ABSTRACT
The pattern of vascular remodelling in relation to recovery after stroke remains largely unclear. We used steady-state contrast-enhanced magnetic resonance imaging to assess the development of cerebral blood volume and microvascular density in perilesional and exofocal areas from (sub)acutely to chronically after transient stroke in rats. Microvascular density was verified histologically after infusion with Evans Blue dye. At day 1, microvascular cerebral blood volume and microvascular density were reduced in and around the ischemic lesion (intralesional borderzone: microvascular cerebral blood volume = 72 ± 8%; microvascular density = 76 ± 8%) (P < 0.05), while total cerebral blood volume remained relatively unchanged. Perilesional microvascular cerebral blood volume and microvascular density subsequently normalized (day 7) and remained relatively stable (day 70). In remote ipsilateral areas in the thalamus and substantia nigra - not part of the ischemic lesion - microvascular density gradually increased between days 1 and 70 (thalamic ventral posterior nucleus: microvascular density = 119 ± 9%; substantia nigra: microvascular density = 122 ± 8% (P < 0.05)), which was confirmed histologically. Our data indicate that initial microvascular collapse, with maintained collateral flow in larger vessels, is followed by dynamic revascularization in perilesional tissue. Furthermore, progressive neovascularization in non-ischemic connected areas may offset secondary neuronal degeneration and/or contribute to non-neuronal tissue remodelling. The complex spatiotemporal pattern of vascular remodelling, involving regions outside the lesion territory, may be a critical endogenous process to promote post-stroke brain reorganization.
Subject(s)
Brain/diagnostic imaging , Magnetic Resonance Imaging/methods , Stroke/diagnostic imaging , Stroke/physiopathology , Vascular Remodeling/physiology , Acute Disease , Animals , Brain/blood supply , Cerebrovascular Circulation/physiology , Chronic Disease , Disease Models, Animal , Male , Microvessels/diagnostic imaging , Microvessels/physiopathology , Neovascularization, Physiologic , Rats, Wistar , Substantia Nigra/diagnostic imaging , Substantia Nigra/physiopathology , Thalamus/diagnostic imaging , Thalamus/physiopathologyABSTRACT
Impaired actin filament dynamics have been associated with cellular senescence. Microglia, the resident immune cells of the brain, are emerging as a central pathophysiological player in neurodegeneration. Microglia activation, which ranges on a continuum between classical and alternative, may be of critical importance to brain disease. Using genetic and pharmacological manipulations, we studied the effects of alterations in actin dynamics on microglia effector functions. Disruption of actin dynamics did not affect transcription of genes involved in the LPS-triggered classical inflammatory response. By contrast, in consequence of impaired nuclear translocation of phospho-STAT6, genes involved in IL-4 induced alternative activation were strongly downregulated. Functionally, impaired actin dynamics resulted in reduced NO secretion and reduced release of TNFalpha and IL-6 from LPS-stimulated microglia and of IGF-1 from IL-4 stimulated microglia. However, pathological stabilization of the actin cytoskeleton increased LPS-induced release of IL-1beta and IL-18, which belong to an unconventional secretory pathway. Reduced NO release was associated with decreased cytoplasmic iNOS protein expression and decreased intracellular arginine uptake. Furthermore, disruption of actin dynamics resulted in reduced microglia migration, proliferation and phagocytosis. Finally, baseline and ATP-induced [Ca(2+)]int levels were significantly increased in microglia lacking gelsolin, a key actin-severing protein. Together, the dynamic state of the actin cytoskeleton profoundly and distinctly affects microglia behaviours. Disruption of actin dynamics attenuates M2 polarization by inhibiting transcription of alternative activation genes. In classical activation, the role of actin remodelling is complex, does not relate to gene transcription and shows a major divergence between cytokines following conventional and unconventional secretion.
