Middle Cerebral Artery Occlusion Control Research Articles

Neuroprotective effects of methanolic extract from Chuanxiong Rhizoma in mice with middle cerebral artery occlusion-induced ischemic stroke: suppression of astrocyte- and microglia-related inflammatory response

BackgroundIn traditional Asian medicine, dried rhizomes of Ligusticum chuanxiong Hort. (Chuanxiong Rhizoma [CR]) have long been used to treat pain disorders that affect the head and face such as headaches. Furthermore, they have been used primarily for blood circulation improvement or as an analgesic and anti-inflammatory medicine. This study aimed to investigate the neuroprotective effects of a methanol extract of CR (CRex) on ischemic stroke in mice caused by middle cerebral artery occlusion (MCAO).MethodsC57BL/6 mice were given a 1.5-h transient MCAO (MCAO control and CRex groups); CRex was administered in the mice of the CRex group at 1,000–3,000 mg/kg either once (single dose) or twice (twice dose) before MCAO. The mechanism behind the neuroprotective effects of CRex was examined using the following techniques: brain infarction volume, edema, neurological deficit, novel object recognition test (NORT), forepaw grip strength, and immuno-fluorescence staining.ResultsPretreating the mice with CRex once at 1,000 or 3,000 mg/kg and twice at 1,000 mg/kg 1 h before MCAO, brought about a significantly decrease in the infarction volumes. Furthermore, pretreating mice with CRex once at 3,000 mg/kg 1 h before MCAO significantly suppressed the reduction of forepaw grip strength of MCAO-induced mice. In the MCAO-induced group, preadministration of CRex inhibited the reduction in the discrimination ratio brought on by MCAO in a similar manner. CRex exhibited these effects by suppressing the activation of astrocytes and microglia, which regulated the inflammatory response.ConclusionsThis study proposes a novel development for the treatment of ischemic stroke and provides evidence favoring the use of L. chuanxiong rhizomes against ischemic stroke.

159 PD-1 Agonism as a Strategy to Prevent Neuroinflammation After Acute Large Vessel Occlusion

INTRODUCTION: Stroke is a leading cause of death and long-term disability worldwide. Acute ischemia results in a profound inflammatory response that is counterproductive to short- and long-term recovery. Programmed Death Protein 1 (PD-1) is an immune checkpoint that has been shown to be upregulated after acute cerebral injury, and—upon binding its ligand PD-L1—negatively regulates immune cells to limit the inflammatory response. METHODS: C57BL/6J mice underwent middle cerebral artery occlusion (MCAO) and were randomized into three arms: (1)sham surgery, (2)MCAO control, and (3)MCAO+PD-L1. Survival, brain water content, and volumetric MRI analysis of cerebral edema were measured. Brains were analyzed by flow cytometry to quantify PD-1 expression on infiltrating immune cells. Peripheral blood was harvested for single-cell RNA-sequencing. Blood from stroke patients was collected, and the frequency of PD-1+ monocytes was correlated to clinical outcomes. RESULTS: PD-L1 significantly decreased brain edema as measured by brain water content (p = 0.0277) and high-resolution MRI (p < 0.0001), and improved overall survival (p = 0.0022). PD-1+ monocytes are upregulated in the ischemic hemisphere, and appear to be critical mediators of edema. PD-L1 treatment decreased PD-1+ monocyte infiltration , while global PD-1 (PD-1-/-) and myeloid-specific knockout of PD-1 (PD-1f/fLysMcre) abrogated the treatment effect on edema. Single-cell RNA sequencing and differential expression analysis of peripheral monocytes revealed downregulation of multiple pro-inflammatory pathways in the treated mice versus the controls. Human correlative studies confirmed that PD-1+ monocytes were present in the peripheral blood of MCA stroke patients, and the frequency of this cell population correlated with post-stroke cerebral edema. CONCLUSIONS: Our results support PD-L1 as a novel therapeutic strategy for neuroinflammation and cerebral edema after acute ischemic stroke.

Roles of isometric contraction training in promoting neuroprotection and angiogenesis after stroke in adult rats.

