laminaran and Sepsis

The diagnosis of neonatal invasive fungal disease (IFD) is difficult and often delayed. The platelet parameters and (1, 3)-β-D-Glucan (BG) may be useful for diagnosing IFD, but their diagnostic performance are not well characterized in neonates. We studied 63 preterm infants with IFD, 160 preterm infants without sepsis (preterm control), and 41 preterm infants with bacterial sepsis. Platelet parameters at the first day of onset of IFD and at the fourteenth day after antifungal treatment were evaluated. Serum BG was measured. Platelet count (PC), plateletcrit (PCT), and platelet distribution width (PDW) values were significantly lower, and mean platelet volume (MPV) values significantly higher in the IFD versus preterm control infants. PC and PCT values were much lower in infants with IFD versus bacterial sepsis, and there were significant differences in BG value between the two groups. After 14 days of antifungal treatment, significant elevations in PC, PCT, PDW and reductions in MPV levels in IFD group were observed. Receiver operating characteristic (ROC) curves showed that PC and PCT were strong predictors of IFD. The PC and PCT cut-offs for predicting IFD were 119.5 (sensitivity 78%, specificity 95%) and 0.21 (sensitivity 83%, specificity 85%), respectively. There were significant differences in PC and PCT levels between deceased and survived patients. The PC and PCT cut-offs for predicting deceased IFD were 39 (sensitivity 62%, specificity 86%) and 0.04 (sensitivity 50%, specificity 95%), respectively. The sensitivity in diagnosing IFD by a BG cutoff of ≥10 pg/ml was 68.3% and specificity was 75.6%. PC and PCT levels in the BG ≥400 pg/ml group were significantly lower compared to the BG<400 pg/ml group. Platelet parameters and BG could be useful biomarkers for the diagnosis and prognosis of neonatal IFD.

Topics: beta-Glucans; Biomarkers; Blood Platelets; Case-Control Studies; Female; Fluconazole; Humans; Infant, Newborn; Infant, Premature; Male; Mean Platelet Volume; Mycoses; Platelet Count; Prognosis; ROC Curve; Sensitivity and Specificity; Sepsis

Procalcitonin serum level has been recommended as a new marker of bacterial infectious diseases. The aim of this prospective, multicenter study was to determine the clinical usefulness of procalcitonin in differentiating patients with sepsis from those with severe sepsis. Eighty-two patients were enrolled: 20 without systemic inflammatory response syndrome (SIRS), 9 with SIRS, 34 with sepsis, and 19 with severe sepsis. The patients with severe sepsis had significantly higher procalcitonin levels (median, 36.1 ng/ml) than those with sepsis (median, 0.6 ng/ml). With a procalcitonin cutoff value of 2.0 ng/ml, sensitivity for the detection of severe sepsis and specificity for the detection of sepsis were 94.7% and 78.1%, respectively. A good correlation was found between the serum procalcitonin level and the Sepsis-Related Organ Failure Assessment (SOFA) score (r = 0.680), although no correlation was found between the C-reactive protein (CRP) level and the SOFA score. In conclusion, the procalcitonin serum level may be useful not only for aiding the diagnosis of sepsis but also for discriminating between sepsis and severe sepsis.

Topics: APACHE; beta-Glucans; Biomarkers; C-Reactive Protein; Calcitonin; Calcitonin Gene-Related Peptide; Endotoxins; Glycoproteins; Humans; Interleukin-6; Predictive Value of Tests; Prospective Studies; Protein Precursors; Sepsis; Severity of Illness Index

