丝素蛋白/明胶/壳聚糖三维多孔软骨组织支架的制备及体外评价  

Preparation and in vitro evaluation of a three-dimensional porous cartilage scaffold made of silk fibroin/gelatin/chitosan

作　　者：谷明西 王常成 田丰德[2] 安宁[2] 郝瑞胡 郭林[2] Gu Mingxi;Wang Changcheng;Tian Fengde;An Ning;Hao Ruihu;Guo Lin(Shenzhen Hospital of Peking University,Shenzhen 518000,Guangdong Province,China;Second Department of Orthopedics,Zhongshan Hospital of Dalian University,Dalian 116001,Liaoning Province,China)

机构地区：[1]北京大学深圳医院,广东省深圳市518000 [2]大连大学附属中山医院骨二科,辽宁省大连市116001

出　　处：《中国组织工程研究》2024年第3期366-372,共7页Chinese Journal of Tissue Engineering Research

摘　　要：背景:软骨缺损是骨科医生面临的主要临床挑战之一,组织工程是一种结合了工程学和细胞生物学知识的跨学科方法,为软骨缺损的修复提供了新思路与途径。目的:基于丝素蛋白、明胶和壳聚糖制备多组分复合支架,通过评估其理化性质和生物学性能,筛选能够适合软骨再生的三维多孔支架。方法:以丝素蛋白、明胶和壳聚糖为基础材料,通过真空冷冻干燥法制备4组多孔支架,分别为明胶/壳聚糖支架、丝素蛋白/壳聚糖支架、丝素蛋白/明胶支架和丝素蛋白/明胶/壳聚糖支架,通过扫描电镜、X射线衍射、孔隙率、吸水膨胀率和生物降解率及力学性能等检测筛选出合适的软骨支架。然后将软骨支架与骨关节炎患者软骨细胞共培养,通过细胞黏附率、活死染色和增殖活性等检测体外评估多孔支架用于软骨损伤修复的可行性。结果与结论:①4组支架均具有多孔结构,综合物理性能检测结果得出丝素蛋白/明胶/壳聚糖支架更符合软骨缺损修复的要求,该支架的孔径为(176.00±53.68)μm,孔隙率为(80.15±2.57)%,吸水溶胀率为(3712±358)%,体外浸泡于含溶菌酶的PBS中28 d后的生物降解速率为(46.87±3.25)%,且具有良好的机械性能;②软骨细胞可在丝素蛋白/明胶/壳聚糖支架上良好黏附,且随着时间的延长,细胞黏附率增加;CCK8和活/死细胞双染检测结果显示,丝素蛋白/明胶/壳聚糖支架具有良好的生物相容性和较低的细胞毒性;③结果表明,丝素蛋白/明胶/壳聚糖支架具有高度水合3D结构、合适的孔径和孔隙率、良好的生物降解性能和优越的机械性能,可以为营养物质的转运和软骨细胞的附着、增殖提供良好的网状骨架和微环境。BACKGROUND:Cartilage defects are one of the major clinical challenges faced by orthopedic surgeons.Tissue engineering is an interdisciplinary approach that combines knowledge of engineering and cell biology to provide new ideas and approaches for the repair of cartilage defects.OBJECTIVE:To prepare a multi-component composite scaffold based on silk fibroin,gelatin,and chitosan to screen for a three-dimensional porous scaffold suitable for cartilage regeneration by evaluating its physicochemical properties and biological performance.METHODS:Four groups of porous scaffolds were prepared by vacuum freeze-drying method using silk fibroin,gelatin and chitosan as the base materials,namely chitosan/gelatin scaffold,silk fibroin/chitosan scaffold,silk fibroin/gelatin scaffold and silk fibroin/chitosan/gelatin scaffold.The suitable cartilage scaffolds were screened by scanning electron microscopy,X-ray diffractometer,porosity,water absorption and swelling rate,biodegradation rate and mechanical property detection.Then cartilage scaffolds were co-cultured with chondrocytes isolated and extracted from patients with osteoarthritis.The feasibility of porous scaffolds for cartilage injury repair was evaluated in vitro by cell adhesion rate assay,cell live-dead staining and cell activity proliferation assay.RESULTS AND CONCLUSION:(1)All four groups of scaffolds had porous structures.The comprehensive physical performance test results showed that the silk fibroin/gelatin/chitosan scaffold was more in line with the requirements of cartilage defect repair.This scaffold had a pore size of(176.00±53.68)μm,the porosity of(80.15±2.57)%,and water absorption and swelling rate of(3712±358)%.After immersion in PBS containing lysozyme for 28 days in vitro,the biodegradation rate was(46.87±3.25)%,and it had good mechanical properties.(2)Chondrocytes could adhere well on the silk fibroin/gelatin/chitosan scaffold,and the cell adhesion rate increased with time.CCK8 and live/dead cell double staining results showed that silk fibroin/gelat

关 键 词：软骨 软骨缺损 组织工程 丝素蛋白 明胶 壳聚糖 

分 类 号：R459.9[医药卫生—治疗学] R318.08[医药卫生—临床医学] R-331