In vitro and in vivo TNFalpha synthesis modulation by methylguanidine, an uremic catabolyte.

This study was performed in order to examine whether the uraemic toxin, methylguanidine (MG), can modulate tumor necrosis factor alpha (TNF alpha) release by activated macrophages. In this study we have evaluated the ability of MG to influence TNF alpha release in vitro, in Escherichia coli lypopolysaccharide- (LPS)-stimulated J774 cells preincubated overnight with MG, and in vivo in rats treated with MG before and after LPS challenge. Parallel experiments employing N(G)-nitro-L-arginine methyl esther (L-NAME) were also carried out for comparison. The effect of LPS (6 x 10(3) u/ml) on TNF alpha release by J774, following overnight incubation with MG or L-NAME (1 mM), was examined 3 hours after LPS challenge. LPS-stimulated J774 released 287.83+/-88 u/ml TNF alpha into the culture medium. MG (1 mM) significantly inhibited TNF alpha release by 73% (P<0.05). L-NAME (1 mM) significantly inhibited TNF alpha release too by 72.88% (P<0.05). The effect of MG and L-NAME have been also studied in vivo. Serum TNF alpha levels in LPS treated rats 2 h after LPS challenge were 88.33+/-31.7 u/ml as compared to the serum TNF alpha levels of control rats (undetectable). Treatment of rats with MG (30 mg/kg, i.p.) strongly and significantly reduced TNF alpha release (98.71% inhibition; with P<0.001); in the same experimental setting L-NAME (10 mg/kg, i.p.) also significantly reduced TNF alpha serum levels (76.47% inhibition; with P<0.01). These results could indicate that immune disfunction related to uremia may be related to the inhibitory capability of uremic catabolyte, MG, on TNF alpha synthesis and release.     

Macrophages rapidly internalize their tumor necrosis factor receptors in response to bacterial lipopolysaccharide

The effect of bacterial lipopolysaccharide (LPS) on macrophage receptors for tumor necrosis factor/cachectin (TNF-R) was studied. At equilibrium, iodinated recombinant human TNF alpha (rTNF alpha) bound to 1100 +/- 200 sites/cell on macrophage-like RAW 264.7 cells with a Kd of 1.3 +/- 0.1 x 10(-9) M. Preexposure of RAW 264.7 cells to 10 ng/ml LPS for 1 h at 37 degrees C resulted in complete loss of cell surface TNF alpha binding sites. 50% loss ensued after 1 h with 0.6 ng/ml LPS, or after 15 min with 10 ng/ml LPS. Complete loss of TNF alpha binding sites occurred without change in numbers of complement receptor type 3. No decrease in TNF-R followed preexposure to LPS at 4 degrees C, nor could LPS displace 125I-rTNF alpha from its binding sites. Although TNF-R disappeared from the surface of intact macrophages following exposure to LPS, specific TNF alpha binding sites were unchanged in permeabilized macrophages, indicating that TNF-R were rapidly internalized. Conditioned media from LPS-treated RAW 264.7 cells induced 30% down-regulation of TNF-R on macrophages from LPS-hyporesponsive mice (C3H/HeJ), suggesting that a soluble macrophage product may be responsible for a minor portion of the LPS effect. Additional evidence against endogenous TNF alpha being the major cause of TNF-R internalization was the rapid onset of the effect of LPS on TNF-R compared to the reported onset of TNF alpha production, the relatively high concentrations of exogenous rTNF alpha required to mimic the effect of LPS, and the inability of TNF alpha-neutralizing antibody to block the effect of LPS. LPS-induced down-regulation of TNF-R was complete or nearly complete not only in RAW 264.7 cells, but also in primary macrophages of both human and murine origin, was less marked in human endothelial cells, and was absent in human granulocytes and melanoma cells and mouse L929 cells. Thus, in situ, macrophages and some other host cells may be resistant to the actions of TNF alpha produced during endotoxinemia, because such cells may internalize their TNF-R in response to LPS before TNF alpha is produced.     

