Platelet transfusions in a murine model of neonatal polymicrobial sepsis: Divergent effects on inflammation and mortality.

BACKGROUND: Platelet transfusions (PTxs) are often given to septic preterm neonates at high platelet count thresholds in an attempt to reduce bleeding risk. However, the largest randomized controlled trial (RCT) of neonatal transfusion thresholds found higher mortality and/or major bleeding in infants transfused at higher thresholds. Using a murine model, we investigated the effects of adult PTx on neonatal sepsis-induced mortality, systemic inflammation, and platelet consumption. STUDY DESIGN AND METHODS: Polymicrobial sepsis was induced via intraperitoneal injection of cecal slurry preparations (CS1, 2, 3) into P10 pups. Two hours after infection, pups were transfused with washed adult Green Flourescent Protein (GFP+) platelets or control. Weights, platelet counts, and GFP% were measured before 4 and 24 h post-infection. At 24 h, blood was collected for quantification of plasma cytokines. RESULTS: The CS batches varied in 24 h mortality (11%, 73%, and 30% in CS1, 2, and 3, respectively), due to differences in bacterial composition. PTx had differential effects on sepsis-induced mortality and systemic inflammatory cytokines, increasing both in mice infected with CS1 (low mortality) and decreasing both in mice infected with CS2 and 3. In a mathematical model of platelet kinetics, the consumption of transfused adult platelets was higher than that of endogenous neonatal platelets, regardless of CS batch. DISCUSSION: Our findings support the hypothesis that transfused adult platelets are consumed faster than endogenous neonatal platelets in sepsis and demonstrate that PTx can enhance or attenuate neonatal inflammation and mortality in a model of murine polymicrobial sepsis, depending on the composition of the inoculum and/or the severity of sepsis.

Discovery of Icenticaftor (QBW251), a Cystic Fibrosis Transmembrane Conductance Regulator Potentiator with Clinical Efficacy in Cystic Fibrosis and Chronic Obstructive Pulmonary Disease.

Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) ion channel are established as the primary causative factor in the devastating lung disease cystic fibrosis (CF). More recently, cigarette smoke exposure has been shown to be associated with dysfunctional airway epithelial ion transport, suggesting a role for CFTR in the pathogenesis of chronic obstructive pulmonary disease (COPD). Here, the identification and characterization of a high throughput screening hit 6 as a potentiator of mutant human F508del and wild-type CFTR channels is reported. The design, synthesis, and biological evaluation of compounds 7-33 to establish structure-activity relationships of the scaffold are described, leading to the identification of clinical development compound icenticaftor (QBW251) 33, which has subsequently progressed to deliver two positive clinical proofs of concept in patients with CF and COPD and is now being further developed as a novel therapeutic approach for COPD patients.

Lysophospholipid micelles sustain the stability and catalytic activity of diacylglycerol kinase in the absence of lipids.

There has been a renewal of interest in interactions of membrane proteins with detergents and lipids, sparked both by recent results that illuminate the structural details of these interactions and also by the realization that some experimental membrane protein structures are distorted by detergent-protein interactions. The integral membrane enzyme diacylglycerol kinase (DAGK) has long been thought to require the presence of lipid as an obligate "cofactor" in order to be catalytically viable in micelles. Here, we report that near-optimal catalytic properties are observed for DAGK in micelles composed of lysomyristoylphosphatidylcholine (LMPC), with significant activity also being observed in micelles composed of lysomyristoylphosphatidylglycerol and tetradecylphosphocholine. All three of these detergents were also sustained high stability of the enzyme. NMR measurements revealed significant differences in DAGK-detergent interactions involving LMPC micelles versus micelles composed of dodecylphosphocholine. These results highlight the fact that some integral membrane proteins can maintain native-like properties in lipid-free detergent micelles and also suggest that C(14)-based detergents may be worthy of more widespread use in studies of membrane proteins.

Aspirin dosing for the prevention and treatment of ischemic stroke: an indication-specific review of the literature.

OBJECTIVE: To evaluate the efficacy of aspirin for the treatment and prevention of ischemic stroke and identify the minimum dose proven to be effective for each indication. DATA SOURCES: PubMed and MEDLINE searches (up to January 2010) were performed to identify primary literature, using search terms including aspirin, stroke prevention, acute ischemic stroke, acetylsalicylic acid, atrial fibrillation, myocardial infarction, and carotid endarterectomy. Additionally, reference citations from publications identified were reviewed. STUDY SELECTION AND DATA EXTRACTION: Articles published in English were evaluated and relevant primary literature evaluating the efficacy of aspirin in the prevention of stroke was included in this review. DATA SYNTHESIS: Antiplatelet therapy is the benchmark for the prevention of ischemic stroke. Aspirin has been proven to prevent ischemic stroke in a variety of settings. Despite the frequency at which aspirin continues to be prescribed in patients at risk of ischemic stroke, there remains confusion in clinical practice as to what minimum dose is required in various at-risk patients. A thorough review of the primary literature suggests that low-dose (50-81 mg daily) aspirin is insufficient for some indications. Acute ischemic stroke treatment requires 160-325 mg, while atrial fibrillation and carotid arterial disease require daily doses of 325 and 81-325 mg, respectively. CONCLUSIONS: Available evidence suggests that aspirin dosing must be individualized according to indication. Recommendations provided by national guidelines at times recommend lower doses of aspirin than have been proven effective. Higher doses are indicated for stroke prevention in atrial fibrillation (325 mg) and acute ischemic stroke patients (160-325 mg). Aspirin has not yet been proven effective for primary prevention of strokes in men, and a minimum dose for these patients cannot be determined from the available data.

