Chemogenetics with PSAM4-GlyR decreases excitability and epileptiform activity in epileptic hippocampus.

Despite the availability of new drugs on the clinics in recent years, drug-resistant epilepsy remains an unresolved challenge for healthcare, and one-third of epilepsy patients remain refractory to anti-seizure medications. Gene therapy in experimental models has emerged as effective treatment targeting specific neuronal populations in the epileptogenic focus. When combined with an external chemical activator using chemogenetics, it also becomes an "on-demand" treatment. Here, we evaluate a targeted and specific chemogenetic therapy, the PSAM/PSEM system, which holds promise as a potential candidate for clinical application in treating drug-resistant epilepsy. We show that the inert ligand uPSEM817, which selectively activates the chloride-permeable channel PSAM4-GlyR, effectively reduces the number of depolarization-induced action potentials in vitro. This effect is likely due to the shunting of depolarizing currents, as evidenced by decreased membrane resistance in these cells. In organotypic slices, uPSEM817 decreased the number of bursts and peak amplitude of events of spontaneous epileptiform activity. Although administration of uPSEM817 in vivo did not significantly alter electrographic seizures in a male mouse model of temporal lobe epilepsy, it did demonstrate a strong trend toward reducing the frequency of interictal epileptiform discharges. These findings indicate that PSAM4-GlyR-based chemogenetics holds potential as an anti-seizure strategy, although further refinement is necessary to enhance its efficacy.

Alteration of gene expression and protein solubility of the PI 5-phosphatase SHIP2 are correlated with Alzheimer's disease pathology progression.

A recent large genome-wide association study has identified EGFR (encoding the epidermal growth factor EGFR) as a new genetic risk factor for late-onset AD. SHIP2, encoded by INPPL1, is taking part in the signalling and interactome of several growth factor receptors, such as the EGFR. While INPPL1 has been identified as one of the most significant genes whose RNA expression correlates with cognitive decline, the potential alteration of SHIP2 expression and localization during the progression of AD remains largely unknown. Here we report that gene expression of both EGFR and INPPL1 was upregulated in AD brains. SHIP2 immunoreactivity was predominantly detected in plaque-associated astrocytes and dystrophic neurites and its increase was correlated with amyloid load in the brain of human AD and of 5xFAD transgenic mouse model of AD. While mRNA of INPPL1 was increased in AD, SHIP2 protein undergoes a significant solubility change being depleted from the soluble fraction of AD brain homogenates and co-enriched with EGFR in the insoluble fraction. Using FRET-based flow cytometry biosensor assay for tau-tau interaction, overexpression of SHIP2 significantly increased the FRET signal while siRNA-mediated downexpression of SHIP2 significantly decreased FRET signal. Genetic association analyses suggest that some variants in INPPL1 locus are associated with the level of CSF pTau. Our data support the hypothesis that SHIP2 is an intermediate key player of EGFR and AD pathology linking amyloid and tau pathologies in human AD.

An update on susceptibility-weighted imaging in brain gliomas.

Susceptibility-weighted imaging (SWI) has become a standard component of most brain MRI protocols. While traditionally used for detecting and characterising brain hemorrhages typically associated with stroke or trauma, SWI has also shown promising results in glioma assessment. Numerous studies have highlighted SWI's role in differentiating gliomas from other brain lesions, such as primary central nervous system lymphomas or metastases. Additionally, SWI aids radiologists in non-invasively grading gliomas and predicting their phenotypic profiles. Various researchers have suggested incorporating SWI as an adjunct sequence for predicting treatment response and for post-treatment monitoring. A significant focus of these studies is on the detection of intratumoural susceptibility signals (ITSSs) in gliomas, which are indicative of microhemorrhages and vessels within the tumour. The quantity, distribution, and characteristics of these ITSSs can provide radiologists with more precise information for evaluating and characterising gliomas. Furthermore, the potential benefits and added value of performing SWI after the administration of gadolinium-based contrast agents (GBCAs) have been explored. This review offers a comprehensive, educational, and practical overview of the potential applications and future directions of SWI in the context of glioma assessment. CLINICAL RELEVANCE STATEMENT: SWI has proven effective in evaluating gliomas, especially through assessing intratumoural susceptibility signal changes, and is becoming a promising, easily integrated tool in MRI protocols for both pre- and post-treatment assessments. KEY POINTS: • Susceptibility-weighted imaging is the most sensitive sequence for detecting blood and calcium inside brain lesions. • This sequence, acquired with and without gadolinium, helps with glioma diagnosis, characterisation, and grading through the detection of intratumoural susceptibility signals. • There are ongoing challenges that must be faced to clarify the role of susceptibility-weighted imaging for glioma assessment.

