Integrative Tissue Engineering in the Spinal Cord

描述（由申请人提供）：将在大鼠横断模型中评估治疗脊髓损伤的综合组织工程方法。以聚乳酸-羟基乙酸（PLGA）为原料，研制了一种具有可控微结构的可生物降解植入物。植入物的结构将通过扫描电子显微镜、共聚焦显微镜和x射线显微计算机断层扫描进行评估和表征。软骨素酶ABC (C-ABC)，一种被证明可以分解损伤导致的富含蛋白聚糖的疤痕的酶，将被包裹在PLGA微球中，并将在体外评估其生物活性和释放动力学。将C-ABC纳入聚合物种植体的技术将被研究，并将描述C-ABC在种植体中的分布和释放。雪旺细胞将从新生大鼠幼仔坐骨神经中获取，在细胞培养中扩增，并悬浮在Matrigel中，以纳入聚合物植入物。成年Sprague-Dawley大鼠在接受胸8-9椎板切除术和脊髓横断手术前将被麻醉。各种植入物将被插入椎管，硬脑膜将其封闭。脊髓将在多个时间点采集，纵向分裂，并固定用于组织学，免疫组织化学和扫描电子显微镜（SEM）。组织修复的形态学评估将在组织切片上进行，而功能分析将用于评估神经恢复。将对实验组和对照组进行评估，以确定每种植入成分及其各种组合对轴突再生和功能恢复的影响。

DESCRIPTION (provided by applicant): An integrative, tissue-engineering approach to treat spinal cord injury will be evaluated in a rat transection model. A biodegradable implant with controlled microarchitecture will be developed and fabricated from poly(lactic-co-glycolic acid) (PLGA). Implant architecture will be evaluated and characterized with scanning electron microscopy, confocal microscopy, and X-ray micro-computed tomography. Chondroitinase ABC (C-ABC), an enzyme shown to break down the proteoglycan-rich scar that results from injury, will be encapsulated in PLGA microspheres, and its bioactivity and release kinetics will be evaluated in vitro. Techniques for incorporating C-ABC into the polymer implant will be investigated, and the distribution and release of C-ABC from the implant will be described. Schwann cells will be harvested from neonatal rat pup sciatic nerves, expanded in cell culture, and suspended in Matrigel for incorporation into the polymer implant. Adult Sprague-Dawley rats will be anesthetized before receiving thoracic 8-9 laminectomies and spinal cord transections. Various implants will be inserted into the spinal canals and the dura closed over them. Spinal cords will be harvested at multiple time points, split longitudinally, and fixed for histology, immunohistochemistry, and scanning electron microscopy (SEM). Morphologic evaluation of tissue repair will be performed on tissue sections, while functional assays will be employed for assessment of neurological recovery. Experimental groups and controls will be evaluated for determining the effects of each implant component, as well as their various combinations, on axon regeneration and functional recovery.