Notch signalling regulates cardiomyocyte proliferation. (A) ET33-mi60a;myl7:mRFP ventricle, volume rendering and optical section of a region of the inner injury border. Endocardial cells surround injury-adjacent cardiomyocytes (arrows) and precede into the wound. (B) IHC showing endocardial cells (GFP+) in contact with myocardial protrusions (tropomyosin+; blue arrowheads). (C,D) IHC against BrdU and Mef2 (C). Heat shock regime is indicated at the top. Scatter plot (D) of percentage of BrdU+ wound-adjacent cardiomyocytes (Mef2+; arrowheads between dotted lines). Mean±s.d.; t-test,**P<0.01. (E,F) Mef2 IHC showing a higher density of wound-adjacent cardiomyocytes (between the dotted lines) in Tg(UAS:NICD) than in control hearts (E), as quantified in F. Mean±s.d.; t-test, *P<0.05. (G) ISH showing higher numbers of nkx2.5+ wound-adjacent cardiomyocytes (between dotted lines) in Tg(UAS:NICD) hearts. (H) RNA-seq data showing differential myocardial gene expression in RO-treated hearts at 3 dpci. (I) ISH showing mycb expression in injury-adjacent cardiomyocytes (arrowheads) in DMSO-treated but not in RO-treated hearts (asterisks). (J) qPCR levels of fosab in the injured heart. Mean±s.d.; t-test, *P<0.05. LOF, loss of function. (K) MF20 IHC and mylk3 ISH on heart sections. RO treatment decreased the numbers of mylk3− cardiomyocytes adjacent to the injury site (arrowheads). (L) Scatter plot showing the percentage of mylk3− wound-adjacent (50 µm) cardiomyocytes. Mean±s.d.; t-test, **P<0.01. (M) qPCR analysis of mylk3 and tcap levels in the injured heart. Mean±s.d.; t-test, ***P<0.001. GOF, gain of function. Dotted lines delineate the injury site (is). Boxed areas are magnified in insets. Scale bars: 100 µm, except 25 µm in magnified views.
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