Department of Cardiology, Xijing Hospital, Fourth Military Medical University, Xi'an, Shaanxi 710032, China.
Chin Med J (Engl). 2013;126(18):3528-33.
Bone marrow derived mesenchymal stem cells (BMdMSCs) can differentiate into cardiomyocyte-like cells induced by different inductors individually or collectively. In this study, by inducing BMdMSCs with p53 inhibitor (p-fifty three inhibitor-alpha, PFT-α), 5-azacytidine (5-AZA), angiotensin-II (Ang-II) and bone morphogenic protein-2 (BMP-2) we compared the influences of four inductors on the differentiation of rat BMdMSCs into caridomyocyte like-cells.
BMdMSCs were collected from the bone marrow of Sprague Dawley rats and after the fourth generation, the purified cells were divided into five groups: 5-AZA (10 µmol/L), Ang-II (0.1 µmol/L), PFT-α (20 µmol/L), BMP-2 (10 µg/L) and control. The purity of the BMdMSCs and the cardiac differentiation rates were obtained by flow cytometry. The expressions of cTnT in the BMdMSCs after four weeks of induction were detected by immunofluorescence and the expressions of cTnI and Cx43 detected by Western blotting. The green fluorescent levels reflecting intracellular calcium transient function were determined by laser scanning confocal microscopy. The total potassium current levels of cells were measured on patch clamp.
All inductors affected to a different degree the differentiation of BMdMSCs into cardiomyocyte-like cells and the expressions of cTnT, cTnI and Cx43 suggesting that the combination of inductors could be an improved method for cardiac regenerative medicine. In addition, the total potassium current level and calcium transient in PFT-α cardiomyocyte-like cells were higher than other groups.
The cardiac differentiation of BMdMSCs induced by PFT-α, 5-AZA, Ang-II and BMP-2 has been improved at different levels. PFT-α has an advantage of differentiation rate and electrophysiological function over other inductors.
骨髓间充质干细胞（BMdMSCs）可在不同诱导剂的单独或共同作用下分化为心肌样细胞。本研究通过 p53 抑制剂（p53 抑制剂-α，PFT-α）、5-氮杂胞苷（5-AZA）、血管紧张素-II（Ang-II）和骨形态发生蛋白-2（BMP-2）诱导 BMdMSCs，比较了四种诱导剂对大鼠 BMdMSCs 向心肌样细胞分化的影响。
从 Sprague Dawley 大鼠的骨髓中分离出 BMdMSCs，在第 4 代后，将纯化的细胞分为 5 组：5-AZA（10μmol/L）、Ang-II（0.1μmol/L）、PFT-α（20μmol/L）、BMP-2（10μg/L）和对照组。通过流式细胞术获得 BMdMSCs 的纯度和心脏分化率。免疫荧光法检测 4 周诱导后 cTnT 的表达，Western blot 法检测 cTnI 和 Cx43 的表达。激光扫描共聚焦显微镜测定反映细胞内钙瞬变功能的绿色荧光强度。通过膜片钳法测量细胞的总钾电流水平。
所有诱导剂均不同程度地影响 BMdMSCs 向心肌样细胞的分化以及 cTnT、cTnI 和 Cx43 的表达，提示诱导剂的联合应用可能是心脏再生医学的一种改进方法。此外，PFT-α 心肌样细胞的总钾电流水平和钙瞬变高于其他组。
PFT-α、5-AZA、Ang-II 和 BMP-2 诱导的 BMdMSCs 的心脏分化在不同程度上得到了改善。PFT-α 在分化率和电生理功能方面优于其他诱导剂。
