巨噬细胞源性乙酰肝素酶在晚期糖基化终产物加重肾小球内皮细胞功能障碍中的作用和机制研究

Minimal expression of heparanase(HPA) was noted in normal glomeruli. Both of marked increased HPA expression and infiltration of macrophages, an HPA-rich cell, makes glomerular endothelial cell dysfunction in diabetic glomeruli. Our study shows that advanced glycation end products (AGEs) induced macrophage-derived HPA expression to damage the adhesion and permeability of glomerular endothelial cells, suggesting that macrophage-derived HPA may be the main source of increased HPA in diabetic glomeruli and the key of glomerular endothelial cell dysfunction induced by AGEs. In this study, we will investigate the role of macrophage-derived HPA in glomerular HPA increased expression using diabetic rat which macrophage function will be cleared by liposomal clodronate. Furthermore, we will investigate the change of glomerular endothelial cell dysfunction and macrophage-derived HPA in glomeruli of diabetic rats pretreated with AGEs and liposomal clodronate. Meanwhile, glomerular endothelial cells are cultured with the supernatant from macrophage pretreated with HPA SiRNA and AGEs to investigate the effect and mechanism of the glomerular endothelial cell damage induced by macrophage-derived HPA activity and signal transduction to provide a new idea and target for treatment of glomerular endothelial cell dysfunction in diabetes.

糖尿病时，正常肾小球中仅为微量的乙酰肝素酶（HPA）高表达及自身高丰度HPA的巨噬细胞在肾小球浸润聚集均导致肾小球内皮细胞功能障碍。我们研究表明，晚期糖基化终产物（AGEs）诱导巨噬细胞源性HPA高表达从而损害肾小球内皮细胞黏附功能和通透性，提示巨噬细胞源性HPA很可能是糖尿病肾小球高表达HPA的主要来源，是AGEs加重糖尿病肾小球内皮细胞功能障碍的关键环节。本研究拟利用氯膦酸二钠脂质体去除巨噬细胞功能的糖尿病大鼠，证实巨噬细胞源性HPA在糖尿病肾小球HPA高表达来源中的地位；并以AGEs干预，观察肾小球巨噬细胞源性HPA表达和肾小球内皮细胞功能的变化；在体外，以AGEs干预HPA SiRNA预处理的巨噬细胞，取上清液作用于肾小球内皮细胞，从HPA酶活性和信号转导功能两方面探讨巨噬细胞源性HPA损害肾小球内皮细胞功能的作用和机制，旨在为糖尿病肾小球内皮细胞功能障碍的治疗提供新的思路和靶点。

糖尿病肾病已经成为我国新进入血液透析患者的主要病因。晚期糖基化终产物（AGEs）积聚作为糖尿病肾病的重要致病因素，损伤肾小球内皮细胞，导致尿白蛋白增多、肾功能恶化。同时，肾小球乙酰肝素酶（HPA）表达增高和巨噬细胞聚集加重了糖尿病肾小球内皮细胞功能障碍。但是巨噬细胞源性乙酰肝素酶在AGEs加重肾小球内皮细胞功能障碍中的作用和机制尚不清楚。因此，我们建立了氯膦酸二钠脂质体剔除巨噬细胞的糖尿病鼠模型，观察其对肾小球乙酰肝素酶表达和尿蛋白的影响；同时，在体外，以AGEs干预巨噬细胞，并作用于肾小球内皮细胞，研究了巨噬细胞源性乙酰肝素酶损害肾小球内皮细胞功能的作用和机制。结果表明，AGEs促使巨噬细胞源性HPA表达显著增高，通过乙酰肝素酶的酶活性和信号转导作用导致肾小球内皮细胞功能障碍，加重糖尿病蛋白尿、损害肾脏。因此，巨噬细胞源性乙酰肝素酶异常表达很可能是AGEs加重糖尿病肾小球内皮细胞功能障碍的重要环节。我们的研究为糖尿病肾小球内皮细胞功能障碍的治疗提供了新的思路。