A unified hydrophobicity scale for multispan membrane proteins.

The concept of hydrophobicity is critical to our understanding of the principles of membrane protein (MP) folding, structure, and function. In the last decades, several groups have derived hydrophobicity scales using both experimental and statistical methods that are optimized to mimic certain natural phenomena as closely as possible. The present work adds to this toolset the first knowledge-based scale that unifies the characteristics of both alpha-helical and beta-barrel multispan MPs. This unified hydrophobicity scale (UHS) distinguishes between amino acid preference for solution, transition, and trans-membrane states. The scale represents average hydrophobicity values of amino acids in folded proteins, irrespective of their secondary structure type. We furthermore present the first knowledge-based hydrophobicity scale for mammalian alpha-helical MPs (mammalian hydrophobicity scale--MHS). Both scales are particularly useful for computational protein structure elucidation, for example as input for machine learning techniques, such as secondary structure or trans-membrane span prediction, or as reference energies for protein structure prediction or protein design. The knowledge-based UHS shows a striking similarity to a recent experimental hydrophobicity scale introduced by Hessa and coworkers (Hessa T et al., Nature 2007;450:U1026-U1032). Convergence of two very different approaches onto similar hydrophobicity values consolidates the major differences between experimental and knowledge-based scales observed in earlier studies. Moreover, the UHS scale represents an accurate absolute free energy measure for folded, multispan MPs--a feature that is absent from many existing scales. The utility of the UHS was demonstrated by analyzing a series of diverse MPs. It is further shown that the UHS outperforms nine established hydrophobicity scales in predicting trans-membrane spans along the protein sequence. The accuracy of the present hydrophobicity scale profits from the doubling of the number of integral MPs in the PDB over the past four years. The UHS paves the way for an increased accuracy in the prediction of trans-membrane spans.

Effect of preceptor-initiated referral system on advanced pharmacy practice experience (APPE) students' rotation progress and graduation rate.

INTRODUCTION: The objective of this study was to determine whether the referral and monitoring system developed at Butler University College of Pharmacy and Health Sciences (BUCOPHS) for advanced pharmacy practice experience (APPE) students is successful in improving on-time and overall graduation rates and decreasing rotation failures. METHODS: In May 2014, a formalized process for referral and monitoring of APPE students was established. This allows preceptors to recommend (via the electronic final evaluation form) additional monitoring or intervention in the areas of professionalism, time management, drug information, communication, and therapeutic/drug knowledge. The experiential education director subsequently meets with referred students and develops a customized longitudinal plan to help each student improve in the identified areas. Data collected for students who were referred included the rotation block, rotation type, preceptor type, and the specific areas recommended for monitoring and referral. In addition, each student's quarterly progress, specific assessments outlined in each student's customized longitudinal plan, number of failed rotations, on-time graduation, and overall graduation status was recorded. The university's institutional review board approved the study. RESULTS: Between May 2014 and April 2017, a total of 36 students were referred to the experiential education office for monitoring and/or intervention. Of these, 35/36 (97.2%) graduated on time. There were eight students who failed one APPE; no students received more than one failure. CONCLUSIONS: Development of a referral and monitoring system for preceptors aids in retention of professional pharmacy students and increases on-time and overall graduation rates.

Global Transcriptomic Analysis of the Candida albicans Response to Treatment with a Novel Inhibitor of Filamentation.

The opportunistic pathogenic fungus Candida albicans can cause devastating infections in immunocompromised patients. Its ability to undergo a morphogenetic transition from yeast to filamentous forms allows it to penetrate tissues and damage tissues, and the expression of genes associated with a number of pathogenetic mechanisms is also coordinately regulated with the yeast-to-hypha conversion. Therefore, it is widely considered that filamentation represents one of the main virulence factors of C. albicans We have previously identified N-[3-(allyloxy)-phenyl]-4-methoxybenzamide (compound 9029936) as the lead compound in a series of small-molecule inhibitors of C. albicans filamentation and characterized its activity both in vitro and in vivo This compound appears to be a promising candidate for the development of alternative antivirulence strategies for the treatment of C. albicans infections. In this study, we performed RNA sequencing analysis of samples obtained from C. albicans cells grown under filament-inducing conditions in the presence or absence of this compound. Overall, treatment with compound 9029936 resulted in 618 upregulated and 702 downregulated genes. Not surprisingly, some of the most downregulated genes included well-characterized genes associated with filamentation and virulence such as SAP5, ECE1 (candidalysin), and ALS3, as well as genes that impact metal chelation and utilization. Gene ontology analysis revealed an overrepresentation of cell adhesion, iron transport, filamentation, biofilm formation, and pathogenesis processes among the genes downregulated during treatment with this leading compound. Interestingly, the top upregulated genes suggested an enhancement of vesicular transport pathways, particularly those involving SNARE interactions.IMPORTANCE These results from whole-genome transcriptional profiling provide further insights into the biological activity and mode of action of a small-molecule inhibitor of C. albicans filamentation. This information will assist in the development of novel antivirulence strategies against C. albicans infections.

