Inflammatory Chemokines: A Novel Target in Early Diabetic Retinopathy

描述(申请人提供)：糖尿病是21世纪的全球流行病，影响着全世界2.85亿人。糖尿病患者视力丧失的主要原因是黄斑水肿(DME)，其特征是血-视网膜屏障的改变。尽管使用了针对血管内皮生长因子(VEGF)的分子疗法，但大多数患者无法恢复良好的功能性视力，并出现残留的水肿，需要反复玻璃体内注射。科学上的差距是由于目前无法完全阻断与DME发病机制有关的所有炎症介质。有证据表明，糖尿病视网膜病变是一种炎症性疾病。趋化因子和白细胞启动炎症级联并最终破坏这一屏障的确切机制尚不清楚。我们的初步发现表明，CC趋化因子(CCL2、5和7)在糖尿病动物的视网膜中显著上调。此外，糖尿病时单核细胞进入视网膜的运输增加，视网膜小胶质细胞也被激活。使用CCL2基因缺陷小鼠的研究显示，糖尿病小鼠的视网膜血管渗漏明显减少，表明这种趋化因子在血-视网膜屏障的改变中发挥了重要作用。这些发现将趋化因子定义为糖尿病视网膜病变的一种新的特异性分子治疗靶点。我们的中心假设是糖尿病视网膜中炎性CC趋化因子的增加通过免疫细胞的募集改变了血-视网膜屏障。这一假设将在三个具体目标上得到检验。I)在这项研究中，我们将用细胞和分子方法研究CCL2及其受体在糖尿病动物视网膜和分离的视网膜细胞中的表达。我们将验证趋化因子和受体在人类供体眼中的表达，关联不同阶段人类糖尿病视网膜病变患者的血清趋化因子水平，并将趋化因子作为早期诊断的生物标志物。Ii)我们将确定CCL2和免疫细胞与血管生成素-2和蛋白水解酶等其他因素共同改变糖尿病血视网膜屏障的分子机制。Iii)我们将在糖尿病动物模型中进一步验证靶向CCL2通路作为一种治疗策略，以防止或逆转血视网膜屏障的改变。趋化因子途径在协调白细胞在糖尿病视网膜病变中的涌入最终导致黄斑水肿方面至关重要，是治疗这种毁灭性失明的良好未来靶点。这种方法是创新的，因为它解决了一种新的机制，稳定白细胞-内皮相互作用的初始步骤，这是糖尿病黄斑水肿的关键上游瓶颈。这项拟议的研究项目具有重要意义，因为它可以通过以下方式改变临床实践范式 介绍了一种以趋化因子为靶点的新疗法，以及一种潜在的用于DME患者早期识别的生物标志物。该项目的成功完成将对DME的临床试验产生很高的翻译影响，并可能导致与目前的激光和抗血管内皮生长因子疗法一起的“联合疗法”。

DESCRIPTION (provided by applicant): Diabetes, a global epidemic of the 21st century, affects 285 millions worldwide. The overwhelming cause of vision loss in diabetics is macular edema (DME) that is characterized by alteration of the blood-retinal barrier. Despite the use of molecular therapeutics targeting vascular endothelial growth factor (VEGF), the majority of patients do not recover good, functional vision, and show residual edema, necessitating repeated intravitreal injections. The scientific gap is due to the present inability to completely block all the inflammatory mediators implicated in the pathogenesis of DME. Evidence suggests that diabetic retinopathy is an inflammatory disease. The precise mechanism by which the chemokines and leukocytes initiate the inflammatory cascade and finally damages this barrier is not clear. Our preliminary findings show that CC chemokines (CCL2, 5 and 7) are significantly up regulated in the retinas of diabetic animals. Also, there is increased trafficking of monocytes into the retina along with activation of retinal microglia in diabetes. Studies using CCL2 gene- deficient mice made diabetic show significantly less retinal vascular leakage, indicating the important role of this chemokine in alteration of the blood-retinal barrier. These findings define chemokines as a novel specific molecular therapeutic target in diabetic retinopathy. Our central hypothesis is that increased inflammatory CC chemokines in the diabetic retina alter the blood-retinal barrier through the recruitment of immune cells. This hypothesis will be tested in three specific aims. i) In this proposal, we will characterize CCL2 and its receptor expression in the retina of diabetic animals, and in isolated retinal cells using cellular and molecular approaches. We will validate expression of chemokines and receptors in human donor eyes, correlate chemokine levels in serum of patients with different stages of human diabetic retinopathy and use chemokines as an early diagnostic bio-marker. ii) We will determine the molecular mechanisms by which the CCL2 and immune cells alter the blood-retinal barrier in diabetes in cooperation with other factors like angiopoietin-2 and proteinases. iii) We will further validate targeting of the CCL2 pathway as a therapeutic strategy in a diabetic animal model to prevent or reverse the alteration of the blood-retinal barrier. The chemokine pathway is critical in orchestrating the influx of leukocytes in diabetic retinopathy ultimately leading to macular edema, and is an excellent future target for the treatment of this devastating blindness. This approach is innovative as it addresses a novel mechanism of stabilizing the initial step of leukocyte-endothelial interaction, the critical upstream bottleneck for diabetic macular edema. This proposed research project is significant as it can transform the clinical practice paradigm by introducing a novel therapy using chemokines as targets, and a potential biomarker in early identification of patients with DME. Successful completion of this project will have the high translational impact for a clinical trial for DME, and potentially lead to a "combination therapy" along with the current laser and anti-VEGF therapies.