间接接触共培养下骨髓间充质干细胞分化为心肌样细胞及相关调控基因的时序表达

发布时间：2018-04-20 11:36

本文选题：骨髓间充质干细胞 + 间接接触　；参考：《心脏杂志》2010年06期

【摘要】：目的:研究间接接触共培养条件下骨髓间充质干细胞(mesenchymal stem cells,MSCs)向心肌细胞(myocadium-lkce cells,CMs)的分化及相关调控基因的时序表达;筛选MSCs定向分化为CMs的重要调控基因。方法:将MSCs与CMs按1∶5的比例进行间接接触共培养,连续观察两周,在相差显微镜下观察MSCs的形态变化。采用免疫荧光染色法检测心肌特征性肌动蛋白α(α-actin)和心脏肌钙蛋白T(cTnT)的表达。应用半定量RT-PCR分析TGF-β、Nkx-2.5、GATA-4、MEF-2C及TEF-1等相关调控基因在分化过程中的时序表达。结果:共培养后,MSCs的体积增大,其梭形形态逐渐变短变粗近似棒状或椭圆形,细胞之间形成连接,排列方向趋于一致。共培养两周时,α-actin及cTnT阳性细胞的比例分别为29.63%和27.38%。TGF-β、Nkx-2.5、GATA-4和MEF-2C基因在共培养后1 d表达开始增强,诱导后7 d达高峰,以后虽有所下降但仍维持在较高水平;TEF-1基因在诱导过程中表达无明显变化。结论:间接接触共培养条件下,MSCs可分化为心肌细胞。在此过程中,TGF-β、Nkx-2.5、GATA-4和MEF-2C基因可能是调控MSCs定向分化为心肌样细胞的重要调控基因。
[Abstract]:Aim: to study the differentiation of mesenchymal stem cells from bone marrow mesenchymal stem cells (MSCs) to myocadium-lkce cells and the expression of related regulatory genes in vitro, and to screen the important regulatory genes for the differentiation of MSCs into CMs. Methods: MSCs and CMs were cocultured at 1:5 and observed for two weeks. The morphological changes of MSCs were observed under phase contrast microscope. The expression of cardiac characteristic actin 伪 (伪 -actinin) and cardiac troponin TcTnT were detected by immunofluorescence staining. Semi-quantitative RT-PCR was used to analyze the temporal expression of related regulatory genes, such as TGF- 尾 -nkx-2.5, GATA-4, MEF-2C and TEF-1, during differentiation. Results: after co-culture, the volume of MSCs increased, the fusiform of MSCs became shorter and thicker, similar to rod or ellipse, the cells formed connection, and the direction of arrangement tended to be the same. After two weeks of co-culture, the rates of 伪 -actin and cTnT positive cells were 29.63% and 27.38%, respectively. The expression of 伪 -actin and cTnT gene began to increase on the 1st day after co-culture, and reached the peak at 7 days after induction. However, the expression of TEF-1 gene remained unchanged at a higher level during induction. Conclusion: MSCs can differentiate into cardiomyocytes under indirect contact co-culture. During this process, TGF- 尾 -nkx-2.5 MEF-2C gene and GATA-4 gene may be the important regulatory genes to regulate the directional differentiation of MSCs into cardiomyocyte-like cells.
【作者单位】：南昌大学第一附属医院心血管内科;南昌大学第一附属医院泌尿外科研究所;
【基金】：江西省卫生厅重大招标项目资助(200602)
【分类号】：R329.21

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