Analysis of pathophysiology of pemphigus foliaceus by using immunoelectron microscopy

天疱疮是一种自身免疫性水疱病，由抗桥粒芯糖蛋白1和/或桥粒蛋白3的自身抗体引起。本研究的目的是用免疫电子显微镜来阐明自身抗体结合导致水泡形成的机制。因此，桥粒分裂和角蛋白回缩是各型天疱疮的共同特征。桥粒的分裂是棘层松解和角蛋白回缩的最初改变，随后导致细胞分离。在接下来的一年里，我们试图免疫定位正常人表皮中的PF自身抗体结合部位，但没有成功。我们目前正在尝试另一种免疫电子显微镜方法。我们还试图在狗身上描述PF的特征。犬肺功能衰竭可作为人类肺功能衰竭的动物模型，值得进一步阐明。我们成功地鉴定了犬PF自身抗体结合部位的超微结构结合部位。包埋后的免疫金EM使其分布在桥粒复合体的胞内和胞外.通过桥粒蛋白抗体的双重染色，细胞内标记的PF血清与桥粒蛋白共定位。通过免疫印迹分析，用PH犬血清检测到一条与桥粒蛋白分子量相似的条带。此外，与人桥粒蛋白1免疫印迹阴性的血清与人桥粒蛋白抗体结合部位的细胞外标记结果表明，犬PF自身抗体识别桥粒分子，包括桥粒蛋白和除桥粒蛋白1以外的未知跨膜分子。

Pemphigus is an autoimmune blistering disease caused by autoantibodies against desmoglein 1 and/ or desmoglein 3. The purpose of this study is to elucidate the mechanism whereby autoantibody binding lead to the blister formation using immunoelectron microscopy.In the first year, we have collected more than 10 cases of pemphigus vulgaris and foliaceus (PF) and characterized their ultrastructural findings according to the clinical subtypes. As results, split of desmosomes and keratin retraction were the shared common features among all types of pemphigus. Split of desmosomes was the initial change of acantholysis and keratin retraction followed resulting in cell separation.In the next year, we tried to immunolocalize the PF autoantibody binding site in the normal human epidermis, but it was unsuccessful. We are currently trying the other method for immunoelectron microscopy. We also tried to characterize PF in dogs. Canine PF can be an animal model of human PF and is worth elucidating. We have succeeded in identifying the ultrastructural binding site of the canine PF autoantibody binding site. It' was distributed around desmosome complex, both intra- and extracellularly by postembedding immunogold EM. By double staining with desmoplakin antibody, intracellular labeling of the PF serum co-localized with desmoplakin. By immunoblot analysis, a band with similar molecular weight to desmoplakin was detected using the canine PH serum. In addition, extracellular labeling of the serum co-localized with human PF antibody binding site, although immunoblotting was negative with human desmoglein 1. These results suggested that Canine PF autoantibody recognized desmosomal molecules including desmoplakin and unknown transmembrane molecule other than desmoglein 1.