人恒牙牙髓干细胞体外定向诱导分化为成骨细胞的研究

Definitive differentiation from human dental pulp stem cells to osteoblast in vitro

目的研究人恒牙牙髓组织来源的牙髓干细胞在体外分化为成骨细胞的能力,探讨其作为骨组织工程种子细胞的可行性。方法从正畸治疗减数拔除的恒前磨牙中分离牙髓组织,应用酶消化法获得牙髓细胞。单抗Stro-1标记、免疫磁珠阳性分选系统分选获得牙髓干细胞,第3代牙髓干细胞用成骨向诱导培养基向成骨细胞诱导分化。用碱性磷酸酶染色和Von Kossa染色鉴定成骨向分化,RT-PCR检测成骨细胞的特异相关或标志基因。结果人恒牙牙髓干细胞经成骨向培养基诱导后表现出成骨细胞特性,碱性磷酸酶和Von Kossa染色结果均为阳性,RT-PCR检测成骨向分化相关基因Ⅰ型胶原、骨涎蛋白、骨钙素、Runx2和Osterix均有阳性表达。结论恒牙牙髓干细胞具有向成骨细胞分化的潜能,有望成为骨组织工程的种子细胞。

Objective To investigate the feasibility of directionally induced differentiation of human dental pulp stem cells（HDPSCs） toward osteoblasts in vitro.Methods Human dental pulp cells（HDPCs） were separated and cultured from dental pulp of healthy premolars extracted for orthodontic purpose.HDPSCs were isolated from cultured HDPCs by magnetic-activated cell sorting′s（MACS） indirect magnetic cell labeling and positive selection strategy with antibody Stro-1,and then they were induced to differentiation by osteogenic medium.Osteogenic differentiation was assessed by Von Kossa and alkaline phosphatase staining,and expressions of osteoblast specific genes were examined by RT-PCR.Results DPSCs ould differentiate into osteoblasts.ALP was expressed and mineralized nodules were observed after induction of HDPSCs.The genes related to osteogenic differentiation were expressed including COLⅠ,BSP,OC,Runx2 and Osterix.Conclusion DPSCs are promising seed cells for bone tissue engineering.