Ribosomal P protein P0 as a candidate for the target antigen of anti-endothelial cell antibodies in mixed connective tissue disease.
OBJECTIVE
To evaluate the presence of anti-endothelial cell antibodies (AECA) in patients with mixed connective tissue disease (MCTD) compared to those with systemic sclerosis (SSc) and to determine the candidates for the endothelial auto-antigen that reacts with AECA in patients with MCTD using a molecular cloning strategy.


METHODS
AECA were measured by a cellular enzyme-linked immunosorbent assay (ELISA) using fixed human umbilical vein endothelial cells (HUVEC) in 47 MCTD patients, 68 SSc patients, and 52 normal controls. A HUVEC cDNA expression library was immunoscreened with pooled sera from 6 patients with high AECA levels determined by cellular ELISA to explore the endothelial autoantigens in MCTD. An ELISA assay for anti-ribosomal protein P0 antibodies was used to assess the correlation with AECA levels.


RESULTS
The candidate target proteins recognized by AECA in MCTD included: (i) ribosomal protein P0; (ii) a putative oncogene derived from dek mRNA; (iii) SS-B/La protein; (iv) U1 RNA-associated 70K protein; and (v) DNA-binding protein B. A significant correlation between the levels of AECA and anti-ribosomal protein P0 antibodies was demon-strated in MCTD, but not in systemic sclerosis. The sera containing high levels of AECA from patients with MCTD frequently cross-reacted with ribosomal protein P0. On the other hand, sera without AECA activity from patients with MCTD never reacted with ribosomal protein P0.


CONCLUSIONS
AECA were more frequently seen in patients with MCTD than in patients with SSc. Ribosomal protein P0 may be one of the major target antigens of AECA in patients with MCTD.
Naniwa
T
T
Division of Rheumatology, Nagoya City University Hospital, and Department of Medical Oncology and Immunology, Nagoya City University Graduate School of Medical Sciences, Nagoya, Japan. tnaniwa@med.nagoya-cu.ac.jp
Sugiura
Y
Y
Banno
S
S
Yoshinouchi
T
T
Matsumoto
Y
Y
Ueda
R
R
D000328:Adult; D000941:Antigens / Q000276:immunology*; D001323:Autoantibodies / Q000276:immunology*; D001324:Autoantigens / Q000032:analysis; D003001:Cloning, Molecular; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D008947:Mixed Connective Tissue Disease / Q000276:immunology*; D009857:Oncogenes / Q000276:immunology; D012261:Ribonucleoproteins / Q000032:analysis; D012269:Ribosomal Proteins / Q000276:immunology*; D012595:Scleroderma, Systemic / Q000276:immunology; D000097585:SS-B Antigen
D000941:Antigens; D001323:Autoantibodies; D001324:Autoantigens; D012261:Ribonucleoproteins; D012269:Ribosomal Proteins; D000097585:SS-B Antigen; C087589:anti-endothelial cell antibody; C054675:ribosomal protein P0