miR-15a/-16 Inhibit Angiogenesis by Targeting the Tie2 Coding Sequence: Therapeutic Potential of a miR-15a/16 Decoy System in Limb Ischemia

MicroRNA-15a (miR-15a) and miR-16, which are transcribed from the miR-15a/miR-16-1 cluster, inhibit post-ischemic angiogenesis. MicroRNA (miRNA) binding to mRNA coding sequences (CDSs) is a newly emerging mechanism of gene expression regulation. We aimed to (1) identify new mediators of the anti-ang...

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Published in:Molecular therapy. Nucleic acids 2019-09, Vol.17, p.49-62
Main Authors: Besnier, Marie, Shantikumar, Saran, Anwar, Maryam, Dixit, Parul, Chamorro-Jorganes, Aranzazu, Sweaad, Walid, Sala-Newby, Graciela, Madeddu, Paolo, Thomas, Anita C., Howard, Lynsey, Mushtaq, Sobia, Petretto, Enrico, Caporali, Andrea, Emanueli, Costanza
Format: Article
Language:English
Subjects:
Amputation
Angiogenesis
angiopoietins
Binding sites
Endothelial cells
Gene expression
Gene regulation
Grants
Hypoxia
Investigations
Ischemia
IVIS
MicroRNAs
miR decoy
miR-15a
miR-16a
miRNA
mouse ischemia model
Musculoskeletal system
Perfusion
Proteins
R&D
Research & development
Skeletal muscle
Tie2
Umbilical vein
Vascular endothelial growth factor
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Summary:MicroRNA-15a (miR-15a) and miR-16, which are transcribed from the miR-15a/miR-16-1 cluster, inhibit post-ischemic angiogenesis. MicroRNA (miRNA) binding to mRNA coding sequences (CDSs) is a newly emerging mechanism of gene expression regulation. We aimed to (1) identify new mediators of the anti-angiogenic action of miR-15a and -16, (2) develop an adenovirus (Ad)-based miR-15a/16 decoy system carrying a luciferase reporter (Luc) to both sense and inhibit miR-15a/16 activity, and (3) investigate Ad.Luc-Decoy-15a/16 therapeutic potential in a mouse limb ischemia (LI) model. LI increased miR-15a and -16 expression in mouse muscular endothelial cells (ECs). The miRNAs also increased in cultured human umbilical vein ECs (HUVECs) exposed to serum starvation, but not hypoxia. Using bioinformatic tools and luciferase activity assays, we characterized miR-15a and -16 binding to Tie2 CDS. In HUVECs, miR-15a or -16 overexpression reduced Tie2 at the protein, but not the mRNA, level. Conversely, miR-15a or -16 inhibition improved angiogenesis in a Tie2-dependent manner. Local Ad.Luc-Decoy-15a/16 delivery increased Tie2 levels in ischemic skeletal muscle and improved post-LI angiogenesis and perfusion recovery, with reduced toe necrosis. Bioluminescent imaging (in vivo imaging system [IVIS]) provided evidence that the Ad.Luc-Decoy-15a/16 system responds to miR-15a/16 increases. In conclusion, we have provided novel mechanistic evidence of the therapeutic potential of local miR-15a/16 inhibition in LI.
ISSN:2162-2531
2162-2531
DOI:10.1016/j.omtn.2019.05.002