骨骺软骨细胞的培养和保存方法

The methods of culture and storage of the epiphyseal chondrocytes

目的建立大量生产骨骺软骨细胞的培养方法和保存方法,为组织工程骺软骨的培养提供种子细胞。方法2周龄兔胫骨上端骨骺软骨经机械剪切和化学消化后,接种、培养及液氮冻存。通过显微镜观察其生物学表现,并通过蕃红花“O”染色和Ⅱ型胶原免疫组化染色对其生物学特征进行鉴定。结果在体外观察到了骨骺软骨细胞从静止、增殖到肥大这个近似体内的生长过程,经多次传代后增殖能力降低,经过冻存的细胞生物学特征不变。蕃红花“O”和Ⅱ型胶原免疫组化染色阳性。结论我们成功地建立了骨骺软骨细胞的培养和冻存方法,并证明培养出的细胞是骨骺软骨细胞。

Objective To establish a kind of epiphyseal chondrocyte culture and storage method so that we can obtain the seed cells in large quantity for the usage of engineered growth plate tissue.Methods The cells we used were harvested from proximal growth plates of tibia of rabbits and cultured. Their biological phenomena were observed and their biological characteristics were identified by using safrarine O staining and immunohistochemistry staining for type Ⅱ collagen.Results We observed that epiphyseal chondrocytes had the ability of proliferation and differentiation in vitro. Frozen storage didn't change the cells' biological characteristics.Conclusions We have established a methods of culture and storage of epiphyseal chondrocyte and the cells we cultured were verified as epiphyseal chondrocyte.