Fig. 3

HAND2 knockout impaired SHF-derived cardiomyocyte differentiation. A The percentage of NKX2.5-eGFP⁺ cells in WT and H2-KO cells at different time points of cardiomyocyte differentiation (n = 3). B, C. Expression of FHF (B) and SHF (C) markers in early cardiomyocyte differentiation of WT and H2-KO cells. Relative to GAPDH expression (n = 3). Unpaired t test. D, E Expression of NKX2.5 (D) and ventricular cardiomyocyte (VCM) markers (E) in WT and H2-KO-derived differentiation day 30 cardiomyocytes. Relative to GAPDH expression (n = 3). Unpaired t test. F Western blot analysis of MYL2 expression in WT and H2-KO-derived differentiation day 30 cardiomyocytes. GAPDH served as loading control. Corresponding uncropped full-length gels and blots are presented in Additional file 8: Fig. S9. G The percentage of MYL2⁺ cardiomyocytes in WT and H2-KO cells at differentiation day 30 (n ≥ 3). H Expression of atrial and OFT cardiomyocyte (ACM, OFT) markers in WT and H2-KO-derived differentiation day 30 cardiomyocytes. Relative to GAPDH expression (n = 3). Unpaired t test. I Immunofluorescence staining of NR2F2 in WT and H2-KO-derived differentiation day 30 cardiomyocytes. Scale bar = 100 μm. J The field potential and simulated action potential recorded by MEA in WT and H2-KO-derived differentiation day 30 cardiomyocytes. Black lines represented the field potential while red lines represented the simulated action potential. K Comparison of corrected APD90 and APD50 in WT and H2-KO-derived differentiation day 30 cardiomyocytes (n ≥ 8). Unpaired t test. L The action potential paced by 1Hz recorded by whole-cell patch clamp in WT and H2-KO-derived day 30 cardiomyocytes. M Comparison of APD90 and APD50 in WT and H2-KO-derived differentiation day 30 cardiomyocytes (n ≥ 11). Unpaired t test. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. n.s: non-significant