Extracellular Kir2.1C122Y Mutant Upsets Kir2.1-PIP2 Bonds and Is Arrhythmogenic in Andersen-Tawil Syndrome.

BACKGROUND: Andersen-Tawil syndrome type 1 is a rare heritable disease caused by mutations in the gene coding the strong inwardly rectifying K+ channel Kir2.1. The extracellular Cys (cysteine)122-to-Cys154 disulfide bond in the channel structure is crucial for proper folding but has not been associated with correct channel function at the membrane. We evaluated whether a human mutation at the Cys122-to-Cys154 disulfide bridge leads to Kir2.1 channel dysfunction and arrhythmias by reorganizing the overall Kir2.1 channel structure and destabilizing its open state. METHODS: We identified a Kir2.1 loss-of-function mutation (c.366 A>T; p.Cys122Tyr) in an ATS1 family. To investigate its pathophysiological implications, we generated an AAV9-mediated cardiac-specific mouse model expressing the Kir2.1C122Y variant. We employed a multidisciplinary approach, integrating patch clamping and intracardiac stimulation, molecular biology techniques, molecular dynamics, and bioluminescence resonance energy transfer experiments. RESULTS: Kir2.1C122Y mice recapitulated the ECG features of ATS1 independently of sex, including corrected QT prolongation, conduction defects, and increased arrhythmia susceptibility. Isolated Kir2.1C122Y cardiomyocytes showed significantly reduced inwardly rectifier K+ (IK1) and inward Na+ (INa) current densities independently of normal trafficking. Molecular dynamics predicted that the C122Y mutation provoked a conformational change over the 2000-ns simulation, characterized by a greater loss of hydrogen bonds between Kir2.1 and phosphatidylinositol 4,5-bisphosphate than wild type (WT). Therefore, the phosphatidylinositol 4,5-bisphosphate-binding pocket was destabilized, resulting in a lower conductance state compared with WT. Accordingly, on inside-out patch clamping, the C122Y mutation significantly blunted Kir2.1 sensitivity to increasing phosphatidylinositol 4,5-bisphosphate concentrations. In addition, the Kir2.1C122Y mutation resulted in channelosome degradation, demonstrating temporal instability of both Kir2.1 and NaV1.5 proteins. CONCLUSIONS: The extracellular Cys122-to-Cys154 disulfide bond in the tridimensional Kir2.1 channel structure is essential for the channel function. We demonstrate that breaking disulfide bonds in the extracellular domain disrupts phosphatidylinositol 4,5-bisphosphate-dependent regulation, leading to channel dysfunction and defects in Kir2.1 energetic stability. The mutation also alters functional expression of the NaV1.5 channel and ultimately leads to conduction disturbances and life-threatening arrhythmia characteristic of Andersen-Tawil syndrome type 1.

A decay-modeled compressed sensing reconstruction approach for non-Cartesian hyperpolarized 129Xe MRI.

PURPOSE: Hyperpolarized 129Xe MRI benefits from non-Cartesian acquisitions that sample k-space efficiently and rapidly. However, their reconstructions are complex and burdened by decay processes unique to hyperpolarized gas. Currently used gridded reconstructions are prone to artifacts caused by magnetization decay and are ill-suited for undersampling. We present a compressed sensing (CS) reconstruction approach that incorporates magnetization decay in the forward model, thereby producing images with increased sharpness and contrast, even in undersampled data. METHODS: Radio-frequency, T1, and T 2 * $$ {\mathrm{T}}_2^{\ast } $$ decay processes were incorporated into the forward model and solved using iterative methods including CS. The decay-modeled reconstruction was validated in simulations and then tested in 2D/3D-spiral ventilation and 3D-radial gas-exchange MRI. Quantitative metrics including apparent-SNR and sharpness were compared between gridded, CS, and twofold undersampled CS reconstructions. Observations were validated in gas-exchange data collected from 15 healthy and 25 post-hematopoietic-stem-cell-transplant participants. RESULTS: CS reconstructions in simulations yielded images with threefold increases in accuracy. CS increased sharpness and contrast for ventilation in vivo imaging and showed greater accuracy for undersampled acquisitions. CS improved gas-exchange imaging, particularly in the dissolved-phase where apparent-SNR improved, and structure was made discernable. Finally, CS showed repeatability in important global gas-exchange metrics including median dissolved-gas signal ratio and median angle between real/imaginary components. CONCLUSION: A non-Cartesian CS reconstruction approach that incorporates hyperpolarized 129Xe decay processes is presented. This approach enables improved image sharpness, contrast, and overall image quality in addition to up-to threefold undersampling. This contribution benefits all hyperpolarized gas MRI through improved accuracy and decreased scan durations.

