The vasodilator-stimulated phosphoprotein-phosphorylation (VASP-P) flow-cytometric assay is mainly used in clinical trials to measure thienopyridine effects. However, there are remarkable differences in the reported optimal cut-offs, ranging from 48-61% platelet reactivity index (PRI). We therefore investigated whether a lack of standardisation might explain the differences in the cut-offs. We measured VASP-P in 62 individuals. PRI was calculated using the mean, geometric mean and median fluorescence intensities (FI). Stability of the blood-samples (time-to-assay, 0-2 days) and stability of the processed samples (0-120 minutes) within the recommended time-span were tested. Time-to-assay significantly influenced the PRI (p<0.001): the PRI from mean FI after two days was lower compared to values on day 1 (52 ± 22.9 vs. 57.7 ± 24.1, p<0.001). The PRI from the geometric mean FI after two days was lower compared to day 0 as well as day 1 (51.3 ± 23 vs. 58.2 ± 24.2 and vs. 59.1 ± 23.7, both p<0.001). The PRI from median FI was stable over time (day 0: 59.1 ± 25%, day 1: 59.7 ± 24.1% and day 2: 56.4 ± 23.9%, all p=ns). Furthermore, the lag time of the processed samples significantly altered the PRI (all p<0.001) with a maximum difference for PRI based on geometric mean FI after 90 minutes compared to baseline (Δ=3.92%PRI, p<0.001). The differences in the reported cut-offs might be explained by a lack of standardisation. More precise standardisation is inevitable, as the PRI significantly depends on the method of calculation, the time-to-assay as well as on the lag time after processing. Tolerably stable results were obtained for the PRI from the median FI.

Background: Prognostic values of genotyping and phenotyping for assessment of clopidogrel responsiveness have been shown in independent studies. Objectives: To compare different assays for prediction of events during long-term follow-up.Methods: In this prospective cohort study polymorphisms of CYP2C19*2 and CYP2C19*17 alleles, vasodilator-stimulated phosphoprotein phosphorylation (VASP) assay, Multiple Electrode Aggregometry (MEA), Cone and Platelet Analyser (CPA) and Platelet Function Analyser (PFA-100) were performed in 416 patients undergoing percutaneous coronary intervention. The rates of events were recorded during a 12-month follow-up. Results: Platelet aggregation by MEA predicted stent thrombosis (2.4%) better (c-index=0.90; p<0.001; sensitivity=90%; specificity=83%) than the VASP assay, CPA or PFA-100 (c-index<0.70; p>0.05; sensitivity<70%; specificity<70% for all) or even the CYP2C19*2 polymorphism (c-index<0.56; p>0.05; sensitivity=30%; specificity=71%). Survival analysis indicated that patients classified as poor-responders by MEA had a substantially higher risk to develop stent thrombosis or MACE than clopidogrel responders (12.5% vs. 0.3%; p<0.001 and 18.5% vs. 11.3%; p=0.022; respectively), whereas poor metabolisers (CYP2C19*1/*2 or *2/*2 carriers) were not at increased risks (stent thrombosis: 2.7% vs. 2.5%; p>0.05; MACE: 13.5% vs. 12.1%; p=0.556; respectively). The incidence of major bleedings (2.6%) was numerically higher in patients with an enhanced vs. poor response to clopidogrel assessed by MEA (4% vs. 0%) or in ultra-metabolisers vs. regular-metabolisers (CYP2C19*17/*17 vs. CYP2C19*1/*1; 9.5% vs. 2%). The classification tree analysis demonstrated that acute coronary syndrome at hospitalisation and diabetes mellitus were the best discriminators for clopidogrel responder status. Conclusions: Phenotyping of platelet response to clopidogrel was a better predictor of stent thrombosis than genotyping.

