Neuroprotective effect of Smilacis chinae rhizome on NMDA-induced neurotoxicity in vitro and focal cerebral ischemia in vivo

Previous work has shown that the Smilacis chinae rhizome (SCR) markedly inhibits amyloid beta protein (25-35)-induced neuronal cell damage in cultured rat cortical neurons. The present study was conducted to further verify the neuroprotective effect of SCR on excitotoxic and cerebral ischemic injury using both in vitro and in vivo studies. Exposure of cultured cortical neurons to 1 mM N-methyl-D-aspartate (NMDA) for 12 h induced neuronal cell death. SCR (10 and 50 microg/ml) inhibited NMDA-induced neuronal death, elevation of intracellular calcium ([Ca(2+)](i)), and generation of reactive oxygen species (ROS) in primary cultures of rat cortical neurons. In vivo, SCR prevented cerebral ischemic injury induced by 3-h middle cerebral artery occlusion (MCAO) and 24-h reperfusion. The ischemic infarct was significantly reduced in rats that received SCR (30 and 50 mg/kg, orally), with a corresponding improvement in neurological function. Moreover, SCR treatment significantly decreased the histological changes observed following ischemia. Oxyresveratrol and resveratrol isolated from SCR also inhibited NMDA-induced neuronal death, increase in [Ca(2+)](i), and ROS generation in cultured cortical neurons, suggesting that the neuroprotective effect of SCR may be attributable to these compounds. Taken together, these results suggest that the neuroprotective effect of SCR against focal cerebral ischemic injury is due to its anti-excitotoxic effects and that SCR may have a therapeutic role in neurodegenerative diseases such as stroke.     

二维三七桂利嗪胶囊对大鼠局灶性脑缺血再灌注损伤的保护作用

目的:观察二维三七桂利嗪胶囊对大鼠局灶性脑缺血再灌注损伤的影响.方法:SD大鼠分别用292,146和73 mg·kg-1的二维三七桂利嗪胶囊灌胃10 d,线拴法制备大鼠大脑中动脉缺血模型(middle cerebral arteryocclusion,MCAO),缺血3 h后再灌注3 h,分别取脑组织测定脑梗死范围、脑指数、脑含水量并观察脑组织形态结构,取血测定血清乳酸脱氢酶(LDH)和超氧化物歧化酶(S0D)活性.结果:3个剂量的二维三七桂利嗪胶囊能够不同程度地缩小缺血再灌注损伤后脑梗死范围、降低脑指数、脑含水量和血清LDH活性,提高SOD活性并减轻光镜下脑组织缺血性损伤.结论:二维三七桂利嗪胶囊对大鼠局灶性脑缺血再灌注损伤具有保护作用.     

脑心清片防治脑动脉硬化研究进展

脑心清片对脑动脉硬化具有良好的防治效果。近十多年国内外对脑心清片防治脑动脉硬化的临床、药理作用及其物质基础的广泛研究表明,脑心清片含有黄酮、有机酸和香豆素类等化学活性成分,脑心清片及其活性成分能降血压,降血脂,改善血液流变性,抑制血管内皮增生,抑制血小板聚集;调节免疫功能和抑制淋巴细胞与神经细胞的粘附,抑制炎症反应,改善神经细胞氧化还原状态,上调抗氧化基因表达,抗氧化清除自由基,抑制脂质过氧化,减轻缺血脑组织自由基损伤;此外,还能调节钙离子稳态,抗兴奋性谷氨酸毒性,抑制神经细胞凋亡和脑水肿,保护缺血脑组织抗脑缺血损伤;保护神经组织并促进神经细胞功能恢复等多方面、多层次、多靶点作用来阻断脑动脉硬化发生发展,从而发挥良好的防治脑动脉硬化症作用。     

A study of the neuroprotective effect of the phenolic glucoside gastrodin during cerebral ischemia in vivo and in vitro

