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Summary: Background: While many patients seem to recover from SARS-CoV-2 infections, many patients report experiencing SARS-CoV-2 symptoms for weeks or months after their acute COVID-19 ends, even developing new symptoms weeks after infection. These long-term effects are called post-acute sequelae of SARS-CoV-2 (PASC) or, more commonly, Long COVID. The overall prevalence of Long COVID is currently unknown, and tools are needed to help identify patients at risk for developing long COVID. Methods: A working group of the Rapid Acceleration of Diagnostics-radical (RADx-rad) program, comprised of individuals from various NIH institutes and centers, in collaboration with REsearching COVID to Enhance Recovery (RECOVER) developed and organized the Long COVID Computational Challenge (L3C), a community challenge aimed at incentivizing the broader scientific community to develop interpretable and accurate methods for identifying patients at risk of developing Long COVID. From August 2022 to December 2022, participants developed Long COVID risk prediction algorithms using the National COVID Cohort Collaborative (N3C) data enclave, a harmonized data repository from over 75 healthcare institutions from across the United States (U.S.). Findings: Over the course of the challenge, 74 teams designed and built 35 Long COVID prediction models using the N3C data enclave. The top 10 teams all scored above a 0.80 Area Under the Receiver Operator Curve (AUROC) with the highest scoring model achieving a mean AUROC of 0.895. Included in the top submission was a visualization dashboard that built timelines for each patient, updating the risk of a patient developing Long COVID in response to clinical events. Interpretation: As a result of L3C, federal reviewers identified multiple machine learning models that can be used to identify patients at risk for developing Long COVID. Many of the teams used approaches in their submissions which can be applied to future clinical prediction questions. Funding: Research reported in this RADx® Rad publication was supported by the National Institutes of Health. Timothy Bergquist, Johanna Loomba, and Emily Pfaff were supported by Axle Subcontract: NCATS-STSS-P00438.

Summary: Background: The HVTN 705 Imbokodo trial of 2636 people without HIV and assigned female sex at birth, conducted in southern Africa, evaluated a heterologous HIV-1 vaccine regimen: mosaic adenovirus 26-based vaccine (Ad26.Mos4.HIV) at Months 0, 3, 6, 12 and alum-adjuvanted clade C gp140 at Months 6, 12. Per-protocol vaccine efficacy (VE) against HIV-1 diagnosis from seven to 24 months was 14.1% (95% CI: −22.0% to 39.5%). Immune correlates analysis was performed for markers selected based on prior evidence in efficacy trials and/or nonhuman primate models. Methods: Humoral and cellular immune response markers at Month 7 were evaluated as immune correlates of risk and of protection in a breakthrough case–control cohort (n = 52 cases, 246 non-cases). Primary markers were IgG binding to vaccine-strain gp140, IgG3 binding to diverse Env antigens (IgG3 Env breadth), IgG3 binding to diverse V1V2 antigens (IgG3 V1V2 breadth), antibody-dependent phagocytosis against the vaccine-strain gp140, Env-specific CD4+ and CD8+ T-cell responses, and multi-epitope functions. Findings: No immune markers were statistically significant correlates of risk. IgG3 V1V2 breadth trended toward an inverse association: hazard ratio 0.70 (95% CI: 0.36 to 1.35; p = 0.29) per 10-fold increase and 0.51 (95% CI: 0.21 to 1.24; p = 0.14) in a Cox model with all primary markers. The VE estimate was 11.8% (95% CI: −17.9% to 34.0%) at all IgG3 V1V2 breadth values below 667 weighted geometric mean net MFI; just above this value, the VE estimate sharply increased to 62.6% (95% CI: −17.9% to 89.6%), and further increased to 80.9% (95% CI: −17.9% to 99.5%) at 1471 MFI, the 95th percentile of the marker distribution. Mediation analysis yielded a VE of 35.7% (95% CI: 15.0% to 51.3%) attributable to the vaccine's impact on this marker. Interpretation: The trend in association of greater IgG3 V1V2 antibody breadth with lower likelihood of HIV acquisition is consistent with the identification of antibodies against V1V2 as immune correlates in three other HIV vaccine efficacy trials and suggests that a greater emphasis should be placed on studying this region in the HIV-1 envelope as a vaccine immunogen. Funding: National Institute of Allergy and Infectious Diseases and Janssen Vaccines & Prevention BV.

Study region: The Steenkoppies Catchment is located approximately 75 km southwest from Pretoria, South Africa (RSA). Study focus: This study tested a framework for downscaling Global Land Data Assimilation System (GLDAS-2.2) groundwater storage anomaly (GWSA) estimates from 0.25° to 0.05°. This was achieved in Google Earth Engine using the Random Forest algorithm with only precipitation and actual evapotranspiration (ETa) as input variables. Additionally, the study assessed whether accounting for temporal lags could minimise residuals and enhance model performance. New hydrological insights for the region: The greater range of downscaled GWSA values indicated that the product effectively captured local recharge (precipitation) and discharge (ETa) variations while maintaining conservation of mass. Optimising the temporal correlation (r) between input variables resulted in lower residuals and fewer outliers. Groundwater level measurements and downscaled estimates for the hard rock aquifer showed larger amplitudes and seasonality and yielded the highest r (0.6) and lowest RMSE (40 mm) and MAE (31 mm). Measurements near the spring and in the karst aquifer showed less evident amplitude and seasonality. The in situ derived and downscaled GWSA comparison demonstrated the effectiveness of the product for monitoring storage declines. When applied over aquifers experiencing significant land use change or below-average precipitation, the approach could monitor groundwater storage changes, even with limited in situ observations. The adaptable code is available for application in other study areas.

ABSTRACT: This study investigated the diet–additive interactions of a Lactobacilli-based probiotic (Pro) and postbiotic (Post) on immune parameters and cecal microbiota composition, with subsequent effects on the metabolome in broilers. A completely randomized block design was employed with 2 diets [standard (SD), and challenge (CD)] and 3 additive conditions (Control, Pro, Post) involving 1,368 one-day-old male Ross 308 broilers equally distributed among 36 pens in a 42 d study. Diets were formulated to contain identical nutrient levels, with CD higher than SD in non-starch polysaccharide content by including rye and barley. Total non-specific serum Ig A, M and G concentrations were determined weekly from d14 to 35. Following vaccination, titres of specific antibodies binding Newcastle disease virus (NDV) and infectious bursal disease virus (IBDV) were measured. Microbiota composition was analyzed by 16S rRNA gene sequencing at d14 and 35, and α- and β-diversity indexes (Observed, Chao1, Bray, Jaccard) were calculated. Cecal short-chain fatty acids and the semi-polar metabolome were determined in the Control SD and all CD groups at d35. At d35, a diet–additive interaction was observed on cecal microbiota composition. Within SD, Pro and Post did not affect operational taxonomic units (OTU) abundance (adjusted-P > 0.05) and diversity indexes (P > 0.05). Within CD, Pro and Post affected the relative abundances of 37 and 44 OTUs, respectively (adjusted-P < 0.05), with Post but not Pro affecting β-diversity indexes (P = 0.041 and 0.064 for Bray and Jaccard, respectively). Within CD, Post increased cecal acetate (21%; P = 0.007) and butyrate (41%; P = 0.002) concentration and affected the concentration of 2 metabolites (adjusted-P < 0.05), while Pro affected 240 metabolites (adjusted-P < 0.05). No diet–additive interactions were observed on serum Ig (P > 0.05), except for IgM at d14 (P = 0.004). Diet composition, but not the additives, affected immune status parameters. The Pro and Post affected cecal microbiota composition only under dietary challenging conditions as previously reported for growth.