Impact of hydrophobic and hydrophilic surface properties on Pseudomonas aeruginosa adhesion in materials used in mineral water wells.

Microbiologically contaminated water is a significant source of infections in humans and animals, with Pseudomonas aeruginosa (PSA) being particularly concerning due to its ability to thrive in water environments and its resistance to many disinfectants. Therefore, this study investigates the adhesion potential of PSA strains on various materials used in mineral water extraction wells, focusing on hydrophobic and hydrophilic properties. Mineral water samples were collected from three wells (P-01, P-07, and P-08) within the Guarani Aquifer System and Fractured Aquifer System (SAF) in Brazil. The physicochemical properties of the water, including concentrations of Sr (strontium), Fe (iron), Si (silicon), SO42- (sulfate ions), Cl- (chloride ions), and ORP (oxidation-reduction potential), were analyzed. Results indicated higher PSA adhesion on hydrophobic materials, particularly high-density polyethylene (HDPE) and geomechanically plasticized polyvinyl chloride (PVC). Multiple correlation analyses revealed positive correlations between PSA adhesion on hydrophilic materials and Sr, Fe, Si, SO42-, and Cl- concentrations. Conversely, ORP negatively correlated with bacterial adhesion on PVC surfaces, suggesting higher ORP values reduced PSA attachment. These findings highlight the importance of water composition and material properties in influencing bacterial adhesion and potential biofilm formation in mineral water extraction systems.

Impact of enhancement filters of a CMOS system on halo artifact expression at the bone-to-implant interface.

OBJECTIVE: To assess the impact of enhancement filters on the formation of halo artifacts in radiographs of dental implants obtained with a complementary metal oxide semiconductor (CMOS) system. METHODS: Digital radiographs of dental implants placed in dry human mandibles were processed with the Noise Reduction smoothing filter, as well as the Sharpen 1, Sharpen 4, and Sharpen UM high-pass filters available in the CLINIVIEW™ software (Instrumentarium Dental, Tuusula, Finland). Subjective analysis involved evaluating the left, right, and apical surfaces of each implant for the presence of much, few, or no halo. The objective analysis involved measurement of the halo area using the Trainable Weka Segmentation plugin (ImageJ, National Institutes of Health, Bethesda, MD, USA). Data were analyzed using Friedman's test (subjective analysis) and ANOVA (objective analysis) (α = 5%). RESULTS: In the subjective evaluation, the Sharpen 4 filter produced more radiographs with much halo present, and in the objective evaluation, a bigger halo area when compared to the original images and the Noise Reduction filter for all surfaces (p < 0.05). CONCLUSIONS: When evaluating dental implants, priority should be given to original images and those enhanced with smoothing filters since they exhibit fewer halo artifacts. CLINICAL RELEVANCE: Post-processing tools, such as enhancement filters, may improve the image quality and assist some diagnostic tasks. However, little is known regarding the impact of enhancement filters in halo formation on CMOS systems, which have been increasingly used in dental offices.

Differences in DNA Methylation in Genes Involved in Vitamin D Metabolism Are Related to Insulin Requirement in Pregnant Women with Gestational Diabetes Mellitus.

