Effects of dietary polyunsaturated fatty acids and oxylipins on human EA.hy926 endothelial cells
Endothelial cells line the inner surface of all blood vessels and play a central role in maintaining vasculature homeostasis. Dietary polyunsaturated fatty acids (PUFAs) and their oxygenated derivatives, oxylipins, have been shown to modulate endothelial cell function in every aspect. The overall hypothesis of this thesis is that dietary PUFAs and oxylipins affect the viability and proliferation of EA.hy926 human endothelial cells through a differential activation of mitogen-activated protein kinases (MAPKs) in growing and confluent (quiescent) states. Testing of this hypothesis was achieved by examining the effects of major PUFAs (linoleic acid [LA], arachidonic acid [AA], α-linolenic acid [ALA], eicosapentaenoic acid [EPA] and docosahexaenoic acid [DHA]) and the effects of lipoxygenase (LOX) inhibition on growing and confluent EA.hy926 human endothelial cell viability and proliferation, and MAPK-related cell signaling. The results have shown that both substrate and growth state are important factors influencing the differential responses of endothelial cells to PUFAs. Specifically, cells became quiescent after reaching confluence when they were grown on Matrigel-coated plates, but not collagen-coated or plastic plates. AA, EPA, and DHA decreased the viability and proliferation rate of both growing and confluent cells in a dose-dependent manner, while LA and ALA had no effect. DHA induced apoptosis of confluent but not growing endothelial cells through a MAPK and caspase-3 dependent mechanism. Activation of MAPKs (p38, ERK1/2, JNK) by DHA followed a temporal sequence and was growth state-dependent. Treatment of the cells with PD146176 resulted in a reduction of viability in both growing and confluent cells, and a decline of proliferation in growing cells. However, these effects were not due to inhibition of 15-LOX, the reported target of this compound. Rather, the oxylipin profile of PD146176-treated cells suggested CYP-derived oxylipins may mediate the cellular action of DHA. Interestingly, PD146176 operated through a p38MAPK, Akt and PPARα-dependent pathway. In conclusion, the major finding of this thesis is that endothelial cell responses to PUFAs differ depending their growth state and presence of extracellular matrix. This may have significant clinical implications since dietary PUFA consumption could differentially affect healthy and dysfunctional endothelium, thus, influencing vascular homeostasis.
Dietary polyunsaturated fatty acids, Oxylipins, Human endothelial cells