Course Of Endotoxemia Research Articles

Recombinant ADAMTS-13 Improves Survival of Mice Subjected to Endotoxemia.

When stimulated by proinflammatory mediators, endothelial cells release ultra-large von Willebrand factor (ULVWF) multimers that are hyperactive in activating and aggregating platelets. These ULVWF multimers can accumulate in the circulation and on the inflamed endothelium because they are insufficiently cleaved by the metalloprotease ADAMTS-13, which becomes moderately deficient under conditions of systemic inflammation. This moderate ADAMTS-13 deficiency may lead to thrombotic complications that contribute to ischemic tissue injury and organ failure that are associated with severe infections. To test this hypothesis, we investigated whether recombinant ADAMTS-13 improves the pathological course of endotoxemia in lipopolysaccharide (LPS)-treated mice. C57BL/J6 mice received a bolus infusion of either 5 µg/mouse of ADAMTS-13 or vehicle control 30 min after LPS challenge and were monitored for seven-day survival. During the monitoring period, platelet counts, VWF antigen, and ADAMTS-13 activity were measured. Thrombosis was also examined by the immunohistochemistry in the liver. We found that ADAMTS-13 reduced mortality from 66% to 34.9%. The improved survival was associated with a greater recovery from thrombocytopenia, higher plasma ADAMTS-13 activity, and less thrombotic vascular occlusion. These results suggest that systemic inflammation could result in deficient ULVWF proteolysis by ADAMTS-13 and that ADAMTS-13 improves the outcomes of endotoxemia-induced inflammation.

Defining TLR4-independent molecular pathways activated during endotoxemia

Abstract Sepsis is the leading cause of nosocomial mortality, accounting for almost 20% of annual global deaths. Patients who recover from septic shock maintain a significantly increased risk for mortality due to secondary infection. Toll-like receptor 4 (TLR4) recognizes Gram-negative bacterial lipopolysaccharides (LPS) and initiates robust immune activation that can be detrimental to the host. Towards the end of septicemia, cells self-regulate and remove surface TLR4 which contributes to immune ataxia. TLR4 deficient (TLR4 −/−) C57BL6 mice are used as a model for the immune refractory period following septic shock and compared to wildtype (WT) mice. In the serum of both WT and TLR4 −/−mice, 51 proteins were previously found to be dysregulated over the course of endotoxemia. We hypothesize that these 51 proteins represent TLR4 independent activation mechanisms. This study further investigates protein and mRNA distribution during endotoxemia. Liver, spleen, kidney, heart, and brain were collected at 0, 6, 12, and 18 hours post-intraperitoneal injection of 30 mg/kg E. coli LPS. Using qPCR and Western Blot to quantify gene and protein expression, we observed down regulation of genes involved with metabolism (Adsl, Mdh1) and erythrocyte cytoskeleton stabilization (Dmtn). In the same samples we observe activation of genes and proteins involved in T-cell function (Coro1a), and the complement cascades (Masp1, C1qb, Cfd, F5). These results further define TLR4 independent activation during endotoxemia as a model for immune ataxia that occurs after primary septic shock. This project could provide proof of principle to support the development of more targeted treatments to address immune activation during secondary infection after septic shock. This project was supported by a Towson University FDRC grant to EMH and a Fisher College of Science and Mathematics undergraduate research award to JCH, HA, PC, and AA.

Scrutinizing Mechanisms of the 'Obesity Paradox in Sepsis': Obesity Is Accompanied by Diminished Formation of Neutrophil Extracellular Traps (NETs) Due to Restricted Neutrophil-Platelet Interactions.

Systemic inflammation is a detrimental condition associated with high mortality. However, obese individuals seem to have higher chances of surviving sepsis. To elucidate what immunological differences exist between obese and lean individuals we studied the course of endotoxemia in mice fed high-fat diet (HFD) and ob/ob animals. Intravital microscopy revealed that neutrophil extracellular trap (NET) formation in liver vasculature is negligible in obese mice in sharp contrast to their lean counterparts (ND). Unlike in lean individuals, neutrophil influx is not driven by leptin or interleukin 33 (IL-33), nor occurs via a chemokine receptor CXCR2. In obese mice less platelets interact with neutrophils forming less aggregates. Platelets transfer from ND to HFD mice partially restores NET formation, and even further so upon P-selectin blockage on them. The study reveals that in obesity the overexaggerated inflammation and NET formation are limited during sepsis due to dysfunctional platelets suggesting their targeting as a therapeutic tool in systemic inflammation.

