Effects of soluble TNF receptor treatment on lipopolysaccharide-induced myocardial cytokine expression

Tumor necrosis factor (TNF)-alpha plays a key role in the pathogenesis of septic shock syndrome, and myocardial TNF-alpha expression may contribute to this pathophysiology. We examined the myocardial expression of TNF-alpha-related cytokines and chemokines in mice exposed to lipopolysaccharide (LPS) and tested the effects of anti-TNF therapy on myocardial cytokine expression. Cytokine mRNA levels were measured by RNase protection assay, and protein levels in the plasma and myocardium were assessed by enzyme-linked immunosorbent assays. LPS (4 microg/g body wt ip) induced marked cytokine expression, including TNF-alpha, interleukin (IL)-1beta, IL-6, and monocyte chemotactic protein (MCP)-1, in both the plasma and myocardium. Pretreatment with adenovirus-mediated TNF receptor fusion protein (AdTNFR1; 10(9) plaque-forming units iv) decreased plasma cytokine levels. In contrast, whereas myocardial IL-1beta expression was also suppressed, expression of IL-6 and MCP-1 was not inhibited by AdTNFR1. In summary, anti-TNF treatment differentially altered the cytokine expression in the plasma and myocardium during endotoxemia. Inability to block myocardial expression of IL-6 and MCP-1 suggests a possible mechanism for the failure of anti-TNF therapies in the treatment of endotoxin shock.     

Inducible nitric oxide synthase mediates cytokine release: the time course in conscious and septic rats

Nitric oxide (NO), tumor necrosis factor-alpha (TNF-alpha), and interleukin 1-beta (IL-1beta) are postulated to play a key pathophysiologic role during sepsis. In this study, we examined the time course of inducible NO synthase (iNOS) mRNA expression and the plasma TNF-alpha and IL-1beta in lipopolysaccharide (LPS)-treated conscious rats. The hemodynamic pattern in septic shock is more similar to clinical conditions without anesthesia. The data showed that a significant increase in iNOS mRNA levels was found in the spleen, lung, liver, with slight elevation in the heart and kidney at 3 h after LPS administration. However, iNOS mRNA levels were not elevated significantly in all tissues examined at 24 h. In the plasma, TNF-alpha and IL-1beta culminated within 1 h, and reduced gradually to baseline levels in a relatively short period (within 9 h). The results suggest that local NO production by activation of iNOS mRNA expression and cytokine release may contribute to LPS-induced organ dysfunction at various time points.     

Selective macrophage suppression during sepsis

Polymicrobial sepsis induces suppression of macrophage function as determined by a reduction of pro-inflammatory cytokine production upon re-exposure to lipopolysaccharide (LPS) in vitro. We examined whether macrophages were refractory to only LPS challenge or if they were immunoparalyzed and unable to respond to other stimuli such as lipoteichoic acid (LTA) or zymosan (ZYM). This study evaluated the capacity of peritoneal macrophages to produce pro-inflammatory and anti-inflammatory cytokines as well as chemokines following mild or severe sepsis induced by cecal ligation and puncture (CLP). Peritoneal macrophages were isolated 29 h after CLP and challenged with different stimuli. LPS was a more potent stimulus for cytokine induction than LTA or ZYM in both mild and severe sepsis. In mild sepsis, the macrophage cytokine response to LPS was selective and less refractory than in severe sepsis. While production of IL-6 and KC was reduced, secretion of TNF-alpha and MIP-1alpha was enhanced in those cells isolated from mice with mild sepsis. Production of IL-10 and the IL-1 receptor antagonist , MIP-2, and MCP-1 in response to LPS stimulation was equivalent to the amount produced by naive macrophages. Our results indicate that macrophages are not immunoparalyzed during sepsis and may still be induced to secrete some inflammatory mediators.     