We examined the effect of modulating phosphoinositide 3-kinase (PI3K) activity in a murine model of cecal ligation and puncture-induced polymicrobial sepsis. Inhibition of PI3K activity with wortmannin increased serum cytokine levels and decreased survival time in septic mice. We have reported that an immunomodulator, glucan phosphate, induces protection in murine polymicrobial sepsis. We observed that glucan stimulated tissue PI3K activity, which positively correlated with increased survival in septic mice. We investigated the effect of PI3K inhibition on survival in septic mice treated with glucan. Treatment of mice with the PI3K inhibitors, wortmannin and LY294002, completely eliminated the protective effect of glucan, indicating that protection against septic mortality was mediated through PI3K. Inhibition of PI3K resulted in increased serum levels of IL1-beta, IL-2, IL-6, IL-10, IL-12, and TNF-alpha in septic mice. Apoptosis is thought to play a central role in the response to septic injury. We observed that inhibition of PI3K activity in septic mice resulted in increased splenocyte apoptosis and a change in the anatomic distribution of splenocyte apoptosis. We conclude that PI3K is a compensatory mechanism that suppresses proinflammatory and apoptotic processes in response to sepsis and/or inflammatory injury. Thus, PI3K may play a pivotal role in the maintenance of homeostasis and the integrity of the immune response during sepsis. We also observed that glucan phosphate decreased septic morbidity and mortality through a PI3K-dependent mechanism. This suggests that stimulation of the PI3K pathway may be an effective approach for preventing or treating sepsis and/or septic shock.

Topics: Androstadienes; Animals; Apoptosis; beta-Glucans; Cecum; Chromones; Cytokines; Disease Susceptibility; Enzyme Inhibitors; Glucans; Immunity, Innate; Injections, Intraperitoneal; Ligation; Male; Mice; Mice, Inbred ICR; Morpholines; Organ Specificity; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Punctures; Sepsis; Signal Transduction; Spleen; Survival Analysis; Wortmannin

To determine whether there was a correlation between induction of polymicrobial sepsis, modulation of tissue Toll-like receptor (TLR) gene, and protein expression and survival outcome.. Prospective, randomized animal study.. University medical school research laboratory.. Age- and weight-matched ICR/HSD mice.. Sepsis was induced by cecal ligation and puncture (CLP). No-surgery and sham (laparotomy)-operated mice were controls. We also examined tissue TLR2 and TLR4 messenger RNA and TLR4 protein levels in mice treated with an immunomodulator that increases survival in polymicrobial sepsis. In the immunomodulator study, mice were treated with glucan phosphate (50 mg/kg, intraperitoneally) 1 hr before CLP. No-surgery, sham surgery, glucan + no-surgery, sham surgery + glucan, and CLP groups were employed as controls.. Total RNA was isolated from liver, lung, and spleen at 0, 1, 3, 6, 8, and 24 hrs after CLP. TLR gene expression was assessed by reverse transcription-polymerase chain reaction. Tissue TLR4 protein levels were evaluated at 24 hrs by Western blot and immunohistochemistry. CLP sepsis increased (p <.05) liver and lung TLR2 and TLR4 gene expression compared with controls. TLR4 protein concentrations also were increased. Increased TLR2/4 gene and TLR4 protein expression correlated with mortality. Immunoprophylaxis with glucan phosphate increased (p <.001) long-term survival (20% vs. 70%) but inhibited (p <.05) CLP-induced increases in tissue TLR2 and TLR4 messenger RNA expression as well as TLR4 protein expression.. Early increases in TLR2/4 gene and TLR4 protein expression correlated with mortality, whereas blunting TLR gene and protein expression correlated with improved long-term survival. This suggests that early up-regulation of tissue TLR2/4 may play a role in the proinflammatory response and pathophysiology of polymicrobial sepsis.

Topics: Adjuvants, Immunologic; Analysis of Variance; Animals; beta-Glucans; Gene Expression Regulation; Glucans; Liver; Lung; Male; Membrane Glycoproteins; Mice; Mice, Inbred ICR; Proportional Hazards Models; Random Allocation; Receptors, Cell Surface; RNA, Messenger; Sepsis; Survival Analysis; Time Factors; Toll-Like Receptor 2; Toll-Like Receptor 4; Toll-Like Receptors

Topics: Adjuvants, Immunologic; Animals; beta-Glucans; Gene Expression Regulation; Glucans; Membrane Glycoproteins; Mice; Receptors, Cell Surface; Sepsis; Toll-Like Receptors