Circulatory effects of dexmedetomidine in early sepsis: a randomised controlled experimental study.

We designed this experimental study with a view to evaluate the effects of dexmedetomidine (DEX) on cardiac performance and systemic and peripheral hemodynamics in healthy and early-stage endotoxemia swine models. Our study hypothesis was that DEX can ensure hemodynamic stability during the course of endotoxemia. Thirty-two male pigs (25-27kg) were assigned into four groups: (1) no intervention (group A), (2) DEX 0.8μg/kg was administered in non-septic animals (group B), (3) sepsis induced by intravenous Escherichia coli endotoxin (group C) and (4) DEX 0.8μg/kg was administered in septic animals (group D). Hemodynamic parameters such as heart rate, mean blood pressure, central venous pressure, pulmonary artery pressures, pulmonary artery occlusion pressure, pulmonary vascular resistance and cardiac output were continuously recorded. Central venous oxygen saturation was also measured in order to obtain a complete evaluation of cardiovascular response to sepsis. Heart rate was decreased, whilst mean arterial pressure decrease was alleviated after DEX administration in septic animals. In addition, central venous pressure was stable in animals with sepsis after DEX infusion. Sepsis dramatically elevated pulmonary function indicators but DEX succeeded in ameliorating this effect. The important decrease measured in central venous oxygen saturation in both sepsis groups reflected the decreased perfusion of tissues that takes place at the end of early sepsis. Our findings support the hypothesis that DEX has beneficial effects on heart rate and pulmonary artery pressure, whilst reduction in systemic blood pressure occurs at acceptable levels.

Preventive Leptin Administration Protects Against Sepsis Through Improving Hypotension, Tachycardia, Oxidative Stress Burst, Multiple Organ Dysfunction, and Increasing Survival.

Sepsis syndrome is the most important cause of mortality in critically ill patients admitted to intensive care units (ICUs). However, current therapies for its prevention and treatment are still unsatisfactory, and the mortality rate is still high. Non-septic ICU patients are vulnerable to acquire sepsis syndrome. Thus, a preventive treatment for this population is needed. During sepsis syndrome and endotoxemia, severe hypotension, tachycardia, oxidative and immune response increase, multiple organ dysfunction syndrome (MODS) and decreased survival are observed. Leptin administration protects against negative effects of sepsis syndrome and endotoxemia. Furthermore, it is has been reported that leptin elevates blood pressure mediated by sympathetic nervous system activation. However, whether leptin administration before sepsis induction mediates its protective effects during sepsis through blood pressure regulation is not known. Therefore, we investigated whether pre-treatment of leptin improves blood pressure and MODS, resulting in survival increase during endotoxemia. The results showed that leptin administration before endotoxemia induction reduced both the hypotension and tachycardia characteristically observed during endotoxemia. Notably, this protective effect was observed early and late in the course of endotoxemia. Endotoxemia-induced MODS decreased in leptin-treated rats, which was reflected in normal values for liver and kidney function, inhibition of muscle mass wasting and maintenance of glycemia. Furthermore, leptin pre-treatment decreased the oxidative stress burst in blood and blunted the increased pro-inflammatory cytokines TNF-α, IL-1β, and IL-6 observed during endotoxemia. Remarkably, according to the leptin-induced increase in survival, leptin pre-administration decreased the risk for death associated with sepsis syndrome at early and late times after endotoxemia induction. These results show a potential preventive therapy against sepsis syndrome and endotoxemia in vulnerable patients, based in the beneficial actions of leptin.

