Soluble epoxide hydrolase regulates hydrolysis of vasoactive epoxyeicosatrienoic acids.

The cytochrome P450-derived epoxyeicosatrienoic acids (EETs) have potent effects on renal vascular reactivity and tubular sodium and water transport; however, the role of these eicosanoids in the pathogenesis of hypertension is controversial. The current study examined the hydrolysis of the EETs to the corresponding dihydroxyeicosatrienoic acids (DHETs) as a mechanism for regulation of EET activity and blood pressure. EET hydrolysis was increased 5- to 54-fold in renal cortical S9 fractions from the spontaneously hypertensive rat (SHR) relative to the normotensive Wistar-Kyoto (WKY) rat. This increase was most significant for the 14,15-EET regioisomer, and there was a clear preference for hydrolysis of 14, 15-EET over the 8,9- and 11,12-EETs. Increased EET hydrolysis was consistent with increased expression of soluble epoxide hydrolase (sEH) in the SHR renal microsomes and cytosol relative to the WKY samples. The urinary excretion of 14,15-DHET was 2.6-fold higher in the SHR than in the WKY rat, confirming increased EET hydrolysis in the SHR in vivo. Blood pressure was decreased 22+/-4 mm Hg (P:<0.01) 6 hours after treatment of SHRs with the selective sEH inhibitor N:, N:'-dicyclohexylurea; this treatment had no effect on blood pressure in the WKY rat. These studies identify sEH as a novel therapeutic target for control of blood pressure. The identification of a potent and selective inhibitor of EET hydrolysis will be invaluable in separating the vascular effects of the EET and DHET eicosanoids.

Increased CYP2J expression and epoxyeicosatrienoic acid formation in spontaneously hypertensive rat kidney.

Epoxyeicosatrienoic acids (EETs) are major products of cytochrome P450 (CYP)-catalyzed metabolism of arachidonic acid in the kidney. The potent effect of EETs on renal vascular tone and tubular ion and water transport implicates their role in the regulation of renal function and blood pressure. The present study was designed to test the hypothesis that CYP-catalyzed EET formation was altered in the spontaneously hypertensive rat (SHR) kidney. The formation of 14,15- and 11,12-EET was approximately 2-fold higher in incubations of arachidonic acid with SHR renal cortical microsomes relative to microsomes from normotensive Wistar-Kyoto (WKY) rats. This was consistent with increased expression of a CYP2J2 immunoreactive protein in the SHR cortex and outer medulla. In contrast, there was no significant difference in the levels of the CYP2E and CYP2C epoxygenases in SHR and WKY kidneys. Protein and RNA analysis suggests that the CYP2J2 immunoreactive protein that is overexpressed in the SHR kidney is distinct from the known rat CYP2J isoforms. EET formation also was documented in vivo from measurements of urinary EET excretion. Importantly, the excretion rates of 14,15-, and 11,12-EETs were 2.5- and 1.8-fold higher, respectively, in SHR than WKY kidney. These studies provide both in vitro and in vivo evidence for increased EET formation in the SHR kidney and identify a novel CYP2J2 immunoreactive protein that is differentially expressed in the hypertensive kidney. In light of the known biological properties of the EETs, these findings may be important in elucidating the mechanisms that control renal vascular tone and tubular ion transport in the SHR.

Developmentally regulated expression of the CYP4A genes in the spontaneously hypertensive rat kidney.

The CYP4A enzymes catalyze the formation of 20-hydroxyeicosatetraenoic acid (20-HETE), which has potent effects on the renal vasculature and tubular ion transport. Based on an increased 20-HETE formation in renal microsomes from spontaneously hypertensive rats, it has been proposed that increased expression of the CYP4A genes is an early event in the development of hypertension in these animals. To test this hypothesis, we developed RNase protection assays for specific detection of the individual CYP4A genes in the kidneys of spontaneously hypertensive and Wistar-Kyoto rats. Distinct age-dependent patterns of expression were observed for the individual CYP4A genes, with only CYP4A3 mRNA measurable in the kidneys of 1-week-old rats. CYP4A1 and CYP4A8 mRNA were detectable by 3 weeks of age and CYP4A2 mRNA at 5 weeks of age. The expression of CYP4A1 and CYP4A3 varied 4-5-fold throughout development and was highest between 3 and 5 weeks of age, declining steadily thereafter to 20% of their maximal level by 9 weeks of age. CYP4A2 mRNA levels increased steadily between 5 and 9 weeks of age, whereas CYP4A8 mRNA levels were relatively constant throughout development. The CYP4A3 mRNA level was significantly increased 1. 6-2-fold in the cortex and outer medulla of 1-4-week-old spontaneously hypertensive rat kidneys relative to the corresponding level in the Wistar-Kyoto. A similar 1.4-1.7-fold increase in CYP4A8 mRNA was also found in 3- and 4-week-old spontaneously hypertensive kidneys. Accompanying the increased expression of CYP4A3 and CYP4A8 mRNA in the prehypertensive rats were corresponding changes in functional CYP4A measured as either arachidonic acid or lauric acid omega-hydroxylase activity (1.4-2.0-fold increases) and CYP4A protein levels. After 4 weeks of age, the level of CYP4A mRNA, enzyme activity, and protein were similar in the kidneys of Wistar-Kyoto and spontaneously hypertensive rats. The findings suggest that the expression of CYP4A3 and CYP4A8 may be critical to the early changes in eicosanoid formation and renal function in the young spontaneously hypertensive rat.