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. 2016 Nov 29;7:13650. doi: 10.1038/ncomms13650

Figure 7. PI3K inhibition rescues established retinal AVMs.

Figure 7

(a) Schematic representation of the experimental strategy to assess Alk1 deletion or BMP9/10 blockade. Arrowhead indicates injection of 50 μg Tx or BMP9/10 blAB. (b) Quantification of AV shunt number 24 h after Tx or BMP9/10 blAB administration. n=8–10 Retinas per group. Error bars: s.e.m. **P<0.01 and ***P<0.001, Mann–Whitney U-test. (c–e) Ve-Cadherin and Erg1/2/3 staining of retinal flat mounts from Ctrl (c), Alk1iΔEC- (d) and BMP9/10 blAB- (e) treated P4 mice. Yellow arrowheads in d,e indicate AV shunts. Numbers of ERG1/2/3+ endothelial cell nuclei at branches of retinal veins and arteries are indicated in yellow. (f,g) Quantification of the number of nuclei at the connection of capillaries with veins (f) or arteries (g) in P4 and P5 retinal vessels. n=6 retinas per group. Error bars: s.e.m., **P<0.01 and ***P<0.001, Mann–Whitney U-test. (h) Schematic representation of the experimental strategy to assess the effects of PI3K inhibition on Alk1iΔEC vasculature 24 h post Tx injection. Arrowheads indicate the time course of Tx (50 μg) and PI3Ki injections. (i,j) IB4 staining of retinal flat mounts from Alk1iΔEC P5 mice treated with PBS (i) or PI3Ki 24 h post Tx injection (j). Yellow arrowheads in i,m indicate AV shunts. (k) Quantification of AV shunt number, vascular density and number of branchpoints. n=10–11 Retinas per group. Error bars: s.e.m. **P<0.01 and ***P<0.001, Mann–Whitney U-test. (l) Schematic representation of the experimental strategy to assess the effects of PI3K inhibition on the retinal vasculature 24 h post BMP9/10 blAB-treated mice. Arrowheads indicate the time course of BMP9/10 blAB and PI3Ki administration. (m,n) IB4 staining of retinal flat mounts from mice treated with BMP9/10 blAB and 24 h later with PBS (m) or PI3Ki (n). Yellow arrowheads indicate AV shunts. (o) Quantification of AV shunt number, vascular density and number of branch points. n=6 Retinas per group. Error bars: s.e.m., *P<0.05 and **P<0.01, Mann–Whitney U-test. Scale bars, 100 μm in c–e,i,j,m,n. a, artery in red; v, vein in blue.