Investigating the mechanisms of cardiac patterning and morphogenesis using a heart formation assay

Doctoral thesis English OPEN
Caporilli, Simona
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Vertebrate heart development involves a precise sequence of morphogenetic events from which a complex structure is formed from a linear heart tube. To study the heart development in mammals is difficult because most alterations of heart structure are lethal. Therefore we use alternative model, Xenopus laevis embryos. The aim of this project is try to establish a new experimental model to help understanding the mechanism that regulates cardiac cell diversification and heart morphogenesis. In order to achieve these goals we use two assays. The cardiogenesis assay involves the use of animal cap explants excised from the animal pole of blastula embryos. It has been previously established that it is possible to induce differentiation of cardiac tissue in the same explants via the injection of GATA-4 mRNA. Here it is shown that GATA-4 reliably induces the expression of ventricular and proepicardial markers, providing an assay to study the mechanisms of cardiac cell fate diversification. However, despite these, cardiomyocytes generated in animal pole explants they do not undergo significant morphogenesis and physiological maturation. In order to study these later aspects of heart development we required a different assay in which was possible to generate a structure similar to the heart. Using GATA-4 injected AC explants transplanted into host embryos we obtained secondary beating hearts in which regionally restricted cardiac gene expression was observed in addition to growth and a limited degree of morphogenesis. We demonstrated that the host plays an essential role as it provides a wide range of permissive regions which allow the development of the SH. Moreover, we also showed that the competence to generate a secondary heart is lost in reaggregates transplanted at stage 28. The host cells however do not contribute to the SH indicating that the role of the host is providing signals which allow the development of the SH. In the future we aim to investigate the signalling pathways which mediate the host-SH interaction and the mechanism by which they allow the development of the secondary structure.
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