Tumor Necrosis Factor Bioactivity Research Articles

Bioactive tumour necrosis factor alpha but not granulocyte-macrophage colony-stimulating factor correlates inversely with Langerhans's cell numbers in skin tumours.

Langerhans' cells (LCs) are thought to play an important role in presentation of tumour antigens for the induction of anti-tumour immunity. Epidermis overlying some transplanted murine skin tumours contains increased numbers of LCs; however, alterations in LC numbers are not related to tumour antigenicity or host immunity, suggesting that another factor(s), such as tumour-produced cytokines, influences LC density. It has been postulated that dendritic epidermal T cells (DETCs) play a role in immunosurveillance within the normal epidermis. Two cytokines which potentially alter LC numbers or function include granulocyte-macrophage colony-stimulating factor (GM-CSF) and tumour necrosis factor-alpha (TNF-alpha). GM-CSF maintains LC viability in culture, and there are reports that it can increase LC density. There is evidence that TNF-alpha induces LC to migrate from the epidermis. In the present study, LC densities in regressor and non-regressor murine skin tumours and overlying epidermis were enumerated, and bioactive GM-CSF and TNF-alpha present in the tumours were measured. We found significantly increased epidermal LC numbers above non-regressor, but not regressor, tumours. DETC numbers were significantly increased above some tumours. Although all tumour types produced TNF-alpha, the regressors, which did not increase LC numbers, produced the most TNF-alpha. In contrast, tumour production of GM-CSF did not correlate with any pattern of alteration of LC density or tumour growth. Tumour production of neither cytokine nor tumour growth correlated with DETC numbers overlying tumours. Our results suggest that TNF-alpha may be associated with skin tumour regression and may prevent LC accumulation by tumours.

Bioactive tumour necrosis factor alpha but not granulocyte‐ macrophage colony‐stimulating factor correlates inversely with Langerhans' cell numbers in skin tumours

Bioactive tumour necrosis factor alpha but not granulocyte‐ macrophage colony‐stimulating factor correlates inversely with Langerhans' cell numbers in skin tumours

Tumour Necrosis Factor and Associated Cytokines in the host's response to malaria

Tumour Necrosis Factor (TNF) is produced at the initiation of malaria infections (pre-erythrocytic phase), as demonstrated by the release of bioactive TNF by peripheral blood mononuclear cells from individuals residing in endemic areas after stimulation with stage specific sporozoite antigens. During the erythrocytic phase, TNF production is greatly augmented by parasite antigens at the time of schizont rupture and merozoite release from infected erythrocytes. Some of the strongest inducers of TNF synthesis and release are malaria toxins, e.g. glycosylphosphatidylinositol moieties and malaria pigment. Because of TNF's well-known cytotoxic activity it was originally hypothesized that it alone was responsible for killing parasites directly or within host cells. Though earlier reports of the capability of serum containing TNF to kill plasmodia supported this idea, later experiments with recombinant TNF showed a lack of significant parasiticidal activity. Recent studies investigating related factors showed that they were involved with TNF in the control of infection. These factors included other cytokines, such as interleukin (IL)-1, IL-6, IL-12, interferon-gamma (IFN γ) as well as nitric oxide intermediates (NOI) and reactive oxygen intermediates (ROI). This positioned TNF as a key regulator of the immune response against the malaria parasite. However, it must be noted that TNF and its associated factors are also responsible for the fever, aches and pains of acute illness, as well as the hypoglycemia, shock, bleeding and reversible coma of severe malaria seen in approximately 1 percent of individuals with malaria. Therein lies the rub; factors important in the control of malaria also appear to have detrimental properties. Research presented in this review characterizes TNF and associated cytokines' importance in the immune response to malaria.