In a previous study performed by our group, pregnant women with Gestational Diabetes (GDM) showed higher vitamin D (VitD) levels in the last trimester, particularly in those requiring insulin. This phenomenon was not linked to factors like season or supplementation. This study aimed to investigate if insulin treatment in GDM affects DNA methylation in VitD metabolism genes. Thirty-two pregnant women were selected, half of whom had GDM, and were divided into insulin-treated and lifestyle groups. The DNA methylation levels in CpGs from 47 VitD metabolism-related genes were analyzed at the diagnostic visit (24-28 weeks) and before delivery. At week 36-38 of pregnancy, twenty-six CpG sites were differentially methylated (DMPs) in the insulin-treated group compared with the control group and the lifestyle group. Twenty-two of these DMPs were not different at the diagnostic visit. Six CpGs (cg18276810 (CTNNB1), cg03919554 (FGFR3), cg03984919 (NCOA1), cg19218509 (ASIP), cg09922639 (SMAD3), and cg25356935 (PDZD3)) showed significant correlations with VitD levels, not only before childbirth, but also in the postpartum period and at one year later. This suggests that insulin treatment in GDM could influence DNA methylation in genes involved in vitamin D metabolism, affecting VitD levels during and after pregnancy. Further research is warranted to elucidate these findings' clinical implications.

Environmental Exposure to Persistent Organic Pollutants and Its Association with Endometriosis Risk: Implications in the Epithelial-Mesenchymal Transition Process.

We aimed to explore the relationship of adipose tissue concentrations of some persistent organic pollutants (POPs) with the risk of endometriosis and the endometriotic tissue expression profile of genes related to the endometriosis-related epithelial-mesenchymal transition (EMT) process. This case-control study enrolled 109 women (34 cases and 75 controls) between January 2018 and March 2020. Adipose tissue samples and endometriotic tissues were intraoperatively collected to determine concentrations of nine POPs and the gene expression profiles of 36 EMT-related genes, respectively. Associations of POPs with endometriosis risk were explored with multivariate logistic regression, while the relationship between exposure and gene expression profiles was assessed through Spearman correlation or Mann-Whitney U tests. After adjustment, increased endometriosis risk was associated with p,p'-DDT, PCB-180, and ΣPCBs. POP exposure was also associated with reduced gene expression levels of the CLDN7 epithelial marker and increased levels of the ITGB2 mesenchymal marker and a variety of EMT promoters (HMGA1, HOXA10, FOXM1, DKK1, CCR1, TNFRSF1B, RRM2, ANG, ANGPT1, and ESR1). Our findings indicate that exposure to POPs may increase the risk of endometriosis and might have a role in the endometriosis-related EMT development, contributing to the disease onset and progression. Further studies are warranted to corroborate these findings.

Comparative analysis of the physical, chemical, and microbiological properties of Ti-6Al-4V disks produced by different methods and subjected to surface treatments.

STATEMENT OF PROBLEM: To improve the osseointegration of dental implants and reduce microbiological growth, different micro- and nanoscale surface topographies can be used. PURPOSE: The purpose of this in vitro study was to evaluate the influence of Ti-6Al-4V with 4 surfaces, machined (DU), machined+hydroxyapatite (DUHAp), machined+acid-alkali treatment (DUAA), and additive manufacturing (DMA), on the physical, chemical, and microbiological properties. MATERIAL AND METHODS: The topography of Ti-6Al-4V disks with the 4 surfaces was evaluated by scanning electron microscopy (SEM), the chemical composition by energy dispersive X-ray spectroscopy (EDS), and the crystalline structure by X-ray diffraction (XRD). Physical and chemical properties were analyzed by using wettability and surface free energy, roughness, and microbial adhesion against Staphylococcus aureus by colony forming units (CFU). One-way ANOVA analysis of variance and the Tukey multiple comparisons test were applied to evaluate the data, except CFU, which was submitted to the Kruskal-Wallis nonparametric test (α=.05). RESULTS: DU photomicrographs showed a topography characteristic of a polished machined surface, DUHAp and DUAA exhibited patterns corresponding to the surface modifications performed, and in DMA the presence of partially fused spherical particles was observed. The EDS identified chemical elements inherent in the Ti-6Al-4V, and the DUHAp and DUAA disks also had the ions from the treatments applied. XRD patterns revealed similarities between DU and DMA, as well as characteristic peaks of hydroxyapatite (HA) in the DUHAp disk and the DUAA. Compared with DU and DMA the DUHAp and DUAA groups showed hydrophilic behavior with smaller contact angles and higher surface free energy (P<.05). DMA showed a higher mean value of roughness, different from the others (P<.05), and a higher CFU for S. aureus (P=.006). CONCLUSIONS: DUHAp and DUAA showed similar behaviors regarding wettability, surface free energy, and bacterial adhesion. Among the untreated groups, DMA exhibited higher roughness, bacterial adhesion, and lower wettability and surface free energy.