Summary: Fibroblastic reticular cells (FRCs) are mesenchymal stromal cells in human lymph nodes (LNs) playing a pivotal role in adaptive immunity. Several FRC subsets have been identified, yet it remains to be elucidated if their heterogeneity is maintained upon culture. Here, we established a protocol to preserve and culture FRCs from human LNs and characterized their phenotypic profile in fresh LN suspensions and upon culture using multispectral flow cytometry. We found nine FRC subsets in fresh human LNs, independent of donor, of which four persisted in culture throughout several passages. Interestingly, the historically FRC-defining marker podoplanin (PDPN) was not present on all FRC subsets. Therefore, we propose that CD45negCD31neg human FRCs are not restricted by PDPN expression, as we found CD90, BST1, and CD146/MCAM to be more widely expressed. Together, our data provide insight into FRC heterogeneity in human LNs, enabling further investigation into the function of individual FRC subsets.

ABSTRACT: Individual differences in free-range chicken systems are important factors influencing how birds use the range (or not), even if individuals are reared in the same environmental conditions. Here, we investigated how various aspects of the birds' behavioral and cognitive tendencies, including their optimism/pessimism, cognitive flexibility, sociability, and exploration levels, are associated with range use and how they may change over time (before and after range access). To achieve this, 100 White Leghorn laying hen chicks underwent three distinct behavioral/cognitive tests—the cognitive bias test, the detour test, and the multivariate test—prior to gaining access to the range, between 9 and 39 days of age. After range access was allowed (from day 71), birds' range use was evaluated over 7 nonconsecutive days (from 74–91 days of age). Subsequently, a subset of birds, classified as high rangers (n = 15) and low rangers (n = 15) based on their range use, underwent retesting on the same three previous tests between 94 and 108 days of age. Our results unveiled a negative correlation trend between birds' evaluation of the ambiguous cue and their subsequent range use (rho = -0.19, p = 0.07). Furthermore, low rangers were faster to learn the detour task (χ2 = 7.34, df = 1, p = 0.006), coupled with increased sociability during the multivariate test (rho = -0.23, p = 0.02), contrasting with their high-ranging counterparts, who displayed more exploratory behaviors (F[1,27] = 3.64, p = 0.06). These behavioral patterns fluctuated over time (before and after range access); however, conclusively attributing these changes to birds' aging and development or the access to the range remains challenging. Overall, our results corroborate that behavioral and cognitive individual differences may be linked to range use and offer novel perspectives on the early behavioral and cognitive traits that may be linked to range use. These findings may serve as a foundation for adapting environments to meet individual needs and improve animal welfare in the future.

ABSTRACT: Dietary ingredient and nutrient composition may affect the efficacy of additives in broilers. Specific feed ingredients can represent dietary challenging conditions for broilers, resulting in impaired performances and health, which might be alleviated by dietary probiotics and postbiotics. We assessed the effects of a Lactobacilli probiotic (Pro) and postbiotic (Post) when added to a standard (SD) and challenge (CD) diet. A completely randomized block study with 2 diets (SD, CD) and 3 additive conditions (Control, Pro and Post) involving 1,368 one-day-old Ross male broilers, equally distributed among 36 pens, from d1 to d42 was conducted. Both diets were formulated to contain identical levels of nutrients, with CD formulated to be richer than SD in nonstarch polysaccharides using rye and barley as ingredients. Readout parameters included growth performance parameters, footpad lesions score, blood minerals and biochemical parameters, and tibia health, strength, and composition. Compared to SD, CD decreased BW (1,936 vs. 2,033 g; p = 0.001), increased FCR (p < 0.01) and impaired tibia health and strength (p < 0.05) at d35, thereby confirming the challenging effect of CD. Pro and Post increased BW in CD (+4.7 and +3.2%, respectively, at d35; P < 0.05) but not in the SD group, without affecting FCR. Independently of the diet, Pro increased plasma calcium, phosphorus and uric acid at d21 (+6.2, +7.4, and +15.5%, respectively) and d35 (+6.6, +6.2 and +21.0%, respectively) (P < 0.05) while Post increased plasma magnesium only at d21 (+11.3%; P = 0.037). Blood bile acids were affected by additives in an age- and diet-dependent manner, with some opposite effects between dietary conditions. Diet composition modulated Pro and Post effects on broiler growth performance. Additionally, Pro and Post affected animal metabolism and leg health diet-dependently for some but not all investigated parameters. Our findings show that the effects of pro- and postbiotics on the growth performance and physiology of broilers can be dependent on diet composition and thus possibly other factors affecting diet characteristics.

Basic oxygen furnace slag (BOF slag), also known as Linz-Donauwitz slag, is an industrially by-product of the steel industry and produced during the converter process. In this research we have investigated the effect of grinding aids and particle fineness on the hydration of potassium citrate activated BOF slag. It was investigated the effects of five different grinding aids on the grindability and hydration of activated BOF slag in laboratory batch milling experiments. The grinding aids generally improve the grindability of BOF slag and therefore increase the particle fineness of BOF slag. This had the consequence that the hydration of activated BOF slag was improved. Only one grinding aid, triethanolamine (TEA) also improved the hydration of BOF slag besides providing higher particle fineness. The combination of the higher particle fineness and additional effect of TEA on the hydration improved the compressive strength to 60 and 78 MPa of activated BOF slag. An additional important of this research is that the hydration of BOF slag is significantly more sensitive on particle fineness compared to ordinary Portland cement.

ABSTRACT: Vaccination is one of the most effective strategies for preventing infectious diseases but individual vaccine responses are highly heterogeneous. Host genetics and gut microbiota composition are 2 likely drivers of this heterogeneity. We studied 94 animals belonging to 4 lines of laying hens: a White Leghorn experimental line genetically selected for a high antibody response against the Newcastle Disease Virus (NDV) vaccine (ND3) and its unselected control line (CTR), and 2 commercial lines (White Leghorn [LEG] and Rhode Island Red [RIR]). Animals were reared in the same conditions from hatching to 42 d of age, and animals from different genetic lines were mixed. Animals were vaccinated at 22 d of age and their humoral vaccine response against NDV was assessed by hemagglutination inhibition assay and ELISA from blood samples collected at 15, 19, and 21 d after vaccination. The immune parameters studied were the 3 immunoglobulins subtypes A, M, and Y and the blood cell composition was assessed by flow cytometry. The composition of the cecal microbiota was assessed at the end of the experiment by analyzing amplified 16S rRNA gene sequences to obtain amplicon sequence variants (ASV). The 4 lines showed significantly different levels of NDV vaccine response at the 3 measured points, with, logically, a higher response of the genetically selected ND3 line, and intermediate and low responses for the unselected CTR control line and for the 2 commercial lines, respectively. The ND3 line displayed also a higher proportion of immunoglobulins (IgA, IgM, and IgY). The RIR line showed the most different blood cell composition. The 4 lines showed significantly different microbiota characteristics: composition, abundances at all taxonomic levels, and correlations between genera and vaccine response. The tested genetic lines differ for immune parameters and gut microbiota composition and functions. These phenotypic differences can be attributed to genetic differences between lines. Causal relationships between both types of parameters are discussed and will be investigated in further studies.

