Antibodies to cachectin/tumor necrosis factor reduce interleukin 1 beta and interleukin 6 appearance during lethal bacteremia

Cytokines secreted in response to invading micro-organisms are important mediators of detrimental hemodynamic and metabolic changes in the host. To test whether cachectin/TNF plays a role in triggering release of other cytokines in the setting of infection, anesthetized baboons were passively immunized against systemic cachectin/TNF before infusion of a LD100 dose of live Escherichia coli. Bacteremia led to significant increases in circulating levels of cachectin/TNF, IL-1 beta, and IL-6. Although bacterial endotoxin/lipopolysaccharide is a potent stimulus for the synthesis and release of IL-1 and IL-6 in vitro, specific neutralization of cachectin/TNF in vivo with mAb pretreatment significantly attenuated both the IL-1 beta and the IL-6 responses despite fulminant overwhelming bacteremia. These data suggest that cachectin/TNF is essential for the initiation or amplification of IL-1 and IL-6 release during lethal gram-negative septic shock syndrome.     

Role of endotoxemia in cardiovascular dysfunction and mortality. Escherichia coli and Staphylococcus aureus challenges in a canine model of human septic shock.

Using different types of bacteria and a canine model simulating human septic shock, we investigated the role of endotoxin in cardiovascular dysfunction and mortality. Either Escherichia coli (a microorganism with endotoxin) or Staphylococcus aureus (a microorganism without endotoxin) were placed in an intraperitoneal clot in doses of viable or formalin-killed bacteria. Cardiovascular function of conscious animals was studied using simultaneous radionuclide heart scans and thermodilution cardiac outputs. Serial plasma endotoxin levels were measured. S. aureus produced a pattern of reversible cardiovascular dysfunction over 7-10 d that was concordant (P less than 0.01) with that of E. coli. Although this cardiovascular pattern was not altered by formalin killing (S. aureus and E. coli), formalin-killed organisms produced a lower mortality and less myocardial depression (P less than 0.01). S. aureus, compared to E. coli, produced higher postmortem concentrations of microorganisms and higher mortality (P less than 0.025). E. coli produced significant endotoxemia (P less than 0.01), though viable organisms (versus nonviable) resulted in higher endotoxin blood concentrations (P less than 0.05). Significant endotoxemia did not occur with S. aureus. Thus, in the absence of endotoxemia, S. aureus induced the same cardiovascular abnormalities of septic shock as E. coli. These findings indicate that structurally and functionally distinct microorganisms, with or without endotoxin, can activate a common pathway resulting in similar cardiovascular injury and mortality.     

Staphylococcus epidermidis induces complement activation, tumor necrosis factor and interleukin-1, a shock-like state and tissue injury in rabbits without endotoxemia. Comparison to Escherichia coli.

Tumor necrosis factor (TNF) and IL-1 are thought to mediate many of the pathophysiologic changes of endotoxemia and Gram-negative bacteremia. In these studies, heat-killed Staphylococcus epidermidis were infused into rabbits to determine whether an endotoxin (LPS)-free microorganism also elicits cytokinemia and the physiologic abnormalities seen in Gram-negative bacteremia. S. epidermidis induced complement activation, circulating TNF and IL-1, and hypotension to the same degree as did one-twentieth the number of heat-killed Escherichia coli. Circulating IL-1 beta levels had a greater correlation coefficient (r = 0.81, P less than 0.001) with the degree of hypotension than TNF levels (r = 0.48, P less than 0.02). Leukopenia, thrombocytopenia, diffuse pulmonary capillary aggregation of neutrophils, and hepatic necrosis with neutrophil infiltration were observed to the same extent after either S. epidermidis or E. coli infusion. However, S. epidermidis infusion did not induce significant (less than 60 pg/ml) endotoxemia, whereas E. coli infusion resulted in high (11,000 pg/ml) serum endotoxin levels. S. epidermidis, E. coli, LPS, or S. epidermidis-derived lipoteichoic acid (LTA) induced TNF and IL-1 from blood mononuclear cells in vitro. E. coli organisms and LPS were at least 100-fold more potent than S. epidermidis or LTA. Thus, a shock-like state with similar levels of complement activation as well as circulating levels of IL-1 and TNF were observed following either S. epidermidis or E. coli. These data provide further evidence that host factors such as IL-1 and TNF are common mediators of the septic shock syndrome regardless of the organism.     

