丁基苯酞对局部脑缺血大鼠神经元迟发性损伤及细胞内钙的影响

观察了丁基苯酞(NBP)对局部脑缺血迟发性脑梗塞和神经功能缺损的影响。结果表明,大鼠大脑中动脉阻断(MCAO)后2h分别poNBP80,160和240mg·kg^-1,均能明显降低脑梗塞面积,抑制率分别为49.0%,69.5%及85.1%,并明显改善神经功能缺失。在MCAO前1h予防给NBP(160mg·kg^-1po),也可明显缩小脑梗塞面积和改善神经功能缺失;在MCAO前连续口服7d,NBP剂量为80mg·kg^-1·d^-1,对上述指标均有改善作用。NBP对高K⁺引起大鼠脑突触体内Ca²⁺含量升高无影响。以上结果提示NBP对迟发性神经元损伤有保护作用。     

丁基苯酞对小鼠全脑缺血的保护作用

观察了丁基苯酞(NBP)对小鼠全脑缺血脑保护作用和脑能量代谢的影响。用Lowry法测定脑乳酸,ATP和磷酸肌酸(PCr)的含量。结果表明,NBP112.5或250mg·kg^-1sc能明显延长断头或iv饱和MgCl₂后喘息的时间;NBP15O或200mg·kg^-1sc能明显减少断头缺血后乳酸的升高和ATP及PCr的降低。实验结果提示,NBP对缺血脑有保护和改善脑能量代谢的作用。     

丁基苯酞对蛛网膜下腔出血后脑血流的改善及血脑屏障的保护作用

侧脑室注入自体血液造成蛛网膜下腔出血模型,观察丁基苯酞(dl-NBP)对局部脑血流的改善作用及血脑屏障的保护作用。结果表明,dl-NBP5～20mg·kg^-1明显提高蛛网膜下腔出血后3h内尾状核的血流量,但无明显的剂量效应关系。0.25mg·kg^-1的尼莫地平亦明显提高脑血流。并发现dl-NBP10mg·kg^-1及尼莫地平0.25mg·kg^-1(分别于蛛网膜下腔出血后5min和3hip)均能明显降低蛛网膜下腔出血6h后皮层组织中伊文氏蓝的摄取量,提示对血脑屏障有明显的保护作用。     

小檗碱对2型糖尿病大鼠视网膜PPARα/δ/γ表达的影响

糖尿病视网膜病是最常见的糖尿病并发症之一，是导致失明的主要原因，因此有必要寻找新的治疗药物。给大鼠腹腔注射链脲菌素(35 mg·kg^-1) 2周后用高糖高脂饲料喂养14周，之后连续16周每天分别拌食给予小檗碱75、150及300 mg·kg^-1、非诺贝特100 mg·kg^-1和罗格列酮4 mg·kg^-1。用HE染色检查视网膜结构和免疫组化检测过氧化物酶体增殖物激活受体(PPARs) α/δ/γ的表达。结果发现，正常对照组大鼠的视网膜厚度大于其他各组，经小檗碱(150及300 mg·kg^-1)和罗格列酮(4 mg·kg^-1)的治疗能增加糖尿病大鼠视网膜的厚度，但视网膜的结构在各组间无差别；小檗碱(150及300 mg·kg^-1)和罗格列酮(4 mg·kg^-1)能明显降低糖尿病大鼠视网膜中PPARγ蛋白表达，小檗碱(150及300 mg·kg^-1)和非诺贝特(100 mg·kg^-1)能显著增加糖尿病大鼠视网膜中PPARα和PPARδ的表达。小檗碱调控视网膜PPAR α/δ/γ蛋白表达可能是其改善糖尿病视网膜病的机制之一，可能成为比罗格列酮和非诺贝特更有效的用于治疗糖尿病视网膜病的药物。     

