Role of P2X7 purinoceptors in neuroprotective mechanism of ischemic postconditioning in mice

Cerebral ischemia–reperfusion (I–R) injury is one of the primary causes of ischemic stroke. Ischemic postconditioning (iPoCo) is evolving as an important adaptive technique to contain I–R injury. Some recent studies have shown neuroprotective effect of iPoCo. However, neuroprotective mechanism of iPoCo is not clear. So, the present study has been undertaken to investigate the possible role of P2X7 purinoceptors in neuroprotective mechanism of iPoCo in mice. Bilateral carotid artery occlusion for 12 min followed by R for 24 h produced a significant rise in cerebral infarct size and neurological severity score (NSS) along with impairment of memory and motor coordination. iPoCo, involving three episodes of 10-s carotid artery occlusion with intermittent R of 10 s applied just after ischemic insult of 12 min, produced a significant decrease in cerebral infarct size and NSS along with reversal of I–R-induced impairment of memory and motor coordination. iPoCo induced neuroprotective effects were significantly abolished by pretreatment with selective purinergic P2X7 receptor blocker Brilliant Blue G (40 mg/kg intraperitoneal). It may be concluded that neuroprotective effect of iPoCo probably involves in activation of purinergic P2X7 receptors.     

7,8-dihydroxyflavone,a small-molecule tropomyosin-related kinase B (TrkB) agonist,attenuates cerebral ischemia and reperfusion injury in rats

7,8-dihydroxyflavone (7,8-DHF) is a recently identified potent agonist of tropomyosin-related kinase B that can cross the blood–brain barrier after oral or intraperitoneal administration. The aim of the present study was to determine whether 7,8-DHF has neuroprotective effects against cerebral ischemia and reperfusion (I/R) injury and, if so, to investigate the possible underlying mechanisms. Cerebral I/R injury rats were induced by middle cerebral artery occlusion for 90 min followed by reperfusion for 24 h. 7,8-DHF was administered intraperitoneally at a dose of 5 mg/kg immediately after ischemia. Our results showed that 7,8-DHF significantly reduced neurological deficit scores, infarct volumes, and neuronal apoptosis in brains of I/R rats. Meanwhile, 7,8-DHF also increased Bcl-2 expression, decreased expression of cleaved caspase-3, Bax and inducible nitric oxide synthase, and inhibited nuclear factor-κB activation in ischemic cortex. Finally, malondialdehyde and nitric oxide contents were reduced, but activities of glutathione, glutathione peroxidase and superoxide dismutase were restored in ischemic cortex treated with 7,8-DHF. Taken together, our findings demonstrated that 7,8-DHF is able to protect against cerebral I/R injury, which may be, at least in part, attributable to its anti-apoptotic, anti-oxidative and anti-inflammatory actions.     

The Involvement of Mitochondrial Biogenesis in Selenium Reduced Hyperglycemia-Aggravated Cerebral Ischemia Injury

Selenium has been shown to possess antioxidant and neuroprotective effects by modulating mitochondrial function and activating mitochondrial biogenesis. Our previous study has also suggested that selenium protected neurons against glutamate toxicity and hyperglycemia-induced damage by regulating mitochondrial fission and fusion. However, it is still not known whether the mitochondrial biogenesis is involved in selenium alleviating hyperglycemia-aggravated cerebral ischemia reperfusion (I/R) injury. The object of this study is to define whether selenium protects neurons against hyperglycemia-aggravated cerebral I/R injury by promoting mitochondrial biogenesis. In vitro oxygen deprivation plus high glucose model decreased cell viability, enhanced reactive oxygen species production, and meanwhile stimulated mitochondrial biogenesis signaling. Pretreated with selenium significantly decreased cell death and further activated the mitochondrial biogenesis signaling. In vivo 30 min of middle cerebral artery occlusion in the rats under hyperglycemic condition enhanced neurological deficits, enlarged infarct volume, exacerbated neuronal damage and oxidative stress compared with normoglycemic ischemic rats after 24 h reperfusion. Consistent to the in vitro results, selenium treatment alleviated ischemic damage in hyperglycemic ischemic animals. Furthermore, selenium reduced the structural changes of mitochondria caused by hyperglycemic ischemia and further promoted the mitochondrial biogenesis signaling. Selenium activates mitochondrial biogenesis signaling, protects mitochondrial structure integrity and ameliorates cerebral I/R injury in hyperglycemic rats.

