Scutellarin, a bioactive flavone isolated from Scutellaria baicalensis, has anti-inflammatory, anti-neurotoxic, anti-apoptotic and anti-oxida- tive effects and has been used to treat cardiovascular and cerebrovascular...Scutellarin, a bioactive flavone isolated from Scutellaria baicalensis, has anti-inflammatory, anti-neurotoxic, anti-apoptotic and anti-oxida- tive effects and has been used to treat cardiovascular and cerebrovascular diseases in China. However, the mechanisms by which scutellarin mediates neuroprotection in cerebral ischemia remain unclear. The interaction between scutellarin and nicotinamide adenine dinucleotide phosphate oxidase 2 （NOX2） was assessed by molecular docking study, which showed that scutellarin selectively binds to NOX2 with high affinity. Cultures of primary astrocytes isolated from the cerebral cortex of neonatal Sprague-Dawley rats were pretreated with 2, 10 or 50 μM scutellarin for 30 minutes. The astrocytes were then subjected to oxygen/glucose deprivation by incubation for 2 hours in glucose-free Dulbecco＇s modified Eagle＇s medium in a 95% N2/5% CO2 incubator, followed by simulated reperfusion for 22 hours. Cell viability was assessed by cell counting kit-8 assay. Expression levels of NOX2, connexin 43 and caspase-3 were assessed by western blot assay. Reactive oxygen species were measured spectrophotometrically. Pretreatment with 10 or 50 μM scutellarin substantially increased viability, reduced the expression of NOX2 and caspase-3, increased the expression of connexin 43, and diminished the levels of reactive oxygen, species in astrocytes subjected to ischemia-＇reperfusion. We also assessed the effects of scutellarin in vivo in the rat transient middle cerebral artery occlusion model of cerebral ischemia-reperfusion injury. Rats were given intraperitoneal injection of 100 mg/kg scutellarin 2 hours before surgery. The Bederson scale was used to assess neurological deficit, and 2,3,5-triphenyltetrazolium chloride staining was used to measure infarct size. Western blot assay was used to assess expression of NOX2 and connexin 43 in brain tissue. Enzyme-linked immunosorbent assay was used to detect 8-hydroxydeoxyguanosine （8-OHdG）, 4-hydroxy-2-nonenal （4-HNE） and 3-nitrotyrosin （3-NT） in brain tissue. Immunofluorescence double staining was used to determine the co-expression of caspase-3 and NeuN. Pretreatment with scutellarin im- proved the neurological function of rats with focal cerebral ischemia, reduced infarct size, diminished the expression of NOX2, reduced levels of 8-OHdG, 4-HNE and 3-NT, and reduced the number of cells co-expressing caspase-3 and NeuN in the injured brain tissue. Furthermore, we examined the effect of the NOX2 inhibitor apocynin. Apocynin substantially increased connexin 43 expression in vivo and in vitro. Collectively, our findings suggest that scutellarin protects against ischemic injury in vitro and in vivo by downregulating NOX2, upregulating connexin 43, decreasing oxidative damage, and reducing apoptotic cell death.展开更多
MicroRNAs are a family of small, genome-encoded endogenous RNAs that are transcribed but are not translated into proteins. They serve essential roles in virtually every aspect of brain function, including neurogenesis...MicroRNAs are a family of small, genome-encoded endogenous RNAs that are transcribed but are not translated into proteins. They serve essential roles in virtually every aspect of brain function, including neurogenesis, neural development, and cellular responses leading to changes in synaptic plasticity. They are also implicated in neurodegeneration and neurological disorders, in responses to hypoxia and ischemia, and in ischemic tolerance induced by ischemic preconditioning. In recent developments, miRNA expres- sion profiling has been examined in stroke, and these studies indicate that miRNAs have emerged as key mediators in ischemic stroke biology. Both increased and decreased miRNA levels may be needed either as prevention or treatment of stroke. Novel approaches are being developed to get miRNA related therapeu- tics into the brain across an intact blood-brain barrier, including chemical modification, use of targeting molecules and methods to disrupt the blood-brain barrier.展开更多
Cardiovascular disease is the leading cause of death worldwide.Stroke is the second leading cause of death above the age of 60 years and the leading cause of acquired disability in adults.The main type of stroke is is...Cardiovascular disease is the leading cause of death worldwide.Stroke is the second leading cause of death above the age of 60 years and the leading cause of acquired disability in adults.The main type of stroke is ischemic stroke（80%）and it is subclassified as thrombotic or embolic in nature.展开更多
BACKGROUND: Activation of N-methyl-D-aspartate receptor (NMDAR) is a key link of exitotoxicity at the phase of cerebral ischemic injury. Because NMDAR is a main way to mediate internal flow of Ca2+ among glutamic acid...BACKGROUND: Activation of N-methyl-D-aspartate receptor (NMDAR) is a key link of exitotoxicity at the phase of cerebral ischemic injury. Because NMDAR is a main way to mediate internal flow of Ca2+ among glutamic acid receptors, over-excitation can cause neuronal apoptosis. Calcitonin gene related peptide has a strongly biological activity. On one hand, it can protect ischemic neurons through inhibiting the expression of NMDAR1 mRNA; on the other hand, it can play the protective effect through down-regulating the expression of NMDAR1 mRNA by exogenous calcitonin gene related peptide. OBJECTIVE: To observe the expression of NMDAR1 and the regulatory effect of calcitonin gene related peptide on the expression of NMDAR1 mRNA and protein in the cerebral cortex of rats with focal cerebral ischemia/reperfusion (I/R). DESIGN: Randomized controlled animal study. SETTING: China Medical University. MATERIALS: A total of 216 healthy male Wistar rats, general grade, weighing 250-280 g, were selected in this study. Twelve rats were randomly selected to regard as control group; meanwhile, other 204 rats were used to establish middle cerebral artery occlusion/reperfusion (MACO) models. The main reagents were detailed as follows: calcitonin gene related peptide (Sigma Company); calcitonin gene related peptide kit (Boster Company); antibody Ⅰ, Ⅱ and antibody β-actin Ⅰ, Ⅱ of NMDAR1 mRNA and chemiluminescence reagent (Santa Cruz Company, USA). METHODS: The experiment