Abstract
The vascular wall is a source of progenitor cells that are able to induce skeletal repair, primarily by paracrine mechanisms. Here, the paracrine role of extracellular vesicles (EVs) in bone healing was investigated. First, purified human perivascular stem cells (PSCs) were observed to induce mitogenic, pro-migratory, and pro-osteogenic effects on osteoprogenitor cells while in non-contact co-culture via elaboration of EVs. PSC-derived EVs shared mitogenic, pro-migratory, and pro-osteogenic properties of their parent cell. PSC-EV effects were dependent on surface-associated tetraspanins, as demonstrated by EV trypsinization, or neutralizing antibodies for CD9 or CD81. Moreover, shRNA knockdown in recipient cells demonstrated requirement for the CD9/CD81 binding partners IGSF8 and PTGFRN for EV bioactivity. Finally, PSC-EVs stimulated bone repair, and did so via stimulation of skeletal cell proliferation, migration, and osteodifferentiation. In sum, PSC-EVs mediate the same tissue repair effects of perivascular stem cells, and represent an 'off-the-shelf' alternative for bone tissue regeneration.
Highlights
In order to better understand the paracrine effects of perivascular stem cells (PSCs) in bone repair, we first defined a set of replicable in vitro paracrine effects of human PSCs on recipient human Bone marrow mesenchymal stem cell (BMSC) (Figure 1)
Cell surface antigens of human PSCs and BMSCs were typified by flow cytometry, and the multilineage differentiation potential of BMSCs was confirmed (Figure 1—figure supplement 2 and Figure 1—figure supplement 3)
Co-culture results showed that PSCs induced significant mitogenic (Figure 1A, MTS assay), promigratory (Figure 1B, scratch wound healing assay), and pro-osteogenic effects (Figure 1C, alkaline phosphatase activity) on BMSCs when placed in non-contact co-culture conditions
Summary
Stromal progenitor cells within vessel walls have multipotent properties (Cathery et al, 2018; Corselli et al, 2012; Covas et al, 2008; Crisan et al, 2008; Dellavalle et al, 2007; FarringtonRock et al, 2004), are native forerunners of mesenchymal stem cells (MSCs), and participate in endogenous bone repair (Diaz-Flores et al, 1991; Diaz-Flores et al, 1992; Grcevic et al, 2012). When purified based on expression of CD146 (Mel-CAM) and CD34, human perivascular stem cells (PSCs) from adipose tissue or other tissue compartments speed bone repair (Askarinam et al, 2013; Chung et al, 2014; James et al, 2017a; James et al, 2012a; James et al, 2012b; Tawonsawatruk et al, 2016). Direct incorporation of human PSCs into chondroblasts, osteoblasts and osteocytes occurs (James et al, 2012b), PSCs induce bone healing either primarily or exclusively via paracrine stimulation of the resident host cells within the defect niche (Chung et al, 2014; Tawonsawatruk et al, 2016).
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