Despite the success of tissue engineered nerve guidance conduits (NGCs) for the treatment of small peripheral nerve injuries, autografts remain the clinical gold standard for larger injuries. The delivery of neurotrophic factors from conduits might enhance repair for more effective treatment of larger injuries but the efficacy of such systems is dependent on a safe, effective platform for controlled and localised therapeutic delivery. Gene therapy might offer an innovative approach to control the timing, release and level of neurotrophic factor production by directing cells to transiently sustain therapeutic protein production in situ. In this study, a gene-activated NGC was developed by incorporating non-viral polyethyleneimine-plasmid DNA (PEI-pDNA) nanoparticles (N/P 7 ratio, 2μg dose) with the pDNA encoding for nerve growth factor (NGF), glial derived neurotrophic factor (GDNF) or the transcription factor c-Jun. The physicochemical properties of PEI-pDNA nanoparticles, morphology, size and charge, were shown to be suitable for gene delivery and demonstrated high Schwann cell transfection efficiency (60±13%) in vitro. While all three genes showed therapeutic potential in terms of enhancing neurotrophic cytokine production while promoting neurite outgrowth, delivery of the gene encoding for c-Jun showed the greatest capacity to enhance regenerative cellular processes in vitro. Ultimately, this gene-activated NGC construct was shown to be capable of transfecting both Schwann cells (S42 cells) and neuronal cells (PC12 and dorsal root ganglia) in vitro, demonstrating potential for future therapeutic applications in vivo.

Statement of Significance

The basic requirements of biomaterial-based nerve guidance conduits have now been well established and include being able to bridge a nerve injury to support macroscopic guidance between nerve stumps, while being strong enough to withstand longitudinal tension and circumferential compression, in addition to being mechanically sound to facilitate surgical handling and implantation. While meeting these criteria, conduits are still limited to the treatment of small defects clinically and might benefit from additional biochemical stimuli to enhance repair for the effective treatment of larger injuries. In this study, a gene activated conduit was successfully developed by incorporating non-viral nanoparticles capable of efficient Schwann cell and neuronal cell transfection with therapeutic genes in vitro, which showed potential to enhance repair in future applications particularly when taking advantage of the transcription factor c-Jun. This innovative approach may provide an alternative to conduits used as platforms for the delivery neurotrophic factors or genetically modified cells (viral gene therapy), and a potential solution for the unmet clinical need to repair large peripheral nerve injury effectively.

中文翻译：

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掺有携带NGF，GDNF和c-Jun编码基因的非病毒性PEI-pDNA纳米粒子的基因激活神经引导导管的体外功效

尽管组织工程化的神经引导导管（NGC）在治疗周围神经小损伤方面取得了成功，但自体移植仍是较大损伤的临床金标准。从导管中输送神经营养因子可能会增强修复效果，以更有效地治疗较大的损伤，但此类系统的功效取决于安全，有效的平台，可控制并进行局部治疗。基因疗法可能通过指导细胞瞬时维持原位治疗性蛋白的产生来提供控制神经营养因子产生的时间，释放和水平的创新方法。在这项研究中，通过将非病毒性聚乙烯亚胺质粒DNA（PEI-pDNA）纳米颗粒（N / P 7比，2μg剂量）与编码神经胶质来源的神经生长因子（NGF）的pDNA结合，开发了一种基因激活的NGC。神经营养因子（GDNF）或转录因子c-Jun。PEI-pDNA纳米粒子的理化性质，形态，大小和电荷被证明适合基因传递，并在体外表现出很高的雪旺氏细胞转染效率（60±13％）。尽管所有三个基因在增强神经营养细胞因子产生，促进神经突向外生长方面均显示出治疗潜力，但编码c-Jun的基因的传递显示出最大的增强体外再生细胞过程的能力。最终，该基因活化的NGC构建体被证明是能够转染两个雪旺氏细胞（S42细胞）和神经元细胞（PC12和背根神经节）的体外，表明潜在为未来的治疗应用在体内。

重要声明

基于生物材料的神经引导导管的基本要求现已得到很好的确立，其中包括能够桥接神经损伤以支持神经残端之间的宏观引导，同时除了机械上健全外，还应足以承受纵向张力和圆周压缩。以便进行外科手术处理和植入。在满足这些标准的同时，导管仍局限于临床上对小缺陷的治疗，并且可能会受益于额外的生化刺激来增强修复，以有效治疗较大的损伤。在这项研究中，通过将能够有效地将雪旺氏细胞和神经元细胞转染治疗性基因的非病毒纳米粒子与体外治疗性基因结合，成功开发了基因激活导管 显示了在未来应用中增强修复的潜力，尤其是在利用转录因子c-Jun的情况下。这种创新的方法可以为用作输送神经营养因子或转基因细胞（病毒基因疗法）的平台的导管提供替代方案，并为尚未满足的有效修复大型周围神经损伤的临床需求提供潜在的解决方案。