Heparan sulfate phage display antibodies recognise epitopes defined by a combination of sugar sequence and cation binding.

Phage display antibodies are widely used to follow heparan sulfate (HS) expression in tissues and cells. We demonstrate by ELISA, that cations alter phage display antibody binding profiles to HS and this is mediated by changes in polysaccharide conformation, demonstrated by circular dichroism spectroscopy. Native HS structures, expressed on the cell surfaces of neuroblastoma and fibroblast cells, also exhibited altered antibody binding profiles following exposure to low mM concentrations of these cations. Phage display antibodies recognise conformationally-defined HS epitopes, rather than sequence alone, as has been assumed, and resemble proteins in being sensitive to changes in both charge distribution and conformation following binding of cations to HS polysaccharides

Are there foetal extracellular vesicles in maternal blood? Prospects for diagnostic biomarker discovery

Prenatal diagnosis of congenital disease improves clinical outcomes; however, as many as 50% of congenital heart disease cases are missed by current ultrasound screening methods. This indicates a need for improved screening technology. Extracellular vesicles (EVs) have attracted enormous interest in recent years for their potential in diagnostics. EVs mediate endocrine signalling in health and disease and are known to regulate aspects of embryonic development. Here, we critically evaluate recent evidence suggesting that EVs released from the foetus are able to cross the placenta and enter the maternal circulation. Furthermore, EVs from the mother appear to be transported in the reverse direction, whilst the placenta itself acts as a source of EVs. Experimental work utilising rodent models employing either transgenically encoded reporters or application of fluorescent tracking dyes provide convincing evidence of foetal-maternal crosstalk. This is supported by clinical data demonstrating expression of placental-origin EVs in maternal blood, as well as limited evidence for the presence of foetal-origin EVs. Together, this work raises the possibility that foetal EVs present in maternal blood could be used for the diagnosis of congenital disease. We discuss the challenges faced by researchers in translating these basic science findings into a clinical non-invasive prenatal test

Subjective executive function deficits in hazardous alcohol drinkers.

BACKGROUND: Dependent alcohol drinkers exhibit differences in the structure and function of the brain, and impairments in cognitive function, including executive functions (EFs). Less is known about the impact of non-dependent but hazardous use (that which raises the risk of harm), and it is also unclear to what extent executive impairments in this cohort affect real-world function. The current study examines the relationship between alcohol use, EF and alcohol-related problems, in the general population. METHODS: A between-groups cross-sectional design assessed EF across two levels of drinking; hazardous (Alcohol Use Disorders Identification Test (AUDIT) score of ⩾8) and non-hazardous. Alcohol drinkers (n = 666; 136 male; 524 female; six not disclosed; aged 28.02 ± 10.40 years) completed validated questionnaires online assessing subjective EF, alcohol use and alcohol-related problems. RESULTS: Organisation, Strategic Planning, Impulse Control and overall function were significantly impaired in hazardous drinkers. Furthermore, the effect of alcohol on EF, partially mediated the relationship between alcohol use and alcohol-related problems. CONCLUSION: Hazardous drinking was associated with lower subjective EF, and this mediated the effect of alcohol on alcohol-related problems. This may be due to changes in prefrontal brain regions, which could indicate greater risk for the development of alcohol dependence (AD). Future research should use additional means to assess EF in hazardous drinkers, including recovery of function, development of AD and the relationship between cognition and alcohol-related daily problems

An octahedral tetrachlorido Fe(II) complex with aminopyrazinium ligands from a serendipitous redox synthesis exhibiting magnetic exchange through non-covalent 3-D architectures

An air stable, neutral Fe(II) complex with four equatorial chlorido ligands has been stabilised through a serendipitous redox process and in situ ligand protonation. A three dimensional non-covalent network composed of halogen bonding and π-π stacking propagates magnetic exchange interactions though the lattice. The electronic structure has been investigated using DFT

MicroRNA-184 Is Induced By Store-Operated Calcium Entry And Regulates Early Keratinocyte Differentiation

