Promoting axon regeneration by activation of the Smad1 signaling pathway

描述(申请人提供)：脊髓损伤会切断大脑和脊髓之间的连接，导致灾难性的后果，如损伤部位以下的感觉和运动控制丧失。造成永久性神经缺陷的原因是成年哺乳动物的神经元在损伤后不能再生。成功再生轴突的一个策略是增强受损神经元的内在轴突生长能力。这需要定义可以促进轴突生长的分子途径，并建立治疗方法来启动它们。最近，我们已经证明了转录因子Smad1在体外将成年感觉神经元转换到活跃的生长状态中的关键作用。这项建议侧重于利用脊髓损伤的啮齿动物模型来研究Smad1通路的体内功能。我们建议检验中枢假说，即在成人感觉神经元中激活Smad1促进上行感觉通路的轴突再生。这一假设得到了我们的初步研究的支持，这些研究表明，在DRG中，Smad1被激活，从而实现了强大的感觉神经轴突再生。这是通过我的实验室专门为活体脊髓研究设计的微创和临床适用的体内基因操作方法实现的。基于这些令人兴奋的结果，Aim 1将研究DRG神经元中Smad1的激活机制。这包括骨形态发生蛋白依赖和非依赖途径。一种新的激酶，Mps1，在调节轴突生长中的作用，以及Smad1-在连接区和C末端的差异磷酸化的功能将被研究。将确定Smad1的下游目标。目的2利用啮齿动物脊髓损伤模型进行体内实验，比较BMP依赖和非依赖途径激活Smad1的再生表型。AAV-BMP的治疗潜力将通过在损伤后注射AAV来进一步评估。综上所述，这些研究有望为操纵Smad1通路对脊髓损伤后轴突再生的治疗潜力提供见解。此外，该建议还旨在确定激活Smad1或作为下游效应器的新的分子靶点。最后，体内研究将验证一种微创和临床适用的治疗方法，这种方法可以很容易地转化为对脊髓损伤患者的临床试验。 公共卫生相关性：脊髓损伤会导致感觉和运动功能丧失。然而，到目前为止，还没有临床治疗方法来促进轴突的功能再生。本文提出的研究有望为操纵Smad1通路促进轴突再生的治疗潜力提供重要的见解，并验证一种临床上相关且易于翻译的治疗策略，以促进脊髓损伤患者的功能恢复。

DESCRIPTION (provided by applicant): Spinal cord injury severs the connections between the brain and the spinal cord, which causes devastating outcomes such as loss of sensory and motor controls below the lesion site. The reason behind the permanent neurological deficit is that adult mammalian neurons fail to regenerate after injury. One strategy for successful axon regeneration is to enhance the intrinsic axon growth capability of injured neurons. This requires defining molecular pathways that can promote axon outgrowth and establishing therapeutic approaches to turn them on. Recently, we have demonstrated a key role of Smad1, a transcription factor, in switching adult sensory neurons into an active growth state in vitro. This proposal focuses on the in vivo function of the Smad1 pathway using rodent models of spinal cord injury. We propose to test the central hypothesis that activation of Smad1 in adult sensory neurons promotes axon regeneration of ascending sensory pathway. This hypothesis is supported by our pilot studies that demonstrate robust sensory axon regeneration with activation of Smad1 in DRGs. This was achieved using a minimally invasive and clinically applicable in vivo gene manipulation method designed in my laboratory specifically for in vivo spinal cord research. Based on these exciting results, Aim 1 will investigate the Smad1 activation mechanisms in DRG neurons. This includes BMP-dependent and -independent pathways. The role of a novel kinase, Mps1, in mediating axonal growth and the function of the differential phosphorylation of Smad1-at the linker area and C- terminus will be studied. The downstream targets of Smad1 will be identified. Aim 2 focuses on in vivo assays using rodent models of spinal cord injury to compare the regenerative phenotypes of Smad1 activation by BMP-dependent and -independent pathways. Therapeutic potential of AAV-BMP will be further evaluated by injecting AAV after injury. In summary, these studies are expected to provide insights into the therapeutic potential of manipulating the Smad1 pathway on axon regeneration after spinal cord injury. Furthermore, this proposal also aims at identifying novel molecular targets that activate Smad1 or act as downstream effectors. Finally the in vivo studies will validate a minimally invasive and clinically applicably therapeutic approach that can be readily translated into clinical trials for spinal cord injury patients. PUBLIC HEALTH RELEVANCE: Spinal cord injury results in loss of sensory and motor function. Yet, to date, no clinical treatment exists to promote functional regeneration of axons. The studies proposed here are expected to provide important insights into the therapeutic potential of manipulating the Smad1 pathway on axonal regeneration, and to validate a clinically relevant and readily translatable therapeutic strategy to promote functional recovery in spinal cord injury patients.