Effects of IL-18 and IL-10 pre-treatment on the alteration of endogenous cytokines in liver and spleen of mice with experimental endotoxemia

Mechanisms of interleukin-18 (IL-18) and interleukin-10 (IL-10) in lipopolysaccharide (LPS) induced endotoxemia are not clear; their protective role is being investigated so that they may effectively modulate the host cytokine levels during endotoxemia. The aim of the study was to evaluate protective effects of IL-18 and IL-10 in experimentally induced endotoxemia in mice correlating the changes in tissue anti-oxidant enzymes and circulating cytokines. Liver injury was determined by estimation of serum glutamate oxalate transaminase (SGOT) and serum glutamate pyruvate transaminase (SGPT), serum nitric oxide (NOx), hepatic anti-oxidant enzyme and cytokine content in LPS (250 microg/kg) induced endotoxemic mice receiving either IL-18 (500 ng/mouse) or IL-10 (600 ng/mouse) treatment. Mice (87% of IL-10 treated and 74% of IL-18 treated) survived when administered prior to LPS challenge. Pre-treatment of mice with either IL-10 or IL-18 followed by LPS, lead to reduction in SGPT and SGOT level, serum NOx, and altered hepatic anti-oxidant enzymes activity and myeloperoxidase activity than the only LPS treated group. Marked reduction in the amounts of LPS-induced hepatic and splenic TNF-u content has been observed after IL-10 pre-treatment. Results suggested that attenuating the induction of TNF-alpha and IFN-gamma and subsequent induction of nitric oxide formation in response to LPS may in part account for efficient protection by IL-18 and IL-10 in the reduction of LPS-induced liver injury.     

Carbohydrate composition of meningococcal lipopolysaccharide modulates the interaction of Neisseria meningitidis with human dendritic cells

Meningococcal lipopolysaccharide (LPS) is of crucial importance for the pathogenesis of invasive infection. We show that sialylation and elongation of the alpha-chain effectively shields viable unencapsulated Neisseria meningitidis from recognition by human dendritic cells (DC). In contrast, beta- and gamma- chain of the LPS carbohydrate moiety play only a minor role in the interaction with DC. The protective function of the LPS for the bacteria can be counteracted in vivo by phase variation of the lgtA gene encoding LPS glycosyltransferase A. Capsule expression protects N. meningitidis efficiently from recognition and phagocytosis by DC independent of the LPS structure. Despite the significant impact of LPS composition on the adhesion and phagocytosis of N. meningitidis no differences were found in terms of cytokine levels secreted by DC for IL1-beta, IL-6, IL-8, TNF-alpha, IFN-gamma and GM-CSF. However, significantly lower levels of the regulatory mediator IL-10 were induced by encapsulated strains in comparison to isogenic unencapsulated derivatives. IL-10 secretion was shown to depend on phagocytosis because poly alpha-2,8 sialic acid did not influence IL-10 secretion. The use of truncated LPS isoforms in vaccine preparations can therefore not only result in attenuation but also in more efficient targeting of DC.     

Evidence for IFN-gamma as a mediator of the lethality of endotoxin and tumor necrosis factor-alpha.

Current evidence indicates that endogenously produced peptide cytokines, most notably TNF-alpha and IL-1, mediate the lethality of experimental endotoxemia. Because circulating serum levels of IFN-gamma can be detected soon after TNF-alpha and IL-1 in response to endotoxin, we investigated the role of IFN-gamma in endotoxin and TNF-alpha lethality. Specific neutralizing antibodies to murine TNF-alpha (anti-TNF-alpha Ab) or murine IFN gamma (anti-IFN-gamma Ab) produced in our laboratory protected mice against the lethality of Escherichia coli endotoxin (LPS) administered 6 h later. Serum IFN-gamma levels 2 h after i.v. LPS were lower in mice treated with anti-TNF-alpha Ab compared to mice that received nonimmune IgG (median less than 2.5 vs 3.0 U/ml, P2 less than 0.05). In contrast, serum TNF-alpha levels 1 h after i.v. LPS peaked more than fourfold higher in mice treated with anti-IFN-gamma Ab compared to controls (median greater than 6400 vs 1405 pg/ml, p2 less than 0.05). Doses of TNF-alpha (300 micrograms/kg) and IFN-gamma (50,000 U) which were well tolerated when given individually were synergistically lethal in combination (0% lethality vs 100% lethality, P2 less than 0.001), and were associated with higher serum levels of IL-6 than with either cytokine alone. Anti-IFN-gamma Ab provided complete protection against exogenous human rTNF-alpha at the LD100 dose (1400 micrograms/kg, p2 less than 0.001), and in fact prevented lethality at doses four- to fivefold greater than the LD100 human rTNF-alpha (up to 6000 micrograms/kg). We conclude that IFN-gamma is synergistic with TNF-alpha, is essential for the lethality of LPS and TNF-alpha, and may have modulating effects on the negative control of serum levels of TNF-alpha after LPS in mice.     

