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The identification of autoantigens in mucous membrane pemphigoid using immortalized oral mucosal keratinocytes
Author(s) -
Kamaguchi Mayumi,
Iwata Hiroaki,
Miyauchi Toshinari,
Ujiie Hideyuki,
Ujiie Inkin,
Nomura Toshifumi,
Ohga Noritaka,
Shimizu Hiroshi,
Kitagawa Yoshimasa
Publication year - 2019
Publication title -
journal of oral pathology and medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.887
H-Index - 83
eISSN - 1600-0714
pISSN - 0904-2512
DOI - 10.1111/jop.12780
Subject(s) - laminin , immortalised cell line , autoantibody , matrix metalloproteinase , biology , pemphigoid , microbiology and biotechnology , cell culture , basement membrane , immunology , bullous pemphigoid , pathology , antibody , extracellular matrix , medicine , biochemistry , genetics
Background Mucous membrane pemphigoid ( MMP ) is a rare chronic autoimmune subepithelial blistering disorder, targeting multiple basement membrane zone ( BMZ ) proteins including collagen XVII ( COL 17). Circulating autoantibodies of MMP are often undetected due to their lower titers. The oral mucosa is a valuable substrate for the detection of autoantibodies in MMP patients. However, obtaining normal human oral mucosa is more difficult than obtaining normal human skin. We established immortalized normal human oral mucosal keratinocytes ( OMK s) and performed immunoblotting using immortalized OMK lysate for detecting autoantigens in MMP . Methods Immortalized OMK s were generated from primary OMK s using E6/E7 proteins of HPV . We compared the protein expression levels of major BMZ proteins between primary OMK s and immortalized OMK s. We performed immunoblotting to detect autoantigens using cell lysates from immortalized OMK s in 30 MMP patients. Results There were no significant differences between primary OMK s and immortalized OMK s in terms of protein expression levels of the BMZ proteins, including COL 17, laminin 332, integrin α6/β4, collagen VII , and collagen IV . Cell lysates of immortalized OMK s effectively identified MMP autoantigens in 60% (18/30) of MMP sera. We found an interesting case of MMP whose autoantibodies preferentially reacted to the 120‐ kD protein that is an ectodomain of COL 17. Conclusion We demonstrated that a cell lysate of immortalized OMK s is a reliable substrate for the detection of MMP autoantigens. This newly developed immunoblotting analysis method promises to contribute to the diagnosis of MMP .