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Apolipoprotein (apo) E-deficient transgenic mice develop marked hyperlipidemia and progressive atherosclerotic lesions. To explore whether oxidative modification of lipoproteins is involved in atherogenesis in this murine model, we performed extensive immunocytochemical studies. Atherosclerotic lesions ranging from early fatty streaks to very advanced plaques were examined from the aortic valve region and the thoracic and abdominal aorta. Using guinea pig antisera against malondialdehyde (MDA)-lysine and 4-hydroxynonenal-lysine, two epitopes generated during the oxidative modification of low-density lipoprotein (LDL), we demonstrated the presence of these "oxidation-specific epitopes" in atherosclerotic lesions. In early lesions, oxidation-specific epitopes were found predominantly in macrophage-rich areas, whereas diffuse extracellular staining predominated in necrotic areas of advanced lesions. We have previously shown that autoantibodies against MDA-lysine are present in the circulation of humans and rabbits and that the immunoglobulin fraction extracted from their lesions contains autoantibodies against several "oxidation-specific" epitopes. Sera from apoE-deficient mice also contained circulating autoantibodies to MDA-lysine, and both early and advanced lesions were rich in murine immunoglobulins. Titers of serum autoantibodies were significantly higher in apoE-deficient mice than in C57BL/6 mice. Autoantibodies in murine plasma recognized MDA-lysine epitopes in atherosclerotic lesions of rabbits, and the immunostaining was competitively inhibited by excess human MDA-LDL. Similar findings were obtained by competitive radioimmunoassay. Finally, a morphometric technique was developed and tested in these mice that allows a quantitative assessment of aortic atherosclerosis. These findings suggest that in apoE-deficient mice, lipoprotein oxidation is involved in atherogenesis and that these transgenic mice constitute an appropriate model with which to study the antiatherogenic effect of antioxidant intervention.

The results of prospective cohort studies indicate that consuming fish or fish oil containing the n-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) is associated with decreased cardiovascular death, whereas consumption of the vegetable oil-derived n-3 fatty acid a-linolenic acid is not as effective. Randomized control trials (RCTs) in the context of secondary prevention also indicate that the consumption of EPA plus DHA is protective at doses <1 g/d. The therapeutic effect appears to be due to suppression of fatal arrhythmias rather than stabilization of atherosclerotic plaques. At doses >3 g/d, EPA plus DHA can improve cardiovascular disease risk factors, including decreasing plasma triacylglycerols, blood pressure, platelet aggregation, and inflammation, while improving vascular reactivity. Mainly on the basis of the results of RCTs, the American Heart Association recommends that everyone eat oily fish twice per week and that those with coronary heart disease eat 1 g/d of EPA plus DHA from oily fish or supplements. Directions for future research include (1) RCTs to confirm the initial trials showing that EPA plus DHA decreases cardiovascular death and additional studies to determine whether this effect is due to EPA, DHA, or the combination; the dosage of the effective components; and whether the mechanism of action in humans is prevention of fatal arrhythmias. (2) Clinical studies to determine whether the reduction in cardiovascular disease risk factors is due to EPA, DHA, or the combination and the dosage of the effective components. (3) Clinical studies to determine whether vegetable oil-derived alpha-linolenic acid added to a diet enriched in n-6 fatty acids can effectively substitute for fish oil-derived EPA plus DHA. Copyright © 2006 American Society for Nutrition [ABSTRACT FROM AUTHOR]

We examined the relation of alcoholic beverage type and risk of myocardial infarction (MI) in a case-control study among 340 cases of MI and an equal number of age-, sex-, and community-matched controls. Alcohol consumption was estimated using a food frequency questionnaire, with alcohol drinkers defined as those consuming > or = 1/2 drink/day on average of any alcoholic beverage. Beer, wine, and liquor drinkers had at least half of their consumption from 1 beverage type. Fasting venous blood samples were obtained and analyzed for lipid profiles. Compared with nondrinkers, after adjustment for age and sex, reductions in risk of MI were similar for regular drinkers of any type of alcoholic beverage (relative risk [RR] 0.54; 95% confidence interval [CI] 0.37 to 0.79; p = 0.001), beer (RR 0.55; 95% CI 0.31 to 0.97; p <0.05), wine (RR 0.48; 95% CI 0.27 to 0.87; p <0.05), and liquor (RR 0.59; 95% CI 0.38 to 0.91; p <0.05) drinkers. Comparable benefits remained apparent even after multivariate adjustment for a wide range of nonlipid coronary risk factors. High-density lipoprotein (HDL) levels were significantly higher in all 4 beverage categories when compared with levels in nondrinkers, and as expected, adjustment for total HDL, a major direct effect of alcohol, substantially attenuated the protective effect in all 4 beverage categories. Relative risks were 0.94 for any beverage, 1.09 for beer, 0.97 for wine, and 0.83 for liquor after further adjustment. This strongly suggests that the protective effect of each beverage type is, in large part, mediated by increased HDL. These data indicate that regular consumption of small to moderate amounts of alcoholic beverages, regardless of the type, reduces the risk of MI, and further suggest that there is benefit, in large part, from increases in HDL levels. [ABSTRACT FROM AUTHOR]

Obesity has reached epidemic proportions in the United States, but little is known about the mechanisms of weight gain and weight loss. Integration of omics data is becoming a popular tool to increase understanding in such complex phenotypes. Biomarkers come in abundance, but small sample size remains a serious limitation in clinical trials. In the present study, we developed a strategy to screen predictors from a multiomics, high-dimensional, and longitudinal dataset from a small cohort of 10 women with obesity who were provided an identical very-low calorie diet. Our proposal explores the combinatorial space of potential predictors from transcriptomics, microbiome, metabolome, fecal bile acids, and clinical data with the application of the first-order Spearman partial correlation coefficient. Two statistics are proposed for screening predictors, the partial association score, and the persistent significance. We applied our strategy to predict rates of weight loss in our sample of participants in a hospital metabolic facility. Our method reduced an initial set of 42,000 biomarker candidates to 61 robust predictors. The results show baseline fecal bile acids and regulation in RT-polymerase chain reaction as the most predictive data sources in forecasting the rate of weight-loss. In summary, the present study proposes a strategy based on nonparametric statistics for ranking and screening predictors of weight loss from a multiomics study. The proposed biomarker screening strategy warrants further translational clinical investigation in obesity and other complex clinical phenotypes.

