Vibrio cholerae vexH Encodes a Multiple Drug Efflux Pump That Contributes to the Production of Cholera Toxin and the Toxin Co-Regulated Pilus

The resistance-nodulation-division (RND) efflux systems are ubiquitous transporters that function in antimicrobial resistance. Recent studies showed that RND systems were required for virulence factor production in Vibrio cholerae. The V. cholerae genome encodes six RND efflux systems. Three of the RND systems (VexB, VexD, and VexK) were previously shown to be redundant for in vitro resistance to bile acids and detergents. A mutant lacking the VexB, VexD, and VexK RND pumps produced wild-type levels of cholera toxin (CT) and the toxin co-regulated pilus (TCP) and was moderately attenuated for intestinal colonization. In contrast, a RND negative mutant produced significantly reduced amounts of CT and TCP and displayed a severe colonization defect. This suggested that one or more of the three uncharacterized RND efflux systems (i.e. VexF, VexH, and VexM) were required for pathogenesis. In this study, a genetic approach was used to generate a panel of V. cholerae RND efflux pump mutants in order to determine the function of VexH in antimicrobial resistance, virulence factor production, and intestinal colonization. VexH contributed to in vitro antimicrobial resistance and exhibited a broad substrate specificity that was redundant with the VexB, VexD, and VexK RND efflux pumps. These four efflux pumps were responsible for in vitro antimicrobial resistance and were required for virulence factor production and intestinal colonization. Mutation of the VexF and/or VexM efflux pumps did not affect in vitro antimicrobial resistance, but did negatively affect CT and TCP production. Collectively, our results demonstrate that the V. cholerae RND efflux pumps have redundant functions in antimicrobial resistance and virulence factor production. This suggests that the RND efflux systems contribute to V. cholerae pathogenesis by providing the bacterium with protection against antimicrobial compounds that are present in the host and by contributing to the regulated expression of virulence factors

Validity and reproducibility of an interviewer-administered food frequency questionnaire for healthy French-Canadian men and women

OBJECTIVE: To evaluate the validity (study 1) and the reproducibility (study 2) of an interviewer-administered food frequency questionnaire (FFQ). METHOD: The FFQ was designed at Laval University and contains 91 items and 33 subquestions. Study 1: The FFQ was compared against a 3-day food record (2 week-days and 1 weekend-day), at week 0, 6 and 12 of a nutritional intervention. Study 2: In order to evaluate the reproducibility of the FFQ, 2 registered dietitians administered the FFQ 4-weeks apart among subjects who were not part of the nutritional intervention. RESULTS: Study 1: Mean values for intake of most nutrients assessed by the FFQ and by the 3-day food record were not statistically different. Energy-adjusted correlation coefficients for major macronutrients ranged from 0.36 for proteins to 0.60 for carbohydrates (p ≤ 0.01). Agreement analysis revealed that on average, 35% of the subjects were classified in the same quartile when nutrients were assessed by either the 3-day food record or the FFQ. Study 2: Significant associations were observed between dietary measurements derived from the two FFQs administered 4 weeks apart. Correlation coefficients for the reproducibility of macronutrients ranged from 0.66 for carbohydrates to 0.83 for lipids after energy adjustment. On average, 46% of the subjects were classified in the same quartile when nutrient intakes were assessed by either FFQ. CONCLUSION: These data indicated that the FFQ developed has a good validity and is reproducible

Programmable Ligand Detection System in Plants through a Synthetic Signal Transduction Pathway

