Estrogen promotes regeneration in zebrafish heart. (A and B) qRT-PCR showing the expression of estrogen receptor genes in the heart of female fish (A) and male zebrafish (B) with DMSO, tamoxifen or E2 treatment at 7 days after cryoinjury. n = 3. (C) PCNA immunofluorescence (red) in the heart of male Tg (cmlc2: eGFP) zebrafish exposed to 1 nM E2 for 7 days after CI. Scale bar: 100 μm. (D). Quantification of proliferating cardiomyocytes in panel C (mean ± s.d., n = 5). One-way ANOVA with LSD post hoc test, *P = 0.05, ***P < 0.001. (E) embCMHC immunofluorescence (red) in the heart of untreated females, females treated with 1 μM tamoxifen (Tam.), untreated males, and males treated with 1 nM E2 for 7 days after CI. Scale bar: 200μm. (F) Quantification of embCMHC staining in panel E (mean ± s.d., n = 4~5) in the injured area. One-way ANOVA with LSD post hoc test, *P = 0.05, **P = 0.01, ***P < 0.001. (G) Expression of embCMHC in the sample shown in E, as detected by Western blotting. (H) Vitellogenin (VTG) immunofluorescence (red) in the heart of untreated females, 7 dpc females, 7 dpc females treated with 1 μM Tamoxifen, untreated males, 7 dpc males, and 7 dpc males treated with 1nM E2. Scale bar: 100 μm. (I) Quantification of VTG staining (mean ± s.d., n = 4~5) in panel H. One-way ANOVA with LSD post hoc test, ***P < 0.001. (J) Expression of VTG in samples shown in panel H, as detected by Western blotting. Un, untreated, SO, sham-operation, and CI, cryoinjury.
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