【作者】 樊鹏程；

【导师】 贾正平；

【作者基本信息】 第四军医大学 ， 药理学， 2013， 博士

【摘要】 研究背景我国高原面积辽阔，是世界上高原面积最大、边境线最长的国家。缺氧是高原环境医学中的首要问题。随着我国西部大开发战略的实施、青藏铁路的开通，越来越多的人到达高海拔地区。高原相关疾病正日渐成为一个严重影响公共健康的问题。因此，研发新型高效、低毒的抗高原缺氧损伤防治药物具有重要的现实意义。通过文献回顾及前期实验基础，课题组发现高原缺氧相关疾病的发生和体内自由基释放增加密切相关。由此提出研究假设：即高原缺氧导致机体氧化应激，线粒体氧化还原途径激活，抗氧化体系失代偿，自由基产生增加是导致高原相关疾病的主要原因之一，自由基清除剂类药物可能通过清除机体产生的过多自由基而对高原缺氧具有保护作用。研究目的1.通过研究高原缺氧动物模型，证实课题假设，阐明高原缺氧与体内自由基释放之间的关系，明确自由基清除剂对高原缺氧的保护作用。2.筛选出全新的抗高原缺氧自由基清除剂，并探讨其作用机制。3.研究兼具自由基清除和NO释放作用的硝酸酯类化合物的抗高原缺氧作用，并探讨其保护机制。实验内容及结果1.自由基清除剂的抗高原缺氧研究(1)4’-羟基-2-苯基咪唑啉自由基的抗高原缺氧研究通过常压密闭缺氧筛选试验对多个可能具有抗缺氧活性的化合物和中药提取物进行了筛选。选择了抗缺氧效果较为显著的咪唑啉类自由基清除剂4’-羟基-2-苯基咪唑啉自由基(HPN)进行抗高原缺氧药效及保护机制研究。结果显示HPN具有较好的抗高原缺氧作用，从而证实了课题假设并得出结论，即通过自由基清除剂类化合物对高原缺氧动物模型的干预可以保护机体免受缺氧引起氧化应激所致自由基产生增加的损伤，防止脂质过氧化，保护机体抗氧化酶系维持正常活性，维持线粒体能量代谢功能正常，稳定了线粒体膜，维持线粒体内氧化磷酸化的正常进行，维持Na/K-ATP酶活性，最终防止AMS及HACE的症状出现。以HPN为母核合成的咪唑啉类自由基化合物在常压密闭缺氧实验中也显示了较好的抗缺氧活性。(2) TEMPOL的抗高原缺氧研究对TEMPOL的抗高原缺氧作用进行了研究。结果显示TEMPOL同样具有较好的抗高原缺氧作用，能够保护海拔8000m实验动物的大脑及心脏。其保护机制同HPN相似，主要是通过清除过量产生的ROS，抑制脂质过氧化，保护机体抗氧化酶系维持正常活性。该结果再次证明了自由基清除剂类化合物具有抗高原缺氧作用，并首次证实了TEMPOL具有潜在的抗高原缺氧作用。2. NO供体乙酰阿魏酸单硝酸异山梨酯的抗高原缺氧研究对兼具自由基清除和NO释放作用的NO供体类化合物乙酰阿魏酸单硝酸异山梨酯(ACMI)进行抗高原缺氧药效及保护机制研究。结果显示，ACMI具有较好的抗高原缺氧作用，能够保护高海拔机体免受缺氧损伤。其机制为清除体内过多的自由基，维持了药物所释放的NO在体内的药理活性，抑制脂质过氧化的同时抑制了因自由基而导致的体内NO失活。以单硝酸异山梨酯为母核，合成的NO供体类自由基清除化合物，同样显示了较好的抗缺氧活性。本课题通过对两个氮氧自由基类化合物HPN、TEMPOL及NO供体类自由基清除化合物ACMI的抗缺氧作用及机制研究，证实了自由基清除剂可通过清除机体产生的过多自由基而对高原缺氧具有保护作用。同时筛选出了具有潜在药用价值的自由基清除剂及NO供体化合物，为进一步将其开发成为缺氧损伤防护药物奠定了基础，同时也为新型抗缺氧药物研发提供了实验依据和理论根据。更多还原