双苯氟嗪抑制FasL分子表达的基因转录机制

研究双苯氟嗪对Fas配体分子表达抑制的基因转录机制的影响。通过四血管阻断法建立大鼠全脑缺血再灌注损伤模型， 所有实验动物缺血15 min， 再灌注72 h。实验大鼠随机分为4组： 假手术组、 缺血再灌注组、 双苯氟嗪组及环孢素A组。药物干预于再灌注后2 h内给药， 每日1次， 连续3 d。双苯氟嗪按20 mg·kg^-1灌胃给药， 环孢素A 10 mg·kg^-1腹腔注射。应用蛋白质印迹和电泳迁移率改变分析技术检测海马CA I区Fas配体(FasL)、 转录因子NFATc、 I-κB-α、 phospho-I-κB-α蛋白表达以及测定FasL分子启动子远端及FasL分子启动子近端NFAT FasL-DNA结合活性。结果表明， 双苯氟嗪明显降低FasL、 NFATc的蛋白表达并显著减少FasL分子启动子远端及FasL分子启动子近端NFAT结合位点的NFAT-DNA结合活性。各组之间I-κB-α蛋白表达无显著区别。未观察到各组phospho-I-κB-α蛋白表达。由此可见， 双苯氟嗪通过降低转录因子NFATc的FasL分子的基因转录功能， 从而抑制FasL分子的基因表达。     

氧化苦参碱对病毒性心肌炎小鼠心肌细胞凋亡及相关因子的影响

目的 探讨氧化苦参碱(oxymatrine,OMT)对病毒性心肌炎小鼠心肌细胞凋亡及其凋亡相关因子蛋白表达的影响。方法 42只BALB/c小鼠随机分为正常对照组(NC)、病毒性心肌炎模型组(VM)、OMT高剂量组(OMT-H,25?mg·kg^-1·d^-1)、OMT中剂量组(OMT-M,12.5 mg·kg^-1·d^-1)、OMT低剂量组(OMT-L,6.25 mg·kg^-1·d^-1)、OMT极低剂量组(OMT-EL,3.125 mg·kg^-1·d^-1)、利巴韦林对照组(RB,100 mg·kg^-1·d^-1)。病毒性心肌炎小鼠由柯萨奇病毒B3型腹腔注射感染所致,各治疗组从末次给予病毒24 h后开始,腹腔注射,每日1次。于治疗第12天,每组处死小鼠6只,留取心肌标本。TUNEL法检测心肌细胞凋亡情况,免疫印迹法和免疫组织化学法检测Bcl-2和Bax蛋白的表达情况。结果 OMT治疗显著降低了病毒性心肌炎小鼠凋亡心肌细胞的数量,与病毒性心肌炎模型组比较,OMT-L组效果最佳(P<0.01)。与病毒性心肌炎模型组小鼠比较,OMT治疗组的小鼠心肌组织中Bax蛋白表达降低,而Bcl-2蛋白表达没有明显改变。结论 氧化苦参碱可减少病毒性心肌炎小鼠心肌细胞的凋亡,该作用与下调Bax蛋白表达有关。     

香菇多糖的免疫调节作用

研究香菇多糖(LTN)的免疫调节作用。结果表明,LTN1及5mg·kg^-1·d^-1×6,ip可促进正常小鼠由ConA(2.5mg·kg^-1)刺激的脾脏T淋巴细胞增殖反应。1,5及10mg·kg^-1·d^-1×8或5,ip能分别纠正由环磷酰胺(Cy,200mg·kg^-1和80mg·kg^-1,ip)诱导的免疫亢进或低下状态。此外,LTN(1,5和10mg·kg^-1·d^-1×6,ip),促使小鼠胸腺L3T4+(Th)和Lyt2+(Ts)细胞数减少,外周脾脏L3T4+和Lyt2+细胞数增加,腹腔巨噬细胞释出肿瘤坏死因子(TNF)也明显增加。这些作用均以LTN5mg·kg^-1·d^-1作用最佳。提示LTN可能通过影响T细胞及其亚型,促进TNF活性调节机体的免疫功能。     

