Inhibition of apoptosis and caspase-3 in vascular smooth muscle cells by plasminogen activator inhibitor type-1.
Increased expression of plasminogen activator inhibitor type 1 (PAI-1) is associated with decreased apoptosis of neoplastic cells. We sought to determine whether PAI-1 alters apoptosis in vascular smooth muscle cells (VSMC) and, if so, by what mechanisms. A twofold increase in the expression of PAI-1 was induced in VSMC from transgenic mice with the use of the SM-22alpha gene promoter (SM22-PAI+). Cultured VSMC from SM22-PAI+ mice were more resistant to apoptosis induced by tumor necrosis factor plus phorbol myristate acetate or palmitic acid compared with VSMC from negative control littermates. Both wild type (WT) and a stable active mutant form of PAI-1 (Active) inhibited caspase-3 amidolytic activity in cell lysates while a serpin-defective mutant (Mut) PAI-1 did not. Similarly, both WT and Active PAI-1 decreased amidolytic activity of purified caspase-3, whereas Mut PAI-1 did not. WT but not Mut PAI-1 decreased the cleavage of poly-[ADP-ribose]-polymerase (PARP), the physiological substrate of caspase-3. Noncovalent physical interaction between caspase-3 and PAI-1 was demonstrable with the use of both qualitative and quantitative in vitro binding assays. High affinity binding was eliminated by mutations that block PAI-1 serpin activity. Accordingly, attenuated apoptosis resulting from elevated expression of PAI-1 by VSMC may be attributable, at least in part, to reversible inhibition of caspase-3 by active PAI-1.
Chen
Yabing
Y
Department of Medicine, The University of Vermont, Burlington, Vermont 05405, USA. ybchen@zoo.uvm.edu
Kelm
Robert J
RJ
Budd
Ralph C
RC
Sobel
Burton E
BE
Schneider
David J
DJ
D000818:Animals; D001011:Aorta / Q000166:cytology / Q000378:metabolism; D017209:Apoptosis; D053148:Caspase 3; D061945:Caspase Inhibitors; D018929:Cell Culture Techniques; D051379:Mice; D008822:Mice, Transgenic; D008840:Microfilament Proteins / Q000235:genetics; D009124:Muscle Proteins / Q000235:genetics; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism* / Q000648:ultrastructure; D017395:Plasminogen Activator Inhibitor 1 / Q000235:genetics / Q000378:metabolism*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D015854:Up-Regulation / Q000235:genetics
D061945:Caspase Inhibitors; D008840:Microfilament Proteins; D009124:Muscle Proteins; D017395:Plasminogen Activator Inhibitor 1; D014409:Tumor Necrosis Factor-alpha; C051615:transgelin; D011065:Poly(ADP-ribose) Polymerases; C505409:Casp3 protein, mouse; D053148:Caspase 3