title: Human alpha-enolase from endothelial cells as a target antigen of anti-endothelial cell antibody in Behçet's disease.
abstract: OBJECTIVE
  To identify and recombine a protein of the human dermal microvascular endothelial cell (HDMEC) that specifically reacts with anti-endothelial cell antibody (AECA) in the serum of patients with Behçet's disease (BD), and to evaluate the usefulness of this protein in BD.
  
  
  METHODS
  The proteomics technique, with 2-dimensional gel electrophoresis and matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF) mass spectrometry, was used to identify and recombine HDMEC antigen. Western blotting and enzyme-linked immunosorbent assay (ELISA) of recombinant protein isolated by gene cloning were performed on serum from healthy controls, patients with BD, and patients with other rheumatic diseases (rheumatoid arthritis, systemic lupus erythematosus, and Wegener's granulomatosis).
  
  
  RESULTS
  Eighteen of 40 BD patients had serum IgM antibody to HDMEC antigen. The purified protein that reacted with AECA in BD patient sera was found to be alpha-enolase by 2-dimensional gel electrophoresis followed by immunoblotting and MALDI-TOF mass spectrometry. Recombinant alpha-enolase protein was isolated and refined by gene cloning. On Western blots, AECA-positive IgM from the sera of patients with active BD reacted strongly with recombinant human alpha-enolase. BD patient sera positive for anti-alpha-enolase did not react with human gamma-enolase. On dot-blotting, reactivity to human alpha-enolase was detected only in the IgM-positive group. Fifteen of the 18 AECA-positive sera that were positive for the HDMEC antigen showed reactivity to recombinant alpha-enolase IgM antibody by ELISA.
  
  
  CONCLUSIONS
  The alpha-enolase protein is the target protein of serum AECA in BD patients. This is the first report of the presence of IgM antibodies to alpha-enolase in endothelial cells from the serum of BD patients. Although further studies relating this protein to the pathogenesis of BD will be necessary, alpha-enolase and its antibody may prove useful in the development of new diagnostic and treatment modalities in BD.
authors.affiliation: Cutaneous Biology Research Institute, and Brain Korea 21 Project for Medical Science, Yonsei University College of Medicine, Seoul, South Korea.
authors.forename: Kwang Hoon
authors.initials: KH
authors.lastname: Lee
authors.forename: Hae-Shin
authors.initials: HS
authors.lastname: Chung
authors.forename: Hyoung Sup
authors.initials: HS
authors.lastname: Kim
authors.forename: Sang-Ho
authors.initials: SH
authors.lastname: Oh
authors.forename: Moon-Kyung
authors.initials: MK
authors.lastname: Ha
authors.forename: Ja-Hyun
authors.initials: JH
authors.lastname: Baik
authors.forename: Sungnack
authors.initials: S
authors.lastname: Lee
authors.forename: Dongsik
authors.initials: D
authors.lastname: Bang
mesh_terms: D001323:Autoantibodies / Q000097:blood*; D001324:Autoantigens / Q000235:genetics / Q000276:immunology*; D001528:Behcet Syndrome / Q000276:immunology*; D015153:Blotting, Western; D002478:Cells, Cultured; D003001:Cloning, Molecular; D000939:Epitopes; D006801:Humans; D007075:Immunoglobulin M / Q000097:blood; D010751:Phosphopyruvate Hydratase / Q000235:genetics / Q000276:immunology*; D040901:Proteomics; D011994:Recombinant Proteins / Q000276:immunology; D012867:Skin / Q000166:cytology
chemical_list: D001323:Autoantibodies; D001324:Autoantigens; D000939:Epitopes; D007075:Immunoglobulin M; D011994:Recombinant Proteins; D010751:Phosphopyruvate Hydratase