Abstract 20117: Exosomes From Hypoxia-Preconditioned Adipose-Derived Stem Cells Play a Novel Pro-Angiogenic Role via Angiogenic MicroRNAs
Adipose-derived stem cells (ASCs) have been reported to be beneficial in therapeutic angiogenesis and myocardial repair. However, as all other stem cells, the application of ASCs in the treatment of cardiovascular disease is limited by the safety concerns such as tumorigenesis and microvasculature occlusion. Our previous studies have demonstrated that exosomes/microvesicles released from ASCs promote angiogenesis. The objectives of this study are to designate role for and determine the molecular mechanisms of exosomes released from hypoxia-preconditioned ASCs in angiogenesis process. The extracted exosomes from normoxia and hypoxia-preconditioned ASCs were identified with nanoparticle tracking analysis and transmission electron microscope. Our results showed that the shape of exosomes were spheroids ranging in 30 - 100 nm with a peak at 56 nm in diameter. Comparison of protein concentration of exosomes and the exosomal marker, Alix, using immunoblot analysis, showed that hypoxia preconditioning of ASCs promoted exosome secretion in 2.6-fold increase. An exosome delivery assay was used to determine the efficiency of exosomal uptake in human microvascular endothelial cells (HMVECs) and quantified using flow cytometry. The results demonstrated that the percentage of delivery to recipient HMVECs happened in a dose-dependent manner with a maximum exosome concentration of 1х107 particles/ml. Furthermore, an increase in HMVEC migration and tube formation was shown after incubation with normoxic ASC-exosomes, which was further enhanced by hypoxic ASC-exosomes. To determine the molecular mechanisms of the angiogenic effect seen by ASC-exosomes, total RNA was isolated from exosomes and subjected to miRNA profiling. The result from miRNA profiling showed that the expressions of pro-angiogenic microRNAs (miRs) were significantly increased in hypoxic ASC-exosome compared to normoxic ASC-exosome, such as miR-146a, -18a, -194, -296, -93, -196b, -143, -107 and -19a. Meanwhile the expression of miR-497, -100, -29 and -34b, which are anti-angiogenic miRs were decreased in hypoxic ASC-exosome. These findings suggest that angiogenic miRNAs as important cargo of hypoxic ASC-exosome may contribute to the pro-angiogenic effect seen in HMVECs.
Author Disclosures: L. Zhao: None. T. Johnson: None. T. Kang: None. D. Liu: None.
- © 2016 by American Heart Association, Inc.