Accuracy of ultrasound signs on two-dimensional transvaginal ultrasound in prediction of adenomyosis: prospective multicenter study.

OBJECTIVE: Two-dimensional (2D) transvaginal ultrasound (TVS) is an accessible and cost-effective diagnostic tool for the detection of adenomyosis. Different ultrasound features related to adenomyosis have been described, but the predictive value of each ultrasound sign and their combinations requires further investigation. We aimed to analyze the accuracy of 2D-TVS and describe possible combinations of ultrasound signs with a high predictive value in the diagnosis of adenomyosis. METHODS: This was a prospective multicenter study of patients scheduled for laparoscopic hysterectomy who had been examined using standardized 2D-TVS at nine expert centers specializing in the diagnosis and treatment of endometriosis. 2D-TVS examination included nine typical adenomyosis ultrasound features, comprising heterogeneous myometrium, myometrial linear striations, myometrial cysts, subendometrial microcysts, asymmetrical myometrial thickening, uterine enlargement, the 'question mark sign', thickening of the junctional zone and hyperechoic myometrial spots, in order to predict or exclude the presence of adenomyosis. Ultrasound examination results were compared with histology after hysterectomy. The diagnostic reliability of the nine ultrasound signs and their combinations, and the influence of concurrent fibroids on the accuracy of the results, were analyzed. RESULTS: A total of 202 patients were enrolled into the study. Histopathological examination revealed adenomyosis in 130 patients (64.4%). The accuracy of prediction of adenomyosis by 2D-TVS examination using all signs was 63.4% (positive predictive value, 71.5%; negative predictive value, 48.6%; sensitivity, 71.5%; specificity, 48.6%). Heterogeneous myometrium, myometrial cysts, subendometrial microcysts and hyperechoic myometrial spots showed the highest accuracy (55.7-62.1%) as individual ultrasound signs for the prediction of adenomyosis. The combination of the most accurate ultrasound signs (subendometrial microcysts, myometrial cysts and heterogeneous myometrium) improved the specificity of prediction (86.1%) when compared with that of these three single markers (35.2-81.7%). Uterine enlargement and asymmetry showed both low sensitivity (60.8% and 52.3%, respectively) and specificity (41.7% and 49.3%, respectively) as individual sonographic signs. CONCLUSIONS: Heterogeneous myometrium, myometrial cysts, subendometrial microcysts and hyperechoic myometrial spots showed the highest accuracy for the detection of adenomyosis in this study, while uterine enlargement and asymmetry led to high false-positive and false-negative results. A combination of ultrasound features including the most accurate signs increases specificity. © 2023 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of International Society of Ultrasound in Obstetrics and Gynecology.

Shark depredation: future directions in research and management.

Shark depredation is a complex social-ecological issue that affects a range of fisheries worldwide. Increasing concern about the impacts of shark depredation, and how it intersects with the broader context of fisheries management, has driven recent research in this area, especially in Australia and the United States. This review synthesises these recent advances and provides strategic guidance for researchers aiming to characterise the occurrence of depredation, identify the shark species responsible, and test deterrent and management approaches to reduce its impacts. Specifically, the review covers the application of social science approaches, as well as advances in video camera and genetic methods for identifying depredating species. The practicalities and considerations for testing magnetic, electrical, and acoustic deterrent devices are discussed in light of recent research. Key concepts for the management of shark depredation are reviewed, with recommendations made to guide future research and policy development. Specific management responses to address shark depredation are lacking, and this review emphasizes that a "silver bullet" approach for mitigating depredation does not yet exist. Rather, future efforts to manage shark depredation must rely on a diverse range of integrated approaches involving those in the fishery (fishers, scientists and fishery managers), social scientists, educators, and other stakeholders.

Multiple mechanisms contribute to fluorometry signals from the voltage-gated proton channel.

Voltage-clamp fluorometry (VCF) supplies information about the conformational changes of voltage-gated proteins. Changes in the fluorescence intensity of the dye attached to a part of the protein that undergoes a conformational rearrangement upon the alteration of the membrane potential by electrodes constitute the signal. The VCF signal is generated by quenching and dequenching of the fluorescence as the dye traverses various local environments. Here we studied the VCF signal generation, using the Hv1 voltage-gated proton channel as a tool, which shares a similar voltage-sensor structure with voltage-gated ion channels but lacks an ion-conducting pore. Using mutagenesis and lipids added to the extracellular solution we found that the signal is generated by the combined effects of lipids during movement of the dye relative to the plane of the membrane and by quenching amino acids. Our 3-state model recapitulates the VCF signals of the various mutants and is compatible with the accepted model of two major voltage-sensor movements.

