Pravastatin Therapy and Biomarker Changes in Children and Young Adults with Autosomal Dominant Polycystic Kidney Disease.

BACKGROUND AND OBJECTIVES: Disease-specific treatment options for autosomal dominant polycystic kidney disease are limited. Clinical intervention early in life is likely to have the greatest effect. In a 3-year randomized double-blind placebo-controlled phase 3 clinical trial, the authors recently showed that pravastatin decreased height-corrected total kidney volume (HtTKV) progression of structural kidney disease over a 3-year period. However, the underlying mechanisms have not been elucidated. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: Participants were recruited nationally from July 2007 through October 2009. Plasma and urine samples collected at baseline, 18 months, and 36 months from 91 pediatric patients enrolled in the above-mentioned clinical trial were subjected to mass spectrometry-based biomarker analysis. Changes in biomarkers over 3 years were compared between placebo and pravastatin-treated groups. Linear regression was used to evaluate the changes in biomarkers with the percent change in HtTKV over 3 years. RESULTS: Changes in plasma concentrations of proinflammatory and oxidative stress markers (9- hydroxyoctadecadienoic acid, 13-hydroxyoctadecadienoic acid, and 15-hydroxyeicosatetraenoic acid [HETE]) over 3 years were significantly different between the placebo and pravastatin-treated groups, with the pravastatin group showing a lower rate of biomarker increase. Urinary 8-HETE, 9-HETE, and 11-HETE were positively associated with the changes in HtTKV in the pravastatin group. CONCLUSIONS: Pravastatin therapy diminished the increase of cyclooxygenase- and lipoxygenase-derived plasma lipid mediators. The identified biomarkers and related molecular pathways of inflammation and endothelial dysfunction may present potential targets for monitoring of disease severity and therapeutic intervention of autosomal dominant polycystic kidney disease.

n-3 fatty acids reduce plasma 20-hydroxyeicosatetraenoic acid and blood pressure in patients with chronic kidney disease.

BACKGROUND: Metabolism of arachidonic acid by cytochrome P450 ω-hydroxylase leads to the formation of 20-hydroxyeicosatetraenoic acid (20-HETE) that regulates vascular function, sodium homeostasis and blood pressure (BP). Supplementation with n-3 fatty acids is known to alter arachidonic acid metabolism and reduce the formation of the lipid peroxidation products F2-isoprostanes, but the effect of n-3 fatty acids on 20-HETE has not been studied. METHOD: We previously reported a significant effect of n-3 fatty acids but not coenzyme Q10 (CoQ) to reduce BP in a double-blind, placebo-controlled intervention, wherein patients with chronic kidney disease (CKD) were randomized to n-3 fatty acids (4 g), CoQ (200 mg), both supplements or control (4 g olive oil), daily for 8 weeks. This study examined the effect of n-3 fatty acids on plasma and urinary 20-HETE in the same study, as well as plasma and urinary F2-isoprostanes, and relate these to changes in BP. RESULTS: Seventy-four patients completed the 8-week intervention. n-3 fatty acids but not CoQ significantly reduced plasma 20-HETE (P = 0.001) and F2-isoprostanes (P < 0.001). In regression models adjusted for BP at baseline, postintervention plasma 20-HETE was a significant predictor of the fall in SBP (P < 0.0001) and DBP (P < 0.0001) after n-3 fatty acids. CONCLUSION: This is the first report that n-3 fatty acid supplementation reduces plasma 20-HETE in humans and that this associates with reduced BP. These results provide a plausible mechanism for the reduction in BP observed in patients with CKD following n-3 fatty acid supplementation.

The effect of a single nucleotide polymorphism of the CYP4F2 gene on blood pressure and 20-hydroxyeicosatetraenoic acid excretion after weight loss.