Introduction: Non-alcoholic fatty liver disease (NAFLD) is an increasing cause of chronic liver disease that accompanies obesity and the metabolic syndrome. Excess fructose consumption can initiate or exacerbate NAFLD in part due to a consequence of impaired hepatic fructose metabolism. Preclinical data emphasized that fructose-induced altered gut microbiome, increased gut permeability, and endotoxemia play an important role in NAFLD, but human studies are sparse. The present study aimed to determine if two weeks of excess fructose consumption significantly alters gut microbiota or permeability in humans., Methods: We performed a pilot double-blind, cross-over, metabolic unit study in 10 subjects with obesity (body mass index [BMI] 30-40 mg/kg/m 2 ). Each arm provided 75 grams of either fructose or glucose added to subjects' individual diets for 14 days, substituted isocalorically for complex carbohydrates, with a 19-day wash-out period between arms. Total fructose intake provided in the fructose arm of the study totaled a mean of 20.1% of calories. Outcome measures included fecal microbiota distribution, fecal metabolites, intestinal permeability, markers of endotoxemia, and plasma metabolites., Results: Routine blood, uric acid, liver function, and lipid measurements were unaffected by the fructose intervention. The fecal microbiome (including Akkermansia muciniphilia ), fecal metabolites, gut permeability, indices of endotoxemia, gut damage or inflammation, and plasma metabolites were essentially unchanged by either intervention., Conclusions: In contrast to rodent preclinical findings, excess fructose did not cause changes in the gut microbiome, metabolome, and permeability as well as endotoxemia in humans with obesity fed fructose for 14 days in amounts known to enhance NAFLD., Competing Interests: The authors have no conflicts of interest to declare., (© The Association for Clinical and Translational Science 2021.)

Obesity is accompanied by dysfunction of many organs, but effects on the skin have received little attention. We studied differences in epithelial thickness by histology and gene expression by Affymetrix gene arrays and PCR in the skin of 10 obese (BMI 35-50) and 10 normal weight (BMI 18.5-26.9) postmenopausal women paired by age and ethnicity. Epidermal thickness did not differ with obesity but the expression of genes encoding proteins associated with skin blood supply and wound healing were altered. In the obese, many gene expression pathways were broadly downregulated and subdermal fat showed pronounced inflammation. There were no changes in skin microbiota or metabolites. African American subjects differed from European Americans with a trend to increased epidermal thickening. In obese African Americans, compared to obese European Americans, we observed altered gene expression that may explain known differences in water content and stress response. African Americans showed markedly lower expression of the gene encoding the cystic fibrosis transmembrane regulator characteristic of the disease cystic fibrosis. The results from this preliminary study may explain the functional changes found in the skin of obese subjects and African Americans.

Background: Microbiota and bile acids in the gastrointestinal tract profoundly alter systemic metabolic processes. In obese subjects, gradual weight loss ameliorates adipose tissue inflammation and related systemic changes. We assessed how rapid weight loss due to a very low calorie diet (VLCD) affects the fecal microbiome and fecal bile acid composition, and their interactions with the plasma metabolome and subcutaneous adipose tissue inflammation in obesity., Methods: We performed a prospective cohort study of VLCD-induced weight loss of 10% in ten grades 2-3 obese postmenopausal women in a metabolic unit. Baseline and post weight loss evaluation included fasting plasma analyzed by mass spectrometry, adipose tissue transcription by RNA sequencing, stool 16S rRNA sequencing for fecal microbiota, fecal bile acids by mass spectrometry, and urinary metabolic phenotyping by 1 H-NMR spectroscopy. Outcome measures included mixed model correlations between changes in fecal microbiota and bile acid composition with changes in plasma metabolite and adipose tissue gene expression pathways., Results: Alterations in the urinary metabolic phenotype following VLCD-induced weight loss were consistent with starvation ketosis, protein sparing, and disruptions to the functional status of the gut microbiota. We show that the core microbiome was preserved during VLCD-induced weight loss, but with changes in several groups of bacterial taxa with functional implications. UniFrac analysis showed overall parallel shifts in community structure, corresponding to reduced abundance of the genus Roseburia and increased Christensenellaceae;g&#95;&#95; (unknown genus). Imputed microbial functions showed changes in fat and carbohydrate metabolism. A significant fall in fecal total bile acid concentration and reduced deconjugation and 7-α-dihydroxylation were accompanied by significant changes in several bacterial taxa. Individual bile acids in feces correlated with amino acid, purine, and lipid metabolic pathways in plasma. Furthermore, several fecal bile acids and bacterial species correlated with altered gene expression pathways in adipose tissue., Conclusions: VLCD dietary intervention in obese women changed the composition of several fecal microbial populations while preserving the core fecal microbiome. Changes in individual microbial taxa and their functions correlated with variations in the plasma metabolome, fecal bile acid composition, and adipose tissue transcriptome. Trial Registration ClinicalTrials.gov NCT01699906, 4-Oct-2012, Retrospectively registered. URL- https://clinicaltrials.gov/ct2/show/NCT01699906.

Background: Vitamin D deficiency, defined as a serum 25-hydroxyvitamin D [25(OH)D] concentration <20 ng/mL, is correlated with a more atherogenic lipid profile. However, oral vitamin D supplementation does not lower LDL-cholesterol concentrations or raise HDL-cholesterol concentrations. This uncoupling between association and causation may result from a failure of oral vitamin D to mimic the effect of dermally synthesized vitamin D in response to ultraviolet type B (UVB) light. Objective: We tested the hypothesis that, in vitamin D-deficient adults, the replenishment of vitamin D with UVB exposure would lower LDL-cholesterol concentrations compared with the effect of oral vitamin D 3 supplementation. Design: We performed a randomized clinical trial in vitamin D-deficient adults and compared vitamin D replenishment between subjects who received oral vitamin D 3 ( n = 60) and those who received narrow-band UVB exposure ( n = 58) ≤6 mo. Results: There was no difference in the change from baseline LDL-cholesterol concentrations between oral vitamin D 3 and UVB groups (difference in median of oral vitamin D 3 minus that of UVB: 1.5 mg/dL; 95% CI: -5.0, 7.0 mg/dL). There were also no differences within groups or between groups for changes in total or HDL cholesterol or triglycerides. Transcriptional profiling of skin and blood, however, revealed significant upregulation of immune pathway signaling with oral vitamin D 3 but significant downregulation with UVB. Conclusions: Correcting vitamin D deficiency with either oral vitamin D 3 or UVB does not improve the lipid profile. Beyond cholesterol, these 2 modalities of raising 25(OH)D have disparate effects on gene transcription. This trial was registered at clinicaltrials.gov as NCT01688102., (© 2017 American Society for Nutrition.)

