Welcome to the WSC 2022 Interactive Program

Background and Aims

Stroke leads to disturbances in cerebral functions.Chronic kidney disease (CKD) is often correlated with an enhanced risk of stroke. The cerebro-renal interaction in stroke has been explored limitedly, hence, it is imperative to explore it in depth for developing therapeutic interventions.Present study aim to decipher the molecular mechanism by which Chronic Kidney Disease (CKD) may exacerbate
ischemic stroke pathology.

Methods

CKD was induced in Sprague Dawley Rats by 0.5% adenine diet for 28 days followed by a middle cerebral artery occlusion (MCAo) for ischemic stroke.LCMS/MS was performed to assess homocysteine levels in brain.Rats were evaluated for neurodeficits, motor functions and were sacrificed at 24 hours post MCAo. Brains were harvested for infarct size estimations, biochemical analysis, mitochondrial respiration and protein expression studies.

Results

Functional outcome was impaired to a greater extent in stroke+ CKD as compared to sham, stroke and CKD animals.Levels of MDA and nitrite were increased, and glutathione decreased in stroke+CKD animals as compared to sham, stroke and CKD. Mitochondrial respiration was compromised in stroke+CKD group as compared to other groups. Increased DRP1, calcineurin and caspase3 levels were observed. Increased brain homocysteine levels were detected in CKD and CKD+Stroke groups by LCMS/MS

Conclusions

Mitochondrial dysfunction following CKD contributes towards exacerbation of ischemic stroke pathology

Background and Aims

Less than 50% of rehabilitation professionals consistently use outcome measures in practice despite being emphasized in foundational training and clinical practice guidelines. Few studies in Asia and none in Singapore have evaluated the barriers to routine use of performance-based outcome measures. Our translational study aimed to use a theory-based approach to identify the barriers and facilitators to routine use of the Fugl-Meyer Assessment (FMA) for post-stroke upper limb function among occupational therapists.

Methods

We used a mixed-method sequential explanatory study design underpinned by the Normalisation Process Theory (NPT). Quantitative data was first collected using a validated survey instrument, followed by focus groups to collect qualitative data to explain and elaborate on survey findings. Occupational therapists employed in four acute and subacute hospitals in Singapore were invited to participate.

Results

The online survey (n = 34) revealed barriers mostly related to two NPT constructs (Collective Action and Coherence), while facilitators were related to the Cognitive Participation construct. Key barriers described by focus groups (of survey participants) included the lack of coaching when learning to administer the FMA, competing tasks and priorities, and limited value of FMA scores to therapists’ immediate clinical reasoning process. Key facilitators were legitimation of therapists’ role in outcome measurement and openness towards novel ways of using the FMA.

Conclusions

We identified unique challenges and enablers using a theory-based approach, and gained insights on designing new implementation strategies to embed the use of FMA into routine clinical practice.

Background and Aims

CKD patients are reported to have impaired cerebral autoregulation.This impairment is associated with higher risk of stroke in CKD patients. However, the mechanism by which stroke outcomes are worsened in CKD are still under debate. At the same time, lack of proper treatment options to limit or intervene the condition make it more urgent to understand the intricate mechanisms underlying the CKD-stroke pathology. In view of this, the aim of the present study is to explore the involvement of endoplasmic reticulum (ER) in the worsening of stroke outcome in CKD and to understand how altered ER homeostasis plays a role towards functional impairment.

Methods

CKD was induced in male SD rats with 0.5% adenine diet for 28 days followed by a middle cerebral artery occlusion (MCAo) to generate ischemic stroke. Rats were evaluated for neurodeficit scores, motor functions and sacrificed after 24 hours for performing molecular studies. In another group of animals, a long-term study was performed for 28 days post-stroke to evaluate cognition changes. Brains were harvested for estimation of infarct size.Cortical brain regions were obtained for further biochemical and molecular studies.

