Fig. 3

Nanosafety assessment of CeONZs in hESCs and hESC-CMs. A and B Relative cell viability of H9 hESCs (A) and H9-CMs (B) after 24 h exposure to varying concentrations of CeONZs (0 to 120 μM) and 0.5 mM AA. C Comet assay of H9 hESCs and H9-CMs treated with different concentrations of CeONZs for 24 h, positive control means cells treated with 200 μM H₂O₂ for 2 h (Scale bar = 100 μm). Arrows indicate damaged cells. D Heatmap of pluripotency and lineage markers expression in H9 hESCs treated with CeONZs for 24 h (Passage 0, P0) and passaged 2 times after CeONZs withdrawal (Passage 2, P2). Relative to GAPDH expression (n = 3). E Immunofluorescence staining of pluripotency markers OCT4 and SOX2 in CeONZs treated H9 hESCs at P0 and P2 (Scale bar = 100 μm). F SSEA-4⁺ cells in CeONZs treated H9 hESCs at P0 and P2. G Immunofluorescence staining of cTNT in H9-CMs after 24 h exposure to CeONZs (Scale bar = 25 μm). H, I Contraction curve (H) and contraction amplitude (I) of H9-CMs after 1 day of CeONZs treatment (n ≥ 30). J–M Electrophysiological characteristics of H9-CMs determined by field potential morphology (J), field potential duration (FPD) (K), action potential morphology (L), and action potential duration (APD) (M), H9-CMs were treated with low (0.375 μM), medium (6 μM), and high (120 μM) concentrations of CeONZs for 24 h before MEA measurement. Data are presented as means ± SEM (n ≥ 3) and statistical significance was determined by one-way ANOVA with a Tukey post-test. ns means no significance