Assessing Outcomes of Patients Subject to Intensive Care to Facilitate Organ Donation: A Spanish Multicenter Prospective Study.

Intensive Care to facilitate Organ Donation (ICOD) consists of the initiation or continuation of intensive care measures in patients with a devastating brain injury (DBI) in whom curative treatment is deemed futile and death by neurological criteria (DNC) is foreseen, to incorporate organ donation into their end-of-life plans. In this study we evaluate the outcomes of patients subject to ICOD and identify radiological and clinical factors associated with progression to DNC. In this first prospective multicenter study we tested by multivariate regression the association of clinical and radiological severity features with progression to DNC. Of the 194 patients, 144 (74.2%) patients fulfilled DNC after a median of 25 h (95% IQR: 17-44) from ICOD onset. Two patients (1%) shifted from ICOD to curative treatment, both were alive at discharge. Factors associated with progression to DNC included: age below 70 years, clinical score consistent with severe brain injury, instability, intracranial hemorrhage, midline shift ≥5 mm and certain types of brain herniation. Overall 151 (77.8%) patients progressed to organ donation. Based on these results, we conclude that ICOD is a beneficial and efficient practice that can contribute to the pool of deceased donors.

The spike protein of SARS-CoV-2 induces endothelial inflammation through integrin α5ß1 and NF-κB signaling.

Vascular endothelial cells (ECs) form a critical interface between blood and tissues that maintains whole-body homeostasis. In COVID-19, disruption of the EC barrier results in edema, vascular inflammation, and coagulation, hallmarks of this severe disease. However, the mechanisms by which ECs are dysregulated in COVID-19 are unclear. Here, we show that the spike protein of SARS-CoV-2 alone activates the EC inflammatory phenotype in a manner dependent on integrin ⍺5ß1 signaling. Incubation of human umbilical vein ECs with whole spike protein, its receptor-binding domain, or the integrin-binding tripeptide RGD induced the nuclear translocation of NF-κB and subsequent expression of leukocyte adhesion molecules (VCAM1 and ICAM1), coagulation factors (TF and FVIII), proinflammatory cytokines (TNFα, IL-1ß, and IL-6), and ACE2, as well as the adhesion of peripheral blood leukocytes and hyperpermeability of the EC monolayer. In addition, inhibitors of integrin ⍺5ß1 activation prevented these effects. Furthermore, these vascular effects occur in vivo, as revealed by the intravenous administration of spike, which increased expression of ICAM1, VCAM1, CD45, TNFα, IL-1ß, and IL-6 in the lung, liver, kidney, and eye, and the intravitreal injection of spike, which disrupted the barrier function of retinal capillaries. We suggest that the spike protein, through its RGD motif in the receptor-binding domain, binds to integrin ⍺5ß1 in ECs to activate the NF-κB target gene expression programs responsible for vascular leakage and leukocyte adhesion. These findings uncover a new direct action of SARS-CoV-2 on EC dysfunction and introduce integrin ⍺5ß1 as a promising target for treating vascular inflammation in COVID-19.

Solid Dispersion Incorporated into Dissolving Microneedles for Improved Antifungal Activity of Amphotericin B: In Vivo Study in a Fungal Keratitis Model.