Combined Enteral and Parenteral Glutamine Supplementation in Endotoxemic Swine: Effects on Portal and Systemic Circulation Levels

Objective: To measure plasma glutamine (GLN) levels in systemic and portal circulation after combined enteral and parenteral administration in early endotoxemic swine. We hypothesized that this combination will be more efficient than intravenous administration alone in restoring plasma levels during the course of endotoxemia. Materials and Methods: Endotoxemia was induced with Escherichia coli O111:B4 lipopolysaccharide (LPS) (250 μg/kg body weight) in 16 anes­thetized, fasted swine and maintained by constant infusion (2 μg/kg/h) over 180 min. Another 16 swine served as controls. After infusion with LPS or placebo, GLN was administered intravenously, enterally or in combination (0.5 g/kg i.v. plus 0.5 g/kg enterally) over 30 min. At 0, 15, 30, 45, 60, 120 and 180 min, blood was drawn from the systemic and portal circulation for colorimetric assessment of GLN. Results: In healthy, placebo-alone swine, GLN levels remained stable throughout the study. Intravenous and combined infusion increased systemic levels (p = 0.001), but after enteral administration alone, a smaller effect was observed (p = 0.026). Portal levels were increased after combined, enteral and intravenous administration (p = 0.001). In endotoxemia, systemic and portal levels decreased significantly. Intravenous and, to a greater extent, combined administration increased systemic levels (p = 0.001), while enteral administration only had a small effect (p = 0.001). In the portal vein, intravenous and combined treatment increased plasma levels (p = 0.001), whereas enteral supplementation alone had again a small, yet significant effect (p = 0.001). Conclusions: The findings indicate that combined GLN supplementation is superior to intravenous treatment alone, in terms of enhanced availability in systemic and portal circulations. Thus, combined treatment at the onset of endotoxemia is a beneficial practice, ensuring adequate GLN to compensate for the resulting intracellular shortage.

Effects of choline treatment in concentrations of serum matrix metalloproteinases (MMPs), MMP tissue inhibitors (TIMPs) and immunoglobulins in an experimental model of canine sepsis

The aim of the present study was to investigate effects of intravenous (i.v.) choline treatment on serum matrix metalloproteinases (MMP), MMP tissue inhibitors (TIMP) and immunoglobulins (Igs), and to determine if there were relations between serum MMPs/TIMPs and C-reactive protein (CRP) (as a marker of the acute phase response), immunoglobulin G and M (IgG and IgM) (as a maker of the Ig responses) and markers of organ damage such as muscular damage (creatine phosphokinase, [CPK]), liver damage (alanine aminotransferase [ALT]) and renal dysfunction (blood urea nitrogen [BUN] and creatinine, [Cr]) in dogs with endotoxemia. Healthy dogs (n=24) were randomized to Saline, Choline (C), Lipopolysaccharide (LPS), and LPS+C groups and received 0.9% NaCl (5mL/i.v.), choline chloride (20mg/kg/i.v.), LPS (0.02mg/kg/i.v.) and LPS (0.02mg/kg/i.v.) plus choline chloride (20mg/kg/i.v.), respectively. Serum MMPs and TIMPs concentrations were analyzed by commercial ELISA kits. MMP and TIMP increased at 1–48h (P<0.05), whereas IgG and IgM decreased at 24–48h in LPS group, compared to their baselines. Choline treatment reduced changes in serum MMPs, TIMPs and markers of organ damage, and prevented the hypoimmunoglobulinemia in LPS+C. MMPs and TIMPs were correlated positively (P<0.05) with serum CRP, CPK, ALT, BUN and Cr, but not with serum Igs. Our findings suggest that the serum MMPs, TIMPs and Igs are involved in the pathophysiology of endotoxemia, and MMPs and TIMPs are correlated with the acute phase reaction and multi-organ failure. In addition, we demonstrated a direct effect of choline administration in decreasing serum MMPs and TIMPs, and preserving serum Igs in the course of endotoxemia.

Features of course of endotoxemia and "metabolic" intoxication in athletes depending on the degree of exercise stress

Modern sports more often associated with the occurrence of immunodeficiency - a reduction in the functional activity of the main components of the immune system. This leads to disturbances in the system of antibiotic resistance of the organism and the growth of infectious diseases of athletes. Therefore, the aim of the study was to explore the features of endotoxemia and metabolic intoxication in athletes depending on the exercise stress. 43 athletes were examined during long-term training. Results. On the basis of the study we found that endotoxemia course of light and medium degree to proportional stress levels is determined in the first and second group. The endotoxemia course is associated with a decrease in the toxin-associated ability of serum albumin, which is a measure of cytolytic activity of albumin fractions and accumulation of toxins of medium size (10-200 nm) of microbial origin and hydrophobic nature in plasma of peripheral blood. Calculation methods for determining the severity of endotoxemia can be used to identify different mechanisms of its formation and give the opportunity to some extent, reflect the complex toxicometry parameters of athletes to determine the levels of adaptive reactions in response to exercise stress

Hypothermia attenuates NO production in anesthetized rats with endotoxemia.

