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. 2021 May 6;12(3):746–768. doi: 10.1002/jcsm.12700

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© 2021 The Authors. Journal of Cachexia, Sarcopenia and Muscle published by John Wiley & Sons Ltd on behalf of Society on Sarcopenia, Cachexia and Wasting Disorders

This is an open access article under the terms of the http://creativecommons.org/licenses/by-nc/4.0/ License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.

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Dysregulation of lipid and iron metabolism activates ferroptosis in injured transverse abdominal (TA) muscle of Tfr1 ^SC/KO mice. (A) Gene Set Enrichment Analysis (GSEA) analysis of adipogenesis pathway between Tfr1 ^SC/WT and Tfr1 ^SC/KO mice upon cardiotoxin (CTX)‐induced regeneration at 30 dpi. (B) qPCR analysis of Adipoq, Fasn, and Cd36 mRNA expression in TA muscle from adult Tfr1 ^SC/WT and Tfr1 ^SC/KO mice before or after CTX‐induced injury (n = 5 per group). (C) Representative images of TA muscle sections from Tfr1 ^SC/WT and Tfr1 ^SC/KO mice upon CTX‐induced injury at 30 dpi (n = 5 per group). Oil Red O (ORO) staining and perilipin (green) and laminin B2 (pink) immunofluorescent staining revealed adipogenesis and lipid accumulation in TA muscle of Tfr1 ^SC/KO mice. (D) qPCR analysis of Pgc1α, Cox7a1, and Cox8b (mitochondrial genes), Tfr1, Slc11a2, Slc40a1, Ftl, and Fth1 (iron homeostasis‐related genes) mRNA expression in TA muscle from adult Tfr1 ^SC/WT and Tfr1 ^SC/KO mice before or after CTX‐induced injury (n = 5 per group). (E) Representative images of TA muscle section with Prussian blue staining (n = 5 per group) and transmission electron microscope images for TA muscle section from adult Tfr1 ^SC/WT and Tfr1 ^SC/KO mice after CTX‐induced injury at 30 dpi. (F) Heatmap of ferroptosis‐related gene expression in TA muscle from adult Tfr1 ^SC/WT and Tfr1 ^SC/KO mice after CTX‐induced injury at 30 dpi (n = 5 per group). (G) KEGG pathway enrichment analysis of up‐regulated genes in TA muscle from adult Tfr1 ^SC/WT and Tfr1 ^SC/KO mice after CTX‐induced injury at 30 dpi. (H) qPCR analysis of Gpx4 and Ptgs2 expression in TA muscle of adult Tfr1 ^SC/WT and Tfr1 ^SC/KO mice before or after CTX‐induced injury (n = 5 per group). (I) Representative western blot images of TA muscle between Tfr1 ^SC/WT and Tfr1 ^SC/KO mice after CTX‐induced injury at 30 dpi. (J) Heatmap of unsaturated fatty acid biosynthesis‐related gene expression in TA muscle from adult Tfr1 ^SC/WT and Tfr1 ^SC/KO mice after CTX‐induced injury at 30 dpi (n = 5 per group). (K) GSEA analysis of unsaturated fatty acid biosynthesis pathway between Tfr1 ^SC/WT and Tfr1 ^SC/KO mice upon CTX‐induced regeneration at 30 dpi. (L) qPCR analysis of Fasn, Elvol5, Elvol6, Scd1, Fads1, and Fads2 expression in TA muscle of adult Tfr1 ^SC/WT and Tfr1 ^SC/KO mice after CTX‐induced injury at 30 dpi (n = 5 per group). (M) Saturated fatty acid (SFA), monounsaturated fatty acid (MUFA), and polyunsaturated fatty acid (PUFA) levels (nmol/g) in TA muscle of adult Tfr1 ^SC/WT and Tfr1 ^SC/KO mice after CTX‐induced injury at 30 dpi (n = 4 per group). N.S., not significant, *P < 0.05, **P < 0.01, and ***P < 0.005, by two‐sided Student's t‐test. Data represent the mean ± standard error of the mean.