     

Hydroxysafflor Yellow A Protects Against Cerebral Ischemia–Reperfusion Injury by Anti-apoptotic Effect Through PI3K/Akt/GSK3β Pathway in Rat

Hydroxysafflor yellow A (HSYA) is the major active chemical component of the flower of the safflower plant, Carthamus tinctorius L. Previously, its neuroprotection against cerebral ischemia–reperfusion (I/R) injury was reported by anti-oxidant action and suppression of thrombin generation. Here, we investigate the role of HSYA in cerebral I/R-mediated apoptosis and possible signaling pathways. Male Wistar rats were subjected to transient middle cerebral artery occlusion for 2 h, followed by 24 h reperfusion. HSYA was administered via tail-vein injection just 15 min after occlusion. The number of apoptotic cells was measured by TUNEL assay, apoptosis-related proteins Bcl-2, Bax and the phosphorylation levels of Akt and GSK3β in ischemic penumbra were assayed by western blot. The results showed that administration of HSYA at the doses of 4 and 8 mg/kg significantly inhibited the apoptosis by decreasing the number of apoptotic cells and increasing the Bcl-2/Bax ratio in rats subjected to I/R injury. Simultaneously, HSYA treatment markedly increased the phosphorylations of Akt and GSK3β. Blockade of PI3K activity by wortmannin dramatically abolished its anti-apoptotic effect and lowered both Akt and GSK3β phosphorylation levels. Taken together, these results suggest that HSYA protects against cerebral I/R injury partly by reducing apoptosis via PI3K/Akt/GSK3β signaling pathway.     

Neuroprotective effects of hydroxyethylpuerarin against focal cerebral ischemia-reperfusion in rats

Our present study was performed to investigate whether hydroxyethylpuerarin (HEP) has a neuroprotective effect on brain injury after focal cerebral ischemia/reperfusion by middle cerebral artery occlusion (MCAO) in adult male Wistar rats. Animals were subjected to one hour of middle cerebral artery occlusion and 48 hours of reperfusion with the pretreatment of drugs (HEP 15, 30, 60 mg/ kg or nimodipine 0.4 mg/kg i.v.) or vehicle. The behavioral tests were used to evaluate the damage to central nervous system. The percentage of brain infarct area was assessed in the brain slices stained with 2% solution of 2, 3, 5-triphenyl tetrazolium chloride (TTC). The pathologic histological changes were observed by H&E staining and the occurrence of apoptosis was determined by flow cytometry. The results showed that pretreatment with HEP at doses of 15, 30, 60 mg/kg exhibited significant neuroprotective effects on rats against focal cerebral ischemia-reperfusion injury by markedly decreasing neurological deficit scores and the percentage of infarct area, reducing necrosis and apoptosis of neurons. All these findings suggest that HEP might provide neuroprotection against focal cerebral ischemia/reperfusion injury probably through its antioxidant and anti-inflammatory property.     

Effects of increased accumulation of doxorubicin due to emodin on efflux transporter and LRP1 expression in lung adenocarcinoma and colorectal carcinoma cells

We investigated the eplerenone-induced, PI3K/Akt- and GSK-3β-mediated cardioprotection against ischemia/reperfusion (I/R) injury in diabetic rats. The study groups comprising diabetic rats were treated for 14 days with 150 mg/kg/day eplerenone orally and 1 mg/kg wortmannin (PI3K/Akt antagonist) intraperitoneally with eplerenone. On the 15th day, the rats were exposed to I/R injury by 20-min occlusion of the left anterior descending coronary artery followed by 30 min of reperfusion. The hearts were processed for biochemical, molecular, and histological investigations. The I/R injury in diabetic rats inflicted a significant rise in the oxidative stress and apoptosis along with a decrease in the arterial and ventricular function and the expressions of PI3K/Akt and GSK-3β proteins. Eplerenone pretreatment reduced the arterial pressure, cardiac inotropy, and lusitropy. It significantly reduced apoptosis and cardiac injury markers. The histology revealed cardioprotection in eplerenone-treated rats. Eplerenone up-regulated the PI3K/Akt and reduced the GSK-3β expression. The group receiving wortmannin with eplerenone was deprived eplerenone-induced cardioprotection. Our results reveal the eplerenone-induced cardioprotection against I/R injury in diabetic rats and substantiate the involvement of PI3K/Akt and GSK-3β pathways in its efficacy.     

