TY - JOUR
T1 - Enhanced neutrophil extracellular trap generation in rheumatoid arthritis
T2 - Analysis of underlying signal transduction pathways and potential diagnostic utility
AU - Sur Chowdhury, Chanchal
AU - Giaglis, Stavros
AU - Walker, Ulrich A.
AU - Buser, Andreas
AU - Hahn, Sinuhe
AU - Hasler, Paul
N1 - Funding Information:
We are grateful to Peter Erb, Ed Palmer and Alan Tyndall for helpful suggestions and comments. We thank Swarna Machineni for performing the ROS analyses, and Maria Stoikou for assistance with the IgG depletion studies. A. Schoetzau, Eudox, Basel, Switzerland provided statistical supervision. Norman Bandelow, Christoph Hemmeler, Eric Deman, Rheumaklinik, Kantonsspital Aarau, and Robyn Benz, Department of Rheumatology, University Hospital, Basel, provided patient samples. This project was supported by a grant from the Fonds W & W of the Cantonal Hospital Aarau. The position of Chanchal Sur Chowdhury was supported by the University Women's Hospital Basel.
PY - 2014/6/13
Y1 - 2014/6/13
N2 - Introduction: Neutrophil extracellular traps (NETs) have recently been implicated in a number of autoimmune conditions, including rheumatoid arthritis (RA). We examined the underlying signaling pathways triggering enhanced NETosis in RA and ascertained whether the products of NETosis had diagnostic implications or usefulness.Methods: Neutrophils were isolated from RA patients with active disease and from controls. Spontaneous NET formation from RA and control neutrophils was assessed in vitro with microscopy and enzyme-linked immunosorbent assay (ELISA) for NETosis-derived products. The analysis of the signal-transduction cascade included reactive oxygen species (ROS) production, myeloperoxidase (MPO), neutrophil elastase (NE), peptidyl arginine deiminase 4 (PAD4), and citrullinated histone 3 (citH3). NET formation was studied in response to serum and synovial fluid and immunoglobulin G (IgG) depleted and reconstituted serum. Serum was analyzed for NETosis-derived products, for which receiver operator characteristic (ROC) curves were calculated.Results: Neutrophils from RA cases exhibited increased spontaneous NET formation in vitro, associated with elevated ROS production, enhanced NE and MPO expression, nuclear translocation of PAD4, PAD4-mediated citrullination of H3, and altered nuclear morphology. NET formation in both anti-citrullinated peptide antibody (ACPA)-positive and -negative RA was abolished by IgG depletion, but restored only with ACPA-positive IgG. NETosis-derived products in RA serum demonstrated diagnostic potential, the ROC area under the curve for cell-free nucleosomes being >97%, with a sensitivity of 91% and a specificity of 92%. No significant difference was observed between ACPA-positive and -negative cases.Conclusions: Signaling elements associated with the extrusion of NETs are significantly enhanced to promote NETosis in RA compared with healthy controls. NETosis depended on the presence of ACPA in ACPA-positive RA serum. The quantitation of NETosis-derived products, such as cell-free nucleosomes in serum, may be a useful complementary tool to discriminate between healthy controls and RA cases.
AB - Introduction: Neutrophil extracellular traps (NETs) have recently been implicated in a number of autoimmune conditions, including rheumatoid arthritis (RA). We examined the underlying signaling pathways triggering enhanced NETosis in RA and ascertained whether the products of NETosis had diagnostic implications or usefulness.Methods: Neutrophils were isolated from RA patients with active disease and from controls. Spontaneous NET formation from RA and control neutrophils was assessed in vitro with microscopy and enzyme-linked immunosorbent assay (ELISA) for NETosis-derived products. The analysis of the signal-transduction cascade included reactive oxygen species (ROS) production, myeloperoxidase (MPO), neutrophil elastase (NE), peptidyl arginine deiminase 4 (PAD4), and citrullinated histone 3 (citH3). NET formation was studied in response to serum and synovial fluid and immunoglobulin G (IgG) depleted and reconstituted serum. Serum was analyzed for NETosis-derived products, for which receiver operator characteristic (ROC) curves were calculated.Results: Neutrophils from RA cases exhibited increased spontaneous NET formation in vitro, associated with elevated ROS production, enhanced NE and MPO expression, nuclear translocation of PAD4, PAD4-mediated citrullination of H3, and altered nuclear morphology. NET formation in both anti-citrullinated peptide antibody (ACPA)-positive and -negative RA was abolished by IgG depletion, but restored only with ACPA-positive IgG. NETosis-derived products in RA serum demonstrated diagnostic potential, the ROC area under the curve for cell-free nucleosomes being >97%, with a sensitivity of 91% and a specificity of 92%. No significant difference was observed between ACPA-positive and -negative cases.Conclusions: Signaling elements associated with the extrusion of NETs are significantly enhanced to promote NETosis in RA compared with healthy controls. NETosis depended on the presence of ACPA in ACPA-positive RA serum. The quantitation of NETosis-derived products, such as cell-free nucleosomes in serum, may be a useful complementary tool to discriminate between healthy controls and RA cases.
UR - http://www.scopus.com/inward/record.url?scp=84902057700&partnerID=8YFLogxK
U2 - 10.1186/ar4579
DO - 10.1186/ar4579
M3 - Article
C2 - 24928093
AN - SCOPUS:84902057700
SN - 1478-6354
VL - 16
JO - Arthritis Research and Therapy
JF - Arthritis Research and Therapy
IS - 3
M1 - R122
ER -