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目的:分离获取大鼠耻骨联合软骨细胞,体外培养并进行形态学观察。方法;采用机械及胰蛋白酶、胶原酶序贯消化法,从大鼠耻骨联合软骨组织中分离出软骨细胞,用含15%FBS的DMEM/F12培养液原代和传代培养,倒置显微镜下动态观察细胞形态及生长情况,阿尔辛蓝组织化学、Ⅱ型胶原免疫组化染色鉴定,通过扫描及透射电镜观察培养细胞的超微结构。结果:体外培养的大鼠耻骨联合软骨细胞呈短梭形、三角形,有短细胞突起,细胞浆内质网发达,线粒体丰富,富含分泌颗粒。细胞胞浆糖胺聚糖、Ⅱ型胶原染色均阳性。结论:耻骨联合细胞体外培养过程中,较好地保持了生长期软骨细胞的形态特征。
OBJECTIVE: To isolate and obtain rat pubic symphysis chondrocytes in vitro and observe the morphology. Methods: Chondrocytes were isolated from rat pubic symphysis cartilage by using mechanical and trypsin and collagenase sequence digestion. Primary and subcultured DMEM / F12 medium containing 15% FBS was used for dynamic observation under inverted microscope Cell morphology and growth, Alcian blue histochemistry, type Ⅱ collagen immunohistochemical staining, scanning electron microscopy and scanning electron microscopy of cultured cells in the ultrastructure. RESULTS: The rat pubic symphysis chondrocytes were short spindle-shaped, triangular with short protuberances. The cytoplasm endoplasmic reticulum was well developed and the mitochondria were rich and the secretory granules were abundant. Cell cytoplasm glycosaminoglycan, type Ⅱ collagen staining were positive. CONCLUSION: During the process of in vitro culture of pubic symphysis cells, the morphology of chondrocytes in growing period is well preserved.