Long-Term Intracerebroventricular Infusion of Corticotropin-Releasing Hormone Alters Neuroendocrine, Neurochemical, Autonomic, Behavioral, and Cytokine Responses to a Systemic Inflammatory Challenge

Corticotropin-releasing hormone (CRH) was infused intracerebroventricularly into rats for 7 d via a miniosmotic pump (1 microg . microl-1 . hr-1). Body temperature and locomotor activity were recorded during the treatment using biotelemetry, whereas hippocampal serotonergic neurotransmission and free corticosterone levels were monitored using in vivo microdialysis on day 7 of CRH treatment. During the microdialysis experiment, behavioral activity was scored by assessing the time during which rats were active (locomotion, grooming, eating, drinking). Continuous intracerebroventricular infusion of CRH produced a transient increase in body temperature and locomotion. Moreover, intracerebroventricularly CRH-treated rats showed elevated free corticosterone levels with no apparent diurnal rhythm. Intraperitoneal administration of bacterial endotoxin -lipopolysaccharide (LPS); 100 microg/kg body weight- on day 7 of CRH/vehicle treatment produced a marked fever response in control animals, which was significantly blunted in intracerebroventricularly CRH-treated rats. Although free corticosterone levels reached similar peak concentrations in both intracerebroventricularly vehicle- and CRH-infused groups after LPS, this response was delayed significantly by approximately 1 hr in the intracerebroventricularly CRH-treated animals. Microdialysis experiments showed no changes in basal extracellular levels of serotonin and 5-hydroxyindoleacetic acid in intracerebroventricularly CRH-infused animals. Injection of LPS in intracerebroventricularly CRH-treated rats produced a blunted 5-HT response and a delayed onset of behavioral inhibition and other signs of sickness behavior. Assessment of the endotoxin-induced cytokine responses showed significantly enhanced plasma interleukin-1 (IL-1) and IL-6 bioactivities in the intracerebroventricularly CRH-infused animals 3 hr after injection of LPS, whereas tumor necrosis factor bioactivity responses were not different. Our data demonstrate that chronically elevated brain CRH levels produce marked changes in basal (largely CRH regulated) physiological and behavioral processes accompanied by aberrant responses to an acute challenge. The present study provides evidence that chronic CRH hypersecretion is an important factor in the etiology of stress-related disorders.

Elevated levels of type II soluble tumor necrosis factor receptors in the bronchoalveolar lavage fluids of patients with sarcoidosis.

Since tumor necrosis factor (TNF) is known to be involved in granuloma formation in sarcoidosis, and soluble TNF receptors (sTNF-Rs) inhibit TNF action in vivo, we evaluated the levels of sTNF-Rs in the bronchoalveolar lavage fluids (BALF) of 31 subjects using an enzyme-linked immunosorbent assay. Our group consisted of 13 patients with sarcoidosis (7 sarcoidosis patients who received no treatment and 6 who received corticosteroid therapy) and 18 control subjects (11 healthy nonsmokers and 7 asymptomatic smokers). Type II (75-kDa), but not type I (55 kDa) sTNF-R in BALF was elevated significantly in patients with sarcoidosis compared with the healthy nonsmokers (type I: 126.7 +/- 17.6 pg/ml BALF vs 79.4 +/- 16.5 pg/ml BALF, p > 0.05; type II: 98.3 +/- 27.8 pg/ml BALF vs 26.7 +/- 4.9 pg/ml BALF, p < 0.05). Although levels of type I sTNF-R in BALF from sarcoidosis patients were not correlated with any cellular profiles of BALF, concentrations of type II correlated significantly with the numbers of lymphocytes in BALF. We concluded that sTNF-R is a normal constituent of the epithelial lining fluids and that levels of type II sTNF-R are elevated significantly in the BALF from individuals with sarcoidosis. This suggests that sTNF-Rs may influence the local bioactivity of TNF and may also contribute to the pathogenesis of sarcoidosis.

Regulation of the release of tumour necrosis factor (TNF)alpha and soluble TNF receptor by gamma irradiation and interferon gamma in Ewing's sarcoma/peripheral primitive neuroectodermal tumour cells.

