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Background: The epithelial Na + channel (ENaC) is intrinsically linked to fluid volume homeostasis and blood pressure. Specific rare mutations in SCNN1A , SCNN1B , and SCNN1G , genes encoding the α, β, and γ subunits of ENaC, respectively, are associated with extreme blood pressure phenotypes. No associations between blood pressure and SCNN1D , which encodes the δ subunit of ENaC, have been reported. A small number of sequence variants in ENaC subunits have been reported to affect functional transport in vitro or blood pressure. The effects of the vast majority of rare and low-frequency ENaC variants on blood pressure are not known. Methods: We explored the association of low frequency and rare variants in the genes encoding ENaC subunits, with systolic blood pressure, diastolic blood pressure, mean arterial pressure, and pulse pressure. Using whole-genome sequencing data from 14 studies participating in the Trans-Omics in Precision Medicine Whole-Genome Sequencing Program, and sequence kernel association tests. Results: We found that variants in SCNN1A and SCNN1B were associated with diastolic blood pressure and mean arterial pressure ( P SCNN1D is poorly expressed in human kidney tissue, SCNN1D variants were associated with systolic blood pressure, diastolic blood pressure, mean arterial pressure, and pulse pressure ( P SCNN1B and SCNN1D ) were also associated with estimated glomerular filtration rate ( P Conclusions: Our results suggest that variants in extrarenal ENaCs, in addition to ENaCs expressed in kidneys, influence blood pressure and kidney function.

Background: Plasma proteins are critical mediators of cardiovascular processes and are the targets of many drugs. Previous efforts to characterize the genetic architecture of the plasma proteome have been limited by a focus on individuals of European descent and leveraged genotyping arrays and imputation. Here we describe whole genome sequence analysis of the plasma proteome in individuals with greater African ancestry, increasing our power to identify novel genetic determinants. Methods: Proteomic profiling of 1301 proteins was performed in 1852 Black adults from the Jackson Heart Study using aptamer-based proteomics (SomaScan). Whole genome sequencing association analysis was ascertained for all variants with minor allele count ≥5. Results were validated using an alternative, antibody-based, proteomic platform (Olink) as well as replicated in the Multi-Ethnic Study of Atherosclerosis and the HERITAGE Family Study (Health, Risk Factors, Exercise Training and Genetics). Results: We identify 569 genetic associations between 479 proteins and 438 unique genetic regions at a Bonferroni-adjusted significance level of 3.8×10 -11 . These associations include 114 novel locus-protein relationships and an additional 217 novel sentinel variant-protein relationships. Novel cardiovascular findings include new protein associations at the APOE gene locus including ZAP70 (sentinel single nucleotide polymorphism [SNP] rs7412-T, β=0.61±0.05, P =3.27×10 -30 ) and MMP-3 (β=-0.60±0.05, P =1.67×10 -32 ), as well as a completely novel pleiotropic locus at the HPX gene, associated with 9 proteins. Further, the associations suggest new mechanisms of genetically mediated cardiovascular disease linked to African ancestry; we identify a novel association between variants linked to APOL1-associated chronic kidney and heart disease and the protein CKAP2 (rs73885319-G, β=0.34±0.04, P =1.34×10 -17 ) as well as an association between ATTR amyloidosis and RBP4 levels in community-dwelling individuals without heart failure. Conclusions: Taken together, these results provide evidence for the functional importance of variants in non-European populations, and suggest new biological mechanisms for ancestry-specific determinants of lipids, coagulation, and myocardial function.

