Extended Data Fig. 5: The hepatocyte-secreted factor FetuA provides genome protection for HSPCs.
From: Fetal hepatocytes protect the HSPC genome via fetuin-A
a, b, Representative fluorescence images and positive proportions of γ-H2AX induced by Eto in E12.5 FL LT-HSCs, ST-HSCs and MPPs cultured with or without hepatocytes (refer to Fig. 3a) (n = 3). Scale bar, 20 µm. c, d, Representative fluorescence images and positive proportions of γ-H2AX Eto-induced in E12.5 FL-HSPCs in cultures of E16.5 or E12.5 hepatocyte-conditioned medium (co-SHI) or basic medium (SHI) (SHI-control, n = 6, E12.5 Co-SHI, n = 3; E16.5 Co-SHI, n = 5). Scale bar, 20 µm. e, Western-blots analysis of FetuA in SHI and co-SHI media and in E12.5 and E16.5 FL tissues; Coomassie blue was used as a loading control (n = 3). f, Representative fluorescence images of FetuA in E12.5 FL, E16.5 FL and E19 FL (n = 3). Scale bar, 500 µm. g, Representative fluorescence images of FetuA and hepatocytes (E-cadherin+), haematopoietic (CD45/Ter119+), endothelial (Scal-1 or CD144+), mesenchymal (Nestin+) cells in E16.5 FL (n = 3). Scale bars, 10 and 5 µm. h, Representative fluorescence images of FetuA (green) and HSPCs (Lin–CD41–CD48–CD150+) in E16.5 FL (n = 3). Scale bars, 10 and 5 µm. i, j, Representative comet-tail DNA images and the proportions of Eto-induced tail DNA in HSPCs cultured with or without FetuA (n = 3). The medians (red dashed lines) and quartiles (black dashed lines) of each group are shown. Scale bar, 100 µm. k, Representative fluorescence images of FetuA in the E19 bone marrow (BM) of Fetua+/– and Fetua–/– mice. Scale bar, 500 µm. l, m, Representative fluorescence images and positive cell proportions of γ-H2AX (green) in Kit+ cells (red) in the E19 BM of Fetua+/– or Fetua–/– mice (n = 3). Scale bar, 10 µm. n, Representative fluorescence images of FetuA in the E12.5 FL of mice with or without intraplacental (IP) injection of FetuA. Scale bar, 300 µm. o, p, Representative fluorescence images and positive cell proportions of γ-H2AX (green) in Kit+ cells (red) in the E12.5 FL of mice with or without IP injection of FetuA followed by Eto-treatment in vivo (IP FetuA–, n = 4; IP FetuA+, n = 3). Scale bar, 10 µm. q, Representative fluorescence images of FetuA in the E16.5 FL of Alb-cre; ROSA26-LSL-DTA mice with or without IP injection of FetuA. Scale bar, 500 µm. r, s, Representative fluorescence images and positive cell proportions of γ-H2AX (green) in Kit+ cells (red) in the E16.5 FL of Alb-cre; ROSA26-LSL-DTA mice with or without IP injection of FetuA followed by Eto-treatment in vivo (IP FetuA–, n = 3; IP FetuA+, n = 4). Scale bar, 10 µm. The n represents independent experiments (a-j) and fetuses (k-s) from 3 independent experiments. The mean ± s.d. is shown (b, d, m, p, s). Statistical tests: unpaired two-sided Student’s t-test (b, d, j, m, p, s).
