title: Epitopes, avidity and IgG subclasses of tyrosine hydroxylase autoantibodies in vitiligo and alopecia areata patients.
abstract: BACKGROUND
  We previously detected antibodies against tyrosine hydroxylase (TH) in 23% of patients with nonsegmental vitiligo and in 19% of patients with alopecia areata (AA).
  
  
  OBJECTIVE
  To identify TH epitopes recognized by TH antibodies in patients with vitiligo and AA.
  
  
  METHODS
  Recombinant plasmids containing defined fragments of TH cDNA were constructed. The cloned TH cDNA fragments were subsequently translated in vitro to produce a series of [(35) S]-labelled TH protein fragments which were then used in radioimmunoassays to analyse the immunoreactivity of sera from 18 TH antibody-positive patients with vitiligo and so initially define TH epitope domains. Further localization of TH epitopes was investigated by antibody absorption experiments using synthetic TH peptides and nonradiolabelled, in vitro-expressed TH protein fragments. Antibody binding to identified epitopes was confirmed in TH peptide enzyme-linked immunosorbent assays.
  
  
  RESULTS
  Analysis of the results obtained indicated the presence of two major antibody-binding sites on TH between amino acids 1 and 14 (epitope 1-14) and between amino acids 61 and 80 (epitope 61-80). Of 18 patients with vitiligo and six with AA, 17 (94%) and five (83%), respectively, had antibodies against epitope 1-14. In addition, 11/18 (61%) vitiligo and 2/6 (33%) AA patient sera displayed immunoreactivity against epitope 61-80.
  
  
  CONCLUSIONS
  Two major binding sites for human TH antibodies are located at the N-terminus of the protein. The humoral immune response to TH in vitiligo and AA is heterogeneous in nature in that patients may have antibodies to more than one TH epitope. TH antibodies from patients with vitiligo or AA can recognize identical epitopes.
authors.affiliation: Department of Human Metabolism, The Medical School, University of Sheffield, Sheffield S10 2RX, UK.
authors.forename: S F E
authors.initials: SF
authors.lastname: Rahoma
authors.forename: H K
authors.initials: HK
authors.lastname: Sandhu
authors.forename: A J G
authors.initials: AJ
authors.lastname: McDonagh
authors.forename: D J
authors.initials: DJ
authors.lastname: Gawkrodger
authors.forename: A P
authors.initials: AP
authors.lastname: Weetman
authors.forename: E H
authors.initials: EH
authors.lastname: Kemp
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000506:Alopecia Areata / Q000276:immunology*; D001323:Autoantibodies / Q000378:metabolism*; D001665:Binding Sites; D002648:Child; D002675:Child, Preschool; D018076:DNA, Complementary / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D018985:Epitopes, B-Lymphocyte / Q000378:metabolism*; D005260:Female; D006801:Humans; D007074:Immunoglobulin G / Q000145:classification / Q000378:metabolism*; D008297:Male; D008875:Middle Aged; D011863:Radioimmunoassay; D014446:Tyrosine 3-Monooxygenase / Q000276:immunology*; D014820:Vitiligo / Q000276:immunology*; D055815:Young Adult
chemical_list: D001323:Autoantibodies; D018076:DNA, Complementary; D018985:Epitopes, B-Lymphocyte; D007074:Immunoglobulin G; D014446:Tyrosine 3-Monooxygenase