Protective effects of combined treatment with ciprofol and mild therapeutic hypothermia during cerebral ischemia-reperfusion injury

Despite improvement in cardiopulmonary resuscitation(CPR)performance,cardiac arrest(CA)is still associated with poor prognosis.The high mortality rate is due to multi-organ dysfunction caused by cerebral ischemia and reperfusion injury(I/R).The guidelines for CPR suggest the use of therapeutic hypothermia(TH)as an effective treatment to decrease mortality and the only approach confirmed to reduce I/R injury.During TH,sedative agents(propofol)and analgesia agents(fentanyl)are commonly used to prevent shiver and pain.However,propofol has been associated with a number of serious adverse effects such as metabolic acidosis,cardiac asystole,myocardial failure,and death.In addition,mild TH alters the pharmacokinetics of agents(propofol and fentanyl)and reduces their systemic clearance.For CA patients undergoing TH,propofol can be overdosed,leading to delayed awakening,prolonged mechanical ventilation,and other subsequent complications.Ciprofol(HSK3486)is a novel anesthetic agent that is convenient and easy to administer intravenously outside the operating room.Ciprofol is rapidly metabolized and accumulates at low concentrations after continuous infusion in a stable circulatory system compared to propofol.Therefore,we hypothesized that treatment with HSK3486 and mild TH after CA could protect the brain and other organs.

Acacetin protects against cerebral ischemia-reperfusion injury via the NLRP3 signaling pathway

Acacetin(5,7-dihydroxy-4′-methoxyflavone), a potential neuroprotective agent, has an inhibitory effect on lipopolysaccharide-induced neuroinflammatory reactions. However, whether acacetin has an effect on inflammatory corpuscle 3(NLRP3) after cerebral ischemia-reperfusion injury has not been fully determined. This study used an improved suture method to establish a cerebral ischemia-reperfusion injury model in C57BL/6 mice. After ischemia with middle cerebral artery occlusion for 1 hour, reperfusion with intraperitoneal injection of 25 mg/kg of acacetin(acacetin group) or an equal volume of saline(0.1 mL/10 g, middle cerebral artery occlusion group) was used to investigate the effect of acacetin on cerebral ischemia-reperfusion injury. Infarct volume and neurological function scores were determined by 2,3,5-triphenyltetrazolium chloride staining and the Zea-Longa scoring method. Compared with the middle cerebral artery occlusion group, neurological function scores and cerebral infarction volumes were significantly reduced in the acacetin group. To understand the effect of acacetin on microglia-mediated inflammatory response after cerebral ischemia-reperfusion injury, immunohistochemistry for the microglia marker calcium adapter protein ionized calcium-binding adaptor molecule 1(Iba1) was examined in the hippocampus of ischemic brain tissue. In addition, tumor necrosis factor-α, interleukin-1β, and interleukin-6 expression in ischemic brain tissue of mice was quantified by enzyme-linked immunosorbent assay. Expression of Iba1, tumor necrosis factor-α, interleukin-1β and interleukin-6 was significantly lower in the acacetin group compared with the middle cerebral artery occlusion group. Western blot assay results showed that expression of Toll-like receptor 4, nuclear factor kappa B, NLRP3, procaspase-1, caspase-1, pro-interleukin-1β, and interleukin-1β were significantly lower in the acacetin group compared with the middle cerebral artery occlusion group. Our findings indicate that acacetin has a protective effect on cerebral ischemia-reperfusion injury, and its mechanism of action is associated with inhibition of microglia-mediated inflammation and the NLRP3 signaling pathway.

Salvianolate increases heat shock protein expression in a cerebral ischemia-reperfusion injury model

Stroke remains a worldwide health problem. Salvianolate exerts a protective effect in various mi- crocirculatory disturbance-related diseases, but studies of the mechanisms underlying its protective action have mainly focused on the myocardium, whereas little research has been carried out in brain tissue following ischemia-reperfusion. We assessed the neuroprotective effects of salvianolate in a rat model of cerebral ischemia-reperfusion injury induced using the suture method. At onset and 24 and 48 hours after reperfusion, rats were intraperitoneally injected with salvianolate （18 mg/kg） or saline. Neurological deficit scores at 72 hours showed that the neurological functions of rats that had received salvianolate were significantly better than those of the rats that had received saline. 2,3,5-Triphenyltetrazolium chloride was used to stain cerebral tissue to determine the extent of the infarct area. A significantly smaller infarct area and a significantly lower number of apoptotic cells were observed after treatment with salvianolate compared with the saline treatment. Expression of heat shock protein 22 and phosphorylated protein kinase B in ischemic brain tissue was significantly greater in rats treated with salvianolate compared with rats treated with saline. Our findings suggest that salvianolate provides neuroprotective effects against cerebral ischemia-reperfusion injury by upregulating heat shock protein 22 and phosphorylated protein kinase B expression.