There is an unmet need to monitor human and natural environments for substances that are intentionally or unintentionally introduced. A long-sought goal is to adapt plants to sense and respond to specific substances for use as environmental monitors. Computationally re-designed periplasmic binding proteins (PBPs) provide a means to design highly sensitive and specific ligand sensing capabilities in receptors. Input from these proteins can be linked to gene expression through histidine kinase (HK) mediated signaling. Components of HK signaling systems are evolutionarily conserved between bacteria and plants. We previously reported that in response to cytokinin-mediated HK activation in plants, the bacterial response regulator PhoB translocates to the nucleus and activates transcription. Also, we previously described a plant visual response system, the de-greening circuit, a threshold sensitive reporter system that produces a visual response which is remotely detectable and quantifiable.We describe assembly and function of a complete synthetic signal transduction pathway in plants that links input from computationally re-designed PBPs to a visual response. To sense extracellular ligands, we targeted the computational re-designed PBPs to the apoplast. PBPs bind the ligand and develop affinity for the extracellular domain of a chemotactic protein, Trg. We experimentally developed Trg fusions proteins, which bind the ligand-PBP complex, and activate intracellular PhoR, the HK cognate of PhoB. We then adapted Trg-PhoR fusions for function in plants showing that in the presence of an external ligand PhoB translocates to the nucleus and activates transcription. We linked this input to the de-greening circuit creating a detector plant.Our system is modular and PBPs can theoretically be designed to bind most small molecules. Hence our system, with improvements, may allow plants to serve as a simple and inexpensive means to monitor human surroundings for substances such as pollutants, explosives, or chemical agents

Maternal Undernutrition and Long-term Effects on Hepatic Function

Undernutrition in utero, regardless of the source, can impair proper liver development leading to long-term metabolic dysfunction. Understanding the molecular mechanisms underlying how nutritional deficits during perinatal life lead to permanent alterations in hepatic gene expression will provide better therapeutic strategies to alleviate the undernourished liver in postnatal life. This chapter addresses the different experimental models of undernutrition in utero, and highlights the direct and indirect mechanisms involved leading to metabolic diseases in the liver. These include hypoxia, oxidative stress, epigenetic alterations, and endoplasmic reticulum (ER) stress. In addition, promising perinatal nutritional and pharmaceutical interventions are highlighted which illustrate how the placidity of the developing liver can be exploited to prevent the onset of long-term metabolic disease

Atherogenic Dyslipidemia: Cardiovascular Risk and Dietary Intervention

Atherogenic dyslipidemia comprises a triad of increased blood concentrations of small, dense low-density lipoprotein (LDL) particles, decreased high-density lipoprotein (HDL) particles, and increased triglycerides. A typical feature of obesity, the metabolic syndrome, insulin resistance, and type 2 diabetes mellitus, atherogenic dyslipidemia has emerged as an important risk factor for myocardial infarction and cardiovascular disease. A number of genes have now been linked to this pattern of lipoprotein changes. Low-carbohydrate diets appear to have beneficial lipoprotein effects in individuals with atherogenic dyslipidemia, compared to high-carbohydrate diets, whereas the content of total fat or saturated fat in the diet appears to have little effect. Achieving a better understanding of the genetic and dietary influences underlying atherogenic dyslipidemia may provide clues to improved interventions to reduce the risk of cardiovascular disease in high-risk individuals

Keratin 8/18 Regulation of Cell Stiffness-Extracellular Matrix Interplay through Modulation of Rho-Mediated Actin Cytoskeleton Dynamics

Cell mechanical activity generated from the interplay between the extracellular matrix (ECM) and the actin cytoskeleton is essential for the regulation of cell adhesion, spreading and migration during normal and cancer development. Keratins are the intermediate filament (IF) proteins of epithelial cells, expressed as pairs in a lineage/differentiation manner. Hepatic epithelial cell IFs are made solely of keratins 8/18 (K8/K18), hallmarks of all simple epithelia. Notably, our recent work on these epithelial cells has revealed a key regulatory function for K8/K18 IFs in adhesion/migration, through modulation of integrin interactions with ECM, actin adaptors and signaling molecules at focal adhesions. Here, using K8-knockdown rat H4 hepatoma cells and their K8/K18-containing counterparts seeded on fibronectin-coated substrata of different rigidities, we show that the K8/K18 IF-lacking cells lose their ability to spread and exhibit an altered actin fiber organization, upon seeding on a low-rigidity substratum. We also demonstrate a concomitant reduction in local cell stiffness at focal adhesions generated by fibronectin-coated microbeads attached to the dorsal cell surface. In addition, we find that this K8/K18 IF modulation of cell stiffness and actin fiber organization occurs through RhoA-ROCK signaling. Together, the results uncover a K8/K18 IF contribution to the cell stiffness-ECM rigidity interplay through a modulation of Rho-dependent actin organization and dynamics in simple epithelial cells

