The Use Of Indocyanine Green Fluorescence In A Preclinical Model Of Human Wound Healing
Aos Karim1, Aiping Liu1, Christie Lin2, Adam Uselmann2, Kevin Eliceiri1, Matthew Brown1, Angela Gibson1.
1University of Wisconsin, Madison, WI, USA, 2OnLume, Madison, WI, USA.
BACKGROUND: Indocyanine Green (ICG) fluorescence is used to evaluate microperfusion in both murine models and humans. We characterized graft take in a mouse model of human skin xenograft utilizing ICG and immunohistochemistry to evaluate the utility of a murine-human xenograft as a model for human skin wound healing.
METHODS: Normal human skin was obtained from reconstructive operations. 1 x 2 cm full thickness (FT; thickness = 2mm) or partial thickness (PT; thickness = 0.75mm) pieces of human skin tissue were grafted onto the dorsal flank of immunocompromised mice. Three groups (N=2 per group) were evaluated. Group 1 mice received PT human skin xenografts and were imaged 8 weeks after grafting. Groups 2 and 3 mice received FT human xenografts and were imaged 8 weeks and 12 weeks after grafting, respectively. All mice received retro-orbital injections of ICG (1 mg/kg). ICG fluorescence and white light images were captured immediately post-injection using a novel, ambient light compatible fluorescence imaging system. Mice were sacrificed and the xenografts were harvested for histology.
RESULTS: Gross examination shows contraction of the skin in both FT and PT grafts (area reduction ~50% and ~30%, respectively). Eight weeks post-graft, PT grafts grossly appeared well-engrafted, while FT grafts exhibited a central necrotic region with surrounding well-engrafted human skin adjacent to mouse skin. Twelve weeks post-graft, FT grafts grossly appeared fully engrafted without necrosis. The extent of ICG fluorescence indicated that perfusion was much greater in PT compared to FT (95% vs 10% of graft area) 8 weeks after grafting. Twelve weeks after grafting, FT grafts showed complete perfusion. After 8 weeks, cell viability and revascularization, evaluated histologically with Lactate Dehydrogenase and CD31 respectively, were greater in the PT grafts compared to FT grafts. By 12 weeks, cell viability and revascularization were noted throughout both PT and FT graft tissue.
CONCLUSIONS: Delayed revascularization to the skin tissue during engraftment leads to ischemia-induced cellular death. Once a graft is fully reperfused, necrotic tissue is shed and the resultant wound undergoes full regeneration. We show the feasibility of ICG fluorescence as a marker for monitoring of human skin xenograft engraftment for future wound healing mechanistic studies.
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