SummaryA number of recent studies have demonstrated that cellular responses to tumor necrosis factor (TNF) mediated by the p55 and the p75 TNF receptors are distinct. To evaluate the relative in vivo toxicities of wild-type TNFot (wtTNFc~) and a novel p55 TNF selective receptor agonist, healthy, anesthetized baboons (Papio sF) were infused with a near-lethal dose of either wtTNFc~ or a TNFc~ double mutant (dmTNFo 0 that binds specifically to the p55, but not to the p75, TNF receptor. Both wtTNFc~ and dmTNFcr produced comparable acute hypotension, tachycardia, increased plasma lactate, and organ dysfunction in Papio. However, administration of wt TNFo~ produced a marked granulocytosis and loss of granulocyte TNF receptors, whereas little if any changes in neutrophil number or cell surface TNF receptor density were seen after dmTNFcx mutant administration. Infusion of dmTNFcx resulted in a plasma endogenous TNFc~ response that peaked after 90-120 min. We conclude that selective p55 TNF receptor activation is associated with early hemodynamic changes and the autocrine release of endogenous TNFc~. Significant systemic toxicity results from p55 TNF receptor activation, but the role of the p75 TNF receptor in systemic TNF toxicity requires further study.

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ObjectiveThe in vivo neutralizing activities of an anti-lipopolysaccharide (LPS) antibody HA-lA (Centoxin [Centocor, Malvern, PA]), a human immunoglobulin M monoclonal antibody, and of bactericidal/ permeability-increasing protein (BPI), an endogenously produced human LPS-neutralizing protein, were studied in a primate model of lethal Escherichia coli bacteremia. Summary Background DataHA-1A has been used with variable success against LPS activity in some animal models and in a recently reported clinical trial. However, no data assessing the efficacy of this agent in subhuman primates is available. Bactericidal/permeability-increasing protein is a product of polymorphomononuclear cells (PMNs) that is stored in azurophilic granules and exhibits LPSneutralizing activity in vitro and in some in vivo models. MethodsImmediately after E. coli infusion and in a blinded fashion, three baboons were treated with BPI (5 mg/kg bolus infusion and 95 ,ug/kg/min infusion over 4 hr). Three animals received 3 mg/kg BW of HA-1 A, whereas another three baboons received a placebo treatment. ResultsThe BPI-treated animals demonstrated significantly (p < 0.03) lower circulating LPS-limulus amoebocyte lysate (LAL) activity compared with the control animals, but this reduction in LPS-LAL activity was not associated with improved survival. HA-1 A treatment did not reduce LPS-LAL activity. However, both BPI and HA-1 A treatment did aftenuate the pro-inflammatory cytokine response. ConclusionThe current data suggests that incomplete neutralization of endotoxin activity does not alter mortality from severe bacteremia. Given the diversity of mediator production under such circumstances, a strategy of combination therapy in the form of anti-lipopolysaccharide and anticytokine treatment may be necessary to achieve optimal survival. 77

Triglyceride-rich lipoproteins can inhibit endotoxin activity in vitro and in rodents. We sought to determine whether Intralipid, a triglyceride-rich fat emulsion which in contact with plasma functions similarly to endogenous lipoproteins, can alter the human response to endotoxin. Intralipid inhibited endotoxin-induced cytokine production in human whole blood in vitro in a dose-dependent manner, with maximal inhibition (up to 70%) being achieved at a concentration of 10 g/liter. In healthy men, a bolus intravenous injection of endotoxin (lot EC-5; 20 U/kg of body weight) was given midway through a 4-h infusion (125 ml/h) of either 5% glucose (n ‫؍‬ 5) or 20% Intralipid (n ‫؍‬ 5). The infusion of Intralipid led to an increase in triglyceride levels in serum from 95 ؎ 16 to 818 ؎ 135 mg/dl prior to endotoxin administration, i.e., levels that importantly reduced cytokine production in endotoxin-stimulated whole blood. However, in vivo hypertriglyceridemia did not influence inflammatory responses to endotoxin (fever, release of tumor necrosis factor and soluble tumor necrosis factor receptors, and leukocytosis) or even potentiated endotoxin responses (release of interleukins 6 and 8 and neutrophil degranulation). Hypertriglyceridemia does not inhibit the in vivo responses to endotoxin in humans.

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Human neutrophil azurophilic granules contain an approximately 55-kDa protein, known as bactericidal/permeability-increasing protein (BPI), which possesses a high-affinity binding domain for the lipid A component of lipopolysaccharide (LPS). The in vivo LPS neutralizing activity of exogenous BPI was studied in a model of lethal Escherichia coli bacteremia. Five baboons were treated with BPI (5 mg/kg bolus injection followed by a 95 micrograms/kg/min BPI infusion over 4 hr), while four additional animals received a genetically engineered variant of BPI (NCY103). Five animals received a placebo treatment and served as controls. Both wild-type rhBPI and NCY103 significantly (P < 0.05) decreased blood levels of LPS throughout an 8-hr evaluation period following live bacterial challenge. Two hours following E. coli administration, LPS levels peaked in the controls, at 6.86 +/- 3.22 ng/ml, whereas LPS levels were 3.39 +/- 2.1 ng/ml in the BPI group and 2.04 +/- 1.18 ng/ml in the NCY103 group. Tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 levels likewise were attenuated in the treatment groups, whereas circulating sTNFR I was significantly (P < 0.05) reduced only in the BPI group. Leukocytopenia and granulocytopenia were significantly (P < 0.02) lessened in the BPI group, by an average of 59% leukocytopenia and 65% granulocytopenia, respectively. This study supports the concept of E. coli LPS neutralization by BPI in vivo and demonstrates that a moderate (70%) reduction in peak LPS-LAL activity is sufficient to alter some hematologic and cytokine manifestations of bacteremia.

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