中国组织工程研究

中国组织工程研究 ›› 2017, Vol. 21 ›› Issue (34): 5423-5429.doi: 10.3969/j.issn.2095-4344.2017.34.003

• 组织工程骨及软骨材料 tissue-engineered bone and cartilage materials • 上一篇    下一篇

单一生物材料聚乳酸/羟基乙酸共聚物构建组织工程骨软骨的可行性

段平国1，郭润生1，余兴元2，潘  震3，李晓峰1，李  虎1，刘  俊1，姚浩群1
  

  1. 1南昌大学第一附属医院骨科，江西省南昌市  330006； 2南昌大学医学院，江西省南昌市  330006；3复旦大学高分子科学系，上海市  200433
  • 收稿日期:2017-09-19 出版日期:2017-12-08 发布日期:2018-01-04
  • 通讯作者: 姚浩群，博士，副教授，南昌大学第一附属医院骨科，江西省南昌市 330006
  • 作者简介:段平国，男，1979年生，江西省南昌市人，汉族，主治医师，2013年复旦大学毕业，博士，主要从事骨与软骨组织工程研究。
  • 基金资助:

    国家自然科学基金项目(81401790)；江西省自然科学青年基金重点项目(20171ACB21057)；江西省自然科学基金项目(20151BAB205053，20161BAB205235)；江西省教育厅科技项目重点项目(GJJ160028)

Feasibility of constructing a scaffold for osteochondral tissue engineering using poly(lactide-co-glycolide) alone 

Duan Ping-guo1, Guo Run-sheng1, Yu Xing-yuan2, Pan Zhen3, Li Xiao-feng1, Li Hu1, Liu Jun1, Yao Hao-qun1 
  

  1. 1Department of Orthopedics, First Affiliated Hospital of Nanchang University, Nanchang 330006, Jiangxi Province, China; 2Medical School of Nanchang University, Nanchang 330006, Jiangxi Province, China; 3Department of Polymer Science, Fudan University, Shanghai 200433, China
  • Received:2017-09-19 Online:2017-12-08 Published:2018-01-04
  • Contact: Yao Hao-qun, M.D., Associate professor, Department of Orthopedics, First Affiliated Hospital of Nanchang University, Nanchang 330006, Jiangxi Province, China
  • About author:Duan Ping-guo, M.D., Attending physician, Department of Orthopedics, First Affiliated Hospital of Nanchang University, Nanchang 330006, Jiangxi Province, China
  • Supported by:
    the National Natural Science Foundation of China, No. 81401790; the Natural Science Foundation for the Youth in Jiangxi Province, No. 20171ACB21057; the Natural Science Foundation of Jiangxi Province, No. 20151BAB205053, 20161BAB205235; the Science and Technology Project of Jiangxi Provincial Education Department, No. GJJ160028

PDF

187

被引次数

1

摘要：

文章快速阅读：

 

文题释义：
双层多孔支架：上、下层孔径分别为100-200 μm和300-450 μm，理论上满足了软骨和软骨下骨形成的孔径需求，上层的小孔径利于细胞聚集，细胞的密度更高，产生了低氧微环境；而下层的大孔径更利于细胞的迁移和渗透，分布更均匀。
DiI：是一种亲脂性的荧光染料，可用来染细胞膜和其他脂溶性生物结构，进入细胞膜后，DiI在整个细胞膜上扩散，最佳浓度时可使整个细胞膜染色。DiI在进入细胞膜之前荧光非常弱，当与细胞膜结合后其荧光强度明显增强，DiI被激发后可发出橙红色的荧光，具有很高的淬灭常数、中等强度的光量子和很短的激发态寿命。
 
