生物衍生骨材料与骨髓间充质干细胞体外的生物相容性

背景：生物衍生骨材料已在临床应用多年，但其作为组织工程骨支架材料的可行性还需验证。 目的：评价异种生物衍生骨材料与骨髓间充质干细胞的生物相容性。 设计、时间及地点：单一样本体外观察性实验，于2007-08/2008-01在中国辐射防护研究院医用组织库组织工程实验室完成。 材料：山西省医用组织库生产的人松质骨；体外分离培养1周龄新西兰兔骨髓间充质干细胞。 方法：经脱脂、脱钙、去抗原等步骤制备生物衍生骨材料，扫描电镜观察其表面超微结构。直接贴壁法分离培养兔骨髓间充质干细胞，MTT法和流式细胞仪检测生物衍生骨材料对骨髓间充质干细胞增殖及凋亡的影响。PKH26标记骨髓间充质干细胞与生物衍生骨材料复合培养，荧光显微镜动态观察细胞在骨材料上的生长情况；扫描电镜观察细胞在骨材料上的贴附、生长情况。 主要观察指标：①生物衍生骨材料的形态结构及骨髓间充质干细胞在其上的生长、增殖情况。②生物衍生骨材料的细胞毒性。 结果：所制备的生物衍生骨材料呈白色，扫描电镜观察呈规则的疏松多孔结构，孔隙间有微孔相互连通。其对骨髓间充质干细胞的增殖和凋亡无明显的影响；荧光显微镜下观察，在材料表面及孔隙内可见PKH26标记的细胞，显示红色荧光；扫描电镜观察可见细胞在材料上附着生长，细胞形态呈梭型或多角形，与材料贴附紧密。 结论：实验制备的异种生物衍生骨材料细胞毒性低，与骨髓间充质干细胞具有良好的生物相容性。

Biocompatibility of bio-derived bone to bone marrow mesenchymal stem cells in vitro

BACKGROUND: The bio-derived bone has been applied in clinics for many years. But its feasibility as the bone tissue engineering scaffolds still needs to be verified. OBJECTIVE: To evaluate the biocompatibility of heterogeneic bio-derived bone to bone marrow mesenchymal stem cells (BMSCs). DESIGN, TIME AND SETTING: An observational experiment of single sample in vitro was performed in Tissue Engineering Laboratory of China Institute for Radiation Protection from August 2007 to January 2008. MATERIALS: Human cancellous bone was produced by Medical Tissue Bank, China Institute for Radiation Protection. BMSCs drawn from one week old New Zealand rabbits were cultured in vitro. METHODS: The bio-derived bone was prepared by degrease, decalcification and antigen extraction. The Ultrastructure of the bio-derived bone was observed by the scanning electron microscope(SEM). The rabbit BMSCs were cultured by direct adherence. The influence of the bio-derived bone on the proliferation and apoptosis of the BMSCs were measured by MTT method and the flow cytometry. The BMSCs labeled by PKH26 were co-cultured with the bone materials in vitro. The growth of BMSCs on the bio-derived bone was dynamically observed by fluorescence microscope. The adhesion and growth of BMSCs were observed by SEM. MAIN OUTCOME MEASURES: The morphological structure and cytotoxicity of bio-derived bone were observed. The growth and proliferation of the BMSCs on the surface of the bio-derived bone were observed. RESULTS: The bio-derived bones were white. Under the SEM, the bio-derived bones had a regular porosity structure with well interconnected pores. There was no influence of the leaching liquor of bio-derived bone on the proliferation and apoptosis of the BMSCs. Under the fluorescence microscope, the BMSCs labeled by PKH26 grew on the surface as well as into the internal of the materials, and emitted red fluorescence. Under the SEM, the BMSCs, which grew well, displayed polygon shape or fusiform shape, and they adhered on the surface of the materials tightly. CONCLUSION: The cytotoxicity of the heterogeneic bio-derived bones are very low, and they have good cytocompatibility to BMSCs.