Molecular biological approach to the abnormalities of extracelluar matrix of the diabetic nephropathy.

细胞外基质由胶原、层粘连蛋白和蛋白聚糖组成。在糖尿病肾病中，IV型胶原的生化特性改变和蛋白聚糖的表达减少被认为是疾病的主要病理改变，并与肾小球细胞的表型改变有关。糖尿病肾病合并肾病综合征患者的免疫电镜检查显示肾小球基底膜（GBM）增厚，IV型胶原数量增加，密度降低，硫酸肝素蛋白聚糖减少。这些细胞外基质成分的改变似乎有助于GBM的大小和电荷屏障的损害和蛋白尿。此外，我们克隆了一条新的IV型胶原蛋白链，即α - 4链，其cDNA （NK4）包含NC-1结构域，长度为1.9 Kb。Northern blot分析显示-9.5 Kb的转录本。免疫荧光检测alpha1、alpha2、alpha3、alpha4和基底膜相关胶原（BAC）显示alpha1和alpha2定位于肾小球系膜区和肾小球毛细血管，而alpha3和alpha4主要沿毛细血管壁定位。在系膜区观察BAC免疫反应性。糖尿病患者肾脏系膜基质扩大，GBM增厚，alpha1-alpha4链和BAC表达增加。此外，我们还分析了间质性胶原如III、V、VI的表达情况。III型胶原只在肾间质表达，V型和VI型胶原在肾系膜上表达。在糖尿病肾中，这些胶原蛋白在肾系膜区增加。

Extracellular matrix has been shown to be consisted of collagens, laminins and proteoglycans. In diabetic nephropathy, the altertions of biochemical properties of type IV collagen and the decreased expression of proteoglycans are considered to be major pathological changes of the disease and related to the phenotypic changes of glomerular cells. Immunoelectron microscopic studies of diabetic nepropathy patients with nephrotic syndrome showed the thicking of glomerular basement membranes (GBM), increased amount of type IV collagens with decreased density, and decreased heparan sulfate proteoglycan. These alterations of extracellular matrix components seems to contribute to the the impairment of size and charge barrier of the GBM and proteinuria. Furthermore, we cloned a novel chain of type IV collagen, i.e.alpha4 chain, which cDNA (NK4) included NC-1 domain and expands 1.9 Kb. Northern blot analyzes showed -9.5 Kb transcript. Immunofluorescence studies of alpha1, alpha2, alpha3, alpha4, and basement membrane associated-collagen (BAC) showed alpha1 and alpha2 localized to the glomerular mesangial area and glomerular capillary, while alpha3 and alpha4 localized mainly along the capillary wall. BAC immunoreactivity was observed in the mesangial area. In the kidney of diabetic patients, expanded mesangial matrix and thickened GBM showed increased expression of alpha1-alpha4 chains and BAC.Furthermore, we analyzed the expression of the interstitial collagens, such as III,V,VI collagens. Type III collagen exclusively expressed in the interstinum of the kidney and type V and VI collagens had mesangial expressions. In diabetic kidney, these collagens were increased in the mesangial area.