was carried out in the Laboratory of Neurobiology of China Medical University from August 2005 to June 2006. ① Right MCAO models of rats were established to cause focal ischemia and scored based on Zea Longa five-grade scale. If the scores were 1, 2 and 3 after wakefulness, the MACO models were established successfully and involved in the experiment. A total of 120 rats with successful modeling were randomly divided into I/R group and administration group with 60 in each group. All rats in the both groups were observed at five time points, including 6, 12, 24, 48 and 72 hours after reperfusion and after 2-hour ischemia, with 12 experimental animals at each time point. Six rats were prepared for detection of hybridization in situ, and the other 6 were used for Western blotting histochemical detection. Rats in the control group were opened their skin to separate common carotid artery and not treated with line and drugs. In addition, rats in the I/R group were treated with 1 mL saline at 2 hours after focal cerebral ischemia, and then, rats in the administration group were treated with 1 mL (1 g/L) calcitonin gene related peptide at 2 hours after focal cerebral ischemia. ② The expression of NMDAR1 mRNA was detected with hybridization in situ at various time points; moreover, the expression of NMDAR1 protein was measured with Western blotting method at various time points. The results were analyzed with Metamoph imaging analytical system. MAIN OUTCOME MEASURES: The expression of NMDAR1 mRNA and its protein in cortical neurons of rats at various time points. RESULTS: A total of 84 rats were excluded because of non-symptoms, exanimation or death; and then, 132 rats were involved in the final analysis. The expression of NMDAR1 mRNA and its protein in cortical neurons of rats in the control group was 0.205±0.001 and 0.184±0.001, respectively; after I/R, expression of NMDAR1 mRNA and its protein was up-regulated, especially, expression of mRNA at 6, 12, 24, 48 and 72 hours was 0.245±0.003, 0.287±0.004, 0.354±0.008, 0.284±0.002 and 0.217±0.006, respectively; moreover, expression of protein at 6, 12, 24, 48 and 72 hours was 0.222±0.003, 0.261±0.028, 0.311±0.004, 0.259±0.013 and 0.210±0.008, respectively. There was significant difference between the two groups (0.205±0.001, P < 0.01). The expression was up-related in the former 24 hours, reached peak at 24 hours, down-regulated, and decreased to the level of control group at 72 hours. Except 72 hours, the expression of NMDAR1 mRNA and its protein was lower in administration group than that in I/R group at other four time points. In addition, the expression of mRNA at 6, 12, 24, 48 and 72 hours was 0.223±0.005, 0.243±0.001, 0.292±0.002, 0.250±0.003 and 0.213±0.003, respectively; moreover, the expression of protein at 6, 12, 24, 48 and 72 hours was 0.216±0.006, 0.245±0.025, 0.276±0.003, 0.241±0.045 and 0.202±0.013, respectively. There was significant difference at various time points (P < 0.05). CONCLUSION: The expressions of NMDAR1 mRNA and its protein of peripheral cortical neurons are up-related in ischemic area after focal cerebral I/R. Meanwhile, exogenous calcitonin gene related peptide can protect cortical neurons through inhibiting expression of NMDAR1 mRNA and its protein after focal cerebral I/R.展开更多
BACKGROUND： Previous researches have proved that aminoguanidin can cure cerebral ischemic injury remarkably as a selective induced nitricoxide synthase （iNOS） inhibitor. However, whether nonselective NOS inhibitor ...BACKGROUND： Previous researches have proved that aminoguanidin can cure cerebral ischemic injury remarkably as a selective induced nitricoxide synthase （iNOS） inhibitor. However, whether nonselective NOS inhibitor could protect cerebral ischemic injury or not is unclear. OBJECTIVE： To investigate the effects of NG-nitro-L-arginine （L-NA）, a nonselective nitricoxide synthase （NOS） inhibitor, on cerebral ischemic injury of rats and the possible mechanism.DESIGN： Randomized controlled study.SETTING ： Pharmacological Department of Medical Academy of Science of Hebei Province.MATERIALS： A total of 56 male healthy SD rats, of grade Ⅱ, weighting 250-290 g, were provided by the Experimental Animal Center of Hebei Province （certification： 04036）. METHODS： The experiment was completed in the Pharmacological Department of Medical Academy of Science of Hebei Province from March 2005 to January 2006.① Grouping： Rats were randomly divided into 3 groups： sham operation group （n=8）, model group （n=24） and L-NA group （n=24）.② Modeling： Middle cerebral artery （MCA） was established on rats in model group and L-NA group with intreluminal line occlusion methods, and rats in sham operation group were separated their external carotid arteries without occlusion of internal carotid artery. ③ Intervention study： Rats in model group and L-NA group were injected intreperitoneally with 10 mL/kg and 20 mg/kg L-NA at 2, 6 and 12 hours respectively after ischemia twice a day for 3 consecutive days. ④ Rats were sacrificed on the third day for measuring volume of cerebral infarction with image analysis and swelling degrees and activities of mitochondria with electron microscope. Effect of L-NA on ultrastructural changes of neurons in cortex was observed after ischemia. MAIN OUTCOME MEASURES：① Volume of cerebral infarction; ②Swelling degrees, contents of nitric oxide （NO） and malondialdehyde （MDA） and activities of adenosine triphosphatase （ATPase）, superoxide dismutase （SOD） and glutathione peroxidase （GSH-Px） in mitochondria;③ Ultrastructural changes of mitochondria in brain tissue after cerebral ischemia. RESULTS： ① At 12 hour after ischemia, volume of cerebral infarction in L-NA group was lower than that in model group （P 〈 0.01）. ② Content of NO in mitochondria in L-NA group was decreased as compared with that in model group at 2, 6 and 12 hours after ischemia （P 〈 0.05）; swelling degree of mitochondria in brain tissue was relieved in L-NA group at 12 hour after ischemia, and content of MDA was decreased （P 〈 0.05）; mitochondrial activity in L-NA group was increased at 12 hour after ischemia, and activities of ATPase, SOD and GSH-Px in mitochondria were increased （P 〈 0.05）.③ Degrees of mitochondrial injury in brain tissue were relieved in L-NA group at 12 hour after ischemia as compared with those in model group and L-NA group at 2 and 6 hours after ischemia. CONCLUSION ： ①L-NA can beneficially inhibit NO production, but not protect brain against damage in ischemia acute stage. ②L-NA might have protective effects on cerebral injury through inhibiting the production of oxygen free radical, increasing antioxidation, ameliorating energy metabolism, beneficially improving the integrity of form and function of mitochondria in brain tissue during postischemia in rats.展开更多
