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Abstract Wolfram syndrome (WS) is a rare childhood disease characterized by diabetes mellitus, diabetes insipidus, blindness, deafness, neurodegeneration and eventually early death, due to autosomal recessive mutations in the WFS1 (and WFS2) gene. While it is categorized as a neurodegenerative disease, it is increasingly becoming clear that other cell types besides neurons may be affected and contribute to the pathogenesis. MRI studies in patients and phenotyping studies in WS rodent models indicate white matter/myelin loss, implicating a role for oligodendroglia in WS-associated neurodegeneration. In this study, we sought to determine if oligodendroglia are affected in WS and whether their dysfunction may be the primary cause of the observed optic neuropathy and brain neurodegeneration. We demonstrate that 7.5-month-old Wfs1 ∆exon8 mice display signs of abnormal myelination and a reduced number of oligodendrocyte precursor cells (OPCs) as well as abnormal axonal conduction in the optic nerve. An MRI study of the brain furthermore revealed grey and white matter loss in the cerebellum, brainstem, and superior colliculus, as is seen in WS patients. To further dissect the role of oligodendroglia in WS, we performed a transcriptomics study of WS patient iPSC-derived OPCs and pre-myelinating oligodendrocytes. Transcriptional changes compared to isogenic control cells were found for genes with a role in ER function. However, a deep phenotyping study of these WS patient iPSC-derived oligodendroglia unveiled normal differentiation, mitochondria-associated endoplasmic reticulum (ER) membrane interactions and mitochondrial function, and no overt signs of ER stress. Overall, the current study indicates that oligodendroglia functions are largely preserved in the WS mouse and patient iPSC-derived models used in this study. These findings do not support a major defect in oligodendroglia function as the primary cause of WS, and warrant further investigation of neurons and neuron-oligodendroglia interactions as a target for future neuroprotective or -restorative treatments for WS.

The machine perfusion (MP) of transplantable grafts has emerged as an upcoming field in Cardiothoracic (CT) transplantation during the last decade. This technology carries the potential to assess, preserve, and even recondition thoracic grafts before transplantation, so it is a possible game-changer in the field. This technology field has reached a critical turning point, with a growing number of publications coming predominantly from a few leading institutions, but still need solid scientific evidence. Due to the increasing need to expand the donor pool, especially in Europe, where the donor age is steeply increased, a consensus has been established to address the growing need and knowledge of machine perfusion in cardiothoracic transplantation, targeting the unmet scientific need in this growing field but also, priorities for development, and regional differences in utilization rates and organizational issues. To address MP in CT, the European Society of Organ Transplantation (ESOT) convened a dedicated Working group comprised of experts in CT to review literature about MP to develop guidelines that were subsequently discussed and voted on during the Consensus Conference that took place in person in Prague during the TLJ 3.0 in November 2022. The findings and recommendations of the Cardiothoracic Working Group on MP are presented in this article.

Background: Donor organ shortage is a barrier to lung transplantation. Donation after circulatory death (DCD) may offer a solution, although it is underutilized. The objective of this study was to compare survival and other postoperative outcomes between DCD and donation after brain death (DBD). Methods: We performed a multicenter analysis of Multi-Institutional Extracorporeal Life Support (ECLS) Registry data from 11 lung transplant centers in the United States and Europe. Demographics and clinical parameters were compared using chi-square test and Fisher's exact test. Survival was assessed by Kaplan-Meier curves and compared by log-rank test with propensity score matching. Results: Of 1,585 patients included in the study, 135 (8.5%) received DCD lungs and 1,450 (91.5%) received DBD lungs. DCD recipients had higher rates of obstructive lung disease (p = 0.042), longer total ischemic time (p

ABSTRACT Accurately estimating effective population size (Ne) is essential for understanding evolutionary processes and guiding conservation efforts. This study investigates Ne estimation methods in spatially structured populations using a population of moor frog (Rana arvalis) as a case study. We assessed the behaviour of Ne estimates derived from the linkage disequilibrium (LD) method as we changed the spatial configuration of samples. Moor frog eggs were sampled from 25 breeding patches (i.e., separate vernal ponds, ditches or parts of larger fens) within a single population, revealing an isolation‐by‐distance pattern and a local spatial genetic structure. Varying buffer sizes around each patch were used to examine the impact of sampling window size on the estimation of effective number of breeders (Nb). Our results indicate a downward bias in LD Nb estimates with increasing buffer size, suggesting an underestimation of Nb. The observed bias is attributed to LD resulting from including genetically divergent individuals (mixture‐LD) confounding LD due to drift. This emphasises the significance of considering even subtle spatial genetic patterns. The implications of these findings are discussed, emphasising the need to account for spatial genetic structure to accurately assess population viability and inform conservation efforts. This study contributes to our understanding of the challenges associated with Ne estimation in spatially structured populations and underscores the importance of refining methodologies to address population‐specific spatial dynamics for effective conservation planning and management.

