ADAM9 Is Involved in Pathological Retinal Neovascularization

Abstract

Pathological ocular neovascularization, caused by diabetic retinopathy, age-related macular degeneration, or retinopathy of prematurity, is a leading cause of blindness, yet much remains to be learned about its underlying causes. Here we used oxygen-induced retinopathy (OIR) and laser-induced choroidal neovascularization (CNV) to assess the contribution of the metalloprotease-disintegrin ADAM9 to ocular neovascularization in mice. Pathological neovascularization in both the OIR and CNV models was significantly reduced in Adam9 ؊/؊ mice compared to wild-type controls. In addition, the level of ADAM9 expression was strongly increased in endothelial cells in pathological vascular tufts in the OIR model. Moreover, tumor growth from heterotopically injected B16F0 melanoma cells was reduced in Adam9 ؊/؊ mice compared to controls. In cell-based assays, the overexpression of ADAM9 enhanced the ectodomain shedding of EphB4, Tie-2, Flk-1, CD40, VCAM, and VE-cadherin, so the enhanced expression of ADAM9 could potentially affect pathological neovascularization by increasing the shedding of these and other membrane proteins from endothelial cells. Finally, we provide the first evidence for the upregulation of ADAM9-dependent shedding by reactive oxygen species, which in turn are known to play a critical role in OIR. Collectively, these results suggest that ADAM9 could be an attractive target for the prevention of proliferative retinopathies, CNV, and cancer.Ocular neovascularization is one of the leading causes of blindness in humans and is found in diverse eye diseases including diabetic retinopathy, age-related macular degeneration, and retinopathy of prematurity (3,4,6). In addition, pathological neovascularization also has critical roles in other diseases such as cancer and rheumatoid arthritis (12,14). Although proteins with crucial functions in pathological neovascularization are considered to be important targets for the treatment of tumor growth (5), proliferative retinopathies (19), and rheumatoid arthritis (12), much remains to be learned about the identity of these molecules and the mechanisms underlying their function. In this study, we focused on the contribution of a disintegrin and metalloprotease, ADAM9, to pathological neovascularization. ADAM9, one of the first ADAM proteins to be identified and characterized, is a membrane-anchored metalloproteinase containing an N-terminal prodomain followed by a metalloprotease domain, a disintegrin domain and cysteine-rich region, an epidermal growth factor (EGF) repeat, a transmembrane domain, and a cytoplasmic tail with potential SH3 ligand domains (25). ADAM9 is catalytically active in both biochemical and cell-based assays and can cleave several membrane proteins including EGF and FGFR2iiib when it is overexpressed together with these substrates (10, 15, 16). In addition, ADAM9 is thought to participate in cell-cell interactions by binding to integrins (13,30). Mice lacking ADAM9 have no evident major abnormalities during development or adult life (24) but show redu...

β-Site Amyloid Precursor Protein (APP)-cleaving Enzyme 1 (BACE1)-deficient Mice Exhibit a Close Homolog of L1 (CHL1) Loss-of-function Phenotype Involving Axon Guidance Defects

“…3A). The broad spectrum matrix metalloprotease inhibitor GM6001 blocks ectodomain shedding of EphA4 (39), and ADAM8 and ADAM9 metalloproteases have been shown to process another member of the Eph receptor family, EphB4, to produce an NTF and a CTF (40,41). Therefore, we reasoned that the fragments we observed with EphA4 transfection alone were likely derived from ADAM (A Disinteg-rin and Metalloprotease) processing, in analogy to the ␣-secretase shedding of APP (42).…”

The Functional Maturation of A Disintegrin and Metalloproteinase (ADAM) 9, 10, and 17 Requires Processing at a Newly Identified Proprotein Convertase (PC) Cleavage Site

“…1), or the Glu Ͼ Ala mutant in the catalytic site as an inactive negative control. To monitor shedding, we co-transfected alkaline phosphatase (AP)-tagged cell surface substrates; TGF␣, an EGFR-ligand to monitor the activity of ADAM17 (28,34), BTC to study ADAM10 (28,35), or Ephrin receptor B4 (EphB4), which is a substrate for overexpressed ADAM9 (36). The cells were stimulated with either phorbol 12-myristate 13-acetate (PMA) to stimulate ADAM17, or ionomycin to enhance the activity of ADAM10 (37-39), or left unstimulated in experiments with overexpressed ADAM9.…”

Shedding of Collagen XVII/BP180 in Skin Depends on Both ADAM10 and ADAM9

“…Moreover the activity of ADAM10 can be up-regulated by calcium influx and potentially also other stimuli, whereas ADAM9-dependent collagen XVII shedding is increased in the presence of reactive oxygen species through transcriptional up-regulation of ADAM9 expression (50). Because reactive oxygen species are generated through insults to the skin, such as by infiltrating activated leukocytes during inflammation, by exposure to UV irradiation, or in cutaneous neoplasias, we propose that such insults affect the relative contribution of ADAMs 9 and 10 to collagen XVII shedding in healthy and diseased skin (51).…”