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Hypertension and obesity in the young population are major risk factors for renal and cardiovascular events, which could arise in adulthood. A candidate-gene approach was applied in a cohort observational study, in which we collected data from 2638 high school adolescent students. Participants underwent anthropometric and blood pressure (BP) measurements, as well as saliva and urine sample collection for genomic DNA extraction and renal function evaluation, respectively. We tested whether candidate genes previously implicated in salt-sensitive hypertension in adults impact BP also among adolescents. Since inflammatory mechanisms may be involved in pathophysiology of hypertension and in endothelial dysfunction and atherosclerosis through reactive oxygen species, the baseline urinary excretion of inflammatory and oxidative stress markers in a subgroup of adolescents stratified according to ADD1 (alpha adducin) rs4961 genotypes was assessed. Regression analysis of BP values with genetic polymorphisms, highlighted an association with a missense variant of LSS (lanosterol synthase, rs2254524), a gene coding for an enzyme involved in endogenous ouabain synthesis. Higher diastolic and systolic BP were associated with LSS A allele ( P =0.011 and P =0.023, respectively). BP resulted associated with 5 more SNPs. The KL (klotho) rs9536314 missense variant was associated with 24 hour urinary Na + excretion ( P =0.0083). Urinary protein tests showed a greater excretion of IL1β (interleukin 1β) and interleukin 10 ( P <0.0001) in carriers of the ADD1 rs4961 T allele. In conclusion, 3 missense gene variants already implicated in adult hypertension impact BP or Na + excretion among adolescents, and, together with activated pro-inflammatory pathways, might predispose to early cardiovascular damage.

This study aimed to identify novel loci influencing the antihypertensive response to hydrochlorothiazide monotherapy. A genome-wide meta-analysis of blood pressure (BP) response to hydrochlorothiazide was performed in 1739 white hypertensives from 6 clinical trials within the International Consortium for Antihypertensive Pharmacogenomics Studies, making it the largest study to date of its kind. No signals reached genome-wide significance (P<5×10 - 8 ), and the suggestive regions (P<10 -5 ) were cross-validated in 2 black cohorts treated with hydrochlorothiazide. In addition, a gene-based analysis was performed on candidate genes with previous evidence of involvement in diuretic response, in BP regulation, or in hypertension susceptibility. Using the genome-wide meta-analysis approach, with validation in blacks, we identified 2 suggestive regulatory regions linked to gap junction protein α1 gene (GJA1) and forkhead box A1 gene (FOXA1), relevant for cardiovascular and kidney function. With the gene-based approach, we identified hydroxy-delta-5-steroid dehydrogenase, 3 β- and steroid δ-isomerase 1 gene (HSD3B1) as significantly associated with BP response (P<2.28×10 - 4 ). HSD3B1 encodes the 3β-hydroxysteroid dehydrogenase enzyme and plays a crucial role in the biosynthesis of aldosterone and endogenous ouabain. By amassing all of the available pharmacogenomic studies of BP response to hydrochlorothiazide, and using 2 different analytic approaches, we identified 3 novel loci influencing BP response to hydrochlorothiazide. The gene-based analysis, never before applied to pharmacogenomics of antihypertensive drugs to our knowledge, provided a powerful strategy to identify a locus of interest, which was not identified in the genome-wide meta-analysis because of high allelic heterogeneity. These data pave the way for future investigations on new pathways and drug targets to enhance the current understanding of personalized antihypertensive treatment., Competing Interests: Conflicts of interest/Disclosures None., (© 2016 American Heart Association, Inc.)

Circulating levels of endogenous ouabain (EO), a vasopressor hormone of adrenocortical origin, are increased by sodium depletion. Furthermore, lanosterol synthase, an enzyme involved in cholesterol biosynthesis, has a missense polymorphism (rs2254524 V642L) that affects EO biosynthesis in adrenocortical cells. Here, we investigated the hypothesis that lanosterol synthase rs2254524 alleles in vivo impact the blood pressure (BP) and EO responses evoked by a low dietary Na intake (<100 mEq/d, 2 weeks) among patients with mild essential hypertension. During the low salt diet, the declines in both systolic BP (SBP: -8.7±1.7 versus -3.0±1.5; P=0.013) and diastolic BP (DBP: -5.1±0.98 versus -1.4±0.94 mm Hg; P<0.05), and the slope of the long-term pressure-natriuresis relationship affected significantly the presence of the lanosterol synthase rs2254524 A variant (AA: 0.71±0.22, AC 0.09±0.13, and CC 0.04±0.11 mEq/mm Hg/24 h; P=0.028). In addition, BP rose in ≈25% of the patients in response to the low salt diet and this was associated with increased circulating EO. Lanosterol synthase gene polymorphisms influence both the salt sensitivity of BP and changes in circulating EO in response to a low salt diet. The response of BP and EO to the low salt diet is markedly heterogeneous. Approximately 25% of patients experienced adverse effects, that is, increased BP and EO when salt intake was reduced and may be at increased long-term risk. The augmented response of EO to the low salt diet further supports the view that adrenocortical function is abnormal in some essential hypertensives., (© 2015 American Heart Association, Inc.)

