Neutrophils in aging and aging-related pathologies.

Over the past millennia, life expectancy has drastically increased. While a mere 25 years during Bronze and Iron ages, life expectancy in many European countries and in Japan is currently above 80 years. Such an increase in life expectancy is a result of improved diet, life style, and medical care. Yet, increased life span and aging also represent the most important non-modifiable risk factors for several pathologies including cardiovascular disease, neurodegenerative diseases, and cancer. In recent years, neutrophils have been implicated in all of these pathologies. Hence, this review provides an overview of how aging impacts neutrophil production and function and conversely how neutrophils drive aging-associated pathologies. Finally, we provide a perspective on how processes of neutrophil-driven pathologies in the context of aging can be targeted therapeutically.

Stroke in Patients with Bacterial Meningitis: A Cohort Study and Meta-Analysis.

OBJECTIVE: The purpose of this study was to characterize patients with ischemic stroke due to bacterial meningitis. METHODS: In a single-center retrospective study, we analyzed 102 patients with bacterial meningitis of which 19 had an ischemic stroke. Clinical characteristics, cerebrospinal fluid (CSF) analyses, and spatiotemporal distribution of infarcts were assessed. In addition, we searched PubMed from database inception to August 2021 for observational studies on ischemic stroke in patients with bacterial meningitis, and performed a meta-analysis to investigate the frequency and timing of stroke as well as its effect on mortality. RESULTS: In our cohort, 15 (78.9%) patients with stroke had an modified Rankin scale (mRS)  ≥  3 at discharge compared to 33 (39.8%) in patients without stroke (p < 0.01). Of 1,692 patients with bacterial meningitis from 15 cohort studies included in our meta-analysis, cerebral infarcts were found in 332 (16%, 95% confidence interval [CI] = 0.13-0.20) patients. The occurrence of stroke was strongly associated with a higher mortality (odds ratio [OR] = 2.38, 95% CI = 1.70-3.34, p < 0.0001). There was no association of any specific causative pathogen with the occurrence of stroke. Infarcts were mainly distributed in territories of arteries located in the vicinity to the infection focus and peaked at 3 to -7 days and at 2 weeks after onset of meningitis. In patients with ischemic stroke, vasculopathy was found in 63.2% and additional intracerebral hemorrhage in 15.8%. INTERPRETATION: This study found that ischemic stroke due to bacterial meningitis is caused by cerebral vasculopathy located in the vicinity of the infection focus, and that the time course of infarctions might enable a therapeutic intervention. ANN NEUROL 2023;93:1094-1105.

Bcl6 controls meningeal Th17-B cell interaction in murine neuroinflammation.

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.

Endurance Exercise Attenuates Established Progressive Experimental Autoimmune Encephalomyelitis and Is Associated with an Amelioration of Innate Immune Responses in NOD Mice.

Multiple sclerosis (MS) is a chronic inflammatory autoimmune disease causing axonal degeneration and demyelination. Exercise in mice with active monophasic experimental autoimmune encephalomyelitis (EAE) attenuates disease severity associated with diverse impacts on T cell-mediated immunity. However, studies have so far focused on preventive approaches. In this study, we investigated the impact of endurance exercise on established EAE disease in a model of secondary progressive MS. When the exercise program on motorized running wheels was started at disease manifestation, the disease course was significantly ameliorated. This was associated with a significant decrease in B cell, dendritic cell, and neutrophil cell counts in the central nervous system (CNS). Furthermore, we observed an increased expression of major histocompatibility complex class II (MHC-II) as well as alterations in costimulatory molecule expression in CNS B cells and dendritic cells. In contrast, T cell responses were not altered in the CNS or periphery. Thus, exercise training is capable of attenuating the disease course even in established secondary progressive EAE, potentially via modulation of the innate immune compartment. Further studies are warranted to corroborate our findings and assess the potential of this lifestyle intervention as a complementary therapeutic strategy in secondary progressive MS patients.

Characterization of Extracranial Giant Cell Arteritis with Intracranial Involvement and its Rapidly Progressive Subtype.