The phenolic glucoside gastrodin (Gas) is a main component extracted from the rhizome of Gastrodia elata, a Chinese herbal medicine, which has long been used for treating dizziness, epilepsy, stroke and dementia. In this study, we investigated the neuroprotective effects of Gas on cerebral ischemic injury in rats caused by transient middle cerebral arterial occlusion (MCAO), oxygen/glucose deprivation (OGD) and glutamate-induced injury in cultured rat hippocampal neurons. Additionally, the effects of Gas on the extracellular glutamate level and changes in intracellular Ca (2+) and the generation of nitric oxide (NO) were examined in cultured hippocampal neurons subjected to OGD in vitro. The results showed that the high dose of Gas (100 mg/kg) markedly decreased the infarct volume and edema volume, and improved the neurological functions after MCAO. Gas treatment (15 microg/mL, 30 microg/mL) also significantly inhibited OGD- and glutamate-induced neuronal cell death and reduced the extracellular glutamate level following OGD. Moreover, Gas treatment significantly inhibited the OGD-induced Ca (2+) and NO increases. In conclusion, the present study indicates that Gas has a neuroprotective action.     

仙人掌多糖对大鼠局灶性脑缺血的神经保护作用

摘要目的观察仙人掌多糖对大脑中动脉栓塞（MCAO）诱导的大鼠缺血 再灌注损伤的神经保护作用。方法雄性SD大鼠随机分为假手术组、缺血组、0.9%氯化钠溶液组、仙人掌多糖治疗组（200 mg·kg－1·d －1,ip）。采用MCAO法制作大鼠缺血 再灌注损伤模型。分别于大鼠缺血2 h再灌注3,6,8 h后进行神经行为学评分;8 h后2,3,5 三苯基氯化四氮唑（TTC）染色测定脑梗死体积;缺血2 h再灌注22 h后脑组织切片苏木精 伊红（HE）染色显微镜下形态学观察。结果与0.9%氯化钠溶液组相比,再灌注8 h后仙人掌多糖治疗组神经行为学评分平均下降（2.35±0.47）分（P＜0.05）,梗死灶体积减少（P＜0.05）;大鼠皮质及海马组织神经细胞丢失、神经胶质增生、核固缩、核深染等形态学均有明显改善（P＜0.05）。结论仙人掌多糖可以缓解大鼠大脑中动脉栓塞症状,具有一定的神经保护作用。     

丹酚酸B对脑缺血再灌注大鼠神经细胞损伤和神经发生的影响

本文旨在观察丹酚酸B对脑缺血再灌注大鼠神经发生和神经细胞损伤的影响，探讨丹酚酸B促进机体功能恢复的作用环节。研究采用大鼠局灶性脑缺血再灌注模型，治疗给药，用BrdU法观察海马齿状回颗粒下层(sub-granular zone，SGZ)和侧脑室下层(sub-ventricular zone，SVZ)神经发生的变化；尼氏体染色观察神经细胞损伤；平衡杆法观察肢体功能恢复。结果显示，缺血后再灌注7 d，模型组SGZ和SVZ的BrdU细胞明显多于假手术组(P<0.05)，丹酚酸B(10 mg·kg^-1)显著增加SGZ和SVZ的BrdU细胞数目(P<0.01 vs模型组)；缺血再灌注14 d，模型动物缺血侧海马CA1区和皮层神经细胞明显减少，丹酚酸B(10 mg·kg^-1)明显改善神经细胞损伤(P<0.01 vs模型组)；同时，丹酚酸B(10 mg·kg^-1)明显促进缺血动物肢体功能恢复。以上结果表明，丹酚酸B能够增加脑缺血大鼠SVZ和SGZ的BrdU细胞数目，改善缺血区神经细胞损伤，促进肢体功能恢复，提示促进神经发生是丹酚酸B改善脑功能的重要环节。     

蝎毒活性多肽对大鼠脑缺血-再灌注后IL-1B蛋白表达和神经功能的影响

目的:研究蝎毒活性多肽对大鼠脑缺血-再灌注损伤的保护作用,并观察该保护作用对白细胞介素-1B(interleukin- 1B,IL-1B)蛋白表达的影响。方法:采用左侧大脑中动脉插入栓线(Middle cerebral artery occlusion,MCAO)方法制作大鼠脑缺血-再灌注模型,观察蝎毒活性多肽对脑缺血大鼠神经功能的影响;用免疫组化方法标记各组大鼠脑组织海马中IL-1B阳性细胞数量。结果:蝎毒活性多肽可降低大鼠局灶性脑缺血再灌注损伤行为学评分,改善因缺血所致神经行为功能缺失,且随着剂量的增加效应增强,并可以显著减少海马IL-1B阳性细胞的数量(P<0．05)。结论:蝎毒活性多肽对脑缺血损伤有一定的保护作用,且这种作用与其减少脑缺血早期IL-1B表达有关。     