BACKGROUND: Genetic background partly determines the efficacy of interventions to lower blood pressure (BP). The CYP4F2 and CYP4A11 enzymes are renal 20-hydroxyeicosatetraenoic acid (20-HETE) synthases that regulate BP. Gene variants of CYP4F2 and CYP4A11 associate with hypertension and stroke. We showed that a gene variant of CYP4F2 but not CYP4A11 was associated with increased 20-HETE excretion and BP. AIM: To compare BP and 20-HETE responses in carriers of the CYP4F2 1347G/A polymorphism and controls CYP4F2-GG (wildtype), during weight loss. METHODS: Volunteers genotyped as CYP4F2GA/AA (n = 26) and controls genotyped as CYP4F2 GG (n = 27) were counselled to reduce weight for 12 weeks, followed by 4 weeks of weight stabilization. Weight, 24-h BP, pulse pressure and urinary 20-HETE were measured at baseline, 12 and 16 weeks. RESULTS: At baseline, SBP was (+1.7 mmHg, P = 0.047) in the CYP4F2 GA/AA genotype. Compared with baseline, weight fell by 3.9 kg, P = 0.0001, in both genotypes, and was maintained to 16 weeks. SBP fell by (-7.6 mmHg, P = 0.004) in both genotypes after 12 weeks. However, after weight stabilization, SBP was +3.6 mmHg, P = 0.004 in CYP4F2 GA/AA genotype. DBP and heart rate changed similarly over time. Pulse pressure fell with weight loss (P < 0.001), but was elevated in the CYP4F2 GA/AA genotype at all time-points (+3.1 mmHg, P < 0.001). Urinary 20-HETE was similar at baseline and 12 weeks but elevated in the CYP4F2 GA/AA genotype (P = 0.017) after weight stabilization. CONCLUSION: Maintenance of lower BP after weight loss is more difficult for carriers of the CYP4F2 G1347A polymorphism and may be related to increased arterial stiffness and increased 20-HETE synthesis.

Lipoxygenase products in the urine correlate with renal function and body temperature but not with acute transplant rejection.

Acute transplant rejection is the leading cause of graft loss in the first months after kidney transplantation. Lipoxygenase products mediate pro- and anti-inflammatory actions and thus we aimed to correlate the histological reports of renal transplant biopsies with urinary lipoxygenase products concentrations to evaluate their role as a diagnostic marker. This study included a total of 34 kidney transplant recipients: 17 with an acute transplant rejection and 17 controls. LTE4, LTB4, 12-HETE and 15-HETE concentrations were measured by enzyme immunoassay. Urinary lipoxygenase product concentrations were not significantly changed during an acute allograft rejection. Nevertheless, LTB4 concentrations correlated significantly with the body temperature (P ≤ 0.05) 3 months after transplantation, and 12- and 15-HETE concentrations correlated significantly with renal function (P ≤ 0.05) 2 weeks after transplantation. In conclusion, our data show a correlation for LTB4 with the body temperature 3 months after transplantation and urinary 12- and 15-HETE concentrations correlate positively with elevated serum creatinine concentrations but do not predict acute allograft rejection.

Disturbed ratio of renal 20-HETE/EETs is involved in androgen-induced hypertension in cytochrome P450 4F2 transgenic mice.

We have previously established a cytochrome P450 4F2 (CYP4F2) transgenic mouse model. The present study elucidated the molecular foundation of hypertension by androgen-induction in this model. The renal expression of CYP4F2 in transgenic mice was highly expressed and strongly induced with 5α-dihydrotestosterone (DHT) treatment determined by Western blot. DHT also increased the renal arachidonic acid ω-hydroxylation and urinary 20-hydroxyeicosatetraenoic acid (20-HETE) excretion (P<0.01), and furthermore elevated the systolic blood pressure by 10 and 22 mm Hg (P<0.05) in female and castrated male transgenic mice, respectively. HET0016 completely eliminated the androgen-induced effects (P<0.01). Endogenous Cyp4a ω-hydroxylases, evaluated by real-time quantitative PCR, were significantly suppressed in transgenic mice (P<0.05). Importantly, transgenic mice with increased 20-HETE showed decreased epoxyeicosatrienoic acids (EETs) and increased dihydroxyeicosatetraenoic acids determined by liquid chromatography-tandem mass spectrometry, contributing to significantly raised ratio of 20-HETE/EETs in the urine and kidney homogenate (P<0.01). These data demonstrate that the androgen aggravated hypertension possibly through an altered ratio of 20-HETE/EETs in CYP4F2 transgenic mice.