OBJECTIVE: The objective of this study was to characterize patients with extracranial giant cell arteritis with intracranial involvement. METHODS: In a multicenter retrospective study, we included 31 patients with systemic giant cell arteritis (GCA) with intracranial involvement. Clinical characteristics, pattern of arterial involvement, and cytokine profiles were assessed. Patients with GCA without intracranial involvement (n = 17), and with intracranial atherosclerosis (n = 25) served as controls. RESULTS: Erythrocyte sedimentation rate (ESR) was elevated in 18 patients (69.2%) with and in 16 patients (100%) without intracranial involvement (p = 0.02). Headache was complained by 15 patients (50.0%) with and 13 patients (76.5%) without intracranial involvement (p = 0.03). Posterior circulation arteries were affected in 26 patients (83.9%), anterior circulation arteries in 17 patients (54.8%), and both territories in 12 patients (38.7%). Patients with GCA had vertebral artery stenosis proximal and, in contrast, patients with atherosclerosis distal to the origin of posterior inferior cerebellar artery (PICA). Among patients with GCA with intracranial involvement, 11 patients (37.9%) had a rapid progressive disease course characterized by short-term recurrent ischemic events. The median modified Rankin Scale (mRS) at follow-up in these patients was 4 (interquartile range [IQR] = 2.0-6.0) and 4 patients (36.4%) died. Vessel wall expression of IL-6 and IL-17 was significantly increased in patients with rapid progressive course. INTERPRETATION: Typical characteristics of GCA, headache, and an elevated ESR, are frequently absent in patients with intracranial involvement. However, differentiation of intracranial GCA from atherosclerosis can be facilitated by the typical pattern of vertebral artery stenosis. About one-third of patients with intracranial GCA had a rapid progressive course with poor outcome. IL-17 and IL-6 may represent potential future treatment targets. ANN NEUROL 2021;90:118-129.

Rivaroxaban Reduces Arterial Thrombosis by Inhibition of FXa-Driven Platelet Activation via Protease Activated Receptor-1.

RATIONALE: A reduced rate of myocardial infarction has been reported in patients with atrial fibrillation treated with FXa (factor Xa) inhibitors including rivaroxaban compared with vitamin K antagonists. At the same time, low-dose rivaroxaban has been shown to reduce mortality and atherothrombotic events in patients with coronary artery disease. Yet, the mechanisms underlying this reduction remain unknown. OBJECTIVE: In this study, we hypothesized that rivaroxaban's antithrombotic potential is linked to a hitherto unknown rivaroxaban effect that impacts on platelet reactivity and arterial thrombosis. METHODS AND RESULTS: In this study, we identified FXa as potent, direct agonist of the PAR-1 (protease-activated receptor 1), leading to platelet activation and thrombus formation, which can be inhibited by rivaroxaban. We found that rivaroxaban reduced arterial thrombus stability in a mouse model of arterial thrombosis using intravital microscopy. For in vitro studies, atrial fibrillation patients on permanent rivaroxaban treatment for stroke prevention, respective controls, and patients with new-onset atrial fibrillation before and after first intake of rivaroxaban (time series analysis) were recruited. Platelet aggregation responses, as well as thrombus formation under arterial flow conditions on collagen and atherosclerotic plaque material, were attenuated by rivaroxaban. We show that rivaroxaban's antiplatelet effect is plasma dependent but independent of thrombin and rivaroxaban's anticoagulatory capacity. CONCLUSIONS: Here, we identified FXa as potent platelet agonist that acts through PAR-1. Therefore, rivaroxaban exerts an antiplatelet effect that together with its well-known potent anticoagulatory capacity might lead to reduced frequency of atherothrombotic events and improved outcome in patients.

Why Most Acute Stroke Studies Are Positive in Animals but Not in Patients: A Systematic Comparison of Preclinical, Early Phase, and Phase 3 Clinical Trials of Neuroprotective Agents.

