title: ANCA antigens, proteinase 3 and myeloperoxidase, are not expressed in endothelial cells.
abstract: BACKGROUND
  One hypothesis for the pathogenesis of vasculitis associated with antineutrophil cytoplasmic autoantibodies (ANCAs) proposes that ANCAs bind to ANCA antigens, such as proteinase 3 (PR3) or myeloperoxidase (MPO), which are produced by endothelial cells and expressed on their surfaces. There are conflicting reports, however, on whether endothelial cells express the ANCA antigen PR3, and there are no reports on endothelial expression of MPO. The aim of this study was to determine the presence or absence of PR3 and MPO mRNA in both venous and arterial endothelial cells, employing standard reverse transcription-polymerase chain reaction (RT-PCR) techniques and also the quantitative and highly specific method, TaqMan PCR.
  
  
  METHODS
  RT-PCR (with 3 primer sets) and TaqMan PCR, a method for detecting low copy transcripts, were used to probe for PR3 and MPO transcripts in human endothelial cells from umbilical vein (HUVEC) and artery (HUAEC) and from lung microvascular (HLMVEC). Cells were treated with interferon-gamma (200 units/mL) or tumor necrosis factor-alpha (3 or 10 ng/mL) or both.
  
  
  RESULTS
  Transcripts for PR3 and/or MPO were not detected in HUVEC, HUAEC, and HLMVEC by standard RT-PCR. Analyses for PR3 protein confirmed that PR3 is not expressed in HUVEC. HUVEC and HUAEC were negative for PR3 and MPO by TaqMan PCR.
  
  
  CONCLUSIONS
  PR3 and MPO are not expressed in HUVEC, HUAEC, or HLMVEC. Endothelial cell presentation of endogenous PR3 and MPO antigens is not involved in the pathogenesis of ANCA-associated vasculitis. Alternative explanations need to be explored to determine the pathogenic effect of ANCAs.
authors.affiliation: Division of Nephrology and Hypertension, Department of Medicine, University of North Carolina at Chapel Hill, 27599, USA.
authors.forename: W F
authors.initials: WF
authors.lastname: Pendergraft
authors.forename: D A
authors.initials: DA
authors.lastname: Alcorta
authors.forename: M
authors.initials: M
authors.lastname: Segelmark
authors.forename: J J
authors.initials: JJ
authors.lastname: Yang
authors.forename: R
authors.initials: R
authors.lastname: Tuttle
authors.forename: J C
authors.initials: JC
authors.lastname: Jennette
authors.forename: R J
authors.initials: RJ
authors.lastname: Falk
authors.forename: G A
authors.initials: GA
authors.lastname: Preston
mesh_terms: D019268:Antibodies, Antineutrophil Cytoplasmic / Q000235:genetics*; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D053146:Myeloblastin; D009195:Peroxidase / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D012697:Serine Endopeptidases / Q000032:analysis / Q000235:genetics*; D014657:Vasculitis / Q000209:etiology
chemical_list: D019268:Antibodies, Antineutrophil Cytoplasmic; D012333:RNA, Messenger; D009195:Peroxidase; D012697:Serine Endopeptidases; D053146:Myeloblastin