Results

Increased infarct and impaired motor function were observed in CKD+Stroke as compared to CKD and Stroke.Increased MDA,nitrite while decreased glutathione,SOD were observed in CKD+Stroke as compared CKD and Stroke.Cognitive impairment was higher in CKD+Stroke as compared to CKD and Stroke. Increased peIF2α, Caspase12, CHOP and decreased in myelin basic protein were observed.

Conclusions

ER stress following stroke in CKD may contribute towards exacerbating ischemic stroke pathology.

Background and Aims

Following acute ischemic stroke (AIS), there is an increase in endoplasmic reticulum (ER) stress which is responsible for activation of unfolded protein response (UPR), in turn causing protein kinase RNA-like endoplasmic reticulum kinase (PERK) pathway activation. Activated PERK pathway inhibits translation along with ATF4 activation, responsible for cell homeostasis disruption and apoptosis. We investigated the neuroprotective effect of PERK inhibitor, GSK2606414 in adult wistar rats following middle cerebral artery occlusion (MCAO) induced stroke.

Methods

In rats, stroke was induced by middle cerebral artery occlusion using nylon filament (4-0) for a duration of 90 minutes. Stroke model was validated by assessment of neurological deficits (ND) score, brain infarct volume and a battery of neurobehavioral parameters including grip strength, rotarod, and morris water maze test. Three different doses of GSK2606414 (50, 100 and 150 mg/kg) were selected and administered at 3 hrs. post MCAO/reperfusion via oral gavage in four doses at 12 hrs. interval. ND scoring, infarct volume, neurobehavioral parameters, RT-PCR, pro-inflammatory cytokines (ELISA), oxidative stress and histopathological changes were measured in all groups.

Results

GSK2606414 had significantly reduced ND score and infarct volume, improved neurobehavioral deficits, decreased mRNA expression (GFAP, GAP43, Bax, Bad, Bcl2, PERK, ATF4 and CHOP), decreased inflammatory cytokines (IL-1β, IL-6 and TNF-α), decreased oxidative stress and reduced pyknotic nuclei in brain of rat’s post MCAO.

Conclusions

GSK2606414 may be taken as a potential neuroprotective agent in the treatment of AIS.

Background and Aims

We studied the colocalization of p53 with different isoforms of histone deacetylases and acetyltransferases in cells of the perifocal region 24 hours after photothrombotic stroke in rats.

Methods

Photothrombotic stroke (PTS) was induced in the rat cerebral cortex after intravenous injection of photosensitizer Rose Bengal, followed by laser irradiation of the sensorimotor cortex. The colocalization of p53 with histone deacetylases 1, 2 classes and sirtuins 1 and 2, as well as HAT1 and PCAF acetyltransferases in the cells of the perifocal region of photothrombotic stroke was assessed by using immunofluorescence microscopy and Duolink PLA (Proximity Ligation Assay).

Results

Immunofluorescence microscopy showed that the intracellular localization of p53 coincided with class 1 histone deacetylases - HDAC1 and HDAC2; class 2 histone deacetylases - HDAC5 and HDAC6; as well as class 3 histone deacetylase - Sirt1. Duolink PLA followed by fluorescence microscopy showed the interaction between p53 and HDAC2.

Conclusions

p53 directly interacts with HDAC2, which can affect the activity of the protein and its intracellular localization after stroke.

The study was supported by the Russian Science Foundation (grant No. 21-15-00188).

Background and Aims

We studied the expression and intracellular localization of α-, β-, and γ-secretases involved in amyloid precursor protein (APP) proteolysis, as well as the effect of their inhibitors on the volume of photothrombotic infarction and apoptosis of cells in the peri-infarct zone in mice.

Methods

Expression and localization of α-, β-, and γ-secretases were assessed using immunofluorescence microscopy and Western blotting. The volume of photothrombotic damage was assessed after staining the sections with 2,3,5-triphenyltetrazolium chloride. Apoptosis was assessed using the TUNEL assay.

