IF a recombinant MyoAAV vector encoding a nuclear-optimized MOTS-c transgene placed under the transcriptional control of a synthetic Antioxidant Response Element (ARE)/NRF2-responsive promoter (MyoAAV-ARE-MOTS-c), delivered at ≥5×10¹³ vg/kg via bilateral intramuscular injection into the gastrocnemius and tibialis anterior, is administered to aged (22–24 month), male C57BL/6J mice with established sarcopenia,

THEN the following measurable outcomes will be observed compared to vehicle-injected aged controls at 12 weeks post-injection:

• ≥20% improvement in grip strength (force normalized to body mass)
• ≥15% increase in hindlimb muscle mass (gastrocnemius wet weight)
• Restored AMPK phosphorylation (pAMPK/AMPK ratio, western blot) to levels approximating 4–6 month reference animals
• Improved mitochondrial oxygen consumption rate (OCR, Seahorse XF) in isolated myofibers
• Re-engagement of stress-responsive nuclear gene programs (NRF2 target genes: HMOX1, NQO1, GCLC; quantified by RT-qPCR) will be observed,

BECAUSE the following step-by-step causal chain connects the intervention to the outcome:

1. Aged skeletal muscle accumulates a chronic MOTS-c signaling deficit. Endogenous MOTS-c levels decline markedly with age due to progressive deterioration in mitochondrial transcriptional and translational fidelity, eroding the retrograde nuclear-mitochondrial communication axis. This accumulated deficit — not merely future decline — underpins the established bioenergetic and proteostatic dysfunction of sarcopenic muscle. (MOTS-c age-related decline and sarcopenia association)[Reynolds et al., Nature Communications, 2021, as cited in Evidence Set]

2. Aged sarcopenic muscle is chronically enriched in NRF2-activating stimuli. Mitochondrial ROS accumulation, lipid peroxidation byproducts (4-HNE), and electrophilic stress in aged muscle constitutively activate NRF2/KEAP1 signaling. An ARE-driven transgene in this environment will be preferentially and autonomously activated in already-damaged aged myofibers — not in healthy young tissue — ensuring that MOTS-c is produced precisely where and when retrograde signaling has collapsed. [SPECULATIVE: ARE promoter activity in aged vs. young muscle requires direct empirical comparison, though chronic oxidative stress in aged muscle is well-established in the Evidence Set]

3. The nuclear-encoded MOTS-c transgene bypasses the failing mitochondrial translation machinery. Because endogenous MOTS-c is encoded within the 12S rRNA locus of the mitochondrial genome, its production depends on intact mitochondrial ribosomes and transcriptional apparatus — both of which deteriorate with age. By re-encoding MOTS-c in the nuclear genome (codon-optimized for cytoplasmic ribosomes, with a signal sequence removed to allow cytoplasmic release and re-entry into the nucleus), the intervention repairs the signaling output of a dysfunctional organelle without requiring mtDNA repair. (Mitochondrial translational deteriora...

SENS category: RepleniSENS