100 rats were randomly divided into a sham-operated group and middle cerebral artery occlusion (MCAO) modeling groups. The sham group after surgery was observed for 14 days. After MCAO, some rats received isometric contraction training (ICT) which was as follows: an atraumatic tourniquet was placed around left or right hind limb to achieve hind limb ischemia for 5 min, followed by 5 min of reperfusion, 4 cycles for one time, once a day, and five days per week. The MCAO modeling groups included the following four groups: i) a group only received MCAO, and was observed for seven days (MCAO-7d), ii) a group only received MCAO, and was observed for 14 days (MCAO-14d), iii) a group, after MCAO, received ICT for seven days (ICT-7d), and iv) a group, after MCAO, received ICT for 14 days (ICT-14d). Brain infarct area, behavioral outcomes, the number of neurons, apoptosis, cerebral edema and cerebral water content were assessed, respectively. The mRNA expression of vascular endothelial growth factor (VEGF) was assayed with RT-PCR, and protein expression of VEGF was quantified with western blot. compared with MCAO controls, cerebral infarction, neurological deficits and neuronal apoptosis were reduced significantly in the ICT groups, while the number of neurons was increased. Moreover, the mRNA expression of VEGF and protein expression of VEGF were enhanced after 1 and 2 weeks of ICT. ICT may promote angiogenesis and neuroprotection after ischemic stroke and this new remodeling method provide a novel strategy for rehabilitation of stroke patients.

Neurite Orientation Dispersion and Density Imaging of Rat Brain Microstructural Changes due to Middle Cerebral Artery Occlusion at a 3T MRI.

The purpose of this work was to demonstrate the feasibility of neurite orientation dispersion and density imaging (NODDI) in characterizing the brain tissue microstructural changes of middle cerebral artery occlusion (MCAO) in rats at 3T MRI, and to validate NODDI metrics with histology. A multi-shell diffusion MRI protocol was performed on 11 MCAO rats and 10 control rats at different post-operation time points of 0.5, 2, 6, 12, 24 and 72 h. NODDI orientation dispersion index (ODI) and intracellular volume fraction (Vic) metrics were compared between MCAO group and control group. The evolution of NODDI metrics was characterized and validated by histology. Infarction was consistent with significantly increased ODI and Vic in comparison to control tissues at all time points (P<0.001). Lesion ODI increased gradually from 0.5 to 72 h, while its Vic showed a more complicated and fluctuated evolution. ODI and Vic were significantly different between hyperacute and acute stroke periods (P<0.001). The NODDI metrics were found to be consistent with the histological findings. In conclusion, NODDI can reflect microstructural changes of brain tissues in MCAO rats at 3T MRI and the metrics are consistent with histology. This study helps to prepare NODDI for the diagnosis and management of ischemic stroke in translational research and clinical practice.

Alterations of Iron Level in the Bilateral Basal Ganglia Region in Patients With Middle Cerebral Artery Occlusion.

Background and Purpose: The purpose of this study was to explore the changes of iron level using quantitative susceptibility mapping (QSM) in the bilateral basal ganglia region in middle cerebral artery occlusion (MCAO) patients with long-term ischemia.Methods: Twenty-seven healthy controls and nine patients with MCAO were recruited, and their QSM images were obtained. The bilateral caudate nucleus (Cd), putamen (Pt), and globus pallidus (Gp) were selected as the regions of interest (ROIs). Susceptibility values of bilateral ROIs were calculated and compared between the affected side and unaffected side in patients with MCAO and between patients with MCAO and healthy controls. In addition, receiver operating characteristic (ROC) curves were performed to evaluate the diagnostic capability of susceptibility values in differentiating healthy controls and patients with MCAO by the area under the curve (AUC).Results: The susceptibility values of bilateral Cd were asymmetric in healthy controls; however, this asymmetry disappeared in patients with MCAO. In addition, compared with healthy controls, the average susceptibility values of the bilateral Pt in patients with MCAO were increased (P < 0.05), and the average susceptibility value of the bilateral Gp was decreased (P < 0.05). ROC curves showed that the susceptibility values of the Pt and Gp had a larger AUC (AUC = 0.700 and 0.889, respectively).Conclusion: As measured by QSM, the iron levels of the bilateral basal ganglia region were significantly changed in patients with MCAO. Iron dyshomeostasis in the basal ganglia region might be involved in the pathophysiological process of middle cerebral artery stenosis and occlusion. These findings may provide a novel insight to profoundly address the pathophysiological mechanisms of MCAO.