This study analyses the production of tumour necrosis factor (TNF)alpha and soluble TNF receptor (sTNF-R) before and after exposure to gamma irradiation and interferon gamma (IFN gamma) in 12 cell lines derived from Ewing's sarcoma (ES)/peripheral primitive neuroectodermal tumours (pPNET). Supernatants from ES/pPNET cell cultures were tested in a TNF alpha-specific amplified enzyme-linked immunosorbent assay (ELISA), a bioassay, and sTNF-Rp55 and sTNF-Rp75 ELISA. The tumour cell lines released minimal amounts of TNF alpha, prominent amounts of sTNF-Rp55 (7/12 cell lines) and no sTNF-Rp75. Exposure to gamma irradiation (5 Gy) either induced (3/12) cell lines) or up-regulated (3/12 cell lines) TNF alpha release without changing sTNF-Rp55 and sTNF-Rp75 levels. Priming of cultures with recombinant human IFN gamma (rhIFN gamma) markedly enhanced TNF alpha secretion in the radiation-responsive cell lines and had no influence on sTNF-Rp55 and sTNF-Rp75 levels. rhIFN gamma affected the magnitude rather than the sensitivity of the radiation response. The TNF alpha secreted was bioactive, as shown by its cytotoxic effect of WEHI-164 cells, and neutralization of its activity by anti-TNF alpha monoclonal antibody. Herbimycin A (a tyrosine-specific protein kinase inhibitor) but not calphostin C (a protein kinase C inhibitor), H89 (a protein kinase A inhibitor), AA-COCF3 (a specific inhibitor of phospholipase A2) and MK-886 (a specific inhibitor of 5-lipoxygenase) abrogated gamma-irradiation-stimulated TNF alpha release. The antioxidants N-acetylcysteine, nordihydroguaiaretic acid and mepacrine dose-dependently inhibited gamma-irradiation-mediated TNF alpha production. Collectively our findings indicate that IFN gamma priming potentiates the secretion of bioactive TNF alpha by ES/pPNET cells in response to gamma irradiation without affecting sTNF-R release. The data suggest a requirement for protein tyrosine kinase activity and a role for reactive oxygen species in the gamma-irradiation-mediated intracellular signalling pathway leading to TNF alpha production.

Preclinical Safety Assessment of the Recombinant TNF Receptor–Immunoglobulin Fusion Protein

The tumor necrosis factor (TNF) is now recognized as one of the most pleiotropic mediators of host defense, immune regulation, and inflammatory response. Due to its broad spectrum of effects, TNF has been implicated as a key mediator in the pathogenesis of acute and chronic inflammatory conditions. The inhibition of bioactive TNF could therefore provide a substantial therapeutic benefit. One approach to a potent TNF antagonist is to use recombinant protein technology in the design of a molecule in which the heavy-chain sequences of an immunoglobulin are fused with the ligand-binding region of the TNF receptor. This paper describes some aspects of the preclinical safety evaluation of the recombinant human TNF receptor–immunoglobulin fusion protein (TENEFUSE). Specific emphasis was placed on transgenic and gene-deleted mice models as central sources of information in the safety evaluation of this TNF antagonist.

Randomized placebo-controlled clinical trial of high-dose interleukin-2 in combination with a soluble p75 tumor necrosis factor receptor immunoglobulin G chimera in patients with advanced melanoma and renal cell carcinoma.

