Local delivery of cell surface-targeted immunocytokines programs systemic antitumor immunity.

Systemically administered cytokines are potent immunotherapeutics but can cause severe dose-limiting toxicities. To overcome this challenge, cytokines have been engineered for intratumoral retention after local delivery. However, despite inducing regression of treated lesions, tumor-localized cytokines often elicit only modest responses at distal untreated tumors. In the present study, we report a localized cytokine therapy that safely elicits systemic antitumor immunity by targeting the ubiquitous leukocyte receptor CD45. CD45-targeted immunocytokines have lower internalization rates relative to wild-type counterparts, leading to sustained downstream cis and trans signaling between lymphocytes. A single intratumoral dose of αCD45-interleukin (IL)-12 followed by a single dose of αCD45-IL-15 eradicated treated tumors and untreated distal lesions in multiple syngeneic mouse tumor models without toxicity. Mechanistically, CD45-targeted cytokines reprogrammed tumor-specific CD8+ T cells in the tumor-draining lymph nodes to have an antiviral transcriptional signature. CD45 anchoring represents a broad platform for protein retention by host immune cells for use in immunotherapy.

Myelin dysfunction drives amyloid-ß deposition in models of Alzheimer's disease.

The incidence of Alzheimer's disease (AD), the leading cause of dementia, increases rapidly with age, but why age constitutes the main risk factor is still poorly understood. Brain ageing affects oligodendrocytes and the structural integrity of myelin sheaths1, the latter of which is associated with secondary neuroinflammation2,3. As oligodendrocytes support axonal energy metabolism and neuronal health4-7, we hypothesized that loss of myelin integrity could be an upstream risk factor for neuronal amyloid-ß (Aß) deposition, the central neuropathological hallmark of AD. Here we identify genetic pathways of myelin dysfunction and demyelinating injuries as potent drivers of amyloid deposition in mouse models of AD. Mechanistically, myelin dysfunction causes the accumulation of the Aß-producing machinery within axonal swellings and increases the cleavage of cortical amyloid precursor protein. Suprisingly, AD mice with dysfunctional myelin lack plaque-corralling microglia despite an overall increase in their numbers. Bulk and single-cell transcriptomics of AD mouse models with myelin defects show that there is a concomitant induction of highly similar but distinct disease-associated microglia signatures specific to myelin damage and amyloid plaques, respectively. Despite successful induction, amyloid disease-associated microglia (DAM) that usually clear amyloid plaques are apparently distracted to nearby myelin damage. Our data suggest a working model whereby age-dependent structural defects of myelin promote Aß plaque formation directly and indirectly and are therefore an upstream AD risk factor. Improving oligodendrocyte health and myelin integrity could be a promising target to delay development and slow progression of AD.

Brain motor and fear circuits regulate leukocytes during acute stress.

The nervous and immune systems are intricately linked1. Although psychological stress is known to modulate immune function, mechanistic pathways linking stress networks in the brain to peripheral leukocytes remain poorly understood2. Here we show that distinct brain regions shape leukocyte distribution and function throughout the body during acute stress in mice. Using optogenetics and chemogenetics, we demonstrate that motor circuits induce rapid neutrophil mobilization from the bone marrow to peripheral tissues through skeletal-muscle-derived neutrophil-attracting chemokines. Conversely, the paraventricular hypothalamus controls monocyte and lymphocyte egress from secondary lymphoid organs and blood to the bone marrow through direct, cell-intrinsic glucocorticoid signalling. These stress-induced, counter-directional, population-wide leukocyte shifts are associated with altered disease susceptibility. On the one hand, acute stress changes innate immunity by reprogramming neutrophils and directing their recruitment to sites of injury. On the other hand, corticotropin-releasing hormone neuron-mediated leukocyte shifts protect against the acquisition of autoimmunity, but impair immunity to SARS-CoV-2 and influenza infection. Collectively, these data show that distinct brain regions differentially and rapidly tailor the leukocyte landscape during psychological stress, therefore calibrating the ability of the immune system to respond to physical threats.

Pre-Transplant Frequencies of FoxP3+CD25+ in CD3+CD8+ T Cells as Potential Predictors for CMV in CMV-Intermediate Risk Kidney Transplant Recipients.

