The effect on consent rates for deceased organ donation in Wales after the introduction of an opt-out system.

Organ transplantation saves and transforms lives. Failure to secure consent for organ retrieval is widely regarded as the single most important obstacle to transplantation. A soft opt-out system of consent for deceased organ donation was introduced into Wales in December 2015, whilst England maintained the existing opt-in system. Cumulative data on consent rates in Wales were compared with those in England, using a two-sided sequential procedure that was powered to detect an absolute difference in consent rates between England and Wales of 10%. Supplementary risk-adjusted logistic regression analysis examined whether any difference in consent rates between the two nations could be attributed to variations in factors known to influence UK consent rates. Between 1 January 2016 and 31 December 2018, 8192 families of eligible donors in England and 474 in Wales were approached regarding organ donation, with overall consent rates of 65% and 68%, respectively. There was a steady upward trend in the proportion of families consenting to donation after brain death in Wales as compared with England and after 33 months, this reached statistical significance. No evidence of any change in the donation after circulatory death consent rate was observed. Risk-adjusted logistic regression analysis revealed that by the end of the study period the probability of consent to organ donation in Wales was higher than in England (OR [95%CI] 2.1 [1.26-3.41]). The introduction of a soft opt-out system of consent in Wales significantly increased organ donation consent though the impact was not immediate.

Structural and electronic determinants of lytic polysaccharide monooxygenase reactivity on polysaccharide substrates.

Lytic polysaccharide monooxygenases (LPMOs) are industrially important copper-dependent enzymes that oxidatively cleave polysaccharides. Here we present a functional and structural characterization of two closely related AA9-family LPMOs from Lentinus similis (LsAA9A) and Collariella virescens (CvAA9A). LsAA9A and CvAA9A cleave a range of polysaccharides, including cellulose, xyloglucan, mixed-linkage glucan and glucomannan. LsAA9A additionally cleaves isolated xylan substrates. The structures of CvAA9A and of LsAA9A bound to cellulosic and non-cellulosic oligosaccharides provide insight into the molecular determinants of their specificity. Spectroscopic measurements reveal differences in copper co-ordination upon the binding of xylan and glucans. LsAA9A activity is less sensitive to the reducing agent potential when cleaving xylan, suggesting that distinct catalytic mechanisms exist for xylan and glucan cleavage. Overall, these data show that AA9 LPMOs can display different apparent substrate specificities dependent upon both productive protein-carbohydrate interactions across a binding surface and also electronic considerations at the copper active site.

Effect of irradiation, asbestos and chemical cocarcinogens on incidence of sarcoma on implants.

An implant can act as a physical carcinogen. Chemicals applied to its surface can augment that. To explore if cocarcinogenesis would affect the incidence of tumour on implants, we tested several physical (asbestos, beta irradiation) and chemical (acridine orange, phorbol methyl ester, cigarette smoke tar, extract of Agaricus bisporus mushroom) carcinogens on 25 mm diameter nitrocellulose filters implanted in groups of BALB/c mice in 9 experiments. Saline was applied in 287 concurrent controls. Irradiation by 540 kBq I-125 fibrinogen on foreign bodies of surface area 20sq cm induced tumour in 6/6 pilot animals (expected 2/6) and in 25/36 animals (controls 13/29; p < 0.04). The mean dose (beta and gamma) to cells within 20 microm of the filter surface was estimated to be 782 mGy. Exposures to 270kBq or less were not significantly cocarcinogenic with single implants. Radiation from the paired implants was detectible up to 80 weeks. Asbestos, however, 2.3 or 4.6 mgm on implants did not in 3 trials increase sarcoma yield significantly, but did induce mesothelial-type growths in the peritoneum. Asbestos (2.3 mg) injected without an implant provoked no tumour at the site. The chemical carcinogen acridine orange gave ambiguous results, and the others reduced tumour incidence insignificantly. The irradiation history may be germane in the human cases that arise.

Directional selectivity and frequency tuning of midbrain cells in the oyster toadfish, Opsanus tau.

