The Weibel-Palade Body Localized SNARE (Soluble NSF Attachment Protein Receptor) Syntaxin-3 Modulates Von Willebrand Factor Secretion From Endothelial Cells

Objective Endothelial cells store von Willebrand factor (VWF) in rod-shaped secretory organelles, called Weibel-Palade bodies (WPBs). WPB exocytosis is coordinated by a complex network of Rab GTPases, Rab-effectors and SNARE proteins. We have previously identified STXBP1 as the link between the Rab27A-Slp4-a complex on WPBs and the SNARE proteins syntaxin-2 and -3. In this study we investigate the function of syntaxin-3 in VWF secretion. Approach and Results In human umbilical vein endothelial cells (HUVECs) and in blood outgrowth endothelial cells (BOECs) from healthy controls endogenous syntaxin-3 immunolocalized to WPBs. A detailed analysis of BOECs isolated from a patient with variant microvillus inclusion disease (MVID), carrying a homozygous mutation in STX3 (STX3-/-), showed a loss of syntaxin-3 protein and absence of WPB-associated syntaxin-3 immunoreactivity. Ultrastructural analysis revealed no detectable differences in morphology or prevalence of immature or mature WPBs in control versus STX3-/- BOECs. VWF multimer analysis showed normal patterns in plasma of the MVID patient, and media from STX3-/- BOECs, together indicating WPB formation and maturation are unaffected by absence of syntaxin-3. However, a defect in basal as well as Ca2+ - and cAMP-mediated VWF secretion was found in the STX3-/- BOECs. We also show that syntaxin-3 interacts with the WPB-associated SNARE protein VAMP8. Conclusions Our data reveal syntaxin-3 as a novel WPB-associated SNARE protein that controls WPB exocytosis

Nuclear Receptor Rev-erb Alpha (Nr1d1) Functions in Concert with Nr2e3 to Regulate Transcriptional Networks in the Retina

The majority of diseases in the retina are caused by genetic mutations affecting the development and function of photoreceptor cells. The transcriptional networks directing these processes are regulated by genes such as nuclear hormone receptors. The nuclear hormone receptor gene Rev-erb alpha/Nr1d1 has been widely studied for its role in the circadian cycle and cell metabolism, however its role in the retina is unknown. In order to understand the role of Rev-erb alpha/Nr1d1 in the retina, we evaluated the effects of loss of Nr1d1 to the developing retina and its co-regulation with the photoreceptor-specific nuclear receptor gene Nr2e3 in the developing and mature retina. Knock-down of Nr1d1 expression in the developing retina results in pan-retinal spotting and reduced retinal function by electroretinogram. Our studies show that NR1D1 protein is co-expressed with NR2E3 in the outer neuroblastic layer of the developing mouse retina. In the adult retina, NR1D1 is expressed in the ganglion cell layer and is co-expressed with NR2E3 in the outer nuclear layer, within rods and cones. Several genes co-targeted by NR2E3 and NR1D1 were identified that include: Nr2c1, Recoverin, Rgr, Rarres2, Pde8a, and Nupr1. We examined the cyclic expression of Nr1d1 and Nr2e3 over a twenty-four hour period and observed that both nuclear receptors cycle in a similar manner. Taken together, these studies reveal a novel role for Nr1d1, in conjunction with its cofactor Nr2e3, in regulating transcriptional networks critical for photoreceptor development and function

Factors associated with herbal use among urban multiethnic primary care patients: a cross-sectional survey