Evaluation of CMV and KAP promoters for driving the expression of human CYP4F2 in transgenic mice.

A transgenic mouse model in which cytochrome P450 4F2 (CYP4F2) was expressed in multiple organs was expected to clarify the role of 20-hydroxyeicosatetraenoic acid (20-HETE) in the regulation of blood pressure, compared with our previously established kidney androgen-regulated protein (KAP) promoter CYP4F2 transgenic mouse model which predominantly showed renal overexpression of CYP4F2. A novel CYP4F2 transgenic mouse model driven by the cyto-megalovirus (CMV) promoter was generated and identified by PCR and subsequent sequencing. Extensive study of CMV-CYP4F2 transgenic mice demonstrated that CYP4F2 was exclusively expressed in the liver, while 20-HETE levels in the urine, kidney and blood were not affected, and there was no resulting change in the systolic blood pressure. This was in contrast to KAP-CYP4F2 transgenic mice which exerted prohypertensive action of 20-HETE resulting from the renal overexpression of CYP4F2. In addition, CYP4F2 overwhelmed the endogenous renal Cyp4a family mRNA levels in the KAP-CYP4F2 but not in the CMV-CYP4F2 transgenic mice. These results support the idea that overexpression of renal CYP4F2, leading to high 20-HETE in the urine and blood, may account for the elevated blood pressure. The CMV promoter did not direct CYP4F2 expression into extensive tissues and organs in an attempt to clarify the action of 20-HETE.

Biomarkers of oxidative damage in cigarette smokers: which biomarkers might reflect acute versus chronic oxidative stress?

Cigarette smoking predisposes to the development of multiple diseases involving oxidative damage. We measured a range of oxidative damage biomarkers to understand which differ between smokers and nonsmokers and if the levels of these biomarkers change further during the act of smoking itself. Despite overnight abstinence from smoking, smokers had higher levels of plasma total and esterified F(2)-isoprostanes, hydroxyeicosatetraenoic acid products (HETEs), F(4)-neuroprostanes, 7-ketocholesterol, and 24- and 27-hydroxycholesterol. Levels of urinary F(2)-isoprostanes, HETEs, and 8-hydroxy-2'-deoxyguanosine were also increased compared with age-matched nonsmokers. Several biomarkers (plasma free F(2)-isoprostanes, allantoin, and 7ß-hydroxycholesterol and urinary F(2)-isoprostane metabolites) were not elevated. The smokers were then asked to smoke a cigarette; this acute smoking elevated plasma and urinary F(2)-isoprostanes, plasma allantoin, and certain cholesterol oxidation products compared to presmoking levels, but not plasma HETEs or urinary 8-hydroxy-2'-deoxyguanosine. Smokers showed differences in plasma fatty acid composition. Our findings confirm that certain oxidative damage biomarkers are elevated in smokers even after a period of abstinence from smoking, whereas these plus some others are elevated after acute smoking. Thus, different biomarkers do not measure identical aspects of oxidative stress.

Peripheral and central augmentation indexes in relation to the CYP4F2 polymorphisms in Chinese.

OBJECTIVE: Cytochrome (CYP) 4F2 isoform is a key metabolizing enzyme for the renal 20-hydroxyeicosatetraenoic acid (20-HETE), which, as an endogenous vasoconstrictor, may influence properties of the peripheral muscular arteries and arterioles. We, therefore, investigated the CYP4F2 polymorphisms in relation to arterial wave reflections, as measured by augmentation indexes (AIx) in Chinese. METHODS: We performed arterial measurements by SphygmoCor and genotyped three CYP4F2 polymorphisms (V433M, rs3093089, and rs3093098) by PCR-restriction fragment length polymorphism in 1421 participants enrolled in the JingNing Population study. A replication study for the V433M polymorphism was performed in 924 Chinese recruited from a workplace setting. Urinary 20-HETE concentration was determined by ELISA in a randomly selected subsample of 318 JingNing individuals. RESULTS: In spite of the fact that genetic associations were not significant (P ≥ 0.12) in all JingNing participants, there was significant (Pint ≤ 0.02) interaction of the V433M polymorphism with sex and pulse rate in relation to peripheral and central AIx. M433 allele carriers, compared with V433V homozygotes, had significantly greater peripheral (+5.0%, P = 0.0002) and central AIx (+3.2%, P = 0.001) in 693 men. The corresponding values were +2.7% (P = 0.04) and +1.9% (P = 0.04) in 490 individuals of the top tertile of pulse rate (≥ 76 beats/min), and were +4.0% (P = 0.02) and +3.3% (P = 0.02) in 315 replication participants with a pulse rate at least 76 beats/min. Urinary 20-HETE concentration was significantly higher (P = 0.002) in M433M (2.06 ng/ml) and V433M (1.13 ng/ml) individuals than in V433V homozygotes (0.98 ng/ml). CONCLUSION: The CYP4F2 V433M polymorphism is associated with the size of arterial wave reflections in male Chinese, or individuals with a faster pulse rate.