Short-term effects of glipizide (an adenosine triphosphate-sensitive potassium channel inhibitor) on cardiopulmonary hemodynamics and global oxygen transport in healthy and endotoxemic sheep

In severe sepsis and septic shock, hemodynamic support is often complicated by a tachyphylaxis against exogenous catecholamines. Because activation of adenosine triphosphate (ATP)-sensitive potassium (K(ATP)) channels plays a pivotal role in the pathogenesis of hyperdynamic vasodilatory shock, we hypothesized that it may be beneficial to administer a specific K(ATP) channel inhibitor to prevent, or at least attenuate, hemodynamic dysfunction in sepsis. The present study was designed as a prospective and controlled laboratory experiment to elucidate the short-term effects of glipizide, a specific K(ATP) channel inhibitor, on cardiopulmonary hemodynamics and global oxygen transport in healthy sheep and sheep with endotoxemia. Ten adult ewes were anesthetized and operatively instrumented with a pulmonary artery, a femoral artery, and a foley catheter. After 24 h of recovery, healthy sheep received glipizide as a bolus infusion (4 mg/kg over 15 min). After 24 h of recovery, a continuous infusion of endotoxin (Salmonella typhosa, 10 ng.kg.(-1)min) was started in the same sheep and administered for the next 17 h. After 16 h of endotoxemia, glipizide was given as described above. Administration of glipizide was followed by a transient, but significant, increase in mean arterial pressure in both healthy controls (95 +/- 3 mmHg vs. 101 +/- 2 mmHg, P < 0.05) and sheep with endotoxemia (86 +/- 3 mmHg vs. 93 +/- 3 mmHg, P < 0.05). However, the increase in mean arterial pressure was longer lasting in ewes with endotoxemia. Cardiac index, oxygen delivery index, arterial lactate concentrations, and arterial pH were not significantly affected by glipizide. Therefore, administration of glipizide may represent a beneficial therapeutic option to treat arterial hypotension resulting from sepsis and systemic inflammatory response syndrome. Additional studies are required to determine the effects of continuous infusion of glipizide in the presence of systemic inflammation.     

Heat stress and/or endotoxin effects on cytokine expression by human whole blood

Immune system cytokines induce vascular shock. Tumor necrosis factor-alpha (TNF-alpha), interleukin 1beta (IL-1beta), and bacterial endotoxin (E) circulate in human heatstroke to suggest that E release from a heat-damaged gut may stimulate cytokines that contribute to hypovolemia. However, immune activation by heat-induced tissue necrosis might stimulate cytokine generation in the absence of E. To evaluate this potential and heat stress effects on the anti-inflammatory cytokines, IL-1 receptor antagonist (IL-1ra) and IL-1 soluble receptor II (IL-1srII), a human whole blood (HWB) model was employed in which the presence or absence of E could be controlled. Using thermoelectric technology to regulate the HWB heat exposures, the temperature modulations of lethal heatstroke were precisely replicated (maximum temperature = 42.4 degrees C +/- 0.04 degrees C; thermal area = 52.3 degrees C +/- 1.5 degrees C per min). Cytokine and mRNA measurements employed enzyme-linked immunosorbant-based assay systems. Significant elevations in TNF-alpha, IL-1beta, interleukin 6 (IL-6), and IL-1ra resulted when HWB was exposed to E concentrations (10 ng/ml) reported to circulate in heatstroke. While E-stimulated IL-1ra was significantly decreased by the presence of prior heat stress (PPHS), E-stimulated IL-1beta, TNF-alpha, and IL-6 were not significantly altered by PPHS, but tended to be elevated. IL-1srII expression was unchanged by PPHS and/or E. PPHS in the absence of E did not induce cytokine responses, nor were there elevations in TNF-alpha or IL-1beta mRNA. Thus, some factor normally absent under in vitro conditions, like endotoxin, was required to provoke HWB cytokine expressions and the heat stress and E conditions that characterize heatstroke affected HWB cytokine metabolism to favor a proinflammatory environment.     

Differential induction of pro- and anti-inflammatory cytokines in whole blood by bacteria: effects of antibiotic treatment.