Neuroprotective effect of penehyclidine hydrochloride on focal cerebral ischemia-reperfusion injury

Penehyclidine hydrochloride can promote microcirculation and reduce vascular permeability. However, the role of penehyclidine hydrochlodde in cerebral ischemia-reperfusion injury remains unclear. In this study, in vivo middle cerebral artery occlusion models were established in experimental rats, and penehyclidine hydrochloride pretreatment was given via intravenous injection prior to model establishment. Tetrazolium chloride, terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling and immunohistochemical staining showed that, penehyclidine hydrochloride pretreatment markedly attenuated neuronal histopathological changes in the cortex, hippocampus and striatum, reduced infarction size, increased the expression level of BcI-2, decreased the expression level of caspase-3, and inhibited neuronal apoptosis in rats with cerebral ischemia-reperfusion injury. Xanthine oxidase and thiobarbituric acid chromogenic results showed that penehyclidine hydrochloride upregulated the activity of superoxide dismutase and downregulated the concentration of malondialdehyde in the ischemic cerebral cortex and hippocampus, as well as reduced the concentration of extracellular excitatory amino acids in rats with cerebral ischemia-reperfusion injury. In addition, penehyclidine hydrochloride inhibited the expression level of the NR1 subunit in hippocampal nerve cells in vitro following oxygen-glucose deprivation, as detected by PCR. Experimental findings indicate that penehyclidine hydrochloride attenuates neuronal apoptosis and oxidative stress injury after focal cerebral ischemia-reperfusion, thus exerting a neuroprotective effect.

Experimental Study on the Protection of Agrimony Extracts from Different Extracting Methods against Cerebral Ischemia-Reperfusion Injury

Objective To study the protective effect of agrimony extracts from different extracting methods on cerebral ischemia-reperfusion injury in rats, in order to optimize the extraction scheme of agrimony.Methods Male rats were randomly assigned into seven groups: 1. Sham-operated group, 2. Untreated MCAO group (MCAO), 3. Petroleum ether extract of Agrimonia pilosa treated MCAO group (PEA), 4. Ethyl acetate extract of Agrimonia pilosa treated MCAO group (EAEA), 5. Ethanol extract of Agrimonia pilosa treated MCAO group (EEA), 6. Water extract of Agrimonia pilosa treated MCAO group (WEA), 7. Nimodipine treated MCAO group (NP). Intragastrical drug administration (i.g) was performed at 0 and 6 hours after MCAO.Neurological function tests were performed after reperfusion for 24 hours, then the brain was removed for the evaluations of the cerebral infarction volume (percentage of total brain volume) by immunohistochemistry,histological changes (hematoxylin-eosin staining), Na+/K+-ATPase, Ca2+-ATPase (modified method of Svoboda and Mosinger), mRNA expression of Tumor suppressor gene (P53) and hot shock protein (HSP70)(quantitative real-time PCR).Results The neurological function of MCAO group had significantly higher scores than the sham group (P<0.01). The WEA group showed a significantly lower neurological score than the MCAO group (P<0.05),indicating the protective effect of WEA on neurological deficits. The mean infarction volumes of WEA (13.5±6.6%, F=4.75, P<0.01), EEA (19.90±6.90%, F=5.23, P<0.01), PEA (20.40±5.30%, F=4.68,P<0.01) and EAEA (22.50±10.50%, F=6.25, P<0.05) group were all significantly smaller than that of MCAO group (29.40±6.50%). HE staining demonstrated that, compared to the treated groups, the infarcted cerebral tissue of MCAO group had more swelling neural cells, lighter stained nucleus, fewer and irregularly distributed neurons. The activity of Na+/K+-ATPase and Ca2+-ATPase reduced in the MCAO group (3.67±0.48 U/mg,1.28±0.26 U/mg, respectively), and were significantly higher in WEA group (7.56±0.85 U/mg, F=12.65,P=0.010; 3.59±0.22 U/mg, F=8.32, P=0.041, respectively). The MCAO group showed significantly elevated P53 and HSP70 mRNA expressions compared to the sham group (P<0.01, P<0.05). P53 mRNA expressions in Agrimony extracts treated groups were significantly lower than that of the MCAO group (all P<0.01), with the WEA group showing the greatest difference from MCAO group. The HSP70 mRNA level of the treated groups were not significantly different from that of the MCAO group.Conclusions Treatment using water extracts of agrimony can promote the best functional and metabolic recovery for rat model of cerebral ischemia-reperfusion injury, which maybe relate with the upregulation of energy metabolism in nerve cells after MCAO.

