Frequency and Patterns of Brain Infarction in Patients With Embolic Stroke of Undetermined Source: NAVIGATE ESUS Trial.

BACKGROUND AND PURPOSE: The spectrum of brain infarction in patients with embolic stroke of undetermined source (ESUS) has not been well characterized. Our objective was to define the frequency and pattern of brain infarcts detected by magnetic resonance imaging (MRI) among patients with recent ESUS participating in a clinical trial. METHODS: In the NAVIGATE ESUS trial (New Approach Rivaroxaban Inhibition of Factor Xa in a Global Trial Versus ASA to Prevent Embolism in Embolic Stroke of Undetermined Source), an MRI substudy was carried out at 87 sites in 15 countries. Participants underwent an MRI using a specified protocol near randomization. Images were interpreted centrally by those unaware of clinical characteristics. RESULTS: Among the 918 substudy cohort participants, the mean age was 67 years and 60% were men with a median (interquartile range) of 64 (26-115) days between the qualifying ischemic stroke and MRI. On MRI, 855 (93%) had recent or chronic brain infarcts that were multiple in 646 (70%) and involved multiple arterial territories in 62% (401/646). Multiple brain infarcts were present in 68% (510/755) of those without a history of stroke or transient ischemic attack before the qualifying ESUS. Prior stroke/transient ischemic attack (P<0.001), modified Rankin Scale score >0 (P<0.001), and current tobacco use (P=0.01) were associated with multiple infarcts. Topographically, large and/or cortical infarcts were present in 89% (757/855) of patients with infarcts, while in 11% (98/855) infarcts were exclusively small and subcortical. Among those with multiple large and/or cortical infarcts, 57% (251/437) had one or more involving a different vascular territory from the qualifying ESUS. CONCLUSIONS: Most patients with ESUS, including those without prior clinical stroke or transient ischemic attack, had multiple large and/or cortical brain infarcts detected by MRI, reflecting a substantial burden of clinical stroke and covert brain infarction. Infarcts most frequently involved multiple vascular territories. Registration: URL: https://www.clinicaltrials.gov; Unique identifier: NCT02313909.

Fecal Transplantation from db/db Mice Treated with Sodium Butyrate Attenuates Ischemic Stroke Injury.

The complication of type 2 diabetes (T2D) exacerbates brain infarction in acute ischemic stroke (AIS). Because butyrate-producing bacteria are decreased in T2D and butyrate has been reported to be associated with attenuated brain injury in AIS, we hypothesize that administering butyrate could ameliorate T2D-associated exacerbation of brain infarction in AIS. Therefore, we first validated that Chinese AIS patients with T2D comorbidity have significantly lower levels of fecal butyrate-producing bacteria and butyrate than AIS patients without T2D. Then, we performed a 4-week intervention in T2D mice receiving either sodium butyrate (SB) or sodium chloride (NaCl) and found that SB improved the diabetic phenotype, altered the gut microbiota, and ameliorated brain injury after stroke. Fecal samples were collected from T2D mice after SB or NaCl treatment and were transplanted into antibiotic-treated C57BL/6 mice. After 2 weeks of transplantation, the gut microbiota profile and butyrate level of recipient mice were tested, and then the recipient mice were subjected to ischemic stroke. Stroke mice that received gut microbiota from SB-treated mice had a smaller cerebral infarct volume than mice that received gut microbiota from NaCl-treated mice. This protection was also associated with improvements in gut barrier function, reduced serum levels of lipopolysaccharide (LPS), LPS binding protein (LBP), and proinflammatory cytokines, and improvements in the blood-brain barrier. IMPORTANCE Ischemic stroke is a major global health burden, and T2D is a well-known comorbidity that aggravates brain injury after ischemic stroke. However, the underlying mechanism by which T2D exacerbates stroke injury has not been completely elucidated. A large amount of evidence suggests that the gut microbiota composition affects stroke outcomes. Our results showed that the gut microbiota of T2D aggravated brain injury after ischemic stroke and could be modified by SB to afford neuroprotection against stroke injury. These findings suggest that supplementation with SB is a potential therapeutic strategy for T2D patients with ischemic stroke.

