Synthesis and screening of a library of Lewisx deoxyfluoro-analogues reveals differential recognition by glycan-binding partners.

Glycan-mediated interactions play a crucial role in biology and medicine, influencing signalling, immune responses, and disease pathogenesis. However, the use of glycans in biosensing and diagnostics is limited by cross-reactivity, as certain glycan motifs can be recognised by multiple biologically distinct protein receptors. To address this specificity challenge, we report the enzymatic synthesis of a 150-member library of site-specifically fluorinated Lewisx analogues ('glycofluoroforms') using naturally occurring enzymes and fluorinated monosaccharides. Subsequent incorporation of a subset of these glycans into nanoparticles or a microarray revealed a striking spectrum of distinct binding intensities across different proteins that recognise Lewisx. Notably, we show that for two proteins with unique binding sites for Lewisx, glycofluoroforms exhibited enhanced binding to one protein, whilst reduced binding to the other, with selectivity governed by fluorination patterns. We finally showcase the potential diagnostic utility of this approach in glycofluoroform-mediated bacterial toxin detection by lateral flow.

Three-dimensional fluid-structure interaction simulation of the Wheatley aortic valve.

Valvular heart diseases (such as stenosis and regurgitation) are recognized as a rapidly growing cause of global deaths and major contributors to disability. The most effective treatment for these pathologies is the replacement of the natural valve with a prosthetic one. Our work considers an innovative design for prosthetic aortic valves that combines the reliability and durability of artificial valves with the flexibility of tissue valves. It consists of a rigid support and three polymer leaflets which can be cut from an extruded flat sheet, and is referred to hereafter as the Wheatley aortic valve (WAV). As a first step towards the understanding of the mechanical behavior of the WAV, we report here on the implementation of a numerical model built with the ICFD multi-physics solver of the LS-DYNA software. The model is calibrated and validated using data from a basic pulsatile-flow experiment in a water-filled straight tube. Sensitivity to model parameters (contact parameters, mesh size, etc.) and to design parameters (height, material constants) is studied. The numerical data allow us to describe the leaflet motion and the liquid flow in great detail, and to investigate the possible failure modes in cases of unfavorable operational conditions (in particular, if the leaflet height is inadequate). In future work the numerical model developed here will be used to assess the thrombogenic properties of the valve under physiological conditions.

Reverse thiophosphorylase activity of a glycoside phosphorylase in the synthesis of an unnatural Manß1,4GlcNAc library.

ß-Mannosides are ubiquitous in nature, with diverse roles in many biological processes. Notably, Manß1,4GlcNAc a constituent of the core N-glycan in eukaryotes was recently identified as an immune activator, highlighting its potential for use in immunotherapy. Despite their biological significance, the synthesis of ß-mannosidic linkages remains one of the major challenges in glycoscience. Here we present a chemoenzymatic strategy that affords a series of novel unnatural Manß1,4GlcNAc analogues using the ß-1,4-d-mannosyl-N-acetyl-d-glucosamine phosphorylase, BT1033. We show that the presence of fluorine in the GlcNAc acceptor facilitates the formation of longer ß-mannan-like glycans. We also pioneer a "reverse thiophosphorylase" enzymatic activity, favouring the synthesis of longer glycans by catalysing the formation of a phosphorolysis-stable thioglycoside linkage, an approach that may be generally applicable to other phosphorylases.

Obtaining Pure 1H NMR Spectra of Individual Pyranose and Furanose Anomers of Reducing Deoxyfluorinated Sugars.