pH-Modulating Poly(ethylene glycol)/Alginate Hydrogel Dressings for the Treatment of Chronic Wounds.

The development of chronic wounds has been frequently associated with alkaline pH values. The application of pH-modulating wound dressings can, therefore, be a promising treatment option to promote normal wound healing. This study reports on the development and characterization of acidic hydrogel dressings based on interpenetrating poly(ethylene glycol) diacrylate/acrylic acid/alginate networks. The incorporation of ionizable carboxylic acid groups results in high liquid uptake up to 500%. The combination of two separate polymer networks significantly improves the tensile and compressive stability. In a 2D cell migration assay, the application of hydrogels (0% to 1.5% acrylic acid) results in complete "wound" closure; hydrogels with 0.25% acrylic acid significantly increase the cell migration velocity to 19.8 ± 1.9 µm h-1 . The most promising formulation (hydrogels with 0.25% acrylic acid) is tested on 3D human skin constructs, increasing keratinocyte ingrowth into the wound by 164%.

Alkaline poly(ethylene glycol)-based hydrogels for a potential use as bioactive wound dressings.

The number of patients with chronic wounds is increasing constantly in today's aging society. However, little work is done so far tackling the associated disadvantageous shift of the wound pH. In our study, we developed two different approaches on pH-modulating wound dressing materials, namely, bioactive interpenetrating polymer network hydrogels based on poly(ethylene glycol) diacrylate/N-vinylimidazole/alginate (named VIx ) and poly(ethylene glycol) diacrylate/2-dimethylaminoethyl methacrylate/N-carboxyethylchitosan (named DMAEMAx ). Both formulations showed a good cytocompatibility and wound healing capacity in vitro. The developed dressing materials significantly increased the cell ingrowth in wounded human skin constructs; by 364% and 313% for the VIx and the DMAEMAx hydrogel formulation, respectively. Additionally, VIx hydrogels were found to be suitable scaffolds for superficial cell attachment. Our research on the material properties suggests that ionic interactions and hydrogen bonds are the driving forces for the mechanical and swelling properties of the examined hydrogels. High amounts of positively charged amino groups in DMAEMAx hydrogels caused increased liquid uptake (around 190%), whereas VIx hydrogels showed a 10-fold higher maximum compressive stress in comparison to hydrogels without ionizable functional groups. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 3360-3368, 2017.

Improved prediction of trans-membrane spans in proteins using an Artificial Neural Network.

Tools for the identification of trans-membrane spans from the protein sequence are widely used in the experimental community. Computational structural biology seeks to increase the prediction accuracy of such methods since they represent a first step towards membrane protein tertiary structure prediction from the amino acid sequence. We introduce a predictor that is able to identify trans-membrane spans from the sequence of a protein. The novelty of the approach presented here is the simultaneous prediction of trans-membrane spanning α-helices and ß-strands within a single tool. An artificial neural network was trained on databases of 102 membrane proteins and 3499 soluble proteins. Prediction accuracies of up to 92% for soluble residues, 75% for residues in the interface, and 73% for TM residues are achieved. On average the algorithm predicts 79% of the residues correctly which is a substantial improvement from a previously published implementation which achieved 57% accuracy (Koehler et al., Proteins: Structure, Function, and Bioinformatics, 2008). The algorithm was applied to four membrane proteins to illustrate the applicability to both α-helical bundles and ß-barrels.

Patient-directed medicine labeling: text differences between the United States and Spain.

As part of a larger project to examine the complexities of patient adherence to medical directions, the current study compares written information given with prescribed medicines to patients in the United States (US) and Spain suffering from two chronic diseases. First, the legal context of the voluntary nature of the provision of such information in the US, and the mandatory nature of provision in Spain, was explored. Then, 30 drugs in common use in both countries, 19 for cardiovascular disease and 11 for endocrine disorders, were identified and the texts of the corresponding patient-directed written information were included in an electronic corpus and analysed. Ten rhetorical features common to both the US and Spanish texts were identified. Differences were found in the placement of two of the features in relation to the complete text: information on 'side effects' and concerning 'the need to seek medical advice' appeared more frequently throughout the US texts than in the Spanish texts. Detailed lexical analysis showed more technical vocabulary in use in the Spanish texts.