Cheng Ying, Wang Lu, Tang Ming, Yin Mengmeng, Cui Yurong, Liang Huamin, Song Yuanlong, Hu Xinwu, Luo Hongyan, Gao Ying, Wang Jiayu, Hescheler Jürgen, Xi Jiaoya
Department of Physiology and Chinese-German Stem Cell Center, Tongji Medical College, Huazhong University of Science and Technology, The Key Laboratory for Drug Target Researches and Pharmacodynamic Evaluation of Hubei Province, Wuhan, China.
Cell Physiol Biochem. 2013;32(4):789-800. doi: 10.1159/000354480. Epub 2013 Sep 20.
It is important to screen and identify chemical compounds to improve the efficiency of cardiac differentiation and specialization of embryonic stem (ES) cells. The objective of this study was to investigate the effect of puerarin, a natural phytoestrogen, on the in vitro cardiac differentiation and ventricular specialization of murine ES cells.
Cardiac differentiation of murine ES cells was performed by embryoid body (EB)-based differentiation method. Quantitative RT-PCR, flow cytometry and immunofluorescence were employed to identify cardiomyocytes (CMs) derived from murine ES cells (mES-CMs). Patch clamp was used to study the electrophysiological properties of CMs.
We found that continuous puerarin treatment significantly increased the population of ES-CMs which express typical cardiac markers and are electrophysiological intact. Puerarin treatment shifted the cardiac phenotype from pacemaker-like cells to ventricular-like cells, which were Mlc2v-positive and present typical ventricular-like AP. Puerarin up-regulated transcripts involved in cardiac differentiation and ventricular specialization of ES cells.
Our results suggest that puerarin promotes cardiac differentiation, and significantly enhances the specialization of mES cells into ventricular-like CMs. Puerarin may be used to increase the yield of ventricular mES-CMs during in vitro differentiation.
筛选和鉴定化合物对于提高胚胎干细胞（ES细胞）向心脏细胞分化及特化的效率至关重要。本研究旨在探讨天然植物雌激素葛根素对小鼠ES细胞体外心脏分化及心室特化的影响。
采用基于胚状体（EB）的分化方法诱导小鼠ES细胞向心脏细胞分化。运用定量逆转录聚合酶链反应（qRT-PCR）、流式细胞术和免疫荧光法鉴定源自小鼠ES细胞的心肌细胞（mES-CMs）。采用膜片钳技术研究心肌细胞的电生理特性。
我们发现持续给予葛根素处理可显著增加表达典型心脏标志物且电生理功能完整的ES-CMs数量。葛根素处理使心脏表型从起搏样细胞转变为心室样细胞，这些细胞Mlc2v呈阳性且具有典型的心室样动作电位（AP）。葛根素上调了参与ES细胞心脏分化和心室特化的转录本。
我们的结果表明，葛根素可促进心脏分化，并显著增强小鼠ES细胞向心室样心肌细胞的特化。葛根素可用于提高体外分化过程中心室mES-CMs的产量。