Cytomegalovirus (CMV) infection detrimentally influences graft survival in kidney transplant recipients, with the risk primarily determined by recipient and donor serostatus. However, recipient CD8+ T cells play a crucial role in CMV control. The optimal preventive strategy (prophylaxis vs. pre-emptive treatment), particularly for seropositive (intermediate risk) recipients, remains uncertain. We investigated CD8+ T cell subpopulation dynamics and CMV occurrence (DNAemia ≥ 100 IU/mL) in 65 kidney transplant recipients, collecting peripheral blood mononuclear cells before (T1) and 1 year after transplantation (T2). Comparing the two timepoints, we found an increase in granulocyte, monocyte and CD3+CD8+ T cells numbers, while FoxP3+CD25+, LAG-3+ and PD-1+ frequencies were reduced at T2. CMV DNAemia occurred in 33 recipients (55.8%) during the first year. Intermediate risk patients were disproportionally affected by posttransplant CMV (N = 29/45, 64.4%). Intermediate risk recipients developing CMV after transplantation exhibited lower leukocyte, monocyte, and granulocyte counts and higher FoxP3+CD25+ frequencies in CD3+CD8+ T cells pre-transplantation compared to patients staying CMV negative. Pre-transplant FoxP3+CD25+ in CD3+CD8+ T cells had the best discriminatory potential for CMV infection prediction within the first year after transplantation (AUC: 0.746). The FoxP3+CD25+ CD3+CD8+ T cell subset may aid in selecting intermediate risk kidney transplant recipients for CMV prophylaxis.

Changes in nutrient composition and diet quality in the first 6 months following bariatric surgery: An observational cohort study.

BACKGROUND: Bariatric surgery (BS) may result in inadequate nutrient intake and poor diet quality, which can lead to nutritional complications. The present study aimed to evaluate changes in macro- and micronutrient composition and diet quality in the first 6 months following BS. METHODS: One hundred seven participants undergoing BS (Roux-en-Y gastric bypass: n = 87, sleeve gastrectomy: n = 20) completed 3-day food records before and 6 months after surgery. Changes in energy, macronutrient (carbohydrates, protein, fat, dietary fibre) and micronutrient intake (folate, vitamin B12, vitamin D, calcium, iron) were evaluated. Diet quality was assessed by adherence to the Dutch food-based dietary guidelines. RESULTS: After BS, we observed a significant decrease in intake of energy and all macro- and micronutrients (p < 0.01 for all), except for calcium (-39.0 ± 404.6 mg; p = 0.32). Overall, nutrient composition slightly changed with an increase in the relative intake of protein (+1.1 ± 4.3 energy percentage [en%]; p = 0.01) and mono- and disaccharides (+4.2 ± 6.4 en%; p < 0.001) post-surgery. Consumption (median [Q1, Q3]) of vegetables (-50 [-120, 6] g day-1 ), wholegrain products (-38 [-81, -8] g day-1 ), liquid fats (-5 [-13, 2] g day-1 ), red meat (-3 [-30, 4] g day-1 ), processed meat (-32 [-55, 13] g day-1 ), sodium (-0.7 [-1.1, -0.2] g day-1 ) and unhealthy food choices (-2.4 [-5.0, 0.6] serves week-1 ) significantly decreased after BS (p < 0.01 for all). CONCLUSIONS: Our results demonstrate both favourable and unfavourable changes in macro- and micronutrient composition and diet quality in the first 6 months following BS. Insight into these changes can improve dietary counselling in this population. Future research into underlying causes, consequences and long-term changes in dietary intake is needed.

Introducing the brain erythropoietin circle to explain adaptive brain hardware upgrade and improved performance.

PREFACE: Executive functions, learning, attention, and processing speed are imperative facets of cognitive performance, affected in neuropsychiatric disorders. In clinical studies on different patient groups, recombinant human (rh) erythropoietin (EPO) lastingly improved higher cognition and reduced brain matter loss. Correspondingly, rhEPO treatment of young rodents or EPO receptor (EPOR) overexpression in pyramidal neurons caused remarkable and enduring cognitive improvement, together with enhanced hippocampal long-term potentiation. The 'brain hardware upgrade', underlying these observations, includes an EPO induced ~20% increase in pyramidal neurons and oligodendrocytes in cornu ammonis hippocampi in the absence of elevated DNA synthesis. In parallel, EPO reduces microglia numbers and dampens their activity and metabolism as prerequisites for undisturbed EPO-driven differentiation of pre-existing local neuronal precursors. These processes depend on neuronal and microglial EPOR. This novel mechanism of powerful postnatal neurogenesis, outside the classical neurogenic niches, and on-demand delivery of new cells, paralleled by dendritic spine increase, let us hypothesize a physiological procognitive role of hypoxia-induced endogenous EPO in brain, which we imitate by rhEPO treatment. Here we delineate the brain EPO circle as working model explaining adaptive 'brain hardware upgrade' and improved performance. In this fundamental regulatory circle, neuronal networks, challenged by motor-cognitive tasks, drift into transient 'functional hypoxia', thereby triggering neuronal EPO/EPOR expression.

