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价治疗脊髓损伤的综合性组织工程方法。一种具有可控微结构的生物可降解植入物将由聚乳酸-羟基乙酸共聚物（PLGA）开发和制造。将使用扫描电子显微镜、共聚焦显微镜和X射线显微计算机断层扫描对植入物结构进行评价和表征。软骨素酶ABC（C-ABC），一种被证明能分解由损伤引起的富含蛋白聚糖的瘢痕的酶，将被封装在PLGA微球中，并将在体外评价其生物活性和释放动力学。将C-ABC纳入聚合物植入物的技术进行了研究，并将描述C-ABC从植入物的分布和释放。将从新生大鼠幼仔坐骨神经中收获许旺细胞，在细胞培养物中扩增，并悬浮在基质胶中以掺入聚合物植入物中。成年Sprague-Dawley大鼠将在接受胸部8-9椎板切除术和脊髓横断术之前麻醉。各种植入物将被插入椎管内，并在其上封闭硬脑膜。将在多个时间点采集脊髓，纵向切开并固定，用于组织学、免疫组织化学和扫描电子显微镜（SEM）检查。将在组织切片上进行组织修复的形态学评价，而功能测定将用于评估神经恢复。将对实验组和对照组进行评价，以确定每种植入物组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.