Gene transfer of the urokinase-type plasminogen activator receptor-targeted matrix metalloproteinase inhibitor TIMP-1.ATF suppresses neointima formation more efficiently than tissue inhibitor of metalloproteinase-1.
Proteases of the plasminogen activator (PA) and matrix metalloproteinase (MMP) system play an important role in smooth muscle cell (SMC) migration and neointima formation after vascular injury. Inhibition of either PAs or MMPs has previously been shown to result in decreased neointima formation in vivo. To inhibit both protease systems simultaneously, a novel hybrid protein, TIMP-1.ATF, was constructed consisting of the tissue inhibitor of metalloproteinase-1 (TIMP-1) domain, as MMP inhibitor, linked to the receptor-binding amino terminal fragment (ATF) of urokinase. By binding to the u-PA receptor this protein will not only anchor the TIMP-1 moiety directly to the cell surface, it will also prevent the local activation of plasminogen by blocking the binding of urokinase-type plasminogen activator (u-PA) to its receptor. Adenoviral expression of TIMP-1.ATF was used to inhibit SMC migration and neointima formation in human saphenous vein segments in vitro. SMC migration was inhibited by 65% in Ad.TIMP-1.ATF-infected cells. Infection with adenoviral vectors encoding the individual domains, Ad.TIMP-1 and Ad.ATF, reduced migration by 32% and 52%, respectively. Neointima formation in saphenous vein organ cultures infected with Ad.TIMP-1.ATF was inhibited by 72% compared with 42% reduction after Ad.TIMP-1 infection and 34% after Ad.ATF infection. These data show that binding of TIMP-1.ATF hybrid protein to the u-PA receptor at the cell surface strongly enhances the inhibitory effect of TIMP-1 on neointima formation in human saphenous vein cultures.
Lamfers
M L M
ML
Gaubius Laboratory TNO-PG, Leiden, The Netherlands.
Grimbergen
J M
JM
Aalders
M C
MC
Havenga
M J
MJ
de Vries
M R
MR
Huisman
L G M
LG
van Hinsbergh
V W M
VW
Quax
P H A
PH
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D016466:CHO Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D002462:Cell Membrane / Q000378:metabolism; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D006224:Cricetinae; D017077:Culture Media, Conditioned / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D005434:Flow Cytometry; D018014:Gene Transfer Techniques; D006801:Humans; D066298:In Vitro Techniques; D053509:Matrix Metalloproteinase 13; D061965:Matrix Metalloproteinase Inhibitors; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D017434:Protein Structure, Tertiary / Q000502:physiology; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D011956:Receptors, Cell Surface / Q000378:metabolism; D055293:Receptors, Urokinase Plasminogen Activator; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology; D012501:Saphenous Vein / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000235:genetics* / Q000494:pharmacology; D017539:Tunica Intima / Q000187:drug effects / Q000378:metabolism*; D014568:Urokinase-Type Plasminogen Activator / Q000235:genetics*
D017077:Culture Media, Conditioned; D061965:Matrix Metalloproteinase Inhibitors; C524898:PLAUR protein, human; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D011993:Recombinant Fusion Proteins; D019715:Tissue Inhibitor of Metalloproteinase-1; D014568:Urokinase-Type Plasminogen Activator; C507079:MMP13 protein, human; D053509:Matrix Metalloproteinase 13