Single-unit recordings were made from areas in the midbrain (torus semicircularis) of the oyster toadfish. We evaluated frequency tuning and directional responses using whole-body oscillation to simulate auditory stimulation by particle motion along axes in the horizontal and mid-sagittal planes. We also tested for bimodality in responses to auditory and hydrodynamic stimuli. One recording location in each animal was marked by a neurobiotin injection to confirm the recording site. Recordings were made in nucleus centralis, nucleus ventrolateralis, and the deep cell layer. Most units were frequency-selective with best frequencies between 50 and 141 Hz. Suppression of activity was apparent in 10% of the cells. Bimodality was common, including inhibition and suppression of background activity by auditory or hydrodynamic stimulation. The majority of the cells were directionally selective with directional response patterns that were sharpened compared with those of primary saccular afferents. The best directional axes were arrayed widely in spherical space, covering most azimuths and elevations. This representation is adequate for the computation of the motional axis of an auditory stimulus for sound source localization.

DHEA treatment of Alzheimer's disease: a randomized, double-blind, placebo-controlled study.

OBJECTIVE: To compare the efficacy and tolerability of dehydroepiandrosterone (DHEA) vs placebo in AD. METHOD: Fifty-eight subjects with AD were randomized to 6 month's treatment with DHEA (50 mg per os twice a day; n = 28) or placebo (n = 30) in a multi-site, double-blind pilot trial. Primary efficacy measures assessed cognitive functioning (the AD Assessment Scale-Cognitive [ADAS-Cog]) and observer-based ratings of overall changes in severity (the Clinician's Interview-Based Impression of Change with Caregiver Input [CIBIC-Plus]). At baseline, 3 months, and 6 months, the ADAS-Cog was administered, and at 3 and 6 months, the CIBIC-Plus was administered. The 6-month time point was the primary endpoint. RESULTS: Nineteen DHEA-treated subjects and 14 placebo-treated subjects completed the trial. DHEA was relatively well-tolerated. DHEA treatment, relative to placebo, was not associated with improvement in ADAS-Cog scores at month 6 (last observation carried forward; p = 0.10); transient improvement was noted at month 3 (p = 0.014; cutoff for Bonferroni significance = 0.0125). No difference between treatments was seen on the CIBIC-Plus at either the 6-month or the 3-month time points. CONCLUSIONS: DHEA did not significantly improve cognitive performance or overall ratings of change in severity in this small-scale pilot study. A transient effect on cognitive performance may have been seen at month 3, but narrowly missed significance.

The effect of head position, electrode site, movement and smoothing window in the determination of a reliable maximum voluntary activation of the upper trapezius muscle.

Quantitative measures derived from raw myoelectric signal (MES) data must be normalized to allow for comparisons both within and between subjects. The most common method of normalization involves dividing the root mean square (RMS) amplitude of the MES recorded during a given activity by the RMS of the MES elicited during a maximal voluntary isometric contraction (MVIC) of that particular muscle. The objective of this study was to use surface-recorded MES amplitude to determine the combination of electrode site, test position, head posture, and smoothing window that elicits the highest and most reliable MES amplitudes during an MVIC of the upper trapezius (UT) muscle. Ten volunteers had surface electrodes positioned at five sites on the UT of their dominant side. Three trials of each of three MVIC test positions were performed both with the head in neutral and rotated 45 degrees to the contralateral side. A repeated-measures ANOVA was used for statistical hypothesis testing. Coefficients of variation were used to quantify the between-factor variability introduced in each case. In addition, the data were re-analyzed using moving windows of 100 to 500 ms in length, and an ANOVA was used to determine the effect of window length on both the amplitude and variability of the estimates of maximum voluntary activation (MVE). Head position had no significant effect on RMS amplitude of the MVIC in any of the test positions. There was a significant electrode site by test position interaction effect. Bonferroni post-hoc analyses were performed on this interaction by fixing test position and testing electrode site, revealing that Sites 1 (2 cm lateral to the midpoint between C7 spinous process and the posterolateral border of the acromion) and 4 (2 cm posterior to Site 1) recorded significantly higher RMS values for all test positions, and were not significantly different from each other. Fixing electrode site, the test position analysis revealed that abduction of the humerus, and abduction with external rotation of the humerus produced significantly higher RMS values than shoulder elevation at both Sites 1 and 4, and that abduction produced a significantly higher RMS amplitude than abduction in external rotation at Site 1. The results confirmed that Sites 1 and 4 consistently produced the highest MES amplitudes for all movements. Pure abduction consistently elicited maximal RMS values; however there is concern regarding supraspinatus cross talk during this movement. Site 1 was found to produce the most reliable data. A moving window of 100 ms was found to generate MVE estimates that were significantly higher than windows ranging from 200 ms to 500 ms in length. There was no effect of window length on the reliability of the MVEs. Based on this study, it was concluded that abduction or abduction with the arms in lateral rotation should be used as normalization contraction positions for the upper trapezius muscle. During this movement, Site 1 data smoothed with a moving window of 100 ms produces the highest amplitude MVE data but window lengths greater than 200 ms produce more stable estimates in terms of being able to compare studies in which moving windows are used to compute RMS.

