Ralph V. Shohet

Genetic Determinants of Lipid Traits in Diverse Populations from the Population Architecture using Genomics and Epidemiology (PAGE) Study

For the past five years, genome-wide association studies (GWAS) have identified hundreds of common variants associated with human diseases and traits, including high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), and triglyceride (TG) levels. Approximately 95 loci associated with lipid levels have been identified primarily among populations of European ancestry. The Population Architecture using Genomics and Epidemiology (PAGE) study was established in 2008 to characterize GWAS–identified variants in diverse population-based studies. We genotyped 49 GWAS–identified SNPs associated with one or more lipid traits in at least two PAGE studies and across six racial/ethnic groups. We performed a meta-analysis testing for SNP associations with fasting HDL-C, LDL-C, and ln(TG) levels in self-identified European American (∼20,000), African American (∼9,000), American Indian (∼6,000), Mexican American/Hispanic (∼2,500), Japanese/East Asian (∼690), and Pacific Islander/Native Hawaiian (∼175) adults, regardless of lipid-lowering medication use. We replicated 55 of 60 (92%) SNP associations tested in European Americans at p<0.05. Despite sufficient power, we were unable to replicate ABCA1 rs4149268 and rs1883025, CETP rs1864163, and TTC39B rs471364 previously associated with HDL-C and MAFB rs6102059 previously associated with LDL-C. Based on significance (p<0.05) and consistent direction of effect, a majority of replicated genotype-phentoype associations for HDL-C, LDL-C, and ln(TG) in European Americans generalized to African Americans (48%, 61%, and 57%), American Indians (45%, 64%, and 77%), and Mexican Americans/Hispanics (57%, 56%, and 86%). Overall, 16 associations generalized across all three populations. For the associations that did not generalize, differences in effect sizes, allele frequencies, and linkage disequilibrium offer clues to the next generation of association studies for these traits.

Conditional HIF-1α Expression Produces a Reversible Cardiomyopathy

Background The response to hypoxia in tissues is regulated by the heterodimeric transcription factor Hypoxia Inducible Factor-1 (HIF-1). Methodology/Principal Findings We have created a strain of mice with inducible cardiomyocyte-specific expression of a mutated, oxygen-stable, form of HIF-1α. Cardiac function steadily decreased with transgene expression, but recovered after the transgene was turned off. Using long-oligo microarrays, we identified 162 transcripts more than 3-fold dysregulated in these hearts after transgene expression. Among the down-regulated genes the transcript for SERCA was reduced 46% and the protein 92%. This led us to an evaluation of calcium flux that showed diminished reuptake of cytoplasmic calcium in myocytes from these hearts, suggesting a mechanism for cardiac dysfunction. Conclusions/Significance These results provide a deeper understanding of transcriptional activity of HIF in the heart, and show that enhanced HIF-1 activity is sufficient to cause contractile dysfunction in the adult heart. HIF is stabilized in the myocardium of patients with ischemic cardiomyopathy, and our results suggest that HIF could be contributing directly to the contractile dysfunction in this disease.

Down-regulation of Caveolin-1, an Inhibitor of Transforming Growth Factor-β Signaling, in Acute Allergen-induced Airway Remodeling

Asthma can progress to subepithelial airway fibrosis, mediated in large part by transforming growth factor-β (TGF-β). The scaffolding protein caveolin-1 (cav1) can inhibit the activity of TGF-β, perhaps by forming membrane invaginations that enfold TGF-β receptors. The study goals were 1) to evaluate how allergen challenge affects lung expression of cav1 and the density of caveolae in vivo 2) to determine whether reduced cav1 expression is mediated by interleukin (IL)-4 and 3) to measure the effects of decreased expression of cav1 on TGF-β signaling. C57BL/6J, IL-4-deficient mice, and cav1-deficient mice, sensitized by intraperitoneal injections of phosphate-buffered saline or ovalbumin (OVA) at days 0 and 12, received intranasal phosphate-buffered saline or OVA challenges at days 24, 26, and 28. Additionally, another group of C57BL/6J mice received IL-4 by intratracheal instillation for 7 days. We confirmed that the OVA-allergen challenge increased eosinophilia and T-helper type 2-related cytokine levels (IL-4, IL-5, and IL-13) in bronchoalveolar lavage. Allergen challenge reduced lung cav1 mRNA abundance by 40%, cav1 protein by 30%, and the number of lung fibroblast caveolae by 50%. Administration of IL-4 in vivo also substantially decreased cav1 expression. In contrast, the allergen challenge did not decrease cav1 expression in IL-4-deficient mice. The reduced expression of cav1 was associated with activation of TGF-β signaling that was further enhanced in OVA-sensitized and challenged cav1-deficient mice. This study demonstrates a previously unknown modulation of TGF-β signaling by IL-4, via cav1, suggesting novel therapeutic targets for controlling the effects of TGF-β and thereby ameliorating pathological airway remodeling.