Context: Obesity is associated with subclinical white adipose tissue inflammation, as defined by the presence of crown-like structures (CLSs) consisting of dead or dying adipocytes encircled by macrophages. In humans, bariatric surgery-induced weight loss leads to a decrease in CLSs, but the effects of rapid diet-induced weight loss on CLSs and metabolism are unclear., Objective: To determine the effects of rapid very-low-calorie diet-induced weight loss on CLS density, systemic biomarkers of inflammation, and metabolism in obese postmenopausal women., Design: Prospective cohort study., Setting: Rockefeller University Hospital, New York, NY., Participants: Ten obese, postmenopausal women with a mean age of 60.6 years (standard deviation, ±3.6 years)., Main Outcome Measures: Effects on CLS density and gene expression in abdominal subcutaneous adipose tissue, cardiometabolic risk factors, white blood count, circulating metabolites, and oxidative stress (urinary isoprostane-M) were measured., Results: Obese subjects lost approximately 10% body weight over a mean of 46 days. CLS density increased in subcutaneous adipose tissue without an associated increase in proinflammatory gene expression. Weight loss was accompanied by decreased fasting blood levels of high-sensitivity C-reactive protein, glucose, lactate, and kynurenine, and increased circulating levels of free fatty acids, glycerol, β -hydroxybutyrate, and 25 hydroxyvitamin D. Levels of urinary isoprostane-M declined., Conclusion: Rapid weight loss stimulated lipolysis and an increase in CLS density in subcutaneous adipose tissue in association with changes in levels of circulating metabolites, and improved systemic biomarkers of inflammation and insulin resistance. The observed change in levels of metabolites ( i.e. , lactate, β -hydroxybutyrate, 25 hydroxyvitamin D) may contribute to the anti-inflammatory effect of rapid weight loss.

Adipose tissue inflammation is associated with obesity comorbidities. Reducing such inflammation may ameliorate these comorbidities. n-3 fatty acids have been reported to have anti-inflammatory properties in obesity, which may modulate this inflammatory state. In the current study a 1 gram per day oral supplement of the n-3 fatty acid docosahexaenoic acid (DHA) was administered for 12 weeks to 10 grade 1-2 obese postmenopausal women and markers of adipose tissue and systemic inflammation measured and compared before and after supplementation. DHA administration resulted in approximately a doubling of plasma and red cell phospholipid and adipose tissue DHA content but no change in systemic markers of inflammation, such as circulating C-reactive protein (CRP) or interleukins (IL) 6, 8 and 10 (IL-6, IL-8, IL-10). DHA supplementation did not alter the adipose tissue marker of inflammation crown-like structure density nor did it affect any gene expression pathways, including anti-inflammatory, hypoxic and lipid metabolism pathways. The obese postmenopausal women studied were otherwise healthy, which leads us to suggest that in such women DHA supplementation is not an effective means for reducing adipose tissue or systemic inflammation. Further testing is warranted to determine if n-3 fatty acids may ameliorate inflammation in other, perhaps less healthy, populations of obese individuals.

Chemokines and their receptors play a critical role in immune function by directing cell-specific movement. C-C chemokine receptor 7 (CCR7) facilitates entry of T cells into lymph nodes. CCR7-dependent chemotaxis requires either of the cognate ligands C-C chemokine ligand 19 (CCL19) or CCL21. Although CCR7-dependent chemotaxis can be augmented through receptor up-regulation or by increased chemokine concentrations, we found that chemotaxis is also markedly enhanced by serum in vitro. Upon purification, the serum cofactor activity was ascribed to domain 5 of high-molecular-weight kininogen. This peptide was necessary and sufficient for accelerated chemotaxis. The cofactor activity in serum was dependent on coagulation factor XIIa, a serine protease known to induce cleavage of high-molecular-weight kininogen (HK) at sites of inflammation. Within domain 5, we synthesized a 24-amino acid peptide that could recapitulate the activity of intact serum through a mechanism distinct from up-regulating CCR7 expression or promoting chemokine binding to CCR7. This peptide interacts with the extracellular matrix protein thrombospondin 4 (TSP4), and antibodies to TSP4 neutralize its activity. In vivo, an HK domain 5 peptide stimulated homing of both T and B cells to lymph nodes. A circulating cofactor that is activated at inflammatory foci to enhance lymphocyte chemotaxis represents a powerful mechanism coupling inflammation to adaptive immunity., Competing Interests: Conflict of interest statement: The authors note that The Rockefeller University has filed two provisional patent applications for small molecule inhibitors of Factor XIIa; however, none of these inhibitors were used in this paper.

White adipose tissue inflammation (WATi) has been linked to the pathogenesis of obesity-related diseases, including type 2 diabetes, cardiovascular disease, and cancer. In addition to the obese, a substantial number of normal and overweight individuals harbor WATi, putting them at increased risk for disease. We report the first technique that has the potential to detect WATi noninvasively. Here, we used Raman spectroscopy to detect WATi with excellent accuracy in both murine and human tissues. This is a potentially significant advance over current histopathological techniques for the detection of WATi, which rely on tissue excision and, therefore, are not practical for assessing disease risk in the absence of other identifying factors. Importantly, we show that noninvasive Raman spectroscopy can diagnose WATi in mice. Taken together, these results demonstrate the potential of Raman spectroscopy to provide objective risk assessment for future cardiometabolic complications in both normal weight and overweight/obese individuals.

Context: Vitamin D deficiency, defined by the total serum 25-hydroxyvitamin D [25(OH)D] level, is common and more prevalent among Blacks than whites. Vitamin D-binding protein (DBP) levels vary with race and may modulate "bioavailable" levels of 25(OH)D., Objective: To determine the effect of DBP levels on the functional response to vitamin D., Setting and Design: A randomized, placebo-controlled trial of vitamin D repletion for 2 mo, which took place at an outpatient research unit. Participants included 150 vitamin D-deficient (25(OH)D <20 ng/mL) adults. Participants were randomly assigned to receive either 50,000 IU of vitamin D3 or placebo weekly for 8 weeks. This is a post-hoc analysis using DBP, 25(OH)D, PTH, and calcium levels., Results: Blacks had lower total 25(OH)D (12 vs 15 ng/mL, P < .001) and DBP levels (119 vs 234 μg/mL, P < .001) than non-Blacks. DBP levels were similar before and after vitamin D3 or placebo treatment (r = 0.98, P < .001). Baseline total 25(OH)D levels were a significant determinant of baseline PTH levels (P < .001). The change in total 25(OH)D was associated with the change in PTH (P < 0.001) and calcium levels (P < .05). In contrast, DBP levels were not a determinant of baseline PTH (P = .57) nor significantly related to changes in either PTH (P = .53) or calcium levels (P = .88)., Conclusions: DBP levels are stable in Blacks and non-Blacks, and do not change with correction of vitamin D deficiency. Even for individuals with total 25(OH)D levels < 20 ng/mL, Blacks have significantly lower DBP levels than non-Blacks. However, within this range of total 25(OH)D, DBP levels do not influence the effect of vitamin D repletion on PTH or calcium levels.

