Prevention of endotoxin-induced mortality by antitissue factor immunization
A. P. Dackiw, I. D. McGilvray, M. Woodside, A. B. Nathens, J. C. Marshall and O. D. Rotstein
Department of Surgery, Toronto Hospital, University of Toronto, Ontario.
BACKGROUND: Microvascular thrombosis with intravascular fibrin deposition
is a characteristic pathologic alteration during endotoxic shock. This
effect is predominantly mediated by expression of the cellular procoagulant
tissue factor by endothelial cells and cells of monocyte or macrophage
lineage, resulting in acceleration of the coagulation cascade and fibrin
deposition. OBJECTIVE: To determine whether modulation of this response by
treatment with an antitissue factor antibody might have beneficial effects.
DESIGN: A polyclonal antibody to murine tissue factor was prepared by
injecting rabbits with a synthesized peptide sequence of murine tissue
factor. To determine the activity of the antibody, elicited murine
peritoneal macrophages were treated for 4 hours with 10-micrograms/mL
lipopolysaccharide (LPS), and procoagulant activity was determined via a
clotting assay (milliunits of activity per 10(6) macrophages). RESULTS: The
tissue factor antibody abrogated LPS-induced macrophage procoagulant
activity, confirming activity of the antibody (macrophages, 236 +/- 28
mU/10(6) macrophages; macrophages/LPS, 3801 +/- 190* mU/10(6) macrophages;
macrophages/LPS/alpha-tissue factor, 753 +/- 92* mU/10(6) macrophages; n =
3; the asterisk indicates P < .05 by an analysis of variance).
Additionally, antibody-protein affinity was confirmed by Western blot
analysis. Having determined the activity of the antibody in vitro, we
tested its efficacy in vivo in a lethal endotoxemia model. Mice were
immunized with 200 microL of antiserum intraperitoneally 2 hours before
injection with 250 micrograms of LPS intraperitoneally and 24 hours later.
Control animals received 200 microL of saline solution. All animals
initially exhibited lethargy and piloerection, characteristic of the
predicted response to LPS. However, immunized animals had a significantly
(P < .05) reduced mortality compared with control animals. Fibrinogen
levels were significantly (P < .05) higher in the immunized mice,
suggesting decreased consumption of coagulation factors, a finding
consistent with an antitissue factor effect. Further, plasma tumor necrosis
factor levels 90 minutes after LPS injection were similar in both groups,
suggesting normal induction of the cytokine cascade. CONCLUSIONS:
Modulation of microvascular fibrin deposition by abrogating tissue
factor-mediated coagulation significantly (P < .05) improved survival in
this model without attenuating the initiation of the cytokine cascade.
These findings suggest a pathogenic role for coagulation in the induction
of acute organ injury during sepsis.