Fungal keratitis (FK) is a fungal infection of the cornea, which is part of the eye and causes corneal ulcers and an increased risk of permanent blindness, which is often found in Candida albicans species. Amphotericin B (AMB), which is a group of polyenes as the first-line treatment of FK, is effective in annihilating C. albicans. However, AMB preparations such as eye drops and ointments have major drawbacks, for instance, requiring more frequent administrations, loss of the drug by the drainage process, and rapid elimination in the precornea, which result in low bioavailability of the drug. An ocular dissolving microneedle containing the solid dispersion amphotericin B (DMN-SD-AMB) had been developed using a mixture of poly(vinyl alcohol) (PVA) and poly(vinylpyrrolidone) (PVP) polymers, while the solid dispersion AMB (SD-AMB) was contained in the needle as a drug. This study aims to determine the most optimal and safest DMN-SD-AMB formula for the treatment of FK in the eye as well as a solution to overcome the low bioavailability of AMB eye drops and ointment preparations. SD-AMB had been successfully developed, which was characterized by increased antifungal activity and drug release in vitro compared to other treatments. Furthermore, DMN-SD-AMB studies had also been successfully performed with the best formulation, which exhibited the best ex vivo corneal permeation profile and antifungal activity as well as being safe from eye irritation. In addition, an in vivo antifungal activity using a rabbit infection model shows that the number of fungal colonies was 0.98 ± 0.11 log10 CFU/mL (F3), 5.76 ± 0.32 log10 CFU/mL (AMB eye drops), 4.01 ± 0.28 log10 CFU/mL (AMB ointments), and 9.09 ± 0.65 log10 CFU/mL (control), which differed significantly (p < 0.05). All of these results evidence that DMN-SD-AMB is a new approach to developing intraocular preparations for the treatment of FK.

Extrusion-based technologies for 3D printing: a comparative study of the processability of thermoplastic polyurethane-based formulations.

Thermoplastic polyurethanes (TPU) offer excellent properties for a wide range of dosage forms. These polymers have been successfully utilized in personalized medicine production using fused deposition modeling (FDM) 3D printing (3DP). However, direct powder extrusion (DPE) has been introduced recently as a challenging technique since it eliminates filament production before 3DP, reducing thermal stress, production time, and costs. This study compares DPE and single-screw extrusion for binary (drug-TPU) and ternary (drug-TPU-magnesium stearate [MS]) mixtures containing from 20 to 60% w/w of theophylline. Powder flow, mechanical properties, fractal analysis, and percolation theory were utilized to analyze critical properties of the extrudates. All the mixtures could be processed at a temperature range between 130 and 160 °C. Extrudates containing up to 50% w/w of drug (up to 30% w/w of drug in the case of single-screw extrusion binary filaments) showed toughness values above the critical threshold of 80 kg/mm2. MS improved flow in mixtures where the drug is the only percolating component, reduced until 25 °C the DPE temperature and decreased the extrudate roughness in high drug content systems. The potential of DPE as an efficient one-step additive manufacturing technique in healthcare environments to produce TPU-based tailored on-demand medicines has been demonstrated.

The HGR motif is the antiangiogenic determinant of vasoinhibin: implications for a therapeutic orally active oligopeptide.

The hormone prolactin acquires antiangiogenic and antivasopermeability properties after undergoing proteolytic cleavage to vasoinhibin, an endogenous prolactin fragment of 123 or more amino acids that inhibits the action of multiple proangiogenic factors. Preclinical and clinical evidence supports the therapeutic potential of vasoinhibin against angiogenesis-related diseases including diabetic retinopathy, peripartum cardiomyopathy, rheumatoid arthritis, and cancer. However, the use of vasoinhibin in the clinic has been limited by difficulties in its production. Here, we removed this barrier to using vasoinhibin as a therapeutic agent by showing that a short linear motif of just three residues (His46-Gly47-Arg48) (HGR) is the functional determinant of vasoinhibin. The HGR motif is conserved throughout evolution, its mutation led to vasoinhibin loss of function, and oligopeptides containing this sequence inhibited angiogenesis and vasopermeability with the same potency as whole vasoinhibin. Furthermore, the oral administration of an optimized cyclic retro-inverse vasoinhibin heptapeptide containing HGR inhibited melanoma tumor growth and vascularization in mice and exhibited equal or higher antiangiogenic potency than other antiangiogenic molecules currently used as anti-cancer drugs in the clinic. Finally, by unveiling the mechanism that obscures the HGR motif in prolactin, we anticipate the development of vasoinhibin-specific antibodies to solve the on-going challenge of measuring endogenous vasoinhibin levels for diagnostic and interventional purposes, the design of vasoinhibin antagonists for managing insufficient angiogenesis, and the identification of putative therapeutic proteins containing HGR.

