【作者】 陈静波；

【导师】 金满文；

【作者基本信息】 华中科技大学 ， 药理学， 2006， 硕士

【摘要】 芪丹中药提取物(QDPTP)是按照中医理论组方、经现代工艺加工而成的复方中药提取物。该组方中黄芪为君药,丹参、川芎和红花为臣药,银杏叶、三七和红景天为佐使药。已有研究表明QDPTP具有抗心肌缺血-再灌注损伤的作用。本课题分别采用大鼠局灶性和全脑缺血-再灌注模型,研究QDPTP的抗大鼠实验性脑缺血-再灌注损伤作用及机制,为此药的开发研究提供实验依据。一、QDPTP抗脑缺血-再灌注损伤作用1. QDPTP对局灶性脑缺血-再灌注大鼠神经功能的影响SD大鼠,给生理盐水(10ml/kg/d,7d,i.g.)、QDPTP低、中、高剂量(450、900、1800mg/kg/d,7d,i.g.),川芎嗪(33mg/kg/d,缺血前30min,i.v.)。阻塞右侧大脑中动脉造成局灶性缺血模型,2小时后再灌注,观察QDPTP对再灌注后神经功能的影响。模型组、QDPTP低、中、高剂量组和川芎嗪组的神经功能缺陷等级评分分别为3.7±1.2、1.4±0.5(vs模型组,P<0.01)、1.8±0.4(P<0.01)、1.8±0.5(P<0.01)和1.9±0.5(P<0.01)。此结果表明:QDPTP低、中、高剂量组均能显著改善缺血-再灌注引起的大鼠神经功能缺陷症状。2. QDPTP对大鼠局灶性脑缺血-再灌注损伤脑梗死范围的影响在大鼠局灶性脑缺血-再灌注损伤模型,采用TTC染色,见模型组、QDPTP低、中、高剂量组和川芎嗪组的脑梗死范围分别为20.9±7.3(%)、6.6±4.4(%)、6.4±4.8(%)、4.4±2.6(%)、和6.6±4.3(%),所有用药组均明显缩小脑梗死范围(P<0.01,vs模型组)。3. QDPTP对大鼠局灶性脑缺血-再灌注损伤脑含水量和脑指数的影响在大鼠局灶性脑缺血-再灌注模型,模型组动物脑组织含水量(81.8±0.8,%)较伪手术组明显增加(P<0.01);各给药组均明显降低局灶性缺血-再灌注引起的脑组织含水量升高(P<0.01或P<0.05),分别为:QDPTP低剂量(78.8±1.4,%)、中剂量(78.1±1.8,%)、高剂量(80.5±0.6,%)和川芎嗪(79.0±2.0,%)。与模型组比较,各给药组脑指数均表现出降低的趋势,但统计学意义不显著。二、QDPTP抗脑缺血-再灌注损伤的机制研究(一) QDPTP对大鼠全脑缺血-再灌注所致氧化损伤和炎症反应的影响1.制备大鼠全脑缺血-再灌注损伤模型和实验分组采用Pulsinelli四血管闭塞的方法略加改良建立大鼠急性全脑缺血-再灌注损伤模型。SD大鼠,随机分为伪手术组、模型组(NS,10ml/kg/d,7d,i.g.)、QDPTP低、中剂量组(450、900mg/kg/d, 7d,i.g.),川芎嗪组(33mg/kg,i.v.,缺血前30min和再灌后各1次)。2. QDPTP对全脑缺血-再灌注大鼠脑组织SOD活性和MDA含量的影响采用黄嘌呤氧化酶法测定SOD活力,采用硫代巴比妥显色法测定MDA含量,组织蛋白测定用考马斯亮蓝蛋白测定法。模型组中脂质过氧化产物MDA的含量(4.53±0.92nmol/mg prot)显著升高(P<0.01 vs伪手术组),过氧化物歧化酶SOD活性(79.4±7.3U/mg prot)显著降低(P<0.01 vs伪手术组);各给药组均能显著减少MDA含量(P<0.01 vs模型组),增加SOD活性(P<0.01 vs模型组),分别为:QDPTP低剂量(2.10±0.17,116.9±11.2)、中剂量(2.35±0.09,111.5±6.2)和川芎嗪(1.66±0.11,95.6±13.3)。3. QDPTP对全脑缺血-再灌注大鼠脑组织及血清IL1-β含量的影响采用双抗体夹心酶联免疫吸附法测定。结果表明:模型组脑组织及血清IL1-β含量分别为1006.4±247.1 pg/ml和370.8±76.6 pg/ml,与之相比,QDPTP中剂量(550.7±99.8,268±64.6)组中脑组织及血清中IL1-β的含量显著降低(P<0.01),川芎嗪脑也明显减少组织IL1-β的含量(627.8±144.7,P<0.05)。(二) QDPTP对麻醉大鼠局部脑血流量和动脉血压的影响采用激光多普勒血流仪测量大鼠软脑膜局部的血灌注量,其被视为局部的脑血流量(rCBF)。单次十二指肠给予生理盐水(10ml/kg)、QDPTP低、中、高剂量(450、900、1800mg/kg),尼莫地平(20mg/kg),复方丹参滴丸(60mg/kg),或静脉注射川芎嗪(33mg/kg),记录给药前后rCBF的变化。采用电生理记录仪监测给药前后外周动脉平均收缩压、平均舒张压、平均压的变化。结果表明:持续观察120min,溶剂对照组的脑血流量和血压各参数无明显变化。QDPTP低、中、高剂量、川芎嗪、尼莫地平和复方丹参滴丸均能不同程度增加大鼠局部脑血流(P<0.01或P<0.05),外周动脉血压出现不同程度的降低。结论1. QDPTP可减轻缺血-再灌注引起的神经功能损伤,减轻脑水肿,缩小脑梗死范围。2. QDPTP抗脑缺血-再灌注损伤作用可能与抑制自由基损伤、减轻细胞因子介导的炎症反应、扩张脑血管增加脑血流等因素有关。更多还原

