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Fig. 3 Ect2 gene inactivation lowers cardiomyocyte endowment and is lethal in mice.
Ect2 gene inactivation lowers cardiomyocyte endowment and is lethal in mice. (A to H) Ect2flox gene inactivation with αMHC-Cre in mice. (A) Immunostaining and quantification of binucleated cardiomyocytes at P1 (Ect2F/Wtn = 6, Ect2F/Fn = 6 hearts). (B) DNA content per nucleus (Ect2F/Wtn = 642 cardiomyocytes, Ect2F/Fn = 647 cardiomyocytes). (C) Cardiomyocyte endowment, quantified by counting of fixation-digested hearts (Ect2F/Wtn = 12, Ect2F/Fn = 5 hearts). (D) Immunostaining and quantification of hypertrophy (cardiomyocyte size). (E) Quantification of mono- and binucleated cardiomyocyte size. (D and E) (Ect2F/Wtn = 1138 cardiomyocytes, 1101 mono, 37 bi, from 6 hearts; Ect2F/Fn = 1015 cardiomyocytes, 892 mono, 123 bi, from 6 hearts) and (F) heart weight (Ect2F/Wtn = 14, Ect2F/Fn = 6 hearts). (G) Echocardiographic analysis of myocardial dysfunction at P0 (LV endocardium outlined in yellow, Ect2Wt/Wtn = 4, Ect2F/Fn = 3 mice; movies S5 and S6). (H) Pup survival (fig. S9B and movie S7). (I) Immunostaining and quantification of cardiomyocyte binucleation after Ect2 rescue (n = 3 cell isolations). (J) Immunostaining and quantification of cell cycle entry in cardiomyocytes after Ect2flox gene inactivation with αMHC-MerCreMer, tamoxifen P0, P1, and P2, followed by 3 days of culture in the presence of BrdU (Ect2Wt/Wtn = 3, Ect2F/Fn = 2 cell isolations). (K) Immunostaining and quantification of cell cycle progression to the M phase in vivo after αMHC-Cre inactivation of Ect2flox (P1, Ect2F/Wtn = 6, Ect2F/Fn = 6 hearts). Statistical significance tested with Student’s t test (A to D, F, G, and I to K), one-way ANOVA with Bonferroni’s multiple comparisons (E), and Fisher’s exact test (H). Scale bars, 20 μm (A and I), 30 μm (J and K), and 100 μm (D). Honghai Liu et al., Sci Transl Med 2019;11:eaaw6419 Copyright © 2019 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works
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