Soluble beta-1,3-glucan has been demonstrated to protect against infection and shock in rats and mice, and clinical studies suggest that administration of soluble glucans to trauma/surgical patients decreases septic complications and improves survival. However, little is known about the precise mechanisms by which glucans influence the state of activation of blood cells, which are responsible for the fulminant cytokine production and the activation of the coagulation system observed in serious gram-negative infection. We studied therefore the effect of an underivatized, soluble yeast beta-1,3-glucan and lipopolysaccharide (LPS), either alone or in combination, on tumor necrosis factor-alpha (TNFalpha), interleukin-6 (IL-6), IL-8 and IL-10 secretion and monocyte tissue factor (TF) expression in human whole blood. As expected, LPS induced the secretion of substantial amounts of all measured parameters, whereas only minor amounts of TNFalpha, IL-6, and IL-10 were induced by beta-glucan itself. However, beta-glucan itself induced the production of significant amounts of IL-8 and TF. Soluble beta-1,3-glucan had a strong synergistic effect on the LPS-induced secretion of IL-8, IL-10, and on monocyte TF activity, but not on TNFalpha and 1L-6 production. On the other hand, soluble beta-glucan strongly primed LPS stimulation of all parameters, including TNFalpha and IL-6. beta-Glucan also induced detectable neutrophil degranulation within 15 min, whereas a response to LPS was first detected after 90 min. In conclusion, soluble beta-1,3-glucan upregulated leukocyte activity, both on its own and in concert with LPS.

Topics: beta-Glucans; Biomarkers; Blood Coagulation; Cytokines; Glucans; Humans; In Vitro Techniques; Indicators and Reagents; Lipopolysaccharides; Platelet Activation; Sepsis; Thromboplastin

Glucans are fungal cell wall polysaccharides which stimulate innate immune responses. We determined the minimum unit ligand that would bind to glucan receptors on human U937 cells using laminarin-derived pentaose, hexaose, and heptaose glucan polymers. When U937 membranes were pretreated with the oligosaccharides and passed over a glucan surface, only the heptasaccharide inhibited the interaction of glucan with membrane receptors at a K(d) of 31 microM (95% CI 20-48 microM) and 100% inhibition. However, the glucan heptasaccharide did not stimulate U937 monocyte NFkappaB signaling, nor did it increase survival in a murine model of polymicrobial sepsis. Laminarin, a larger and more complex glucan polymer (M(w) = 7700 g/mol), only partially inhibited binding (61 +/- 4%) at a K(d) of 2.6 microM (99% CI 1.7-4.2 microM) with characteristics of a single binding site. These results indicate that a heptasaccharide is the smallest unit ligand recognized by macrophage glucan receptors. The data also indicate the presence of at least two glucan-binding sites on U937 cells and that the binding sites on human monocyte/macrophages can discriminate between glucan polymers. The heptasaccharide and laminarin were receptor antagonists, but they were not receptor agonists with respect to activation of NFkappaB-dependent signaling pathways or protection against experimental sepsis.

Topics: Animals; Binding Sites; Dose-Response Relationship, Drug; Glucans; Humans; Ligands; Male; Mice; Mice, Hairless; Mice, Inbred ICR; Monocytes; NF-kappa B; Polysaccharides; Protein Binding; Receptors, Immunologic; Sepsis; U937 Cells

Growing evidence supports the role of transcription factor activation in the pathophysiology of inflammatory disorders, sepsis, ARDS, SIRS, and shock. Kinase mediated phosphorylation of IkappaBalpha is a crucial step in the NFkappaB activation pathway. We investigated IKBalpha phosphorylation in murine liver and lung extracts after cecal ligation and puncture (CLP) in the presence and absence of a glucan ligand. ICR mice were subjected to CLP. Unoperated and sham-operated mice served as the controls. Glucan phosphate (50 mg/kg) was administered 1 h before or 15 min after CLP. CLP increased hepatic and pulmonary levels of phospho-IkappaBalpha by 48-192%. Pre- or post-treatment with glucan phosphate decreased (P < 0.05) tissue phospho-IkappaBalpha levels in CLP mice. Phospho-IkappaBalpha in the glucan-CLP group were not significantly different from the unoperated controls. To investigate mechanisms we examined IKKbeta kinase activity, IkappaBalpha phosphorylation and degradation, and NFkappaB activity in a murine macrophage cell line, J774a.1, treated with LPS (1 microg/mL) and/or glucan phosphate (1 microg/mL) for up to 120 min. The glucan ligand blunted LPS-induced IKKbeta kinase activity, phosphorylation and degradation of IkappaBalpha, and NFkappaB nuclear binding activity. The data indicate that one mechanism by which (1-->3)-beta-D-glucan may alter the response to endotoxin or polymicrobial sepsis involves modulation of IKK3 kinase activity with subsequent decreases in IkappaBalpha phosphorylation and NFkappaB activation.

