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Ischemic stroke is a common cause of morbidity and mortality worldwide. The current treatment fails to achieve satisfactory results, because interventional therapy as first-line treatment management has a strict time window. In recent years, a large number of studies have confirmed that adenosine, as an inhibitory neurotransmitter, has a protective effect on cerebral ischemic injury. Nevertheless, direct administration of adenosine has many side effects. Previous studies showed that adenosine exerted neuroprotective effects mainly through adenosine receptor A1 (A1 receptor). Therefore, further study on the mechanism of A 1 receptor induced neuroprotection may find new targets for stroke treament. Mitochondrial biogenesis (MB) is a therapeutic target for ischemic stroke, and the nuclear-encoded peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC-1α) is a major regulator of MB. However, the influence of A1 receptor on MB and PGC-1α is unclear. In this study, using the middle cerebral artery occlusion (MCAO) model of mice, we evaluated the temporal and spatial effects of A1 receptor after ischemic stroke and verified the neuroprotection of A1 receptor. Neurological scores were used to assess functional changes in mice. At the same time, we observed the effect of activating A1 receptor on MB and PGC-1α, and the effect of knockdown PGC-1α on A1 receptor induced MB in vitro. WB and immunofluorescence were used to detect relevant indicators of MB. In addition, we downregulated PGC-1α in vivo to observe the effects on A1 receptor induced MB and neuroprotection. The findings indicated that A1 receptor was increased and mainly expressed on neurons in the penumbra, further activated A1 receptor after stroke had neuroprotection. In vitro, activation of A1 promotes MB and increases the expression level of PGC-1α, while downregulation of PGC-1α partially reverses the effect of A1 receptor after OGD/R. Down regulation of PGC-1α in the penumbra neurons can reverse the effects of activation of A1 receptor on MB and neuroprotection. Taken together, these findings indicated that A1receptor promotes MB and improves neurological function after ischemic stroke via PGC-1α. [ABSTRACT FROM AUTHOR]

Postmenopausal women have a higher incidence of stroke compared to the age-matched males, and the estrogen was thought to be the main cause of such difference. However, estrogen replacement therapy for the prevention of postmenopausal stroke shows controversial results and is widely disputed because of its serious side effects after chronic administration. Genistein (Gen), a natural phytestrogen with fewer side effects, has a protective effect against cerebral ischemia damage. However, whether Gen could effectively prevent postmenopausal stroke has not been elucidated. In the current study, reproductively senescent mice were treated with Gen (10 mg/kg) for 2 weeks before having transient cerebral ischemia insults. Neurological scores, infarct volumes, and cell apoptosis were evaluated 24 h after reperfusion. The levels of inflammatory factors and nod-like receptor protein 3 (NLRP3) inflammasome-related proteins were also examined. The results showed that Gen treatment reduced infarct volumes, improved neurological scores, attenuated apoptosis, and decreased inflammatory factor release. The expression of NLRP3 inflammasome-related proteins in microglia was downregulated by Gen. However, the overexpression of NLRP3 in microglia abrogated the Gen-induced inhibition of inflammatory factor release and reversed the neuroprotective effect of Gen. Taken together, the results suggest that Gen treatment could attenuate the acute injury induced by cerebral ischemia in reproductively senescent mice via the inhibition of the NLRP3 inflammasome in microglia, indicating that Gen could be a candidate drug for the treatment of stroke in postmenopausal women. [ABSTRACT FROM AUTHOR]

Therapeutics targeting the Nogo-A signal pathway hold promise to promote recovery following brain injury. Based on the temporal characteristics of Nogo-A expression in the process of cerebral ischemia and reperfusion, we tested a novel asynchronous treatment, in which TAT-M9 was used in the early stage to decrease neuronal loss, and TAT-NEP1-40 was used in the delayed stage to promote neurite outgrowth after bilateral common carotid artery occlusion (BCCAO) in mice. Both TAT-M9 and TAT-NEP1-40 were efficiently delivered into the brains of mice by intraperitoneal injection. TAT-M9 treatment promoted neuron survival and inhibited neuronal apoptosis. Asynchronous therapy with TAT-M9 and TAT-NEP1-40 increased the expression of Tau, GAP43 and MAP-2 proteins, and enhanced short-term and long-term cognitive functions. In conclusion, the asynchronous treatment had a long-term neuroprotective effect, which reduced neurologic injury and apoptosis, promoted neurite outgrowth and enhanced functional recovery after ischemia. It suggests that this asynchronous treatment could be a promising therapy for cerebral ischemia in humans. [ABSTRACT FROM AUTHOR]