Neuroprotection of total steroid saponins from Dioscorea zingiberensis C.H.Wright against transient focal cerebral ischemia-reperfusion injury in rats via its anti-inflammatory and anti-apoptotic effects

OBJECTIVE The total steroid saponins(TSSN) isolated from Dioscorea zingiberensis C. H. Wright(D.zingiberensis) has shown a variety of beneficial bioactivities. However, there are no reports about the neuroprotective effects of the TSSN until now. Therefore, we explored the neuroprotective effects of TSSN on rats against transient focal cerebral ischemia-reperfusion(I/R) and the underlying mechanisms. METHODS The healthy adult Sprague-Dawley rats were randomly assigned into six groups. After pre-treatment with the TSSN intragastrically for six days, the rats were subjected to the ischemia injury by the surgery of middle cerebral artery occlusion(MCAO) for 90 min. Some indexes were evaluated and detected. RESULTS As compared to the I/R group, TSSN group of rats, especial y given the 30 mg·kg~(-1) of TSSN, not only marked reduction in the neurological deficit scores, cerebral infarct volume, and brain edema, but also an increase in neuron survival(Nissl bodies) in the hippocampal cornuammons 1(CA1) and cortex hemisphere of the ipsilateral ischemia. At the same time, the inflammatory cytokines in serum induced by MCAO were significantly alleviated by the TSSN pre-administration. What′s more, the increase of caspase-3 was evidently reduced in the CA1 and cortex of the hemisphere injured brain. Final y, the down-regulating anti-apoptotic Bcl-2 and up-regulating pro-apoptotic Bax proteins were obviously suppressed. CONCLUSION TSSN plays a potential neuroprotective role against a severe injury induced by transient focal cerebral ischemic reperfusion in a rat experimental model, and this role may be mediated by its antiinflammatory and anti-apoptotic actions.     

复方蒲黄颗粒对局灶性脑缺血再灌注大鼠神经元凋亡及Caspase-3 mRNA表达的影响

目的:观察复方蒲黄颗粒(PH)对局灶性脑缺血再灌注大鼠神经元凋亡的影响,并探讨其作用机制。方法:采用大脑中动脉线栓法建立局灶性脑缺血再灌注大鼠模型。75只大鼠分为假手术、模型和PH高、中、低剂量组。采用Nissl染色观察神经元形态变化,TUNEL法检测神经元凋亡,原位杂交检测Caspase-3mRNA的表达。结果:PH高、中剂量组少量细胞核固缩深染,细胞丢失、凋亡细胞数及Caspase-3mRNA阳性细胞数均显著减少。结论:PH可下调Caspase-3的表达,从而抑制神经元凋亡,对脑缺血再灌注损伤有保护作用。     

芪参胶囊对大鼠局灶性脑缺血再灌注损伤的保护作用

目的研究芪参胶囊对大鼠局灶性脑缺血再灌注损伤的保护作用。方法用线栓法建立大鼠局灶性脑缺血模型,大鼠于缺血1h再灌注2h断头取脑,检测大脑组织Ca2 -ATPase,Na ,K -ATPase,NOS活性和NO,水的含量及大脑皮层神经元内游离钙离子浓度,行为学评价及梗死面积,常规石蜡切片,HE染色,作病理学检查。结果芪参胶囊显著降低缺血再灌注后大脑皮层神经元内游离钙离子浓度及大脑组织NOS活性,NO的含量和水肿程度及梗死面积;显著增强Ca2 -ATPase,Na ,K -ATPase的活性;病理学检查显示芪参胶囊能明显减轻脑水肿及神经元坏死程度。结论芪参胶囊对大鼠局灶性脑缺血再灌注损伤有明显保护作用。     

