Biology subjects

Hagemann, N.

Publications and source records attributed to Hagemann, N..

8 recordsLinked to original sources

bioRxiv2026

Hyperlipidemia abolishes, but immune balancing by DNase-I restores neuroprotection by MSC-derived extracellular vesicles

BackgroundOwing to their potent immunomodulatory properties, mesenchymal stromal cell (MSC)-derived small extracellular vesicles (EVs) have emerged as promising neuroprotective treatments for ischemic stroke. Preclinical studies using MSC-EVs have mainly been performed in young, otherwise healthy rodents. Stroke patients frequently carry vascular risk factors and comorbidities. We herein investigated whether MSC-EVs retain neuroprotective activity in hyperlipidemic mice on cholesterol-rich Western diet. MethodsMale C57BL/6J mice were exposed to regular normal diet or Western diet for 6 weeks. At the age of 9-10 weeks, mice were exposed to transient intraluminal middle cerebral artery occlusion (MCAO). Vehicle or MSC-EVs (2x106 or 6x106 cell equivalents) were intravenously administered immediately after reperfusion, and vehicle or rosuvastatin (5 mg/kg/day) were intraperitoneally applied starting immediately after or seven days before MCAO. Neurological deficits, ischemic injury, and immune responses were evaluated up to 72 hours post-ischemia. To investigate the hyperlipidemia-associated immune dysregulation, mice received DNase-I before or immediately after MCAO. In defined subgroups, monocytes/ macrophages or neutrophils were additionally depleted by clodronate liposomes or anti-Ly6G antibodies, respectively. ResultsIn contrast to normolipidemic control mice, MSC-EVs failed to induce post-ischemic neuroprotection in hyperlipidemic mice. Neither MSC-EV dose escalation nor rosuvastatin co-treatment restored the therapeutic efficacy of MSC-EVs. Hyperlipidemia induced systemic innate immune dysregulation characterized by reduced monocyte/ macrophage activation, increased neutrophil activation, and elevated circulating cell-free DNA. DNase-I treatment before, but not after MCAO reversed these immune abnormalities and restored neuroprotection by MSC-EVs, decreasing neurological deficits, infarct volume and brain edema. Depletion of either monocytes/ macrophages or neutrophils abolished the neuroprotective effects of MSC-EVs in DNase-I-pretreated hyperlipidemic mice. ConclusionsImmune dysregulation abolishes MSC-EV-induced neuroprotection after ischemic stroke in hyperlipidemic mice. DNase-I priming restores MSC-EV responsiveness through mechanisms critically involving monocyte/ macrophage and neutrophil rebalancing. Our data highlight the host immune status as determinant of EV therapeutic efficacy.

neuroscience↗

bioRxiv2026

Monocytes shape the neuroprotective and immunomodulatory effects of mesenchymal stromal cell-derived extracellular vesicles

BACKGROUNDMesenchymal stromal cell-derived extracellular vesicles (MSC-EVs) exert neuroprotective effects in ischemic stroke largely through immunomodulatory mechanisms. Monocytes are first-line responders to MSC-EVs. Their contribution to MSC-EV-induced neuroprotection remains poorly understood. This study investigated the role of monocytes in shaping neuroprotective responses to MSC-EVs after ischemic stroke. METHODSMale C57BL/6J mice were exposed to transient middle cerebral artery occlusion (MCAO). Monocytes were depleted using pharmacological (clodronate liposomes), immunological (anti-CCR2), or genetic (Mrp8-Cre+/- Nr4a1fl/fl) approaches removing total, CCR2+, or Ly6Clow monocytes, respectively. In additional cohorts, neutrophils and T cells were simultaneously depleted by anti-Ly6G or anti-CD4/CD8 antibodies. Small EVs from clonally expanded immortalized MSCs were administered intravenously. Neurological deficits, ischemic injury, and immune responses were analyzed up to 72 hours post-MCAO. Complementary ex vivo studies were performed, in which MSC-EVs were administered to monocyte-depleted or non-depleted peripheral blood mononuclear cells (PBMCs) obtained from acute ischemic stroke patients. RESULTSIn ischemic mice with intact monocyte compartment, MSC-EVs reduced neurological deficits, infarct volume, neuronal injury, and brain leukocyte infiltrates. These protective effects were abolished in monocyte-depleted mice, particularly following CCR2+ monocyte depletion. Under these conditions, MSC-EV treatment exacerbated neurological deficits, ischemic injury, and leukocyte infiltration, accompanied by neutrophil and T cell expansion and overactivation. Depletion of neutrophils or T cells prevented the EV-induced worsening of stroke outcome in monocyte-deficient mice. Ly6Clow monocytes played a crucial role in orchestrating immune responses to MSC-EVs. Their depletion abolished EV-induced neuroprotection. In stroke patient PBMCs, MSC-EVs induced phenotypic reprogramming of monocytes, whereas they promoted CD4+ and CD8+ T cell activation in the absence of monocytes. CONCLUSIONSMonocytes shape the immunomodulatory actions of MSC-EVs. In their absence, MSC-EVs trigger neutrophil and T cell overactivation that worsens stroke outcome. These findings highlight the importance of monocyte- and T cell-related potency assays for the clinical translation of MSC-EV therapies.