Tumor necrosis factor release from lipopolysaccharide-stimulated human monocytes: Lipopolysaccharide tolerance in vitro

Human peripheral blood monocytes secrete tumor necrosis factor (TNF) in response to stimulation with bacterial lipopolysaccharide (LPS). We have shown that isolated human monocytes pretreated with LPS for 24 h secrete lower levels of TNF on a second stimulation with LPS than monocytes that have been stimulated with a single dose of LPS either immediately after isolation or 24 h after isolation. The levels of TNF released by monocytes after the second stimulation with LPS are proportional to the LPS concentration over a range from 1 ng/mL to 10 micrograms/mL. Increasing concentrations of LPS used during the first 24-h stimulation induce greater suppression of TNF release after a second stimulation with LPS. After an initial stimulus of 10 micrograms/mL LPS, a second stimulation of monocytes even with 10 micrograms/mL LPS will result in TNF secretion similar to that of unstimulated cells. This in vitro tolerance apparently can be overcome by stimulating previously activated cells with phorbol myristate acetate. We have also shown that neither prostaglandin E2 nor dexamethasone added during the initial stimulation with LPS had an effect on the subsequent reduction in TNF release on a second stimulation of monocytes with LPS.     

Role of LPS and receptor subtypes in the uptake of TNF by the murine lung

Tumor necrosis factor alpha (TNF) is an important mediator in lung injury. The kinetics of TNF uptake by the lung are not completely understood. In this study, we evaluated the role that lipopolysaccharide (LPS) and the two types of TNF receptor (p55 and p75) play in the uptake of circulating murine TNF by the murine lung. TNF radioactively labeled with 125I (I-mTNF) was administered intravenously (2 x 10(6) cpm/mouse) to mice with both receptors (wild-type) or to mice missing one (p55-/- or p75-/-) or both (p55-/- and p75-/-) TNF receptors. Blood to lung non-reversible sequestration (Ki) and reversible uptake (Vi) were measured with multiple-time regression analysis. Uptake by lung of I-mTNF in wild-type mice had reversible and non-reversible components. This uptake was decreased by intratracheal, but not by intravenous, LPS, suggesting modulation by local, rather than systemic, inflammation. The p75-/- deficient mice retained the Ki (saturable, non-reversible) component of TNF uptake, whereas p55-/- deficient mice retained the Vi (saturable, reversible) component of TNF uptake. Both Ki and Vi components of TNF uptake were absent in the lungs of p55-/- p75-/- deficient mice. These studies show that local inflammation inhibits the uptake of circulating I-mTNF by lung and that uptake consists of two distinguishable compartments: reversible uptake mediated by the p75 receptor and non-reversible sequestration mediated by the p55 receptor.     

Reversal of defective IL-6 production in lipopolysaccharide-tolerant mice by phorbol myristate acetate

The development of LPS tolerance has been suggested to be mediated by an inhibition of cytokine synthesis. Here we have studied serum IL-6 and TNF levels in mice after LPS administration. Repeated administration of LPS (35 micrograms daily for 4 days) to mice induced a refractoriness (tolerance) to subsequent administrations of LPS in terms of induction of circulating IL-6 and TNF. To investigate the mechanism by which LPS down-regulates its own induction of cytokine synthesis and the relationship between IL-6 and TNF production, we attempted to revert the inhibition of IL-6 and TNF production using agents like PMA or IFN-gamma, previously reported to activate macrophage production of cytokines. Pretreatment with PMA (4 micrograms, 10 min before LPS) partially restored IL-6 production in LPS-tolerant mice given 2 micrograms LPS. On the other hand, PMA did not restore TNF induction in LPS-tolerant mice, even when administered with high doses of LPS (up to 200 micrograms). A similar reversal of LPS resistance to IL-6, but not TNF, induction by PMA was observed in genetically LPS-resistant C3H/HeJ mice. IFN-gamma also restored, although to a lesser extent than PMA, IL-6 production. However, unlike PMA, IFN-gamma could also partially restore TNF production in LPS-tolerant mice, although only when LPS was administered at high doses. By contrast with PMA, IFN-gamma was clearly more active in restoring TNF synthesis than that of IL-6. Similar results were obtained in genetically LPS-unresponsive C3H/HeJ mice. These data suggest that different mechanisms are implicated in the inhibition of IL-6 and TNF synthesis in LPS-tolerant mice and that part of this inhibition can be overcome by PMA or IFN-gamma.     