Defective pressure-natriuresis related to abnormalities in the natriuretic response has been associated with hypertension development. A major signaling pathway mediating pressure natriuresis involves the cGMP-dependent protein kinase 1 (PRKG1) that, once activated by Src kinase, inhibits renal Na(+) reabsorption via a direct action on basolateral Na-K ATPase and luminal Na-H exchanger type 3, as shown in renal tubuli of animals. Because a clear implication of PRKG1 in humans is still lacking, here we addressed whether PRKG1 polymorphisms affect pressure-natriuresis in patients. Naive hypertensive patients (n = 574), genotyped for PRKG1 rs1904694, rs7897633, and rs7905063 single nucleotide polymorphisms (SNPs), underwent an acute Na(+) loading, and the slope of the pressure-natriuresis relationship between blood pressure and Na(+) excretion was calculated. The underlying molecular mechanism was investigated by immunoblotting protein quantifications in human kidneys. The results demonstrate that the PRKG1 risk haplotype GAT (rs1904694, rs7897633, rs7905063, respectively) associates with a rightward shift of the pressure-natriuresis curve (0.017 ± 0.004 μEq/mm Hg per minute) compared with the ACC (0.0013 ± 0.003 μEq/mm Hg per minute; P = 0.001). In human kidneys, a positive correlation of protein expression levels between PRKG1 and Src (r = 0.83; P<0.001) or α1 Na-K ATPase (r = 0.557; P<0.01) and between α1 Na-K ATPase and Na-H exchanger type 3 (r = 0.584; P<0.01) or Src (r = 0.691; P<0.001) was observed in patients carrying PRKG1 risk GAT (n = 23) but not ACC (n = 14) variants. A functional signaling complex among PRKG1, α1 Na-K ATPase, and Src was shown by immunoprecipitation from human renal caveolae. These findings indicate that PRKG1 risk alleles associate with salt-sensitivity related to a loss of the inhibitory control of renal Na(+) reabsorption, suggestive of a blunt pressure-natriuresis response.

To identify novel genes influencing blood pressure response to thiazide diuretic therapy for hypertension, we conducted genome-wide association meta-analyses of ≈1.1 million single-nucleotide polymorphisms in a combined sample of 424 European Americans with primary hypertension treated with hydrochlorothiazide from the Pharmacogenomic Evaluation of Antihypertensive Responses study (n=228) and the Genetic Epidemiology of Responses to Antihypertensive study (n=196). Polymorphisms associated with blood pressure response at P<10(-5) were tested for replication of the associations in independent samples of hydrochlorothiazide-treated European hypertensives. The rs16960228 polymorphism in protein kinase C, α replicated for same-direction association with diastolic blood pressure response in the Nordic Diltiazem study (n=420) and the Genetics of Drug Responsiveness in Essential Hypertension study (n=206), and the combined 4-study meta-analysis P value achieved genome-wide significance (P=3.3 × 10(-8)). Systolic or diastolic blood pressure responses were consistently greater in carriers of the rs16960228 A allele than in GG homozygotes (>4/4 mm Hg) across study samples. The rs2273359 polymorphism in the GNAS-EDN3 region also replicated for same-direction association with systolic blood pressure response in the Nordic Diltiazem study, and the combined 3-study meta-analysis P value approached genome-wide significance (P=5.5 × 10(-8)). The findings document clinically important effects of genetic variation at novel loci on blood pressure response to a thiazide diuretic, which may be a basis for individualization of antihypertensive drug therapy and identification of new drug targets.

