Developmental outcome of electroencephalographic findings in SYNGAP1 encephalopathy.

SYNGAP1 haploinsufficiency results in a developmental and epileptic encephalopathy (DEE) causing generalized epilepsies accompanied by a spectrum of neurodevelopmental symptoms. Concerning interictal epileptiform discharges (IEDs) in electroencephalograms (EEG), potential biomarkers have been postulated, including changes in background activity, fixation-off sensitivity (FOS) or eye closure sensitivity (ECS). In this study we clinically evaluate a new cohort of 36 SYNGAP1-DEE individuals. Standardized questionnaires were employed to collect clinical, electroencephalographic and genetic data. We investigated electroencephalographic findings, focusing on the cortical distribution of interictal abnormalities and their changes with age. Among the 36 SYNGAP1-DEE cases 18 presented variants in the SYNGAP1 gene that had never been previously reported. The mean age of diagnosis was 8 years and 8 months, ranging from 2 to 17 years, with 55.9% being male. All subjects had global neurodevelopmental/language delay and behavioral abnormalities; 83.3% had moderate to profound intellectual disability (ID), 91.7% displayed autistic traits, 73% experienced sleep disorders and 86.1% suffered from epileptic seizures, mainly eyelid myoclonia with absences (55.3%). A total of 63 VEEGs were revised, observing a worsening of certain EEG findings with increasing age. A disorganized background was observed in all age ranges, yet this was more common among older cases. The main IEDs were bilateral synchronous and asynchronous posterior discharges, accounting for ≥50% in all age ranges. Generalized alterations with maximum amplitude in the anterior region showed as the second most frequent IED (≥15% in all age ranges) and were also more common with increasing age. Finally, diffuse fast activity was much more prevalent in cases with 6 years or older. To the best of our knowledge, this is the first study to analyze EEG features across different age groups, revealing an increase in interictal abnormalities over infancy and adolescence. Our findings suggest that SYNGAP1 haploinsufficiency has complex effects in human brain development, some of which might unravel at different developmental stages. Furthermore, they highlight the potential of baseline EEG to identify candidate biomarkers and the importance of natural history studies to develop specialized therapies and clinical trials.

Analysis of Poly(ethylene terephthalate) degradation kinetics of evolved IsPETase variants using a surface crowding model.

Poly(ethylene terephthalate) (PET) is a major plastic polymer utilized in the single-use and textile industries. The discovery of PET-degrading enzymes (PETases) has led to an increased interest in the biological recycling of PET in addition to mechanical recycling. IsPETase from Ideonella sakaiensis is a candidate catalyst, but little is understood about its structure-function relationships with regards to PET degradation. To understand the effects of mutations on IsPETase productivity, we develop a directed evolution assay to identify mutations beneficial to PET film degradation at 30 °C. IsPETase also displays enzyme concentration-dependent inhibition effects, and surface crowding has been proposed as a causal phenomenon. Based on total internal reflectance fluorescence microscopy and adsorption experiments, IsPETase is likely experiencing crowded conditions on PET films. Molecular dynamics simulations of IsPETase variants reveal a decrease in active site flexibility in free enzymes and reduced probability of productive active site formation in substrate-bound enzymes under crowding. Hence, we develop a surface crowding model to analyze the biochemical effects of three hit mutations (T116P, S238N, S290P) that enhanced ambient temperature activity and/or thermostability. We find that T116P decreases susceptibility to crowding, resulting in higher PET degradation product accumulation despite no change in intrinsic catalytic rate. In conclusion, we show that a macromolecular crowding-based biochemical model can be used to analyze the effects of mutations on properties of PETases and that crowding behavior is a major property to be targeted for enzyme engineering for improved PET degradation.

Flexible active-site loops fine-tune substrate specificity of hyperthermophilic metallo-oxidases.

