| 刘 娜,李文磊,贾 鹏,朱东亚,叶 民,丁新生.Bmi-1基因表达与人骨髓间充质干细胞增殖关系的探讨[J].南京医科大学学报,2008,28(6):717~721 |
| Bmi-1基因表达与人骨髓间充质干细胞增殖关系的探讨 |
| Relationship of Bmi-1 gene expression and human mesenchymal stem cell replication |
| 投稿时间：2008-01-13 |
| DOI：10.7655 |
| 中文关键词: 骨髓间充质干细胞 贴壁培养 表型 Bmi |
| 英文关键词: human mesenchymal stem cells adherent culture phenotype Bmi-1 |
| 基金项目:南京市卫生局基金资助(2006YKK06119) |
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| 中文摘要: |
| 目的:建立人骨髓间充质干细胞(hMSCs)的体外培养体系,研究hMSCs表面抗原的表达情况,以及体外传代培养对Bmi-1基因表达变化的影响,为进一步探讨Bmi-1基因在hMSCs增殖中的作用及调控机制提供依据?方法:骨穿抽取骨髓,以1.077 g/ml的Ficoll分离液梯度密度离心,收集单个核细胞进行培养和传代,观察细胞形态和生长周期?取P4代的hMSCs,流式细胞仪检测细胞表面抗原的表达情况?通过SA-β-gal的染色检测细胞的老化程度,通过抽提各代次细胞的RNA,逆转录PCR定量Bmi-1基因量的变化,分析与细胞倍增代数的关系?结果:体外培养的hMSCs贴壁生长,呈长梭形,可增殖形成克隆?经分离纯化培养后,流式细胞仪显示细胞表型为CD13,CD105,少量表达CD33,不表达造血细胞的标志CD34及CD117?SA-β-gal的染色显示随细胞培养时间延长,SA-β-gal染色更明显和清晰?逆转录PCR显示随传代次数的增加,hMSCs的增殖能力逐渐下降,Bmi-1基因表达随培养时间而减少?结论:本培养体系可获得纯度较高的hMSCs,具有强大的增殖能力?Bmi-1基因极可能是影响MSC增殖能力的一个重要基因? |
| 英文摘要: |
| Objective:To establish the cultural system of hMSCs in vitro,investigate the changes of Bmi-1 gene expression in different passages,and explore the effect of Bmi-1 in hMSCs replicative senescence. Methods:hMSCs were isolated from the femur bone marrow using adherence method and cultured in L-DMEM with 10% fetal bovine serum(FBS). The 4 th passage hMSCs were picked up for analyzing expressions of cell surface phenotype,and the 1 st,3 rd,6 th,9 th,12 th,15 th and 20 th passages were examined of Bmi-1 gene expression by RT-PCR. Results:hMSCs were successfully isolated from the femur bone marrow which were positive for CD13,CD105,CD33 and negative for CD34,CD38,CD117,CD45. The SA-β-gal staining showed that the senescent cells were more suffused than the vigorous cells. The Bmi-1 mRNA was detected in hMSCs, and the gene expression decreased related to the cell passage. Conclusion:hMSCs that can be successfully isolated from femur bone marrow are easy to proliferate. The replicative senescence of hMSCs has correlation with the expression level of Bmi-1 gene. |
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