Mesenchymal stromal cells deliver HS-enhanced Nrf2 via extracellular vesicles to mediate mitochondrial homeostasis for repairing hypoxia-ischemia brain damage.

Mesenchymal stromal cells (MSCs) are considered a therapeutic approach for neurological diseases via extracellular vesicles (EVs). Modified EVs contain active components with enhanced therapeutic potential. In this study, we aimed to explore the role and underlying mechanism of EVs from MSCs preconditioned by NaHS (an Hydrogen sulfide donor) (HS-EVs) in hypoxia-ischemia (HI) brain damage. Our results showed that HS-EVs treatment via the non-invasive intranasal route in HI mice was able to reduce oxidative stress and mitochondrial dysfunction compared to EVs treatment. Mechanistic studies demonstrated that NaHS promoted nuclear factor erythroid-2 related factor 2 (Nrf2) expression in the cytoplasm by inducing Parkinson disease protein 7 (PARK7)-dependent disintegration of Nrf2/Keap-1 complex in MSCs. In particular, the free Nrf2 was loaded into the EVs as a result of its KFERQ motif being recognized by 70-kDa heat shock proteins and lysosomal-associated membrane protein 2A. Subsequently, HS-EVs were internalized into neurons in the ipsilateral hemisphere, thus delivering abundant Nrf2 to accumulate in the mitochondria and remodeling mitochondrial function following HS-EVs treatment in HI mice. Moreover, Nrf2 knockdown in MSCs remarkably impaired HS-EVs-mediated therapeutic effects on HI mice. In brief, the present study for the first time demonstrated that HS-modified MSCs significantly accumulated higher Nrf2 in EVs via upregulating PARK7 expression, revealing the mechanism through which antioxidant protein Nrf2 delivered by HS-EVs protect against mitochondrial dysfunction in HI brain damage.