The in vitro production of interleukin-1beta (IL-1beta), IL-6, and the IL-1 receptor antagonist (IL-1ra) in whole blood upon stimulation with different bacterial strains was measured to study the possible relationship between disease severity and the cytokine-inducing capacities of these strains. Escherichia coli, Neisseria meningitidis, Neisseria gonorrhoeae, Bacteroides fragilis, Capnocytophaga canimorsus, Staphylococcus aureus, Enterococcus faecalis, Streptococcus pneumoniae, and Streptococcus pyogenes induced the cytokines IL-1beta, IL-6, and IL-1ra. Gram-negative bacteria induced significantly higher levels of proinflammatory cytokine production than gram-positive bacteria. These differences were less pronounced for the anti-inflammatory cytokine IL-1ra. In addition, blood was stimulated with E. coli killed by different antibiotics to study the effect of the antibiotics on the cytokine-inducing capacity of the bacterial culture. E. coli treated with cefuroxime and gentamicin induced higher levels of IL-1beta and IL-6 production but levels of IL-1ra production similar to that of heat-killed E. coli. In contrast, ciprofloxacin- and imipenem-cilastatin-mediated killing showed a decreased or similar level of induction of cytokine production as compared to that by heat-killed E. coli; polymyxin B decreased the level of production of the cytokines.     

The thrombin antagonist hirudin fails to inhibit endotoxin-induced leukocyte/endothelial cell interaction and microvascular perfusion failure

Besides its central role in coagulatory pathways, thrombin is known to be a key mediator of macrophage and granulocyte activation in vitro. During recent years the concept of thrombin inhibition by the specific thrombin inhibitor, hirudin, has been established to treat septic disorders. Since basic mechanisms of sepsis include leukocyte/endothelial cell interaction and deterioration of capillary perfusion, we hypothesized that hirudin modulates leukocyte activation and microvascular injury. Severe endotoxemia was induced in Syrian hamsters by intravenous administration of endotoxin (lipopolysaccharide [LPS], E. coli, 2mg/kg) at 0 h. Hirudin (0.25 mg/kg/h) was substituted intravenously during the 4 h after the induction of endotoxemia (n = 7, hirudin). In control animals (n = 6, control) LPS was given without hirudin substitution. In skinfold chamber preparations leukocyte/endothelial cell interaction and functional capillary density (FCD, measure of capillary perfusion) were analyzed during a 24-h period after LPS injection using intravital fluorescence microscopy. Hirudin effectively normalized thromboplastin time and antithrombin activity when compared to controls (P < 0.05, ANOVA). However, hirudin did not attenuate LPS-induced arteriolar and venular leukocyte adherence, and even tended to increase leukocyte adherence after 24 h (P > 0.05, MANOVA). In parallel, addition of hirudin led to a significant deterioration of FCD over time when compared to controls (hirudin: baseline = 171 +/- 19 cm(-1) versus 16 +/- 9 at 24 h; control: baseline = 150 +/- 20 cm(-1) versus 62 +/- 18 at 24 h; P < 0.05). The fall in FCD in hirudin animals was associated with a significant increase of wet-to-dry weight ratios in lung, kidney, muscle, and small intestine (P < 0.05 versus control, ANOVA). Thus our study does not indicate a protective effect of hirudin on microcirculation during endotoxemia, despite an improvement of coagulatory parameters. This result may at least in part explain the lack of efficacy of hirudin on lethality during endotoxemia and sepsis.     