Due to tautomeric equilibria, NMR spectra of reducing sugars can be complex with many overlapping resonances. This hampers coupling constant determination, which is required for conformational analysis and configurational assignment of substituents. Given that mixtures of interconverting species are physically inseparable, easy-to-use techniques that enable facile full 1H NMR characterization of sugars are of interest. Here, we show that individual spectra of both pyranoside and furanoside forms of reducing fluorosugars can be obtained using 1D FESTA. We discuss the unique opportunities offered by FESTA over standard sel-TOCSY and show how it allows a more complete characterization. We illustrate the power of FESTA by presenting the first full NMR characterization of many fluorosugars, including of the important fluorosugar 2-deoxy-2-fluoroglucose. We discuss in detail all practical considerations for setting up FESTA experiments for fluorosugars, which can be extended to any mixture of fluorine-containing species interconverting slowly on the NMR frequency-time scale.

Synthesis and Structural Characteristics of all Mono- and Difluorinated 4,6-Dideoxy-d-xylo-hexopyranoses.

Protein-carbohydrate interactions are implicated in many biochemical/biological processes that are fundamental to life and to human health. Fluorinated carbohydrate analogues play an important role in the study of these interactions and find application as probes in chemical biology and as drugs/diagnostics in medicine. The availability and/or efficient synthesis of a wide variety of fluorinated carbohydrates is thus of great interest. Here, we report a detailed study on the synthesis of monosaccharides in which the hydroxy groups at their 4- and 6-positions are replaced by all possible mono- and difluorinated motifs. Minimization of protecting group use was a key aim. It was found that introducing electronegative substituents, either as protecting groups or as deoxygenation intermediates, was generally beneficial for increasing deoxyfluorination yields. A detailed structural study of this set of analogues demonstrated that dideoxygenation/fluorination at the 4,6-positions caused very little distortion both in the solid state and in aqueous solution. Unexpected trends in α/ß anomeric ratios were identified. Increasing fluorine content always increased the α/ß ratio, with very little difference between regio- or stereoisomers, except when 4,6-difluorinated.

Introducing affinity and selectivity into galectin-targeting nanoparticles with fluorinated glycan ligands.

Galectins are potential biomarkers and therapeutic targets. However, galectins display broad affinity towards ß-galactosides meaning glycan-based (nano)biosensors lack the required selectivity and affinity. Using a polymer-stabilized nanoparticle biosensing platform, we herein demonstrate that the specificity of immobilised lacto-N-biose towards galectins can be 'turned on/off' by using site-specific glycan fluorination and in some cases reversal of specificity can be achieved. The panel of fluoro-glycans were obtained by a chemoenzymatic approach, exploiting BiGalK and BiGalHexNAcP enzymes from Bifidobacterium infantis which are shown to tolerate fluorinated glycans, introducing structural diversity which would be very laborious by chemical methods alone. These results demonstrate that integrating non-natural, fluorinated glycans into nanomaterials can encode unprecedented selectivity with potential applications in biosensing.

Assembling the Dead, Gathering the Living: Radiocarbon Dating and Bayesian Modelling for Copper Age Valencina de la Concepción (Seville, Spain).

The great site of Valencina de la Concepción, near Seville in the lower Guadalquivir valley of southwest Spain, is presented in the context of debate about the nature of Copper Age society in southern Iberia as a whole. Many aspects of the layout, use, character and development of Valencina remain unclear, just as there are major unresolved questions about the kind of society represented there and in southern Iberia, from the late fourth to the late third millennium cal BC. This paper discusses 178 radiocarbon dates, from 17 excavated sectors within the c. 450 ha site, making it the best dated in later Iberian prehistory as a whole. Dates are modelled in a Bayesian statistical framework. The resulting formal date estimates provide the basis for both a new epistemological approach to the site and a much more detailed narrative of its development than previously available. Beginning in the 32nd century cal BC, a long-lasting tradition of simple, mainly collective and often successive burial was established at the site. Mud-vaulted tholoi appear to belong to the 29th or 28th centuries cal BC; large stone-vaulted tholoi such as La Pastora appear to date later in the sequence. There is plenty of evidence for a wide range of other activity, but no clear sign of permanent, large-scale residence or public buildings or spaces. Results in general support a model of increasingly competitive but ultimately unstable social relations, through various phases of emergence, social competition, display and hierarchisation, and eventual decline, over a period of c. 900 years.