The labelling of human serum transferrin with 99mTc and a study concerning uptake of the complex by tumour cells.

The direct labelling of serum transferrin (sTf) with 99mTc on high-affinity binding sites, producing a complex of excellent stability, is described. The high-affinity binding sites were prepared by pre-treating sTf with 2-mercaptoethanol. For the radiolabelling step, thiourea was used as an exchange ligand and a filtration procedure used to remove 99mTc that had not complexed with the protein. RT112 bladder tumour cells incubated in the presence of labelled sTf showed a rapid initial uptake of 99mTc, reaching a plateau after about 20 min. Radiolabelling was also carried out without a pre-reduction step in an attempt to form a co-ordination complex between 99mTc and the Fe3+-binding site of sTf, analogous to that formed by Fe3+. The tumour cell uptake of sTf labelled without pre-reduction was then examined. In contrast to 59Fe3+ and other radio-metals co-ordinated with the Fe3+-binding site which show a continuous increase in incorporation with time, the uptake of 99mTc rapidly reached a plateau.

Short-term infusion of LongR(3) insulin-like growth factor (IGF)-I decreases hepatic IGF-I mRNA but not IGF binding protein-3 mRNA expression in pigs.

Infusion of pigs with an insulin-like growth factor-I (IGF-I) analogue (LongR(3)IGF-I) that does not bind to IGF-binding proteins decreases growth rate and the plasma concentration of growth hormone (GH), IGF-I, IGFBP-3, and insulin. This study was designed to determine whether the decrease is due to changes in IGF-I and IGFBP-3 gene expression. IGF-I or LongR(3)IGF-I (180 microg/kg/day) was infused into 55-kg finisher pigs for 4 days using Travenol infuser pumps. Plasma IGF-I concentration was measured by radioimmunoassay and plasma IGFBP-3 and IGFBP-2 were estimated by Western ligand blotting. Steady-state levels of IGF-I and IGFBP-3 mRNA were measured by RNase protection assay. Neither IGF-I nor LongR(3)IGF-I had a significant effect on hepatic IGF-I class 1 mRNA expression, whereas hepatic IGF-I class 2 mRNA expression was significantly reduced by both peptides. Plasma IGFBP-3 levels were unaffected by IGF-I treatment but were reduced by LongR(3)IGF-I treatment. The decrease in IGFBP-3 was not due to decreased gene expression in porcine liver or kidney, since neither IGF-I nor LongR(3)IGF-I treatment altered IGFBP-3 mRNA. This study infers a direct effect of the IGF analogue LongR(3)IGF-I on GH through its inhibition of plasma IGF-I concentration and class 2 IGF-I mRNA. The decrease in plasma IGFBP-3 was not accompanied by a decrease in hepatic or renal IGFBP-3 mRNA, suggesting that in this case, plasma IGFBP-3 protein levels are posttranslationally regulated or are derived from tissues other than liver or kidney.

In situ measurements of the pH of mammalian peroxisomes using the fluorescent protein pHluorin.

