Framingham Heart Study 100K Project: genome-wide associations for blood pressure and arterial stiffness

Background

About one quarter of adults are hypertensive and high blood pressure carries increased risk for heart disease, stroke, kidney disease and death. Increased arterial stiffness is a key factor in the pathogenesis of systolic hypertension and cardiovascular disease. Substantial heritability of blood-pressure (BP) and arterial-stiffness suggests important genetic contributions.

Methods

In Framingham Heart Study families, we analyzed genome-wide SNP (Affymetrix 100K GeneChip) associations with systolic (SBP) and diastolic (DBP) BP at a single examination in 1971–1975 (n = 1260), at a recent examination in 1998–2001 (n = 1233), and long-term averaged SBP and DBP from 1971–2001 (n = 1327, mean age 52 years, 54% women) and with arterial stiffness measured by arterial tonometry (carotid-femoral and carotid-brachial pulse wave velocity, forward and reflected pressure wave amplitude, and mean arterial pressure; 1998–2001, n = 644). In primary analyses we used generalized estimating equations in models for an additive genetic effect to test associations between SNPs and phenotypes of interest using multivariable-adjusted residuals. A total of 70,987 autosomal SNPs with minor allele frequency ≥ 0.10, genotype call rate ≥ 0.80, and Hardy-Weinberg equilibrium p ≥ 0.001 were analyzed. We also tested for association of 69 SNPs in six renin-angiotensin-aldosterone pathway genes with BP and arterial stiffness phenotypes as part of a candidate gene search.

Results

In the primary analyses, none of the associations attained genome-wide significance. For the six BP phenotypes, seven SNPs yielded p values < 10^-5. The lowest p-values for SBP and DBP respectively were rs10493340 (p = 1.7 × 10^-6) and rs1963982 (p = 3.3 × 10^-6). For the five tonometry phenotypes, five SNPs had p values < 10^-5; lowest p-values were for reflected wave (rs6063312, p = 2.1 × 10^-6) and carotid-brachial pulse wave velocity (rs770189, p = 2.5 × 10^-6) in MEF2C, a regulator of cardiac morphogenesis. We found only weak association of SNPs in the renin-angiotensin-aldosterone pathway with BP or arterial stiffness.

Conclusion

These results of genome-wide association testing for blood pressure and arterial stiffness phenotypes in an unselected community-based sample of adults may aid in the identification of the genetic basis of hypertension and arterial disease, help identify high risk individuals, and guide novel therapies for hypertension. Additional studies are needed to replicate any associations identified in these analyses.

     

A role for Hemolectin in coagulation and immunity in Drosophila melanogaster

Hemolectin has been identified as a candidate clotting factor in Drosophila. We reassessed the domain structure of Hemolectin (Hml) and propose that instead of C-type lectin domains, the two discoidin domains are most likely responsible for the protein's lectin activity. We also tested Hml's role in coagulation and immunity in Drosophila. Here we describe the isolation of a new hml allele in a forward screen for coagulation mutants, and our characterization of this and two other hml alleles, one of which is a functional null. While loss of Hml had strong effects on larval hemolymph coagulation ex vivo, mutant larvae survived wounding. Drosophila thus possesses redundant hemostatic mechanisms. We also found that loss of Hml in immune-handicapped adults rendered them more sensitive to Gram(−) bacterial infection. This demonstrates an immunological role of this clotting protein and reinforces the importance of the clot in insect immunity.     

Surveillance cultures and duration of carriage of multidrug-resistant Acinetobacter baumannii

Isolating carriers of multidrug-resistant (MDR) Acinetobacter baumannii is the main measure to prevent its spread. Identification of carriers accompanied by contact precautions is essential. We aimed to determine the appropriate surveillance sampling sites and the duration of carriage of MDR A. baumannii. We studied prospectively two groups of patients from whom MDR A. baumannii was previously isolated: (i) those with recent clinical isolation (or=6 months). Screening for carriage was conducted from six sites: nostrils, pharynx, skin, rectum, wounds, and endotracheal aspirates. Strains recovered concurrently from different sites were genotyped using pulsed-field gel electrophoresis. Twelve of 22 with recent clinical isolation of MDR A. baumannii had >or=1 positive screening culture, resulting in a sensitivity of 55% when six body sites were sampled. Sensitivities of single sites ranged from 13.5% to 29%. Among 30 patients with remote clinical isolation, screening cultures were positive in 5 (17%), with a mean duration of 17.5 months from the last clinical culture. Remote carriers had positive screening cultures from the skin and pharynx but not from nose, rectum, wounds, or endotracheal aspirates. Eleven strains from five patients were genotyped. In all but one case, isolates from different sites in a given patient were clonal. Current methodology is suboptimal to detect MDR A. baumannii carriage. The sensitivity of surveillance cultures is low, even when six different body sites are sampled. The proportion of individuals with previous MDR A. baumannii isolation who remain carriers for prolonged periods is substantial. These data should be considered when designing measures to limit the spread of MDR A. baumannii.     

