Jeffrey E. Grossman

Prolonged weakness after infusion of atracurium in two intensive care unit patients.

dministration of neuromuscular blocking (NMB) drugs to patients in the intensive care A unit (ICU) is used to facilitate mechanical ventilation, control intracranial hypertension, eliminate shivering, decrease oxygen consumption, and facilitate diagnostic procedures and studies (1,2). In the United States, the steroid-based NMB drugs, vecuronium and pancuronium, are most frequently selected for ICU administration (1,2). However, numerous reports suggest that prolonged administration of these drugs to ICU patients may result in sustained weakness, persisting days to weeks after withdrawal of the drug (3-5). Three features are common to these reports: 1) the prolonged, usually continuous infusion of steroid-based NMB agents, most often without peripheral nerve twitch monitoring, 2) concurrent administration of drugs which may alter neuromuscular transmission such as aminoglycoside antibiotics, calcium-channel blockers, magnesium, and especially high-dose corticosteroids, and 3 ) end-organ dysfunction such as significant renal failure (3-11). Because of concern about administration of steroidbased NMB drugs in patients receiving concurrent exogenous corticosteroids, recommended strategies include the use of routine peripheral nerve twitch monitoring and the selection of neuromuscular blocking agents with an alternative (nonsteroid) pharmacologic structure (12). Atracurium, an intermediate-acting benzylisoquinolinium compound, is structurally unrelated to the aminosteroid NMB agents (13). Introduced in 1983, atracurium is a unique NMB drug that undergoes ester hydrolysis and Hofmann elimination (independent of hepatic or renal function) to laudanosine and monoacrylate metabolites (131, which are inactive at the neuromuscular junction.

Response of plasma fibronectin to major body burn.

The ability of macrophages of phagocytize particulate matter is largely dependent on fibronectin, a nonspecific opsonin found in plasma. Fibronectin depletion, resulting in reticuloendothelial system (RES) depression, occurs following a variety of physical insults. RES depression may contribute to postinjury sequelae such as respiratory distress syndrome and septicemia. Fibronectin concentration was measured in the plasma of sheep with chronic lung lymph fistulas subjected to controlled thermal injury. Fibronectin levels were significantly (p < 0.05) decreased at 4, 24, 48, and 72 hours following burning. Fibronectin concentration decreased in parallel with serum albumin concentration; serum globulin concentrations did not decrease. Fibronectin concentration had an inverse relationship to lung lymph flow, a reflection of pulmonary transvascular fluid filtration rate. The role of fibronectin in the pathogenesis of postinjury respiratory distress syndrome deserves further exploration.

Plasma fibronectin concentration in animal models of sepsis and endotoxemia

Plasma fibronectin is a nonspecific opsonin which mediates phagocytosis of particulate matter by macrophages. Fibronectin depletion results in depression of reticuloendothelial system phagocytic function. This may potentiate microvascular embolization and sludging in critical illness. It has been hypothesized that sepsis is a major cause of fibronectin depletion. To explore this hypothesis, plasma fibronectin concentrations were measured in rats with intraabdominal abscesses and in rabbits subjected to the generalized Shwartzman reaction (spaced doses of endotoxin). In both groups of animals there was a significant increase (P less than 0.05) rather than decrease in fibronectin concentrations at times when sepsis and disseminated intravascular coagulation were manifest. This study does not support the hypothesized relationship between sepsis and fibronectin depletion. Until the kinetics of fibronectin production and utilization are further delineated, caution must be exercised in the interpretation of immunoreactive plasma fibronectin levels.

ATTENUATION OF ENDOTOXIN-INDUCED PATHOPHYSIOLOGY BY A NEW POTENT PAF RECEPTOR ANTAGONIST

The role of platelet-activating factor (PAF) as a mediator of endotoxin-induced pathophysiology has been studied in several animal models with conflicting results. We evaluated the effect of a new, potent, and specific PAF receptor antagonist, ABT-299 (Abbott Laboratories) against endotoxin (lipopolysaccharide; LPS)-induced cardiopulmonary dysfunction in a porcine model. In initial experiments, the potency of ABT-299 was confirmed in vitro by its ability to inhibit PAF-induced porcine platelet aggregation at an IC50 of .047 ± .01 μM, and in vivo by the ability of low doses (.12 mg/kg + .03 mg/kg/h) to block the cardiopulmonary pathologic response to exogenous PAF infusion. To evaluate the effect of ABT-299 administration during endotoxemia, pigs were randomly assigned to one of three groups: controls (n = 7), LPS (n = 9), or ABT-299 + LPS (n = 7). ABT-299 was given at 1.0 mg/kg from −0.5 to 0 h plus .3 mg/kg/h from 0 to 6 h. LPS was given at .5 μg/kg/hr from 0 to 6 h. ABT-299 reduced the early LPS-induced fall in cardiac index and stroke volume, pulmonary hypertension and vasoconstriction, bronchoconstriction, and hypoxemia. Administration of LPS resulted in 44% mortality (before 6 h), which was blocked by ABT-299. Results with this antagonist indicate that PAF contributes to endotoxin-induced cardiopulmonary dysfunction in the pig, and is associated with mortality in this model.