Hyperthermophilic ('superheat-loving') archaea found in high-temperature environments such as Pyrobaculum aerophilum contain multicopper oxidases (MCOs) with remarkable efficiency for oxidizing cuprous and ferrous ions. In this work, directed evolution was used to expand the substrate specificity of P. aerophilum McoP for organic substrates. Six rounds of error-prone PCR and DNA shuffling followed by high-throughput screening lead to the identification of a hit variant with a 220-fold increased efficiency (kcat/Km) than the wild-type for 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) without compromising its intrinsic activity for metal ions. The analysis of the X-ray crystal structure reveals four proximal mutations close to the T1Cu active site. One of these mutations is within the 23-residues loop that occludes this site, a distinctive feature of prokaryotic MCOs. The increased flexibility of this loop results in an enlarged tunnel and one additional pocket that facilitates bulky substrate-enzyme interactions. These findings underscore the synergy between mutations that modulate the dynamics of the active-site loop enabling enhanced catalytic function. This study highlights the potential of targeting loops close to the T1Cu for engineering improvements suitable for biotechnological applications.

Longitudinal Changes and Associations Between Quantitative Sensory Testing and Psychological Factors in Whiplash-Associated Disorders: A Systematic Review and Meta-Analyses-Based Data Synthesis.

Whiplash-associated disorders (WAD) represent a multifactorial condition often accompanied by altered nociceptive processing and psychological factors. This systematic review on acute and chronic WAD aimed to investigate the relationship between quantitative sensory testing (QST) and psychological factors and quantify whether their trajectories over time follow a similar pattern to disability levels. Eight databases were searched until October 2022. When 2 prospective studies examined the same QST or psychological variable, data synthesis was performed with random-effects meta-analysis by pooling within-group standardized mean differences from baseline to 3-, 6-, and 12-month follow-ups. From 5,754 studies, 49 comprising 3,825 WAD participants were eligible for the review and 14 for the data synthesis. Altered nociceptive processing in acute and chronic WAD, alongside worse scores on psychological factors, were identified. However, correlations between QST and psychological factors were heterogeneous and inconsistent. Furthermore, disability levels, some QST measures, and psychological factors followed general positive improvement over time, although there were differences in magnitude and temporal changes. These results may indicate that altered psychological factors and increased local pain sensitivity could play an important role in both acute and chronic WAD, although this does not exclude the potential influence of factors not explored in this review. PERSPECTIVE: Acute WAD show improvements in levels of disability and psychological factors before significant improvements in nociceptive processing are evident. Facilitated nociceptive processing might not be as important as psychological factors in chronic WAD-related disability, which indicates that chronic and acute WAD should not be considered the same entity although there are similarities. Nonetheless, pressure pain thresholds in the neck might be the most appropriate measure to monitor WAD progression.

Geo-Spatial Economic Assessment of the Potential Development of Bioenergy Combined with Direct Air Carbon Capture (BEDAC) in the USA.

This study presents a geo-spatial and economic framework to localize future bioenergy power plants combined with direct air capture (BEDAC). This framework is applied to two regions in the USA to assess the optimal use of forest biomass and in situ carbon sequestration under three specific short-term sequestration targets. Results show that there are many locations that have both the necessary biomass and geology required for storage. The Southeast has greater potential for forestry biomass due to both the rate of growth and forested areas, but the sequestration potential is mostly limited to a CO2 solution in saline aquifers. The Pacific Northwest has more sequestration potential than the Southeast given the location of managed forests and storage sites in carbonate mineralization in bedrock. The two combined regions have a total potential sequestration of 9.3 GtCO2 for the next 20 years that can be achieved under an implicit carbon value of $249/tCO2.

Mechanistic insights into glycoside 3-oxidases involved in C-glycoside metabolism in soil microorganisms.

C-glycosides are natural products with important biological activities but are recalcitrant to degradation. Glycoside 3-oxidases (G3Oxs) are recently identified bacterial flavo-oxidases from the glucose-methanol-coline (GMC) superfamily that catalyze the oxidation of C-glycosides with the concomitant reduction of O2 to H2O2. This oxidation is followed by C-C acid/base-assisted bond cleavage in two-step C-deglycosylation pathways. Soil and gut microorganisms have different oxidative enzymes, but the details of their catalytic mechanisms are largely unknown. Here, we report that PsG3Ox oxidizes at 50,000-fold higher specificity (kcat/Km) the glucose moiety of mangiferin to 3-keto-mangiferin than free D-glucose to 2-keto-glucose. Analysis of PsG3Ox X-ray crystal structures and PsG3Ox in complex with glucose and mangiferin, combined with mutagenesis and molecular dynamics simulations, reveal distinctive features in the topology surrounding the active site that favor catalytically competent conformational states suitable for recognition, stabilization, and oxidation of the glucose moiety of mangiferin. Furthermore, their distinction to pyranose 2-oxidases (P2Oxs) involved in wood decay and recycling is discussed from an evolutionary, structural, and functional viewpoint.