Effect of rosuvastatin on outcomes in chronic haemodialysis patients – design and rationale of the AURORA study

BACKGROUND: Patients with end-stage renal disease (ESRD) are at high risk of cardiovascular events. Multiple risk factors for atherosclerosis are present in ESRD and may contribute to the increased risk of cardiovascular mortality in this population. In contrast to patients with normal renal function, the benefits of modifying lipid levels on cardiovascular outcomes in patients with ESRD on haemodialysis have yet to be confirmed in large prospective randomised trials. A study to evaluate the Use of Rosuvastatin in subjects On Regular haemodialysis: an Assessment of survival and cardiovascular events (AURORA) will be the first large-scale international trial to assess the effects of statin therapy on cardiovascular morbidity and mortality in ESRD patients on chronic haemodialysis. METHODS: More than 2,750 ESRD patients who have been receiving chronic haemodialysis treatment for at least 3 months have been randomised (1:1), irrespective of baseline lipid levels, to treatment with rosuvastatin 10 mg or placebo. The primary study endpoint is the time to a major cardiovascular event (first occurrence of cardiovascular death, non-fatal myocardial infarction or non-fatal stroke). Secondary endpoints include all-cause mortality, major cardiovascular event-free survival time, time to cardiovascular death, time to non-cardiovascular death, cardiovascular interventions, tolerability of treatment and health economic costs per life-year saved. Study medication will be given until 620 subjects have experienced a major cardiovascular event. CONCLUSION: Our hypothesis is that results from AURORA will establish the clinical efficacy and tolerability of rosuvastatin in patients with ESRD receiving chronic haemodialysis and guide the optimal management of this expanding population

Assessing the evolution of primary healthcare organizations and their performance (2005-2010) in two regions of Québec province: Montréal and Montérégie

<p>Abstract</p> <p>Background</p> <p>The Canadian healthcare system is currently experiencing important organizational transformations through the reform of primary healthcare (PHC). These reforms vary in scope but share a common feature of proposing the transformation of PHC organizations by implementing new models of PHC organization. These models vary in their performance with respect to client affiliation, utilization of services, experience of care and perceived outcomes of care.</p> <p>Objectives</p> <p>In early 2005 we conducted a study in the two most populous regions of Quebec province (Montreal and Montérégie) which assessed the association between prevailing models of primary healthcare (PHC) and population-level experience of care. The <b>goal </b>of the present research project is to track the <it>evolution </it>of PHC organizational models and their relative performance through the reform process (from 2005 until 2010) and to assess factors at the organizational and contextual levels that are associated with the transformation of PHC organizations and their performance.</p> <p>Methods/Design</p> <p>This study will consist of three interrelated surveys, hierarchically nested. The first survey is a population-based survey of randomly-selected adults from two populous regions in the province of Quebec. This survey will assess the current affiliation of people with PHC organizations, their level of utilization of healthcare services, attributes of their experience of care, reception of preventive and curative services and perception of unmet needs for care. The second survey is an organizational survey of PHC organizations assessing aspects related to their vision, organizational structure, level of resources, and clinical practice characteristics. This information will serve to develop a taxonomy of organizations using a mixed methods approach of factorial analysis and principal component analysis. The third survey is an assessment of the organizational context in which PHC organizations are evolving. The five year prospective period will serve as a natural experiment to assess contextual and organizational factors (in 2005) associated with migration of PHC organizational models into new forms or models (in 2010) and assess the impact of this evolution on the performance of PHC.</p> <p>Discussion</p> <p>The results of this study will shed light on changes brought about in the organization of PHC and on factors associated with these changes.</p