BACKGROUND: The use of herbal supplements in the United States has become increasingly popular. The prevalence of herbal use among primary care patients varies in previous studies; the pattern of herbal use among urban racially/ethnically diverse primary care patients has not been widely studied. The primary objectives of this study were to describe the use of herbs by ethnically diverse primary care patients in a large metropolitan area and to examine factors associated with such use. The secondary objective was to investigate perceptions about and patterns of herbal use. METHODS: Data for a cross-sectional survey were collected at primary care practices affiliated with the Southern Primary-care Urban Research Network (SPUR-Net) in Houston, Texas, from September 2002 to March 2003. To participate in the study, patients had to be at least 18 years of age and visiting one of the SPUR-Net clinics for routine, nonacute care. Survey questions were available in both English and Spanish. RESULTS: A total of 322 patients who had complete information on race/ethnicity were included in the analysis. Overall, 36% of the surveyed patients (n = 322) indicated use of herbs, with wide variability among ethnic groups: 50% of Hispanics, 50% of Asians, 41% of Whites, and 22% of African-Americans. Significant factors associated with an individual's herbal use were ethnicity other than African-American, having an immigrant family history, and reporting herbal use by other family members. About 40% of survey respondents believed that taking prescription medications and herbal medicines together was more effective than taking either alone. One-third of herbal users reported using herbs on a daily basis. More Whites (67%) disclosed their herbal use to their health-care providers than did African-Americans (45%), Hispanics (31%), or Asians (31%). CONCLUSIONS: Racial/ethnic differences in herbal use were apparent among this sample of urban multiethnic adult primary care patients. Associated factors of herbal use were non-African-American ethnicity, immigrant family history, and herbal use among family members. Whereas Hispanics and Asians reported the highest rates of herbal use, they were the least likely to disclose their use to health-care professionals. These findings are important for ensuring medication safety in primary care practices

Comparing the efficacy of a web-assisted calprotectin-based treatment algorithm (IBD-live) with usual practices in teenagers with inflammatory bowel disease:study protocol for a randomized controlled trial

Background: To prevent clinical relapse in teenagers with inflammatory bowel disease (IBD) there is a need to monitor disease activity continuously. Timely optimisation of medical treatment may nip a preclinical relapse in the bud and change the natural course of IBD. Traditionally, disease monitoring is done during scheduled visits, but this is when most teenagers report full control. IBD care could be more efficient if patients were seen at times of clinical need. This study aims to examine the effectiveness of a web-assisted calprotectin-based treatment algorithm (IBD-live) compared with usual practices in teenagers with IBD. Methods/design: A randomized trial of web-based disease monitoring versus usual care is conducted at 10 Dutch IBD care centers. We plan to recruit 180 patients between 10-and 19-years old with quiescent IBD at baseline. Teenagers assigned to IBD-live will use the flarometer -an automatic cumulation of disease activity and fecal calprotectin measurements-to estimate probability of relapse. In case the flarometer indicates high risk the patient requires treatment intensification in accordance with national guidelines; low risk means that maintenance therapy is unchanged; and intermediate risk requires optimisation of drug adherence. Patients assigned to usual practice get the best accepted medical care with regular health checks. Primary outcome is the frequency of relapse at 52 weeks of follow-up. The diagnosis of relapse is based on a clinical activity index score >10 points necessitating remission induction therapy. Secondary outcomes include quality of life and cost-effectiveness. Discussion: Web-assisted monitoring of disease activity with rapid access for those with acute relapse may allow teenagers to develop skills that are required of adult patients (including communication and self-determination). Similar monitoring systems have been introduced for teenagers with asthma and diabetes, with a positive effect on disease control, but the intervention has not been evaluated in teenagers with IBD. A randomized trial in adult patients with ulcerative colitis showed that a web-assisted treatment algorithm is feasible, safe and cost-effective. Results of the current trial are expected to have important implications for teenagers with IBD that incurs substantial health burdens and economic costs

Comparative Analysis of Gene Regulation by the Transcription Factor PPARα between Mouse and Human

Studies in mice have shown that PPARalpha is an important regulator of hepatic lipid metabolism and the acute phase response. However, little information is available on the role of PPARalpha in human liver. Here we set out to compare the function of PPARalpha in mouse and human hepatocytes via analysis of target gene regulation

The integration of lipid-sensing and anti-inflammatory effects: how the PPARs play a role in metabolic balance

