Mechanism of Skin Improvement in Radiation Wounds Following Fat Grafting: The Fate of Adipose-derived Stem Cells and Role of Stromal Vascular Fraction

Abstract

PURPOSE: Consequences of radiation include thickened, fibrotic and inelastic skin. Fat grafting helps alleviate this damage by decreasing epidermal thickness, increasing vascularization, and decreasing fibrosis. The exact mechanism is not understood and carries many hypotheses including the effects of adipose-derived stem cells (ADSCs) within fat. The first aim of this study was to determine which components of fat cause benefits seen with postradiation fat grafting with the hypothesis that pure ADSC group would have the greatest difference in epidermal thickness. The second aim was to determine the mechanism by which these skin changes are mediated with the hypothesis that human ADSCs can differentiate into epithelial stem cells to regenerate the skin. METHODS: The dorsal skin of nude mice was directly radiated. Four weeks postradiation, injections were performed under the radiated skin with either human lipoaspirate, stromal vascular fraction, or pure ADSCs. The pure ADSCs were confirmed as p63− with flow cytometry before injection. The mice were euthanized at 2 and 4 weeks postinjection. Epidermal thickness was measured to determine treatment effect. Immunohistochemistry was performed using an antibody specific for human epithelial stem cell marker p63 and imaged using confocal microscopy. Nuclei positive for DAPI or p63 were quantified using ImageJ. RESULTS: At 2 weeks, all experimental groups that were injected with human cells (ADSC, SVF, and lipoaspirate) had statistically thinner epidermis compared to the radiation-only control group without statistical differences between experimental groups. At 4 weeks, lipoaspirate and SVF groups remained statistically thinner than control groups with no statistical difference between the two. At 4 weeks, the epidermal thickness of the ADSC group was not statistically different than controls. There was a significant decrease in epidermal thickness from week 2 to week 4 in the lipoaspirate, SVF, and matrigel-only groups. Immunohistochemistry showed the presence of p63+ human cells in all experimental groups and absence in control groups. At 2 weeks, there is a statistically higher percent of p63+ cells in the ADSC and SVF groups compared to all other groups. From week 2 to week 4, there was a significant increase in the percent of p63+ cells present in the lipoaspirate group. At week 4, all experimental groups had a statistically higher percentage of p63+ cells than control groups without statistical differences between the experimental groups. CONCLUSIONS: These findings suggest that improvements seen in radiated skin after fat grafting are due to presence of transferred ADSCs. ADSCs are likely not the only factor necessary to mediate the changes and the other components present within the SVF and lipoaspirate are likely important because these 2 groups maintained significantly thinner epidermis at 4 weeks, whereas the pure ADSC group did not. The ADSCs seem to convert into epithelial stem cells as evidenced by the presence of p63+ human cells within the epidermis of the experimental groups and absence in control groups. The increase in percentage of p63+ cells from week 2 to week 4 suggests that these stem cells are continuing to divide and regenerate the skin.