beta2-Adrenoceptor blockade partially restores ex vivo TNF production following hemorrhagic shock

The aim of the study was to assess the mechanisms through which leukocyte deactivation occurs upon hemorrhagic shock. In particular, the influence of beta-adrenergic tone was evaluated. BALB/c mice were hemorrhaged and resuscitated 60 min after hemorrhage. Animals were sacrificed 60 min later by exsanguination. Blood from exsanguination was cultured ex vivo with lipopolysaccharide (LPS) and heat-killed Staphylococcus aureus Cowan I (SAC). Hemorrhage resulted in a major decrease of LPS-induced TNF production whereas IL-10 production was significantly enhanced. Selective beta(2)-adrenoceptor antagonists (ICI 118,551) attenuated the decrease in TNF production and further enhanced IL-10 production. Hemorrhage did not alter SAC-induced TNF production levels whereas IL-10 production was increased. ICI 118,551 further increased the production of both TNF and IL-10. These data suggest that leukocyte deactivation after LPS stimulation is not a generalized phenomenon since TNF production was maintained when another microbial activator was used. IL-10 production was enhanced after hemorrhagic shock, independently of the nature of the triggering agent. Finally, this study demonstrates that beta(2)-adrenoceptor ligands play an important role in blood leukocyte deactivation to LPS after hemorrhagic shock.     

MNK kinases regulate multiple TLR pathways and innate proinflammatory cytokines in macrophages

The MNK kinases are downstream of both the p38 and ERK MAP kinase pathways and act to increase gene expression. MNK inhibition using the compound CGP57380 has recently been reported to inhibit tumor necrosis factor (TNF) production in macrophage cell lines stimulated with Escherichia coli lipopolysaccharide (LPS). However, the range of receptors that signal through the MNK kinases and the extent of the resultant cytokine response are not known. We found that TNF production was inhibited in RAW264.7 macrophage cells by CGP57380 in a dose-responsive manner with agonists for Toll-like receptor (TLR) 2 (HKLM), TLR4 (Salmonella LPS), TLR6/2 (FSL), TLR7 (imiquimod), and TLR9 (CpG DNA). CGP57380 also inhibited the peak of TNF mRNA production and increased the rate of TNF mRNA decay, effects not due to the destabilizing RNA binding protein tristetraprolin (TTP). Similar to its effects on TNF, CGP57380 caused dose-responsive inhibition of TTP production from stimulation with either LPS or CpG DNA. MNK inhibition also blocked IL-6 but permitted IL-10 production in response to LPS. Studies using bone marrow-derived macrophages (BMDM) isolated from a spontaneous mouse model of Crohn's disease-like ileitis (SAMP1/YitFc strain) revealed significant inhibition by CGP57380 of the proinflammatory cytokines TNF, IL-6, and monocyte chemoattractant protein-1 at 4 and 24 h after LPS stimulation. IL-10 production was higher in CGP53870-treated BMDM at 4 h but was similar to the controls by 24 h. Taken together, these data demonstrate that MNK kinases signal through a variety of TLR agonists and mediate a potent innate, proinflammatory cytokine response.     

Nonsteroidal Anti-Inflammatory Drugs Increase Tumor Necrosis Factor Production in the Periphery but Not in the Central Nervous System in Mice and Rats

Abstract: Nonsteroidal anti-inflammatory drugs (NSAIDs), which inhibit prostaglandin (PG) synthesis, augment production of tumor necrosis factor (TNF) in most experimental models. We investigated the effect of two NSAIDs, indomethacin and ibuprofen, on the production of TNF in the CNS induced by intracerebroventricular injection of lipopolysaccharide (LPS). Indomethacin and ibuprofen, administered intraperitoneally, augmented (three- to ninefold) the levels of TNF in serum and peripheral organs of mice injected intraperitoneally with LPS and in rats with adjuvant arthritis (up to a sevenfold increase). However, NSAIDs (intraperitoneally or intracerebroventricularly) did not increase brain TNF production induced by intravenous LPS. In fact, indomethacin decreased (1.4–1.8-fold) TNF levels in the spinal cord of rats with experimental autoimmune encephalomyelitis and in the cortex of rats with focal cerebral ischemia. Systemic administration of iloprost inhibited serum TNF levels after intraperitoneal LPS, whereas intracerebroventricular injection of iloprost or PGE₂ did not inhibit brain TNF induced by intracerebroventricular LPS. Both peripheral and central TNF productions were inhibited by cyclic AMP level-elevating agents or dexamethasone. Thus, a PG-driven negative feedback controls TNF production in the periphery but not in the CNS.     