NMDAR1 autoantibodies amplify behavioral phenotypes of genetic white matter inflammation: a mild encephalitis model with neuropsychiatric relevance.

Encephalitis has an estimated prevalence of ≤0.01%. Even with extensive diagnostic work-up, an infectious etiology is identified or suspected in <50% of cases, suggesting a role for etiologically unclear, noninfectious processes. Mild encephalitis runs frequently unnoticed, despite slight neuroinflammation detectable postmortem in many neuropsychiatric illnesses. A widely unexplored field in humans, though clearly documented in rodents, is genetic brain inflammation, particularly that associated with myelin abnormalities, inducing primary white matter encephalitis. We hypothesized that "autoimmune encephalitides" may result from any brain inflammation concurring with the presence of brain antigen-directed autoantibodies, e.g., against N-methyl-D-aspartate-receptor NR1 (NMDAR1-AB), which are not causal of, but may considerably shape the encephalitis phenotype. We therefore immunized young female Cnp-/- mice lacking the structural myelin protein 2'-3'-cyclic nucleotide 3'-phosphodiesterase (Cnp) with a "cocktail" of NMDAR1 peptides. Cnp-/- mice exhibit early low-grade inflammation of white matter tracts and blood-brain barrier disruption. Our novel mental-time-travel test disclosed that Cnp-/- mice are compromised in what-where-when orientation, but this episodic memory readout was not further deteriorated by NMDAR1-AB. In contrast, comparing wild-type and Cnp-/- mice without/with NMDAR1-AB regarding hippocampal learning/memory and motor balance/coordination revealed distinct stair patterns of behavioral pathology. To elucidate a potential contribution of oligodendroglial NMDAR downregulation to NMDAR1-AB effects, we generated conditional NR1 knockout mice. These mice displayed normal Morris water maze and mental-time-travel, but beam balance performance was similar to immunized Cnp-/-. Immunohistochemistry confirmed neuroinflammation/neurodegeneration in Cnp-/- mice, yet without add-on effect of NMDAR1-AB. To conclude, genetic brain inflammation may explain an encephalitic component underlying autoimmune conditions.

Preadult polytoxicomania-strong environmental underpinnings and first genetic hints.

Considering the immense societal and personal costs and suffering associated with multiple drug use or "polytoxicomania", better understanding of environmental and genetic causes is crucial. While previous studies focused on single risk factors and selected drugs, effects of early-accumulated environmental risks on polytoxicomania were never addressed. Similarly, evidence of genetic susceptibility to particular drugs is abundant, while genetic predisposition to polytoxicomania is unexplored. We exploited the GRAS data collection, comprising information on N~2000 deep-phenotyped schizophrenia patients, to investigate effects of early-life environmental risk accumulation on polytoxicomania and additionally provide first genetic insight. Preadult accumulation of environmental risks (physical or sexual abuse, urbanicity, migration, cannabis, alcohol) was strongly associated with lifetime polytoxicomania (p = 1.5 × 10-45; OR = 31.4), preadult polytoxicomania with OR = 226.6 (p = 1.0 × 10-33) and adult polytoxicomania with OR = 17.5 (p = 3.4 × 10-24). Parallel accessibility of genetic data from GRAS patients and N~2100 controls for genome-wide association (GWAS) and phenotype-based genetic association studies (PGAS) permitted the creation of a novel multiple GWAS-PGAS approach. This approach yielded 41 intuitively interesting SNPs, potentially conferring liability to preadult polytoxicomania, which await replication upon availability of suitable deep-phenotyped cohorts anywhere world-wide. Concisely, juvenile environmental risk accumulation, including cannabis and alcohol as starter/gateway drugs, strongly predicts polytoxicomania during adolescence and adulthood. This pivotal message should launch more effective sociopolitical measures to prevent this deleterious psychiatric condition.

Autoantibodies against NMDA receptor 1 modify rather than cause encephalitis.