Lung injury and lung lysosomal enzyme release during endotoxemia

Abstract We measured the release of the lysosomal enzymes β-glucuronidase and aryl sulfatase A into lung lymph and plasma after Escherichia coli endotoxin and compared these changes with the pulmonary vascular injury produced. We used 18 unanesthetized sheep, infusing doses of endotoxin from 1 to 10 αg/kg. We identified an early injury phase characterized by severe pulmonary hypertension, a three- to fivefold increase in protein-poor lung lymph flow, and a marked decrease in blood leukocyte count. This was followed several hours later by an increase in vascular permeability with a three- to fivefold increase in protein-rich lung lymph. Lymph lysosomal enzymes increased 200–800% over baseline with a sharp increase occuring during the early hypertension phase. Lymph β-glucuronidase correlated well with the change in vascular permeability measured by lymph flow, r = 0.7. Plasma enzyme values increased, but to a much lesser degree, correlating poorly, r = 0.2, with the lung injury. However, increases over baseline greater than 200% corresponded into with later systemic shock and mortality. We conclude that lysosomal enzymes are released into the lung after endotoxin, probably from sequestered leukocytes, with the degree of release corresponding to the degree of vascular injury.

REGULATION OF LEUKOCYTE FUNCTION BY NITRIC OXIDE DONORS: THE EFFECT OF S-NITROSO-THIOL COMPLEXES

The aim of this study was to evaluate the effectiveness of a novel protein-bound Snitroso-thiol, S-nitroso-albumin (S-NO-alb) , in modulating neutrophil–endothelial cell adhesion, activation, and interactions. Due to the highly variable kinetics of NO release from the low-molecular-weight thiol adducts S-nitroso-N-acetylpenicillamine (SNAP) and S-nitroso-glutathione (GSNO) , we expected S-NO-alb to be a more effective modulator of inflammatory interactions through its slow, steady, and prolonged release of NO. Human umbilical-vein endothelial cells (HUVECs) challenged with lipopolysaccharide (LPS) demonstrated upregulated adhesion of neutrophils that was significantly attenuated by pretreatment with S-NO-alb (1.0–100 μM) (p < .05), but not SNAP or GSNO. Pretreatment with S-NO-alb, SNAP, or GSNO attenuated tumor necrosis factor-a primed O2 – release from neutrophils and increased neutrophil cGMP accumulation. On a molar basis, S-NO-alb expressed a 10-fold greater potency than SNAP or GSNO at modulating these effects. Kinetics studies confirmed the relative stability of spontaneous NO release from S-NO-alb compared with highly variable kinetic profiles of SNAP and GSNO. Our results demonstrate that S-NO-alb more effectively modulates endothelial-cell and neutrophil immunoinflammatory responses versus its related low-molecular-weight thiol complexes.The aim of this study was to evaluate the effectiveness of a novel protein-bound S-nitroso-thiol, S-nitroso-albumin (S-NO-alb), in modulating neutrophil-endothelial cell adhesion, activation, and interactions. Due to the highly variable kinetics of NO release from the low-molecular-weight thiol adducts S-nitroso-N-acetylpenicillamine (SNAP) and S-nitroso-glutathione (GSNO), we expected S-NO-alb to be a more effective modulator of inflammatory interactions through its slow, steady, and prolonged release of NO. Human umbilical-vein endothelial cells (HUVECs) challenged with lipopolysaccharide (LPS) demonstrated upregulated adhesion of neutrophils that was significantly attenuated by pretreatment with S-NO-alb (1.0-100 microM) (p < .05), but not SNAP or GSNO. Pretreatment with S-NO-alb, SNAP, or GSNO attenuated tumor necrosis factor-alpha primed *O2- release from neutrophils and increased neutrophil cGMP accumulation. On a molar basis, S-NO-alb expressed a 10-fold greater potency than SNAP or GSNO at modulating these effects. Kinetics studies confirmed the relative stability of spontaneous NO release from S-NO-alb compared with highly variable kinetic profiles of SNAP and GSNO. Our results demonstrate that S-NO-alb more effectively modulates endothelial-cell and neutrophil immunoinflammatory responses versus its related low-molecular-weight thiol complexes.

