Figure 1

Loss of cpt1b function inhibits cardiomyocyte proliferation in zebrafish embryos. (A) A schematic representation of the design for generating the cpt1b knockout zebrafish. (B) A representative agarose gel electrophoresis image for the genotyping of cpt1b^+/+, cpt1b^+/−, and cpt1b^−/− zebrafish. (C) Genomic sequences of wild-type (WT) and cpt1b knockout zebrafish around cpt1b guide RNA target sites. The guide RNA target sequences are denoted by red letters, with the protospacer adjacent motif (PAM) sequences underscored, and the coding sequences are represented by capital letters. (D) Whole-mount in situ hybridization analysis of cpt1b expression in cpt1b^+/+ and cpt1^−/− zebrafish embryos at 4 dpf. The black triangle indicates muscle, the yellow triangle indicates liver, and the red triangle indicates heart. Scale bar: 500 μm. (E) Quantitative RT-PCR analysis of cpt1b mRNA expression in 4 dpf cpt1b^+/+, cpt1b^+/−, and cpt1b^−/− embryos. (F) Quantitative RT-PCR analysis of cpt1b mRNA expression in the hearts of adult cpt1b^+/+, cpt1b^+/−, and cpt1b^−/− zebrafish. ** p < 0.01, *** p < 0.001. (G) Representative maximum intensity projection confocal images of dissected hearts from 4 dpf Tg(pcna^mGFP;myl7:H2A-mCherry;cpt1b^+/+) and Tg(pcna^mGFP;myl7:H2A-mCherry;cpt1b^−/−) zebrafish. Enlarged single optical section images of the boxed regions are shown on the right. White triangles indicate proliferating cardiomyocytes. Scale bar: 50 µm. (H) Quantitative analysis of proliferating cardiomyocytes in G. n = 9–20. ** p < 0.01. (I) Quantification of ventricular areas of adult cpt1b^+/+ and cpt1b^−/− zebrafish. n = 9–10. *** p < 0.001.