Jo Nobuo, Ju Meihua, Nishijima Kazuaki, Robinson Gregory S, Adamis Anthony P, Shima David T, Mailhos Carolina
OSI Eyetech, Eyetech Research Center, Lexington, MA 02421, USA.
Mol Vis. 2006 Oct 26;12:1243-9.
The wet form of age-related macular degeneration (AMD) occurs as a consequence of abnormal blood vessel growth from the choroid into the retina. Pathological angiogenesis during tumor growth and ocular disease has been associated with specific exposure of cryptic extracellular matrix epitopes. We investigated the presence of cryptic collagen IV epitopes in a murine model of choroidal neovascularization (CNV), and tested the effect on blood vessel growth of H8, a humanized antibody directed against a cryptic collagen type IV epitope.
To induce experimental CNV in adult C57BL/6 mice, Bruch's membrane was ruptured using a diode laser. Subsequently, mice were treated with daily intraperitoneal (i.p.) injections of either H8 (10 mg/kg or 30 mg/kg) or an isotype-matched antibody control. Two weeks postinjection, choroidal flat mounts were immunostained with the blood vessel marker platelet/endothelial cell adhesion molecule-1 (PECAM-1) and H8. CNV was visualized using fluorescence microscopy and the CNV lesion area measured using Open Lab software.
Collagen type IV and the cryptic epitope were observed at the site of laser-induced lesions. Staining with H8 was first observed three days post injury, two days after MMP2 expression in CNV lesions, becoming most intense five days following laser injury and extending beyond the area of neovascularization. At 14 days post injury, H8 staining was reduced in intensity, colocalized with the area of CNV, and was nearly absent from the underlying choroidal vessels. In addition, mice treated with H8 had a significant dose-dependent decrease in the area of CNV as compared to isotype-matched antibody controls.
Results suggest that exposure of cryptic collagen type IV epitopes is associated with the incidence of CNV and that the humanized antibody H8 may provide a new treatment for CNV.
湿性年龄相关性黄斑变性（AMD）是脉络膜血管异常生长至视网膜的结果。肿瘤生长和眼部疾病中的病理性血管生成与隐蔽的细胞外基质表位的特定暴露有关。我们在脉络膜新生血管（CNV）小鼠模型中研究了隐蔽的IV型胶原表位的存在，并测试了针对隐蔽的IV型胶原表位的人源化抗体H8对血管生长的影响。
为在成年C57BL/6小鼠中诱导实验性CNV，使用二极管激光破坏布鲁赫膜。随后，小鼠每天腹腔注射H8（10mg/kg或30mg/kg）或同型匹配的抗体对照。注射后两周，用血管标记物血小板/内皮细胞黏附分子-1（PECAM-1）和H8对脉络膜扁平标本进行免疫染色。使用荧光显微镜观察CNV，并使用Open Lab软件测量CNV病变面积。
在激光诱导损伤部位观察到IV型胶原和隐蔽表位。损伤后三天首次观察到H8染色，在CNV病变中MMP2表达两天后出现，激光损伤五天后最为强烈，并延伸至新生血管区域之外。损伤后14天，H8染色强度降低，与CNV区域共定位，在下方脉络膜血管中几乎不存在。此外，与同型匹配的抗体对照相比，用H8治疗的小鼠CNV面积有显著的剂量依赖性减少。
结果表明，隐蔽的IV型胶原表位的暴露与CNV的发生有关，人源化抗体H8可能为CNV提供一种新的治疗方法。