Inhibition of 20-hydroxyeicosatetraenoic acid synthesis using specific plant lignans: in vitro and human studies.

Sesamin, the major lignan found in sesame, has been shown to increase vitamin E levels by inhibiting its metabolism via the cytochrome P450 isozyme CYP4F2. CYP4F2 and CYP4A11 are the predominant human isoforms that synthesize 20-hydroxyeicosatetraenoic acid (20-HETE) from arachidonic acid. Considerable evidence suggests that 20-HETE may play a role in the pathogenesis of hypertension. We hypothesized that sesamin could be an inhibitor of 20-HETE synthesis. This study investigated the effects of sesamin on 20-HETE synthesis in vitro and the effect of sesame supplementation on plasma and urinary 20-HETE concentrations in humans. Human microsomes were used to investigate the potency and selectivity of sesamin inhibition of 20-HETE synthesis. Sesamin inhibited human renal and liver microsome 20-HETE synthesis with IC50 <20 micromol/L. It was selective toward CYP4F2 (IC50: 1.9 micromol/L) and had reduced activity toward CYP4A11 (IC50: >150 micromol/L), as well as cytochrome P epoxygenation of arachidonic acid (IC50: >50 micromol/L). In a randomized, controlled crossover trial, overweight men and women (n=33) consumed 25 g/d of sesame (approximately 50 mg/d of sesame lignan) or an isocaloric matched control for 5 weeks each. Relative to control, sesame supplementation resulted in a 28% decrease in plasma and a 32% decrease in urinary 20-HETE (P<0.001). Urinary sodium, potassium, and blood pressure were not affected. This study demonstrates for the first time that sesame supplementation in humans reduces the plasma and urinary levels of 20-HETE, likely via inhibition of CYP4F2 by sesame lignans. These results suggest that sesame lignans could be used for the investigation of potential roles of 20-HETE in humans.

Oxidative damage in dengue fever.

Oxidative stress may be important in the pathogenesis of dengue infection. Using accurate markers of oxidative damage, we assessed the extent of oxidative damage in dengue patients. The levels of hydroxyeicosatetraenoic acid products (HETEs), F(2)-isoprostanes (F(2)-IsoPs), and cholesterol oxidation products (COPs) were measured in 28 adult dengue patients and 28 age-matched study controls during the febrile, defervescent, and convalescent stages of infection. We compared the absolute and the percentage change in these markers in relation to key clinical parameters and inflammatory markers. The levels of total HETEs and total HETEs/arachidonate, total F(2)-IsoPs/arachidonate, and COPs/cholesterol were higher during the febrile compared to the convalescent level. Total HETEs correlated positively with admission systolic blood pressure (r=0.52, p<0.05), whereas an inverse relationship was found between 7beta-hydroxycholesterol and systolic and diastolic blood pressure (r=-0.61 and -0.59, respectively, p<0.01). The urinary F(2)-IsoP level was higher in urine during the febrile stage compared to the convalescent level. Despite lower total cholesterol levels during the febrile stage compared to convalescent levels, a higher percentage of cholesterol was found as COPs (7beta-, 24-, and 27-hydroxycholesterol). The levels of platelet-activating factor-acetylhydrolase activity, vascular cellular adhesion molecule-1, tumor necrosis factor-alpha, and high-sensitivity C-reactive protein were higher during the febrile stage compared to their convalescent levels (p<0.01). Markers of oxidative damage are altered during the various stages of dengue infection.