Inflammation-induced hepatotoxicity in humans

Because severe sepsis is frequently complicated by multiple organ failure, it is of importance to monitor organ function. Unfortunately, conventional liver function markers are either relatively unspecific or have a long half-life, which make them poor predictors of acute liver injury. Glutathione S-transferase A1-1 (GSTA1-1) has a relatively short half-life (1 h), is more specific, and is rapidly released into the blood after liver damage. In the present study, we measured plasma GSTA1-1 levels by enzyme-linked immunosorbent assay in seven healthy volunteers after repeated experimental endotoxemia induced by 2 ng kg Escherichia coli endotoxin per day (to investigate inflammation-induced hepatic injury) and in 21 patients within 12 h after the occurrence of severe sepsis/septic shock (to investigate its ability to predict an increase of transaminases on day 7). During repeated experimental endotoxemia in healthy volunteers, TNF-alpha and IL-6 levels increased from undetectable levels to 1,425 (474-1,949) and 1,739 (989-2,047) pg mL, respectively, whereas GSTA1-1 levels did not exceed the normal range, indicating that no (sub)clinical liver injury occurs in this model of inflammation. In septic patients, GSTA1-1 levels had a specificity of 88%, resulting in a positive predictive value for liver injury of 86% and a positive likelihood ratio of 6 to indicate an increase in transaminases on day 7. Furthermore, GSTA1-1 levels did not correlate with IL-6 levels but did with dobutamine infusion rate (Spearman r = 0.94; P = 0.02), suggesting that the extent of hemodynamic instability and not the degree of inflammation could be of importance for the occurrence of liver damage. In septic shock patients, GSTA1-1 may represent a useful marker for early liver injury.     

Interleukin-1 receptor antagonist ameliorates experimental anti-glomerular basement membrane antibody-associated glomerulonephritis.

The contribution of IL-1 to leukocyte infiltration in anti-glomerular basement membrane (GBM) antibody (Ab) glomerulonephritis (GN) was examined by the administration of a specific IL-1 receptor antagonist (IL-1ra). Lewis rats received anti-GBM Ab or normal rabbit serum and were treated with either 0.9% saline or 6 mg IL-1ra over a 24-h time period. Plasma IL-1ra concentration was 2,659 +/- 51 ng/ml 4 h after anti-GBM Ab and IL-1ra administration. PMN and monocyte/macrophage infiltration declined 39% (9.8 +/- 1.9 to 6.0 +/- 1.5 PMN/glomerulus, P < 0.001) and 29% (4.9 +/- 0.8 to 3.5 +/- 0.8 ED-1 cells/glomerulus, P = 0.002) with IL-1ra treatment at 4 h, respectively. Similarly, the number of glomerular cells staining for lymphocyte function-associated molecule-1 beta (CD18) declined 39% from 16.7 +/- 1.9 to 10.7 +/- 1.6 cells/glomerulus at 4 h (P = 0.0001). This was associated with a decrease in glomerular intracellular adhesion molecule-1 expression. The mean glomerular intracellular adhesion molecule-1 score in anti-GBM Ab GN rats treated with IL-1ra was less than that of rats administered anti-GBM Ab and 0.9% saline at 4 (2.0 +/- 0.2 vs 2.5 +/- 0.2, P < 0.05) and 24 (2.5 +/- 0.1 vs 3.1 +/- 0.2, P = 0.0001) h. These immunopathologic changes correlated with a 50% reduction in proteinuria from 147 +/- 34 to 75 +/- 25 mg/d (P < 0.002). Treatment with IL-1ra did not affect the steady state mRNA expression of either IL-1 beta or TNF alpha. An increase in the IL-1ra dose to 30 mg given within the initial 4 h provided no additional benefit. The decline in PMN and monocyte/macrophage infiltration of the glomerulus at 4 h was similar to that found in the initial study. Furthermore, the protective benefit of IL-1ra was abrogated by doubling the dose of the anti-GBM Ab GN, despite administering high dose IL-1ra (30 mg). In these studies, detectable IL-1ra was found in the serum of untreated anti-GBM Ab GN controls. These data suggest a positive yet limited role for IL-1ra in the therapeutic intervention of anti-GBM Ab GN.     

Somatostatin infusion increases intestinal ischemia and does not improve vasoconstrictor response to norepinephrine in ovine endotoxemia