A randomized, double-blind, placebo-controlled trial was performed to compare the toxicity and biologic effects of treatment with high-dose intravenous (IV) bolus interleukin-2 (IL-2) plus the recombinant human soluble p75 tumor necrosis factor (TNF) receptor immunoglobulin G (IgG) chimera (rhuTNFR:Fc) with high-dose IL-2 alone in patients with advanced melanoma and renal cell carcinoma. Twenty patients with advanced melanoma or renal cell carcinoma were randomized to receive IL-2 (Chiron, Emeryville, CA) 600,000 IU/kg every 8 hours on days 1 to 5 and 15 to 19 (maximum, 28 doses) combined with placebo or the rhuTNFR:Fc fusion protein (Immunex, Seattle, WA) 10 mg/m2 on days 1 and 15 and 5 mg/m2 on days 3, 5, 17, and 19. The impact of rhuTNFR:Fc on IL-2 toxicity and biologic effects was evaluated. No clinically significant difference in toxicity was observed in the two treatment arms. The adjusted median number of IL-2 doses administered during cycle 1 was 24.5 (range, seven to 28) and 21.5 (range, five to 27) for the placebo and rhuTNFR:Fc arms, respectively (P = .544). IL-2-induced TNF bioactivity, neutrophil chemotactic defect, and serum IL-6, IL-8, and IL-1 receptor antagonist (IL-1RA) induction were suppressed by rhuTNFR:Fc. Two of nine assessable patients (22%) on IL-2/placebo and three of 10 patients (30%) on IL-2/rhuTNFR:Fc responded. Despite evidence of in vitro neutralization of TNF functional activity and partial inhibition of other secondary biologic effects of IL-2, rhuTNFR:Fc does not reduce the clinical toxicity associated with high-dose IL-2 therapy. These results suggest that the toxicity and antitumor effects of IL-2 treatment are independent of circulating TNF.

Intense exercise impairs the integrity of the pulmonary blood-gas barrier in elite athletes.

The blood-gas barrier must be very thin to allow gas exchange and it is therefore subjected to high mechanical stresses when the capillary pressure rises. In some animals, such as the thoroughbred race-horse during intense exercise, the stresses are so large that the capillaries fail and bleeding occurs. We tested the hypothesis that, in elite human athletes, the high capillary pressure that occurs during severe exercise alters the structure and function of the blood-gas barrier. We performed bronchoalveolar lavage (BAL) in six healthy athletes, who had a history suggestive of lung bleeding, 1 h after a 7-min cycling race simulation and four normal sedentary control subjects who did not exercise before BAL. The athletes had higher (p < 0.05) concentrations of red blood cells (0.51 x 10(5) versus 0.01 x 10(5).ml-1), total protein (128.0 versus 94.1 micrograms/ml), albumin (65.6 versus 53.0 micrograms/ml), and leukotriene B4 (LTB4) (243 versus 0 pg/ml) in BAL fluid than control subjects. The proportion of neutrophils was similar in athletes and control subjects but the proportion of lymphocytes in BAL fluid was reduced (p < 0.05). There were no differences in levels of surfactant apoprotein A, tumor necrosis factor bioactivity, lipopolysaccharide, or interleukin-8 (IL-8) between groups. These results show that brief intense exercise in athletes with a history suggestive of lung bleeding alters blood-gas barrier function resulting in higher concentrations of red cells and protein in BAL fluid. The lack of activation of proinflammatory pathways (except LTB4) in the airspaces supports the hypothesis that the mechanism for altered blood-gas barrier function is mechanical stress.

Local cytokine induction by LPS in the rat air pouch and its relationship to the febrile response.

Peripheral induction of cytokines is a critical event in the induction of febrile responses. The sequence of induction and site of action of these cytokines, however, remain unclear. The objective of the present study was to investigate the kinetics of local and systemic production of interleukin (IL)-1, IL-6, and tumor necrosis factor (TNF), with the aim of identifying the relationship between these cytokines and the febrile response induced by injection of lipopolysaccharide (LPS) into a subcutaneous air pouch in the rat. Intrapouch injection of LPS induced dose-dependent fevers and increases in the concentration of bioactive IL-6 in the plasma. Further studies using 100 microg/kg LPS demonstrated significant increases in local (air pouch) concentrations of bioactive IL-1, TNF and IL-6, and circulating IL-6. No significant increases in TNF or IL-1 were detected in the plasma of the same animals. Local TNF was induced rapidly and peaked 1 h after LPS injection. The kinetics of local IL-1 and IL-6 induction were similar and both peaked after 3 h. The rise in local IL-6 preceded that of plasma IL-6 and reached a peak concentration that was 25-fold higher than that observed in the plasma. The data indicate that IL-1 and TNF act locally at the site of inflammation and that locally induced IL-6 is the important systemic mediator of the response.