Modulation of endotoxin-induced cardiopulmonary dysfunction by S-nitroso-albumin

Nitric oxide (NO) is an endogenous vasodilator and modulator of inflammation. During endotoxemia, the beneficial effects of NO are overwhelmed by the inflammatory cascade, resulting in a functional depletion of NO. S-nitroso-albumin (S-NO-alb) exists as a novel and highly stable NO thiol complex that slowly releases NO into the vascular micro-environment. Using a porcine model, we examined the ability of intravenous S-NO-alb to modulate cardiopulmonary dysfunction characteristic of endotoxemia. Pigs were anesthetized, instrumented for standard cardiopulmonary function measurements, and randomly assigned to receive: (i) albumin + saline; (ii) albumin + LPS; or (iii) S-NO-alb + LPS. Cardiopulmonary parameters were evaluated every 30 min and ex vivo phorbol myristate acetate (PMA)-stimulated superoxide release was serially determined as a marker of in vivo neutrophil priming. Lung myeloperoxidase (MPO) activity was measured as a marker of neutrophil migration into the lung. LPS-induced cardiopulmonary dysfunction was characterized by a sustained elevation in mean pulmonary arterial pressure, pulmonary vascular resistance, and peak intratracheal pressure, as well as a reduction in cardiac index, stroke volume index and PaO2 over 6 h. Pretreatment with S-NO-alb attenuated LPS-induced cardiopulmonary dysfunction without adversely affecting systemic hemodynamics. Moreover, S-NO-alb blunted the LPS-induced hypoxemic response and reduced neutrophil activation. S-NO-alb did not, however, attenuate LPS-induced increases in lung MPO. Our results suggest that S-NO-alb can selectively modulate endotoxin-induced pulmonary dysfunction, attenuate neutrophil priming and block the early mortality (40%) in this model.

Low molecular weight heparin alters porcine neutrophil responses to platelet-activating factor.

Because platelet-activating factor (PAF) is an important mediator of inflammation and heparin has anti-inflammatory effects, we hypothesized that low molecular weight heparin (LMWH) would inhibit PAF-induced activation and chemotaxis in porcine neutrophils. Citrated blood was obtained from pentobarbital-anesthetized pigs, and neutrophils were isolated over a 55%/65% Percoll gradient. The effect of LMWH on basal phorbol myristate acetate (PMA)-induced superoxide (SO) release, as well as its effect on PAF priming for PMA-induced SO release, were investigated. Additionally, the effect of LMWH on PAF-induced chemotaxis of neutrophils across transwell membranes was evaluated. Baseline SO release in response to PMA was .351+/-.046 nmol/10(6) cells/min, and this was decreased to .289+/-.034 nmol/10(6) cells/min by pretreatment with 50 U/mL LMWH. PMA-induced SO production was increased by .240+/-.042 nmol/10(6) cells/min when cells were primed with 10 microM PAF. This priming effect of PAF was reduced significantly by pretreatment of neutrophils with LMWH at 10 and 50 U/mL. Chemotaxis of neutrophils in response to 100 microM PAF was significantly decreased to 70.02+/-6.4% (n = 8) of the control response by pretreatment of cells with 50 U/mL LMWH. We conclude that LMWH has anti-inflammatory effects on porcine neutrophils, which includes attenuation of cell activation and chemotaxis in response to the lipid-derived inflammatory mediator, PAF.

Clinical use of fibronectin

SummaryThere is considerable experimental evidence that the RES is an important host defense system in critical illness. The role of fibronectin in the control of RES phagocytic function and the potential for fibronetin therapy to ameliorate or reverse multisystem organ failure in critically ill patients are less well documented. This documentation may be difficult to accrue in the clinical setting because of the multifactorial problems of individual critically ill patients and the heterogeneity of underlying diseases in such patients. More work needs to be done in animal models in which these variables can be controlled. Large scale production of pasteurized biologically active fibronection should be possible32 if the cryoprecipitate studies prove the clinical utility of fibronectin concentrates. Fibronectin will certainly not be a panacea for critically ill patients, but the addition of any useful substance to the now meager armamentarium would be most welcome.

An Immunocytochemical Method for Quantification of Lung Tissue Fibronectin

Abstract A technique to quantify tissue fibronectin was developed, using peroxidase-anti-peroxidase immunocytochemistry and automated scanning light microscopy. This technique was developed using isolated perfused rat lungs, some of which were subjected to acute oxidant lung injury. Both injured and control lungs were perfused with solutions containing heterologous fibronectin. The technique clearly demonstrated differences in the amount of tissue fibronectin in injured and noninjured lung as well as differences between lungs exposed to fibronectin and those not exposed. The described technique offers a reliable method for quantifying tissue fibronectin and is sensitive enough to detect differences in tissue fibronectin under experimental conditions of acute lung injury.