Benchmarking global fisheries discards.

Discarding by fisheries is one of the most wasteful human marine activities, yet we have few estimates of its scale. Reliable estimates of global discards are essential for sustainable fisheries management. Using United Nations Food and Agriculture Organization databases on country-specific landings, we estimated the discard rate and magnitude for global marine and estuarine capture fisheries using fishery-specific discard rates derived from direct observations and global gear-specific discard rates estimated within a Bayesian modelling framework. An estimated 9.1 million tonnes are discarded annually (95% uncertainty interval: 7-16 M t)-or 10.8% of the global catch (95% UI: 10-12%). Encouragingly, this is about half of the annual global discard rate estimated in the late 1980s. Trawl fisheries, especially demersal otter trawls, warrant intensified efforts to reduce discards. Periodic benchmarks of global discards are needed to assess the performance of reduction efforts.

Natural history and clinical characteristics of inhibitors in previously treated haemophilia A patients: a case series.

BACKGROUND: Development of inhibitors is the most serious complication in haemophilia A treatment. The assessment of risk for inhibitor formation in new or modified factor concentrates is traditionally performed in previously treated patients (PTPs). However, evidence on risk factors for and natural history of inhibitors has been generated mostly in previously untreated patients (PUPs). The purpose of this study was to examine cases of de novo inhibitors in PTPs reported in the scientific literature and to the EUropean HAemophilia Safety Surveillance (EUHASS) programme, and explore determinants and course of inhibitor development. METHODS: We used a case series study design and developed a case report form to collect patient level data; including detection, inhibitor course, treatment, factor VIII products used and events that may trigger inhibitor development (surgery, vaccination, immune disorders, malignancy, product switch). RESULTS: We identified 19 publications that reported 38 inhibitor cases and 45 cases from 31 EUHASS centres. Individual patient data were collected for 55/83 (66%) inhibitor cases out of 12 330 patients. The median (range) peak inhibitor titre was 4.4 (0.5-135.0), the proportion of transient inhibitors was 33% and only two cases of 12 undergoing immune tolerance induction failed this treatment. In the two months before inhibitor development, surgery was reported in nine (22%) cases, and high intensity treatment periods reported in seven (17%) cases. CONCLUSIONS: By studying the largest cohort of inhibitor development in PTPs assembled to date, we showed that inhibitor development in PTPs, is on average, a milder event than in PUPs.

Whole-GUV patch-clamping.

Studying how the membrane modulates ion channel and transporter activity is challenging because cells actively regulate membrane properties, whereas existing in vitro systems have limitations, such as residual solvent and unphysiologically high membrane tension. Cell-sized giant unilamellar vesicles (GUVs) would be ideal for in vitro electrophysiology, but efforts to measure the membrane current of intact GUVs have been unsuccessful. In this work, two challenges for obtaining the "whole-GUV" patch-clamp configuration were identified and resolved. First, unless the patch pipette and GUV pressures are precisely matched in the GUV-attached configuration, breaking the patch membrane also ruptures the GUV. Second, GUVs shrink irreversibly because the membrane/glass adhesion creating the high-resistance seal (>1 GΩ) continuously pulls membrane into the pipette. In contrast, for cell-derived giant plasma membrane vesicles (GPMVs), breaking the patch membrane allows the GPMV contents to passivate the pipette surface, thereby dynamically blocking membrane spreading in the whole-GMPV mode. To mimic this dynamic passivation mechanism, beta-casein was encapsulated into GUVs, yielding a stable, high-resistance, whole-GUV configuration for a range of membrane compositions. Specific membrane capacitance measurements confirmed that the membranes were truly solvent-free and that membrane tension could be controlled over a physiological range. Finally, the potential for ion transport studies was tested using the model ion channel, gramicidin, and voltage-clamp fluorometry measurements were performed with a voltage-dependent fluorophore/quencher pair. Whole-GUV patch-clamping allows ion transport and other voltage-dependent processes to be studied while controlling membrane composition, tension, and shape.