Investigating the Suitability of Carbon Nanotube Reinforced Polymer in Transcatheter Valve Applications.

The current delivery size of transcatheter aortic valves, limited by the thickness of their pericardial leaflets, correlates with a high prevalence of major vascular complications. Polyurethane valves can be developed to a fraction of the thickness of pericardial valves through the addition of carbon nanotubes to reinforce their leaflets. This study investigates the suitability of a novel carbon nanotube reinforced leaflet to reduce the delivery profile of transcatheter aortic valves. Carbon nanotube polyurethane composites were developed with thicknesses of 50 µm and their mechanical properties were determined in relation to various environmental effects. The composites demonstrated improvements to the material stiffness, particularly at increasing strain rates compared to the neat polymer. However, increasing nanotube concentrations significantly decreased the fatigue life of the composites. Key findings highlighted a potential for carbon nanotube reinforcement in valve replacement which experience very high strain rates during the cardiac cycle. Further testing is needed to achieve a strong nanotube-matrix interface which will prolong the cyclic fatigue life and further strengthen tensile properties. Testing on the durability and haemocompatibility of these composite heart valves are ongoing.

Universal label-free in-process quantification of influenza virus-like particles.

Virus-like particles (VLPs) are becoming established as vaccines, in particular for influenza pandemics, increasing the interest in the development of VLPs manufacturing bioprocess. However, for complex VLPs, the analytical tools used for quantification are not yet able to keep up with the bioprocess progress. Currently, quantification for Influenza relies on traditional methods: hemagglutination assay or Single Radial Immunodiffusion. These analytical technologies are time-consuming, cumbersome, and not supportive of efficient downstream process development and monitoring. Hereby we report a label-free tool that uses Biolayer interferometry (BLI) technology applied on an Octet platform to quantify Influenza VLPs at all stages of bioprocess. Human (α2,6-linked sialic acid) and avian (α2,3-linked sialic acid) biotinylated receptors associated with streptavidin biosensors were used, to quantify hemagglutinin content in several mono- and multivalent Influenza VLPs. The applied method was able to quantify hemagglutinin from crude samples up to final bioprocessing VLP product. BLI technology confirmed its value as a high throughput analytical tool with high sensitivity and improved detection limits compared to traditional methods. This simple and fast method allowed for real-time results, which are crucial for in-line monitoring of downstream processing, improving process development, control and optimization.

Sir Donald Ross, pioneer aortic valve surgeon.

Tribute to Sir Donald Ross by David Wheatley, as read by Robert Kleinloog, President, Society of Cardiothoracic Surgeons of South Africa at the Annual Congress of the South African Heart Association 19 October 2014.

Characterisation of H2S···CuCl and H2S···AgCl isolated in the gas phase: a rigidly pyramidal geometry at sulphur revealed by rotational spectroscopy and ab initio calculations.

Pure rotational spectra of the ground vibrational states of eight isotopologues of H(2)S···CuCl and twelve isotopologues of H(2)S···AgCl have been analysed allowing rotational constants and hyperfine coupling constants to be determined. The molecular structures have been determined from the measured rotational constants and are presented alongside the results of calculations at the CCSD(T) level. Both molecules have C(s) symmetry at equilibrium and are pyramidal at the sulphur atom. The chlorine, metal, and sulphur atoms are collinear while the local C(2) axis of the hydrogen sulphide molecule intersects the axis defined by the heavy atoms at an angle, φ = 74.46(2)° for Cu and φ = 78.052(6)° for Ag. The molecular geometries are rationalised using simple rules that invoke the electrostatic interactions within the complexes. Centrifugal distortion constants, Δ(J), and nuclear quadrupole coupling constants, χ(aa)(Cu) and χ(aa)(Cl) for H(2)S···CuCl are presented for the first time. The geometry of H(2)S···AgCl is determined with fewer assumptions and greater precision than previously.

