A Novel Pathogenic Pathway for Diabetic Keratopathy

项目摘要/摘要 糖尿病角膜病变是糖尿病的一种并发症，也是导致视力丧失的主要原因。没有 可以预防或逆转与糖尿病相关的角膜缺陷的有效药物。两个独立的纵向 临床研究表明，非诺贝特是一种特殊的过氧化物酶激动剂，具有很强的治疗效果。 增殖物激活受体α(PPARα)，在糖尿病视网膜病变中的作用。我们使用糖尿病患者的初步研究 人类供体角膜和动物模型表明，PPARα在维持角膜神经完整性方面发挥了作用。在……里面 我们的初步研究，我们已经建立了一个神经化的3D体外人类角膜模型，证明了 在解剖学和生理学上与活体角膜组织基本相似。使用这种新颖的模型，我们开始 揭示PPARα在糖尿病角膜病变中的S作用PPARα表达显著下调 来自T1 DM和T2 DM人类供体的细胞，与PPARα水平的降低一致，如 糖尿病人的角膜。我们在体内的初步研究表明，非糖尿病的PPARα基因敲除 (PPARα-/-)小鼠基础下神经纤维密度降低，角膜敏感度降低，类似 与糖尿病人身上看到的一样。此外，令我们惊讶的是，老年、非糖尿病的PPARα基因敲除小鼠 与年龄匹配的WT小鼠相比，自然发展成更严重的角膜溃疡。治疗 糖尿病大鼠服用非诺贝特的活性代谢物非诺贝酸减轻大鼠角膜神经变性 糖尿病大鼠。海马氏分析显示，PPARα-/-视网膜线粒体功能受损。基于 这些初步研究，我们假设糖尿病诱导的PPARα表达下调在 在糖尿病角膜病变中的关键病理作用，并代表了一个新的药物靶点。我们提出以下建议 针对这一假说的研究。首先，我们将在PPARα-/-小鼠和PPARα转基因小鼠中诱导糖尿病 在角膜中过表达PPARα，以确定PPARαKO是否恶化，而PPARα过表达 减轻糖尿病引起的角膜神经密度和敏感度的下降。我们还将治疗糖尿病小鼠 使用非诺贝特来确定激活PPARα是否可以阻止角膜神经纤维变性的进展。 第二，我们将确定PPARα的神经保护作用是否通过减弱氧化应激和 PPARα-/-在炎症、线粒体功能保护和神经营养因子上调中的作用 小鼠和PPARα转基因小鼠以及神经支配的体外3D人角膜模型。第三，到 将神经保护的PPARα功能转化为治疗，我们将评估外用的治疗效果 专利非诺贝特滴眼液在糖尿病神经纤维变性中的应用。这项研究有 鉴定PPARα在角膜中新功能的可能性。这些研究有可能建立一种新的 糖尿病角膜病变的发病机制和导致新的治疗方法的发展。

PROJECT SUMMARY / ABSTRACT Diabetic keratopathy is a complication of diabetes and a major cause of vision loss. There are no effective drugs that can prevent or reverse corneal defects related to diabetes. Two independent longitudinal clinical studies have shown robust therapeutic effects of fenofibrate, a specific agonist of Peroxisome Proliferator-Activated Receptor-α (PPARα), on diabetic retinopathy. Our preliminary studies using diabetic human donor corneas and animal models suggest a role of PPARα in maintaining corneal nerve integrity. In our preliminary studies, we have fabricated an innervated 3D in vitro human corneal model that demonstrates basic anatomical and physiological similarities to the corneal tissue in vivo. Using this novel model we began unravelling PPARα’s role in diabetic keratopathy. Significant downregulation of PPARα expression was seen in cells from both T1DM and T2DM human donors, in agreement with decreased PPARα levels as shown in diabetic human corneas. Our in vivo preliminary studies have shown that non-diabetic PPARα knockout (PPARα-/-) mice have decreased densities of the sub-basal nerve fibers and reduced corneal sensitivity, similar to what is seen in diabetic humans. Furthermore, to our surprise, aged, non-diabetic PPARα knockout mice naturally developed more severe corneal ulcerations compared to that in age-matched WT mice. Treatment of diabetic rats with fenofibric acid, an active metabolite of fenofibrate, alleviates corneal nerve degeneration in diabetic rats. As shown by Seahorse analysis, mitochondrial function is impaired in PPARα-/- retina. Based on these preliminary studies, we hypothesize that diabetes-induced down-regulation of PPARα expression plays a key pathological role in diabetic keratopathy and represents a novel drug target. We propose the following studies to address the hypothesis. First, we will induce diabetes in PPARα-/- mice and PPARα transgenic mice over-expressing PPARα in the cornea, to determine if PPARα KO exacerbates while PPARα over-expression alleviates diabetes-induced decreases of corneal nerve density and sensitivity. We will also treat diabetic mice with fenofibrate to determine if activation of PPARα arrests progression of corneal nerve fiber degeneration. Second, we will determine if the neuroprotective effect of PPARα is through attenuation of oxidative stress and inflammation, protection of mitochondrial functions and up-regulation of neurotrophic factors using PPARα-/- mice and PPARα transgenic mice as well as the innervated in vitro 3D human corneal model. Third, to translate the neuroprotective PPARα function into a therapy, we will evaluate therapeutic efficacy of topical application of a proprietary fenofibrate eyedrop on diabetes-induced nerve fiber degeneration. This study has potential to identify a new function of PPARα in the cornea. These studies have potential to establish a novel pathogenic mechanism for diabetic keratopathy and to lead to the development of a novel therapy.