On the Molecular Architecture of the Epitherial Desmosom and Its an Etiological Role

上皮桥粒的分子结构及其病因学作用

基本信息

批准号：
02670474
负责人：
SHIDA Hisato
金额：
$ 1.34万
依托单位：
Yamanashi Medical College (Yamanashi Ikadaigaku)
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1990
资助国家：
日本
起止时间：
1990 至 1991
项目状态：
已结题

项目摘要

For the purpose of investigating the molecular organization of the desmosome, quantitative analysis of the immunoelecton microscopy (IEM) using immuno-gold method with anti-D mono/polyclonal antibodies (Abs) was carried out. According to the analysis, such useful tool as position of peaks, shape of curves obtained from length/frequency curves which were specific to the DG and DP molecules are recognized in the first year. However, theoretical meanings were remained to be uncertain. From the second year, computer aided distributions were investigated as the simulated models of immunoelectron microscopy. By applying fitting analysis to the histogram, a combined curve of two normal distribution curves with the different value of half width and the same value of peak position were appeared as a least square curve and from this combined curve, the resolution was estimated as 60 A. Basing on the theoretical analysis and the quantitative immunoelectron microscopy of desmosome, a three dimensional model of cytoskeletal-D proteins complex at the desmosome was proposed.A two dimensional architecture of D proteins and E-cadherin (ECAD) was also analyzed by IEM using isolated D pre-treated with 6 M guanidine HCl (G-HCl). At the proper region of D, density of DG1, DP1/2, DG2/3 was decreased in this order forming a random distribution. The considerable amount of DP3 was extracted by the G-HCl treatment. Only a few amount of E-CAD was detected at the peripheral region of D. The difference of the localization pattern between the D molecules and E-CAD was also confirmed in cultured keratinocytes. Though the break-down of cell-to-cell adhesion of the epitherial cells was observed under the presence of anti-DG1 and/or DG2/3 Abs for 27 hs incubation, non of the effect of pepstatin was observed.

为了研究脱骨体的分子组织，使用抗D单/多克隆抗体（ABS）的免疫金方法对免疫电信显微镜显微镜（IEM）进行定量分析。根据分析，诸如峰位置的有用工具，从长度/频率曲线获得的曲线形状，这些曲线是特定于DG和DP分子的曲线。但是，理论含义尚不确定。从第二年开始，将计算机辅助分布研究为免疫电子显微镜的模拟模型。 By applying fitting analysis to the histogram, a combined curve of two normal distribution curves with the different value of half width and the same value of peak position were appeared as a least square curve and from this combined curve, the resolution was estimated as 60 A. Basing on the theoretical analysis and the quantitative immunoelectron microscopy of desmosome, a three dimensional model of cytoskeletal-D proteins complex at the desmosome was提出的。还使用IEM使用与6 m鸟苷HCl（G-HCl）预处理的隔离D进行了IEM分析D蛋白和E-钙粘蛋白（ECAD）的二维结构。在D的适当区域，DG1，DP1/2，DG2/3的密度减少了，形成随机分布。通过G-HCL处理提取了大量DP3。在培养的角质形成细胞中也证实了D分子和E-CAD之间的定位模式的差异。尽管在27 HS孵育的抗DG1和/或DG2/3 ABS的存在下观察到了e腹细胞的细胞对细胞粘附的分解，但观察到pepstatin的作用并非被观察到pepstatin的作用。