目的:探索体外培养组织工程骨-软骨复合组织种子细胞的条件,并观察其部分生物学活性。方法:采用机械-酶消化法对3周龄新西兰大白兔耳软骨和关节软骨消化来获得软骨细胞,采用全骨髓贴壁法来获得骨髓间充质干细胞,对两种细胞进行原代和传代培养,通过倒置相差显微镜观察细胞形态、绘制生长曲线、免疫组化染色等对细胞进行生物学特性分析。结果:原代培养的软骨细胞以多角形或三角形为主,传代3次以后出现去分化。形态学和免疫组化显示细胞3代以内可以保持表型稳定,具有较强的增殖及分泌细胞外基质的能力,而且耳软骨及关节软骨细胞在实验中没有表现出明显的生物学差别。采用贴壁筛选法获得的BMSCs呈长梭形或多边形,生长曲线呈“S”形,Ⅱ型胶原免疫组化染色阳性,细胞生长增殖能力旺盛。结论:体外分离培养的软骨细胞和骨髓间充质干细胞符合组织工程要求,能够作为骨-软骨复合组织的种子细胞。 Objective: To explore the conditions of culturing seed cells of tissue-engineered bone-cartilage composite tissue in vitro and to observe some of its biological activities. Methods: Chondrocytes were obtained by digestion of ear cartilage and articular cartilage of 3-week-old New Zealand rabbits by mechanical-enzymatic digestion. Bone marrow-derived mesenchymal stem cells were obtained by whole bone marrow adherent method. Primary and subculture of both cells were performed The morphology of the cells was observed by inverted phase contrast microscope, and the growth curve and immunohistochemical staining were used to analyze the biological characteristics of the cells. Results: Primary cultured chondrocytes were mainly polygons or triangles. After three passages, dedifferentiation occurred. Morphology and immunohistochemistry showed that the cells maintained stable phenotype within 3 generations and had a strong ability of proliferation and secretion of extracellular matrix. Moreover, no obvious biological differences were observed in the cartilage of ear and cartilage. BMSCs obtained by adherence screening showed long spindle or polygon, the growth curve was “S” shape, type Ⅱ collagen was positive by immunohistochemistry, and cell growth and proliferation ability was strong. CONCLUSION: Chondrocytes and BMSCs cultured in vitro are in accordance with the requirements of tissue engineering and can be used as seed cells of bone-cartilage complex.