Rationale: Quantitative trait locus mapping of an intercross between C57.Apoe⁻/⁻ and FVB.Apoe⁻/⁻ mice revealed an atherosclerosis locus controlling aortic root lesion area on proximal chromosome 10, Ath11. In a previous work, subcongenic analysis showed Ath11 to be complex with proximal (10a) and distal (10b) regions., Objective: To identify the causative genetic variation underlying the atherosclerosis modifier locus Ath11 10b., Methods and Results: We now report subcongenic J, which narrows the 10b region to 5 genes, Myb, Hbs1L, Aldh8a1, Sgk1, and Raet1e. Sequence analysis of these genes revealed no amino acid coding differences between the parental strains. However, comparing aortic expression of these genes between F1.Apoe⁻/⁻ Chr10SubJ((B/F)) and F1.Apoe⁻/⁻ Chr10SubJ((F/F)) uncovered a consistent difference only for Raet1e, with decreased, virtually background, expression associated with increased atherosclerosis in the latter. The key role of Raet1e was confirmed by showing that transgene-induced aortic overexpression of Raet1e in F1.Apoe⁻/⁻ Chr10SubJ((F/F)) mice decreased atherosclerosis. Promoter reporter constructs comparing C57 and FVB sequences identified an FVB mutation in the core of the major aortic transcription start site abrogating activity., Conclusions: This nonbiased approach has revealed Raet1e, a major histocompatibility complex class 1-like molecule expressed in lesional aortic endothelial cells and macrophage-rich regions, as a novel atherosclerosis gene and represents one of the few successes of the quantitative trait locus strategy in complex diseases.

Objective: Obesity and diabetes are particularly high in indigenous populations exposed to a Western diet and lifestyle. The prevalence of obesity, diabetes, hyperglycemia, dyslipidemia, and hypertension in one such population, the Micronesian island of Kosrae was described., Design and Methods: Longitudinal screenings for metabolic traits were conducted on adult Kosraens ≥ 20 years of age in 1994 and again in 2001. Data was obtained on 3,106 Kosraens, comprising ∼80% of the adult population. Diabetes was diagnosed using World Health Organization guidelines. Prevalences of obesity, hyperglycemia, dyslipidemia, and hypertension were assessed., Results: The overall age-adjusted prevalence of diabetes increased from 14 to 21%. The most significant change observed in the population was increases in obesity and hyperglycemia, especially among young Kosraens and women. Obesity age-adjusted prevalence increased from 45 to 62%, and hyperglycemia age-adjusted prevalence increased from 19 to 44%. Of note, Kosraens as a group had unusually low high density lipoprotein (HDL) levels with 80% classified as low HDL by NCEP-ATPIII criteria, despite lacking the usually accompanying increase in triglycerides. Comparison to reports from other populations shows that Kosrae experiences one of the highest rates of obesity, hyperglycemia, and low HDL globally while maintaining relatively healthy levels of triglycerides., Conclusion: Our study shows a dramatic increase in obesity and hyperglycemia in Kosrae in just 7 years and forebodes significantly increased health risks for this part of the world., (Copyright © 2012 The Obesity Society.)

Statins are highly effective drugs prescribed to millions of people to lower LDL-cholesterol and decrease cardiovascular risk. The benefits of statin therapy seen in randomized clinical trials will only be replicated in real-life if patients adhere to the prescribed treatment regimen. But, about half of patients discontinue statin therapy within the first year, and adherence decreases with time. Patient, physician and healthcare system-related factors play a role in this problem. Recent studies have focused more on the patients' perspectives on non-adherence. Adverse events are cited as the most common cause of statin discontinuation; thus, the healthcare provider must be willing to ally and dialogue with patients to address concerns and assess the risks and benefits of continued statin therapy.

Background: Vitamin D deficiency is common in the general population and even more prevalent in patients with chronic kidney disease (CKD). Low 25-hydroxyvitamin D [25(OH)D] levels have been associated with cardiovascular disease, though a definitive mechanistic link has not been established. Further, it is unclear if repleting vitamin D mitigates the excess risk observed in epidemiologic studies. Because vitamin D may regulate innate immunity and gut epithelial differentiation, we hypothesized that oral cholecalciferol (D3) would result in decreased blood endotoxin activity, a potential risk factor for cardiovascular disease. STUDY DESIGN, SETTING & PARTICIPANTS, INTERVENTION: We studied 12 stable outpatients with CKD stage 3 and 25(OH)D deficiency, who received D3 30,000 units weekly for 8 weeks. The primary endpoint was the change in blood endotoxin activity., Results: Baseline endotoxin activity correlated with 25(OH)D levels (r = -0.60, p = 0.04). Endotoxin activity decreased by 25% from baseline (p = 0.03). Despite the decrease in endotoxin activity, there was no change in intestinal permeability., Conclusion: The results of this study suggest that vitamin D repletion therapy may have an effect on endotoxin activity in early CKD. Further intervention studies using vitamin D in the CKD population are required.

STARD4, a member of the evolutionarily conserved START gene family, has been implicated in the nonvesicular intracellular transport of cholesterol. However, the direction of transport and the membranes with which this protein interacts are not clear. We present studies of STARD4 function using small hairpin RNA knockdown technology to reduce STARD4 expression in HepG2 cells. In a cholesterol-poor environment, we found that a reduction in STARD4 expression leads to retention of cholesterol at the plasma membrane, reduction of endoplasmic reticulum-associated cholesterol, and decreased ACAT synthesized cholesteryl esters. Furthermore, D4 KD cells exhibited a reduced rate of sterol transport to the endocytic recycling compartment after cholesterol repletion. Although these cells displayed normal endocytic trafficking in cholesterol-poor and replete conditions, cell surface low density lipoprotein receptor (LDLR) levels were increased and decreased, respectively. We also observed a decrease in NPC1 protein expression, suggesting the induction of compensatory pathways to maintain cholesterol balance. These data indicate a role for STARD4 in nonvesicular transport of cholesterol from the plasma membrane and the endocytic recycling compartment to the endoplasmic reticulum and perhaps other intracellular compartments as well.

