Ceramide-mediated mitochondrial damage in diabetic retinopathy investigated by novel microfluidic O2 sensing and bio-mimetic electrochemistry

通过新型微流体 O2 传感和仿生电化学研究神经酰胺介导的糖尿病视网膜病变线粒体损伤

基本信息

批准号：
10132325
负责人：
Julia V Busik
金额：
$ 36.12万
依托单位：
MICHIGAN STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-04-01 至 2023-03-31
项目状态：
已结题

项目摘要

Despite recent success in treatment approaches, diabetic retinopathy (DR) remains a leading cause of progressive vision loss and blindness. Conceptual and technical breakthroughs to identify novel targets and strategies to cure this complication are paramount. We believe that such a breakthrough is offered by recent evidence of mitochondrial dysfunction in diabetic retinopathy in combination with the data from large clinical trials demonstrating a strong association between lipid abnormalities and DR progression, and the discovery that ceramide affects mitochondrial function. Mitochondria play cornerstone role in cellular metabolism and even slight modification of mitochondrial function can lead to pathology. Indeed, mitochondrial damage precedes histopathological abnormalities in DR. Recent studies demonstrate that there is an intricate connection between ceramide and mitochondrial function. Mitochondria have been shown to contain many sphingolipids including sphingomyelin and ceramide, as well as enzymes of sphingolipid pathway. Ceramide-induced restriction of respiratory chain function at the level of complex III, as well as succinate accumulation has been shown to be a causative factor in ischemia/reperfusion and stroke- induced tissue damage. In addition to effects on respiratory enzymes, ceramides were shown to contribute to mitochondrial outer membrane permeability either through S1P and hexadecenal production and activation of BAX/BAK, or directly through the formation of protein-permeable ceramide channels in mitochondrial outer membranes. These channels are shown to play a key role in the induction of apoptosis through the release of cytochrome c into the cytoplasm. We have previously demonstrated that activation of acid sphingomyelinase is an important early event in the pathogenesis of diabetic retinopathy. In this proposal we will test the overall hypothesis that increased levels of mitochondrial ceramide upon ASM activation leads to a) cytochrome c release and apoptosis and b) restriction on mitochondrial respiratory chain function in REC and RPE cells in diabetes. As traditional polarographic or fluorescence quenching (Seahorse) methods are not conducive for studies on limited amounts of available retinal tissue and cells, we are developing a novel microfluidic method for functional mitochondrial studies. We will utilize this novel methodology to assess the role of ASM activation and ceramide production in mitochondrial damage in diabetic retina.

尽管最近在治疗方法上取得了成功，但糖尿病性视网膜病（DR）仍然是渐进视力丧失和失明的主要原因。概念和技术突破确定新的目标和治愈这种并发症的策略至关重要。我们相信糖尿病中线粒体功能障碍的最新证据提供了这种突破视网膜病与大型临床试验的数据结合使用，证明很强脂质异常与DR进展之间的关联以及神经酰胺的发现影响线粒体功能。线粒体在细胞代谢中扮演基石的角色，甚至对线粒体功能可以导致病理。确实，线粒体损伤先于 DR的组织病理学异常。最近的研究表明，神经酰胺与线粒体功能。线粒体已显示包含许多鞘脂鞘胺和神经酰胺，以及鞘脂途径的酶。神经酰胺引起的在复合物III水平以及琥珀酸酯水平上限制呼吸链功能积累已被证明是缺血/再灌注和中风 - 诱导组织损伤。除了对呼吸酶的影响外，神经酰胺还显示出有助于线粒体外膜通透性要么通过S1P和六核生产和Bax/Bak的激活，或直接通过蛋白质可渗透神经酰胺的形成线粒体外膜中的通道。这些渠道被证明在通过将细胞色素C释放到细胞质中，凋亡诱导。我们有以前证明，酸鞘磷脂酶的激活是重要的早期事件糖尿病性视网膜病的发病机理。在此提案中，我们将检验总体假设 ASM激活时线粒体神经酰胺的水平增加导致A）细胞色素C释放和凋亡以及b）对线粒体呼吸链的限制在糖尿病中的REC和RPE细胞中功能。作为传统的光学或荧光淬火（海马）方法不利于对有限量的可用研究视网膜组织和细胞，我们正在开发一种新型的微流体方法来进行功能线粒体研究。我们将利用这种新颖的方法来评估ASM的作用糖尿病性视网膜线粒体损伤中的激活和神经酰胺的产生。