Effects of the methanol fraction of modified Seonghyangjeongki-san water extract on transient ischaemic brain injury in mice

Context Recently in Korean medicine, the antioxidant and anti-inflammatory activities of Seonghyangjeongki-san (SHJKS) were reported. However, studies on the specific mechanisms of action of SHJKS for the treatment of ischaemic stroke are still lacking. Objective This study investigates the mechanism of action of the water extract methanol fraction of modified SHJKS (SHJKSmex) on cerebral ischaemic injury. Materials and methods C57BL/6 male mice were orally administered SHJKSmex (30, 100, or 300 mg/kg) for 3 consecutive days (2 days, 1 day, and 1 h, respectively) before middle cerebral artery occlusion (MCAO). Twenty-four hours after MCAO, the infarct volumes were measured, brain edoema indices were calculated, and neurological deficit scores were determined. Inflammation-related substances in the ipsilateral hemisphere were determined by western blotting, dichlorofluorescin diacetate, thiobarbituric acid-reactive substances assay, and enzyme-linked immunosorbent assay. Results SHJKSmex pre-treatment at 300 mg/kg decreased infarct volume by 87% and mean brain water content by 90% of the MCAO control group. Moreover, SHJKSmex effectively suppressed the expression of inducible nitric oxide synthase, reactive oxygen species, interleukin 1, and caspases-8 and −9 and increased the B-cell lymphoma 2/Bcl-2-associated X protein ratio (Bcl-2/Bax) in ischaemic mouse brain. The hippocampal pyramidal cell densities were significantly increased in the 300 mg/kg SHJKSmex-administered group compared to the MCAO control group. Discussion and conclusions SHJKSmex protected the brain from ischaemic stroke in mice through its antioxidant, anti-inflammatory, and antiapoptotic activities. Our findings suggest that SHJKSmex is a promising therapeutic candidate for the development of a new formulation for ischaemia-induced brain damage.

Angelica gigas root ameliorates ischaemic stroke-induced brain injury in mice by activating the PI3K/AKT/mTOR and MAPK pathways

Context Traditionally, the root of Angelica gigas Nakai (Umbelliferae), has long been used to treat ischaemic diseases and is considered safe in humans. Objective To investigate the neuroprotective effects of a methanol extract of A. gigas root (AGmex) on the middle cerebral artery occlusion (MCAO)-induced brain injury in mice, and the underlying mechanisms. Materials and methods Two hours of transient MCAO (tMCAO) was induced in C57BL/6 mice (MCAO control group and AGmex groups), AGmex was administered to the AGmex group at 300-3,000 mg/kg bw at 1, 1, and 24 h before tMCAO or at 1000 mg/kg bw at 1 h before and after tMCAO. Infarction volumes, tissue staining, and western blotting were used to investigate the mechanism underlying the neuroprotective effects of AGmex. Results The median effective dose (ED50) could not be measured because the AGmex treatment did not reduce the infarction volume caused by 2 h of tMCAO to within 50%; however, pre-treatment with AGmex twice at 1,000 mg/kg bw before tMCAO significantly reduced the infarction volumes. The proteins related to cell growth, differentiation, and death were upregulated by this treatment, and the major recovery mechanisms appeared to involve the attenuation of the mitochondrial function of Bcl-2/Bax and activation of the PI3K/AKT/mTOR and MAPK signalling pathways in ischaemic neurons. Conclusions This study provides evidence supporting the use of A. gigas root against ischaemic stroke and suggests a novel developmental starting point for the treatment of ischaemic stroke.

An increase in AMPK/e-NOS signaling and attenuation of MMP-9 may contribute to remote ischemic perconditioning associated neuroprotection in rat model of focal ischemia

