Adult Langerhans Cells Derive Directly From Embryonic Macrophages.

Abstract 3788

Dendritic cells (DCs) represent the most potent antigen-presenting cells in vertebrates. Upon immune challenge, DCs can uptake antigens and migrate to lymphoid organs to activate naïve T cells. Therefore, understanding their biology is important for designing new treatments. Langerhans cells (LCs) are a subset of DCs residing in the adult epidermis. In adult teleosts, many evidences point to the existence of DCs, but no tools have been generated to enable in vivo imaging, prospective isolation and functional analyses of this subset.

By combining two new transgenic lines, cd45:DsRED and mhc2dab:eGFP, we could isolate mononuclear phagocytes (macrophages/DCs) in the CD45⁺DAB⁺ fraction from adult tissues. Zebrafish DC-like cells displayed cytological features that resembled those of mammalian DCs. In the skin, the majority of CD45⁺DAB⁺ cells exhibited a typical morphology of LCs. Thus, we can isolate a pure population of LCs from the adult skin.

Most DC subtypes are constantly replenished from hematopoietic stem cell derived progenitors. However, recent studies indicate that LCs self-renew independently from the bone marrow in steady-state conditions, suggesting the existence of a local progenitor in the skin. Recent reports showed that mouse LCs derive from embryonic myeloid precursors that seed the skin before undergoing proliferation. By combining imaging and gene expression analyses, we could show that mhc2dab is first expressed in myeloid cells (CD45⁺) at 7 days post fertilization (dpf). Accordingly, we can first observe CD45⁺DAB⁺ cells at 8 dpf, in the skin of developing embryos, suggesting that LCs derive from early CD45⁺ cells. In order to unravel their origin, we traced CD45⁺ cells by using the tamoxifen-inducible cd45:CRE-ERt2 transgenic line, combined to a switch reporter that allows us to permanently label cells. This switch-reporter consists of the b-actin promoter driving the expression of DsRED after a STOP cassette is excised by the CRE recombinase. By exposing double-transgenic embryos to tamoxifen between 20 and 26 hpf, we could induce CRE expression in primitive macrophages only. Several weeks after tamoxifen treatment, we could identify the progeny of primitive macrophages in the skin, showing LC morphology. This strongly suggests that primitive macrophages can persist to adulthood and contribute to the LC network. Additional experiments will be required to prove if they are the only source of LCs in steady-state conditions.