Abstract
BackgroundThe paracrine effects of multipotent mesenchymal stromal cells (MSCs) are mediated by their secretome composed by soluble factors (i.e., cytokines, growth factors, hormones) and extracellular vesicles (EVs). EVs promote intercellular communication, and the EV cargoes [e.g., proteins, soluble factors, microRNAs (miRNAs), messenger RNA (mRNA), DNA] reflect the molecular and functional characteristics of their parental cells. MSC-derived EVs (MSC-EVs) are currently evaluated as subcellular therapeutics. A key function of the MSC secretome is its ability to promote immune tolerance (i.e., immunopotency), a property that is enhanced by priming approaches (e.g., cytokines, hypoxia, chemicals) and inversely correlates with the age of the MSC donors. We evaluated mechanisms underlying MSC vesiculation and the effects of inflammation and aging on this process.MethodsWe evaluated the effects of interferon gamma (IFN-γ) and tumor necrosis factor alpha (TNF-α) on human adipose-derived MSC: (a) vesiculation (custom RT2 Profiler PCR Array), (b) EV profiles (Nanoparticle Tracking Analysis and Nanoparticle Flow Cytometry), (c) EV cargo (proteomic analysis and Western blot analysis), and (d) immunopotency (standard MSC:CD4 T cell proliferation inhibition assay). We confirmed the role of RAB27B on MSC vesiculation (RAB27B siRNA) and assessed its differential contribution to vesiculation in adult and pediatric MSCs (qPCR).ResultsCytokine priming upregulated RAB27B in adipose-derived MSCs increasing their secretion of exosome-like small EVs (sEVs; < 200 nm) containing two key mediators of immunopotency: A20 and TSG-6. These EVs inhibited T cell proliferation in a dose-dependent manner. RAB27B siRNA inhibited MSC vesiculation. Adipose-derived MSCs isolated from pediatric donors exhibited higher RAB27B expression and secreted more sEVs than adult MSCs.ConclusionsCytokine priming is a useful strategy to harvest anti-inflammatory MSC-sEVs for clinical applications. Of relevance, donor age should be considered in the selection of MSC-sEVs for clinical applications.
Highlights
The paracrine effects of multipotent mesenchymal stromal cells (MSCs) are mediated by their secretome composed by soluble factors and extracellular vesicles (EVs)
Cytokine priming enhances the immunopotency and increases exosome-like EV release by MSCs In order to quantitatively evaluate the effect of cytokine priming on the immunopotency of the MSC secretome, we designed an in vitro assay the sensitivity of which allowed us to test the immunosuppressive ability of MSC Multipotent mesenchymal stromal cell-conditioned media (CM) against T cell targets
Because the biological activity of MSC secretome is often attributed to the EV content, we assessed the effect of MSC cytokine priming on EV release by comparing the size distribution and concentration of nanoparticles in the MSC-conditioned media (MSC CM) using nanoparticle tracking analysis (NTA)
Summary
The paracrine effects of multipotent mesenchymal stromal cells (MSCs) are mediated by their secretome composed by soluble factors (i.e., cytokines, growth factors, hormones) and extracellular vesicles (EVs). A key function of the MSC secretome is its ability to promote immune tolerance (i.e., immunopotency), a property that is enhanced by priming approaches (e.g., cytokines, hypoxia, chemicals) and inversely correlates with the age of the MSC donors. MSCs possess functional properties (e.g., high expansion capacity, low immunogenicity, antiinflammatory/pro-angiogenic, and anti-fibrotic effects) that motivate their use in ongoing clinical trials of immune-mediated, inflammatory, and degenerative diseases [1, 2]. The functional improvements post-MSC infusion are in part mediated by paracrine mechanisms (e.g., cytokines, chemokines, and growth factors) which modulate the MSC microenvironment and influence the activity of resident cells [8]. On the other hand, stimulate MSCs to secrete pro-angiogenic factors (e.g., IGF-1, TGF-β1, VEGF, angiogenin, EGF, and bFGF) [18]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