Tumour necrosis factor and the lysosomal enzymes of macrophages or macrophage-like cell line

Summary The relationship between tumour necrosis factor (TNF) and macrophages or macrophage-like cell line, especially the lysosomal enzymes was investigated. The serum lysosomal enzymes and LDH activities were increased in proportion to the TNF production even in different strains of mice. Lysosomal enzymes and TNF activity were released into the supernatant of the culture medium of macrophage-enriched peritoneal exudate cells (PEC) or spleen cells derived from Propionibacterium acnes-primed mice after addition of lipopolysaccharide (LPS). After passage through a Sephadex G-10 column, TNF activity could not be detected in the supernatant of these spleen cells after addition of LPS. Also TNF activity could not be detected in the supernatant following destruction of PEC. These results suggest that TNF producibility is strongly related to the degree of activation of macrophages, especially the lysosomal enzymes. The murine macrophage-like cell line, J 774, also released TNF activity and lysosomal enzymes after addition of LPS.     

Nitric oxide suppression of cellular proliferation depends on cationic amino acid transporter activity in cytokine-stimulated pulmonary endothelial cells

Inducible nitric oxide (NO) synthase (iNOS) is a stress response protein upregulated in inflammatory conditions, and NO may suppress cellular proliferation. We hypothesized that preventing L-arginine (L-arg) uptake in endothelial cells would prevent lipopolysaccharide/tumor necrosis factor-α (LPS/TNF)-induced, NO-mediated suppression of cellular proliferation. Bovine pulmonary arterial endothelial cells (bPAEC) were treated with LPS/TNF or vehicle (control), and either 10 mM L-leucine [L-leu; a competitive inhibitor of L-arg uptake by the cationic amino acid transporter (CAT)] or its vehicle. In parallel experiments, iNOS or arginase II were overexpressed in bPAEC using an adenoviral vector (AdiNOS or AdArgII, respectively). LPS/TNF treatment increased the expression of iNOS, arginase II, CAT-1, and CAT-2 mRNA in bPAEC, resulting in greater NO and urea production than in control bPAEC, which was prevented by L-leu. LPS/TNF treatment resulted in fewer viable cells than in controls, and LPS/TNF-stimulated bPAEC treated with L-leu had more viable cells than LPS/TNF treatment alone. LPS/TNF treatment resulted in cleaved caspase-3 and cleaved poly(ADP-ribose) polymerase expression, which was attenuated by L-leu. AdiNOS reduced viable cell number, and treatment of AdiNOS transfected bPAEC with L-leu preserved cell number. AdArgII increased viable cell number, and treatment of AdArgII transfected bPAEC with L-leu prevented the increase in cell number. These data demonstrate that iNOS expression in pulmonary endothelial cells leads to decreased cellular proliferation, which can be attenuated by preventing cellular L-arg uptake. We speculate that CAT activity may represent a novel therapeutic target in inflammatory lung diseases characterized by NO overproduction.     

