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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.

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.

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/m2). 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.

Background and AimsNon-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 due in part to a consequence of impaired hepatic fructose metabolism. Pre-clinical data have 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 explore the relevance of these pre-clinical studies to observations in humans.MethodsWe performed a classical double-blind metabolic unit study in 10 obese subjects (BMI 30-40 mg/kg/m2) providing 75gms. of either fructose or glucose in their individual diets substituted isocalorically for complex carbohydrates in a cross-over study. Excess fructose intake was provided in the fructose arm of the study and totaled a mean of 22.7% of calories.ResultsRoutine 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.ConclusionsAlthough pre-clinical rodent studies have shown that excess fructose causes pronounced changes in the gut microbiome, metabolome, and permeability as well as endotoxemia, this did not occur in obese individuals fed fructose in amounts known to enhance NAFLD. Therapeutic efforts to improve NAFLD through changes in the gut microbiome and gut homeostasis may not be beneficial.

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 from high-throughput experiments, but small sample size is still is a serious limitation in clinical trials. It makes assessment of more realistic assumptions for complex relationships such as nonlinearity, interaction and normality more difficult. In the present study, we developed a strategy to screen predictors of weight loss from a multi-omics, high-dimensional and longitudinal dataset from a small cohort of subjects. Our proposal explores the combinatorial space of candidate biomarkers from different data sources with the use of first-order Spearman partial correlation coefficients. Statistics derived from the sample correlations are used to rank and select biomarkers, and to evaluate the relative importance of each data source. We tackle the small sample size problem by combining nonparametric statistics and dimensionality reduction techniques useful for omics data. We applied the proposed strategy to assess the relative importance of biomarkers from 6 different data sources: RNA-seq, RT-qPCR, metabolomics, fecal microbiome, fecal bile acid, and clinical data used to predict the rate of weight loss in 10 obese subjects provided an identical low-calorie diet in a hospital metabolic facility. The strategy has reduced an initial set of more than 40K biomarkers to a set of 61 informative ones across 3 time points: pre-study, post-study and changes from pre- to post-study. Our study sheds light on the relative importance of different omics to predict rates of weight loss. We showed that baseline fecal bile acids, and changes in RT-qPCR biomarkers from pre- to post-study are the most predictive data sources for the rate of weight loss.Competing Interest StatementThe authors have declared no competing interest.AbbreviationsBPSBiomarker Predictive ScoreGSEAGene Set Enrichment AnalysisGSVAGene Set Variation AnalysisRT-qPCRReal Time quantitative Polymerase Chain ReactionSATSubcutaneous Adipose TissueVLCDVery Low Calorie DietWLWeight LossView Full Text

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 race. 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 Caucasians with a trend to increased epidermal thickening. In obese African Americans, compared to obese Caucasians, 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.

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.

Background: Vitamin D deficiency, defined as a serum 25-hydroxyvitamin D [25(OH)D] concentration

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., Rapid diet-induced weight loss increases subcutaneous adipose crown-like structure density in obese postmenopausal women.

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.

BackgroundMicrobiota 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.MethodsWe 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 by1H-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.ResultsAlterations 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 genusRoseburiaand increasedChristensenellaceae;g__(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.ConclusionsVLCD 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 RegistrationClinicalTrials.gov NCT01699906, 4-Oct-2012, Retrospectively registered. URL-https://clinicaltrials.gov/ct2/show/NCT01699906

OBJECTIVES/SPECIFIC AIMS: The objectives of this study are to 1) expand upon the paradigm of HK-D5 accelerated immune cell chemotaxis; 2) characterize the role of FXII in murine models of immune-mediated disease using FXII KO mice and a small molecule inhibitor of FXIIa. METHODS/STUDY POPULATION: To test whether the addition of HK-D5 peptide would accelerate C-C chemokine receptor type 2 (CCR2)-mediated chemotaxis in vitro, a real-time transwell chemotaxis assay was developed utilizing human THP-1 monocyte cell line (Fig 1). For in vivo studies, both pharmacologic FXIIa antagonism and FXll KO mice were used. Genotyping, histopathological review, FXll protein expression, and active partial thromboplastin time (aPTT) measurements were used to phenotypically characterize FXII KO mice compared to C57BL/6 wild type mice (Fig 2). Subsequently, the FXll KO mice were studied in varied models of immune-mediated disease, including LPS-induced sepsis and experimental autoimmune encephalitis (EAE), to see if and how the absence of FXll can mitigate disease severity. The EAE model involved active immunization with myelin oligodendrocyte glycoprotein (MOG) and measurement of established clinical disease severity scores. The LPS sepsis model involved an intraperitoneal injection of LPS followed by 48-hour monitoring of core body temperature using subdermal temperature transponders as a proxy for inflammatory events related to septic shock (Fig 3). RESULTS/ANTICIPATED RESULTS: HK-D5 significantly accelerates CCR2-mediated chemotaxis compared to chemokine alone (p = 0.001) similar to HK-D5’s ability to accelerate CCR7-mediated chemotaxis as previously established. The FXll KO mice were backcrossed to the C57BL/6J background and confirmed by genotyping and complete absence of FXII protein in plasma. Compared to the control, FXII KO mice have a significantly prolonged aPTT without evidence of bleeding abnormalities, which confirms the expected phenotype previously described and recapitulates what is observed in Factor XII deficiency in humans. KO mice showed no significant gross or histopathological differences in secondary lymphoid structures compared to the control. Immunohistochemistry confirmed well-organized lymphoid structures with intact B- and T-cell populations. FXll KO mice are protected in LPS-induced septic shock and EAE models. Regarding the EAE model, FXIIa inhibition significantly reduced disease severity compared to control. In the LPS model, FXll KO mice recover within 24 hours after LPS-challenge measured subjectively and objectively by core body temperature measurement. DISCUSSION/SIGNIFICANCE OF IMPACT: The current study and previous findings suggest a novel immune signaling mechanism by which a peptide fragment of high molecular-weight kininogen (HK-D5) acts as an accelerant of both innate and adaptive immune cell chemotaxis in multiple immune contexts. This has broad implications regarding a mechanism of immune-mediated inflammation in a variety of disease states, which might be amenable to the targeting this pathway for therapeutic intent.