Glycogen Synthase Kinase-3β Is Involved in Electroacupuncture Pretreatment via the Cannabinoid CB1 Receptor in Ischemic Stroke

We have previously shown that electroacupuncture (EA) pretreatment produces neuroprotective effects, which were mediated through an endocannabinoid signal transduction mechanism. Herein, we have studied the possible contribution of the phosphorylated form of glycogen synthase kinase-3β (GSK-3β) in EA pretreatment-induced neuroprotection via the cannabinoid CB1 receptor (CB1R). Focal transient cerebral ischemia was induced by middle cerebral artery occlusion in rats. Phosphorylation of GSK-3β at Ser-9 [p-GSK-3β (Ser-9)] was evaluated in the penumbra tissue following reperfusion. Infarct size and neurological score were assessed in the presence of either PI3K inhibitors or a GSK-3β inhibitor 72 h after reperfusion. Cellular apoptosis was evidenced by TUNEL staining and determination of the Bax/Bcl-2 ratio 24 h after reperfusion. The present study showed that EA pretreatment increased p-GSK-3β(Ser-9) 2 h after reperfusion in the ipsilateral penumbra. Augmented phosphorylation of GSK-3β induced similar neuroprotective effects as did EA pretreatment. By contrast, inhibition of PI3K dampened the levels of p-GSK-3β(Ser-9), and reversed not only the neuroprotective effect but also the anti-apoptotic effect following EA pretreatment. Regulation of GSK-3β by EA pretreatment was abolished following treatment with a CB1R antagonist and CB1R knockdown, whereas two CB1R agonists enhanced the phosphorylation of GSK-3β. Therefore we conclude that EA pretreatment protects against cerebral ischemia/reperfusion injury through CB1R-mediated phosphorylation of GSK-3β.     

Neuroprotective Effects of <Emphasis Type="Italic">Lycium barbarum</Emphasis> Polysaccharide on Focal Cerebral Ischemic Injury in Mice

Increasing evidence demonstrates inflammation contributes to neuronal death following cerebral ischemia. Lycium barbarum polysaccharide (LBP) has been reported to prevent scopolamine-induced cognitive and memory deficits. We recently indicated that LBP exerts neuroprotective effect against focal cerebral ischemic injury in mice via attenuating the mitochondrial apoptosis pathway. The aim of this study was to investigate the neuroprotective effects of LBP against the behavioral dysfunction induced by focal cerebral ischemia injury in mice. Following 7 successive days of pretreatment with LBP (10, 20 and 40 mg/kg) and nimodipine (4 mg/kg) by intragastric gavage, mice were subjected to middle cerebral artery occlusion (MCAO). Following reperfusion, cerebral blood flows, the total power of the spontaneous EEG, and morphological changes were estimated. Learning and memory ability, and motor coordination were determined by Morris water maze task, rotarod and grip test. Western blot analysis, Real-Time fluorogenic PCR assays, and immunofluorescence staining were used to examine the expression of proinflammatory mediators and activation of microglia. The present study showed that LBP pretreatment significantly enhanced regional cortical blood flow and the total power of the spontaneous EEG, improved memory and motor coordination impairments, and inhibited over-activation of microglia and astrocytes after MCAO. Further study demonstrated LBP suppressed MCAO-induced activations of P65 NF-κB and P38 MAPK, and prevented up-regulations of proinflammatory mediators in hippocampus. Our data suggest that LBP can exert functional recovery of memory and motor coordination deficits and neuroprotective effect against cerebral ischemic injury in mice.     

