Abstract 18915: Modulation of Exosomal Pro-angiogenic MicroRNAs from CD34+ Progenitor Cells by Hypoxic-Preconditioning Enhances its Angiogenic Potency and Post-ischemic Tissue Repair
Exosomes, membrane bound nano-vesicles secreted by the cells, has great potential to advance cell-based therapies for ischemic cardiovascular diseases by overcoming its limitations- such as viability and retention, difficulty in isolation of adequate number of cells, or, appropriate storage of the cells. Recently, we have discovered that adult human CD34+ stem cells, one of the most successful cell types for treating cardiovascular diseases, secrete exosomes (Exo) that mediates most of the angiogenic and therapeutic activity of the stem cells via transfer of pro-angiogenic microRNAs. We hypothesize that the regenerative efficacy of the cell-free Exo derived from human CD34+ cells can be improved by hypoxic pre-conditioning, modulating its pro-angiogenic miRNA content.
Methods and Results: CD34+ Exo isolated from hypoxic human CD34+ cells were significantly proliferative, anti-apoptotic and angiogenic both in vitro and in vivo, as compared to CD34+ Exo from normoxic cells. Treatment of hypoxic Exo significantly improved perfusion and prevented ischemic limb amputation, compared with normoxic Exo, in a mouse model of hind limb ischemia. We investigated the possibility of involvement of proteins by comparing the total proteins from hypoxic and normoxic Exo using 2-D differential gel electrophoresis and mass spectrometry; we did not detect any difference in the proteins expressions quantified. Interestingly, bioanalyzer analyses indicated that hypoxic cells and Exo were enriched for miRNAs. The differential expression of miRNAs between hypoxic and normoxic Exo are profiled using miRNA microarray and validated by qRT-PCR. We detected significantly high expression of proangiogenic miRNAs in hypoxic Exo. Flow cytometry and in vitro trafficking by confocal microscopy demonstrates transfer of Cy3 labeled miRNA to recipient endothelial cells. We are studying the involvement of transferred pro-angiogenic miRNAs on hypoxia-Exo-induced angiogenic activity and modulation of target endothelial cell gene expression by loss of function-gain of function studies.
Conclusion: Our results demonstrate that hypoxia modulates the miRNA content of adult human CD34+ progenitor cell derived exosomes improving its angiogenic and therapeutic potency.
- © 2012 by American Heart Association, Inc.