Long-term outcome following lung transplantation remains one of the poorest of all solid organ transplants with a 1- and 5-year survival of 85% and 59% respectively for adult lung transplant recipients and with 50% of patients developing chronic lung allograft dysfunction (CLAD) in the first 5 years following transplant. Reducing the risk of inflammatory type primary graft dysfunction (PGD) is vital for improving both short-term survival following lung transplantation and long-term outcome due to the association of early inflammatory-mediated damage to the allograft and the risk of CLAD. PGD has a multifactorial aetiology and high-grade inflammatory-type PGD is the result of cumulative insults that may be incurred in one or more of the three variables of the transplantation continuum: the donor lungs, the recipient and intraoperative process. We set out a conceptual framework which uses a fully integrated approach to this transplant continuum to attempt to identify and, where possible, modify specific donor, recipient and intraoperative PGD risk with the goal of reducing inflammatory-type PGD risk for an individual recipient. We also consider the concept and risk-benefit of matching lung allografts and recipients on the basis of donor and recipient PGD-risk compatibility. The use of ex vivo lung perfusion (EVLP) and the extended preservation of lung allografts on EVLP will be explored as safe, non-injurious EVLP may enable extensive inflammatory testing of specific donor lungs and has the potential to provide a platform for targeted therapeutic interventions on lung allografts.

IntroductionCD39 plays an important role in the immunoregulation and inhibition of effector cells. It is expressed on immune cells, including Tregs, and on extracellular vesicles (EVs) budding from the plasma membrane. Platelet transfusion may induce alloimmunization against HLA-I antigens, leading to refractoriness to platelet transfusion with severe consequences for patients. Tregs may play a key role in determining whether alloimmunization occurs in patients with hematologic disorders. We hypothesized that CD39+ EVs might play an immunoregulatory role, particularly in the context of platelet transfusions in patients with hematologic disorders. Such alloimmunization leads to the production of alloantibodies and is sensitive to the regulatory action of CD39.MethodsWe characterized CD39+ EVs in platelet concentrates by flow cytometry. The absolute numbers and cellular origins of CD39+ EVs were evaluated. We also performed functional tests to evaluate interactions with immune cells and their functions.ResultsWe found that CD39+ EVs from platelet concentrates had an inhibitory phenotype that could be transferred to the immune cells with which they interacted: CD4+ and CD8+ T lymphocytes (TLs), dendritic cells, monocytes, and B lymphocytes (BLs). Moreover, the concentration of CD39+ EVs in platelet concentrates varied and was very high in 10% of concentrates. The number of these EVs present was determinant for EV-cell interactions. Finally, functional interactions were observed with BLs, CD4+ TLs and CD39+ EVs for immunoglobulin production and lymphoproliferation, with potential implications for the immunological management of patients.

IntroductionCompared with traditional static ice storage, controlled hypothermic storage (CHS) at 4–10°C may attenuate cold-induced lung injury between procurement and implantation. In this study, we describe the first European lung transplant (LTx) experience with a portable CHS device.MethodsA prospective observational study was conducted of all consecutively performed LTx following CHS (11 November 2022 and 31 January 2024) at two European high-volume centers. The LUNGguard device was used for CHS. The preservation details, total ischemic time, and early postoperative outcomes are described. The data are presented as median (range: minimum–maximum) values.ResultsA total of 36 patients underwent LTx (i.e., 33 bilateral, 2 single LTx, and 1 lobar). The median age was 61 (15–68) years; 58% of the patients were male; 28% of the transplantations had high-urgency status; and 22% were indicated as donation after circulatory death. In 47% of the patients, extracorporeal membrane oxygenation (ECMO) was used for perioperative support. The indications for using the CHS device were overnight bridging (n = 26), remote procurement (n = 4), rescue allocation (n = 2), logistics (n = 2), feasibility (n = 1), and extended-criteria donor (n = 1). The CHS temperature was 6.5°C (3.7°C–9.3°C). The preservation times were 11 h 18 (2 h 42–17 h 9) and 13 h 40 (4 h 5–19 h 36) for the first and second implanted lungs, respectively, whereas the total ischemic times were 13 h 38 (4 h 51–19 h 44) and 15 h 41 (5 h 54–22 h 48), respectively. The primary graft dysfunction grade 3 (PGD3) incidence rates were 33.3% within 72 h and 2.8% at 72 h. Intensive care unit stay was 8 (4–62) days, and the hospital stay was 28 (13–87) days. At the last follow-up [139 (7–446) days], three patients were still hospitalized. One patient died on postoperative day 7 due to ECMO failure. In-hospital Clavien–Dindo complications of 3b were observed in six (17%) patients, and 4a in seven (19%).ConclusionCHS seems safe and feasible despite the high-risk recipient and donor profiles, as well as extended preservation times. PGD3 at 72 h was observed in 2.8% of the patients. This technology could postpone LTx to daytime working hours. Larger cohorts and longer-term outcomes are required to confirm these observations.