What role geoparks play improving the health and well-being of senior tourists?

In recent years geoparks, helped by governmental policies, have become tourist destinations especially among senior visitors. The paper aimed to analyse whether geoparks contribute to improving the health of tourists older than 65 years and what were their main motives to visit geoparks. The data were collected from 398 senior tourists who visited the Villuerca- Ibores-Jara Geopark (Spain) in 2023, presenting our results using SmartPLS version 4. The results showed that senior tourists are very interested in visiting this geopark for psychotherapeutic reasons, given its high environmental and geological interest. In addition, they consider geoparks as spaces where they can socialise, which is beneficial considering the isolation that many often experience during the year. These findings are highly relevant for public authorities to protect, maintain and promote geoparks among senior tourists.

Influence of chronic kidney disease and its severity on the efficacy of semaglutide in type 2 diabetes patients: a multicenter real-world study.

Objectives: Semaglutide is a glucagon-like peptide 1 receptor agonist that improves glycemic control and achieves weight loss in type 2 diabetes (T2D) patients. Subcutaneous (s.c.) semaglutide at 1 mg once weekly (OW) is safe in T2D patients with chronic kidney disease (CKD). Whether or not CKD and its severity influence treatment response remains undetermined. Method: This is an observational, ambispective, multicenter, nationwide, real-world study designed to compare safety/efficacy of OW s.c. 1 mg semaglutide in T2D patients with or without CKD. The influence of CKD severity was also addressed. Patients were followed up for 12 months. Primary end-points were glycosylated hemoglobin (HbA1c), weight, and renal outcomes. Secondary end-points included insulin resistance, atherogenic and hepatic steatosis indexes, and changes in antihyperglycemic medications. Results: A total of 296 and 190 T2D patients without or with CKD, respectively, were recruited. Baseline CKD risk was moderate, high, or very high in 82, 53, and 45 patients, respectively. Treatment reduced HbA1c by 0.90%-1.20%. Relevant differences were seen neither between non-CKD and CKD patients nor among CKD subgroups. Notable weight losses were achieved in both non-CKD and CKD patients. The median reduction was higher in the former at 6 months (5.90 kg vs. 4.50 kg, P = 0.008) and at end of study (6.90 kg vs. 5.00 kg, P = 0.087). A trend toward slightly lower weight losses as CKD severity increased was observed. CKD markers improved across all CKD subgroups. Relevant differences were not observed for other variables, either between non-CKD and CKD patients, or among CKD subgroups. Safety concerns were not reported. Conclusion: The safety/efficacy of OW s.c. semaglutide to improve glycemic control and weight in T2D patients with CKD is not notably lower than that in T2D patients without renal failure. CKD severity barely influences treatment response. OW s.c. semaglutide can be useful to manage T2D patients with CKD in daily clinical practice.

TOR complex 1 negatively regulates NDR kinase Cbk1 to control cell separation in budding yeast.

The target of rapamycin (TOR) signalling pathway plays a key role in the coordination between cellular growth and the cell cycle machinery in eukaryotes. The underlying molecular mechanisms by which TOR might regulate events after anaphase remain unknown. We show for the first time that one of the 2 TOR complexes in budding yeast, TORC1, blocks the separation of cells following cytokinesis by phosphorylation of a member of the NDR (nuclear Dbf2-related) protein-kinase family, the protein Cbk1. We observe that TORC1 alters the phosphorylation pattern of Cbk1 and we identify a residue within Cbk1 activation loop, T574, for which a phosphomimetic substitution makes Cbk1 catalytically inactive and, indeed, reproduces TORC1 control over cell separation. In addition, we identify the exocyst component Sec3 as a key substrate of Cbk1, since Sec3 activates the SNARE complex to promote membrane fusion. TORC1 activity ultimately compromises the interaction between Sec3 and a t-SNARE component. Our data indicate that TORC1 negatively regulates cell separation in budding yeast by participating in Cbk1 phosphorylation, which in turn controls the fusion of secretory vesicles transporting hydrolase at the site of division.

