COBRE: UL: PKC REGULATION OF REGENERATION ACROSS CSPG AFTER SCI

COBRE：UL：PKC SCI 后 CSPG 再生调节

• 批准号: 7381132
• 负责人: CHRISTOPHER B SHIELDS
• 金额: $ 23.34万
• 依托单位: UNIVERSITY OF LOUISVILLE
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-06-01 至 2007-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7381132
• 关键词: COBRE; UL; PKC; REGULATION; REGENERATION

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Glial scar formation after spinal cord injury (SCI) is central in the failure of axonal regeneration. Chondroitin sulfate proteoglycan (CSPG) is a class of major inhibitory molecules that is upregulated after SCI. We hypothesize that CSPG triggers PKC activation which induces a downstream activation of a small GTPase Rho and Rho-associated kinase (ROK) in sensory axons following SCI. We found that: 1) chondroitinase ABC, an enzyme that removes CSPG's glycosaminoglycan (GAG) side chains, had limited beneficial effect on DRG axon regeneration, 2) the Rho inhibitor bacterial exotoxin C3 transferase applied to the spinal cord was neurotoxic at high doses, 3) PKC phosphorylation is required for RhoA activation, and 4) inhibitation of conventional PKC isoforms resulted in neurite outgrowth in vitro and regeneration of DRG axons in vivo. Understanding interactions between PKC and Rho/ROK activation and how they affect neurite outgrowth in vitro and axonal regeneration in vivo are mechanistic issues related to CNS injury and regeneration. We will use contusion and laceration SCI models as well as KO transgenic mice to investigate effects of PKC and its downstream Rho/ROK inhibition on DRG axon regeneration. Aim 1 will characterize the effect of conventional PKC and Rho/ROK inhibition on DRG neurite outgrowth on a CSPG substrate using an in vitro cell culture system. Aim 2 will investigate neurite outgrowth of engrafted DRG neurons within a CSPG-enriched glial scar environment developed after a contusive SCI in mice. Aim 3 will explore axonal regeneration of engrafted DRGs towards a CSPG-enriched gradient developed after a dorsal spinal cord laceration injury in adult mice in vivo. Aim 4 will study sensory axon regeneration of host DRGs through and beyond a spinal cord dorsal laceration injury under various pharmacological treatments including the PKC and RhoA/ROK inhibitors. We hope that understanding regulation of PKC-Rho-ROK signaling pathway may provide new insights into axon regeneraton through a glial scar and provide new directions for SCI repair.

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。脊髓损伤(SCI)后胶质瘢痕的形成是轴突再生失败的核心。硫酸软骨素蛋白多糖(CSPG)是脊髓损伤后表达上调的一类主要抑制分子。我们假设CSPG触发PKC激活，从而诱导脊髓损伤后感觉轴突中一个小GTP酶Rho和Rho相关激酶(ROK)的下游激活。我们发现：1)去除CSPG糖胺聚糖(GAG)侧链的软骨素酶ABC对DRG轴突再生的有利作用有限；2)Rho抑制剂细菌外毒素C3转移酶在高剂量下对脊髓有神经毒性；3)PKC磷酸化是RhoA激活所必需的；4)抑制传统的PKC亚型导致体外突起生长和体内DRG轴突再生。了解PKC和Rho/ROK激活之间的相互作用及其对体外轴突生长和体内轴突再生的影响是与中枢神经系统损伤和再生相关的机制问题。我们将使用挫伤和撕裂伤脊髓损伤模型以及KO转基因小鼠来研究PKC及其下游的Rho/ROK抑制对DRG轴突再生的影响。目的1利用体外细胞培养系统研究常规PKC和Rho/ROK抑制对背根神经节突起生长的影响。目的2将研究在小鼠挫伤脊髓损伤后形成的富含CSPG的神经胶质瘢痕环境中移植的DRG神经元的轴突生长。目的3将探索植入背根节的轴突再生朝向CSPG丰富的梯度，在成年小鼠脊髓背部撕裂损伤后形成的体内。目的4将研究在包括PKC和RhoA/ROK抑制剂在内的各种药物治疗下，宿主DRG通过脊髓背部撕裂伤和超越脊髓背侧撕裂伤的感觉神经元轴突再生。我们希望通过了解PKC-Rho-ROK信号通路的调控，能够对神经胶质瘢痕的轴突再生提供新的认识，为脊髓损伤的修复提供新的方向。