The Role of O-GlcNAcylation in Diabetic Nephropathy

 描述(申请人提供)：糖尿病肾病是慢性肾脏疾病的主要原因，也是糖尿病患者发病率和死亡率方面最重要的长期并发症之一。几个超微结构的变化发生在 糖尿病患者的肾小球，如肾小球高滤过、因细胞外基质成分积聚而导致的系膜扩张，以及足细胞凋亡。虽然糖尿病肾病发生的具体机制尚不清楚，但高血糖被认为是该病发生和发展的重要因素。研究表明，葡萄糖代谢的变化通过氨基己糖生物合成途径刺激通量，从而启动超微结构的变化。氨基己糖生物合成途径的主要终点是形成UDP-GlcNAc，它是O-GlcNAc转移酶(OGT)和糖胺聚糖(GAG)合成的底物。硫酸乙酰肝素蛋白多糖(HSPGs)是肾小球基底膜(GBM)的一部分，被认为在维持电荷选择性滤过屏障和GBM成分的组织维持中起重要作用。足细胞被认为与基底膜一起工作，起到阻止蛋白质丢失的动态屏障的作用。最近，基底膜蛋白多糖的丢失被认为是导致蛋白尿的原因，目前的研究集中在足细胞作为代谢环境在糖尿病蛋白尿发生发展中的作用的中心靶点。已有研究表明，一些蛋白质，如足细胞细胞骨架蛋白α-肌动蛋白4[22]，以及转录因子，如Sp1和FOXO-1，都是O-GlcN酰化的。这些修饰可能会改变蛋白质的功能和启动子的特异性。因此，我们试图通过以下方法来表征O-GlcNAc在糖尿病相关肾脏疾病中的直接作用：1)利用qPCR阵列阐明O-GlcN酰化对ECM蛋白和蛋白多糖(PG)基因表达的影响，2)检测O-GlcN酰化是否通过改变高血糖环境中HSPGs的表达、生物合成或裂解而促进超滤，以及3)在他莫昔芬诱导的PODOCKO的2型糖尿病转基因模型中确定O-GlcN酰化在肾脏疾病的肾脏生物学中的作用。

 DESCRIPTION (provided by applicant): Diabetic nephropathy (DN) is the leading cause of chronic kidney disease and one of the most significant long-term complications in terms of morbidity and mortality for individuals with diabetes. Several ultrastructural changes occur in the glomeruli of a diabetic patient, such as glomerular hyperfiltration, mesangial expansion as a result of accumulation of the extracellular matrix components, and podocyte apoptosis. While the specific mechanisms underlying the development of DN remain unknown, hyperglycemia is considered an important contributing factor in the development and the progression of this disease. Changes in glucose metabolism have been shown to initiate ultrastructural changes by stimulating flux through the hexosamine biosynthetic pathway. The major endpoint for the hexosamine biosynthetic pathway is the formation of UDP-GlcNAc, which is a substrate for O-GlcNAc transferase (OGT) and glycosaminoglycan (GAG) synthesis. Heparan sulfate proteoglycans (HSPGs), part of the glomerular basement membrane (GBM), are thought to have an important role in the maintenance of the charge selective filtration barrier and the organizational maintenance of the GBM constituents. Podocytes are thought to work together with the GBM to function as a dynamic barrier to protein loss. The loss of proteoglycans in the GBM has been recently questioned as causative of the proteinuria and current research has focused on the podocyte as a central target for the effects of the metabolic milieu in the development and progression of diabetic proteinuria. It has been shown that several proteins, such as podocyte cytoskeletal protein a-actinin 4 [22], and transcription factors, such Sp1 and FOXO-1, are O-GlcNAcylated. These modifications may alter the function of proteins and the promoter specificity. Thus, we sought to characterize the direct roles of O-GlcNAc in kidney disease associated with diabetes by: 1) elucidating effects of O-GlcNAcylation on ECM protein and proteoglycan (PG) gene expression utilizing qPCR arrays, 2) examining whether O-GlcNAcylation contributes to hyperfiltration by altering the expression, biosynthesis, or cleavage of HSPGs in a hyperglycemic environment, and 3) defining the role of O-GlcNAcylation in the nephrobiology of renal disorders in a type-2 diabetes transgenic model of tamoxifen-inducible podocyte specific knockout of O-GlcNAc transferase (OGT), leading to specific OGT KO in podocytes.