Hemodynamic support of patients with septic shock is often complicated by a tachyphylaxis against exogenous catecholamines. Because an increase in somatotropic hormones may play a pivotal role in the regulation of the inflammatory response to endotoxin, intravenous supplementation of the neuroendocrine hormone somatostatin (SOMA) may attenuate hemodynamic dysfunction resulting from endotoxemia. The objective of the present study was to assess the short-term effects of SOMA alone and in combination with norepinephrine (NE) on cardiopulmonary hemodynamics, global oxygen transport, plasma nitrate/nitrite levels, and intestinal integrity compared with single NE therapy in ovine endotoxemia. After a baseline measurement in healthy sheep (n = 16) had been performed, Salmonella typhosa endotoxin was centrally infused (10 ng x kg(-1) x min(-1)) to induce a hypotensive-hyperdynamic circulation using an established protocol. Animals surviving 16 h of endotoxemia were randomly assigned to one of the two groups (each n = 6). Sheep allocated to the SOMA + NE group received SOMA as a loading dose of 10.5 microg x kg(-1) x min(-1) for 1 h, followed by a continuous infusion of 3.5 microg x kg(-1) x min(-1) for the next 2 h. After the SOMA loading dose had been given, NE was concurrently infused (0.3 microg x kg(-1) x min(-1)) for 2 h. In the NE group (control), NE (0.3 microg x kg(-1) x min(-1)) was continuously infused for 3 h. Endotoxemia caused a decrease in MAP and systemic vascular resistance index in both groups, but to a greater extent in the NE group. Arterial hypotension persisted despite administration of the study drugs. Infusion of SOMA alone and in combination with NE did not significantly increase systemic vascular resistance index. Neither SOMA nor NE infusion alone affected pulmonary vasoregulation. Plasma nitrate/nitrite levels did not differ between groups. However, combined infusion of SOMA and NE significantly increased arterial lactate concentrations, oxygen consumption index, and oxygen extraction rate (P < 0.05) and aggravated ileal mucosal injury. In conclusion, short-term treatment with SOMA failed to attenuate cardiocirculatory shock resulting from endotoxemia and did not improve vasopressor response to NE. In addition, combined SOMA and NE therapy resulted in intestinal injury. Therefore, SOMA does not seem to represent a therapeutic option to treat arterial hypotension resulting from sepsis and systemic inflammatory response syndrome.     

Anti-ovine interleukin-1beta monoclonal antibody immunotherapy in an ovine model of gram-negative septic shock

OBJECTIVE: To investigate the efficacy of an anti-ovine interleukin-1beta monoclonal antibody to ameliorate pathophysiological derangements and improve survival in an ovine model of gram-negative septic shock. DESIGN: Prospective, placebo-controlled, interventional study (24-hr study period). SETTING: University hospital animal research laboratory. SUBJECTS: Ten awake, mature female sheep. INTERVENTIONS: Seven milligrams per kilogram of intravenous anti-ovine interleukin-1beta immunoglobin G1 monoclonal antibody (anti-interleukin-1beta group, n = 5) or equivalent amount of protein (5% human albumin; control group, n = 5) was infused over 1 hr (time-zero minus 1 hr to time-zero) and followed by an intravenous LD100 live Escherichia coli infusion (time-zero to time-zero plus 1 hr). Normal saline, maintenance and boluses to maintain baseline filling pressures, and gentamicin, 3 mg/kg intravenous, at time-zero plus 2 and time-zero plus 13 hrs. MEASUREMENTS AND MAIN RESULTS: Hemodynamic and oxygen transport indexes as well as hematological, biochemical, cytokine (interleukin-1beta, tumor necrosis factor-alpha), and endotoxin measurements were performed at baseline (time-zero minus 1 hr), on completion of the monoclonal antibody/placebo (time-zero) and E. coli (time-zero plus 1 hr) infusions, and at multiple time points thereafter (time-zero plus 1.5 hrs to time-zero plus 24 hrs). Baseline data were not different between the treatment groups. From time-zero plus 1.5 hrs onward, in the anti-interleukin-1beta group, there was a sustained increase in mean arterial pressure, decreased peripheral vasodilation, and an attenuated metabolic acidosis, relative to the control group (p < or = .01, repeated-measures analysis of variance). Predicted percentage increases in mean arterial pressure and systemic vascular resistance index relative to the control group were 35% and 40%, respectively. Resuscitation fluid requirements were also decreased: anti-interleukin-1beta group, 4.1 +/- 2.9 mL x kg(-1) x hr(-1); control group, 10.6 +/- 1.8 mL x kg(-1) x hr(-1) (p < or = .01, Student's t-test). Survival was not different (anti-interleukin-1beta group, 40%; control group, 0%; p > .01, log-rank test). CONCLUSIONS: Adjunctive therapy with anti-ovine interleukin-1beta monoclonal antibody in ovine gram-negative septic shock was associated with improved hemodynamic performance. However, the beneficial effects were incomplete and survival was not significantly improved.     