Sepsis is often associated with upregulation of nitric oxide production and fever, and it is common to control an excessive febrile response with antipyretic therapy and external cooling. Our aim was to evaluate the effect of hypothermia on NO production in a model of septic shock. Rats were anesthetized, ventilated, and instrumented for hemodynamic monitoring and divided into four groups. Normothermic controls (NC) received saline intravenously and were maintained at 37 °C. Hypothermic controls (HC) received saline but were allowed to become hypothermic. Normothermic endotoxic (NE) received Escherichia Coli lipopolysaccharides (LPS) intravenously to induce endotoxic shock and was maintained at 37 °C. Hypothermic endotoxic (HE) received LPS intravenously and was allowed to become hypothermic. Exhaled NO (NOe) was measured from mixed expired gas at time zero and every 30 min, for 5 h. After injection of LPS, NOe increased substantially in the NE group (700 ± 24 ppb), but increased only to 25 ± 2 ppb in the HE group. NOe increased to 90 ± 3 ppb in the NC group, and to 17.6 ± 3.1 ppb in the HC group after 5 h (P < 0.05), whilst blood pressure remained stable. In the HE group, blood pressure fell immediately after injection of the LPS, but thereafter remained stable despite the rise in NOe. In the NE group, the blood pressure fell gradually, and the animals became hypotensive. During the natural course of endotoxemia in anesthetized rats, allowing severe hypothermia to ensue by not actively managing temperature and hemodynamics resulted in significantly reduced expired NO concentrations, lung injury, and prolonged survival. The clinical benefits of such a finding currently remain unclear and merit further investigation.

Regional Sympathetic Blockade Attenuates Activation of Intestinal Macrophages and Reduces Gut Barrier Failure

Endotoxin-induced activation of monocytes may lead to extravasation of cells, excessive production of nitric oxide, and subsequent epithelial injury in the gut. Regional sympathetic blockade by means of thoracic epidural anesthesia has been implicated to protect the epithelial barrier. This study tested the hypothesis that thoracic epidural anesthesia decreases epithelial permeability by attenuating monocytic production of nitric oxide and nitrosative stress. Rats were anesthetized, hemodynamically monitored, and mechanically ventilated. Endotoxemia was induced by an intravenous bolus injection of Escherichia coli lipopolysaccharide. Either lidocaine 2% or normal saline was injected as a bolus, followed by a continuous infusion via an epidural catheter. Three hundred minutes after injection of lipopolysaccharide or normal saline, gut epithelial permeability to fluorescein isothiocyanate-dextran (4 kDa), intestinal expression of inducible nitric oxide synthase by macrophages, and lipid peroxidation represented by 8-isoprostane tissue concentration were quantified. Thoracic epidural anesthesia significantly attenuated the endotoxin-induced increases in gut epithelial permeability (437 [293, 492] vs. 628 [532, 1,042] ng/ml, median [quartiles], P = 0.03), expression of nitric oxide synthase (2 [1,2] vs. 7 [5,8] cells per 384 µm(2), P = 0.003), macrophage infiltration, and lipid peroxidation (22,460 ± 11,476 vs. 37,840 ± 17,551 pg/ml, mean ± SD, P = 0.05). Thoracic epidural anesthesia attenuates endotoxin-induced gut epithelial injury. This is likely due to a decrease in monocytic extravasation and intestinal nitrosative stress. As possible mechanisms, direct nerve-immune interplay, a reduction in plasma catecholamines, or a systemic lidocaine effect has to be considered.