OBJECTIVE: To analyze why numerous acute stroke treatments were successful in the laboratory but failed in large clinical trials. METHODS: We searched all phase 3 trials of medical treatments for acute ischemic stroke and corresponding early clinical and experimental studies. We compared the overall efficacy and assessed the impact of publication bias and study design on the efficacy. Furthermore, we estimated power and true report probability of experimental studies. RESULTS: We identified 50 phase 3 trials with 46,008 subjects, 75 early clinical trials with 12,391 subjects, and 209 experimental studies with >7,141 subjects. Three (6%) phase 3, 24 (32%) early clinical, and 143 (69.08%) experimental studies were positive. The mean treatment effect was 0.76 (95% confidence interval [CI] = 0.70-0.83) in experimental studies, 0.87 (95% CI = 0.71-1.06) in early clinical trials, and 1.00 (95% CI = 0.95-1.06) in phase 3 trials. Funnel plot asymmetry and trim-and-fill revealed a clear publication bias in experimental studies and early clinical trials. Study design and adherence to quality criteria had a considerable impact on estimated effect sizes. The mean power of experimental studies was 17%. Assuming a bias of 30% and pre-study odds of 0.5 to 0.7, this leads to a true report probability of <50%. INTERPRETATION: Pivotal study design differences between experimental studies and clinical trials, including different primary end points and time to treatment, publication bias, neglected quality criteria and low power, contribute to the stepwise efficacy decline of stroke treatments from experimental studies to phase 3 clinical trials. Even under conservative estimates, less than half of published positive experimental stroke studies are truly positive. ANN NEUROL 2020;87:40-51.

Heart Failure Results in Inspiratory Muscle Dysfunction Irrespective of Left Ventricular Ejection Fraction.

BACKGROUND: Exercise intolerance in heart failure with reduced ejection fraction (HFrEF) or heart failure with preserved ejection fraction (HFpEF) results from both cardiac dysfunction and skeletal muscle weakness. Respiratory muscle dysfunction with restrictive ventilation disorder may be present irrespective of left ventricular ejection fraction and might be mediated by circulating pro-inflammatory cytokines. OBJECTIVE: To determine lung and respiratory muscle function in patients with HFrEF/HFpEF and to determine its associations with exercise intolerance and markers of systemic inflammation. METHODS: Adult patients with HFrEF (n = 22, 19 male, 61 ± 14 years) and HFpEF (n = 8, 7 male, 68 ± 8 years) and 19 matched healthy control subjects underwent spirometry, measurement of maximum mouth occlusion pressures, diaphragm ultrasound, and recording of transdiaphragmatic and gastric pressures following magnetic stimulation of the phrenic nerves and the lower thoracic nerve roots. New York Heart Association (NYHA) class and 6-min walking distance (6MWD) were used to quantify exercise intolerance. Levels of circulating interleukin 6 (IL-6) and tumor necrosis factor-α (TNF-α) were measured using ELISAs. RESULTS: Compared with controls, both patient groups showed lower forced vital capacity (FVC) (p < 0.05), maximum inspiratory pressure (PImax), maximum expiratory pressure (PEmax) (p < 0.05), diaphragm thickening ratio (p = 0.01), and diaphragm strength (twitch transdiaphragmatic pressure in response to supramaximal cervical magnetic phrenic nerve stimulation) (p = 0.01). In patients with HFrEF, NYHA class and 6MWD were both inversely correlated with FVC, PImax, and PEmax. In those with HFpEF, there was an inverse correlation between amino terminal pro B-type natriuretic peptide levels and FVC (r = -0.77, p = 0.04). In all HF patients, IL-6 and TNF-α were statistically related to FVC. CONCLUSIONS: Irrespective of left ventricular ejection fraction, HF is associated with respiratory muscle dysfunction, which is associated with increased levels of circulating IL-6 and TNF-α.

Respiratory Muscle and Lung Function in Lung Allograft Recipients: Association with Exercise Intolerance.

BACKGROUND: In lung transplant recipients (LTRs), restrictive ventilation disorder may be present due to respiratory muscle dysfunction that may reduce exercise capacity. This might be mediated by pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). OBJECTIVE: We investigated lung respiratory muscle function as well as circulating pro-inflammatory cytokines and exercise capacity in LTRs. METHODS: Fifteen LTRs (6 female, age 56 ± 14 years, 63 ± 45 months post-transplantation) and 15 healthy controls matched for age, sex, and body mass index underwent spirometry, measurement of mouth occlusion pressures, diaphragm ultrasound, and recording of twitch transdiaphragmatic (twPdi) and gastric pressures (twPgas) following magnetic stimulation of the phrenic nerves and the lower thoracic nerve roots. Exercise capacity was quantified using the 6-min walking distance (6MWD). Plasma IL-6 and TNF-α were measured using enzyme-linked immunosorbent assays. RESULTS: Compared with controls, patients had lower values for forced vital capacity (FVC; 81 ± 30 vs.109 ± 18% predicted, p = 0.01), maximum expiratory pressure (100 ± 21 vs.127 ± 17 cm H2O, p = 0.04), diaphragm thickening ratio (2.2 ± 0.4 vs. 3.0 ± 1.1, p = 0.01), and twPdi (10.4 ± 3.5 vs. 17.6 ± 6.7 cm H2O, p = 0.01). In LTRs, elevation of TNF-α was related to lung function (13 ± 3 vs. 11 ± 2 pg/mL in patients with FVC ≤80 vs. >80% predicted; p < 0.05), and lung function (forced expiratory volume after 1 s) was closely associated with diaphragm thickening ratio (r = 0.81; p < 0.01) and 6MWD (r = 0.63; p = 0.02). CONCLUSION: There is marked restrictive ventilation disorder and respiratory muscle weakness in LTRs, especially inspiratory muscle weakness with diaphragm dysfunction. Lung function impairment relates to elevated levels of circulating TNF-α and diaphragm dysfunction and is associated with exercise intolerance.