Objective: Vitamin D deficiency is common and associated with dyslipidemia. However, it is unclear whether oral vitamin D supplementation improves the lipid profile. Therefore, we conducted a randomized, placebo-controlled trial to determine the short-term effects of vitamin D repletion on the lipid profile., Methods and Results: One hundred fifty-one vitamin D-deficient (25-hydroxyvitamin D <20 ng/mL) adults with elevated risk for cardiovascular disease were randomized to receive either 50 000 IU of vitamin D3 weekly for 8 weeks or placebo. The primary outcome was the change in small low-density lipoprotein (LDL) particle number. Secondary outcomes included changes in other nuclear magnetic resonance-based and chemical lipid fractions. Vitamin D failed to improve the lipid profile. Compared with the placebo, vitamin D repletion did not change small LDL particle number (mean change, +18 nmol/L; 95% CI [-80 to +116 nmol/L]; P=0.63). There were also no changes in the chemical lipid profile: total cholesterol (+5.8 mg/dL, 95% CI [-1.4 to +13.0 mg/dL], P=0.14); LDL cholesterol (+3.8 mg/dL, 95% CI [-2.5 to +10.2 mg/dL], P=0.13); high-density lipoprotein cholesterol (+0.4 mg/dL 95% CI [-1.6 to +2.6 mg/dL], P=0.71); and triglycerides (+7.9 mg/dL 95% CI [-6.5 to +22.3 mg/dL]). In the vitamin D repletion group, exploratory multivariate regression analysis demonstrates that changes in LDL cholesterol were positively correlated with the changes in serum calcium (P<0.001) and inversely with the changes in serum parathyroid hormone (P=0.02)., Conclusions: In contrast to the association between low 25-hydroxyvitamin D levels and dyslipidemia, correcting vitamin D deficiency in the short-term does not improve the lipid profile. Repletion of 25-hydroxyvitamin D levels raised serum calcium levels and decreased serum parathyroid hormone levels. These expected physiological responses to vitamin D therapy were correlated with a significant increase in LDL cholesterol. Clinical Trial Registration- URL: http://www.clinicaltrials.gov. Unique identifier: NCT01008384.

Lowering of LDL cholesterol, predominantly accomplished clinically by statins, is one of the key components of both the prevention and medical management of coronary atherosclerosis; however, additional or alternative cholesterol lowering agents are needed for patients who fail to achieve goals or have adverse effects on statins. Owing to relatively rapid translation of basic science research on a novel regulatory pathway of the LDL receptor by PCSK9, a new class of such drugs with a different mode of action, and potentially better tolerance and less off-target effects may be just over the horizon.

Background: Vitamin D deficiency is highly prevalent and is associated with dyslipidemia and cardiovascular disease. The impact of correcting vitamin D deficiency on blood lipids, strong cardiovascular disease prognostic factors, is unknown., Methods and Results: To determine relationships between 25-hydroxyvitamin D levels and lipids, we analyzed 4.06 million deidentified patient laboratory test results from September 2009 through February 2011. We performed a cross-sectional study of this population to determine associations between 25-hydroxyvitamin D levels and lipids across clinically defined strata. We also conducted a retrospective cohort study of vitamin D deficient patients to investigate how changes in 25-hydroxyvitamin D levels relate to changes in lipid levels. After exclusions, 107 811 patients with serial testing were selected for cross-sectional analysis. Compared with vitamin D deficient patients (<20 ng/mL), those with optimal levels (≥30 ng/mL) had lower mean total cholesterol (-1.9 mg/dL; 95% confidence interval [95% CI], -1.2 to -2.7; P<0.0001), lower low-density lipoprotein cholesterol (-5.2 mg/dL; 95% CI, -4.5 to -5.8; P<0.0001), higher high-density lipoprotein cholesterol (4.8 mg/dL; 95% CI, 4.5-5.0; P<0.0001), and lower triglycerides (-7.5 mg/dL; 95% CI, -6.2 to -8.7; P<0.0001). For the retrospective cohort analysis, raising vitamin D levels from <20 to ≥30 ng/mL (n=6260), compared with remaining at <20 ng/mL (n=2332), was associated with a mean increase in total cholesterol (0.77 mg/dL; 95% CI, 0.18-1.36; P=0.01) and high-density lipoprotein cholesterol (0.42 mg/dL; 95% CI, 0.08-0.76; P=0.02) but nonsignificant changes in low-density lipoprotein cholesterol (0.32 mg/dL; 95% CI, -0.01 to 0.66; P=0.06) and triglycerides (0.04 mg/dL; 95% CI, -2.16 to 2.23 mg/dL; P=0.97)., Conclusions: Although vitamin D deficiency is associated with an unfavorable lipid profile in cross-sectional analyses, correcting for a deficiency might not translate into clinically meaningful changes in lipid concentrations; however, data from intervention trials are required to confirm these findings.

Whole-genome sequencing in an isolated population with few founders directly ascertains variants from the population bottleneck that may be rare elsewhere. In such populations, shared haplotypes allow imputation of variants in unsequenced samples without resorting to complex statistical methods as in studies of outbred cohorts. We focus on an isolated population cohort from the Pacific Island of Kosrae, Micronesia, where we previously collected SNP array and rich phenotype data for the majority of the population. We report identification of long regions with haplotypes co-inherited between pairs of individuals and methodology to leverage such shared genetic content for imputation. Our estimates show that sequencing as few as 40 personal genomes allows for inference in up to 60% of the 3000-person cohort at the average locus. We ascertained a pilot data set of whole-genome sequences from seven Kosraean individuals, with average 5× coverage. This assay identified 5,735,306 unique sites of which 1,212,831 were previously unknown. Additionally, these variants are unusually enriched for alleles that are rare in other populations when compared to geographic neighbors (published Korean genome SJK). We used the presence of shared haplotypes between the seven Kosraen individuals to estimate expected imputation accuracy of known and novel homozygous variants at 99.6% and 97.3%, respectively. This study presents whole-genome analysis of a homogenous isolate population with emphasis on optimal rare variant inference.

