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-  2015 

人脂肪来源干细胞成软骨分化过程中成软骨相关微小RNA表达的初步研究

DOI: doi:10.7507/1002-1892.20150017

Keywords: 微小RNA, 人脂肪来源干细胞, 成软骨分化, 基因芯片, 靶基因

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Abstract:

目的探讨微小RNA(microRNA,miRNA)在人脂肪来源干细胞(human adipose-derived stem cells,hADSCs)诱导成软骨分化过程中的表达规律及其影响软骨分化的可能机制。 方法取行抽脂术或其他腹部手术患者自愿捐赠的脂肪组织,分离、培养hADSCs并鉴定。取第3代细胞成软骨分化,倒置相差显微镜下观察细胞形态,于诱导21 d行阿尔新蓝染色观察软骨形成情况,诱导0、7、14、21 d行ELISA检测成软骨相关蛋白Ⅱ型胶原蛋白(collagen type Ⅱ,Col2a1)、蛋白聚糖(Aggrecan)、Col10a1以及硫酸软骨素表达。采用基因芯片技术筛选hADSCs成软骨诱导前及诱导后21 d差异性表达miRNA,并预测筛选出的miRNA靶基因。 结果实验成功培养hADSCs,经成软骨诱导培养后,随时间延长可形成软骨球;21 d阿尔新蓝染色呈阳性;hADSCs成软骨诱导后7、14、21d,Col2a1、Aggrecan、Col10a1及硫酸软骨素表达水平均较成软骨诱导前hADSCs显著增高,差异均有统计学意义(P<0.05)。基因芯片技术共筛选出11个差异性表达miRNA,其中7个miRNA表达上调,4个miRNA表达下调。筛选出的成软骨相关miRNAs预测靶基因可能参与了干细胞成软骨分化、增殖、凋亡、细胞周期调控,以及介导细胞内级联反应和自我更新等。 结论实验筛选出11个成软骨分化差异性表达超过2倍的miRNAs,并对其靶基因进行预测,加深了对hADSCs成软骨分化机制的理解,为定向控制hADSCs成软骨分化及筛选组织工程改良种子细胞提供了理论依据

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