Extracellular Matrix Metalloproteinase 2 Levels Are Regulated by the Low Density Lipoprotein-related Scavenger Receptor and Thrombospondin 2

Abstract

We have recently shown that the adhesive defect observed in dermal fibroblasts derived from thrombospondin 2 (TSP2)-null mice results from an increase in matrix metalloproteinase 2 (MMP2) levels (Yang, Z., Kyriakides, T. R., and Bornstein, P. (2000) Mol. Biol. Cell 11, 3353-3364). Adhesion was restored by replacement of TSP2 and by inhibitors of MMP2 activity. In pursuing the observation that TSP2 and MMP2 interact, we now demonstrate that this interaction is required for optimal clearance of extracellular MMP2 by fibroblasts. Since TSP2 is known to be endocytosed by the scavenger receptor, low density lipoprotein receptor-related protein (LRP), we determined whether interference with LRP function affected fibroblast adhesion and/or extracellular MMP2 levels. Addition of heparin, which competes for the binding of TSP2 to LRP coreceptor proteoglycans, inhibited adhesion of control but not TSP2-null cells, and a blocking antibody to LRP as well as the LRP inhibitor, receptor-associated protein, also inhibited adhesion and increased MMP2 levels only in control fibroblasts. TSP2 did not inhibit active MMP2 directly and did not inhibit the activation of pro-MMP2. Finally, the internalization of 125 I-MMP2 was reduced in TSP2-null compared with control fibroblasts. We propose that clearance of MMP2-TSP2 complexes by LRP is an important mechanism for the regulation of extracellular MMP2 levels in fibroblasts, and perhaps in other cells. Thus, some features of the phenotype of TSP2-null mice, such as abnormal collagen fibrillogenesis, accelerated wound healing, and increased angiogenesis, could result in part from increased MMP2 activity. Thrombospondins (TSP)1 1 and 2 are large extracellular macromolecules whose diverse functions reflect their ability to bind to multiple cell-surface receptors, cytokines, growth factors, and proteases, and to structural components of the matrix (1, 2). TSP2-null mice display a complex phenotype that is characterized by changes in connective tissues, particularly in response to injury, an increase in vascular density and endosteal bone growth, and a bleeding defect (3-5). Dermal fibroblasts, isolated from adult animals, also show an adhesive defect in vitro that is most evident when cells are plated on a variety of pure protein substrates in the absence of serum. Adhesion was restored by prolonged (48 h) incubation of TSP2-null cells with recombinant mouse TSP2 or by transfection with a TSP2 cDNA gene (6). The basis for this adhesive defect was recently investigated and was shown by zymography to result from an increase in matrix metalloproteinase 2 (MMP2) levels in both the conditioned media and cell layers of cultured cells (6). Although virtually all of the enzyme that was analyzed was in the zymogen or pro-MMP2 form, an increase in active MMP2 in TSP2-null cells was inferred from the observation that inhibitors of MMP2, including TIMP2 and a neutralizing anti-MMP2 antibody, corrected the adhesive defect (6).Both TSP2 and its close relative, TSP1, are known to interact with the scave...

Thrombospondins Use the VLDL Receptor and a Nonapoptotic Pathway to Inhibit Cell Division in Microvascular Endothelial Cells

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“…To our surprise, RAP completely inhibited the stimulation of thymidine incorporation by VEGF, both in the presence or absence of TSP2 or TSP1 (Figures 5, A and B). We initially considered the possibility that LRP1 was the target of RAP, because this member of the LDL receptor family is known to bind both TSP2 and TSP1, functions as a clearance receptor for TSP-MMP2 complexes and participates in the focal adhesion disassembly that results from the interaction of TSPs with calreticulin (Mikhailenko et al, 1997;Yang et al, 2001;Elzie and Murphy-Ullrich, 2004). However, mouse microvascular endothelial cells (MVECs) have been reported to express little or no LRP1 (Lillis et al, 2005), but rather express the VLDLR, which had been presumed to perform primarily a nutritional function by endocytosis of lipoproteins (Wyne et al, 1996).…”

Gelatinase-mediated migration and invasion of cancer cells

“…The inhibitor-MMP complexes are internalized into cells via the low-density lipoprotein receptor-related protein (LRP) and are eventually degraded. However, MMP-9 can also be internalized as a monomer or TIMP-1 complex and MMP-2 with thrombospondin-2 apparently in the absence of α2-macroglobulin (Hahn- Dantona et al, 2001;Yang et al, 2001).…”

Thrombospondin 2 Inhibits Microvascular Endothelial Cell Proliferation by a Caspase-independent Mechanism

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