Wound Healing| Volume 158, ISSUE 3, P846-856, September 2015

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Angiopoietin-1 improves endothelial progenitor cell–dependent neovascularization in diabetic wounds

  • Swathi Balaji
    Laboratory for Regenerative Wound Healing, Division of Pediatric, General, Thoracic and Fetal Surgery, Cincinnati Children's Hospital Medical Center, Cincinnati, OH
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  • Nate Han
    Laboratory for Regenerative Wound Healing, Division of Pediatric, General, Thoracic and Fetal Surgery, Cincinnati Children's Hospital Medical Center, Cincinnati, OH
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  • Chad Moles
    Laboratory for Regenerative Wound Healing, Division of Pediatric, General, Thoracic and Fetal Surgery, Cincinnati Children's Hospital Medical Center, Cincinnati, OH
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  • Aimen F. Shaaban
    Laboratory for Regenerative Wound Healing, Division of Pediatric, General, Thoracic and Fetal Surgery, Cincinnati Children's Hospital Medical Center, Cincinnati, OH
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  • Paul L. Bollyky
    Division of Infectious Diseases, Department of Medicine, Stanford University School of Medicine, Stanford, CA
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  • Timothy M. Crombleholme
    Laboratory for Regenerative Wound Healing, Division of Pediatric, General, Thoracic and Fetal Surgery, Cincinnati Children's Hospital Medical Center, Cincinnati, OH

    Center for Children's Surgery, Children's Hospital Colorado and the University of Colorado School of Medicine, Aurora, CO
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  • Sundeep G. Keswani
    Reprint requests: Sundeep G. Keswani, MD, Laboratory for Regenerative Tissue Repair, Division of Pediatric Surgery, Texas Children's Hospital and Baylor College of Medicine, 6701 Fannin St. Suite 1210, Houston, TX 77030.
    Laboratory for Regenerative Wound Healing, Division of Pediatric, General, Thoracic and Fetal Surgery, Cincinnati Children's Hospital Medical Center, Cincinnati, OH
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      The diabetic phenotype of wound healing is in part characterized by impaired neovascularization and deficient endothelial progenitor cell (EPC) recruitment. Angiopoietin-1 (Ang-1) is a potent mobilizer of EPCs from the bone marrow (BM). A suggested mechanism for EPC mobilization from the BM is mediated by matrix metalloproteinase 9 (MMP-9) and stem cell factor (SCF). Taken together, we hypothesized that overexpression of Ang-1 in diabetic wounds will recruit EPCs and improve neovascularization and wound healing.


      An endothelial lineage BM-labeled murine model of diabetes was developed to track BM-derived EPCs. FVBN mice were lethally irradiated and then reconstituted with BM from syngeneic Tie2/LacZ donor mice. Diabetes was induced with streptozotocin. Dorsal wounds in BM-transplanted mice were treated with Ad-Ang-1, Ad-GFP, or phosphate-buffered saline. At day 7 after injury, wounds were harvested and analyzed. A similar experiment was conducted in EPC mobilization deficient MMP-9 –/– mice to determine whether the effects of Ang-1 were EPC-dependent.


      Overexpression of Ang-1 resulted in greatly improved re-epithelialization, neovascularization, and EPC recruitment in diabetic BM-transplanted wounds at day 7. Ang-1 treatment resulted in increased serum levels of proMMP-9 and SCF but had no effect on vascular endothelial growth factor levels. According to our FACS results, peripheral blood EPC (CD34+/Cd133+/Flk1+) counts at day 3 after wounding showed impaired EPC mobilization in MMP-9 –/– mice compared with those of wild-type controls. EPC mobilization was rescued by SCF administration, validating this model for EPC-mobilization–deficient mechanistic studies. In MMP-9 –/– mice, Ad-Ang-1 accelerated re-epithelialization in a similar manner, but had no effect on neovascularization.


      Our results show that Ang-1 administration results in improved neovascularization which is dependent on EPC recruitment and has direct effects on wound re-epithelialization. These data may represent a novel strategy to correct the phenotype of impaired diabetic neovascularization and may improve diabetic wound healing.
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