The need to integrate more clearly societal expectations on livestock farming has led the authors of this article to consider that livestock farming systems must be redesigned to position health and welfare at the heart of their objectives. This article proposes a vision of the advances in knowledge required at different scales to contribute to this transformation. After defining health and welfare of animals, the article emphasises the need to consider health in a broader perspective, to deepen the question of positive emotional experiences regarding welfare, and raises the question of how to assess these two elements on farms. The positive interactions between health and welfare are presented. Some possible tensions between them are also discussed, in particular when improving welfare by providing a more stimulating and richer environment such as access to outdoor increases the risk of infectious diseases. Jointly improving health and welfare of animals poses a number of questions at various scales, from the animal level to the production chain. At the animal level, the authors highlight the need to explore: the long-term links between better welfare and physiological balance, the role of microbiota, the psycho-neuro-endocrine mechanisms linking positive mental state and health, and the trade-off between the physiological functions of production, reproduction and immunity. At the farm level, in addition to studying the relationships at the group level between welfare, health and production, the paper supports the idea of co-constructing innovative systems with livestock farmers, as well as analysing the cost, acceptability and impact of improved systems on their working conditions and well-being. At the production chain or territory levels, various questions are raised. These include studying the best strategies to improve animal health and welfare while preserving economic viability, the labelling of products and the consumers’ willingness to pay, the consequences of heterogeneity in animal traits on the processing of animal products, and the spatial distribution of livestock farming and the organisation of the production and value chain. At the level of the citizen and consumer, one of the challenges is to better inter-relate sanitary and health perspectives on the one hand, and welfare concerns on the other hand. There is also a need to improve citizens' knowledge on livestock farming, and to develop more intense and constructive exchanges between livestock farmers, the livestock industry and citizens. These difficult issues plead for interdisciplinary and transdisciplinary research involving various scientific disciplines and the different stakeholders, including public policy makers through participatory research.

Summary: The mitochondrial inner membrane plays central roles in bioenergetics and metabolism and contains several established membrane protein complexes. Here, we report the identification of a mega-complex of the inner membrane, termed mitochondrial multifunctional assembly (MIMAS). Its large size of 3 MDa explains why MIMAS has escaped detection in the analysis of mitochondria so far. MIMAS combines proteins of diverse functions from respiratory chain assembly to metabolite transport, dehydrogenases, and lipid biosynthesis but not the large established supercomplexes of the respiratory chain, ATP synthase, or prohibitin scaffold. MIMAS integrity depends on the non-bilayer phospholipid phosphatidylethanolamine, in contrast to respiratory supercomplexes whose stability depends on cardiolipin. Our findings suggest that MIMAS forms a protein-lipid mega-assembly in the mitochondrial inner membrane that integrates respiratory biogenesis and metabolic processes in a multifunctional platform.

Objective: Continuation of smoking after a cancer diagnosis increases the burdensome side effects from cancer treatment, and decreases the chances of cure. Smoking cessation may improve oncological outcomes in cancer patients. This study aims to evaluate if radiation oncologists can be motivated by a smoking cessation awareness intervention to discuss smoking status more frequently and increase the referral rate for smoking cessation-support.Study design and Setting:A multifaceted approach was used to improve awareness: First, current practice was evaluated by a retrospective chart review of 282 patients referred for radiotherapy to ascertain smoking status, discussion of smoking cessation support and referral rates. Secondly, radiation oncologists were interviewed about their motives and barriers to discuss smoking status and smoking cessation support. The results were fed back in a teaching lecture to the doctors involved. Finally, the effect of this smoking cessation awareness intervention was prospectively evaluated in 100 patients. Results: After the smoking cessation awareness intervention, smoking cessation was more frequently discussed compared to baseline (77% (10/13)) and 39.5% (17/43) respectively. The referral rate for smoking cessation increased from 2.3% (1/43) to 69.2% (9/13) Conclusion: Without an active smoking prevention awareness policy, referral for smoking cessation support for cancer patients by radiation oncologists is low. A relatively short and simple smoking awareness intervention for radiation oncologist may result in a more frequent discussion with patients about smoking cessation and an even larger increase in referrals for smoking cessation support.

Background and aims: Patients who underwent carotid endarterectomy (CEA) still have a residual risk of 13% of developing a major adverse cardiovascular event (MACE) within 3 years. Inflammatory processes leading up to MACE are not fully understood. Therefore, we examined blood cell characteristics (BCCs), possibly reflecting inflammatory processes, in relation to MACE to identify BCCs that may contribute to an increased risk. Methods: We analyzed 75 pretreatment BCCs from the Sapphire analyzer, and clinical data from the Athero-Express biobank in relation to MACE after CEA using Random Survival Forests, and a Generalized Additive Survival Model. To understand biological mechanisms, we related the identified variables to intraplaque hemorrhage (IPH). Results: Of 783 patients, 97 (12%) developed MACE within 3 years after CEA. Red blood cell distribution width (RDW) (HR 1.23 [1.02, 1.68], p = 0.022), CV of lymphocyte size (LACV) (HR 0.78 [0.63, 0.99], p = 0.043), neutrophil complexity of the intracellular structure (NIMN) (HR 0.80 [0.64, 0.98], p = 0.033), mean neutrophil size (NAMN) (HR 0.67 [0.55, 0.83], p

Introduction: Technical advances and the increasing role of interdisciplinary decision-making may warrant formal definitions of expertise in surgical neuro-oncology. Research question: The EANS Neuro-oncology Section felt that a survey detailing the European neurosurgical perspective on the concept of expertise in surgical neuro-oncology might be helpful. Material and methods: The EANS Neuro-oncology Section panel developed an online survey asking questions regarding criteria for expertise in neuro-oncological surgery and sent it to all individual EANS members. Results: Our questionnaire was completed by 251 respondents (consultants: 80.1%) from 42 countries. 67.7% would accept a lifetime caseload of >200 cases and 86.7% an annual caseload of >50 as evidence of neuro-oncological surgical expertise. A majority felt that surgeons who do not treat children (56.2%), do not have experience with spinal fusion (78.1%) or peripheral nerve tumors (71.7%) may still be considered experts. Majorities believed that expertise requires the use of skull-base approaches (85.8%), intraoperative monitoring (83.4%), awake craniotomies (77.3%), and neuro-endoscopy (75.5%) as well as continuing education of at least 1/year (100.0%), a research background (80.0%) and teaching activities (78.7%), and formal interdisciplinary collaborations (e.g., tumor board: 93.0%). Academic vs. non-academic affiliation, career position, years of neurosurgical experience, country of practice, and primary clinical interest had a minor influence on the respondents’ opinions. Discussion and conclusion: Opinions among neurosurgeons regarding the characteristics and features of expertise in neuro-oncology vary surprisingly little. Large majorities favoring certain thresholds and qualitative criteria suggest a consensus definition might be possible.

Summary: Genome-wide association studies (GWASs) have identified hundreds of risk loci for coronary artery disease (CAD). However, non-European populations are underrepresented in GWASs, and the causal gene-regulatory mechanisms of these risk loci during atherosclerosis remain unclear. We incorporated local ancestry and haplotypes to identify quantitative trait loci for expression (eQTLs) and splicing (sQTLs) in coronary arteries from 138 ancestrally diverse Americans. Of 2,132 eQTL-associated genes (eGenes), 47% were previously unreported in coronary artery; 19% exhibited cell-type-specific expression. Colocalization revealed subgroups of eGenes unique to CAD and blood pressure GWAS. Fine-mapping highlighted additional eGenes, including TBX20 and IL5. We also identified sQTLs for 1,690 genes, among which TOR1AIP1 and ULK3 sQTLs demonstrated the importance of evaluating splicing to accurately identify disease-relevant isoform expression. Our work provides a patient-derived coronary artery eQTL resource and exemplifies the need for diverse study populations and multifaceted approaches to characterize gene regulation in disease processes.

