Lipophilic Chemicals from Diesel Exhaust Particles Trigger Calcium Response in Human Endothelial Cells via Aryl...

Exposure to diesel exhaust particles (DEPs) affects endothelial function and may contribute to the development of atherosclerosis and vasomotor dysfunction. As intracellular calcium concentration [Ca²⁺] is considered important in myoendothelial signalling, we explored the effects of extractable organic matter from DEPs (DEP-EOM) on [Ca²⁺] and membrane microstructure in endothelial cells. DEP-EOM of increasing polarity was obtained by pressurized sequential extraction of DEPs with n-hexane (n-Hex-EOM), dichloromethane (DCM-EOM), methanol, and water. Chemical analysis revealed that the majority of organic matter was extracted by the n-Hex- and DCM-EOM, with polycyclic aromatic hydrocarbons primarily occurring in n-Hex-EOM. The concentration of calcium was measured in human microvascular endothelial cells (HMEC-1) using micro-spectrofluorometry. The lipophilic n-Hex-EOM and DCM-EOM, but not the more polar methanol- and water-soluble extracts, induced rapid [Ca²⁺] increases in HMEC-1. n-Hex-EOM triggered [Ca²⁺] increase from intracellular stores, followed by extracellular calcium influx consistent with store operated calcium entry (SOCE). By contrast, the less lipophilic DCM-EOM triggered [Ca²⁺] increase via extracellular influx alone, resembling receptor operated calcium entry (ROCE). Both extracts increased [Ca²⁺] via aryl hydrocarbon receptor (AhR) non-genomic signalling, verified by pharmacological inhibition and RNA-interference. Moreover, DCM-EOM appeared to induce an AhR-dependent reduction in the global plasma membrane order, as visualized by confocal fluorescence microscopy. DCM-EOM-triggered [Ca²⁺] increase and membrane alterations were attenuated by the membrane stabilizing lipid cholesterol. In conclusion, lipophilic constituents of DEPs extracted by n-hexane and DCM seem to induce rapid AhR-dependent [Ca²⁺] increase in HMEC-1 endothelial cells, possibly involving both ROCE and SOCE-mediated mechanisms. The semi-lipophilic fraction extracted by DCM also caused an AhR-dependent reduction in global membrane order, which appeared to be connected to the [Ca²⁺] increase.