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Objectives: This study coupled 2 strategies - trait extremes and genome-wide pooling - to discover a novel blood pressure (BP) locus that encodes a previously uncharacterized thiamine transporter. Background: Hypertension is a heritable trait that remains the most potent and widespread cardiovascular risk factor, although details of its genetic determination are poorly understood. Methods: Representative genomic deoxyribonucleic acid (DNA) pools were created from male and female subjects in the highest- and lowest-fifth percentiles of BP in a primary care population of >50,000 patients. The peak associated single-nucleotide polymorphisms were typed in individual DNA samples, as well as in twins/siblings phenotyped for cardiovascular and autonomic traits. Biochemical properties of the associated transporter were evaluated in cellular assays. Results: After chip hybridization and calculation of relative allele scores, the peak associations were typed in individual samples, revealing an association between hypertension, systolic BP, and diastolic BP and the previously uncharacterized solute carrier SLC35F3. The BP genetic association at SLC35F3 was validated by meta-analysis in an independent sample from the original source population, as well as the International Consortium for Blood Pressure Genome-Wide Association Studies (across North America and western Europe). Sequence homology to a putative yeast thiamine (vitamin B1) transporter prompted us to express human SLC35F3 in Escherichia coli, which catalyzed [3H]-thiamine uptake. SLC35F3 risk-allele homozygotes (T/T) displayed decreased erythrocyte thiamine content on microbiological assay. In twin pairs, the SLC35F3 risk allele predicted heritable cardiovascular traits previously associated with thiamine deficiency, including elevated cardiac stroke volume with decreased vascular resistance, and elevated pressor responses to environmental (cold) stress. Allelic expression imbalance confirmed that cis variation at the human SLC35F3 locus influenced expression of that gene, and the allelic expression imbalance peak coincided with the hypertension peak. Conclusions: Novel strategies were coupled to position a new hypertension-susceptibility locus, uncovering a previously unsuspected thiamine transporter whose genetic variants predicted several disturbances in cardiac and autonomic function. The results have implications for the pathogenesis and treatment of systemic hypertension. © 2014 by the American College of Cardiology Foundation.

Objectives The goal of this study was to understand the role of genetic variation in the catecholamine biosynthetic pathway for control of human heart rate (HR). Background Human HR is an integrated cardiovascular trait predictive of morbidity and survival. Because the autonomic pathway exerts rapid control over the heart, we probed the role of heredity in the control of HR, focusing on a component of the autonomic sympathetic pathway already predictive of outflow responses: cytochrome b561 (CYB561), the electron shuttle in catecholamine vesicle membranes for transmitter biosynthesis. Methods We studied hereditary control of HR with the twin pair design, at rest and during environmental (cold) stress. Single nucleotide polymorphism disruption of a microribonucleic acid (microRNA) recognition motif in the human CYB561 3′-UTR was identified computationally, and its differential effect on gene expression was demonstrated in a transfected luciferase reporter/3′-UTR variant. We exposed stem cell–derived human embryoid bodies to the microRNA mimic or antagomir oligonucleotides, and we observed the effects on contraction rate in proto-hearts. Results Substantial heritability (h2) was demonstrated by using twin pair variance components for both basal/resting HR (h2 50.9 ± 6.4% of trait variation, p = 2.47 × 10−10) and stress-augmented HR (h2 55.1 ± 5.9%, p = 8.79 × 10−13), and the 2 HR traits shared genetic determination (genetic covariance ρG 0.747 ± 0.058, p = 2.85 × 10−9). CYB561 displayed 1 common genetic variant in the transcript region: A+1485G (rs3087776), in the 3′-UTR, 1485 bp downstream of the termination codon, in a conserved region, with the A-allele ancestral in primates. In a twin/sibling sample (n = 576), A+1485G influenced HR, both at rest (p = 0.010) and after environmental stress (p = 0.002), with the minor (A) allele displaying a recessive effect with lower HR. The effect of A+1485G on HR was extended by meta-analysis into 2 additional population samples (total n = 2,579), and the influence remained directionally consistent and significant (p = 0.007). A+1485G disrupted a microRNA (human microribonucleic acid-1294 [hsa-miR-1294]) recognition motif in the 3′-UTR, as demonstrated by a transfected luciferase reporter/human 3′-UTR variant system in 2 different neuronal/neuroendocrine cell types. The microRNA effect was further documented by cotransfection of an hsa-miR-1294 mimic, yielding an exaggerated decline in expression of the A-allele (better match) reporter (p = 4.3 × 10−5). Similar findings of differential 3′-UTR allelic susceptibility to hsa-miR-1294 were noted during expression of the full-length human CYB561 messenger ribonucleic acid with its cognate 3′-UTR. Finally, exposure of stem cell–derived human embryoid bodies to hsa-miR-1294 mimic or antagomir oligonucleotides yielded directionally opposite effects on contraction rate in proto-hearts. Conclusions HR is a substantially heritable trait, with genetic influence by variation in the adrenergic pathway, here shown for messenger ribonucleic acid translational control at the CYB561 step of transmitter formation. The results have implications for potentially modifiable autonomic pathways that influence this risk trait in the population.