Effect of minocycline on cerebral ischemia-reperfusion injury

Minocylcine, a tetracycline derivate, has been shown to cross the blood-brain barrier and enter the central nervous system. In this study, cerebral ischemia-reperfusion injury models were established using the suture method, and minocycline was immediately injected intraperitoneally after cerebral ischemia-repeffusion （22.5 mg/kg, initially 45 mg/kg） at a 12-hour interval. Results showed that after minocycline treatment, the volume of cerebral infarction was significantly reduced, the number of surviving cell in the hippocampal CA1 region increased, the number of apoptotic cells decreased, the expression of caspase-3 and poly（adenosine diphosphate-ribose） polymerase-1 protein was down-regulated, and the escape latency in the water maze test was significantly shortened compared with the ischemia-reperfusion group. Our experimental findings indicate that minocycline can protect against neuronal injury induced by focal ischemia-reperfusion, which may be mediated by the inhibition of caspase-3 and poly（adenosine diphosphate-ribose） polymerase-1 protein expression.

Amyloid beta-peptide worsens cognitive impairment following cerebral ischemia-reperfusion injury

Amyloid 13-peptide, a major component of senile plaques in Alzheimer＇s disease, has been implicated in neuronal cell death and cognitive impairment. Recently, studies have shown that the pathogenesis of cerebral ischemia is closely linked with Alzheimer＇s disease. In this study, a rat model of global cerebral ischemia-reperfusion injury was established via occlusion of four arteries; meanwhile, fibrillar amyloid [3-peptide was injected into the rat lateral ventricle. The Morris water maze test and histological staining revealed that administration of amyloid 13-peptide could further aggravate impairments to learning and memory and neuronal cell death in the hippocampus of rats subjected to cerebral ischemia-reperfusion injury. Western blot showed that phosphorylation of tau protein and the activity of glycogen synthase kinase 313 were significantly stronger in cerebral ischemia-reperfusion injury rats subjected to amyloid [3-peptide administration than those undergo- ing cerebral ischemia-repetfusion or amyloid 13-peptide administration alone. Conversely, the activ- ity of protein phosphatase 2A was remarkably reduced in rats with cerebral ischemia-reperfusion injury following amyloid 13-peptide administration. These findings suggest that amyloid 13-peptide can potentiate tau phosphorylation induced by cerebral ischemia-reperfusion and thereby aggravate cognitive impairment.

Expression of nerve growth factor precursor, mature nerve growth factor and their receptors during cerebral ischemia-reperfusion injury

We investigated nerve growth factor precursor （proNGF） and mature NGF expression in ischemic and non-ischemic cortices after cerebral ischemia-reperfusion injury. In both ischemic and non-ischemic cortices, proNGF was found to be present in the extracellular space and cytoplasm. In addition, mature NGF was expressed in extracellular space, but with a very low signal. In ischemic cortex only, proNGF was significantly decreased, reaching a minimal level at 1 day. Mature NGF was increased at 4 hours, then reached a minimal level at 3 days. The p75 neurotrophin receptor （p75NTR） was significantly decreased after ischemia, and increased at 3 days after ischemia. These results confirmed that proNGF was the predominant form of NGF during the pathological process of cerebral ischemia-repeffusion injury. In addition, our findings suggest that ischemic injury may influence the conversion of proNGF to mature NGF, and that proNGF/p75NTR may be involved in reperfusion injury.

Acupuncture effects on serum myelin basic protein and remyelination following 30 minutes and 2 hours of ischemia in a rat model of cerebral ischemia-reperfusion injury