Chronic exposure to Helicobacter pylori impairs dendritic cell function and inhibits Th1 development

Helicobacter pylori causes chronic gastric infection that affects the majority of the world's population. Despite generating an inflammatory response, the immune system usually fails to clear the infection. Since dendritic cells (DCs) play a pivotal role in shaping the immune response, we investigated the effects of H. pylori on DC function. We have demonstrated that H. pylori increased the expression of activation markers on DCs while upregulating the inhibitory B7 family molecule, PD-L1. Functionally, H. pylori-treated DCs resulted in the production of interleukin-10 (IL-10) and IL-23 but not of alpha interferon (IFN-alpha). While very little or no IL-12 was produced to H. pylori alone, simultaneous ligation of CD40 on DCs induced IL-12 release. We also demonstrated that DCs treated with H. pylori-induced IFN-gamma production by allogeneic naive T cells. However, stimulation of DCs with H. pylori for an extended period of time impaired their ability to produce cytokines after CD40 ligation and limited their ability to promote IFN-gamma release, suggesting that the DCs had become exhausted by the prolonged stimulation. The effect of chronic infection with H. pylori on DC function was further investigated by focusing on DC development. Demonstrating that monocytes differentiated into DCs in the presence of H. pylori exhibited an exhausted phenotype with an impaired ability to produce IL-12 and a downregulation of CD1a. Our results raise the possibility that in chronic H. pylori infection DCs become exhausted after prolonged antigen exposure leading to suboptimal Th1 development. This effect may contribute to persistence of H. pylori infection.     

Kinetic and expression analyses of seven novel mutations in mitochondrial acetoacetyl-CoA thiolase (T2): identification of a Km mutant and an analysis of the mutational sites in the structure

Mitochondrial acetoacetyl-CoA thiolase (T2) deficiency is an inborn error of metabolism that affects isoleucine catabolism and ketone body metabolism. We identified 7 novel and 2 previously reported mutations in six T2-deficient patients. Transient expression analysis of wild-type and eight mutant cDNAs was performed at 40, 37 and 30 degrees C. Although no significant residual activity was detected, mutant proteins were detected in the N158D, N158S, R208Q, Y219H and N282H mutants. Accumulation of these mutant proteins was temperature-sensitive with the highest expression levels at lower temperatures. Expression of Q73P and N353K cDNAs yielded neither residual T2 protein nor enzyme activity. An E252del mutant T2 was detected with a relative protein amount and enzyme activity of 30% and 25%, respectively, in comparison to the wild-type at 37 degrees C. The E252del mutant protein was more stable at 30 degrees C expression than 37 degrees C, but was essentially undetectable at 40 degrees C, indicating its temperature-sensitive instability. Kinetic studies revealed a twofold K(m) elevation for substrates coenzyme A and acetoacetyl-CoA in the E252del mutant, while V(max) was comparable to the wild-type. We conclude that the E252del is a temperature-sensitive K(m) mutant. This correlates well with the effect predicted from the T2 tertiary structure analysis, using the crystal structure of the human T2 homotetramer. The probable effect of the other mutations on the T2 tertiary structure was also evaluated.     

Comparative efficacy of exenatide versus insulin glargine on glycemic control in type 2 diabetes mellitus patients inadequately treated with metformin monotherapy

PurposeComparative efficacy of exenatide versus insulin glargine primarily on glucemic control, and secondarily on body mass index (BMI), lipid profile and blood pressure, in type 2 diabetes mellitus (T2DM) patients suboptimally treated with metformin monotherapy.Material/MethodsForty-seven inadequately treated T2DM patients on metformin assigned to exenatide (n=18) or insulin glargine (n=29) for 26 weeks. Glycosylated hemoglobin (HbA1c), serum lipids, BMI, systolic and diastolic blood pressure, and adverse events, including episodes of hypoglycemia and gastrointestinal symptoms, were recorded.ResultsEither treatment had a similar favorable mean reduction in HbA1c. However, more patients in exenatide group achieved HbA1c ≤ 7% at the 26th week compared with insulin glargine group (p=0.036). Insulin glargine group had significantly more episodes of hypoglycemia compared with exenatide group (p=0.039). Gastrointestinal adverse events were non-significantly higher in the exenatide group. A significantly greater BMI reduction was observed in exenatide group, whereas BMI was not altered in insulin glargine group. Total and LDL cholesterol (p=0.012), and triglycerides (p=0.016) significantly decreased, whereas HDL cholesterol increased (p=0.021) in the exenatide group, whereas only total cholesterol decreased in insulin glargine group. Changes in systolic and diastolic blood pressure were insignificant in both groups.ConclusionsExenatide provided similar reduction in HbA1c, but fewer episodes of hypoglycemia, compared with insulin glargine. Exenatide had also a favorable effect on weight loss, although more gastrointestinal adverse events. Exenatide may provide a justified alternative in second line treatment of T2DM, but more trials are required to elucidate its long-term safety and cost-effectiveness.     