Summary: Coronary artery disease (CAD) is characterized by atherosclerotic plaque formation in the arterial wall. CAD progression involves complex interactions and phenotypic plasticity among vascular and immune cell lineages. Single-cell RNA-seq (scRNA-seq) studies have highlighted lineage-specific transcriptomic signatures, but human cell phenotypes remain controversial. Here, we perform an integrated meta-analysis of 22 scRNA-seq libraries to generate a comprehensive map of human atherosclerosis with 118,578 cells. Besides characterizing granular cell-type diversity and communication, we leverage this atlas to provide insights into smooth muscle cell (SMC) modulation. We integrate genome-wide association study data and uncover a critical role for modulated SMC phenotypes in CAD, myocardial infarction, and coronary calcification. Finally, we identify fibromyocyte/fibrochondrogenic SMC markers (LTBP1 and CRTAC1) as proxies of atherosclerosis progression and validate these through omics and spatial imaging analyses. Altogether, we create a unified atlas of human atherosclerosis informing cell state-specific mechanistic and translational studies of cardiovascular diseases.

Summary: Bile ducts are essential for bile transport and consist of complex branching tubular networks. Human patient-derived cholangiocyte develops a cystic rather than branching duct morphology. Here, we present a protocol to establish branching morphogenesis in cholangiocyte and cholangiocarcinoma organoids. We describe steps for the initiation, maintenance, and expansion of intrahepatic cholangiocyte organoids branching morphology. This protocol enables the study of organ-specific and mesenchymal-independent branching morphogenesis and provides an improved model to study biliary function and diseases.For complete details on the use and execution of this protocol, please refer to Roos et al. (2022).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Background: The evidence on prophylactic use of negative pressure wound therapy on primary closed incisional wounds (iNPWT) for the prevention of surgical site infections (SSI) is confusing and ambiguous. Implementation in daily practice is impaired by inconsistent recommendations in current international guidelines and published meta-analyses. More recently, multiple new randomised controlled trials (RCTs) have been published. We aimed to provide an overview of all meta-analyses and their characteristics; to conduct a new and up-to-date systematic review and meta-analysis and Grading of Recommendations Assessment, Development and Evaluation (GRADE) assessment; and to explore the additive value of new RCTs with a trial sequential analysis (TSA). Methods: PubMed, Embase and Cochrane CENTRAL databases were searched from database inception to October 24, 2022. We identified existing meta-analyses covering all surgical specialties and RCTs studying the effect of iNPWT compared with standard dressings in all types of surgery on the incidence of SSI, wound dehiscence, reoperation, seroma, hematoma, mortality, readmission rate, skin blistering, skin necrosis, pain, and adverse effects of the intervention. We calculated relative risks (RR) with corresponding 95% confidence intervals (CI) using a Mantel-Haenszel random-effects model. We assessed publication bias with a comparison-adjusted funnel plot. TSA was used to assess the risk of random error. The certainty of evidence was evaluated using the Cochrane Risk of Bias-2 (RoB2) tool and GRADE approach. This study is registered with PROSPERO, CRD42022312995. Findings: We identified eight previously published general meta-analyses investigating iNPWT and compared their results to present meta-analysis. For the updated systematic review, 57 RCTs with 13,744 patients were included in the quantitative analysis for SSI, yielding a RR of 0.67 (95% CI: 0.59–0.76, I2 = 21%) for iNPWT compared with standard dressing. Certainty of evidence was high. Compared with previous meta-analyses, the RR stabilised, and the confidence interval narrowed. In the TSA, the cumulative Z-curve crossed the trial sequential monitoring boundary for benefit, confirming the robustness of the summary effect estimate from the meta-analysis. Interpretation: In this up-to-date meta-analysis, GRADE assessment shows high-certainty evidence that iNPWT is effective in reducing SSI, and uncertainty is less than in previous meta-analyses. TSA indicated that further trials are unlikely to change the effect estimate for the outcome SSI; therefore, if future research is to be conducted on iNPWT, it is crucial to consider what the findings will contribute to the existing robust evidence. Funding: Dutch Association for Quality Funds Medical Specialists.

Summary: Background: Ischemia of the bile duct is a common feature in liver disease and transplantation, which represents a major cause of morbidity and mortality, especially after liver transplantation. Detailed knowledge of its pathogenesis remains incomplete due to the lack of appropriate in vitro models. Methods: To recapitulate biliary damage induced by ischemia and reperfusion in vitro, human intrahepatic cholangiocyte organoids (ICOs) were grown at low oxygen levels of 1% up to 72 h, followed by re-oxygenation at normal levels. Findings: ICOs stressed by ischemia and subsequent re-oxygenation represented the dynamic change in biliary cell proliferation, upregulation of epithelial–mesenchymal transition (EMT)-associated markers, and the evocation of phase-dependent cell death programs similar to what is described in patients. Clinical-grade alpha-1 antitrypsin was identified as a potent inhibitor of both ischemia-induced apoptosis and necroptosis. Interpretation: These findings demonstrate that ICOs recapitulate ischemic cholangiopathy in vitro and enable drug assessment studies for the discovery of new therapeutics for ischemic cholangiopathies. Funding: Dutch Digestive Foundation MLDS D16-26; TKI-LSH (Topconsortium Kennis en Innovatie-Life Sciences & Health) grant RELOAD, EMC-LSH19002; Medical Delta program “Regenerative Medicine 4D”; China Scholarship Council No. 201706230252.

Background & Aims: Liver and bile duct diseases often are associated with extensive cell death of cholangiocytes. Necroptosis represents a common mode of programmed cell death in cholangiopathy, however, detailed mechanistic knowledge is limited owing to the lack of appropriate in vitro models. To address this void, we investigated whether human intrahepatic cholangiocyte organoids (ICOs) can recapitulate cholangiopathy-associated necroptosis and whether this model can be used for drug screening. Methods: We evaluated the clinical relevance of necroptosis in end-stage liver diseases and liver transplantation by immunohistochemistry. Cholangiopathy-associated programmed cell death was evoked in ICOs derived from healthy donors or patients with primary sclerosing cholangitis or alcoholic liver diseases by the various stimuli. Results: The expression of key necroptosis mediators, receptor-interacting protein 3 and phosphorylated mixed lineage kinase domain-like, in cholangiocytes during end-stage liver diseases was confirmed. The phosphorylated mixed lineage kinase domain-like expression was etiology-dependent. Gene expression analysis confirmed that primary cholangiocytes are more prone to necroptosis compared with primary hepatocytes. Both apoptosis and necroptosis could be specifically evoked using tumor necrosis factor α and second mitochondrial-derived activator of caspases mimetic, with or without caspase inhibition in healthy and patient-derived ICOs. Necroptosis also was induced by ethanol metabolites or human bile in ICOs from donors and patients. The organoid cultures further uncovered interdonor variable and species-specific drug responses. Dabrafenib was identified as a potent necroptosis inhibitor and showed a protective effect against ethanol metabolite toxicity. Conclusions: Human ICOs recapitulate cholangiopathy-associated necroptosis and represent a useful in vitro platform for the study of biliary cytotoxicity and preclinical drug evaluation.