Elucidating the Impact of Surfactants on the Performance of Dissolving Microneedle Array Patches.

The need for biocompatible polymers capable of dissolving in the skin while exhibiting reasonable mechanical features and delivery efficiency limits the range of materials that could be utilized in fabricating dissolving microneedle array patches (MAPs). The incorporation of additives, such as surfactants, during microneedle fabrication might be an alternative solution to overcome the limited range of materials used in fabricating dissolving MAPs. However, there is a lacuna in the knowledge on the effect of surfactants on the manufacture and performance of dissolving MAPs. The current study explores the role of surfactants in the manufacture and performance of dissolving MAPs fabricated from poly(vinyl alcohol) (PVA) and poly(vinyl pyrrolidone) (PVP) loaded with the model drugs, ibuprofen sodium and itraconazole. Three nonionic surfactants, Lutrol F108, Pluronic F88, and Tween 80, in solutions at varying concentrations (0.5, 1.0, and 2.0% w/w) were loaded into these dissolving MAPs. It was discovered that all of the dissolving MAPs that incorporated surfactant displayed a lower reduction in the microneedle height (≈10%) relative to the control formulation (≈20%) when subjected to a compressive force of 32 N. In addition, the incorporation of surfactants in some instances enhanced the insertion profile of these polymeric MAPs when evaluated using ex vivo neonatal porcine skin. The incorporation of surfactant into ibuprofen sodium-loaded dissolving MAPs improved the insertion depth of MAPs from 400 µm down to 600 µm. However, such enhancement was not apparent when the MAPs were loaded with the model hydrophobic drug, itraconazole. Skin deposition studies highlighted that the incorporation of surfactant enhanced the delivery efficiency of both model drugs, ibuprofen sodium and itraconazole. The incorporation of surfactant enhanced the amount of ibuprofen sodium delivered from 60.61% up to ≈75% with a majority of the drug being delivered across the skin and into the receptor compartment. On the other hand, when surfactants were added into MAPs loaded with the model hydrophobic drug itraconazole, we observed enhancement in intradermal delivery efficiency from 20% up to 30%, although this did not improve the delivery of the drug across the skin. This work highlights that the addition of nonionic surfactant is an alternative formulation strategy worth exploring to improve the performance and delivery efficiency of dissolving MAPs.

The Role of Manganese in Very Low Birth Weight Infants.

OBJECTIVES: High manganese (Mn) levels during fetal growth or prolonged parenteral nutrition (PN) may have adverse effects on neurodevelopment. We aim to report on Mn levels and their short-term impact on clinical course in very low birth weight infants. METHODS: An observational study including newborns with a gestational age (GA) ≤32 weeks and/or ≤1500 g of birth weight (BW). Newborns received intravenous supplementation of Mn at 1 µg/kg/day (Peditrace ® ) in PN and continued with fortified breast milk. Mothers answered surveys about dietary and other habits and blood levels of Mn in newborns were analyzed at days 1, 15, and 30 of life. Associations of Mn levels with mothers' and newborns' data were evaluated and adjusted for multiple comparisons. RESULTS: One hundred and sixty premature infants were recruited. Median blood Mn levels at birth were 43.0 and 24.5 µg/L at day 30. No important association with mothers' data was found. Median [interquartile range (IQR)] duration of PN was 8 days (7-14). A prolonged PN and late oral feeding showed a nonsignificant association with lower blood Mn levels at day 30 ( P = 0.010, P threshold 0.003). Mn levels at day 15 and 30 were associated with increasing GA ( P < 0.001). Low Mn was not a significant predictor of adverse outcomes such as retinopathy of prematurity, bronchopulmonary dysplasia, or respiratory distress syndrome after adjusting for potential confounders and multiple testing. CONCLUSIONS: Mn showed lower levels with decreasing GA and prolonged PN. Using a low Mn PN solution may not raise blood Mn levels in premature infants.

A New and Sensitive HPLC-UV Method for Rapid and Simultaneous Quantification of Curcumin and D-Panthenol: Application to In Vitro Release Studies of Wound Dressings.

