Hemoglobin infusion augments the tumor necrosis factor response to bacterial endotoxin (lipopolysaccharide) in mice

Objective: To determine whether cell-free hemoglobin augments the inflammatory cascade, as detected by production of tumor necrosis factor (TNF) elicited by bacterial endotoxin (lipopolysaccharide [LPS]), Design: In vivo and ex vivo study, using a mouse model of sepsis, Setting: Animal research facility Subjects: Female Swiss Webster mice Interventions: For the in vivo experiments, an LD50 dose (500 mu g) of Escherichia coli LPS was injected intraperitoneally into mice. Cell free crosslinked hemoglobin (60 mg/mouse) or saline was administered intravenously 10 hrs before or coincident with LPS, For the ex vivo experiments, hemoglobin (60 mg/mouse) or saline was administered intravenously to mice, and, 10 hrs later, hepatic Kupffer cells, peripheral blood mononuclear cells, or peritoneal macrophages were isolated. Measurements and Main Results: Intravenous infusion of hemoglobin either 10 hrs before or coincident with intraperitoneal LPS resulted in a peak of plasma TNF that was greater than in control mice administered LPS only. Cultured Kupffer cells, isolated from mice that had received hemoglobin in vivo 10 hrs before cell collection, produced more TNF in response to LPS in vitro than cells from normal mice. A trend toward greater TNF production in vitro by peripheral blood mononuclear cells obtained from hemo globin treated mice also was observed. Enhanced sensitivity to LPS was not observed with cultured peritoneal macrophages from mice that had received hemoglobin. Conclusions: Intravenous hemoglobin increased the sensitivity of hepatic macrophages to subsequent stimulation by LPS, This effect may contribute to the increased mortality that we have observed in animals that have received both LPS and hemoglobin.