Three articles conceming the molecular biology and proteomics study of the mechanism underlying the effects of acupuncture on the repair of cerebral ischemic injury were published in the Neural Regeneration Research. ...Three articles conceming the molecular biology and proteomics study of the mechanism underlying the effects of acupuncture on the repair of cerebral ischemic injury were published in the Neural Regeneration Research. We hope that our readers find these papers useful to their research.展开更多
BACKGROUND： Diffusion tensor imaging （DTI） is one of the noninvasive methods to study the morphological structure of brain white matter fibrous bands in vivo, and it has been applied primarily in clinic. DTI is ack...BACKGROUND： Diffusion tensor imaging （DTI） is one of the noninvasive methods to study the morphological structure of brain white matter fibrous bands in vivo, and it has been applied primarily in clinic. DTI is acknowledged as the more effective imaging method to diagnose ultra-acute and/or acute cerebral infarction.OB_3ECTIVE： To observe the anisotropic characters of cerebral white matter fibrous bands in patients with ischemic stroke by using DTI, and investigate the correlation between the damage of corticospinal tract and muscle strength in patients with ischemic stroke at acute period.DESIGN： A case-control observationSEFIING： Department of Medical Imaging, Fuzhou General Hospital of Nanjing Military Area Command of Chinese PLA.PARTICIPANTS： Nine inpatients with injury of motor function induced by acute ischemic stroke （patient group） at 6 hours to 2 weeks after the attack were selected from the Department of Neurology, Fuzhou General Hospital of Nanjing Military Area Command of Chinese PLA from September 2005 to March 2006, and they all accorded with the present diagnostic standard for cerebrovascular disease in China. There were 5 males and 4 females, aged 16-87 years. At the same time, nine healthy right-handed physical examinees matched by age and sex with the patients were taken as the control group, and they all had no nervous disease, mental diseases, cerebrovascular abnormalities and injury history, etc. All the subjects were informed with the detected items and agreed to participate in the study.METHODS： All the 9 patients with ischemic stroke at acute period and 9 healthy subjects were examined with MRI, T1 weighted imaging, T2 weighted imaging and DTI. And the data were processed offline with dTV.II software, the images of fractional anisotropy and directional encoded color （DEC） were obtained, and the three-dimensional fibrous band images of bilateral corticospinal tracts were reconstructed. In the control group, the values of fractional anisotropy of main white matter fibrous bands were measured in the region of interest （ROI） of the anterior limb, knee and posterior limb of internal capsule. In the patient group, the values of fractional anisotropy of white matter were measured in the infarcted sites and corresponding contralateral sites of the patients. The ROI was set in bilateral cerebral peduncles to reconstruct three-dimensionally the bilateral corticospinal tracts. The muscle strength of the affected hand was assessed with Brunnstorm standard in the stroke patients.MAIN OUTCOME MEASURES ： The characters of DTI and images of the value of fractional anisotropy, and the manifestations of three-dimensional corticospinal tracts were observed in the two groups.RESULTS： All the data from the 9 patients and 9 healthy volunteers were involved in the analysis of results. In the control group, the white matter and gray matter could be distinguished clearly in the image of fractional anisotropic values, the fibers of different directions were shown by different colors in DEC picture, which clearly demonstrated the normal anatomic structure and direction of white matter fibers. In the patient group, the infarctions occurred in the gray matter or white matter could be distinguished in the images of fractional anisotropic values, DEC picture could clearly show the direct influence of the infarcted site on the white matter fibers. The fractional anisotropic values in different white matter structure of the same side were significantly different in the control group （t=-3.12, P 〈 0.05）, and the reconstructed images fractional anisotropic values and DEC picture could show most of the main white matter fibrous bands. The fractional anisotropic values of the infarcted sites were significantly lower than the contralateral ones in the patient group （t=-5.570, P 〈 0.01）. ② The reconstructed bilateral corticospinal tracts showed that the anatomic forms of the contralateral corticospinal tract of the patients were almost identical to those of normal people, it started from precentral gyrus, downward to the nternal capsule, and extended to pontine and medulla oblongata, each fibrous band was continuous, and the form had good consistency. Because of the involvement of infarction of different severity, the ipsilateral corticospinal tract manifested as continuous interruption and the loss of consistent anatomic structural form. The involved severity of corticospinal tract had significant correlation with that of muscle strength of the ipsilateral hand （r=-1.30, P 〈 0.01）.CONCLUSION： ① DTI can display the direction and distribution of cerebral white matter fibrous bands.② DTI images of fractional anisotropic values and DEC can show the directions and anisotropic degree of white matter fibers in the infarcted sites of stroke patients. ③ The three-dimensional images of fibrous bands can show the conditions of pyramidal tracts more directly. ④ The damaged severity of corticospinal tracts is correlated with that of muscle strength.展开更多