Curcumin (CUR) and D-panthenol (DPA) have been widely investigated for wound-healing treatment. In order to analyse these two compounds from a dosage form, such as polymer-based wound dressings or creams, an analytical method that allows the quantification of both drugs simultaneously should be developed. Here, we report for the first time a validated high-performance liquid chromatographic (HPLC) method coupled with UV detection to quantify CUR and DPA based on the standards set by the International Council on Harmonization (ICH) guidelines. The separation of the analytes was performed using a C18 column that utilised a mobile phase consisting of 0.001% v/v phosphoric acid and methanol using a gradient method with a run time of 15 min. The method is linear for drug concentrations within the range of 0.39-12.5 µg mL-1 (R2 = 0.9999) for CUR and 0.39-25 µg mL-1 for DPA (R2 = 1). The validated method was found to be precise and accurate. Moreover, the CUR and DPA solution was found to be stable under specific storage conditions. We, therefore, suggest that the HPLC-UV method developed in this study may be very useful in screening formulations for CUR and DPA within a preclinical setting through in vitro release studies.

Finite element simulations of hyperpolarized gas DWI in micro-CT meshes of acinar airways: validating the cylinder and stretched exponential models of lung microstructural length scales.

PURPOSE: This work assesses the accuracy of the stretched exponential (SEM) and cylinder models of lung microstructural length scales that can be derived from hyperpolarized gas DWI. This was achieved by simulating 3 He and 129 Xe DWI signals within two micro-CT-derived realistic acinar airspace meshes that represent healthy and idiopathic pulmonary fibrosis lungs. METHODS: The healthy and idiopathic pulmonary fibrosis acinar airway meshes were derived from segmentations of 3D micro-CT images of excised human lungs and meshed for finite element simulations of the Bloch-Torrey equations. 3 He and 129 Xe multiple b value DWI experiments across a range of diffusion times (3 He Δ = 1.6 ms; 129 Xe Δ = 5 to 20 ms) were simulated in each mesh. Global SEM mean diffusive length scale and cylinder model mean chord length value was derived from each finite element simulation and compared against each mesh's mean linear intercept length, calculated from intercept length measurements within micro-CT segmentation masks. RESULTS: The SEM-derived mean diffusive length scale was within ±10% of the mean linear intercept length for simulations with both 3 He (Δ = 1.6 ms) and 129 Xe (Δ = 7 to 13 ms) in the healthy mesh, and with 129 Xe (Δ = 13 to 20 ms) for the idiopathic pulmonary fibrosis mesh, whereas for the cylinder model-derived mean chord length the closest agreement with mean linear intercept length (11.7% and 22.6% difference) was at 129 Xe Δ = 20 ms for both healthy and IPF meshes, respectively. CONCLUSION: This work validates the use of the SEM for accurate estimation of acinar dimensions and indicates that the SEM is relatively robust across a range of experimental conditions and acinar length scales.

An Approach to Faculty Development for Underrepresented Minorities in Medicine.

OBJECTIVES: The diversity of the US physician workforce lags significantly behind the population, and the disparities in academic medicine are even greater, with underrepresented in medicine (URM) physicians accounting for only 6.8% of all US medical school faculty. We describe a "for URM by URM" pilot approach to faculty development for junior URM Family Medicine physicians that targets unique challenges faced by URM faculty. METHODS: A year-long fellowship was created for junior URM academic clinician faculty with funding through the Society of Teachers of Family Medicine Project Fund. Seven junior faculty applied and were accepted to participate in the fellowship, which included conference calls and an in-person workshop covering topics related to writing and career advancement. RESULTS: The workshop included a mix of prepared programming on how to move from idea to project to manuscript, as well as time for spontaneous mentorship and manuscript collaboration. Key themes that emerged included how to address the high cost of the minority tax, the need for individual passion as a pathway to success, and how to overcome imposter syndrome as a hindrance to writing. CONCLUSIONS: The "for URM by URM" approach for faculty development to promote writing skills and scholarship for junior URM Family Medicine physicians can address challenges faced by URM faculty. By using a framework that includes the mentors' lived experiences and creates a psychological safe space, we can address concerns often overlooked in traditional skills-based faculty development programs.

Poly(caprolactone)-Based Coatings on 3D-Printed Biodegradable Implants: A Novel Strategy to Prolong Delivery of Hydrophilic Drugs.

