Role of Abnormal lysyl oxidase in the pathogenesis of diabetic retinopathy

赖氨酰氧化酶异常在糖尿病视网膜病变发病机制中的作用

• 批准号: 9043896
• 负责人: Sayon Roy
• 金额: $ 43.5万
• 依托单位: BOSTON MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-04-01 至 2020-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9043896
• 关键词: Affinity; Age; Apoptosis; Attenuated; Basement membrane; Binding; Biochemical; Biogenesis; Blindness; Blood Vessels; Blood capillaries; Cell Culture Techniques; Cell Death; Cell Proliferation; Cell Survival; Cell physiology; Cells; Characteristics; Clinical; Clinical Trials; Data; Development; Diabetes Mellitus; Diabetic Retinopathy; Disease; Endothelial Cells; Enzyme Precursors; Enzymes; Extracellular Matrix; Extravasation; FGF2 gene; Feedback; Fibronectins; Figs - dietary; Glucose; Goals; Health; Human; In Vitro; Incidence; Inhibition of Apoptosis; Knockout Mice; LOXL2 gene; Lead; Lesion; Literature; Maps; Mediating; Modality; Molecular Genetics; Mus; Pathogenesis; Pathway interactions; Patients; Pericytes; Permeability; Phosphotransferases; Population; Prevalence; Process; Protein Isoforms; Protein-Lysine 6-Oxidase; Proteins; Public Health; Research; Retina; Retinal; Role; Signal Transduction; Testing; Up-Regulation; Vascular Permeabilities; Work; animal facility; base; capillary; crosslink; diabetic; diabetic rat; extracellular; genetic approach; insight; monolayer; novel therapeutics; overexpression; prevent

 DESCRIPTION (provided by applicant): The overall goal of this project is to test the hypothesis that increased lysyl oxidase (LOX) expression and activation triggered by high glucose/diabetes compromises basement membrane (BM) function and thereby induces excess permeability and vascular cell death in diabetic retinopathy (DR). The hypothesis is based on findings that diabetes increases expression of LOX, which is required for the formation and maturation of the BM, and that HG-induced LOX upregulation and over-activation compromises endothelial barrier function. Importantly, inhibition of HG-induced LOX overexpression results in reduced cell monolayer permeability. While LOX is known for its cross-linking activity, LOX propeptide (LOX-PP), released during LOX biogenesis, is known for regulating cell proliferation and survival through modulation of Akt and Erk pathways. Our preliminary data indicates that HG-induced LOX overexpression promotes apoptosis in retinal endothelial cells, and that the process may be exacerbated by LOX-PP, which is upregulated by HG. Furthermore, preliminary data indicates that LOX binds with high affinity to extracellular fibronectin (FN) under HG condition, and interferes with LOX internalization into the cell, a process unique to LOX. Thus, the rationale for the proposed studies is that (i) HG-induced LOX overexpression contributes to capillary leakage, and together with increased LOX-PP promotes apoptosis by modulating Akt and Erk pathways, and (ii) HG-induced increased binding of LOX with ECM proteins, decreases LOX internalization, attenuates negative feedback signaling and thereby promotes LOX overexpression. The proposed studies will be performed using biochemical, molecular, and genetic approaches utilizing cell culture models, diabetic rats, and LOX heterozygous KO mice (LOX+/-). Towards this goal, we have generated LOX+/- mice colonies at our animal facility that are now available for the proposed studies. The specific aims of this proposal are to determine (i) whether inhibition of LOX overexpression and activation prevents apoptosis in retinal endothelial cells in vitro, and reduces vascular cell loss and permeability in diabetic rat retinas (ii) whether the decreased LOX level in LOX+/- mice is protective against diabetes-induced retinal vascular lesions, and (iii) mechanism(s) by which HG induces LOX overexpression and promotes retinal vascular cell loss. Findings from these studies are expected to identify a novel therapeutic strategy for preventing retinal vascular cell loss and capillary leakage, and provide mechanistic insights underlying HG-induced LOX overexpression in DR.

 描述（由申请人提供）：该项目的总体目标是检验以下假设：高血糖/糖尿病引发的赖氨酰氧化酶（LOX）表达和激活增加会损害基底膜（BM）功能，从而诱导糖尿病视网膜病变（DR）的通透性过高和血管细胞死亡。该假设基于以下发现：糖尿病会增加 LOX 的表达，这是 BM 形成和成熟所必需的，并且 HG 诱导的 LOX 上调和过度激活会损害内皮屏障功能。重要的是，抑制 HG 诱导的 LOX 过度表达会导致细胞单层通透性降低。 LOX 以其交联活性而闻名，而 LOX 生物发生过程中释放的 LOX 前肽 (LOX-PP) 则通过调节 Akt 和 Erk 途径来调节细胞增殖和存活。我们的初步数据表明，HG 诱导的 LOX 过度表达促进视网膜内皮细胞凋亡，并且该过程可能会因 HG 上调的 LOX-PP 而加剧。此外，初步数据表明，在 HG 条件下，LOX 以高亲和力与细胞外纤连蛋白 (FN) 结合，并干扰 LOX 内化到细胞中，这是 LOX 独有的过程。因此，本研究的基本原理是：(i) HG 诱导的 LOX 过度表达会导致毛细血管渗漏，并与增加的 LOX-PP 一起通过调节 Akt 和 Erk 途径促进细胞凋亡；(ii) HG 诱导的 LOX 与 ECM 蛋白的结合增加，减少 LOX 内化，减弱负反馈信号，从而促进 LOX 过度表达。拟议的研究将使用生物化学、分子和遗传方法，利用细胞培养模型、糖尿病大鼠和 LOX 杂合 KO 小鼠 (LOX+/-) 进行。为了实现这一目标，我们在我们的动物设施中生成了 LOX+/- 小鼠群体，现在可用于拟议的研究。该提案的具体目的是确定（i）抑制LOX过表达和激活是否可以在体外防止视网膜内皮细胞凋亡，并减少糖尿病大鼠视网膜的血管细胞损失和通透性（ii）LOX+/-小鼠中LOX水平降低是否可以预防糖尿病引起的视网膜血管病变，以及（iii）机制 HG 诱导 LOX 过度表达并促进视网膜血管细胞损失。这些研究的结果有望确定一种预防视网膜血管细胞丢失和毛细血管渗漏的新治疗策略，并提供 DR 中 HG 诱导的 LOX 过度表达的机制见解。