Overexpression of cytochrome P450 4F2 in mice increases 20-hydroxyeicosatetraenoic acid production and arterial blood pressure.

Cytochrome P450 4F2 (CYP4F2) activity is thought to be a factor in the pathogenesis of hypertension through its bioactive metabolite 20-hydroxyeicosatetraenoic acid (20-HETE). We previously found that a gain-in-function CYP4F2 variant in a Chinese cohort was associated with elevated urinary 20-HETE and hypertension. To further explore this association we generated a transgenic mouse model expressing CYP4F2 driven by a modified mouse kidney androgen-regulated protein promoter. This heterologous promoter regulated the expression of luciferase and his-tagged CYP4F2 in transfected HEK 293 cells. In the kidney of transgenic mice, CYP4F2 was localized to renal proximal tubule epithelia and was expressed at a higher level than in control mice, leading to increased urinary 20-HETE excretion. Assessment of CYP4F2 activity by an arachidonic acid hydroxylation assay showed that 20-HETE production was significantly higher in kidney microsomes of transgenic mice compared to control mice, as was their systolic blood pressure. There was a positive correlation of blood pressure with urinary 20-HETE levels. Our results show that increased expression of CYP4F2 in mice enhanced 20-HETE production and elevated blood pressure.

Coenzyme Q10 improves endothelial dysfunction in statin-treated type 2 diabetic patients.

OBJECTIVE: The vascular benefits of statins might be attenuated by inhibition of coenzyme Q(10) (CoQ(10)) synthesis. We investigated whether oral CoQ(10) supplementation improves endothelial dysfunction in statin-treated type 2 diabetic patients. RESEARCH DESIGN AND METHODS: In a double-blind crossover study, 23 statin-treated type 2 diabetic patients with LDL cholesterol <2.5 mmol/l and endothelial dysfunction (brachial artery flow-mediated dilatation [FMD] <5.5%) were randomized to oral CoQ(10) (200 mg/day) or placebo for 12 weeks. We measured brachial artery FMD and nitrate-mediated dilatation (NMD) by ultrasonography. Plasma F(2)-isoprostane and 24-h urinary 20-hydroxyeicosatetraenoic acid (HETE) levels were measured as systemic oxidative stress markers. RESULTS: Compared with placebo, CoQ(10) supplementation increased brachial artery FMD by 1.0 +/- 0.5% (P = 0.04), but did not alter NMD (P = 0.66). CoQ(10) supplementation also did not alter plasma F(2)-isoprostane (P = 0.58) or urinary 20-HETE levels (P = 0.28). CONCLUSIONS: CoQ(10) supplementation improved endothelial dysfunction in statin-treated type 2 diabetic patients, possibly by altering local vascular oxidative stress.

20-HETE and F2-isoprostanes in the metabolic syndrome: the effect of weight reduction.

20-Hydroxyeicosatetraenoic acid (20-HETE) is a cytochrome P450 metabolite of arachidonic acid that regulates vascular function and sodium homeostasis. Studies showing an association between 20-HETE excretion, raised BMI, and oxidative stress suggest that 20-HETE may be important in the development of cardiovascular disease in the metabolic syndrome (MetS). We investigated whether 20-HETE and F(2)-isoprostanes (markers of oxidative stress) were altered in the MetS before and after weight reduction. A case-controlled comparison of 30 participants with the MetS and matched controls showed that plasma and urinary 20-HETE and F(2)-isoprostanes were significantly elevated in the MetS group. There was a significant gender x group interaction such that women with the MetS had higher urinary 20-HETE and F(2)-isoprostanes compared to controls (p<0.0001). In a randomized controlled trial, 42 participants with the MetS were assigned to 16 weeks of weight maintenance or a 12-week weight-loss program followed by 4 weeks weight stabilization. Relative to the weight-maintenance group, a 4-kg loss in weight resulted in a 2-mm Hg fall in blood pressure (BP) but did not alter urinary or plasma 20-HETE or F(2)-isoprostanes. 20-HETE and oxidative stress may be important mediators of cardiovascular disease risk in the MetS. Although a 4% reduction in body weight reduced BP, there were no changes in plasma or urinary 20-HETE or F(2)-isoprostanes.

