Endoglin deficiency results in congestive heart failure in zebrafish. (A) Sanger sequencing chromatograms of gRNA-targeted region on endoglin exon 2 in wild-type (eng+/+), heterozygous (eng+/−) and homozygous (eng−/−) mutant zebrafish. Complementary gRNA sequence is indicated below chromatograms. (B) Imaging of 72 hpf sibling and eng−/− zebrafish blood vessel perfusion. Analysis performed in Tg(kdrl:GFP), Tg(gata1:mRFP) background to highlight blood vessels and erythrocytes, respectively. Images are representative of data from siblings (n=10) and eng−/− fish (n=8). Scale bars: 500 µm. (C) Kaplan–Meier representation of eng+/+, eng+/− and eng−/− fish survival. +/+ versus −/− and +/− versus −/−, ***P<0.0001; +/+ versus +/−, not significant (ns) [log-rank (Mantel–Cox) test]. +/+, n=45; +/−, n=104; −/−, n=56. Arrows point to symptomatic siblings later identified as eng+/−. (D) Influence of genotype over sex ratio in individuals aged 3 months and above. Total number of individuals analyzed is indicated above bars. (E) Representative morphology of 30 dpf sibling and eng−/− fish. Note the enlarged cardiac area and overall paleness in eng−/− fish. Scale bars: 1 mm. (F) H&E-stained heart histological sections reveal enlargement and structural alteration of the ventricle, hypochromic red blood cells and swollen surrounding tissue. a, atrium; ba, bulbus arteriosus; v, ventricle. Scale bars: 100 µm.
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