Arai A, Yamamoto K, Toyama J
Department of Circulation, The Research Institute of Environmental Medicine, Nagoya University, Japan.
Dev Dyn. 1997 Nov;210(3):344-53. doi: 10.1002/(SICI)1097-0177(199711)210:3<344::AID-AJA13>3.0.CO;2-A.
Cardiac progenitor cells in avian and amphibian embryos are known to commit to cardiac lineage during gastrulation or early neurulation. These cells require cell interaction with anterior endoderm for their differentiation into cardiomyocytes. However, little is known about cell interaction in mammalian cardiogenesis. We investigated the staging of murine cardiomyocyte commitment and the role of cell interaction in differentiation of cardiac progenitor cells into cardiomyocytes, using cultures of various embryonic regions at 7.25 and 7.5 days post coitum (p.c.), respectively. To evaluate the terminal differentiation of cardiac progenitor cells, we employed three parameters; expression of spontaneous beating, myosin heavy chain (MHC) protein, and cardiac-specific genes (alpha myosin heavy chain, Csx/Nkx2.5 and myosin light chain 2V genes). mRNAs of cardiac-specific genes were detected in 7.25-day p.c. mesoderm by RT-PCR, suggesting that the genetic specification to cardiac lineage initiated in the mesoderm by 7.25 days p.c. The 7.25-day p.c. isolated mesoderm in 48 hr culture, however, failed to differentiate into spontaneous beating cardiomyocytes and exhibited non-organized MHC protein in 19% of these culture. In contrast, all of the 7.5-day p.c. isolated mesoderm differentiated into beating cardiomyocytes even in 24 hr culture. The 7.25-day p.c. mesoderm associated with primitive streak increased MHC protein expression in 93% of these cultures, although they formed beating foci in 3%. The 7.25-day p.c. explants containing both visceral embryonic endoderm and primitive streak succeeded in terminal differentiation into spontaneous beating cardiomyocytes. Our study suggests that cardiac progenitor cells obtain the potency to complete terminal differentiation autonomously at 7.5 days p.c., as a consequence of the multistep induction by cell interactions with both the primitive streak and visceral embryonic endoderm, following the genetic specification to cardiac lineage in the early gastrula stage.
已知鸟类和两栖类胚胎中的心脏祖细胞在原肠胚形成期或早期神经胚形成期确定为心脏谱系。这些细胞需要与前肠内胚层进行细胞间相互作用才能分化为心肌细胞。然而，关于哺乳动物心脏发生过程中的细胞间相互作用却知之甚少。我们分别使用交配后7.25天和7.5天的不同胚胎区域培养物，研究了小鼠心肌细胞确定的阶段以及细胞间相互作用在心脏祖细胞分化为心肌细胞过程中的作用。为了评估心脏祖细胞的终末分化，我们采用了三个参数：自发搏动的表达、肌球蛋白重链（MHC）蛋白以及心脏特异性基因（α-肌球蛋白重链、Csx/Nkx2.5和肌球蛋白轻链2V基因）。通过逆转录聚合酶链反应（RT-PCR）在交配后7.25天的中胚层中检测到心脏特异性基因的mRNA，这表明到交配后7.25天时，中胚层已开始向心脏谱系进行基因定向。然而，交配后7.25天分离的中胚层在48小时培养后未能分化为自发搏动的心肌细胞，并且在这些培养物中有19%表现出MHC蛋白无序分布。相比之下，交配后7.5天分离的所有中胚层即使在24小时培养后也能分化为搏动的心肌细胞。交配后7.25天与原条相关的中胚层在93%的这些培养物中增加了MHC蛋白表达，尽管它们只有3%形成了搏动灶。含有内脏胚胎内胚层和原条的交配后7.25天的外植体成功地终末分化为自发搏动的心肌细胞。我们的研究表明，由于在原肠胚早期阶段向心脏谱系进行基因定向后，与原条和内脏胚胎内胚层的细胞间相互作用进行了多步骤诱导，心脏祖细胞在交配后7.5天时获得了自主完成终末分化的能力。
