Innate Immune Sensing And Tissue Remodeling Of A Biodegradable Tempering Matrix Supported Islet Graft

Introduction: The subcutaneous site represents an accessible alternative islet transplant site, but suffers from poor vascularisation. Here we test a clinically proven Biodegradable-Temporising-Matrix; NovoSorb™ (IDT) repurposed from use in burn and wound treatment that may provide a platform for subcutaneous islet-transplants. Materials and Methods: Human, neonatal porcine and murine islets were isolated using standard techniques. Islet viability & function was assessed using Annexin/Propodium iodide staining, Glucose Stimulated Insulin Secretion, ELISA and histology. Diabetes was induced using alloxan tetrahydrate i.v. in injectable grade water (20 mg/ml) at a dose of 110 mg/kg body weight. All transplants were surgically placed under the left renal capsule. Microscopy data was obtained using a Zeiss 7MP two-photon microscope. Results and Discussion: When tested in in-vitro co-culture experiments with human islets, IDT showed no impact on islet viability and function. Mouse islets were then transplanted under the renal capsule of diabetic recipients pre-implanted with IDT and resulted in complete recovery and long-term maintenance of euglycemia (>100 days). Also, neonatal porcine islets transplanted with IDT showed increasing porcine insulin production through time (>300 days). These data demonstrate that IDT is non-toxic for human, mouse and porcine islets and does not interfere with normal physiological function of islet grafts. Mice were then transplanted with IDT alone and the in-vivo foreign body reaction and tissue remodelling was assessed using transgenic (LysM-GFP+/tdTomTg) mice: blood vessels express tdTomato red and Lysozyme+ myeloid cells express GFP. Post-operative day (POD) 7 showed LysM-GFP+ myeloid cells infiltrating the graft. By POD 14, LysM-GFP+ myeloid cell-fusion events had occurred evidenced by the presence of giant cells and scattered immature vascular networks with dispersed loose collagen-fibrils. At POD 30, dendritic-like cells were observed, the vascular network was denser and the collagen fibres had increased in thickness. Further; to assess whether innate sensing of the islet graft would occur in the absence of a supportive scaffold, we transplanted islets from whole body tdTomTg mice (Tomato positive islets) into LysM-GFP/+ mice (syngeneic). 24hrs later LysM-GFP+ myeloid cells were migrating over the surface of the islets and were exhibiting neutrophil like ‘swarming’ behaviour with evident disruption of islet tissue. Conclusion: This analysis reveals that IDT can provide a neo-vascularization platform advantageous for subcutaneous islet transplantation and that there is a dynamic interaction between the innate-immune system and the syngeneic islet graft with or without IDT.