人羊膜间充质细胞向成骨细胞诱导分化的实验研究

摘要

目的 观察人羊膜间充质细胞(human amnion mesenchymal cells，hAMCs)体外诱导向成骨细胞分化，为骨组织工程提供种子细胞。方法 从剖宫产后废弃的人羊膜组织分离培养hAMCs，经成骨细胞诱导条件培养基诱导后，对细胞形态特征、碱性磷酸酶、骨桥素、骨钙素表达以及I型胶原分泌进行观察和检测。结果 原代培养的hAMCs形态呈长梭形或不规则形，呈均匀分布生长，传代后细胞体积略变大，约5～7d传代1次。经成骨细胞诱导培养15d后，hAMCs碱性磷酸酶、骨钙素、骨桥素的表达呈阳性，并且检测有I型胶原分泌。结论 hAMCs易于体外分离培养及扩增，体外成骨细胞定向诱导的hAMCs具有典型的成骨细胞的形态和功能性特征，是良好的骨组织工程种子细胞。%ObjectiveTo investigate the feasibility of inducing differentiation of the human amniotic mesenchymal cells (hAMCs) into osteoblasts in vitro, so as to provide the seed cells for bone tissue engineering. Methods The hAMCs were isolated from abandoned human amnion and cultured in osteogenic media to induce the osteogenic differentiation in vitro. After hAMCs were induced by osteogenic media for 15 days, morphological observation, immunocytochemistry and western blot were used to study the cellular morphology and expression of alkaline phosphatase ( ALP), type Ⅰ collagen, osteopontin and osteocalcin. Results The primary cultured hAMCs had long spindle shape or irregular shape, which were distributed evenly. The cells were usually subcultured in 5 or 7 days. After subculture, the cells became larger. After cultured by osteogenic media for 15 days, the hAMCs were detected to express ALP, osteocalcin and osteopontin, and secrete type Ⅰ collagen. Conclusions The hAMCs are isolated,cultured and amplified easily in vitro. The induced differentiated cells by osteogenic media have typical osteoblast morphological and functional characteristics, which can be used as seed cells for bone tissue engineering.