A detailed in-situ scattering study has been carried out on the formation of amorphous calcium carbonate (ACC) particles modulated by the presence of small amounts of sodium polyacrylate chains. The work is aiming at an insight into the modulation of ACC formation by means of two polyacrylate samples differing in their molecular weight by a factor of 50. The ACC formation process was initiated by an in-situ generation of CO32- ions via hydrolysis of 10 mM dimethylcarbonate in the presence of 10 mM CaCl2. Analysis of the formation process by means of time-resolved small angle X-ray and light scattering in the absence of any additives provided evidence for a monomer addition mechanism for the growth of ACC particles. ACC formation under these conditions sets in after a lag-period of some 350 s. In the presence of sodium polyacrylate chains, calcium polyacrylate aggregates are formed during the lag-period, succeeded by a modulated ACC growth in a second step. The presence of anionic polyacrylate chains changed the shape of the growing particles toward loose and less homogeneous entities. In the case of low amounts (1.5-7.5 mg/l) of the long chain additive with 97 kDa, the size of the aggregates is comparable to the size of the successively formed hybrid particles. No variation of the lag-period has been observed in this case. Use of the short chain additive with 2 kDa enabled increase of the additive concentration up to 100 mg/l and resulted in a significant increase of the lag-period. This fact, together with the finding that the resulting hybrid particles remained stable in the latter case identified short chain sodium polyacrylates as more efficient modulators than long chain polyacrylates.

Platelet glycoprotein IIb/IIIa inhibitors (GPI) are antithrombotic agents preventing the binding of fibrinogen to GP IIb/IIIa receptors. Thus, GPI interfere with interplatelet bridging mediated by fibrinogen. Currently, three generic GPI with different antithrombotic properties are available for intravenous administration: abciximab, eptifibatide, and tirofiban. The development of oral GPI was abandoned, whereas intravenous GPI were introduced in various clinical settings during the 1990s, yielding substantial benefit in the treatment of acute coronary syndromes, particularly during percutaneous coronary interventions. Results of the many randomised trials evidenced the efficacy of this drug class, though these trials were conducted prior to the emergence of modern oral antiplatelet therapy with efficient P2Y12 inhibitors. Subsequent trials failed to consolidate the strongly favourable impression of GPI, and indications for their use have been more restricted in recent years. Nonetheless, GPI may still be beneficial during coronary interventions among high-risk patients including acute ST-elevation and non-ST-elevation myocardial infarctions, particularly in the absence of adequate pretreatment with oral antiplatelet drugs or when direct thrombin inhibitors are not utilised. Intracoronary GPI administration has been suggested as adjunctive therapy during primary percutaneous coronary intervention, and the results of larger ongoing trials are expected to elucidate its clinical potential. The present review outlines the key milestones of GPI development and provides an up-to-date overview of the clinical applicability of these drugs in the era of refined coronary stenting, potent antithrombotic drugs, and novel thrombin inhibiting agents.

In ruminant feeding, the reduction of dietary protein is an effective approach for decreasing the excretion of N. In non-ruminant species, the intestinal absorption of Ca was affected when dietary protein was reduced. Therefore, it was the aim of the present study to characterise the intestinal absorption of Ca and inorganic phosphate (Pi) in goats fed different N and Ca diets. Intestinal flux rates of Ca and Pi were determined in goats fed a reduced N and Ca diet by Ussing chamber experiments. For a more mechanistic approach, the uptake of Ca and Pi in intestinal brush-border membrane vesicles (BBMV), the expression levels of the epithelial Ca channel transient receptor potential vanilloid channel type 6 (TRPV6), the sodium-dependent Pi transporter (NaPi) IIb and the vitamin D receptor (VDR) were measured. In goats fed a reduced N and Ca diet, the intestinal flux rates of Ca and Pi were elevated. However, the reduced N and Ca diet had no effect on the uptake of Ca and Pi in intestinal BBMV, while the expression of TRPV6 and NaPi IIb protein in the corresponding intestinal segments was even decreased. The mRNA expression of NaPi IIb and VDR was not affected. Therefore, a post-transcriptional regulation of TRPV6 and NaPi IIb protein was suggested in goats fed a reduced N and Ca diet. From these data, it can be concluded that the intestinal absorption of Ca and Pi in growing goats was affected by changes in dietary N and Ca intake like those in single-stomached animals but differently modulated.

Metabolic flexibility is the body's ability to adapt to changing energy demand and nutrient supply. Maternal undernutrition causes growth restriction at birth and subsequent obesity development. Intriguingly, metabolic flexibility is maintained due to adaptations of muscle tissue. The aim of the present study was to investigate developmental pathways of these adaptive changes. Wistar rats received standard chow at either ad libitum (AD) or 30% of ad libitum intake (UN) throughout pregnancy. At all ages, metabolic status indicated similar insulin sensitivity in AD and UN offspring despite the development of adiposity in UN offspring at weaning. Type IIA fiber size was reduced in soleus muscle of UN offspring at weaning and they had a higher percentage of type I fibers in adulthood with a concomitantly higher oxidative capacity. Plasticity of muscle was present during the postnatal period and proposes novel pathways for the dynamic development of metabolic flexibility throughout postnatal life.

