PATHOLOGY OF BASEMENT MEMBRANE COMPONENTS IN DIABETES

糖尿病具有严重的微血管并发症，导致多系统 基底膜和细胞外基质异常。 糖尿病肾病多表现为肾小球基底膜病变 膜（GBM）功能异常，系膜扩张， 存在GBM的进行性增厚。 GBM和系膜基质是 由IV、V型胶原蛋白、蛋白多糖、纤连蛋白（FN）组成， 层粘连蛋白（LMN）和巢蛋白等。 在众多的分子 糖尿病中肾小球和血浆的异常是：1）增加 系膜中的FN，2）硫酸乙酰肝素蛋白聚糖减少，3） IV型胶原蛋白的非酶糖化增加，和4）增加 FN的非酶糖化。 FN或胶原蛋白的非酶糖化 降低了这些分子的结合亲合力。 的结合 纤连蛋白和胶原蛋白（明胶或IV型）或层粘连蛋白和IV型 胶原，导致3 H-肝素结合的正协同性。 FN、LMN或胶原蛋白的非酶糖化导致严重的 肝素结合的正协同性降低。 糖基化 层粘连蛋白还直接减少与其结合的肝素。 拟议的研究将进行正常和 非酶糖化FN、LMN、肝素、硫酸乙酰肝素蛋白聚糖 来源于肾小球和EHS肿瘤，天然胶原蛋白（IV、I和III）， Clq和纤维蛋白。 我们将定义结合IV型FN的结构域 胶原蛋白，并进一步定义胶原蛋白I（明胶）结合域。 这 将涉及蛋白水解和化学切割以及各种 纤连蛋白的结构域。 我们接下来将决定 放射性标记的FN和非酶糖化的FN， 给药正常和糖尿病动物。 接下来，我们将确定 肾小球中纤维连接蛋白的积累或周转增加， 来自正常和糖尿病动物的培养物。 最后，我们将分离血浆 从控制不佳的糖尿病患者中提取纤连蛋白，以确定其是否 显示出与上述相同的异常。 统称 研究将使我们能够研究可能是一个关键的生化 糖尿病GBM和系膜的异常，即 相关分子的分子缔合。 血浆评价 这种蛋白质可以作为微血管病变的指标， 在糖尿病中发生的肾病变化是非常有吸引力的。 它 似乎很清楚，这些研究也可能揭示的过程， 控制着基底成分的合成、转换和聚集。

Diabetes has profound microvascular complications which lead to multisystem abnormalities involving the basement membrane and extracellular matrix. The diabetic kidney manifests much pathology where: glomerular basement membrane (GBM) function is abnormal, the mesangium becomes expanded and there is a progressive thickening of GBM. The GBM and mesangial matrix are composed of: type IV, V collagens, proteoglycans, fibronectin (FN), laminin (LMN), and entactin, among other things. Among numerous molecular abnormalities in the glomerulus and plasma in diabetes are: 1) increased FN in the mesangium, 2) decreased heparan sulfate proteoglycan, 3) increased nonenzymatic glycation of type IV collagen, and 4) increased nonenzymatic glycation of FN. Nonenzymatic glycation of FN or collagen decreases the binding avidity of these molecules. The binding of fibronectin and collagen (gelatin or type IV) or laminin and type IV collagen, leads to positive cooperativity for 3H-heparin binding. Nonenzymatic glycation of FN, LMN or collagen(s) leads to a profound decrease in positive cooperativity for heparin binding. Glycation of laminin also produces a direct decrease in heparin binding to it. The proposed studies will perform binding assays with normal and nonenzymatically glycated FN, LMN, heparin, heparan sulfate proteoglycan derived from glomeruli and the EHS tumor, native collagens (IV, I and III), Clq and fibrin. We will define the domain of FN that binds type IV collagen and further define the collagen I (gelatin) binding domain. This will involve proteolytic and chemical cleavage and purification of various domains of fibronectin. We will next determine the deposition and turnover of radiolabeled FN and nonenzymatically glycated FN following I.V. administration of normal and diabetic animals. Next, we will determine if there is increased accumulation or turnover of fibronectin in glomerular cultures from normal and diabetic animals. Lastly, we will isolate plasma fibronectin from poorly controlled diabetic humans to determine whether it demonstrates the same abnormalities described above. Collectively these studies will allow us to investigate what may be a key biochemical abnormality in the diabetic GBM and mesangium, namely the perturbed molecular association of relevant molecules. Evaluation of a plasma protein which could serve as an indicator for the microangiopathic and nephropathic changes that are occurring in diabetes is very attractive. It seems clear that these studies may also shed light on the processes governing synthesis, turnover, and accumulation of basement constituents.