Department of Clinical Neurosciences, University of Cambridge, Cambridge CB2 0PY, UK.
Brain. 2010 Feb;133(Pt 2):448-64. doi: 10.1093/brain/awp319.
Retinal pigment epithelial cell malfunction is a causative feature of age-related macular degeneration, and transplantation of new retinal pigment epithelial cells is an attractive strategy to prevent further progression and visual loss. However, transplants have shown limited efficacy, mainly because transplanted cells fail to adhere and migrate onto pathological Bruch's membrane. Adhesion to Bruch's membrane is integrin-mediated. Ageing of Bruch's membrane leads to a decline in integrin ligands and, added to this, wet age-related macular degeneration leads to upregulation of anti-adhesive molecules such as tenascin-C. We have therefore investigated whether manipulation of integrin function in retinal pigment epithelial cells can restore their adhesion and migration on wet age-related macular degeneration-damaged Bruch's membrane. Using spontaneously immortalized human retinal pigment epithelial cells (adult retinal pigment epithelium-19), we show that adhesion and migration on the Bruch's membrane components is integrin-dependent and enhanced by integrin-activating agents manganese and TS2/16. These allowed cells to adhere and migrate on low concentrations of ligand, as would be found in aged Bruch's membrane. We next developed a method for stripping cells from Bruch's membrane so that adhesion and migration assays can be performed on its surface. Integrin activation had a moderate effect on enhancing retinal pigmented epithelial cell adhesion and migration on normal human and rat Bruch's membrane. However, on Bruch's membrane prepared from human wet age-related macular degeneration-affected eyes, adhesion was lower and integrin activation had a much greater effect. A candidate molecule for preventing retinal pigmented epithelial interaction with age-related macular degeneration-affected Bruch's membrane is tenascin-C which we confirm is present at high levels in wet age-related macular degeneration membrane. We show that tenascin-C is anti-adhesive for retinal pigmented epithelial cells, but after integrin activation, they can adhere and migrate on it using alphaVbeta3 integrin. Alternatively, we find that transduction of retinal pigmented epithelial cells with alpha9 integrin, a tenascin-C-binding integrin, led to a large increase in alpha9beta1-mediated adhesion and migration on tenascin-C. Both expression of alpha9 integrin and integrin activation greatly enhanced the ability of retinal pigment epithelial cells to adhere to tenascin-rich wet age-related macular degeneration-affected Bruch's membranes. Our results suggest that manipulation of retinal pigment epithelial cell integrins through integrin activating strategies, or expression of new integrins such as alpha9, could be effective in improving the efficacy of retinal pigment epithelial cell transplantation in wet age-related macular degeneration-affected eyes.
视网膜色素上皮细胞功能障碍是年龄相关性黄斑变性的一个致病特征，而移植新的视网膜色素上皮细胞是一种防止疾病进一步进展和视力丧失的有吸引力的策略。然而，移植的效果有限，主要是因为移植的细胞不能附着并迁移到病理性的布鲁赫膜上。对布鲁赫膜的附着是整合素介导的。布鲁赫膜的老化导致整合素配体的减少，此外，湿性年龄相关性黄斑变性导致抗附着分子如 tenascin-C 的上调。因此，我们研究了是否可以通过操纵视网膜色素上皮细胞中的整合素功能来恢复它们在湿性年龄相关性黄斑变性损伤的布鲁赫膜上的粘附和迁移。我们使用自发永生化的人视网膜色素上皮细胞（成人视网膜色素上皮细胞-19），证明在布鲁赫膜成分上的粘附和迁移是整合素依赖性的，并被整合素激活剂锰和 TS2/16 增强。这些允许细胞在低浓度的配体上附着和迁移，就像在老化的布鲁赫膜中发现的那样。接下来，我们开发了一种从布鲁赫膜上剥离细胞的方法，以便可以在其表面进行粘附和迁移实验。整合素激活对增强正常人和大鼠布鲁赫膜上的视网膜色素上皮细胞粘附和迁移有适度的影响。然而，在来自湿性年龄相关性黄斑变性患者的布鲁赫膜上，粘附力较低，整合素激活的影响更大。一种可能的预防视网膜色素上皮细胞与年龄相关性黄斑变性相关的布鲁赫膜相互作用的候选分子是 tenascin-C，我们证实其在湿性年龄相关性黄斑变性膜中高表达。我们表明 tenascin-C 对视网膜色素上皮细胞具有抗粘附性，但在整合素激活后，它们可以使用 alphaVbeta3 整合素附着和迁移。或者，我们发现用 tenascin-C 结合整合素 alpha9 转导视网膜色素上皮细胞，可大大增加 alpha9beta1 介导的 tenascin-C 上的粘附和迁移。alpha9 整合素的表达和整合素激活都极大地增强了视网膜色素上皮细胞在富含 tenascin-C 的湿性年龄相关性黄斑变性相关布鲁赫膜上的粘附能力。我们的结果表明，通过整合素激活策略操纵视网膜色素上皮细胞整合素，或表达新的整合素，如 alpha9，可能会有效提高湿性年龄相关性黄斑变性患者视网膜色素上皮细胞移植的效果。