畲药地稔全草及其不同部位重金属含量相关性分析

目的 分析地稔全草及其不同部位与重金属含量的关系。方法 参照中国药典2010年版的检测方法,对地稔全草及其根、茎、叶中铜、铅、镉、汞、砷含量进行测定。结果 26批全草中,铅的含量均>5 mg·kg^-1,7批镉的含量>0.3 mg·kg^-1,砷的含量均≤1 mg·kg^-1,汞的含量均<0.1 mg·kg^-1,铜的含量均<20 mg·kg^-1。10批分部位考察的样品中,根中铅含量均>5 mg·kg^-1、9批镉的含量>0.3 mg·kg^-1;茎中5批铅的含量>5 mg·kg^-1、10批镉的含量>0.3 mg·kg^-1;叶中1批铅的含量>5 mg·kg^-1、10批镉的含量均<0.3 mg·kg^-1;各部位砷的含量均≤1 mg·kg^-1、汞的含量均<0.1 mg·kg^-1、铜的含量均<20 mg·kg^-1。结论 以重金属含量为指标进行考察后,依据用药安全性原则,建议地稔以地上部位入药。     

反式白藜芦醇对Aβ25-35诱导的阿尔兹海默病小鼠模型学习记忆的改善作用

目的 观察反式白藜芦醇对阿尔茨海默病(Alzheimer's disease,AD)模型小鼠记忆损伤的改善作用。方法 小鼠随机分为假手术组、模型组和反式白藜芦醇10,20,40 mg·kg^-1组。在小鼠海马CA1区注射Aβ25-35后给予反式白藜芦醇10 d,采用水迷宫试验观察药物对小鼠学习记忆的影响,免疫组化法观察各组小鼠海马CA1区神经元可塑性变化,western-blot法检测神经可塑性相关蛋白的表达变化。结果 40?mg·kg^-1反式白藜芦醇能明显缩短AD小鼠寻找平台的潜伏期,增加原平台所在象限的停留时间和穿越次数;20,40 mg·kg^-1反式白藜芦醇能增加海马CA1区神经元顶端树突的长度和树突的密度;40 mg·kg^-1反式白藜芦醇能增加AD小鼠海马CA1区BDNF、pCREB以及c-fos蛋白的表达。结论 反式白藜芦醇能逆转Aβ引起的小鼠的学习记忆损伤,其机制可能与改善海马神经元的神经可塑性有关。     

阿魏酸对小鼠脑缺血再灌注损伤及大鼠血液流变性的影响

目的观察阿魏酸对小鼠脑缺血再灌注损伤、大鼠血液流变性的影响。方法用结扎双侧颈总动脉法,造成小鼠脑缺血再灌注模型;观察阿魏酸(0.8,1.6 g·kg^-1·d^-1)对小鼠全脑缺血30m in及再灌注60m in后超氧化物歧化酶(SOD)活性、乳酸脱氢酶(LDH)活性、MDA含量的影响。大鼠皮下注射肾上腺素(0.8mg·kg^-1),共2次,间隔4h,在第一次注射后2h将大鼠浸入4℃冰水5m in,造成血瘀模型,观察阿魏酸(50～200mg·kg^-1·d^-1)对血黏度的影响。结果阿魏酸(0.8g·kg^-1·d^-1)可显著提高脑组织LDH活性、可明显降低MDA含量、可显著提高SOD活性;阿魏酸(1.6g·kg^-1·d^-1)可显著降低MDA/SOD比值;对大鼠血黏度无明显影响。结论预防性应用阿魏酸对小鼠脑缺血再灌注损伤有一定程度的保护作用,对大鼠血液流变性无明显影响。     

丁基苯酞对大鼠局灶性脑缺血和重灌后脑内TXB2和6—keto—PGF1α…

目的：观察丁基苯酞（ＮＢＰ）对大鼠局灶性脑缺血及重灌后海马，纹状体和皮层中ＴＸＢ２及６－ｋｅｔｏ－ＰＧＦ１α含量的影响，方法：尼龙线栓塞法造成大鼠局灶性脑缺血模型，ＴＸＢ２和６－ｋｅｔｏ－ＰＧＦ１α用放免法测定。结果：ＮＢＰ１０ｍｇ．ｋｇ＾－１治疗对缺血重灌注后脑组织中ＴＸＢ２的产生具有抑制作用，但对６－ｋｅｔｏ－ＰＧＦ１α的产生无明显作用，ＮＢＰ２０ｍｇ．ｋｇ＾－１治疗后，重灌５ｍｉｎ缺血脑组织     

