Matrix Metalloproteinases -14, -9 and -2 are Localized to the Podosome and Involved in Podosome Development in the A7r5 Smooth Muscle Cell.

Sean E Thatcher ,Lisa A Cassis ,Jason Edward Black ,Kazuhiro Kohama ,M E Fultz ,Hideyuki Tanaka ,Gary L Wright

doi:10.13188/2332-3671.1000020

Abstract

AimThe purpose of the study was to localize matrix metalloproteinase (MMP)-14, -9, and -2 in the A7r5 smooth muscle cell and to understand the interaction between these MMPs and the cytoskeleton. This interaction was observed under non-stimulating and phorbol 12, 13-dibutyrate (PDBu)-stimulating conditions.MethodsConfocal microscopy was utilized to define the localizations of MMPs and tissue inhibitor of matrix metalloproteinases (TIMPs) in the A7r5 cell and to determine interaction between MMPs and the cytoskeleton. Under PDBu-stimulating conditions, the presence of MMP active forms and activity by gel zymography was evaluated in the A7r5 cell. Actin and microtubule-polymerization inhibitors were used to evaluate MMP interaction with the cytoskeleton and the cytoskeleton was observed on matrix and within a Type I collagen gel.ResultsMMP-14, -9, and -2 were localized to the podosome in the A7r5 smooth muscle cell and interactions were seen with these MMPs and the actin cytoskeleton. PDBu-stimulation induced increases in the protein abundance of the active forms of the MMPs and MMP-2 activity was increased. MMPs also interact with a-actin and not β-tubulin in the A7r5 cell. Galardin, also known as GM-6001, was shown to inhibit podosome formation and prevented MMP localization to the podosome. This broad spectrum MMP inhibitor also prevented collagen gel contraction and prevented cell adhesion and spreading of A7r5 cells within this collagen matrix.ConclusionMMPs are important in the formation and function of podosomes in the A7r5 smooth muscle cell. MMPs interact with a-actin and not β-tubulin in the A7r5 cell. Podosomes play an important role in cell migration and understanding the function of podosomes can lead to insights into cancer metastasis and cardiovascular disease.

Highlights

Introduction been shown that mail: seanthatcher@uky.edu metalloproteinase (MMP)2 can degrade myosin light chains during ischemia-reperfusion in the heart indicating that may alsoMatrix metalloproteinases (MMPs) may Matrix metalloproteinases (MMPs) are endopeptidases that help affect contractile proteins [9]
The A7r5 smooth muscle cell and interactions were seen with these MMPs and the actin cytoskeleton
MMPs are important in the formation and function of podosomes in the A7r5 smooth muscle cell

Summary

Introduction

Introduction been shown that MMP2 can degrade myosin light chains during ischemia-reperfusion in the heart indicating that MMPs may Matrix metalloproteinases (MMPs) are endopeptidases that help affect contractile proteins [9]. Positive or outward remodeling is caused macrophages contain podosomes when given certain agonists, such by high levels of MMP activity resulting in a decrease in tensile as phorbol esters (PDBu), transforming growth factor-beta (TGF-β), strength of the vessel wall leading to such conditions as abdominal or other possible cytokines [10,11,12,13,14]. These mechanical perturbations can induce cytoskeletal remodeling in vascular smooth muscle cells resulting in such phenotypes as atherosclerotic plaque destabilization and rupture of the vessel. MMP activity can be controlled at the mRNA/protein levels, tissue inhibitor of metalloproteinases (TIMPs) levels, or through pharmacological intervention [3].

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