Objectives This study sought to understand whether genetic variation at the Neuropeptide Y ( NPY ) locus governs secretion and stress responses in vivo as well as NPY gene expression in sympathochromaffin cells. Background The NPY is a potent pressor peptide co-released with catecholamines during stress by sympathetic axons. Genome-wide linkage on NPY secretion identified a LOD (logarithm of the odds ratio) peak spanning the NPY locus on chromosome 7p15. Methods Our approach began with genomics (linkage and polymorphism determination), extended into NPY genetic control of heritable stress traits in twin pairs, established transcriptional mechanisms in transfected chromaffin cells, and concluded with observations on blood pressure (BP) in the population. Results Systematic polymorphism tabulation at NPY (by re-sequencing across the locus: promoter, 4 exons, exon/intron borders, and untranslated regions; on 2n = 160 chromosomes of diverse biogeographic ancestries) identified 16 variants, of which 5 were common. We then studied healthy twin/sibling pairs (n = 399 individuals), typing 6 polymorphisms spanning the locus. Haplotype and single nucleotide polymorphism analyses indicated that proximal promoter variant ∇−880Δ (2-bp TG/—, Ins/Del, rs3037354) minor/Δ allele was associated with several heritable (h 2 ) stress traits: higher NPY secretion (h 2 = 73 ± 4%) as well as greater BP response to environmental (cold) stress, and higher basal systemic vascular resistance. Association of ∇−880Δ and plasma NPY was replicated in an independent sample of 361 healthy young men, with consistent allelic effects; genetic variation at NPY also associated with plasma NPY in another independent series of 2,212 individuals derived from Australia twin pairs. Effects of allele −880Δ to increase NPY expression were directionally coordinate in vivo (on human traits) and in cells (transfected NPY promoter/luciferase reporter activity). Promoter −880Δ interrupts a novel glucocorticoid response element motif, an effect confirmed in chromaffin cells by site-directed mutagenesis on the transfected promoter, with differential glucocorticoid stimulation of the motif as well as alterations in electrophoretic mobility shifts. The same −880Δ allele also conferred risk for hypertension and accounted for approximately 4.5/approximately 2.1 mm Hg systolic BP/diastolic BP in a population sample from BP extremes. Conclusions We conclude that common genetic variation at the NPY locus, especially in proximal promoter ∇−880Δ, disrupts glucocorticoid signaling to influence NPY transcription and secretion, raising systemic vascular resistance and early heritable responses to environmental stress, eventuating in elevated resting BP in the population. The results point to new molecular strategies for probing autonomic control of the human circulation and ultimately susceptibility to and pathogenesis of cardiovascular and neuropsychiatric disease states.

Objectives The purpose of this study is to better understand the origins and progression of pre-hypertension. Background Pre-hypertension is a risk factor for progression to hypertension, cardiovascular disease, and increased mortality. We used a cross-sectional twin study design to examine the role of heredity in likely pathophysiological events (autonomic or hemodynamic) in pre-hypertension. Methods Eight hundred twelve individuals (337 normotensive, 340 pre-hypertensive, 135 hypertensive) were evaluated in a sample of twin pairs, their siblings, and other family members. They underwent noninvasive hemodynamic, autonomic, and biochemical testing, as well as estimates of trait heritability (the percentage of trait variance accounted for by heredity) and pleiotropy (the genetic covariance or shared genetic determination of traits) by variance components. Results In the hemodynamic realm, an elevation of cardiac contractility prompted increased stroke volume, in turn increasing cardiac output, which elevated blood pressure into the pre-hypertension range. Autonomic monitoring detected an elevation of norepinephrine secretion plus a decline in cardiac parasympathetic tone. Twin pair variance components documented substantial heritability as well as joint genetic determination for blood pressure and the contributory autonomic and hemodynamic traits. Genetic variation at a pathway locus also indicated pleiotropic effects on contractility and blood pressure. Conclusions Elevated blood pressure in pre-hypertension results from increased cardiac output, driven by contractility as well as heart rate, which may reflect both diminished parasympathetic and increased sympathetic tone. In the face of increased cardiac output, systemic vascular resistance fails to decline homeostatically. Such traits display substantial heritability and shared genetic determination, although by loci not yet elucidated. These findings clarify the role of heredity in the origin of pre-hypertension and its autonomic and hemodynamic pathogenesis. The results also establish pathways that suggest new therapeutic targets for pre-hypertension, or approaches to its prevention.

The purpose of this study is to better understand the origins and progression of pre-hypertension.Pre-hypertension is a risk factor for progression to hypertension, cardiovascular disease, and increased mortality. We used a cross-sectional twin study design to examine the role of heredity in likely pathophysiological events (autonomic or hemodynamic) in pre-hypertension.Eight hundred twelve individuals (337 normotensive, 340 pre-hypertensive, 135 hypertensive) were evaluated in a sample of twin pairs, their siblings, and other family members. They underwent noninvasive hemodynamic, autonomic, and biochemical testing, as well as estimates of trait heritability (the percentage of trait variance accounted for by heredity) and pleiotropy (the genetic covariance or shared genetic determination of traits) by variance components.In the hemodynamic realm, an elevation of cardiac contractility prompted increased stroke volume, in turn increasing cardiac output, which elevated blood pressure into the pre-hypertension range. Autonomic monitoring detected an elevation of norepinephrine secretion plus a decline in cardiac parasympathetic tone. Twin pair variance components documented substantial heritability as well as joint genetic determination for blood pressure and the contributory autonomic and hemodynamic traits. Genetic variation at a pathway locus also indicated pleiotropic effects on contractility and blood pressure.Elevated blood pressure in pre-hypertension results from increased cardiac output, driven by contractility as well as heart rate, which may reflect both diminished parasympathetic and increased sympathetic tone. In the face of increased cardiac output, systemic vascular resistance fails to decline homeostatically. Such traits display substantial heritability and shared genetic determination, although by loci not yet elucidated. These findings clarify the role of heredity in the origin of pre-hypertension and its autonomic and hemodynamic pathogenesis. The results also establish pathways that suggest new therapeutic targets for pre-hypertension, or approaches to its prevention.