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. 2022 Sep 27;119(3):813–825. doi: 10.1093/cvr/cvac159

Figure 5.

Figure 5

Nanoparticle-mediated inhibition of TGFBR2 attenuates EndoMT and progression of PAH in vivo. (A) Representative images showing the morphology of control, BMPR1A, TGFBR2, or BMPR1A/TGFBR2 siRNA-treated PAECs. Morphological alteration in BMPR1A siRNA-treated PAECs was restored by concomitant inhibition of TGFBR2 (scale bar = 50 μm). (B) Quantification of elongation index in control, BMPR1A, TGFBR2, or BMPR1A/TGFBR2 siRNA-treated PAECs [**P ≤ 0.005 (unpaired t-test), n = 11]. (C) Western blots showing the expression of EndoMT markers (αSMA, SM22α, and TGFBR2) and BMPR1A in control, BMPR1A, TGFBR2, or BMPR1A/TGFBR2 siRNA-treated PAECs. Inhibiting TGFBR2 effectively suppressed the ectopic expression of EndoMT markers induced by BMPR1A siRNA treatment. (D) Quantification of αSMA (left) and SM22α (right) protein expression in control, BMPR1A, TGFBR2, or BMPR1A/TGFBR2 siRNA-treated PAECs [**P ≤ 0.005 (unpaired t-test), n = 3]. (E) Schematic illustration on the treatment regimen for 7C1 nanoparticles coated with either control or TGFBR2 siRNA. 7C1 nanoparticles were injected into the tail vein of Bmpr1aiECKO mice after tamoxifen administration. Three rounds of 7C1 nanoparticle injections were performed with a 1-week interval. (F) Western blot with whole lung lysate showed that treatment with TGFBR2 siRNA-coated 7C1 nanoparticle attenuated the ectopic expression of TGFBR2, pSMAD2, and pSMAD3 in Bmpr1aiECKO mice. (G) Quantification of TGFBR2, pSMAD2, and pSMAD3 protein level in control or TGFBR2 siRNA-coated 7C1 nanoparticle-injected Bmpr1aiECKO mice [**P ≤ 0.005 (unpaired t-test), n = 3]. (H) Immunostaining showed αSMA expression was significantly reduced in the lung of TGFBR2 siRNA-coated 7C1 nanoparticle-injected Bmpr1aiECKO mice (right) compared with control (left) (scale bar = 2 mm). (I) Quantification of αSMA-positive EC area in the lung of control or TGFBR2 siRNA-coated 7C1 nanoparticle-injected Bmpr1aiECKO mice [**P ≤ 0.005 (unpaired t-test), n = 14]. (J) Elevated right ventricle systolic pressure was restored to normal range in TGFBR2 siRNA-coated 7C1 nanoparticles-injected Bmpr1aiECKO mice. (K) Quantification of the right ventricle systolic pressure in control or TGFBR2 siRNA-coated 7C1 nanoparticles-injected Bmpr1aiECKO mice [**P ≤ 0.005 (unpaired t-test), n = 7]. (L) Quantification of the right ventricle heart wall thickness in control or TGFBR2 siRNA-coated 7C1 nanoparticles-injected Bmpr1aiECKO mice [**P ≤ 0.005 (unpaired t-test), n = 8].