Peroxisomes are metabolically active organelles that participate in the oxidation of long-chain fatty acids and in the biosynthesis of bile acids, cholesterol, and ether phospholipids. Even though maintenance of a stable acid-base milieu is essential for proper peroxisomal function, the determination of the peroxisomal pH (pH(p)) remains inconclusive, and little is known about its regulation. To measure the pH of intact peroxisomes in situ, we used the peroxisome-specific carboxyl-terminal targeting sequence, SKL, to deliver a pH-sensitive mutant of the green fluorescent protein (pHluorin-SKL) selectively into peroxisomes. Proper targeting was verified by colocalization with the peroxisomal marker catalase. Peroxisomes were visualized by imaging fluorescence microscopy, and ratiometric measurements were combined with calibration using ionophores or a null-point method to estimate pH(p). The pH(p) was between 6.9 and 7.1, resembling the cytosolic pH. Manipulation of the cytosolic pH in intact cells or after permeabilization of the plasmalemma with streptolysin O revealed that pH(p) changed in parallel, suggesting that the peroxisomal membrane is highly permeable to H(+) (equivalents). We conclude that peroxisomes do not regulate their pH independently, but instead their large H(+) permeability effectively connects them with the buffer reservoir of the cytoplasm and with the homeostatic mechanisms that control cytosolic pH.

An evaluation of the clinical performance of newly qualified nurses: a competency based assessment.

The clinical performance of newly qualified staff nurses, or their 'fitness for purpose' has become a central professional and corporate issue and highlighted as a central causes for the reforms recommended by the Peach report (Peach 1999). However the ability to gauge the performance of newly qualified nurses remains a largely subjective exercise relying upon anecdotal evidence or general statements of newly qualified nurses' feelings of inadequacy on qualification. This article seeks to address this issue by reporting the findings of project that sought to compare the expectations of senior nurses regarding the level of competence of newly qualified nurses with that of the actual level of competency as assessed by the preceptors after 8 weeks in post. Using a specifically designed instrument to assess clinical competency a comparison has been made between 139 senior nurses' expectations of the competency of newly qualified nurses and the actual competence as assessed by preceptors of 36 newly qualified nurses after 8 weeks in post. The findings indicate that the senior nurses have clear subjective expectations of the competence level of newly qualified nurses. However these expectations are consistently lower than the actual level of competency demonstrated by the newly qualified nurses as assessed by their preceptors. This research demonstrates that through the use of a validated tool newly qualified nurses consistently perform at a higher level of competency than that expected by senior nurses. The ability to estimate competency levels by objective means should be developed. Further research is needed involving a larger sample of Trusts and Schools of Nursing to replicate the results of this study and to compare methods of the assessment of performance in terms of competency in practice on qualification. It is through developments based upon research such as this that a systematic evaluation of the contribution of Schools of Nursing to the competence of newly qualified nurses can be addressed.

Syntheses and structures of M(L)(X)BPh4 complexes (M = Co(II), Zn(II); L = cis-1,3,5-tris[3-(2-furyl)prop-2-enylideneamino]cyclohexane, X = OAc, NO3) n: structural models of the active site of carbonic anhydrase.

The metal coordination geometries in the structures of the zinc(II) and cobalt(II) complexes of the ligand cis-1,3,5-tris[3-(2-furyl)prop-2-enylideneamino]cyclohexane (fr-protach) and with the anions nitrate and acetate are structural models for the active site of carbonic anhydrase. The acetate structures show a striking structural correlation with the metal coordination environments in the known bicarbonate forms of the enzyme. Such structures provide a basis for understanding the marked effect of different metal substitution on the catalytic rate of the enzyme.

Directional encoding by fish auditory systems.