莽草对大鼠大脑中动脉血栓所致局部脑缺血性损伤的拮抗作用

目的：研究莽草酸（ＳＡ）对大脑中动脉血栓所致局部脑缺血的影响。方法：用三氯化铁局部涂抹损伤血管形成的大鼠大脑中动脉血栓模型（ＭＣＡＴ）观察ＳＡ对行为障碍程度、脑梗塞范围、脑水肿和缺血区脑血流的影响。结果：ＳＡ２５、５０ｍｇ．ｋｇ＾－１ｉｐ于ＭＣＡＴ前连续红药３ｄ可分涉及了梗塞范围５１％、４２％,使脑含水量由８０．７％降致７９．８％、７９．９％,并可改善行为障碍程度和缺血区脑血流量的降低,病理检查显     

丁基苯酞对大脑中动脉阻断后皮层组织中花生四烯酸释放及磷脂酶A2基因表达的影响

目的　研究丁基苯酞 (dl NBP) ,d NBP和l NBP对大脑中动脉阻断 (MCAO) 6h后缺血区皮层中花生四烯酸 (AA)释放及磷脂酶A2 (PLA2 )基因表达的影响。方法　阻断大脑中动脉起始部造成局灶性脑缺血模型。HPLC检测AA。Northernblot检测皮层中PLA2 基因表达。结果　MCAO后 6h ,皮层中AA释放明显增加。于脑缺血后 5min和 12 0min ,给dl NBP(10或 2 0mg·kg- 1)和尼莫地平 (0 5mg·kg- 1)可显著抑制AA的释放。d NBP和l NBP作用比较 ,显示d NBP有与dl NBP相似的作用 ,而l NBP则无明显影响。Northern印迹结果表明 ,脑缺血 6h ,皮层中PLA2 的基因表达增强。dl NBP和d NBP(10 ,2 0mg·kg- 1,ip)皆可使表达降低 ,而l NBP对缺血脑组织中PLA2 的基因表达的升高无明显影响。结论　dl NBP和d NBP可抑制MCAO后脑组织中AA释放和PLA2 的基因表达。     

Inhibitory effects of chiral 3-n-butylphthalide on inflammation following focal ischemic brain injury in rats

AIM: To evaluate the degree of neutrophil infiltration into ischemic tissue after transient focal cerebral ischemia, and to examine the effects of chiral 3-n-butylphthalide (NBP) on this inflammatory process. METHODS: After a 24-h reperfusion following transient cerebral ischemia, two different techniques, histologic analysis and modified myeloperoxidase (MPO)-quantification method, were utilized to identify the infiltration of neutrophils into cerebral tissue following ischemia. The expression of intercellular adhesion molecule-1 (ICAM-1) and tumor necrosis factor-alpha(TNF-alpha) in the ischemic zone were observed by immunohistochemistry, Western blot, and in situ hybridization techniques. RESULTS: In cerebral cortex area perfused by middle cerebral artery (MCA), MPO activity was greatly increased after 24 h of reperfusion in the vehicle group, and it correlated well with the infiltration of neutrophils. Administration of dl-, d-, and l-NBP (20 mg.kg-1) partially inhibited both the increase in MPO activity and the appearance of neutrophils in ischemia-reperfusion sites. Up-regulation of ICAM-1 was also observed on the microvessel endothelium in the ischemic territory. In addition, chiral NBP markedly blunted ICAM-1 expression, and decreased the number of TNF-alpha blue purple-positive neurons induced by ischemia-reperfusion injury. CONCLUSION: The results indicate that the increase in neutrophils infiltration into the infarct site implicated postischemic brain injury, and NBP was effective in protecting the ischemic sites following ischemic insult.     

