Lysyl oxidase propeptide inhibits smooth muscle cell signaling and proliferation

Abstract

Lysyl oxidase is required for the normal biosynthesis and maturation of collagen and elastin. It is expressed by vascular smooth muscle cells, and its increased expression has been previously found in atherosclerosis and in models of balloon angioplasty. The lysyl oxidase propeptide (LOX-PP) has more recently been found to have biological activity as a tumor suppressor, and it inhibits Erk1/2 Map kinase activation. We reasoned that LOX-PP may have functions in normal non-transformed cells. We, therefore, investigated its effects on smooth muscle cells, focusing on important biological processes mediated by Erk1/2-dependent signaling pathways including proliferation and matrix metalloproteinase-9 (MMP-9) expression. In addition, we investigated whether evidence for accumulation of LOX-PP could be found in vivo in a femoral artery injury model. Recombinant LOX-PP was expressed and purified, and was found to inhibit primary rat aorta smooth muscle cell proliferation and DNA synthesis by more than 50%. TNF-α-stimulated MMP-9 expression and Erk1/2 activation were both significantly inhibited by LOX-PP. Immunohistochemistry studies carried out with affinity purified anti-LOX-PP antibody showed that LOX-PP epitopes were expressed at elevated levels in vascular lesions of injured arteries. These novel data suggest that LOX-PP may provide a feedback control mechanism that serves to inhibit properties associated with the development of vascular pathology.Tumor necrosis factor-α (TNF-α) contributes to the development of atherosclerotic lesions [15]. Important among its many activities, TNF-α promotes the proliferation of smooth muscle cells (SMCs) [15] and it stimulates matrix metalloproteinase (MMP) production [6]. Thinning of the fibrous cap by MMPs is undesirable due to the release of sequestered factors that leads to thrombosis [15]. TNF-α regulation of MMP-9 is of particular importance in atherosclerotic lesion development and in fibrous cap thinning [3,20]. Thus, up-regulation of MMP-9 expression by TNF-α can promote development of atherosclerotic lesions by multiple mechanisms that include stimulation of SMC proliferation and MMP-9 production.

Lysyl Oxidase Propeptide Inhibits FGF-2-induced Signaling and Proliferation of Osteoblasts

“…The growth inhibitory effects of LOX-PP on osteoblast cultures are in line with recent studies that demonstrate growth inhibitory activity of LOX-PP in some transformed cell lines and in primary vascular cells (33,34,38). In vascular smooth muscle cells, tumor necrosis factor-␣ signaling is inhibited (38), although in breast cancer cells RAS-dependent pathways and FAK activation and haptotaxis are inhibited (63).…”

“…In vascular smooth muscle cells, tumor necrosis factor-␣ signaling is inhibited (38), although in breast cancer cells RAS-dependent pathways and FAK activation and haptotaxis are inhibited (63). Taken together, these findings suggest that LOX-PP has more than one molecular target and mechanism of action.…”

“…Expression and Purification of Recombinant LOX-PP-Recombinant rat LOX-PP was generated and purified to homogeneity as described earlier (38). Detailed characterization of rLOX-PP will be published elsewhere.…”

“…LOX-PP inhibits the phosphatidylinositol 3-kinase/AKT and the ERK1/2 MAP kinase pathways, as well as levels of downstream NF-B and cyclin D1 in breast, pancreatic, and lung cancer cell lines (34,35) and in prostate (36) and oral cancer cell lines (37). In addition, LOX-PP inhibits DNA synthesis in cultures of phenotypically normal, primary rat vascular smooth muscle cells (38).…”

Hypoxia-Inducible Factor 1-Regulated Lysyl Oxidase Is Involved in Staphylococcus aureus Abscess Formation

“…Slides were again washed twice with PBS (5 min each time). Sections were stained either with nonimmune rabbit IgG (10 g/ml) as a negative control or with an affinity-purified rabbit anti-rat lysyl oxidase antibody raised against amino acid residues 78 to 115 contained in the unique lysyl oxidase proregion (38) and were visualized with immunoperoxidase-based technology by using the anti-rabbit Elite VectaStain kit (Vector Laboratories, Burlingame, CA) according to the manufacturer's instructions. Corresponding sections were counterstained with hematoxylin-eosin by using standard laboratory procedures.…”