Abstract WP322: Do Mural Cells Differentiate Into Microglia-Like Cells After Ischemic Stroke?

Introduction: Controversial results exist on whether mural cells can differentiate into microglia-like cells after ischemic stroke. This discrepancy can be due to different experimental methods (immunostaining versus lineage tracing) and/or distinct transgenic animals (RGS5 GFP versus Tbx18-CreERT mice). Methods: To determinate whether mural cells are able to differentiate into microglia-like cells after ischemic stroke, we permanently labeled mural cells with tdTomato by crossing PDGFRβ-Cre and PDGFRβ-CreERT with Ai14 reporter mice. The resulting Ai14:PDGFRβ-Cre + and Ai14:PDGFRβ-CreERT + mice were subjected to 45 minutes of middle cerebral artery occlusion (MCAO) followed by reperfusion. At various time points after injury, the proliferation, apoptosis, and differentiation of PDGFRβ + cells were examined. Results: In both Ai14:PDGFRβ-Cre + and Ai14:PDGFRβ-CreERT + mice, we observed a substantial reduction of PDGFRβ + cells at day 2 after ischemic stroke and a subsequent repopulation (mainly due to proliferation) of PDGFRβ + cells at day 7 after ischemic stroke. We also showed that PDGFRβ + cells changed their morphology and differentiated into microglia-like cells at day 7 after injury, suggesting that PDGFRβ + cells can indeed differentiate into microglia-like cells after ischemic stroke. In addition, we noted that PDGFRβ also labeled Col1α1 + fibroblasts in the brain. Interestingly, high numbers of PDGFRβ + Col1α1 + cells were found in both Ai14:PDGFRβ-Cre + and Ai14:PDGFRβ-CreERT + mice at day 7 after ischemic injury. Conclusions: These results suggest that PDGFRβ is not an ideal marker for mural cells in pathological conditions that involve fibroblast activation. It remains unclear whether mural cells or fibroblasts differentiate into microglia-like cells after ischemic stroke. Future research should focus on answering this important question.

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Abstract 166: Endothelial Overexpression of LOX-1 Increases Stroke Size in vivo

Circulation Research ◽

10.1161/res.113.suppl_1.a166 ◽

2013 ◽

Vol 113 (suppl_1) ◽

Author(s):

Alexander Akhmedov ◽

Remo D Spescha ◽

Francesco Paneni ◽

Giovani G Camici ◽

Thomas F Luescher

Keyword(s):

Endothelial Cells ◽

Ischemic Stroke ◽

Endothelial Dysfunction ◽

Middle Cerebral Artery ◽

Middle Cerebral Artery Occlusion ◽

Cerebral Artery ◽

Oxidized Ldl ◽

Artery Occlusion ◽

Cerebral Artery Occlusion ◽

The Brain

Background— Stroke is one of the most common causes of death and long term disability worldwide primarily affecting the elderly population. Lectin-like oxidized LDL receptor 1 (LOX-1) is the receptor for oxidized LDL identified in endothelial cells. Binding of OxLDL to LOX-1 induces several cellular events in endothelial cells, such as activation of transcription factor NF-kB, upregulation of MCP-1, and reduction in intracellular NO. Accumulating evidence suggests that LOX-1 is involved in endothelial dysfunction, inflammation, atherogenesis, myocardial infarction, and intimal thickening after balloon catheter injury. Interestingly, a recent study demonstrated that acetylsalicylic acid (aspirin), which could prevent ischemic stroke, inhibited Ox-LDL-mediated LOX-1 expression in human coronary endothelial cells. The expression of LOX-1 was increased at a transient ischemic core site in the rat middle cerebral artery occlusion model. These data suggest that LOX-1 expression induces atherosclerosis in the brain and is the precipitating cause of ischemic stroke. Therefore, the goal of the present study was to investigate the role of endothelial LOX-1 in stroke using experimental mouse model. Methods and Results— 12-week-old male LOX-1TG generated recently in our group and wild-type (WT) mice were applied for a transient middle cerebral artery occlusion (MCAO) model to induce ischemia/reperfusion (I/R) brain injury. LOX-1TG mice developed 24h post-MCAO significantly larger infarcts in the brain compared to WT (81.51±8.84 vs. 46.41±10.13, n=7, p < 0.05) as assessed morphologically using Triphenyltetrazolium chloride (TTC) staining. Moreover, LOX-1TG showed higher neurological deficit in RotaRod (35.57±8.92 vs. 66.14±10.63, n=7, p < 0.05) and Bederson tests (2.22±0.14 vs. 1.25±0.30, n=9-12, p < 0.05) - two experimental physiological tests for neurological function. Conclusions— Thus, our data suggest that LOX-1 plays a critical role in the ischemic stroke when expressed at unphysiological levels. Such LOX-1 -associated phenotype could be due to the endothelial dysfunction. Therefore, LOX-1 may represent novel therapeutic targets for preventing ischemic stroke.

