Kim Soung-Min, Lee Suk-Keun, Lee Jong-Ho
Department of Oral and Maxillofacial Surgery, College of Dentistry, Kangnung National University, Gangneung, South Korea.
J Craniofac Surg. 2007 May;18(3):475-88. doi: 10.1097/01.scs.0000249362.41170.f3.
The use of artificial nerve conduit containing viable Schwann cells is one of the most promising strategies to repair peripheral nerve injury. To fabricate an effective nerve conduit whose microstructure and internal environment are more favorable in nerve regeneration than those currently existing, a new three-dimensional (3D) Schwann cell culture technique using Matrigel and dorsal root ganglion (DRG) was developed. Nerve conduit of 3D arranged Schwann cells was fabricated using direct seeding of freshly harvested DRG into Matrigel-filled silicone tubes (inner diameter 1.98 mm, 14 mm length) and in vitro rafting culture for 2 weeks. The nerve regeneration efficacy of 3D cultured Schwann cell conduit (3D conduit group, n = 6) was assessed using an Sprague-Dawley rat sciatic nerve defect of 10 mm and compared with that of a silicone conduit filled with Matrigel and Schwann cells prepared with the conventional plain culture method (two-dimensional [2D] conduit group, n = 6). After 12 weeks, sciatic function was evaluated with sciatic function index (SFI) and gait analysis, and histomorphology of nerve conduit and the innervated tissues of sciatic nerve were examined using image analyzer and electromicroscopic methods. The SFI and ankle stance angle in the functional evaluation were -60.1 +/- 13.9, 37.9 degrees +/- 5.4 degrees in the 3D conduit group (n = 5) and -87.0 +/- 12.9, 32.2 degrees +/- 4.8 degrees in the 2D conduit group (n = 4). The myelinated axon was 44.91% +/- 0.13% in the 3D conduit group and 13.05% +/- 1.95% in the 2D conduit group. In the transmission electron microscope study, the 3D conduit group showed more abundant myelinated nerve fibers with well-organized and thickened extracellular collagen than the 2D conduit group, and the gastrocnemius muscle and biceps femoris tendon in the 3D conduit group were less atrophied and showed decreased fibrosis with less fatty infiltration than the 2D conduit group. A new 3D Schwann cell culture technique was established, and nerve conduit fabricated using this technique showed much improved nerve regeneration capacity than the silicone tube filled with Matrigel and Schwann cells prepared from the conventional plain culture method.
使用含有活雪旺细胞的人工神经导管是修复周围神经损伤最有前景的策略之一。为了制造一种有效的神经导管，其微观结构和内部环境在神经再生方面比现有导管更有利，开发了一种使用基质胶和背根神经节（DRG）的新型三维（3D）雪旺细胞培养技术。通过将新鲜收获的DRG直接接种到填充有基质胶的硅胶管（内径1.98mm，长14mm）中并进行体外漂浮培养2周，制造出3D排列雪旺细胞的神经导管。使用10mm的Sprague-Dawley大鼠坐骨神经缺损评估3D培养雪旺细胞导管（3D导管组，n = 6）的神经再生效果，并与用传统平面培养方法制备的填充有基质胶和雪旺细胞的硅胶导管（二维[2D]导管组，n = 6）进行比较。12周后，用坐骨神经功能指数（SFI）和步态分析评估坐骨神经功能，并用图像分析仪和电镜方法检查神经导管和坐骨神经支配组织的组织形态学。功能评估中的SFI和踝关节站立角度在3D导管组（n = 5）中为-60.1±13.9、37.9°±5.4°，在2D导管组（n = 4）中为-87.0±12.9、32.2°±4.8°。3D导管组中有髓轴突为44.91%±0.13%，2D导管组中为13.05%±1.95%。在透射电子显微镜研究中，3D导管组显示出比2D导管组更多的有髓神经纤维，细胞外胶原组织有序且增厚，并且3D导管组中的腓肠肌和股二头肌肌腱萎缩程度较轻，纤维化程度降低，脂肪浸润也比2D导管组少。建立了一种新的3D雪旺细胞培养技术，使用该技术制造的神经导管显示出比用传统平面培养方法制备的填充有基质胶和雪旺细胞的硅胶管有显著改善的神经再生能力。
