Engineering Extracellular Matrix-bound Nanovesicles Secreted by Three-dimensional Human Mesenchymal Stem Cells.

Extracellular matrix (ECM) in the human tissue contains vesicles, which were defined as matrix-bound nanovesicles (MBVs). MBVs serve as one of the functional components in ECM, recapitulating part of the regulatory roles and in vivo microenvironment. In this study, extracellular vesicles from culture supernatants (SuEVs) and MBVs were isolated from the conditioned medium or ECM respectively of 3D human mesenchymal stem cells (hMSCs). Nanoparticle tracking analysis showed MBVs were smaller than SuEVs (100-150 nm). Transmission electron microscopy captured the typical cup shape morphology for both SuEVs and MBVs. Western blot revealed that MBVs have low detection of some SuEV markers such as syntenin-1. miRNA analysis of MBVs showed that 3D microenvironment enhanced the expression of miRNAs such as miR-19a and miR-21. In vitro functional analysis showed that MBVs can facilitate human pluripotent stem cell-derived forebrain organoid recovery after starvation and promote high passage fibroblast proliferation. In macrophage polarization 2D MBVs tended to suppress the pro-inflammatory cytokine IL-12β, while 3D MBVs tended to enhance the anti-inflammatory cytokine IL-10. This study has the significance in advancing our understanding of the bio-interface of nanovesicles with human tissue and the design of cell-free therapy for treating neurological disorders such as ischemic stroke. This article is protected by copyright. All rights reserved.