BACKGROUND: Asthma is a common respiratory disorder with a highly heterogeneous nature that remains poorly understood. The objective was to use whole genome sequencing (WGS) data to identify regions of common genetic variation contributing to lung function in individuals with a diagnosis of asthma. METHODS: WGS data were generated for 1,053 individuals from trios and extended pedigrees participating in the family-based Genetic Epidemiology of Asthma in Costa Rica study. Asthma affection status was defined through a physician’s diagnosis of asthma, and most participants with asthma also had airway hyperresponsiveness (AHR) to methacholine. Family-based association tests for single variants were performed to assess the associations with lung function phenotypes. RESULTS: A genome-wide significant association was identified between baseline FEV(1)/FVC ratio and a single-nucleotide polymorphism in the top hit cysteine-rich secretory protein LCCL domain-containing 2 (CRISPLD2) (rs12051168; P = 3.6 × 10(−8) in the unadjusted model) that retained suggestive significance in the covariate-adjusted model (P = 5.6 × 10(−6)). Rs12051168 was also nominally associated with other related phenotypes: baseline FEV(1) (P = 3.3 × 10(−3)), postbronchodilator (PB) FEV(1) (7.3 × 10(−3)), and PB FEV(1)/FVC ratio (P = 2.7 × 10(−3)). The identified baseline FEV(1)/FVC ratio and rs12051168 association was meta-analyzed and replicated in three independent cohorts in which most participants with asthma also had confirmed AHR (combined weighted z-score P = .015) but not in cohorts without information about AHR. CONCLUSIONS: These findings suggest that using specific asthma characteristics, such as AHR, can help identify more genetically homogeneous asthma subgroups with genotype-phenotype associations that may not be observed in all children with asthma. CRISPLD2 also may be important for baseline lung function in individuals with asthma who also may have AHR.

Background: Shorter telomere length is associated with an increased risk of coronary artery disease (CAD). A prior genomewide association study (GWAS) on 37,684 European ancestry individuals identified seven loci determining leukocyte telomere length (LTL). A genetic risk score across all 7 loci showed an association with CAD. In this study we sequenced these loci to determine whether the same risk variants could be replicated in African admixed individuals. Methods: We used whole genome sequence data (WGS) in 127 healthy African American subjects from GeneSTAR, a family study of early-onset CAD. LTL was calculated from WGS raw bam data (>30x coverage on the Illumina HiSeq platform) for 7 contiguous repeats of the telomere motif (TTAGGG or CCCTAA, Ding et al., 2014). Tests for association for LTL adjusting for age and sex were performed for a total of 55,821 called variants from ~1Mb subset regions of WGS genotype data centered on each of the 7 European peak GWAS SNPs. Results: We identified a total of 17 variants with p-4 mapping to 6 of the 7 regions examined; none of the prior European GWAS peak SNPs were significant. The peak SNP per region in the African Americans included intronic variants rs77138331 in ACYP2 (p=0.0005, MAF = 7%), rs149577640 in NAF1 (p=0.002, MAF=1%), rs35387865 in TERT (p=0.003, MAF = 1%) and rs186486116 in OBFC1 (p=0.004, MAF=1%). Additionally, novel intronic variants not previously observed in dbSNP located at chr19: 22190184 (p=0.005, MAF=1%) and chr2:62445438 (0.005, MAF=2%), mapping to ZNF208 and RTEL1 , were also identified as determinants of LTL. Discussion: This is the first study of telomere length in African Americans using a sequencing approach. We are unable to confirm the European-based GWAS variants but identify several associations mapping to genes of importance in telomere biology. Our results provide evidence that the set of SNPs to be included in calculation of a telomeric genetic CAD risk score may be different in populations of European and African ancestry.

