Anti-citrullinated Protein Antibodies Activated ERK1/2 and JNK Mitogen-activated Protein Kinases via Binding to Surface-
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ORIGINAL RESEARCH
Anti-citrullinated Protein Antibodies Activated ERK1/2 and JNK Mitogen-activated Protein Kinases via Binding to Surface-expressed Citrullinated GRP78 on Mononuclear Cells Ming-Chi Lu & Ning-Sheng Lai & Wen-Yao Yin & Hui-Chun Yu & Hsien-Bin Huang & Chien-Hsueh Tung & Kuang-Yung Huang & Chia-Li Yu
Received: 6 August 2012 / Accepted: 16 November 2012 / Published online: 28 November 2012 # Springer Science+Business Media New York 2012
Abstract In a previous study, we found that anti-citrullinated protein antibodies (ACPAs) enhance nuclear factor (NF)-κB activity and tumor necrosis factor (TNF)-α production by normal human peripheral blood mononuclear cells (PBMCs) and U937 cells via binding to surface-expressed citrullinated glucose-regulated protein 78 (cit-GRP78). However, the downstream signaling pathways remain unclear after binding. In the present study, we firstly measured the effects of different M.-C. Lu : N.-S. Lai : H.-C. Yu : C.-H. Tung : K.-Y. Huang Division of Allergy, Immunology and Rheumatology, Buddhist Dalin Tzu Chi General Hospital, Chia-Yi, Taiwan M.-C. Lu : N.-S. Lai : K.-Y. Huang School of Medicine, Tzu Chi University, Hualien, Taiwan W.-Y. Yin Division of General Surgery, Buddhist Dalin Tzu Chi General Hospital, Chia-Yi, Taiwan H.-C. Yu : H.-B. Huang Department of Life Science and Institute of Molecular Biology, National Chung Cheng University, Chia-Yi, Taiwan C.-L. Yu Department of Internal Medicine, National Taiwan University Hospital and National Taiwan University College of Medicine, Taipei, Taiwan C.-L. Yu (*) Department of Internal Medicine, National Taiwan University Hospital, No. 7 Chung-Shan South Road, Taipei 100, Taiwan e-mail: [email protected] W.-Y. Yin Division School of Medicine, Tzu Chi University, Hualien, Taiwan
kinase inhibitors on ACPA-mediated TNF-α production from normal PBMCs and monocytes. Then, the native and phosphorylated mitogen-activated protein kinases (MAPKs) were detected in ACPA-activated U937 cells by Western blotting. We also explored the role of the phosphoinositide 3-kinase (PI3K)-Akt pathway in activating IκB kinase alpha (IKK-α) in ACPA-stimulated U937 cells. Finally, we measured the amount of cit-GRP78 from PBMC membrane extracts in RA patients and controls. We found that MAPK and Akt inhibitors, but not PI3K inhibitor, remarkably suppressed ACPAmediated TNF-α production. Interestingly, ACPAs selectively activated extracellular signal-regulated kinase 1/2 (ERK1/2) and c-jun N-terminal kinase (JNK), but not p38 MAPK, in U937 cells. This activation was suppressed by cit-GRP78, but not GRP78. The JNK activation further enhanced the phosphorylation of Akt and IKK-α. The expression of cit-GRP78 on cell membrane was higher in RA than normal PBMCs. Taken together; these results suggest that through binding to surface, over-expressed cit-GRP78 on RA PBMCs, ACPAs selectively activate ERK1/2 and JNK signaling pathways to enhance IKK-α phosphorylation, which leads to the activation of NF-κB and the production of TNF-α . Keywords ACPAs . rheumatoid
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