Fig 2: Enrichment and optimization of in vitro mESC-derived CPC enrichment.A Representative microphotographs depicting stages of differentiation of murine ground state Tbx5Cre;R26ReYFP/eYFP mESC cultured under specified cardiomyocyte differentiation conditions, either as monolayers or as embryoid bodies. B Flow cytometric analysis using three surface markers Pdgfra, Kdr and Gfra2 on different days of cardiomyocyte differentiation indicates an CPC enrichment window between days 7 and 9. C The expression of Kdr is dynamic and defines two waves of CPC in the current differentiation protocol. N = 1–4. Error bars = SEM.

Fig 3: CPC surface analysis of YFP+ cells.A Real-time PCR analysis of Yfp and Tbx5 transcripts in the ventricles of the adult heart in different time-points. B Flow cytometry acquisition of YFP+ cells from different time-points following cardiac injury. N = 3–6 hearts. C, D Representative two- and three-dimensional graphs from flow cytometric analysis of Pdgfra+Kdrlow/+Gfra2+ adult, postnatal days 6 and 9 heart and their co-expression with YFP (Tbx5-tracing), Sca-1 and c-Kit. N = 9. Key-CP; cardiomyocyte precursor, CM; cardiomyocyte, EC; endothelial cell. E YFP+ cells were collected 7 days after chemical injury and cultured in CM differentiating conditions for 5 and 8 days. Only YFP+ from the injured heart were able to differentiate into CM-like cells. Measurement of YFP+/cTnT+ mononucleated and binucleated cells 5 and 8 days in vitro culture. N = 5–6 independent cultures. Mann–Whitney test, p = 0.0519 (ns = not statistically significant) for D5 binucleated/mononucleated ratio.