Accelerated atherosclerosis is the leading cause of death in type 1 diabetes, but the mechanism of type 1 diabetes-accelerated atherosclerosis is not well understood, in part due to the lack of a good animal model for the long-term studies required. In an attempt to create a model for studying diabetic macrovascular disease, we have generated type 1 diabetic Akita mice lacking the low density lipoprotein receptor (Ins2(Akita)Ldlr⁻/⁻). Ins2(Akita)Ldlr⁻/⁻ mice were severely hyperglycemic with impaired glucose tolerance. Compared with Ldlr⁻/⁻ mice, 20-week-old Ins2(Akita)Ldlr⁻/⁻ mice fed a 0.02% cholesterol AIN76a diet showed increased plasma triglyceride and cholesterol levels, and increased aortic root cross-sectional atherosclerotic lesion area [224% (P < 0.001) in males and 30% (P < 0.05) in females]. Microarray and quantitative PCR analyses of livers from Ins2(Akita)Ldlr⁻/⁻ mice revealed altered expression of lipid homeostatic genes, including sterol-regulatory element binding protein (Srebp)1, liver X receptor (Lxr)α, Abca1, Cyp7b1, Cyp27a1, and Lpl, along with increased expression of pro-inflammatory cytokine genes, including interleukin (Il)1α, Il1β, Il2, tumor necrosis factor (Tnf)α, and Mcp1. Immunofluorescence staining showed that the expression levels of Mcp1, Tnfα, and Il1β were also increased in the atherosclerotic lesions and artery walls of Ins2(Akita)Ldlr⁻/⁻ mice. Thus, the Ins2(Akita)Ldlr⁻/⁻ mouse appears to be a promising model for mechanistic studies of type 1 diabetes-accelerated atherosclerosis.

Rare variants affecting phenotype pose a unique challenge for human genetics. Although genome-wide association studies have successfully detected many common causal variants, they are underpowered in identifying disease variants that are too rare or population-specific to be imputed from a general reference panel and thus are poorly represented on commercial SNP arrays. We set out to overcome these challenges and detect association between disease and rare alleles using SNP arrays by relying on long stretches of genomic sharing that are identical by descent. We have developed an algorithm, DASH, which builds upon pairwise identical-by-descent shared segments to infer clusters of individuals likely to be sharing a single haplotype. DASH constructs a graph with nodes representing individuals and links on the basis of such segments spanning a locus and uses an iterative minimum cut algorithm to identify densely connected components. We have applied DASH to simulated data and diverse GWAS data sets by constructing haplotype clusters and testing them for association. In simulations we show this approach to be significantly more powerful than single-marker testing in an isolated population that is from Kosrae, Federated States of Micronesia and has abundant IBD, and we provide orthogonal information for rare, recent variants in the outbred Wellcome Trust Case-Control Consortium (WTCCC) data. In both cohorts, we identified a number of haplotype associations, five such loci in the WTCCC data and ten in the isolated, that were conditionally significant beyond any individual nearby markers. We have replicated one of these loci in an independent European cohort and identified putative structural changes in low-pass whole-genome sequence of the cluster carriers., (Copyright © 2011 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

The potential benefits of using population isolates in genetic mapping, such as reduced genetic, phenotypic and environmental heterogeneity, are offset by the challenges posed by the large amounts of direct and cryptic relatedness in these populations confounding basic assumptions of independence. We have evaluated four representative specialized methods for association testing in the presence of relatedness; (i) within-family (ii) within- and between-family and (iii) mixed-models methods, using simulated traits for 2906 subjects with known genome-wide genotype data from an extremely isolated population, the Island of Kosrae, Federated States of Micronesia. We report that mixed models optimally extract association information from such samples, demonstrating 88% power to rank the true variant as among the top 10 genome-wide with 56% achieving genome-wide significance, a >80% improvement over the other methods, and demonstrate that population isolates have similar power to non-isolate populations for observing variants of known effects. We then used the mixed-model method to reanalyze data for 17 published phenotypes relating to metabolic traits and electrocardiographic measures, along with another 8 previously unreported. We replicate nine genome-wide significant associations with known loci of plasma cholesterol, high-density lipoprotein, low-density lipoprotein, triglycerides, thyroid stimulating hormone, homocysteine, C-reactive protein and uric acid, with only one detected in the previous analysis of the same traits. Further, we leveraged shared identity-by-descent genetic segments in the region of the uric acid locus to fine-map the signal, refining the known locus by a factor of 4. Finally, we report a novel associations for height (rs17629022, P< 2.1 × 10⁻⁸).

Fish oils containing both EPA and DHA have been shown to have beneficial cardiovascular effects, but less is known about the independent effects of DHA. This study was designed to examine the effects of DHA on plasma lipid and lipoprotein concentrations and other biomarkers of cardiovascular risk in the absence of weight loss. In this randomized, controlled, double-blind trial, 36 overweight or obese adults were treated with 2 g/d of algal DHA or placebo for 4.5 mo. Markers of cardiovascular risk were assessed before and after treatment. In the DHA-supplemented group, the decrease in mean VLDL particle size (P ≤ 0.001) and increases in mean LDL (P ≤ 0.001) and HDL (P ≤ 0.001) particle sizes were significantly greater than changes in the placebo group. DHA supplementation also increased the concentrations of large LDL (P ≤ 0.001) and large HDL particles (P = 0.001) and decreased the concentrations of small LDL (P = 0.009) and medium HDL particles (P = 0.001). As calculated using NMR-derived data, DHA supplementation reduced VLDL TG (P = 0.009) and total TG concentrations (P = 0.006). Plasma IL-10 increased with DHA supplementation to a greater extent than placebo (P = 0.021), but no other significant changes were observed in glucose metabolism, insulin sensitivity, blood pressure, or markers of inflammation with DHA. In summary, DHA supplementation resulted in potentially beneficial changes in some markers of cardiometabolic risk, whereas other markers were unchanged.

Recent genome-wide association studies have identified a genetic locus at human chromosome 8q24 as having minor alleles associated with lower levels of plasma triglyceride (TG) and LDL cholesterol (LDL-C), higher levels of HDL-C, as well as decreased risk for myocardial infarction. This locus contains only one annotated gene, tribbles homolog 1 (TRIB1), which has not previously been implicated in lipoprotein metabolism. Here we demonstrate a role for Trib1 as a regulator of lipoprotein metabolism in mice. Hepatic-specific overexpression of Trib1 reduced levels of plasma TG and cholesterol by reducing VLDL production; conversely, Trib1-knockout mice showed elevated levels of plasma TG and cholesterol due to increased VLDL production. Hepatic Trib1 expression was inversely associated with the expression of key lipogenic genes and measures of lipogenesis. Thus, we provide functional evidence for what we believe to be a novel gene regulating hepatic lipogenesis and VLDL production in mice that influences plasma lipids and risk for myocardial infarction in humans.