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P2Y6 receptor inhibition aggravates ischemic brain injury by reducing microglial phagocytosis

10.21203/rs.2.17542/v1 ◽

2019 ◽

Author(s):

Ruoxue Wen ◽

Hui Shen ◽

Shuxian Huang ◽

Liping Wang ◽

Zongwei Li ◽

...

Keyword(s):

Ischemic Stroke ◽

Myosin Light Chain ◽

Artery Occlusion ◽

Walking Test ◽

Microglial Phagocytosis ◽

Tnf Α ◽

Severity Scores ◽

P2y6 Receptor ◽

Cerebral Artery Occlusion ◽

The Brain

Abstract Background Clearance of damaged cells is beneficial for the functional recovery after brain injury. Phagocytosis of tissue and cell debris is an important function of microglia during the development and pathological diseases. However, which specific phagocytic receptor mediates microglial phagocytosis after ischemic stroke is obscure. Methods ICR mice (n=59) underwent 90 minutes transient middle cerebral artery occlusion. P2Y6R, Iba1, GFAP and Tuj-1 double immunostainings were performed to determine P2Y6 receptor location. MRS2578 was injected into mice to inhibit P2Y6 receptor activity. Iba1 and TUNEL staining were performed to examine microglia phagocytosis. Modified neurological severity scores and Grid walking test were used to evaluate the neurological function after ischemic stoke. The expression of IL-1 α, IL-1 β, IL-6, IL-10, TNF-α, TGF-β and MPO was used to examine the inflammation response in the brain. Results The expression of P2Y6 receptor in microglia increased within three days after transient middle cerebral artery occlusion. Inhibition of microglial phagocytosis by the selective inhibitor MRS2578 enlarged the brain atrophy and edema volume after ischemic stroke, subsequently aggravated neurological function using modified neurological severity scores and Grid walking test. MRS2578 treatment had no effect on the expression of IL-1α, IL-1β, IL-6, IL-10, TNF-α, TGF-β and MPO after ischemic stroke, which suggested that it had no effect on the inflammation in the brain. Finally, we found that the expression of myosin light chain kinase decreased after microglial phagocytosis inhibition in ischemic mice, which suggested that myosin light chain kinase was involved in P2Y6 receptor mediated phagocytosis. Conclusion Our results indicated that the P2Y6 receptor mediated microglial phagocytosis played an important role during the acute stage of ischemic stroke, which was a potential target for ischemic stroke treatment.

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Image-based stroke rat brain atrophy volume and infarct volume computation

The Journal of Supercomputing ◽

10.1007/s11227-020-03224-y ◽

2020 ◽

Vol 76 (12) ◽

pp. 10090-10121

Author(s):

Yung-Kuan Chan ◽

Chun-Fu Hong ◽

Meng-Hsiun Tsai ◽

Ya-Lan Chang ◽

Ping-Hsuan Sun

Keyword(s):

Ischemic Stroke ◽

Rat Brain ◽

Brain Slice ◽

Infarct Volume ◽

Brain Slices ◽

Artery Occlusion ◽

Tetrazolium Chloride ◽

Rat Brain Slice ◽

Cerebral Artery Occlusion ◽

The Brain

Abstract Stroke is one of the leading causes of death as well as results in a massive economic burden for society. Stroke is a cerebrovascular disease mainly divided into two types: ischemic stroke and hemorrhagic stroke, which, respectively, refer to the partial blockage and bleeding inside brain blood vessels. Both stroke types lead to nutrient and oxygen deprivation in the brain, which ultimately cause brain damage or death. This study focuses on ischemic stroke in rats with middle cerebral artery occlusion (MCAO) as experimental subjects, and the volumes of infarct and atrophy are calculated based on the brain slice images of rat brains stained with 2,3,5-triphenyl tetrazolium chloride. In this study, a stroke rat brain infarct and atrophy volumes computation system (SRBIAVC system) is developed to segment the infarcts and atrophies from the rat brain slice images. Based on the segmentation results, the infarct and atrophy volumes of a rat brain can be computed. In this study, 168 images of brain slices cut from 28 rat brains with MCAO are used as the test samples. The experimental results show that the segmentation results obtained by the SRBIAVC system are close to those obtained by experts.

