共培养诱导小鼠ES细胞向软骨细胞分化的初步研究

The Preliminary Study of Chondrogenic Differentiation of Mouse Embryonic Stem Cells by Co -Culture with Chondrocytes

目的探讨软骨共培养体系诱导小鼠ES细胞向软骨细胞分化的可行性。方法GFP标记的小鼠ES细胞初步分化为EB后,将EB消化为单个细胞,同猪关节软骨细胞按一定比例(1:3)混合后接种于PGA材料,体外培养1周后植入裸鼠皮下3周取材。对照组为EB细胞接种组及软骨细胞接种组。取材后行连续冰冻切片,切片分别做荧光拍照,HE染色及甲苯胺蓝染色。结果组织学结果显示,EB细胞接种组形成畸胎瘤;软骨细胞对照组形成软骨组织;实验组形成软骨组织和畸胎瘤的混合体。甲苯胺蓝染色结果和荧光照片对照结果显示,部分软骨组织GFP阳性,由小鼠ES细胞分化而来。结论软骨其培养体系可以诱导小鼠ES细胞向软骨细胞分化,但得到的软骨组织不纯,混有畸胎瘤组织。

Purpose To induce the mouse embryonic stem (ES) cells differentiate into chondrocytes by co-culture with chondrocytes. Methods GFP-labeled mouse ES cells were firstly cultured in suspension to form embryoid bodies (EBs). EBs were digested into single cells, mixed with pig joint chondrocytes at the ratio of 1:3, and seeded into poly glycolic acid (PGA) scaffolds. The groups seeded with EB cells or chondrocytes alone were used as control. The cell-PGA complex were cultured for 1 week in vitro and then implanted subcutaneously into nude mouse for 3 weeks. The specimens were froze, serially sectioned, and stained with HE and toludine blue. Results The control groups which seeded with EB cells or chondrocytes alone formed teratoma and cartilage, respectively. The experimental group formed a mixed tissue complex containing both teratoma and cartilage, Toludine blue staining and fluorescence microscopy examination show that part of chondrocytes with green fluorescence, indicating that those chondrocytes were differentiated from mouse ES cells. Conclusion Mouse ES cells could be differentiated into chondrocytes by co-culture with chondrocytes. However, further purification process should be taken to formed pure cartilage.