Role of the kallikrein-kinin system in diabetic retinopathy

描述(申请人提供)：糖尿病视网膜病变是发达国家视力丧失的主要原因。糖尿病视网膜病变进展到威胁视力的阶段，包括糖尿病黄斑水肿(DME)，通常与潜在的视网膜血管功能障碍和疾病的恶化有关，包括视网膜出血的数量和严重程度的增加。近年来的临床研究表明，玻璃体腔内注射抗血管内皮生长因子治疗DME能有效提高视力和缓解黄斑水肿，FDA最近批准玻璃体腔内注射雷尼比单抗治疗DME。虽然这种治疗对许多DME患者非常有效，但几项大型临床研究显示，约40%至50%的研究对象在视力改善和视网膜增厚的解决方案中出现屈光不正或对抗血管内皮生长因子指导的治疗没有完全反应。这些临床发现表明，血管内皮生长因子非依赖性机制也可能是导致大量DME患者视网膜水肿的原因之一。在最近的报道和更多的初步数据中，我们发现DME患者玻璃体中血浆激肽释放酶(Pkal)、FXII因子和激肽原水平升高。我们还发现玻璃体中的Pkal水平与血管内皮生长因子水平没有相关性，这揭示了DME患者中分子的异质性的一个组成部分。利用基因敲除和药理学方法，我们发现Pkal促进了啮齿动物视网膜血管的高通透性和视网膜增厚，并且我们已经开始描述促成这种反应的缓激肽依赖和非独立的机制。我们的研究表明，糖尿病增加了Pkal对血管功能障碍和视网膜水肿的影响(Clermont等人糖尿病2011年，Liu等人2011年自然医学)。我们最近还发现，自体血液进入玻璃体会增加视网膜炎症和视网膜血管通透性(Liu等人IOVS 2013)，而且激肽释放酶系统在诱导视网膜增厚方面与血管内皮生长因子一样有效。此外，我们还利用蛋白质组学研究了缓激肽诱导的视网膜水肿，并鉴定出一组血浆蛋白在视网膜水肿时显著增加，同时视网膜中间丝蛋白的一个子集显著减少。这项资助将确定PK诱导的导致视网膜血管高通透性和视网膜增厚的机制，并检验Pkal-Bradykinin系统是导致DME的视网膜功能障碍的不依赖于VEGF的介质的假设。其具体目的是：1)研究和比较缓激肽受体依赖和独立的Pkal在糖尿病小鼠视网膜水肿中的作用；2)比较Pkal和血管内皮生长因子诱导的视网膜水肿，并探讨这两种途径之间的相互作用；3)研究一氧化氮合酶和钙蛋白酶介导的中间丝降解/重塑在视网膜水肿中的作用。进一步确定视网膜中血浆激肽释放酶系统的作用和调节机制将有助于 评估靶向PKAL系统治疗DME的潜在临床机会。

DESCRIPTION (provided by applicant): Diabetic retinopathy is a leading cause of vision loss in developed countries. Progression to sight threatening stages of diabetic retinopathy, including diabetic macular edema (DME), is usually associated with worsening of underlying retinal vascular dysfunction and disease, including an increase in the number and severity of retinal hemorrhages. Recent clinical advances have demonstrated that intravitreal injection of anti-VEGF directed therapies are effective in improving visual acuity and resolving macular edema in DME, and the FDA has recently approved intravitreal injection of ranibizumab for DME. While this treatment is highly effective for many patients with DME, several large clinical studies have revealed that about 40 to 50% of study subjects appear refractive or do not fully respond in the improvement of visual acuity and the resolution of retinal thickening to anti-VEGF directed therapies. These clinical findings suggest that VEGF-independent mechanisms might also contribute to retinal edema in a large number of DME patients. In recent reports and additional preliminary data, we show that plasma kallikrein (Pkal), factor FXII, and kininogen are increased in the vitreous of patients with DME. We also show that Pkal levels in the vitreous do not correlate with VEGF levels, revealing a component of molecular heterogeneity among DME patients. Using both knockout and pharmacological approaches, we show that Pkal contributes to retinal vascular hyperpermeability and retinal thickening in rodents and we have begun to characterize both bradykinin-dependent and -independent mechanisms that contribute to this response. Our studies have revealed that diabetes increases the effects of Pkal on vascular dysfunction and retinal edema (Clermont et al Diabetes 2011, Liu et al Nature Medicine 2011). We have also recently shown that autologous blood introduced into the vitreous increases retinal inflammation and retinal vascular permeability (Liu et al IOVS 2013) and the kallikrein system is as potent as VEGF in inducing retinal thickening. In addition we have used proteomics to characterize bradykinin-induced retinal edema and have identified a set of plasma proteins that are robustly increased in retinal edema coupled with striking decreases in a subset of retinal intermediate filament proteins. This grant will identify the PK- induced mechanisms that cause retinal vascular hyperpermeability and retinal thickening, and examine the hypothesis that the Pkal-bradykinin system is a VEGF-independent mediator of retinal dysfunction that contributes to DME. The specific aims are 1) to characterize and compare the bradykinin receptor-dependent and independent effects of Pkal on retinal edema in diabetic mice, 2) to compare Pkal and VEGF-induced retinal edema and investigate interactions between these pathways, and 3) characterize the roles of nitric oxide synthase and calpain-mediated intermediate filament degradation/remodeling in retina edema. Further identification of the mechanism of action and regulation of the plasma kallikrein system in the retina will be helpful in evaluating potential clinical opportunities for targeting the Pkal system for the treatment of DME.