The etiology and pathogenesis of "anti-N-methyl-D-aspartate-receptor (NMDAR) encephalitis" and the role of autoantibodies (AB) in this condition are still obscure. While NMDAR1-AB exert NMDAR-antagonistic properties by receptor internalization, no firm evidence exists to date that NMDAR1-AB by themselves induce brain inflammation/encephalitis. NMDAR1-AB of all immunoglobulin classes are highly frequent across mammals with multiple possible inducers and boosters. We hypothesized that "NMDAR encephalitis" results from any primary brain inflammation coinciding with the presence of NMDAR1-AB, which may shape the encephalitis phenotype. Thus, we tested whether following immunization with a "cocktail" of 4 NMDAR1 peptides, induction of a spatially and temporally defined sterile encephalitis by diphtheria toxin-mediated ablation of pyramidal neurons ("DTA" mice) would modify/aggravate the ensuing phenotype. In addition, we tried to replicate a recent report claiming that immunizing just against the NMDAR1-N368/G369 region induced brain inflammation. Mice after DTA induction revealed a syndrome comprising hyperactivity, hippocampal learning/memory deficits, prefrontal cortical network dysfunction, lasting blood brain-barrier impairment, brain inflammation, mainly in hippocampal and cortical regions with pyramidal neuronal death, microgliosis, astrogliosis, modest immune cell infiltration, regional atrophy, and relative increases in parvalbumin-positive interneurons. The presence of NMDAR1-AB enhanced the hyperactivity (psychosis-like) phenotype, whereas all other readouts were identical to control-immunized DTA mice. Non-DTA mice with or without NMDAR1-AB were free of any encephalitic signs. Replication of the reported NMDAR1-N368/G369-immunizing protocol in two large independent cohorts of wild-type mice completely failed. To conclude, while NMDAR1-AB can contribute to the behavioral phenotype of an underlying encephalitis, induction of an encephalitis by NMDAR1-AB themselves remains to be proven.

Hippocampal neurons respond to brain activity with functional hypoxia.

Physical activity and cognitive challenge are established non-invasive methods to induce comprehensive brain activation and thereby improve global brain function including mood and emotional well-being in healthy subjects and in patients. However, the mechanisms underlying this experimental and clinical observation and broadly exploited therapeutic tool are still widely obscure. Here we show in the behaving brain that physiological (endogenous) hypoxia is likely a respective lead mechanism, regulating hippocampal plasticity via adaptive gene expression. A refined transgenic approach in mice, utilizing the oxygen-dependent degradation (ODD) domain of HIF-1α fused to CreERT2 recombinase, allows us to demonstrate hypoxic cells in the performing brain under normoxia and motor-cognitive challenge, and spatially map them by light-sheet microscopy, all in comparison to inspiratory hypoxia as strong positive control. We report that a complex motor-cognitive challenge causes hypoxia across essentially all brain areas, with hypoxic neurons particularly abundant in the hippocampus. These data suggest an intriguing model of neuroplasticity, in which a specific task-associated neuronal activity triggers mild hypoxia as a local neuron-specific as well as a brain-wide response, comprising indirectly activated neurons and non-neuronal cells.

Multiple inducers and novel roles of autoantibodies against the obligatory NMDAR subunit NR1: a translational study from chronic life stress to brain injury.

Circulating autoantibodies (AB) of different immunoglobulin classes (IgM, IgA, and IgG), directed against the obligatory N-methyl-D-aspartate-receptor subunit NR1 (NMDAR1-AB), belong to the mammalian autoimmune repertoire, and appear with age-dependently high seroprevalence across health and disease. Upon access to the brain, they can exert NMDAR-antagonistic/ketamine-like actions. Still unanswered key questions, addressed here, are conditions of NMDAR1-AB formation/boosting, intraindividual persistence/course in serum over time, and (patho)physiological significance of NMDAR1-AB in modulating neuropsychiatric phenotypes. We demonstrate in a translational fashion from mouse to human that (1) serum NMDAR1-AB fluctuate upon long-term observation, independent of blood-brain barrier (BBB) perturbation; (2) a standardized small brain lesion in juvenile mice leads to increased NMDAR1-AB seroprevalence (IgM + IgG), together with enhanced Ig-class diversity; (3) CTLA4 (immune-checkpoint) genotypes, previously found associated with autoimmune disease, predispose to serum NMDAR1-AB in humans; (4) finally, pursuing our prior findings of an early increase in NMDAR1-AB seroprevalence in human migrants, which implicated chronic life stress as inducer, we independently replicate these results with prospectively recruited refugee minors. Most importantly, we here provide the first experimental evidence in mice of chronic life stress promoting serum NMDAR1-AB (IgA). Strikingly, stress-induced depressive-like behavior in mice and depression/anxiety in humans are reduced in NMDAR1-AB carriers with compromised BBB where NMDAR1-AB can readily reach the brain. To conclude, NMDAR1-AB may have a role as endogenous NMDAR antagonists, formed or boosted under various circumstances, ranging from genetic predisposition to, e.g., tumors, infection, brain injury, and stress, altogether increasing over lifetime, and exerting a spectrum of possible effects, also including beneficial functions.