Introduction: Although lipoprotein(a) (Lp(a)) is atherogenic, little is known about how it is transported from the plasma. Scavenger receptor class B type 1 (SR-B1), encoded by the gene SCARB1, binds high density lipoproteins (HDL). Recently we found that SR-B1 is also a receptor for Lp(a), mediates cellular uptake of Lp(a) in vitro, and promotes clearance of Lp(a) in vivo in mice. Genetic variants in SCARB1 have been associated with HDL level in humans, but no SCARB1 variants affecting Lp(a) have been reported. We hypothesized that loss of function mutations in SCARB1 would be associated with a phenotype of combined high HDL-C and Lp(a). Methods and Results: An index subject was identified in the NIH Lipid Clinic with high levels of both HDL-C and Lp(a). SCARB1 was sequenced and demonstrated a missense mutation at c.386 resulting in an S129L substitution in exon 3. To follow up on this finding, two cohorts of subjects (n=543 in the Genetic Study of Atherosclerosis Risk (GeneSTAR) and n=5907 in the Copenhagen City Heart Study) were screened for combined elevations of HDL-C and Lp(a). Subjects having HDL >80 mg/dl and Lp(a) >100 nmol/L in GeneSTAR, n=8, and >100 mg/dl in CCHS, n=9, underwent sequencing of all SCARB1 exons and splice junctions; 15 from the combined population of 18 were found to have variants predicted or demonstrated to have reduced function of the translated protein. Functional studies showed that 4 of the SCARB1 variants (c.386C>T; c.631-14T>G, c.4G>A and c.631-53mC>T & c.726+55mCG>CA, all heterozygous) had decreased receptor function in vitro, and the latter two were novel and not previously reported. Six of the subjects were homozygous for a common synonymous SCARB1 polymorphism (c.1050C>T) which has been reported to be associated with reduced protein expression. Conclusion: Human SCARB1 gene variants are associated with a new lipid phenotype, characterized by increases in both HDL-C and Lp(a). We demonstrate that SCARB1 exonic variants often result in diminished function of the translated SR-B1 lipid receptor with reduced binding/intracellular transport of Lp(a).

Remote ischemic preconditioning (RIPC) induces a prolonged late phase of multi-organ protection against ischemia-reperfusion (IR) injury. In the present study, we tested the hypothesis that RIPC confers late protection against myocardial IR injury by upregulating expression of IL-10. Mice were exposed to lower limb RIPC or sham leg ischemia. After 24 h, mice with RIPC demonstrated decreased myocardial infarct size and improved cardiac contractility following 30-min ischemia and 120-min reperfusion (I-30/R-120), accompanied by increased phosphorylation of Akt and eNOS in the heart. These effects of RIPC were completely blocked by anti-IL-10 receptor antibodies. Moreover, in IL-10 KO mice, RIPC was no longer able to provide cardioprotection against IR injury. IL-10 receptors were expressed in cardiomyocytes under basal conditions. Stimulation with exogenous IL-10 increased Akt phosphorylation and decreased myocardial infarct size after I-30/R-120 in isolated perfused hearts. Wildtype mice with RIPC demonstrated increased plasma IL-10 levels, while in the preconditioned gastrocnemius muscle, the phosphatase PTEN was inactivated and expression of IL-10 was increased through Stat3. Myocyte-specific PTEN inactivation led to increased Stat3 phosphorylation and IL-10 protein expression in the gastrocnemius muscle. Taken together, these results suggest that RIPC induces late protection against myocardial IR injury by increasing expression of IL-10 in the remote muscle, followed by release of IL-10 into the circulation, and activation of protective signaling pathways in the heart. This study provides a scientific basis for the use of RIPC to confer systemic protection against IR injury.

This study was designed to determine whether sex differences in platelet function exist in families with premature atherosclerosis. Compared with men, women have a greater risk of death and recurrent events following myocardial infarction and coronary artery bypass graft surgery. The reasons for this sex discrepancy are unknown. Because blood platelets play a central role in the formation of pathologic thrombi at sites of ruptured atheromatous plaques, we postulated that sex differences in platelet function exist that may be partly responsible for the sex difference in coronary artery disease (CAD) outcomes.We compared platelet reactivity in 400 asymptomatic men and women with family histories of premature CAD. Subjects were participants in the Johns Hopkins Sibling Study, a prospective investigation of coronary risk factors in asymptomatic, apparently healthy siblings of people with documented CAD.The platelets from women bound more fibrinogen in response to low and high concentrations of adenosine diphosphate. This sex difference was greater in whites than in African Americans. Age did not have an impact on these findings, although platelets from women 48 to 59 years old tended to bind less fibrinogen than those younger than 48 years. Female platelets also demonstrated greater spontaneous aggregation compared with male platelets, and women had higher levels of plasma thromboxane than men did. These differences were independent of smoking, hypertension, diabetes, hypercholesterolemia, or aspirin use.In asymptomatic individuals with family histories of premature CAD, platelets from women are more reactive than platelets from men. This observation cannot be explained by differences in cardiac risk factors.