BACKGROUND： Acupuncture treatment on injured cerebral axons has shown to provide efficacy in clinical practice. It is unknown whether acupuncture produces therapeutic effects by protecting injured cerebral myelin in ischemic stroke. OBJECTIVE： To test whether acupuncture provides protection for injured cerebral myelin, based on quantitative data from cerebral ischemia-reperfusion rats, and to compare the effects of early and late acupuncture on serum myelin basic protein （MBP） content and remyelination of the ischemic internal capsule.DESIGN, TIME AND SETTING： A randomized, controlled experiment was performed at the Neurobiological Laboratory, Sichuan University from March 2005 to March 2006. MATERIALS： ＂Hua Tuo＂ Brand filiform needles were produced by the Medical Instrument Factory of Suzhou, China.METHODS： A total of 52 adult, healthy, male, Sprague Dawley rats were randomly assigned to four groups： control （n = 4）, model （n = 16）, early acupuncture （n = 16）, and late acupuncture （n = 16）. The focal cerebral ischemia-reperfusion model was established by middle cerebral artery occlusion in the right hemisphere using the modified thread embolism method in the latter three groups. Early and late acupuncture groups underwent acupuncture after ischemia for 30 minutes and 2 hours using the Xingnaokaiqiao needling method, respectively. Acupoints were ＂Neiguarf＇ （PC 6） and ＂Sanyinjiao＂ （SP 6） on the bilateral sides, as well as ＂Shuigou＇ （DU 26） and ＂Baihui＂ （DU 20） with stimulation for 1 minute at each acupoint. Acupuncture at all acupoints was performed two or three times while the needle was retained, once per day. No special handling was administered to the control clroup.MAIN OUTCOME MEASURES： For each group, remyelination of the internal capsule was observed by Pal-Weigert＇s myelin staining and serum MBP content was detected using enzyme-linked immunosorbent assay method on days 1,3, 5, and 7 following ischemia-reperfusion injury.RESULTS： Compared with the control group, massive demyelination of the internal capsule occurred, and serum MBP content increased in the model group （P 〈 0.05）. Compared with the model group, the extent of demyelination in the internal capsule was less distinct and serum MBP content was significantly less in the early and late acupuncture group （P 〈 0.01 ）. Compared with the late acupuncture group, serum MBP content reached a peak later and the peak value was less in the early acupuncture group. CONCLUSION： Results suggest that acupuncture exerts a protective effect on injured cerebral myelin in ischemia-reperfusion rats by reducing serum MBP content and promoting remyelination. The study also suggests that the effect of early acupuncture is superior to late acupuncture.

Effects of ligustrazine on somatosensory evoked potential in normal rabbits and rabbits with cerebral ischemia-reperfusion injury

BACKGROUND： Somatosensory evoked potential （SEP） has become a method with higher sensitivity and specificity than electroencephalogram in detecting the brain function and the region, range and degree of ischemia. However, the effects of ligustrazine on SEP is still not clear. OBJECTIVE ： To study the protective effects of ligustrazini injection on cerebral ischemia-reperfusion injury.DESIGN： Auto-control study, random grouping.SETTING： Qilu Hospital of Shandong University.MATERIALS： The experiment was completed in the Cerebral Functional Room of Qilu Hospital Affiliated to Shandong University from March 2002 to June 2004. A totally of 24 healthy Harbin rabbits were randomly divided into blank control group （n=8）, model control group （n=8） and ligustrazine treatment group （n=8）. Hydrochloric ligustrazine injection, 40 mg/2 mL each ampoule, was provided by the Third Pharmaceutical Factory of Beijing （certification： 93035236273）. The main component was hydrochloric ligustrazine and the chemical name was 2, 3, 5, 6-tetramethyl pyrazine hydrochloride. METHODS：① Modeling method： The bilateral common carotid artery ligation was adopted to make the model. ② Index of cerebral functional lesion evaluated with SEP during ischemia-reperfusion： DISA 2000C neuromyoeletrometer provided by Dantec Electronics Ltd, Denmark was used to detect SEP. ③ Interventional process： Blank control group： The latencies and amplitudes of SEP were measured before injection with 1.5 mg/kg ligustrazine and at the points of 15 minutes, 20 minutes, 30 minutes, 60 minutes, 90 minutes and 120 minutes after injection. Ligustrazine treatment group： Rabbits were injected with 1.5 mg/kg ligustrazine, and those of model control group were injected the same volume of saline. Thirty minutes later, the bilateral common carotid artery of the rabbits all had been ligated for 30 minutes, and then reperfused for 120 minutes. The latencies and amplitudes of SEP were measured before injection, before ligation, at the points of 1 minute, 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes and 30 minutes after ligation, and at the points of 5 minutes, 10 minutes, 15 minutes, 20 minutes, 30 minutes, 60 minutes, 90 minutes and 120 minutes after reperfusion.④ Evaluating criteria： Normal values of P-wave latencies and amplitudes were （19.34±3.18） ms and （4.55±1.43）μV. Average value before injection in blank control group and average values before injection, after injection and before ligation in ischemiareperfusion group were regarded as control criteria to evaluate changes of P-wave latencies and amplitudes after experiment. MAIN OUTCOME MEASURES： P-wave latencies and amplitudes of SEP in the three groups.RESULTS ： A total of 24 rabbits were involved in the final analysis without any loss.① Blank control group： The P-wave latencies delayed markedly at each time point after injection. Compared with that before injection, there was a significant difference （P 〈 0.05-0.01）. The P-wave amplitudes did not fluctuate noticeably all the time after injection, but significantly decreased when compared with those before injection （P 〈 0.05-0.01）. ② Ischemia-reperfusion group： The P-wave latencies delayed and amplitudes decreased in the rabbits with cerebral ischemia-reperfusion at all points of time during cerebral ischemia-reperfusion, and there was significant difference when compared with the levels before ischemia （P 〈 0.05）. When ligustrazine was injected, the latencies and amplitudes changed less, and as compared with the levels before ischemia, the difference was not significant （P〉 0.05）.CONCLUSION：① Ligustrazine can inhibit P-wave latencies and amplitudes of SEP of normal rabbits.②Ligustrazine can improve P-wave latencies and amplitudes of SEP of rabbits with cerebral ischemia-reperfusion injury.

