|
|
| |
Fig. 3. : Loss of Rcsd1 phenocopies the loss of Wnt11a. A. Depletion of Rcsd1 results in a severe cardiac phenotype characterized by cardiac edema (lateral view, red arrowheads) and heart malformations (ventral view, dotted lines; isolated hearts) at indicated stages. Scale bars: lateral view: 1000 μm; ventral view: 250 μm; isolated hearts 100 μm. B. Quantitative presentation of the data shown in A at stage 43. Rcsd1 MO injection leads to a cardiac phenotype in a dose dependent manner (red columns) compared to control MO injected embryos (blue column). Co-injection of full-length Xenopus rcsd1 RNA rescues the cardiac phenotype upon Rcsd1 deficiency (black column). C. Bilateral injection of Rcsd1 MO leads to a reduced heart rate (beats per minute, bpm) at stage 43, which is rescued by co-injecting full-length Xenopus rcsd1 mRNA. D-G. Analysis of cardiac Troponin T stained Rcsd1 morphant hearts using OPT. Atrial width, ventricular width and a-v length were measured as indicated in E resulting in data provided in F. Measurement of the OFT was done as indicated in E. Data are given in G. Scale bar: 100 μm. H. Cardiac Troponin T (Tnnt2) staining and sectioning of Control and Rcsd1 depleted embryos at stage 43. Loss of Rcsd1 leads to malformed hearts with disturbed ventricular trabeculation (t, white arrowheads). Rcsd1 injection restores the Rcsd1 MO induced phenotype. Scale bars: 100 μm. a, atrium; n, number of independent experiments; N, number of analysed embryos; ng, nanogram; oft, outflow tract; v, ventricle. Error bars indicate standard error of the means (s.e.m.). *, p≤0.05; **, p≤0.01; ***, p≤0.001; ****, p≤0.0001; calculated by a non-parametric Mann-Whitney rank sum test. |
|
|
| |
Fig. 7. : Cardiac phenotype after loss of Rcsd1 and after rescue with murine Rcsd1 constructs. A. Cardiac phenotype after Rcsd1 depletion and after rescue with murine Rcsd1 constructs at stage 43. The three upper rows show embryos from lateral and ventral view and the respective isolated hearts. The three lower rows show analysed ventricular trabeculation by cardiac Troponin T (Tnnt2) staining. Rows top to bottom: ventral view of the embryos, close-up of the respective hearts and section through the hearts cardiac phenotype after loss of Rcsd1 was rescued by co-injection with the full-length murine Rcsd1 construct and Rcsd1δNLS, although ventricular trabeculation was only partially restored to normal physical appearances. Rcsd1δCPI however was not able to restore cardiac anatomy and physiology. a, atrium; oft, outflow tract; t, trabeculae; v, ventricle, NLS: nuclear localization signal. (Lateral view: red arrowhead highlights cardiac edema upon loss of Rscd1. White arrow heads highlight the cardiac region in embryos of different rescue experiments. Scale bars: upper part: lateral view: 1000 μm; ventral view and isolated hearts: 250 μm; lower part Tnnt2: ventral view: 1000 μm; hearts and sections: 100 μm. B. Quantitative presentation of data shown in A. C. Injection of Rcsd1 MO lead to a reduced heart rate (beats per minute, bpm), which is rescued by co-injecting murine full-length Rcsd1 or Rcsd1δNLS mRNA but not by Rcsd1δCPI mRNA. n, number of independent experiments; N, number of analysed embryos; ng, nanogram. Error bars indicate standard error of the means (s.e.m.). n.s., not significant; **, p≤0.01; ****, p≤0.0001; calculated by a non-parametric Mann-Whitney rank sum test. |