Development of both potent and selective kinase inhibitors depicts a challenging task in modern drug discovery. The innate promiscuity of kinase inhibitors largely results from ATP-mimetic binding to the kinase hinge region. We present a novel class of substituted 7,8-dichloro-1-oxo-ß-carbolines based on the distinct structural features of the alkaloid bauerine C whose kinase inhibitory activity does not rely on canonical ATP mimetic hinge interactions. Intriguingly, cocrystal structures revealed an unexpected inverted binding mode and the presence of halogen bonds with kinase backbone residues. The compounds exhibit excellent selectivity over a comprehensive panel of human protein kinases whilst inhibiting selected kinases such as the oncogenic PIM1 at low nanomolar concentrations. Together our biochemical and structural data suggest that this scaffold may serve as a valuable template for the design and development of specific inhibitors of various kinases including the PIM family of kinases, CLKs, DAPK3 (ZIPK), BMPK2, and others.

The kinetics and mechanism of crystallization of the dense zinc imidazolate framework with zni topology, from comparatively dilute methanol solutions containing Zn(NO(3))·6H(2)O and imidazole with variation of the zinc-to-imidazole ratio, were followed in situ by time-resolved static and dynamic light scattering. The light scattering data revealed that metastable primary particles of about 100 nm in diameter form rapidly upon mixing the component solutions. After a lag time that is dependent on the imidazole concentration, the primary particles aggregate into secondary particles by a monomer addition mechanism with the primary particles as the monomers. Complementary scanning electron microscopy revealed that further evolution of the secondary particles is a complex process involving polycrystalline intermediates, the non-spherical morphologies of which depend on the initial zinc-to-imidazole ratio. Time and location of the first appearance of crystalline order could so far not be established. The pure-phase ZIF-zni crystals obtained after 240 min are twins. The aspect ratio of the tetragonal crystals can be controlled via the zinc-to-imidazole ratio.

This study sought to identify the methods and associations of water storage, treatment and use among residents in the Kimana Fenced Area, Oloitokitok, Kenya for comparison with current best practices in order to develop recommendations to improve water sanitation issues in this area. In a cross-sectional study design, 330 households were randomly selected and interviewed on water storage, treatment, and use practices. Eighty two percent of observed containers met CDC guidelines for improved water storage containers. Fifty seven percent of survey respondents reported not treating their drinking water, of which 49% indicated that they believed the water was already clean. Logistic regression showed that people who believed their water was unsafe were twice more likely to treat their water than those who perceived their water to be somewhat safe (p = 0.058). Those living outside the furrows were 56% less likely to treat their water in the home compared to those living along the furrow (p = 0.023). Respondents with a pastoral lifestyle were 69% less likely to treat their water than those with a non-pastoral lifestyle (p =.009). In terms of tribe, the largest treatment disparity was noted amongst the Maasai, with only 37.7% reporting any form of treatment. Tribe, pastoral lifestyle, proximity to the furrow and socio-economic status were found to contribute to water storage method and treatment within the Kimana fence. It is critical that these factors be addressed in future water storage and treatment interventions in this area.

In this executive summary of a Consensus Document from the European Heart Rhythm Association, endorsed by the European Society of Cardiology Working Group on Thrombosis, we comprehensively review the published evidence and propose a consensus on bleeding risk assessments in atrial fibrillation (AF) patients. The main aim of the document was to summarise 'best practice' in dealing with bleeding risk in AF patients when approaching antithrombotic therapy, by addressing the epidemiology and size of the problem, and review established bleeding risk factors. We also summarise definitions of bleeding in the published literature. Patient values and preferences balancing the risk of bleeding against thromboembolism as well as the prognostic implications of bleeding are reviewed. We also provide an overview of published bleeding risk stratification and bleeding risk schema. Brief discussion of special situations (e.g. periablation, peri-devices such as implantable cardioverter defibrillators [ICD] or pacemakers, presentation with acute coronary syndromes and/or requiring percutanous coronary interventions/stents and bridging therapy) is made, as well as a discussion of the prevention of bleeds and managing bleeding complications. Finally, this document puts forwards consensus statements that may help to define evidence gaps and assist in everyday clinical practice.