Tissue Factor Activity in Dialysis Access Grafts.

Background: Intimal hyperplasia at the venous anastomosis of dialysis grafts causes early failure. We developed a sheep model of arteriovenous prosthetic grafts that fail rapidly due to intimal hyperplasia with histologic features nearly identical to human access grafts. A prominent feature of lesion development in this model is formation of luminal thrombus that becomes organized into stenosing lesions by macrophage and myofibroblast infiltration.

Control of smooth muscle cell function by membrane-type matrix metalloproteinases.

Vascular smooth muscle cells (SMCs) express membrane-type matrix metalloproteinases-1 and -3 (MT1- and MT3-MMPs). Expression is induced by PDGF in culture or by balloon injury in rat carotid arteries. In this study, we tried to define their functions in SMCs by transducing MT1- and MT3-MMP cDNAs into baboon-cultured SMCs, using adenoviral vectors.

Membrane-type matrix metalloproteinase-1 and -3 activity in primate smooth muscle cells.

Membrane-type matrix metalloproteinases-1 and -3 (MT1- and MT3-MMPs) are expressed by activated smooth muscle cells (SMCs) both in vitro and in vivo (19). To define their functions in SMCs, we transduced MT1- and MT3-MMP cDNAs into baboon SMCs by using adenoviral vectors. Overexpression of MT1-MMP increased the conversion of proMMP-2 to the intermediate and active forms.

Tissue factor overexpression in rat arterial neointima models thrombosis and progression of advanced atherosclerosis.

Background: Tissue factor located in the atherosclerotic plaque might cause the clinically significant thrombotic events associated with end-stage disease. It might also affect intimal area by increasing matrix accumulation and stimulating smooth muscle cell (SMC) migration and proliferation. To test this hypothesis, we overexpressed tissue factor in a rat model of the human fibrous plaque.

Local plasminogen activator inhibitor type 1 overexpression in rat carotid artery enhances thrombosis and endothelial regeneration while inhibiting intimal thickening.

Elevated levels of plasminogen activator inhibitor type 1 (PAI-1) are found in advanced atherosclerotic plaque compared with normal vessel and may contribute to plaque progression and complications associated with plaque rupture. Increased expression of PAI-1 probably contributes to the thrombotic properties of advanced atherosclerotic plaque by impeding plasmin generation and degradation of fibrin. To test this hypothesis, we have deliberately created synthetic neointimas by seeding onto the denuded luminal surface of rat carotid arteries smooth muscle cells transduced with replication-defective retrovirus encoding rat PAI-1.

Matrix metalloproteinase-9 overexpression enhances vascular smooth muscle cell migration and alters remodeling in the injured rat carotid artery.

Matrix metalloproteinase-9 (MMP-9) has been implicated in the pathogenesis of atherosclerosis as well as intimal hyperplasia after vascular injury. We used Fischer rat smooth muscle cells (SMCs) overexpressing MMP-9 to determine the role of MMP-9 in migration and proliferation as well as in vessel remodeling after balloon denudation. Fischer rat SMCs were stably transfected with a cDNA for rat MMP-9 under the control of a tetracycline-regulatable promoter.

Prevention of aneurysm development and rupture by local overexpression of plasminogen activator inhibitor-1.

Background: Arterial aneurysms exhibit a loss of elastin and an increase in the plasminogen activators urokinase plasminogen activator (u-PA) and tissue plasminogen activator (t-PA). Because u-PA, t-PA, and plasmin have a limited proteolytic activity against elastin, the role of plasminogen activators in the aneurysmal disease is unclear. To investigate this question, we overexpressed plasminogen activator inhibitor-1 (PAI-1), an inhibitor of t-PA and u-PA, in a rat model of aortic aneurysm.

Metalloproteinase blockade by local overexpression of TIMP-1 increases elastin accumulation in rat carotid artery intima.

We have recently demonstrated that the blockade of matrix metalloproteinases by local overexpression of the intrinsic inhibitor tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) reduces intimal hyperplasia. We now report a major change in the elastin content of the intima of rat carotid arteries seeded with TIMP-1-overexpressing smooth muscle cells. To understand the mechanism responsible for elastin accumulation, synthesis and degradation of elastin in TIMP-1 and control cell-seeded rats were measured.

Plasminogen activator inhibitor type 1 and tissue inhibitor of metalloproteinases-2 increase after arterial injury in rats.

Vascular injury induced by angioplasty causes smooth muscle cells to migrate, proliferate, and form a neointima. The neointima is further enlarged by the accumulation of matrix molecules synthesized by smooth muscle cells. Smooth muscle cell migration and matrix accumulation are associated with an increase in the expression of matrix-degrading enzymes and might be regulated by the balance of protease and anti-protease activity.

Overexpression of tissue inhibitor of matrix metalloproteinase-1 inhibits vascular smooth muscle cell functions in vitro and in vivo.

Arterial smooth muscle cells (SMCs) are in a quiescent growth state under normal physiological conditions, but they can be stimulated to proliferate and migrate from one tissue compartment to another if the vessel is injured. This response might require a selective and focal increase in tissue degradation, which might be mediated through the increased production of matrix metalloproteinases (MMPs). Blockade of MMP activity might therefore inhibit the SMC response to injury.

Generating antibodies against secreted proteins using vascular smooth muscle cells transduced with replication-defective retrovirus.

The traditional method of antibody (Ab) generation requires repeated injections of antigen (Ag). We have developed an alternative method that allows an investigator to generate a polyclonal antiserum with only a cDNA in hand. We cloned a cDNA encoding the coding frame for baboon tissue inhibitor of matrix metalloproteinase-1 (TIMP-1).

Cloning and characterization of a cDNA encoding the baboon tissue inhibitor of matrix metalloproteinase-1 (TIMP-1).

A baboon aortic smooth muscle cell (SMC) cDNA library was screened for the presence of tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) by polymerase chain reaction (PCR); oligodeoxyribonucleotide primers corresponding to the coding frame of the known human TIMP-1 gene were used as primers. Sequencing of the PCR-amplified baboon cDNA demonstrated only eight single-nucleotide (nt) mismatches, when compared with the coding frame of human TIMP-1. The authenticity of the PCR-amplified TIMP-1 cDNA was further confirmed by clonal screening of the library with the PCR probe and sequencing of positive clones.