新型乳酸-羟基乙酸共聚物(PLGA)/胶原复合材料用于软骨再生的体外研究

摘要

Objective To design and fabricate a novel poly (lactic-co-glycolic acid) (PLGA)/collagen hybrid scaffold and to investigate its effect on in vitro chondrogenesis. Methods Novel PLGA/ collagen hybrid scaffold was prepared by forming collagen microsponge in the openings of a PLGA knitted mesh. Surface morphologies were observed by scanning electron microscopy. Pore sizes were calculated with an image analysis software. Bovine articular chondrocytes (BACs) were isolated, cultured and examined by phase contrast microscopy before seeded on the scaffold, and then seeding efficiency was investigated. Cell growth on the scaffold was observed by scanning electron microscopy (SEM). Quantification of DNA and glycosaminoglycan (GAG) was carried out after the samples were cultured in vitro for 1 week. Real-time PCR analysis was done to evaluate the expressions of type I collagen, type II collagen and aggrecan mRNA. BACs cultured in polystyrene cell culture plates for 1 week and native cartilage were used as controls. Results Ovel PLGA/collagen scaffold was successfully fabricated. The surface pore size was (136. 4 ?11. 8) 祄. BACs were isolated and multiplied. The seeding efficiency was 87. 8% ?1.6%. Homogenous cell distribution and cell growth were confirmed by SEM observation. DNA and GAG amount, type II collagen and aggrecan mRNA expression were significantly higher than that in control group (P<0. 05). Conclusions The novel PLGA/collagen scaffold we prepared enhanced in vitro chondrogenesis greatly, and can be applied in the future cartilage regeneration research.%目的 设计和制备新型乳酸-羟基乙酸共聚物[poly(lactic-co-glycolic acid),PLGA]/胶原(collagen)复合材料,研究其在体外对软骨再生的促进作用,为软骨组织工程提供新型支架材料.方法 利用冷冻干燥技术将胶原多孔海绵复合于PLGA编织网膜；扫描电镜观察材料表面微观结构,利用图像软件统计孔径大小；分离与培养牛膝关节软骨细胞(bovine articular chondrocyte,BAC),接种于PLGA/胶原材料,检测细胞接种效率,扫描电镜观察细胞在材料内部生长情况；体外培养1周后检测DNA和糖胺聚糖(glycosaminoglycan,GAG)含量,real-time PCR检测Ⅰ型胶原、Ⅱ型胶原和聚集蛋白聚糖(aggrecan) mRNA表达强度.牛膝关节软骨组织和体外单层培养的BACs做对照.结果 成功构建新型PLGA/胶原复合材料,表面孔径为(136.4±11.8) μm;细胞接种效率为87.8％±1.6％；BACs在材料表面和中心生长活跃,培养1周后的DNA、GAG含量,Ⅱ型胶原和聚集蛋白聚糖mRNA表达强度均显著高于对照组(P＜0.05).结论 新设计制备的PLGA/胶原复合材料能促进体外软骨再生,可作为支架材料用于软骨组织工程研究.