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.To determine the effect of DBP levels on the functional response to vitamin D.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)D20 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.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 (P0.001) 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).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 levels20 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.

OBJECTIVES/SPECIFIC AIMS: To build a multisite de-identified database of female adolescents, aged 12–21 years (January 2011–December 2012), and their subsequent offspring through 24 months of age from electronic health records (EHRs) provided by participating Community Health. METHODS/STUDY POPULATION: We created a community-academic partnership that included New York City Community Health Centers (n=4) and Hospitals (n=4), The Rockefeller University, The Sackler Institute for Nutrition Science and Clinical Directors Network (CDN). We used the Community-Engaged Research Navigation model to establish a multisite de-identified database extracted from EHRs of female adolescents aged 12–21 years (January 2011–December 2012) and their offspring through 24 months of age. These patients received their primary care between 2011 and 2015. Clinical data were used to explore possible associations among specific measures. We focused on the preconception, prenatal, postnatal periods, including pediatric visits up to 24 months of age. RESULTS/ANTICIPATED RESULTS: The analysis included all female adolescents (n=122,556) and a subset of pregnant adolescents with offspring data available (n=2917). Patients were mostly from the Bronx; 43% of all adolescent females were overweight (22%) or obese (21%) and showed higher systolic and diastolic blood pressure, blood glucose levels, hemoglobin A1c, total cholesterol, and triglycerides levels compared with normal-weight adolescent females (p

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.

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, SETTINGPARTICIPANTS, 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.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.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.

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.

Background HDL cholesterol levels are inversely correlated with coronary heart disease risk in humans, and in animal studies, HDL elevation decreases formation and progression of foam-cell lesions. The potential for HDL to affect preexisting advanced atherosclerotic lesions is not known. To approach this issue, we used a novel mouse aortic transplantation model. Methods and Results ApoE-deficient (EKO) mice were fed a Western-type diet for 6 months, and thoracic aortic segments containing advanced lesions replaced segments of the abdominal aorta of 4-month-old EKO syngeneic mice not expressing (plasma HDL cholesterol ≈26 mg/dL) or expressing (HDL ≈64 mg/dL) a human apoAI (hAI) transgene. Both types of recipients had comparable non-HDL cholesterol levels. Five months after transplantation, mice were killed and grafts analyzed. Compared with lesion area in pretransplant mice (0.14±0.04 mm 2 , mean±SEM), there was progression in the EKO recipients (0.39±0.06 mm 2 , P 2 , P 80% ( P 300% ( P Conclusions Increasing HDL cholesterol levels in EKO mice retards progression of advanced atherosclerotic lesions and remodels them to a more stable-appearing phenotype.

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.

Effect of a Monoclonal Antibody to PCSK9 on LDL Cholesterol Stein et al New Eng J Med . 2012;366:1108–1118. 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. Proprotein convertase subtilisin/kexin 9 (PCSK9) binds to LDL receptors, resulting in their accelerated degradation and increased LDL cholesterol levels. In a recent New England Journal of Medicine article, Stein et al reported the use of a monoclonal antibody (REGN727) to PCSK9 with the goal of lowering LDL cholesterol levels in 3 randomized double-blind placebo controlled phase I studies.1 The first 2 studies were single-dose escalation studies performed in healthy volunteers, 1 by the intravenous and the other by the subcutaneous route, and each led to dose-dependent reductions in LDL cholesterol levels (28%–65% and 32%–46%, respectively) that were rapid and prolonged. The third study was a multiple-dose escalation study, with drug delivered subcutaneously on days 1, 29, and 43, performed in patients with familial hypercholesterolemia (FH) and non-FH receiving atorvastatin and a small group of 10 patients with non-FH on a modified diet alone (baseline LDL cholesterol, >130 mg per deciliter [3.4 mmol per liter]). In statin-treated FH and non-FH patients, subcutaneous REGN727 lowered LDL cholesterol levels an additional 40% to 60%, depending on dose. Interestingly, REGN727 lowered LDL cholesterol levels by a similar degree in non-FH patients not taking a statin compared to those taking a statin. These studies strongly suggest PCSK9 inhibition as a promising …