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The KCa3.1 Blocker TRAM-34 Reduces Infarction and Neurological Deficit in a Rat Model of Ischemia/Reperfusion Stroke

Journal of Cerebral Blood Flow & Metabolism ◽

10.1038/jcbfm.2011.101 ◽

2011 ◽

Vol 31 (12) ◽

pp. 2363-2374 ◽

Cited By ~ 60

Author(s):

Yi-Je Chen ◽

Girija Raman ◽

Silke Bodendiek ◽

Martha E O'Donnell ◽

Heike Wulff

Keyword(s):

Ischemic Stroke ◽

Rat Model ◽

Neurological Deficit ◽

Ischemia Reperfusion ◽

Artery Occlusion ◽

Beneficial Effects ◽

Inflammatory Damage ◽

Male Wistar Rats ◽

Cerebral Artery Occlusion ◽

The Brain

Microglia and brain infiltrating macrophages significantly contribute to the secondary inflammatory damage in the wake of ischemic stroke. Here, we investigated whether inhibition of KCa3.1 (IKCa1/KCNN4), a calcium-activated K+ channel that is involved in microglia and macrophage activation and expression of which increases on microglia in the infarcted area, has beneficial effects in a rat model of ischemic stroke. Using an HPLC/MS assay, we first confirmed that our small molecule KCa3.1 blocker TRAM-34 effectively penetrates into the brain and achieves micromolar plasma and brain concentrations after intraperitoneal injection. Then, we subjected male Wistar rats to 90 minutes of middle cerebral artery occlusion (MCAO) and administered either vehicle or TRAM-34 (10 or 40 mg/kg intraperitoneally twice daily) for 7 days starting 12 hours after reperfusion. Both compound doses reduced infarct area by ∼50% as determined by hematoxylin & eosin staining on day 7 and the higher dose also significantly improved neurological deficit. We further observed a significant reduction in ED1+-activated microglia and TUNEL-positive neurons as well as increases in NeuN+ neurons in the infarcted hemisphere. Our findings suggest that KCa3.1 blockade constitutes an attractive approach for the treatment of ischemic stroke because it is still effective when initiated 12 hours after the insult.

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A Sphingosine 1-Phosphate Gradient Is Linked to the Cerebral Recruitment of T Helper and Regulatory T Helper Cells during Acute Ischemic Stroke

International Journal of Molecular Sciences ◽

10.3390/ijms21176242 ◽

2020 ◽

Vol 21 (17) ◽

pp. 6242

Author(s):

Alexandra Lucaciu ◽

Hannah Kuhn ◽

Sandra Trautmann ◽

Nerea Ferreirós ◽

Helmuth Steinmetz ◽

...

Keyword(s):

Ischemic Stroke ◽

Expression Patterns ◽

Treg Cells ◽

Artery Occlusion ◽

T Helper ◽

Sphingosine 1 Phosphate ◽

Cerebral Artery Occlusion ◽

Kinetics Of ◽

The Brain

Emerging evidence suggests a complex relationship between sphingosine 1-phosphate (S1P) signaling and stroke. Here, we show the kinetics of S1P in the acute phase of ischemic stroke and highlight accompanying changes in immune cells and S1P receptors (S1PR). Using a C57BL/6 mouse model of middle cerebral artery occlusion (MCAO), we assessed S1P concentrations in the brain, plasma, and spleen. We found a steep S1P gradient from the spleen towards the brain. Results obtained by qPCR suggested that cells expressing the S1PR type 1 (S1P1+) were the predominant population deserting the spleen. Here, we report the cerebral recruitment of T helper (TH) and regulatory T (TREG) cells to the ipsilateral hemisphere, which was associated with differential regulation of cerebral S1PR expression patterns in the brain after MCAO. This study provides insight that the S1P-S1PR axis facilitates splenic T cell egress and is linked to the cerebral recruitment of S1PR+ TH and TREG cells. Further insights by which means the S1P-S1PR-axis orchestrates neuronal positioning may offer new therapeutic perspectives after ischemic stroke.