Results

On the first day after photothrombotic stroke, α-secretase ADAM10 decreased in neurons. Intracellular localization of β-secretase BACE1 was limited to the cytoplasm and cytoplasmic membrane of nerve cells, while the protein expression in nerve cells did not change. The expression of the main proteins of γ-secretase complex: Presenilin-1 and Nicastrin increased in astrocytes, but not in penumbra neurons on the first day after stroke and in the early recovery period. Administration of γ-secretase inhibitor DAPT (N-[N- (3,5-difluorophenacetyl)-1-alanyl]-S-phenylglycine t-butyl ester) after photothrombotic stroke reduced the apoptosis of cells in the perifocal region. α-secretase inhibitor Batimastat (BB-94) and β-secretase inhibitor LY2886721 had no effect on the apoptosis of peri-infarct cells.

Conclusions

Probably, inhibition of γ-secretase reduces inflammation and activation of astrocytes, which contributes to a decrease in the level of apoptosis and, consequently, the amount of damage after ischemia. Thus, DAPT may be considered as a potential drug for stroke therapy.

This work was supported by the Ministry of Science and Higher Education of the Russian Federation (grant No. 0852-2020-0028).

Background and Aims

In this work, we studied the intracellular localization and expression pattern of the transcription factors c-Myc, E2F1, p53, involved in the regulation of apoptosis, in neurons, astrocytes, macrophages, and microglia in the first 24 hours and after 7 days in the perifocal region of photothrombotic stroke in rats.

Methods

Photothrombotic stroke (PTS) in the rat cerebral cortex was induced by laser irradiation of the sensorimotor cortex after intravenous injection of Rose Bengal photosensitizer. The main research methods were immunofluorescence microscopy and Western blot analysis.

Results

The most significant changes in p53 and E2F1 expression occurred 4 hours after PTS in both the nuclear and cytoplasmic fractions. A high level of p53 in the cytoplasmic fraction of brain cells persisted up to 7 days after PTS. The c-Myc level did not change. Ischemia affected the level of p53 in neurons and microglial cells, but not in astrocytes or macrophages. Changes in E2F1 expression after PTS were observed in neurons and astrocytes.

Conclusions

Probably, E2F1 stimulates apoptosis of ischemic penumbra neurons by increasing p53 expression.

The study was supported by the Russian Science Foundation (grant No. 21-15-00188).

Background and Aims

In this work, we studied the localization of the C- and N-terminal fragments of the amyloid precursor protein (APP) in cell compartments on the first day after photothrombotic stroke.

Methods

Photothrombotic stroke (PTS) in the cerebral cortex of rats and mice was induced by laser irradiation of the sensorimotor cortex after intravenous administration of the Rose Bengal photosensitizer. The main research method was electron immunohistochemistry.

Results

PTS caused an increase in the level of the N- and C-terminal fragments of APP after 24 hours, but not earlier. Both fragments were localized in the cytoplasm of ischemic penumbra cells. The localization of APP fragments was associated with microtubules and neurofilaments, the rough endoplasmic reticulum (RER), and ribosomes. A significant number of APPs were visualized both in thin unmyelinated compartments of neurons and in large compartments including mitochondria, as well as in the zones of chemical synapses, mainly in the postsynaptic part, and were absent in the nuclei. 24 hours after photothrombosis, some elements of the damaged tissue accumulated a significant amount of APP, but there were cells that did not contain APP fragments.

Conclusions

Thus, APP fragments were present in the cytoplasm, but not in the nuclei of the cells of the photothrombotic penumbra. The level of the N- and C-terminal fragments of APP increased 24 hours after a stroke. The reason why some penumbra cells did not contain APP needs to be studied.

This work was supported by the Ministry of Science and Higher Education of the Russian Federation (grant No. 0852-2020-0028).

