VIB-KU Leuven Center for Brain & Disease Research, KU Leuven, Belgium.
Department of Human Genetics, KU Leuven, Belgium.
Sci Rep. 2018 Mar 16;8(1):4722. doi: 10.1038/s41598-018-22971-w.
Atrioventricular septal defects often result from impaired endocardial cushion development. Endothelial-to-mesenchymal transition (EndoMT) is a critical event in endocardial cushion development that initiates in the atrioventricular canal (AVC). In ex vivo EndoMT studies, mouse AVCs are flat-mounted on a collagen gel. In the explant outgrowths, the ratio of elongated spindle-like mesenchymal cells over cobblestone-shaped cells, generally considered as endothelial cells, reflects EndoMT. Using this method, several key signalling pathways have been attributed important functions during EndoMT. Using genetic lineage tracing and cell-type-specific markers, we show that monolayers of cobblestone-shaped cells are predominantly of epicardial rather than endothelial origin. Furthermore, this epicardium is competent to undergo mesenchymal transition. Contamination by epicardium is common and inherent as this tissue progressively attaches to AVC myocardium. Inhibition of TGFβ signalling, previously shown to blunt EndoMT, caused an enrichment in epicardial monolayers. The presence of epicardium thus confounds interpretations of EndoMT signalling pathways in this assay. We advocate to systematically use lineage tracers and cell-type-specific markers on stage-matched AVC explants. Furthermore, a careful reconsideration of earlier studies on EndoMT using this explant assay may identify unanticipated epicardial effects and/or the presence of epicardial-to-mesenchymal transition (EpiMT), which would alter the interpretation of results on endothelial-to-mesenchymal transition.
房室间隔缺损通常是由于心内膜垫发育不良引起的。内皮-间质转化（EndoMT）是心内膜垫发育过程中的一个关键事件，起始于心房-心室管（AVC）。在体外 EndoMT 研究中，将小鼠 AVC 平铺在胶原蛋白凝胶上。在培养物的外生突中，长梭形间充质细胞与鹅卵石状细胞（通常被认为是内皮细胞）的比例反映了 EndoMT。使用这种方法，几个关键的信号通路在 EndoMT 中被认为具有重要作用。通过遗传谱系追踪和细胞类型特异性标志物，我们表明鹅卵石状细胞的单层主要来源于心外膜而不是内皮细胞。此外，这种心外膜具有进行间质转化的能力。由于这种组织逐渐附着于 AVC 心肌，因此心包的污染是常见且固有的。先前已显示抑制 TGFβ 信号通路可阻止 EndoMT，这导致心包单层的富集。因此，心包的存在使该测定中 EndoMT 信号通路的解释变得复杂。我们主张在匹配阶段的 AVC 外植体上系统地使用谱系示踪剂和细胞类型特异性标志物。此外，使用这种外植体测定对早期关于 EndoMT 的研究进行仔细的重新考虑，可能会发现意想不到的心外膜效应和/或心外膜-间质转化（EpiMT）的存在，这将改变对内皮-间质转化结果的解释。