Quantitative Assessment of Viral Dispersion Associated with Respiratory Support Devices in a Simulated Critical Care Environment.

Rationale: Patients with severe coronavirus disease (COVID-19) require supplemental oxygen and ventilatory support. It is unclear whether some respiratory support devices may increase the dispersion of infectious bioaerosols and thereby place healthcare workers at increased risk of infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).Objectives: To quantitatively compare viral dispersion from invasive and noninvasive respiratory support modalities.Methods: This study used a simulated ICU room with a breathing-patient simulator exhaling nebulized bacteriophages from the lower respiratory tract with various respiratory support modalities: invasive ventilation (through an endotracheal tube with an inflated cuff connected to a mechanical ventilator), helmet ventilation with a positive end-expiratory pressure (PEEP) valve, noninvasive bilevel positive-pressure ventilation, nonrebreather face masks, high-flow nasal oxygen (HFNO), and nasal prongs.Measurements and Main Results: Invasive ventilation and helmet ventilation with a PEEP valve were associated with the lowest bacteriophage concentrations in the air, and HFNO and nasal prongs were associated with the highest concentrations. At the intubating position, bacteriophage concentrations associated with HFNO (2.66 × 104 plaque-forming units [PFU]/L of air sampled), nasal prongs (1.60 × 104 PFU/L of air sampled), nonrebreather face masks (7.87 × 102 PFU/L of air sampled), and bilevel positive airway pressure (1.91 × 102 PFU/L of air sampled) were significantly higher than those associated with invasive ventilation (P < 0.05 for each). The difference between bacteriophage concentrations associated with helmet ventilation with a PEEP valve (4.29 × 10-1 PFU/L of air sampled) and bacteriophage concentrations associated with invasive ventilation was not statistically significant.Conclusions: These findings highlight the potential differential risk of dispersing virus among respiratory support devices and the importance of appropriate infection prevention and control practices and personal protective equipment for healthcare workers when caring for patients with transmissible respiratory viral infections such as SARS-CoV-2.

A Quantitative Study of Particle Dispersion due to Respiratory Support Modalities in PC-12 Aircraft: Prehospital Patient Transport.

OBJECTIVE: It is unclear whether supplemental oxygen and noninvasive ventilation respiratory support devices increase the dispersion of potentially infectious bioaerosols in a pressurized air medical cabin. This study quantitatively compared particle dispersion from respiratory support modalities in an air medical cabin during flight. METHODS: Dispersion was measured in a fixed wing air ambulance during flight with a breathing medical mannequin simulator exhaling nebulized saline from the lower respiratory tract with the following respiratory support modalities: a nasal cannula with a surgical mask, high-flow nasal oxygen (HFNO) with a surgical mask, and noninvasive bilevel positive airway pressure (BiPAP) ventilation. RESULTS: Nasal cannula oxygen with a surgical mask was associated with the highest particle concentrations. In the absence of mask seal leaks, BiPAP was associated with 1 order of magnitude lower particle concentration compared with a nasal cannula with a surgical mask. Particle concentrations associated with HFNO with a surgical mask were lower than a nasal cannula with a surgical mask but higher than BiPAP. CONCLUSIONS: Particle dispersion associated with the use of BiPAP and HFNO with a surgical mask is lower than nasal cannula oxygen with a surgical mask. These findings may assist air medical organizations with operational decisions where little data exist about respiratory particle dispersion.

Blockade of tumor necrosis factor superfamily members CD30 and OX40 abrogates disease activity in murine immune-mediated glomerulonephritis.

