犬自体血清培养骨髓基质细胞的实验研究

目的:探讨犬自体血清培养骨髓基质细胞的可行性方法:获取犬自体血清,配制成含10%和20%自体血清的培养液,与含10%胎牛血清的培养液在相同条件下培养犬骨髓基质细胞(BMSCs)。通过倒置显微镜下观察细胞形态,细胞计数以及MTT法对细胞增殖情况进行检测。并且对第2代细胞用3种不同血清成分培养液(均加入β-甘油磷酸钠、抗坏血酸及地塞米松)进行成骨诱导分化,采用免疫组化检测碱性磷酸酶及钙结节形成。对检测到的量化数据做统计学分析,比较各组间的差异,以评价不同血清成分培养液对细胞增殖分化的影响。结果:3组细胞的生长及形态无明显差别,经过诱导培养均能形成钙结节。对3组间量化检测做统计学分析,各组间差异无统计学意义。结论:制备的犬自体血清能完全替代胎牛血清对骨髓基质细胞进行诱导培养。

Experimental Study of Canine Bone Marrow Stromal Cells Cultured by Autologous Serum in vitro

Objective：The purpose of this study was to investigate whether cell culture medium of autologus serum is more effective on expansion and differentiation of canine bone marrow stromal cells（BMSCs）.Methods：One adult beagle dog was used in this study.10%and 20%autologous serum medium were prepared.BMSCs were obtained from bone marrow of the canine.Experiments were divided into three groups：A,BMSCs cultured in 10%autologus serum;B,BMSCs cultured in 20%autologus serum;C,BMSCs cultured in 10%fetal bovine serum（FBS）.From the second generation of cells,β-glycerophosphate,ascorbic acid and dexamethaso were added to the three culture medium for osteogenic induction.The alkaline phosphatase and calcium nodules were examined by immunohistochemistry.The morphology of BMSCs,cells counting and MTT assay were detected.Quantitative data were expressed as means±SD,SPSS software version 13.0 for Windows was used.Results：Morphology of the BMSCs in A,B and C groups were similar.Calcium nodules could be observed in all groups after induction.There were no significant differences between groups in statistical analysis.Conclusion：Preparation of autologous serum can completely replace FBS for culture of bone marrow stromal cells.