Extracellular calcium (Ca2+) and store‐operated Ca2+ entry (SOCE) govern homoeostasis in the mammalian epidermis. Multiple microRNAs (miRNA) also regulate epidermal differentiation, and raised external Ca2+ modulates the expression of several such miRNAs in keratinocytes. However, little is known about the regulation of miR‐184 in keratinocytes or the roles of miR‐184 in keratinocyte differentiation. Here we report that exogenous Ca2+ stimulates miR‐184 expression in primary epidermal keratinocytes and that this occurs in a SOCE‐dependent manner. Levels of miR‐184 were raised by about 30‐fold after exposure to 1.5 mM Ca2+ for 5 days. In contrast, neither phorbol ester nor 1,25‐dihydroxyvitamin D3 had any effect on miR‐184 levels. Pharmacologic and genetic inhibitors of SOCE abrogated Ca2+‐dependent miR‐184 induction by 70% or more. Ectopic miR‐184 inhibited keratinocyte proliferation and led to a fourfold increase in the expression of involucrin, a marker of early keratinocyte differentiation. Exogenous miR‐184 also triggered a threefold rise in levels of cyclin E and doubled the levels of γH2AX, a marker of DNA double‐strand breaks. The p21 cyclin‐dependent kinase inhibitor, which supports keratinocyte growth arrest, was also induced by miR‐184. Together our findings point to an SOCE:miR‐184 pathway that targets a cyclin E/DNA damage regulatory node to facilitate keratinocyte differentiation

Incorporation of porcine adenovirus 4 fiber protein enhances infectivity of adenovirus vector on dendritic cells: Implications for immune-mediated cancer therapy

One strategy in cancer immunotherapy is to capitalize on the key immunoregulatory and antigen presenting capabilities of dendritic cells (DCs). This approach is dependent on efficient delivery of tumor specific antigens to DCs, which subsequently induce an anti-tumor T-cell mediated immune response. Human adenovirus serotype 5 (HAdV5) has been used in human studies for gene delivery, but has limited infection in DCs, which lack the proper receptors. Addition of the porcine fiber knob (PK) from porcine adenovirus type 4 to HAdV5 allows the virus to deliver genetic material via binding to glycosylated surface proteins and bypasses the coxsackie-and-adenovirus receptor required by wild-type HAdV5. In this study we explored the potential therapeutic applications of an adenovirus with PK-based tropism against cancers expressing mesothelin. Infectivity and gene transfer assays were used to compare Ad5-PK to wild-type HAdV5. Mouse models were used to demonstrate peptide specificity and T-cell responses. We show that the PK modification highly augmented infection of DCs, including the CD141+ DC subset, a key subset for activation of naïve CD8+ T-cells. We also show that Ad5-PK increases DC infectivity and tumor specific antigen expression. Finally, vaccination of mice with the Ad5-PK vector resulted in enhanced T-cell-mediated interferon gamma (IFN-γ) release in response to both mesothelin peptide and a tumor line expressing mesothelin. Ad5-PK is a promising tool for cancer immunotherapy as it improves infectivity, gene transfer, protein expression, and subsequent T-cell activation in DCs compared to wild-type HAdV5 viruses

Intrauterine death following intraamniotic triiodothyronine and thyroxine therapy for fetal goitrous hypothyroidism associated with polyhydramnios and caused by a thyroglobulin mutation

In the absence of maternal thyroid disease or iodine deficiency, fetal goitre is rare and usually attributable to dyshormonogenesis, for which genetic ascertainment is not always undertaken in the UK. Mechanical complications include tracheal and oesophageal compression with resultant polyhydramnios, malpresentation at delivery and neonatal respiratory distress. We report an Indian kindred in which the proband (first-born son) had congenital hypothyroidism (CH) without obvious neonatal goitre. His mother's second pregnancy was complicated by fetal hypothyroid goitre and polyhydramnios, prompting amniotic fluid drainage and intraamniotic therapy (with liothyronine, T3 and levothyroxine, T4). Sadly, intrauterine death occurred at 31 weeks. Genetic studies in the proband demonstrated compound heterozygous novel (c.5178delT, p.A1727Hfs*26) and previously described (c.7123G > A, p.G2375R) thyroglobulin (TG) mutations which are the likely cause of fetal goitre in the deceased sibling. TG mutations rarely cause fetal goitre, and management remains controversial due to the potential complications of intrauterine therapy however an amelioration in goitre size may be achieved with intraamniotic T4, and intraamniotic T3/T4 combination has achieved a favourable outcome in one case. A conservative approach, with surveillance, elective delivery and commencement of levothyroxine neonatally may also be justified, although intubation may be required post delivery for respiratory obstruction. Our observations highlight the lethality which may be associated with fetal goitre. Additionally, although this complication may recur in successive pregnancies, our case highlights the possibility of discordance for fetal goitre in siblings harbouring the same dyshormonogenesis-associated genetic mutations. Genetic ascertainment may facilitate prenatal diagnosis and assist management in familial cases. LEARNING POINTS: CH due to biallelic, loss-of-function TG mutations is well-described and readily treatable in childhood however mechanical complications from associated fetal goitre may include polyhydramnios, neonatal respiratory compromise and neck hyperextension with dystocia complicating delivery.CH due to TG mutations may manifest with variable phenotypes, even within the same kindred.Treatment options for hypothyroid dyshormogenic fetal goitre in a euthyroid mother include intraamniotic thyroid hormone replacement in cases with polyhydramnios or significant tracheal obstruction. Alternatively, cases may be managed conservatively with radiological surveillance, elective delivery and neonatal levothyroxine treatment, although intubation and ventilation may be required to support neonatal respiratory compromise.Genetic ascertainment in such kindreds may enable prenatal diagnosis and anticipatory planning for antenatal management of further affected offspring.This work was supported by funding from the Wellcome Trust (grant number 100585/Z/12/Z to N S) and the National Institute for Health Research Biomedical Research Centre Cambridge (N S). The Genomics/ Transcriptomics Core Facility is supported by the UK Medical Research Council (MRC) Metabolic Disease Unit (MRC_MC_UU_12012/5) and a Wellcome Trust Strategic Award (100574/Z/12/Z)