Co-stimulation is a prerequisite for pathogenic activity in T cell-mediated diseases and has been demonstrated to achieve tolerance in organ-specific autoimmunity as a therapeutic target. Here, we evaluated the involvement of the tumor necrosis factor family members CD30 and OX40 in immune-complex mediated kidney disease. In vitro stimulation and proliferation studies were performed with CD4+ cells from wild type and CD30/OX40 double knock-out (CD30OX40-/-) mice. In vivo studies were performed by induction of nephrotoxic serum nephritis in wild type, CD30OX40- /- , CD30-/-, OX40-/-, reconstituted Rag1-/- and C57Bl/6J mice treated with αCD30L αOX40L antibodies. CD30, OX40 and their ligands were upregulated on various leukocytes in nephrotoxic serum nephritis. CD30OX40-/- mice, but not CD30-/- or OX40-/- mice were protected from nephrotoxic serum nephritis. Similar protection was found in Rag1-/- mice injected with CD4+ T cells from CD30OX40-/- mice compared to Rag1-/- mice injected with CD4+ T cells from wild type mice. Furthermore, CD4+ T cells deficient in CD30OX40-/- displayed decreased expression of CCR6 in vivo. CD30OX40-/- cells were fully capable of differentiating into disease mediating T helper cell subsets, but showed significantly decreased levels of proliferation in vivo and in vitro compared to wild type cells. Blocking antibodies against CD30L and OX40L ameliorated nephrotoxic serum nephritis without affecting pan-effector or memory T cell populations. Thus, our results indicate disease promotion via CD30 and OX40 signaling due to facilitation of exaggerated T cell proliferation and migration of T helper 17 cells in nephrotoxic serum nephritis. Hence, co-stimulation blockade targeting the CD30 and OX40 signaling pathways may provide a novel therapeutic strategy in autoimmune kidney disease.

Glucagon-Like Peptide-1 Receptor Agonism Improves Nephrotoxic Serum Nephritis by Inhibiting T-Cell Proliferation.

Glucagon-like peptide (GLP)-1 analogs such as liraglutide improved albuminuria in patients with type 2 diabetes in large randomized controlled trials. One of the suspected mechanisms is the anti-inflammatory potential of GLP-1 receptor (Glp1r) agonism. Thus, the anti-inflammatory action of Glp1r agonism was tested in a nondiabetic, T-cell-mediated murine model of nephrotoxic serum nephritis (NTS). The role of Glp1r in NTS was evaluated by using Glp1r-/- mice or C57BL/6 mice treated with liraglutide. In vitro, murine T cells were stimulated in the presence of liraglutide or vehicle. Glp1r-/- mice displayed increased renal infiltration of neutrophils and T cells after induction of NTS. Splenocyte proliferation and TH1 cytokine transcription were increased in spleen and lymph nodes of Glp1r-/- mice. Liraglutide treatment significantly improved the renal outcome of NTS in C57BL/6 mice by decreasing renal infiltration and proliferation of T cells, which resulted in decreased macrophage infiltration. In vitro, T cells stimulated in the presence of liraglutide showed decreased proliferation of TH1 and TH17 cells. Liraglutide blocked glycolysis in T cells and decreased their Glut1 mRNA expression. Together, Glp1r agonism protects mice from a T-cell-dependent glomerulonephritis model by inhibition of T-cell proliferation, possibly by interacting with their metabolic program. This mechanism may explain in part the renoprotective effects of Glp1r agonism in diabetic nephropathy.

Autoantibodies against the NMDAR subunit NR1 are associated with neuropsychiatric outcome after ischemic stroke.