BACKGROUND： Some researches report that He-Ne laser can activate function of erythrocytes and increase content of blood and oxygen via bio-stimulating effect; therefore, it suspects that laser radiation at Baihui and...BACKGROUND： Some researches report that He-Ne laser can activate function of erythrocytes and increase content of blood and oxygen via bio-stimulating effect; therefore, it suspects that laser radiation at Baihui and Dazhui can partially increase blood circulation for oxygen-supplying content of brain and improve functional status of neurons. OBJECTIVE： To verify the effects of laser radiation at Baihui and Dazhui on the expression of Nissl body of brain tissue neurons and brain-derived neurotrophic factor （BDNF） in newborn rats with ischemic/hypoxic cerebral injury. DESIGN： Randomized controlled animal study. SETTING： Department of Neurological Histochemistry, Xianning University. MATERIALS： Forty Wistar rats of 7 - 8 days old, weighing 15 - 20 g and of both genders, were selected from Wuhan Experimental Animal Center. All the rats were randomly divided into sham operation group （n =8）, model group （n =16） and radiation group （n = 16）. The experimental animals were disposed according to ethical criteria. BDNF kit was provided by Wuhan Boster Bioengineering Co., Ltd. METHODS： The experiment was carried out in the Department of Neurological Histochemistry, Xianning University from April 2005 to October 2006. Rats in the radiation group and model group were performed with ligation of left common carotid artery, recovered at room temperature for 1 - 6 days, maintained in self-made hypoxic cabin under normal pressure and injected mixture gas （0.05 volume fraction of 02 and 0.92 volume fraction of N2） for 2 hours. In addition, rats in the sham operation group were treated with separation of left common carotid artery but not ligation and hypoxia. Rats in the model group were not given any treatment; while, rats in the radiation group were exposed with He-Ne laser of 63.28 nm in the wave length at Baihui and Dazhui acupoints on the second day after ischemia-hypoxia. The radiation was given for 10 minutes per day and once a day. Ten days were regarded as a course and the rats were exposed for 2 courses in total. At 20 days after routine breeding, left hemisphere tissues of rats in the three groups were collected for staining of Nissl body and immunohistochemistry of BDNF. MAIN OUTCOME MEASURES： Nissl body staining in left hemisphere tissue and expression of immune positive cells of BDNF. RESULTS： All 40 rats were involved in the final analysis. （1） Nissl body staining： Neuronal cytoplasm of brain tissue was full of blue granule Nissl bodies in the sham operation group; while, Nissl body in neuronal cytoplasm in the model group was stained slightly and had a certain degree of degeneration; meanwhile, there were a lot of blank area in ischemic region. Nissl body in neuron cytoplasm was gradually recovered in the radiation group and relieved as compared with that in the model group. （2） Positive cells of BDNF： Number of immune positive cells of BDNF which were ligated in lateral cerebral hemisphere of rats in the model group was higher than that in the sham operation group （P 〈 0.05）; while, BDNF expression in the radiation group was increased as compared with that in the model group （P 〈 0.05）. CONCLUSION： After laser acupoint radiation, Nissl body is increased and BDNF expression is also increased. This suggests that laser acupoint radiation has neuroprotective effect on brain tissue after ischemia-hypoxia injury.展开更多
Nicotiflorin is a flavonoid extracted from Carthamus tinctorius.Previous studies have shown its cerebral protective effect,but the mechanism is undefined.In this study,we aimed to determine whether nicotiflorin protec...Nicotiflorin is a flavonoid extracted from Carthamus tinctorius.Previous studies have shown its cerebral protective effect,but the mechanism is undefined.In this study,we aimed to determine whether nicotiflorin protects against cerebral ischemia/reperfusion injury-induced apoptosis through the JAK2/STAT3 pathway.The cerebral ischemia/reperfusion injury model was established by middle cerebral artery occlusion/reperfusion.Nicotiflorin（10 mg/kg） was administered by tail vein injection.Cell apoptosis in the ischemic cerebral cortex was examined by hematoxylin-eosin staining and terminal deoxynucleotidyl transferase d UTP nick end labeling assay.Bcl-2 and Bax expression levels in ischemic cerebral cortex were examined by immunohistochemial staining.Additionally,p-JAK2,p-STAT3,Bcl-2,Bax,and caspase-3 levels in ischemic cerebral cortex were examined by western blot assay.Nicotiflorin altered the shape and structure of injured neurons,decreased the number of apoptotic cells,down-regulates expression of p-JAK2,p-STAT3,caspase-3,and Bax,decreased Bax immunoredactivity,and increased Bcl-2 protein expression and immunoreactivity.These results suggest that nicotiflorin protects against cerebral ischemia/reperfusion injury-induced apoptosis via the JAK2/STAT3 pathway.展开更多
Buyanghuanwu decoction has been shown to protect against cerebral ischemia/reperfusion injury,but the underlying mechanisms remain unclear.In this study,rats were intragastrically given Buyanghuanwu decoction,15 m L/k...Buyanghuanwu decoction has been shown to protect against cerebral ischemia/reperfusion injury,but the underlying mechanisms remain unclear.In this study,rats were intragastrically given Buyanghuanwu decoction,15 m L/kg,for 3 days.A rat model of cerebral ischemia/reperfusion injury was established by middle cerebral artery occlusion.In rats administered Buyanghuanwu decoction,infarct volume was reduced,serum vascular endothelial growth factor and integrin αvβ3 levels were increased,and brain tissue vascular endothelial growth factor and CD34 expression levels were increased compared with untreated animals.These effects of Buyanghuanwu decoction were partially suppressed by an angiogenesis inhibitor（administered through the lateral ventricle for 7 consecutive days）.These data suggest that Buyanghuanwu decoction promotes angiogenesis,improves cerebral circulation,and enhances brain tissue repair after cerebral ischemia/reperfusion injury.展开更多