Subject(s)
Actins/metabolism , Microglia/metabolism , Animals , Calcium/metabolism , Cell Proliferation , Cells, Cultured , Chemotaxis , Cytokines/metabolism , Encephalitis/chemically induced , Encephalitis/metabolism , Gelsolin/genetics , Interleukin-4/metabolism , Lipopolysaccharides , Mice , Mice, Knockout , Nitrogen Oxides/metabolism , Phagocytosis , STAT6 Transcription Factor/metabolismABSTRACT
Ambivalent effects of interleukin-6 on the pathogenesis of ischaemic stroke have been reported. However, to date, the long-term actions of interleukin-6 after stroke have not been investigated. Here, we subjected interleukin-6 knockout (IL-6(-/-)) and wild-type control mice to mild brain ischaemia by 30-min filamentous middle cerebral artery occlusion/reperfusion. While ischaemic tissue damage was comparable at early time points, IL-6(-/-) mice showed significantly increased chronic lesion volumes as well as worse long-term functional outcome. In particular, IL-6(-/-) mice displayed an impaired angiogenic response to brain ischaemia with reduced numbers of newly generated endothelial cells and decreased density of perfused microvessels along with lower absolute regional cerebral blood flow and reduced vessel responsivity in ischaemic striatum at 4 weeks. Similarly, the early genomic activation of angiogenesis-related gene networks was strongly reduced and the ischaemia-induced signal transducer and activator of transcription 3 activation observed in wild-type mice was almost absent in IL-6(-/-) mice. In addition, systemic neoangiogenesis was impaired in IL-6(-/-) mice. Transplantation of interleukin-6 competent bone marrow into IL-6(-/-) mice (IL-6(chi)) did not rescue interleukin-6 messenger RNA expression or the early transcriptional activation of angiogenesis after stroke. Accordingly, chronic stroke outcome in IL-6(chi) mice recapitulated the major effects of interleukin-6 deficiency on post-stroke regeneration with significantly enhanced lesion volumes and reduced vessel densities. Additional in vitro experiments yielded complementary evidence, which showed that after stroke resident brain cells serve as the major source of interleukin-6 in a self-amplifying network. Treatment of primary cortical neurons, mixed glial cultures or immortalized brain endothelia with interleukin 6-induced robust interleukin-6 messenger RNA transcription in each case, whereas oxygen-glucose deprivation did not. However, oxygen-glucose deprivation of organotypic brain slices resulted in strong upregulation of interleukin-6 messenger RNA along with increased transcription of key angiogenesis-associated genes. In conclusion, interleukin-6 produced locally by resident brain cells promotes post-stroke angiogenesis and thereby affords long-term histological and functional protection.
Subject(s)
Infarction, Middle Cerebral Artery/complications , Interleukin-6/metabolism , Neovascularization, Pathologic/etiology , Analysis of Variance , Angiogenic Proteins/genetics , Angiogenic Proteins/metabolism , Animals , Bone Marrow Transplantation/methods , Brain/pathology , Calcium-Binding Proteins/metabolism , Cells, Cultured , Cerebral Cortex/cytology , Cytokine Receptor gp130/genetics , Cytokine Receptor gp130/metabolism , Cytokines/genetics , Cytokines/metabolism , Disease Models, Animal , Embryo, Mammalian , Endothelial Cells/pathology , Endothelin-1/genetics , Endothelin-1/metabolism , Enzyme-Linked Immunosorbent Assay , Gait Disorders, Neurologic/etiology , Gene Expression Profiling , Gene Expression Regulation/genetics , Glucose/deficiency , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Hypoxia/complications , Infarction, Middle Cerebral Artery/pathology , Infarction, Middle Cerebral Artery/physiopathology , Infarction, Middle Cerebral Artery/surgery , Interleukin-6/genetics , Mice , Mice, Knockout/genetics , Microfilament Proteins/metabolism , Neovascularization, Pathologic/metabolism , Neuroglia/physiology , Neurons/drug effects , Oligonucleotide Array Sequence Analysis , Perfusion Imaging , Receptor, trkB/genetics , Receptor, trkB/metabolism , Rotarod Performance Test , STAT3 Transcription Factor/genetics , STAT3 Transcription Factor/metabolism , Signal Transduction/genetics , Signal Transduction/physiology , Tetrazolium Salts , ThiazolesABSTRACT
OBJECTIVES: The purpose of this study was to evaluate vascular effects of diet supplementation with plant sterol esters (PSE). BACKGROUND: Plant sterol esters are used as food supplements to reduce cholesterol levels. Their effects on endothelial function, stroke, or atherogenesis are not known. METHODS: In mice, plasma sterol concentrations were correlated with endothelial function, cerebral lesion size, and atherosclerosis. Plasma and tissue sterol concentrations were measured by gas-liquid chromatography-mass spectrometry in 82 consecutive patients with aortic stenosis. RESULTS: Compared with those fed with normal chow (NC), wild-type mice fed with NC supplemented with 2% PSE showed increased plant sterol but equal cholesterol plasma concentrations. The PSE supplementation impaired endothelium-dependent vasorelaxation and increased cerebral lesion size after middle cerebral artery occlusion. To test the effects of cholesterol-lowering by PSE, apolipoprotein E (ApoE)-/- mice were randomized to Western-type diet (WTD) with the addition of PSE or ezetimibe (EZE). Compared with WTD, both interventions reduced plaque sizes; however, WTD + PSE showed larger plaques compared with WTD + EZE (20.4 +/- 2.1% vs. 10.0 +/- 1.5%). Plant sterol plasma concentration strongly correlated with increased atherosclerotic lesion formation (r = 0.50). Furthermore, we examined plasma and aortic valve concentrations of plant sterol in 82 consecutive patients with aortic stenosis. Patients eating PSE-supplemented margarine (n = 10) showed increased plasma concentrations and 5-fold higher sterol concentrations in aortic valve tissue. CONCLUSIONS: Food supplementation with PSE impairs endothelial function, aggravates ischemic brain injury, effects atherogenesis in mice, and leads to increased tissue sterol concentrations in humans. Therefore, prospective studies are warranted that evaluate not only effects on cholesterol reduction, but also on clinical endpoints.