Proliferation of Ly6C+ monocytes/macrophages contributes to their accumulation in mouse skin wounds

Monocytes and macrophages (Mo/MΦ) play critical roles in all phases of skin wound healing. The majority of these cells are thought to be recruited from blood Mo; however, the role local proliferation of Mo/MΦ in the wound has not been defined. Therefore, we tested the hypothesis that local proliferation of Mo and/or MΦ contributes to their accumulation during wound healing. Male C57Bl/6 mice (N = 4-9/group) were subjected to excisional skin wounding. Proliferating Mo/MΦ (F4/80+Ki67+) were observed in wound cryosections, peaking on day 5 post-wounding. Cell cycle analysis on cells isolated from skin tissue revealed that wounding increased both the number and percentage of inflammatory Ly6C+F4/80lo/- Mo/MΦ in the S/G2/M phases, peaking on day 6 post-wounding. In contrast, more mature Ly6C-F4/80+ cells were found predominantly in the G0 phase with less than 1% cells in S/G2/M phase following injury. In peripheral blood, Mo were very rarely found in the S/G2/M phase, suggesting that the wound environment triggered the Ly6C+F4/80lo/- Mo proliferative response. Furthermore, injury induced several potential regulators of proliferation in wounds, including IL-1β and IL-6, and wound Mo/MΦ expressed surface receptors for these cytokines. However, wound Mo/MΦ proliferation was not altered in IL-1R1 knockout (KO) or IL-6 KO mice. In summary, our findings indicate that proliferation contributes to Mo/MΦ accumulation in wounds and, contrary to findings in other pathophysiologic conditions, Ly6C+/F4/80lo/- Mo/MΦ proliferate during skin wound healing whereas mature Ly6C-F4/80+ MΦ do not.