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The Hsp90 Inhibitor 17-DMAG Attenuates Hyperglycemia-Enhanced Hemorrhagic Transformation in Experimental Stroke

BioMed Research International ◽

10.1155/2021/6668442 ◽

2021 ◽

Vol 2021 ◽

pp. 1-7

Author(s):

Jiaming Zhang ◽

Kai Wang ◽

Jia Qi ◽

Xiaodong Cao ◽

Feng Wang

Keyword(s):

Ischemic Stroke ◽

Hsp90 Inhibitor ◽

Hemorrhagic Transformation ◽

Artery Occlusion ◽

Vital Role ◽

Experimental Stroke ◽

Acute Hyperglycemia ◽

Inflammatory Molecules ◽

Cerebral Artery Occlusion ◽

The Brain

Introduction. Hemorrhagic transformation (HT) is one of the most common complications of ischemic stroke which is exacerbated by hyperglycemia. Oxidative stress, inflammatory response, and matrix metalloproteinases (MMPs) have been evidenced to play a vital role in the pathophysiology of HT. Our previous study has reported that 17-DMAG, an Hsp90 inhibitor, protects the brain against ischemic injury via inhibiting inflammation and reducing MMP-9 after ischemia. However, whether 17-DMAG would attenuate HT in hyperglycemic middle cerebral artery occlusion (MCAO) rats is still unknown. Methods. Acute hyperglycemia was induced by an injection of 50% dextrose. Rats were pretreated with 17-DMAG before MCAO. Infarction volume, hemorrhagic volume neurological scores, expressions of inflammatory molecules and tight junction proteins, and activity of MMP-2 and MMP-9 were assessed 24 h after MCAO. Results. 17-DMAG was found to reduce HT, improve neurological function, and inhibit expressions of inflammatory molecules and the activation of MMPs at 24 h after MCAO. Conclusion. These results implicated that Hsp90 could be a novel therapeutic target in HT following ischemic stroke.

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Improved reperfusion following alternative surgical approach for experimental stroke in mice

F1000Research ◽

10.12688/f1000research.22594.2 ◽

2020 ◽

Vol 9 ◽

pp. 188

Author(s):

Melissa Trotman-Lucas ◽

Raymond Wong ◽

Stuart M. Allan ◽

Claire L. Gibson

Keyword(s):

Ischemic Stroke ◽

Surgical Approach ◽

Artery Occlusion ◽

Laser Speckle ◽

Experimental Stroke ◽

Contrast Imaging ◽

Ipsilateral Hemisphere ◽

Cerebral Artery Occlusion ◽

The Brain

Background: Following ischemic stroke, recanalisation and restoration of blood flow to the affected area of the brain is critical and directly correlates with patient recovery. In vivo models of ischemic stroke show high variability in outcomes, which may be due to variability in reperfusion. We previously reported that a surgical refinement in the middle cerebral artery occlusion (MCAO) model of stroke, via repair of the common carotid artery (CCA), removes the reliance on the Circle of Willis for reperfusion and reduced infarct variability. Here we further assess this refined surgical approach on reperfusion characteristics following transient MCAO in mice. Methods: Mice underwent 60 min of MCAO, followed by either CCA repair or ligation at reperfusion. All mice underwent laser speckle contrast imaging at baseline, 24 h and 48 h post-MCAO. Results: CCA ligation reduced cerebral perfusion in the ipsilateral hemisphere compared to baseline (102.3 ± 4.57%) at 24 h (85.13 ± 16.09%; P < 0.01) and 48 h (75.04 ± 12.954%; P < 0.001) post-MCAO. Repair of the CCA returned perfusion to baseline (94.152 ± 2.44%) levels and perfusion was significantly improved compared to CCA ligation at both 24 h (102.83 ± 8.41%; P < 0.05) and 48 h (102.13 ± 9.34%; P < 0.001) post-MCAO. Conclusions: Our findings show CCA repair, an alternative surgical approach for MCAO, results in improved ischemic hemisphere perfusion during the acute phase.

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Accumulation of Amyloid Beta (Aβ) Peptide on Blood Vessel Walls in the Damaged Brain after Transient Middle Cerebral Artery Occlusion

Biomolecules ◽

10.3390/biom9080350 ◽

2019 ◽

Vol 9 (8) ◽

pp. 350 ◽

Cited By ~ 3

Author(s):

Antonio Martins ◽

Astrid Zayas-Santiago ◽

Yancy Ferrer-Acosta ◽

Solianne Martinez-Jimenez ◽

Lidia Zueva ◽

...

