脑红蛋白对创伤性脑损伤的保护作用研究.pdf

军医进修学院博士学位论文脑红蛋白对创伤性脑损伤的保护作用研究中文摘要脑红蛋白(Ngb)是2000年被发现的人体内第三类携氧球蛋白,主要位于脑组织内。随即人们对它是否对脑缺血缺氧性损害有保护作用及其作用机理产生强烈兴趣并进行了深入研究。目前多个研究显示:在脑缺血/缺氧时Ngb 的表达升高,而且通过转基因技术使脑内Ngb表达升高后可以显著减小脑梗死体积,这表明Ngb对脑缺血缺氧性损伤有保护作用。然而至今未见关于Ngb 是否对创伤性脑损伤有保护作用的研究报道。本课题的目的在于应用重组Ngb腺病毒载体使大鼠脑内高表达Ngb,通过观察大鼠脑创伤后的病理变化,探讨Ngb对创伤性脑损伤有无保护作用。第一部分,成功建立硬膜外撞击中度局灶性脑损伤模型,观察损伤后的病理变化并证实伤灶周围皮质及同侧海马CAl区Ngb的表达升高,确定上述区域为重点观察对象。第二部分,对重组腺病毒载体在大鼠脑内的转导效率进行研究,:腺病毒注射在胼胝体通过逆轴浆运输可以高效转柒伤侧皮质神经元;通过多点注射可以使伤侧CAl区全部被转染,表明腺病毒载体可以满足实验要求。第三部分,建立重组Ngb腺病毒载体,并通过细胞培养和动物实验,应用PCR、免疫组织化学和Western Blot技术证明它能高效转染细胞或大鼠脑组织且正确表达Ngb。第四部分,将重组Ngb腺病毒载体立体定向注入脑内,使Ngb高表达, 然后施以大鼠局灶性中度脑损伤。结果显示:损伤灶体积较对照组明显减小; 伤侧海马CAl区细胞丢失减少;凋亡细胞数明显减少。结论:脑组织内高表达Ngb可以减轻创伤性脑损伤的程度,即在创伤时 Ngb有神经保护作用。关键词:脑红蛋白:创伤性脑损伤;腺病毒载体;神经保护剂 3 军医进修学院博士学位论文 Neuroglobin Protects theBrain from Traumatic Inj ury Abstract Neuroglobin fNgb)is the tllirdkind of 02一binding globins,expressed preferentially in nervous ,it was speculated thatNgb maybe play a neuroprotected role inhypoxia tothepresent day,many studies revealed thattheexpression ofNgb Was increased aftertraumatic braininjury (Tin),and over expressing Ngb bytransgenic techniques could reduce thevolume ofbrain infarct theseresult support thestandpoint thatN’gb protect brain from ischemia or hypoxia ,there is no any research report on rolesofNgb inTBI until tonow. Our previous study showed thatexpression ofNgb aftertrauma Wasincreased significantly and relatedto someapoptosis suggests thatNgb may play aroleofneuroprotection in ,the aim ofthisstudy Wasto explore whether Ngb protects brain ,we Bse binant adenovirus vector containing GFP--tagged Ngb(Ad—·GFP-Ngb)to transfer Ngb gene ,TBI Was last,thechanges of injured brain pathology were assessed. establish Feeney’S weight—dropping model ofmoderate traumatic brain injury and observe thechanges ofexpression ofNgb foun