Salvianolate lyophilized injection regulates the autophagy-lysosomal pathway in cerebral ischaemia/reperfusion rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Activation of autophagy has been implicated in cerebral ischiemia/reperfusion (I/R) injury. Salvianolate lyophilized injection (SLI) has been widely used in the clinical treatment of cerebrovascular disease in China. Whether SLI has any influence on the activation of autophagy in cerebral I/R injury remains elusive. AIM OF THE STUDY: The aim of this study were to assess whether SLI attenuates I/R-induced brain injury and evaluate its associated mechanisms. MATERIALS AND METHODS: Focal cerebral ischaemia was induced by middle cerebral artery occlusion (MCAO). SLI (21 mg/kg) was injected intravenously at the beginning of the reperfusion period and 24 and 48 h after ischaemia. The effects of SLI on brain injury were detected according to infarct volume, neurological score, brain oedema, and HE and TUNEL staining at 72 h post-MCAO. Western blotting was used to detect alterations in the autophagy-relevant proteins LC3, Beclin-1, mTOR, p62, Lamp-1, and CTSD in the ipsilateral cortex at 24 or 72 h post-MCAO. RESULTS: We first demonstrated that SLI significantly alleviated the infarct volume, neurological deficits, and brain oedema, and reduced the number of TUNEL-positive cells in rats with cerebral I/R injury. Next, we found that SLI has a bidirectional regulatory effect on autophagy: early-stage (24 h) cerebral ischaemia promotes the activation of autophagy and developmental-stage (72 h) cerebral ischaemia has an inhibitory effect. SLI enhanced I/R-induced autophagy as evidenced by the increased expression level of the autophagy marker protein LC3Ⅱ, as well as the decreased expression of mTOR and the autophagy substrate protein p62, but there was no change in lysosomal activity at 24 h after I/R-induced injury. Moreover, SLI also inhibited excessive activation of autophagy at 72 h after I/R-induced injury, which manifested as downregulating LC3Ⅱ expression, upregulating mTOR and p62 expression, and inhibiting lysosomal activity. CONCLUSION: SLI has a protective effect on cerebral ischaemia/reperfusion injury, which may be mediated by the autophagy-lysosome pathway.

Asiaticoside Alleviates Cerebral Ischemia-Reperfusion Injury via NOD2/Mitogen-Activated Protein Kinase (MAPK)/Nuclear Factor kappa B (NF-κB) Signaling Pathway.

BACKGROUND Cerebral ischemia-reperfusion injury (CIRI) remains a serious health problem. Centella asiatica formulations are used to treat central nervous system disorders. In the present study, asiaticoside, an extract of the plant Centella asiatica, was investigated in CIRI in vivo and vitro. MATERIAL AND METHODS We made a CIRI model in vivo in SD rats treated by middle cerebral artery occlusion, and a cell model of ischemia-reperfusion injury was made in PC12 cells treated by deprivation of oxygen and glucose/restoration. CIRI in vivo was assessed by scores of neurological functions, encephaledema, and cerebral infarction area. Inflammation level and oxidative stress level were detected by the appropriate kits. TUNEL assay was performed for assessment of cell apoptosis and Western blot analysis was performed to assess protein expression levels. CCK8 assay was performed for evaluation of cell survival and flow cytometer was used to detect cell apoptosis in vitro. RESULTS Nervous function injury, brain edema, cell apoptosis, infarct size, apoptosis-related protein expressions, and protein expressions of the NOD2/MAPK/NF-kappaB signaling pathway in the CIRI model were all reversed by asiaticoside in rats. The cell apoptosis, inflammation level, and oxidative stress level in the model of cerebral ischemia-reperfusion injury were reduced by asiaticoside. The effects of asiaticoside on CIRI were reversed by NOD 2 agonists. CONCLUSIONS Asiaticoside showed a protective effect against cerebral ischemia-reperfusion injury via the NOD2/MAPK/NF-kappaB signaling pathway. These findings are vital for future research on use of asiaticoside in CIRI, providing a new avenue for alleviating CIRI.