Methamphetamine‐Associated Cardiomyopathy

Methamphetamine and related compounds are now the second most commonly used illicit substance worldwide, after cannabis. Reports of methamphetamine‐associated cardiomyopathy (MAC) are increasing, but MAC has not been well reviewed. This analysis of MAC will provide an overview of the pharmacology of methamphetamine, historical perspective and epidemiology, a review of case and clinical studies, and a summary of the proposed mechanisms for MAC. Clinically, many questions remain, including the appropriate therapeutic interventions for MAC, the incidence and prevalence of cardiac pathology in methamphetamine users, risk factors for developing MAC, and prognosis of these patients. In conclusion, recognition of the significance of MAC among physicians and other medical caregivers is important given the growing use of methamphetamine and related stimulants worldwide.

Cardioprotective Signaling by Endothelin

The endothelin axis promotes vasoconstriction, suggesting that antagonists of endothelin signaling might be useful in treatment of heart failure. However, promising results from animal trials have not been recapitulated in heart failure patients. Here we review the role of major signaling pathways in the heart that are involved in cell survival initiated by ET-1. These pathways include mitogen-activated protein kinase, phosphatidyl inositol-1,4,5-triphosphate kinase (PI3K-AKT), nuclear factor-kappaB (NF-kappaB), and calcineurin signaling. A better understanding of endothelin-mediated signaling in cardiac cell survival may allow a reevaluation of endothelin receptor antagonists (ETRAs) in the treatment of heart failure.

Cubozoan venom-induced cardiovascular collapse is caused by hyperkalemia and prevented by zinc gluconate in mice.

Chironex fleckeri (Australian box jellyfish) stings can cause acute cardiovascular collapse and death. We developed methods to recover venom with high specific activity, and evaluated the effects of both total venom and constituent porins at doses equivalent to lethal envenomation. Marked potassium release occurred within 5 min and hemolysis within 20 min in human red blood cells (RBC) exposed to venom or purified venom porin. Electron microscopy revealed abundant ∼12-nm transmembrane pores in RBC exposed to purified venom porins. C57BL/6 mice injected with venom showed rapid decline in ejection fraction with progression to electromechanical dissociation and electrocardiographic findings consistent with acute hyperkalemia. Recognizing that porin assembly can be inhibited by zinc, we found that zinc gluconate inhibited potassium efflux from RBC exposed to total venom or purified porin, and prolonged survival time in mice following venom injection. These findings suggest that hyperkalemia is the critical event following Chironex fleckeri envenomation and that rapid administration of zinc could be life saving in human sting victims.

Caveolin-1 modulates TGF-β1 signaling in cardiac remodeling

The cardiac response to myocardial injury includes fibrotic and hypertrophic processes and a key mediator in this response is transforming growth factor-β1 (TGF-β1). Caveolin-1 (cav1), the main structural protein of caveolae, is an inhibitor of the TGF-β1 signaling pathway. To examine the role of cav1 in cardiac repair, cav1 deficient (Cav1(-/-)) and wild type (WT) mice were subjected to cryoinjury of the left ventricle (LV). At baseline the two groups exhibited no inflammation, similar collagen content, and similar cardiac function. After injury, Cav1(-/-) animals displayed enhanced TGF-β1 signaling, as reflected by a 3-fold increase in the activation of the Smad2-dependent pathway and more widespread collagen deposition in the heart. Qualitative and quantitative analyses indicated that collagen deposition peaked in the WT LV 14days after injury, accompanied by increased mRNA abundance for procol1a2 (2-fold) and procol3a1 (3-fold). Collagen deposition was further enhanced in Cav1(-/-) mice, which was accompanied by reduced expression of matrix metalloproteinases MMP-8 (3-fold) and -13 mRNA (2-fold). The levels of expression of inflammatory markers of acute phase were similar between the strains However, macrophage clearance in the damaged region was delayed in Cav1(-/-) mice. We observed a 4-fold decrease in collagen deposition in Cav1(-/-) mice injected with a cav1 scaffolding domain peptide (CSD) and a 2-fold decrease in WT mice treated with the CSD. We conclude that cav1 has a direct role in reducing TGF-β1 signaling and as such might be an appropriate target for therapies to influence cardiac remodeling.