In the central nervous system, the formation of fibrotic scar after injury inhibits axon regeneration and promotes repair. However, the mechanism underlying fibrotic scar formation and regulation remains poorly unders...In the central nervous system, the formation of fibrotic scar after injury inhibits axon regeneration and promotes repair. However, the mechanism underlying fibrotic scar formation and regulation remains poorly understood. M2 macrophages regulate fibrotic scar formation after injury to the heart, lung, kidney, and central nervous system. However, it remains to be clarified whether and how M2 macrophages regulate fibrotic scar formation after cerebral ischemia injury. In this study, we found that, in a rat model of cerebral ischemia induced by middle cerebral artery occlusion/reperfusion, fibrosis and macrophage infiltration were apparent in the ischemic core in the early stage of injury(within 14 days of injury). The number of infiltrated macrophages was positively correlated with fibronectin expression. Depletion of circulating monocyte-derived macrophages attenuated fibrotic scar formation. Interleukin 4(IL4) expression was strongly enhanced in the ischemic cerebral tissues, and IL4-induced M2 macrophage polarization promoted fibrotic scar formation in the ischemic core. In addition, macrophage-conditioned medium directly promoted fibroblast proliferation and the production of extracellular matrix proteins in vitro. Further pharmacological and genetic analyses showed that sonic hedgehog secreted by M2 macrophages promoted fibrogenesis in vitro and in vivo, and that this process was mediated by secretion of the key fibrosis-associated regulatory proteins transforming growth factor beta 1 and matrix metalloproteinase 9. Furthermore, IL4-afforded functional restoration on angiogenesis, cell apoptosis, and infarct volume in the ischemic core of cerebral ischemia rats were markedly impaired by treatment with an sonic hedgehog signaling inhibitor, paralleling the extent of fibrosis. Taken together, our findings show that IL4/sonic hedgehog/transforming growth factor beta 1 signaling targeting macrophages regulates the formation of fibrotic scar and is a potential therapeutic target for ischemic stroke.展开更多
基金financially supported by the National Natural Science Foundation of China,No.81303115,81774042,81771353the Natural Science Foundation of Guangdong Province of China,No.S2013040016915+3 种基金the Science and Technology Program of Guangzhou City of China,No.201508020050,201604020003the Pearl River S&T Nova Program of Guangzhou,the Postdoctoral Foundation of China,No.BBK42913K09,201003345,BBH429151701a grant from the Hong Kong Scholar Program,Guangzhou University of TCM 2017 High Level University Construction Program,No.A1-AFD018171Z11096a grant from the Specialty Program of Guangdong Province Hospital of Traditional Chinese Medicine of China,No.YN2016MJ07,YN2015QN16,YN2015B2025
文摘Scutellarin, a bioactive flavone isolated from Scutellaria baicalensis, has anti-inflammatory, anti-neurotoxic, anti-apoptotic and anti-oxida- tive effects and has been used to treat cardiovascular and cerebrovascular diseases in China. However, the mechanisms by which scutellarin mediates neuroprotection in cerebral ischemia remain unclear. The interaction between scutellarin and nicotinamide adenine dinucleotide phosphate oxidase 2 （NOX2） was assessed by molecular docking study, which showed that scutellarin selectively binds to NOX2 with high affinity. Cultures of primary astrocytes isolated from the cerebral cortex of neonatal Sprague-Dawley rats were pretreated with 2, 10 or 50 μM scutellarin for 30 minutes. The astrocytes were then subjected to oxygen/glucose deprivation by incubation for 2 hours in glucose-free Dulbecco＇s modified Eagle＇s medium in a 95% N2/5% CO2 incubator, followed by simulated reperfusion for 22 hours. Cell viability was assessed by cell counting kit-8 assay. Expression levels of NOX2, connexin 43 and caspase-3 were assessed by western blot assay. Reactive oxygen species were measured spectrophotometrically. Pretreatment with 10 or 50 μM scutellarin substantially increased viability, reduced the expression of NOX2 and caspase-3, increased the expression of connexin 43, and diminished the levels of reactive oxygen, species in astrocytes subjected to ischemia-＇reperfusion. We also assessed the effects of scutellarin in vivo in the rat transient middle cerebral artery occlusion model of cerebral ischemia-reperfusion injury. Rats were given intraperitoneal injection of 100 mg/kg scutellarin 2 hours before surgery. The Bederson scale was used to assess neurological deficit, and 2,3,5-triphenyltetrazolium chloride staining was used to measure infarct size. Western blot assay was used to assess expression of NOX2 and connexin 43 in brain tissue. Enzyme-linked immunosorbent assay was used to detect 8-hydroxydeoxyguanosine （8-OHdG）, 4-hydroxy-2-nonenal （4-HNE） and 3-nitrotyrosin （3-NT） in brain tissue. Immunofluorescence double staining was used to determine the co-expression of caspase-3 and NeuN. Pretreatment with scutellarin im- proved the neurological function of rats with focal cerebral ischemia, reduced infarct size, diminished the expression of NOX2, reduced levels of 8-OHdG, 4-HNE and 3-NT, and reduced the number of cells co-expressing caspase-3 and NeuN in the injured brain tissue. Furthermore, we examined the effect of the NOX2 inhibitor apocynin. Apocynin substantially increased connexin 43 expression in vivo and in vitro. Collectively, our findings suggest that scutellarin protects against ischemic injury in vitro and in vivo by downregulating NOX2, upregulating connexin 43, decreasing oxidative damage, and reducing apoptotic cell death.
文摘MicroRNAs are a family of small, genome-encoded endogenous RNAs that are transcribed but are not translated into proteins. They serve essential roles in virtually every aspect of brain function, including neurogenesis, neural development, and cellular responses leading to changes in synaptic plasticity. They are also implicated in neurodegeneration and neurological disorders, in responses to hypoxia and ischemia, and in ischemic tolerance induced by ischemic preconditioning. In recent developments, miRNA expres- sion profiling has been examined in stroke, and these studies indicate that miRNAs have emerged as key mediators in ischemic stroke biology. Both increased and decreased miRNA levels may be needed either as prevention or treatment of stroke. Novel approaches are being developed to get miRNA related therapeu- tics into the brain across an intact blood-brain barrier, including chemical modification, use of targeting molecules and methods to disrupt the blood-brain barrier.
文摘Cardiovascular disease is the leading cause of death worldwide.Stroke is the second leading cause of death above the age of 60 years and the leading cause of acquired disability in adults.The main type of stroke is ischemic stroke（80%）and it is subclassified as thrombotic or embolic in nature.