Physical basis of apparent pore dilation of ATP-activated P2X receptor channels.

The selectivity of ion channels is fundamental for their roles in electrical and chemical signaling and in ion homeostasis. Although most ion channels exhibit stable ion selectivity, the prevailing view of purinergic P2X receptor channels, transient receptor potential V1 (TRPV1) channels and acid-sensing ion channels (ASICs) is that their ion conduction pores dilate upon prolonged activation. We investigated this mechanism in P2X receptors and found that the hallmark shift in equilibrium potential observed with prolonged channel activation does not result from pore dilation, but from time-dependent alterations in the concentration of intracellular ions. We derived a physical model to calculate ion concentration changes during patch-clamp recordings, which validated our experimental findings and provides a quantitative guideline for effectively controlling ion concentration. Our results have fundamental implications for understanding ion permeation and gating in P2X receptor channels, as well as more broadly for using patch-clamp techniques to study ion channels and neuronal excitability.

Reconstitution of a transmembrane protein, the voltage-gated ion channel, KvAP, into giant unilamellar vesicles for microscopy and patch clamp studies.

Giant Unilamellar Vesicles (GUVs) are a popular biomimetic system for studying membrane associated phenomena. However, commonly used protocols to grow GUVs must be modified in order to form GUVs containing functional transmembrane proteins. This article describes two dehydration-rehydration methods - electroformation and gel-assisted swelling - to form GUVs containing the voltage-gated potassium channel, KvAP. In both methods, a solution of protein-containing small unilamellar vesicles is partially dehydrated to form a stack of membranes, which is then allowed to swell in a rehydration buffer. For the electroformation method, the film is deposited on platinum electrodes so that an AC field can be applied during film rehydration. In contrast, the gel-assisted swelling method uses an agarose gel substrate to enhance film rehydration. Both methods can produce GUVs in low (e.g., 5 mM) and physiological (e.g., 100 mM) salt concentrations. The resulting GUVs are characterized via fluorescence microscopy, and the function of reconstituted channels measured using the inside-out patch-clamp configuration. While swelling in the presence of an alternating electric field (electroformation) gives a high yield of defect-free GUVs, the gel-assisted swelling method produces a more homogeneous protein distribution and requires no special equipment.

Membrane shape modulates transmembrane protein distribution.

Although membrane shape varies greatly throughout the cell, the contribution of membrane curvature to transmembrane protein targeting is unknown because of the numerous sorting mechanisms that take place concurrently in cells. To isolate the effect of membrane shape, we used cell-sized giant unilamellar vesicles (GUVs) containing either the potassium channel KvAP or the water channel AQP0 to form membrane nanotubes with controlled radii. Whereas the AQP0 concentrations in flat and curved membranes were indistinguishable, KvAP was enriched in the tubes, with greater enrichment in more highly curved membranes. Fluorescence recovery after photobleaching measurements showed that both proteins could freely diffuse through the neck between the tube and GUV, and the effect of each protein on membrane shape and stiffness was characterized using a thermodynamic sorting model. This study establishes the importance of membrane shape for targeting transmembrane proteins and provides a method for determining the effective shape and flexibility of membrane proteins.

Causes and methods to estimate cryptic sources of fishing mortality.

Cryptic, not readily detectable, components of fishing mortality are not routinely accounted for in fisheries management because of a lack of adequate data, and for some components, a lack of accurate estimation methods. Cryptic fishing mortalities can cause adverse ecological effects, are a source of wastage, reduce the sustainability of fishery resources and, when unaccounted for, can cause errors in stock assessments and population models. Sources of cryptic fishing mortality are (1) pre-catch losses, where catch dies from the fishing operation but is not brought onboard when the gear is retrieved, (2) ghost-fishing mortality by fishing gear that was abandoned, lost or discarded, (3) post-release mortality of catch that is retrieved and then released alive but later dies as a result of stress and injury sustained from the fishing interaction, (4) collateral mortalities indirectly caused by various ecological effects of fishing and (5) losses due to synergistic effects of multiple interacting sources of stress and injury from fishing operations, or from cumulative stress and injury caused by repeated sub-lethal interactions with fishing operations. To fill a gap in international guidance on best practices, causes and methods for estimating each component of cryptic fishing mortality are described, and considerations for their effective application are identified. Research priorities to fill gaps in understanding the causes and estimating cryptic mortality are highlighted.

