Exogenous fatty acids modulate the functional and cytotoxic responses of cultured pulmonary artery endothelial cells to oxidant stress

We previously reported that supplementation with exogenous fatty acids modulated the susceptibility of cultured pulmonary artery endothelial cells (PAEC) to oxidant-mediated cytotoxicity. The current study investigates the effects of fatty acids with increasing degrees of unsaturation on oxidant-mediated dysfunction and cytotoxicity in cultured porcine pulmonary artery and aortic endothelial cells (AEC). Monolayers supplemented with 0.1 mmol/L oleic (18:1), linoleic (18:2), or γ-linolenic (18:3) acids were exposed to oxidant stress (100 μmol/L hydrogen peroxide [H₂O₂)) or to control conditions for 30 minutes. Gas chromatographic analysis of the PAEC fatty acids confirmed incorporation of supplemental fatty acids into PAEC lipids. Cytotoxicity, measured as the release of intracellular lactate dehydrogenase (LDH), and PAEC monolayer barrier function, assessed by measuring the monolayer clearance of Evans blue dye bound to albumin, were determined for 1 to 3 hours after oxidant stress. The PAEC and AEC demonstrated comparable responses to H₂O₂. Hydrogen peroxide caused increases in monolayer permeability and detachment of cells from the monolayer that were most attenuated by supplementation with 18:2 or 18:3, and to a lesser degree with 18:1. In contrast, H₂O₂-mediated LDH release was attenuated by supplementation with 18:1, whereas 18:2 and 18:3 potentiated cytotoxicity after exposure to H₂O₂. These results indicate that the relationship between PAEC lipid composition and oxidant susceptibility is complex and that the extent of fatty acid unsaturation does not predict the functional or cytotoxic responses of PAEC to oxidant stress. Furthermore, these results suggest that functional derangements may not correlate with traditional assays of cytotoxicity induced by oxidant injury in cultured endothelium.