Curcumin Improves Outcomes and Attenuates Focal Cerebral Ischemic Injury via Antiapoptotic Mechanisms in Rats

Curcumin, a member of the curcuminoid family of compounds, is a yellow colored phenolic pigment obtained from the powdered rhizome of C. longa Linn. Recent studies have demonstrated that curcumin has protective effects against cerebral ischemia/reperfusion injury. However, little is known about its mechanism. In the present study, we tested the effects of curcumin in focal cerebral ischemia in rats and the possible mechanisms. Adult male Sprague–Dawley rats were treated with curcumin (100, 300 and 500 mg/kg) administered intraperitoneally after 60 min of occlusion (beginning of reperfusion). Neurological score and infarct volume were assessed at 24 and 72 h. Oxidative stress was evaluated by malondialdehyde assay and the apoptotic mechanisms were studied by Western blotting. Curcumin treatment significantly reduced infarct volume and improved neurological scores at different time points compared with the vehicle-treated group. Curcumin treatment decreased malondialdehyde levels, cytochrome c, and cleaved caspase 3 expression and increased mitochondrial Bcl-2 expression. Inhibition of oxidative stress with curcumin treatment improves outcomes after focal cerebral ischemia. This neuroprotective effect is likely exerted by antiapoptotic mechanisms.     

Neuroprotective effect of curcumin in middle cerebral artery occlusion induced focal cerebral ischemia in rats

Free radical induced neuronal damage is implicated in cerebral ischemia reperfusion (IR) injury and antioxidants are reported to have neuroprotective activity. Several in vitro and in vivo studies have proved the antioxidant potential of curcumin and its metabolites. Hence, in the present study the neuroprotective potential of curcumin was investigated in middle cerebral artery occlusion (MCAO) induced focal cerebral IR injury. 2 h of MCAO and 22 h of reperfusion resulted in the infarct volume of 210.39 +/- 31.25 mm3. Administration of curcumin 100 and 300 mg/kg, i.p. 30 min. after MCAO produced 37.23 +/- 5.10% and 46.39 +/- 10.23% (p < 0.05) reduction in infarct volume, respectively. Ischemia induced cerebral edema was reduced in a dose dependent manner. Curcumin at 300 mg/kg, i.p. produced 50.96 +/- 6.04% reduction in edema (p < 0.05) volume. Increase in lipid peroxidation after MCAO in ipsilateral and contralateral hemisphere of brain was observed, which was reduced by curcumin (300 mg/kg, i.p.)-treatment. Decrease in superoxide dismutase and glutathione peroxidase activity was observed in ipsilateral hemisphere of MCAO animal. Curcumin-treatment (300 mg/kg, i.p.) prevented IR injury mediated fall in glutathione peroxide activity. Peroxynitrite measured using rhodamine123 fluorescence and anti-nitrotyrosine immunofluorescence indicated increased peroxynitrite formation after IR insult. Curcumin-treatment reduced peroxynitrite formation and hence the extent of tyrosine nitration in the cytosolic proteins. These results suggest the neuroprotective potential of curcumin in cerebral ischemia and is mediated through its antioxidant activity.     

Shogaol potentiates sevoflurane mediated neuroprotection against ischemia/reperfusion-induced brain injury via regulating apoptotic proteins and PI3K/Akt/mTOR/s6K signalling and HIF-1α/HO-1 expression

The current research was intended to evaluate the impact of 6-shogaol in rodent model of ischemic-reperfusion induced- brain injury and also assessed 6-shogaol enhanced sevoflurane''s neuroprotective effects. Ischemic-Reperfusion (I/R) injury was induced by middle cerebral artery occlusion (MCAO) method in Sprague-Dawley rats. A separate group of animal was exposed to sevoflurane (2.5%) post-conditioning for 1 h immediately after reperfusion. The 6-shogaol (25 mg or 50 mg/kg body weight) was orally administered to treatment group rats for 14 days and then subjected to I/R. The 6-shogaol treatment along with/without sevoflurane post-conditioning reduced the number of apoptotic cell counts, brain edema and cerebral infarct volume. The western blotting analysis revealed a significant stimulation of the PI3K/Akt/mTOR signal pathway. RT-PCR and western blotting studies revealed improved expressions of HIF-1α and HO-1 at both gene level and protein levels. I/R induced neurological deficits were also alleviated on sevoflurane post-conditioning with/without 6-shogaol treatment. The present findings revealed that pre-treatment with 6-shogoal enhanced the neuroprotective properties of sevoflurane post-conditioning, illustrated the efficacy of the compound against I/R injury.     