Splenectomy differentially influences immune responses in various tissue compartments of the body

Patients without spleens have an increased risk of infection. Previous studies have shown that splenectomy (Spx) influences Kupffer cells (KC), peritoneal macrophages (pMphi) and alveolar macrophages (aMphi) phagocytosis and bactericidal activity. This study examined the effect of Spx on the cytokine production by peripheral blood monocular cells (PBMC), KC, aMphi, pMphi and cells from mesenteric lymph nodes (MLN). We also determined if Spx influences survival following sepsis induced by cecal ligation and puncture (CLP). C57BL/6J male mice (8-10 weeks old) underwent sham operation or Spx. Two weeks after sham or Spx, KC, pMphi, aMphi, PBMC and cells from MLN were isolated and stimulated with LPS. Cell-free supernatants were analyzed for TNF-alpha, IL-6, and IL-10. A significant increase in KC, pMphi and aMphi TNF-alpha and IL-6 release was observed following Spx. The production of IL-10 was also significantly higher in KC under those conditions. In contrast, the release of TNF-alpha, IL-6 and IL-10 was significantly decreased in PBMC after Spx. Similarly, TNF-alpha and IL-6 was also decreased in MLN after Spx. Overall survival after CLP was not different in mice subjected to either sham or Spx. However, the mean time to death was significantly decreased in mice subjected to Spx compared to sham injured mice. These findings suggest that Spx modulates immune cell functions in various tissue compartments of the body and results in early deaths following sepsis.     

Synergistic induction of interleukin-6 production and gene expression in human thymic epithelial cells by LPS and cytokines

We examined the ability of LPS and several cytokines (TNF-alpha, IL-1-beta, IFN-gamma, IL-4) to modulate IL-6 production by cultured human thymic epithelial cells (TEC). IL-6 activity was measured by the hybridoma growth factor biological activity. Moderate but detectable IL-6 activity was spontaneously produced in the presence of serum proteins. LPS as well as the cytokines TNF-alpha and IL-1-beta was a potent inducer of IL-6, increasing, respectively, IL-6 levels by 9-, 28-, and 75-fold (mean values) while IL-4 and IFN-gamma provoked no significant effect. Interestingly, clearly different kinetics were observed for IL-6 induction by the various activation agents, the maximal effect being reached at 24, 48, and 72 hr, respectively for LPS, TNF-alpha, and IL-1-beta. Moreover, a synergistic effect of TNF-alpha and either LPS or IL-1-beta was observed. Indeed, TEC incubated with the cytokines in combination at optimal doses produced 5- to 170-fold more IL-6 than TEC stimulated with the cytokines individually. Neutralizing anti-IL-6 polyclonal and monoclonal antibodies completely blocked hybridoma proliferation stimulating activity of TEC supernatants; thus, implying that this activity is essentially due to IL-6. In situ hybridization analysis of cytocentrifuged TEC with an mRNA antisense probe specific for human IL-6 and labeled with 35S demonstrated that up to 90% of TEC could be induced to express the IL-6 gene. Computer-aided quantification of IL-6 mRNA levels indicated that upon stimulation with TNF-alpha combined to LPS, both the numbers of cells expressing IL-6 mRNA and the amounts of cytoplasmic IL-6 mRNA per cell were increased. Taken altogether these results demonstrate that LPS and/or cytokines can modulate and synergistically stimulate IL-6 production. In addition to a possible role in regulating normal thymic T cell activation, the IL-6 produced by TEC could be of pathophysiological relevance in disregulated situations such as in hyperplastic thymuses from patients with myasthenia gravis.     