Venous thromboembolism prophylaxis guideline implementation is improved by nurse directed feedback and audit.

BACKGROUND: Venous thromboembolism (VTE) is a major health and financial burden. VTE impacts health outcomes in surgical and non-surgical patients. VTE prophylaxis is underutilized, particularly amongst high risk medical patients. We conducted a multicentre clinical audit to determine the extent to which appropriate VTE prophylaxis in acutely ill hospitalized medical patients could be improved via implementation of a multifaceted nurse facilitated educational program. METHODS: This multicentre clinical audit of 15 Australian hospitals was conducted in 2007-208. The program incorporated a baseline audit to determine the proportion of patients receiving appropriate VTE prophylaxis according to best practice recommendations issued by the Australian and New Zealand Working Party on the Management and Prevention of Venous Thromboembolism (ANZ-WP recommendations), followed by a 4-month education intervention program and a post intervention audit. The primary endpoint was to compare the proportion of patients being appropriately managed based on their risk profile between the two audits. RESULTS: A total of 8774 patients (audit 1; 4399 and audit 2; 4375) were included in the study, most (82.2% audit 1; and 81.0% audit 2) were high risk based on ANZ-WP recommendations. At baseline 37.9% of high risk patients were receiving appropriate thromboprophylaxis. This increased to 54.1% in the post intervention audit (absolute improvement 16%; 95% confidence interval [CI] 11.7%, 20.5%). As a result of the nurse educator program, the likelihood of high risk patients being treated according to ANZ-WP recommendations increased significantly (OR 1.96; 1.62, 2.37). CONCLUSION: Utilization of VTE prophylaxis amongst hospitalized medical patients can be significantly improved by implementation of a multifaceted educational program coordinated by a dedicated nurse practitioner.

Biomechanical analysis of a synthetic, biodegradable impaction graft substitute.

Impaction bone grafting in revision arthroplasty is a common and successful procedure to restore primary bone stock. Reducing the amount of bone needed to fill large grafts has been a driving force for the use of synthetic materials that can act as extenders or substitutes. To this end, we evaluated the mechanical properties of a new class of biodegradable polymer beads with and without donor bone to determine its suitability for use in impaction grafting. Biodegradable methacrylated anhydride beads were synthesized using thermal polymerization techniques. The mechanical properties of the beads were then tested in an impaction grafting test chamber and compared with morsellised porcine allograft. The beads, porcine allograft and a 50/50 combination all had similar mechanical properties, both in compression and relaxation. Pure polymer beads compacted significantly less than pure allograft and retained macroporosity after impaction. Our results suggest that the biodegradable beads have sufficient mechanical properties to be considered as an impaction grafting substitute or extender. Their ability to fill space whilst retaining macroporosity may be advantageous for tissue ingrowth and remodeling.

18F-FDG dual-headed gamma camera PET in detection of axillary nodal disease in patients with large or locally advanced breast cancer: possible alternative staging of axilla.

Positron emission tomography (PET) has been used in staging the axilla. Gamma Camera PET (GCPET) is a cost effective alternative, but poorly studied. The aim of this study was to assess GCPET in demonstrating metastatic deposits in axillary nodes in patients with a high likelihood of nodal disease. Twenty-seven women with large (T2, T3 or T4) or advanced breast cancer (N1, N2 or M1) were recruited. All patients underwent axillary lymph node removal or biopsy (fine needle aspiration cytology (FNAC) or core cut) and whole body GCPET imaging. Images were reported anonymously and compared with the histological findings. Twenty-one patients proceeded to surgery and 10 had tumour-involved axillary nodes; GCPET was positive in 8 of these. The remaining 6 patients underwent core cut or FNAC of the axillary nodes, 2 of which contained a tumour. GCPET was positive in both cases. Thus, the diagnostic values were: sensitivity 83%, specificity 100%, positive predictive value 100%, negative predictive value 88% and accuracy 93%. In conclusion, GCPET is a reliable method and can be performed in a district general hospital and detecting disease in axillary nodes in certain patients, possibly obviating the need for surgery.