BACKGROUND AND PURPOSE: Preexisting autoantibodies against N-methyl-D-aspartate-receptor subunit NR1 (NMDAR1-AB) in acute ischemic stroke patients with previously intact blood-brain-barrier were associated with smaller evolution of lesion size. Effects of chronic exposure to NMDAR1-AB long after stroke, however, have remained unclear. We investigated in a prospective follow-up study whether long-term neuropsychiatric outcome after stroke differs depending on NMDAR1-AB status. METHODS: Blood samples for NMDAR1-AB analysis were collected within 24 h after ischemic stroke from n = 114 patients. Outcome was assessed 1-3 years later using NIHSS, modified Rankin-scale, Barthel-Index, RBANS (Repeatable Battery for the Assessment of Neuropsychological Status) subcategories (immediate/delayed memory, attention, visuoconstruction), anamnesis evaluating neuropsychiatric symptoms (e.g. hallucinations, psychomotor slowing, reduced alertness, depressiveness, fatigue) and questionnaires (Beck's Depression Inventory-BDI, Fatigue Impact Scale-FIS). Scores were generated to cover RBANS plus neuropsychiatric symptoms (Score A; n = 96) or only neuropsychiatric symptoms (Score B; n = 114, including patients unable to conduct RBANS). Depression/fatigue were measured in patients, capable to perform questionnaires (n = 86). RESULTS: NMDAR1-AB (IgM, IgA, IgG) were detected in n = 27 patients (23.7%). NMDAR1-AB seropositive patients showed inferior results in Score A (p = 0.006), Score B (p = 0.004), BDI (p = 0.013) and FIS (p = 0.018), compared to seronegative patients. Multiple regression analysis including covariates age, NIHSS at day 7 post-stroke, and days from stroke to follow-up, showed NMDAR1-AB seropositivity associated with worse outcome in Scores A (b: 1.517, 95%CI: 0.505-2.529, p = 0.004) and B (b: 0.803, 95%CI: 0.233-1.373; p = 0.006). Also FIS was unfavorably associated with NMDAR1-AB seropositivity (binary logistic regression: OR: 3.904, 95%CI: 1.200-12.695; p = 0.024). CONCLUSIONS: Even though the numbers of included patients are low, our data apparently indicate that NMDAR1-AB seropositivity at the time point of acute ischemic stroke is associated with neuropsychiatric symptoms including cognitive dysfunction and fatigue years after stroke. Preclinical proof of a causal relation provided, targeted immunosuppression may be a future prophylactic option to be clinically evaluated.

CYLD is a causative gene for frontotemporal dementia - amyotrophic lateral sclerosis.

Frontotemporal dementia and amyotrophic lateral sclerosis are clinically and pathologically overlapping disorders with shared genetic causes. We previously identified a disease locus on chromosome 16p12.1-q12.2 with genome-wide significant linkage in a large European Australian family with autosomal dominant inheritance of frontotemporal dementia and amyotrophic lateral sclerosis and no mutation in known amyotrophic lateral sclerosis or dementia genes. Here we demonstrate the segregation of a novel missense variant in CYLD (c.2155A>G, p.M719V) within the linkage region as the genetic cause of disease in this family. Immunohistochemical analysis of brain tissue from two CYLD p.M719V mutation carriers showed widespread glial CYLD immunoreactivity. Primary mouse neurons transfected with CYLDM719V exhibited increased cytoplasmic localization of TDP-43 and shortened axons. CYLD encodes a lysine 63 deubiquitinase and CYLD cutaneous syndrome, a skin tumour disorder, is caused by mutations that lead to reduced deubiquitinase activity. In contrast with CYLD cutaneous syndrome-causative mutations, CYLDM719V exhibited significantly increased lysine 63 deubiquitinase activity relative to the wild-type enzyme (paired Wilcoxon signed-rank test P = 0.005). Overexpression of CYLDM719V in HEK293 cells led to more potent inhibition of the cell signalling molecule NF-κB and impairment of autophagosome fusion to lysosomes, a key process in autophagy. Although CYLD mutations appear to be rare, CYLD's interaction with at least three other proteins encoded by frontotemporal dementia and/or amyotrophic lateral sclerosis genes (TBK1, OPTN and SQSTM1) suggests that it may play a central role in the pathogenesis of these disorders. Mutations in several frontotemporal dementia and amyotrophic lateral sclerosis genes, including TBK1, OPTN and SQSTM1, result in a loss of autophagy function. We show here that increased CYLD activity also reduces autophagy function, highlighting the importance of autophagy regulation in the pathogenesis of frontotemporal dementia and amyotrophic lateral sclerosis.

CaMKIIα Expressing Neurons to Report Activity-Related Endogenous Hypoxia upon Motor-Cognitive Challenge.