Computed diffusion-weighted imaging in patients with transient neurovascular symptoms with and without ischemic infarction.

PURPOSE: Detection of ischemic lesions in patients with transient neurovascular symptoms is relevant for the estimation of the risk of a subsequent stroke and etiological classification. To improve detection rates, different technical approaches have been used, such as diffusion-weighted imaging (DWI) with high b-values or higher magnetic field strength. Here, we sought to investigate the value of computed DWI (cDWI) with high b-values in these patients. METHODS: From an MRI report database we identified patients with transient neurovascular symptoms who underwent repeated MRI including DWI. cDWI was calculated with a monoexponential model with high b-values (2000, 3000, and 4000 s/mm2) and compared to the routinely used standard DWI with regard to presence of ischemic lesions and lesion detectability. RESULT: Overall 33 patients with transient neurovascular symptoms (71 [IQR 57-83.5] years; 21 [63.6%] male) were included. On DWI, acute ischemic lesions were observed in 22 (78.6%). Acute ischemic lesions were observed in 17 (51.5%) patients on initial DWI, and in 26 (78.8%) patients on follow-up DWI. Lesion detectability was rated significantly better on cDWI at 2000s/mm2 compared to standard DWI. In 2 (9.1%) patients, cDWI at 2000s/mm2 revealed an acute ischemic lesion proven on follow-up standard DWI which was not detected with certainty on the initial standard DWI. CONCLUSION: cDWI might be a valuable addition to routinely acquired standard DWI in patients with transient neurovascular symptoms since its use might result in improved ischemic lesion detection. A b-value of 2000s/mm2 seems most promising for clinical practice.

The Benefits of an Integral HAMMAM Experience Combining Hydrotherapy and Swedish Massage on Pain, Subjective Well-Being and Quality of Life in Women with Endometriosis-Related Chronic Pelvic Pain: A Randomized Controlled Trial.

Background and Objectives: To evaluate the effectiveness of an integral HAMMAM experience, a 4-week therapeutic program that combined hydrotherapy and Swedish massage, applied in a multisensorial immersive environment, on pain, well-being and quality of life (QoL) in women with endometriosis-related chronic pelvic pain that is unresponsive to conventional treatment. Materials and Methods: This randomized controlled trial included 44 women with endometriosis. They were randomly allocated to either the 'HAMMAM' group (n = 21) or to a control group (n = 23). The primary outcome, pain intensity, was evaluated using numeric rating scales (NRSs). The secondary outcomes were pain interference, pain-related catastrophic thoughts, pressure pain thresholds (PPTs), subjective well-being, functional capacity and QoL, which were evaluated using the brief pain inventory (BPI), the pain catastrophizing scale (PCS), algometry, the subjective well-being scale-20 (EBS-20), the Patient-Reported Outcomes Measurement Information System-29 (PROMIS-29) and the Endometriosis Health Profile-30 Questionnaire (EHP-30), respectively. The primary and secondary outcomes were measured at the baseline and after the intervention. The statistical (between-group analyses of covariance) and clinical effects were analyzed by the intention to treat. Results: The adherence rate was 100.0% and the mean (± standard deviation) satisfaction was 9.71 ± 0.46 out of 10. No remarkable health problems were reported during the trial. The 'HAMMAM' intervention improved dysmenorrhea and dyspareunia after the intervention with large and moderate effect sizes, respectively. Improvements in pain interference during sleep and PPTs in the pelvic region were also observed in women allocated to the 'HAMMAM' group. No effects were observed in catastrophizing thoughts, well-being nor QoL, except for the sleep subscale. Conclusions: A 4-week program of an integral 'HAMMAM' experience combining hydrotherapy and massage in a multisensorial immersive environment is a feasible and effective intervention to alleviate pain during menstruation and sexual intercourse as well as pain interference with sleep in women with endometriosis.

From cells to form: A roadmap to study shape emergence in vivo.