The effects of 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) on tumor necrosis factor (TNF) production by peritoneal cells

Recent studies in mice have demonstrated that TNF plays a critical role in mediating the TCDD-induced enhanced inflammatory response to intraperitoneal (i.p.) sheep red blood cells. The current studies were designed to evaluate the effects of TCDD on TNF production by ex-vivo peritoneal cells and a peritoneal macrophage cell line (IC-21) stimulated with LPS. In support of the hypothesis that TCDD can act directly on the peritoneal macrophage to increase TNF production, following pretreatment with TCDD, both ex-vivo peritoneal cells and IC-21 cells produced increased levels of bioactive TNF when stimulated with LPS. Flow cytometric analyses of IC-21 cells indicate that TCDD exposure increases intracellular production and secretion of TNF but does not alter levels of membrane associated TNF.

Soluble tumor necrosis factor (TNF) receptors conserve TNF bioactivity in meningitis patient spinal fluid.

The content of tumor necrosis factor (TNF)-alpha antigen and the bioactivity of soluble TNF receptor type II (sTNF-RII) in cerebrospinal fluid (CSF) from 29 patients with meningeal symptoms and fever were examined. Immunoreactive TNF was demonstrated in CSF from 4 of 7 patients with bacterial meningitis. In 3 of 8 patients with aseptic meningitis, CSF also contained TNF, but TNF bioactivity was confined to samples from patients with bacterial meningitis. Bioactive TNF was exclusively in high-performance liquid chromatography fractions containing 30- to 60-kDa proteins. Lipopolysaccharide induced down-regulation, possibly after shedding of granulocyte surface membrane TNF-RII. Consistently, there was a statistically significant correlation between sTNF-RII and CSF leukocyte counts. Bioactive TNF was found only in CSF containing >1 ng of sTNF-RII/mL; samples without TNF bioactivity contained less sTNF-RII. Thus, a stabilizing effect of sTNF-RII on the oligomeric cytokine in vivo is plausible.

Evidence for hormonally regulated changes in ovarian and circulating levels of bioactive tumor necrosis factor alpha in the gonadotropin-treated immature rat.

Evidence for hormonally regulated changes in ovarian and circulating levels of bioactive tumor necrosis factor alpha in the gonadotropin-treated immature rat.

Plasma Inhibitory Activity against Tumour Necrosis Factor in Fulminant Hepatic Failure

1. Soluble tumour necrosis factor receptors released into the circulation inhibit the effects of excess tumour necrosis factor-alpha and represent an important protective response. 2. In this study we have measured the levels of tumour necrosis factor and soluble tumour necrosis factor receptors p55 and p75 in the plasma of 10 patients with fulminant hepatic failure and 10 healthy control subjects. The capacity of the plasmas at varying dilutions to inhibit the biological activity of 1000 pg/ml of recombinant tumour necrosis factor in a tumour necrosis factor cytotoxicity assay in vitro was also determined. 3. The mean plasma levels of tumour necrosis factor in patients with fulminant hepatic failure (48.4 +/- 10.9 pg/ml) were significantly increased compared with normal control subjects (6.1 +/- 1.04 pg/ml, P < 0.01). Plasma soluble tumour necrosis factor receptors p55 and p75 were also significantly elevated in patients with fulminant hepatic failure (18.16 +/- 9.94 ng/ml and 16.06 +/- 9.93 ng/ml respectively) when compared with normal control subjects (1.28 +/- 0.24 ng/ml and 1.62 +/- 0.91 ng/ml, P < 0.001). 4. Fulminant hepatic failure plasma had a much lower capacity to inhibit tumour necrosis factor bioactivity in vitro, with a statistically significant difference between the inhibitory capacity of the fulminant hepatic failure and normal plasma seen at plasma dilutions of 1:5 and 1:20 (P < 0.05). 5. The reduced tumour necrosis factor neutralization capacity observed in fulminant hepatic failure, despite the increased levels of soluble tumour necrosis factor receptors, suggests enhanced susceptibility to the potential deleterious effects of tumour necrosis factor in fulminant hepatic failure.

Antiserum to tumor necrosis factor and failure to prevent murine colitis.