【Abstract】 QDPTP is an compound extractive of several kinds of the traditional Chinese medicine including Astragalus mongholicus, salvia miltiorrhiza, szechwan lovage rhizome, Carthamus tinctorius and so forth. It has been proved that QDPTP protect heart damaged by ischemia / reperfusion (I/R). And the aims of this study were to confirm the protect effect of QDPTP in experimental cerebral ischemia / reperfusion model and further clarify the mechanisms of actions by using different experimental methods. The main results are as follows:Part I QDPTP decreased cerebral injury induced by focal cerebral ischemia / reperfusion in rats1. The effects of QDPTP on the neurological deficits scores after I/R in ratsFocal cerebral ischemia was induced by middle cerebral artery occlusion (MCAO) for 2 hours and followed reperfusion for 4 hours. After that, we estimated deficit scores grading on a scale of 0-5. QDPTP (450, 900, 1800mg/kg) and Lig (33mg/kg) were found significantly decreasing neurological deficit scores after I/R (P<0.01).2. The effects of QDPTP on cerebral infarction size after I/R in ratsThe cerebral infarction size was measured by TTC staining technique after I/R. QDPTP (450, 900, 1800mg/kg) and Lig (33mg/kg) evidently decreased the cerebral infarction size after I/R (P<0.01).3. The effects of QDPTP on cerebral water contents and cerebral index after I/R in ratsWe measured the cerebral water contents with wet/dry weighing method and surveryed cerebral index after I/R in rats. The results showed that QDPTP (450, 900, 1800mg/kg) groups and Lig (33mg/kg) cut down both cerebral water contents (P<0.01 or P<0.05) and cerebral index.Part II Mechanism of protection of QDPTP from cerebral ischemia/ reperfusion in rats1. The effects of QDPTP on oxidative damage and inflammatory reaction after global cerebral I/R in ratsA. The effects of QDPTP on cerebral SOD activities and MDA contents after global cerebral I/R in ratsThe model of global ischemia / reperfusion was established by improved Pulsinelli’s method for 20 minutes’occlusion and 24 hours’reperfusion. The activities of SOD and the contents of MDA in cerebral tissue after I/R were tested respectively. Comparing with NS in model group, QDPTP (450, 900mg/kg) lowered the contents of MDA (P<0.01) and reserved the activities of SOD(P<0.01). These effects of QDPTP (450, 900mg/kg) were comparable with Lig (33mg/kg).②The effects of QDPTP on IL1-βcontents of cerebral tissue and blood serum after global cerebral I/R in ratsIn the model group the IL1-βcontents of cerebral tissue and blood serum messured by ELISA were markedly increased after global cerebral I/R in rats. Comparing with the model group, the IL1-βcontents of cerebral tissue and blood serum decreased significantly in QDPTP (900mg/kg) group (P<0.01) and Lig (33mg/kg) (P<0.05) .2. The effects of QDPTP on cerebral blood flow and artery blood pressure in anesthetized ratsThe regional cerebral blood flow (rCBF) in cerebral pia mater was examined by Laser Doppler Flowmeter and the artery blood pressure (including mean systolic pressure, mean diastolic pressure and mean blood pressure) were monitored by the electrophysiological polygraph before and after drug administration. Comparing with the solvent, QDPTP (450, 900, 1800mg/kg), Lig (33mg/kg), Nim (20mg/kg) and DSDW (60mg/kg) expanded rCBF (P<0.01 or P<0.05) and simultaneously decreased the artery blood pressure (P<0.01 or P<0.05).Conclusion1. QDPTP had protective effects on the brain damaged by acute local and global cerebral ischemia/reperfusion in rats.2. QDPTP improved neurological function and decreased the infarction size and cerebral edema induced by acute local cerebral ischemia /reperfusion in rats.3. The mechanism of anti-cerebral injury of QDPTP in cerebral ischemia / reperfusion model may be related to the inhibition of the damages caused by oxidative free radicals, the extenuation of inflammatory reaction mediated by cytokines, the extension of cerebral vessels and the increase of cerebral blood flow.更多还原