Proinflammatory cytokines increase in sepsis after anti-adhesion molecule therapy

Cytokine mediators and leukocyte-endothelial cell adhesion molecules are critical and interdependent components of the acute inflammatory response in sepsis. We hypothesized that the administration of monoclonal antibodies to intercellular adhesion molecule-1 (CD54) or E- and L-selectin (CD62E/L) would decrease serum levels of the proinflammatory cytokines interleukin-1beta (IL-1), IL-6, and IL-8 and tumor necrosis factor receptor (TNFR-1) in baboons during sepsis. Adult male baboons received infusions of 1 x 10(9) colony forming units (CFU)/kg heat-killed Escherichia coli (E. coli) followed 12 h later by live E. coli (1 x 10(10) CFU/kg). At the time of live bacterial infusion, six septic animals were treated with a monoclonal antibody to CD54 and six with an antibody to CD62E and L (1 mg/kg). Eight untreated septic animals served as controls. Sequentially drawn serum samples were assayed for IL-1, IL-6, IL-8, and TNFR-1 using enzyme-linked immunoassay (ELISA). Data were compared using Mann-Whitney U tests and Chi-square analyses. Median survival was decreased in both treatment groups compared to controls (P < 0.05). Peak IL-1 level was higher than controls in septic animals treated with anti-CD54 but not anti-CD62E/L (P < 0.05, P = NS, respectively). Elevations in IL-6, IL-8, and TNFR-1 were increased and prolonged in both antibody treated groups compared to controls (P < 0.05). These results provide the first in vivo evidence that leukocyte-endothelial adhesion molecules CD54 and CD62E/L regulate cytokine production in sepsis.     

Terlipressin as rescue therapy for intractable hypotension due to septic shock in children

Intractable hypotension due to septic shock is associated with high mortality rates in critically ill children worldwide. The use of terlipressin (triglycyl-lysine-vasopressin), an analog of vasopressin with a longer duration of action, recently emerged as a treatment of hypotension not responsive to vasopressors and inotropes. This was a retrospective study set in an 18-bed pediatric critical care department in a tertiary care children's hospital. We reviewed the files of all children with septic shock who were treated with terlipressin between January 2003 and February 2004. Fourteen children (mean age, 5.6 years; range, 4 days to 17.7 years) were treated with terlipressin in 16 septic shock episodes. Significant improvements in respiratory and hemodynamic indices were noted shortly after treatment. Mean arterial blood pressure increased significantly from 54 +/- 3 to 72 +/- 5 mmHg 10 min after terlipressin administration (P = 0.001). Heart rate decreased from 153.0 +/- 6.5 beats/min to 138.0 +/- 7.5 beats/min 12 h after treatment onset (P = 0.003). Epinephrine infusion was decreased or stopped in eight patients after terlipressin administration. Urine output increased from 1.6 +/- 0.5 mL/kg/h to 4.3 +/- 1.2 mL/kg/h 1 h after treatment onset (P = 0.011). PaO2 increased from 95.1 +/- 12.3 mmHg to 110.1 +/- 20.5 mmHg, and the oxygenation index decreased from 10.2 +/- 2.2 to 9.2 +/- 1.7. Terlipressin treatment of hypotension due to septic shock was successful in eight out of 16 episodes. Six of the 14 patients with poor prognosis for survival recovered. We conclude that terlipressin improves hemodynamic indices and renal function in critically ill children. Terlipressin should be considered as a rescue therapy in intractable shock not responsive to catecholamines in children.     

Interleukin-18 plasma levels are increased in patients with sepsis compared to severely injured patients.

Interleukin-18 (IL-18) appears to play a critical role in cytokine-induced organ failure during endotoxemia in animal models. Therefore, plasma samples from patients with severe trauma and sepsis were examined for the presence of IL-18. Significantly elevated plasma IL-18 concentrations were found in patients with sepsis compared to severely injured patients and healthy humans. Septic patients who died and patients with septic shock exhibited higher levels of IL-18 than survivors and septic patients without shock. In addition, septic patients with gram-positive infections had significantly higher IL-18 plasma levels than patients with gram-negative infection. These findings were confirmed by whole blood assay from healthy humans where Staphylococcus aureus markedly (P < 0.05) increased the release of IL-18 in whole blood ex vivo, while endotoxin was ineffective. Although obtained from a small patient group, these results suggest that IL-18 production may discriminate between gram-positive and gram-negative sepsis, and that increased IL-18 appearance may be associated with an adverse outcome in septic patients.     

