Extended Data Fig. 6: Downregulation of endothelial G6PD and PGD in vivo and in vitro.
a-b, qRT-PCR expression analysis of G6PD and PGD in control, G6PDKD and PGDKD in HUVEC (n=4) (a) and HAEC (n=3) (b), respectively from three independent experiments were used. Data are shown as mean ± SEM. Statistics were done using one-way ANOVA followed by Tukey’s multiple comparison test. c, Western blot analysis of G6PD and PGD expression in control, G6PDKD and PGDKD HUVEC. VINCULIN (VCL) is used as loading control. d-e, Densitometric analysis of G6PD and PGD expression in HUVEC (d) and HAEC (e) relative to VINCULIN. In HUVEC (n=4 for Ab-G6PD and n=6 for Ab-PGD) and in HAEC (n=3) from three independent experiments. Data are shown as mean ± SEM. Statistics were done using one-way ANOVA followed by Tukey’s multiple comparison test. f, qRT-PCR expression analysis of LAMA4, FN1, COL4A1 and ELN in control, G6PDKD and PGDKD in HAEC. LAMa4, FN1, COL4A1 (n=3) and ELN (n=5) from three independent experiments were used. Data are shown as mean ± SEM. Statistics were done using one-way ANOVA followed by Tukey’s multiple comparison test. g, Confocal images of DA sections of control and Pgd𝝙EC mouse embryos at E11 that were immunosignal with anti-ELN (red) and anti-CD31 (green) antibodies. DAPI (blue) is used for nuclei staining. Scale bars, 50 μm. Images are representative of at least two independent experiments. h, Timeline for the analysis of postnatal mouse retina angiogenesis in Pgd KO mouse. Control and Pgdi𝝙EC mice were injected with tamoxifen at P2-P3-P4 and retinas analyzed at P6. i, qRT-PCR expression analysis of Pgd and Cdh5 in EC from mouse liver in control and Pgdi𝝙EC pups at P6. n=4 from three independent experiments. Data are shown as mean ± SEM. Statistics were done using unpaired Student’s t-test. j, Western Blot of PGD on MPEC isolated from control and Pgdi𝝙EC pups at P6. β-ACTIN is used as loading control. And CDH5 as EC marker. Images are representative of at least two independent experiments.