neuroscience↗

bioRxiv2026

Histone neutralization protects the ischemic brain against stroke-associated pneumonia

Bacterial pneumonia aggravates ischemic stroke via mechanisms that still remain to be determined. In ischemic stroke patients and mice exposed to middle cerebral artery occlusion, we show that stroke-associated pneumonia markedly worsens clinical stroke outcome. In mice, pneumonia induced 3 days after stroke impaired neurological recovery and increased brain neutrophil infiltrates, blood-brain barrier breakdown, cerebral microvascular thrombosis, and progressive brain atrophy. The antibiotic amoxicillin only partially ameliorated pneumonia-associated neurological deficits and neutrophil infiltrates. Neutrophils were critical mediators of pneumonia-induced blood-brain barrier breakdown and microvascular thrombosis. Notably, administration of a neutralizing anti-histone antibody during pneumonia--unlike degradation or blockade of neutrophil extracellular trap formation or myeloperoxidase inhibition--restored long-term neurological recovery and prevented brain atrophy in stroke-associated pneumonia mice. This study identifies extracellular histones as key drivers of secondary inflammatory brain injury and establishes histone neutralization as a therapeutic strategy with an extended treatment window in the post-acute stroke phase. One Sentence SummaryNeutralizing extracellular histones reverses pneumonia-driven secondary brain injury and restores long-term recovery after ischemic stroke.

neuroscience↗

bioRxiv2025

The nuclear receptor NR4A1 serves as a neutrophil-intrinsic regulator mitigating stroke severity

Ischemic stroke is accompanied by recruitment and activation of immune cells which play an important role in the progression of the brain damage. The nuclear receptor NR4A1 emerged as a key regulator within the inflammatory response of several immune diseases by regulating immune cell activation. In this study, we investigated the role of NR4A1 in the activation and recruitment of brain resident and peripheral immune cells after cerebral ischemia. Here, we show that NR4A1 mediates an anti-inflammatory and damage-limiting effect after stroke. This effect is largely mediated by neutrophil recruitment and importantly, NR4A1 activation with its ligand Cytosporone B improves functional outcome and reduces brain damage. Modulation of NR4A1 is therefore a promising therapeutic target for the treatment of the nuclear receptor NR4A1 in the activation and recruitment of peripheral and brain resident immune cells after cerebral ischemia and its consequences for stroke outcome. We demonstrate that NR4A1 ablation augments neutrophil activation and CNS recruitment within days after stroke thereby increasing infarct size, CNS inflammation, neuronal damage and deteriorating functional outcome. This effect is mediated via modulation of cell-intrinsic neutrophil function and maturation as illustrated by neutrophil-specific NR4A1 ablation and mixed bone-marrow chimera experiments. Notably, the NR4A1 agonist Cytosporone B reduced CNS neutrophil infiltration, infarct size and functional outcome after stroke in a bicentric preclinical stroke trial, demonstrating that NR4A1-mediated control of neutrophil reactivity is amenable to pharmacological modulation. In humans, NR4A1 expressing neutrophils are present in the peripheral blood of stroke patients and neutrophil NR4A1 expression correlates with improved long-term outcome after 3 months. Furthermore, NR4A1 expression in brain parenchyma neutrophils is negatively correlated with neuronal cell loss, illustrating a role of NR4A1 in regulating neutrophil mediated neuronal cell death in human stroke. Together our data reveal the nuclear factor NR4A1 as a brake of intrinsic neutrophil activity controlling neutrophil-mediated brain inflammation and neurotoxicity in stroke which may serve as a novel therapeutic target to limit inflammation-associated augmentation of ischemic damage after stroke.

neuroscience↗

bioRxiv2025

Lifetime physical activity and network attack tolerance contribute to the preservation of motor function in Parkinson's disease