Keyword(s):

Ischemic Stroke ◽

Ion Channels ◽

Blood Vessel ◽

Middle Cerebral Artery ◽

Middle Cerebral Artery Occlusion ◽

Amyloid Beta ◽

Artery Occlusion ◽

Aβ Peptide ◽

Cerebral Artery Occlusion ◽

The Brain

It is well known that amyloid beta (Aβ) peptides are generated in blood vessels, released into the brain during thrombosis, and temporarily accumulate in this organ after injury. Here we demonstrate that 24 h after transient middle cerebral artery occlusion (tMCAO), one of the standard models of focal ischemic stroke, Aβ peptide accumulates in the brain, concentrating on the blood vessel walls. Because Aβ oligomers are known to induce significant damage to brain cells, they act as an additional damaging factor during ischemic stroke. Considering that they have been shown to form ion channels in cells, affecting osmotic balance, we used an Aβ peptide channel blocker, tromethamine (2-amino-2-(hydroxymethyl) propane-1,3-diol), to prevent this additional injury. Tromethamine injected 0.1 g/100 g body weight intraperitoneally at 5 min before tMCAO decreased water content in the damaged hemisphere, as measured by dry brain weight. Congo red staining, which binds only to Aβ oligomer plaques (amyloid), showed that there was no significant presence of plaques. Therefore, we suggest that Aβ peptide oligomers are responsible for some of the brain damage during stroke and that blockage of the ion channels that they form could be beneficial in treating this complex neurological syndrome.

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Improved reperfusion following alternative surgical approach for experimental stroke in mice

F1000Research ◽

10.12688/f1000research.22594.3 ◽

2020 ◽

Vol 9 ◽

pp. 188

Author(s):

Melissa Trotman-Lucas ◽

Raymond Wong ◽

Stuart M. Allan ◽

Claire L. Gibson

Keyword(s):

Ischemic Stroke ◽

Surgical Approach ◽

Artery Occlusion ◽

Laser Speckle ◽

Experimental Stroke ◽

Contrast Imaging ◽

Ipsilateral Hemisphere ◽

Cerebral Artery Occlusion ◽

The Brain

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Alterations of the pulmonary immunity are independent of spontaneous pneumonia in an experimental model of ischemic stroke

10.1101/618314 ◽

2019 ◽

Author(s):

Breanne Y. Farris ◽

Kelly L. Monaghan ◽

Courtney D. Amend ◽

Wen Zheng ◽

Heng Hu ◽

...

Keyword(s):

T Cells ◽

Ischemic Stroke ◽

Immune Cell ◽

Artery Occlusion ◽

Cell Fates ◽

Cytokines And Chemokines ◽

Lung Infections ◽

Pulmonary Immunity ◽

Cerebral Artery Occlusion ◽

The Brain

AbstractStroke-associated pneumonia (SAP) is a major cause of mortality in patients who have suffered from severe ischemic stroke. Although multi-factorial in nature, stroke-induced immunosuppression plays a key role in the development of SAP. Previous studies of focal ischemic stroke induction, using a murine model of transient middle cerebral artery occlusion (tMCAO) have shown that severe brain damage results in massive apoptosis and functional defects of lymphocytes in the spleen, thymus, and peripheral blood. However, how immune alternations in remote tissues lead to a greater susceptibility to lung infections is not well-understood. Importantly, how ischemic stroke alters immune-cell fates, and the expression of cytokines and chemokines in the lungs that directly impact pulmonary immunity, has not been characterized. We report here that ischemic stroke increases the percentage of alveolar macrophages, neutrophils, and CD11b+ dendritic cells (DCs), but reduces the percentage of CD4+ T cells, CD8+ T cells, B cells, natural killer (NK) cells, and eosinophils in the lungs. The depletion of immune cells in the lungs is not caused by apoptosis, cell infiltration to the brain, or spontaneous pneumonia following ischemic stroke as previously described, but correlates with a significant reduction in the levels of multiple chemokines in the lungs, including: CCL3, CCL4, CCL5, CCL17, CCL20, CCL22, CXCL5, CXCL9, and CXCL10. These findings suggest that ischemic stroke negatively impacts pulmonary immunity to become more susceptible for SAP development. Further investigation into the mechanisms that control pulmonary immune alternations following ischemic stroke may identify novel diagnostic or therapeutic targets for SAP.

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