Remote ischemic perconditioning (RIPerC) results in collateral enhancement and a reduction in middle cerebral artery occlusion (MCAO) induced ischemia. RIPerC likely activates multiple metabolic protective mechanisms, including effects on matrix metalloproteinases (MMPs) and protein kinases. Here we explore if RIPerC improves neuroprotection and collateral flow by modifying the activities of MMP-9 and AMPK/e-NOS. Age matched adult male Sprague Dawley rats were subjected to MCAO followed one hour later by RIPerC (3 cycles of 15 min ischemia). Animals were euthanized 24 h post-MCAO. Haematoxylin and Eosin (H&E) staining 24 h post-MCAO revealed a significant (p < 0.02) reduction in the infarction volume in RIPerC treated animals (24.9 ± 5.4%) relative to MCAO controls (42.5 ± 4.2, %). TUNEL staining showed a 42.6% reduction in the apoptotic cells with RIPerC treatment (p < 0.01). Immunoblotting in congruence with RT-PCR and Zymography showed that RIPerC significantly reduced MMP-9 expression and activity in RIPerC + MCAO group compared to MCAO group (218.3 ± 19.1% vs. 148.9 ± 12.05% (p < 0.01). Immunoblotting revealed that RIPerC was associated with a significant 2.5-fold increase in activation of p-AMPK compared to the MCAO group (p < 0.01) which was also associated with a significant increase in the e-NOS activity (p < 0.01). RIPerC resulted in reduction of infarction volume, decreased apoptotic cell death and attenuated MMP-9 activity. This together with the increased activity of p-AMPK and increase in p-eNOS may, in part explain the neuroprotection and sustained increase in blood flow observed with RIPerC following acute stroke.

1 H NMR metabolomic profiling elucidated attenuation of neurometabolic alterations by lercanidipine in MCAo model in rats.

Stroke is a leading cause of death and disability worldwide with limited therapeutic interventions. The current study explored proton nuclear magnetic resonance spectroscopy (1 H NMR)-based metabolomic approach to elucidate the effect of lercanidipine on neurometabolic alterations in transient model of ischaemic stroke in rats. In the present investigation, male Wistar rats were subjected to middle cerebral artery occlusion (MCAo) for 2h followed by reperfusion using intraluminal filament method. Rats were randomly divided into three groups as vehicle-treated sham control, vehicle-treated MCAo control and lercanidipine-treated MCAo. Vehicle or lercanidipine (0.5mg/kg, i.p.) was administered 120min post-reperfusion. The rat brain cortex tissues were isolated 24h post-MCAo and were investigated by 1 H NMR spectroscopy through perchloric extraction method. A total of 23 metabolites were altered significantly after cerebral ischaemic-reperfusion injury in MCAo control as compared to sham control rats. Lercanidipine significantly reduced the levels of valine, alanine, lactate, acetate and tyrosine, while N-acetylaspartate, glutamate, glutamine, aspartate, creatine/phosphocreatine, choline, glycerophosphorylcholine, taurine, myo-inositol and adenosine di-phosphate were elevated as compared to MCAo control. Present study illustrates effect of lercanidipine on neurometabolic alterations which might be mediated through its antioxidant, anti-inflammatory, vasodilatory and anti-apoptotic property in MCAo model of stroke.

Abstract TP102: Human Placental Mesenchymal Stem Cells Derived Exosome-Angiotensin Converting Enzyme-2 Dependent Protection in Ischemic Stroke Injury

Following thrombolytic therapy for stroke, ischemia/reperfusion (I/R) mediated inflammation often disrupts the blood brain barrier (BBB). This can enhance expression of endothelial adhesion markers and perturb normal blood flow regulation. Proposed benefits of stem cell therapy (SCT) in stroke, besides long-term trans-differentiation into neural cells, include secretion of protective factors, which partly depends on exosomes released by stem cells. We evaluated human placenta mesenchymal stem cells (hPMSC) as potential ameliorative SCT in an acute ischemic stroke model. We hypothesize that hPMSC would achieve site-specific suppression of post-ischemic immune cell transmigration, preservation of the BBB and maintenance of blood flow via ‘paracrine’ signaling pathways in acute stroke injury.We found that intraperitoneal (IP) administration of hPMSC at the time of reperfusion, using the middle cerebral artery occlusion (MCAO)/reperfusion model, produced significant protection ( p =0.0001) of the ipsilateral hemisphere. We also demonstrated that hPMSC-treated MCAO mice exhibited significantly greater neurological recovery ( p &lt;0.0001) compared to untreated MCAO, an effect which was accompanied by significant restoration of blood flow ( p &lt;0.01) to the MCAO-stressed brains. Using Evans Blue dye assay, we also observed significant ( p =0.004) improvement of BBB integrity in ipsilateral hemispheres of hPMSC-treated mice vs MCAO controls. Furthermore, we determined that hPMSC-derived exosomes contribute to paracrine based protection of hPMSC in MCAO model. Importantly, we found that hPMSC/exosome protection is mediated partly by the function of angiotensin converting enzyme 2 (ACE2). To evaluate the contribution of ACE2 in protection of the brain after ischemic stroke, we first demonstrated that hPMSC and their exosomes express ACE2. Second, mice injected with hPMSC which had been pre-treated with the specific ACE2 inhibitor (10μM) MLN-4760, showed tissue injury and neurological behavior similar to that seen in untreated MCAO.We conclude that pleiotropic factors associated with hPMSC administration can have a favorable impact on blood flow, BBB integrity potentially alleviating the detrimental effects of ischemic stroke.