背景：为仿生软骨和软骨下骨的生理和功能需求，近年来一些研究通过制备双层支架构建组织工程骨软骨来修复软骨缺损。但大多研究使用了不同的复合生物材料，很少文献报道使用单一生物材料制备双层多孔支架。
目的：研究一体化双层聚乳酸/羟基乙酸共聚物(poly(lactic-co-glycolide)，PLGA)多孔支架材料的生物相容性，探讨其构建组织工程骨软骨的可行性。
方法：使用PLGA作为生物材料，采用室温模压/粒子浸出法制备一体化双层多孔支架。①体外实验：将第3代兔骨髓间充质干细胞接种于双层多孔支架上，接种1周后进行扫描电镜观察，接种48 h后进行LIVE/DEAD荧光染色；②体内实验：将骨髓间充质干细胞-双层多孔支架复合物、双层多孔支架分别植入裸鼠皮下，4，8周后取出植入物，进行苏木精-伊红染色和DAPI染色。
结果与结论：①成功获得一体化双层多孔支架，支架上层(软骨层)孔径为100-200 μm，下层(软骨下骨层)孔径为300-450 μm，孔隙率为85%；②体外实验：LIVE/DEAD染色显示，骨髓间充质干细胞可在支架良好存活；扫描电镜可见细胞黏附在支架孔壁上，并可见大量沉积的细胞外基质；③体内实验：植入后8周苏木精-伊红染色显示，实验组支架上层纤维组织中夹杂有少量软骨细胞，下层可见大量骨小梁样组织形成，上下层组织紧密结合，支架部分降解，DAPI染色显示骨髓间充质干细胞可在裸鼠体内存活8周；对照组支架上下层可见少量纤维组织，未见软骨细胞及明显的小梁骨组织形成，支架部分降解；④结果表明：单一生物材料PLGA制备的一体化双层支架生物相容性良好，构建组织工程骨软骨具有可行性。

关键词: 生物材料, 软骨组织工程, 双层支架, PLGA, 骨髓间充质干细胞, 生物相容性, 国家自然科学基金

Abstract:

BACKGROUND: To mimic physiological and functional requirements of cartilage and subcondral bone, some recent studies have fabricated bilayered scaffolds for osteochondral tissue engineering in cartilage repair. However, in cartilage repair, and little is reported on the use of a single biomaterial instead of composite biomaterials to fabricate bilayered porous scaffolds.
OBJECTIVE: To investigate the biocompatibility of an integrated bilayered porous scaffold fabricated with poly(lactic-co-glycolide) (PLGA), and then to discuss its feasibility of constructing tissue-engineered osteochondral graft.
METHODS: The integrated bilayered porous scaffolds were fabricated with PLGA by changing the ultrastructure of scaffold. In vitro, bone marrow mesenchymal stem cells (BMSCs) isolated from rabbits were seeded into the bilayered porous PLGA scaffold, co-cultured for 1 week and then observed under scanning electron microscope. After co-culturing 48 hours, the cells were stained by LIVE/DEAD Kit. In vivo, the composite of DiI-labeled BMSCs and the scaffold or the scaffold alone were implanted subcutaneously into the skin of nude mice, and the implants were taken out and stained by hematoxylin-eosin and DAPI staining at 4 and 8 weeks after implantation.
RESULTS AND CONCLUSION: (1) We successfully obtained the bilayered porous scaffolds with different pore sizes (the upper layer: 100-200 μm and the lower layer: 300-450 μm) and 85% porosity. (2) In the in vitro experiment, the LIVE/DEAD staining showed that BMSCs survived well in the scaffold, and the cells adhered well to the wall of pores in all the scaffolds and extracellular matrix deposition was found under the scanning electron microscope. (3) In the in vivo experiment, shown by the hematoxylin-eosin staining, a small amount of chondrocytes formed in the fibrous tissue in the upper layer and a large amount of trabecular bones in the lower layer were found at 8 weeks after implantation in the experimental group. The close integration between the upper layer and the lower layer appeared. BMSCs could survive for 8 weeks in vivo shown by the DAPI staining. However, in the control group, a little fibrous tissue was found without chondrocytes and apparent trabecular bone, and the scaffold was partially degraded. To conclude, the integrated bilayered porous scaffold fabricated with PLGA alone has good biocompatibility, which is feasibly used in osteochondral tissue engineering. 

Key words: Cartilage, Stem Cells, Tissue Engineering

中图分类号: