PDGF ENHANCES THE PRO-REGENERATIVE PROPERTIES OF EXTRACELLULAR VESICLES RELEASED FORM ADIPOSE MESENCHYMAL STEM CELLS

Abstract

Full Text

INTRODUCTION: Adipose mesenchymal stem cells (ASCs) promote angiogenesis and tissue regeneration through paracrine mechanisms. We have previously shown that platelet derived growth factor (PDGF) stimulate ASCs to secret extracellular vesicles (PDGF-EVs) with a stronger pro-angiogenic potential than EVs secreted in basic conditions (EVs). The aim of the present study was to investigate the molecular mechanism involved in angiogenic, regenerative and immunomodulatory activity of PDGF-EVs. METHODS: For this purpose we studied in vitro the effects of PDGF-EVs on the secretion of inflammatory factors by peripheral blood mononuclear cells (PBMCs) as well as their influence on PBMC adhesion on endothelial cells. EVs were used for comparison. In vivo we have also studied the effects of EVs and PDGF-EVs in an acute limb ischemia pre-clinical model. The molecular differences between EVs and PDGF-EVs were also investigated. RESULTS: In vivo results demonstrate that PDGF-EVs was significantly more effective in restoring large vessel reperfusion and muscle tissue regeneration. More of these, PDGF-EVs inhibited inflammatory cell recruitment in injured tissue, then EVs stimulate immune cell infiltration. In vitro control EVs but not PDGF-EVs enhanced PBMC adhesion on endothelium, confirming our in vivo results. In addition, PDGF-EVs were able to stimulate nitric oxide production in endothelium cells that could be implicated in PBMC adhesion. Direct stimulation of PBMC with control EVs induced secre Гены & Клетки Том XII, № 3, 2017 18 материалы III национального конгресса по регенеративной медицине tion of pro-inflammatory factors such as IFN-γ, IL-1, IL-17 and TNF-α. Whereas PDGF-EVs not only attenuated the secretion of pro-inflammatory factors, but also significantly enhanced the expression of TGF-β1, well-described anti-inflammatory factors, implicated in the differentiation of regulatory T cells. Indeed we have shown that stimulation of PBMC with PDGF-EVs increased the population of Treg in vitro. Proteomic analysis demonstrated differences in pro-angiogenic and pro-inflammatory protein content between PDGF-EVS and control EVs. In particular PDGF-EVs were enriched in HGF, TGFα/β and their receptors, IL-1 ra, VEGF, Tie, OSM, uPA, uPAR, MMPs, Thrombospondins, BDNF, ICAM, IGF. While bEVs carried high levels of CD80, G-CSF, GM-CSF and CD40/TNFRSF5. PDGF-EVs were also enriched in pro-regenerative microRNAs, such as miR-130a, miR-19a, miR-296, miR-17, miR-21, miR-92a, miR-34b, miR-520d, miR-377, miR-146b and long non-coding RNA such as MALAT1. CONCLUSIONS: This study demonstrates that PDGF stimulates ASCs to secrete EVs enriched in anti-inflammatory and proregenerative factors, which could account for PDGF-EV-mediated protection against ischemia reperfusion injury.