Uncovering the origins of myocardial cells is important for understanding and treating heart diseases. Cai et al. suggest that Tbx18-expressing epicardium provides a substantial contribution to myocytes in the ventricular septum and the atrial and ventricular walls. Here we show that the T-box transcription factor gene 18 (Tbx18) itself is expressed in the myocardium, showing that their genetic lineage tracing system does not allow conclusions of an epicardial origin of cardiomyocytes in vivo to be drawn.

Abstract-The primary myocardium of the embryonic heart, including the atrioventricular canal and outflow tract, is essential for septation and valve formation. In the chamber-forming heart, the expression of the T-box transcription factor Tbx2 is restricted to the primary myocardium. To gain insight into the cellular contributions of the Tbx2 ϩ primary myocardium to the components of the definitive heart, genetic lineage tracing was performed using a novel Tbx2 Cre allele. These analyses revealed that progeny of Tbx2 ϩ cells provide an unexpectedly large contribution to the Tbx2-negative ventricles. Contrary to common assumption, we found that the embryonic left ventricle only forms the left part of the definitive ventricular septum and the apex. The atrioventricular node, but not the atrioventricular bundle, was found to derive from Tbx2 ϩ cells. The Tbx2 ϩ outflow tract formed the right ventricle and right part of the ventricular septum. In Tbx2-deficient embryos, the left-sided atrioventricular canal was found to prematurely differentiate to chamber myocardium and to proliferate at increased rates similar to those of chamber myocardium. As a result, the atrioventricular junction and base of the left ventricle were malformed. Together, these observations indicate that Tbx2 temporally suppresses differentiation and proliferation of primary myocardial cells. A subset of these Tbx2Cre -marked cells switch off expression of Tbx2, which allows them to differentiate into chamber myocardium, to initiate proliferation, and to provide a large contribution to the ventricles. These findings imply that errors in the development of the early atrioventricular canal may affect a much larger region than previously anticipated, including the ventricular base. Key Words: atrioventricular canal Ⅲ lineage Ⅲ fate Ⅲ patterning Ⅲ transgenic Ⅲ Cre T he embryonic heart tube is composed of "primary" myocardium and rapidly elongates by addition of progenitor cells to its poles. During looping, specific regions in the embryonic tubular heart differentiate to chamber myocardium and expand to form the future working myocardium of the ventricles and atria. In contrast, the region in between these expanding chambers does not differentiate or expand and becomes visible as an atrioventricular constriction. 1 During prenatal life, the atrioventricular canal (AVC) myocardium conducts the electric impulse between the atrial and ventricular chambers in a slow manner, reminiscent of the function of the mature atrioventricular (AV) node. This slow conducting feature allows the AVC to act as a sphincter preventing backflow from the ventricles to the atria, analogous to the AV valves. Furthermore, the primary myocardium provides the signals that initiate formation of the cushions, which subsequently will form the valves and partake in septation. 2,3 The primary myocardial AVC is extensively remodeled to properly connect and align both atria and ventricles and to coordinate the formation of the fibrous insulation. 4 Given all these roles of the AV...

Rationale:The clinically important atrioventricular conduction axis is structurally complex and heterogeneous, and its molecular composition and developmental origin are uncertain.Objective: To assess the molecular composition and 3D architecture of the atrioventricular conduction axis in the postnatal mouse heart and to define the developmental origin of its component parts. Methods and Results:We generated an interactive 3D model of the atrioventricular junctions in the mouse heart using the patterns of expression of Tbx3, Hcn4, Cx40, Cx43, Cx45, and Nav1.5, which are important for conduction system function. We found extensive figure-of-eight rings of nodal and transitional cells around the mitral and tricuspid junctions and in the base of the atrial septum. The rings included the compact node and nodal extensions. We then used genetic lineage labeling tools (Tbx2 ؉/Cre , Mef2c-AHF-Cre, Tbx18 ؉/Cre ), along with morphometric analyses, to assess the developmental origin of the specific components of the axis. The majority of the atrial components, including the atrioventricular rings and compact node, are derived from the embryonic atrioventricular canal. The atrioventricular bundle, including the lower cells of the atrioventricular node, in contrast, is derived from the ventricular myocardium. No contributions to the conduction system myocardium were identified from the sinus venosus, the epicardium, or the dorsal mesenchymal protrusion. Key Words: atrioventricular canal Ⅲ atrioventricular node Ⅲ three-dimensional reconstruction Ⅲ lineage analysis Ⅲ heart development Ⅲ transgenic mice T he atria and ventricles are separated by the connective tissues of the atrioventricular junction that insulate the atrial and ventricular muscle masses. A small part of the musculature, however, the atrioventricular conduction axis, crosses the plane of insulation, thus allowing conduction of the impulse generated by the sinus node to the ventricles. The axis has atrial parts, including the atrioventricular node and atrioventricular ring bundles, and ventricular parts, the atrioventricular bundle and the bundle branches. 1,2 The atrioventricular node delays the electric impulse, thus permitting the ventricles to fill before ventricular contraction. The node can also function as a subsidiary pacemaker. Several arrhythmias, such as atrioventricular block and reentrant tachycardia, have their anatomic substrates within the axis, 2-5 which is complex and heterogeneous in terms of its morphology. [2][3][4][5][6] Insight into the mechanisms of the arrhythmias can be provided by an understanding of development. Of the transcription factors implicated in the regulation of the developmental process, transcription factor Tbx3 is expressed specifically in the central conduction system, thus providing a key marker with which to delineate these tissues throughout development and in the adult. 7-9 The origin and lineages of the atrioventricular junctions, however, have still to be clarified. It is currently thought that the atrial component...

Our data indicate that the cardiogenic mesoderm contains an additional progenitor subpopulation that contributes to the sinus venosus myocardium. After patterning of the cardiogenic mesoderm, this progenitor population remains spatially separated and genetically distinctive from the second heart field subpopulation.