(682d) Three-Dimensional Dynamic Culture Improves Extracellular Vesicle Production and Cargo Profile of Human Mesenchymal Stem Cells through Altered Biogenesis
In current study, dynamic three-dimensional (3D) aggregation is applied to hMSC to improve the EV production by biogenesis. hMSC were cultured as 3D aggregates under wave motion to facilitate aggregation and EV secretion. A modified isolation strategy also was utilized to enrich EV efficiently. Compared to monolayer culture, hMSC-EV quantity was improved significantly under 3D dynamic culture. Moreover, the derived hMSC-EV from 3D culture exhibited smaller sizes compared to monolayer culture, indicating higher enrichment of the exosome population among hMSC-EV. Analysis of EV biogenesis markers from 3D hMSC aggregates demonstrated increased activation of the endosomal sorting complexes required for transport (ESCRT) pathway, which may contribute to the altered biogenesis. For functional evaluation, EV from 3D hMSC aggregates enhanced the stress resistance of adult stem cells and rejuvenated aged stem cells expressing cellular senescence after EV application to those cultures. Moreover, 2D and 3D hMSC-EV were shown to modulate the immune response in different patterns determined by T lymphocyte and macrophage phenotype assays. MicroRNA sequencing of EV revealed in detail the cargo profile differences for EV derived from 3D dynamic culture versus monolayer culture, which may explain some of the functional differences. In summary, this study provides a promising strategy for high quality EV production from hMSC with enhanced therapeutic potentials compared to conventional monolayer culture.
This work is in part supported by NIH (R01 NS102395) and NSF (#1743426).