Transcriptomic analysis of skeletal muscle regeneration across mouse lifespan identifies altered stem cell states

Skeletal muscle regeneration relies on the orchestrated interaction of
myogenic and non-myogenic cells with spatial and temporal coordination.
The regenerative capacity of skeletal muscle declines with aging due to
alterations in myogenic stem/progenitor cell states and functions,
non-myogenic cell contributions, and systemic changes, all of which accrue
with age. A holistic network-level view of the cell-intrinsic and
-extrinsic changes influencing muscle stem/progenitor cell contributions
to muscle regeneration across the lifespan remains poorly resolved. To
provide a comprehensive atlas of regenerative muscle cell states across
mouse lifespan, we collected a compendium of 273,923 single-cell
transcriptomes from hindlimb muscles of young, old, and geriatric (4-7,
20, and 26 months old, respectively) mice at six closely sampled
time-points following myotoxin injury. We identified 29 muscle-resident
cell types, eight of which exhibited accelerated or delayed dynamics in
their abundances between age groups, including T and NK cells and multiple
macrophage subtypes, suggesting that the age-related decline in muscle
repair may arise from temporal miscoordination of the inflammatory
response. We performed a pseudotime analysis of myogenic cells across the
regeneration timespan and found age-specific myogenic stem/progenitor cell
trajectories in old and geriatric muscles. Given the critical role that
cellular senescence plays in limiting cell contributions in aged tissues,
we built a series of tools to bioinformatically identify senescence in
these single-cell data and assess their ability to identify senescence
within key myogenic stages. By comparing single-cell senescence scores to
co-expression of hallmark senescence genes Cdkn2a and Cdkn1a, we found
that an experimentally derived gene list derived from a muscle foreign
body response (FBR) fibrosis model accurately (receiver-operator curve AUC
= 0.82-0.86) identified senescent-like myogenic cells across mouse ages,
injury time-points, and cell-cycle states, in a manner comparable to
curated gene-lists. Further, this scoring approach in both single-cell and
spatial transcriptomic datasets pinpointed transitory senescent-like
subsets within the myogenic stem/progenitor cell trajectory that are
associated with stalled MuSC self-renewal states across all ages of mice.
This new resource on mouse skeletal muscle aging provides a comprehensive
portrait of the changing cellular states and interactions underlying
skeletal muscle regeneration across the mouse lifespan.