Fig. 8: Inhibition of the upstream TIE2 receptor signaling limits Pik3caH1047R -driven VM growth.
From: Angiopoietin–TIE2 feedforward circuit promotes PIK3CA-driven venous malformations
a, Experimental scheme for therapeutic treatment of advanced Pik3ca-driven VM with the TIE2 inhibitor BAY-826 (50 mg per kg body weight by oral gavage) and/or rapamycin (rapa, 10 mg per kg body weight by intraperitoneal (i.p.) injection). b, Whole-mount immunofluorescence of ear skin from Pik3caH1047R;Vegfr1-CreERT2 mice with advanced VMs after a 2-week treatment period. c,d, Quantification of the treatment outcome. Bar plots show the increase in EMCN+ vessel area relative to Cre− littermate Ctrl mice, mean ± s.d. (n = 5 (vehicle), n = 5 (BAY-826), n = 4 (rapa), n = 7 (BAY-826 + rapa) mice; c); or increase in vessel diameter relative to Cre− littermate controls, mean ± s.d. (n = 5 (vehicle), n = 5 (BAY-826), n = 4 (rapa), n = 4 (BAY-826 + rapa) mice (d). e, Experimental scheme for the induction of VMs and inhibition of TIE2 signaling by intraperitoneal injection of AAV vectors encoding a ligand-neutralizing soluble TIE2 extracellular domain (TIE2-ECD). f–h, Whole-mount immunofluorescence of ear skin from Pik3caH1047R;Vegfr1-CreERT2 mice (f) and quantification of the treatment outcome (g and h). Bar plots show the increase in EMCN+ vessel area relative to Cre− littermate Ctrl mice, mean ± s.d. (n = 14 (untreated), n = 8 (TIE2-ECD), n = 4 (rapa), n = 7 (TIE2-ECD + rapa) mice; g); or increase in vessel diameter relative to Cre− littermate Ctrl mice, mean ± s.d. (n = 18 (Ctrl); Cre+ cohorts: n = 14 (untreated), n = 8 (TIE2-ECD), n = 4 (rapa), n = 7 (TIE2-ECD + rapa) mice (h). i, Scheme for assessing BAY-826 treatment response in Pik3caH1047R;R26-iChr2-Mosaic;Vegfr1-CreERT2 mice. j, Top, Intravital two-photon (2P) microscopy images of the dermal microvasculature stained using intravenous PECAM1 antibody injection, showing clonal lesions expressing EGFP or mCherry, at the start of treatment period. Below, Confocal images of the same lesions after a 2-week treatment period. Boxed regions are magnified on the right. k, Quantification of clonal expansion showing pretreatment and post-treatment nuclei counts (n = 42 lesions from six Ctrl mice and n = 24 lesions from three BAY-836-treated mice). In c, d, g, h and k, *P < 0.05, **P < 0.01, ****P < 0.0001, ordinary one-way ANOVA and Tukey’s multiple-comparison test (c, d, g and h) and paired two-tailed Student’s t-test (k). Scale bars, 100 μm (b and j, magnification) and 500 μm (b, f and j, overview).
