Figure 4: Analysis of Representative Human CPC Clones

(A) Flow cytometric analysis of CD31, CD34, CD45, CD73, CD90, CD105, CD117, CXCR4, CD140a, CD146, and NG2 expression in 1 CPC clone. Histograms show control (gray) and specific fluorescent intensity signal (white). (B) Analysis of cardiac transcription factor expression by immunostaining. NKX2.5 (red); MEF2C (green). Nuclei were stained with DAPI (blue). (C) Expression of cardiac transcription factors (GATA4, MEF2C, NKX2.5), and sarcomeric proteins (MYH6, MYH7, MYH11) in proliferating adult CPCs (Expansion; black) and in adult CPCs exposed to DLL1 (gray), and differentiated in the absence (upper panels) or presence of DAPT (lower panels). Data represent mean ± SEM; *p < 0.05 as compared with proliferating CPCs; ‡p < 0.05 as compared with differentiating unstimulated (None; white) CPCs in absence of DAPT (n = 3). (D) Analysis of SM-MHC and α-ACTININ expression by immunostaining in CPCs exposed or not to DLL1 and differentiated in the absence or presence of DAPT. Nuclei were stained with DAPI (blue). Scale bars: 20 μm. (E) Analysis of α-ACTININ (green) and TNNI (magenta), or MLC2V (magenta) expression by immunostaining in CPCs-derived cardiomyocytes. Nuclei were stained with DAPI (blue). Scale bars: 20 μm. (F) Cytosolic Ca2+ were recorded in CPCs-derived cardiomyocytes. Cells were either evaluated under resting conditions, electrical stimulation (30 V, 2 ms at 5 Hz), or perfusion with caffeine. Spontaneous (top panel) and evoked (middle panel) Ca2+ oscillations were detected. Quantification shows the percentage of cells with spontaneous Ca2+ activity or responding to caffeine. Abbreviations as in Figure 1.