Abstract 12726: Differential miRNA and Target Gene Expression in Embryonic Stem Cells Lacking the Notch1 Receptor
Cardiomyocytes derived from embryonic stem (ES) cells could represent an attractive source of cells for replacement therapies in cardiac disease. However, cardiogenic differentiation of ES cells requires a complete understanding of the complex molecular mechanisms controlling the differentiation process. We showed previously that differentiation of ES cells into cardiomyocytes was favored by inactivation of the Notch1 receptor pathway. In the present study, we compared two ES cell lines, one with normal Notch1 expression (WT ES cells) and the other carrying two deleted Notch1 alleles (N1del ES cells). Quantitative RT-PCR analysis of pluripotent (Oct4, nanog, sox2), mesodermal (brachyury, mesp1) and cardiac gene (GATA4, Nkx2.5, α-MHC) expression during differentiation in embryoid bodies (EBs) demonstrated that N1del ES cells produce significantly higher numbers of mesodermal and cardiac precursors, and in turn of cardiomyocytes, than WT ES cells. We then determined the whole transcriptome of undifferentiated WT and N1del ES cells using microarray analysis. In parallel, we also identified differentially modulated miRNAs in each cell lines. In particular, we found twenty miRNAs, which were highly (up to 10 folds) up or downregulated in N1del ES cells as compared to WT ES cells. Moreover, among the modulated genes, a hundred predicted targets of the differentially expressed miRNAs were identified using computer assignment. We focused our analysis on pairs of miRNAs and target genes, which were inversely regulated in N1del ES cells. The differential expression of each pairs in undifferentiated cells was confirmed by quantitative methods. For instance, we showed that miR-127 was significantly downregulated and miR-17 significantly upregulated in differentiating N1del ES cells. Moreover, their putative targets, i.e. the surface marker CD24a and cyclin D2 were inversely and coordinately expressed in EBs. Together, these data suggest that Notch1 regulates specific miRNA subsets that could control the commitment of ES cells toward the cardiac lineage. These microRNAs could represent useful tools to increase the number of cardiac precursors during ES cell differentiation.
- © 2011 by American Heart Association, Inc.