Defining mechanisms of neural plasticity after brainstem ischemia in rats.

OBJECTIVE: Neurological recovery after stroke mainly depends on the location of the lesion. A substantial portion of strokes affects the brainstem. However, patterns of neural plasticity following brainstem ischemia are almost unknown. METHODS: Here, we established a rat brainstem ischemia model that resembles key features of the human disease and investigated mechanisms of neural plasticity, including neurogenesis and axonal sprouting as well as secondary neurodegeneration. RESULTS: Spontaneous functional recovery was accompanied by a distinct pattern of axonal sprouting, for example, an increased bilateral fiber outgrowth from the corticorubral tract to the respective contralesional red nucleus suggesting a compensatory role of extrapyramidal pathways after damage to pyramid tracts within the brainstem. Using different markers for DNA replication, we showed that the brainstem displays a remarkable ability to undergo specific plastic cellular changes after injury, highlighting a yet unknown pattern of neurogenesis. Neural progenitor cells proliferated within the dorsal brainstem and migrated toward the lesion, whereas neurogenesis in classic neurogenic niches, the subventricular zone of the lateral ventricle and the hippocampus, remained, in contrast to what is known from hemispheric stroke, unaffected. These beneficial changes were paralleled by long-term degenerative processes, that is, corticospinal fiber loss superior to the lesion, degeneration of spinal tracts, and a decreased neuron density within the ipsilesional substantia nigra and the contralesional red nucleus that might have limited further functional recovery. INTERPRETATION: Our findings provide knowledge of elementary plastic adaptions after brainstem stroke, which is fundamental for understanding the human disease and for the development of new treatments. Ann Neurol 2018;83:1003-1015.

Targeting Different Monocyte/Macrophage Subsets Has No Impact on Outcome in Experimental Stroke.

BACKGROUND AND PURPOSE: Peripheral immune cell infiltration contributes to neural injury after ischemic stroke. However, in contrast to lymphocytes and neutrophils, the role of different monocyte/macrophage subsets remains to be clarified. Therefore, we evaluated the effects of selective and unselective monocyte/macrophage depletion and proinflammatory (M1-) and anti-inflammatory (M2-) macrophage transfer on the outcome after experimental cerebral ischemia. METHODS: To assess short-term effects of monocytes/macrophages in acute ischemic stroke, mice underwent transient middle cerebral artery occlusion and received either clodronate liposomes for unselective macrophage depletion, MC-21-antibody for selective depletion of proinflammatory Ly-6Chigh monocytes, or proinflammatory (M1-) or anti-inflammatory (M2-) macrophage transfer. In addition, the impact of MC-21-antibody administration and M2-macrophage transfer on long-term neural recovery was investigated after photothrombotic stroke. Neurobehavioral tests were used to analyze functional outcomes, infarct volumes were determined, and immunohistochemical analyses were performed to characterize the postischemic inflammatory reaction. RESULTS: Selective and unselective monocyte/macrophage depletion and M1- and M2-macrophage transfer did not influence tissue damage and neurobehavioral outcomes in the acute phase after middle cerebral artery occlusion. Beyond, selective depletion of Ly-6Chigh monocytes and M2-macrophage transfer did not have an impact on neural recovery after photothrombotic stroke. CONCLUSIONS: Targeting different monocyte/macrophage subsets has no impact on outcome after ischemic stroke in mice. Altogether, our study could not identify monocytes/macrophages as relevant therapeutic targets in acute ischemic stroke.