Topics: Animals; beta-Glucans; Cell Line; Cell Nucleus; Cells, Cultured; Cytosol; DNA-Binding Proteins; Gene Expression Regulation; Glucans; I-kappa B Kinase; I-kappa B Proteins; Intestinal Perforation; Ligands; Lipopolysaccharides; Liver; Lung; Macrophages; Male; Mice; Mice, Inbred ICR; NF-kappa B; NF-KappaB Inhibitor alpha; Phosphorylation; Protein Processing, Post-Translational; Protein Serine-Threonine Kinases; Sepsis; Shock, Septic

Recent data implicate the activation of nuclear factor-kappa B (NF-kappa B) and nuclear factor interleukin 6 (NF-IL6) as important steps in the pathophysiologic mechanisms of adult respiratory distress syndrome and systemic inflammatory response syndrome.. This study evaluated the effect of immunomodulating polysaccharides on transcription factor activation, cytokine expression, and mortality in a murine cecal ligation and puncture (CLP) model. ICR/HSD mice were treated with glucan (50 mg/kg) 1 hour before or 15 minutes after CLP. Liver and lung tissue were harvested at 3 hours and mortality trends were observed for 20 days.. CLP increased liver and lung NF-kappa B and NF-IL6 nuclear binding activity as well as tumor necrosis factor-alpha and interleukin 6 messenger RNA levels at 3 hours. Pretreatment or posttreatment with glucans inhibited tissue NF-kappa B and NF-IL6 nuclear binding activity and tissue cytokine messenger RNA levels. Prophylaxis with glucan phosphate or scleroglucan increased (P < .001) long-term survival (20% CLP vs 65% glucan phosphate, 75% scleroglucan). Posttreatment with glucan phosphate also increased (P < .05) long-term survival (20% vs 65%).. Pretreatment or posttreatment with biologic response modifiers decreased tissue transcription factor nuclear binding activity and cytokine message in liver and lung of septic mice. Inhibiting early transcription factor activation and cytokine message expression correlates with improved outcome in polymicrobial sepsis as denoted by increased long-term survival.

Topics: Animals; beta-Glucans; CCAAT-Enhancer-Binding Proteins; DNA-Binding Proteins; Glucans; Interleukin-6; Male; Mice; Mice, Inbred ICR; NF-kappa B; Nuclear Proteins; RNA, Messenger; Sepsis; Tumor Necrosis Factor-alpha

Beta-1,3-D-polyglucose derivatives protect mice against otherwise lethal bacterial infections. This protective effect has been considered to be mediated through mononuclear phagocytes. By using radioactive labelling, we localized the beta-1,3-D-polyglucose derivatized microbeads (GDM) during the period following injection. The GDM was recovered mainly in the milky spots of the omentum. In animals treated with GDM, the total white cell number was significantly increased in peritoneal fluid of mice before and after challenge with E. coli. Bacterial counts in peritoneal fluid of GDM treated animals declined to zero after 24 h. In untreated animals there was a slight increase in bacterial counts until the animals died after about 12 h. Mouse peritoneal macrophages stimulated with GDM released significant amounts of IL-1 and PGE2. There was no significant release of TNF. Levels of IL-1 and PGE2 in peritoneal fluid increased significantly during the first 48 h after treatment with GDM. There was no increase of levels of TNF. After challenge with E. coli, the levels of IL-1, TNF, and PGE2 were significantly lower compared with control animals. In untreated animals the levels of IL-1 and TNF remained elevated until the animals died after about 12 h. These studies demonstrate that the raised levels of arachidonic acid metabolites after pretreatment with GDM or AG seems to inhibit the otherwise lethal elevation of IL-1 and TNF in body fluids which is seen in untreated animals.