Combined Therapy With Hyperbaric Oxygen and Melatonin Effectively Reduce Brain Infarct Volume and Preserve Neurological Function After Acute Ischemic Infarct in Rat.

This study tested the hypothesis that combined hyperbaric oxygen (HBO) and melatonin (Mel) was superior to either one for protecting the brain functional and parenchymal integrity from acute ischemic stroke (IS) injury. Adult-male Sprague-Dawley rats were divided into groups 1 (sham-operated control), 2 (IS), 3 (IS + HBO), 4 (IS + Mel), and 5 (IS + HBO-Mel). By day 28 after IS, the brain infarct area (BIA) was lowest in group 1, highest in group 2, significantly higher in groups 3 and 4 than in group 5, but not different between groups 3 and 4. The neurological function at day 7, 14, and 28 exhibited an opposite pattern to BIA among the 5 groups. The protein expressions of inflammatory (IL-1ß/IL-6/iNOS/TNF-α/p-NF-κB), apoptotic (cleaved-caspase3/cleaved-PARP/mitochondrial Bax), mitochondrial/DNA-damaged (cytochrome-C/γ-H2AX), oxidative stress (NOX-1/NOX-2), and autophagy (i.e. ratio of CL3B-II/CL3B-I) biomarkers displayed an identical pattern of BIA among 5 groups. Cellular expressions of inflammation (F4/80+/GFAP+) and DNA-damaged biomarker (γ-H2AX+) exhibited an identical pattern, whereas the integrities of myelin sheath/neuron (MPB+/NeuN+), endothelial cell (CD31+/vWF+), and number of small vessels exhibited an opposite pattern of BIA among the 5 groups. Combined HBO-Mel therapy offered an additional benefit in protecting the brain against IS injury.

Rationale, design, and baseline participant characteristics in the MRI and cognitive substudy of the cardiovascular outcomes for people using anticoagulation strategies trial.

BACKGROUND: Covert vascular disease of the brain manifests as infarcts, white matter hyperintensities, and microbleeds on MRI. Their cumulative effect is often a decline in cognition, motor impairment, and psychiatric disorders. Preventive therapies for covert brain ischemia have not been established but represent a huge unmet clinical need. AIMS: The MRI substudy examines the effects of the antithrombotic regimens in COMPASS on incident covert brain infarcts (the primary outcome), white matter hyperintensities, and cognitive and functional status in a sample of consenting COMPASS participants without contraindications to MRI. METHODS: COMPASS is a randomized superiority trial testing rivaroxaban 2.5 mg bid plus acetylsalicylic acid 100 mg and rivaroxaban 5 mg bid against acetylsalicylic acid 100 mg per day for the combined endpoint of MI, stroke, and cardiovascular death in individuals with stable coronary artery disease or peripheral artery disease. T1-weighted, T2-weighted, T2*-weighted, and FLAIR images were obtained close to randomization and near the termination of assigned antithrombotic therapy; biomarker and genetic samples at randomization and one month, and cognitive and functional assessment at randomization, after two years and at the end of study. RESULTS: Between March 2013 and May 2016, 1905 participants were recruited from 86 centers in 16 countries. Of these participants, 1760 underwent baseline MRI scans that were deemed technically adequate for interpretation. The mean age at entry of participants with interpretable MRI was 71 years and 23.5% were women. Coronary artery disease was present in 90.4% and 28.1% had peripheral artery disease. Brain infarcts were present in 34.8%, 29.3% had cerebral microbleeds, and 93.0% had white matter hyperintensities. The median Montreal Cognitive Assessment score was 26 (interquartile range 23-28). CONCLUSIONS: The COMPASS MRI substudy will examine the effect of the antithrombotic interventions on MRI-determined covert brain infarcts and cognition. Demonstration of a therapeutic effect of the antithrombotic regimens on brain infarcts would have implications for prevention of cognitive decline and provide insight into the pathogenesis of vascular cognitive decline.