Combining Growth Factor and Bone Marrow Cell Therapy Induces Bleeding and Alters Immune Response After Stroke in Mice.

BACKGROUND AND PURPOSE: Bone marrow cell (BMC)-based therapies, either the transplantation of exogenous cells or stimulation of endogenous cells by growth factors like the granulocyte colony-stimulating factor (G-CSF), are considered a promising means of treating stroke. In contrast to large preclinical evidence, however, a recent clinical stroke trial on G-CSF was neutral. We, therefore, aimed to investigate possible synergistic effects of co-administration of G-CSF and BMCs after experimental stroke in mice to enhance the efficacy compared with single treatments. METHODS: We used an animal model for experimental stroke as paradigm to study possible synergistic effects of co-administration of G-CSF and BMCs on the functional outcome and the pathophysiological mechanism. RESULTS: G-CSF treatment alone led to an improved functional outcome, a reduced infarct volume, increased blood vessel stabilization, and decreased overall inflammation. Surprisingly, the combination of G-CSF and BMCs abrogated G-CSFs' beneficial effects and resulted in increased hemorrhagic infarct transformation, altered blood-brain barrier, excessive astrogliosis, and altered immune cell polarization. These increased rates of infarct bleeding were mainly mediated by elevated matrix metalloproteinase-9-mediated blood-brain barrier breakdown in G-CSF- and BMCs-treated animals combined with an increased number of dilated and thus likely more fragile vessels in the subacute phase after cerebral ischemia. CONCLUSIONS: Our results provide new insights into both BMC-based therapies and immune cell biology and help to understand potential adverse and unexpected side effects.

Progressive cognitive deficits in a mouse model of recurrent photothrombotic stroke.

BACKGROUND AND PURPOSE: In spite of its high disease burden, there is no specific treatment for multi-infarct dementia. The preclinical evaluation of candidate drugs is limited because an appropriate animal model is lacking. Therefore, we aimed to evaluate whether a mouse model of recurrent photothrombotic stroke is suitable for the preclinical investigation of multi-infarct dementia. METHODS: Recurrent photothrombotic cortical infarcts were induced in 25 adult C57BL/6 mice. Twenty-five sham-operated animals served as controls. The object recognition test and the Morris water maze test were performed >6 weeks to assess cognitive deficits. Afterward, histological analyses were performed to characterize histopathologic changes associated with recurrent photothrombotic infarcts. RESULTS: After the first infarct, the object recognition test showed a trend toward an impaired formation of recognition memories (P=0.08), and the Morris Water Maze test revealed significantly impaired spatial learning and memory functions (P<0.05). After recurrent infarcts, the object recognition test showed significant recognition memory deficits (P<0.001) and the Morris water maze test demonstrated persisting spatial learning and memory deficits (P<0.05). Histological analyses revealed remote astrogliosis in the hippocampus. CONCLUSIONS: Our results show progressive cognitive deficits in a mouse model of recurrent photothrombotic stroke. The presented model resembles the clinical features of human multi-infarct dementia and enables the investigation of its pathophysiological mechanisms and the evaluation of treatment strategies.

Cortical photothrombotic infarcts impair the recall of previously acquired memories but spare the formation of new ones.

BACKGROUND AND PURPOSE: Despite a high incidence of poststroke dementia, there is no specific treatment for this condition. Because the evaluation of poststroke cognitive deficits in animal models of stroke is exceedingly challenging, the preclinical evaluation of candidate drugs is limited. We aimed to explore the impact of small cortical photothrombotic strokes on poststroke cognition, thereby assessing the suitability of this experimental stroke model for the investigation of cognitive impairment after stroke. METHODS: Photothrombotic cortical infarcts were induced in 19 adult male Wistar rats. Nineteen sham-operated animals served as controls. Using the Morris water maze, we analyzed the impact of photothrombotic stroke on both the acquisition of new memories and the recall of previously acquired memories. The cylinder test, the adhesive tape removal test, and the rotarod test were performed to investigate sensorimotor deficits. RESULTS: Photothrombotic stroke significantly impaired the recall of previously acquired memories (P<0.05), whereas the acquisition of new memories remained largely intact. The analysis of the animals' swimming speed in the water maze and the rotarod test showed no confounding motor impairments after photothrombotic stroke. The adhesive tape removal test and the cylinder test revealed mild sensorimotor deficits in lesioned animals (P<0.05). CONCLUSIONS: Photothrombotic cortical infarcts impair the recall of memories acquired before stroke, whereas the formation of new memories remains unimpaired. The observed deficits in the water maze are not confounded by disturbed motor functions. Overall, experimental photothrombotic strokes are well suited for the investigation of specific cognitive impairments after stroke.