The mechanism of electroacupuncture (EA) pretreatment-induced neuroprotection remains unclear. In this study, we found that neuronal Triggering receptor expressed on myeloid cells 2 (TREM2) expression was increased and peaked at 48 h and 72 h after ischemia/reperfusion. After specific knockdown of TREM2 in excitatory neurons, neurological function was damaged, and the infarct volume was enlarged. Furthermore, the expression of LC3II/LC3I and Beclin1 was decreased, while the expression of p62 was increased. EA pretreatment enhanced TREM2, LC3II/LC3I and Beclin1 expression while reducing p62 in the ischemic penumbra area. The EA-induced neuroprotective effects and improvements in autophagic flux were abolished by specific knockdown of TREM2 in excitatory neurons. Taken together, our findings provide novel mechanistic insight into EA-induced ischemic tolerance and suggest a promising therapeutic strategy of targeting neuronal TREM2 to treat brain ischemia. • TREM2 expression on excitatory neurons play a role in cerebral ischemia/reperfusion. • Electroacupuncture pretreatment enhanced TREM2 expression and autophagic flux. • Neuronal TREM2 knockdown abolished EA-induced neuroprotective effects. [ABSTRACT FROM AUTHOR]

• Rosmarinic acid (a- O -caffeoyl-3,4-dihydroxyphenyllacticacid, RA) is one of the most potent polyphenols. Which is commercially available with high (>96%) purification and much less expensive compared to other polyphenol compounds. • RA administration alleviated neurological deficits and reduced infarct volume. • The results revealed RA treatment alleviated the post-stroke depressive behaviors. Rosmarinic acid (RA), a natural polyphenol, possesses potent antioxidant and anti-inflammatory activities. To evaluate the ability of RA to cure ischemic stroke and post-stroke depression (PSD), rats were treated with various doses of RA after cerebral ischemia. Neurological deficits and infarct volume of the brain were measured. The activities of superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH) were examined at different time points. In addition, a forced swimming test and sucrose preference test were performed to detect the anti-depressive effects of RA. Our results revealed RA administration significantly alleviated neurological deficits and reduced infarct volumes. RA attenuated the decrease of SOD, CAT activities and GSH levels in the ischemic penumbra of the brain. Most importantly, RA treatment alleviated the depression behaviors. Increased expression of Nrf2 was also induced by RA, while down regulation Nrf2 by Nrf2-short-hairpin RNA sequences reversed the increasing activity of SOD and CAT induced by RA, as well as the protection against PSD. The present study indicates that RA exerts a potent neuroprotective effect against stroke and PSD, which could be a promising therapeutic intervention for stroke. [ABSTRACT FROM AUTHOR]

Background: The steroid hormone estrogen (17-β-estradiol, E2) provides neuroprotection against cerebral ischemic injury by activating estrogen receptors. The novel estrogen receptor G protein-coupled receptor 30 (GPR30) is highly expressed in the brain and provides acute neuroprotection against stroke. However, the underlying mechanisms remain unclear.Methods: In this study, ovariectomized female mice were subjected to middle cerebral artery occlusion (MCAO), and E2, G1, and ICI182780 were administered immediately upon reperfusion. The infarction volume, neurological scores, and neuronal injuries were examined. Primary microglial cells were subjected to oxygen-glucose deprivation (OGD), and the drugs were administered immediately upon reintroduction. The pro-inflammatory cytokines TNF-α, IL-1β, and IL-6 in penumbra and microglia were assessed by ELISA. The cell viability and lactose dehydrogenase (LDH) release of neurons co-cultured with microglia were analyzed using cell counting kit-8 (CCK8) and LDH release assays. Microglial activation as well as GPR30, Iba1, and Toll-like receptor 4 (TLR4) protein expression and TLR4 mRNA expression were detected. Additionally, NF-κB activity was detected in lipopolysaccharide (LPS)-activated microglia after the activation of GPR30.Results: GPR30 was highly expressed in microglia and significantly increased after ischemic injury. The activation of GPR30 significantly reduced the infarction volume, improved the neurological deficit, and alleviated neuronal injuries. Moreover, GPR30 activation significantly reduced the release of TNF-α, IL-1β, and IL-6 from ischemic penumbra and microglia subjected to OGD and alleviated neuronal injury as assessed using the CCK8 and LDH assays. Finally, the activation of GPR30 relieved microglial activation, reduced Iba1 and TLR4 protein expression and TLR4 mRNA levels, and inhibited NF-κB activity.Conclusions: Microglial GPR30 exerts acute neuroprotective effects by inhibiting TLR4-mediated microglial inflammation, which indicates that GPR30 may be a potential target for the treatment of ischemic stroke. [ABSTRACT FROM AUTHOR]

