Expression Patterns Of Nkx2.5, ISL-1 During Early Development Of The Mouse Embryonic Heart

Background:The embryonic heart morphogenesis is regulated by different signal system and transcriptional factors. Under the regulation of these signals and factors, cardiagenic plate which is called primary heart field is fused to form the primary heart tube with the embryonic disc folding. A body of investigations have demonstrated that the primary heart tube only developes into the left ventricle, while the outflow tract, right ventricle and atrium of the embryonic heart are formed by addition of cardiomyocytes, which are originated from the mesenchyme ventral to the foregut called the second heart field, into the primary heart tube at both venous and arterial pole. With the experiments of transgene and gene deletion, transcriptional factors Nkx2.5 and ISL-1 are considered to be the important proteins to regulate the differentiation of the cardiac precursor cells into the cardiomyocytes in the primary heart field and second heart field. Nkx2.5 and ISL-1 knockout or mutation result in the congenital heart malformations or embryo death. The expression patterns of Nkx2.5 and ISL-1 during early development of the embryonic heart, as well as in the second heart field have not been reported. Elucidation of the expression patterns of Nkx2.5 and ISL-1 during normal heart morphogenesis can provide insight into the mechanism of heart development.Objective:To investigate the relationship of Nkx2.5 and ISL-1 expression with early development of the mouse embryonic heart.Methods:Serial sections of mouse embryos from embryonic day(ED) 7.5 to embryonic day(ED) 13 were stained with antibodies against a-SMA, MHC, Nkx2.5 and ISL-1.Results:At ED 7.5, strong expression of Nkx2.5 was found in the cardiogenic plate, earlier than that of heart specific proteins, MHC and SMA. From ED 8 to ED 12, following the morphological establishment of the heart, the expression of Nkx2.5 and MHC in heart became similar and strong. The initial expression of ISL-1 was detected in the dorsal mesocardium and the splanchnic mesoderm of dorsal pericardial cavity wall ventral to the foregut at ED 8. At ED 9, the Nkx2.5 expression could be seen extending from arterial pole of primary heart tube to the splanchnic mesoderm dorsal to the pericardial cavity and paraxial mesoderm around the primary pharynx to involve in the development of the outflow tract myocardium. After ED 10, most dorsal mesocardium have disappeared, and the number of ISL-1 positive cells increased evidently, forming a continuous band ventral to the foregut that connected the distal pole of the outflow tract with the dorsal wall of the atrium. At ED 11-12, with the progression of heart looping and morphogenesis, the cells in the distal pole of the outflow tract gradually shed Nkx2.5 and MHC expression with increasing SMA and ISL-1 expression. The number of ISL-1 positive cells reached highest level at ED 11-12, especially in arterious pole. After ED 13, the expression intensity of Nkx2.5 in heart and ISL-1 in mesenchymal cells ventral to the foregut gradually decreased.Conclusion:Nkx2.5 is an early marker of the cardiac precursor cells in the primary heart field. Its expression in the second heart field subset is confined to the splanchnic mesoderm and paraxial mesoderm surrounding the pharynx at ED 9 to involve in the development of the outflow tract myocardium. The initial expression of ISL-1 in the dorsal mesocardium and in the splanchnic mesoderm ventral to the foregut was found at ED 8 and the number of ISL-1 positive cells reached the highest level at ED 11-12, suggesting that it is during the period of ED 8 to ED 12 that the addition of myocardial cells into the developing heart from the second heart field drives the heart tube to complete heart looping and morphogenesis. At ED 11-12, the mesenchymal cells derived from the second heart field began differentiating into smooth muscle in the walls of the aorta and pulmonary trunk.