Abstract Changes in the aged brain that occur with stroke, head injury or Alzheimer’s Disease Related Dementias (ADRD) result in chronic cognitive and motor deficits. Mesenchymal stem cells (MSCs) have recently received attention to reverse or slow neurodegenerative and injury-related changes in the aging brain as they suppress inflammation and facilitate tissue repair and remyelination. We have now completed studies as part of R21- NS102991 and R56-NS112207 that have shown that MSC-derived extracellular vesicles (EVs), the active product of MSCs, accelerate and enhance recovery of motor function following cortical injury in aged female monkeys. EV-treated monkeys exhibited full recovery by 3-5 weeks post-injury and untreated monkeys reached a plateau in recovery by 8-12 weeks post-injury. At a chronic recovery stage (16-weeks post-injury, R21- NS102991), MSC-EVs reduced injury-induced microglial neuroinflammation, neuronal excitotoxicity, synapse loss, oligodendrocyte damage and myelination deficits. However, the precise nature of how MSC-EVs act within both acute or chronic stages of recovery after injury remains unknown and is important to decipher in order to fine tune the efficacious use of MSC-EVs for age-related injury and neurodegenerative diseases. Our preliminary analysis of injury- and MSC-EV-associated changes in blood and brain tissue collected at an earlier stage of recovery (6 weeks post-injury, R56-NS112207) showed early treatment-related differences in microglial phenotypes and neuronal excitability that are distinct from treatment-related differences observed at 16-week post-injury. Therefore, we now propose to continue and expand these studies to combine in vivo behavioral, MRI and proteomic CSF and blood biomarker analyses with comprehensive proteomic and single-cell transcriptomic profiling and physiological and histological assessments of brain tissue harvested at distinct time points during recovery (3, 6, 9 weeks post- injury) from male and females aged monkeys. Furthermore, using advance human- derived in-vitro models, we will elucidate specific encapsulated miRNAs or/and proteins of MSC-EVs that ameliorate injury-related inflammatory and oxidative responses, and facilitate recovery and neuroprotection. We hypothesize that MSC-EVs contain miRNA and protein signals that modulate the acute response to injury and mitigate immediate damage, leading to reduced secondary chronic inflammation and degeneration, promoting a restorative microenvironment that will facilitate neural plasticity later in recovery. These studies will elucidate the temporal progression, sex-dependent dynamics and cell-specific molecular mechanism(s) of action of MSC-EV mediated recovery, that will pave the way for its potential development as a therapeutic for age-related injury and neurodegenerative diseases in humans.