Background and Aims

Ferroptosis is a newly identified form of programmed cell death. We aim to determine the expression patterns and role of 15-lipoxygenase-1 (ALOX15) in subarachnoid hemorrhage (SAH), and to investigate whether cepharanthine (CEP) can inhibit ferroptosis by inhibiting ALOX15 in specific cell types.

Methods

A mouse SAH model was developed by endovascular perforation method. bEend.3 endothelial cells and BV2 microglial cells as well as RSL3 and hemin were used to simulated SAH in vitro. Mice and cell lines were treated with CEP and a group of specific oxygenase inhibitors to explore the protection effect from ferroptosis. Lipid peroxidation staining with BODIPY 581/591 C11 and transmission electron microscopy were used to identify ferroptosis.

Results

We found the accumulation of lipid peroxide, as well as highly expressed ALOX15 in microglia and endothelium, and ferroptotic changes in microglial mitochondria confirmed the occurrence of ferroptosis after SAH. Further, CEP was shown to inhibit ferroptosis by downregulating the expression of ALOX15. In addition, we found that M2 type microglia are more sensitive to RSL3-induced ferroptosis than M1 type microglia, and that hemin probably induced ferroptosis in M2 type microglia by increasing ALOX15 levels and decreasing GPx4 levels. The effect of CEP treatment was also demonstrated in vitro.

Conclusions

In summary, to the best of our knowledge, this is the first study demonstrating that ferroptosis occurred in the microglia and endothelium after SAH, and this process was facilitated by increased ALOX15 levels. More importantly, treatment with CEP could inhibit ferroptosis through downregulating the expression of ALOX15.

Background and Aims

To observe the effect of edaravone dexborneol on post stroke anxiety and depression in rats, and to explore its possible mechanism.

Methods

Male SD rats were subjected to middle cerebral artery occlusion (MCAO) and treated with ED (10 mg/kg/d, edaravone: dexborneol=4:1), edaravone (8 mg/kg/d) and vehicle for 14 days. Open field test was performed 14 days after surgery. After euthanization, brain tissue samples were collected for western blot and RT-qPCR to detect the expression of proinflammatory mediators, and immunofluorescence to detect the expression of microglia M1 markers and neuron markers.

Results

The administration of edaravone dexborneol resulted effectively in enhancing the number of rats entering into the open arm, the time spent in the open arm and the time spent in the central area in open field behavior test (p＜0.05). Immunofluorescence staining results demonstrated that the ratio of polarized M1 microglia is reduced and the neuron number is protected in the peri-infarct cortex and striatum area in ED group (p＜0.05). Moreover, the protein levels of nuclear factor kappa-B (NF-κB), tumor necrosis factor α (TNF-α), interleukin 1β (IL-1β) were significantly decreased in ED group (p＜0.05). Besides, mRNA levels of TNF-α, IL-1β, inducible nitric oxide synthase (iNOS), cluster of differentiation 86 (CD86) were reduced in ED group, while CD206 were increased in ED group.

Conclusions

Edaravone dexborneol can alleviate anxiety and depression behavior in rats with cerebral ischemia-reperfusion, which may be associated with the inhibition of M1 microglial polarization, the down-regulation of NF-κB signaling pathway and the enhancement of neuronal structural stability.

Background and Aims

Stem cell-based therapy of stroke is a promising approach, offering hope for preservation of damage brain tissue. There are various ways of cells administration, among which intra-arterial delivery has shown to be one of the most effective and challenging. Technical inaccuracies with intra-arterial transplantation can lead to the development of serious complications, such as thromboembolism. In this study we examine several methodological aspects of intra-arterial catheter placement in rats with experimantal stroke model, including catheter position, maintenance of blood flow around it and possible development of vasospasm after surgery.

Methods

Wistar rats 250-300g were subjected to transient 90min MCAO and after 24h microcatheter was inserted via the ECA into the lumen of the ICA in the direction of blood flow (n=4) or into the CCA (n=44) in the opposite direction of blood flow. After catheter placement experimental animals were transferred into MR-scaner for visualization of the catheter and cerebral arteries.