IL-4 pretreatment selectively enhances cytokine and chemokine production in lipopolysaccharide-stimulated mouse peritoneal macrophages

Although well recognized for its anti-inflammatory effect on gene expression in stimulated monocytes and macrophages, IL-4 is a pleiotropic cytokine that has also been shown to enhance TNF-alpha and IL-12 production in response to stimulation with LPS. In the present study we expand these prior studies in three areas. First, the potentiating effect of IL-4 pretreatment is both stimulus and gene selective. Pretreatment of mouse macrophages with IL-4 for a minimum of 6 h produces a 2- to 4-fold enhancement of LPS-induced expression of several cytokines and chemokines, including TNF-alpha, IL-1alpha, macrophage-inflammatory protein-2, and KC, but inhibits the production of IL-12p40. In addition, the production of TNF-alpha by macrophages stimulated with IFN-gamma and IL-2 is inhibited by IL-4 pretreatment, while responses to both LPS and dsRNA are enhanced. Second, the ability of IL-4 to potentiate LPS-stimulated cytokine production appears to require new IL-4-stimulated gene expression, because it is time dependent, requires the activation of STAT6, and is blocked by the reversible protein synthesis inhibitor cycloheximide during the IL-4 pretreatment period. Finally, IL-4-mediated potentiation of TNF-alpha production involves specific enhancement of mRNA translation. Although TNF-alpha protein is increased in IL-4-pretreated cells, the level of mRNA remains unchanged. Furthermore, LPS-stimulated TNF-alpha mRNA is selectively enriched in actively translating large polyribosomes in IL-4-pretreated cells compared with cells stimulated with LPS alone.     

Regulation of synovial cell proliferation and prostaglandin E2 production by combined action of cytokines

To study the causes of synovitis in rheumatoid arthritis (RA), we have analyzed the effect of several cytokines known to be secreted in RA joints, on synovial cell proliferation and prostaglandin E2 (PGE2) production. Recombinant interleukin-1-beta (IL-1-beta) and tumor necrosis factor-alpha (TNF-alpha) stimulated moderately the DNA synthesis and markedly the production of PGE2. Interferon-gamma (IFN-gamma) was often mitogenic but never induced PGE2 secretion. The association of IL-1-beta and TNF-alpha showed an additive effect on both parameters, whereas addition of IFN-gamma to either monokine reduced the proliferation and increased PGE2 release. Incubation with a crude T cell supernatant or a mixture of cytokines including IL-1-beta, TNF-alpha and IFN-gamma enhanced synovial cell growth and PGE2 production as compared to the effect elicited by each single cytokine. In contrast, interleukin-2 (IL-2) down regulated the synovial cell activation induced by the combined action of the three other cytokines. Taken together, our findings indicate that synovial cell proliferation is weakly stimulated, reaching a two-fold increase over background levels, whatever cytokines are used. Furthermore, proliferation can vary independently of PGE2 production. Nevertheless, the monokines IL-1-beta and TNF-alpha both exert agonistic effects on synovial cell activation, thus contributing to cartilage damage in RA, whereas IFN-gamma, IL-6 or IL-2 may rather play a regulatory role.     

Endotoxin and Staphylococcus aureus enterotoxin A induce different patterns of cytokines.