This paper reviews and discusses several investigations of the peripheral neural code for the directional axis of acoustical particle motion in the saccule of two fishes: goldfish (Carassius auratus) and toadfish (Opsanus tau). Most saccular afferents are directional in the manner of hair cells, having a cosine-shaped directional response pattern. The saccular sensory epithelia are orientated almost vertically in a parasagittal plane. In the horizontal plane, these epithelia are orientated obliquely with respect to the midline. Hair-cell stereocilia project perpendicularly. Thus, directional response patterns of saccular afferents tend to be orientated in azimuth parallel to the orientation of the epithelia in the head. The oblique angle of the toadfish saccule is greater than that of the goldfish, and the range of best directions in the horizontal plane for each species reflects those differing orientations. The azimuth of acoustical particle motion could be computed by comparing the relative activation of the two saccules, as is the case for the ears of most terrestrial vertebrates. The spatial patterns of saccular hair-cell orientation of most fishes thus appear to have little function in azimuthal source location, but for toadfish are probably most important for determining the elevation of monopole sources.

Growth hormone stimulation testing in both short and normal statured children: use of an immunofunctional assay.

Accurate interpretation of the results of GH stimulation tests is of pivotal importance not only in the evaluation of the etiology of growth retardation in children but also in the selection of the best candidates for GH therapy. We performed this study to test a novel immunofunctional GH ( IFGH) assay that makes use of the concept that one GH molecule dimerizes two GH receptors and compared the results with those obtained using two GH assays, the Diagnostic Systems Laboratories ELISA and a Hybritech immunoradiometric assay in 19 children with short stature undergoing routine GH stimulation testing. We also tested 13 normally statured control children to revisit the issue of what constitutes normal GH responses to stimuli, using all three assays and arginine and either L-dopa or insulin-induced hypoglycemia as secretagogues. Concentrations of IGF-I, IGF binding protein-3, and acid labile subunit were measured as well. There was a significant correlation between peak IFGH and Diagnostic Systems Laboratories ELISA GH responses to stimuli (r(2) = 0.93) as well as between the Diagnostic Systems Laboratories ELISA and Hybritech immunoradiometric assay (r(2) = 0.91). There were no significant differences between the short stature and normal group in peak or mean GH concentrations regardless of the assay used; however, the IGF-I, IGF binding protein-3, and acid labile subunit concentrations were substantially lower in the short stature group. There was a wide spectrum of GH concentrations in the normal group; approximately 50% of the children had peak GH concentrations <7 ng/mL, approximately 30% <5 ng/mL, and two pubertal normal subjects peaked to only 2 ng/mL with use of both the ELISA and IFGH assays. We conclude that 1) sensitive GH assays, ELISA and immunoradiometric assay, accurately detect a GH capable of generating a biologic signal comparable to an IFGH and 2) that normal GH stimulation test results can be substantially lower than previously accepted. GH-dependent growth factors may be more sensitive indicators of GH sufficiency than GH concentrations in response to pharmacologic stimuli.

Measurement of the acid-labile subunit of the insulin-like growth factor binding protein complex in human serum: a comparison of four immunoassays.

The acid-labile subunit (ALS) of the high molecular weight insulin-like growth factor binding protein complex is a liver-derived glycoprotein which is regulated by growth hormone and serves as a serum marker of growth hormone action. We have compared the measurement of ALS by four immunoassay methods (two RIAs, two ELISAs) utilizing various polyclonal and monoclonal antibodies raised against natural or recombinant human ALS, or synthetic ALS peptides. Despite the variety of methodologies and reagents, results obtained by the four methods were highly correlated for 125 sera from various patient groups, and when compared for individual groups of sera from healthy children and adults, growth hormone-deficient children and adults, and subjects with acromegaly. Some weaker correlations among methods were seen when measuring ALS levels in groups of sera from pregnant subjects and subjects with chronic renal failure. An assay using antibodies raised against recombinant ALS yielded lower apparent values than the other methods in patient sera, the discrepancy probably being attributable to a difference in standardization. We conclude that a variety of assay formats and reagents can yield serum ALS values of potential clinical utility.

Dendritic arbors on the saccule and lagena in the ear of the goldfish, Carassius auratus.

