Fig. 2 TGF-β1 signaling drives fibrosis in Neu1−/− muscle, which is rescued by AAV-mediated gene therapy. TGF-β1 signaling drives fibrosis in Neu1−/− muscle,

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Fig. 2 TGF-β1 signaling drives fibrosis in Neu1−/− muscle, which is rescued by AAV-mediated gene therapy. TGF-β1 signaling drives fibrosis in Neu1−/− muscle, which is rescued by AAV-mediated gene therapy. (A to C) Immunohistochemical (IHC) analysis of WT and Neu1−/− gastrocnemius (GA) muscle sections performed with anti–TGF-β1 (A), anti-SMAD2 (B), and anti-pSMAD2 (C) antibodies. Scale bars, 25 μm. (D) Immunoblot analysis of WT and Neu1−/− total GA muscle lysates probed with anti–TGF-β1, anti-SMAD2, and anti-pSMAD2 antibodies. (E and F) Transverse sections of the WT and Neu1−/− muscles were costained with anti-vimentin (green) and anti–collagen IV (red) (E) and anti-TCF4/TCF7 (red) and anti-αSMA (green) (F) antibodies. (G) Transverse sections of the WT and Neu1−/− muscles were costained with anti-TCF4 (green) and anti-TGF-β1 (red) antibodies. (H) Immunoblot analysis of WT and Neu1−/− (myo)fibroblast lysates probed with anti–TGF-β1, anti-SMAD2, and anti-pSMAD2 antibodies. (I) Immunoblots of WT and Neu1−/− (myo)fibroblasts were probed with and anti-vimentin, anti–collagen IV, anti-αSMA, and anti-TCF4 antibodies. (J) RT-qPCR of the indicated mRNAs in WT, Neu1−/−, and Neu1−/−-AAV muscles (n = 7). mRNA levels are normalized by 18S ribosomal RNA (rRNA) and relative to WT control. (K) Coronal gastrocnemius skeletal muscle sections of WT, Neu1−/−, and Neu1−/−-AAV (scAAV2/8-CMV-huNEU and scAAV2/8-CMV-huPPCA) treated mice stained with Masson’s trichrome, showing complete rescue of the muscle fibrosis. Scale bars, 200 μm. Values are expressed as means ± SD. Statistical analysis was performed using the Student t test; ****P < 0.0001. Diantha van de Vlekkert et al. Sci Adv 2019;5:eaav3270 Copyright © 2019 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).