Thoracic Epidural Anesthesia Attenuates Endotoxin-induced Impairment of Gastrointestinal Organ Perfusion

Systemic inflammation can be associated with a redistribution of organ blood flow and a decrease in gastrointestinal perfusion. Regional sympathetic blockade by means of thoracic epidural anesthesia (TEA) has been shown to improve intestinal microcirculation during systemic inflammation. This study tests the hypothesis that during systemic inflammation, TEA attenuates the impairment of gastrointestinal organ perfusion without compromising blood flow to vital organs. Eighteen rats were anesthetized, hemodynamically monitored, and mechanically ventilated with room air. By using fluorescent microspheres, organ perfusion was quantified at baseline, 30 min after the start of epidural infusion of either 2% lidocaine (TEA) or normal saline (control), and after 60 and 120 min of intravenous Escherichia coli lipopolysaccharide infusion in TEA and control animals. Blood pressure initially was lower in TEA animals, but it was comparable to controls during endotoxemia. Gastrointestinal organ perfusion significantly decreased after 120 min of endotoxemia in the controls but not in the TEA animals (-23 +/- 27% vs. -6 +/- 26%, mean +/- SD, P < 0.05). Perfusion of the vital organs such as the heart, brain, liver, and kidneys was comparable between controls and TEA after 120 min of endotoxemia. TEA attenuates the impairment of gastrointestinal organ perfusion during endotoxemia. Hence, the protective effects of TEA on intestinal microcirculation during endotoxemia may be due to a higher total organ blood flow compared with endotoxemic control animals. Furthermore, in the course of endotoxemia, TEA provides hemodynamic stability and does not compromise blood flow to vital organs.

Pleiotropic functions of TNF-α determine distinct IKKβ-dependent hepatocellular fates in response to LPS

TNF-alpha influences morbidity and mortality during the course of endotoxemia. However, the complex pleiotropic functions of TNF-alpha remain poorly understood. We evaluated how hepatic induction of NF-kappaB and TNF-alpha influence survival and hepatocellular death in a lethal murine model of endotoxic shock. Using dominant-negative viral vectors to inhibit the IKK complex, we demonstrate through this study that the liver is a major source of TNF-alpha during the course of lethal endotoxemia and that IKKbeta (but not IKKalpha) is predominantly responsible for activating NF-kappaB and TNF-alpha in the liver after LPS administration. Using TNF-alpha knockout mice and hepatic-specific inhibition of IKKbeta, we demonstrate that the status of TNF-alpha and NF-kappaB balances necrotic and apoptotic fates of hepatocytes in the setting of endotoxemia. In the presence of TNF-alpha, inhibiting hepatic IKKbeta resulted in increased survival, reduced serum proinflammatory cytokines, and reduced hepatocyte necrosis in response to a lethal dose of endotoxin. In contrast, inhibiting hepatic IKKbeta in TNF-alpha knockout mice resulted in decreased survival and increased caspase 3-mediated hepatocyte apoptosis after endotoxin challenge, despite a reduced proinflammatory cytokine response. In the presence of TNF-alpha, NF-kappaB-dependent hepatocellular necrosis predominated, while in the absence of TNF-alpha, NF-kappaB primarily influenced apoptotic fate of hepatocytes. Changes in JNK phosphorylation after LPS challenge were also dynamically affected by both IKKbeta and TNF-alpha; however, this pathway could not solely explain the differential outcomes in hepatocellular fates. In conclusion, our studies demonstrate that induction of NF-kappaB and TNF-alpha balances protective (antiapoptotic) and detrimental (proinflammatory) pathways to determine hepatocellular fates during endotoxemia.

Nitric oxide modifies the sarcoplasmic reticular calcium release channel in endotoxemia by both guanosine-3′,5′ (cyclic) phosphate-dependent and independent pathways*