Introduction: In the last decades, the application of stereotactic laser ablation (SLA) for the treatment of intracranial tumours has been growing, even though comparative trials are lacking. Our aim was to investigate the familiarity with SLA of neurosurgeons in Europe and their opinion regarding potential neuro-oncological indications. Furthermore, we investigated treatment preferences and variability for three exemplar neuro-oncological cases and willingness to refer for SLA. Material and methods: A 26-questions survey was mailed to members of the EANS neuro-oncology section. We presented three clinical cases of respectively deep-seated glioblastoma, recurrent metastasis and recurrent glioblastoma. Descriptive statistics was applied to report results. Results: 110 respondents completed all questions. Recurrent glioblastoma and recurrent metastases were regarded as the most feasible indications for SLA (chosen by 69% and 58% of the respondents) followed by newly diagnosed high-grade gliomas (31%). Seventy percent of respondents would refer patients for SLA. The majority of respondents would consider SLA as a treatment option for all three presented cases: 79% for the deep-seated glioblastoma case, 65% for the recurrent metastasis case and 76% for the recurrent glioblastoma case. Among respondents who wouldn't consider SLA, preference for standard treatment and lack of clinical evidence were reported as the main reasons. Conclusions: Most of respondents considered SLA as a treatment option for recurrent glioblastoma, recurrent metastases and newly diagnosed deep-seated glioblastoma. At the moment the current evidence to support such a treatment is very low. Comparative prospective trials are needed to support the use of SLA and determine proper indications.

With population growth and dwindling freshwater sources, protecting such sources has come to the forefront of water resource management. Historically, society’s response to a problem is based on funding availability, current threat, and public outcry. Achieving this is largely dependent on the knowledge of the factors that are resulting in compromised water sources. These factors are constantly changing as novel contaminants are introduced into surface water sources. As we are in the information age, the interest in contaminants of emerging concern (CEC) is gaining ground. Whilst research is being conducted to identify contaminants in South African water sources, the research outputs and available information is not collated and presented to the science community and stakeholders in readily available formats and platforms. Current research outcomes need to be made known to regulators in order to develop environmental laws. By using fourth industrial revolution technology, we were able to collate available data in literature and display these in a user-friendly online format to regulatory bodies as well as researchers. A standardized excel spreadsheet was developed and uploaded to a PostgreSQL, running a PostGIS extension and was then processed in the GeoServer to allow for visualization on an interactive map which can be continuously updated. The near real-time access to information will reduce the possibility of duplication of research efforts, enhance collaboration in the discipline, and act as a CEC early warning system.

Summary: Background: Statin use could benefit patients with non-alcoholic fatty liver disease (NAFLD), but the evidence is segmented and inconclusive. This multidimensional study comprehensively investigated the potential benefits and mechanism-of-action of statins in NAFLD. Methods: A cross-sectional investigation was performed within the Rotterdam Study (general population; n = 4.576) and the PERSONS cohort (biopsy-proven NAFLD patients; n = 569). Exclusion criteria were secondary causes for steatosis and insufficient data on alcohol, dyslipidemia or statin use. Associations of statin use with NAFLD (among entire general population), fibrosis and NASH (among NAFLD individuals and patients) were quantified. These results were pooled with available literature in meta-analysis. Last, we assessed statins’ anti-lipid and anti-inflammatory effects in 3D cultured human liver organoids and THP-1 macrophages, respectively. Findings: Statin use was inversely associated with NAFLD in the Rotterdam study compared to participants with untreated dyslipidemia. In the PERSONS cohort, statin use was inversely associated with NASH, but not with fibrosis. The meta-analysis included 7 studies and indicated a not significant inverse association for statin use with NAFLD (pooled-Odds Ratio: 0.69, 95% Confidence Interval: 0.46–1.01) and significant inverse associations with NASH (pooled-OR: 0.59, 95% CI: 0.44–0.79) and fibrosis (pooled-OR: 0.48, 95% CI: 0.33–0.70). In vitro, statins significantly reduced lipid droplet accumulation in human liver organoids and downregulated expression of pro-inflammatory cytokines in macrophages. Interpretation: Pooled results demonstrated that statin use was associated with a lower prevalence of NASH and fibrosis and might prevent NAFLD. This may be partially attributed to the anti-lipid and anti-inflammatory characteristics of statins. Given their under-prescription, adequate prescription of statins may limit the disease burden of NAFLD. Funding: ZonMw, KWF, NWO, SLO, DGXII, RIDE, National and regional government, Erasmus MC and Erasmus University.

Abstract Background Atrial fibrillation (AF) is associated with profound structural and functional changes in the atria. In the present study, we investigated the association between left atrial (LA) phasic function and the extent of LA fibrosis using advanced cardiovascular magnetic resonance (CMR) imaging techniques, including 3-dimensional (3D) late gadolinium enhancement (LGE) and feature tracking. Methods Patients with paroxysmal and persistent AF (n = 105) underwent CMR in sinus rhythm. LA global reservoir strain, conduit strain and contractile strain were derived from cine CMR images using CMR feature tracking. The extent of LA fibrosis was assessed from 3D LGE images. Healthy subjects underwent CMR and served as controls (n = 19). Results Significantly lower LA reservoir strain, conduit strain and contractile strain were found in AF patients, as compared to healthy controls (− 15.9 ± 3.8% vs. − 21.1 ± 3.6% P

Background: Acute acoustic trauma (AAT) is an acute hearing impairment caused by intense noise-impact. The current management strategy for AAT with substantial hearing loss in the Dutch military is the combination therapy with corticosteroids and hyperbaric oxygen therapy (HBOT). In a previous study, early initiation of the combination therapy was associated with better outcomes. Therefore, we performed a new analysis to assess the difference in hearing outcome between patients in whom combination therapy was started within two days, versus after more than two days. Methods: A retrospective analysis was performed on military patients diagnosed with AAT with substantial hearing loss who presented between February 2018 and March 2020. Absolute and relative hearing improvement between first and last audiograms were calculated for all affected frequencies (defined as loss of ≥20 dB on initial audiogram). We also determined the amount of patients who recovered to the level of Dutch military requirement, and performed speech discrimination tests. Results: In this analysis, 30 male patients (49 ears) with AAT were included. The median age was 24.5 years (IQR 23–29). The median time to initiation of therapy with corticosteroids and HBOT were one and two days, respectively. HBOT was started within two days in 31 ears, and after more than two days in 18 ears. The mean absolute and relative hearing gains were 18.8 dB (SD 14.6) and 46.8% (SD 31.3) on all affected frequencies. The 100% discrimination/speech perception level improved from 64.0 dB to 51.7 dB (gain 12.3 dB ± 14.1). There was significantly more improvement in absolute and relative hearing improvement when HBOT was started in ≤2 days, compared to >2 days. Conclusion: Our analysis shows results in favor of early initiation (≤2 days) of the combination treatment of HBOT and corticosteroids in patients with AAT.