A single nucleotide polymorphism in the CYP4F2 but not CYP4A11 gene is associated with increased 20-HETE excretion and blood pressure.

Arachidonic acid is a major fatty acid that can be metabolized by the cytochrome P450 enzyme to a number of bioactive eicosanoids. A major metabolite of this oxidation is 20-hydroxyeicosatetraenoic acid, which acts as a potent vasoconstrictor. However, in the kidney, its vasoconstrictor actions can be offset by its natriuretic properties. A guanine-to-adenine polymorphism in the CYP4F2 gene was associated with a reduction in 20-hydroxyeicosatetraenoic acid production in vitro. A thymidine-to-cytosine polymorphism in the CYP4A11 gene reduced catalytic activity by >50% in vitro and was associated with hypertension. The aim was to determine whether these 2 mutations are associated with urinary 20-hydroxyeicosatetraenoic acid excretion and blood pressure in humans. For the CYP4F2, 51% were homozygous for the G allele, 40% were carriers, and 9% were homozygous for the A allele. For CYP4A11, 72% were homozygous for the T allele, 25% were carriers, and 3% were homozygous for the C allele. The CYP4F2 GA/AA genotype was significantly associated with an increase in both 20-hydroxyeicosatetraenoic acid excretion and systolic blood pressure. The CYP4A11 CC/TC genotype was significantly associated with a reduction in 20-hydroxyeicosatetraenoic acid excretion but was not associated with blood pressure. We have demonstrated for the first time in humans that polymorphisms of the CYP4F2 and CYP4A11 genes have opposite effects on 20-hydroxyeicosatetraenoic acid excretion. The positive association between the CYP4F2 GA/AA genotype and both systolic blood pressure and 20-hydroxyeicosatetraenoic acid excretion strengthens a role for 20-hydroxyeicosatetraenoic acid in the modulation of blood pressure.

Association of a functional cytochrome P450 4F2 haplotype with urinary 20-HETE and hypertension.

Cytochrome P450 4F2 (CYP4F2) catalyzes the omega-hydroxylation of arachidonic acid to 20-hydroxyeicosatetraenoic acid (20-HETE), a natriuretic and vasoactive eicosanoid that participates in the development of hypertension. The relationship among CYP4F2 genetic variants in the regulatory region, formation of renal 20-HETE, and hypertension is unknown. Here are reported seven genetic variants around the CYP4F2 intronic regulatory region. Four of these variants made up two common haplotypes, Hap I (c.-91T/c.-48G/c.-13T/c.+34T) and Hap II (c.-91C/c.-48C/c.-13C/c.+34G). Hap I included a major functional variant, c.-91T-->C, which was identified by reporter assay and electrophoretic mobility shift assay. Transfected into HEK293 cells, the Hap I construct showed a trend toward higher basal transcriptional activity and exhibited significantly greater LPS-stimulated activity than Hap II; these findings were the result of different NF-kappaB binding affinity between the two constructs. In vivo, a case-control study demonstrated that homozygosity for Hap I doubled the risk for hypertension in a Chinese population, even after adjustment for risk factors including age, gender, and body mass index. This association was confirmed in a family-based association study. In addition, Hap I was associated with elevated urinary 20-HETE. These results indicate that a functional variant of the CYP4F2 regulatory region, which increases the binding affinity of NF-kappaB, increases the risk for hypertension, likely by modulating the production of 20-HETE.

Impact of genetic polymorphisms in CYP2C8 and rosiglitazone intake on the urinary excretion of dihydroxyeicosatrienoic acids.

