20-HETE Upregulates Endothelial Angiotensin Converting Enzyme (ACE): Contribution to 20-HETEDependent Hypertension, Vascular Dysfunction and Microvascular Remodeling

Increased vascular 20-hydroxyeicosatetraenoic acid (20-HETE), a cytochrome P450-arachidonic acid metabolite, promotes vascular dysfunction and hypertension that is dependent, in part, on the renin angiotensin system (RAS). Studies in our laboratory identified 20-HETE as an inducer of angiotensin converting enzyme (ACE) in cultured human microvascular endothelial cells (HMVEC) and suggested that 20-HETE-mediated induction of ACE contributes to 20-HETE-mediated hypertension, vascular dysfunction and microvascular remodeling. The objective of this dissertation was to determine the mechanisms by which 20-HETE induces ACE in vitro and the contribution of ACE and RAS to 20-HETE-mediated hypertension and vascular complications. We showed that in HMVEC 20-HETE induces ACE mRNA, protein and activity via a distinct EGFR-MAPK-IKK signaling pathway. These effects were abrogated by co-treatment with 20-HEDE, a 20-HETE antagonist. Additional experiments further demonstrated that 20-HETE transcriptionally activates ACE via NF-kB binding to the somatic and germinal promoters of the ACE genes. The 20-HETE-stimulated promoter activation was also abrogated by 20-HEDE and was dependent on EGFR, MAPK, IKK? and NF-kB activation. Animal studies using the Cyp4a12tg mice, whose Cyp4a12-20-HETE synthase expression is under the control of a tetracycline (doxycycline, DOX) promoter, were used to assess the contribution of ACE/RAS to vascular dysfunction and remodeling in 20-HETE-dependent hypertension. Treatment of Cyp4a12tg mice with DOX increased systolic blood pressure (SBP, 136+/-2 vs 102+/-1 mmHg). This increase was prevented by administration of either the 20-HETE antagonist 20-HEDGE, lisinopril or losartan. DOX-induced hypertension was associated with dysfunction and remodeling of preglomerular microvessels, which was prevented by 20-HEDGE, yet only lessened but not prevented by lisinopril or losartan illustrating 20-HETE's actions independent of ACE activation. In ACE 3/3 mice, which lack vascular endothelial ACE, administration of 5?-dihydrotestosterone (DHT), a known inducer of 20-HETE production, increased SBP; however, the increase was about 50% of that in WT mice (103+/-4 vs 151+/-1 and 104+/-2 vs 126+/-1 mmHg WT and ACE 3/3 treated mice, respectively) Losartan and 20-HEDGE prevented the DHT-induced increase in SBP in WT and ACE 3/3 mice. DHT treatment increased 20-HETE production and microvascular remodeling in both WT and ACE 3/3 mice; however, remodeling in the ACE 3/3 mice was tempered compared to WT mice. 20-HEDGE prevented microvascular remodeling in WT and ACE 3/3 mice, while losartan had no effect on remodeling in ACE 3/3. The Cyp4a14(-/-) mouse model is a model of male-specific androgen-driven 20-HETE-mediated hypertension. Male Cyp4a14(-/-) mice are hypertensive and exhibit increased vascular ACE and p-IkB expression compared to female Cyp4a14(-/-) mice. Administration of the water soluble 20-HETE antagonist, 20-SOLA, reduced blood pressure in Cyp4a14(-/-) male mice in 10 days. The reduction in blood pressure was associated with decreases in vascular ACE and p-IkB expression. Taken together, these results indicate that 20-HETE transcriptionally activates ACE and that the RAS partially contributes to the 20-HETE-mediated microvascular dysfunction and remodeling in hypertension. In addition, 20-HETE-driven microvascular dysfunction and remodeling take place independent of blood pressure elevation and does not fully rely on ACE activity in the vascular endothelium. Lastly, the use of new novel water-soluble 20-HETE antagonists such as 20-SOLA can serve as new tools to explore 20-HETE's biological functions and roles in disease states such as hypertension.