Cardiac anterior-posterior patterning by retinoic acid: developmental mechanisms for the raldh2 caudorostral wave.

Abstract

Congenital heart defects (CHD) constitute the most common type of birth defects and are an important cause of early death and increased risk of cardiac problems in adulthood. Increasing knowledge of heart organogenesis can improve CHD diagnosis and treatment. Early establishment of anteroposterior (AP) orientation of precursors is a key step in cardiogenesis because anterior, posterior, and intermediate segments give rise to cardiac outflow tract, atria, and ventricles, respectively. Previous findings from the laboratory indicated that a wave of RALDH2 promotes asymmetric synthesis of retinoic acid (RA) in posterior cardiac mesoderm, conferring AP polarity to the cardiac field. In this project, we propose to investigate the developmental mechanisms responsible for this Raldh2 caudorostral wave progression in the cardiac field. First, we aim to confirm in a quantitative fashion, our preliminary qualitative results indicating that the RALDH2 wave depends on the induction of Raldh2 in anterior mesoderm cells, Father than to a cephalic migration of Raldh2 expressing cells from the posterior region. This will be achieved with culture of isolated or combined anterior and posterior mesoderm explants from chicken and quail embryos. Genetic polymorphisms and antigen specificity between chicken and quail will allow us to ascertain, by in situ hybridization, immunohistochemistry and Real-Time PCR, the species origin of Raldh2 expression in the anterior, induced fragment. Second, we aim to identify candidate regulators of RALDH2 expression by analyzing differential expression in anterior versus posterior cardiac precursors using a microarray approach, as well as, by analyzing the expression patterns of possible RALDH2-interactors spotted out from literature. Over expression or silencing of candidate genes will be produced by chicken electroporation. We expect to identify key players in the early steps of heart development and major actors of cardiac AP patterning with the potential for causative or modifier roles in CHD. This project will develop the second item of the FAPESP project 06/50843-0 "Evolução e Desenvolvimento das Câmaras Cardíacas". (AU)