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The incorporation of nuclear quadrupole interaction into electron spin-echo modulation spectra by second-order perturbation methods has been developed for both two-pulse and three-pulse echo sequences. The results have been compared with exact numerical diagonalization of the Hamiltonian and with a first-order perturbation treatment. Model calculations have been carried out for deuterium (I=1) and aluminum (I=5/2) nuclei. It is shown that the second-order expressions can be used to obtain relatively accurate values for the number and distance of interacting nuclei at electron–nuclear distances greater than 0.26 nm. The procedure is more limited when the quadrupolar interaction exceeds the dipolar interaction when neither can be neglected. [ABSTRACT FROM AUTHOR]

Fibrates, which are agonists of peroxisome proliferator-activated receptor alpha, have received increasing attention in the treatment of primary biliary cholangitis. Reduced alkaline phosphatase levels and improved clinical outcomes were observed in patients with primary biliary cholangitis with an inadequate response to ursodeoxycholic acid (UDCA) monotherapy4 when treated with bezafibrate or fenofibrate combined with UDCA. In contrast to obeticholic acid, which exacerbates pruritus in patients, fibrates have been shown to relieve pruritus. Clinical trial outcomes show potential for the treatment of primary biliary cholangitis by targeting peroxisome proliferator-activated receptors. It is currently agreed that primary biliary cholangitis is an autoimmune-mediated cholestatic liver disease, and peroxisome proliferator-activated receptor is a nuclear receptor that regulates the functions of multiple immune cells, thus playing an important role in regulating innate and adaptive immunity. Therefore, this review focuses on the immune disorder of primary biliary cholangitis and summarizes the regulation of hepatic immunity when peroxisome proliferator-activated receptors are targeted for treating primary biliary cholangitis.

Liver fibrosis (LF) is a major disease that threatens human health. Hepatic stellate cells (HSCs) contribute directly to LF via extracellular matrix (ECM) secretion. Moreover, RXRα is an important nuclear receptor that plays a key regulatory role in HSC activation. Meanwhile, microRNAs (miRNAs) have been identified as significant regulators of LF development. In particular, miR-654-5p is involved in cellular migration and proliferation, and via bioinformatics analysis, has been identified as a potential factor that targets RXRα in humans and in mice. However, the precise relationship between miR-654-5p and RXRα in the context of LF, remains unknown and is the primary focus of the current study. To establish in vitro activated cell model human primary HSCs were cultured in vitro and LX-2 cells were stimulated with recombinant human TGF-β1. mRNA and protein levels of RXRα, miR-654-5p and fibrogenic genes were compared in quiescent and activated HSCs. Moreover, after transfected with miR-654-5p mimics, the expression changes of above related genes in LX-2 cells were estimated. Meanwhile, cell proliferation and apoptosis were detected in miR-654-5p overexpressed LX-2 cells. Simultaneously, the targeted binding between miR-654-5p and RXRα was verified in LX-2 cells. Carbon tetrachloride (CCl4)-induced mouse model with liver fibrosis was use to research the role of the miR-654-5p in vitro. Our results show that miR-654-5p expression levels increased in activated human HSCs and TGFβ-treated LX-2 cells. Moreover, miR-654-5p mimics markedly promoted LX-2 cell proliferation while inhibiting their apoptosis. Accordingly, the expression levels of RXRα are decreased in activated HSCs and LX-2 cells. Additionally, dual-luciferase reporter assay results reveal direct targeting of RXRα by miR-654-5p. Similarly, in vivo miR-654-5p overexpression aggravates LF in mice that are intraperitoneally injected with CCl4. Taken together, our findings elucidated a novel molecular mechanism with potential use for treatment of LF.

Abstract Background Hepatitis C virus (HCV) infection is a public health issue for which an effective universal screening method is urgently needed. An oral anti-HCV test could provide a noninvasive and rapid screening strategy for HCV infection. This study evaluated the performance of a new point-of-care oral assay developed by Well for the detection of HCV antibody. Methods Individuals from three centers with and without HCV infection were enrolled. All participants were tested for oral HCV antibody using the Well assay and for serum HCV antibody using established tests (ARCHITECT i2000 anti-HCV assay and InTec serum anti-HCV assay). For participants who obtained positive results, HCV RNA was tested for verification. Some patients underwent the OraQuick HCV test at the same time, and some self-tested with the Well assay during the same period. Results A total of 1179 participants, including 486 patients with chronic HCV infection, 108 patients with other liver diseases, and 585 individuals who underwent physical examination, were enrolled. The Well anti-HCV test had a sensitivity of 91.88% (95% confidence interval [CI]: 88.97–94.09%) and a specificity of 98.00% (96.58–98.86%) for oral HCV antibody detection. The consistency between the Well and InTec assays was 97.02% (1138/1179). The consistency between the Well and OraQuick assays was 98.50% (197/200). Furthermore, the results of self-testing were highly consistent with those of researcher-administered tests (Kappa = 0.979). In addition, the HCV RNA results also showed that HCV RNA could only be detected on 1 of the 39 false-negative samples, and for 172 positive HCV RNA results, 171 could be detected by the Well oral anti-HCV assay. Conclusions The Well oral anti-HCV test offers high sensitivity and specificity and performed comparably to both the OraQuick assay and InTec assay for HCV diagnosis. Thus, the Well test represents a new tool for universal HCV screening to identify infected patients, particularly in regions with limited medical resources.

