碱性成纤维细胞生长因子对离心管内培养骺板细胞生成软骨组织的作用

Effects of basic fibroblast growth factor on cartilaginous tissue formation from epiphyseal plate chondrocytes cultured in centrifuge tube

背景:采用离心管技术体外培养骺板细胞的报道已很多。目的:观察碱性成纤维细胞生长因子对离心管内培养的骺板细胞生成类骺板组织的影响。方法:获取3周龄新西兰白兔股骨远端的骺板组织,利用组织块纱巾培养法获得骺板细胞,加入含有10μg/L碱性成纤维细胞生长因子的DMEM培养液,连续培养4周。结果与结论:离心管内聚集的骺板细胞在含有碱性成纤维细胞生长因子的DMEM培养液中形成类骺板样组织块。在组织块外周形成类似骺软骨的生发层,中心的骺板细增殖情况良好,向肥大细胞方向分化。类骺板样组织甲苯胺蓝及番红"O"染色阳性,Ⅱ型胶原免疫组织化学染色呈强阳性。说明碱性成纤维细胞生长因子能够促进离心管中的骺板细胞形成富含蛋白聚糖及Ⅱ型胶原等细胞外基质的类骺板样软骨组织。

BACKGROUND：There have been many reports regarding epiphyseal plate cells cultured in the centrifuge tube technique. OBJECTIVE：To observe the effects of basic fibroblast growth factor （bFGF） on cartilaginous tissue formation from epiphyseal plate chondrocytes cultured in centrifuge tube. METHODS：Chondrocytes were isolated from 3-week-old New Zealand rabbits using the method of tissue-yarn. A total of 5×106 chondrocytes obtained by centrifugation were cultured in a plastic centrifuge tube （15 mL） with the DMEM culture fluid including 10 μg/L bFGF for 4 weeks continuously. Cell morphology was investigated with light and inverted microscope, and the histological structure of new cartilaginous tissue was observed by histological staining. RESULTS AND CONCLUSIONS：Cartilaginous tissue regenerated from growth plate chondrocytes cultured in centrifuge tube with bFGF in the DMEM culture medium. The periphery of the cartilaginous tissue was resembled with the germinal layer of growth plate, consisting of several cellular layers. The chondrocytes in the centre of tissue grew well, and several were differentiating to the mast chondrocytes. The cartilaginous tissue appeared strongly positive for safranine ＂O＂ and toluidine blue staining, which showed that the chondrocytes could synthesize the proteoglycans. Type II collagen immunohistochemistry appeared strongly metachromatic. bFGF could promote the formation of cartilaginous tissue, rich in proteoglycans and type II collagen, from epiphyseal plate cells cultured in centrifuge tune.