Cytokines regulate many aspects of disease and have been implicated as mediators of the inflammatory reactions in patients with both ulcerative (UC) and Crohn's colitis. We examined the local and systemic appearance of tumor necrosis factor (TNF) and interleukin 6 (IL-6) in an experimental animal model of inflammatory bowel disease. Colitis was induced in CBA/J mice by adding dextran sulfate sodium (DSS), 5% (wt/vol), to their water. DSS-induced colitis is a reproducible animal model for evaluating the role of cytokines in the pathology of colitis. Animals were weighed daily, and stools were checked for the presence of blood. Groups of mice were killed daily, blood samples were taken for measurement of plasma cytokine levels, and colonic samples were taken for histology and measurement of TNF and IL-6 bioactivity. Mice fed DSS developed colitis with bloody diarrhea, weight loss, and colonic inflammation by days 5-9. Histologic examination of the colons showed focal crypt destruction and ulceration. In mice with DSS-induced colitis no TNF was detectable in colonic tissue extracts or in plasma. In contrast, plasma IL-6 was detectable from days 4 to 9 and was detectable in colonic tissue in only a few (two of four) terminally ill animals on day 9. Animals were injected with a neutralizing, polyclonal anti-TNF antiserum that maintained high in vivo neutralizing titers for > or = 48 h. This anti-TNF antiserum failed to block or modify the severity of colitis induced by DSS. Failure to detect local or systemic TNF and failure to prevent colonic inflammation with anti-TNF antiserum showed that TNF is not an inflammatory mediator in DSS-induced murine colitis.

Growth hormone activation of human monocytes for superoxide production but not tumor necrosis factor production, cell adherence, or action against Mycobacterium tuberculosis

We have previously demonstrated that growth hormone (GH) is a human macrophage-activating factor which primes monocytes for enhanced production of H2O2 in vitro. This report extends our observations to other monocyte functions relevant to infection. We find that GH also primes monocytes for O2- production, to a degree similar to the effect of gamma interferon. Neither macrophage-activating factor alone stimulates monocytes to release bioactive tumor necrosis factor. However, GH, unlike gamma interferon, does not synergize with endotoxin for enhanced tumor necrosis factor production. In further contrast, GH does not alter monocyte adherence or morphology, while phagocytosis and killing of Mycobacterium tuberculosis by GH-treated monocytes are also unaffected. Therefore, despite the multiplicity of the effects of GH on the immune system in vivo, its effects on human monocytes in vitro appear to be limited to priming for the release of reactive oxygen intermediates.

Inflammatory cytokine expression in swine experimentally infected with Actinobacillus pleuropneumoniae

An Actinobacillus pleuropneumoniae infection model in swine was established to study the expression of inflammatory cytokines during acute respiratory disease. Lavage fluid, lavage cells consisting primarily of alveolar macrophages, and lung tissue were analyzed for the presence of various cytokines at 2, 4, 8, and 24 h following endotracheal inoculation of A. pleuropneumoniae. Interleukin-1 beta (IL-1) and IL-8 mRNA levels were elevated within 2 h in lavage cells of animals inoculated with A. pleuropneumonia but not in cells from controls treated with saline-bovine serum albumin, based on Northern (RNA blot) analysis. Tumor necrosis factor (TNF) mRNA was present at low levels in all animals, and the level was not increased at any time point. In situ hybridization was more sensitive than Northern blotting and revealed elevations of all three cytokines in lavage cells within 2 to 4 h of A. pleuropneumoniae inoculation. IL-6 was detected in lavage cells by in situ hybridization but not by Northern blotting. In lung tissue obtained 18 to 24 h after A. pleuropneumoniae instillation, all cytokine mRNAs, including that of IL-6, were detected by Northern blot analysis. The levels of bioactive IL-1 and IL-6 in lavage fluids increased approximately 1,000-fold following A. pleuropneumoniae inoculation, but TNF bioactivity was not detected. Morphological localization of cytokine mRNAs by in situ hybridization indicated markedly increased levels of TNF, IL-1, and IL-8 mRNAs at the periphery of focal lung lesions. These findings indicate that inflammatory cytokines, particularly IL-1 and IL-8, are associated with the development of pleuropneumonia and may contribute to disease severity.

