Characterization of F-actin polymerisation as a major toxic event induced by coal fly-ash

P Brown, T Jones, H Al-Bulushi, R Adams & K BéruBé

A multi-disciplinary approach was taken to study whether differences in particle composition and morphology have significant effects on particle-cell interactions. Coal fly-ash (CFA), a by-product from the combustion of coal, was chosen as the subject of interest along with a range of controls including carbon black (CB), DQ12 quartz, and M60 mullite. The CFA used in this study was sourced from coal-burning power stations in the UK, Poland and China. Prior to toxicological investigation, the particles were first physicochemically characterised by Inductively Coupled Plasma-Mass Spectroscopy (ICP-MS), Field Emission-Scanning Electron Microscopy/Energy Dispersive X-ray Microanalysis (FE-SEM/EDX) and X-ray Diffraction (XRD) analysis. Particle suspensions (0-0.1mg/ml in phosphate buffered saline; PBS) were incubated with whole human blood for 2 minutes, prior to red cell lysis, and fixation and permeabilisation of leukocytes (white blood cells; WBCs).  Intracellular F-actin was stained with the fluorescent molecular dye-FITC-phalloidin. Intracellular F-actin content was determined by measurement of mean fluorescence intensity (MFI) using a flow cytometer. All measurements were compared to a basal level of activity seen in the absence of particles, and the results confirmed that an acute response had occurred within the short exposure time for all particle types. Confocal microscopy supported these findings.

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