兔羊膜上皮细胞向兔软骨细胞诱导的研究

目的 观察兔羊膜上皮细胞(AECs)向兔软骨细胞诱导分化的特性.方法 获得兔新鲜羊膜组织,去除外膜组织,并剪碎,胶原酶消化法进行原代培养.经传代培养、细胞形态观察后,用软骨细胞诱导培养液使AECs向软骨细胞分化,并以组织化学和免疫组织化学染色进行鉴定,电镜观察细胞形态检测软骨细胞特征性表现、细胞活性噻唑蓝(MTT)比色法检测.结果 兔羊膜上皮细胞具有前期胚胎干细胞的多向分化潜能,采用酶消化法可快速分离培养原代细胞.MTT比色法显示第3代传代细胞活性、增殖能力较高,未进行软骨细胞诱导的细胞未显示出软骨细胞的特征性表现,经软骨细胞诱导分化后,组织化学和免疫组织化学检测表明其能较好的表现软骨细胞特征.结论 兔AECs无伦理学限制,且具有前期胚胎干细胞的多向分化潜能,有望成为软骨细胞组织工程学新型的种子细胞.

Chondrogenic induction from amniotic epithelial cells in rabbits

Objective To investigate the characteristics of rabbit's amniotic epithelial cells (AECs) and possibility of chondrogenic induction. Methods The AECs in rabbits were obtained, external membrane tissue was removed, and the remaining tissue was cut into pieces and digested with collagenase of type Ⅰ with concentration of 0. 2%. The rabbit' s AECs were passaged. Phenotypes were detected by flow cytometry and the viability of AECs was determined by MTT. The AECs were cultured in chondrogenic media and detected by using histochemistry and immunohistochemistry. The morphologic changes of chondrocytes were observed under the inverted microscopy and electron microscopy. Results The rabbit' s AECs had capability of differentiation potential such as embryonic stem cells (MSCs). The isolation of these cells had been proved easily when collagenase was used to detach the cells. They also promoted the cell viability at the 3rd passages by MTT. The histochemistry and immunohistochemistry revealed that the rabbit's AECs strongly expressed chondrocytes markers after induction. Conclusion The rabbit' s AECs are abundant without limitation of ethics problems. The rabbit' s AECs with capability of differentiation potential may be the newly stem cells for tissue engineering of cartilage.