Fig. 4

Bmp7 loss- and gain-of-function respectively reduces and increases cardiomyocyte proliferation during zebrafish heart regeneration (A) Experimental design for the analysis of cardiomyocyte cell cycling in bmp7a homozygous mutant fish at 7 days post cryoinjury (dpi). (B) Immunofluorescence analysis of EdU incorporation and MHC (to identify cardiomyocytes) on cryoinjured hearts of bmp7a mutant fish and wild-type siblings. n (wild-type) = 7 hearts, three sections per heart, 1158 cardiomyocytes, n (bmp7a−/−) = 6 hearts, three sections per heart, 472 cardiomyocytes. Representative pictures are provided; arrows point at cycling cardiomyocytes; scale bars, 100 μm. (C) Experimental design depicting the use of a transgenic line allowing for heat-shock-inducible overexpression of bmp7b together with constitutive expression of GFP in cardiomyocytes. (D) Immunofluorescence analysis of PCNA (proliferating cell nuclear antigen) and myosin light chain 7 (MYL7, to identify cardiomyocytes) in heat-shocked hsp70L:bmp7b, myl7:eGFPaf5Tg transgenic and wild-type siblings at 7 dpi. Note that GFP fluorescence is not shown. n (wild-type) = 12 hearts, two sections per heart, 421 cardiomyocytes. n (hsp70L:bmp7b, myl7:eGFP) = 9 hearts, two sections per heart, 472 cardiomyocytes. EdU- and PCNA-positive cardiomyocytes in (B) and (D), respectively, were counted manually within the border zone of control and transgenic fishes. Each dot represents a different heart (biological replicate), which is the average of the analysis of two to three sections. Representative pictures are provided; arrows point at cycling cardiomyocytes; scale bars, 50 μm. The values in (B) and (D) are presented as mean, error bars show SEM, statistical significance was determined using two-sided Student’s t test.