手性3-n丁基苯酞对大鼠局灶性脑缺血诱导的凋亡的影响

目的:观察和比较手性3-n丁基苯酞(NBP)对大鼠局灶性脑缺血再灌注诱导的凋亡的影响。方法:插线法制备大脑中动脉阻断(MCAO)模型,原位末端标记(TUNEL)和琼脂糖凝胶电泳检测DNA断裂,蛋白质印迹和免疫组化方法检测细胞色素c及caspase-3蛋白的表达。结果:大鼠局灶性脑缺血2小时,再灌注开始后24小时可见明显的TUNEL阳性染色和DNA ladder.s-(-)-NBP 5,10 mg/kg显著减少TUNEL阳性细胞数和DNA ladder,s-(-)-NBP 10 mg/kg几乎完全抑制DNA片段化,而r-(+)-NBP 10 mg/kg仅有轻度抑制作用。(±)-NBP(20 mg/kg)对DNA片段化的抑制作用介于s-(-)-NBP(10 mg/kg)和r-(+)-NBP(10 mg/kg)之间。脑缺血过程中可见到线粒体细胞色素c的释放及caspase-3的激活,s-(-)-NBP能明显抑制这一效应,r-(+)-NBP和(±)-NBP对细胞色素c和caspase-3作用的强弱与它们抑制DNA片段化的作用强度相似。结论:NBP,尤其是s-(-)-NBP能够抑制脑缺血诱导的DNA片段化,细胞色素c释放和caspase-3激活。     

丁基苯酞对线粒体呼吸链复合酶活性的影响

目的：阐明正丁基苯酞(dl-3-n-butylphthalide, dl-NBP)改善缺血脑能量代谢的机制。方法：用分光光度法测定局灶性脑缺血大鼠脑线粒体呼吸链复合酶I,II,III和IV活性的改变,并观察dl-NBP对这些变化的影响。结果：缺血时复合酶II活性升高,IV的活性显著降低;再灌后, 复合酶I活性升高,II的活性降低。在缺血前10 min给予dl-NBP(5 mg.kg^-1或10 mg.kg^-1, ip)能逆转缺血-再灌引起的上述复合酶活性改变,尤其是使缺血后急剧降低的复合酶IV活性得到明显提高。同时NBP(d-,l-,dl-)还能逆转低糖低氧造成的原代培养大鼠皮质细胞呼吸链复合酶IV活性降低。结论：NBP能够改善缺血脑内能量状态是直接作用于脑线粒体的结果,而d-NBP起着主要作用。     

Orally administered oleoylethanolamide protects mice from focal cerebral ischemic injury by activating peroxisome proliferator-activated receptor α

Oleoylethanolamide (OEA) is a high-affinity agonist of peroxisome proliferator-activated receptor α (PPARα) which may act as an endogenous neuroprotective factor. However, it is not clear whether orally administered OEA is effective against ischemic brain injury. In our study, transient focal cerebral ischemia was induced by middle cerebral artery occlusion for 90 min followed by reperfusion. To evaluate its preventive effects, OEA (10, 20 or 40 mg/kg, ig) was administered for 3 days before ischemia. To evaluate its therapeutic effects, OEA (40 mg/kg, ig) was administered at 0.5 or 1h before reperfusion or at 0 or 1h after reperfusion. In some experiments, the PPARα antagonist MK886 (10mg/kg, ig) was administered 0.5h before OEA. Neurological deficit score, infarct volume and brain edema degree were determined at 24h after reperfusion. Blood-brain barrier (BBB) disruption was evaluated by Evans blue (EB) leakage at 6h after reperfusion. Real-time RT-PCR and western blot were performed to detect PPARα mRNA and protein expression. Oral OEA pretreatment improved neurological dysfunction reduced infarct volume and alleviated brain edema in a dose-dependent manner; the most effective dose was 40 mg/kg. The therapeutic time is within 1h after reperfusion. OEA also increased PPARα mRNA and protein expression in the ischemic brain. The PPARα antagonist MK886 abolished the protective effects of OEA. In conclusion, our results indicate that orally administered OEA protects against acute cerebral ischemic injury in mice, at least in part by activating PPARα.     