Functional reconstitution of a voltage-gated potassium channel in giant unilamellar vesicles.

Voltage-gated ion channels are key players in cellular excitability. Recent studies suggest that their behavior can depend strongly on the membrane lipid composition and physical state. In vivo studies of membrane/channel and channel/channel interactions are challenging as membrane properties are actively regulated in living cells, and are difficult to control in experimental settings. We developed a method to reconstitute functional voltage-gated ion channels into cell-sized Giant Unilamellar Vesicles (GUVs) in which membrane composition, tension and geometry can be controlled. First, a voltage-gated potassium channel, KvAP, was purified, fluorescently labeled and reconstituted into small proteoliposomes. Small proteoliposomes were then converted into GUVs via electroformation. GUVs could be formed using different lipid compositions and buffers containing low (5 mM) or near-physiological (100 mM) salt concentrations. Protein incorporation into GUVs was characterized with quantitative confocal microscopy, and the protein density of GUVs was comparable to the small proteoliposomes from which they were formed. Furthermore, patch-clamp measurements confirmed that the reconstituted channels retained potassium selectivity and voltage-gated activation. GUVs containing functional voltage-gated ion channels will allow the study of channel activity, distribution and diffusion while controlling membrane state, and should prove a powerful tool for understanding how the membrane modulates cellular excitability.

Mobility in geometrically confined membranes.

Lipid and protein lateral mobility is essential for biological function. Our theoretical understanding of this mobility can be traced to the seminal work of Saffman and Delbrück, who predicted a logarithmic dependence of the protein diffusion coefficient (i) on the inverse of the size of the protein and (ii) on the "membrane size" for membranes of finite size [Saffman P, Delbrück M (1975) Proc Natl Acad Sci USA 72:3111-3113]. Although the experimental proof of the first prediction is a matter of debate, the second has not previously been thought to be experimentally accessible. Here, we construct just such a geometrically confined membrane by forming lipid bilayer nanotubes of controlled radii connected to giant liposomes. We followed the diffusion of individual molecules in the tubular membrane using single particle tracking of quantum dots coupled to lipids or voltage-gated potassium channels KvAP, while changing the membrane tube radius from approximately 250 to 10 nm. We found that both lipid and protein diffusion was slower in tubular membranes with smaller radii. The protein diffusion coefficient decreased as much as 5-fold compared to diffusion on the effectively flat membrane of the giant liposomes. Both lipid and protein diffusion data are consistent with the predictions of a hydrodynamic theory that extends the work of Saffman and Delbrück to cylindrical geometries. This study therefore provides strong experimental support for the ubiquitous Saffman-Delbrück theory and elucidates the role of membrane geometry and size in regulating lateral diffusion.

EUHASS: The European Haemophilia Safety Surveillance system.

Pharmacovigilance is an essential element of any drug treatment and considering the history of adverse events due to products used to treat inherited bleeding disorders, it should be an integral component of modern haemophilia treatment. Because inherited bleeding disorders and adverse events are rare, a multicentre, preferably multinational, adverse event reporting scheme for all clotting factor products is required. EUHASS is a European, prospective, multicentre adverse event reporting scheme in the field of inherited bleeding disorders.

Lipid cosorting mediated by shiga toxin induced tubulation.

To maintain cell membrane homeostasis, lipids must be dynamically redistributed during the formation of transport intermediates, but the mechanisms driving lipid sorting are not yet fully understood. Lowering sphingolipid concentration can reduce the bending energy of a membrane, and this effect could account for sphingolipid depletion along the retrograde pathway. However, sphingolipids and cholesterol are enriched along the anterograde pathway, implying that other lipid sorting mechanisms, such as protein-mediated sorting, can dominate. To characterize the influence of protein binding on the lipid composition of highly curved membranes, we studied the interactions of the B-subunit of Shiga toxin (STxB) with giant unilamellar vesicles containing its glycosphingolipid receptor [globotriaosylceramide (Gb3)]. STxB binding induced the formation of tubular membrane invaginations, and fluorescence microscopy images of these highly curved membranes were consistent with co-enrichment of Gb3 and sphingolipids. In agreement with theory, sorting was stronger for membrane compositions close to demixing. These results strongly support the hypothesis that proteins can indirectly mediate the sorting of lipids into highly curved transport intermediates via interactions between lipids and the membrane receptor of the protein.