文摘BACKGROUND: Activation of N-methyl-D-aspartate receptor (NMDAR) is a key link of exitotoxicity at the phase of cerebral ischemic injury. Because NMDAR is a main way to mediate internal flow of Ca2+ among glutamic acid receptors, over-excitation can cause neuronal apoptosis. Calcitonin gene related peptide has a strongly biological activity. On one hand, it can protect ischemic neurons through inhibiting the expression of NMDAR1 mRNA; on the other hand, it can play the protective effect through down-regulating the expression of NMDAR1 mRNA by exogenous calcitonin gene related peptide. OBJECTIVE: To observe the expression of NMDAR1 and the regulatory effect of calcitonin gene related peptide on the expression of NMDAR1 mRNA and protein in the cerebral cortex of rats with focal cerebral ischemia/reperfusion (I/R). DESIGN: Randomized controlled animal study. SETTING: China Medical University. MATERIALS: A total of 216 healthy male Wistar rats, general grade, weighing 250-280 g, were selected in this study. Twelve rats were randomly selected to regard as control group; meanwhile, other 204 rats were used to establish middle cerebral artery occlusion/reperfusion (MACO) models. The main reagents were detailed as follows: calcitonin gene related peptide (Sigma Company); calcitonin gene related peptide kit (Boster Company); antibody Ⅰ, Ⅱ and antibody β-actin Ⅰ, Ⅱ of NMDAR1 mRNA and chemiluminescence reagent (Santa Cruz Company, USA). METHODS: The experiment was carried out in the Laboratory of Neurobiology of China Medical University from August 2005 to June 2006. ① Right MCAO models of rats were established to cause focal ischemia and scored based on Zea Longa five-grade scale. If the scores were 1, 2 and 3 after wakefulness, the MACO models were established successfully and involved in the experiment. A total of 120 rats with successful modeling were randomly divided into I/R group and administration group with 60 in each group. All rats in the both groups were observed at five time points, including 6, 12, 24, 48 and 72 hours after reperfusion and after 2-hour ischemia, with 12 experimental animals at each time point. Six rats were prepared for detection of hybridization in situ, and the other 6 were used for Western blotting histochemical detection. Rats in the control group were opened their skin to separate common carotid artery and not treated with line and drugs. In addition, rats in the I/R group were treated with 1 mL saline at 2 hours after focal cerebral ischemia, and then, rats in the administration group were treated with 1 mL (1 g/L) calcitonin gene related peptide at 2 hours after focal cerebral ischemia. ② The expression of NMDAR1 mRNA was detected with hybridization in situ at various time points; moreover, the expression of NMDAR1 protein was measured with Western blotting method at various time points. The results were analyzed with Metamoph imaging analytical system. MAIN OUTCOME MEASURES: The expression of NMDAR1 mRNA and its protein in cortical neurons of rats at various time points. RESULTS: A total of 84 rats were excluded because of non-symptoms, exanimation or death; and then, 132 rats were involved in the final analysis. The expression of NMDAR1 mRNA and its protein in cortical neurons of rats in the control group was 0.205±0.001 and 0.184±0.001, respectively; after I/R, expression of NMDAR1 mRNA and its protein was up-regulated, especially, expression of mRNA at 6, 12, 24, 48 and 72 hours was 0.245±0.003, 0.287±0.004, 0.354±0.008, 0.284±0.002 and 0.217±0.006, respectively; moreover, expression of protein at 6, 12, 24, 48 and 72 hours was 0.222±0.003, 0.261±0.028, 0.311±0.004, 0.259±0.013 and 0.210±0.008, respectively. There was significant difference between the two groups (0.205±0.001, P < 0.01). The expression was up-related in the former 24 hours, reached peak at 24 hours, down-regulated, and decreased to the level of control group at 72 hours. Except 72 hours, the expression of NMDAR1 mRNA and its protein was lower in administration group than that in I/R group at other four time points. In addition, the expression of mRNA at 6, 12, 24, 48 and 72 hours was 0.223±0.005, 0.243±0.001, 0.292±0.002, 0.250±0.003 and 0.213±0.003, respectively; moreover, the expression of protein at 6, 12, 24, 48 and 72 hours was 0.216±0.006, 0.245±0.025, 0.276±0.003, 0.241±0.045 and 0.202±0.013, respectively. There was significant difference at various time points (P < 0.05). CONCLUSION: The expressions of NMDAR1 mRNA and its protein of peripheral cortical neurons are up-related in ischemic area after focal cerebral I/R. Meanwhile, exogenous calcitonin gene related peptide can protect cortical neurons through inhibiting expression of NMDAR1 mRNA and its protein after focal cerebral I/R.
基金the Natural Science Foundation of Hebei Province, No. C2005000840
文摘BACKGROUND： Previous researches have proved that aminoguanidin can cure cerebral ischemic injury remarkably as a selective induced nitricoxide synthase （iNOS） inhibitor. However, whether nonselective NOS inhibitor could protect cerebral ischemic injury or not is unclear. OBJECTIVE： To investigate the effects of NG-nitro-L-arginine （L-NA）, a nonselective nitricoxide synthase （NOS） inhibitor, on cerebral ischemic injury of rats and the possible mechanism.DESIGN： Randomized controlled study.SETTING ： Pharmacological Department of Medical Academy of Science of Hebei Province.MATERIALS： A total of 56 male healthy SD rats, of grade Ⅱ, weighting 250-290 g, were provided by the Experimental Animal Center of Hebei Province （certification： 04036）. METHODS： The experiment was completed in the Pharmacological Department of Medical Academy of Science of Hebei Province from March 2005 to January 2006.① Grouping： Rats were randomly divided into 3 groups： sham operation group （n=8）, model group （n=24） and L-NA group （n=24）.② Modeling： Middle cerebral artery （MCA） was established on rats in model group and L-NA group with intreluminal line occlusion methods, and rats in sham operation group were separated their external carotid arteries without occlusion of internal carotid artery. ③ Intervention study： Rats in model group and L-NA group were injected intreperitoneally with 10 mL/kg and 20 mg/kg L-NA at 2, 6 and 12 hours respectively after ischemia twice a day for 3 consecutive days. ④ Rats were sacrificed on the third day for measuring volume of cerebral infarction with image analysis and swelling degrees and activities of mitochondria with electron microscope. Effect of L-NA on ultrastructural changes of neurons in cortex was observed after ischemia. MAIN OUTCOME MEASURES：① Volume of cerebral infarction; ②Swelling degrees, contents of nitric oxide （NO） and malondialdehyde （MDA） and activities of adenosine triphosphatase （ATPase）, superoxide dismutase （SOD） and glutathione peroxidase （GSH-Px） in mitochondria;③ Ultrastructural changes of mitochondria in brain tissue after cerebral ischemia. RESULTS： ① At 12 hour after ischemia, volume of cerebral infarction in L-NA group was lower than that in model group （P 〈 0.01）. ② Content of NO in mitochondria in L-NA group was decreased as compared with that in model group at 2, 6 and 12 hours after ischemia （P 〈 0.05）; swelling degree of mitochondria in brain tissue was relieved in L-NA group at 12 hour after ischemia, and content of MDA was decreased （P 〈 0.05）; mitochondrial activity in L-NA group was increased at 12 hour after ischemia, and activities of ATPase, SOD and GSH-Px in mitochondria were increased （P 〈 0.05）.③ Degrees of mitochondrial injury in brain tissue were relieved in L-NA group at 12 hour after ischemia as compared with those in model group and L-NA group at 2 and 6 hours after ischemia. CONCLUSION ： ①L-NA can beneficially inhibit NO production, but not protect brain against damage in ischemia acute stage. ②L-NA might have protective effects on cerebral injury through inhibiting the production of oxygen free radical, increasing antioxidation, ameliorating energy metabolism, beneficially improving the integrity of form and function of mitochondria in brain tissue during postischemia in rats.
