Abstract 17861: Identification of Cardiovascular Lineage Descendants at Single Cell Resolution

BACKGROUND: The transcriptional profile of cardiac cells derived from murine embryos and from mouse embryonic stem cells (mESCs) has primarily been studied as a cell population. However, the characterization of gene expression in these cells at a single cell level may demonstrate unique variations not appreciated as a pool. METHODS AND RESULTS: To establish a single cell quantitative PCR platform and perform side-by-side comparison between cardiac progenitors cells (CPCs) and cardiomyocytes (CMs) derived from our previously described Nkx2.5 enhancer-eGFP mESC and mouse embryos, we generated a reference map for single cardiovascular cells through quantifying lineage-defining genes for CPCs, CMs, smooth muscle cells (SMCs), endothelial cells (EDCs), fibroblasts, and mESCs using the Fluidigm microfluidic-enabled multiplex PCR assay. This panel was then applied against day 10.5 embryonic heart single cells to demonstrate its ability to identify chamber-specific CMs, endocardial cells, and fibroblasts. In addition, we compared the gene expression profiles of Nkx2.5 enhancer-eGFP embryo- and mESC-derived CPCs and CMs at different developmental stages and showed that single mESC-derived CM is transcriptionally similar to embryo-derived CM up to the neonatal stage. Furthermore, we show that time-lapse microscopy coupled with single cell expression assay can resolve the identity and the lineage relationship of progenies of single cultured CPC. With this approach, we found differential propensity for mESC-derived CPCs to become SMCs and CMs, whereas embryo-derived CPC to become CMs or EDCs. CONCLUSIONS: Our single cell expression profiling assays demonstrate the transcriptional similarity between mESC and embryo-derived CPC and CM up to the neonatal stage of development as well as differences in the propensity of CPCs to differentiate into cardiovascular cells. Single cell expression analysis appears to be a powerful tool to address the unique behavior of individual embryo- or mESC-derived cardiovascular cells.