Objective: Ath11, an atherosclerosis susceptibility locus on proximal chromosome 10 (0 to 21 cM) revealed in a cross between apolipoprotein E deficient C57BL/6 (B6) and FVB mice, was recently confirmed in congenic mice. The objectives of this study were to assess how Ath11 affects lesion development and morphology, to determine aortic gene expression in congenics, and to narrow the congenic interval., Methods and Results: Assessing lesion area over time in congenic mice showed that homozygosity for the FVB allele increased lesion area at 6 weeks persisting through to 24 weeks of age. Staining of aortic root sections at 16 weeks did not reveal obvious differences between congenics. Aortic expression-array analysis at 6 weeks revealed 97 genes that were >2-fold regulated, including 1 gene in the quantitative trait locus interval, Aldh8a1, and 2 gene clusters regulated by Hnf4alpha and Esr1. Analysis of lesion area in 11 subcongenic strains revealed 2 narrowed regions, 10a (21 genes), acting in females, and 10b (7 genes), acting in both genders., Conclusions: Ath11 appears to act early in lesion formation, with significant effects on aortic gene expression. This quantitative trait locus is genetically complex, containing a female-specific region 10a from 0 to 7.3 megabases (21 genes) and a gender-independent region 10b from 20.1 to 21.9 megabases (7 genes).

Steroidogenic acute regulatory protein (StAR)D4 is a member of the StAR related lipid transfer family. Homology comes from the approximately 210 amino acid lipid binding domain implicated in intracellular transport, cell signaling, and lipid metabolism. StARD4 was identified as a gene downregulated 2-fold by dietary cholesterol (Soccio, R. E., R. M. Adams, K. N. Maxwell, and J. L. Breslow. 2005. Differential gene regulation of StarD4 and StarD5 cholesterol transfer proteins. Activation of StarD4 by sterol regulatory element-binding protein-2 and StarD5 by endoplasmic reticulum stress. J. Biol. Chem. 280: 19410-19418). A mouse knockout was created to investigate StARD4's functionality and role in lipid metabolism. Homozygous knockout mice exhibited normal Mendelian mating genetics, but weighed less than wild-type littermates, an effect not accounted for by energy metabolism or food intake. Body composition as analyzed by DEXA scan showed no significant difference. No significant alterations in plasma or liver lipid content were observed on a chow diet, but female knockout mice showed a decrease in gallbladder bile cholesterol and phospholipid concentration. When challenged with a 0.2% lova-statin diet, StARD4 homozygous mice exhibited no changes. However, when challenged with a 0.5% cholesterol diet, female StARD4 homozygous mice showed a moderate decrease in total cholesterol, LDL, and cholesterol ester concentrations. Microarray analysis of liver RNA found few changes. However, NPC1's expression, a gene not on the microarray, was decreased approximately 2.5-fold in knockouts. These observations suggest that StARD4's role can largely be compensated for by other intracellular cholesterol transporters.

The architecture of natural variation present in a contemporary population is a result of multiple population genetic forces, including population bottleneck and expansion, selection, drift, and admixture. We seek to untangle the contribution of admixture to genetic diversity on the Micronesian island of Kosrae. Toward this goal, we used a complete genetic approach by combining a dense genome-wide map of 100,000 single-nucleotide polymorphisms (SNPs) with data from uniparental markers from the mitochondrial genome and the nonrecombining portion of the Y chromosome. These markers were typed in approximately 3200 individuals from Kosrae, representing 80% of the adult population of the island. We developed novel software that uses SNP data to delineate ancestry for individual segments of the genome. Through this analysis, we determined that 39% of Kosraens have some European ancestry. However, the vast majority of admixed individuals (77%) have European alleles spanning less than 10% of their genomes. Data from uniparental markers show most of this admixture to be male, introduced in the late nineteenth century. Furthermore, pedigree analysis shows that the majority of European admixture on Kosrae is because of the contribution of one individual. This approach shows the benefit of combining information from autosomal and uniparental polymorphisms and provides new methodology for determining ancestry in a population.

Rationale: The complications of atherosclerosis are a major cause of death and disability in type 2 diabetes. Defective clearance of apoptotic cells by macrophages (efferocytosis) is thought to lead to increased necrotic core formation and inflammation in atherosclerotic lesions., Objective: To determine whether there is defective efferocytosis in a mouse model of obesity and atherosclerosis., Methods and Results: We quantified efferocytosis in peritoneal macrophages and in atherosclerotic lesions of obese ob/ob or ob/ob;Ldlr(-/-) mice and littermate controls. Peritoneal macrophages from ob/ob and ob/ob;Ldlr(-/-) mice showed impaired efferocytosis, reflecting defective phosphatidylinositol 3-kinase activation during uptake of apoptotic cells. Membrane lipid composition of ob/ob and ob/ob;Ldlr(-/-) macrophages showed an increased content of saturated fatty acids (FAs) and decreased omega-3 FAs (eicosapentaenoic acid and docosahexaenoic acid) compared to controls. A similar defect in efferocytosis was induced by treating control macrophages with saturated free FA/BSA complexes, whereas the defect in ob/ob macrophages was reversed by treatment with eicosapentaenoic acid/BSA or by feeding ob/ob mice a fish oil diet rich in omega-3 FAs. There was also defective macrophage efferocytosis in atherosclerotic lesions of ob/ob;Ldlr(-/-) mice and this was reversed by a fish oil-rich diet., Conclusions: The findings suggest that in obesity and type 2 diabetes elevated levels of saturated FAs and/or decreased levels of omega-3 FAs contribute to decreased macrophage efferocytosis. Beneficial effects of fish oil diets in atherosclerotic cardiovascular disease may involve improvements in macrophage function related to reversal of defective efferocytosis and could be particularly important in type 2 diabetes and obesity.

The nuclear hormone receptor pregnane X receptor (PXR; also called SXR) functions as a xenobiotic sensor to coordinately regulate xenobiotic metabolism via transcriptional regulation of xenobiotic-detoxifying enzymes and transporters. Although many clinically relevant PXR ligands have been shown to affect cholesterol levels, the role of PXR in cholesterol homeostasis and atherosclerosis has not been thoroughly investigated. Here, we report that activation of PXR by feeding the PXR agonist pregnenolone 16alpha-carbonitrile (0.02%) for 2 weeks to wild-type (WT) mice significantly increased total cholesterol levels and atherogenic lipoproteins VLDL and LDL levels, but had no effect in PXR knockout (PXR(-/-)) mice. Chronic PXR activation in atherosclerosis prone apolipoprotein E deficient (ApoE(-/-)) mice was found to decrease HDL levels and increase atherosclerotic cross-sectional lesion area at both the aortic root and in the brachiocephalic artery by 54% (P < 0.001) and 116% (P < 0.01), respectively. PXR activation significantly regulated genes in the liver involved in lipoprotein transportation and cholesterol metabolism, including CD36, ApoA-IV, and CYP39A1, in both WT and ApoE(-/-) mice. Furthermore, PXR activation can increase CD36 expression and lipid accumulation in peritoneal macrophages of ApoE(-/-) mice. In summary, PXR activation in WT mice increases levels of the atherogenic lipoproteins VLDL and LDL, whereas in ApoE(-/-) mice, PXR increases atherosclerosis, perhaps by diminishing levels of the antiatherogenic ApoA-IV and increasing lipid accumulation in macrophages.