The effects of Staphylococcus aureus enterotoxin A (SEA) and lipopolysaccharide (LPS) in cytokine production were assessed at the single cell level in cells obtained from healthy blood donors. Cytokine production was studied with UV-microscopy of fixed and permeabilized cells stained with cytokine specific monoclonal antibodies. The cytokines evaluated included tumour necrosis factor (TNF)-alpha, interleukin (IL)-1 alpha, IL-1 beta, IL-6, IL-8, IL-10, IL-2, IL-4, interferon (IFN)-gamma and TNF-beta. LPS exhibited marked production of IL-1 alpha, IL-1 beta, TNF-alpha, IL-6 and IL-8. After LPS stimulation IL-1 alpha, IL-1 beta, TNF-alpha and IL-8 were the dominating products, all peaking at or before 4 hours after cell stimulation. In addition, IL-10 production was evident after 12 hours of cell stimulation. The T-lymphocyte-derived cytokines TNF-beta, IL-2, IFN-gamma and IL-4 were never detected in the cultures. All cytokine production, except IL-8, was downregulated at 96 hours. In contrast, peak production of IL-1 alpha, IL-1 beta and IL-8, which were the dominant products, occurred after 12 hours in the SEA-stimulated cultures. Further, a significant T-lymphocyte production of TNF-beta, TNF-alpha, IFN-gamma and IL-2 was found with peak production 12-48 hours after initiation. Only low amounts of IL-6 were evident. The two types of cytokine pattern and kinetics found may correspond to the different clinical conditions after invasive Gram-negative Escherichia coli vs Gram-positive Staphylococcus aureus infections in humans, with a much more rapid onset of disease after E. coli infections.(ABSTRACT TRUNCATED AT 250 WORDS)     

IL-8 in septic shock, endotoxemia, and after IL-1 administration

Much effort has been directed toward elucidating the host response to sepsis and inflammation, resulting in the definition of a cascade of endogenous mediators that direct metabolic and immunological responses. Here we report that IL-8, a novel cytokine produced by a variety of cells in vitro in response to stimulation with bacterial LPS and the proinflammatory cytokines, appears in the circulation of primates in vivo during septic shock, sublethal endotoxemia, and after the administration of IL-1 alpha. The magnitude of the IL-8 response correlates with the severity of the insult, and levels of IL-8 peak relatively late, after those of TNF-alpha and IL-1 beta, and simultaneously with those of IL-6. IL-8 has been primarily defined as a selective activator and chemoattractant of neutrophils, and we demonstrate that after LPS or IL-1 alpha infusion, circulating neutrophil numbers rapidly recover from an initial neutropenia while IL-8 concentrations are maximal, supporting the hypothesis that IL-8 influences circulating leukocyte populations in vivo. We conclude that IL-8 is another participant in the cytokine cascade elicited by sepsis and inflammation and, as such, may play a significant role in host defense and disease.     

TNF-alpha is a potent inducer for IFN-inducible protein-10 in hepatocytes and unaffected by GM-CSF in vivo, in contrast to IL-1beta and IFN-gamma

We have recently shown that IFN-inducible protein 10 (IP-10), a member of the CXC chemokine family, is induced in hepatocytes surrounded by infiltrative mononuclear cells in human livers with chronic hepatitis. Hence, we examined the kinds of stimuli that can induce IP-10 expression in hepatocytes in vivo. While the liver expressed three chemokine genes (IP-10, JE/MCP-1, KC/GRO) in a tissue-specific fashion following systemic treatment with pro-inflammatory cytokines, IP-10 mRNA expression showed the most marked liver-specificity. Pretreatment with GM-CSF selectively inhibited IL-1beta, but not TNF-alpha-induced IP-10 mRNA expression. In situ hybridization analysis in the liver and Northern hybridization analysis in isolated liver cell fractions from rodents treated with pro-inflammatory cytokines revealed cellular sources of chemokine expression. IP-10 mRNA expression in hepatocytes was induced by i.v. administration of TNF-alpha, and to a much lesser extent in response to IL-1beta and IFN-gamma, whereas Kupffer cells and endothelial cells expressed IP-10 mRNA equivalently in response to these three stimuli. On the other hand, JE/MCP-1 mRNA expression was detected only in non-parenchymal cells in response to TNF-alpha and IL-1beta, but not in response to IFN-gamma. KC/GRO mRNA expression was also induced mainly in sinusoidal cells by treatment with TNF-alpha or IL-1beta, although it was detected to a lesser extent in hepatocytes. Our results demonstrated that chemokine induction is stimulus-, tissue- and cell type-specific and that IP-10 (but not MCP-1) is inducible in hepatocytes by TNF-alpha most potently, even in the presence of GM-CSF, suggesting the specific role of TNF-alpha-induced IP-10 on intralobular mononuclear infiltration in chronic hepatitis.     