Shogaol potentiates sevoflurane mediated neuroprotection against ischemia/reperfusion-induced brain injury via regulating apoptotic proteins and PI3K/Akt/mTOR/s6K signalling and HIF-1α/HO-1 expression

The current research was intended to evaluate the impact of 6-shogaol in rodent model of ischemic-reperfusion induced- brain injury and also assessed 6-shogaol enhanced sevoflurane's neuroprotective effects. Ischemic-Reperfusion (I/R) injury was induced by middle cerebral artery occlusion (MCAO) method in Sprague-Dawley rats. A separate group of animal was exposed to sevoflurane (2.5%) post-conditioning for 1 h immediately after reperfusion. The 6-shogaol (25 mg or 50 mg/kg body weight) was orally administered to treatment group rats for 14 days and then subjected to I/R. The 6-shogaol treatment along with/without sevoflurane post-conditioning reduced the number of apoptotic cell counts, brain edema and cerebral infarct volume. The western blotting analysis revealed a significant stimulation of the PI3K/Akt/mTOR signal pathway. RT-PCR and western blotting studies revealed improved expressions of HIF-1α and HO-1 at both gene level and protein levels. I/R induced neurological deficits were also alleviated on sevoflurane post-conditioning with/without 6-shogaol treatment. The present findings revealed that pre-treatment with 6-shogoal enhanced the neuroprotective properties of sevoflurane post-conditioning, illustrated the efficacy of the compound against I/R injury.     

Nicotine-Induced Neuroprotection Against Ischemic Injury Involves Activation of Endocannabinoid System in Rats

Nicotine has been reported to exert certain protective effect in the Parkinson’s and Alzheimer’s diseases. Whether it has a similar action in focal cerebral ischemia was unclear. In the present study, rats received either an injection of (?)-nicotine hydrogen tartrate salt (1.2 mg/kg, i.p.) or the vehicle 2 h before the 120 min middle cerebral artery occlusion. Neurological deficits and histological injury were assessed at 24 h after reperfusion. The content of endocannabinoids and the expression of cannabinoid receptor CB1 in brain tissues were determined at different time points after nicotine administration. Results showed that nicotine administration ameliorated neurological deficits and reduced infarct volume induced by cerebral ischemia in the rats. The neuroprotective effect was partially reversed by CB1 blockage. The content of the endocannabinoids N-arachidonylethanolamine and 2-arachidonoylglycerol, as well as the expression of cannabinoid receptor CB1 were up-regulated in brain tissues after nicotine delivery. These results suggest that endogenous cannabinoid system is involved in the nicotine-induced neuroprotection against transient focal cerebral ischemia.     

Donepezil provides neuroprotective effects against brain injury and Alzheimer's pathology under conditions of cardiac ischemia/reperfusion injury

Cardiac ischemia/reperfusion (I/R) injury induces brain pathology. Donepezil, a well-known acetylcholine esterase (AChE) inhibitor, has been proven to exert neuroprotective effects against several neurodegenerative diseases. However, the comprehensive mechanism regarding the therapeutic potential of donepezil on the brain under cardiac I/R injury remains obscure. Here, we hypothesized that treatment with donepezil ameliorates brain pathology following cardiac I/R injury by decreasing blood brain barrier (BBB) breakdown, oxidative stress, neuroinflammation, mitochondrial dysfunction, mitochondrial dynamics imbalance, microglial activation, amyloid-beta (Aβ) accumulation, neuronal apoptosis, and dendritic spine loss. Forty-eight adult male Wistar rats were subjected to surgery for cardiac I/R injury. Then, rats were randomly divided into four groups to receive either (1) saline (vehicle group), donepezil 3 mg/kg via intravenously administered (2) before ischemia (pretreatment group), (3) during ischemia (ischemia group), or (4) at the onset of reperfusion (reperfusion group). At the end of cardiac I/R paradigm, the brains were evaluated for BBB breakdown, brain inflammation, oxidative stress, mitochondrial function, mitochondrial dynamics, microglial morphology, Aβ production, neuronal apoptosis, and dendritic spine density. Administration of donepezil at all time points equally showed an attenuation of brain damage in response to cardiac I/R injury, as indicated by increased expression of BBB junction protein, reduced brain inflammation and oxidative stress, improved mitochondrial function and mitochondrial dynamics, and alleviated Aβ accumulation and microglial activation, resulting in protection of neuronal apoptosis and preservation of dendritic spine number. These findings suggest that donepezil potentially protects brain pathology caused by cardiac I/R injury regardless the timing of treatment.     