Adjuvant Activity of Naturally Occurring Monophosphoryl Lipopolysaccharide Preparations from Mucosa-Associated Bacteria

Natural heterogeneity in the structure of the lipid A portion of lipopolysaccharide (LPS) produces differential effects on the innate immune response. Gram-negative bacterial species produce LPS structures that differ from the classic endotoxic LPS structures. These differences include hypoacylation and hypophosphorylation of the diglucosamine backbone, both differences known to decrease LPS toxicity. The effect of decreased toxicity on the adjuvant properties of many of these LPS structures has not been fully explored. Here we demonstrate that two naturally produced forms of monophosphorylated LPS, from the mucosa-associated bacteria Bacteroides thetaiotaomicron and Prevotella intermedia, function as immunological adjuvants for antigen-specific immune responses. Each form of mucosal LPS increased vaccination-initiated antigen-specific antibody titers in both quantity and quality when given simultaneously with vaccine antigen preparations. Interestingly, adjuvant effects on initial T cell clonal expansion were selective for CD4 T cells. No significant increase in CD8 T cell expansion was detected. MyD88/Toll-like receptor 4 (TLR4) and TRIF/TLR4 signaling pathways showed equally decreased signaling with the LPS forms studied here as with endotoxic LPS or detoxified monophosphorylated lipid A (MPLA). Natural monophosphorylated LPS from mucosa-associated bacteria functions as a weak but effective adjuvant for specific immune responses, with preferential effects on antibody and CD4 T cell responses over CD8 T cell responses.     

Social reasoning in Tourette syndrome

Introduction. Tourette syndrome (TS) is thought to be associated with striatal dysfunction. Changes within frontostriatal pathways in TS could lead to changes in abilities reliant on the frontal cortex. Such abilities include executive functions and aspects of social reasoning.

Methods. This study aimed to investigate executive functioning and Theory of Mind (ToM; the ability to reason about mental states, e.g., beliefs and emotions), in 18 patients with TS and 20 controls. A range of tasks involving ToM were used. These required participants to make judgements about mental states based on pictures of whole faces or the eyes alone, reason about humour in cartoons that featured sarcasm, irony or “slapstick” style humour, and make economic decisions. The executive measures assessed inhibition and verbal fluency.

Results. Patients with TS exhibited significantly poorer performance than controls on all four tasks involving ToM, even when patients with comorbid obsessive-compulsive disorder were excluded. These difficulties were despite no inhibitory deficits. Patients with TS exhibited impairment on the verbal fluency task but their performance on executive and ToM tasks was not related.

Conclusions. We propose that TS is associated with changes in ToM. The observed deficits could reflect dysfunction in frontostriatal pathways involving ventromedial prefrontal cortex.     

To tell or not to tell – what to do about p.C282Y heterozygotes identified by HFE screening

Hereditary hemochromatosis (HH) is a common preventable disorder of iron overload that can result in liver cirrhosis and reduced lifespan. Most HH is due to homozygosity for the HFE p.C282Y substitution. We conducted a study of screening for p.C282Y in high schools where p.C282Y heterozygotes (CY) individuals were informed of their genotype by letter. We studied whether these individuals understood the implications of their genotype, whether this resulted in anxiety or reduced health perception and whether cascade testing was higher in families of CY than wild‐type homozygous (CC) individuals. We found 586 of 5757 (1 in 10) screened individuals were CY. One month after receiving their result, 83% correctly answered that they have one copy of p.C282Y. There was no adverse change in anxiety or health perception from prior to screening to 1 month after receiving results. Significantly more family members of CY individuals than CC individuals were informed about HH and had testing for HH. In conclusion, we found that informing CY individuals of their genotype does not increase anxiety and the implications are generally well understood. This leads to cascade testing in a minority of families. CY individuals should be informed of their genetic status when identified by population screening.