Assessment of Blood-Brain Barrier Function and the Neuroinflammatory Response in the Rat Brain by Using Cerebral Open Flow Microperfusion (cOFM)

Blood-brain barrier (BBB) impairment in systemic inflammation leads to neuroinflammation. Several factors including cytokines, chemokines and signal transduction molecules are implicated in BBB dysfunction in response to systemic inflammation. Here, we have adopted a novel in vivo technique; namely, cerebral open flow microperfusion (cOFM), to perform time-dependent cytokine analysis (TNF-alpha, IL-6 and IL-10) in the frontal cortex of the rat brain in response to a single peripheral administration of lipopolysaccharide (LPS). In parallel, we monitored BBB function using sodium fluorescein as low molecular weight reporter in the cOFM sample. In response to the systemic LPS administration, we observed a rapid increase of TNF-alpha in the serum and brain, which coincides with the BBB disruption. Brain IL-6 and IL-10 synthesis was delayed by approximately 1 h. Our data demonstrate that cOFM can be used to monitor changes in brain cytokine levels and BBB disruption in a rat sepsis model.     

Modulation of astrocyte inducible nitric oxide synthase and cytokine expression by interferon beta is associated with induction and inhibition of interferon gamma-activated sequence binding activity

Although interferon (IFN)-beta is firmly established as a therapeutic agent for multiple sclerosis, information regarding its role in astrocyte cytokine production is limited. In primary cultures of human astrocytes, we determined the effects of IFN-beta on astrocyte cytokine [tumor necrosis factor-alpha (TNF-alpha) and interleukin (IL)-6] and inducible nitric oxide synthase (iNOS) expression by ribonuclease protection assay and ELISA. We found that IFN-beta inhibited astrocyte cytokine/iNOS induced by IL-1 plus IFN-gamma, but in the absence of IFN-gamma, IFN-beta enhanced IL-1-induced cytokine/iNOS expression. Electrophoretic mobility shift analysis (EMSA) demonstrated that IFN-gamma induced sustained IFN-gamma-activated sequence (GAS) binding, while IFN-beta induced transient GAS binding. When used together, IFN-beta inhibited IFN-gamma-induced GAS binding activity. Nuclear factor-kappa B (NF-kappaB) activation was not altered by either IFNs, whereas IFN stimulated response element (ISRE) was only activated by IFN-beta and not IFN-gamma. These results suggest that IFN-beta can both mimic and antagonize the effect of IFN-gamma by modulating induction of nuclear GAS binding activity. Our results demonstrating differential regulation of astrocyte cytokine/iNOS induction by IFN-beta are novel and have implications for inflammatory diseases of the human CNS.     

Post-treatment with N-acetylcysteine ameliorates endotoxin shock-induced organ damage in conscious rats