Pinpointing culprit causal variants along signal peaks of genome-wide association studies (GWAS) is challenging. To overcome confounding effects of multiple independent variants at such a locus and narrow the interval for causal allele capture, we developed an approach that maps local shared haplotypes harboring a putative causal variant. We demonstrate our method in an extreme isolate founder population, the pacific Island of Kosrae. We analyzed plasma plant sterol (PPS) levels, a surrogate measure of cholesterol absorption from the intestine, where previous studies have implicated 2p21 mutations in the ATP binding cassette subfamily G members 5 or 8 (ABCG5 or ABCG8) genes. We have previously reported that 11.1% of the islanders are carriers of a frameshift ABCG8 mutation increasing PPS levels in carriers by 50%. GWAS adjusted for this mutation revealed genomewide significant signals along 11 Mb around it. To fine-map this signal, we detected pairwise identity-by-descent haplotypes using our tool GERMLINE and implemented a clustering algorithm to identify haplotypes shared across multiple samples with their unique shared boundaries. A single 526-kb haplotype mapped strongly to PPS levels, dramatically refining the mapped interval. This haplotype spans the ABCG5/ABCG8 genes, is carried by 1.8% of the islanders, and results in a striking 100% increase of PPS in carriers. Resequencing of ABCG5 in these carriers found a D450H missense mutation along the associated haplotype. These findings exemplify the power of haplotype analysis for mapping mutations in isolated populations and specifically for dissecting effects of multiple variants of the same locus.

Objective: We have previously identified a quantitative trait locus (QTL) for atherosclerosis susceptibility on proximal chromosome 10 (Chr10) (Ath11) in independent crosses of FVB and C57BL/6 (B6) mice on the apolipoprotein E (ApoE-/-) and LDL receptor (LDLR-/-) deficient backgrounds. The aims of the current study were to (1) test a novel strategy for validating QTLs using interval-specific congenic strains that were heterozygous (F1) across the genome, (2) validate the Chr10 QTL, and (3) to assess whether the phenotype is transferable by bone marrow transplantation., Methods and Results: We generated Chr10 (0 to 21 cM) interval-specific mice on the F1.ApoE-/- background by crossing congenic FVB.ApoE-/-Chr10(B6/FVB) with B6.ApoE-/-, and B6.ApoE-/-Chr10(B6/FVB) with FVB.ApoE-/- mice. Lesion size was significantly larger in the resultant F1.ApoE-/-Chr10(FVB/FVB) mice compared to F1.ApoE-/-Chr10(B6/FVB) and F1.ApoE-/-Chr10(B6/B6) mice, validating the Chr10 QTL. The effect of the congenic interval was more robust on the F1.ApoE-/- than on the FVB.ApoE-/- and B6.ApoE-/- backgrounds. Bone marrow transplantation in congenic mice showed that the effect of the proximal Chr10 interval was not transferable by bone marrow-derived cells., Conclusions: A novel strategy of congenic strains on an F1 background proved useful to validate an atherosclerosis susceptibility QTL on mouse proximal Chr10.

Background: Cardiac conduction, as assessed by electrocardiographic PR interval and QRS duration, is an important electrophysiological trait and a determinant of arrhythmia risk., Objective: We sought to identify common genetic determinants of these measures., Methods: We examined 1604 individuals from the island of Kosrae, Federated States of Micronesia, an isolated founder population. We adjusted for covariates and estimated the heritability of quantitative electrocardiographic QRS duration and PR interval and, secondarily, its subcomponents, P-wave duration and PR segment. Finally, we performed a genome-wide association study (GWAS) in a subset of 1262 individuals genotyped using the Affymetrix GeneChip Human Mapping 500K microarray., Results: The heritability of PR interval was 34% (standard error [SE] 5%, P = 4 x 10(-18)); of PR segment, 31% (SE 6%, P = 3.2 x 10(-13)); and of P-wave duration, 17% (SE 5%, P = 5.8 x 10(-6)), but the heritablility of QRS duration was only 3% (SE 4%, P = .20). Hence, GWAS was performed only for the PR interval and its subcomponents. A total of 338,049 single nucleotide polymorphisms (SNPs) passed quality filters. For the PR interval, the most significantly associated SNPs were located in and downstream of the alpha-subunit of the cardiac voltage-gated sodium channel gene SCN5A, with a 4.8 ms (SE 1.0) or 0.23 standard deviation increase in adjusted PR interval for each minor allele copy of rs7638909 (P = 1.6 x 10(-6), minor allele frequency 0.40). These SNPs were also associated with P-wave duration (P = 1.5 x 10(-4)) and PR segment (P = .01) but not with QRS duration (P > or =.22)., Conclusions: The PR interval and its subcomponents showed substantial heritability in a South Pacific islander population and were associated with common genetic variation in SCN5A.

It has been argued that the limited genetic diversity and reduced allelic heterogeneity observed in isolated founder populations facilitates discovery of loci contributing to both Mendelian and complex disease. A strong founder effect, severe isolation, and substantial inbreeding have dramatically reduced genetic diversity in natives from the island of Kosrae, Federated States of Micronesia, who exhibit a high prevalence of obesity and other metabolic disorders. We hypothesized that genetic drift and possibly natural selection on Kosrae might have increased the frequency of previously rare genetic variants with relatively large effects, making these alleles readily detectable in genome-wide association analysis. However, mapping in large, inbred cohorts introduces analytic challenges, as extensive relatedness between subjects violates the assumptions of independence upon which traditional association test statistics are based. We performed genome-wide association analysis for 15 quantitative traits in 2,906 members of the Kosrae population, using novel approaches to manage the extreme relatedness in the sample. As positive controls, we observe association to known loci for plasma cholesterol, triglycerides, and C-reactive protein and to a compelling candidate loci for thyroid stimulating hormone and fasting plasma glucose. We show that our study is well powered to detect common alleles explaining >/=5% phenotypic variance. However, no such large effects were observed with genome-wide significance, arguing that even in such a severely inbred population, common alleles typically have modest effects. Finally, we show that a majority of common variants discovered in Caucasians have indistinguishable effect sizes on Kosrae, despite the major differences in population genetics and environment., Competing Interests: The authors have declared that no competing interests exist.