磷酸肌酸钠预给药对大鼠局灶性脑缺血再灌注损伤的影响

目的:观察不同浓度磷酸肌酸钠预给药对大鼠脑缺血再灌注损伤(I/R)模型脑梗死体积的影响。方法:模型建立采用大脑中动脉线栓法。50只SD大鼠随机分为假手术组(Sham组),I/R组及低、中、高剂量磷酸肌酸钠预给药组(即L、M、H组),每组10只。再灌注末,测血清超氧化物歧化酶(SOD)、丙二醛(MDA)水平,脑组织中Na+-K+-ATP酶及髓过氧化物酶(MPO)活性。实验结束对脑组织进行HE、TTC染色,观察脑组织损伤情况。结果:再灌注24h末,各组间血清SOD、MDA及脑组织中MPO、Na+-K+-ATP酶活性比较差异有统计学意义(P<0.05);Bederson神经功能缺损评分显示Sham组无损伤,其余各组均出现不同程度的神经损伤。脑梗死体积百分比比较,Sham组脑组织无梗死,I/R组脑梗死体积百分比较L、M、H组明显增大,L组大于M组、H组;M组脑梗死体积大于H组,差异均有统计学意义(P<0.05)。结论:低、中、高浓度磷酸肌酸钠预给药均可有效地减少局灶性脑缺血再灌注损伤后大鼠脑梗死体积,减轻脑缺血再灌注后的损伤。     

双苯氟嗪对大鼠全脑缺血再灌注损伤的保护作用

目的研究双苯氟嗪(D ip)对大鼠急性全脑缺血再灌注损伤的保护作用,并初步探讨其作用机制。方法采用Pu lsinelli等的四动脉结扎法(4-VO)造成大鼠全脑缺血再灌注损伤模型,观察大鼠全脑缺血再灌注损伤后早期脑组织水分的变化、生化指标的改变,再灌注后期行为学和组织形态学的改变。结果缺血30 m in再灌注1 h脑组织水分及丙二醛(MDA)含量升高,乳酸脱氢酶(LDH)和超氧化物歧化酶(SOD)活性下降;缺血20 m in再灌注5 d后海马CA1区椎体细胞层破裂,胞核固缩或溶解,间质也变得疏松。行为学实验表明大鼠记忆力明显受损。D ip可不同程度地抑制上述变化,能对抗自由基损伤和脑水肿,并能保护海马CA1区神经元免受缺血损伤,提高大鼠对空间辨别的记忆能力。结论D ip对大鼠全脑缺血再灌注早期损伤有明显的保护作用,并能保护海马CA1区神经元免受缺血损伤,提高大鼠对空间辨别的记忆能力,对迟发性神经元死亡有一定的保护作用。这可能与其抗脂质过氧化产物产生有关。     

异丙酚对大鼠局灶性脑缺血和蛋白激酶C γ亚单位的作用

目的观察异丙酚对大鼠局灶性脑缺血和脑内蛋白激酶Cγ亚单位(PKCγ)变化的作用。方法♂SD大鼠随机分为Ⅰ:假手术组,Ⅱ:损伤组,Ⅲ:异丙酚(25mg·kg-1)+损伤组,Ⅳ:异丙酚(50mg·kg-1)+损伤组,Ⅴ:脂肪乳剂+损伤组,每组8例。采用大脑中动脉线栓法缺血3h再灌注24h。异丙酚和脂肪乳剂于再灌注前30min腹腔注射。通过原位末端脱氧核苷酸转移酶标记法观察局灶性脑缺血产生的神经细胞凋亡和异丙酚作用效果,免疫细胞化学法观察各组PKCγ蛋白在脑内表达变化的差异。结果再灌注24h后Ⅱ、Ⅲ、Ⅳ、Ⅴ组大鼠体重较缺血前降低(P<0.01),和Ⅰ组比较:Ⅱ、Ⅲ、Ⅳ、Ⅴ组大鼠再灌注24h后出现明显的神经体征缺陷(P<0.01),Ⅱ组大鼠额叶皮层和纹状体区域大量神经细胞凋亡,纹状体PKCγ蛋白表达明显下降。Ⅱ、Ⅲ、Ⅳ、Ⅴ组大鼠纹状体区域凋亡细胞密度与PKCγ染色阳性面积均没有差异(P>0.05)。结论再灌注前30min腹腔注射异丙酚对大鼠局灶性脑缺血再灌注损伤所致的神经细胞凋亡和PKCγ表达降低无保护作用。     