Addition of alinidine, a specific bradycardic agent, to dobutamine in a canine model of endotoxic shock.

BACKGROUND AND METHODS: Alinidine is a recently developed antiarrhythmic medication that acts directly on the cardiac pacemaker cells to reduce heart rate (HR). At effective doses, alinidine might have cardiodepressant actions that could be hazardous in the presence of hemodynamic instability. On the other hand, one limitation of the use of catecholamines is tachycardia, and alinidine could be beneficial in situations such as septic shock, where adrenergic agents are commonly required. The present study explored the hemodynamic and gasometric effects of alinidine during dobutamine administration in a canine model of septic shock induced by endotoxin administration. In ten pentobarbital-anesthetized, mechanically ventilated dogs (weight 28 +/- 4 kg), Escherichia coli endotoxin (3 mg/kg) injection was followed 30 mins later by saline infusion to restore and maintain pulmonary artery occlusion pressure at the baseline value. Sixty minutes after the endotoxin administration, a dobutamine infusion was started at a rate of 10 micrograms/kg/min. Thirty minutes later, alinidine was administered as a bolus dosage of 1 mg/kg in five dogs; the other five dogs served as a control group. RESULTS: Alinidine administration resulted in a decrease in HR from 157 +/- 20 to 138 +/- 27 beats/min (p less than .01) and a nonsignificant increase in cardiac output from 5.2 +/- 3.0 to 6.8 +/- 2.8 L/min, as a consequence of increases in stroke volume from 31.9 +/- 15.3 to 49.2 +/- 13.9 mL (p less than .01) and in left ventricular stroke work from 32.1 +/- 20.0 to 57.4 +/- 32.1 g.m (p less than .05). CONCLUSIONS: During experimental septic shock, alinidine administration can reverse dobutamine-induced tachycardia and simultaneously improve ventricular function.     

Methylprednisolone reverses vasopressin hyporesponsiveness in ovine endotoxemia

Tachyphylaxis against catecholamines often complicates hemodynamic support in patients with septic shock. Recent experimental and clinical research suggests that the hemodynamic response to exogenous arginine vasopressin (AVP) infusion may also be blunted. The purpose of the present study was therefore to clarify whether the efficacy of a continuous AVP infusion (0.04 U x min(-1)) decreases over time in ovine endotoxemia. An additional objective was to determine whether the anticipated hyporesponsiveness can be counteracted by corticosteroids. Fourteen adult ewes (37 +/- 1 kg) were instrumented for chronic hemodynamic monitoring. All ewes received a continuous endotoxin infusion that contributed to a hypotensive-hyperdynamic circulation. After 16 h of endotoxemia, the sheep were randomized to receive either AVP (0.04 U x min(-1)) or the vehicle (normal saline; n = 7 each). After 6 h of AVP or placebo infusion, respectively, methylprednisolone (30 mg x kg(-1)) was injected. Arginine vasopressin infusion increased mean arterial pressure and systemic vascular resistance index at the expense of a reduced cardiac index (P < 0.05 each). Supraphysiologic AVP plasma levels in the treatment group (298 +/- 15 pg x mL(-1)) were associated with increased surrogate parameters of liver, mesenterial, and myocardial dysfunction. After 6 h of continuous AVP infusion, the vasopressor effect was significantly reduced. Interestingly, a bolus infusion of methylprednisolone (30 mg x kg(-1)) reestablished mean arterial pressure by increasing both cardiac index and systemic vascular resistance index. The present study demonstrates that in endotoxemia, (a) the vasopressor effect of AVP infusion may be reduced, (b) corticosteroids may potentially be useful to increase the efficacy of AVP infusion, and (c) even moderate AVP doses may potentially impair myocardial and hepatic function.     

