Role of Myelin in Spinal Cord Regeneration

描述（由申请人提供）：成人中枢神经系统轴突再生的主要障碍之一是髓磷脂相关抑制剂，如MAG。然而，如果神经元cAMP水平升高，在培养和体内的抑制性环境中，可以通过类似物如db cAMP或通过神经营养素启动神经元来促进轴突生长。自发性中枢神经系统轴突再生发生的一种情况是背根神经节轴突，如果同一神经元的外周分支，背根神经节（DRG）神经元，在一周前受损-条件反射损伤（CL）效应，这是cAMP依赖的。cAMP和CL的作用都依赖于转录，其中一个被上调的基因是精氨酸酶I （Arg I）的基因，这是合成多胺的关键。多胺亚精胺能在体外克服MAG的抑制作用，促进体内视神经再生。此外，亚精胺通过抑制真核起始因子5A （eIF5A）激活CDK5激酶，从而增加CDK5激活因子p35的翻译，从而促进再生。在Aim 1a中，多胺激活的CDK5底物将被鉴定并表征其在体内克服抑制和促进再生中的作用。Aim 1b将解决神经营养因子和MAG信号通路之间的串扰，重点关注MAG阻断神经营养因子激活小GTPase Rap1的能力。强有力的初步数据表明，cAMP和CL效应都需要轴突的局部翻译来促进抑制环境下的再生。在Aim 2中，将确定在CL和db cAMP治疗后过程中增加的mRNA和microrna。在Aim 3中，在这两种情况下增加的mrna和microrna将被表征（过表达，敲低），以在培养物中促进MAG存在的再生和在体内促进再生中可能发挥作用。通过本提案中描述的实验，我们不仅会对已知促进体内再生的药物的作用机制有更深入的了解，而且还会发现新的药物。这反过来将揭示新的干预目标和药物开发，以促进体内轴突再生。

DESCRIPTION (provided by applicant): One of the major obstacles to axonal regeneration in the adult CNS is inhibitors associated with myelin, such as MAG. However, axonal growth can be encouraged in an inhibitory environment both in culture and in vivo if the neuronal cAMP levels are elevated, either with analogues such as db cAMP or by priming neurons with neurotrophins. One situation where spontaneous CNS axon regeneration does occur is of dorsal root ganglion axons if the peripheral branch of the same neuron, the dorsal root ganglion (DRG) neuron, is lesioned one week before - the conditioning lesion (CL) effect, which is cAMP dependent. Both the cAMP and the CL effects are dependent on transcription and one gen that is up-regulated is for the enzyme Arginase I (Arg I), which is key in the synthesis of polyamines. The polyamine, spermidine, can overcome inhibition by MAG in culture and promotes optic nerve regeneration in vivo. Furthermore, spermidine promotes regeneration by activating the kinase CDK5 by hypusinating the eukaryotic initiation factor 5A (eIF5A), resulting in an increase in translation of the CDK5 activator p35. In Aim 1a the CDK5 substrates that are activated in response to polyamine will be identified and characterized for their role in overcoming inhibition and promoting regeneration in vivo. Aim 1b will address the cross-talk between the neurotrophin and MAG signaling pathway, focusing on the ability of MAG to block the activation of the small GTPase, Rap1, by neurotrophin. Strong preliminary data suggest that both the cAMP and CL effects require local translation in the axon to promote regeneration in an inhibitory environment. In Aim 2 mRNA and microRNAs that increase in the processes after both a CL and treatment with db cAMP will be identified. In Aim 3 those mRNAs and microRNAs that increase after both conditions will be characterized (over-expression, knock-down) for a possible role in promoting regeneration in the presence of MAG in culture and in promoting regeneration in vivo. Through the experiments described in this proposal not only will our understanding be advanced of the mechanism of action of agents known to promote regeneration in vivo but novel agents will be identified. This in turn will reveal novel targets for intervention and drug development to promote axonal regeneration in vivo.