The ear of the goldfish (Carassius auratus) contains three otolithic endorgans: the saccule, lagena, and utricle. The saccule has an auditory function in most teleost fishes for whom data are available, and there is evidence that the lagena is also an auditory endorgan in the goldfish. This study was conducted to compare the innervation of the saccule and the lagena to one another and to previously published data from goldfish and other species. We placed cobaltous-lysine in saccular and lagenar nerves in vivo and permitted uptake over 18-24 h. A total of 59 saccular and 59 lagenar dendritic arbors were labeled in 10 fishes. Our data indicate that arbors on the saccule and lagena have similar morphologies, but differ in relative size. Saccular arbors tend to be smaller than lagenar arbors, with median arbor widths of 50 micrometer on the saccule and 74 micrometer on the lagena. Fiber diameters on the two endorgans are similar. A regional analysis of the saccule indicated that a wide range of arbor sizes are found along the rostral-caudal axis, with larger arbors more common caudally. Our data do not support the presence of two distinct categories of saccular afferents with non-overlapping distributions. Moderate arbor widths (50-99 micrometer) were most common in all regions of the lagena. Maximum arbor width and hair cell density do not appear to be correlated with one another on either the saccule or the lagena. Comparisons with published data from goldfish and oscar revealed similarities and differences that may be attributable to variations in label uptake or transport as well as potential species differences.

Transport of circulating IGF-I and LR3IGF-I from blood to extracellular wound fluid sites in rats.

Significant levels of IGF-I are found in wound fluid. The contribution that systemic IGF-I makes to the total IGF-I pool in wounds and the influence of IGF binding proteins (IGFBPs) on the delivery of systemic IGF-I to these wound sites has not been established. In the present series of experiments, we have shown that IGF-I flux across a model endothelial cell barrier is decreased in the presence of IGFBPs, whereas flux of Long R(3)IGF-I (LR(3)IGF-I, an IGF-I analogue with low affinity for IGFBPs) is unaffected. On the basis of these findings, the transport of IGF-I and LR(3)IGF-I from blood to extracellular wound fluid was assessed. Wound chambers were implanted subcutaneously in the backs of adult male rats and left in place for 14 days. A single i.v. bolus of either (125)I-IGF-I or (125)I-LR(3)IGF-I (10x10(6) c.p.m.) was administered via a jugular catheter and wound fluid and plasma samples taken at sequential time points between 5 and 240 min. (125)I-LR(3)IGF-I was removed from the circulation more rapidly than (125)I-IGF-I in both sham control and chamber implanted rats. Although implantation of the chambers did not alter the pharmacokinetic parameters of (125)I-IGF-I, significant increases in the steady state volume of distribution, clearance rate and half-life were recorded for (125)I-LR(3)IGF-I. In addition, significantly more intact (125)I-LR(3)IGF-I was recovered in wound fluid than (125)I-IGF-I at each time point, although only 0.08% of administered (125)I-LR(3)IGF-I was recovered per ml of wound fluid at 240 min. Compared with plasma, a greater proportion of wound fluid IGF-I radioactivity had distributed to the lower molecular weight IGFBPs or existed as free peptide. However, a small amount of wound fluid (125)I-IGF-I was detected in the 150 kDa region 30 min after injection. A greater proportion of (125)I-LR(3)IGF-I was associated with the lower molecular weight IGFBPs or existed as free peptide in both wound fluid and plasma. These data point to the importance of IGFBPs in determining the pharmacokinetic parameters of IGF-I in an extracellular fluid-expanded state. They also suggest only a minor role for endocrine IGF-I in surface wound repair.

Monitoring the quality of pre-registration education: development, validation and piloting of competency based performance indicators for newly qualified nurses.

The clinical competence or 'fitness for purpose' of newly qualified nurses continues to be an important professional and corporate issue that as yet has no objective means of assessment. A mixed group of managers, clinicians and educationalists was commissioned to develop a method for the measurement and evaluation of performance during the first year of employment of newly qualified nurses. Two instruments were developed and the results of the initial pilot study are demonstrated in this article. The results are preceded by a review of the relevant literature. The initial pilot study results indicate that in the case of both instruments the tests designed do give clear results on the small numbers used. A complete picture of the validity of the audit tool will not be seen until the results of the full validation study, including the above exercise, are known. However, the results to date indicate that the instruments have the potential to demonstrate the clinical competency of newly qualified staff on employment in their first post and their development over the first year of employment.