Topics: Adjuvants, Immunologic; Animals; beta-Glucans; Cells, Cultured; Cytokines; Dinoprostone; Dose-Response Relationship, Immunologic; Escherichia coli; Female; Glucans; Interleukin-1; Leukocyte Count; Macrophages; Mice; Mice, Inbred Strains; Peritoneum; Sepsis; Tumor Necrosis Factor-alpha

The influences of pretreatment with beta-1,3-D-polyglucose derivatives on levels of cytokines and arachidonic acid metabolites in body fluids in experimental peritonitis in mice are reported. Peritonitis was induced by an intraperitoneal injection of 10(8) live Escherichia coli. Pretreated animals survived the infection, untreated animals died about 12 h after inoculation with E. coli. Levels of IL-1 in plasma and peritoneal fluid, measured by cytotoxicity assay of the HT-2 cell line, increased significantly during the first 48 h after intraperitoneal treatment with beta-1,3-D-polyglucose-derivatized microbeads (GDM) or soluble, aminated beta-1,3-D-polyglucose (AG). After subsequent challenge with E. coli, the levels of IL-1 were significantly lower than in untreated animals. There was no increase in levels of TNF after treatment with GDM or AG, measured by cytotoxicity assay of the WEHI clone 13 cell line. After challenge with E. coli, TNF in plasma and peritoneal fluid was significantly lower compared with untreated animals. Both PGE2 and LTB4, measured by radioimmunoassay kits, were increased in peritoneal fluid after treatment with GDM and AG. After challenge with E. coli, PGE2 and LTB4 in peritoneal fluid increased to about half the concentration of infected control animals. Intraperitoneal injection of indomethacin to pretreated animals resulted in increased levels of IL-1 and TNF and decreased levels of PGE2 following challenge with E. coli. The levels of IL-1 and TNF remained elevated until the animals died after about 12 h. These studies demonstrate that the raised levels of arachidonic acid metabolites after pretreatment with GDM or AG seem to inhibit the otherwise lethal elevation of IL-1 and TNF in body fluids which is seen in untreated animals.

Topics: Animals; Arachidonic Acids; Ascitic Fluid; beta-Glucans; Cytokines; Dinoprostone; Escherichia coli Infections; Female; Glucans; Indomethacin; Interleukin-1; Leukocyte Count; Leukotriene B4; Mice; Mice, Inbred CBA; Sepsis; Tumor Necrosis Factor-alpha

Beta-1,3-D-polyglucose derivatives protect mice against otherwise lethal bacterial infections. This protective effect has previously been considered to be mediated through mononuclear phagocytes. We have now investigated the cellular composition in blood and peritoneal fluid after administration of the beta-1,3-D-polyglucose before and after challenge with Escherichia coli. In animals treated with beta-1,3-D-polyglucose derivatives, the total white cell number was significantly increased in both blood and peritoneal fluid before and after challenge with E. coli. The increased total cell number was mainly the result of raised levels of granulocytes. The effects of beta-1,3-D-polyglucose-derivatized microbeads (GDM) and soluble aminated beta-1,3-D-polyglucose (AG) were similar. Bacterial counts in peripheral blood in GDM- and AG-treated animals increased with 6 h after challenge and approached zero after 24 h. In untreated animals the bacterial counts increased gradually until the animals died after about 12 h. Bacterial counts in peritoneal fluid of GDM- and AG-treated animals declined to zero after 24 h. In untreated animals there was a slight increase in bacterial counts until the animals died after about 12 h. By using radioactive labelling, we localized the bacterial as well as the beta-1,3-D-polyglucose derivatives during the period following injection. Particle-bound beta-1,3-D-polyglucose was recovered mainly in the milky spots of the omentum. A conspicuous number of bacteria were also recovered in the milky spots. The soluble aminated beta-1,3-D-polyglucose was recovered mainly in the liver. However, on a weight basis, the greatest concentration of radioactivity was in the milky spots.

Topics: Animals; Ascitic Fluid; beta-Glucans; Colony Count, Microbial; Escherichia coli Infections; Female; Glucans; Immunity, Innate; Leukocyte Count; Mice; Mice, Inbred C3H; Mice, Inbred CBA; Microspheres; Omentum; Peritoneal Diseases; Sepsis