THE GUT

The aim of this study was to test the hypothesis that the gut is capable of becoming a cytokine-generating organ following either a lethal or nonlethal inflammatory insult. Adult male rats were given an intraperitoneal challenge with saline, or with a nonlethal (.1 mg/g) or LD50 (.5 mg/g) dose of zymosan. Mesenteric lymph nodes, efferent mesenteric lymph, liver, spleen, and blood (portal and systemic) were obtained at 2, 4, 6, 8, or 10 h post challenge. Organs, lymph, and blood were tested for bacterial translocation (BT); blood and lymph were assayed for tumor necrosis factor (TNF) and IL-6. After .1 mg/g zymosan, BT was limited to the mesenteric lymph node complex only; .5 mg/g zymosan promoted BT to blood, mesenteric lymph, and organs (p < .05 vs. control or .1 mg/g zymosan). The magnitude of portal bacteremia was greater than systemic bacteremia (p < .003). Serum TNF peaked at 2 h (p < .05 vs. control), and serum IL-6 peaked at 4-6 h (p < .05 vs. control) post zymosan challenge. Portal and systemic bioactivity was similar for either cytokine, and serum bioactivity did not correlate with zymosan dose. TNF bioactivity was increased in the mesenteric lymph at 2 h post challenge with .5 mg/g zymosan only (p < .05 vs. control or .1 mg/g zymosan). IL-6 bioactivity was increased in the mesenteric lymph at 4 through 10 h post zymosan challenge (p < .05 vs. control), but was similar with either dosage of zymosan. In conclusion, the gut may be capable of producing cytokines in response to an inflammatory stimulus, even in the absence of portal or systemic spread of bacteria. The magnitude of the cytokine response does not correlate with the magnitude of bacterial translocation.

Bioactive tumor necrosis factor in the sputa of cystic fibrosis patients

Eighty sputum samples from 37 cystic fibrosis patients were assayed for tumor necrosis factor alpha (TNF-alpha) in the L929 bioactivity assay. In addition, the presence of TNF-alpha-specific mRNA in the sputum was determined by using TNF-alpha-specific DNA probes in a PCR amplification step after reverse transcription of the mRNA to cDNA. Bioactive TNF-alpha was detected in 69 (86%) of the 80 sputum samples tested and, on at least one occasion, in 35 (95%) of the 37 cystic fibrosis patients sampled. The highest levels of TNF-alpha were observed in sputa obtained from patients presenting with an exacerbation of their disease.

Differential effects of in vivo ethanol on LPS-induced TNF and nitric oxide production in the lung

Alcohol (EtOH) has been shown to suppress lipopolysaccharide (LPS)-induced nitric oxide (NO) generation and tumor necrosis factor (TNF) production in the lung in vivo. We have previously reported that EtOH suppressed gene expression for inducible nitric oxide synthase (iNOS) with a subsequent decrease in release of reactive nitrogen intermediates by alveolar macrophages and recruited lung neutrophils. We hypothesized that a similar mechanism may be involved in EtOH-induced suppression of LPS-stimulated TNF production. In contrast to what we found with iNOS, EtOH had no effect on TNF mRNA in alveolar macrophages or recruited lung neutrophils. However, immunoreactive and bioactive TNF was reduced by 72%. EtOH treatment resulted in an increased level of the membrane-bound 26-kDa form of TNF, which suggested that proteolytic cleavage of this prohormone was affected by EtOH. Experiments with t-butyl alcohol, a tertiary alcohol that is not metabolized to acetaldehyde, yielded similar results. Thus EtOH appears to be the active substance in suppression of TNF in the lung in vivo. Pretreatment with intratracheal interferon-gamma 24 h before intratracheal LPS increased TNF bioactivity partly due to increased TNF mRNA and by increasing TNF processing, as evidenced by a decrease in the 26-kDa TNF prohormone and an increase in immunoreactive and bioactive TNF.