文摘Three articles conceming the molecular biology and proteomics study of the mechanism underlying the effects of acupuncture on the repair of cerebral ischemic injury were published in the Neural Regeneration Research. We hope that our readers find these papers useful to their research.
文摘BACKGROUND： Diffusion tensor imaging （DTI） is one of the noninvasive methods to study the morphological structure of brain white matter fibrous bands in vivo, and it has been applied primarily in clinic. DTI is acknowledged as the more effective imaging method to diagnose ultra-acute and/or acute cerebral infarction.OB_3ECTIVE： To observe the anisotropic characters of cerebral white matter fibrous bands in patients with ischemic stroke by using DTI, and investigate the correlation between the damage of corticospinal tract and muscle strength in patients with ischemic stroke at acute period.DESIGN： A case-control observationSEFIING： Department of Medical Imaging, Fuzhou General Hospital of Nanjing Military Area Command of Chinese PLA.PARTICIPANTS： Nine inpatients with injury of motor function induced by acute ischemic stroke （patient group） at 6 hours to 2 weeks after the attack were selected from the Department of Neurology, Fuzhou General Hospital of Nanjing Military Area Command of Chinese PLA from September 2005 to March 2006, and they all accorded with the present diagnostic standard for cerebrovascular disease in China. There were 5 males and 4 females, aged 16-87 years. At the same time, nine healthy right-handed physical examinees matched by age and sex with the patients were taken as the control group, and they all had no nervous disease, mental diseases, cerebrovascular abnormalities and injury history, etc. All the subjects were informed with the detected items and agreed to participate in the study.METHODS： All the 9 patients with ischemic stroke at acute period and 9 healthy subjects were examined with MRI, T1 weighted imaging, T2 weighted imaging and DTI. And the data were processed offline with dTV.II software, the images of fractional anisotropy and directional encoded color （DEC） were obtained, and the three-dimensional fibrous band images of bilateral corticospinal tracts were reconstructed. In the control group, the values of fractional anisotropy of main white matter fibrous bands were measured in the region of interest （ROI） of the anterior limb, knee and posterior limb of internal capsule. In the patient group, the values of fractional anisotropy of white matter were measured in the infarcted sites and corresponding contralateral sites of the patients. The ROI was set in bilateral cerebral peduncles to reconstruct three-dimensionally the bilateral corticospinal tracts. The muscle strength of the affected hand was assessed with Brunnstorm standard in the stroke patients.MAIN OUTCOME MEASURES ： The characters of DTI and images of the value of fractional anisotropy, and the manifestations of three-dimensional corticospinal tracts were observed in the two groups.RESULTS： All the data from the 9 patients and 9 healthy volunteers were involved in the analysis of results. In the control group, the white matter and gray matter could be distinguished clearly in the image of fractional anisotropic values, the fibers of different directions were shown by different colors in DEC picture, which clearly demonstrated the normal anatomic structure and direction of white matter fibers. In the patient group, the infarctions occurred in the gray matter or white matter could be distinguished in the images of fractional anisotropic values, DEC picture could clearly show the direct influence of the infarcted site on the white matter fibers. The fractional anisotropic values in different white matter structure of the same side were significantly different in the control group （t=-3.12, P 〈 0.05）, and the reconstructed images fractional anisotropic values and DEC picture could show most of the main white matter fibrous bands. The fractional anisotropic values of the infarcted sites were significantly lower than the contralateral ones in the patient group （t=-5.570, P 〈 0.01）. ② The reconstructed bilateral corticospinal tracts showed that the anatomic forms of the contralateral corticospinal tract of the patients were almost identical to those of normal people, it started from precentral gyrus, downward to the nternal capsule, and extended to pontine and medulla oblongata, each fibrous band was continuous, and the form had good consistency. Because of the involvement of infarction of different severity, the ipsilateral corticospinal tract manifested as continuous interruption and the loss of consistent anatomic structural form. The involved severity of corticospinal tract had significant correlation with that of muscle strength of the ipsilateral hand （r=-1.30, P 〈 0.01）.CONCLUSION： ① DTI can display the direction and distribution of cerebral white matter fibrous bands.② DTI images of fractional anisotropic values and DEC can show the directions and anisotropic degree of white matter fibers in the infarcted sites of stroke patients. ③ The three-dimensional images of fibrous bands can show the conditions of pyramidal tracts more directly. ④ The damaged severity of corticospinal tracts is correlated with that of muscle strength.
