Cultures of peripheral blood mononuclear cells (MNCs) give rise to at least two different variants of endothelial progenitor cells (EPCs), early and late outgrowth EPCs. We investigated whether microRNAs in early and late EPCs could serve as markers of internal processes that can be exploited to distinguish cell identity and functional capacity. We hypothesized that as MNCs give rise to early and late EPCs, there is a gradual change in total microRNA profile, reflecting a total change in processes within the predominant cell population. Using a candidate microRNA array, early and late EPCs showed vastly different microRNA expression profiles. MiR-146a expression increased progressively as early EPCs emerged around 5-7 days (p<0.05). Through targeting TRAF6 and IRAK1, miR-146a conferred inflammatory tolerance in early EPCs, likely contributing to their purported ability to suppress inflammation. MiR-146a knock down (KD) in endotoxin-stimulated early EPCs reduced anti-inflammatory cytokine IL-1RA (p<0.001), and increased expression of pro-inflammatory cytokines IL-1 (p<0.001) and IL-8 (p<0.01). Interestingly, the microRNA expression profile of late EPCs was highly congruent to mature endothelial cells, with 100-fold greater miR-126 expression than monocytes and early EPCs (p<0.01). MiR-126KD in late EPCs abolished matrigel-network formation (p<0.05); while overexpression (OE) in early EPC augmented network formation (p<0.05) and chemotactic migration (p<0.001). We also found that the melanoma cell adhesion molecule or MCAM (CD146) identified late EPC precursors. Only MCAM+MNCs from adult blood (<5% of total MNCs) yielded late EPC-like colonies. Robust miR-126 expression in these cells predicted the generation of late EPCs. Overall, our results suggest that miR-146a in early EPCs likely contributes to repair by suppressing inflammation during cardiovascular injury; while in late EPCs, miR-126 directly promotes angiogenesis and vascular repair. Finally, we highlight a unique method for the efficient generation of late EPCs by using MCAM selection and screening for miR-126.
| Identifer | oai:union.ndltd.org:uottawa.ca/oai:ruor.uottawa.ca:10393/35080 |
| Date | January 2016 |
| Creators | Behbahani, John |
| Contributors | Stewart, Duncan |
| Publisher | Université d'Ottawa / University of Ottawa |
| Source Sets | Université d’Ottawa |
| Language | English |
| Detected Language | English |
| Type | Thesis |
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