Results

In the group with the location of the catherer in the ICA in most animals there was no blood flow in the CCA and proximal part of the ICA. Vasospams was observed in both experimental groups. For the group with the location of the catherer in the CCA the following ratio of vasospasm was determined: 41% weak, 41% local and 18% strong.

Conclusions

Placement of the intra-arterial catheter into the CCA reduces the chance of brachiocephalic arteries occlusion and ensures cells administration with maintenance of blood flow. During transplantation, it is worth considering the possibility of angiospasm as a result of endovascular interventions.

Background and Aims

Clopidogrel, one of the most prescribed antiplatelet medications in the world, is given to stroke survivors for the prevention of secondary cardiovascular events. Clopidogrel exerts its antiplatelet activity via antagonism of the P2Y12 receptor (P2Y12R). Although not widely known, or considered during clinical trials, P2Y12R is also expressed on brain microglia, where the receptor facilitates the microglial chemotaxis toward cellular damage and repair processes. If microglial P2Y12R is blocked, microglia lose the ability to migrate to damaged sites.

Determine whether the post-stroke administration of clopidogrel is associated with; (i) extravascular leakage, (ii) altered microglial morphology, and (iii) impaired cognitive recovery.

Methods

Male C57BL/6 mice (n=7-8) underwent photothrombotic occlusion. From 24 h post-stroke, mice were treated daily for 14 days with either clopidogrel or vehicle. Cognitive performance (memory and learning) was assessed using a mouse touchscreen platform (paired associated learning task). At day 15, animal brains were collected for immunohistochemistry analysis.

Results

Post-stroke clopidogrel treatment significantly increased vascular leakage and significantly altered microglia morphology across multiple regions of the brain. Additionally, clopidogrel treatment significantly impaired learning and memory recovery post-stroke.

Conclusions

These data suggest that clopidogrel enters the brain due to stroke-induced vascular leakage, where it hampers the ability of microglia to facilitate brain repair and enable cognitive recovery.

Background and Aims

To investigate if arterial thrombosis per se was associated with alterations in complete blood count (CBC) and white blood cell (WBC) differential count in a mouse model of thrombosis.

Methods

Twelve-week-old male C57Bl/6 mice were used for FeCl₃-mediated carotid thrombosis (n=79) or sham-operation (n=72), followed by a CBC test with differential at one of the following time-points: 30-min, 1-day, and 4-days. Non-operated mice (n=26) also underwent a CBC test with WBC differential.

Results

Monocyte count (/µL) at 30-min after thrombosis (mean±SEM, 196±13) was ~1.4-fold higher compared with 30-min after sham-operation (139±14), and was about 2.6-fold higher compared with non-operated mice (75±6). At day-1 and -4 post-thrombosis, compared with 30-min, monocyte count decreased by ~21% (155±2) and 39% (119±9), which however were about 2.2-fold and 2.1-fold higher than in sham-operated mice at the corresponding time-points (71±5 and 58±5), respectively. Lymphocyte counts (/µL) at 1- and 4-days after thrombosis (3513±253 and 2590±192) were ~60% and ~116% lower compared with the time-points in sham mice (5630±350 and 5596±314), and were ~65% and ~124% lower compared with non-operated mice (5791±264). Monocyte-lymphocyte-ratio (MLR) was substantially higher at all three time-points (0.050±0.003, 0.046±0.007, and 0.050±0.005) than in sham mice (0.003±0.003, 0.013±0.001, and 0.010±0.001). MLR was 0.013±0.001 in non-operated mice. Neutrophil-lymphocyte-ratio (NLR) and platelet-lymphocyte-ratio (PLR) were higher at 4-days after thrombosis (0.281±0.041 and 374.4±19.4), compared with sham mice (0.077±0.005 and 197.5±14.4) and non-operated mice (0.071±0.004 and 181.4±11.03), respectively.