Melting experiments on ultramafic rocks rich in the hydrous minerals phlogopite or phlogopite + K-richterite, some including 5% of accessory phases, have been conducted at 15 and 50 kbar. The assemblages represent probable source components that contribute to melts in cratonic regions, but whose melt compositions are poorly known. A main series of starting compositions based on MARID xenoliths consisted of a third each of clinopyroxene (CPX), phlogopite (PHL) and K-richterite (KR) with or without 5% ilmenite, rutile or apatite. Additional experiments were run without KR and with higher proportions of accessory phases. Melt traps were used at near-solidus temperatures to facilitate accurate analysis of well-quenched melts, for which reversal experiments demonstrate equilibrium.Results show that KR melts rapidly and completely within 50 °C of the solidus, so that melts reflect the composition of the amphibole and its melting reaction. Melts have high SiO2 and especially K2O but low CaO and Al2O3 relative to basaltic melts produced from peridotites at similar pressures. They have no counterparts amongst natural rocks, but most closely resemble leucite lamproites at 15 kbar. KR and PHL melt incongruently to form olivine (OL) and CPX at 15 kbar, promoting SiO2 contents of the melt, whereas orthopyroxene OPX is increasingly stable at lower lithosphere pressures, leading to an increase in MgO and decrease in SiO2 in melts, which resemble olivine lamproites. Melts of mica pyroxenites without KR are richer in CaO and Al2O3 and do not resemble lamproites. These experiments show that low CaO and Al2O3 in igneous rocks is not necessarily a sign of a depleted peridotite source. Accessory phases produce melts exceptionally rich in P2O5 or TiO2 depending on the phases present and are unlike any melts seen at the Earth’s surface, but may be important agents of metasomatism seen in xenoliths. The addition of the 5% accessory phases ilmenite, rutile or apatite result in melting temperatures a few ten of degrees lower; at least two of these appear essential to explain the compositions of many alkaline igneous rocks on cratons.Melting temperatures for CPX + PHL + KR mixtures are close to cratonic geotherms at depths > 130 km: minor perturbations of the stable geotherm at >150 km will rapidly lead to 20% melting. Melts of hydrous pyroxenites with a variety of accessory phases will be common initial melts at depth, but will change if reaction with wall-rocks occurs, leading to volcanism that contains chemical components of peridotite even though the temperature in the source region remains well below the melting point of peridotite. At higher temperatures, extensive melting of peridotite will dilute the initial alkaline melts: this is recognizable as alkaline components in basalts and, in extreme cases, alkali picrites. Hydrous pyroxenites are, therefore, components of most mantle-derived igneous rocks: basaltic rocks should not be oversimplified as being purely melts of peridotite or of mixtures of peridotite and dry pyroxenite without hydrous phases.

Objectives: Administrative claims data are prone to underestimate the burden of non-tuberculous mycobacterial pulmonary disease (NTM-PD). Methods: We developed machine learning-based algorithms using historical claims data from cases with NTM-PD to predict patients with a high probability of having previously undiagnosed NTM-PD and to assess actual prevalence and incidence. Adults with incident NTM-PD were classified from a representative 5% sample of the German population covered by statutory health insurance during 2011–2016 by the International Classification of Diseases, 10th revision code A31.0. Pre-diagnosis characteristics (patient demographics, comorbidities, diagnostic and therapeutic procedures, and medications) were extracted and compared to that of a control group without NTM-PD to identify risk factors. Results: Applying a random forest model (area under the curve 0.847; total error 19.4%) and a risk threshold of >99%, prevalence and incidence rates in 2016 increased 5-fold and 9-fold to 19 and 15 cases/100,000 population, respectively, for both coded and non-coded vs. coded cases alone. Conclusions: The use of a machine learning-based algorithm applied to German statutory health insurance claims data predicted a considerable number of previously unreported NTM-PD cases with high probabilty.

Summary: The presequence translocase (TIM23 complex) imports precursor proteins into the mitochondrial inner membrane and matrix. The presequence translocase-associated motor (PAM) provides a driving force for transport into the matrix. The J-protein Pam18 stimulates the ATPase activity of the mitochondrial Hsp70 (mtHsp70). Pam16 recruits Pam18 to the TIM23 complex to ensure protein import. The Pam16-Pam18 module also associates with components of the respiratory chain, but the function of the dual localization of Pam16-Pam18 is largely unknown. Here, we show that disruption of the Pam16-Pam18 heterodimer causes redistribution of Pam18 to the respiratory chain supercomplexes, where it forms a homodimer. Redistribution of Pam18 decreases protein import into mitochondria but stimulates mtHsp70-dependent assembly of respiratory chain complexes. We conclude that coupling to Pam16 differentially controls the dual function of Pam18. It recruits Pam18 to the TIM23 complex to promote protein import but attenuates the Pam18 function in the assembly of respiratory chain complexes.

Epigenetic aging biomarkers are associated with increased morbidity and mortality. We evaluated if occupational exposure to three established chemical carcinogens is associated with acceleration of epigenetic aging. We studied workers in China occupationally exposed to benzene, trichloroethylene (TCE) or formaldehyde by measuring personal air exposures prior to blood collection. Unexposed controls matched by age and sex were selected from nearby factories. We measured leukocyte DNA methylation (DNAm) in peripheral white blood cells using the Infinium HumanMethylation450 BeadChip to calculate five epigenetic aging clocks and DNAmTL, a biomarker associated with leukocyte telomere length and cell replication. We tested associations between exposure intensity and epigenetic age acceleration (EAA), defined as the residuals of regressing the DNAm aging biomarker on chronological age, matching factors and potential confounders. Median differences in EAA between exposure groups were tested using a permutation test with exact p-values. Epigenetic clocks were strongly correlated with age (Spearman r > 0.8) in all three occupational studies. There was a positive exposure-response relationship between benzene and the Skin-Blood Clock EAA biomarker: median EAA was −0.91 years in controls (n = 44), 0.78 years in workers exposed to

In this paper, the synthesis of dicalcium silicate (C2S) via sol-gel (acid-catalyzed) process including aqueous route, non-aqueous route, and the Pechini method is reported. The composition of C2S (α, β, and γ) polymorphs, by-products, and amorphous content is established by employing QXRD (quantitative X-ray diffraction) studies. The attention has been focused to comprehend assay-amorphous relationships of C2S yield. The intermediate dried gels have been investigated via thermal analysis to monitor changes in the gel structures and precursors at low temperatures. The synthetic parameters including calcination time and temperature, Ca/Si molar ratio and mode of cooling have been optimized to get pure β-C2S with low amorphous content. The dependency of β → γ C2S polymorphic transformation on mean crystallite size (Dcryst) is studied. Overall, the Pechini method exhibits the most promising results for the purity and tuning of β-C2S polymorph. Moreover, the non-aqueous and aqueous routes require calcining the dried gel at a temperature higher than 1200 ℃ due to the presence of CaO precursors as CaCO3. The theoretical calculations of amorphous content have revealed that the change in the stoichiometry from 2.0 to 1.7 Ca/Si ratio is not a viable solution to improve the C2S product yield.

Summary: Invaginations of the mitochondrial inner membrane, termed cristae, are hubs for oxidative phosphorylation. The mitochondrial contact site and cristae organizing system (MICOS) and the dimeric F1Fo-ATP synthase play important roles in controlling cristae architecture. A fraction of the MICOS core subunit Mic10 is found in association with the ATP synthase, yet it is unknown whether this interaction is of relevance for mitochondrial or cellular functions. Here, we established conditions to selectively study the role of Mic10 at the ATP synthase. Mic10 variants impaired in MICOS functions stimulate ATP synthase oligomerization like wild-type Mic10 and promote efficient inner membrane energization, adaptation to non-fermentable carbon sources, and respiratory growth. Mic10's functions in respiratory growth largely depend on Mic10ATPsynthase, not on Mic10MICOS. We conclude that Mic10 plays a dual role as core subunit of MICOS and as partner of the F1Fo-ATP synthase, serving distinct functions in cristae shaping and respiratory adaptation and growth.

