Effect of 2D and 3D Culture Microenvironments on Mesenchymal Stem Cell-Derived Extracellular Vesicles Potencies

• DOI: 10.3389/fcell.2022.819726
• Publication Date: 2022-02-14T04:46:09Z
• Authors: Gina D. Kusuma; Anqi Li; Dandan Zhu; Hannah McDonald; Ishmael M. Inocencio; Daniel C. Chambers; Kenneth Sinclair; Haoyun Fang; David W. Greening; Jessica E. Frith; Rebecca Lim
• Keywords: Myofibroblast; 3D cell culture; Versican; Exosome; Proteome

Therapeutic benefits of mesenchymal stem cells (MSCs) are now widely believed to come from their paracrine signalling, i.e. secreted factors such as cytokines, chemokines, and extracellular vesicles (EVs). Cell-free therapy using EVs is an active emerging field in regenerative medicine. Typical 2D cultures on tissue culture plastic far removed the physiological environment MSCs. The application 3D cell allows MSCs adapt cellular which, turn, influences signalling activity. In this study we evaluated impact secretion, cargo proteome composition, functional assessment immunomodulatory, anti-inflammatory anti-fibrotic properties. MSC-EVs expressed classical EV markers CD81, CD63, CD9 with particle diameter <100 nm. There were distinct changes immunomodulatory potencies where exhibited reduced indoleamine 2,3-dioxygenase (IDO) activity significantly macrophage phagocytosis. Administration following double dose bleomycin challenge aged mice showed a marked increase bodyweight loss group throughout days 7-28. Histopathological observations lung tissues increased collagen deposition, myofibroblast differentiation leukocytes infiltrations. Assessment mechanics did not improve function instead resistance damping. Proteome profiling MSC-EV composition revealed molecular enrichment (compared parental cells) differential between condition associated immune-based fibrosis/extracellular matrix/membrane organization function. This provides insight into variation protein dependent microenvironment cells, which could have implications therapeutic effect potency. Overall, work suggests that produced MSC enhance typical properties expected (immunomodulation, anti-fibrotic, anti-inflammatory). outcome highlights critical differences obtained different microenvironments, should be considered when scaling up for clinical manufacturing.