【作者】 陈其昕；

【导师】 杨迪生；

【作者基本信息】 浙江大学 ， 外科学， 2002， 博士

【摘要】 研究背景资料 机械性的压迫及随后的继发性损伤可破坏脊髓内的精细结构而导致神经功能的丧失。以往的观点认为成年哺乳动物中枢神经系统损伤后将发生神经轴突的退变，而不会发生轴突的再生。近年来的研究则发现成年脊髓轴突在损伤后可通过生长锥样的轴突末端生长而再生。提出中枢神经系统的轴突具有内在再生潜能的观点。寻找中枢神经系统的轴突再生不如周围神经系统有效的原因，激发有效的脊髓神经再生已成为目前研究的主要焦点。 生长相关蛋白43(Growth associated protein 43，GAP43)是一种膜相关的钙调结合磷蛋白。目前的研究已经证明这种蛋白可大量的存在于神经轴突的生长锥中，在胚胎及新生大鼠的大脑和脊髓的生长神经元中也有大量的表达，同时在由神经生长因子分离的PC12细胞中也有高表达。在成年动物的中枢神经系统中GAP43表达可急剧下降并消失。但在大脑的一些高度集成区域如新皮质、海马回等仍有GAP-43的表达。上述观察结果提示GAP43的高表达与中枢神经系统的轴突生长及神经可塑性有关。近来的研究表明在神经受到机械性损伤后该蛋白及其mRNA可发生再次的高表达。 W。wit 47Hh 博士学位论文 魁搽 神经系统的功能主要依靠神经元之间以及神经元与周围靶细胞之间的特殊的 联接来完成。建立这一联接通路需要有突触前神经元的轴突生长，使其最终与突触 后神经元相联接。这一轴突的生长过程主要涉及到微管蛋白的装配和稳定。微管联 络蛋白IB（Microtubule－Associated Protein IB，MAPIB）则参与了微管蛋白的装 配和稳定。因此研究微管联接蛋白m的表达能够间接反映神经轴索的生长状态，从 而提示中枢神经系统中的轴索的生长。 神经细胞粘着分于（Neural Cell Adhesion Molecule，NCAN）是一种介导神 经元之间神经元与其他细胞之间粘着的糖蛋白。在神经系统发育过程中，对神经轴 突生长及轴突伸展的引导起着重要作用。试管内研究表明NCth主要集中分布在轴突 的生长锥及神经丝状突起（fi1Op。i。）中。有研究表明NCAM所介导的细胞间的粘 着效应在靶器官神经支配和随后的突触形成过程中具有定向选择性作用。在成年哺 乳动物脊髓的Rexed、11、X区仍能发现有NCW的表达，这一现象被认为与脊髓 的神经可塑性有关。 胆碱能神经元是脊髓内主要的神经细胞。因该类神经元内的主要功能成分乙酞 胆碱由胆碱乙酞转移酶（Choline Acetyltransferase，ChAT）合成，因此神经x 内ChAT活性改变能反应脊髓神经的传导功能。胆碱乙酞转移酶的活性可作为神经元 传导功能的重要指标。 根据损伤的类型，脊髓损伤可分为下列几种类型：脊髓震荡、脊髓挫伤、脊髓 压迫及脊髓撕裂伤。临床上以脊髓压迫伤最为常见。与脊髓损伤的横切动物模型比 较，脊髓压迫性损伤具有不同的神经生理学和神经病理学特点。临床上，脊髓减压 是用于治疗脊髓压迫损伤的常规方法。因此，可将脊髓压迫损伤分为以下2种类型： 持续性脊髓压迫，该类模型可代表减压前的脊髓受压状态；一过性脊髓压迫，可代 表脊髓减压后状态。临床上这2类脊髓压迫状态具有不同的临床特点和转归。然而 迄今为止，脊髓减压促进神经功能恢复的生物学机理尚未阐明。 本研究的目的： 1、探讨神经传导功能标志物胆碱乙酞化酶活性在脊髓压迫性损伤后神经功能 恢复中的可能的作用。 －3－ 加户人纱 乙巳b 博士学位论文 电枉汐 2、探讨生长相关因子GAP－43（生长相关蛋白43），微管联络蛋白＊（MAPIB）， 神经细胞粘着分子（NCAM）等神经再生相关物质的变化与神经功能恢复的关系。 3、研究脊髓减压对上述各种因子的影响，并探讨减压对脊髓压迫后神经功能 恢复的作用机理。 材料和方法 一、大鼠脊髓压迫摸型的制作和分组 体重为30H009的Wistar大白鼠用于本研究，共计大白鼠108只。豆％己戊 巴比妥腹腔内麻醉后，做以Th12为中心的后正中切口暴露椎板，切除ThlZ棘突后， 用磨钻制出直径为2．7n的孔，直达硬膜外，将一直经为3．orm，螺距为0．4llllll的螺 丝垂直旋入该孔。通过计算残留在椎板外的螺纹数控制螺丝进入椎管内更多还原