Behavioral Changes in Combination Therapy of Ethanol and Modafinil on Rats Focal Cerebral Ischemia.

Introduction:Ethanol is considered as an effective agent in reducing brain stroke injury. In this study, we assessed the effects of modafinil along with ethanol as a combination therapy on behavioral function in Wistar rats.Methods:The right Middle Cerebral Artery Occlusion (MCAO) was performed and the rats were divided into nine groups (n=8 per group). The animal groups in this study were as follows: 1. MCAO control group (ischemia without treatment); 2. Vehicle group; 3. Modafinil group that was randomly subdivided into three groups receiving different doses of modafinil (10, 30, and 100 mg/kg) for 7 days before MCAO; 4. Ethanol group receiving 1.5 g/kg ethanol at the time of reperfusion; 5. Modafinil + ethanol group that was further subdivided into three groups receiving modafinil at different doses (10, 30, and 100 mg/kg) for 7 days before MCAO and ethanol at the time of reperfusion. The motor behavior was measured using the Garcia test 24, 48, and 72 h after the ischemia, and the elevated body swing test was performed 48 and 72 h after the ischemia. The anxiety and locomotor activity were analyzed by open field test 48 and 72 h post-ischemia.Results:The results showed that the neurological deficit score, locomotor activity, and unexpected thigmotaxis (anxiety) in the ethanol, modafinil (in a dose-dependent manner), and ethanol+modafinil treatment groups were significantly higher than the MCAO control group.Conclusion:It seems that the combination therapy of modafinil (100 mg/kg) and ethanol (1.5 g/kg) significantly enhanced neuroprotection via an improvement in locomotor activity and neurological functions.

Intracerebral transplantation of erythropoietin-producing fibroblasts facilitates neurogenesis and functional recovery in an ischemic stroke model.

IntroductionErythropoietin (EPO) can enhance neurogenesis and fibroblasts can secrete growth factors; together, they may benefit ischemic stroke. We transplanted EPO‐producing fibroblasts into the rodent infarcted brain to test their effect on neurogenesis and functional recovery.MethodsA total of 106 cells of EPO‐producing NIH/3T3 fibroblasts (EPO/EGFP/3T3) or enhanced green fluorescence protein (EGFP)‐expressing fibroblasts (EGFP/3T3) were stereotaxically injected into the infarcted striatum of adult rats that received transient middle cerebral artery occlusion (MCAO) surgery 1 day poststroke. On day 14 after MCAO, the animals were euthanized for the evaluation of neurogenesis via immunohistochemistry and of the expression of growth factors using enzyme‐linked immunosorbent assay. The infarct volume was analyzed using magnetic resonance imaging and the neurological behavior was assessed using the neurological severity scoring performed within 14 days after MCAO.ResultsThe MCAO rats with EPO/EGFP/3T3 treatment showed high EPO expression in the infarcted brain for at least 1 week. The concentration of brain‐derived neurotrophic factor was higher in both hemispheres of MCAO rats with either EGFP/3T3 or EPO/EGFP/3T3 treatment at 14 days poststroke compared with untreated MCAO rats. The number of Ki‐67‐, nestin‐, or doublecortin‐immunoreactive cells in bilateral subventricular zones was higher in EPO/EGFP/3T3‐treated MCAO rats than it was in untreated MCAO control animals, indicating the enhancement of neurogenesis after EPO/EGFP/3T3 treatment. Notably, post‐MCAO EPO/EGFP/3T3 treatment significantly reduced infarct size and improved functional recovery.ConclusionThe intracerebral transplantation of EPO‐producing fibroblasts benefited an ischemic stroke model probably via the enhancement of neurogenesis.

