Cardiovascular Department, Xin Hua Hospital, Affiliated Hospital of Shanghai Jiaotong University, Shanghai 200092, P.R. China.
Mol Med Rep. 2017 Nov;16(5):5887-5893. doi: 10.3892/mmr.2017.7373. Epub 2017 Aug 28.
As a novel type of seed cell, amniotic fluid‑derived mesenchymal stem cells (AFMSCs) are promising for the regeneration of myocardial cells. A focus of cardiovascular regenerative medicine is to improve the efficiency of AFMSC differentiation. The present study replaced the traditional method of AFMSC differentiation with a combined induction method, in order to improve the efficiency of directional differentiation. AFMSCs were obtained from rabbit amniotic fluid samples, and western blot analysis was performed to analyze the expression of octamer‑binding transcription factor 4 (OCT4), and tumorigenicity experiments were conducted. AFMSCs were divided into the following 4 groups: Induction with transforming growth factor β1 (TGFβ1); induction with 5‑azacytidine (5Aza); induction with TGFβ1 and 5Aza combined; and untreated controls. Reverse transcription‑quantitative polymerase chain reaction was performed to analyze the expression of cardiac‑specific GATA binding protein 4 (GATA4), and immunofluorescence was employed to analyze the expression of cardiac troponin T (cTnT). In addition, western blotting was performed to analyze the expression of connexin 43, and transmission electron microscopy was used to observe the ultrastructure of the differentiated cells. AFMSCs exhibited positive OCT4 expression and were not observed to induce tumor development in nude mice. The expression levels of GATA4, cTnT, and connexin 43 in the combined induction group were markedly higher when compared with the remaining groups. Transmission electron microscopy analysis revealed that differentiated cells exhibited myocardial cell characteristics. In conclusion, AFMSCs are multipotent, non‑tumorigenic cells that are capable of differentiating into cardiomyocyte‑like cells. This combined induction method may improve the efficiency of directed differentiation.
作为一种新型的种子细胞，羊水来源间充质干细胞（AFMSCs）在心肌细胞再生方面具有广阔的应用前景。心血管再生医学的研究重点是提高 AFMSC 分化效率。本研究用联合诱导法取代传统的 AFMSC 分化方法，以提高定向分化的效率。本研究从兔羊水样本中获取 AFMSCs，采用 Western blot 分析检测八聚体结合转录因子 4（OCT4）的表达，并进行致瘤性实验。将 AFMSCs 分为以下 4 组：TGFβ1 诱导组；5-氮杂胞苷（5Aza）诱导组；TGFβ1 和 5Aza 联合诱导组；未处理对照组。采用反转录-定量聚合酶链反应分析心脏特异性 GATA 结合蛋白 4（GATA4）的表达，采用免疫荧光法分析心肌肌钙蛋白 T（cTnT）的表达。此外，采用 Western blot 分析分析连接蛋白 43 的表达，采用透射电子显微镜观察分化细胞的超微结构。AFMSCs 呈 OCT4 阳性表达，且在裸鼠中未观察到致瘤性。与其余各组相比，联合诱导组的 GATA4、cTnT 和连接蛋白 43 的表达水平明显更高。透射电子显微镜分析显示分化细胞具有心肌细胞特征。综上所述，AFMSCs 是多能、无致瘤性的细胞，能够分化为心肌样细胞。这种联合诱导方法可能提高定向分化的效率。