Our inability to predict thromboembolic events after prosthetic valve surgery.

BACKGROUND AND AIM OF THE STUDY: Thromboembolic and bleeding complications detract from outcome for patients with prosthetic heart valves. The study aim was to investigate whether measurement of coagulation activation markers and transcranial Doppler ultrasound microembolic signals (MES) could identify patients at subsequent higher risk of thromboembolism or bleeding events. METHODS: A total of 526 patients (mean age 66 years; 266 males, 260 females) who underwent elective valve replacement surgery was enrolled between April 1999 and October 2002. Clinical assessment and blood sampling for coagulation activation markers was performed preoperatively and at three and 12 months postoperatively. Transcranial Doppler MES were recorded in the first 144 patients. Status was reviewed between 21st April and 9th June 2005, with 99.4% follow up. RESULTS: Among patients, 62% had an aortic valve replaced, and mechanical valves constituted 60% of all implants. The mean follow up was 3.61 years; total follow up was 1,899.2 patient-years (pt-yr). In total, 115 patients died, while 61 experienced a total of 80 thromboembolic events: linearized event rates were 3.94% (mechanical valves) and 4.4% (bioprostheses). There was no difference between mitral and aortic implants, or among bileaflet, tilting-disc mechanical and porcine valves. Atrial fibrillation was not influential. Coagulation activation markers were not associated with thromboembolic events, except for an elevated von Willebrand factor (vWF), which was associated with a five-fold increase in embolic event rate. Fifty-one patients experienced 59 bleeding events; eight patients experienced multiple events. Linearized event rates were 3.37% (mechanical valves) and 2.49% (bioprostheses). The INR was suboptimal in 44-58% of patients. Transcranial Doppler MES were not associated with blood coagulation markers or thromboembolic events. CONCLUSION: Coagulation activation markers (except vWF) and MES did not predict thromboembolic events in valve replacement patients. Thromboembolic and bleeding event rates for West of Scotland patients generally exceeded reported rates: suboptimal anticoagulation appeared common and most likely influenced thromboembolic and bleeding event rates more than any other factor.

Autologous blood transfusion for cardiopulmonary bypass: effects of storage conditions on platelet function.

OBJECTIVES: Cardiopulmonary bypass impairs formation of large stable platelet aggregates (macroaggregation), although formation of small aggregates (microaggregation) is preserved. A factor in the uncertain benefits of intraoperative autologous blood transfusion may be the effects of storage on platelet function. The effects of citrate preservative and heparinization before storage on platelet function was therefore assessed. METHODS: Twenty-seven patients undergoing elective coronary artery bypass grafting were randomly allocated to have 450 to 1,000 mL of blood taken into CPDA anticoagulant bags either before (n = 14) or after heparinization (n = 13). Samples from the patients and stored blood were anticoagulated with rhirudin, 200 U/mL. The macroaggregatory response to submaximal collagen was measured by impedance aggregometry and microaggregation by single platelet counting. RESULTS: During macroaggregation, before cardiopulmonary bypass, the ex vivo median (interquartile range) response was 16.3 (12.4-18.7) Omega. This decreased 10 minutes after heparin to 8.9 (3.3-11.0) Omega (p < 0.0001). In the blood bags (in vitro), the initial response for nonheparinized blood was 4.8 (0.1-7.5) Omega (p < 0.002 v ex vivo) and at end-cardiopulmonary bypass was 2.4 (1.6-8.2) Omega. During microaggregation, in vivo heparinization decreased microaggregation both ex vivo and in vitro in CPDA blood; the in vitro response of nonheparinized blood at end-cardiopulmonary bypass was greater than that seen after in vivo heparinization (p < 0.007). No difference in bleeding or transfusion requirements was seen. CONCLUSIONS: Collecting blood into CPDA anticoagulant caused a marked deterioration in platelet function. This was worse after in vivo heparinization and included depression of microaggregation.