We previously introduced the brain erythropoietin (EPO) circle as a model to explain the adaptive 'brain hardware upgrade' and enhanced performance. In this fundamental circle, brain cells, challenged by motor-cognitive tasks, experience functional hypoxia, triggering the expression of EPO among other genes. We attested hypoxic cells by a transgenic reporter approach under the ubiquitous CAG promoter, with Hif-1α oxygen-dependent degradation-domain (ODD) fused to CreERT2-recombinase. To specifically focus on the functional hypoxia of excitatory pyramidal neurons, here, we generated CaMKIIα-CreERT2-ODD::R26R-tdTomato mice. Behavioral challenges, light-sheet microscopy, immunohistochemistry, single-cell mRNA-seq, and neuronal cultures under normoxia or hypoxia served to portray these mice. Upon complex running wheel performance as the motor-cognitive task, a distinct increase in functional hypoxic neurons was assessed immunohistochemically and confirmed three-dimensionally. In contrast, fear conditioning as hippocampal stimulus was likely too short-lived to provoke neuronal hypoxia. Transcriptome data of hippocampus under normoxia versus inspiratory hypoxia revealed increases in CA1 CaMKIIα-neurons with an immature signature, characterized by the expression of Dcx, Tbr1, CaMKIIα, Tle4, and Zbtb20, and consistent with accelerated differentiation. The hypoxia reporter response was reproduced in vitro upon neuronal maturation. To conclude, task-associated activity triggers neuronal functional hypoxia as a local and brain-wide reaction mediating adaptive neuroplasticity. Hypoxia-induced genes such as EPO drive neuronal differentiation, brain maturation, and improved performance.

Genetically induced brain inflammation by Cnp deletion transiently benefits from microglia depletion.

Reduced expression of 2'-3'-cyclic nucleotide 3'-phosphodiesterase (Cnp) in humans and mice causes white matter inflammation and catatonic signs. These consequences are experimentally alleviated by microglia ablation via colony-stimulating factor 1 receptor (CSF1R) inhibition using PLX5622. Here we address for the first time preclinical topics crucial for translation, most importantly 1) the comparison of 2 long-term PLX5622 applications (prevention and treatment) vs. 1 treatment alone, 2) the correlation of catatonic signs and executive dysfunction, 3) the phenotype of leftover microglia evading depletion, and 4) the role of intercellular interactions for efficient CSF1R inhibition. Based on our Cnp-/- mouse model and in vitro time-lapse imaging, we report the unexpected discovery that microglia surviving under PLX5622 display a highly inflammatory phenotype including aggressive premortal phagocytosis of oligodendrocyte precursor cells. Interestingly, ablating microglia in vitro requires mixed glial cultures, whereas cultured pure microglia withstand PLX5622 application. Importantly, 2 extended rounds of CSF1R inhibition are not superior to 1 treatment regarding any readout investigated (magnetic resonance imaging and magnetic resonance spectroscopy, behavior, immunohistochemistry). Catatonia-related executive dysfunction and brain atrophy of Cnp-/- mice fail to improve under PLX5622. To conclude, even though microglia depletion is temporarily beneficial and worth pursuing, complementary treatment strategies are needed for full and lasting recovery.-Fernandez Garcia-Agudo, L., Janova, H., Sendler, L. E., Arinrad, S., Steixner, A. A., Hassouna, I., Balmuth, E., Ronnenberg, A., Schopf, N., van der Flier, F. J., Begemann, M., Martens, H., Weber, M. S., Boretius, S., Nave, K.-A., Ehrenreich, H. Genetically induced brain inflammation by Cnp deletion transiently benefits from microglia depletion.

Roadmap for cardiovascular education across the European Society of Cardiology: inspiring better knowledge and skills, now and for the future.

AIMS: The provision of high-quality education allows the European Society of Cardiology (ESC) to achieve its mission of better cardiovascular practice and provides an essential component of translating new evidence to improve outcomes. METHODS AND RESULTS: The 4th ESC Education Conference, held in Sophia Antipolis (December 2016), brought together ESC education leaders, National Directors of Training of 43 ESC countries, and representatives of the ESC Young Community. Integrating national descriptions of education and cardiology training, we discussed innovative pathways to further improve knowledge and skills across different training programmes and health care systems. We developed an ESC roadmap supporting better cardiology training and continued medical education (CME), noting: (i) The ESC provides an excellent framework for unbiased and up-to-date cardiovascular education in close cooperation with its National Societies. (ii) The ESC should support the harmonization of cardiology training, curriculum development, and professional dialogue and mentorship. (iii) ESC congresses are an essential forum to learn and discuss the latest developments in cardiovascular medicine. (iv) The ESC should create a unified, interactive educational platform for cardiology training and continued cardiovascular education combining Webinars, eLearning Courses, Clinical Cases, and other educational programmes, along with ESC Congress content, Practice Guidelines and the next ESC Textbook of Cardiovascular Medicine. (v) ESC-delivered online education should be integrated into National and regional cardiology training and CME programmes. CONCLUSION: These recommendations support the ESC to deliver excellent and comprehensive cardiovascular education for the next generation of specialists. Teamwork between international, national and local partners is essential to achieve this objective.