a) To determine whether decreased sarcoplasmic calcium release channel (CRC) activity is a mechanism by which myocardial contractility is reduced in endotoxemia; b) to determine whether nitric oxide modulates CRC activity in endotoxemia; and c) to examine two nitric oxide signaling pathways in relation to CRC function in endotoxemia. Randomized, prospective using a rat model of endotoxemia. : Research laboratory. Sprague-Dawley rats. Endotoxemia was induced by lipopolysaccharide administration. The effects of nitric oxide were studied using the highly selective inducible nitric oxide synthase inhibitor N-(3-(aminomethyl)benzyl)acetamidine dihydrochloride (1400W) and the specific guanylyl cyclase inhibitor 1-H (1, 2, 4)oxadiazolo[4,3-a]quinoxalin-1-one (ODQ). We assessed myocardial contractility, myocardial nitric oxide content, and guanosine-3',5' (cyclic) phosphate (cGMP) content. We determined CRC activity by calcium release and ryanodine binding assays. We followed these variables at four time points through the course of endotoxemia. We found that myocardial contractility and CRC activity were decreased in late but not in early endotoxemia. Furthermore, inducible nitric oxide synthase inhibition with 1400W restored contractility and CRC activity in late endotoxemia but paradoxically worsened these variables in early endotoxemia. Through the use of the guanylyl cyclase inhibitor ODQ, we demonstrate that nitric oxide acts through cGMP-mediated mechanisms in early and late endotoxemia. We investigated cGMP-independent pathways by assessing the oxidative status of the CRC. We found that in late endotoxemia, nitric oxide decreased the number of free thiols, demonstrating that nitric oxide also acts through cGMP-independent pathways. Nitric oxide has a dual effect on the CRC in endotoxemia. At low concentrations, as measured in early endotoxemia, nitric oxide stabilizes the CRC through cGMP-mediated mechanisms. In late endotoxemia, high nitric oxide concentrations decrease channel activity through both cGMP-dependent and cGMP-independent mechanisms.

Treatment of Endotoxic Shock: Glucocorticoids, Lazaroids, Nonsteroidals, Others

Review of the literature would indicate that a therapy inhibiting a single step in the inflammatory mediator cascade, although potentially beneficial, cannot serve as the "magic bullet" in endotoxic shock. Thus, combination therapy using drugs that target multiple steps in the mediator cascade may prove useful in the future. Until controlled clinical studies have been performed in veterinary medicine using the agents discussed in this manuscript (or others), many questions will remain regarding the best possible pharmacologic therapies to use in supplementing traditional supportive care (e.g., fluids, broad-spectrum antimicrobials) measures. Many of the therapies discussed in this manuscript hold tremendous potential, but must be administered before endotoxic shock occurs or early in the course of endotoxic shock to elicit a beneficial response. Unfortunately this is not usually possible in the clinical situation. Further, the possibility of eliciting a detrimental response by use of drugs at inappropriate dosages or too late in the course of the disease must be considered. In time, therapies may be grouped into those that hold promise for being beneficial in the patient at risk for endotoxic shock and those that are beneficial for treatment of established endotoxic shock. Therapies that may hold promise for the patient with established endotoxic shock possibly include the lazaroids, certain NSAIDs, pentoxifylline, and selective inhibitors of iNOS activity. Although still controversial, methylprednisolone and other glucocorticoids are probably best reserved for the patient at risk for endotoxic shock, or those identified very early in the course of endotoxemia and endotoxic shock, and should never be used in conjunction with NSAIDs. Since no one therapy is likely to provide the "magic bullet" for the patient in endotoxic shock, treatment may ultimately involve combination therapy using some of these drugs that target different pathways and mediators and may also include antibodies against endotoxin or its mediators.

Limulus antilipopolysaccharide factor prevents mortality late in the course of endotoxemia.

Limulus antilipopolysaccharide factor (LALF) can neutralize bacterial endotoxin, but its ability to prevent mortality following prolonged endotoxemia is unknown. Mice were challenged with an LD50 dose of intraperitoneal E. coli lipopolysaccharide (LPS) and then received LALF at various times after administration of LPS. Survival at 72 h was significantly improved by the administration of LALF at 4, 10, and even 24 h after LPS (73%, 78%, and 65% survival, respectively, vs. 15% survival in controls). Following intravenous administration of LALF at either 10 or 24 h after LPS, plasma levels of biologically active LPS abruptly fell (> 1000-fold lower than pre-LALF levels). Plasma LALF concentrations fell much more gradually in LPS-treated mice (t1/2 = 120 min) than in control mice (t1/2 = 2.5 min). In conclusion, LALF markedly decreased plasma concentrations of biologically active LPS and protected mice from lethality even when LALF was not administered until long after the onset of continuous endotoxemia.