松龄血脉康对大鼠局灶性脑缺血-再灌注损伤和细胞凋亡的影响

目的探讨松龄血脉康对局灶性脑缺血-再灌注损伤和细胞凋亡的作用及机制。方法线栓法制作大鼠大脑中动脉阻塞(MCAO)模型。观察松龄血脉康对脑缺血-再灌注损伤大鼠行为学、海马CA1区凋亡细胞数、细胞超微结构的影响;运用免疫组织化学法检测海马凋亡相关基因Bcl-2、Bax蛋白免疫反应阳性细胞表达。结果松龄血脉康显著降低MCAO模型大鼠神经功能缺损评分,明显减少海马组织凋亡细胞数,促进海马神经元修复,松龄血脉康组海马Bcl-2的表达明显增加、Bax的表达降低。结论松龄血脉康可减少脑缺血-再灌注损伤后神经元的凋亡而发挥脑保护作用,机制可能与增强Bcl-2的表达及抑制Bax的表达有关。     

Neuroprotective effects of hydroxysafflor yellow A: in vivo and in vitro studies

Previous work has shown that hydroxysafflor yellow A (HSYA), extracted from Carthamus tinctorius L. markedly extended the coagulation time in mice and exhibited a significant antithrombotic effect in rats. The present study was conducted to demonstrate further its neuroprotective effects on cerebral ischemic injury in both in vivo and in vitro studies. In vivo, male Wistar-Kyoto (WKY) rats with middle cerebral artery occlusion (MCAO) were evaluated for neurological deficit scores followed by the treatment with a single dose of HSYA. Furthermore, the infarction area of the brain was assessed in the brain slices. In vitro, the effect of HSYA was tested in cultured fetal cortical cells exposed to glutamate and sodium cyanide (NaCN) to identify its neuroprotection against neurons damage. The results in vivo showed that sublingular vein injection of HSYA at doses of 3.0 mg/kg and 6.0 mg/kg exerted significant neuroprotective effects on rats with focal cerebral ischemic injury by significantly decreasing neurological deficit scores and reducing the infarct area compared with the saline group, HSYA at a dose of 6.0 mg/kg showed a similar potency as nimodipine at a dose of 0.2 mg/kg. Sublingular vein injection of HSYA at the dose of 1.5 mg/kg showed a neuroprotective effect, however, with no significant difference when compared with the saline group. Results in vitro showed that HSYA significantly inhibited neuron damage induced by exposure to glutamate and sodium cyanide (NaCN) in cultured fetal cortical cells. Noticeably, the neuroprotective action of HSYA on glutamate-mediated neuron injury was much better than that of HSYA on NaCN-induced neuron damage. All these findings suggest that HSYA might act as a potential neuroprotective agent useful in the treatment in focal cerebral ischemia. Abbreviations. HSYA:hydroxysafflor yellow A TTC:2,3,5-triphenyltetrazolium chloride MTT:3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide DMEM:Dulbecco's modified Eagle medium FCS:Fetal calf serum MCAO:middle cerebral artery occlusion ECA:external carotid artery ICA:internal carotid artery LDH:lactate dehydrogenase NMDA: N-methyl- D-aspartate     

