Utilization of a continuous flow reactor to study the lipoprotein-associated coagulation inhibitor (LACI) that inhibits tissue factor.
A microperfusion system containing a glass capillary, the inner surface of which is coated with a phospholipid bilayer containing tissue factor, was used to explore the requirement for factors VIIa and Xa in the complex formed with the lipoprotein-associated coagulation inhibitor (LACI). Various combinations of factors VIIa, Xa, and LACI were perfused together or sequentially at a wall shear rate of 300 sec-1; a final perfusion of factors X and VIIa was performed to evaluate the residual tissue factor activity. Factor Xa concentration at the outlet of the tube was determined using a chromogenic substrate. In the presence of factors VIIa, Xa, and LACI, complete inhibition of tissue factor was observed on both phosphatidylcholine (neutral surfaces) and on phosphatidylserine/phosphatidylcholine (acidic) surfaces; omission of factors Xa or LACI resulted in no inhibition. The absence of factor VIIa in the initial perfusion steps resulted in no inhibition on neutral surfaces whereas about 90% inhibition was observed on acidic surfaces. Initial perfusion with factor Xa, but not LACI, followed by the remaining protein components, resulted in an inhibitory complex. Thus, it appears that a tissue factor:factor Xa:LACI complex can form in the absence of factor VIIa on acidic surfaces; moreover, our data imply a tissue factor binding site for factor Xa, but not for LACI.
Gemmell
C H
CH
Department of Medicine, Mt Sinai Medical Center, New York, NY 10029.
Broze
G J
GJ
Turitto
V T
VT
Nemerson
Y
Y
D002118:Calcium / Q000494:pharmacology; D002197:Capillary Action; D004305:Dose-Response Relationship, Drug; D005167:Factor VII / Q000008:administration & dosage / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D015942:Factor VIIa / Q000378:metabolism / Q000494:pharmacology; D015951:Factor Xa / Q000378:metabolism / Q000494:pharmacology; D005898:Glass; D007700:Kinetics; D008074:Lipoproteins / Q000008:administration & dosage / Q000494:pharmacology*; D010477:Perfusion; D010713:Phosphatidylcholines; D010718:Phosphatidylserines; D011480:Protease Inhibitors / Q000494:pharmacology*; D013925:Thromboplastin / Q000008:administration & dosage / Q000037:antagonists & inhibitors* / Q000494:pharmacology
D008074:Lipoproteins; D010713:Phosphatidylcholines; D010718:Phosphatidylserines; D011480:Protease Inhibitors; C051928:lipoprotein-associated coagulation inhibitor; D005167:Factor VII; D013925:Thromboplastin; D015942:Factor VIIa; D015951:Factor Xa; D002118:Calcium