FIGURE 6

FIGURE 6

Smad3 inhibition affects vimentin and N-cadherin expression during regeneration. (A–D) Whole-mount in situ hybridization showing the expression level change of vimentin during regeneration. Smad3 inhibition via SIS3 treatment reduced the upregulation of vimentin at 5 dpf/2 dpt. Dashed lines outline the hearts. (E–H″) Maximal projection images of Tg(vmhc:mCherry-NTR) hearts showing surface view of N-cadherin immunostaining (green) in the control or ablated hearts without or with SIS3 treatment at 5 dpf/2 dpt. (E′–H′) Enlargement of box area in panels (E–H). (E″–H″) Green channel only. Yellow dashed lines outline the ablated areas; open arrowheads point to N-cadherin-absent CMs adjacent to the ablated areas. (I–L″′) Optical section images of Tg(vmhc:mCherry-NTR) hearts showing the cross-section view of N-cadherin immunostaining (green) in the ventricle. (I″–L″) Enlargement of box area in panels (I–L). (I′–L′,I″′–L″′) Green channel only. Arrowheads point to regular punctate N-cadherin distribution; arrows point to irregularly dispersed N-cadherin distribution; white lines outline the outer layer without CM marker. (M) Quantification of average fluorescence intensity of N-cadherin immunostaining in the ablated hearts without or with SIS3 treatment at 5 dpf/2 dpt. N = 7 and 5, respectively. Mean ± s.e.m., Student’s t-test, two-tailed, **P < 0.01. (N) Quantification of the outer layer width of N-cadherin immunostaining in the ablated hearts without or with SIS3 treatment at 5 dpf/2 dpt. N = 7 and 5, respectively. Mean ± s.e.m., Student’s t-test, two-tailed, *P < 0.05. Scale bars, (A–D) 50 μm, (E–H) 20 μm, (E′–H″,I″–L″′) 5 μm, (I–L′) 10 μm. dpf, days post-fertilization, dpt, days post-treatment; atr., atrium; oft., out flow tract; vent., ventricle; CM, cardiomyocyte.