MicroRNAs (miRNAs) participate in hepatic stellate cell (HSC) activation, which drives liver fibrosis initiation and progression. We aimed to identify novel hepatic fibrosis targets using miRNA sequencing (miRNA-seq) of human primary HSCs. Surgically resected liver tissues were used to extract HSCs. Based on next-generation sequencing, miRNA-seq was performed on four pairs of HSCs before and after in vitro culture. Additionally, we compared our data with open access miRNA-seq data derived from fourteen cirrhotic and nine healthy liver tissues. Selected miRNAs associated with fibrosis were verified by quantitative real-time PCR. Target mRNAs of differentially expressed (DE) miRNAs were predicted to construct co-expression networks. We identified 230 DEmiRNAs (118 upregulated and 112 downregulated) upon HSC activation. Of the 17 miRNAs with the most significant differences in expression, liver disease-related miRNAs included miR-758-3p, miR-493-5p, miR-409-3p, miR-31-5p, miR-1268a, and miR-381-3p, which might play roles in hepatic fibrosis. Moreover, let-7g-5p, miR-107, miR-122-5p, miR-127-3p, miR-139-5p, miR-148a-3p, miR-194-5p, miR-215-5p, miR-26a-5p, miR-340-5p, miR-451a, and miR-99a-5p were common between our data and the publicly available sequencing data. A co-expression network comprising 1891 matched miRNA–mRNA pairs representing 138 DEmiRNAs and 1414 DEmRNAs was constructed. MiR-1268a and miR-665, possessing the richest target DEmRNAs, may be vital in HSC activation. The targeted genes were involved in collagen metabolism, extracellular matrix structural constituent, cytoskeletal protein binding, and cell adhesion. The miRNAs we identified may provide a basis and reference for the selection of diagnostic and therapeutic targets for hepatic fibrosis.

Background/Aims: The present study aims to explore the protective role and mechanism of ginsenoside Rg1 combined with bone marrow mesenchymal stem cell (BMSC) transplantation for cerebral ischemia reperfusion injury (CIRI) in rat brain. Methods: One hundred twenty male SD rats were randomly divided into a sham group, an Ischemia Reperfusion (IR) group, an IR group treated with BMSC transplantation (IR+BMSCs), an IR group treated with Rg1 (IR+Rg1), and an IR group treated with BMSC transplantation and Rg1 (IR+Rg1+BMSCs). To establish a CIRI model, right middle cerebral artery embolization was used. The neurological score, 2,3,5-triphenyltet-razolium chloride monohydrate (TTC) staining and brain water content were detected to assess the treatment efficiency. HE staining and TUNEL were used to explore the pathologic changes and apoptosis. To explore the protein levels of neuron-specific enolase (NSE) and glial fibrillary acidic protein (GFAP), immunofluoresence was utilized. Western blotting was used to explore apoptosis-related proteins such as Bcl-2 and Bax. Results: Compared with the sham group, the IR group demonstrated obvious ischemic changes, such as significant neurologic defects and enhanced brain water content. The Rg1 treatment resulted in an obvious decrease in cell apoptosis and improved ischemic conditions. By BMSC transplantation, the transplanted cells could be differentiated into neurons and glial cells, which also improved cerebral ischemia. More importantly, the IR+Rg1+BMSCs group showed the best treatment efficiency with reduced cell apoptosis and better cerebral recovery. Conclusions: The Rg1 treatment resulted in an obvious decrease in cell apoptosis, while the transplanted cells could be differentiated into neurons and glial cells, which also improved cerebral ischemia.

Glycerine is a by-product of the biodiesel industry. Owing to the exponential growth of biodiesel production worldwide and particularly in Europe, the glycerine market has been driven during the last few years by the oversupply being generated by the biodiesel industry. Prices of both refined and crude glycerine have fallen to all-time lows. Consequently, there are clear signs that demand for glycerine is growing strongly with new usages both for crude and refined glycerine now to the fore. This article attempts to analyse to what extent a « substitution/new-usage breakthrough » for glycerine is occurring and will propose a supply/demand scenario for the coming 3 years. It will also attempt to predict how glycerine prices will develop over the same period.

