Novel Gene-Modified Mesenchymal Stem Cell Therapy Reverses Impaired Wound Healing in Ischemic Limbs

Abstract

Objective: Here, we report a new method to increase the therapeutic potential of mesenchymal stem/stromal cells (MSCs) for ischemic wound healing. We tested biological effects of MSCs modified with E-selectin, a cell adhesion molecule capable of inducing postnatal neovascularization, on a translational murine model. Background: Tissue loss significantly worsens the risk of extremity amputation for patients with chronic limb-threatening ischemia. MSC-based therapeutics hold major promise for wound healing and therapeutic angiogenesis, but unmodified MSCs demonstrate only modest benefits. Methods: Bone marrow cells harvested from FVB/ROSA26SormTmG donor mice were transduced with E-selectin-green fluorescent protein (GFP)/AAV-DJ or GFP/AAV-DJ (control). Ischemic wounds were created via a 4 mm punch biopsy in the ipsilateral limb after femoral artery ligation in recipient FVB mice and subsequently injected with phosphate-buffered saline or 1×106 donor MSCGFP or MSCE-selectin-GFP. Wound closure was monitored daily for 7 postoperative days, and tissues were harvested for molecular and histologic analysis and immunofluorescence. Whole-body DiI perfusion and confocal microscopy were utilized to evaluate wound angiogenesis. Results: Unmodified MSCs do not express E-selectin, and MSCE-selectin-GFP gain stronger MSC phenotype yet maintain trilineage differentiation and colony-forming capability. MSCE-selectin-GFP therapy accelerates wound healing compared with MSCGFP and phosphate-buffered saline treatment. Engrafted MSCE-selectin-GFP manifest stronger survival and viability in wounds at postoperative day 7. Ischemic wounds treated with MSCE-selectin-GFP exhibit more abundant collagen deposition and enhanced angiogenic response. Conclusions: We establish a novel method to potentiate regenerative and proangiogenic capability of MSCs by modification with E-selectin/adeno-associated virus. This innovative therapy carries the potential as a platform worthy of future clinical studies.