Department of Biomedical Engineering, University of Minnesota - Twin Cities, 312 Church St. SE, Minneapolis, MN, 55455, USA.
Department of Biochemical Engineering, University of Applied Sciences Technikum Wien, Höchstädtplatz 5, 1200, Vienna, Austria.
Stem Cell Rev Rep. 2016 Dec;12(6):731-743. doi: 10.1007/s12015-016-9678-0.
Methods to generate murine cardiomyocytes from pluripotent stem cells (PSCs) in vitro are resource and time intensive. All current protocols require exogenously provided soluble factors and almost all utilize embryoid body formation to modulate pathways associated with mesoderm specification and cardiomyocyte differentiation. Here, we developed a simple protocol without EBs and without exogenous soluble factors that enabled cardiomyocyte differentiation of a murine induced PSC line based on controlled nutrient deprivation in 2D monolayer cultures. We showed that this protocol reproducibly imposed metabolic stress and consequently modulated active β-catenin levels to yield functional cardiomyocytes. The yield of cardiomyocytes and calcium handling kinetics were comparable to existing approaches. However, this approach did not produce consistent results between murine PSC lines suggesting signaling pathways linking nutrient deprivation to β-catenin activation are not universally conserved and may be a remnant of the parent population from which the induced PSCs were derived.
从多能干细胞(PSCs)体外生成鼠心肌细胞的方法既耗费资源又耗时。目前所有的方案都需要外源性提供可溶性因子,几乎所有方案都利用胚状体形成来调节与中胚层特化和心肌细胞分化相关的途径。在这里,我们开发了一种简单的方案,无需 EB 且无需外源性可溶性因子,该方案能够在 2D 单层培养物中基于受控的营养剥夺使鼠诱导 PSC 系分化为心肌细胞。我们表明,该方案可重复性地施加代谢应激,从而调节活性 β-连环蛋白水平以产生功能正常的心肌细胞。心肌细胞的产量和钙处理动力学与现有的方法相当。然而,这种方法在鼠 PSC 系之间没有产生一致的结果,这表明将营养剥夺与β-连环蛋白激活联系起来的信号通路并非普遍保守,并且可能是诱导 PSCs 来源的亲本群体的残余物。