UNLABELLED: The cytochrome P450 enzymes CYP2C8, CYP2C9 and CYP2J2 generate 8,9-, 11,12-, and 14,15-epoxyeicosatrienic acid (EET) from arachidonic acid, and these EETs are then hydrolyzed to dihydroxyeicosatrienoic acids (DHET) before excretion into the urine. It is unknown how genetic polymorphisms affect formation of these diuretic, vasodilatory and anti-inflammatory eicosanoids, and whether the CYP2C8 substrate rosiglitazone inhibits their formation. METHODS: A panel of 14, 13 and four carriers of the CYP2C8 genotypes *1/*1, *1/*3 and *3/*3, respectively was preselected for this study. Daily morning oral doses of 8 mg rosiglitazone were administered for 15 days. Urine was collected prior to rosiglitazone, and for 24 h after the first and last administration of rosiglitazone. Urinary EETs and DHETs were analyzed by tandem mass spectrometry. RESULTS: Carriers of the high-activity CYP2C8*3 allele had higher excretion of all three DHETs (p < 0.01 for 11,12-DHET, p < 0.05 for 14,15-DHET), whereas carriers of the low-activity CYP2C8 haplotype C (genotypes GCGA at positions rs2275622, rs7909236, rs1113129 and rs11572080) had lower DHET excretion in urine before and during rosiglitazone. Rosiglitazone intake leads to a decrease in DHET excretion by approximately 10% (p < 0.02). Urinary excretion of unhydrolyzed EETs was below the limit of quantification of 50 pg/ml in all samples. CONCLUSION: The data consistently indicate that genetic variation in CYP2C8 moderately modulates-EET formation as reflected in urinary DHET excretion. This might impact cardiovascular functions, and may be one mechanism explaining the influence of CYP polymorphisms on myocardial infarction and hypertension.

The T8590C polymorphism of CYP4A11 and 20-hydroxyeicosatetraenoic acid in essential hypertension.

A role for a deficit in transport actions of 20-hydroxyeicosatetraenoic acid (20-HETE) in hypertension is supported by the following: (1) diminished renal 20-HETE in Dahl-S rats; (2) altered salt- and furosemide-induced 20-HETE responses in salt-sensitive hypertensive subjects; and (3) increased population risk for hypertension in C allele carriers of the T8590C polymorphism of CYP4A11, which encodes an enzyme with reduced catalytic activity. We determined T8590C genotypes in 32 hypertensive subjects, 25 of whom were phenotyped for salt sensitivity of blood pressure and insulin sensitivity. Urine 20-HETE was lowest in insulin-resistant, salt-sensitive subjects (F=5.56; P<0.02). Genotypes were 13 TT, 2 CC, and 17 CT. C allele frequency was 32.8% (blacks: 38.9%; whites: 25.0%). C carriers (CC+CT) and TT subjects were similarly distributed among salt- and insulin-sensitivity phenotypes. C carriers had higher diastolic blood pressures and aldosterone:renin and waist:hip ratios but lower furosemide-induced fractional excretions of Na and K than TT. The T8590C genotype did not relate to sodium balance or pressure natriuresis. However, C carriers, compared with TT, had diminished 20-HETE responses to salt loading after adjustment for serum insulin concentration and resetting of the negative relationship between serum insulin and urine 20-HETE to a 1-microg/h lower level of 20-HETE. The effect of C was insulin independent and equipotent to 18 microU/mL of insulin (Delta20-HETE= 2.84-0.054xinsulin-0.98xC; r(2)=0.53; F=11.1; P<0.001). Hence, genetic (T8590C) and environmental (insulin) factors impair 20-HETE responses to salt in human hypertension. We propose that genotype analyses with sufficient homozygous CC will establish definitive relationships among 20-HETE, salt sensitivity of blood pressure, and insulin resistance.

A reduction in alcohol consumption is associated with reduced plasma F2-isoprostanes and urinary 20-HETE excretion in men.