Clinical utility of an antibody-free LC-MS method to detect brain amyloid deposition in cognitively unimpaired individuals from the screening visit of the A4 Study.

INTRODUCTION: This study explored the ability of plasma amyloid beta (Aß)42/Aß40 to identify brain amyloid deposition in cognitively unimpaired (CU) individuals. METHODS: Plasma Aß was quantified with an antibody-free high-performance liquid chromatography tandem mass spectrometry method from Araclon Biotech (ABtest-MS) in a subset of 731 CU individuals from the screening visit of the Anti-Amyloid Treatment in Asymptomatic Alzheimer's (A4) Study, to assess associations of Aß42/Aß40 with Aß positron emission tomography (PET). RESULTS: A model including Aß42/Aß40, age, apolipoprotein E ε4, and recruitment site identified Aß PET status with an area under the curve of 0.88 and an overall accuracy of 81%. A plasma-based pre-screening step could save up to 42% of the total number of Aß PET scans. DISCUSSION: ABtest-MS accurately identified brain amyloid deposition in a population of CU individuals, supporting its implementation in AD secondary prevention trials to reduce recruitment time and costs. Although a certain degree of heterogeneity is inherent to large and multicentric trials, ABtest-MS could be more robust to pre-analytical bias compared to other immunoprecipitation mass spectrometry methods. HIGHLIGHTS: Plasma amyloid beta (Aß)42/Aß40 accurately identified brain Aß deposition in cognitively unimpaired individuals from the Anti-Amyloid Treatment in Asymptomatic Alzheimer's (A4) Study.The inclusion of the recruitment site in the predictive models has a non-negligible effect.A plasma biomarker-based model could reduce recruitment costs in Alzheimer's disease secondary prevention trials.Antibody-free liquid chromatography mass spectrometry methods may be more robust to pre-analytical variability than other platforms.

Enzyme immobilization studied through molecular dynamic simulations.

In recent years, simulations have been used to great advantage to understand the structural and dynamic aspects of distinct enzyme immobilization strategies, as experimental techniques have limitations in establishing their impact at the molecular level. In this review, we discuss how molecular dynamic simulations have been employed to characterize the surface phenomenon in the enzyme immobilization procedure, in an attempt to decipher its impact on the enzyme features, such as activity and stability. In particular, computational studies on the immobilization of enzymes using i) nanoparticles, ii) self-assembled monolayers, iii) graphene and carbon nanotubes, and iv) other surfaces are covered. Importantly, this thorough literature survey reveals that, while simulations have been primarily performed to rationalize the molecular aspects of the immobilization event, their use to predict adequate protocols that can control its impact on the enzyme properties is, up to date, mostly missing.

Results of semaglutide in patients older than 70 years, a real-world study of efficacy and safety.

BACKGROUND: The aim of this study is to investigate the use of once-weekly semaglutide in a real population of patients with type 2 diabetes mellitus (T2DM) over 70 years in two Spanish hospitals. METHODS: An observational, retrospective, and multicenter clinical study was designed. It included 60 patients with T2DM, with a mean age of 76.5 years, 63.3% women, and a mean of 15.5 years of evolution of T2DM, all managed in the outpatient clinical setting. The primary endpoint was the change in HbA1c from baseline to the end of the study. The secondary endpoints included changes in body weight and the proportion of patients achieving HbA1c <7.0% and body weight loss >5%. RESULTS: After 12 months of follow-up, the reductions in HbA1c were -0.61±0.9% (P<0.0001) in the total cohort. Body weight reductions were -8.2±5.3 kg (P<0.0001). Overall, 67% reached the objective of an HbA1c level of <7%, and 73% achieved a weight loss of ≥5%. CONCLUSIONS: In routine clinical practice in Spain, the use of semaglutide once a week was associated with statistically significant and clinically relevant improvements in HbA1c and body weight in adults aged over 70 years with T2DM, without notable adverse effects, which supports real-world use.

