Janus effect of glucocorticoids on differentiation of muscle fibro/adipogenic progenitors

Andrea Cerquone Perpetuini,Alessandro Palma,Gianni Cesareni,Roberta Stefanelli,Marisabella Santoriello,Giulio Giuliani,Steven Harper,Alessio Reggio,Luisa Castagnoli,Alberto Bresciani,Mauro Cerretani,Simone Vumbaca

doi:10.1038/s41598-020-62194-6

Abstract

Muscle resident fibro-adipogenic progenitors (FAPs), support muscle regeneration by releasing cytokines that stimulate the differentiation of myogenic stem cells. However, in non-physiological contexts (myopathies, atrophy, aging) FAPs cause fibrotic and fat infiltrations that impair muscle function. We set out to perform a fluorescence microscopy-based screening to identify compounds that perturb the differentiation trajectories of these multipotent stem cells. From a primary screen of 1,120 FDA/EMA approved drugs, we identified 34 compounds as potential inhibitors of adipogenic differentiation of FAPs isolated from the murine model (mdx) of Duchenne muscular dystrophy (DMD). The hit list from this screen was surprisingly enriched with compounds from the glucocorticoid (GCs) chemical class, drugs that are known to promote adipogenesis in vitro and in vivo. To shed light on these data, three GCs identified in our screening efforts were characterized by different approaches. We found that like dexamethasone, budesonide inhibits adipogenesis induced by insulin in sub-confluent FAPs. However, both drugs have a pro-adipogenic impact when the adipogenic mix contains factors that increase the concentration of cAMP. Gene expression analysis demonstrated that treatment with glucocorticoids induces the transcription of Gilz/Tsc22d3, an inhibitor of the adipogenic master regulator PPARγ, only in anti-adipogenic conditions. Additionally, alongside their anti-adipogenic effect, GCs are shown to promote terminal differentiation of satellite cells. Both the anti-adipogenic and pro-myogenic effects are mediated by the glucocorticoid receptor and are not observed in the presence of receptor inhibitors. Steroid administration currently represents the standard treatment for DMD patients, the rationale being based on their anti-inflammatory effects. The findings presented here offer new insights on additional glucocorticoid effects on muscle stem cells that may affect muscle homeostasis and physiology.

Highlights

Www.nature.com/scientificreports machinery, MyoD and BAF60C, a subunit of the SWI/SNF chromatin-remodeling complex promoting the switch from a fibro-adipogenic to a pro-myogenic phenotype[10,11,12]
We developed a fluorescent microscopy-based protocol for the screening of compounds that modulate adipogenic differentiation of fibro-adipogenic progenitors (FAPs) isolated from mdx mice, a model of Duchenne muscular dystrophy
By magnetic bead separation, from 45-day-old mdx mice as CD31-/CD45-/ITGA7-/SCA1 + cells and as shown in Fig. S1a,b, purified cells were strongly enriched for CD140a (PDGFRα) which is a known marker of FAPs6

Summary

Introduction

Www.nature.com/scientificreports machinery, MyoD and BAF60C, a subunit of the SWI/SNF chromatin-remodeling complex promoting the switch from a fibro-adipogenic to a pro-myogenic phenotype[10,11,12]. To alleviate the unfavorable consequences of fat infiltrations in myopathies it would be desirable to enrich our toolbox of drugs controlling FAP adipogenesis by alternative mechanisms. To this end we performed a screening looking for new modulators of the fibro-adipogenic differentiation of FAPs isolated from the mdx mice model of Duchenne muscular dystrophy. Steroids presently represent the standard pharmacological treatment for DMD patients[13]. Despite their moderate beneficial effect on disease progression, their etiological role is not well understood. We present results suggesting that FAPs are a glucocorticoid target and that the anti-adipogenic effect of this class of molecules may contribute to their beneficial impact in delaying DMD progression

Methods

Results

Conclusion