Objective: Tumour progression drives profound alterations in host metabolism, such as adipose tissue depletion, an early event of cancer cachexia. As fatty acid consumption by cancer cells increases upon acidosis of the tumour microenvironment, we reasoned that fatty acids derived from distant adipose lipolysis may sustain tumour fatty acid craving, leading to the adipose tissue loss observed in cancer cachexia. Methods: To evaluate the pro-lipolytic capacities of acid-exposed cancer cells, primary mouse adipocytes from subcutaneous and visceral adipose tissue were exposed to pH-matched conditioned medium from human and murine acid-exposed cancer cells (pH 6.5), compared to naive cancer cells (pH 7.4). To further address the role of tumoral acidosis on adipose tissue loss, a pH-low insertion peptide was injected into tumour-bearing mice, and tumoral acidosis was neutralised with a sodium bicarbonate buffer. Prolipolytic mediators were identified by transcriptomic approaches and validated on murine and human adipocytes. Results: Here, we reveal that acid-exposed cancer cells promote lipolysis from subcutaneous and visceral adipocytes and that dampening acidosis in vivo inhibits adipose tissue depletion. We further found a set of well-known prolipolytic factors enhanced upon acidosis adaptation and unravelled a role for β-glucuronidase (GUSB) as a promising new actor in adipocyte lipolysis. Conclusions: Tumoral acidosis promotes the mobilization of fatty acids derived from adipocytes via the release of soluble factors by cancer cells. Our work paves the way for therapeutic approaches aimed at tackling cachexia by targeting the tumour acidic compartment.

The gut microbiota makes critical contributions to host homeostasis, and its role in the treatment of acute myeloid leukaemia (AML) has attracted attention. We investigated whether the gut microbiome is affected by AML, and whether such changes are associated with cachectic hallmarks. Biological samples and clinical data were collected from 30 antibiotic-free AML patients at diagnosis and matched volunteers (1:1) in a multicenter cross-sectional prospective study. The composition and functional potential of the faecal microbiota were analyzed using shotgun metagenomics. Faecal, blood, and urine metabolomics analyses were performed. AML patients displayed muscle weakness, anorexia, signs of altered gut function, and glycaemic disorders. The composition of the faecal microbiota differed between patients with AML and control subjects, with an increase in oral bacteria. Alterations in bacterial functions and faecal metabolome support an altered redox status in the gut microbiota, which may contribute to the altered redox status observed in patients with AML. Eubacterium eligens, reduced 3-fold in AML patients, was strongly correlated with muscle strength and citrulline, a marker of enterocyte mass and function. Blautia and Parabacteroides, increased in patients with AML, were correlated with anorexia. Several bacterial taxa and metabolites (e.g. Blautia, Prevotella, phenylacetate, and hippurate) previously associated with glycaemic disorders were altered. Our work revealed important perturbations in the gut microbiome of AML patients at diagnosis, which are associated with muscle strength, altered redox status, and anorexia. These findings pave the way for future mechanistic work to explore the function and therapeutic potential of the bacteria identified in this study.

IntroductionPeople living with HIV (PLWH) now benefit from combined antiviral treatments that durably control viral replication. These antiretroviral treatments decrease mortality and improve quality of life in PLWH, but do not completely control the excessive non-specific activation of the immune system in PLWH. This chronic immune activation is a key element of HIV immunopathology that contributes to the pathophysiology of inflammatory comorbid conditions, such as cardiovascular disorders, cancer and autoimmune diseases. Circulating non-exosomal extracellular vesicles, also known as microparticles (MPs) are detected in these diseases and have been linked to immune activation. The objective of this study was to characterize the MPs present in PLWH and to assess their association with chronic immune activation.MethodsWe performed flow cytometry for the complete phenotypic characterization of MPs from fresh plasma from PLWH and from people without HIV as the control group. The absolute number, size and cellular origin of MPs were evaluated. The immunoregulatory profile was determined by cell origin, for MPs derived from platelets (PMPs), monocytes (MMPs) and T lymphocytes (LMPs).ResultsPLWH had significantly more circulating MPs than controls, for MPs of all sizes originating from T lymphocytes, red blood cells, neutrophils, dendritic cells, B lymphocytes and endothelial cells. PMPs and MMPs were not more numerous in PLWH, but the immunoregulatory phenotypes of these MPs differed between PLWH and controls. These differences in immunoregulatory molecule expression profile were also observed for LMPs. PDL1, ICOSL, CCR5, TGFβ1, MHC classes I and II, TRAIL, CXCR4, OX40, DC-SIGN, CTLA4 and PDL2 were more strongly expressed on the surface of MPs from PLWH than on those from controls.ConclusionMPs are an important element in intercellular communication, making it possible to transfer phenotypes and functions to immune cells. The significantly higher numbers of MPs expressing diverse immunomodulatory molecules in PLWH may make a major contribution to the maintenance and/or the development of immune-cell activation in these individuals.

