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Abstract
The purpose of this study was to identify the endothelial cell antigens that react with circulating antiendothelial antibody (AECA) in mixed connective tissue disease (MCTD). We screened serum AECA reactivity in 23 patients with MCTD using a human glomerular endothelial cell (HGEC) cellular ELISA. Proteomics, two-dimensional gel electrophoresis and matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry were used to identify the endothelial cell antigens of HGECs that reacted with serum antibodies from MCTD patients. Sera from 12 patients (52.0%) were positive for anti-HGEC antibody based on cellular ELISA. MALDI-TOF mass spectrometry used in combination with immunoblotting using serum antibody revealed one protein spot that represented a 36-kDa cell component of HGECs, with an isoelectric point (IP) of about 9, which had a high homology with the voltage-dependent anion-selective channel 1 (VDAC-1). This protein spot was confirmed to react with the antibody specific to VDAC-1. This is the first report of the presence of antibody to VDAC-1 from HGECs in the sera from MCTD patients. Although future studies will be needed to clarify the disease specificity of the a-VDAC-1 antibody in MCTD, the results show that modern proteomics technology is useful for identifying antigens that react with AECA in autoimmune diseases such as MCTD.