Ipsilateral Ptosis and Contralateral Ataxic Hemiparesis as Initial Symptoms of Combined Tuberothalamic and Paramedian Artery Infarction.

Thalamic infarcts, accounting for approximately 14% of lacunar infarcts, exhibit varied clinical manifestations due to complex anatomy of nuclei and varying blood supply. Pure and combined types of thalamic infarctions have been summarized in some paper, but information of cerebral angiography was not mentioned. Here we report a rare case of combined tuberothalamic and paramedian artery occlusion presenting with ipsilateral ptosis and contralateral ataxic hemiparesis.

Celastrol treatment protects against acute ischemic stroke-induced brain injury by promoting an IL-33/ST2 axis-mediated microglia/macrophage M2 polarization.

BACKGROUND: Acute ischemic stroke (AIS) is the most common type of cerebrovascular disease and is a leading cause of disability and death worldwide. Recently, a study suggested that transformation of microglia from the pro-inflammatory M1 state to the anti-inflammatory and tissue-reparative M2 phenotype may be an effective therapeutic strategy for ischemic stroke. Celastrol, a traditional oriental medicine, may have anti-inflammatory and neuroprotective effects. However, the underlying mechanisms remain unknown. METHODS: We first determined the expression levels of inflammatory factors in patients and rodent models associated with AIS; we then determined the anti-inflammatory effects of celastrol in AIS, both in vivo and in vitro, using animal models of middle cerebral artery occlusion (MCAO) and cell models of oxygen-glucose deprivation (OGD) treatment with or without celastrol, respectively. RESULTS: The results indicated that expression of both inflammatory (interleukin (IL)-1ß, IL-6, and tumor necrosis factor (TNF)-α) cytokines, as well as the anti-inflammatory cytokine, IL-33, and IL-10, were increased following AIS in patients and in animal models. Furthermore, in vitro experiments confirmed that celastrol treatment decreased inflammatory cytokine expression induced by OGD through an IL-33/ST2 axis-mediated M2 microglia/macrophage polarization. Finally, celastrol is protected against ischemic-induced nerve injury, both in vivo and in vitro. CONCLUSIONS: Taken together, these data suggest that celastrol post-treatment reduces ischemic stroke-induced brain damage, suggesting celastrol may represent a novel potent pharmacological therapy.

Preclinical Development of a Prophylactic Neuroprotective Therapy for the Preventive Treatment of Anticipated Ischemia-Reperfusion Injury.

Ischemia-reperfusion brain injury can be iatrogenically induced secondary to life-saving procedures. Prophylactic treatment of these patients offers a promising prevention for lifelong complications. We postulate that a cytosine-guanine (CpG) oligodeoxynucleotide (ODN) can provide robust antecedent protection against cerebral ischemic injury with minimal release of pro-inflammatory cytokines, making it an ideal candidate for further clinical development. Mouse and nonhuman primate (NHP) models of cerebral ischemic injury were used to test whether an A-type CpG ODN, which induces minimal systemic inflammatory cytokine responses, can provide prophylactic protection. Extent of injury in the mouse was measured by histological staining of live tissue. In the NHP, injury was assessed 2 and 7 days post-occlusion from T2-weighted magnetic resonance images and neurological and motor deficits were cataloged daily. Plasma cytokine levels were measured using species-specific Luminex assays. Prophylactic administration of an A-type CpG ODN provided robust protection against cerebral ischemic injury in the mouse with minimal systemic inflammation. Rhesus macaques treated with D192935, a mixture of human optimized A-type CpG ODNs, had smaller infarcts and demonstrated significantly less neurological and motor deficits following ischemic injury. Our findings demonstrate the translational potential of D192935 as a prophylactic treatment for patients at risk of cerebral ischemic injury.

Complex neurological symptoms in bilateral thalamic stroke due to Percheron artery occlusion.