The peroxisomal proliferating-activated receptors (PPARs) are lipid-sensing transcription factors that have a role in embryonic development, but are primarily known for modulating energy metabolism, lipid storage, and transport, as well as inflammation and wound healing. Currently, there is no consensus as to the overall combined function of PPARs and why they evolved. We hypothesize that the PPARs had to evolve to integrate lipid storage and burning with the ability to reduce oxidative stress, as energy storage is essential for survival and resistance to injury/infection, but the latter increases oxidative stress and may reduce median survival (functional longevity). In a sense, PPARs may be an evolutionary solution to something we call the 'hypoxia-lipid' conundrum, where the ability to store and burn fat is essential for survival, but is a 'double-edged sword', as fats are potentially highly toxic. Ways in which PPARs may reduce oxidative stress involve modulation of mitochondrial uncoupling protein (UCP) expression (thus reducing reactive oxygen species, ROS), optimising forkhead box class O factor (FOXO) activity (by improving whole body insulin sensitivity) and suppressing NFkB (at the transcriptional level). In light of this, we therefore postulate that inflammation-induced PPAR downregulation engenders many of the signs and symptoms of the metabolic syndrome, which shares many features with the acute phase response (APR) and is the opposite of the phenotype associated with calorie restriction and high FOXO activity. In genetically susceptible individuals (displaying the naturally mildly insulin resistant 'thrifty genotype'), suboptimal PPAR activity may follow an exaggerated but natural adipose tissue-related inflammatory signal induced by excessive calories and reduced physical activity, which normally couples energy storage with the ability to mount an immune response. This is further worsened when pancreatic decompensation occurs, resulting in gluco-oxidative stress and lipotoxicity, increased inflammatory insulin resistance and oxidative stress. Reactivating PPARs may restore a metabolic balance and help to adapt the phenotype to a modern lifestyle

Systematic Evaluation of Pleiotropy Identifies 6 Further Loci Associated With Coronary Artery Disease

Background: Genome-wide association studies have so far identified 56 loci associated with risk of coronary artery disease (CAD). Many CAD loci show pleiotropy; that is, they are also associated with other diseases or traits. Objectives: This study sought to systematically test if genetic variants identified for non-CAD diseases/traits also associate with CAD and to undertake a comprehensive analysis of the extent of pleiotropy of all CAD loci. Methods: In discovery analyses involving 42,335 CAD cases and 78,240 control subjects we tested the association of 29,383 common (minor allele frequency >5%) single nucleotide polymorphisms available on the exome array, which included a substantial proportion of known or suspected single nucleotide polymorphisms associated with common diseases or traits as of 2011. Suggestive association signals were replicated in an additional 30,533 cases and 42,530 control subjects. To evaluate pleiotropy, we tested CAD loci for association with cardiovascular risk factors (lipid traits, blood pressure phenotypes, body mass index, diabetes, and smoking behavior), as well as with other diseases/traits through interrogation of currently available genome-wide association study catalogs. Results: We identified 6 new loci associated with CAD at genome-wide significance: on 2q37 (KCNJ13-GIGYF2), 6p21 (C2), 11p15 (MRVI1-CTR9), 12q13 (LRP1), 12q24 (SCARB1), and 16q13 (CETP). Risk allele frequencies ranged from 0.15 to 0.86, and odds ratio per copy of the risk allele ranged from 1.04 to 1.09. Of 62 new and known CAD loci, 24 (38.7%) showed statistical association with a traditional cardiovascular risk factor, with some showing multiple associations, and 29 (47%) showed associations at p < 1 × 10−4 with a range of other diseases/traits. Conclusions: We identified 6 loci associated with CAD at genome-wide significance. Several CAD loci show substantial pleiotropy, which may help us understand the mechanisms by which these loci affect CAD risk

Increased Systemic Th17 Cytokines Are Associated with Diastolic Dysfunction in Children and Adolescents with Diabetic Ketoacidosis

Diastolic dysfunction suggestive of diabetic cardiomyopathy is established in children with T1DM, but its pathogenesis is not well understood. We studied the relationships of systemic inflammatory cytokines/chemokines and cardiac function in 17 children with T1DM during and after correction of diabetic ketoacidosis (DKA). Twenty seven of the 39 measured cytokines/chemokines were elevated at 6–12 hours into treatment of DKA compared to values after DKA resolution. Eight patients displayed at least one parameter of diastolic abnormality (DA) during acute DKA. Significant associations were present between nine of the cytokine/chemokine levels and the DA over time. Interestingly, four of these nine interactive cytokines (GM-CSF, G-CSF, IL-12p40, IL-17) are associated with a Th17 mediated cell response. Both the DA and CCL7 and IL-12p40, had independent associations with African American patients. Thus, we report occurrence of a systemic inflammatory response and the presence of cardiac diastolic dysfunction in a subset of young T1DM patients during acute DKA