Institut für Molekularbiologie, Medizinische Hochschule Hannover, Hannover, Germany.
Circ Res. 2012 Jul 6;111(2):165-9. doi: 10.1161/CIRCRESAHA.112.273946. Epub 2012 Jun 12.
The embryonic epicardium is a crucial cell source of the cardiac fibrous skeleton as well as of the coronary system. Genetic lineage tracing systems based on Wt1 regulatory sequences provided evidence that epicardium-derived cells also adopt a myocardial fate in the mouse.
To define the adequacy of Wt1-based lineage tracing systems for epicardial fate mapping.
Using in situ hybridization analysis and immunofluorescence on tissue sections, we detected endogenous expression of Wt1 mRNA and Wt1 protein in the proepicardium and epicardium and also in endothelial cells throughout cardiogenesis. Expression analysis of a sensitive GFP reporter showed that recombination mediated by cre recombinase in the Wt1(creEGFP) line occurs randomly and sporadically in all cells of the embryo. Recombination in cardiomyocytes was found in the linear heart tube before establishment of a (pro)epicardium. In contrast, the tamoxifen-inducible Wt1(creERT2) mouse line mediated poor and variable recombination in the epicardium. Recombination in cardiomyocytes was not detected in this case.
Frequently used Wt1 based cre-mediated lineage tracing systems are not suitable for epicardial fate mapping because of endogenous endothelial expression of Wt1, ectopic recombination (Wt1(creEGFP)), and poor recombination efficiency (Wt1(creERT2)) in the developing heart. We conclude that claims of a cardiomyocyte fate of epicardial cells in the mouse are not substantiated.
胚胎的心外膜是心脏纤维骨架和冠状系统的重要细胞来源。基于 Wt1 调控序列的遗传谱系追踪系统提供的证据表明，心外膜衍生细胞在小鼠中也具有心肌命运。
确定基于 Wt1 的谱系追踪系统在心外膜命运图谱中的充分性。
通过原位杂交分析和组织切片免疫荧光，我们检测到 Wt1 mRNA 和 Wt1 蛋白在内胚层和心外膜中的内源性表达，以及在整个心脏发生过程中的内皮细胞中。对敏感 GFP 报告基因的表达分析表明，在 Wt1(creEGFP)系中，cre 重组酶介导的重组随机且散在地发生在胚胎的所有细胞中。在（前）心外膜建立之前，在线性心脏管中发现了心肌细胞中的重组。相比之下，他莫昔芬诱导的 Wt1(creERT2)小鼠系在心外膜中介导的重组较差且可变。在这种情况下，未检测到心肌细胞中的重组。
由于 Wt1 的内皮细胞表达、异位重组（Wt1(creEGFP)）和发育心脏中的重组效率差（Wt1(creERT2)），常用的基于 Wt1 的 cre 介导的谱系追踪系统不适合心外膜命运图谱。我们得出结论，小鼠心外膜细胞具有心肌细胞命运的说法没有得到证实。