后适应对大鼠脑缺血神经细胞凋亡和内皮型一氧化氮合酶与诱导型一氧化氮合酶的影响

目的：研究缺血耐受（后适应和预适应）对脑缺血再灌注神经细胞凋亡及内皮型一氧化氮合酶（eNOS）和诱导型一氧化氮合酶（iNOS）蛋白表达的影响。方法：50只雄性SD大鼠随机分为假手术组（sham）、脑缺血再灌注模型组（MCAO）、预适应组（MCAO＋preconditioning）、后适应组（MCAO＋postconditioning）和尼莫地平组（MCAO＋nimodipine），每组10只大鼠。预适应组大鼠在MCAO前24h，双侧颈总动脉夹闭2min，再灌注20min，循环两次。后适应组大鼠在脑缺血再灌注开始时，再灌注20s，栓塞20s，循环3次。大鼠大脑中动脉阻断1．5h，再灌注24h。用TUNEL法和免疫组化染色法分别检测缺血半暗带凋亡细胞和iNOS、eNOS蛋白表达。结果：与MCAO组比较，后适应组大鼠脑缺血半暗带的凋亡细胞显著减少，eNOS蛋白表达显著增加，iNOS蛋白表达显著减少，差异有统计学意义（P〈0．05）。结论：缺血后适应能诱导脑缺血耐受，对脑缺血／再灌注损伤产生保护作用，其保护作用与促进eNOS蛋白的表达，抑制iNOS蛋白的表达有关。     

盐酸非那嗪奈对大鼠局灶性脑缺血损伤的保护作用

目的探讨盐酸非那嗪奈(fenazinel dihydrochloride,FD)对大鼠局灶性脑缺血损伤的保护作用及其机制。方法线拴法制备大鼠中动脉局灶性脑缺血(24h)模型。测定脑梗死面积;检测缺血组织中超氧化物歧化酶(SOD)活性以及丙二醛(MDA)、乳酸(LA)含量;观察组织病理学改变。制备大鼠原代神经元细胞撤血清损伤模型及N-甲基-D-门冬氨酸(NMDA)损伤模型,检测FD对损伤细胞死亡率、SOD活性及MDA、LA含量的影响。结果FD可明显降低脑梗死面积,升高组织中SOD活性,降低MDA、LA含量,减轻脑组织病理学损害;在细胞水平上,FD降低损伤细胞死亡率,升高SOD活性,降低MDA、LA含量。结论FD对大鼠局灶性脑缺血损伤具有明显的保护作用,其机制与降低兴奋性氨基酸损伤、清除氧自由基及改善能量代谢有关。     

阿司匹林预处理对大鼠脑缺血再灌注损伤的神经保护作用

目的:观察阿司匹林(ASA)预处理对大鼠局灶性脑缺血再灌注损伤(I/R)的神经保护作用,并对其作用机制进行探讨。方法:复制大鼠大脑中动脉缺血再灌注模型,分别采用TTC染色法、神经功能缺损评分法观察ASA预处理对大鼠脑梗死体积和神经功能评分的影响,以及对脑组织中超氧化物歧化酶(SOD)、丙二醛(MDA)和血前列环素I2(PGI2)/血栓素A2(TXA2)的影响。结果:ASA能够降低I/R大鼠脑梗死体积和神经功能评分,增加脑组织中SOD的活性,降低MDA的含量,升高血PGI/TXA2的比值。结论:ASA对大鼠脑缺血再灌注损伤有一定保护作用,其作用机制与增加脑组织中SOD的活性、降低MDA的含量、提高血PGI2/TXA2的比值有关。     

Blood brain barrier permeability and therapeutic time window of Ginkgolide B in ischemia–reperfusion injury

The goal of this study was to estimate the blood brain barrier (BBB) permeability of Ginkgolide B in normal condition and models of ischemia both in vivo and in vitro. A sensitive LC-MS/MS analytical method was developed to determinate accurately the concentration of Ginkgolide B in cell, plasma and brain tissue. The injured rat brain microvessel endothelial cells (RBMECs) induced by Na₂S₂O₄ served as a hypoxia/reoxygenation model in vitro. Intracellular concentration of Ginkgolide B increased in injured cells in a concentration-dependent manner. As a model of in vivo—ischemia/reperfusion, we performed middle cerebral artery occlusion (MCAO) in rats. Concentration of Ginkgolide B in the brain tissues showed higher in cerebral ischemia-reperfused animals than that in normal rats. To evaluate potential clinical effect of Ginkgolide B, we determined therapeutic time window in MCAO rats. Up to i.v. administration at 2 h after reperfusion of rats, Ginkgolide B could decrease infarction volume and brain edema, exerting significant protective effect in cerebral ischemia injury. In conclusion, Ginkgolide B could pass through BBB, especially after ischemia–reperfusion injury of brain, and might be therapeutically effective for ischemia/reperfusion injury of human brain.