Implantable devices are versatile and promising drug delivery systems, and their advantages are well established. Of these advantages, long-acting drug delivery is perhaps the most valuable. Hydrophilic compounds are particularly difficult to deliver for prolonged times. This work investigates the use of poly(caprolactone) (PCL)-based implant coatings as a novel strategy to prolong the delivery of hydrophilic compounds from implantable devices that have been prepared by additive manufacturing (AM). Hollow implants were prepared from poly(lactic acid) (PLA) and poly(vinyl alcohol) (PVA) using fused filament fabrication (FFF) AM and subsequently coated in a PCL-based coating. Coatings were prepared by solution-casting mixtures of differing molecular weights of PCL and poly(ethylene glycol) (PEG). Increasing the proportion of low-molecular-weight PCL up to 60% in the formulations decreased the crystallinity by over 20%, melting temperature by over 4 °C, and water contact angle by over 40°, resulting in an increased degradation rate when compared to pure high-molecular-weight PCL. Addition of 30% PEG to the formulation increased the porosity of the formulation by over 50% when compared to an equivalent PCL-only formulation. These implants demonstrated in vitro release rates for hydrophilic model compounds (methylene blue and ibuprofen sodium) ranging from 0.01 to 34.09 mg/day, depending on the drug used. The versatility of the devices produced in this work and the range of release rates achievable show great potential. Implants could be specifically developed in order to match the specific release rate required for a number of drugs for a wide range of conditions.

One-step synthesis of carbon nanospheres with an encapsulated iron-nickel nanoalloy and its potential use as an electrocatalyst.

For many years, in electrochemical processes, carbon nanostructures with metal support have been employed as electrodes due to their high surface area, chemical stability, and excellent performance as catalyst support by allowing a better electronic transfer. Nevertheless, on the surface, metallic nanoparticles are susceptible to corrosion. Instead, by encapsulating individual nanoparticles, they are protected. Among the carbon nanostructures, the most common are graphene, carbon nanotubes (CNTs), and carbon nanospheres (CNSs). Unlike CNTs and CNSs, graphene is difficult to obtain in mass production, limiting their applications. Regarding CNTs and CNSs, the latter presents better catalytic activity. Nonetheless, the process of synthesis of CNSs with metal inside is commonly made by time-consuming autoclave processes, some involving more than 43 h, and hence are expensive. Here, we suggest an advantageous synthesis of CNSs with an iron-nickel alloy encapsulated inside, by using a one-step chemical vapor deposition (CVD) process in less than 3 h. This material has potential applications for environmental and energy processes. According to the authors, the uses of iron-nickel alloys as an electrocatalyst for the ammonia oxidation reaction has not been proved. Thus, we evaluate the composite as an electrocatalyst for the ammonia oxidation reaction, an electrochemical process that offers environmental remediation and hydrogen as a fuel. The electrochemical characterization shows that the use of a bimetallic electrode improves the catalytic activity. In this case, nickel is the active specie and iron is the metal added which reduces the reaction potential. Besides, the composite presents high specific capacitance, better than other materials proposed such as graphene decorated with FeNi alloys. This behavior can be related to the variation of the catalyst morphology (supported vs. encapsulated) by improving the catalyst dispersion and particle size stabilization.

Nickel supported on carbon nanotubes and carbon nanospheres for ammonia oxidation reaction.

Here it is proposed the use of nickel supported on carbon nanotubes and carbon nanospheres as a cheaper catalyst alternative to the platinum electrodes for the oxidation process of ammonia. Using scanning electron microscopy and Raman spectroscopy confirmed the presence of the Ni on the surface of the carbon nanostructures and based on the electrochemical results is established that the redox process between Ni+2 and Ni+3 plays the role of intermediate in the ammonia oxidation reaction. This process causes an increment in the anodic current density that is dependent on the pH and ammonia concentration. Even more, using the rotating disk electrode technique, the ammonia oxidation, is defined as a diffusion-controlled process that follows first-order kinetics in respect of the concentration of ammonia and suggests the formation of nitrogen as the principal reaction product, where the carbon nanospheres present the best results.

Thoughts and expectations of young professionals about the European Federation of Clinical Chemistry and Laboratory Medicine (EFLM).

