-linked immunosorbent assay. However, IL-1 production by LPS-stimulated monocytes from RA patients cultured in medium containing indomethacin, an inhibitor of PGE2 synthesis, was significantly greater than that of monocytes from normal controls. In addition, the levels of PGE2 in culture supernatants of unstimulated or LPS-stimulated monocytes from RA patients were higher than in culture supernatants of monocytes from normal controls. Moreover, the increase of in vitro IL-2 production by RA T cells stimulated by phytohemagglutinin (PHA) was observed when monocytes were removed from peripheral blood mononuclear cells. These results indicated that peripheral blood monocytes from RA patients could produce IL-1 in excess in vitro, but that in vivo IL-1 production by RA monocytes and IL-2 induction by RA T cells might be negatively regulated by endogenous PGE2.     

Spontaneous and LPS-stimulated production of intracellular IL-1 beta by synovial macrophages in rheumatoid arthritis is inhibited by IFN-gamma.

In the peripheral blood (PB) as well as the synovial fluid (SF) of rheumatoid arthritis (RA) patients significantly elevated levels of interleukin 1 beta (IL-1 beta) were determined compared to controls by means of a sensitive and specific ELISA (median values: 280 pg/ml and 325 pg/ml vs. less than 20 pg/ml). In 12-h cell cultures of adherent cells, significantly increased spontaneous intracellular IL-1 beta production was determined in SF macrophage (SFM phi) cultures of RA patients compared to PB monocyte (PBMo) cultures of controls (median values: 91.0 ng/10(6) cells vs. 31.5 ng/10(6) cells). However, secretion must be elicited by additional stimulation with lipopolysaccharide (LPS). Interferon-gamma (IFN-gamma) significantly inhibited the spontaneous intracellular IL-1 beta production in SFM phi 12-h cultures of RA patients.     

Expression of IL-18 in patients with head and neck squamous cell carcinoma

Interleukin-18 (IL-18), a recently described cytokine secreted mainly by macrophages, stimulates interferon-gamma (IFN-gamma) production by natural killer cells and T cells. The purpose of this study was to determine tissue expression and serum levels of IL-18 in head and neck squamous cell carcinoma (HNSCC) and to evaluate ethanol and endotoxin-driven cytokine secretion. In 24 patients with primary HNSCC and 28 healthy controls, PBMC were isolated and incubated with 50 mM ethanol, LPS (doses 25 ng/ml, 250 ng/ml, 2500 ng/ml) and both agents for 24 h. Levels of IL-18 in serum, and cell supernatants were analysed by capture ELISA, IL-18 tissue level by immunoblotting. Serum levels of IL-8, IL-10 and IL-12, IFN-gamma, and endotoxin plasma levels were also determined. Statistical analysis involved Welch t-test and Page's test for trend. The majority of patients with HNSCC had high concentrations of serum IL-18. The level of IL-18 in the sera of these patients had a mean level of 271.7 pg/ml, while the mean IL-18 serum level in healthy controls was 174,0 pg/ml (p<0.001). Levels of IL-10 and IL-12, IFN-gamma were not increased in patients. Endotoxin was not detectable in either group. LPS stimulated dose-dependently IL-18 secretion from PBMC of patients and controls in vitro (p<0.05). Incubation with ethanol alone did not affect basal IL-18 secretion, but ethanol reduced LPS-stimulated IL-18 secretion compared to LPS stimulation alone. The mRNA expression of IL-18 in unstimulated PBMC and the response of PBMC to ethanol and LPS was similar in patients and controls. Our data on elevated serum levels of IL-18 in the majority of HNSCC cancer patients, irrespective of its biological activity, suggest that serum IL-18 might be a candidate for a new marker for HNSCC. The pathways for IL-18 production and its mechanisms of action in patients with HNSCC remain to be determined. Understanding of the immunological pathways might offer new therapeutic options in head and neck cancer in the future.     

The combined effect of interleukin (IL)-10 and IL-12 polymorphisms on induced cytokine production

Interleukin (IL)-12 and IL-10 are immunoregulatory cytokines with an antagonistic effect of the T-helper (Th)1/Th2 cytokine balance and provide a functional link between innate and adaptive immune responses. The aim of the study was to investigate the combined effect of -1082A*G in IL10 and +16974A*C in IL12B single nucleotide polymorphisms (SNPs) on induced cytokine production by stimulated peripheral blood mononuclear cells (PBMCs) isolated from healthy donors. The presence of the high-producer IL-12p40 genotype led to diminished production of IL-10 as determined by the -1082*G allele of SNP in IL10. Significantly decreased IL-10 production was detected in AA+AG/GG in comparison with the low-producer IL-12p40 (AC/CC+AG/GG) genotype combination after stimulation with C3bgp (2 +/- 4 vs. 29 +/- 14.2 pg/ml; p = 0.0003) and LPS (33.4 +/- 13.5 vs. 93.3 +/- 59.6 pg/ml; p = 0.019). IL-12p40 production was independent of IL10 genotype. The present results demonstrated that the production of IL-10 from PBMC depended on both -1082A*G in IL10 and +16974A*C in IL12B polymorphisms.     

