| 摘要: |
| 目的:探讨体外诱导犬骨髓间充质干细胞(bone marrow mesenchymal stem cells, BMSCs)向上皮样细胞分化的可行性和基本条件.方法:抽取成年健康杂种犬的骨髓,用密度梯度离心及贴壁培养法分离纯化BMSCs,培养扩增后,取第2代BMSCs在以下不同培养液中诱导培养:K-SFM+EGF+BPE、DMEM+EGF、K-SFM+BPE、DMEM+FBS,观察细胞形态的变化,绘制细胞的生长曲线,诱导培养后第7、14天,用免疫细胞化学染色和流式细胞仪鉴定上皮细胞特异性标志物细胞角蛋白(cytokeratin, CK)和上皮膜抗原(epithelial membrane antigen, EMA)的表达.结果:K-SFM+EGF+BPE组诱导培养第7天,少量细胞形态变为圆形或椭圆形,有突起,第14天圆形或椭圆形细胞明显增多;诱导培养过程中DMEM+EGF、K-SFM+BPE、DMEM+FBS组细胞形态无明显变化.CK免疫细胞化学染色结果提示:K-SFM+EGF+BPE组诱导培养第7天,少量细胞呈阳性表达,第14天,阳性细胞明显增多;DMEM+EGF、K-SFM+BPE、DMEM+FBS组细胞均呈阴性表达.流式细胞仪检测发现K-SFM+EGF+BPE组诱导培养第14天,部分细胞CK和EMA呈阳性表达(>7﹪),DMEM+EGF、K-SFM+BPE、DMEM+FBS组细胞均为阴性.结论:本实验提示K-SFM+EGF+BPE联合培养液能够在体外诱导犬BMSCs向上皮样细胞分化. |
| 关键词: 骨髓、间质干细胞、上皮样细胞、细胞分化 |
| DOI：10.3724/SP.J.1008.2006.00858 |
| 投稿时间：2006-01-19修订日期：2006-06-12 |
| 基金项目:国家自然科学基金(30271293). |
|
| Differentiation of canine bone marrow mesenchymal stem cells into epithelium-like cells: an in vitro study |
| HAN Gui-lin,DING Fang-bao,MEI Jü,HUANG Sheng-dong,GONG De-jun,LIU Xiao-hong,LIU Yan-ling |
| (第二军医大学长海医院胸心外科,上海,200433) |
| Abstract: |
| Objective：To investigate the feasibility of inducing canine bone marrow mesenchymal stem cells （BMSCs） to differentiate into epithelium-like cells in vitro and the basic inducing conditions. Methods： BMSCs were obtained from healthy adult canine bone marrow aspirates and were purified by density gradient centrifugation and anchoring culture. BMSCs of second passage were cultured with 4 different media.. K-SFM＋ EGF＋ BPE, DMEM4- EGF, K-SFM＋ BPE, and DMEM＋ FBS. The changes in cell morphology were observed daily under phase-contrast microscopy and cell growth curves of every group were drawn by the continual cell counting. The cells were identified by immunocytochemistry staining and flow cytometer through detecting cytokeratin （CK） and epithelial membrane antigen （EMA） on the 7^th and 14^th day of induction. Results.. A few BMSCs in K-SFM＋EGF＋BPE group became round or oval with processes on the 7^th day of induction; more BMSCs became round or oval and their phenotypic markers were CK＋ and EMA＋ on the 14^th day. FACS analysis showed that CK or EMA positive cells were rare in K-SFM＋EGF＋BPE group on the 7^th day after induction （〈1%）, and the number of positive cells were increased （〉 7 %） on the 14^th day. Cell morphology had no obvious change in DMEM＋EGF, K-SFM＋BPE, and DMEM＋FBS groups during the whole culture process. Moreover, cells in DMEM＋ EGF, K-SFM＋ BPE, and DMEM＋ FBS group were negative for CK and EMA. Conclusion： Our experiment indicates that the combination of K-SFM, EGF and BPE can induce canine BMSCs to differentiate into epithelium-like cells in vitro. |
| Key words: bone marrow mesenchymal stem cells epithelium-like cells cell differentiation |