Effects of physiological ischemic training on post-stroke neuroprotection and angiogenesis in adult rats

To examine potentially beneficial effects of physiological ischemic training (PIT) following stroke. Three hundred seventy-five adult male rats were subjected to 90-min transient middle cerebral artery occlusion (MCAO) and randomized into a PIT group and a non-PIT control group. A sham-operated group was used as negative control. After 1- and 2-weeks of PIT, brain infarct volume was measured by Triphenyl tetrazolium chloride (TTC) staining, and behavioral outcomes, presence of intact neurons, apoptosis, and cerebral edema were assessed with modified neurological severity score (mNSS), Nissl staining, TdT-mediated dUTP Nick-End Labeling (TUNEL) staining and cerebral water volume. The mRNA expression of vascular endothelial growth factor (VEGF) was assayed with RT-PCR and protein expression of VEGF was quantified with Western blot. Cerebral infarction, neurological deficits and neuronal apoptosis were reduced significantly in the PIT groups, while the presence of neurons was increased in comparison with MCAO controls. Moreover, mRNA and protein expression of VEGF were enhanced after 1 and 2 weeks of PIT. PIT may promote angiogenesis and neuroprotection during post-stroke rehabilitation and provide a novel strategy for rehabilitation of stroke patients.

Citalopram attenuated neurobehavioral, biochemical, and metabolic alterations in transient middle cerebral artery occlusion model of stroke in male Wistar rats.

Oxidative stress and inflammation are implicated as cardinal mechanisms of neuronal death following stroke. In the present study citalopram (Cit) was investigated in a 2 h middle cerebral artery occlusion (MCAo) model of stroke in male Wistar rats. Pretreatment, posttreatment (Post Cit) and pre plus posttreatment (Pre + Post Cit) with Cit were evaluated for its neuroprotective effect. In pretreatment protocol, effect of Cit at three doses (2, 4, and 8 mg/kg) administered i.p., 1 h prior to MCAo was evaluated using neurological deficit score (NDS), motor deficit paradigms, and cerebral infarction 24 h post-MCAo. In posttreatment and pre plus posttreatment protocol, the effective dose of Cit (4 mg/kg) was administered i.p., 0.5 h post-reperfusion (Post Cit) only, and 1 h prior to MCAo and again at 0.5 h post-reperfusion (Pre + Post Cit), respectively. These two groups were assessed for NDS and cerebral infarction. Though NDS was significantly reduced in both Post Cit and Pre + Post Cit groups, significant reduction in cerebral infarction was evident only in Pre + Post Cit group. Infarct volume assessed by magnetic resonance imaging was significantly attenuated in Pre + Post Cit group (10.6 ± 1.1%) compared to MCAo control group (18.5 ± 3.0%). Further, Pre + Post Cit treatment significantly altered 17 metabolites along with attenuation of malondialdehyde, reduced glutathione, matrix metalloproteinases, and apoptotic markers as compared to MCAo control. These results support the neuroprotective effect of Cit, mediated through amelioration of oxidative stress, inflammation, apoptosis, and altered metabolic profile.

Remote Limb Ischemic Conditioning during Cerebral Ischemia Reduces Infarct Size through Enhanced Collateral Circulation in Murine Focal Cerebral Ischemia

Remote ischemic conditioning (RIC) induces protection in focal cerebral ischemia. The conditioning is divided into pre-, per-, and postconditioning. However, the mechanisms of RIC remain unknown. This study aimed to determine the most effective subtype of RIC. We also examined involvement of collateral circulation on RIC. Transient middle cerebral artery occlusion (MCAO) was performed with nylon sutures in adult C57BL/6 mice under the monitoring of cerebral blood flow (CBF). Fifty mice were divided into 5 groups: MCAO control group, delayed pre-RIC group (RIC 24 hours before MCAO), early pre-RIC group (RIC 5 minutes before MCAO), per-RIC group (RIC during MCAO), and post-RIC group (RIC 5 minutes after MCAO). In other middle cerebral artery (MCA) control and per-RIC groups, collateral circulation was visualized with latex compound perfusion. After MCAO, CBF was reduced by 80% in all groups. At the end of MCAO, relative increase in CBF in per-RIC group was significantly greater than that in MCA control, whereas the infarct volume in per-RIC group was significantly smaller than that in other groups. The diameter of leptomeningeal anastomosis was larger in the per-RIC group than that in the control group. Among the 4 RIC procedures, only the per-RIC group showed clear brain protection. Enhancement of collateral circulation could play a role in the protective effect of per-RIC.

