【作者】 李富盛；

【导师】 贾正平； 张汝学；

【作者基本信息】 兰州大学 ， 生物化学与分子生物学， 2009， 硕士

【摘要】 目的筛选淫羊藿中具有抗缺氧作用的活性部位,研究其抗缺氧药理作用和机制。方法1.采用常压密闭缺氧小鼠模型,从淫羊藿水提物、淫羊藿醇提物和淫羊藿总黄酮中筛选淫羊藿中抗缺氧活性部位,再利用急性减压缺氧模型和免疫抑制模型,研究其抗缺氧药理作用。2.研究淫羊藿抗缺氧活性部位抗缺氧的药理作用机制。采用减压缺氧小鼠模型,清洁级雄性BALB/C小鼠随机分为:正常对照组,HYM组,药物低、中、高组和阳性药对照组,正常对照组和HYM组均给予等体积蒸馏水,药物组的给药剂量分别为300mg·kg^-1·d^-1、600 mg·kg^-1·d^-1和900 mg·kg^-1·d^-1,以上均为灌胃（i.g.）给药。分别从心、脑、肺组织的保护作用,糖代谢,神经内分泌系统等方面研究药理作用机制,所选指标包括:缺氧小鼠的存活时间,大脑、心肌和肺的含水量,大脑和心肌的SOD活性、MDA含量和乳酸含量,肝糖原、肌糖原,肝脏G-6-Pase活性和GK活性,血浆皮质酮、雌二醇和睾酮的含量,外周血象指标和脏器指数,制作大脑、心肌、肾上腺和睾丸组织的病理切片,观察病理变化。结果1.淫羊藿水提物能明显延长缺氧小鼠的存活时间。对淫羊藿的水提物、醇提物和总黄酮的药效对比时,淫羊藿总黄酮（TFE）能显著延长缺氧小鼠的存活时间（P＜0.05）。在做TFE抗缺氧量效关系实验时,给药剂量为900 mg·kg^-1·d^-1的高剂量组的缺氧小鼠存活时间最长;从外周血象的指标来看,TFE能使缺氧小鼠的血红细胞数量和血红蛋白含量增加,说明淫羊藿总黄酮能改善缺氧小鼠的血液成分,提高缺氧小鼠的血液携氧能力;TFE能使血白细胞和淋巴细胞的数量增加,说明淫羊藿总黄酮能提高缺氧小鼠免疫功能。给药剂量为900 mg·kg^-1·d^-1的TFE高剂量组的效果最好。2.TFE能使缺氧小鼠心、脑SOD活性升高,MDA含量下降,乳酸含量下降;脑、心肌和肺含水量的降低。病理学实验结果也表明TFE能够保护缺氧时心、脑组织的细胞结构,减轻细胞水肿。药效最佳的给药剂量为900 mg·kg^-1。3.TFE可显著升高缺氧时小鼠空腹血糖,可使肝糖原水平降低,G-6-Pase活性升高,GK活性降低,脑、心肌蛋白质含量升高。4.在神经内分泌方面,TFE可抑制应激状态下HPA轴的功能,显著降低血浆CORT的含量,肾上腺指数提高,病理实验结果也表明TFE能保护缺氧小鼠的肾上腺组织。TFE可以发挥性激素样作用,调节HPG轴功能,血浆睾酮、雌二醇含量降低,病理学实验结果也表明TFE能保护缺氧小鼠的睾丸组织。药效最佳的给药剂量为900 mg·kg^-1。5.在免疫系统方面,TFE能升高CTX致免疫抑制小鼠的免疫器官指数和外周血白细胞和淋巴细胞数目。药效最佳的给药剂量为900 mg·kg^-1。结论1.淫羊藿具有良好的抗缺氧作用。2.淫羊藿抗缺氧小鼠缺氧应激的主要药效部位是淫羊藿总黄酮,并发现在药物有效的、合理的使用范围内,给药剂量与药效呈线性关系。3.淫羊藿总黄酮抗缺氧机制可能包括:改善缺氧小鼠的心、脑自由基代谢,改善缺氧小鼠心肌、大脑和肺脏的水盐代谢,对心、脑和肺起到保护作用;改善缺氧时的物质代谢水平,以糖代谢为主,减缓蛋白质代谢,保证重要脏器的能量供应,这可能是其对急性缺氧时小鼠心、脑和肺脏组织起保护作用的物质代谢基础。4.淫羊藿总黄酮对缺氧状态下激活的HPA轴显示了一定的抑制作用;TFE能发挥性激素样作用,对缺氧状态下的HPG轴具有调节作用;同时对CTX致免疫抑制小鼠体液免疫有较强的改善作用。这说明淫羊藿总黄酮可能通过神经内分泌免疫网络调节机制减轻缺氧造成的损伤。5.淫羊藿总黄酮对体液免疫及非特异性免疫功能有较强的改善作用。更多还原