Abstract The analysis of environmental DNA (eDNA) is becoming integrated as an established biomonitoring tool, often characterized by detection limits exceeding those of conventional counterparts. However, further improving the sensitivity of these methods may be invaluable for the early detection of invasive species, or for locating remnant populations of endangered species, and the adequate quantification of their abundances. In this study, we provide empirical evidence showing that the implementation of multiple genetic markers targeting different loci is a surprisingly overlooked strategy to increase the sensitivity of single‐species detections and quantification of their abundance, particularly at the lower end of the species abundance range. We analyzed 45 natural eDNA samples obtained from a wide range of water bodies in Belgium, in which either the invasive American bullfrog (Lithobates catesbeianus) or the rare European weather loach (Misgurnus fossilis) occurred under variable abundances, and compared detection success and precision of quantification of simplex (single locus) versus multiplex (multilocus) droplet digital PCR (ddPCR) analyses. Multiplexing different primer/probe assays targeting independent loci resulted in a significantly enhanced detection probability compared to the simplex analyses, gaining a twofold reduction in the limit of detection (LOD). Also the precision of eDNA quantification significantly improved in multiplex reactions, especially in low concentration samples. This was reflected in a significant reduction in the coefficient of variation (CV) among technical replicates, resulting in an associated decrease of the limit of quantification (LOQ). We conclude that the use of multiple markers can significantly improve the analytical sensitivity of eDNA‐based single‐species detections and precision of absolute abundance quantifications.

Abstract Background The aryl hydrocarbon receptor (AHR) is expressed in the intestine and liver, where it has pleiotropic functions and target genes. This study aims to explore the potential implication of AHR in cancer cachexia, an inflammatory and metabolic syndrome contributing to cancer death. Specifically, we tested the hypothesis that targeting AHR can alleviate cachectic features, particularly through the gut–liver axis. Methods AHR pathways were explored in multiple tissues from four experimental mouse models of cancer cachexia (C26, BaF3, MC38 and APCMin/+) and from non‐cachectic mice (sham‐injected mice and non‐cachexia‐inducing [NC26] tumour‐bearing mice), as well as in liver biopsies from cancer patients. Cachectic mice were treated with an AHR agonist (6‐formylindolo(3,2‐b)carbazole [FICZ]) or an antibody neutralizing interleukin‐6 (IL‐6). Key mechanisms were validated in vitro on HepG2 cells. Results AHR activation, reflected by the expression of Cyp1a1 and Cyp1a2, two major AHR target genes, was deeply reduced in all models (C26 and BaF3, P

AbstractInvestigation of environmental DNA (eDNA) is increasingly used to precisely and non‐invasively detect and monitor pathogens. Among these, Tetracapsuloides bryosalmonae is a myxozoan endoparasite that causes proliferative kidney disease (PKD) in salmonid fish. Although the detection of T. bryosalmonae DNA in water samples has been shown to be promising and successful, method comparison and cross‐validation are currently lacking. This study aims to directly compare the sensitivity of different eDNA‐based methods in field and laboratory applications, and to develop an easy‐to‐apply and sensitive protocol to monitor T. bryosalmonae occurrence non‐invasively by its eDNA in water samples. First, we tested three existing probe‐based T. bryosalmonae‐specific detection assays in parallel by comparing the limit of detection (LOD) and limit of quantification (LOQ) using quantitative PCR (qPCR) and digital droplet PCR (ddPCR) platforms. Second, the impact of different filter types and water volumes on the detection probability was tested by sampling water directly from riverbanks with a syringe‐based protocol. The most sensitive detection protocol was the combination of the probe‐based assay published by Bettge et al. run via ddPCR, resulting in a LOD of 1.65 copies/μL input (6.6 copies/reaction) and a LOQ of 3.66 copies/μL input (14.67 copies/reaction). The type of filter (Sterivex™ compared to Millex®) did not significantly influence detection probability, however, the volume of water sampled (600 mL compared to 300 mL) significantly affected the probability of capturing eDNA in a sample. Based on modeled probabilities of eDNA capture and detection, we calculated that using the Bettge et al. assay via the ddPCR platform for data collection, 95% overall detection probability could be achieved with three replicates of 600 mL filtered water with Sterivex™ filters. Based on this cross‐validation of assays and detection platforms, we provide a cost‐effective, straightforward, and highly sensitive laboratory analysis workflow to detect DNA of T. bryosalmonae from water samples.