The artery of Percheron is a rare anatomical variant where a single thalamic perforating artery arises from the proximal posterior cerebral artery (P1 segment) between the basilar artery and the posterior communicating artery and supplies the rostral mesencephalon and both paramedian territories of the thalami. Almost one-third of human brains present this variant. Occlusion of the artery of Percheron mostly results in a bilateral medial thalamic infarction, which usually manifests with altered consciousness (including coma), vertical gaze paresis, and cognitive disturbance. The presentation is similar to the "top of the basilar syndrome", and early recognition should be prompted. We describe the case of a young female with this vessel variant who experienced a bilateral thalamic stroke. Magnetic resonance angiography demonstrated bilateral thalamic infarcts and a truncated artery of Percheron. Occlusion of the vessel was presumably due to embolism from a patent foramen ovale. Thrombolysis was performed, with incomplete symptom remission, cognitive impairment, and persistence of speech disorders. Early recognition and treatment of posterior circulation strokes is mandatory, and further investigation for underlying stroke etiologies is needed.

Refractory OCD Due to Thalamic Infarct With Response to Dronabinol.

Structural brain lesions can be a rare cause of refractory psychiatric symptoms. The analysis of such cases may lead to insights into psychiatric neurobiology. Here we present a case of a dronabinol-responsive obsessive-compulsive syndrome after thalamic infarct.

Suppressive effects of Gua Lou Gui Zhi decoction on MCAO-induced NO and PGE2 production are dependent on the MAPK and NF-κB signaling pathways.

The present study aimed to investigate the inhibitory effects, and underlying mechanisms, of Gua Lou Gui Zhi decoction (GLGZD) in a rat model of neuroinflammation. Sprague-Dawley rats were treated with GLGZD following middle cerebral artery occlusion (MCAO). Neurological function and infarct volume were evaluated to confirm successful generation of the rat model. Subsequently, brain tissues and blood samples were collected for further analysis. Nitric oxide (NO) and prostaglandin E2 (PGE2) were evaluated in peripheral blood samples using the Griess reagent assay and an ELISA, respectively. The relative expression levels of inducible nitric oxide synthase (iNOS) and cylooxygenase­2 (COX­2) were detected by quantitative polymerase chain reaction and immunohistochemistry. The associated pathways, including nuclear factor­κB (NF­κB) and mitogen­activated protein kinases (MAPK) signaling pathways, were detected by electrophoretic mobility shift assay and western blotting. The results demonstrated that treatment with GLGZD significantly inhibited MCAO-induced inflammation; GLGZD suppressed the production of NO and PGE2, and the expression of iNOS and COX­2, by inhibiting NF­κB activation and MAPK phosphorylation. These findings suggest that GLGZD, a potential agent for post­stroke treatment, may exert anti­inflammatory effects, thus providing neuroprotection.

Enhanced cell survival of pH-sensitive bioenergetic nucleotide nanoparticles in energy/oxygen-depleted cells and their intranasal delivery for reduced brain infarction.