A pilot randomised, open, uncontrolled, clinical study of two dosages of St John's wort (Hypericum perforatum) herb extract (LI-160) as an aid to motivational/behavioural support in smoking cessation.

There is an association between smoking and depression, yet the herbal antidepressant St John's wort (Hypericum perforatum L.; SJW) herb extract has not previously been investigated as an aid in smoking cessation. In this open, uncontrolled, pilot study, 28 smokers of 10 or more cigarettes per day for at least one year were randomised to receive SJW herb extract (LI-160) 300 mg once or twice daily taken for one week before and continued for 3 months after a target quit date. In addition, all participants received motivational/behavioural support from a trained pharmacist. At 3 months, the point prevalence and continuous abstinence rates were both 18%, and at 12 months were 0%. Fifteen participants (54%) reported 23 adverse events up to the end of the 3-month follow-up period. There was no statistically significant difference in the frequency of adverse events for participants taking SJW once or twice daily (p > 0.05). Most adverse events were mild, transient and non-serious. This preliminary study has not provided convincing evidence that a SJW herb extract plus individual motivational/behavioural support is likely to be effective as an aid in smoking cessation. However, it may be premature to rule out a possible effect on the basis of a single, uncontrolled pilot study, and other approaches involving SJW extract may warrant investigation.

Medicinal plants for insomnia: a review of their pharmacology, efficacy and tolerability.

A number of medicinal plants are traditionally endowed with anxiolytic or sedative properties and, in the context of this revue, both indications are considered since the former may induce a mood conducive to the latter. For any sleep-inducing drug to be effective, a tranquil ambience needs to be established a priori. Thus, physical ailments (i.e. pain), factors interfering with sleep (i.e. noise), psychological conditions causing stress, psychiatric illnesses (i.e. depression) and other drugs that interfere with sleep (i.e. caffeine) need to be controlled, if possible. Kava-kava is a well-established hypnotic drug, with a rapid onset of effect, adequate duration of action and minimal morning after-effects. However, reports of serious hepatotoxicity with this preparation have led to it being banned in most countries worldwide. On the other hand, side-effects with valerian would appear to be bland indeed. However, it's slow onset of effect (2-3 weeks) renders it unsuitable for short-term use (i.e. 'jet-lag'), but it does have profound beneficial effects on sleep architecture (augments deep sleep) that may make it particularly suitable for long-term use and for the elderly. In a personal trial (not double-blind) in stress-induced insomnia, both kava and valerian improved sleep and the ill-effects of stress, and the combination of the two was even more effective for the control of insomnia. Aromatherapy (lavender, chamomile, Ylang-Ylang) would appear to improve sleep, but how practical a form of treatment this may be remains to be determined. The only other plant drug that may have some effect on sleep is melissa, but reports are too scanty to form any opinion about this. Based on animal experiments, passion flower (passiflora) may have a sedative action, but the sedative action of hops has not been investigated in any detail. In conclusion, there is a need for longer-term controlled studies with some of these compounds (particularly valerian). Aromatherapy constitutes a tantalising possibility. In the interpretation of this review, it should be borne in mind that the evidence on which it is based is often incomplete or missing, but that is all that is available. Consequently some conjecture on the part of the author is inevitable and should be appreciated as such.

Mitral valve replacement with mechanical or bioprosthetic valve.

A presentation on the technique of mitral valve replacement is shown: exposure and excision of the diseased mitral valve is demonstrated along with the use of sub-annular and supra-annular interrupted suture techniques for securing biological and mechanical mitral prostheses. A brief overview of the literature is presented.