High mobility group 1 protein (HMG-1) stimulates proinflammatory cytokine synthesis in human monocytes

Lipopolysaccharide (LPS) is lethal to animals because it activates cytokine release, causing septic shock and tissue injury. Early proinflammatory cytokines (e.g., tumor necrosis factor [TNF] and interleukin [IL]-1) released within the first few hours of endotoxemia stimulate mediator cascades that persist for days and can lead to death. High mobility group 1 protein (HMG-1), a ubiquitous DNA-binding protein, was recently identified as a "late" mediator of endotoxin lethality. Anti-HMG-1 antibodies neutralized the delayed increase in serum HMG-1, and protected against endotoxin lethality, even when passive immunization was delayed until after the early cytokine response. Here we examined whether HMG-1 might stimulate cytokine synthesis in human peripheral blood mononuclear cell cultures. Addition of purified recombinant HMG-1 to human monocyte cultures significantly stimulated the release of TNF, IL-1alpha, IL-1beta, IL-1RA, IL-6, IL-8, macrophage inflammatory protein (MIP)-1alpha, and MIP-1beta; but not IL-10 or IL-12. HMG-1 concentrations that activated monocytes were within the pathological range previously observed in endotoxemic animals, and in serum obtained from septic patients. HMG-1 failed to stimulate cytokine release in lymphocytes, indicating that cellular stimulation was specific. Cytokine release after HMG-1 stimulation was delayed and biphasic compared with LPS stimulation. Computer-assisted image analysis demonstrated that peak intensity of HMG-1-induced cellular TNF staining was comparable to that observed after maximal stimulation with LPS. Administration of HMG-1 to Balb/c mice significantly increased serum TNF levels in vivo. Together, these results indicate that, like other cytokine mediators of endotoxin lethality (e.g., TNF and IL-1), extracellular HMG-1 is a regulator of monocyte proinflammatory cytokine synthesis.     

Pretreatment of mice with clindamycin improves survival of endotoxic shock by modulating the release of inflammatory cytokines

Suppression of endotoxin release and subsequent production of inflammatory cytokines is crucial in the treatment of septic shock. We investigated the effect of clindamycin (CLI) on endotoxic shock induced in mice by Escherichia coli lipopolysaccharide (LPS). Mice were treated with CLI (160 to 600 mg/kg) or saline and then injected with E. coli LPS and D-(+)-galactosamine intraperitoneally 0.5 h after CLI administration. Pretreatment with CLI significantly improved survival in a dose-dependent manner (CLI, at 160, 300, and 440 mg/kg) and significantly lowered the peak concentrations of tumor necrosis factor alpha and interleukin-1beta (IL-1beta) in serum. However, the peak concentrations of IL-6 in the sera of CLI-treated mice were higher than in control mice. Our findings suggest that CLI alters LPS-induced inflammatory cytokine production and suppresses endotoxin-induced mortality in this murine model.     

Delayed neutralization of interferon-gamma prevents lethality in primate Gram-negative bacteremic shock

OBJECTIVE: Whether anticytokine therapies have a place in the treatment of severe sepsis and septic shock remains a question. Although a number of preclinical studies have shown efficacy in primate models of bacteremic shock when administered prophylactically, these same therapies have a significantly diminished effectiveness when administered therapeutically. This study investigated whether delayed administration of a novel anti-human interferon-gamma monoclonal antibody could improve outcome and reduce organ injury in a lethal model of Escherichia coli bacteremia, when administered after the onset of shock. DESIGN: Randomized, prospective, double-blinded intervention study. SUBJECTS: Cynomolgus monkeys. INTERVENTIONS: Treatment with a humanized monoclonal antibody directed against human interferon-gamma (INNO 202), administered after the onset of shock, induced by the infusion of live E. coli. MEASUREMENTS AND MAIN RESULTS: Five of the six vehicle-treated monkeys died or were killed within 24-72 hrs after E. coli administration, and all died within 5 days. In contrast, six of the eight animals treated with the anti-interferon-gamma survived for 7 days, and three of the eight animals survived 14 days (p = .013 vs. vehicle). Delayed treatment with the anti-interferon-gamma monoclonal antibody did not restore hemodynamics or reduce the amount of crystalloid-containing fluid required to resuscitate the animals but did attenuate renal failure (p < .05) and the magnitude of the inflammatory cytokine response (p < .05). CONCLUSIONS: In a primate model of E. coli bacteremic shock, delayed neutralization of interferon-gamma after the onset of shock improved survival and attenuated the pathologic changes associated with the development of organ dysfunction. These findings suggest that interferon-gamma blockade represents a potentially effective mode of late intervention in lethal septic shock.