【Abstract】 Research BackgroundMechanical compression and secondary mechanisms may cause neurological deficit by influencing subtle structures of the cord. In the past, most investigators have considered that the axonal swellings in CMS trauma cases will degenerate. Furthermore, it has been thought that most axons in the CNS of mature mammals will fail to regenerate after trauma. However, recent studies have indicated that injured axons of the spinal cord and brain in fact may show some regenerative responses following injury by a way of growthconelike axonal terminals and aberrant axonal branching. Therefore, the idea is now proposed that axons of the CNS have an intrinsic potential for regeneration. A lot of efforts have been done to explore why axons of CNS cannot repair themselves as efficiently as those of the PNS and to find means by which CNS repair can be improved.Growth associated protein 43 (GAP-43) is a membrane-associated calmodulin-binding phosphoprotein present in the nervous system. It has been demonstrated that the protein is present in large quantities in the axonal growth cone, in outgrowing neurons in fetal and neonatal rat brain and spinal cord, and outgrowing processes of nerve growth factor differentiated PC 12 cells.Furthermore, the protein and its mRNA are re-expressed in regenerating axons following mechanical damage. Moreover, in the mature brain, GAP-43 expression declines sharply, but remains present in higher integrative areas of the brain such as the associative neocortex and the hippocampus. Taken together, these observations indicate that GAP-43 is associated with axonal out-growth and plasticity in the central nervous system (CNS).Microtubules have important structural and functional roles in neurons, particularly during development. They contribute to the generation and maintenance of the shape of nurites (undifferentiated axons and dendrites) and they provide structural support for fast axoplasmic transport. MAPI b is involved in microtubule assembly, cytoskeletal organization an organelle transport, and has recently been implicated in neurite outgrowth. The increased expression of MAP1B is thus considered as a marker for neurite regrowth in CNS.The neural cell adhesion molecule (NCAM), a cell surface glycoprotein, act as an homophilic ligand to promote cell to cell adhsion and thus take part I the regulation of cell-cell and cell-substrate inteactions. During axon outgrowth and axon extension in the developing nervous system, adhensive interactions between axon and glial cells o r between neurons have been shown to be mediated by NCAM. NCAM is highly concentrated in developing neural tissues, and in vitro studies show that it is localized in axonal growth cones and filopodia. In addition, NCAM mediated adhesive interactions have been shown to be important in very selective mechanisms such as target innervation and subsequent synapse formation. In some regions of the adult central nervous system, the embryonic form of NCAM could be detected, which might indicate the plasticity in these regions.It is well established that cholinergic neurons make up a significant proportion of the total neuronal pool in the spinal cord. The extent of spinal cord injury will thus be reflected by a reduction in the activity of choline acetyltransferase (ChAT), the enzyme that synthesizes acetylcholine. Theinjury-induced disappearance of ChAT only reflects a loss of such enzymes rather than actual neuronal death, and the rebound of ChAT activity after CNS injury is temporally associated with the partial recovery of function. Choline acetyitransferase can therefore be used as definitive marker for neuronal function in spinal cord injury.In contrast to experimental transection models, spinal cord compression demonstrates different neurophysiologic and neuropathologic features. Clinically, decompressive surgery is considered routine therapy for spinal cord compression. Accordingly, two types of spinal cord compression were analyzed: continuously compressed spinal cord, which could repre更多还原