Cerebral Venous Thrombosis After Unintentional Dural Puncture: Raising awareness for an uncommon cause of postpartum headache.

Headache is a common symptom in the postpartum period, which can have a varied aetiology. Although rare, cerebral venous thrombosis can be a fatal complication in the parturient. Dural puncture is considered as one of the risk factors for cerebral venous thrombosis and the proposed mechanism pathogenesis can be explained by the components of Virchow's triad: stasis of the blood, hypercoagulability, and endothelial damage. Headache is usually the most frequent symptom and can mimic those of postdural puncture headache, which can delay the diagnosis. We will report a case of an 18-year-old woman that develops a postpartum headache after an accidental dural puncture during epidural catheter placement for labour analgesia. Our patient was initially managed for postdural puncture headache, but later the character changed, which made us look for a differential diagnosis. After a multidisciplinary approach, neuroimaging confirmed the diagnosis of cerebral venous thrombosis. This case report emphasises the importance of a careful differential diagnosis of postpartum headache particularly if the headache persists or changes its character. Brain imaging and multidisciplinary evaluation can lead to prompt diagnosis and initiation of appropriate treatment.

PKD phosphorylation and COP9/Signalosome modulate intracellular Spry2 protein stability.

Spry2 is a molecular modulator of tyrosine kinase receptor signaling pathways that has cancer-type-specific effects. Mammalian Spry2 protein undergoes tyrosine and serine phosphorylation in response to growth factor stimulation. Spry2 expression is distinctly altered in various cancer types. Inhibition of the proteasome functionality results in reduced intracellular Spry2 degradation. Using in vitro and in vivo assays, we show that protein kinase D (PKD) phosphorylates Spry2 at serine 112 and interacts in vivo with the C-terminal half of this protein. Importantly, missense mutation of Ser112 decreases the rate of Spry2 intracellular protein degradation. Either knocking down the expression of all three mammalian PKD isoforms or blocking their kinase activity with a specific inhibitor contributes to the stabilization of Spry2 wild-type protein. Downregulation of CSN3, a component of the COP9/Signalosome that binds PKD, significantly increases the half-life of Spry2 wild-type protein but does not affect the stability of a Spry2 after mutating Ser112 to the non-phosphorylatable residue alanine. Our data demonstrate that both PKD and the COP9/Signalosome play a significant role in control of Spry2 intracellular stability and support the consideration of the PKD/COP9 complex as a potential therapeutic target in tumors where Spry2 expression is reduced.

Contribution of clinical information to the predictive performance of plasma ß-amyloid levels for amyloid positron emission tomography positivity.

Background: Early detection of ß-amyloid (Aß) accumulation, a major biomarker for Alzheimer's disease (AD), has become important. As fluid biomarkers, the accuracy of cerebrospinal fluid (CSF) Aß for predicting Aß deposition on positron emission tomography (PET) has been extensively studied, and the development of plasma Aß is beginning to receive increased attention recently. In the present study, we aimed to determine whether APOE genotypes, age, and cognitive status increase the predictive performance of plasma Aß and CSF Aß levels for Aß PET positivity. Methods: We recruited 488 participants who underwent both plasma Aß and Aß PET studies (Cohort 1) and 217 participants who underwent both cerebrospinal fluid (CSF) Aß and Aß PET studies (Cohort 2). Plasma and CSF samples were analyzed using ABtest-MS, an antibody-free liquid chromatography-differential mobility spectrometry-triple quadrupole mass spectrometry method and INNOTEST enzyme-linked immunosorbent assay kits, respectively. To evaluate the predictive performance of plasma Aß and CSF Aß, respectively, logistic regression and receiver operating characteristic analyses were performed. Results: When predicting Aß PET status, both plasma Aß42/40 ratio and CSF Aß42 showed high accuracy (plasma Aß area under the curve (AUC) 0.814; CSF Aß AUC 0.848). In the plasma Aß models, the AUC values were higher than plasma Aß alone model, when the models were combined with either cognitive stage (p < 0.001) or APOE genotype (p = 0.011). On the other hand, there was no difference between the CSF Aß models, when these variables were added. Conclusion: Plasma Aß might be a useful predictor of Aß deposition on PET status as much as CSF Aß, particularly when considered with clinical information such as APOE genotype and cognitive stage.

