TY - JOUR
T1 - Proteomic identification of heterogeneous nuclear ribonucleoprotein K as a novel cold-associated autoantigen in patients with secondary Raynaud's phenomenon
AU - Yang, Lingli
AU - Fujimoto, Minoru
AU - Murota, Hiroyuki
AU - Serada, Satoshi
AU - Fujimoto, Manabu
AU - Honda, Hiromi
AU - Yamada, Kohji
AU - Suzuki, Katsuya
AU - Nishikawa, Ayumi
AU - Hosono, Yuji
AU - Yoneda, Yoshihiro
AU - Takehara, Kazuhiko
AU - Imura, Yoshitaka
AU - Mimori, Tsuneyo
AU - Takeuchi, Tsutomu
AU - Katayama, Ichiro
AU - Naka, Tetsuji
N1 - Funding Information:
We would like to thank Y. Kanazawa and J. Yamagishi for secretarial assistance and M. Urase and K. Yoshimoto for technical assistance. This study was supported by a grant-in-aid for the Program for Promotion of Fundamental Studies in Health Sciences of the National Institute of Biomedical Innovation and a Grant-in-Aid for Research on Biological Markers for New Drug Development (H20-0005) from the Ministry of Health, Labour and Welfare of Japan.
Publisher Copyright:
© The Author 2014.
PY - 2015/2
Y1 - 2015/2
N2 - Objective. The aim of this study was to identify cold-associated autoantibodies in patients with RP secondary to CTDs. Methods. Indirect immunofluorescence staining was performed on non-permeabilized cold-stimulated normal human dermal microvascular endothelial cells (dHMVECs), using patients' sera. Cold-induced alterations in cell surface proteomes were analysed by isobaric tag for relative and absolute quantitation (iTRAQ) analysis. Serological proteome analysis (SERPA) was applied to screen cold-associated autoantigens. The prevalence of the candidate autoantibody was determined by ELISA in 290 patients with RP secondary to CTDs (SSc, SLE or MCTD), 10 patients with primary RP and 27 healthy controls. Results. Enhanced cell surface immunoreactivity was detected in cold-stimulated dHMVECs when incubated with sera from patients with secondary RP. By iTRAQ analysis, many proteins, including heterogeneous nuclear ribonucleoprotein K (hnRNP-K), were found to be increased on the cell surface of dHMVECs after cold stimulation. By the SERPA approach, hnRNP-K was identified as a candidate autoantigen in patients with secondary RP. Cold-induced translocation of hnRNP-K to the cell surface was confirmed by immunoblotting and flow cytometry. By ELISA analysis, patients with secondary RP show a significantly higher prevalence of anti-hnRNP-K autoantibody (30.0%, 61/203) than patients without RP (9.2%, 8/87, P = 0.0001), patients with primary RP (0%, 0/10, P = 0.0314) or healthy controls (0%, 0/27, P = 0.0001). Conclusion. By comprehensive proteomics, we identified hnRNP-K as a novel cold-associated autoantigen in patients with secondary RP. Anti-hnRNP-K autoantibody may potentially serve as a biomarker for RP secondary to various CTDs.
AB - Objective. The aim of this study was to identify cold-associated autoantibodies in patients with RP secondary to CTDs. Methods. Indirect immunofluorescence staining was performed on non-permeabilized cold-stimulated normal human dermal microvascular endothelial cells (dHMVECs), using patients' sera. Cold-induced alterations in cell surface proteomes were analysed by isobaric tag for relative and absolute quantitation (iTRAQ) analysis. Serological proteome analysis (SERPA) was applied to screen cold-associated autoantigens. The prevalence of the candidate autoantibody was determined by ELISA in 290 patients with RP secondary to CTDs (SSc, SLE or MCTD), 10 patients with primary RP and 27 healthy controls. Results. Enhanced cell surface immunoreactivity was detected in cold-stimulated dHMVECs when incubated with sera from patients with secondary RP. By iTRAQ analysis, many proteins, including heterogeneous nuclear ribonucleoprotein K (hnRNP-K), were found to be increased on the cell surface of dHMVECs after cold stimulation. By the SERPA approach, hnRNP-K was identified as a candidate autoantigen in patients with secondary RP. Cold-induced translocation of hnRNP-K to the cell surface was confirmed by immunoblotting and flow cytometry. By ELISA analysis, patients with secondary RP show a significantly higher prevalence of anti-hnRNP-K autoantibody (30.0%, 61/203) than patients without RP (9.2%, 8/87, P = 0.0001), patients with primary RP (0%, 0/10, P = 0.0314) or healthy controls (0%, 0/27, P = 0.0001). Conclusion. By comprehensive proteomics, we identified hnRNP-K as a novel cold-associated autoantigen in patients with secondary RP. Anti-hnRNP-K autoantibody may potentially serve as a biomarker for RP secondary to various CTDs.
KW - Autoantibody
KW - Heterogeneous nuclear RNP-K
KW - Proteomics
KW - Raynaud's phenomenon
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U2 - 10.1093/rheumatology/keu325
DO - 10.1093/rheumatology/keu325
M3 - Article
C2 - 25172934
AN - SCOPUS:84925582114
VL - 54
SP - 349
EP - 358
JO - Rheumatology and Rehabilitation
JF - Rheumatology and Rehabilitation
SN - 1462-0324
IS - 2
ER -