【Abstract】 BackgroundChina is the world’s largest plateau country with longest high altitude border. Our countryhas vast plateau area. Hypoxia is the most important issue in the plateau environmentalMedicine. With the implementation of the western development strategy in China and theopening of the Qinghai-Tibet Railway, more and more people reach high altitudes. Thehigh altitude related disease is becoming a serious impact on public health problem. Thus,the research and development of new high efficiency, low toxicity anti-altitude hypoxiaimpairment preventive drug has important practical significance.Base on the literature review and preliminary experimental results, the research groupfound the occurrence of high-altitude hypoxia related disease is closely related to theincrease of free radicals release in the body. Thus proposed research hypotheses:high-altitude hypoxia leads to oxidative stress, mitochondrial redox pathway activation,antioxidant system decompensation. increasing radical generating is one of the major factors that lead to high-altitude related diseases. Radical scavenger drugs may have aprotective effect through clearing the body free radicals on the plateau hypoxia.Research purposes1. Through the study of the high altitude anoxic animal models confirmed researchhypothesis, clarified the relationship between the plateau hypoxia and free radicalsrelease and confirmed the protective effect of free radical scavenger compounds toplateau hypoxia.2. Screening new anti high-altitude radical scavenger and discussing the mechanisms.3. Study on the anti-altitude hypoxia protective effect of both radical scavenging and NOrelease NO donor compounds and discussing the mechanisms.Experimental content and results1. Research for the anti-altitude hypoxia protective effect of radical scavenger(1) the anti-altitude hypoxia protective effect of4’-hydroxy-2-phenyl imidazoline radicalScreening the activities of multiple compounds and traditional Chinese medicineextracts which may have anti-hypoxia activity through of normobaric hypoxia tests.Imidazoline radical scavenger4’-hydroxy-2-phenyl-imidazoline radical (HPN) withmore significant anti-hypoxia effect was selected in the anti high-altitude hypoxiapharmacodynamic and protective mechanism study. The results showed that HPN hasgood anti high-altitude hypoxia effect. Thus confirming the subject’s assumptions andgetting concluded conclusion. Intervention in animal models of plateau hypoxiacompounds by radical scavenging agents can protect the body from hypoxia inducedoxidative stress caused by free radicals increased injury, preventing lipid peroxidation,protecting the body antioxidant enzyme system to maintain normal activity, maintainmitochondrial energy metabolism functioning properly, stabilizing the mitochondrialmembrane, maintaining mitochondrial oxidative phosphorylation to normal level,maintaining the Na/K-ATP enzyme activity and ultimately prevent AMS and HACEsymptoms. The HPN nucleus synthesized imidazolines radical compound also shows abetter anti-hypoxic activity in the normobaric hypoxia experiment. (2) the anti-altitude hypoxia protective effect of TEMPOLStudy on the anti Hypoxia effect of of TEMPOL. The results showed that TEMPOLalso had good resistant effect to high altitude hypoxia. TEMPOL can protectexperimental animals’ brain and heart under the altitude of8000m. The protectivemechanism of TEMPOL was similar with HPN, primarily by removing the excessiveproduction of ROS, inhibition of lipid peroxidation and protect the body’s antioxidantenzyme system to maintain normal activity. The results proved once again that theradical scavenger compounds has anti-altitude hypoxia effect, and for the first timeconfirmed the potential protective role of TEMPOL under anti-altitude hypoxia.2. The anti high-altitude hypoxia research of NO donor acetyl ferulic acid isosorbidemononitrateStudy on the anti-altitude hypoxia protective effect of NO donor compounds acetylferulic acid isosorbide mononitrate (ACMI) with both of free radical scavenging andNO release function and discussing the protection mechanisms. The results showedthat ACMI had good resistance to high-altitude hypoxia and could protect the bodyfrom high altitude hypoxic injury. Its proctive mechanism was cleaning excess freeradicals in the body, maintaining the drug release NO in vivo pharmacological activity,Inhibition of lipid peroxidation, while suppressing the inactivation of NO caused byfree radicals in vivo. Using isosorbide mononitrate as parent nucleus structure tosynthesis the radical scavenging NO donor compounds, also showed good anti-hypoxiaactivity.Through the research on anti-hypoxia effect and radical scavenging mechanism oftwo nitroxide radical compounds of HPN and TEMPOL, and the NO donor ACMI, thisissue confirmed that a free radical scavenger could have a protective effect on theplateau hypoxia by clearing too many free radicals produced by the body. Meanwhilethe issue filtered out the radical scavenging and NO donor compounds with potentialmedicinal value. The study laid the foundation for the further development of hypoxicinjury protection drugs, as well as provided an experimental basis and theoretical basisfor the new anti-hypoxia drug development.更多还原