Clinical performance of an antibody-free assay for plasma Aß42/Aß40 to detect early alterations of Alzheimer's disease in individuals with subjective cognitive decline.

BACKGROUND: Accessible and cost-effective diagnostic tools are urgently needed to accurately quantify blood biomarkers to support early diagnosis of Alzheimer's disease (AD). In this study, we investigated the ability of plasma amyloid-beta (Aß)42/Aß40 ratio measured by an antibody-free mass-spectrometric (MS) method, ABtest-MS, to detect early pathological changes of AD. METHODS: This cohort study included data from the baseline and 2-year follow-up visits from the Fundació ACE Healthy Brain Initiative (FACEHBI) study. Plasma Aß42/Aß40 was measured with ABtest-MS and compared to 18F-Florbetaben PET as the reference standard (cutoff for early amyloid deposition of 13.5 centiloids). Cross-validation was performed in an independent DPUK-Korean cohort. Additionally, associations of plasma Aß42/Aß40 with episodic memory performance and brain atrophy were assessed. RESULTS: The FACEHBI cohort at baseline included 200 healthy individuals with subjective cognitive decline (SCD), of which 36 (18%) were Aß-PET positive. Plasma Aß42/Aß40 levels were significantly lower in Aß-PET positive individuals (median [interquartile range, IQR], 0.215 [0.203-0.236]) versus Aß-PET negative subjects (median [IQR], 0.261 [0.244-0.279]) (P < .001). Plasma Aß42/Aß40 was significantly correlated with Aß-PET levels (rho = -0.390; P < .001) and identified Aß-PET status with an area under the receiver operating characteristic curve (AUC) of 0.87 (95% confidence interval [CI], 0.80-0.93). A cutoff for the Aß42/Aß40 ratio of 0.241 (maximum Youden index) yielded a sensitivity of 86.1% and a specificity of 80.5%. These findings were cross-validated in an independent DPUK-Korean cohort (AUC 0.86 [95% CI 0.77-0.95]). Lower plasma Aß42/Aß40 ratio was associated with worse episodic memory performance and increased brain atrophy. Plasma Aß42/Aß40 at baseline predicted clinical conversion to mild cognitive impairment and longitudinal changes in amyloid deposition and brain atrophy at 2-year follow-up. CONCLUSIONS: This study suggests that plasma Aß42/Aß40, as determined by this MS-based assay, has potential value as an accurate and cost-effective tool to identify individuals in the earliest stages of AD, supporting its implementation in clinical trials, preventative strategies and clinical practice.

Corrigendum to "Unveiling molecular details behind improved activity at neutral to alkaline pH of an engineered DyP-type peroxidase" Comput Struct Biotechnol J, 20 (2022), 3899-3910.

[This corrects the article DOI: 10.1016/j.csbj.2022.07.032.].

Bronchobiliary fistula after stenting of biliary duct as the management of iatrogenic bile duct injury during elective cholecystectomy.

BACKGROUND: Bronchobiliary fistula is a rare and complex entity defined by an abnormal communication between the biliary and bronchial systems. The etiopathogenesis is not completely understood, but the most common factors implicated are hepatobiliary tumors, biliary obstruction, iatrogenic damage or trauma. METHODS: Here we present a case of a 69-year-old man that developed a bronchobiliary fistula and a pulmonary abscess after migration of a bile duct stent placed as part of the treatment of an iatrogenic bile duct injury that occurred during elective cholecystectomy. RESULTS: A conservative approach, that included broad-spectrum antibiotic, removal of the stent, and sphincterotomy, was enough for the closure of the fistula and resolution of the symptoms. CONCLUSION: We emphasize the importance of prompt recognition of this entity and a concerted therapeutic strategy to optimize the probability of success, avoiding the destructive consequences of the bile in the pulmonary parenchyma and septic complications.