A bicontinuous double gyroid hybrid solar cell.

We report the first successful application of an ordered bicontinuous gyroid semiconducting network in a hybrid bulk heterojunction solar cell. The freestanding gyroid network is fabricated by electrochemical deposition into the 10 nm wide voided channels of a self-assembled, selectively degradable block copolymer film. The highly ordered pore structure is ideal for uniform infiltration of an organic hole transporting material, and solid-state dye-sensitized solar cells only 400 nm thick exhibit up to 1.7% power conversion efficiency. This patterning technique can be readily extended to other promising heterojunction systems and is a major step toward realizing the full potential of self-assembly in the next generation of device technologies.

Structures of beta-amino ester enolates: new strategies using the method of continuous variation.

The solution structures of four enolates derived from beta-amino esters are investigated using (6)Li NMR spectroscopy in conjunction with the method of continuous variation (method of Job). Ensembles of homo- and heteroaggregated enolates are generated by mixing enantiomers of a single enolate (R/S mixtures), opposite antipodes of two different enolates (R/S' mixtures), and the same antipodes of two different enolates (R/R' mixtures). The numbers of observable aggregates and their dependence on the mole fraction of the two enolates confirm the hexamer assignments. Inherent symmetries observable in the (6)Li NMR spectra show the stereochemistry of chelation about the hexagonal drum.

Liquid-liquid domains in bilayers detected by wide angle X-ray scattering.

Wide angle x-ray scattering (WAXS) from oriented lipid multilayers is used to examine liquid-ordered (Lo)/liquid-disordered (Ld) phase coexistence in the system 1,2-dioleoyl-sn-glycero-3-phosphocholine/1,2-dipalmitoyl-sn-glycero-3-phosphocholine/cholesterol (DOPC/DPPC/Chol), which is a model for the outer leaflet of the animal cell plasma membrane. Using the method of analysis developed in the accompanying work, we find that two orientational distributions are necessary to fit the WAXS data at lower temperatures, whereas only one distribution is needed at temperatures higher than the miscibility transition temperature, T(mix) = 25-35 degrees C (for 1:1 DOPC/DPPC with 15%, 20%, 25%, and 30% Chol). We propose that the necessity for two distributions is a criterion for coexistence of Lo domains with a high S(x-ray) order parameter and Ld domains with a lower order parameter. This criterion is capable of detecting coexistence of small domains or rafts that the conventional x-ray criterion of two lamellar D spacings may not. Our T(mix) values tend to be slightly larger than published NMR results and microscopy results when the fluorescence probe artifact is considered. This is consistent with the sensitivity of WAXS to very short time and length scales, which makes it more capable of detecting small, short-lived domains that are likely close to T(mix).

Order parameters and areas in fluid-phase oriented lipid membranes using wide angle X-ray scattering.

We used wide angle x-ray scattering (WAXS) from stacks of oriented lipid bilayers to measure chain orientational order parameters and lipid areas in model membranes consisting of mixtures of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC)/cholesterol and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC)/cholesterol in fluid phases. The addition of 40% cholesterol to either DOPC or DPPC changes the WAXS pattern due to an increase in acyl chain orientational order, which is one of the main properties distinguishing the cholesterol-rich liquid-ordered (Lo) phase from the liquid-disordered (Ld) phase. In contrast, powder x-ray data from multilamellar vesicles does not yield information about orientational order, and the scattering from the Lo and Ld phases looks similar. An analytical model to describe the relationship between the chain orientational distribution and WAXS data was used to obtain an average orientational order parameter, S(x-ray). When 40% cholesterol is added to either DOPC or DPPC, S(x-ray) more than doubles, consistent with previous NMR order parameter measurements. By combining information about the average chain orientation with the chain-chain correlation spacing, we extended a commonly used method for calculating areas for gel-phase lipids to fluid-phase lipids and obtained agreement to within 5% of literature values.