Assembly of the prothrombin activator complex on rabbit alveolar macrophage high-affinity factor Xa receptors. A kinetic study.
Efficient prothrombin activation occurs after assembly of factors Va, Xa, and phospholipid surface cofactor as a multimolecular complex. These components are provided by platelets and plasma within the vascular space, but molecules and membranes for prothrombin activator assembly in extravascular spaces have not been identified. In the present study, purified alveolar macrophages were found to produce high-affinity factor Xa receptors that mediate formation of enzymatic prothrombinase complexes and rapid prothrombin to thrombin conversion in the absence of exogenous factor V/Va or platelets. Thus, in reaction mixtures with alveolar macrophages cultured for 20 h in serum-free medium, the thrombin formation rate was 152 nM/min/0.66 X 10(6) cells, after adding prothrombin (1.5 microM), Ca2+ (5 mM), and factor Xa (3.7 nM). The observed Kd of factor Xa interaction with macrophage receptors is 2.1 +/- 0.94 X 10(-10) M. Kinetic analysis and inhibition studies using isolated factor V and anti-factor V antibody show that macrophage Xa receptors are functionally and antigenically similar to plasma factor V. By contrast, freshly isolated cells lacked receptors promoting prothrombin conversion at high rates. Inhibitors of protein synthesis and glycosylation, puromycin and monensin, respectively, abrogated production of Xa receptors in culture. Additionally, subcellular fractionation and enzyme-marker studies (alkaline phosphodiesterase I) indicate that internal and external membranes of alveolar macrophages have phospholipid surface cofactor activity required for prothrombinase complexes. Pulmonary surfactant is also shown to express this cofactor activity. Alveolar macrophages and surfactant comprise an efficient prothrombin activator system that is independent of plasma factor V. This system may facilitate rapid extravascular alveolar thrombin formation even at very low concentrations of factor Xa during lung defense reactions to inflammation or edema.
D000818:Animals; D002478:Cells, Cultured; D005170:Factor X / Q000378:metabolism; D015951:Factor Xa; D005260:Female; D007425:Intracellular Membranes / Q000378:metabolism; D007700:Kinetics; D008264:Macrophages / Q000378:metabolism*; D008985:Monensin / Q000494:pharmacology; D010980:Platelet Membrane Glycoproteins; D011516:Prothrombin / Q000378:metabolism*; D011650:Pulmonary Alveoli / Q000166:cytology*; D011663:Pulmonary Surfactants / Q000378:metabolism; D011691:Puromycin / Q000494:pharmacology; D011817:Rabbits; D011956:Receptors, Cell Surface / Q000378:metabolism*; D013925:Thromboplastin / Q000378:metabolism
D010980:Platelet Membrane Glycoproteins; D011663:Pulmonary Surfactants; D011956:Receptors, Cell Surface; C045069:factor X receptors; D011691:Puromycin; D011516:Prothrombin; D005170:Factor X; D013925:Thromboplastin; D008985:Monensin; D015951:Factor Xa