Exhaled nitric oxide as a marker for serum nitric oxide concentration in acute endotoxemia

Purpose: The main aim of this study was to assess the correlation between exhaled nitric oxide (NO) and serum NO concentrations during the course of endotoxemia. We also assessed whether or not the inducible isoform of NO synthase is responsible for the increase in NO production in endotoxemia animals. Materials and Methods: Anesthetized and mechanically ventilated dogs were injected with either saline (control) or Escherichia coli endotoxin (LPS [Lipopolysaccharides]), and the animals were sacrificed 150 minutes later. We measured hemodynamics, exhaled NO, and serum arterial and mixed venous NO concentrations. Western blotting was performed on lung, pulmonary artery, aorta, and kidney tissue samples using anti-inducible NO synthase antibody. Results: Arterial pressure, cardiac output, and pulmonary arterial pressure in the control group remained unchanged, whereas a significant decline in these parameters was observed in the LPS group. Exhaled NO and serum arterial NO concentrations rose significantly within 30 minutes of endotoxin injection and remained higher than baseline values, whereas mixed venous serum NO did not change from baseline values. There was a significant linear relationship between exhaled NO and arterial serum NO concentrations. By comparison, exhaled NO, and arterial and mixed venous serum NO levels remained unchanged in the control group. Western blotting showed no expression of inducible NO synthase (iNOS) isoform in the control or LPS groups. Conclusions: These results suggest that exhaled NO accurately reflects changes in arterial serum NO concentration and that the source of enhanced NO release in acute endotoxemia is not the iNOS isoform.

INTRAOPERATIVE ESTIMATION OF ENDOTOXIN, TNFα, AND IL-6 IN ORTHOTOPIC LIVER TRANSPLANTATION AND THEIR RELATION TO REJECTION AND POSTOPERATIVE INFECTION

The course of endotoxemia, TNF alpha, and IL-6 during orthotopic liver transplantation was studied in 28 transplantations performed in 27 patients to evaluate their impact on early postoperative rejection and infection. The preoperative levels of endotoxin, TNF alpha, and IL-6 were not different in patients who did or did not develop postoperative rejection and/or infection within the first 10 postoperative days. At the end of surgery, TNF alpha levels increased in patients who developed rejection (median 100 pg/ml vs. 11.5 pg/ml, P = 0.004). A TNF alpha level of greater than 100 pg/ml at the end of transplantation predicted rejection in 82% of the patients. During surgery, IL-6 levels increased significantly in patients with subsequent postoperative infection, reaching significance after revascularization of the graft (median 975 pg/ml vs. 185 pg/ml, P = 0.006). An IL-6 cutoff level of 800 pg/ml predicted postoperative infection in 75% of the patients. Endotoxins were elevated intraoperatively in patients with postoperative infection, but the difference did not reach significance. There was no prognostic relevance with respect to the intraoperative values of TNF alpha and infection or IL-6 values and rejection. An intraoperative elevation of TNF alpha seems to precede early postoperative rejection, and highly increased IL-6 may be a predictor of subsequent infection in human liver transplantation.

Radiation induced gram negative bacteremia and endotoxemia in rabbits: Modification by anti-lipopolysaccharide hyperimmune equine plasma

Lethal whole body irradiation damages the gut mucosa and leads to leakage of endotoxin or lipopolysaccharides (LPS) into the systemic circulation. Sixteen rabbits, irradiated with 900 rads 60Co, were randomly divided on day 4 into 2 groups, one of which received an intraperitoneal injection of normal saline (control) and the other (experimental) an equal volume of anti-LPS hyperimmune plasma. The time course of endotoxemia and bacteremia were determined for the duration of the experiment. While rabbits in both groups died within 13 days after irradiation, rabbits given saline died on average 2 days earlier, than rabbits given anti-LPS plasma. Plasma LPS concentrations rose to a small peak on day 2 prior to treatment. Thereafter plasma LPS in rabbits given saline increased forty fold by day 9. In contrast, in rabbits given anti-LPS plasma, LPS concentrations in the plasma remained within pretreatment limits (p 0.01). By day 12 after irradiation, plasma anti-LPS IgG had declined to 5.8% of pretreatment levels in rabbits given saline as compared to 46% in rabbits given anti-LPS plasma (p 0.005). Whilst both groups developed gram-positive bacteremia, rabbits given saline in addition also developed gram-negative bacteremia. No rabbits treated with Anti-LPS showed gram-negative bacteremia. Treatment with Anti-LPS plasma thus significantly protects radiated rabbits from the incidence of gram-negative bacteremia, development of high plasma LPS levels and hence endotoxemia, and prolongs survival to a certain extent.