It is known that preconditional treatment with volatile anesthetics can induce tolerance of the brain to stroke. A previous study demonstrated that the involvement of TREK-1, a two-pore domain K+ channel, in sevoflurane preconditioning induced neuroprotection against focal cerebral ischemia in rats. The present study testified whether TREK-2, another anesthetic-target K+ channel, is also associated with volatile anesthetic-induced neuroprotection, and further explored its potential mechanism. Rats preconditioned with isoflurane were subjected to 1.4vol% isoflurane plus 98% O2 (1.5 L/min) inhalation for 1 hour daily and continuing for 5 consecutive days. Then, these rats were subjected to middle cerebral artery occlusion (MCAO) as focal cerebral ischemia model. The expression of TWIK-related K+ channel 2 (TREK-2) was analyzed by western blotting and quantitative real-time RT-PCR, and its downstream signaling molecules, protein kinase C (PKC) alpha, extracellular signal-regulated kinase 1/2 (ERK1/2), and pERK1/2 were detected by western blotting also. Subsequently, the expression of TREK-2 was regulated by siRNA transfection in the brain to clarify its role in the neuroprotection of isoflurane preconditioning. Neurological scores, infarction volume, and TdT-mediated dUTP Nick-End Labeling (TUNEL) staining were examined to evaluate the outcomes. The impact of TREK-2 on the expression of its downstream signaling molecules was also examined for preliminary analysis of the possible mechanisms. Isoflurane preconditioning reduced the infarct volume, inhibited the cell apoptosis, and improved the neurological outcome in rats subjected to MCAO. These effects were parallel with the increase in TREK-2 protein and inhibition of the ERK1/2 phosphorylation. The downregulation of TREK-2 through siRNA could significantly attenuate the isoflurane preconditioning-induced neuroprotective effects. Isoflurane preconditioning-induced neuroprotective effects against ischemia-reperfusion injury are associated with the increase in TREK-2 channel activation. These effects depend on the attenuation of PKC alpha and inhibition of ERK1/2 phosphorylation. Results enrich our understanding on the mechanism of two-pore domain K+ channel in preconditioning-induced tolerance to focal cerebral ischemia. [ABSTRACT FROM AUTHOR]

Tanshinone IIA (TSA), a principal component derived from the Traditional Chinese Medicine Danshen has been suggested to exert neuroprotective effect against experimental cerebral ischemic/reperfusion injury. But the associated underlying mechanisms still have not been understood. The current study characterized the role of nuclear factor erythroid two-related factor-induced antioxidant response in the neuroprotective efficacy of TSA treatment. The focal cerebral ischemia/reperfusion model was established by 60-minute middle cerebral artery occlusion. At the onset during reperfusion, mice were treated with 10 mg/kg TSA intraperitoneally. The mRNA and nuclear factor erythroid 2 (Nrf2) protein expression, the antioxidant enzymes, and oxidative production levels were measured. To further verify the role of Nrf2 in the neuroprotective effect induced by TSA, the Nrf2 small silenced RNA and Nrf2 knockout mice were used, the neurological function, brain infarct volume, and cellular apoptosis examination were assessed. TSA treatment improved neurological scores, reduced infarct volume, and attenuated the cellular apoptosis. TSA treatment upregulated the expression of Nrf2 mRNA and the contents of Nrf2 protein in nuclear extract. Nrf2 activation by TSA treatment increased the contents of antioxidant enzymes, and reduced the generation of oxidative productions. Either Nrf2 knockdown or Nrf2 knockout abolished the antioxidative and neuroprotective effect of TSA treatment. These results demonstrate that the Nrf2 activation contributes to TSA-induced neuroprotection from experimental ischemic stroke through maintaining antioxidant effect. [ABSTRACT FROM AUTHOR]