Neuroprotective effect of cerebroprotein hydrolysate on cerebral ischemia-reperfusion injury mice

OBJECTIVE To investigate the neuroprotective effect of cerebroprotein hydroly⁃sate(CH)on cerebral ischemia-reperfusion injury in mice.METHODS A total of 60 male SPF Kunming mice were randomly divided,reforming longa method into sham group(sham),model group(tMCAO,reforming longa method),CH 0.2 and 0.5 g·kg-1 groups and positive drug control group(edaravone 0.008 g·kg-1).Neurological deficit score were performed 24 h after opera⁃tion.Mice with scores ranged between 1 and 3 were included in subsequent experiments.Each group had 8 mice.CH edaravone and normal sa⁃line were ip injected for 5 d.The tMCAO group and the sham group were administered the same amount of normal saline as administration groups.TTC staining was used to measure the volume of cerebral infarction;ELISA was per⁃formed to detect the levels of interleukin-6(IL-6),interleukin-1β(IL-1β),brain-derived neurotrophic factor(BDNF)and interferon-γ(IFN-γ)in serum and penumbra.RESULTS TTC staining results showed that there was no infarction in sham group.Compared with tMCAO group,the infarct volume in each administration group was signifi⁃cantly decreased(P<0.01).ELISA results showed that IL-6,IL-1βand IFN-γin serum and penumbra were of significant difference between tMCAO group and sham group(P<0.01),and BDNF was significantly decreased(P<0.01).Compared with tMCAO group,IL-6,IL-1βand IFN-γin serum and ischemic penumbra were sig⁃nificantly decreased in all administration groups(P<0.01),while the content of BDNF was in⁃creased in CH 0.2 g·kg-1 group and edaravone 0.008 g·kg-1 group(P<0.05),and other groups were significantly increased(P<0.01).CONCLU⁃SION CH could reduce the cerebral infarction vol⁃ume and improve the nerve injury caused by cerebral ischemia-reperfusion.The mechanism was related to inhibit the expression of IL-6,IL-1βand IFN-γand increase the expression of BDNF possibly.

Protective Effect of GRK2 and Effect of Sanguis Draconis Flavones on Focal Cerebral Ischemia-Reperfusion Injury in Rats

[Objectives] To explore the protective effect of Sanguis Draconis flavones (SDF) on rat focal cerebral ischemia-reperfusion injury (CIRI) models established by middle cerebral artery occlusion (MCAO).[Methods] A total of 60 healthy adult male Sprague-Dawley rats were selected. They were evenly and randomly divided into sham group, model group, edaravone group (12 mg/kg) and SDF group (360 mg/kg), and administered intragastrically and intraperitoneally. The middle cerebral artery of each rat was blocked by suture-occluded method to establish a CIRI model. After ischemia for 2 h and reperfusion for 48 h, the pathological injury on the ischemic side was observed by HE staining;the neuron and myelin sheath structure was observed by transmission electron microscopy;the expression of G protein-coupled receptor kinase 2 (GRK2) was preserved by immunohistochemistry;and the transfer of GRK2 was detected by western-blot.[Results] After 48 h of CIRI, the nuclei of the penumbral cortical neurons shrank, the chromatin was unevenly distributed, the nuclear membrane was dissolved and the mitochondria in the cytoplasm were swollen and vacuolated. The myelin layer was disordered. With this change, the distribution of GRK2 subcellular cells in the penumbra of the injured lateral cortex transferred from the cytoplasm to the membrane. SDF can effectively restore neuronal and myelin sheath structural damage and reduce the functional (membrane coupling) expression of GRK2.[Conclusions] GRK2 may be an effective target for SDF to protect the impaired blood-brain barrier (BBB) in CIRI.