UNLABELLED: Nucleotides (NTs) (e.g., adenosine triphosphate) are very important molecules in the body. They generate bioenergy through phosphate group release, are involved in various biological processes, and are used to treat various diseases that involve energy depletion. However, their highly anionic characteristics might limit delivery of exogenous NTs into the cell, which is required to realize their functions as bioenergy sources. In this study, ionic complexation between Ca(2+) and NT phosphates was used to form Ca(2+)/NT nanocomplexes (NCs), and branched polyethyleneimine (bPEI1.8kDa) was coated on the surface of Ca(2+)/NT NCs via a simple electrostatic coating. The resultant Ca(2+)/NT/bPEI1.8kDa NCs were approximately 10-25nm in size and had positive zeta-potentials, and their NT loading efficiency and content were approximately 60-75% and 10-20 wt%, respectively. Faster NT release from Ca(2+)/NT/bPEI1.8kDa NCs was induced by lower pH and by NTs with fewer phosphates. Reductions in cell viability in response to low temperature, serum deprivation, or hypoxia were recovered by NT delivery in Ca(2+)/NT/bPEI1.8kDa NCs. In a middle cerebral artery occlusion (MCAO)-induced post-ischemic rat model, the BBB (blood brain barrier)-detoured intranasal administration of Ca(2+)/ATP/bPEI1.8kDa NCs induced a better reduction in infarct volume and neurological deficits than did free ATP. In conclusion, intracellular NT delivery using Ca(2+)/NT/bPEI1.8kDa NCs might potentially enhance cell survival and reduce infarction in energy-/oxygen-depleted environments. STATEMENT OF SIGNIFICANCE: This study describes bioenergetic nucleotide delivery systems and their preparation, physicochemical characterization, and biological characterization both in vitro and in vivo. Nucleotides, such as adenosine triphosphate (ATP) and guanosine triphosphate (GTP), are very important signaling and energy molecules in the body. However, research on these nucleotides using nanosized carriers has been very limited. Liposomal ATP delivery has been reported in heart and renal ischemia studies. Notably, although this delivery system has potential in energy-depleted environments (e.g., low temperature, serum deprivation, and hypoxia) and in brain ischemia, studies are lacking regarding these systems. Thus, we designed polycation-shielded Ca(2+)/nucleotide nanocomplexes using simple mixing, which produced 10- to 25-nm-sized particles. The nanocomplexes released nucleotides in response to acidic pH, and they enhanced cell survival rates under conditions of low temperature, serum deprivation, or hypoxia. Importantly, the nanocomplexes reduced cerebral infarct volumes in a post-ischemic rat model. Thus, our study demonstrates that a novel nucleotide nanocomplex could have potential for preventing or treating diseases that involve energy depletion, such as cardiac, cerebral, and retinal ischemia, and liver failure.

Study on cerebroprotective actions of Clerodendron glandulosumleaves extract against long term bilateral common carotid artery occlusion in rats.

Stroke is a major cause of death and disability worldwide. The resulting burden on the society continues to grow, with increase in the incidence of stroke. Oxidative stress has been involved in the pathogenesis of several neurological diseases including acute stroke.Focal and global cerebral ischemia represents diseases that are common in the human population.In recent years much attention is being paid towards the exploration of herbal preparation, antioxidant agents and combination therapies including COX-2 inhibitors in experimental model of stroke.Possible effect of a hydroalcoholic leaf extract of Clerodendron glandulosumColeb (C. glandulosum)on oxidant-antioxidant status in ischemia-hypoperfusion injury in the rat forebrain has been investigated.Healthy adult male Wistar albino rats were divided into five groups (n=8). Group I was served as Sham control (normal saline 1ml/kg, orally), group II was served hypoperfusion control (normal saline 1ml/kg, orally), group III, group IV were served as hydroalcoholic extract treated (200 and 400mg/kg, orally) and group V was treated with Quercetin (10mg/kg, orally) for 14days to assess preventive and curative effects of C. glandulosum. Flavonoid and phenolic compounds exhibit a broad spectrum of biological activity, including antioxidant. C. glandulosum extract (200 and 400mg/kg, p.o) was administered orally, once daily for a period of 2 weeks after the occlusion of BCCA. After 14th days rats were subjected to behavioral studies. After behavioral studies animals were sacrificed and brain was removed and homogenized. Estimation of Lipid peroxidation (LPO) Myeloperoxidase (MPO), estimation of protein levels and the activities of Superoxide dismutase (SOD), Catalase (CAT), were performed. Infarct size and histopathological changes were observed in treated groups.

Traditional Chinese Patent Medicine for Acute Ischemic Stroke: An Overview of Systematic Reviews Based on the GRADE Approach.