Abstract: Ethnopharmacological relevance: Breviscapine injection is a Chinese herbal medicine standardized product extracted from Erigeron breviscapus (Vant.) Hand.-Mazz. It has been widely used for treating cardiovascular and cerebrovascular diseases. However, the therapeutic time window and the action mechanism of breviscapine are still unclear. The present study was designed to investigate the therapeutic time window and underlying therapeutic mechanism of breviscapine injection against cerebral ischemic/reperfusion injury. Materials and methods: Sprague–Dawley rats were subjected to middle cerebral artery occlusion for 2h followed by 24h of reperfusion. Experiment part 1 was used to investigate the therapeutic time window of breviscapine. Rats were injected intravenously with 50mg/kg breviscapine at different time-points of reperfusion. After 24h of reperfusion, neurologic score, infarct volume, brain water content and serum level of neuron specific enolase (NSE) were measured in a masked fashion. Part 2 was used to explore the therapeutic mechanism of breviscapine. 4-Hydroxy-2-nonenal (4-HNE), 8-hydroxyl-2′- deoxyguanosine (8-OHdG) and the antioxidant capacity of ischemia cortex were measured by ELISA and ferric-reducing antioxidant power (FRAP) assay, respectively. Immunofluorescence and western blot analysis were used to analyze the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1). Results: Part 1: breviscapine injection significantly ameliorated neurologic deficit, reduced infarct volume and water content, and suppressed the levels of NSE in a time-dependent manner. Part 2: breviscapine inhibited the increased levels of 4-HNE and 8-OHdG, and enhanced the antioxidant capacity of cortex tissue. Moreover, breviscapine obviously raised the expression of Nrf2 and HO-1 proteins after 24h of reperfusion. Conclusion: The therapeutic time window of breviscapine injection for cerebral ischemia/reperfusion injury seemed to be within 5h after reperfusion. By up-regulating the expression of Nrf2/HO-1 pathway might be involved in the therapeutic mechanism of breviscapine injection. [ABSTRACT FROM AUTHOR]

Abstract: Purposes: To investigate whether activation of adenosine A1 receptor (A1R) through limb remote ischemic preconditioning (RIPC) by a noninvasive tourniquet contribute neuroprotective effects against rat focal cerebral ischemic injury induced by transient middle cerebral artery occlusion (MCAO). Methods: One hundred twenty-eight Sprague–Dawley (SD) rats were randomly assigned into eight groups (n=16 each): MCAO, Control, 8-cyclopentyl-1,3-dipropulxanthine (DPCPX, Adenosine A1 receptor antagonist), RIPC, DPCPX+RIPC, Vehicle+RIPC, 2-chloro-N6-cyclopentyladenosine (CCPA, Adenosine A1 receptor agonist) and CCPA+DPCPX groups. All animals underwent right middle cerebral artery occlusion (MCAO) for 2h. Limb RIPC consisted of three cycles of 5-minute ischemia followed by 5-minute reperfusion in right hind-limb by tourniquet application. Neurological deficit scores were evaluated 24h after reperfusion, and then the infarct volume was assessed with diffusion weighted imaging (DWI) and 2, 3, 5-triphenyltetrazolium chloride (TTC) staining. Inflammation was assessed by serum tumor necrosis factor α (TNF α ) and NO; oxidative stress was estimated by malondialdehyde (MDA) and 4-hydroxyalkenals (4-HAD), superoxide dismutase (SOD) activity and GSH. Results: Animals in the RIPC, Vehicle+RIPC and CCPA groups developed lower neurological deficit scores and smaller brain infarct volumes than the Control group (P <0.01). Animals in the DPCPX, DPCPX+RIPC and CCPA+DPCPX groups developed higher neurological deficit scores and larger brain infarct volumes than the RIPC, Vehicle+RIPC and CCPA groups (P<0.01). DPCPX abolished the protective effects of RIPC and CCPA. RIPC or CCPA induced a significant increase in brain MnSOD (manganese SOD) activity and NO generation, and this activity was abolished by DPCPX pretreatment. RIPC or CCPA induced a significant reduction (P< 0.05) in the GSH and MDA+4HDA concentration and an accumulation in the GSSG concentration in both compartments (serum and tissue) as compared with the MCAO group. Conclusions: The present study demonstrates that limb RIPC induced by noninvasive tourniquet reduces cerebral ischemic injury in rats, and the effect of neuroprotection may depend on the activation of adenosine A1 receptors. CCPA pretreatment can induce delayed ischemic tolerance against cerebral ischemia/reperfusion injury. These protective effects are associated with a reduction in oxidative stress, inflammation and endogenous antioxidant preservation. [Copyright &y& Elsevier]