Specific Arrest of Cardiogenesis in Cultured Embryonic Stem Cells Lacking Cripto-1

Abstract

The molecular events of cardiac lineage specification and differentiation are largely unknown. Here we describe the involvement of a growth factor with an EGF-like domain, Cripto-1 (Cr-1), in cardiac differentiation. During embryonic development, Cr-1 is expressed in the mouse blastocyst, primitive streak, and later is restricted to the developing heart. To investigate the role of Cr-1, we have generated Cr-1-negative embryonic stem (ES) cell lines by homologous recombination. The resulting double "knockout" ES cells have selectively lost the ability to form beating cardiac myocytes, a process that can be rescued by reintroducing Cr-1 gene back into the Cr(-/-) cells. Furthermore, the lack of functional Cr-1 is correlated with absence of expression of cardiac-specific myosin light and heavy chain genes during differentiation. Differentiation into other cell types including skeletal muscle is not disrupted. These results suggest that Cr-1 is essential for contractile cardiomyocyte formation in vitro.

Cardiac patterning and morphogenesis in zebrafish

“…Although not all of these factors have been studied in the same manner in other organisms, it is likely that their functions are conserved across taxa. For example, many experiments suggest that the roles of the Bmp, Nodal, and RA signaling pathways during heart development are similar in different species (e.g., Schultheiss et al, 1997;Xu et al, 1998Xu et al, , 1999Rosenthal and Xavier-Neto, 2000). Rather than discuss all of the experiments that demonstrate similarities in cardiac patterning mechanisms, this section will simply describe data relevant to the conserved roles of Gata transcription factors during cardiogenesis.…”

Nodal-dependant Cripto signaling in ES cells: from stem cells to tumor biology

“…Recently, a great effort has been undertaken to unravel the role of cripto in development and ES cell differentiation (Ding et al, 1998;Xu et al, 1998). In mouse embryos, cripto is expressed early in the ICM and the throphoblast cells of the blastocyst (Johnson et al, 1994).…”

“…Mouse embryos deficient for the cripto gene die around day 7.5 of embryogenesis due to defects in mesoderm formation and axial organization (Ding et al, 1998;Liguori et al, 2003). Notably, mouse cripto mutants exhibit defects in myocardial development as evidenced by the absence of expression of terminal myocardial differentiation genes such as aMHC and MLC2v (Ding et al, 1998;Xu et al, 1998). Unfortunately, the early lethality observed in cripto À/À mice occurs before a stage in which the role of cripto in cardiogenesis can be evaluated.…”

“…Unfortunately, the early lethality observed in cripto À/À mice occurs before a stage in which the role of cripto in cardiogenesis can be evaluated. In vitro analysis of Cripto-deficient ES cells, by overcoming the complexity of the in vivo situation, led to novel insights into the functional role of cripto in cardiomiogenesis (Xu et al, 1998(Xu et al, , 1999Parisi et al, 2003).…”

“…By using EBs derived from Cripto À/À ES cells, it has been shown that cripto is essential for cardiogenesis in culture (Xu et al, 1998); indeed, Cripto À/À EBs do not produce contracting cardiomyocytes even during extended culture periods (Xu et al, 1998). These cells thus represent a good source to explore the molecular mechanisms underlying this process, and have been successfully used to elucidate the dynamic of Cripto signaling required for differentiation of cardiac precursors from ES cells.…”