In Vitro Conversion of Murine Fibroblasts into Cardiomyocyte-Like Cells.
In Vitro Conversion of Murine Fibroblasts into Cardiomyocyte-Like Cells.
复制标题
鼠的成纤维细胞体外转化为心肌细胞样细胞。
DOI：
10.1007/978-1-0716-0668-1_12
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Qian L
中科院分区：
文献类型：
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作者：
Xu J;Wang L;Liu J;Qian L
将成纤维细胞直接重编程为诱导的心肌细胞（iCM）作为心血管疾病的潜在治疗方法具有很大的前景，其中许多与功能性心肌细胞的巨大损失和同时形成瘢痕组织有关。新兴的研究已经表明，引入三种最小的转录因子Gata 4、Mef 2c和Tbx 5（G/M/T）可以将鼠成纤维细胞转化为在体外和体内都与内源性CM非常相似的iCM。最近关于iCM细胞命运决定的研究表明，去除遗传和表观遗传障碍可以促进iCM重编程。然而，不同的研究小组之间的重编程效率不同，阻碍了其进一步研究和潜在的应用。在这里，我们提供了一个新的更新和详细的协议，在体外生成和评价功能iCM从小鼠胚胎成纤维细胞和新生儿心脏成纤维细胞使用逆转录病毒多顺反子构建编码的G/M/T因子的最佳表达。我们希望这种优化的方案将奠定基础，为未来的机制研究的小鼠iCM和进一步改善iCM的生产。
Direct reprogramming of fibroblasts into induced cardiomyocytes (iCMs) holds great promise as a potential treatment for cardiovascular disease, many of which are associated with tremendous loss of functional cardiomyocytes and simultaneous formation of scar tissue. Burgeoning studies have shown that the introduction of three minimal transcriptional factors, Gata4, Mef2c, and Tbx5 (G/M/T), could convert murine fibroblasts into iCMs that closely resemble endogenous CMs both in vitro and in vivo. Recent studies on iCM cell fate determination have demonstrated that the removal of genetic and epigenetic barriers could facilitate iCM reprogramming. However, varied reprogramming efficiency among research groups hinders its further study and potential applicability. Here, we provide a newly updated and detailed protocol for in vitro generation and evaluation of functional iCMs from mouse embryonic fibroblasts and neonatal cardiac fibroblasts using retroviral polycistronic construct encoding optimal expression of G/M/T factors. We hope that this optimized protocol will lay the foundation for future mechanistic studies of murine iCMs and further improvement of iCM generation.