Differential effects of IL-10 on prostaglandin H synthase-2 expression and prostaglandin E2 biosynthesis between spleen and bone marrow macrophages

Different populations of mononuclear phagocytes (MO) show considerable diversity of cellular function including prostaglandin E2 (PGE2) biosynthesis. Certain bacterial components enhance PGE2 biosynthesis differentially in selected populations of MO. Interleukin (IL)-10 is proposed to inhibit modulation of PGE2 biosynthesis by down-regulating prostaglandin G/H synthase-2 (PGHS-2) expression. To assess whether IL-10 regulates PGE2 biosynthesis and PGHS-2 expression, splenic and bone marrow MO were isolated from IL-10-deficient (IL-10(-/-)), C57Bl/6 [wild-type (WT) control], and Balb/c (comparison control) mice and were treated with lipopolysaccharide (LPS) and/or interferon-gamma (IFN-gamma) as a model of bacterial inflammation. LPS-induced PGHS-2 expression was similar for splenic MO isolated from the three strains of mice. However, PGE2 released by LPS-treated splenic MO was significantly higher in IL-10(-/-) and Balb/c than in WT cells. In the presence of LPS and IFN-gamma, PGHS-2 expression and PGE2 release by IL-10(-/-) and Balb/c splenic MO were enhanced compared with stimulation with LPS alone or IFN-gamma alone. However, there was no significant increase in PGE2 release from WT splenic MO treated with LPS plus IFN-gamma despite increased PGHS-2 expression. In sharp contrast, PGHS-2 expression and PGE2 release by bone marrow MO were greatly enhanced in IL-10(-/-) cells compared with control cells. Our results indicate that IL-10 regulation of MO PGE2 biosynthesis and PGHS-2 expression is compartment-dependent and that PGE2 production is not linked directly to PGHS-2 levels. Furthermore, our findings emphasize strain-specific differences between C57Bl/6 and Balb/c mice, and Balb/c appears more similar to the IL-10(-/-) than to the C57Bl/6 with respect to prostanoid production.     

Role of IFN-gamma on dissociation between nitric oxide and TNF/IL-6 production by murine peritoneal cells after restimulation with bacterial lipopolysaccharide

Murine peritoneal exudate cells (PEC) pre-exposed to bacterial lipopolysaccharide (LPS) show augmented nitric oxide (NO) production by LPS restimulation, in contrast to LPS tolerance with reduced production of tumor necrosis factor alpha (TNF-alpha) and interleukin-6 (IL-6). Significant amounts of interferon-gamma (IFN-gamma) were detected in the PEC cultures on LPS stimulation, and anti-IFN-gamma antibody suppressed the LPS-induced NO, but not TNF-alpha and IL-6, production. Addition of anti-IFN-gamma antibody to the cultures in the LPS pre-exposure step strongly suppressed the augmented NO production on LPS restimulation. Anti-IL-12 antibody, which suppressed the LPS-induced IFN-gamma production, also suppressed the augmented NO production, as did anti-IFN-gamma antibody. Taken together, we propose the following mechanisms: (1) T and NK cells in PEC produce IFN-gamma by the action of IL-12, which is derived from LPS-stimulated macrophages, and (2) the de novo-produced IFN-gamma activates macrophages to augment NO production on LPS restimulation.     

High TNF-alpha and low IL-2 producing T cells characterize active disease in Takayasu's arteritis

We have investigated intracellular production by T cells and plasma levels of TNF-alpha, IL-2 and IFN-gamma in 12 active and 10 inactive Takayasu's arteritis (TA) patients and 12 healthy controls. The active TA compared to inactive TA and controls had higher TNF-alpha (52.7 +/- 22.3% vs. 32.9 +/- 14.2% and 35.2 +/- 14.5%, respectively; P = 0. 020), lower IL-2 (19.6 +/- 13.2% vs. 36.1 +/- 10.1% and 31.2 +/- 10.3%, respectively; P = 0.010) and comparable IFN-gamma (38.6 +/- 13.9% vs. 34.2 +/- 12.4% and 34.9 +/- 11.1%, respectively; P = 0.581) producing CD3+ T cells. There was no difference in the plasma levels of the cytokines between active TA, inactive TA and controls (TNF-alpha: 79.1 +/- 94.5 vs. 72.9 +/- 120.0 and 9.5 +/- 6.7 pg/ml, P = 0.110; IL-2: 4.3 +/- 4.8 vs. 6.6 +/- 4.7 and 8.6 +/- 4.5 pg/ml, P = 0.094 and IFN-gamma: 10.1 +/- 11.3 vs. 8.8 +/- 8.7 and 8.2 +/- 6.5 pg/ml, P = 0.871, respectively). The data show an important role of these high TNF-alpha and low IL-2 producing T cells in TA.