雪旺氏细胞与同种异体骨支架的体外共培养研究

In Vitro Culture of Schwann Cells on Allogeneic Bone Scaffolds

目的:探讨雪旺细胞(Schwann’s cells,SCs)在同种异体骨支架上的生物相容性,体外构建组织工程骨神经化模型。方法:利用新鲜人体骨骼制备同种异体骨支架材料,检测其物理性能;采用优化方法提取新生SD大鼠坐骨、臂丛神经培养SCs,实验分为三维培养实验组(SCs+同种异体骨)、二维培养对照组(SCs+胶原玻片),S-100抗体免疫荧光染色鉴定SCs纯度;细胞计数法检测两组细胞增殖特点;细胞接种后第3、7天取样,扫描电镜观察。结果:同种异体骨支架具有良好的三维孔隙结构,适宜细胞贴附生长;S-100免疫荧光染色证实SCs纯度>95%;扫描电镜检测显示两组SCs均可正常粘附增殖,细胞间排布规律相似,培养早期实验组SCs胞体更加细长,伪足更加明显,随着培养时间的延长表现出较强的迁移能力;细胞增殖检测:两组SCs生长曲线特征基本一致,支架材料对SCs无毒性作用。结论:同种异体骨支架SCs具有良好的生物相容性,其三维立体多孔结构有利于SCs的粘附与迁移,初步构建了体外组织工程骨神经化模型。

Objective： To investigate the biocompatibility between in vitro cultured Schwann cells（SCs） and allogeneic bone scaffolds. Methods： Allograft bone scaffold materials were prepared from fresh human bones and the physical properties were tested. SCs were isolated from extracted sciatic and brachial plexus from newborn SD rats. Two culture conditions were used： three-dimensional（SCs plus allogeneic bone scaffold） and two-dimensional culture model（SCs plus collagen glass）. The purity of SCs was identified with immunofluorescence staining by anti-S-100 antibody. The cells were evaluated for proliferation by cell counting and also imaged with scanning electron microscopy. Results： Allogeneic bone scaffolds had the special three-dimensional pore structure, which supported the adhesion and growth of SCs. The purity of SC was 〉95%. SCs cultured in both three-dimensional and two-dimensional models proliferated well at the similar rate and shared the similar distribution characteristics. Compared with the two-dimensional model, the cells cultured in the three-dimensional model showed slender cell bodies and more cellular pseudopods at the early stage of incubation, and enhanced capability of migration at the later stage of incubation. Conclusions： Allogeneic bone scaffold and SCs have good biocompatibility and the in vitro model of tissue engineering neural bone is successfully established.