Involvement of tumor necrosis factor in cytotoxicity mediated by human monocytes

Human monocytes cultured in a specially prepared medium free of lipopolysaccharide (LPS) constitutively produced a small, though significant, amount of tumor necrosis factor (TNF). Upon addition of LPS, the amount produced remained constant until the LPS concentration reached 1-10 ng/ml, whereupon the production of TNF dramatically increased, eventually becoming 100-fold greater than when the LPS concentration was below 1 ng/ml. Priming the monocytes with recombinant interferon-gamma (rIFN-gamma) before LPS exposure resulted in a 2- to 10-fold increase in TNF production, the highest relative increase being obtained at lower LPS concentrations and in the absence of LPS. Monocyte-produced TNF appears to be the effector molecule in monocyte-mediated killing of some target cell types, since antiserum against recombinant TNF inhibited killing of both actinomycin D-treated and untreated WEHI 164 cells by human monocytes. However, it also appears that TNF may not in all cases be an effector molecule in monocyte-mediated killing, since cytolysis of K562 cells mediated by IFN-gamma/LPS-activated monocytes was not inhibited by antiserum against recombinant TNF. Antiserum which was raised against a monocyte-derived cytotoxic factor and which neutralized recombinant TNF did, however, inhibit monocyte-mediated cytolysis of K562 cells, suggesting that an extracellular factor, perhaps related to TNF, was also involved in monocyte-mediated killing of K562 cells. A TNF-like activity was associated with the monocyte surface membrane, since paraformaldehyde-fixed monocytes expressed cytotoxic activity which was neutralized by antiserum against recombinant TNF. Fixed monocytes activated with rIFN-gamma in addition to LPS before fixation were generally more cytotoxic than those exposed to LPS alone, and those exposed to LPS were much more cytotoxic than those not exposed to LPS. Thus it is possible that high local TNF concentrations may be generated near the target cell upon direct contact between effector and target cells, and that also monocyte-associated TNF may in this way be involved in monocyte-mediated cytotoxicity.     

TNFα-Erk1/2 signaling pathway-regulated SerpinE1 and SerpinB2 are involved in lipopolysaccharide-induced porcine granulosa cell proliferation

Lipopolysaccharide (LPS) is an inhibitory factor that causes hormonal imbalance and subsequently affects ovarian function and fertility in mammals. Previous studies have shown that the exposure of granulosa cells (GC) to LPS leads to steroidogenesis dysfunction. However, the effects of LPS on the viability of GC remain largely unclear. In the present study, we aimed to address this question and unveil the underlying molecular mechanisms using cultured porcine GC. Results showed that GC proliferation and tumor necrosis factor α (TNFα) secretion were significantly increased after exposure to LPS, and these effects were completely reversed by blocking the TNFα sheddase, ADAM17. Moreover, GC proliferation induced by LPS was mimicked by treatment with recombinant TNFα. In addition, SerpinE1 and SerpinB2 expression levels increased in GC after treatment with LPS or recombinant TNFα, whereas blocking the Erk1/2 pathway completely abolished these effects and also inhibited GC proliferation. Further, consistent with the effects of blocking the Erk1/2 pathway, cell proliferation was completely inhibited by knocking down SerpinE1 or SerpinB2 in the presence of LPS or recombinant TNFα. Mitochondrial membrane potential (MMP) polarization in GC was increased by LPS or recombinant TNFα treatment, and these changes were completely negated by Erk1/2 inhibition, but not by SerpinE1 or SerpinB2 knockdown. Taken together, these results suggested that the TNFα-mediated upregulation of SerpinE1 and SerpinB2, through activation of the Erk1/2 pathway plays a crucial role in LPS-stimulated GC proliferation, and the increase in GC MMP may synergistically influence this process.     

Immunostimulating effects of muramyl dipeptide, glucosaminyl muramyl dipeptide and their synthetic derivatives in vitro

The effect of muramyldipeptide (MDP), glucosaminylmuramyldipeptide (GMDP) and their six synthetic derivatives on production of tumor necrosis factor (TNF), interleukin-1 (IL-1) and interleukin-2 (IL-2) by murine spleen cells in vitro was studied. MDP induced insignificant TNF production and did not stimulate production of IL-1 by the murine splenocytes within a 24-hour cultivation period whereas in combination with lipopolysaccharide (LPS) it induced significant production of both the cytokins. GMDP induced marked production of TNF (54 per cent cytotoxic index) and IL-1 (stimulation index 8). Addition of LPS in an amount of 10 ng/ml increased production of TNF by the murine splenocytes under the effect of GMDP but had no effect on production of IL-1. Neither MDP nor GMDP even in combination with LPS induced production of IL-2 by splenocytes of mice DVA/2 and C57B1/6 at activation for 24 hours. All the synthetic derivatives of MDP and GMDP except the MDP polymer activated TNF production by the murine spleen cells. GMDP lysine had the highest effect: 67 per cent cytotoxic index. In combination with LPS its cytotoxic index amounted to 87 per cent. The TNF activity was always higher when LPS in an amount of 10 ng/ml was added to the glycopeptides.     