Background and Objective: Flow cytometry is a widely used technique for identifying cell populations in patient-derived fluids, such as peripheral blood (PB) or cerebrospinal fluid (CSF). Despite its ubiquity in research and clinical practice, the process of gating, i.e., manually identifying cell types, is labor-intensive and error-prone. The objective of this study is to address this challenge by introducing GateNet, a neural network architecture designed for fully end-to-end automated gating without the need for correcting batch effects., Methods: For this study a unique dataset is used which comprises over 8,000,000 events from N = 127 PB and CSF samples which were manually labeled independently by four experts. Applying cross-validation, the classification performance of GateNet is compared to the human experts performance. Additionally, GateNet is applied to a publicly available dataset to evaluate generalization. The classification performance is measured using the F1 score., Results: GateNet achieves F1 scores ranging from 0.910 to 0.997 demonstrating human-level performance on samples unseen during training. In the publicly available dataset, GateNet confirms its generalization capabilities with an F1 score of 0.936. Importantly, we also show that GateNet only requires ≈10 samples to reach human-level performance. Finally, gating with GateNet only takes 15 microseconds per event utilizing graphics processing units (GPU)., Conclusions: GateNet enables fully end-to-end automated gating in flow cytometry, overcoming the labor-intensive and error-prone nature of manual adjustments. The neural network achieves human-level performance on unseen samples and generalizes well to diverse datasets. Notably, its data efficiency, requiring only ∼10 samples to reach human-level performance, positions GateNet as a widely applicable tool across various domains of flow cytometry., Competing Interests: Declaration of competing interest We, the undersigned, confirm that the manuscript represents our own work, is original and has not been copyrighted, published, submitted, or accepted for publication elsewhere. We further confirm that we all have fully read the manuscript and give consent to be co-authors of the manuscript., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Introduction: Pathogenic variants in the XK gene are associated with dysfunction or loss of XK protein leading to McLeod syndrome (MLS), a rare X-linked neuroacanthocytosis syndrome with multisystemic manifestation. Here we present clinical, genetic and immunological data on a patient originally admitted to our clinic for presumed post-COVID-19 syndrome, where thorough clinical work-up revealed a novel frameshift deletion in XK causal for the underlying phenotype. We additionally review the clinicogenetic spectrum of reported McLeod cases in the literature. Methods: We performed in-depth clinical characterization and flow cytometry of cerebrospinal fluid (CSF) in a patient where multi-gene panel sequencing identified a novel hemizygous frameshift deletion in XK . Additionally, Kell (K) and Cellano (k) antigen expression was analysed by Fluorescence-activated Cell Sorting (FACS). KEL gene expression was examined by RNA sequencing. Results: A novel hemizygous frameshift deletion in the XK gene resulting in premature termination of the amino acid chain was identified in a 46-year old male presenting with decrease in physical performance and persisting fatigue after COVID-19 infection. Examinations showed raised creatine kinase (CK) levels, neuropathy and clinical features of myopathy. FACS revealed the K-k+ blood type and reduced Cellano density. CSF flow cytometry showed elevation of activated T Cells. Conclusion: In-depth clinical, genetic, immunological and ribonucleic acid (RNA) expression data revealed axonal neuropathy, myopathy and raised levels of activated CSF-T-lymphocytes in a patient with a previously unpublished frameshift deletion in the XK gene., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Dambietz, Doescher, Heming, Schirmacher, Schlüter, Schulte-Mecklenbeck, Thomas, Wiendl, Meyer zu Hörste and Wiethoff.)

Cryptococcal meningitis (CM) is a severe fungal disease in immunocompromised patients affecting the central nervous system (CNS). Host response and immunological alterations in the cerebrospinal fluid (CSF) after invasion of Cryptococcus neoformans to the central nervous system have been investigated before but rigorous and comprehensive studies examining cellular changes in the CSF of patients with cryptococccal meningitis are still rare. We retrospectively collected CSF analysis and flow cytometry data of CSF and blood in patients with CM (n = 7) and compared them to HIV positive patients without meningitis (n = 13) and HIV negative healthy controls (n = 7). Within the group of patients with CM we compared those with HIV infection (n = 3) or other immunocompromised conditions (n = 4). Flow cytometry analysis revealed an elevation of natural killer cells and natural killer T cells in the CSF and blood of HIV negative patients with CM, pointing to innate immune activation in early stages after fungal invasion. HIV positive patients with CM exhibited stronger blood-CSF-barrier disruption. Follow-up CSF analysis over up to 150 days showed heterogeneous cellular courses in CM patients with slow normalization of CSF after induction of antifungal therapy., (© 2024. The Author(s).)

Neuroinflammation and blood-cerebrospinal fluid barrier (BCB) disruption could be key elements in schizophrenia-spectrum disorderś(SSDs) etiology and symptom modulation. We present the largest two-stage individual patient data (IPD) meta-analysis, investigating the association of BCB disruption and cerebrospinal fluid (CSF) alterations with symptom severity in first-episode psychosis (FEP) and recent onset psychotic disorder (ROP) individuals, with a focus on sex-related differences. Data was collected from PubMed and EMBASE databases. FEP, ROP and high-risk syndromes for psychosis IPD were included if routine basic CSF-diagnostics were reported. Risk of bias of the included studies was evaluated. Random-effects meta-analyses and mixed-effects linear regression models were employed to assess the impact of BCB alterations on symptom severity. Published (6 studies) and unpublished IPD from n = 531 individuals was included in the analyses. CSF was altered in 38.8 % of individuals. No significant differences in symptom severity were found between individuals with and without CSF alterations (SMD = -0.17, 95 %CI -0.55-0.22, p = 0.341). However, males with elevated CSF/serum albumin ratios or any CSF alteration had significantly higher positive symptom scores than those without alterations (SMD = 0.34, 95 %CI 0.05-0.64, p = 0.037 and SMD = 0.29, 95 %CI 0.17-0.41p = 0.005, respectively). Mixed-effects and simple regression models showed no association (p > 0.1) between CSF parameters and symptomatic outcomes. No interaction between sex and CSF parameters was found (p > 0.1). BCB disruption appears highly prevalent in early psychosis and could be involved in positive symptomś severity in males, indicating potential difficult-to-treat states. This work highlights the need for considering BCB breakdownand sex-related differences in SSDs clinical trials and treatment strategies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

5q-associated spinal muscular atrophy (SMA) is a motoneuron disease caused by mutations in the survival motor neuron 1 (SMN1) gene. Adaptive immunity may contribute to SMA as described in other motoneuron diseases, yet mechanisms remain elusive. Nusinersen, an antisense treatment, enhances SMN2 expression, benefiting SMA patients. Here we have longitudinally investigated SMA and nusinersen effects on local immune responses in the cerebrospinal fluid (CSF) - a surrogate of central nervous system parenchyma. Single-cell transcriptomics (SMA: N = 9 versus Control: N = 9) reveal NK cell and CD8+ T cell expansions in untreated SMA CSF, exhibiting activation and degranulation markers. Spatial transcriptomics coupled with multiplex immunohistochemistry elucidate cytotoxicity near chromatolytic motoneurons (N = 4). Post-nusinersen treatment, CSF shows unaltered protein/transcriptional profiles. These findings underscore cytotoxicity's role in SMA pathogenesis and propose it as a therapeutic target. Our study illuminates cell-mediated cytotoxicity as shared features across motoneuron diseases, suggesting broader implications., (© 2024. The Author(s).)