OBJECTIVES/SPECIFIC AIMS: To build a multisite deidentified database of female adolescents, aged 12–21 years (January 2011–December 2012), and their subsequent offspring through 24 months of age from electronic health records (EHRs) provided by participating Community Health. METHODS/STUDY POPULATION: We created a community-academic partnership that included New York City Community Health Centers (n=4) and Hospitals (n=4), The Rockefeller University, The Sackler Institute for Nutrition Science and Clinical Directors Network (CDN). We used the Community-Engaged Research Navigation model to establish a multisite deidentified database extracted from EHRs of female adolescents aged 12–21 years (January 2011–December 2012) and their offspring through 24 months of age. These patients received their primary care between 2011 and 2015. Clinical data were used to explore possible associations among specific measures. We focused on the preconception, prenatal, postnatal periods, including pediatric visits up to 24 months of age. RESULTS/ANTICIPATED RESULTS: The preliminary analysis included all female adolescents (n=49,292) and a subset of pregnant adolescents with offspring data available (n=2917). Patients were mostly from the Bronx; 43% of all adolescent females were overweight (22%) or obese (21%) and showed higher systolic and diastolic blood pressure, blood glucose levels, hemoglobin A1c, total cholesterol, and triglycerides levels compared with normal-weight adolescent females (p

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% (P0.001) in males and 30% (P0.05) 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.

found 95 loci significantly associated with at least one plasma lipid trait [7]. Many of these associations are mapped in or near genes with described functions in lipid metabolism and, in some cases, genes responsible for relatively rare Mendelian lipid disorders. These findings provide strong evidence for the validity of the GWAS approach. More excitingly, some of the newly discovered variants lie in genomic regions containing genes for which there is no existing evidence for a role in lipid metabolism. These loci offer the chance to broaden our understanding of lipid metabolism by discovering new modulator genes and ultimately hold the potential to yield new drug targets for the treatment of lipid disorders. At present, there is active debate as to whether new GWAS loci can fulfill these promises. Critics have questioned the scientific value of the GWAS approach and its capability to identify biologically relevant disease genes, since the identified loci have modest predictive power and explain only a small fraction of the heritability [9,10]. However, publication of the first studies tying novel genes from GWAS loci to functions in lipid metabolism [11–13] gives confidence that one can expect significant advances in lipid biology based on GWAS discoveries. In the following sections, we will take on the example of TRIB1 to illustrate how GWAS have led to the discovery of a novel regulator of lipoprotein metabolism.

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.

In November 2013, the American Heart Association (AHA)/American College of Cardiology (ACC) released a new Guideline on the Treatment of Blood Cholesterol to Reduce Atherosclerotic Cardiovascular Risk in Adults.1 Recommendations were made for several groups of patients. This perspective will focus only on the recommendation for those with no history of atherosclerotic cardiovascular disease (ASCVD) but with a 10-year ASCVD risk of ≥7.5% and a low-density lipoprotein (LDL) cholesterol level of 70 to 189 mg/dL and who are between the ages of 40 to 75 years. The guideline recommends such individuals receive moderate-intensity statin therapy to reduce LDL cholesterol between 30% and 49%. The guideline specifies that the 10-year risk of ASCVD, defined as nonfatal myocardial infarction (MI), coronary heart disease death, and nonfatal and fatal stroke, be calculated using the pooled cohort equations. The pooled cohort equations represent a new algorithm for estimating risk, and the spreadsheet for risk calculation can be downloaded from http://my.americanheart.org/professional/StatementsGuidelines/PreventionGuidelines/Prevention-Guidelines\_UCM\_457698_SubHomePage.jsp. Unfortunately, this algorithm was announced simultaneously with the new guideline without a prior period for external scientific evaluation. In fact, the algorithm has already been severely criticized.2 Ridker and Cook “calculated predicted 10-year risks of the same atherosclerotic events using the new ACC/AHA risk prediction algorithm and compared these estimates with observed event rates in 3 large-scale primary prevention cohorts, the Women’s Health Study, the Physicians’ Health Study, and the Women’s Health Initiative Observational Study.” They found “in all 3 of these primary prevention cohorts that the new ACC/AHA risk prediction algorithm systematically overestimated observed risks by 75% to 150%, roughly doubling the actual observed risk.” Ridker and Cook also found a similar overestimation of risk in 2 additional cohorts, prompting them to observe “that as many as 40% to 50% of the 33 million middle-aged Americans targeted by the new …

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).

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 Hnf4α 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. Conclusion— 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).