白藜芦醇甙对大鼠局灶性脑缺血的保护作用(英文)

Protective effects of polydatin on focal cerebral ischemia in rats

背景:脑缺血时大量的自由基生成,使脑内脂质过氧化作用加强,导致细胞及细胞屏障损害,神经元坏死或凋亡,引起和加重缺血脑组织水肿。目的:通过观察白藜芦醇甙对大鼠局灶性脑缺血后脑组织自由基、脂质过氧化物含量、脑含水量及病理形态学的影响,探讨其对脑缺血的保护作用。设计:随机对照实验。单位:重庆医科大学附属第二医院ICU和重庆医科大学附属儿童医院儿科医学研究所。材料:实验于2001-10／2002-07在重庆医科大学儿科医学研究所完成。将48只健康成年雄性Wistar大鼠随机分为3组,每组16只。缺血前白藜芦醇甙处理组:缺血前30 min,经颈外静脉注射6g/L白藜芦醇甙(12 mg／kg)溶液。假手术组:大鼠仅在麻醉状态下分离右侧颈总动脉, 不予阻断血流。缺虚模型组:建立大鼠右侧大脑中动脉梗死模型。假手术组、缺血模型组与缺血前白藜芦醇甙处理组以同样方式、剂量静脉给予生理盐水。大鼠大脑中动脉阻塞后2 h在麻醉状态下快速断头取脑。每组随机选取8只大鼠测定脑组织含水量,其余8只大鼠测定脑组织自由基及脂质过氧化物含量。方法:应用干-湿重法测脑组织含水量,以硫代巴比妥酸法检测丙二醛水平、黄嘌呤氧化酶法检测超氧化物歧化酶活性、化学比色法测定谷胱甘肽过氧化物酶活性、化学比色法检测一氧化氮合酶活性,以比色法测定过氧化氢酶活性,并按Folin-酚试剂法标定蛋白含量,所有操作均严格按说明书进行。主要观察指标:①各组大鼠脑组织含水量。②各组大鼠脑组织中丙二醛含量,超氧化物歧化酶、谷胱甘肽过氧化物酶、过氧化氢酶及一氧化氮合酶活性。③各组大鼠脑组织病理学观察。结果:48只大鼠均进入结果分析。①缺血前白藜芦醇甙处理组脑组织中超氧化物歧化酶、谷胱甘肽过氧化物酶、过氧化氢酶活性明显高于缺血模型组[(226．43±8．69),(193．37±11．14)NU／mg;(244．38±12.34), (211．71±16．50)μkat／g;(59．85±9．67),(35．51±7．67)μkat/g,(q=4.38-10．45, P<0.01)]。②缺血前白藜芦醇甙处理组脑组织中丙二醛含量、脑含水量明显低于缺血模型组[(6．38±0．54),(8．63±0．78)μmol/g;(78．72±0．43), (80．41±0.64),％,P<0．01]。③白藜芦醇甙有降低缺血脑组织中一氧化氮合酶活性的趋势,但与缺血模型组非常接近[(12．00±1．00),(12．84±1．17)μkat/g, P>0．05]。④病理学组织检查显示,白藜芦醇甙能减轻缺血所导致的脑水肿。结论:白藜芦醇甙能减轻脂质过氧化反应,降低缺血脑组织中丙二醛含量,提高体内超氧化物歧化酶、谷胱甘肽过氧化物酶、过氧化氢酶活性, 抑制或减轻脑水肿形成,保护细胞膜功能,减轻缺血神经元功能损害, 对局灶脑缺血性脑损伤具有明显的保护作用。

BACKGROUND： Free radicals are produced during ischemia, which can strengthen activity of lipid peroxidation; induce lesion of cell and cellular barrier, result in necrosis or apoptosis of neurons; and aggravate edema of ischemic cerebral tissue. OBJECTIVE： To observe the effects of polydatin （PD） on free radicals, lipid peroxidation, water contents and pathological morphology of brain tissue in rats with focal cerebral ischemia so as to explore its protective mechanisms. DESIGN： A randomized controlled trial. SETTING： Intensive Care Unit, Second Affiliated Hospital, Chongqing University of Medical Sciences; Pediatric Research Institute, Children＇s Hospital Chongqing University of Medical Sciences. MATERIALS： The experiment was carried out at the Pediatrics Medicine Institute of Chongqing Medical University from October 2001 to July 2002. Totally 48 healthy adult male Wistar rats were divided into 3 groups randomly, with 16 in each group. Group Ⅰ was sham-operated group： rats were anaesthetized, the fight common carotid arteries were separated instead of being occluded. Group Ⅱ was ischemia group： to establish the fight middle cerebral artery occlusion model of rats. Group Ⅲ was PD pretreatment group： polydatin （6 g/K 12 mg/kg） were intravenously administrated 30 minutes before the onset of ischemia. Saline substituted for PD, besides, were intravenously administrated with the same way and dosage on Group Ⅰ, Group Ⅱand GroupⅢ. The rats were decapitated and the brains were immediately removed after cerebral ischemia 2 hours. In each group, 8 rats were chosen to be determined water contents of brain tissue, the other 8 rats were chosen to be determinedlevels of lipid peroxidation and free radicals in brain tissue. METHODS： According to the formula which was： wet weight-dry weight/ wet weight×100%, water content of cerebral tissue was assayed. Superior liquid was taken to assay MDA with spectrophotometer thiobarbituric acid method （TBA） method, SOD activity assayed by xanthine oxidase enzyme method, the activities of GSH-Px, CAT and NOS determined by colorime- try ,the amount of protein determined by the method of Lowry. All the procedures were carried out strictly according to the instruction. MAIN OUTCOME MEASURES：①Brain water content.② Level of malonaldehyde （MDA） and activities of superoxide dismutase （SOD）, glutathione peroxidase （GSH-Px）, catalase （CAT） and nitric oxide synthase （NOS）. ③ Pathological sections of brain tissue were observed. RESULTS： Data of totally 48 rats was entered the analysis. 0） Before ischemia, contents of SOD, GSH-Px and CAT in cerebral tissue of PD group were obviously higher than those of ichemia model group [（226.43±8.69）, （193.37±11.14） NU/mg; （244.38±12.34）, （211.71±16.50）μkat/g; （59.85±9.67）, （35.51±7.67）μkat/g, q=4.38-10.45, P 〈 0.01].② Contents of MDA and water in cerebral tissue of PD group were obviously lower than those of i- chemia model group [（6.38±0.54）, （8.63±0.78）μmol/g; （78.72±0.43）%, （80.41±0.64） %, P 〈 0.01].③PD had the tendency of decrease NOS activity in ischemic tissue but the results were similar to that in ischemia model group [（12.00±1.00）, （12.84±1.17）μkat/g, P 〉 0.05] in brain tissue. ④ The observation on the pathological morphology of brain tissue indicated that PD alleviated the ischemia edema of cerebral ischemia. CONCLUSION： PD can alleviate the reaction of lipid overoxidation, improve the activities of antioxidant-enxymes, reduce ischemia brain edema, protect the function of cell member, bring down the damage to ischemia neurons. It shows that PD has significant cerebral protective role on focal ischemia brain damage.