Objectives: Young laboratory medicine professionals (YLMPs) are the future of clinical laboratories. Although everyday practice shows significant differences among countries, especially during residency training, most of them face the same challenges. Besides promoting scientific, professional and clinical aspects of laboratory medicine in Europe, the European Federation of Clinical Chemistry and Laboratory Medicine (EFLM) should take into consideration YLMPs' concerns and interests to help them achieve excellence. The aim of this survey was to assess the opinion and expectations of YLMPs about their involvement in the activities of EFLM. Methods: An online survey was distributed to YLMPs in Europe through different channels. The questionnaire consisted of 21 items grouped into five sections: demographic questions, opinion about the current status of YLMPs within EFLM, YLMPs network, suggestions and opportunities, and scientific training and exchange. Where appropriate, responses from residents and specialists were compared. Results: A total of 329 valid responses were obtained from 53 different countries. Countries with the highest number of participants were Spain, Turkey, Croatia and Romania. A significant percentage would like to know more about EFLM and their activities (86%) and wish EFLM promoted networking and scientific exchanges (95%), for instance by means of a European YLMPs network (93%). EFLMLabX project was widely unknown (75%). Conclusions: YLMPs demand better connection to share concerns about daily healthcare duties, to keep updated and to advance professionally. EFLM needs to improve their advertising through national societies to increase YLMPs' participation. In addition to international meetings and congresses, respondents have emphasized that workshops and other small group activities would significantly help promote laboratory medicine practice in Europe.

A CROSS-SECTIONAL STUDY OF THE ASSOCIATION BETWEEN THE 1-HOUR ORAL GLUCOSE TOLERANCE TEST AND THE METABOLIC SYNDROME IN A HIGH-RISK SAMPLE WITH IMPAIRED FASTING GLUCOSE.

Objective: The aim of this study was to evaluate the association between the 1-hour oral glucose tolerance test (OGTT) (≥155 mg/dL) and metabolic syndrome (MS) in a sample with previous impaired fasting glucose (IFG). Methods: Three hundred and twenty four Peruvian subjects with a history of IFG ≥100 mg/dL were selected for a cross-sectional study. They underwent a 75 g OGTT and were assigned to different groups according to the result. We evaluated the association between 1-hour OGTT and MS. Results: The mean age was 56.5 ± 12.6 years and 191 (61.5%) were female. During the OGTT, we found 28 (8.6%) subjects with diabetes, 74 (22.8%) with IGT, and 222 (68.5%) with a normal glucose tolerance test with a 2-hour glucose <140 mg/dL (NGT). In the NGT group, 124 (38.3%) had 1-hour glucose levels <155 mg/dL, while 98 (30.2%) had 1-hour glucose levels ≥155 mg/dL. Evaluating the association between the 1-hour value in the OGTT and MS, we found that subjects with a 1-hour glucose ≥155 mg/dL were more than twice as likely to have MS as those with a 1-hour glucose <155 mg/dL (odds ratio = 2.64, 95% confidence interval: 1.52 to 4.57). In addition, body mass index, fasting glycemia, triglycerides, and waist circumferences were significantly higher in subjects with 1-hour glucose levels ≥155 mg/dL compared to those with 1-hour glucose levels <155 mg/dL (P<.05). Conclusion: Among subjects with IFG, performing an OGTT was helpful to identify subjects with 1-hour glucose levels ≥155 mg/dL and NGT who were significantly more likely to have MS and a worse cardiometabolic risk profile. Abbreviations: AST = aspartate aminotransferase; BMI = body mass index; CI = confidence interval; IFG = impaired fasting glucose; IGT = impaired glucose tolerance; LDL = low-density lipoprotein; MS = metabolic syndrome; NGT = normal glucose tolerance; OGTT = oral glucose tolerance test; OR = odds ratio; T2DM = type 2 diabetes; TG = triglycerides.

Pneumonia versus graft dysfunction as the cause of acute respiratory failure after lung transplant: a 4-year multicentre prospective study in 153 adults requiring intensive care admission.

