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Basic studies for the diagnosis of bullous diseases having pathologic changes at the basement membrane zone

基底膜带病变的大疱性疾病诊断基础研究

基本信息

批准号：
02044130
负责人：
NISHIKAWA Takeji
金额：
$ 2.82万
依托单位：
Keio Gijuku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for international Scientific Research
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

There are a number of skin diseases in which epidermal basement membrane zone (BMZ) is affected. Therefore, the research for epidermal BMZ is important for both diagnostic and therapeutic respects. First, we investigated a series of antibodies against various components of BMZ, including LH7.2 and GB3 monoclonal antibodies and human monoclonal antibody against 23OkD bullous pemphigoid (BP) antigen. One approach is the precise localization of the antigens for these antibodies. For this purpose we have established cryofixation and cryosubstitution for immunoelectron microscopy in collaboration with Professor Eady, England.Another approach is the determination of such antigens by using biochemical and molecular biological techniques. We discussed for these projects with Drs. Stanley and Amagai, U. S. A. and with Dr. Black, England, who also provided us with sera from their BP cases. Using immunoblot and immunoprecipitation method, we first showed that autoantigen profile is almost the same among Japanese, British and U. S. cases. We also found that although 230kD BP antigen is restricted to BMZ, 170kD BP antigen may exist in both BMZ and cell membrane of basal keratinocytes. Furthermore, although 170kD BP antigen was thought not to be detectable by immunoprecipitation, we showed 170kD BP antigen can be detectable by our improved immunoprecipitation method. In addition, we also established ELISA method using recombinant antigen protein produced by CDNA clone for 230kD BP antigen. This method can substitute the conventional immunofluorescence technique to detect anti-BMZ antibodies in BP sera.Another important disease is linear IgA bullous dermatosis (LAD). We have investigated for LAD autoantigen as collaboration work with Professor Zone, U. S. A. and Drs. Black and Wojnarowska, England. We confirmed that some LAD sera specifically react with 97kD protein. However, the real nature of this 97kD protein should be clarified by further confirmative studies.

有许多皮肤病，其中表皮基底膜区（BMZ）受到影响。因此，对表皮BMZ的研究具有重要的诊断和治疗意义。首先，我们研究了一系列针对BMZ不同成分的抗体，包括LH7.2和GB3单克隆抗体以及23OkD大疱性类天疱疮（BP）抗原的人单克隆抗体。一种方法是精确定位这些抗体的抗原。为此，我们与英国的Eady教授合作，建立了免疫电子显微镜的冷冻固定和冷冻替代。另一种方法是利用生物化学和分子生物学技术来测定这些抗原。我们和博士讨论了这些项目。Stanley和Amagai，美国以及英国的Black博士，他也向我们提供了他们的BP病例的血清。利用免疫印迹和免疫沉淀方法，我们首先发现日本、英国和美国病例的自身抗原谱几乎相同。我们还发现，虽然230kD BP抗原局限于BMZ，但170kD BP抗原可能存在于BMZ和基底角化细胞的细胞膜中。此外，尽管170kD BP抗原被认为不能通过免疫沉淀法检测到，但我们发现170kD BP抗原可以通过改进的免疫沉淀法检测到。此外，我们还建立了利用CDNA克隆产生的重组抗原蛋白对230kD BP抗原进行酶联免疫吸附测定的方法。该方法可替代传统的免疫荧光技术检测BP血清中抗bmz抗体。另一种重要疾病是线状IgA大疱性皮肤病（LAD）。我们与美国的Zone教授和dr。Black and Wojnarowska，英格兰。我们证实一些LAD血清与97kD蛋白有特异性反应。然而，这种97kD蛋白的真正性质需要进一步的确证性研究来澄清。