Study of the ER-mitochondria interface as a new target in diabetic retinopathy

描述（申请人提供）：内质网（ER）和线粒体是调节细胞代谢的主要隔室。令人信服的证据表明，这两个细胞器密切相互作用和协调，以维持细胞的代谢完整性。因此，内质网稳态紊乱（或内质网应激）和线粒体功能障碍通常在神经退行性疾病和糖尿病等病理条件下共存。在糖尿病视网膜病变（DR）中，高血糖足以诱导内质网应激和氧化应激，两者都导致视网膜炎症、血管渗漏、细胞凋亡，最终导致新生血管和神经元变性。有趣的是，除了功能上的相互作用，内质网和线粒体通过内质网高度特化的子结构域线粒体相关内质网膜（MAM）在物理和生化上相互连接。尽管MAM的确切作用机制尚不清楚，但新出现的证据表明，MAM是脂质和蛋白质代谢以及钙信号传导的关键位点。几项研究

DESCRIPTION (provided by applicant): The endoplasmic reticulum (ER) and mitochondria are the primary compartments that regulate cellular metabolism. Compelling evidence suggests that these two organelles closely interact and coordinate to maintain the cell's metabolic integrity. Accordingly, disturbed ER homeostasis (or ER stress) and mitochondrial dysfunction often co-exist in pathological conditions such as neurodegenerative diseases and diabetes. In diabetic retinopathy (DR), hyperglycemia is sufficient to induce ER stress and oxidative stress, both contributing to retinal inflammation, vascular leakage, apoptosis, and ultimate neovascularization and neuronal degeneration. Intriguingly, additional to functional interactions, the ER and mitochondria are physically and biochemically interconnect through a highly specialized subdomain of the ER named the mitochondria- associated ER membrane (MAM). Although the exact mechanism of action of MAM remains obscure, emerging evidence suggests that MAM is a critical site for lipid and protein metabolism and calcium signaling. Several studies show that defective MAM function and/or structure negatively affects mitochondrial ATP production, increases ROS generation, exacerbates ER stress and leads to apoptosis. However, the role of MAM in healthy and diseased retina and retinal cells has not been studied. The overall goal of this pilot study is to establish a role of MAM in retinal cell metabolism in diabetes. In Aim 1, we will use innovative approaches to characterize the structural and biochemical changes in MAM in retinal cells exposed to diabetic insults and in the retina from diabetic animals. We will determine the functional consequence of MAM alterations on oxidative stress, ER stress, inflammation, vascular injury, and retinal cell death in DR. In Aim 2, we will use a specifically refined state-of-the-art proteomic technology to analyze the protein profile of MAM from normal and diabetic retinas. This comprehensive and unbiased assay of MAM proteins will confirm the role of MAM in regulation of retinal metabolism and explore the functional implication of MAM in signaling pathways related to DR pathogenesis. We anticipate that this novel and exciting project will generate essential data for future mechanistic study on mitochondria-ER regulation, which may also fill in a gap in our understanding of diabetes- induced metabolic defects in retinal cells and identify new therapeutic targets for DR.