Delayed neuroprotection against cerebral ischemia reperfusion injury: putative role of BDNF and GSK-3β

Aim: Numerous studies have demonstrated the possible neuroprotective role of lithium treatment against neurological disorders. However, the role of lithium in delayed phase of neuronal death against focal ischemia has not been explored. Therefore, the present study was designed to investigate the effect and molecular mechanisms of post-lithium treatment against cerebral ischemic reperfusion (I/R) injury and associated cognitive deficits in rats. Methods: I/R injury was induced by right middle cerebral artery occlusion and lithium (40 and 60?mg/kg) were given intraperitoneally, 24?h after the insult and continued for 1 week with 24-h interval. Using Lasser Doppler, cerebral blood flow was monitored before, during and after MCAO induction. Besides behavioral, biochemical, and histological evaluation, levels of tumor necrosis factor alpha (TNF-α) and brain-derived neurotrophic factor (BDNF) were also estimated. Results: I/R injury resulted in significant elevation of neurological deficits, oxidative stress, neuroinflammation, and cognitive impairments. We found that lithium injection, 24?h after I/R-injury continued for 1 week, dose dependently prevented behavioral abnormality and cognitive impairments. Moreover, lithium attenuated the levels of oxidative stress and pro-inflammatory-cytokines TNF-α level. Further, lithium treatments significantly reduced neuronal damage and augmented healthy neuronal count and improved neuronal density in hippocampus. These neuroprotective effects of delayed lithium treatment were associated with upregulation of neurotrophic factor BDNF levels. Conclusion: Delayed lithium treatment provides neuroprotection against cerebral I/R injury and associated cognitive deficits by upregulating BDNF expression that opens a new avenue to treat I/R injury even after active cell death.     

3-Bromo-7-nitroindazole attenuates brain ischemic injury in diabetic stroke via inhibition of endoplasmic reticulum stress pathway involving CHOP

AimsThe role of nitric oxide (NO) and endoplasmic reticulum (ER) stress has been implicated in the pathogenesis of cerebral ischemic/reperfusion (I/R) injury and diabetes. The aim of the study was to investigate the neuroprotective potential of 3-bromo-7-nitroindazole (3-BNI), a potent and selective neuronal nitric oxide synthase (nNOS) inhibitor against ER stress and focal cerebral I/R injury associated with comorbid type 2 diabetes in-vivo.Main methodsType 2 diabetes was induced by feeding high-fat diet and streptozotocin (35 mg/kg) treatment in rats. Focal cerebral ischemia was induced by 2 h middle cerebral artery occlusion (MCAO) followed by 22 h of reperfusion. Immunohistochemistry and western blotting methods were employed for the detection and expression of ER stress/apoptosis markers [78 kDa glucose regulated protein (GRP78) and CCAAT/enhancer binding protein homologous protein (CHOP)]. TUNEL assay for DNA fragmentation was also performed.Key findingsThe diabetic rats subjected to cerebral I/R had prominent neurological damage and functional deficits compared with sham-operated rats. Massive DNA fragmentation was observed in ischemic penumbral region of diabetic brains. Concomitantly, the enhanced immunoreactivity and expression of ER stress/apoptosis markers were noticed. 3-BNI (30 mg/kg, i.p.) treatment significantly inhibited the cerebral infarct, edema volume and improved functional recovery of neurological deficits. The neuroprotection was further evident by lesser DNA fragmentation with a concomitant reduction of GRP78 and CHOP.SignificanceThe study demonstrates the neuroprotective potential of 3-BNI in diabetic stroke model which may be partly due to inhibition of ER stress pathway involving CHOP.     