The Effect of Sneezing on the Reduction of Infarct Volume and the Improvement of Neurological Deficits in Male Rats.

Background:Sneezing transiently elevates cerebral blood flow. We speculated that induced sneezing, following embolism would restore arterial flow, thereby diminishing infarct volume and improving neurological deficits.Materials and Methods:Male rats were subjected to middle cerebral artery occlusion (MCAO) using prepared clots (embolization) and randomized into four equal groups as follows: (1) pre-MCAO-induced sneezing (PRMIS), (2) post-MCAO-induced sneezing (POMIS), and (3) pre- and POMIS (PRPOMIS) and the control group (eight rats per group). In the treatment groups, rats’ sneezing episodes were induced before MCAO in PRMIS group or before regaining consciousness from surgical anesthesia in other treatment groups by cutting their whiskers during their anesthesia and subsequently inserted them into the rats’ nostrils. Infarct volume was evaluated by 2, 3, 5-triphenyl tetrazolium chloride staining, and neurological deficits and brain edema were assessed by Bederson scale deficit scores 24-h post-MCAO.Results:The infarct volume and brain edema reduced and neurological deficits improved in the induced sneezing groups as compared with the MCAO control group. Compared to the control group, the highest improvements in the infarct volume and neurological deficits were seen in the PRPOMIS group, and POMIS group showed the most significant differences concerning the results of both ischemic and nonischemic brain edema. The highest protective effect was observed in the central region of the MCA territory.Conclusions:The reduction in ischemia-induced brain injury, brain edema, and neurological deficits by sneezing suggest that brief episodes of acute hypertension after stroke can increase blood flow to the ischemic area and improve recovery.

Neuroprotective and Anti-inflammatory Effects of a Dodecamer Peptide Harboring Ninjurin 1 Cell Adhesion Motif in the Postischemic Brain.

It has been reported that the innate immune response plays important roles in brain ischemia and that the infiltration of blood-derived immune cells is a key initiator of this response. Nerve injury-induced protein 1 (Ninjurin 1, Ninj1) is a cell adhesion molecule responsible for cell-to-cell interactions between immune cells and endothelial cells. In the present study, we investigated the proinflammatory and neuroprotective effects of Ninj1 and a dodecamer peptide harboring Ninj1 N-terminal adhesion motif (N-NAM, Pro26~Asn37) in a rat middle cerebral artery occlusion (MCAO) model of stroke. Ninj1 was predominantly induced in neutrophils and endothelial cells in the ischemic hemispheres around 12h to 1day post-MCAO, which coincided with a massive neutrophil influx. We demonstrated that intranasal administration of Ninj1 small interfering RNA (siRNA) or N-NAM significantly blocked neutrophil infiltration in postischemic brains. In addition, intranasal administration of Ninj1 siRNA or N-NAM reduced the mean infarct volume to 46.5±9.2 or 30.6 ± 11.7% of that of the PBS-treated MCAO controls, respectively, which was accompanied by significant amelioration of neurological and motor deficits. We showed that N-NAM or Ninj1 siRNA effectively blocked the adhesion and transendothelial migration of TNF-α-stimulated human myelocytic leukemia cells to human umbilical vein endothelial cells and similarly suppressed adhesion and migration of monocytes. Activations of phosphoinositide 3-kinase and Ras-related C3 botulinum toxin substrate 1 are involved in these Ninj1-mediated processes and can be inhibited by N-NAM or Ninj1 siRNA. These results indicate that Ninj1 plays an important role in neutrophil infiltration in the postischemic brain and N-NAM confers robust neuroprotective and anti-inflammatory effects by inhibiting Ninj1-mediated infiltration of neutrophils.

Anti-Zn2+-Toxicity of 4-Hydroxybenzyl Alcohol in Astrocytes and Neurons Contribute to a Robust Neuroprotective Effects in the Postischemic Brain.

