Supplementary Material for: Macrophages and Fibroblasts Differentially Contribute to Tattoo Stability

Background: Little information is available about the complexity and function of skin cells contributing to the high stability of tattoos. It has been shown that dermal macrophages play an important role in the storage and maintenance of pigment particles. By contrast, the impact of dermal fibroblasts, forming the connective tissue of the skin, on the stability of the tattoo is not known. Method: In this study, we compared the cell number and the particle load in dermal macrophages versus dermal fibroblasts, isolated from tail skin of tattooed mice. Results: Microscopic analysis revealed that both cell populations contained the tattoo particles, although in largely different amounts. A small number of macrophages with high side scatter intensity contained a large quantity of pigment particles, whereas a high number of dermal fibroblasts harbored only a few pigment particles. Using the CD64^dtr mouse model that allows for selective, diphtheria toxin-mediated depletion of macrophages, we have previously shown that macrophages hold the tattoo in place by capture-release and recapture cycles. In the tattooed skin of macrophage-depleted mice, the content of pigment particles in fibroblasts did not change; however, the total number of fibroblasts carrying particles increased. Conclusion: The present study demonstrates that dermal macrophages and fibroblasts contribute in different ways to the tattoo stability and further improves our knowledge on tattoo persistence.