Fig. 1: Spatiotemporally dynamic endocardial ciliation during cushion EndoMT is blood flow dependent.
From: Endocardial primary cilia and blood flow regulate EndoMT during endocardial cushion development
a, Immunofluorescence on mouse embryo sections for cilia (ARL13B, green) on endocardial cells (CD31, white) of the AVC during cushion development (e8.5, e9.5 and e10). Nuclei are shown in blue (Hoechst). Close-ups with and without CD31 are provided; white arrows indicate cilia; luminal endocardial cells are outlined in white, and the entire AVC (including the cushion mesenchyme) is outlined in red. b, Overview of e9.5 AVC with a close-up of AVC endocardial cells (CD31, white) with cilia (ARL13B, green) costained with a centriole marker (γ-tubulin, red). A version without CD31 is also provided (right panel). c, Ciliated endocardial cells as a percentage of all endocardial cells versus distance in the AVC at e8.5 (top, n = 3), e9.5 (middle, n = 4) and e10 (bottom, n = 3). Distance runs from left to right: atrium to left ventricle. d, Endocardial ciliation over time as a percentage of all endocardial cells in either the AVC or OFT (e8.5 (AVC n = 10, OFT n = 10), e9.0 (AVC n = 7, OFT n = 7), e9.5 (AVC n = 6, OFT n = 8), e10 (AVC n = 3, OFT n = 4)) (OFT P = 0.0009, AVC P = 0.0001). e, Immunofluorescence on e9.5 WT and Ncx1−/− mouse heart sections for cilia (ARL13B, green) on endocardial cells (CD31, white). Nuclei are shown in blue (Hoechst). A version without CD31 is provided; white arrows indicate cilia, while the endocardium is outlined in white. f, Endocardial ciliation in the AVC over time as a percentage of all endocardial cells in WT mice (blue) and Ncx1−/− mice (red) (e8.5 (WT n = 8, Ncx1−/− n = 8), e9.0 (WT n = 4, Ncx1−/− n = 4), e9.5 (WT n = 7, Ncx1−/− n = 7)) (e8.5 P = 0.2716, e9.0 P < 0.0001, e9.5 P = 0.0016). g, Ciliated endocardial cells versus distance in the AVC of Ncx1−/− mice at e9.5 (n = 4). For comparison, the WT line is provided in blue (n = 4). Statistics: no significance (NS), P > 0.05; **P ≤ 0.01; ***P ≤ 0.001; ****P ≤ 0.0001. The statistical test used was a two-sided t test with Welch’s correction. Data are represented as mean ± s.e.m. Unless otherwise stated, n is the number of embryos. A, atrium; ECs, endocardial cells; LV, left ventricle.