Chimeric antigen receptor (CAR) T cell immunotherapy for the treatment of neurological autoimmune diseases is promising, but CAR T cell kinetics and immune alterations after treatment are poorly understood. Here, we performed single-cell multi-omics sequencing of paired cerebrospinal fluid (CSF) and blood samples from patients with neuromyelitis optica spectrum disorder (NMOSD) treated with anti-B cell maturation antigen (BCMA) CAR T cells. Proliferating cytotoxic-like CD8 + CAR T cell clones were identified as the main effectors in autoimmunity. Anti-BCMA CAR T cells with enhanced features of chemotaxis efficiently crossed the blood-CSF barrier, eliminated plasmablasts and plasma cells in the CSF, and suppressed neuroinflammation. The CD44-expressing early memory phenotype in infusion products was potentially associated with CAR T cell persistence in autoimmunity. Moreover, CAR T cells from patients with NMOSD displayed distinctive features of suppressed cytotoxicity compared with those from hematological malignancies. Thus, we provide mechanistic insights into CAR T cell function in patients with neurological autoimmune disease.

Introduction: Experiencing acute and chronic stress can contribute to adverse health outcomes. Responses to acute stress differ between individuals (i.e., stress reactivity) and the experience of chronic stress has been discussed to be associated with acute stress responses and stress recovery. This study thus aims to investigate whether hair cortisol concentration (HCC), being an indicator for hypothalamus-pituitary-adrenal (HPA) axis activity over a prolonged period of time, is associated with acute stress responses and recovery in a sample of medical students., Material and Methods: From July 2020 to July 2021, medical students (n = 54) underwent a virtual-reality Trier Social Stress Test in which their blood pressure and heart rate variability (HRV) were measured, and hair samples were taken to determine HCC. Piecewise linear growth analyses were used to investigate whether HCC (categorized into low, medium and high levels) is associated with acute stress responses and recovery regarding blood pressure and HRV., Results: Significant interaction effects in piecewise linear growth analyses showed that participants with higher levels of HCC had lower systolic and diastolic blood pressure responses compared to participants with medium levels of HCC. No significant interaction effects were observed for HRV responses or for recovery measures., Conclusions: The study suggests that higher levels of HCC are associated with a lower cardiovascular response in terms of blood pressure to an acute stressor in medical students. Therefore, long-term HPA-axis activity may contribute to different magnitudes of acute stress responses in the autonomic nervous system. As the shown lower blood pressure responses to acute stress in individuals with increased long-term HPA-axis activity may represent inadequate stress responses, these should be further studied in order to find out more about their interaction and potential subsequent disease risks., Competing Interests: Declaration of Competing Interest On behalf of all authors, there are no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

B-cell maturation antigen (BCMA), expressed in plasmablasts and plasma cells, could serve as a promising therapeutic target for autoimmune diseases. We reported here chimeric antigen receptor (CAR) T cells targeting BCMA in two patients with highly relapsed and refractory myasthenia gravis (one with AChR-IgG, and one with MuSk-IgG). Both patients exhibited favorable safety profiles and persistent clinical improvements over 18 months. Reconstitution of B-cell lineages with sustained reduced pathogenic autoantibodies might underlie the therapeutic efficacy. To identify the possible mechanisms underlying the therapeutic efficacy of CAR-T cells in these patients, longitudinal single-cell RNA and TCR sequencing was conducted on serial blood samples post infusion as well as their matching infusion products. By tracking the temporal evolution of CAR-T phenotypes, we demonstrated that proliferating cytotoxic-like CD8 clones were the main effectors in autoimmunity, whereas compromised cytotoxic and proliferation signature and profound mitochondrial dysfunction in CD8 + Te cells before infusion and subsequently defect CAR-T cells after manufacture might explain their characteristics in these patients. Our findings may guide future studies to improve CAR T-cell immunotherapy in autoimmune diseases., (© 2024. The Author(s).)

Valid approaches to conveniently measure stress reactivity are needed due to the growing evidence of its health-impairing effects. This study examined whether the Perceived Stress Reactivity Scale (PSRS) predicts cardiovascular and psychological responses to psychosocial stressors during daily life and during a virtual reality (VR) Trier Social Stress Test (TSST). Medical students answered a standardized baseline questionnaire to assess perceived stress reactivity by the PSRS. The PSRS asks participants to rate the intensity of their typical affective responses to common stressors during daily life. They were further asked to participate in a VR-TSST and in an ecological momentary assessment (EMA) over a period of three consecutive workdays during daily life. Blood pressure and self-reported stress were repeatedly, heart rate variability (HRV) continuously measured during the VR-TSST and EMA. Furthermore, participants repeatedly assessed task demands, task control and social conflict during the EMA. Data was analysed using multilevel analysis and multiple linear regression. Results indicate that the PSRS moderates associations between blood pressure (but not HRV) and demands and control during daily life. Furthermore, the PSRS directly predicted self-reported stress, but did not moderate associations between self-reported stress and demands, control and social conflict. The PSRS did not predict physiological and self-reported stress responses to the VR-TSST. This study partly confirmed convergent validity of the PSRS to stress reactivity in daily life. Furthermore, the lack of association between the PSRS and stress responses to the VR-TSST calls for future studies to search for reliable and valid ways to assess stress reactivity., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Objective: Hospitals are frequently associated with poor working conditions that can lead to work stress and increase the risk for reduced employee well-being. Managers can shape and improve working conditions and thereby, the health of their teams. Thus, as a prerequisite, managers need to be aware of their employees' stress levels. This study had two objectives: At first, it aimed to test the criterion validity of the Effort-Reward Imbalance (ERI) questionnaire measuring psychosocial workload in hospital employees. Secondly, mean scales of the ERI questionnaire filled in by employees were compared with mean scales of an adapted ERI questionnaire, in which managers assessed working conditions of their employees., Methods: Managers (n = 141) from three hospitals located in Germany assessed working conditions of their employees with an adapted external, other-oriented questionnaire. Employees (n = 197) of the mentioned hospitals completed the short version of the ERI questionnaire to assess their working conditions. Confirmatory factor analyses (CFA) were applied to test factorial validity, using the ERI scales for the two study groups. Criterion validity was assessed with multiple linear regression analysis of associations between ERI scales and well-being among employees., Results: The questionnaires demonstrated acceptable psychometric properties in terms of internal consistency of scales, although some indices of model fit resulting from CFA were of borderline significance. Concerning the first objective, effort, reward, and the ratio of effort-reward imbalance were significantly associated with well-being of employees. With regard to the second objective, first tentative findings showed that managers' ratings of their employees' effort at work was quite accurate, whereas their reward was overestimated., Conclusions: With its documented criterion validity the ERI questionnaire can be used as a screening tool of workload among hospital employees. Moreover, in the context of work-related health promotion, managers' perceptions of their employees' workload deserve increased attention as first findings point to some discrepancies between their perceptions and those provided by employees., (© 2023. The Author(s).)