4-Hydroxybenzyl alcohol (4-HBA) is an important phenolic constituent of Gastrodia elata (GE) Blume, which is used as a traditional herbal medicine in East Asia. Many activities have been reported to underlie the beneficial effects of 4-HBA in brain, such as, anti-oxidative, anti-inflammatory, anti-excitotoxic, and anti-apoptotic effects in neurons and microglia. Here, the authors demonstrate the robust neuroprotective effects of 4-HBA in rat middle cerebral artery occlusion (MCAO) model of stroke, and showed anti-Zn2+ toxicity in neurons and astrocytes as a molecular mechanism contributing to these effects. Intraperitoneal administration of 4-HBA (20mg/kg) in Sprague-Dawley rats 1h after MCAO reduced infarct volumes to 27.1±9.2% of that of MCAO controls and significantly ameliorated motor impairments and neurological deficits. Significant suppressions of Zn2+-induced cell death, ROS generation, and PARP-1 induction by 4-HBA were observed in primary cortical cultures. 4-HBA also protected astrocytes from Zn2+-induced toxicity and suppressing ROS generation by employing slightly different molecular mechanisms, i.e., suppressing PARP-1 induction and NAD depletion under acute Zn2+-treatment and suppressing p67 NADPH oxidase subunit induction under chronic Zn2+-treatment. Results indicate that the protective effects of 4-HBA against Zn2+-toxicity in neurons and astrocytes contribute to its robust neuroprotective effects in the postischemic brain. Considering the pleiotropic effects of 4-HBA, which have been reported in previous reports and added in the present study, it has therapeutic potential for the amelioration of ischemic brain damage.

Reduced bone formation markers, and altered trabecular and cortical bone mineral densities of non-paretic femurs observed in rats with ischemic stroke: A randomized controlled pilot study.

BackgroundImmobility and neural damage likely contribute to accelerated bone loss after stroke, and subsequent heightened fracture risk in humans.ObjectiveTo investigate the skeletal effect of middle cerebral artery occlusion (MCAo) stroke in rats and examine its utility as a model of human post-stroke bone loss.MethodsTwenty 15-week old spontaneously hypertensive male rats were randomized to MCAo or sham surgery controls. Primary outcome: group differences in trabecular bone volume fraction (BV/TV) measured by Micro-CT (10.5 micron istropic voxel size) at the ultra-distal femur of stroke affected left legs at day 28. Neurological impairments (stroke behavior and foot-faults) and physical activity (cage monitoring) were assessed at baseline, and days 1 and 27. Serum bone turnover markers (formation: N-terminal propeptide of type 1 procollagen, PINP; resorption: C-terminal telopeptide of type 1 collagen, CTX) were assessed at baseline, and days 7 and 27.ResultsNo effect of stroke was observed on BV/TV or physical activity, but PINP decreased by -24.5% (IQR -34.1, -10.5, p = 0.046) at day 27. In controls, cortical bone volume (5.2%, IQR 3.2, 6.9) and total volume (6.4%, IQR 1.2, 7.6) were higher in right legs compared to left legs, but these side-to-side differences were not evident in stroke animals.ConclusionMCAo may negatively affect bone formation. Further investigation of limb use and physical activity patterns after MCAo is required to determine the utility of this current model as a representation of human post-stroke bone loss.

Neuroprotective effect of triflusal and its main metabolite, 2-hydroxy-4-trifluoromethylbenzoic acid (HTB), in the postischemic brain

2-Hydroxy-4-trifluoromethylbenzoic acid (HTB) is a metabolite of triflusal (TF), and has been reported to exert anti-inflammatory effect. In this study, the authors investigated whether HTB has a neuroprotective effect against ischemic brain injuries. We showed that intravenous administration of HTB (5mg/kg) 30min before or 1, 3, or 6h after middle cerebral artery occlusion (MCAO) reduced brain infarct to 10.4±3.3%, 16.9±2.3%, 22.2±1.5% and 40.7±7.5%, respectively, of that of treatment-naive MCAO controls, and the therapeutic time window extended to 9h after MCAO (40.7±7.5%). Furthermore, HTB suppressed infarct formation, protected motor activities, and ameliorated neurological deficits more effectively than by TF or salicylic acid (SA). HTB markedly suppressed microglial activation and proinflammatory cytokines expressions in the postischemic brain and in BV2 cells and suppressed LPS-induced nitrite production by inhibiting IkB degradation. In addition, HTB suppressed NMDA-induced neuronal cell death more effectively than TF or SA in primary cortical neuron cultures. Together, these results indicate that HTB has multi-modal protective effects against ischemic brain damage that encompass anti-inflammatory, anti-excitotoxicity, and anti-Zn2+-toxicity effects.