Conclusions

This is the first report on acute arterial thrombosis-related alterations in CBC and WBC differential count.

Background and Aims

Ischemic stroke is one of the main cause of long-term disability. Transcranial magnetic stimulation (TMS) is a rehabilitation therapy that modulates the motoric cortex with the principle of electromagnetic induction to stimulate cortical motoric neurons, so as to generate motoric potential in ischemic stroke patients. The aim of the study was to evaluate the improvement of motoric strength extremities recovery in ischemic stroke, measured using fugl meyer assessment (FMA) by repetitive transcranial magnetic stimulation (rTMS).

Methods

Cohort study method January - december 2020. The study involved 42 subjects of ischemic stroke patients who were divided into treatment and control group. The treatment group performed repetitive transcranial magnetic stimulation for 5 days in day 9-13. FMA was assessed on the day 8 before treatment, and day 14 and day 45 after treatment.

Results

There were siginificances differences in FMA score after rTMS administration in ischemic stroke patients (p< 0.001) and significances differences in improvement of FMA score in day 14-8 and day 45-8 (p< 0.001).

Conclusions

Repetitive transcranial magnetic stimulation can improve motoric function in ischemic stroke.

Background and Aims

Glial Fibrillary Acidic Protein (GFAP) is a protein that responsible for the cytoskeleton structure of glial cells. Neuron Specific Enolase (NSE) is the neuronal form of intracytoplasmic glycolitic enzyme enolase. Increased level of both biomarkers have potensial value to predict neuronal damage that accompanies hemorrhagic stroke. The aim of this study was to evaluate the role of GFAP and NSE with clinical outcome of hemorrhagic stroke patients.

Methods

Prospective cohort study during July-November 2020. Serum blood sampling for GFAP and NSE examination was carried out < 72 hours of stroke onset. Then the third day NIHSS, seventh day NIHSS and 30th day mRS were assessed. Data were analyzed and statistical analyzes were performed to evaluate GFAP and NSE serum levels with neurologic clinical outcome of hemorrhagic stroke patients.

Results

Serum NSE levels did not significantly affect NIHSS changes and 30 day mRS (p > 0.05). Serum GFAP levels were significant for NIHSS changes (OR= 1.350; 95% CI: 1.112-1.640). Significant differences were also obtained at serum GFAP levels with 30 days mRS (OR= 1.603; 95% CI: 1.127-2.231). ROC curve analysis obtained serum GFAP levels with a cut-point of 11.35 ng/mL able to identify patients with poor clinical outcome (30 days mRS 3-6), and has a sensitivity and specificity of 78.9% and 78.6%.

Conclusions

Serum GFAP levels has potential value as predictor of hemorrhagic stroke clinical outcome. The role of serum NSE with clinical outcome of hemorrhagic stroke still requires further research.

Background and Aims

Hemorrhagic transformation is a complication of acute ischemic stroke and especially common after thrombolytic therapy. There is evidence that the blood brain barrier plays a critical role in the prevention of HT, reduces the volume of hemorrhage, alleviates the damage of neurological function. We developed a mouse model for embolic middle cerebral artery occlusion with homologous blood clots. To improve the method of establishing the model, the modified PE50/PE10 tubes and vascular tone were used.

Methods

Twenty Sprague-Dawley rats (SPF grade) were used to establish the rat eMCAO model, the modified PE tubes (outsidediameter: 0.34cm) were fixed and inserted by using the vascular tone with single person.

Results

The modified PE tube which carries the embolis was inserted, by dissolving into the three directions of the carotid vessels, the embolis were injected into the brain. The key technics include the direction of injection of the embolis, the branches of the carotid vessels, and the vascular tone when the PE tubes fixed to the specific site of the brain vessel branches. The brain slices by 2,3,5-Triphenyltetrazolium chloride (TTC) staining showed that the patterns of the brain infarction involving cortical, sub-cortical, and basal ganglia regions. The average time of making the eMCAO model was 30 min, 90% of the rats was successfully made of eMCAO model.