Meta-analysis of the efficacy of different training strategies in animal models of ischemic stroke.

BACKGROUND AND PURPOSE: Although several studies have shown beneficial effects of training in animal stroke models, the most effective training strategy and the optimal time to initiate training have not been identified. The present meta-analysis was performed to compare the efficacy of different training strategies and to determine the optimal time window for training in animal stroke models. METHODS: We searched the literature for studies analyzing the efficacy of training in animal models of ischemic stroke. Training was categorized into forced physical training, voluntary physical training, constraint-induced movement therapy, and skilled reaching training. Two reviewers independently extracted data on study quality, infarct size, and neurological outcome. Data were pooled by means of a meta-analysis. RESULTS: Thirty-five studies with >880 animals were included. A meta-analysis of all treatments showed that training reduced the infarct volume by 14% (95% confidence interval, 2%-25%) and improved the cognitive function by 33% (95% confidence interval, 8%-50%), the neuroscore by 13.4% (95% confidence interval, 1.5%-25.3%), and the running function by 6.6% (95% confidence interval, 1.4%-11.9%). Forced physical training reduced the infarct volume and enhanced the running function most effectively, whereas skilled reaching training improved the limb function most effectively. A meta-regression illustrated that training was particularly efficacious when initiated between 1 and 5 days after stroke onset. CONCLUSIONS: Our meta-analysis confirms that training reduces the infarct volume and improves the functional recovery in animal stroke models. Forced physical training and skilled reaching training were identified as particularly effective training strategies. The efficacy of training is time dependent.

Inhibition of leukocyte migration after ischemic stroke by VE-cadherin mutation in a mouse model leads to reduced infarct volumes and improved motor skills.

AIMS: To distinguish between the genuine cellular impact of the ischemic cascade by leukocytes and unspecific effects of edema and humoral components, two knock-in mouse lines were utilized. Mouse lines Y731F and Y685F possess point mutations in VE-cadherin, which lead to a selective inhibition of transendothelial leukocyte migration or impaired vascular permeability. METHODS: Ischemic stroke was induced by a model of middle cerebral artery occlusion. Analysis contained structural outcomes (infarct volume and extent of brain edema), functional outcomes (survival analysis, rotarod test, and neuroscore), and the extent and spatial distribution of leukocyte migration (heatmaps and fluorescence-activated cell sorting (FACS) analysis). RESULTS: Inhibition of transendothelial leukocyte migration as in Y731F mice leads to smaller infarct volumes (52.33 ± 4719 vs. 70.43 ± 6483 mm3 , p = .0252) and improved motor skills (rotarod test: 85.52 ± 13.24 s vs. 43.06 ± 15.32 s, p = .0285). An impaired vascular permeability as in Y685F mice showed no effect on structural or functional outcomes. Both VE-cadherin mutations did not influence the total immune cell count or spatial distribution in ischemic brain parenchyma. CONCLUSION: Selective inhibition of transendothelial leukocyte migration by VE-cadherin mutation after ischemic stroke in a mouse model leads to smaller infarct volumes and improved motor skills.

Anti-LINGO-1 treatment restores myelination of corticospinal tract neurons and improves functional recovery after stroke.

