详细内容： 【摘要】 目的 采用低温沉积技术 3D 打印制备聚己内酯（polycaprolactone，PCL）/Ⅰ型胶原组织工程半月板支架（以下简称 PCL/Ⅰ型胶原半月板支架），探讨其理化特性。方法 制备 15%PCL/4%Ⅰ型胶原溶液及15%PCL 溶液，利用低温沉积技术 3D 打印制备 PCL/Ⅰ型胶原半月板支架及 PCL 半月板支架。大体及扫描电镜观察支架形态及微观结构，生物力学试验测量支架压缩模量及拉伸模量，红外光谱分析支架成分，测量支架表面接触角；将两种支架及其浸提液分别与兔半月板细胞复合培养，细胞计数试剂盒 8（cell counting kit 8，CCK-8）检测细胞增殖，并以正常培养细胞作对照；扫描电镜观察支架-细胞复合物中细胞黏附及生长情况。结果 大体及扫描电镜观察显示，两种支架均具有取向的三维微观结构及孔隙，但 PCL/Ⅰ型胶原半月板支架表面更粗糙。生物力学测试，两种支架压缩模量及拉伸模量比较，差异均无统计学意义（P>0.05）。红外光谱分析提示，PCL/Ⅰ型胶原半月板支架中 PCL 和Ⅰ型胶原成功混合。PCL/Ⅰ型胶原半月板支架表面接触角为（83.19±7.49）°，较 PCL 半月板支架（111.13±5.70）° 显著减小（t=6.638，P=0.000）。CCK-8 检测显示，随培养时间延长，两种支架浸提液培养的细胞数量呈递增趋势，与对照组比较差异均无统计学意义（P>0.05）。支架-细胞复合物扫描电镜观察示，PCL/Ⅰ型胶原半月板支架表面黏附细胞多于 PCL 半月板支架。结论 低温沉积技术 3D 打印制备的 PCL/Ⅰ型胶原半月板支架具有优良的理化学性能，无细胞毒性，有望作为半月板组织工程支架材料。
【关键词】 半月板组织工程；低温沉积技术；3D 打印技术；支架材料；理化特性
Study on the preparation of polycaprolactone/typeⅠcollagen tissue engineeredmeniscus scaffold by three-dimensional printing and its physiochemical properties
SHEN Shi1, 2, CHEN Mingxue2, GAO Shuang2, GUO Weimin2, WANG Zhenyong2, LI Haojiang2, LI Xu2,ZHANG Bin1, 2, XIAN Hai1, 2, ZHANG Xueliang2, LIU Shuyun2, HAO Libo2, ZHUO Naiqiang1, GUO Quanyi2
1. Department of Orthopedics and Joint Surgery, the Affiliated Hospital of Southwest Medical University, Luzhou Sichuan, 646000,P.R.China
2. Institute of Orthopedics, General Hospital of Chinese PLA, Beijing Key Laboratory of Regenerative Medicine in Orthopedics, Key Laboratory of Musculoskeletal Trauma &?War Injuries, PLA, Beijing, 100853, P.R.China
Corresponding authors: ZHUO Naiqiang, Email: znq0101@163.com; GUO Quanyi, Email: doctorguo_301@163.com
【Abstract】 Objective To manufacture a polycaprolactone (PCL)/type Ⅰ collagen (COL Ⅰ) tissue engineered meniscus scaffold (hereinafter referred to as PCL/COL Ⅰ meniscus scaffold) by three-dimensional (3D) printing with low temperature deposition technique and to study its physicochemical properties. Methods First, the 15% PCL/4% COLⅠ composite solution and 15% PCL simple solution were prepared. Then, 15% PCL/4% COL Ⅰmeniscus scaffold and 15% PCL meniscal scaffold were prepared by using 3D printing with low temperature deposition techniques. The morphology and microstructure of the scaffolds were observed by gross observation and scanning electron microscope. The compression modulus and tensile modulus of the scaffolds were measured by biomechanical test. The components of the scaffolds were analyzed by Fourier transform infrared spectroscopy (FTIR). The contact angle of the scaffold surface was measured. The meniscus cells of rabbits were cultured with the two scaffold extracts and scaffolds, respectively. After cultured, the cell proliferations were detected by cell counting kit 8 (CCK-8), and the normal cultured cells were usedas controls. Cell adhesion and growth of scaffold-cell complex were observed by scanning electron microscope.
Results According to the gross and scanning electron microscope observations, two scaffolds had orientated 3D microstructures and pores, but the surface of the PCL/COLⅠ meniscus scaffold was rougher than the PCL meniscus scaffold. Biomechanical analysis showed that the tensile modulus and compression modulus of the PCL/COL Ⅰ meniscus scaffold were not significantly different from those of the PCL meniscus scaffold (P>0.05). FTIR analysis results showed that COL Ⅰ and PCL were successful mixed in PCL/ COL Ⅰ meniscus scaffolds. The contact angle of PCL/COLⅠ meniscus scaffold [(83.19±7.49)°] was significantly lower than that of PCL meniscus scaffold [(111.13±5.70)°] (t=6.638,P=0.000). The results of the CCK-8 assay indicated that with time, the number of cells cultured in two scaffold extracts showed an increasing trend, and there was no significant difference when compared with the control group (P>0.05).Scanning electron microscope observation showed that the cells attached on the PCL/ COL Ⅰ meniscus scaffold more than that on the PCL scaffold. Conclusion ? ?PCL/COLⅠmeniscus scaffolds are prepared by 3D printing with low temperature deposition technique, which has excellent physicochemical properties without cytotoxicity. PCL/COLⅠ meniscus scaffold is expected to be used as the material for meniscus tissue engineering.
【Key words】 Meniscus tissue engineering; low temperature deposition technique; Three-dimensional printing technology; scaffold material; physicochemical property
Foundation items: National Key Research and Development Plan of China (2017YFC1103404); National Natural Science Foundation of China (81772319, 21134004); Natural Science Foundation of Beijing (7172203)