Background: Medical students often experience high levels of stress due to adverse study conditions, which may have adverse health consequences. Hair cortisol concentration (HCC) has been described as a physiological marker for chronic stress and might thus help to identify students under stress and examine the study conditions being responsible for long-term physiological stress responses. This study therefore investigated the association between study conditions and HCC in a sample of medical students., Methods: Fifty-five students from a medical school in Germany completed a paper-based questionnaire and had hair samples collected between May 2020 and July 2021. Study conditions were assessed with student versions of questionnaires based on the Job-Demand-Control-Support model (StrukStud, 25 items) and Effort-Reward Imbalance model (Student ERI, nine items). HCC of two centimeters closest to the scalp were determined by a cortisol luminescence immunoassay. Linear multiple regression analyses were performed to examine associations between study conditions and HCC., Results: Demands (B = 0.23, p = 0.002), effort (B = 0.12, p = 0.029) and the effort-reward-ratio (B = 0.28, p = 0.007) were positively associated with HCC in separate regression analyses, adjusted for age and sex. Only the association between demands and HCC remained significant when all components of the respective questionnaire were considered in the same model (B = 0.22, p = 0.003)., Conclusion: The results suggest that adverse study conditions may be associated with activation of the hypothalamic-pituitary-adrenal axis stress response as reflected by increased HCC. Longitudinal research is needed to confirm these cross-sectional results and examine effects of more prolonged stress due to adverse study conditions., (© 2023. The Author(s).)

Background and Objectives: Immune responses in the central nervous system (CNS) are highly compartmentalized and cerebrospinal fluid (CSF) in particular often reflects CNS pathology better than peripheral blood. While CSF leukocytes are known to be distinct from blood, the immediate effects of peripheral leukocyte depletion on CSF leukocytes have not been studied in humans., Methods: We here analyzed CSF and blood from two relapsing-remitting multiple sclerosis (RRMS) patients early after peripheral leukocyte depletion with the anti-CD52 antibody alemtuzumab compared to untreated RRMS and control patients using single cell RNA-sequencing., Results: As expected for alemtuzumab, most leukocyte lineages including T cells were synchronously depleted from CSF and blood, while - surprisingly - pDCs were maintained in CSF but depleted from blood by alemtuzumab. Transcriptionally, genes associated with migration were elevated only in the CSF after alemtuzumab. Predicted cellular interactions indicated a central role of pDCs and enhanced migration signaling in the CSF after alemtuzumab., Discussion: The CSF and blood compartments are thus partially uncoupled, emphasizing that the CNS is only partially accessible even for treatments profoundly affecting the blood., Competing Interests: Declaration of Competing Interest GMzH received compensation for serving on scientific advisory boards (LFB) and speaker honoraria (Alexion). HW is acting as a paid consultant for AbbVie, Actelion, Biogen, IGES, Johnson & Johnson, Novartis, Roche, Sanofi-Aventis, and the Swiss Multiple Sclerosis Society. The remaining authors declare no financial interests or conflicts of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Objective: Neurological manifestations of autoimmune connective tissue diseases (CTD) are poorly understood and difficult to diagnose. We here aimed to address this shortcoming by studying immune cell compositions in CTD patients with and without neurological manifestation., Methods: Using flow cytometry, we retrospectively investigated paired cerebrospinal fluid (CSF) and blood samples of 28 CTD patients without neurological manifestation, 38 CTD patients with neurological manifestation (N-CTD), 38 non-inflammatory controls, and 38 multiple sclerosis (MS) patients, a paradigmatic primary neuroinflammatory disease., Results: We detected an expansion of plasma cells in the blood of both N-CTD and CTD compared to non-inflammatory controls and MS. Blood plasma cells alone distinguished the clinically similar entities N-CTD and MS with high discriminatory performance (AUC: 0.81). Classical blood monocytes indicated higher disease activity in systemic lupus erythematosus (SLE) patients. Surprisingly, immune cells in the CSF did not differ significantly between N-CTD and CTD, while CD4 + T cells and the CD4 + /CD8 + ratio were elevated in the blood of N-CTD compared to CTD. Several B cell-associated parameters partially overlapped in the CSF in MS and N-CTD. We built a machine learning model that distinguished N-CTD from MS with high discriminatory power using either blood or CSF., Conclusion: We here find that blood flow cytometry alone surprisingly suffices to distinguish CTD with neurological manifestations from clinically similar entities, suggesting that a rapid blood test could support clinicians in the differential diagnosis of N-CTD., (© 2023. The Author(s).)

Background: The SARS-CoV-2 pandemic and climate change are two simultaneously occurring large scale environmental health crises. This provides an opportunity to compare the risk perception of both crises in the population. In particular, whether experiencing the acute pandemic sensitizes people to the risks of ongoing climate change., Methods: Panel participants answered a web-based questionnaire. The risk perception of SARS-CoV-2 and influencing factors were assessed. Differences of risk perception dimensions regarding SARS-CoV-2 and climate change were analyzed as well as associations between dimensions., Results: The results show that an economic impact by the pandemic is associated with more dimensions of SARS-CoV-2 risk perception than an experienced health impact. Moreover, dimensions of risk perception of the pandemic and climate change are perceived differently. Furthermore, the affective dimension of pandemic risk perception is significantly associated with all dimensions of climate change risk perception., Conclusions: Emotional-based coping with the risks of SARS-CoV-2 is associated with risk perception of climate change as well as various factors that shape the individuals' risk perception. It is currently necessary and will be increasingly necessary in the future to solve coexisting crises, not selectively, but in a common context within the framework of a social-ecological and economic transformation.

Multiple sclerosis (MS) is a chronic and often disabling autoimmune disease of the central nervous system (CNS). Cerebrospinal fluid (CSF) surrounds and protects the CNS. Analysis of CSF can aid the diagnosis of CNS diseases, help to identify the prognosis, and underlying mechanisms of diseases. Several recent studies have leveraged single-cell RNA-sequencing (scRNA-seq) to identify MS-associated changes in CSF cells that are considerably more altered than blood cells in MS. However, not all alterations were replicated across all studies. We therefore integrated multiple available scRNA-seq datasets of CSF cells from MS patients with early relapsing-remitting (RRMS) disease. We provide a searchable and interactive resource of this integrated analysis ( https://CSFinMS.bxgenomics.com ) facilitating diverse visualization and analysis methods without requiring computational skills. In the present joint analysis, we replicated the known expansion of B lineage and the recently described expansion of natural killer (NK) cells and some cytotoxic T cells and decrease of monocytes in the CSF in MS. The previous observation of the abundance of Th1-like Th17 effector memory cells in the CSF was not replicated. Expanded CSF B lineage cells resembled class-switched plasmablasts/-cells (e.g., SDC1/CD138, MZB1) as expected. Our integrative analysis thus validates increased cell type diversity and B cell maturation in the CSF in MS and improves accessibility of available data., (© 2022. The Author(s).)

Microglia, the resident immune cells of the central nervous system (CNS), are derived from yolk-sac macrophages that populate the developing CNS during early embryonic development. Once established, the microglia population is self-maintained throughout life by local proliferation. As a scalable source of microglia-like cells (MGLs), we here present a forward programming protocol for their generation from human pluripotent stem cells (hPSCs). The transient overexpression of PU.1 and C/EBPβ in hPSCs led to a homogenous population of mature microglia within 16 d. MGLs met microglia characteristics on a morphological, transcriptional, and functional level. MGLs facilitated the investigation of a human tauopathy model in cortical neuron-microglia cocultures, revealing a secondary dystrophic microglia phenotype. Single-cell RNA sequencing of microglia integrated into hPSC-derived cortical brain organoids demonstrated a shift of microglia signatures toward a more-developmental in vivo-like phenotype, inducing intercellular interactions promoting neurogenesis and arborization. Taken together, our microglia forward programming platform represents a tool for both reductionist studies in monocultures and complex coculture systems, including 3D brain organoids for the study of cellular interactions in healthy or diseased environments.

Background: Primary central nervous system lymphoma (PCNSL) is a rare lymphoma of the central nervous system, usually of diffuse large B cell phenotype. Stereotactic biopsy followed by histopathology is the diagnostic standard. However, limited material is available from CNS biopsies, thus impeding an in-depth characterization of PCNSL., Methods: We performed flow cytometry, single-cell RNA sequencing, and B cell receptor sequencing of PCNSL cells released from biopsy material, blood, and cerebrospinal fluid (CSF), and spatial transcriptomics of biopsy samples., Results: PCNSL-released cells were predominantly activated CD19 + CD20 + CD38 + CD27 + B cells. In single-cell RNA sequencing, PCNSL cells were transcriptionally heterogeneous, forming multiple malignant B cell clusters. Hyperexpanded B cell clones were shared between biopsy- and CSF- but not blood-derived cells. T cells in the tumor microenvironment upregulated immune checkpoint molecules, thereby recognizing immune evasion signals from PCNSL cells. Spatial transcriptomics revealed heterogeneous spatial organization of malignant B cell clusters, mirroring their transcriptional heterogeneity across patients, and pronounced expression of T cell exhaustion markers, co-localizing with a highly malignant B cell cluster., Conclusions: Malignant B cells in PCNSL show transcriptional and spatial intratumor heterogeneity. T cell exhaustion is frequent in the PCNSL microenvironment, co-localizes with malignant cells, and highlights the potential of personalized treatments., (© 2022. The Author(s).)

The cerebrospinal fluid (CSF) features a unique immune cell composition and is in constant contact with the brain borders, thus permitting insights into the brain to diagnose and monitor diseases. Recently, the meninges, which are filled with CSF, were identified as a neuroimmunological interface, highlighting the potential of exploring central nervous system (CNS) immunity by studying CNS border compartments. Here, we summarize how single-cell transcriptomics of such border compartments advance our understanding of neurological diseases, the challenges that remain, and what opportunities novel multi-omic methods offer. Single-cell transcriptomics studies have detected cytotoxic CD4 + T cells and clonally expanded T and B cells in the CSF in the autoimmune disease multiple sclerosis; clonally expanded pathogenic CD8 + T cells were found in the CSF and in the brain adjacent to β-amyloid plaques of dementia patients; in patients with brain metastases, CD8 + T cell clonotypes were shared between the brain parenchyma and the CSF and persisted after therapy. We also outline how novel multi-omic approaches permit the simultaneous measurements of gene expression, chromatin accessibility, and protein in the same cells, which remain to be explored in the CSF. This calls for multicenter initiatives to create single-cell atlases, posing challenges in integrating patients and modalities across centers. While high-dimensional analyses of CSF cells are challenging, they hold potential for personalized medicine by better resolving heterogeneous diseases and stratifying patients., (© 2022. The Author(s).)

Alemtuzumab is a monoclonal antibody that causes rapid depletion of CD52-expressing immune cells. It has proven to be highly efficacious in active relapsing-remitting multiple sclerosis; however, the high risk of secondary autoimmune disorders has greatly complicated its use. Thus, deeper insight into the pathophysiology of secondary autoimmunity and potential biomarkers is urgently needed. The most critical time points in the decision-making process for alemtuzumab therapy are before or at Month 12, where the ability to identify secondary autoimmunity risk would be instrumental. Therefore, we investigated components of blood and CSF of up to 106 multiple sclerosis patients before and after alemtuzumab treatment focusing on those critical time points. Consistent with previous reports, deep flow cytometric immune-cell profiling (n = 30) demonstrated major effects on adaptive rather than innate immunity, which favoured regulatory immune cell subsets within the repopulation. The longitudinally studied CSF compartment (n = 18) mainly mirrored the immunological effects observed in the periphery. Alemtuzumab-induced changes including increased numbers of naïve CD4+ T cells and B cells as well as a clonal renewal of CD4+ T- and B-cell repertoires were partly reminiscent of haematopoietic stem cell transplantation; in contrast, thymopoiesis was reduced and clonal renewal of T-cell repertoires after alemtuzumab was incomplete. Stratification for secondary autoimmunity did not show clear immununological cellular or proteomic traits or signatures associated with secondary autoimmunity. However, a restricted T-cell repertoire with hyperexpanded T-cell clones at baseline, which persisted and demonstrated further expansion at Month 12 by homeostatic proliferation, identified patients developing secondary autoimmune disorders (n = 7 without secondary autoimmunity versus n = 5 with secondary autoimmunity). Those processes were followed by an expansion of memory B-cell clones irrespective of persistence, which we detected shortly after the diagnosis of secondary autoimmune disease. In conclusion, our data demonstrate that (i) peripheral immunological alterations following alemtuzumab are mirrored by longitudinal changes in the CSF; (ii) incomplete T-cell repertoire renewal and reduced thymopoiesis contribute to a proautoimmune state after alemtuzumab; (iii) proteomics and surface immunological phenotyping do not identify patients at risk for secondary autoimmune disorders; (iv) homeostatic proliferation with disparate dynamics of clonal T- and B-cell expansions are associated with secondary autoimmunity; and (v) hyperexpanded T-cell clones at baseline and Month 12 may be used as a biomarker for the risk of alemtuzumab-induced autoimmunity., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Guarantors of Brain.)

Inflammation triggers secondary brain damage after stroke. The meninges and other CNS border compartments serve as invasion sites for leukocyte influx into the brain thus promoting tissue damage after stroke. However, the post-ischemic immune response of border compartments compared to brain parenchyma remains poorly characterized. Here, we deeply characterize tissue-resident leukocytes in meninges and brain parenchyma and discover that leukocytes respond differently to stroke depending on their site of residence. We thereby discover a unique phenotype of myeloid cells exclusive to the brain after stroke. These stroke-associated myeloid cells partially resemble neurodegenerative disease-associated microglia. They are mainly of resident microglial origin, partially conserved in humans and exhibit a lipid-phagocytosing phenotype. Blocking markers specific for these cells partially ameliorates stroke outcome thus providing a potential therapeutic target. The injury-response of myeloid cells in the CNS is thus compartmentalized, adjusted to the type of injury and may represent a therapeutic target., (© 2022. The Author(s).)

Psychotic disorders are common and disabling mental conditions. The relative importance of immune-related mechanisms in psychotic disorders remains subject of debate. Here, we present a large-scale retrospective study of blood and cerebrospinal fluid (CSF) immune cell profiles of psychosis spectrum patients. We performed basic CSF analysis and multi-dimensional flow cytometry of CSF and blood cells from 59 patients with primary psychotic disorders (F20, F22, F23, and F25) in comparison to inflammatory (49 RRMS and 16 NMDARE patients) and non-inflammatory controls (52 IIH patients). We replicated the known expansion of monocytes in the blood of psychosis spectrum patients, that we identified to preferentially affect classical monocytes. In the CSF, we found a relative shift from lymphocytes to monocytes, increased protein levels, and evidence of blood-brain barrier disruption in psychosis. In fact, these CSF features confidently distinguished autoimmune encephalitis from psychosis despite similar (initial) clinical features. We then constructed machine learning models incorporating blood and CSF parameters and demonstrated their superior ability to differentiate psychosis from non-inflammatory controls compared to individual parameters. Multi-dimensional and multi-compartment immune cell signatures can thus support the diagnosis of psychosis spectrum disorders with the potential to accelerate diagnosis and initiation of therapy., (© 2021. The Author(s).)