丁基苯酞对大脑中动脉阻断后皮层组织中花生四烯酸释放及磷脂酶A2基因表达的影响

目的　研究丁基苯酞（dl-NBP), d-NBP和 l-NBP对大脑中动脉阻断(MCAO) 6 h后缺血区皮层中花生四烯酸(AA)释放及磷脂酶A₂（PLA₂）基因表达的影响。方法　阻断大脑中动脉起始部造成局灶性脑缺血模型。HPLC检测AA。Northern blot检测皮层中PLA₂基因表达。结果　MCAO后6 h,皮层中AA释放明显增加。于脑缺血后5 min 和120 min,给dl-NBP(10或20 mg.kg^-1) 和尼莫地平(0.5 mg.kg^-1) 可显著抑制AA的释放。d-NBP和l-NBP作用比较,显示d-NBP有与dl-NBP相似的作用,而l-NBP则无明显影响。Northern印迹结果表明,脑缺血6 h,皮层中PLA₂的基因表达增强。 dl-NBP和d-NBP(10, 20 mg.kg^-1,ip)皆可使表达降低,而l-NBP对缺血脑组织中PLA₂的基因表达的升高无明显影响。结论　dl-NBP和d-NBP可抑制MCAO后脑组织中AA释放和PLA₂的基因表达。     

The edaravone and 3-n-butylphthalide ring-opening derivative 10b effectively attenuates cerebral ischemia injury in rats

Aim:

Compound 10b is a hybrid molecule of edaravone and a ring-opening derivative of 3-n-butylphthalide (NBP). The aim of this study was to examine the effects of compound 10b on brain damage in rats after focal cerebral ischemia.

Methods:

SD rats were subjected to 2-h-middle cerebral artery occlusion (MCAO). At the onset of reperfusion, the rats were orally treated with NBP (60 mg/kg), edaravone (3 mg/kg), NBP (60 mg/kg)+edaravone (3 mg/kg), or compound 10b (70, 140 mg/kg). The infarct volume, motor behavior deficits, brain water content, histopathological alterations, and activity of GSH, SOD, and MDA were analyzed 24 h after reperfusion. The levels of relevant proteins in the ipsilateral striatum were examined using immunoblotting.

Results:

Administration of compound 10b (70 or 140 mg/kg) significantly reduced the infarct volume and neurological deficits in MCAO rats. The neuroprotective effects of compound 10b were more pronounced compared to NBP, edaravone or NBP+edaravone. Furthermore, compound 10b significantly upregulated the protein levels of the cytoprotective molecules Bcl-2, HO-1, Nrf2, Trx, P-NF-κB p65, and IκB-α, while decreasing the expression of Bax, caspase 3, caspase 9, Txnip, NF-κB p65, and P-IκB-α.

Conclusion:

Oral administration of compound 10b effectively attenuates rat cerebral ischemia injury.     

Cerebral activation and distribution of inducible hsp110 and hsp70 mRNAs following focal ischemia in rat

A potential function for inducible heat shock protein 70 (hsp70i) expression in the pathophysiology of ischemic brain has been well documented. The recently cloned hsp70 superfamily member, hsp110, was shown to be highly expressed in the brain and suggested to have a similar functional property as members of the hsp70 family. In this study, as an initial step to probe for its physiological significance in the ischemic brain, cerebral activation and distribution of hsp110 mRNA was comparatively evaluated with that of hsp70i mRNA by in situ hybridization. A rat focal cerebral ischemia model was employed to examine the distribution and localization of hsp110 and hsp70i mRNAs in both affected (ipsilateral) and unaffected (contralateral) hemispheres of the same animal. Our results demonstrated a significant accumulation of hsp110 as well as hsp70i mRNAs following ischemia; although the magnitude and kinetics of induction differ slightly, spatial expression profiles of hsp110 and hsp70i mRNAs were highly correlated in the affected region. In control brain, limited hybridization signal was observed with 3'-untranslated region (UTR) containing hsp110 probe, suggesting a possible existence of inducible hsp110 and a selective recognition of our 3'-UTR containing probe for the inducible hsp110 mRNA species. Subsequent 2D western analysis with Hsp110 specific Ab was consistent with our view, which resolved constitutive and inducible immunostained spots in rat ischemic brain. Considering a regulatory similarity as well as previously documented structural and functional similarities between hsp110 and hsp70i, we propose that coordinated cerebral activation of hsp110 and hsp70i is likely to be of significant relevance in the context of pathophysiology of ischemic brain. Further study is required to characterize the genetic and biochemical nature of rat inducible hsp110 identified in the current study.