Thrombin Preconditioning Boosts Biogenesis of Extracellular Vesicles from Mesenchymal Stem Cells and Enriches Their Cargo Contents via Protease-Activated Receptor-Mediated Signaling Pathways

• Main Authors: Dong Kyung Sung, Se In Sung, So Yoon Ahn, Yun Sil Chang, Won Soon Park
• Format: Article
• Language: English
• Published: MDPI AG 2019-06-01
• Series: International Journal of Molecular Sciences
• Subjects: mesenchymal stem cell; extracellular vesicle; thrombin; protease activated receptors
• Online Access: https://www.mdpi.com/1422-0067/20/12/2899

We investigated the role of protease-activated receptor (PAR)-mediated signaling pathways in the biogenesis of human umbilical cord blood-derived mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) and the enrichment of their cargo content after thrombin preconditioning. Immunoblot analyses showed that MSCs expressed two PAR subtypes: PAR-1 and PAR-3. Thrombin preconditioning significantly accelerated MSC-derived EV biogenesis more than five-fold and enriched their cargo contents by more than two-fold via activation of Rab5, early endosomal antigen (EEA)-1, and the extracellular signal regulated kinase (ERK)1/2 and AKT signaling pathways. Blockage of PAR-1 with the PAR-1-specific antagonist, SCH79797, significantly suppressed the activation of Rab5, EEA-1, and the ERK1/2 and AKT pathways and subsequently increased EV production and enriched EV cargo contents. Combined blockage of PAR-1 and PAR-3 further and significantly inhibited the activation of Rab5, EEA-1, and the ERK1/2 and AKT pathways, accelerated EV production, and enriched EV cargo contents. In summary, thrombin preconditioning boosted the biogenesis of MSC-derived EVs and enriched their cargo contents largely via PAR-1-mediated pathways and partly via PAR-1-independent, PAR-3-mediated activation of Rab5, EEA-1, and the ERK1/2 and AKT signaling pathways.