Protective effect of ultrashortwave versus radix salviae miltiorrhizae on brains of rats with cerebral ischemia-reperfusion injury

BACKGROUND： HOW to control the effect of oxygen-derived free radicals on development of cerebral injury and cerebral edema is a key factor for treating cerebral ischemia-reperfusion injury. OBJECTIVE： To observe and compare the protective effects, synergistic action and mechanisms of ultrashortwave （USW） and radix salviae miltiorrhizae （RSM） on the focal cerebral ischemia-reperfusion injuries in rats. DESIGN： Randomized controlled animal study SEI-FING： Department of Rehabilitation Medicine, First Hospital affiliated to China Medical University MATERIALS： A total of 160 healthy Wistar rats of both genders and aged 18-20 weeks weighing 250-300 g of clean grade were selected in this study. 5 mL/ampoule RSM injection fluid was produced by the First Pharmaceutical Corporation of Shanghai （batch number： 011019, 0.01 mug）. The USW therapeutic device was produced by Shanghai Electronic Device Factory with the frequency of 40.68 MHz and the maximal export power of 40 W. The first channel of power after modulation was 11 W. METHODS： The experiment was carried out in the Rehabilitation Medicine Department of the First Hospital affiliated to China Medical University from May 2002 to January 2003. Focal ischemia-reperfusion model was established in rats by reversible right middle cerebral artery occlusion with filament. Right cerebral ischemia was for 2 hours and then with 24 hours reperfusion. The scores of neurological deficits were evaluated by 0 to 4 scales. After surgery, 64 successful rats models were divided into four groups according to digital table： control group, USW group, RSM group and RSM ＋ USW group with 16 cases in each group. Rats in control group were intraperitoneally injected with the same volume of saline （0.1 mL/g）; rats in USW group were given small dosage of USW on head for 10 minutes at 6 hours after reperfusion; rats in RSM group were intraperitoneally injected with 0.01 mL/g RSM solution at 30 minutes before reperfusion; rats in RSM ＋ USW group were intraperitoneally injected with 0.01 mL/g RSM parenteral solution at 30 minutes before reperfusion and given small dosage of USW on head for 10 minutes once at 6 hours after reperfusion; sixteen rats in sham operation group did not receive any treatment. All 80 rats were taken brains at 24 hours after reperfusion to measure wet and dry weights to calculate water content： Cerebral water content （%） = （1-dry/wet weight） × 100%. Superoxide dismutase （SOD） activity was measured by hydroxylamine method and malondialdehyde （MDA） content was measured by TBA photometric method. MAIN OUTCOME MEASURES ： Cerebral water content, SOD activity and MDA content RESULTS： All 160 rats except 80 failing in modeling were involved in the final analysis. （① The cerebral water content of left hemisphere made no significant difference （P 〉 0.05）. The cerebral water content of right hemisphere in the control group and the three treatment groups was obviously higher than that of the sham operation group [（81.26±0.77）%, （79.74±0.68）%, （79.76±0.81）%, （79.61±0.79）%, （77.43±0.61）%, P 〈 0.05]. The cerebral water content of right hemisphere in the three treatment groups was obviously lower than that of the control group （P〈 0.05）. There was no significant difference among the three treatment groups （P 〉 0.05）. ② Compared with the control group, SOD activity （right） of the control group decreased obviously （P 〈 0.05）, while MDA content increased obviously （P 〈 0.05）. SOD activity in the three therapeutic groups increased obviously, while MDA content decreased obviously （P 〈 0.05）; there was no significant difference among the three treatment groups （P 〉 0.05）. CONCLUSION： ① USW and RSM therapy have neuroprotective effects against focal cerebral ischemia-reperfusion injuries by means of decreasing cerebral water content and MDA and increasing the activity of SOD. ② Synergistic action was not observed between these two therapeutic methods.