Organogenesis arises from the collective arrangement of cells into progressively 3D-shaped tissue. The acquisition of a correctly shaped organ is then the result of a complex interplay between molecular cues, responsible for differentiation and patterning, and the mechanical properties of the system, which generate the necessary forces that drive correct shape emergence. Nowadays, technological advances in the fields of microscopy, molecular biology, and computer science are making it possible to see and record such complex interactions in incredible, unforeseen detail within the global context of the developing embryo. A quantitative and interdisciplinary perspective of developmental biology becomes then necessary for a comprehensive understanding of morphogenesis. Here, we provide a roadmap to quantify the events that lead to morphogenesis from imaging to image analysis, quantification, and modeling, focusing on the discrete cellular and tissue shape changes, as well as their mechanical properties.

Latiglutenase Protects the Mucosa and Attenuates Symptom Severity in Patients With Celiac Disease Exposed to a Gluten Challenge.

BACKGROUND & AIMS: Gluten ingestion in patients with celiac disease can lead to gastrointestinal symptoms and small intestinal mucosal injury. METHODS: This gluten challenge phase 2 trial was double blind and placebo controlled, and it assessed the efficacy and safety of a 1200-mg dose of IMGX003 in patients with celiac disease exposed to 2 g of gluten per day for 6 weeks. The change in the ratio of villus height to crypt depth was the primary endpoint. Secondary endpoints included density of intraepithelial lymphocytes and symptom severity. These endpoints were evaluated by analysis of covariance. Additional endpoints included serology and gluten-immunogenic peptides in urine. RESULTS: Fifty patients were randomized, and 43 patients completed the study (IMGX003, n = 21; placebo, n = 22). The mean change in the ratio of villus height to crypt depth (primary endpoint) for IMGX003 vs placebo was -0.04 vs -0.35 (P = .057). The mean change in the density of intraepithelial lymphocytes (secondary endpoint) for IMGX003 vs placebo was 9.8 vs 24.8 cells/mm epithelium (P = .018). The mean change (worsening) in symptom severity in relative units (secondary endpoint) for IMGX003 vs placebo was 0.22 vs 1.63 (abdominal pain, P = .231), 0.96 vs 3.29 (bloating, P = .204), and 0.02 vs 3.20 (tiredness, P = .113). The 3 × 2-week trend line significance values for these symptoms, respectively, were P = .014, .030, and .002. CONCLUSIONS: IMGX003 reduced gluten-induced intestinal mucosal damage and symptom severity. (ClinicalTrials.gov, Number: NCT03585478).

Collagen/κ-Carrageenan-Based Scaffolds as Biomimetic Constructs for In Vitro Bone Mineralization Studies.

Tissue engineering offers attractive strategies to develop three-dimensional scaffolds mimicking the complex hierarchical structure of the native bone. The bone is formed by cells incorporated in a molecularly organized extracellular matrix made of an inorganic phase, called biological apatite, and an organic phase mainly made of collagen and noncollagenous macromolecules. Although many strategies have been developed to replicate the complexity of bone at the nanoscale in vitro, a critical challenge has been to control the orchestrated process of mineralization promoted by bone cells in vivo and replicate the anatomical and biological properties of native bone. In this study, we used type I collagen to fabricate mineralized scaffolds mimicking the microenvironment of the native bone. The sulfated polysaccharide κ-carrageenan was added to the scaffolds to fulfill the role of noncollagenous macromolecules in the organization and mineralization of the bone matrix and cell adhesion. Scanning electron microscopy images of the surface of the collagen/κ-carrageenan scaffolds showed the presence of a dense and uniform network of intertwined fibrils, while images of the scaffolds' lateral sides showed the presence of collagen fibrils with a parallel alignment, which is characteristic of dense connective tissues. MC3T3-E1 osteoblasts were cultured in the collagen scaffolds and were viable after up to 7 days of culture, both in the absence and in the presence of κ-carrageenan. The presence of κ-carrageenan in the collagen scaffolds stimulated the maturation of the cells to a mineralizing phenotype, as suggested by the increased expression of key genes related to bone mineralization, including alkaline phosphatase (Alp), bone sialoprotein (Bsp), osteocalcin (Oc), and osteopontin (Opn), as well as the ability to mineralize the extracellular matrix after 14 and 21 days of culture. Taken together, the results described in this study shed light on the potential use of collagen/κ-carrageenan scaffolds to study the role of the structural organization of bone-mimetic synthetic matrices in cell function.