基金the Scientific and Technological Foundation of Hubei Educational Bureau,No. D200528006
文摘BACKGROUND： Some researches report that He-Ne laser can activate function of erythrocytes and increase content of blood and oxygen via bio-stimulating effect; therefore, it suspects that laser radiation at Baihui and Dazhui can partially increase blood circulation for oxygen-supplying content of brain and improve functional status of neurons. OBJECTIVE： To verify the effects of laser radiation at Baihui and Dazhui on the expression of Nissl body of brain tissue neurons and brain-derived neurotrophic factor （BDNF） in newborn rats with ischemic/hypoxic cerebral injury. DESIGN： Randomized controlled animal study. SETTING： Department of Neurological Histochemistry, Xianning University. MATERIALS： Forty Wistar rats of 7 - 8 days old, weighing 15 - 20 g and of both genders, were selected from Wuhan Experimental Animal Center. All the rats were randomly divided into sham operation group （n =8）, model group （n =16） and radiation group （n = 16）. The experimental animals were disposed according to ethical criteria. BDNF kit was provided by Wuhan Boster Bioengineering Co., Ltd. METHODS： The experiment was carried out in the Department of Neurological Histochemistry, Xianning University from April 2005 to October 2006. Rats in the radiation group and model group were performed with ligation of left common carotid artery, recovered at room temperature for 1 - 6 days, maintained in self-made hypoxic cabin under normal pressure and injected mixture gas （0.05 volume fraction of 02 and 0.92 volume fraction of N2） for 2 hours. In addition, rats in the sham operation group were treated with separation of left common carotid artery but not ligation and hypoxia. Rats in the model group were not given any treatment; while, rats in the radiation group were exposed with He-Ne laser of 63.28 nm in the wave length at Baihui and Dazhui acupoints on the second day after ischemia-hypoxia. The radiation was given for 10 minutes per day and once a day. Ten days were regarded as a course and the rats were exposed for 2 courses in total. At 20 days after routine breeding, left hemisphere tissues of rats in the three groups were collected for staining of Nissl body and immunohistochemistry of BDNF. MAIN OUTCOME MEASURES： Nissl body staining in left hemisphere tissue and expression of immune positive cells of BDNF. RESULTS： All 40 rats were involved in the final analysis. （1） Nissl body staining： Neuronal cytoplasm of brain tissue was full of blue granule Nissl bodies in the sham operation group; while, Nissl body in neuronal cytoplasm in the model group was stained slightly and had a certain degree of degeneration; meanwhile, there were a lot of blank area in ischemic region. Nissl body in neuron cytoplasm was gradually recovered in the radiation group and relieved as compared with that in the model group. （2） Positive cells of BDNF： Number of immune positive cells of BDNF which were ligated in lateral cerebral hemisphere of rats in the model group was higher than that in the sham operation group （P 〈 0.05）; while, BDNF expression in the radiation group was increased as compared with that in the model group （P 〈 0.05）. CONCLUSION： After laser acupoint radiation, Nissl body is increased and BDNF expression is also increased. This suggests that laser acupoint radiation has neuroprotective effect on brain tissue after ischemia-hypoxia injury.
基金financially supported by the Natural Science Foundation of Education Department of Sichuan Province of China,No.14ZB0152the Joint Research Program of Luzhou and Southwest Medical University,in China,No.14JC0120
文摘Nicotiflorin is a flavonoid extracted from Carthamus tinctorius.Previous studies have shown its cerebral protective effect,but the mechanism is undefined.In this study,we aimed to determine whether nicotiflorin protects against cerebral ischemia/reperfusion injury-induced apoptosis through the JAK2/STAT3 pathway.The cerebral ischemia/reperfusion injury model was established by middle cerebral artery occlusion/reperfusion.Nicotiflorin（10 mg/kg） was administered by tail vein injection.Cell apoptosis in the ischemic cerebral cortex was examined by hematoxylin-eosin staining and terminal deoxynucleotidyl transferase d UTP nick end labeling assay.Bcl-2 and Bax expression levels in ischemic cerebral cortex were examined by immunohistochemial staining.Additionally,p-JAK2,p-STAT3,Bcl-2,Bax,and caspase-3 levels in ischemic cerebral cortex were examined by western blot assay.Nicotiflorin altered the shape and structure of injured neurons,decreased the number of apoptotic cells,down-regulates expression of p-JAK2,p-STAT3,caspase-3,and Bax,decreased Bax immunoredactivity,and increased Bcl-2 protein expression and immunoreactivity.These results suggest that nicotiflorin protects against cerebral ischemia/reperfusion injury-induced apoptosis via the JAK2/STAT3 pathway.
基金financially supported by the National Natural Science Foundation of China,No.81072799
文摘Buyanghuanwu decoction has been shown to protect against cerebral ischemia/reperfusion injury,but the underlying mechanisms remain unclear.In this study,rats were intragastrically given Buyanghuanwu decoction,15 m L/kg,for 3 days.A rat model of cerebral ischemia/reperfusion injury was established by middle cerebral artery occlusion.In rats administered Buyanghuanwu decoction,infarct volume was reduced,serum vascular endothelial growth factor and integrin αvβ3 levels were increased,and brain tissue vascular endothelial growth factor and CD34 expression levels were increased compared with untreated animals.These effects of Buyanghuanwu decoction were partially suppressed by an angiogenesis inhibitor（administered through the lateral ventricle for 7 consecutive days）.These data suggest that Buyanghuanwu decoction promotes angiogenesis,improves cerebral circulation,and enhances brain tissue repair after cerebral ischemia/reperfusion injury.
基金supported by the National Natural Science Foundation of China,Nos.82171456 (to QY),81971229 (to QY)the Natural Science Foundation of Chongqing,No.cstc2021jcyj-msxmX0263 (to QY)the Postgraduate Research and Innovation Project of Chongqing,Nos.CYB20151 (to QY),CYS19182 (to YC)。
文摘In the central nervous system, the formation of fibrotic scar after injury inhibits axon regeneration and promotes repair. However, the mechanism underlying fibrotic scar formation and regulation remains poorly understood. M2 macrophages regulate fibrotic scar formation after injury to the heart, lung, kidney, and central nervous system. However, it remains to be clarified whether and how M2 macrophages regulate fibrotic scar formation after cerebral ischemia injury. In this study, we found that, in a rat model of cerebral ischemia induced by middle cerebral artery occlusion/reperfusion, fibrosis and macrophage infiltration were apparent in the ischemic core in the early stage of injury(within 14 days of injury). The number of infiltrated macrophages was positively correlated with fibronectin expression. Depletion of circulating monocyte-derived macrophages attenuated fibrotic scar formation. Interleukin 4(IL4) expression was strongly enhanced in the ischemic cerebral tissues, and IL4-induced M2 macrophage polarization promoted fibrotic scar formation in the ischemic core. In addition, macrophage-conditioned medium directly promoted fibroblast proliferation and the production of extracellular matrix proteins in vitro. Further pharmacological and genetic analyses showed that sonic hedgehog secreted by M2 macrophages promoted fibrogenesis in vitro and in vivo, and that this process was mediated by secretion of the key fibrosis-associated regulatory proteins transforming growth factor beta 1 and matrix metalloproteinase 9. Furthermore, IL4-afforded functional restoration on angiogenesis, cell apoptosis, and infarct volume in the ischemic core of cerebral ischemia rats were markedly impaired by treatment with an sonic hedgehog signaling inhibitor, paralleling the extent of fibrosis. Taken together, our findings show that IL4/sonic hedgehog/transforming growth factor beta 1 signaling targeting macrophages regulates the formation of fibrotic scar and is a potential therapeutic target for ischemic stroke.
