Normal stress on surface of mesenchymal stem cells boosts extracellular vesicle secretion and regenerative bioactivity.

Scalable approaches for enhancing therapeutic small extracellular vesicles (sEVs) production can facilitate the transition of sEVs from bench to bedside and beyond. Here, we present a user-friendly method to manipulate the extracellular mechanical microenvironment of umbilical cord-derived mesenchymal stem cells (MSCs), a promising cell type for generating sEVs with therapeutic benefits, to boost sEV secretion and regenerative bioactivity. The bioreactor system, called the programmable controlled rotating platform (PRP), is designed to apply normal stress on cell surface through centrifugal rotation culture. Experimental analyses suggested that the PRP can promote a 4-fold sEV secretion increase without affecting cell viability and sEV size when compared to the traditional static culture condition. More importantly, PRP-induced MSC-sEVs can significantly promote epithelial cell migration in vitro and accelerate corneal wound healing in a murine model, with suppressed inflammatory responses in wound bed tissue. Further mechanistic investigations revealed that this process involves the activation of cellular transcriptional signals implicated in sEV biogenesis. Concurrently, sEV cargo undergoes remodeling to enrich regenerative and immunoregulatory functions. These findings demonstrate the efficacy of our established platform in advancing sEV production and improving clinical performance, providing a novel sEV-based mechanism for ocular treatments, including corneal epithelialization and even retinal neural regeneration.