There is considerable evidence that chronic moderate-to-high alcohol consumption increases blood pressure. The mechanisms by which this occurs are not clear. Alcohol consumption can induce oxidative stress and cytochrome P450 (CYP450) isoforms that are associated with oxidative stress and may influence vascular tone. To study the role of such mechanisms we examined whether reducing alcohol intake in moderate-to-heavy drinkers (40-110 g/day) resulted in changes in urinary excretion of 20-HETE, a CYP450 metabolite of arachidonic acid, and plasma and urinary F(2)-isoprostanes as markers of lipid peroxidation. After a 4-week run-in period during which healthy men maintained their usual drinking pattern they were randomized to a two-way crossover intervention study. In each of the 4-week treatment periods subjects either substituted their usual alcohol intake with a 0.9% alcohol beer or maintained their usual alcohol intake. Plasma and urinary F(2)-isoprostanes and urinary 20-HETE were measured by gas chromatography mass spectrometry, and serum gamma-glutamyl transpeptidase (gamma-GT) was measured as a biomarker of alcohol consumption, at the end of each study period. Sixteen healthy men age 51.0+/-2.7 years and with a BMI of 26.4+/-0.61 kg/m(2) completed the study. The reductions in alcohol intake (72.4+/-5.0 vs 7.9+/-1.6 g/day, p<0.001) and serum gamma-GT (geometric mean 24.4 U/L (95% CI 19.7, 30.2) vs 18.6 U/L (95% CI 15.5, 22.2, p<0.01) were accompanied by a significant fall in blood pressure as well as urinary 20-HETE excretion (158+/-23 vs 109+/-19 pmol/mmol creatinine, p<0.001) and plasma F(2)-isoprostanes (3438+/-158 vs 2929+/-145 pmol/L, p=0.01). A substantial reduction in alcohol consumption in healthy men lowered plasma F(2)-isoprostanes and urinary 20-HETE. Increased oxidative stress and 20-HETE production may be linked, at least in part, to the pathogenesis of alcohol-related hypertension.

Inability to upregulate cytochrome P450 4A and 2C causes salt sensitivity in young Sprague-Dawley rats.

BACKGROUND: Young Sprague-Dawley rats develop high blood pressure (BP) when exposed to a high salt intake, whereas adult ones generally do not. We investigated the role of renal cytochromes P450 4A (CYP 4A) and 2C (CYP 2C) in maintaining normal BP. METHODS: Young (age 5 weeks) and adult (age 53 weeks) Sprague-Dawley rats were given either 20 mmol sodium carbonate (vehicle for clofibrate) or 0.9% saline to drink for 3 weeks. Some young animals received the peroxisome proliferator activated receptor (PPAR)alpha agonist clofibrate (80 mg daily). We measured tail-cuff and intra-arterial BP, weight change, sodium balance, 20-hydroxyeicosatetraenoic acid (20-HETE) excretion (by high-performance liquid chromatography), and renal expression of CYP 4A and CYP 2C (by real-time reverse transcriptase-polymerase chain reaction). RESULTS: Saline-treated adult animals remained normotensive: systolic BP (SBP) 117 +/- 2 mm Hg v 117 +/- 1 mm Hg in control animals. In contrast, young rats given saline developed increased SBP: 134 +/- 2 mm Hg v 115 +/- 2 mm Hg in control animals (P < . 001). Interestingly, clofibrate lowered SBP to 102 +/- 2 mm Hg in saline-treated young rats but had no effect in control animals (114 +/- 2 mm Hg). Adult rats given saline increased renal expression of CYP 4A and 2C and excreted more 20-HETE. However, young rats given saline showed no induction, and even reduced CYP 4A and 2C, decreased urinary 20-HETE excretion, and retained sodium. Clofibrate increased renal CYP and 20-HETE excretion and prevented sodium retention. CONCLUSIONS: The products of renal CYP4A and 2C, including 20-HETE, aid in excreting salt. Animals that are unable to increase renal 20-HETE formation do not excrete sodium and are prone to hypertension.

Elevated 12- and 20-hydroxyeicosatetraenoic acid in urine of patients with prostatic diseases.

The role of eicosanoids (metabolites of arachidonic acid) in prostate diseases is receiving increased attention. We investigated the relationship between the concentrations of urinary free acids of 12- and 20-hydroxyeicosatetraenoic acids (12- and 20-HETE) and the benign prostatic hypertrophy (BPH) and prostate cancer (Pca). Urinary concentrations of 12-HETE and 20-HETE of BPH and Pca patients were significantly higher than normal subjects. After removal of the prostate gland, the urinary concentrations of these eicosanoids decreased to concentrations similar to the normal subjects. These results suggest that urinary free acids of 12-HETE and 20-HETE indicate an abnormality of the prostate gland.