title: Thrombin-treated endothelium primes neutrophil functions: inhibition by platelet-activating factor receptor antagonists.
abstract: We have shown previously that fluid phase platelet-activating factor (PAF) can enhance or "prime" polymorphonuclear (PMN) responses to subsequent stimulation with agonists such as formyl-methionine-leucine-phenylalanine (FMLP). Since thrombin induces PAF production in endothelial cells, we tested whether this thrombin-provoked endothelial PAF primes responses of marginated PMNs. Monolayers of human umbilical vein endothelial cells were exposed to either thrombin (0.5-5.0 units/ml) or buffer for up to 5 min and then PMNs were layered on top of the endothelial cells. After a further 5 min incubation, the PMNs were stimulated with a suboptimal concentration of FMLP (10(-7) M), and their superoxide production, elastase release, adhesion to endothelium, and capacity to cause endothelial cell lysis and detachment were assessed. Thrombin pretreatment significantly enhanced each of these FMLP-stimulated neutrophil responses. The extent of this enhancement correlated with both the dose and duration of thrombin treatment of endothelial cells and also the duration of PMN incubation with thrombin-exposed endothelium. Evidence that the augmentation was due to endothelial-derived PAF was obtained as follows: (1) thrombin induced [3H]acetate incorporation into endothelial PAF (assayed in lipid extracts); (2) antithrombin III conjointly inhibited this [3H]acetate uptake and prevented the priming effect of thrombin-treated endothelium on PMN responses; and (3) the PAF receptor antagonist BN52021, when preincubated with PMNs, also effectively blocked the enhancement of PMN responses. We conclude that thrombin stimulation of endothelial cells initiates a sequence of events culminating in the production of PAF--a membrane phospholipid capable of priming marginated PMNs. We suggest that this coagulation-fostered endothelial/PMN interaction may underlie a paracrine response that may potentiate PMN-mediated endothelial injury during sepsis and other thrombin-generating disorders.
authors.affiliation: Department of Medicine, University of Minnesota Medical School, Minneapolis.
authors.forename: G M
authors.initials: GM
authors.lastname: Vercellotti
authors.forename: N W
authors.initials: NW
authors.lastname: Wickham
authors.forename: K S
authors.initials: KS
authors.lastname: Gustafson
authors.forename: H Q
authors.initials: HQ
authors.lastname: Yin
authors.forename: M
authors.initials: M
authors.lastname: Hebert
authors.forename: H S
authors.initials: HS
authors.lastname: Jacob
mesh_terms: D004730:Endothelium, Vascular / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000502:physiology*; D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface / Q000037:antagonists & inhibitors*; D043562:Receptors, G-Protein-Coupled; D013917:Thrombin / Q000494:pharmacology*
chemical_list: D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; C039351:platelet activating factor receptor; D009240:N-Formylmethionine Leucyl-Phenylalanine; D013917:Thrombin