Annexin A5 stabilizes matrix vesicle-biomimetic lipid membranes: unravelling a new role of annexins in calcification.

Matrix vesicles are a special class of extracellular vesicles thought to actively contribute to both physiologic and pathologic mineralization. Proteomic studies have shown that matrix vesicles possess high amounts of annexin A5, suggesting that the protein might have multiple roles at the sites of calcification. Currently, Annexin A5 is thought to promote the nucleation of apatitic minerals close to the inner leaflet of the matrix vesicles' membrane enriched in phosphatidylserine and Ca2+. Herein, we aimed at unravelling a possible additional role of annexin A5 by investigating the ability of annexin A5 to adsorb on matrix-vesicle biomimetic liposomes and Langmuir monolayers made of dipalmitoylphosphatidylserine (DPPS) and dipalmitoylphosphatidylcholine (DPPC) in the absence and in the presence of Ca2+. Differential scanning calorimetry and dynamic light scattering measurements showed that Ca2+ at concentrations in the 0.5-2.0 mM range induced the aggregation of liposomes probably due to the formation of DPPS-enriched domains. However, annexin A5 avoided the aggregation of liposomes at Ca2+ concentrations lower than 1.0 mM. Surface pressure versus surface area isotherms showed that the adsorption of annexin A5 on the monolayers made of a mixture of DPPC and DPPS led to a reduction in the area of excess compared to the theoretical values, which confirmed that the protein favored attractive interactions among the membrane lipids. The stabilization of the lipid membranes by annexin A5 was also validated by recording the changes with time of the surface pressure. Finally, fluorescence microscopy images of lipid monolayers revealed the formation of spherical lipid-condensed domains that became unshaped and larger in the presence of annexin A5. Our data support the model that annexin A5 in matrix vesicles is recruited at the membrane sites enriched in phosphatidylserine and Ca2+ not only to contribute to the intraluminal mineral formation but also to stabilize the vesicles' membrane and prevent its premature rupture.

Multiple sclerosis imaging in clinical practice: a European-wide survey of 428 centers and conclusions by the ESNR Working Group.

OBJECTIVES: To evaluate compliance with the available recommendations, we assessed the current clinical practice of imaging in the evaluation of multiple sclerosis (MS). METHODS: An online questionnaire was emailed to all members and affiliates. Information was gathered on applied MR imaging protocols, gadolinium-based contrast agents (GBCA) use and image analysis. We compared the survey results with the Magnetic Resonance Imaging in MS (MAGNIMS) recommendations considered as the reference standard. RESULTS: A total of 428 entries were received from 44 countries. Of these, 82% of responders were neuroradiologists. 55% performed more than ten scans per week for MS imaging. The systematic use of 3 T is rare (18%). Over 90% follow specific protocol recommendations with 3D FLAIR, T2-weighted and DWI being the most frequently used sequences. Over 50% use SWI at initial diagnosis and 3D gradient-echo T1-weighted imaging is the most used MRI sequence for pre- and post-contrast imaging. Mismatches with recommendations were identified including the use of only one sagittal T2-weighted sequence for spinal cord imaging, the systematic use of GBCA at follow-up (over 30% of institutions), a delay time shorter than 5 min after GBCA administration (25%) and an inadequate follow-up duration in pediatric acute disseminated encephalomyelitis (80%). There is scarce use of automated software to compare images or to assess atrophy (13% and 7%). The proportions do not differ significantly between academic and non-academic institutions. CONCLUSIONS: While current practice in MS imaging is rather homogeneous across Europe, our survey suggests that recommendations are only partially followed. CLINICAL RELEVANCE STATEMENT: Hurdles were identified, mainly in the areas of GBCA use, spinal cord imaging, underuse of specific MRI sequences and monitoring strategies. This work will help radiologists to identify the mismatches between their own practices and the recommendations and act upon them. KEY POINTS: • While current practice in MS imaging is rather homogeneous across Europe, our survey suggests that available recommendations are only partially followed. • Several hurdles have been identified through the survey that mainly lies in the areas of GBCA use, spinal cord imaging, underuse of specific MRI sequences and monitoring strategies.