Introduction: To prevent and manage the societal and economic burden of occupational diseases (ODs), countries should develop strong prevention policies, health surveillance and registry systems. This study aims to contribute to the improvement of OD surveillance at national level as well as to identify priority actions in Turkey. Methods: The history and current status of occupational health studies were considered from the perspective of OD surveillance. Interpretative research was done through literature review on occupational health at national, regional and international level. Analyses were focused on countries’ experiences in policy development and practice, roles and responsibilities of institutions, multidisciplinary and intersectoral collaboration. OD surveillance models of Turkey, Belgium and the Netherlands were examined through exchange visits. Face-to-face interviews were conducted to explore the peculiarities of legislative and institutional structures, the best and worst practices, and approach principles. Results: Some countries are more focused on exploring OD trends through effective and cost-efficient researches, with particular attention to new and emerging ODs. Other countries try to reach every single case of OD for compensation and rehabilitation. Each practice has advantages and shortcomings, but they are not mutually exclusive, and thus an effective combination is possible. Conclusion: Effective surveillance and registry approaches play a key role in the prevention of ODs. A well-designed system enables monitoring and assessment of OD prevalence and trends, and adoption of preventive measures while improving the effectiveness of redressing and compensation. A robust surveillance does not only provide protection of workers’ health but also advances prevention of economic losses. Keywords: Intersectoral collaboration, Occupational diseases, Occupational health, Prevention, Worker’s health surveillance

Green spaces play an important role in urban areas. We study the accessibility of green urban areas by combining open data sets about green with population size data. We develop a mathematical model to define the population density of a green area and calculate the available green space depending on the location. To this end, we do not only consider walking distance to and size of the green area, but also take into account the local population size. Our model quantifies how the available green space depends on the location in the city, such that heavily populated areas have a small amount of green available, even when closely located to a green area.

Introduction: Therapeutic options for diffuse malignant peritoneal mesothelioma (DMPM) are limited to surgery and locoregional chemotherapy. Despite improvements in survival rates, patients eventually succumb to disease progression. We investigated splicing deregulation both as molecular prognostic factor and potential novel target in DMPM, while we tested modulators of SF3b complex for antitumor activity. Methods: Tissue-microarrays of 64 DMPM specimens were subjected to immunohistochemical assessment of SF3B1 expression and correlation to clinical outcome. Two primary cell cultures were used for gene expression profiling and in vitro screening of SF3b modulators. Drug-induced splicing alterations affecting downstream cellular pathways were detected through RNA sequencing. Ultimately, we established bioluminescent orthotopic mouse models to test the efficacy of splicing modulation in vivo. Results: Spliceosomal genes are differentially upregulated in DMPM cells compared to normal tissues and high expression of SF3B1 correlated with poor clinical outcome in univariate and multivariate analysis. SF3b modulators (Pladienolide-B, E7107, Meayamycin-B) showed potent cytotoxic activity in vitro with IC50 values in the low nanomolar range. Differential splicing analysis of Pladienolide-B-treated cells revealed abundant alterations of transcripts involved in cell cycle, apoptosis and other oncogenic pathways. This was validated by RT-PCR and functional assays. E7107 demonstrated remarkable in vivo antitumor efficacy, with significant improvement of survival rates compared to vehicle-treated controls. Conclusions: SF3B1 emerged as a novel potential prognostic factor in DMPM. Splicing modulators markedly impair cancer cell viability, resulting also in potent antitumor activity in vivo. Our data designate splicing as a promising therapeutic target in DMPM. Keywords: Mesothelioma, Therapy, SF3b modulation, Splicing, RNA-sequencing

ITER foresees applying laser induced breakdown spectroscopy (LIBS) as a tool for quantitative assessment of fuel retention in the first walls. One open problem related to LIBS application is the choice of the laser type. Here we compare two Nd/YAG lasers with different pulse durations, 0.15 and 8 ns, working at λ = 1064 nm for LIBS studies of samples with D-doped W/Al coatings of ≈ 3 μm thickness (Al is used as a proxy of Be) on Mo. Low pressure argon was used as a background gas. Experiments were done in conditions where other factors (broadening of spectral lines, signal-to-noise ratio, limited thickness of coatings etc.) did not shadow the effect of laser pulse duration. For these reasons, low pressure argon was used as the background gas and fluences were kept at comparatively low values. Spectra of laser-produced plasma were recorded as a function of the number of laser pulses.Partially overlapping lines of hydrogen isotopes were fitted with Voigt contours, intensities were fitted and depth profiles of deuterium were reconstructed. The relative standard error of curve-fitting of spectra recorded with the laser of shorter pulse duration was two times smaller than that recorded by the longer pulse laser. The electron density was found from the Stark broadening of Hα line of the laser-produced plasma and the electron temperature of plasma was found on the basis W and Mo lines.It was found that in the case of ps laser an acceptable accuracy of the detection of deuterium was possible at considerably lower values of fluence. Steps needed for comparison of ps and ns lasers in ITER-relevant conditions were discussed. Keywords: ITER-relevant coatings, LIBS diagnostics, Detection of hydrogen isotopes, Elemental depth profiles, Temperature of laser-produced plasma

Study region: Kaleya River Catchment in southern Zambia. Study focus: The ability of a landscape hydrology approach to detect controls on water availability in a fragmented landscape to inform interventions under a changing environment was investigated. Simple and measurable climatic and landscape pattern attributes were analysed using change detection, trend analysis and backward variable elimination with Partial Least Squares Regression (PLSR) to identify controls on seasonal river flows and how landscape components could be enhanced to augment natural river flows. New hydrological insights for the region: Landscape pattern showed increasing fragmentation, expansion of irrigated cropland and reservoirs and loss of forestland. Significant increasing trends (p

Summary: Mitochondrial preproteins contain amino-terminal presequences directing them to the presequence translocase of the mitochondrial inner membrane (TIM23 complex). Depending on additional downstream import signals, TIM23 either inserts preproteins into the inner membrane or translocates them into the matrix. Matrix import requires the coupling of the presequence translocase-associated motor (PAM) to TIM23. The molecular mechanisms coordinating preprotein recognition by TIM23 in the intermembrane space (IMS) with PAM activation in the matrix are unknown. Here we show that subsequent to presequence recognition in the IMS, the Tim50 matrix domain facilitates the recruitment of the coupling factor Pam17. Next, the IMS domain of Tim50 promotes PAM recruitment to TIM23. Finally, the Tim50 transmembrane segment stimulates the matrix-directed import-driving force exerted by PAM. We propose that recognition of preprotein segments in the IMS and transfer of signal information across the inner membrane by Tim50 determine import motor activation. : Caumont-Sarcos et al. report a transmembrane coordination mechanism that promotes import of presequence-containing preproteins into mitochondria. The presequence translocase subunit Tim50 links the preprotein receptor function on the cis side of the mitochondrial inner membrane to functional activation of the import motor on the trans side. Keywords: mitochondria, preprotein import, presequence translocase, import motor, TIM23, Tim50, mitochondrial Hsp70, Pam17