【Abstract】 Objective:To screen the anti-hypoxia active components of Epimedium Herb（HE）,study the anti-hypoxia pharmacological effect and explore its mechanisms.Methods:The anti-hypoxia active component of Epimedium Herb was screened by the closed anoxic model under the condition of ordinary.The mechanisms of its effect were studied by the low pressure and immunization models in mice,respectively.The mechanism of the active components of HE was investigated by the acute anoxic model under the low pressure conditions similar to the plateau hypoxia environment in BALB/C mice,and according to the purpose of the experiment,the mice were divided into four groups,i.e.normal control group,HYM group,drug-treated group and positive control group.All drugs were administered by intragastric administration（i.g.）.The mechanism of the active components of HE was systematically evaluated by determining the following indicators.The content of water in cerebral,myocardium and lung tissues;SOD activity,lactic acid,malondialdehyde（MDA） and protein content in both cerebral and cardiac tissues;liver glycogen and muscle glycogen contents,GK and G-6-P activities in liver;levels of plasmatic corticosterone,plasma glucose level and plasmatic testosterone,estradiol peripheral hemogram and organ index;pathological sections of cerebral, myocardium,adrenal and testis tissues.The mechanism of the active components of HE was also studied by the protective effects on heart,brain and lung,the regulatory effects on glucose and protein metabolism,and the improving effects on nerve-endocrine-immune system. Results:The survival time of mice under ordinary pressure was longer in aqueous extracts of Epimedium Herb treated groups than that of blank control group（P＜0.05）.TFE had a similar effect on increasing the survival time of mice than that of aqueous and alcohol extracts of HE. The mice treated with the high-dose of TFE（900 mg·kg^-（1）·d^-（1）） displayed the longest survival time.The treatment of TFE significantly increased the contents of red blood cell,white blood cell and lymphocytes,and the hemoglobin content as well.In the dose-effect relationship study of TFE anti-hypoxia,high-dose of TFE(900 mg·kg^-1·d^-1) showed the most significantly increasing effects.After the administration of TFE,SOD activity increased,while MDA and lactic acid content decreased in cerebral and cardiac muscle tissue.Meanwhile,plasma glucose level,the liver G-6-Pase activity and protein content in both cerebral tissue and cardiac tissue increased,while liver glycogen content,GK activity and the content of water in cerebral tissue,myocardium and lung tissue decreased when the mice were exposed to hypoxia.The pathological study proved that there was some damage in cerebral and cardiac muscle tissue,which was protected by the treatment of TFE in hypoxia mice.Compared with hypoxia model group,after the administration of TFE,the function of HPA axis was inhibited,the level of plasma corticosterone decreased and organ index of adrenal increased in hypoxia stress mice.The pathological study proved that there was some damage in adrenal tissue,which had been protected by the treatment of TFE in hypoxia stress mice.TFE was also proved to have the effect similar to sex hormone.After the administration of TFE,the function of HPA axis was regulated,the content of plasma estrogen and testosterone increased.The pathological study also demonstrated that testis tissue was damaged while protected by treatment of TFE in hypoxia stress mice.Compared with hypoxia model group,TFE increased immune organ index,leukocyte and lymphocyte numbers of immunosuppressive mice induced by cyclophosphamide.High-dose ofTFE(900mg·kg^-1·d^-1) had the best anti-hypoxia stress effect among the three groups. Conclusion:Epimedium Herb has significant anti-hypoxia pharmacological effect.TFE is the active components of Epimedium Herb in the antihypoxia effect.On the reasonable range of dosage,it showed linear dose-effect relationship.TFE can improve the metabolism of free radical,water and salt content in cerebral and cardiac muscle tissues.It can also reduce edema degree,and thus plays an important role in the protection for cerebral,cardiac and lung in hypoxia mice.TFE can also regulate the metabolism of substance,increase the speed of glucose metabolism,slow down protein metabolism and increase the fasting blood glucose level to supply energy to critical organ protecting the heart and brain of hypoxia mice.TFE had inhibitory effect on HPA axis of hypoxia mice.It acted as a sex hormone for the regulation of HPG axis function and could improve the immune function of immunosuppressive mice induced by cyclophosphamide,suggesting that TFE reduced injury ofhypoxia mice by the mechanism of NIM.TFE can improve blood component,increase the oxygen-carry ability of blood,the number of white blood cell and lymphocyte cell and the immunological competence in hypoxia stress mice.Hence,TFE effectively improve the functions of humoral immunity and nonspecific immuuity.更多还原