Conclusions

The vascular tone can be used in the model making of rat eMCAO with reducing the time and labor cost, in the purpose of evaluation of thrombolysis/hemorrhagic transformation (HT) therapy. While indicates the mechanism of embolis leading to different patterns of brain infarctions.

Background and Aims

Neurointervention with contrast media (CM) has rapidly increased, but the impact of CM extravasation and the related side effects remain controversial. This study investigated the effect of CM on blood–brain barrier (BBB) integrity.

Methods

We established in vitro BBB models using primary cultures of rat BBB-related cells. To assess the effects of CM on BBB functions, we evaluated transendothelial electrical resistance, permeability, and tight junction (TJ) protein expression using immunohistochemistry (IHC) and Western blotting. To investigate the mechanism of iopamidolinduced barrier dysfunction, the role of mitogen-activated protein (MAP) kinases in brain endothelial cells was examined.

Results

Short-term iopamidol exposure on the luminal side induced transient, while on the abluminal side caused persistent BBB dysfunction. IHC and immunoblotting revealed CM decreased the expression of TJ proteins. Iopamidol-induced barrier dysfunction was improved via the regulation of MAP kinase pathways. CM may cause BBB dysfunction.

Conclusions

MAP kinase pathways in brain endothelial cells mediates iopamidol-induced barrier dysfunction. CM extravasation may have negative effects on clinical outcomes in patients.

Background and Aims

Post-ischemic inflammation plays an essential role in the progression of ischemic stroke pathologies. However, the detailed molecular mechanisms underlying the activation of infiltrating immune cells which trigger sterile post-ischemic inflammation have not been sufficiently clarified. We tried to identify the previously unknown damage-associated molecular patterns (DAMPs), inflammatogenic self-molecules derived from damaged tissue.

Methods

Among the candidate proteins which were detected from brain lysate by mass spectrometry, recombinant proteins were generated and added to the culture of bone marrow-derived macrophages (BMMs) to examine the expression of inflammatory cytokines. We finally examined the extracellular release of candidate DAMPs by using a mouse model of transient middle cerebral artery occlusion. Neutralizing antibodies or KO mice were used for compromising DAMP activity in ischemic stroke.

Results

We identified DJ-1 as a novel DAMP in brain lysate. Recombinant DJ-1 protein activated BMMs only through TLR2 and TLR4. The expression of inflammatory cytokines was induced in a DJ-1 dose-dependent manner in vitro. DJ-1 had a unique peptide sequence to trigger the production of inflammatory cytokines. In the ischemic brain, twenty-four hours after stroke onset, DJ-1 was passively released into extracellular space from necrotic brain cells and directly contacted with infiltrating myeloid cells. DJ-1 deficiency reduced the expression of inflammatory cytokines after the stroke. Administration of DJ-1-neutralizing antibody suppressed the expression of inflammatory cytokines and improved neurological deficits.

Conclusions

Extracellularly released DJ-1 functioned as the previously unknown DAMP that directly activated infiltrating myeloid cells and induced sterile inflammation. Thus, extracellular DJ-1 would be a prominent therapeutic target for ischemic stroke.

Background and Aims

Obesity and nicotine are risk factors for stroke. However, obese and smoking patients suffering from stroke may have higher survival rates and better outcomes compared to normal patients. Central (hypothalamic) AMP-activated protein kinase (AMPK) plays a major role in the regulation of energy balance, mediating among others, the effects of nicotine on body weight. The aim of this study has been to investigate whether AMPK could be involved in the neuroprotective effect of obesity and nicotine on cerebral ischemia.