Demyelination of corticospinal tract neurons contributes to long-term disability after cortical stroke. Nonetheless, poststroke myelin loss has not been addressed as a therapeutic target, so far. We hypothesized that an antibody-mediated inhibition of the Nogo receptor-interacting protein (LINGO-1, leucine-rich repeat and immunoglobulin domain-containing Nogo receptor-interacting protein) may counteract myelin loss, enhance remyelination and axonal growth, and thus promote functional recovery following stroke. To verify this hypothesis, mice were subjected to photothrombotic stroke and received either an antibody against LINGO-1 (n = 19) or a control treatment (n = 18). Behavioral tests were performed to assess the effects of anti-LINGO-1 treatment on the functional recovery. Seven weeks after stroke, immunohistochemical analyses were performed to analyze the effect of anti-LINGO-1 treatment on myelination and axonal loss of corticospinal tract neurons, proliferation of oligodendrocytes and neurogenesis. Anti-LINGO-1 treatment resulted in significantly improved functional recovery (p < 0.0001, repeated measures analysis of variance), and increased neurogenesis in the hippocampus and subventricular zone of the ipsilateral hemisphere (p = 0.0094 and p = 0.032, t-test). Notably, we observed a significant increase in myelin (p = 0.0295, t-test), platelet-derived growth factor receptor α-positive oligodendrocyte precursor cells (p = 0.0356, t-test) and myelinating adenomatous polyposis coli-positive cells within the ipsilateral internal capsule of anti-LINGO-1-treated mice (p = 0.0021, t-test). In conclusion, we identified anti-LINGO-1 as the first neuroregenerative treatment that counteracts poststroke demyelination of corticospinal tract neurons, presumably by increased proliferation of myelin precursor cells, and thereby improves functional recovery. Most importantly, our study presents myelin loss as a novel therapeutic target following stroke.

The dilemma of neuroprotection trials in times of successful endovascular recanalization.

Background: The "translational roadblock" between successful animal stroke studies and neutral clinical trials is usually attributed to conceptual weaknesses. However, we hypothesized that rodent studies cannot inform the human disease due to intrinsic pathophysiological differences between rodents and humans., i.e., differences in infarct evolution. Methods: To verify our hypothesis, we employed a mixed study design and compared findings from meta-analyses of animal studies and a retrospective clinical cohort study. For animal data, we systematically searched pubmed to identify all rodent studies, in which stroke was induced by MCAO and at least two sequential MRI scans were performed for infarct volume assessment within the first two days. For clinical data, we included 107 consecutive stroke patients with large artery occlusion, who received MRI scans upon admission and one or two days later. Results: Our preclinical meta-analyses included 50 studies with 676 animals. Untreated animals had a median post-reperfusion infarct volume growth of 74%. Neuroprotective treatments reduced this infarct volume growth to 23%. A retrospective clinical cohort study showed that stroke patients had a median infarct volume growth of only 2% after successful recanalization. Stroke patients with unsuccessful recanalization, by contrast, experienced a meaningful median infarct growth of 148%. Conclusion: Our study shows that rodents have a significant post-reperfusion infarct growth, and that this post-reperfusion infarct growth is the target of neuroprotective treatments. Stroke patients with successful recanalization do not have such infarct growth and thus have no target for neuroprotection.

A dietary intervention with conjugated linoleic acid enhances microstructural white matter reorganization in experimental stroke.

Background: A dietary supplementation with conjugated linoleic acid (CLA) was shown to attenuate inflammation and increase the proportions of circulating regulatory T cells (Tregs) and M2-type macrophages in disease models such as autoimmune encephalitis and arteriosclerosis. Since Tregs and anti-inflammatory (M2-type) macrophages were found to enhance stroke recovery, we hypothesized that CLA-supplementation might improve stroke recovery via immune modulatory effects. Methods: Functional assessment was performed over 90 days after induction of experimental photothrombotic stroke in wild type mice (n = 37, sham n = 10). Subsequently, immunological characterization of different immunological compartments (n = 16), ex vivo magnetic resonance (MR, n = 12) imaging and immunohistochemical staining (n = 8) was performed. Additionally, we tested the effect of CLA in vitro on peripheral blood mononuclear cells from human stroke patients and healthy controls (n = 12). Results: MR diffusion tensor imaging (DTI) demonstrated enhanced microstructural reorganization of interhemispheric white matter tracts, dependent on lesion size. Functional recovery over 90 days remained unaffected. Detailed immunological analyses across various compartments revealed no significant long-term immunological alterations due to CLA. However, analyses of human blood samples post-stroke showed reduced levels of pro-inflammatory interferon-γ (IFN-γ) and tumor necrosis factor alpha (TNF-α) release by T-lymphocytes following in vitro treatment with CLA. Conclusion: We aimed to explore the efficacy of a dietary intervention with minimal known side effects that could be accessible to human stroke patients, regardless of the degree of disability, and without the risks associated with aggressive immunomodulatory therapies. Our main findings include improved microstructural reorganization in small infarcts and a reduced inflammatory response of human T cells in vitro.