E18 - Healing ischemic wounds by revascularization with angiogenic biomaterials

Introduction: : Tissue healing requires vascular expansion to support cellular remodeling and regeneration, yet clinical approaches to localized small vessel angiogenesis and vascular regeneration remain elusive. Further, persistent impaired microcirculation due to injury, ischemia, or disease perpetuates maladaptive remodeling and disrupts tissue function. Inspired by innate regenerative models of the heart in zebrafish and neonatal mice, where a flourishing vascularization response initiates regeneration via cell proliferation and tissue remodeling, we pursued development of a broad angiogenic therapeutic for stimulating tissue regeneration.

Materials and

Methods: : We developed a fully defined biomaterial delivery system using heparin-modified alginate and collagen type 1 and evaluated its angiogenic potential in vivo in a subcutaneous gel plug assay in WT immune competent rats. To identify a highly potent protein cocktail for stimulating angiogenesis in the local environment of the biomaterial, we employed an iterative fractional factorial design of experiments (DOE) approach to evaluate in vivo angiogenesis using combinations of ten proteins (VEGF, bFGF, Shh, PDGF, IGF-1, EGF, TGF-β1, Ang-1, MCP-1 and MMP-9), followed by experiments assessing the top five proteins in cocktails and empirical assessment of 1-3 protein released from the heparinized-alginate-collagen hydrogel. The biomaterial and final cocktail was assessed in a rat model of ischemic dermal wound healing that requires rapid vascular flourishing for wound closure and skin cell regeneration.

Results, Conclusions, and Discussions:: The novel heparinized-alginate-collagen hydrogel biomaterial prolonged protein release up to 14 days in vitro versus Matrigel and unmodified alginate (2-4 days) and increased average lumen diameter, diameter range up to 200 µm, and endothelial cell infiltration 12-fold versus Matrigel in vivo. The top five performing angiogenic factors were VEGF, bFGF, Shh, PDGF, and IGF-1. Empirical evaluation of up to 3 factors delivered in our novel biomaterial identified an optimum combination of VEGF, IGF-1, and PDGF (VIP) for rapid, local vascularization, enhancing perfused vascular density by 6-fold vs. unloaded biomaterial. In ischemic skin wounds, VIP-loaded biomaterial enhanced vascularization, increased both the rate and magnitude of wound closure by 14 days, and enabled restoration of multi-layered skin regeneration with cuboidal skin stem cells in the healed epidermis. In conclusion, we identified highly potent cocktails, including VIP, and leveraged controlled release from a defined heparinized-alginate-collagen biomaterial as a tailored revascularization therapy for local angiogenesis and vascular remodeling to enable tissue regeneration, as shown in vivo in ischemic skin wounds.

Acknowledgements and/or References (Optional):: We gratefully acknowledge funding from NIH R01 HL135091 and NIH R01 HL173938.