The Small Molecule H89 Facilitates Mesenchymal Stem Cell‐derived Extracellular Vesicle Release and Optimizes Therapeutic Efficacy in Liver Regeneration

The role of human umbilical cord mesenchymal stem cell-derived extracellular vesicles (hUCMSC-EVs) in liver regeneration is promising, yet their clinical translation is hampered by insufficient production. Current strategies targeting their secretion are inefficient and lack a clear mechanistic understanding. We isolated and characterized hUCMSC-EVs pretreated with the H89 and other mTORC1 inhibitors. Our findings revealed that H89 effectively enhances the secretion of hUCMSC-EVs across diverse cell types, demonstrating universal efficacy. Importantly, H89 upregulates GABARAPL1 expression, a key negative regulator of the PKA/mTORC1 pathway, to inhibit mTORC1 activity and promote the formation of amphisomes and SNARE-mediated hUCMSC-EVs release. Furthermore, EVs derived from H89-pretreated hUCMSCs (H-EVs) exhibited altered cargo composition, significantly increased proliferative activity, and potentiated liver regeneration via the RELA/miR-29a axis, which regulates the homeostasis of hepatic stellate cells. Our results highlight that H89 enhances hUCMSC-EV secretion through mTORC1 inhibition, with the resulting benefits for liver regeneration mediated by the RELA/miR-29a network. These findings demonstrate the great promise of H89 in EV-based liver regeneration, offering a promising platform for clinical translation.