Phase 3 Multicenter Study of Revusiran in Patients with Hereditary Transthyretin-Mediated (hATTR) Amyloidosis with Cardiomyopathy (ENDEAVOUR)

PURPOSE: The Phase 3 ENDEAVOUR study evaluated revusiran, an investigational RNA interference therapeutic targeting hepatic transthyretin (TTR) production, for treating cardiomyopathy caused by hereditary transthyretin-mediated (hATTR) amyloidosis. / METHODS: Patients with hATTR amyloidosis with cardiomyopathy were randomized 2:1 to receive subcutaneous daily revusiran 500 mg (n = 140) or placebo (n = 66) for 5 days over a week followed by weekly doses. Co-primary endpoints were 6-min walk test distance and serum TTR reduction. / RESULTS: Revusiran treatment was stopped after a median of 6.71 months; the study Sponsor prematurely discontinued dosing due to an observed mortality imbalance between treatment arms. Eighteen (12.9%) patients on revusiran and 2 (3.0%) on placebo died during the on-treatment period. Most deaths in both treatment arms were adjudicated as cardiovascular due to heart failure (HF), consistent with the natural history of the disease. A post hoc safety investigation of patients treated with revusiran found that, at baseline, a greater proportion of those who died were ≥ 75 years and showed clinical evidence of more advanced HF compared with those who were alive throughout treatment. Revusiran pharmacokinetic exposures and TTR lowering did not show meaningful differences between patients who died and who were alive. Revusiran did not deleteriously affect echocardiographic parameters, cardiac biomarkers, or frequency of cardiovascular and HF hospitalization events. / CONCLUSIONS: Causes for the observed mortality imbalance associated with revusiran were thoroughly investigated and no clear causative mechanism could be identified. Although the results suggest similar progression of cardiac parameters in both treatment arms, a role for revusiran cannot be excluded. / CLINICAL TRIAL REGISTRATION: NCT02319005

Extracts of Feijoa Inhibit Toll-Like Receptor 2 Signaling and Activate Autophagy Implicating a Role in Dietary Control of IBD

Inflammatory bowel disease (IBD) is a heterogeneous chronic inflammatory disease affecting the gut with limited treatment success for its sufferers. This suggests the need for better understanding of the different subtypes of the disease as well as nutritional interventions to compliment current treatments. In this study we assess the ability of a hydrophilic feijoa fraction (F3) to modulate autophagy a process known to regulate inflammation, via TLR2 using IBD cell lines

Transformation of UML Activity Diagram for Enhanced Reasoning

IT industry has adopted the unified modelling language activity diagram (UML-AD) as a de facto standard. UML AD facilitates modellers to graphically represent and document business processes to show the flow of activities and behaviour of a system. However, UML AD has many drawbacks such as lack of formal semantics i.e. ontology used for the constructs based on intuition, that vaguely describes processes and no provision for verifiability. Petri Net (PN) has been around for decades and used to model the workflow systems but PNs and its variants are too complex for business process modellers with no prior experience. A logical foundation is desirable to construct a business process with a precision that facilitates in transforming UML AD into a formal mechanism supported by verifiability capabilities for enhanced reasoning. Therefore, in this paper, we will provide a framework that will provide formal definitions for UML AD core terms and constructs used for modelling, and subsequently transform them to formal representation called point graph(PG). This will provide an insight into UML AD and will improve the overall functionality required from a modelling tool. A case study is conducted at King’s College Hospital trust’ to improve their patient flows of an accident and emergency (A&E) department