N-acetylcysteine (NAC) is an antioxidant and cytoprotective agent with scavenging action against reactive oxygen species and inhibitory effects on pro-inflammatory cytokines. In a previous study, we found that pretreatment with NAC attenuated organ dysfunction and damage, reduced the production of free radicals, tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) following endotoxemia elicited by administration of lipopolysaccharide (LPS). In the present study, we tested the effects of post-treatment with NAC on the sepsis-induced change. Post-treatment imitates clinical therapeutic regimen with administration of drug after endotoxemia. Endotoxin shock was induced by intravenous injection of Klebsiella pneumoniae LPS (10 mg/kg) in conscious rats. Mean arterial pressure (MAP) and heart rate (HR) were continuously monitored for 48 h after LPS administration. NAC was given 20 min after LPS. Measurements of biochemical substances were taken to reflect organ functions. Biochemical factors included blood urea nitrogen (BUN), creatinine (Cre), lactate dehydrogenase (LDH), creatine phosphokinase (CPK), aspartate transferase (GOT), alanine transferase (GPT), TNF-alpha, interleukin-6 (IL-6), and interleukin-10 (IL-10). LPS significantly increased blood BUN, Cre, LDH, CPK, GOT, GPT, TNF-alpha, IL-6, IL-10 levels and HR, and decreased MAP. Post-treatment with NAC diminished the decrease in MAP, increased the HR, and decreased the markers of organ injury (BUN, Cre, LDH, CPK, GOT, GPT) and inflammatory biomarkers (TNF-alpha, IL-6, IL-10) after LPS. We conclude that post-treatment with NAC suppresses the release of plasma TNF-alpha, IL-6, and IL-10 in endotoxin shock, and decreases the markers of organ injury. These beneficial effects protect against LPS-induced kidney, heart and liver damage in conscious rats. The beneficial effects may suggest a potential chemopreventive effect of this compound after sepsis.     

Androstenediol ameliorates alterations in immune cells cytokine production capacity in a two-hit model of trauma-hemorrhage and sepsis

Although administration of androstenediol (a metabolite of dehydroepiandrosterone) following trauma-hemorrhage (T-H) produces beneficial effects on inflammatory cytokines and organ function, it remains unknown whether this metabolite has any salutary effects in preventing alterations in immune cell cytokine production following a combined insult of T-H and sepsis. To examine this, male rats underwent laparotomy, hemorrhagic shock (mean BP 40 mmHg for 90 min) and resuscitation or sham operation. Androstenediol (1 mg/kg BW i.v.) or vehicle was administered at the end of resuscitation. Twenty hrs after T-H or sham operation, sepsis was induced by cecal ligation and puncture (CLP). Five hours thereafter, plasma cytokine levels and cytokine production of various immune cells were determined. In a separate set of experiments, survival was monitored for 10 days after the induction of sepsis. Administration of androstenediol markedly decreased plasma IL-6 and TNF-alpha levels following T-H and CLP. Furthermore, it prevented the increased production of IL-6 and TNF-alpha by Kupffer cells and alveolar macrophages and attenuated the decrease in IL-6 and TNF-alpha production by splenic macrophages; however, it had no significant effects on the depressed IL-6 and TNF-alpha production by PBMC following T-H and CLP. The depressed IL-2 and IFN-gamma production by splenocytes under those conditions was attenuated by the administration of androstenediol. Furthermore, survival rate following T-H and subsequent sepsis was improved by androstenediol treatment. Since androstenediol administration following T-H attenuated cytokine production and reduced mortality in a double-hit model of T-H and sepsis, this agent appears to be a novel and useful adjunct for maintaining the immune cell functions following T-H and for decreasing the mortality rate from subsequent susceptibility to sepsis.     

Systemic and local cytokine response of young piglets to oral infection with<Emphasis Type="Italic">Salmonella enterica</Emphasis> serotype typhimurium

One-week-old breast-fed miniature piglets were orally infected either with virulent LT2 strain or with a non-virulent SF1591 rough mutant of Salmonella Typhimurium for 1 d. Both microorganisms were cultivated from mesenteric lymph nodes but not from the blood of infected piglets. Interleukins (IL) 1 beta, 8, 18, tumor necrosis factor alpha (TNF-alpha) and interferon gamma (IFN-gamma) were quantified by ELISA in plasma and washes of a terminal part of the small bowel. In plasma, cytokines were mostly missing in non-infected piglets and either missing or low in infected piglets. In the gut of non-infected piglets, IL-1 beta, IL-8 and IL-18 were detected whereas TNF-alpha and IFN-gamma were mostly missing. IFN-gamma levels highly increased (p < 0.05) after infection with nonvirulent salmonellae. The variability of cytokine levels in the gut of suckling piglets is discussed.