We tested whether network resilience, quantified by network attack tolerance (NAT), is associated with dopamine terminal (DaT) integrity, motor function and lifetime factors in Parkinsons disease (PD). Data from 22 PD patients and 39 healthy controls included information on lifetime physical activity (PA), cognitive/motor performance, putaminal DaT integrity, and resting-state fMRI. NAT was assessed at global and subnetwork level by calculating global efficiency upon iterative node removal. Generalized linear-mixed-effects models were used to test the effects of PA, education, and dopamine integrity on NAT. Next, the moderating effect of lifetime factors on the association between NAT and motor function were assessed, controlling for DaT integrity. Greater putaminal DaT integrity was linked to higher somatomotor NAT. Higher global and somatomotor NAT supported motor function, especially in patients with moderate lifetime PA. Lifestyle factors may thus serve network-specific attack tolerance, thereby promoting motor preservation in PD, independent of dopaminergic impairment.

neuroscience↗

bioRxiv2022

Peyers Patch B cells undergo cell death via neutrophil-released toxic DNA following sterile tissue injury

Lymphocyte contraction (LC) in central immune organs is a concomitant of sterile tissue injury, for example after stroke. Intestinal Peyers patches (PP) harbor large numbers of B cells, but how sterile tissue injury leads to LC in PP has not been explored. We observed rapid and macroscopically evident shrinkage of PP after stroke and myocardial infarction. Light-sheet fluorescence microscopy and flow cytometry revealed a strong reduction in the number of PP-resident B cells. Mechanistically, tissue injury triggered the activation of neutrophils that released B cell-toxic neutrophil extracellular traps (NETs) decorated with citrullinated histone-H3. Antibody-mediated or genetically induced neutrophil-loss, NETs-degradation or blockade of their generation completely reversed B cell loss and preserved the tissue architecture of PP. We also found NET-like elements in human post-stroke plasma. Hence, we propose that targeting NET-generation or -function counteracts post-injury B cell contraction in PP and thereby maintains immune homeostasis at mucosal barriers. In briefHigh numbers of B cells reside in the intestinal Peyers patches. Tuz et al. revealed that in response to sterile tissue injury, activated neutrophils release histone-decorated DNA into the circulation which induces B cell death. The loss of B cells results in the shrinkage of Peyers patches and reduced amounts of secretory IgA. HighlightsO_LIStroke and myocardial infarction induce the melting of Peyers patch C_LIO_LILight-sheet microscopy and cytometry revealed B cell loss in Peyers patch C_LIO_LIPost-injury activated neutrophils release NETs and trigger B cell death C_LIO_LIInhibition of NETs rescues B cell loss and degeneration of Peyers patch C_LI

immunology↗

bioRxiv2022

Rapid and fully automated blood vasculature analysis in 3D light-sheet image volumes of different organs

Blood vasculature represents a complex network of vessels with varying lengths and diameters that are precisely organized in space to allow proper tissue function. Light-sheet fluorescence microscopy (LSFM) is very useful to generate tomograms of tissue vasculature with high spatial accuracy. Yet, quantitative LSFM analysis is still cumbersome and available methods are restricted to single organs and advanced computing hardware. Here, we introduce VesselExpress, an automated software that reliably analyzes six characteristic vascular network parameters including vessel diameter in LSFM data on average computing hardware. VesselExpress is ~100 times faster than other existing vessel analysis tools, requires no user interaction, integrates batch processing, and parallelization. Employing an innovative dual Frangi filter approach we show that obesity induces a large-scale modulation of brain vasculature in mice and that seven other major organs differ strongly in their 3D vascular makeup. Hence, VesselExpress transforms LSFM from an observational to an analytical working tool.

bioinformatics↗

bioRxiv2021

Acid sphingomyelinase deactivation post-ischemia/ reperfusion promotes cerebral angiogenesis and brain remodeling via small extracellular vesicles

Functional inhibitors of acid sphingomyelinase are clinically used as anti-depressants since [~]60 years. Here, we show that acid sphingomyelinase inhibition by the antidepressants amitriptyline, fluoxetine and desipramine protects from ischemia/reperfusion and elicits a profound brain remodeling response with increased angiogenesis, improved blood-brain barrier integrity, reduced brain leukocyte infiltration and increased neuronal survival. Angiogenesis is promoted by small extracellular vesicles with bona fide characteristics of exosomes, which are released from endothelial cells and which constitute an elegant target for the amplification of stroke recovery.

neuroscience↗