Retinal neovascularization is a leading cause of human blindness. However, little is known concerning the molecular mechanisms controlling retinal neovascularization in vivo. Here we provide evidence that exposure of a collagen type IV cryptic epitope detected by monoclonal antibody (mAb) HUIV26, delineates sites of vascular bud formation and represents one of the earliest structural remodeling events required before vessel out-growth. Exposure of these cryptic sites was inhibited in matrix metalloproteinase (MMP)-9-deficient but not MMP-2-deficient mice implicating MMP-9 in their exposure. Retinal endothelial cell interactions with the HUIV26 epitopes induced endothelial cell migration, which was blocked by mAb HUIV26. Importantly, subcutaneous administration of mAb HUIV26 potently inhibited retinal angiogenesis in vivo. Taken together, these findings suggest a novel mechanism in which MMP-9 facilitates exposure of HUIV26 cryptic sites, thereby promoting retinal endothelial cell migration and neovascularization in vivo.
We previously documented protein kinase CK2 involvement in retinal neovascularization. Here we describe retinal CK2 expression and combined effects of CK2 inhibitors with the somatostatin analog octreotide in a mouse model of oxygen-induced retinopathy (OIR). CK2 expression in human and rodent retinas with and without retinopathy and in astrocytic and endothelial cultures was examined by immunohistochemistry, Western blotting, and reverse transcriptase-polymerase chain reaction. A combination of CK2 inhibitors, emodin or 4,5,6,7-tetrabromobenzotriazole, with octreotide was injected intraperitoneally from postnatal (P) day P11 to P17 to block mouse OIR. All CK2 subunits (alpha, alpha', beta) were expressed in retina, and a novel CK2alpha splice variant was detected by reverse transcriptase-polymerase chain reaction. CK2 antibodies primarily reacted with retinal astrocytes, and staining was increased around new intraretinal vessels in mouse OIR and rat retinopathy of prematurity, whereas preretinal vessels were negative. Cultured astrocytes showed increased perinuclear CK2 staining compared to endothelial cells. In the OIR model, CK2 mRNA expression increased modestly on P13 but not on P17. Octreotide combined with emodin or 4,5,6,7-tetrabromobenzotriazole blocked mouse retinal neovascularization more efficiently than either compound alone. Based on its retinal localization, CK2 may be considered a new immunohistochemical astrocytic marker, and combination of CK2 inhibitors and octreotide may be a promising future treatment for proliferative retinopathies.
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