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. Author manuscript; available in PMC: 2021 Jul 6.
Published in final edited form as: Sci Transl Med. 2020 May 13;12(543):eaaz5660. doi: 10.1126/scitranslmed.aaz5660

Fig. 3. ACTRIIA-Fc attenuates PH and vascular remodeling in MCT-exposed rats.

Fig. 3.

(A) Treatment timing of adult rats after vehicle or MCT (40 mg/ kg, sc) with ACTRIIA-Fc (15 mg/kg, ip, twice weekly), sildenafil [30 mg/kg, orally, twice daily], or vehicle (10 mM TBS) for 4 weeks starting 1 day after MCT. (B) Mean pulmonary artery pressure (mPAP) and (C) RV hypertrophy (RVH) in rats treated with ACTRIIA-Fc or sildenafil compared to vehicle (n = 5 to 9 per group). Data represent means ± SEM. *P < 0.05, **P < 0.01, #P < 0.0001, and not significant (ns; P > 0.05) as indicated, one-way ANOVA with Sidak’s post test for mPAP and Kruskal-Wallis test with Dunn’s post test for RVH. (D) Medial wall thickness index and (E) the percentage of fully muscularized vessels in rats treated with ACTRIIA-Fc or sildenafil compared to vehicle (diameter, 10 to 50 μm). Values are shown as means ± SEM. n = 3 to 6 per group (30 to 50 vessels counted per sample). *P < 0.05, **P < 0.01, †P < 0.001, and #P < 0.0001 as indicated, one-way and two-way ANOVA with Tukey’s and Dunnett’s test for multiple comparisons. (F) Immunofluorescence images of pulmonary arteriole medial hypertrophy in lung sections from rats with MCT-induced PH treated with vehicle, ACTRIIA-Fc, or sildenafil. vWF, von Willebrand factor; SMA, α-SMA; DAPI, 4′,6-diamidino-2-phenylindole (nuclear stain). (G) Treatment timing of rats with established PH, with ACTRIIA-Fc (1, 3, and 10 mg/kg, ip, twice weekly) or isotype control with vehicle (mIgG2a, 10 mg/kg, ip, twice weekly) starting on day 28 after MCT. (H) RV systolic pressure (RVSP) and (I) RVH in rats treated with control IgG or ACTRIIA-Fc measured among surviving animals at 42 days (n = 6 to 10 per group). Values are shown as means ± SEM. *P < 0.05 and **P < 0.01, Kruskal-Wallis test with Dunn’s post test for multiple comparisons. (J) Medial wall thickness and (K) percentage of muscularized vessels in rats with established PH treated with vehicle IgG or ACTRIIA-Fc compared to control. Values are shown as means ± SEM. n = 3 to 6 per group (30 to 50 vessels counted per sample). *P < 0.05, **P < 0.01, †P < 0.001, and #P < 0.0001 as indicated, two-way ANOVA with Dunnett’s test for multiple comparisons. (L) Immunofluorescence images of MCT-induced medial hypertrophy in pulmonary arterioles from rats with established PH treated with vehicle IgG or ACTRIIA-Fc. (M) Ki67-positive cells in MCT-PH lungs treated with vehicle IgG or ACTRIIA-Fc. Values are shown as means ± SEM. n = 3 to 6 per group [20 random high-powered fields (HPF) containing >30 to 50 vessels counted per sample]. (N) Western blot and (O) densitometric analysis of phosphorylation of SMAD2 in lung lysates from MCT-exposed rats treated with vehicle IgG or ACTRIIA-Fc in comparison to control rats. Data represent means ± SEM. n = 3 to 4. *P < 0.05 and **P < 0.01, one-way ANOVA with Tukey’s test for multiple comparisons. Scale bars, 50 μm.