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）：本项目的总体目标是检验以下假设：由高糖/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）中的过度通透性和血管细胞死亡。该假设基于以下发现：糖尿病增加了LOX的表达，这是BM形成和成熟所必需的，并且HG诱导的LOX上调和过度活化损害了内皮屏障功能。重要的是，抑制HG诱导的LOX过表达导致细胞单层通透性降低。虽然LOX因其交联活性而为人所知，但在LOX生物发生期间释放的LOX前肽（LOX-PP）已知用于通过调节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过表达促进视网膜内皮细胞的凋亡，并且该过程可能被HG上调的LOX-PP加剧。此外，初步数据表明，LOX在HG条件下以高亲和力结合细胞外纤连蛋白（FN），并干扰LOX内化到细胞中，这是LOX特有的过程。因此，所提出的研究的基本原理是（i）HG诱导的LOX过表达有助于毛细血管渗漏，并且与增加的LOX-PP一起通过调节Akt和Erk途径促进细胞凋亡，以及（ii）HG诱导的LOX与ECM蛋白的结合增加，降低LOX内化，减弱负反馈信号传导，从而促进LOX过表达。将使用细胞培养模型、糖尿病大鼠和LOX杂合KO小鼠（LOX+/-），采用生物化学、分子和遗传学方法进行拟定研究。为了实现这一目标，我们已经在我们的动物设施中产生了LOX+/-小鼠集落，这些集落现在可用于所提出的研究。该建议的具体目的是确定（i）LOX过表达和活化的抑制是否在体外防止视网膜内皮细胞的凋亡，并减少糖尿病大鼠视网膜中的血管细胞损失和渗透性（ii）LOX+/-小鼠中降低的LOX水平是否保护免受糖尿病诱导的视网膜血管损伤，和（iii）HG诱导LOX过表达和促进视网膜血管细胞损失的机制。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LOX过表达的机制见解。