Human toll-like receptor 2 mediates monocyte activation by Listeria monocytogenes, but not by group B streptococci or lipopolysaccharide

Human Toll like receptor (TLR) 2 has been implicated as a signaling receptor for LPS from Gram-negative bacteria and cell wall components from Gram-positive organisms. In this study, we investigated whether TLR2 can signal cell activation by the heat-killed group B streptococci type III (GBS) and Listeria monocytogenes (HKLM). HKLM, but not GBS, showed a time- and dose-dependent activation of Chinese hamster ovary cells transfected with human TLR2, as measured by translocation of NF-kappaB and induction of IL-6 production. A mAb recognizing a TLR2-associated epitope (TL2.1) was generated that inhibited IL-6 production from Chinese hamster ovary-TLR2 cells stimulated with HKLM or LPS. The TL2.1 mAb reduced HKLM-induced TNF production from human monocytes by 60%, whereas a CD14 mAb (3C10) reduced the TNF production by 30%. However, coadministrating TL2.1 and 3C10 inhibited the TNF response by 80%. In contrast to this, anti-CD14 blocked LPS-induced TNF production from monocytes, whereas anti-TLR2 showed no inhibition. Neither TL2.1 nor 3C10 affected GBS-induced TNF production. These results show that TLR2 can function as a signaling receptor for HKLM, possibly together with CD14, but that TLR2 is unlikely to be involved in cell activation by GBS. Furthermore, although LPS can activate transfected cell lines through TLR2, this receptor does not seem to be the main transducer of LPS activation of human monocytes. Thus, our data demonstrate the ability of TLR2 to distinguish between different pathogens.     

Production of lipopolysaccharide-induced tumour necrosis factor during influenza virus infection in mice coincides with viral replication and respiratory oxidative burst

Increased morbidity and mortality occur regularly during influenza epidemics. The exact mechanisms involved are not well defined but bacterial superinfection of influenza virus infected patients is considered to play an important role. In the present study, the effect of influenza virus infection on in vivo production of turnout necrosis factor (TNF) in response to bacterial stimuli was investigated. Release of TNF in mice infected by an aerosol of influenza virus was significant after administration of bacterial lipopolysaccharide (LPS) at 72 h, whereas administration of homologous influenza virus produced only modest amounts of TNF at 96 h. Significant production of TNF was observed 48 h after intravenous administration of infectious influenza in response to LPS but not with the homologous virus. TNF induced after influenza virus infection could be blocked by a specific murine anti-TNF monoclonal antibody. Higher TNF production following aerosol influenza infection correlated with peak titres of influenza virus in the lungs of infected mice and with enhanced generation of luminoldependent chemiluminscence.     

Ciliary neurotrophic factor inhibits brain and peripheral tumor necrosis factor production and, when coadministered with its soluble receptor, protects mice from lipopolysaccharide toxicity.