We aimed to assess the main causes of intensive care unit (ICU) readmissions in lung transplant adults and to identify independent predictors of ICU mortality (primary end-point).This Spanish five-centre prospective cohort study enrolled all lung transplant adults with ICU readmissions after post-transplant ICU discharge between 2012 and 2016. Patients were followed until hospital discharge or death.153 lung transplant recipients presented 174 ICU readmissions at a median (interquartile range) of 6 (2-25) months post-transplant. Chronic lung allograft dysfunction was reported in 39 (25.5%) recipients, 13 of whom (all exitus) had restrictive allograft syndrome (RAS). Acute respiratory failure (ARF) (110 (71.9%)) was the main condition requiring ICU readmission. Graft rejection (six (5.4%) acute) caused only 12 (10.8%) readmissions whereas pneumonia (56 (36.6%)) was the main cause (50 admitted for ARF and six for shock), with Pseudomonas aeruginosa (50% multidrug resistant) being the predominant pathogen. 55 (35.9%) and 69 (45.1%) recipients died in the ICU and the hospital, respectively. Bronchiolitis obliterans syndrome (BOS) stage 2 (adjusted OR (aOR) 7.2 (95% CI 1.0-65.7)), BOS stage 3 (aOR 13.7 (95% CI 2.5-95.3)), RAS (aOR >50) and pneumonia at ICU readmission (aOR 2.5 (95% CI 1.0-7.1)) were identified in multivariate analyses as independent predictors of ICU mortality. Only eight (5.2%) patients had positive donor-specific antibodies prior to ICU readmission and this variable did not affect the model.ARF was the main condition requiring ICU readmission in lung transplant recipients and was associated with high mortality. Pneumonia was the main cause of death and was also an independent predictor. RAS should receive palliative care rather than ICU admission.

Sequence optimization and glycosylation of vasoinhibin: Pitfalls of recombinant production.

Vasoinhibin belongs to a family of proteins with antiangiogenic properties derived by proteolytic cleavage from the hormone prolactin (PRL). Vasoinhibin isoforms range from the first 79 to the first 159 residues of PRL. In an attempt to increase the yield of recombinant vasoinhibin and avoid incorrect intra- and inter-disulfide bond formation, the cDNA sequence comprising the first 123 amino acids of human PRL, in which cysteine 58 was or not mutated to serine, was codon-optimized. The optimized constructs achieved a 6-fold increase in mRNA expression but showed no change in protein production and reduced protein secretion when expressed in human embryo kidney (HEK293T/17) cells. Limited vasoinhibin levels associated with the activation of the unfolded protein response (UPR) and endoplasmic reticulum-associated degradation (ERAD) as revealed by the upregulation of UPR (Bip, Xbp-1, and Chop) and ERAD (Hrd1, Os9, and Sel1l) target genes. Mutation to serine introduced a new N-glycosylation site and associated with increased glycosylation and release of glycosylated vasoinhibin isoforms having reduced antiangiogenic properties. We conclude that overexpression and excessive glycosylation lead to protein degradation and reduced bioactivity, respectively, negatively affecting the production of recombinant vasoinhibin in mammalian cells.

The effect of diet on the physical and mental development of children: views of parents and teachers in four European countries.

Although the impact of diet on physical health is an important public health issue, less attention has been devoted to the relationship between nutrition and children's mental development. The views of parents and teachers about the extent to which diet affects physical and mental development of children were compared in four European countries. An online questionnaire (developed in English and translated) was circulated through a market research agency. Participants were parents or teachers of children aged 4-10 years without learning or behavioural issues. Questionnaires were returned by 1606 parents (401 in England, Germany and Hungary; 403 in Spain) and 403 teachers (100 in each country, except for 103 in Hungary). Teachers were older than parents (35·3 % v. 18·3 % over 45 years; P<0·001) and less likely to smoke (15·9 % v. 26·3 %, P<0·001). There was no difference between the proportions of parents and teachers who felt that a child's physical development depended very much/extremely (v. moderately/slightly/not at all) on diet (overall 79·8 %). Lower proportions of both groups thought that mental development was very much/extremely influenced by diet (67·4 %). In the regression modelling, believing that physical and mental performance was greatly influenced by diet was significantly and positively associated with living in Hungary, scoring higher on a measure of General Health Interest and (parents only) level of education attained. Differences existed among countries in most views. Lower levels of awareness of the importance of diet for brain development and cognition (compared with physical health outcomes) indicate the potential for educating consumers, especially parents with lower educational attainment.