Isolation of multiple types of plasminogen activator inhibitors from vascular smooth muscle cells.
Large arteries have a natural resistance to tumor cell invasion thought to be due to the production of protease inhibitors. Vascular smooth muscle cells (VSMC) representing the major cellular part of arteries were isolated from human aortas and grown in tissue culture. These cells were found to produce large amounts of inhibitors of plasminogen activators (PA). Fractionation of VSMC-conditioned medium by heparin-affigel chromatography separated three immunologically and functionally distinct PA inhibitors (PAI), namely PAI-1, PAI-2 and protease-nexin I. The three inhibitors were characterized by functional assays and immunoblotting. PA inhibitor 2 (PAI-2) had little affinity for heparin, whereas PA inhibitor 1 (PAI-1) bound to heparin and was eluted from the column at NaCl concentrations of 0.1 to 0.35 M. Protease-nexin I, eluted at NaCl concentrations of 0.5 M and higher. Most of the PAI-1 was present in the latent, inactive form. PAI-1 was further purified by ion exchange chromatography on a Mono-Q column. Partial sequencing of the purified PAI-1 confirmed its nature by matching completely with the sequence deduced from the cDNA nucleotide sequence of endothelial cell PAI-1. Thus, human VSMC produce all three presently known PAI and these can be separated in single heparin affinity purification step.
Laug
W E
WE
Division of Hematology-Oncology, Children's Hospital, Los Angeles, California 90027.
Aebersold
R
R
Jong
A
A
Rideout
W
W
Bergman
B L
BL
Baker
J
J
D000595:Amino Acid Sequence; D001011:Aorta; D002846:Chromatography, Affinity; D006801:Humans; D015151:Immunoblotting; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000032:analysis*; D010960:Plasminogen Activators / Q000037:antagonists & inhibitors* / Q000378:metabolism; D015849:Plasminogen Inactivators / Q000302:isolation & purification*; D011480:Protease Inhibitors; D013917:Thrombin / Q000378:metabolism
D009097:Multienzyme Complexes; D015849:Plasminogen Inactivators; D011480:Protease Inhibitors; D010960:Plasminogen Activators; D013917:Thrombin