Abstract 2108: Notch1-Dependent Cardiomyogenesis is an Essential Component of Postnatal Cardiac Growth
The objective of this work was to determine whether activation of cardiac progenitor cells (CPCs), their commitment to the myocyte lineage and cardiomyogenesis contribute to cardiac growth postnatally. In the adult heart following injury, Notch1 controls new myocyte formation. However, the function of Notch1 in neonatal CPCs is unknown. In the developing heart, CPCs expressed Notch1 receptor and, in proximity of interstitial accumulation of the Notch1 ligand Jagged1, CPCs showed nuclear localization of the active fragment of Notch1, N1ICD. In all cases, the presence of N1ICD was coupled with Nkx2.5 expression suggesting that CPC commitment is regulated by Notch1 in the neonatal heart. To test this possibility, the Notch1 pathway was blocked by administration of a γ-secretase inhibitor to newborn mice for 3–7 days. Mice in which Notch1 was inhibited developed a dilated myopathy characterized by decreased muscle mass, wall thinning, increased chamber diameter, and decreased fractional shortening and ejection fraction. Treated mice showed a 50–75% reduction in the percentage of CPCs and myocytes showing N1ICD and Nkx2.5. Myocyte number decreased 40% while myocyte volume increased only 25%. A 62% reduction in replicating Ki67-positive amplifying myocytes was observed in treated mice while myocyte apoptosis did not differ in the two groups of animals. These cellular alterations were accompanied by downregulation of Nkx2.5 mRNA. To establish whether Notch1 is a transcriptional regulator of Nkx2.5, neonatal CPCs were infected with a lentiviral vector carrying N1ICD. By Q-RT-PCR, N1ICD-CPCs showed a marked increase in transcripts for Hes1, Nkx2.5 and MEF2C and a significant decrease in GATA4 mRNA. Notch1-dependent upregulation of Nkx2.5 was confirmed at the protein level. A consensus site for the target gene of Notch, RBP-Jk, was recognized in the Nkx2.5 promoter. The formation of a functional complex between Nkx2.5 and RBP-Jk was demonstrated by chromatin immunoprecipitation and reporter gene assay. In conclusion, cardiomyogenesis is a critical component of the maturation of the postnatal heart structurally and functionally and inhibition of myocyte generation through interference with the Notch1 pathway results in a decompensated dilated myopathy.