Abstract 14208: Versican is Essential for Chamber Formation by Recruiting Cardiac Progenitor Cells from the Outflow Tract in Medaka Fish
Background: Heart tube formation from primary heart field precedes rightward looping and chamber formation. Ventricular chamber growth requires cardiomyocyte proliferation and progenitor cells migration from outside the heart tube. We investigated how these progenitors are integrated into heart.
Methods and Results: Using forward-genetic N-ethyl-N-nitrosourea (ENU) mutagenesis screening, we identified a cardiovascular mutant Medaka fish called big heart tube (bht). Mutant embryos showed hypomorphic noncontracting ventricle and constricted outflow tract, whereby looping failed and development terminated at the linear heart tube stage. Positional cloning and sequencing analyses revealed a mutation in the 3′-untranslated region of versican, a proteoglycan gene. Semiquantitative RT-PCR showed normal versican mRNA levels in the bht mutant, but significantly decreased protein levels. Injection of a versican-specific morpholino recapitulated the bht phenotype, suggesting that bht is a loss-of-function mutant. Cell proliferation was assayed by phosphorylated-histone-H3 staining, and cardiomyocytes were counted by MEF2 staining. Cardiac cell proliferation in the wild-type linear heart tube was rare (n = 3), but cardiomyocyte numbers increased during cardiac looping (n = 301 to 590). A large fraction of the proliferating cells was located at the outflow. Thus, ventricular chamber growth occurs through the addition of new cardiomyocytes. The bht mutant exhibited normal development until the heart tube stage, shown by the expression of early-differentiation markers amhc, vmhc, and cmlc2; however, ventricular chamber formation was disrupted, and cardiomyocyte numbers remained unchanged. New cmlc2-positive cells accumulated around the outflow tract in the bht mutant, but did not integrate into the ventricular chamber. Versican mRNA was expressed in outflow tract during the heart tube stage. Photoconverted Kaede labeled newly differentiated cardiomyocytes in the ventricular chamber from outflow in wild-type fish, but this recruitment of cardiac progenitors was disrupted in the bht mutant.
Conclusions: Versican is essential for ventricular chamber formation through recruitment of cardiac progenitor cells from outflow tract.
- © 2010 by American Heart Association, Inc.