Vitamin D and Cardiovascular Disease: Potential Role in Health Disparities

Cardiovascular disease (CVD), which includes coronary artery disease and stroke, is the leading cause of mortality in the nation. Excess CVD morbidity and premature mortality in the African American community is one of the most striking examples of racial/ ethnic disparities in health outcomes. African Americans also suffer from increased rates of hypovitaminosis D, which has emerged as an independent risk factor for all-cause and cardiovascular mortality. This overview examines the potential role of hypovitaminosis D as a contributor to racial and ethnic disparities in cardiovascular disease (CVD). We review the epidemiology of vitamin D and CVD in African Americans and the emerging biological roles of vitamin D in key CVD signaling pathways that may contribute to the epidemiological findings and provide the foundation for future therapeutic strategies for reducing health disparities.

Specific antioxidant selenoproteins are induced in the heart during hypertrophy.

Selenium (Se) is thought to confer cardioprotective effects through the actions of antioxidant selenoprotein enzymes that directly limit levels of ROS such as hydrogen peroxide (H(2)O(2)) or that reverse oxidative damage to lipids and proteins. To determine how the selenoproteome responds to myocardial hypertrophy, two mouse models were employed: triidothyronine (T3)- or isoproterenol (ISO)-treatment. After 7days of T3- and ISO-treatment, cardiac stress was demonstrated by increased H(2)O(2) and caspase-3 activity. Neither treatment produced significant increases in phospholipid peroxidation or TUNEL-positive cells, suggesting that antioxidant systems were protecting the cardiomyocytes from damage. Many selenoprotein mRNAs were induced by T3- and ISO-treatment, with levels of methionine sulfoxide reductase 1 (MsrB1, also called SelR) mRNA showing the largest increases. MsrB enzymatic activity was also elevated in both models of cardiac stress, while glutathione peroxidase (GPx) activity and thioredoxin reductase (Trxrd) activity were moderately and nonsignificantly increased, respectively. Western blot assays revealed a marked increase in MsrB1 and moderate increases in GPx3, GPx4, and Trxrd1, particularly in T3-treated hearts. Thus, the main response of the selenoproteome during hypertrophy does not involve increased GPx1, but increased GPx3 for reducing extracellular H(2)O(2) and increased GPx4, Trxrd1, and MsrB1 for minimizing intracellular oxidative damage.

Aortic Complications Associated With Pregnancy in Marfan Syndrome: The NHLBI National Registry of Genetically Triggered Thoracic Aortic Aneurysms and Cardiovascular Conditions (GenTAC)

Background The risk of aortic complications associated with pregnancy in women with Marfan syndrome (MFS) is not fully understood. Methods and Results MFS women participating in the large National Registry of Genetically Triggered Thoracic Aortic Aneurysms and Cardiovascular Conditions (GenTAC) were evaluated. Among 184 women with MFS in whom pregnancy information was available, 94 (51%) had a total of 227 pregnancies. Among the women with pregnancies, 10 (10.6%) experienced a pregnancy‐related aortic complication (4 type A and 3 type B dissections, 1 coronary artery dissection, and 2 with significant [≥3 mm] aortic growth). Five of 7 aortic dissections, including all 3 type B, and the coronary dissection (75% of all dissections) occurred in the postpartum period. Only 5 of 8 women with pregnancy‐associated dissection were aware of their MFS diagnosis. The rate of aortic dissection was higher during the pregnancy and postpartum period (5.4 per 100 person‐years vs 0.6 per 100 person‐years of nonpregnancy; rate ratio, 8.4 [95% CI=3.9, 18.4]; P<0.0001). Conclusions Pregnancy in MFS is associated with an increased risk of aortic dissection, both types A and B, particularly in the immediate postpartum period. Lack of knowledge of underlying MFS diagnosis before aortic dissection is a major contributing factor. These findings underscore the need for early diagnosis, prepregnancy risk counseling, and multidisciplinary peripartum management.