title: Therapeutic Angiogenesis by Ultrasound-Mediated MicroRNA-126-3p Delivery.
abstract: OBJECTIVE
  MicroRNAs are involved in many critical functions, including angiogenesis. Ultrasound-targeted microbubble destruction (UTMD) is a noninvasive technique for targeted vascular transfection of plasmid DNA and may be well suited for proangiogenic microRNA delivery. We aimed to investigate UTMD of miR-126-3p for therapeutic angiogenesis in chronic ischemia.
  
  
  RESULTS
  The angiogenic potential of miR-126-3p was tested in human umbilical vein endothelial cells in vitro. UTMD of miR-126-3p was tested in vivo in Fischer-344 rats before and after chronic left femoral artery ligation, evaluating target knockdown, miR-126-3p and miR-126-5p expression, phosphorylated Tie2 levels, microvascular perfusion, and vessel density. In vitro, miR-126-3p-transfected human umbilical vein endothelial cells showed repression of sprouty-related protein-1 and phosphatidylinositol-3-kinase regulatory subunit 2, negative regulators of vascular endothelial growth factor and angiopoietin-1 signaling, increased phosphorylated Tie2 mediated by knockdown of phosphatidylinositol-3-kinase regulatory subunit 2 and greater angiogenic potential mediated by both vascular endothelial growth factor/vascular endothelial growth factor R2 and angiopoietin-1 /Tie2 effects. UTMD of miR-126-3p resulted in targeted vascular transfection, peaking early after delivery and lasting for >3 days, and resulting in inhibition of sprouty-related protein-1 and phosphatidylinositol-3-kinase regulatory subunit 2, with minimal uptake in remote organs. Finally, UTMD of miR-126-3p to chronic ischemic hindlimb muscle resulted in improved perfusion, vessel density, enhanced arteriolar formation, pericyte coverage, and phosphorylated Tie2 levels, without affecting miR-126-5p or delta-like 1 homolog levels.
  
  
  CONCLUSIONS
  UTMD of miR-126 results in improved tissue perfusion and vascular density in the setting of chronic ischemia by repressing sprouty-related protein-1 and phosphatidylinositol-3-kinase regulatory subunit 2 and enhancing vascular endothelial growth factor and angiopoietin-1 signaling, with no effect on miR-126-5p. UTMD is a promising platform for microRNA delivery, with applications for therapeutic angiogenesis.
authors.affiliation: From the Division of Cardiology, Keenan Research Centre for Biomedical Science, Li Ka Shing Knowledge Institute, St Michael's Hospital, University of Toronto, Ontario, Canada.
authors.forename: Wei J
authors.initials: WJ
authors.lastname: Cao
authors.affiliation: From the Division of Cardiology, Keenan Research Centre for Biomedical Science, Li Ka Shing Knowledge Institute, St Michael's Hospital, University of Toronto, Ontario, Canada.
authors.forename: Joshua D
authors.initials: JD
authors.lastname: Rosenblat
authors.affiliation: From the Division of Cardiology, Keenan Research Centre for Biomedical Science, Li Ka Shing Knowledge Institute, St Michael's Hospital, University of Toronto, Ontario, Canada.
authors.forename: Nathan C
authors.initials: NC
authors.lastname: Roth
authors.affiliation: From the Division of Cardiology, Keenan Research Centre for Biomedical Science, Li Ka Shing Knowledge Institute, St Michael's Hospital, University of Toronto, Ontario, Canada.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Kuliszewski
authors.affiliation: From the Division of Cardiology, Keenan Research Centre for Biomedical Science, Li Ka Shing Knowledge Institute, St Michael's Hospital, University of Toronto, Ontario, Canada.
authors.forename: Pratiek N
authors.initials: PN
authors.lastname: Matkar
authors.affiliation: From the Division of Cardiology, Keenan Research Centre for Biomedical Science, Li Ka Shing Knowledge Institute, St Michael's Hospital, University of Toronto, Ontario, Canada.
authors.forename: Dmitriy
authors.initials: D
authors.lastname: Rudenko
authors.affiliation: From the Division of Cardiology, Keenan Research Centre for Biomedical Science, Li Ka Shing Knowledge Institute, St Michael's Hospital, University of Toronto, Ontario, Canada.
authors.forename: Christine
authors.initials: C
authors.lastname: Liao
authors.affiliation: From the Division of Cardiology, Keenan Research Centre for Biomedical Science, Li Ka Shing Knowledge Institute, St Michael's Hospital, University of Toronto, Ontario, Canada.
authors.forename: Paul J H
authors.initials: PJ
authors.lastname: Lee
authors.affiliation: From the Division of Cardiology, Keenan Research Centre for Biomedical Science, Li Ka Shing Knowledge Institute, St Michael's Hospital, University of Toronto, Ontario, Canada. leong-poih@smh.ca.
authors.forename: Howard
authors.initials: H
authors.lastname: Leong-Poi
mesh_terms: D042501:Angiogenic Proteins / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D004195:Disease Models, Animal; D005786:Gene Expression Regulation; D015316:Genetic Therapy / Q000379:methods*; D006614:Hindlimb; D061307:Human Umbilical Vein Endothelial Cells / Q000378:metabolism*; D006801:Humans; D007511:Ischemia / Q000235:genetics / Q000378:metabolism / Q000503:physiopathology / Q000628:therapy*; D008297:Male; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D045423:Microbubbles; D008833:Microcirculation; D055806:Microvessels / Q000503:physiopathology*; D018482:Muscle, Skeletal / Q000098:blood supply*; D018919:Neovascularization, Physiologic; D011916:Rats, Inbred F344; D012039:Regional Blood Flow; D013997:Time Factors; D014162:Transfection / Q000379:methods*; D014465:Ultrasonics
chemical_list: D042501:Angiogenic Proteins; C527111:MIRN126 microRNA, human; D035683:MicroRNAs
keywords: angiogenesis; gene therapy; ischemia; microRNA; ultrasound