Inhibition of the cGAS–STING pathway:contributing to the treatment of cerebral ischemia-reperfusion injury

The cGAS–STING pathway plays an important role in ischemia-reperfusion injury in the heart,liver,brain,and kidney,but its role and mechanisms in cerebral ischemia-reperfusion injury have not been systematically reviewed.Here,we outline the components of the cGAS–STING pathway and then analyze its role in autophagy,ferroptosis,cellular pyroptosis,disequilibrium of calcium homeostasis,inflammatory responses,disruption of the blood–brain barrier,microglia transformation,and complement system activation following cerebral ischemia-reperfusion injury.We further analyze the value of cGAS–STING pathway inhibitors in the treatment of cerebral ischemia-reperfusion injury and conclude that the pathway can regulate cerebral ischemia-reperfusion injury through multiple mechanisms.Inhibition of the cGAS–STING pathway may be helpful in the treatment of cerebral ischemia-reperfusion injury.

Silencing miRNA-324-3p protects against cerebral ischemic injury via regulation of the GATA2/A1R axis

Previous studies have suggested that miR-324-3p is related to the pathophysiology of cerebral ischemia,but the mechanism underlying this relationship is unclea r.In this study,we found that miR-324-3p expression was decreased in patients with acute ischemic stroke and in in vitro and in vivo models of ischemic stro ke.miR-324-3p agomir potentiated ischemic brain damage in rats subjected to middle cerebral artery occlusion,as indicated by increased infarct volumes and cell apoptosis rates and greater neurological deficits.In a PC12 cell oxygen-glucose deprivation/reoxygenation model,a miR-324-3 p mimic decreased cell viability and expression of the anti-apoptotic protein BCL2 and increased expression of the pro-apoptotic protein BAX and rates of cell apoptosis,whereas treatment with a miR-324-3p inhibitor had the opposite effects.Silencing miR-324-3p increased adenosine A1 receptor(A1R)expression thro ugh regulation of GATA binding protein 2(GATA2).These findings suggest that silencing miR-324-3p reduces ischemic brain damage via the GATA2/A1R axis.

The effect of hyperglycemia on blood brain barrier of rats with focal cerebral ischemia/reperfusion injury

Objective： To determine whether hyperglycemia could aggravate the microvascular damage in ischemic stroke. Methods： Hyperglycemia model was made by injection of streptozocin through subcutaneous injection in wistar rats. Using the suture model, the rats were subjected to 3h of focal ischemia and different times of reperfusion, including 6,12,24,48,96h and 7d. TIC dyeing was used to Show the infarction area of rats. The infarctive volume of rats were calculated by computer imaging analysis system;Matrix metalloproteinase （MMP-2） and （MMP-9）were detected by immunohistochemistly and in situ hybridization histochemistly in Wistar rats. Results： The infarctive volume was siginificantly larger in hyperglycemic rats than that of nonhyperglycemic rats. The level of MMP-2, MMP-9 expression in the group of hyperglycemic rats was higher than that of nonhyperglycemic rats. Conclusion： Hyperglycemia aggravated the injury of focal ischmia-repeffusion in wistar rats and the higher expression of MMP-2,MMP-9 might be one of the mechanism in aggravation of focal ischemia/repeffusion injury.

Network Pharmacology-Based and Pharmacological Evaluation of the Effects of Curcumae Radixon Cerebral Ischemia-Reperfusion Injury

Objective: This study aimed to investigate the network pharmacology of curcumae radix(CR, Yujin) and explore the mechanism of CR in the treatment of cerebral ischemia-reperfusion injury(CIRI). Materials and Methods: Network analysis and pharmacological evaluation were performed to explore the protective role of CR to treat CIRI. The potential target genes of the active components and CIRI were identified using SwissTarget Prediction, Bioinformatics Analysis Tool for Molecular mechANism of Traditional Chinese Medicine, GeneCards, and Online Mendelian Inheritance in Man. Furthermore, network analysis was performed using Cytoscape software.Gene ontology analysis and Kyoto Encyclopedia of Genes and Genomes enrichment analysis were performed using the R software. In vivo experiments were performed using the water extract of CR(WECR) on PC12 cells induced by hypoxia/reoxygenation(H/R) to simulate ischemia/reperfusion injury. Results: The results exhibited that 21 active compounds identified in CR were associated with 73 targets of CIRI. Functional analysis showed that multiple pathways, including response to stress, regulation of apoptotic process, and hypoxia-inducible factor 1 signaling pathway, were significantly enriched. In addition, STAT3, IL4, HIFIA, and CTNNB1 were predicted to be the most important genes among the 36 hub genes. Furthermore, WECR treatment significantly improved PC12 cell injury and decreased apoptosis levels in cells induced by H/R, with malondialdehyde contents reduced and superoxide dismutase or glutathione peroxidase levels increased. Conclusions: Network analysis and pharmacological evaluation of CR could provide valuable directions for further research on CR and improve comprehension of CIRI.