The kidney plays an important role in salt and blood pressure (BP) homeostasis. In previous studies, variants in the genes for alpha-adducin (ADD1), WNK1, and NEDD4L, which all regulate renal sodium absorption, have been associated with increased BP. However, findings have been inconsistent. We tested whether this is because of physiological interactions between the effects of variants in these genes. We assessed the single and combined effects of the ADD1 (Gly460Trp), WNK1 (rs880054 A/G), and NEDD4L (rs4149601 G/A) polymorphisms on renal and BP response to an acute Na load (n=344 subjects), BP decrease after 1 month of treatment with 12.5 mg of hydrochlorothiazide (n=193), and ambulatory 24-hour BP (n=690). Individually, the variants showed modest effects on some of the studied phenotypes. We found the ADD1 Trp allele to be permissive for the effects of variants of the other genes. In combination, the same variants (ADD1 Trp/WNK1 GG/Nedd4L GA+AA) showed a consistent effect on renal Na handling (P=0.009) and acute BP response to a saline infusion (P=0.021), BP lowering after thiazide treatment (P=0.008), and nocturnal systolic BP (P=0.044). Physiological interaction between the ADD1 and WNK1-NEDD4L pathways influences the effects of variants in these genes on sodium-related BP regulation. Relatively common alleles in the ADD1, WNK1, and NEDD4L genes when present in combination may have significant effects on renal sodium handling, BP, and antihypertensive response to thiazides.

Renin-angiotensin system reactivity and the constitutive capacity of the renal tubule to reabsorb sodium play a role in the individual response to diuretic therapy; therefore we evaluated the blood pressure (BP) response to hydrochlorothiazide in 87 never-treated individuals with mild essential hypertension, according to ACE gene I/D and alpha-adducin Gly460Trp polymorphism. These genotypes where chosen because previous data showed their interaction in determining the BP response to salt probably was the result of their involvement in the activation of the renin-angiotensin system (ACE) and in the constitutive capacity of the kidney to reabsorb sodium (alpha-adducin) (treatment for 2 months). BP was measured after 3 run-in visits and after the first and second months of treatment by means of a standardized procedure. Data were analyzed by ANOVA, t test, and multivariate ANOVA for repeated measures (covarying for gender, age, and body mass index). Although basal mean BP (MBP) was similar in the different ACE and alpha-adducin genotypes, patients carrying at least one I allele of ACE and one 460Trp allele of alpha-adducin had the largest MBP decrease with treatment (12.7+/-1.9 mm Hg), the effect of the combination of genotypes being additive but not epistatic. These patients had an odds ratio of 15.75 of being a responder to hydrochlorothiazide compared with patients with Gly460Gly+DD, with the least MBP decrease (3.4+/-1.7 mm Hg). Alpha-adducin and ACE I/D polymorphism may be useful to predict the interindividual degree of response to hydrochlorothiazide; the analysis of the combination of the 2 genotypes increases the accuracy of the prediction of response to the drug.

An ouabain-like factor has been implicated repeatedly in salt-sensitive hypertension as a natriuretic agent. However, the response of plasma ouabain-like factor to acute and chronic variation of body sodium is unclear. We studied 138 patients with essential hypertension who underwent an acute volume expansion/contraction maneuver (2 days) and 20 patients who entered a blind randomized crossover design involving chronically controlled sodium intake and depletion (170 to 70 mmol/d; 2 weeks each period). In both studies, plasma levels of ouabain-like factor were higher during sodium depletion (acute: 338.8+/-17.4 and 402.7+/-22.8 pmol/L for baseline and low sodium, respectively, P<0.01; chronic: 320.4+/-32.0 versus 481.0+/-48.1 pmol/L, P=0.01). No significant change in plasma ouabain-like factor was observed after a 2-hour saline infusion (333.4+/-23.9 pmol/L) or controlled sodium (402.1+/-34.9 pmol/L). When patients were divided into salt-sensitive or salt-resistant groups, no differences in plasma ouabain-like factor were observed in the 2 groups at baseline or in response to the 2 protocols: salt resistant (n=69, 340.1+/-25.9 pmol/L) versus salt sensitive (n=69, 337.4+/-23.6 pmol/L) and chronic salt resistant (n=11, 336.0+/-53.2) versus salt sensitive (n=9, 301.1+/-331.4 pmol/L). However, circulating ouabain-like factor was increased by sodium depletion in both groups. These results demonstrate that circulating ouabain-like factor is raised specifically by maneuvers that promote the loss of body sodium. Acute expansion of body fluids with isotonic saline is not a stimulus to plasma ouabain-like factor. Moreover, basal levels of plasma ouabain-like factor do not differ among patients with salt-sensitive or salt-resistant hypertension. Taken together, these new results suggest that ouabain-like factor is involved in the adaptation of humans to sodium depletion and argue against the hypothesis that ouabain-like factor is a natriuretic hormone.