The aim of the study is to conduct an overview of systematic reviews (SRs) to provide a contemporary review of the evidence for delivery of Traditional Chinese Patent Medicine (TCPMs) for patients with acute ischemic stroke.SRs were assessed for quality using the Assessment of Multiple Systematic Reviews (AMSTAR) tool and the Oxman-Guyatt Overview Quality Assessment Questionnaire (OQAQ). We assessed the quality of the evidence of high methodological quality (an AMSTAR score ≥9 or an OQAQ score ≥7) for reported outcomes using the GRADE (the Grading of Recommendations Assessment, Development and Evaluation) approach.(1) Dan Shen agents: tiny trends toward the improvement in different neurological outcomes (RR = 1.16, 1.10, 1.23, 1.08, 1.12); (2) Mailuoning: a tiny trend toward improvement in the neurological outcome (RR = 1.18); (3) Ginkgo biloba: tiny trends toward improvement in the neurological outcome (RR = 1.18, MD = 0.81); (4) Dengzhanhua: a tiny trend toward an improvement in neurological (RR = 1.23); (5) Acanthopanax: a small positive (RR = 1.17, 1.31) result on neurological improvement reported; (6) Chuanxiong-type preparations: neurological functional improved (MD = 2.90);(7) Puerarin: no better effect on the rate of death or disability (OR = 0.81, 95% CI 0.35-1.87); (8) Milk vetch: no better effect on the rate of death (OR = 0.66, 95% CI: 0.11-2.83);(9) Qingkailing: rate of death reduced (OR = 0.66, 95% CI: 0.11-2.83). Limitations in the methodological quality of the RCTs, inconsistency and imprecision led to downgrading of the quality of the evidence, which varied by review and by outcome. Consequently, there are currently only weak evidences to support those TCPMs.The 9 TCPMs may be effective in the treatment of acute ischemic stroke, as the GRADE approach indicated a weak recommendation for those TCPMs' usage.

Omega-3 polyunsaturated fatty acids ameliorate neuroinflammation and mitigate ischemic stroke damage through interactions with astrocytes and microglia.

Omega-3 polyunsaturated fatty acids (PUFA n3) provide neuroprotection due to their anti-inflammatory and anti-apoptotic properties as well as their regulatory function on growth factors and neuronal plasticity. These qualities enable PUFA n3 to ameliorate stroke outcome and limit neuronal damage. Young adult male rats received transient middle cerebral artery occlusion (tMCAO). PUFA n3 were intravenously administered into the jugular vein immediately after stroke and 12h later. We analyzed stroke volume and behavioral performance as well as the regulation of functionally-relevant genes in the penumbra. The extent of ischemic damage was reduced and behavioral performance improved subject to applied PUFA n3. Expression of Tau and growth-associated protein-43 genes were likewise restored. Ischemia-induced increase of cytokine mRNA levels was abated by PUFA n3. Using an in vitro approach, we demonstrate that cultured astroglial and microglia directly respond to PUFA n3 administration by preventing ischemia-induced increase of cyclooxygenase 2, hypoxia-inducible factor 1alpha, inducible nitric oxide synthase, and interleukin 1beta. Cultured cortical neurons also appeared as direct targets, since PUFA n3 shifted the Bcl-2-like protein 4 (Bax)/B-cell lymphoma 2 (Bcl 2) ratio towards an anti-apoptotic constellation. Thus, PUFA n3 reveal a high neuroprotective and anti-inflammatory potential in an acute ischemic stroke model by targeting astroglial and microglial function as well as improving neuronal survival strategies. Our findings signify the potential clinical feasibility of PUFA n3 therapeutic treatment in stroke and other acute neurological diseases.

Sexually dimorphic response of TRPM2 inhibition following cardiac arrest-induced global cerebral ischemia in mice.

Transient global cerebral ischemia due to cardiac arrest followed by resuscitation (CA/CPR) causes significant neurological damage in vulnerable neuron populations within the brain, such as hippocampal CA1 neurons. In recent years, we have implicated the transient receptor potential M2 (TRPM2) channel as a mediator of ischemic injury to neurons. We previously demonstrated that genetic and pharmacological strategies that reduce TRPM2 function preferentially protect male neurons in vitro and reduce infarct volume following experimental stroke. Due to the narrow therapeutic window for intervention following ischemic stroke, it is important to assess the role of TRPM2 in other models of cerebral ischemia. Therefore, this study utilized a modified mouse model of CA/CPR to mimic more accurately the clinical condition by maintaining body and head temperatures near the physiological range throughout. Here, we report that inhibition of TRPM2 activity with clotrimazole reduces hippocampal CA1 neuronal injury when administered 30 min after resuscitation from cardiac arrest. Consistent with our previous observations, neuroprotection was observed in male mice and no effect on injury was observed in the female. These findings provide further evidence for TRPM2 as a target for protection against cerebral ischemia in the male brain.

