Cellular activation and inflammation leading to endothelial dysfunction is associated with cardiovascular disease (CVD). We investigated whether a single cell label-free multi parameter optical interrogation system can detect endothelial cell and endothelial progenitor cell (EPC) activation in vitro and ex vivo, respectively. Cultured human endothelial cells were exposed to increasing concentrations of tumour necrosis factor alpha (TNF-α) or lipopolysaccharide (LPS) before endothelial activation was validated using fluorescence-activated cell sorting (FACS) analysis of inflammatory marker expression (PECAM-1, E-selectin and ICAM-1). A centrifugal microfluidic system and V-cup array was used to capture individual cells before optical measurement of light scattering, immunocytofluorescence, auto-fluorescence (AF) and cell morphology was determined. In vitro, TNF-α promoted specific changes to the refractive index and cell morphology of individual cells concomitant with enhanced photon activity of fluorescently labelled inflammatory markers and increased auto-fluorescence (AF) intensity at three different wavelengths, an effect blocked by inhibition of downstream signalling with Iκβ. Ex vivo, there was a significant increase in EPC number and AF intensity of individual EPCs from CVD patients concomitant with enhanced PECAM-1 expression when compared to normal controls. This novel label-free ‘lab on a disc’ (LoaD) platform can successfully detect endothelial activation in response to inflammatory stimuli in vitro and ex vivo.
Science Foundation Ireland (SFI) and Fraunhofer-Gesellschaft under the SFI Strategic Partnership Programme [Grant Number 16/SPP/3321], Science Foundation Ireland [Grant Number: 10/CE/B1821], ERDF, LiPhos project funded by the European Commission [Grant Number: 317916], Science Foundation Ireland [Grant number: 11/PI/1181], Health Research Board (HRB) of Ireland [Grant Number: HRA-POR-2015-1315].
ID Code:
27460
Deposited On:
29 Jul 2022 16:34 by
Thomas Murtagh
. Last Modified 10 Jan 2023 15:15