Neuroprotection by tetramethylpyrazine against ischemic brain injury in rats

In traditional Chinese medicine, Ligusticum wallichii Franchat (Chuan Xiong) and its active ingredient tetramethylpyrazine (TMP) have been used to treat cardiovascular diseases and to relieve various neurological symptoms such as ischemic deficits. However, scientific evidence related to their effectiveness or precise modes of neuroprotective action is largely unclear. In the current study, we elicited the neuroprotective mechanisms of TMP after focal cerebral ischemic/reperfusion (I/R) by common carotid arteries and middle cerebral artery occlusion model in rats. TMP was administrated 60 min before occlusion via intraperitoneal injection. TMP concentration-dependently exhibited significant neuroprotective effect against ischemic deficits by reduction of behavioral disturbance. Neuronal loss and brain infarction in the ischemic side of rats were markedly lowered by treatment with TMP. Cerebral I/R-induced internucleosomal DNA fragmentation, caspase-8, caspase-9, and caspase-3 activation, and cytochrome c release were reduced by TMP treatment. Western blot analysis revealed the down-regulation of Bcl-2 and Bcl-xL and the up-regulation of Bax and Bad by cerebral I/R insult. Among them, only the alteration in Bcl-xL expression was reversed by TMP treatment. Moreover, the activation of microglia and/or recruitment of inflammatory cells within the ischemic side and the consequent production of monocyte chemoattractant protein 1 (MCP-1) were suppressed by TMP pre-treatment. Our findings suggest that TMP might provide neuroprotection against ischemic brain injury, in part, through suppression of inflammatory reaction, reduction of neuronal apoptosis, and prevention of neuronal loss.     

UPEI-100, a conjugate of lipoic acid and apocynin, mediates neuroprotection in a rat model of ischemia/reperfusion

Previous work in our laboratory has provided evidence that preadministration of apocynin and lipoic acid at subthreshold levels for neuroprotection enhanced the neuroprotective capacity when injected in combination. Therefore, the present investigation was designed to determine whether a co-drug consisting of lipoic acid and apocynin functional groups bound by a covalent bond, named UPEI-100, is capable of similar efficacy using a rodent model of stroke. Male rats were anesthetized with Inactin (100 mg/kg iv), and the middle cerebral artery was occluded for 6 h or allowed to reperfuse for 5.5 h following a 30-min occlusion (ischemia/reperfusion, I/R). Preadministration of UPEI-100 dose-dependently decreased infarct volume in the I/R model (P < 0.05), but not in the middle cerebral artery occlusion model of stroke. Using the optimal dose, we then injected UPEI-100 during the stroke or at several time points during reperfusion, and significant neuroprotection was observed when UPEI-100 was administered up to 90 min following the start of reperfusion (P < 0.05). A time course for this neuroprotective effect showed that UPEI-100 resulted in a decrease in infarct volume following 2 h of reperfusion compared with vehicle. The time course of this neuroprotective effect was also used to study several mediators along the antioxidant pathway and showed that UPEI-100 increased the level of mitochondrial superoxide dismutase and oxidized glutathione and decreased a marker of lipid peroxidation due to oxidative stress (HNE-His adduct formation). Taken together, the data suggest that UPEI-100 may utilize similar pathways to those observed for the two parent compounds; however, it may also act through a different mechanism of action.     

Butylphthalide Suppresses Neuronal Cells Apoptosis and Inhibits JNK–Caspase3 Signaling Pathway After Brain Ischemia /Reperfusion in Rats

Although Butylphthalide (BP) has protective effects that reduce ischemia-induced brain damage and neuronal cell death, little is known about the precise mechanisms occurring during cerebral ischemia/reperfusion (I/R). Therefore, the aim of this study was to investigate the neuroprotective mechanisms of BP against ischemic brain injury induced by cerebral I/R through inhibition of the c-Jun N-terminal kinase (JNK)–Caspase3 signaling pathway. BP in distilled non-genetically modified Soybean oil was administered intragastrically three times a day at a dosage of 15 mg/(kg day) beginning at 20 min after I/R in Sprague–Dawley rats. Immunohistochemical staining and Western blotting were performed to examine the expression of related proteins, and TUNEL-staining was used to detect the percentage of neuronal apoptosis in the hippocampal CA1 region. The results showed that BP could significantly protect neurons against cerebral I/R-induced damage. Furthermore, the expression of p-JNK, p-Bcl2, p–c-Jun, FasL, and cleaved-caspase3 was also decreased in the rats treated with BP. In summary, our results imply that BP could remarkably improve the survival of CA1 pyramidal neurons in I/R-induced brain injury and inhibit the JNK–Caspase3 signaling pathway.