Conclusion
UC-MSCs derived exosomes could inhibit H9C2 cells apoptosis induced by H/SD through regulating autophagy via PI3K/Akt/mTOR pathway. Hypoxia preconditioning could enhance above effects through increasing exosomes secretion of UC-MSCs.
Methods
Exosomes were isolated from normal UC-MSCs culture media and hypoxic preconditioning culture media. Transmission electron microscopy was used to observe the morphology of exosomes. Nanoparticle tracking analysis was used to detect the size distribution and concentration of exosomes. Western blot analysis was used to analyzed the surface marker CD63 of exosomes. H9C2 cells were induced apoptosis by hypoxia and serum deprivation (H/SD) and then were treated respectively by group. Cell Counting Kit-8 assay was used to detect viability of H9C2 cells. Apoptosis was detected by Hochest staining and annexin V-FITC/PI. The expression levels of related proteins of apoptosis, autophagy, and PI3K/Akt/mTOR pathway were analyzed by Western blot analysis. Immunofluorescence was used to analyze LC3B expression.
Results
Hypoxic preconditioning increased the exosomes secretion of UC-MSCs. UC-MSCs derived exosomes could inhibit H/SD-induced H9C2 cells apoptosis. Hypoxic preconditioning strengthened this antiapoptosis effect of UC-MSCs. Hypoxic preconditioning UC-MSCs derived exosomes (H-Exo) downregulated LC3B-II/I and beclin-1 and upregulated P62, p-Akt/Akt and p-mTOR/mTOR. The antiapoptotic effect of H-Exo could be attenuated by treatment with LY294002 and rapamycin.