Introduction: The co-occurrence of inflammatory bowel disease (IBD) in up to 80% of patients with primary sclerosing cholangitis (PSC) suggests a relation between the gut and the liver in patients with both PSC and IBD. One hypothesis suggests that aberrantly expressed homing molecules in the liver drive infiltration of gut-homing memory T-cells that are originally primed in intestinal environment. One of the main findings supporting this hypothesis is the expression of mucosal addressin cell adhesion molecule 1 (MAdCAM-1) in PSC livers. Expression of homing molecules in early PSC remains unclear. The aim of this study was to investigate expression patterns of homing chemokines and adhesion molecules in PSC-IBD colons and livers, and to study whether changes are already present in early stages of PSC. Methods: Needle biopsies from livers of 20 PSC patients with short-term PSC (PSC-IBDST) as well as explant liver biopsies of 8 patients with long-term PSC (PSC-IBDLT) were collected (median disease duration 0 and 22 years, respectively). Only patients with concomitant IBD were included (89% ulcerative colitis and 11% Crohn’s disease). Expression and distribution of MAdCAM-1, VAP-1, integrin β7, CCL25, CCL28, CXCL12, αE (CD103) and E-cadherin were assessed in both liver and colon tissue. Liver tissue collected from obstructive cholangitis in resection specimens for Klatskin tumors or resection specimens from hepatic metastasis, liver tissue of patients with hepatitis C virus (HCV) and of patients with primary biliary cholangitis (PBC) served as controls. Results: MAdCAM-1 expression in livers of PSC-IBDLT patients was increased compared to controls. The proportion of CD3+ T-cells expressing integrin β7 did not differ between PSC-IBDST and control groups, but was higher in liver tissue of PSC-IBDLT patients. There was no difference in αE+ T-cells between PSC-IBDLT and control groups. The chemokine CCL28 was highly expressed in biliary epithelial cells. This intense staining pattern was more pronounced in PSC-IBDST, but overall did not significantly differ from controls. Conclusions: We confirm that aberrant gut lymphocyte homing to the liver exists in PSC, linking gut and liver disease pathology in PSC-IBD. Our data suggests that this phenomenon increases over time in later stages of the disease, worsening ongoing inflammation.

Dravet syndrome (DS) is an early onset refractory epilepsy typically caused by de novo heterozygous variants in SCN1A encoding the α-subunit of the neuronal sodium channel Nav1.1. The syndrome is characterized by age-related progression of seizures, cognitive decline and movement disorders. We hypothesized that the distinct neurodevelopmental features in DS are caused by the disruption of molecular pathways in Nav1.1 haploinsufficient cells resulting in perturbed neural differentiation and maturation. Here, we established DS-patient and control induced pluripotent stem cell derived neural progenitor cells (iPSC NPC) and GABAergic inter-neuronal (iPSC GABA) cells. The DS-patient iPSC GABA cells showed a shift in sodium current activation and a perturbed response to induced oxidative stress. Transcriptome analysis revealed specific dysregulations of genes for chromatin structure, mitotic progression, neural plasticity and excitability in DS-patient iPSC NPCs and DS-patient iPSC GABA cells versus controls. The transcription factors FOXM1 and E2F1, positive regulators of the disrupted pathways for histone modification and cell cycle regulation, were markedly up-regulated in DS-iPSC GABA lines. Our study highlights transcriptional changes and disrupted pathways of chromatin remodeling in Nav1.1 haploinsufficient GABAergic cells, providing a molecular framework that overlaps with that of neurodevelopmental disorders and other epilepsies.

Summary: The two related members of the vasohibin family, VASH1 and VASH2, encode human tubulin detyrosinases. Here we demonstrate that, in contrast to VASH1, which requires binding of small vasohibin binding protein (SVBP), VASH2 has autonomous tubulin detyrosinating activity. Moreover, we demonstrate that SVBP acts as a bona fide activator of both enzymes. Phylogenetic analysis of the vasohibin family revealed that regulatory diversification of VASH-mediated tubulin detyrosination coincided with early vertebrate evolution. Thus, as a model organism for functional analysis, we used Trypanosoma brucei (Tb), an evolutionarily early-branched eukaryote that possesses a single VASH and encodes a terminal tyrosine on both α- and β-tubulin tails, both subject to removal. Remarkably, although detyrosination levels are high in the flagellum, TbVASH knockout parasites did not present any noticeable flagellar abnormalities. In contrast, we observed reduced proliferation associated with profound morphological and mitotic defects, underscoring the importance of tubulin detyrosination in cell division. : van der Laan et al. describe the evolutionary diversification of the VASH-mediated mechanism of tubulin detyrosination. In addition, they show that this modification plays an important role in Trypanosoma parasites, where it regulates cell division and cell morphology. Keywords: detyrosination, cell cycle, activator, peptide, inhibitor, parasite, tubulin, modification, vash, microtubule

Laser-induced breakdown spectroscopy (LIBS) is a promising method for quantifying the fuel content of the plasma-facing components of ITER both in between plasma discharges (in-situ) and after maintenance operations. The aim of the present study is to test the applicability of in-situ LIBS for monitoring deuterium (D) and helium (He) content of W samples exposed to fusion relevant plasma fluxes in the linear plasma device Pilot-PSI.The D loading was performed during 1000s of plasma exposure at low (200-300 °C) surface temperatures. Despite of low intensity and noisy LIBS spectra, H and D lines, at 656.1 and 656.3nm, respectively, could be fitted with Lorentzian contours and reliably resolved at 1.2 mbar background pressure of argon.In the case of He loading, the samples were also exposed to plasma during 1000s while the surface temperature reached values up to 720 °C at the center. Already at 10–2 mbar residual pressure of the device, the He I line at 587.6nm was visible for the first 2–3 laser shots.We demonstrated that in-situ LIBS is a reliable method for detection of He and D retention in ITER-relevant materials. Nevertheless, for measuring relative and absolute concentrations of D and He in the ITER-relevant samples, further studies are needed. Keywords: Plasma surface interaction, LIBS, Fuel retention, Tungsten

The ablation and fuel-retention characteristics of aluminum–tungsten (AlW) and beryllium–tungsten (BeW) samples have been determined using Laser Induced Breakdown Spectroscopy (LIBS) and compared to results obtained using Secondary Ion Mass Spectrometry (SIMS). The measurements have been made both at vacuum (of the order of 10−4Pa) and at 50Pa of argon to especially enhance the intensities of the spectral lines of H and D. For reliable evaluation of the ablation rate of the samples the electron density ne and temperature Te of the LIBS plasma have been determined with the help of selected of spectral lines of Be, Al, and W. The electron density ne has been obtained from Stark broadening lines of Al I (308.7nm and 394.4nm) and Be I (457.3nm) and Te from the Saha–Boltzmann plot using W I and W II spectral lines having a higher value of the energy of upper states in order to prevent the influence of self-absorption on the results. The results indicate similar ablation characteristics between AlW (AlWD) and BeW (BeWD) samples but the inclusion of deuterium in the coating increases the ablation rate by a factor of 10 for both sample types. Concerning fuel retention more than one order of magnitude less D is retained in the AlWD sample than in BeWD. In the presence of background argon, the H and D lines were stronger and more easily distinguishable. This is a positive sign considering the real application in ITER where LIBS measurements are foreseen to be done during maintenance breaks. However the higher pressure gave a better signal, it is still far from the measurement conditions planned for ITER which need to be tested separately. Keywords: LIBS, Beryllium, Aluminum

Mitochondria perform central functions in cellular bioenergetics, metabolism, and signaling, and their dysfunction has been linked to numerous diseases. The available studies cover only part of the mitochondrial proteome, and a separation of core mitochondrial proteins from associated fractions has not been achieved. We developed an integrative experimental approach to define the proteome of east mitochondria. We classified > 3,300 proteins of mitochondria and mitochondria-associated fractions and defined 901 high-confidence mitochondrial proteins, expanding the set of mitochondrial proteins by 82. Our analysis includes protein abundance under fermentable and nonfermentable growth, submitochondrial localization, single-protein experiments, and subcellular classification of mitochondria-associated fractions. We identified mitochondrial interactors of respiratory chain supercomplexes, ATP synthase, AAA proteases, the mitochondrial contact site and cristae organizing system (MICOS), and the coenzyme Q biosynthesis cluster, as well as mitochondrial proteins with dual cellular localization. The integrative proteome provides a high-confidence source for the characterization of physiological and pathophysiological functions of mitochondria and their integration into the cellular environment.