MicroRNA-29a in Adult Muscle Stem Cells Controls Skeletal Muscle Regeneration During Injury and Exercise Downstream of Fibroblast Growth Factor-2

Abstract

The expansion of myogenic progenitors (MPs) in the adult muscle stem cell niche is critical for the regeneration of skeletal muscle. Activation of quiescent MPs depends on the dismantling of the basement membrane and increased access to growth factors such as fibroblast growth factor-2 (FGF2). Here, we demonstrate using microRNA (miRNA) profiling in mouse and human myoblasts that the capacity of FGF2 to stimulate myoblast proliferation is mediated by miR-29a. FGF2 induces miR-29a expression and inhibition of miR-29a using pharmacological or genetic deletion decreases myoblast proliferation. Next generation RNA sequencing from miR-29a knockout myoblasts (Pax7 CE/1 ; miR-29a flox/flox ) identified members of the basement membrane as the most abundant miR-29a targets. Using gain-and loss-of-function experiments, we confirm that miR-29a coordinately regulates Fbn1, Lamc1, Nid2, Col4a1, Hspg2 and Sparc in myoblasts in vitro and in MPs in vivo. Induction of FGF2 and miR-29a and downregulation of its target genes precedes muscle regeneration during cardiotoxin (CTX)-induced muscle injury. Importantly, MP-specific tamoxifen-induced deletion of miR-29a in adult skeletal muscle decreased the proliferation and formation of newly formed myofibers during both CTX-induced muscle injury and after a single bout of eccentric exercise. Our results identify a novel miRNAbased checkpoint of the basement membrane in the adult muscle stem cell niche. Strategies targeting miR-29a might provide useful clinical approaches to maintain muscle mass in disease states such as ageing that involve aberrant FGF2 signaling. STEM CELLS 2016;34:768-780 SIGNIFICANCE STATEMENTSkeletal muscle mass and function is critical for the maintenance of health, and the decline of muscle mass during aging inversely correlates with mortality. Adult muscle stem cells provide an important target for strategies to maintain muscle mass, but the molecular mechanisms that control the activation of these stem cells in vivo are still incompletely understood. We demonstrate that microRNA-29a is a novel downstream target of FGF2, and that miR-29a mediates the dismantling of the basement membrane in the adult muscle stem cell niche that is required for the proliferation of myogenic progenitors during muscle regeneration. We propose the FGF2/miR-29a pathway as a novel target to prevent a decrease in muscle mass in disease states such as ageing where FGF2 signaling is overly activated.

MicroRNA deep sequencing in two adult stem cell populations identifies miR-501 as a novel regulator of myosin heavy chain during muscle regeneration

“…MiRNAs that reside on polycistronic transcripts are processed with different efficiencies despite being co-transcribed (Chakraborty et al, 2012; Chaulk et al, 2014), which could explain the higher abundance of miR-501 compared with miR-362. We have recently described a similar phenomenon for the miR-29a/b cluster in mouse and human muscle cells (Galimov et al, 2016). We currently lack sufficient evidence to conclude that other miRNAs located in the Clcn5-2 intron also share the miR-362/-501 cluster.…”

Uncoupling protein 1 expression in adipocytes derived from skeletal muscle fibro/adipogenic progenitors is under genetic and hormonal control

“…Moreover, a subpopulation of Myf5 + cells from extraocular muscle expresses Sca1 and is adipogenic 50. In our study, we relied on a prospective isolation strategy to obtain MPs from skeletal muscle by FACS based on the absence of CD31, CD45, and Sca1 and the presence of alpha 7‐integrin, which results in highly pure myogenic cultures 35. Analysing both polyclonal and monoclonal cell cultures, we could not identify any MP subpopulation with adipogenic potential.…”

“…After incubation with fluorescent antibodies, pure populations of MPs and FAPs cells, respectively, were obtained using fluorescence‐activated cell sorting (FACS, BD FACSAria III, BD Biosciences, San Diego, CA, USA). Fibro/adipogenic progenitors were sorted based on positive staining for Sca1 and absence of staining for alpha 7‐integrin, CD31, and CD45,30 while MPs were sorted based on positive staining for alpha 7‐integrin and absence of Sca1, CD31, and CD45 35. For analysis of monoclonal cell colonies, single FAPs and MPs were sorted directly into 96‐well cell culture plates containing growth medium.…”

MiR‐19b‐3p regulates osteogenic differentiation of PDGFRα+ muscle cells by specifically targeting PTEN

“…Later, it was also reported by Uezumi et al (2014) that mesenchymal progenitors were identified in the human skeletal muscle interstitium and could be isolated by using PDGFRα as a specific marker. A single miRNA may regulate a large number of target genes acting in a similar pathway, thus playing an important role in biological regulation (Galimov et al, 2016). Some researchers have reported that PDGFRα + muscle cells participate in HO formation (Wosczyna et al, 2012;Oishi et al, 2013;Agarwal et al, 2016Agarwal et al, , 2017, however, the mechanisms that direct these progenitor cells to initiate cartilage and bone formation are still unknown.…”