N-3 fatty acid rich triglyceride emulsions are neuroprotective after cerebral hypoxic-ischemic injury in neonatal mice.

We questioned if acute administration of n-3 fatty acids (FA) carried in n-3 rich triglyceride (TG) emulsions provides neuroprotection in neonatal mice subjected to hypoxic-ischemic (H/I) brain injury. We examined specificity of FA, optimal doses, and therapeutic windows for neuroprotection after H/I. H/I insult was induced in C57BL/6J 10-day-old mice by right carotid artery ligation followed by exposure to 8% O(2) for 15 minutes at 37°C. Intraperitoneal injection with n-3-rich TG emulsions, n-6 rich TG emulsions or saline for control was administered at different time points before and/or after H/I. In separate experiments, dose responses were determined with TG containing only docosahexaenoic acid (Tri-DHA) or eicosapentaenoic acid (Tri-EPA) with a range of 0.1-0.375 g n-3 TG/kg, administered immediately after H/I insult. Infarct volume and cerebral blood flow (CBF) were measured. Treatment with n-3 TG emulsions both before- and after- H/I significantly reduced total infarct volume by a mean of 43% when administered 90 min prior to H/I and by 47% when administered immediately after H/I. In post-H/I experiments Tri-DHA, but not Tri-EPA exhibited neuroprotective effects with both low and high doses (p<0.05). Moreover, delayed post-H/I treatment with Tri-DHA significantly decreased total infarct volume by a mean of 51% when administered at 0 hr, by 46% at 1 hr, and by 51% at 2 hr after H/I insult. No protective effect occurred with Tri-DHA injection at 4 hr after H/I. There were no n-3 TG related differences in CBF. A significant reduction in brain tissue death was maintained after Tri-DHA injection at 8 wk after the initial brain injury. Thus, n-3 TG, specifically containing DHA, is protective against H/I induced brain infarction when administered up to 2 hr after H/I injury. Acute administration of TG-rich DHA may prove effective for treatment of stroke in humans.

Neuroprotection after stroke by targeting NOX4 as a source of oxidative stress.

SIGNIFICANCE: Stroke, a leading cause of death and disability, poses a substantial burden for patients, relatives, and our healthcare systems. Only one drug is approved for treating stroke, and more than 30 contraindications exclude its use in 90% of all patients. Thus, new treatments are urgently needed. In this review, we discuss oxidative stress as a pathomechanism of poststroke neurodegeneration and the inhibition of its source, type 4 nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX4), as a conceptual breakthrough in stroke therapy. RECENT ADVANCES: Among potential sources of reactive oxygen species (ROS), the NOXes stand out as the only enzyme family that is solely dedicated to forming ROS. In rodents, three cerebrovascular NOXes exist: the superoxide-forming NOX1 and 2 and the hydrogen peroxide-forming NOX4. Studies using NOX1 knockout mice gave conflicting results, which overall do not point to a role for this isoform. Several reports find NOX2 to be relevant in stroke, albeit to variable and moderate degrees. In our hands, NOX4 is, by far, the major source of oxidative stress and neurodegeneration on ischemic stroke. CRITICAL ISSUES: We critically discuss the tools that have been used to validate the roles of NOX in stroke. We also highlight the relevance of different animal models and the need for advanced quality control in preclinical stroke research. FUTURE DIRECTIONS: The development of isoform-specific NOX inhibitors presents a precious tool for further clarifying the role and drugability of NOX homologues. This could pave the avenue for the first clinically effective neuroprotectant applied poststroke, and even beyond this, stroke could provide a proof of principle for antioxidative stress therapy.