Murine muscle stem cell response to perturbations of the neuromuscular junction are attenuated with aging.

Jacqueline Larouche ,Gregorio Valdez,Kaitlyn Sabin,Robert Louis Hastings,Sarah J Kurpiers,Wenxuan Liu,Kanishka De Silva,Benjamin Levi,Mahir Mohiuddin,Sethuramasundaram Pitchiaya,Paula Fraczek,Joe V Chakkalakal ,Lemuel A Brown ,James F Markworth ,Sofía D Merajver ,Young C Jang ,Peter Ulintz ,Jesus A Castor-Macias ,Jeongmoon J Choi ,Susan V Brooks ,Carlos A Aguilar

doi:10.7554/elife.66749

Abstract

During aging and neuromuscular diseases, there is a progressive loss of skeletal muscle volume and function impacting mobility and quality of life. Muscle loss is often associated with denervation and a loss of resident muscle stem cells (satellite cells or MuSCs); however, the relationship between MuSCs and innervation has not been established. Herein, we administered severe neuromuscular trauma to a transgenic murine model that permits MuSC lineage tracing. We show that a subset of MuSCs specifically engraft in a position proximal to the neuromuscular junction (NMJ), the synapse between myofibers and motor neurons, in healthy young adult muscles. In aging and in a mouse model of neuromuscular degeneration (Cu/Zn superoxide dismutase knockout - Sod1-/-), this localized engraftment behavior was reduced. Genetic rescue of motor neurons in Sod1-/- mice reestablished integrity of the NMJ in a manner akin to young muscle and partially restored MuSC ability to engraft into positions proximal to the NMJ. Using single cell RNA-sequencing of MuSCs isolated from aged muscle, we demonstrate that a subset of MuSCs are molecularly distinguishable from MuSCs responding to myofiber injury and share similarity to synaptic myonuclei. Collectively, these data reveal unique features of MuSCs that respond to synaptic perturbations caused by aging and other stressors.

Highlights

Skeletal muscle atrophy and weakness are primary features of physical frailty, and are common among the elderly and patients afflicted with neuromuscular disorders[1]
Both young and aged WT FACS-Sorted-muscle stem cells (MuSCs) (FSMs) and Pax7CreER/+; Rosa26mTmG/+ (P7mTmG) mononucleated cells from uninjured hindlimb muscles were subjected to droplet-based single-cell mRNA sequencing[24] and a total of 23,065 single MuSCs were generated (3,725 young FSMs, 6,603 aged FSMs, and 12,737 P7mTmG cells) yielding a mean of 6,150 unique molecular identifiers (UMIs) per cell after basic quality filtering
To further assess the purity of the FSMs, scRNA-Seq profiles of cells isolated from limb muscles across lifespan (3 mos - Tabula Muris[26] and 24 mos - Tabula Muris Senis) were integrated and compared (Fig. 1c; 2,301 Tabula Muris and 11,895 Tabula Muris Senis cells, mean 4,418 UMIs per cell)

Summary

Introduction

Skeletal muscle atrophy and weakness are primary features of physical frailty, and are common among the elderly and patients afflicted with neuromuscular disorders[1]. The decline in the health and repair of skeletal muscle can be partially attributed to decreases in number and function of a population of resident stem cells called satellite cells[2] or muscle stem cells (MuSCs). MuSCs respond to muscle damage via molecular changes[3] that facilitate activation followed by differentiation. MuSCs are lost, undergo exhaustion[4] and inefficiently repair tissue after injury, further contributing to degeneration and sarcopenia[5,6]. The reduced capacity for MuSCs to function in the context of age-associated impairments[7] due to intrinsic molecular changes remains unclear, owing to several factors such as heterogeneity[8,9] of the MuSC pool. Single-cell expression profiling[12] offers a powerful means to explore these actions as well as providing insights into how ensembles of cells act individually and/or together before and after injury[13] to maintain healthy muscle into old age

Methods

Results

Conclusion