Detection of antigen-specific B cells in patients with pemphigus vulgaris by enzyme-linked immunospot assay: Requirement of T cell collaboration for autoantibody production

Koji Nishifuji, Masayuki Amagai, Masataka Kuwana, Toshiro Iwasaki, Takeji Nishikawa

Research output: Contribution to journalArticle

96 Citations (Scopus)

Abstract

Patients with pemphigus vulgaris have circulating IgG autoantibodies against desmoglein 3, which inhibit cell-cell adhesion of keratinocytes and cause blister formation in the skin and mucous membrane. To examine cellular mechanisms underlying the autoantibody production in pemphigus vulgaris patients, we have successfully developed an enzyme-linked immunospot assay which was able to detect desmoglein 3-specific autoimmune B cells quantitatively. Circulating B cells producing anti-desmoglein 3 antibodies were detected exclusively in three patients with severe disease (1.3-2.3/105 peripheral blood mononuclear cells), but not in 10 patients with mild disease or in remission or in seven healthy individuals. When this enzyme-linked immunospot assay was combined with in vitro stimulation of peripheral blood mononuclear cells with pokeweed mitogen and recombinant-desmoglein 3, we could detect circulating desmoglein 3-specific memory B cells in nine of 14 patients (6.3-84.0/105 peripheral blood mononuclear cells), but in none of 10 healthy individuals. We further analyzed the role of CD4+ T cells in promoting anti-desmoglein 3 antibody production. The in vitro anti-desmoglein 3 antibody production was abolished when CD4+ cells were depleted or when anti-HLA-DR or anti-HLA-DQ monoclonal antibody was added to the cultures. Our results demonstrated the quantitative detection of circulating 'activated' and 'memory' desmoglein 3-specific B cells and suggested the important part of HLA class II-restricted CD4+ T cells in the autoantibody production in pemphigus vulgaris. In addition, the enzyme-linked immunospot assay in combination with in vitro stimulation of B cells could be broadly applied to study mechanisms for autoantibody production in various autoimmure diseases.

Original languageEnglish
Pages (from-to)88-94
Number of pages7
JournalJournal of Investigative Dermatology
Volume114
Issue number1
DOIs
Publication statusPublished - 2000

Fingerprint

Desmoglein 3
Enzyme-Linked Immunospot Assay
Pemphigus
T-cells
Autoantibodies
Assays
B-Lymphocytes
Cells
T-Lymphocytes
Antigens
Enzymes
Blood Cells
Blood
Antibody Formation
Antibodies
HLA-DQ Antigens
Data storage equipment
Pokeweed Mitogens
Cell adhesion
HLA-DR Antigens

Keywords

  • Autoimmunity
  • Desmoglein
  • T cell-B cell colloboration

ASJC Scopus subject areas

  • Dermatology

Cite this

Detection of antigen-specific B cells in patients with pemphigus vulgaris by enzyme-linked immunospot assay : Requirement of T cell collaboration for autoantibody production. / Nishifuji, Koji; Amagai, Masayuki; Kuwana, Masataka; Iwasaki, Toshiro; Nishikawa, Takeji.

In: Journal of Investigative Dermatology, Vol. 114, No. 1, 2000, p. 88-94.

Research output: Contribution to journalArticle

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abstract = "Patients with pemphigus vulgaris have circulating IgG autoantibodies against desmoglein 3, which inhibit cell-cell adhesion of keratinocytes and cause blister formation in the skin and mucous membrane. To examine cellular mechanisms underlying the autoantibody production in pemphigus vulgaris patients, we have successfully developed an enzyme-linked immunospot assay which was able to detect desmoglein 3-specific autoimmune B cells quantitatively. Circulating B cells producing anti-desmoglein 3 antibodies were detected exclusively in three patients with severe disease (1.3-2.3/105 peripheral blood mononuclear cells), but not in 10 patients with mild disease or in remission or in seven healthy individuals. When this enzyme-linked immunospot assay was combined with in vitro stimulation of peripheral blood mononuclear cells with pokeweed mitogen and recombinant-desmoglein 3, we could detect circulating desmoglein 3-specific memory B cells in nine of 14 patients (6.3-84.0/105 peripheral blood mononuclear cells), but in none of 10 healthy individuals. We further analyzed the role of CD4+ T cells in promoting anti-desmoglein 3 antibody production. The in vitro anti-desmoglein 3 antibody production was abolished when CD4+ cells were depleted or when anti-HLA-DR or anti-HLA-DQ monoclonal antibody was added to the cultures. Our results demonstrated the quantitative detection of circulating 'activated' and 'memory' desmoglein 3-specific B cells and suggested the important part of HLA class II-restricted CD4+ T cells in the autoantibody production in pemphigus vulgaris. In addition, the enzyme-linked immunospot assay in combination with in vitro stimulation of B cells could be broadly applied to study mechanisms for autoantibody production in various autoimmure diseases.",
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