BACKGROUND: The receptor of ciliary neurotrophic factor (CNTF) contains the signal transduction protein gp130, which is also a component of the receptors of cytokines such as interleukin (IL)-6, leukemia-inhibitory factor (LIF), IL-11, and oncostatin M. This suggests that these cytokines might share common signaling pathways. We previously reported that CNTF augments the levels of corticosterone (CS) and of IL-6 induced by IL-1 and induces the production of the acute-phase protein serum amyloid A (SAA). Since the elevation of serum CS is an important feedback mechanism to limit the synthesis of proinflammatory cytokines, particularly tumor necrosis factor (TNF), we have investigated the effect of CNTF on both TNF production and lipopolysaccharide (LPS) toxicity. MATERIALS AND METHODS: To induce serum TNF levels, LPS was administered to mice at 30 mg/kg i.p. and CNTF was administered as a single dose of 10 micrograms/mouse i.v., either alone or in combination with its soluble receptor sCNTFR alpha at 20 micrograms/mouse. Serum TNF levels were the measured by cytotoxicity on L929 cells. In order to measure the effects of CNTF on LPS-induced TNF production in the brain, mice were injected intracerebroventricularly (i.c.v.) with 2.5 micrograms/kg LPS. Mouse spleen cells cultured for 4 hr with 1 microgram LPS/ml, with or without 10 micrograms CNTF/ml, were also analyzed for TNF production. RESULTS: CNTF, administered either alone or in combination with its soluble receptor, inhibited the induction of serum TNF levels by LPS. This inhibition was also observed in the brain when CNTF and LPS were administered centrally. In vitro, CNTF only marginally affected TNF production by LPS-stimulated mouse splenocytes, but it acted synergistically with dexamethasone (DEX) in inhibiting TNF production. Most importantly, CNTF administered together with sCNTFR alpha protected mice against LPS-induced mortality. CONCLUSIONS: These data suggest that CNTF might act as a protective cytokine against TNF-mediated pathologies both in the brain and in the periphery.     

Regulation of bronchoalveolar macrophage proinflammatory cytokine production by dexamethasone and granulocyte-macrophage colony-stimulating factor after stimulation by Aspergillus conidia or lipopolysaccharide

There is substantial evidence that local production of proinflammatory cytokines are very important in host resistance to aspergillosis. Dexamethasone (DEX) down-regulates production of these cytokines by stimulated bronchoalveolar macrophages (BAM) and constitutes a risk factor for aspergillosis. Granulocyte-macrophage colony-stimulating factor (GM-CSF) antagonizes DEX suppression of antifungal activity by BAM. Here we investigated the possibility that GM-CSF could antagonize DEX down-regulation of interleukin (IL)-1alpha and tumour necrosis factor (TNF)-alpha production by stimulated BAM. Control BAM responded to increasing numbers of conidia of Aspergillus fumigatus with increasing production of IL-1 and TNF. DEX (10(-7)M) significantly suppressed IL-1 and TNF production by BAM+conidia. Although GM-CSF did not enhance IL-1 or TNF production by BAM+conidia, GM-CSF significantly antagonized DEX suppression of IL-1 cytokine production. For comparative purposes, lipopolysaccharide (LPS, 1 microg/ml) was used to stimulate BAM in experiments similar to the above. In contrast to the findings with conidia, GM-CSF enhanced the production of IL-1 (5-fold) and TNF (1.5-fold) by LPS treated BAM. DEX suppression of cytokine production by BAM+LPS was modestly but significantly antagonized by GM-CSF. Moreover, differences between regulation of IL-1 and TNF production by BAM+conidia or LPS and peritoneal macrophages (PM)+conidia or LPS were documented. Finally, the anti-inflammatory cytokine IL-10 was minimally produced by BAM + conidia or LPS, but IL-10 was produced by PM + conidia or LPS. In summary, these data indicate that the risk factor for aspergillosis associated with DEX could be lessened in the pulmonary compartment with GM-CSF. On the other hand, desired effects of DEX could be maintained in other compartments.     

Neurosteroid levels are increased in vivo after LPS treatment and negatively regulate LPS-induced TNF production

Neuroactive steroids such as dehydroepiandrosterone sulfate and pregnenolone inhibit lipopolysaccharide (LPS)-induced tumor necrosis factor (TNF) production. Corticosteroids not only inhibit TNF production but their levels are increased in vivo after endotoxin injection, thus representing a feedback system that limits TNF production. We wondered whether the same could be true for neuroactive steroids. Thus, the possibility that neuroactive steroids might be increased concomitantly to TNF induction in vivo in mice treated with LPS was investigated. Increased plasma and hippocampal levels of allopregnanolone (but not of dehydroepiandrosterone or pregnenolone) were found 90 min after LPS injection. Allopregnanolone and progesterone (IC50 10- 7 and 10- 9 M, respectively) also inhibited TNF production by mouse peritoneal macrophages in vitro at concentrations in the range of those detected in vivo. These findings suggest that neuroactive steroids may act as endogenous inhibitors of cerebral and systemic TNF production.