A novel 4-dimensional live-cell imaging system to study leukocyte-endothelial dynamics in ANCA-associated vasculitis

Walls, C. A., Basu, N. , Hutcheon, G., Erwig, L. P., Little, M. A. and Kidder, D. (2020) A novel 4-dimensional live-cell imaging system to study leukocyte-endothelial dynamics in ANCA-associated vasculitis. Autoimmunity, 53(3), pp. 148-155. (doi: 10.1080/08916934.2019.1704274) (PMID:31865776)

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Neutrophils, monocytes and the endothelium are critical to ANCA-associated vasculitis (AAV) pathogenesis. This study aimed to develop a 4-dimensional (4D) live-cell imaging system that would enable investigation of spatial and temporal dynamics of these cells in health and disease. We further aimed to validate this system using autologous donor serum from AAV patients and polyclonal ANCA IgG, as well as exploring its potential in the pre-clinical testing of putative therapeutic compounds. Neutrophils and monocytes were isolated from peripheral venous blood of AAV patients or healthy controls and co-incubated on an endothelial monolayer in the presence of autologous serum. Alternatively, polyclonal ANCA IgG was used, following TNF-α priming, and imaged in 4-dimensions for 3 h using a spinning disc confocal microscope. Volocity 6.3® analysis software was used for quantification of leukocyte dynamics. The use of autologous serum resulted in increased neutrophil degranulation (p = .002), transmigration (p = .0096) and monocyte transcellular transmigration (p = .0013) in AAV patients. Polyclonal MPO-ANCA IgG induced neutrophil degranulation (p < .001) in this system. C5aR1 antagonism reduced neutrophil degranulation (p < .0002). We have developed a novel 4D in vitro system that allows accurate quantification of multiple neutrophil- and monocyte-endothelial interactions in AAV in a single assay. This system has the potential to highlight dynamics key to pathophysiology of disease, as well investigating the impact of potential therapeutics on these functions.

Item Type:Articles
Keywords:Immunology, immunology and allergy.
Glasgow Author(s) Enlighten ID:Basu, Professor Neil
Authors: Walls, C. A., Basu, N., Hutcheon, G., Erwig, L. P., Little, M. A., and Kidder, D.
College/School:College of Medical Veterinary and Life Sciences > Institute of Infection Immunity and Inflammation
Journal Name:Autoimmunity
Publisher:Taylor & Francis
ISSN (Online):1607-842X
Published Online:23 December 2019
Copyright Holders:Copyright © 2019 Informa UK Limited, trading as Taylor and Francis Group
First Published:First published in Autoimmunity 53(3):148-155
Publisher Policy:Reproduced in accordance with the publisher copyright policy

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