NPP1 and TNAP hydrolyze ATP synergistically during biomineralization.

Matrix vesicles (MVs) are a special class of extracellular vesicles released by mineralizing cells during bone and tooth mineralization that initiate the precipitation of apatitic minerals by regulating the extracellular ratio between inorganic phosphate (Pi), a calcification promoter, and pyrophosphate (PPi), a calcification inhibitor. The Pi/PPi ratio is thought to be controlled by two ecto-phosphatases present on the outer leaflet of the MVs' membrane: ectonucleotide pyrophosphatase/phosphodiesterase 1 (NPP1) that produces PPi as well as Pi from ATP and tissue-nonspecific alkaline phosphatase (TNAP) that hydrolyzes both ATP and PPi to generate Pi. However, if and how these enzymes act in concert in MVs are still unclear. Herein, we investigated the role of NPP1 and TNAP in ATP hydrolysis during MV-mediated biomineralization using proteoliposomes as a biomimetic model for MVs. Proteoliposomes composed by 1,2-dipalmitoylphosphatidylcholine (DPPC) and harboring NPP1 alone, TNAP alone, or both together at different molar ratios (1:1, 10:1, and 1:10) were fabricated. After 48 h of incubation with ATP, TNAP-containing proteoliposomes consumed more ATP than NPP1-containing vesicles (270 and 210 nmol, respectively). Both types of vesicles comparatively formed ADP (205 and 201 nmol, respectively), while NPP1-containing vesicles hydrolyzed AMP less efficiently than TNAP-containing proteoliposomes (10 and 25 nmol, respectively). In vitro mineralization assays showed that in the presence of ATP, TNAP-harboring proteoliposomes mineralized through a sigmoidal single-step process, while NPP1-harboring vesicles displayed a two-step mineralization process. ATR-FTIR analyses showed that the minerals produced by TNAP-harboring proteoliposomes were structurally more similar to hydroxyapatite than those produced by NPP1-harboring vesicles. Our results with proteoliposomes indicate that the pyrophosphohydrolase function of NPP1 and the phosphohydrolase activity of TNAP act synergistically to produce a Pi/PPi ratio conducive to mineralization and the synergism is maximal when the two enzymes are present at equimolar concentrations. The significance of these findings for hypophosphatasia is discussed.

Post-Intensive Care Unit Multidisciplinary Approach in Patients with Severe Bilateral SARS-CoV-2 Pneumonia.

Background: Short and long-term sequelae after admission to the intensive care unit (ICU) for coronavirus disease 2019 (COVID-19) are to be expected, which makes multidisciplinary care key in the support of physical and cognitive recovery. Objective: To describe, from a multidisciplinary perspective, the sequelae one month after hospital discharge among patients who required ICU admission for severe COVID-19 pneumonia. Design: Prospective cohort study. Environment: Multidisciplinary outpatient clinic. Population: Patients with severe COVID-19 pneumonia, post- ICU admission. Methods: A total of 104 patients completed the study in the multidisciplinary outpatient clinic. The tests performed included spirometry, measurement of respiratory muscle pressure, loss of body cell mass (BCM) and BCM index (BCMI), general joint and muscular mobility, the short physical performance battery (SPPB or Guralnik test), grip strength with hand dynamometer, the six-minute walk test (6-MWT), the functional assessment of chronic illness therapy-fatigue scale (FACIT-F), the European quality of life-5 dimensions (EQ-5D), the Barthel index and the Montreal cognitive assessment test (MoCA). While rehabilitation was not necessary for 23 patients, 38 patients attended group rehabilitation sessions and other 43 patients received home rehabilitation. Endpoints: The main sequelae detected in patients were fatigue (75.96%), dyspnoea (64.42%) and oxygen therapy on discharge (37.5%). The MoCA showed a mean score compatible with mild cognitive decline. The main impairment of joint mobility was limited shoulder (11.54%) and shoulder girdle (2.88%) mobility; whereas for muscle mobility, lower limb limitations (16.35%) were the main dysfunction. Distal neuropathy was present in 23.08% of patients, most frequently located in lower limbs (15.38%). Finally, 50% of patients reported moderate limitation in the EQ-5D, with a mean score of 60.62 points (SD 20.15) in perceived quality of life. Conclusions: Our findings support the need for a multidisciplinary and comprehensive evaluation of patients after ICU admission for COVID-19 because of the wide range of sequelae, which also mean that these patients need a long-term follow-up. Impact on clinical rehabilitation: This study provides data supporting the key role of rehabilitation during the follow-up of severe patients, thus facilitating their reintegration in society and a suitable adaptation to daily living.