Methods

We used Sprague-Dawley rats that were subjected to transient occlusion of the middle cerebral artery (tMCAO). Animals were fed with standard (STD) or high-fat diet (HFD) and treated with vehicle or nicotine. Outcome variables were lesion and edema volumes, as well as neurological deficits. Furthermore, AMPK, endoplasmic reticulum (ER) stress, and autophagy/apoptosis pathways were analyzed.

Results

Obese rats showed a reduction in lesion volume and milder neurological deficit compared to normal-weight rats. Moreover, obese rats showed activation of AMPK signaling after cerebral ischemia. Nicotine treatment reduced brain ischemic injury and improved neurological recovery through the inhibition of AMPK in both obese and normal-weight groups. The AMPK activator AICAR reversed the neuroprotective effect of nicotine, indicating dependency on AMPK.

Conclusions

Obese rats showed a reduction in lesion volume and milder neurological deficit compared to normal-weight rats, related to a reduction of AMPK signaling after stroke and decreased ER stress. Furthermore, nicotine treatment reduced ischemic injury and improved neurological recovery after stroke by inhibiting AMPK signaling. Therefore, brain AMPK mediates the neuroprotective effect of nicotine on cerebral ischemia.

Background and Aims

Stroke affects millions across the globe due to its long-term effects. The present study aims to understand the role of MSCs in regulating the brain SIRT1 levels following stroke and the involvement of SIRT1 in regulating mitochondrial transfer and protection.

Methods

Ovariectomized Sprague Dawley rats were infused intraarterially with 1*10⁵ MSCs at 6 hrs post MCAo. Following 24 hrs of MCAo, animals were examined for functional and behavioral outcomes. Brains were harvested for molecular studies.

Results

Significant improvement in motor functional and behavioral outcomes and reduction in infarct size was observed following infusion of MSCs intraarterially at 6 hrs post stroke. Increase in average neuronal length and density were also observed. Increased expression of SIRT1, mitochondrial fusion, biogenesis and tunneling nanotube markers were and concomitant reduction in the expression of mitochondrial fission in the brain cortical regions were observed following MSCs treatment. Improvement in mitochondrial bioenergetics and the activity of various mitochondrial complexes were increased post MSCs infusion.

Conclusions

Results from our study provides primary evidence that IA MSCs therapy post-stroke regulates SIRT1 to confer mitochondrial protection. The intraarterial (IA) approach for administering MSCs is highly relevant in the present clinical scenario.

Background and Aims

Interdisciplinary cooperation and translation approach are frequently needed to develop new treatments in stroke, but it is difficult to establish translational collaborations because different expertise is needed. Here, we present the development of the platform for translational research in stroke in the Czech Republic by creating a STROKEBRNO Cluster. The aim of the STROKEBRNO Cluster is to accelerate translational research in stroke through multidisciplinary and trans-institutional cooperation.

Methods

In 2015 institutions with the expertise needed for preclinical research, such as veterinary, biophysics, protein engineering, molecular biology, histology, biomedical engineering, accepted an invitation to join STROKEBRNO Cluster by International Clinical Research Center Stroke Program within St. Anne’s University Hospital Brno. Teams from involved institutions have been submitting joint grant applications with stroke scientists. In 2020 the STROKEBRNO Cluster was established by a Memorandum of understanding with involved institutions, by developing a brand and a communication platform.

Results

The STROKEBRNO Cluster (www.strokebrno.com) involves 6 research institutions and 1 global industry partner from Brno. 25 researchers directly participate in 7 research projects and 2 comprehensive programs – in vivo and in vitro programs to develop new thrombolytics. There were 6 joint inter/national grants obtained in an amount of >4 mil EUR, 11 students involved, and 15 joint papers already published. Basic researchers and stroke professionals share their know-how and progress on monthly basis.

Conclusions

We have built a functional platform, probably unique in Eastern Europe, which interconnects wide expertise, capacities and technologies. Our goal is to develop better treatment for stroke.