Mechanism Research of Electroacupuncture Stimulation at Baihui and Zusanli in Cerebral Ischemia-reperfusion Injury Using RNA-Sequencing

Background:Electroacupuncture(EA)therapy,as a combination of electric stimulation and traditional acupunc-ture technology,is currently an important mean for treatment of ischemic stroke and post-stroke rehabilitation in China.Because of its remarkable therapeutic effect,it has been widely used in hospitals and clinic,however,the detailed mechanism remains unclear.Objective:This research aims to comprehensively and systematically elucidate the mechanisms of EA treatment at the acupoints of Zusanli(ST36)and Baihui(GV20)on ischemic stroke.Methods:In this study,EA was performed twice at onset of reperfusion and 20 h after reperfusion following cerebral ischemia-reperfusion(I/R)in middle cerebral artery occlusion(MCAO)rats,and transcriptomic changes of various molecules in ischemic hippocampal neurons of rats in Sham,I/R and EA groups were detected by RNA-Sequencing(RNA-Seq).Results:Thus,we detected 18 significantly different genes related to atherosclerosis(AS),with their functions associated with lipid metabolism,thrombosis,monocytes and vascular smooth muscle cells.And,we detected 10 significantly different genes related to oxidative stress and apoptosis and 10 significantly different genes related to calcium overload and excitatory amino acids release.Also,we detected 19 significantly different genes related to blood-brain barrier(BBB)and 22 significantly different genes related to inflammatory response.Conclusion:In conclusion,EA can play a role in treating ischemic stroke through a variety of mechanisms,affecting atherosclerosis,oxidative stress,apoptosis,calcium overload,excitatory amino acids release,blood-brain barrier(BBB)and inflammatory response.

Neuroprotective effect of ischemic preconditioning in focal cerebral infarction: relationship with upregulation of vascular endothelial growth factor

Neuroprotection by ischemic preconditioning has been confirmed by many studies, but the precise mechanism remains unclear. In the present study, we performed cerebral ischemic pre- conditioning in rats by simulating a transient ischemic attack twice （each a 20-minute occlusion of the middle cerebral artery） before inducing focal cerebral infarction （2 hour occlusion-reper- fusion in the same artery）. We also explored the mechanism underlying the neuroprotective effect of ischemic preconditioning. Seven days after ocdusion-reperfusion, tetrazolium chloride staining and immunohistochemistry revealed that the infarct volume was significantly smaller in the group that underwent preconditioning than in the model group. Furthermore, vascular endothelial growth factor immunoreactivity was considerably greater in the hippocampal CA3 region of preconditioned rats than model rats. Our results suggest that the protective effects of ischemic preconditioning on focal cerebral infarction are associated with upregulation of vascu- lar endothelial growth factor.

Effects of repetitive transcranial magnetic stimulation on adenosine triphosphate content and microtubule associated protein-2 expression after cerebral ischemia-reperfusion injury in rat brain

Background Repetitive transcranial magnetic stimulation （rTMS） research has mainly been focused on the therapeutic effect of psychiatric disorders and Parkinson＇s disease. A few studies have shown that rTMS might protect against delayed neuronal death induced by transient ischemia, enhance long-term potentiation in ischemic conditions and affect regional brain blood flow and metabolism. The aim of this study was to determine the effects of repetitive transcranial magnetic stimulation （rTMS） on adenosine triphosphate （ATP） content and microtubule associated protein-2 （MAP-2） expression in rat brain after middle cerebral artery occlusion （MCAO）/reperfusion. Methods To study the effects of different timecourses of rTMS on ATP content and MAP-2 expression, 90 rats were randomly divided into three groups （30 rats in each group）. To study the effects of multiple rTMS parameters on ATP content and MAP-2 expression, the rats in each group were further divided into six subgroups （five rats each）. The rats were sacrificed at 1-hour, 24-hour and 48-hour intervals after reperfusion, and the brain tissues were collected for the detection of ATP and MAP-2. Results rTMS could significantly increase ATP content and MAP-2 expression in the left brain following ischemic insult （P 〈0.01） and different rTMS parameters had different effects on the ATP level and the MAP-2 expression in the left striatum. A high-frequency rTMS played an important role in MAP-2 expression and ATP preservation. Conclusions This study revealed that rTMS induced significant increase of ATP content and MAP-2 expression in the injured area of the brain, suggesting that the regulation of both ATP and MAP-2 may be involved in the biological mechanism of the effect of rTMS on neural recovery. Therefore, rTMS may become a potential adjunctive therapy for ischemic cerebrovascular disease.