Protective effects of epoxyeicosatrienoic acids on human endothelial cells from the pulmonary and coronary vasculature.

In contrast to its constrictor effects on peripheral arteries, 20-hydroxyeicosatetraenoic acid (20-HETE) is an endothelial-dependent dilator of pulmonary arteries (PAs). The present study examined the hypothesis that the vasodilator effects of 20-HETE in PAs are due to an elevation of intracellular calcium concentration ([Ca(2+)](i)) and the release of nitric oxide (NO) from bovine PA endothelial cells (BPAECs). BPAECs express cytochrome P-450 4A (CYP4A) protein and produce 20-HETE. 20-HETE dilated PAs preconstricted with U-46619 or norepinephrine and treated with the cytochrome P-450 inhibitor 17-octadecynoic acid and the cyclooxygenase inhibitor indomethacin. The dilator effect of 20-HETE was blocked by the NO synthase inhibitor N(omega)-nitro-L-arginine methyl ester (L-NAME) or by removal of endothelium. 20-HETE significantly increased [Ca(2+)](i) and NO production in BPAECs. 20-HETE-induced NO release was blunted by removal of extracellular calcium, as well as NO synthase inhibitors (L-NAME). These results suggest that 20-HETE dilates PAs at least in part by increasing [Ca(2+)](i) and NO release in BPAECs.

The arachidonic acid derivative, 20‐hydroxyeicosatetraenoic acid (20‐HETE) dilates small pulmonary arteries (~1–3 mm in diameter). We recently showed that 20‐HETE increases generation of superoxide in bovine pulmonary artery endothelial cells (BPAECs) via activation of NADPH oxidase. Superoxide is known to stimulate growth of endothelial cells and activate protein kinases including ERK, p38MAPK, and Akt. We therefore tested if 20‐HETE promoted survival of BPAEC from apoptotic cell death. 20‐HETE (10 nM – 1 microM) produced a concentration‐dependent increase in numbers of BPAECs after serum deprivation and attenuated increases in caspase 3 activity, indicating protection against apoptosis. This pro‐survival action of 20‐HETE was abolished by pharmacological inhibition of phospho‐inositide 3‐kinase and Akt. Lipopolysaccharide (LPS) induced apoptosis in BPAECs as determined by nuclear fragmentation and increased activity of caspase 3 was effectively inhibited by 20‐HETE (1 microM). Further, 20‐HETE also reduced caspase 3 activity in ex vivo cultures of small pulmonary arteries from mouse and exposed to hypoxia followed by reoxygenation. In summary, 20‐HETE protects the pulmonary vasculature from apoptosis induced my multiple insults such as serum deprivation, LPS and hypoxia/reoxygenation. Funding: NIH/NHLBI 68627 (ERJ), 49294 (ERJ), 69996 (MMM), GM 31278 (JRF).