Mitochondrial dynamics maintain stem cell regenerative competence throughout adult life by regulating metabolism and mitophagy

We profiled by bulk RNA-seq the transcriptional changes upon deletion of Drp1 in murine satellite cells, freshly harvested from young and old homeostatic muscle and during regeneration, after acute cardiotoxin-induced myoinjury Overall design: Hindlimb skeletal muscles from tamoxifen-treated WT ( Pax7-CreER; ROSA26YFP) and conditional Drp1-KO (Pax7-CreER;ROSA26YFP;Drp1fl/fl) mice were collected, manually minced, and mechano-enzymatically dissociated with Liberase/Dispase for 1-1.5 hours at 37ºC. Tissue homogenates were filtered and red blood cells lysed. The final single-suspension cell pellets were stained and processed for FACS-sorting to isolate highly purified satellite cells based on YFP+ expression (QSC, quiescent satellite cells). Concurrently pre-injured (tibialis anterior, quadriceps, and gastrocnemius) muscles, were also harvested from equivalent groups of genotype- and age-matched animals at 1-day ('Activated' satellite cells, ASC) or 3-days ('Proliferating' satellite cells) post-cardiotoxin-injection (DPI). For these timepoint conditions, cells were FACS-sorted from the regenerating bulk homogenates based on YFP+ (1DPI) or CD34-YFP+ (3DPI) immunophenotypes. Ages for the Young and the Old animal groups were 3 and 24 months, respectively.