Uveitis describes a heterogeneous group of inflammatory eye diseases characterized by infiltration of leukocytes into the uveal tissues. Uveitis associated with the HLA haplotype B27 (HLA-B27) is a common subtype of uveitis and a prototypical ocular immune-mediated disease. Local immune mechanisms driving human uveitis are poorly characterized mainly due to the limited available biomaterial and subsequent technical limitations. Here, we provide the first high-resolution characterization of intraocular leukocytes in HLA-B27-positive (n = 4) and -negative (n = 2) anterior uveitis and an infectious endophthalmitis control (n = 1) by combining single-cell RNA-sequencing with flow cytometry and protein analysis. Ocular cell infiltrates consisted primarily of lymphocytes in both subtypes of uveitis and of myeloid cells in infectious endophthalmitis. HLA-B27-positive uveitis exclusively featured a plasmacytoid and classical dendritic cell (cDC) infiltrate. Moreover, cDCs were central in predicted local cell-cell communication. This suggests a unique pattern of ocular leukocyte infiltration in HLA-B27-positive uveitis with relevance to DCs., Competing Interests: MK, MH, DS, XL, TL, MM, DB, KW, HW, KL, AH, GM No competing interests declared, (© 2021, Kasper et al.)

The study investigated the association between onset of workplace violence and onset of sleep disturbances. We used self-reported data from the Swedish Longitudinal Occupational Survey of Health (SLOSH) collected in 2014, 2016, and 2018. A two-wave design was based on participants who had no exposure to workplace violence or sleep disturbances at baseline (n = 6,928). A three-wave design was based on participants who in addition were unexposed to sleep disturbances in the second wave (n = 6,150). Four items of the Karolinska Sleep Questionnaire were used to measure sleep disturbances and one question was used to measure the occurrence of workplace violence or threats of violence. Multivariate logistic regression analyses were performed. In the two-wave approach, onset of workplace violence was associated with onset of sleep disturbances after adjustment for sex, age, occupational position, education, and civil status (adjusted odds ratio 1.41, 95% confidence interval 1.02-1.96). The association was no longer statistically significant after further adjustment for night/evening work, demands, control, and social support at work. In the three-wave approach, results were only suggestive of an association between onset of workplace violence and subsequent onset of sleep disturbances after adjustment for sex, age, occupational position, education, and civil status. Onset of frequent exposure to workplace violence was associated with subsequent onset of sleep disturbances in the adjusted analyses, but these analyses were based on few individuals (13 exposed versus 5,907 unexposed). The results did not conclusively demonstrate that onset of workplace violence predicts development of sleep disturbances. Further research could elucidate the role of other working conditions., (© 2021 The Authors. Journal of Sleep Research published by John Wiley & Sons Ltd on behalf of European Sleep Research Society.)

Ectopic lymphoid tissue containing B cells forms in the meninges at late stages of human multiple sclerosis (MS) and when neuroinflammation is induced by interleukin (IL)-17 producing T helper (Th17) cells in rodents. B cell differentiation and the subsequent release of class-switched immunoglobulins have been speculated to occur in the meninges, but the exact cellular composition and underlying mechanisms of meningeal-dominated inflammation remain unknown. Here, we performed in-depth characterization of meningeal versus parenchymal Th17-induced rodent neuroinflammation. The most pronounced cellular and transcriptional differences between these compartments was the localization of B cells exhibiting a follicular phenotype exclusively to the meninges. Correspondingly, meningeal but not parenchymal Th17 cells acquired a B cell-supporting phenotype and resided in close contact with B cells. This preferential B cell tropism for the meninges and the formation of meningeal ectopic lymphoid tissue was partially dependent on the expression of the transcription factor Bcl6 in Th17 cells that is required in other T cell lineages to induce isotype class switching in B cells. A function of Bcl6 in Th17 cells was only detected in vivo and was reflected by the induction of B cell-supporting cytokines, the appearance of follicular B cells in the meninges, and of immunoglobulin class switching in the cerebrospinal fluid. We thus identify the induction of a B cell-supporting meningeal microenvironment by Bcl6 in Th17 cells as a mechanism controlling compartment specificity in neuroinflammation., Competing Interests: Competing interest statement: A patent application covering the method for reconstructing T cell receptor information from 3′ libraries has been applied with the title “Circulation Method to Sequence Immune Repertoires of Individual Cells” under the filing no. LU101949 by X. L. and G.M.z.H. (date of filing: July 29, 2020)., (Copyright © 2021 the Author(s). Published by PNAS.)

The CNS is ensheathed by the meninges and cerebrospinal fluid, and recent findings suggest that these CNS-associated border tissues have complex immunological functions. Unlike myeloid lineage cells, lymphocytes in border compartments have yet to be thoroughly characterized. Based on single-cell transcriptomics, we here identified a highly location-specific composition and expression profile of tissue-resident leukocytes in CNS parenchyma, pia-enriched subdural meninges, dura mater, choroid plexus and cerebrospinal fluid. The dura layer of the meninges contained a large population of B cells under homeostatic conditions in mice and rats. Murine dura B cells exhibited slow turnover and long-term tissue residency, and they matured in experimental neuroinflammation. The dura also contained B lineage progenitors at the pro-B cell stage typically not found outside of bone marrow, without direct influx from the periphery or the skull bone marrow. This identified the dura as an unexpected site of B cell residence and potentially of development in both homeostasis and neuroinflammation., (© 2021. The Author(s), under exclusive licence to Springer Nature America, Inc.)