Adipogenesis and β-oxidation are differentially regulated in muscle after glycerol or CTX injection.

<p>qPCR analysis of the mRNA levels of different adipogenic (A), or in fatty-acid oxidation (B) regulators. Data are expressed as mean ± s.e.m., n = 5–6/group. * p-value <0.05 <i>vs</i>. control, # p-value <0.05 in Glycerol <i>vs</i>. CTX at same time points. Acadm, acyl-CoA dehydrogenase medium; Acs/l, acyl-CoA synthesase short-/long-chain; Acss, Acetyl-coenzyme A synthetase; Acox, Acyl-coenzyme A oxidase, Palmitoyl; C/EBP: CCAAT/ Enhancer binding protein; Cpt, carnitine palmitoyltransferase; Hadh, hydroxyacyl-CoA dehydrogenase; PPAR, peroxisome proliferator activated receptor.</p

Genomic Profiling Reveals That Transient Adipogenic Activation Is a Hallmark of Mouse Models of Skeletal Muscle Regeneration

<div><p>The marbling of skeletal muscle by ectopic adipose tissue is a hallmark of many muscle diseases, including sarcopenia and muscular dystrophies, and generally associates with impaired muscle regeneration. Although the etiology and the molecular mechanisms of ectopic adipogenesis are poorly understood, fatty regeneration can be modeled in mice using glycerol-induced muscle damage. Using comprehensive molecular and histological profiling, we compared glycerol-induced fatty regeneration to the classical cardiotoxin (CTX)-induced regeneration model previously believed to lack an adipogenic response in muscle. Surprisingly, ectopic adipogenesis was detected in both models, but was stronger and more persistent in response to glycerol. Importantly, extensive differential transcriptomic profiling demonstrated that glycerol induces a stronger inflammatory response and promotes adipogenic regulatory networks while reducing fatty acid β-oxidation. Altogether, these results provide a comprehensive mapping of gene expression changes during the time course of two muscle regeneration models, and strongly suggest that adipogenic commitment is a hallmark of muscle regeneration, which can lead to ectopic adipocyte accumulation in response to specific physio-pathological challenges.</p></div

Gene set enrichment mapping of glycerol- <i>vs.</i> CTX-injected muscle.

<p>Gene set enrichment analysis was performed on glycerol-injected compared to CTX-injected muscles 3 and 7 days after injection, and clustered according to gene set ontology. The size of nodes is proportional to the number of genes contained in the gene set. Red nodes: gene sets upregulated in glycerol <i>vs</i>. CTX model, blue nodes: gene sest downregulated in glycerol <i>vs</i>. CTX model, green bar: link between two gene sets sharing regulated genes.</p

Glycerol and CTX induce similar kinetics of degeneration and regeneration.

<p>Control uninjured tibialis anterior muscle, and tibialis anterior muscles injected with either 25 µl of 50% (v/v) glycerol or 10 µM CTX were sectioned and stained with laminin and DAPI 3, 7, 14 or 21 days after injection (dpi) (A), or with hematoxylin-eosin at 21 dpi (B). Cryosections were performed at the mid-belly part of tibialis anterior. Scale bars, 100 μm. Yellow arrow: immune cell nuclei, white arrow: central nuclei, red circle: fat cell-like structure. (C) Quantitative analysis of total myofibers and of myofibers with at least one central nuclei from laminin/DAPI stained sections. (D) qPCR analysis of the mRNA levels of different markers of muscle regeneration. Data are expressed as mean ± s.e.m., n = 5–6/group. * p-value <0.05 <i>vs</i>. control. MYH, Myosin Heavy Chain.</p

Ectopic adipogenesis occurs in both glycerol- and CTX-induced muscle regeneration.

<p>(A) qPCR analysis of the mRNA level of the platelet-derived growth factor receptor alpha (PDGFRα). (B) Cryosections were performed at the mid-belly part of TA and subjected to H&E and perilipin staining at each time points after injection. Representative perilipin (green) /DAPI (blue) fluorescent stainings at 21 dpi are shown next to an H&E staining of the same region. Scale bars, 50 μm. (C), Quantitative analysis of perilipin expression assessed by counting and measuring the area of all perilipin expressing cells per section. Data are expressed as mean ± s.e.m., n = 5–6/group. * p-value <0.05 <i>vs</i>. control, # p-value <0.05 in Glycerol <i>vs</i>. CTX at same time points.</p

The inflammatory signature is stronger in response to glycerol than to CTX.

<p>qPCR analysis of the mRNA levels of various macrophage markers and cytokines. Data are expressed as mean ± s.e.m., n = 5–6/group. * p-value <0.05 <i>vs</i>. control, # p-value <0.05 in Glycerol <i>vs</i>. CTX at same time points. Emr1; EGF-like module containing mucin-like hormone receptor 1; TNFα, tumor necrosis factor alpha, IL, interleukin; TGF-β1, transforming growth factor beta 1.</p