Report of a case of RAVEN, hair heterochromia and autism in the setting of FGFR2 mutation.

A newborn presented with extensive rounded and velvety epidermal nevus (RAVEN) with a genetic study of the cutaneous lesions revealing a heterozygous mutation in FGFR2 (p.Cys382Arg). By 2 years of age, the patient developed hair heterochromia and autism spectrum disorder. Although RAVEN was initially associated with fibroblast growth factor 3 (FGFR3) mutations, three cases of RAVEN have been identified with mutations in FGFR2 (p.Ser252Trp) and one case of linear keratinocytic epidermal nevi has been identified with the same mutation as the mutation identified in our patient. This strongly supports the pathogenic role of these mutations.

Differences in self-control, self-efficacy and depressive symptoms between active and inactive middle-aged and older adults after 1 year of COVID restrictions.

OBJECTIVES: The psychological impact of the prolonged lockdown measures in the UK as a response to the COVID-19 pandemic is unclear. Our aim was to determine if there are significant differences in self-control, self-efficacy, depressive symptoms and leisure motivation between UK older adults with differing levels of physical activity, and which of these variables can be used to predict activity level after 1 year of lockdown restrictions. METHODS: 521 adults aged 50-92 years completed an online survey consisting of several validated measures relating to physical activity, self-control, self-efficacy, depressive symptoms, and leisure motivation. Participant's responses were grouped into active (≥150minutes activity per week) and inactive (<150minutes activity per week). Data was analysed using ANOVA, Pearson's Correlation and Multiple Regression (forward stepwise). RESULTS: We found significant differences in self-efficacy, self-control, and depressive symptoms between physically active vs inactive subjects. High levels of self-control and self-efficacy were associated with higher levels of activity and fewer depressive symptoms. Self-control, amotivation, depressive symptoms and self-efficacy were predictors of physical activity level. CONCLUSION: Psychological variables including self-control, self-efficacy, depressive symptoms and amotivation can be used to predict physical activity levels in UK middle-aged and older adults following 1 year of Covid restrictions.

Influence of Er:YAG laser irradiation on surface properties of Ti-6Al-4V machined and hydroxyapatite coated.

Surface treatment by laser irradiation can change the topography of titanium; however, little is known about the changes it causes when applied to other coatings. This study aimed to evaluate the influence of Er:YAG laser irradiation on the surface properties of titanium-aluminum-vanadium (Ti-6Al-4V) discs. Four Ti-6Al-4V surfaces were evaluated (n = 10): CON-control, machined without surface treatment; LT-machined + laser treatment; HA-hydroxyapatite coating; and LT-HA-hydroxyapatite coating + laser treatment. For the laser treatment, an Er:YAG laser with a wavelength of 2940 nm, a frequency of 10 Hz, and an energy density of 12.8 J/cm2 was used. The morphology of the coating was investigated by scanning electron microscopy and the surface composition by energy-dispersive X-ray spectroscopy. The influence of laser irradiation treatment on roughness and wettability was also evaluated. The Er:YAG laser promoted a significant reduction in the roughness Sa (p < 0.05) and in the contact angle (p = 0.002) of the LT surface compared to the CON surface. On the LT-HA surface, a significant decrease in roughness was observed only for the Rz parameter (p = 0.015) and an increase in the contact angle (p < 0.001) compared to the HA surface. The use of the Er:YAG laser with the evaluated parameters decreased the surface roughness and improved the wetting capacity of machined without surface treatment. In the group with hydroxyapatite coating, the laser influenced the surface roughness only for the parameter Rz and reduced their wetting capacity.