Cdh5-lineage independent origin of dermal lymphatic vessels revealed by temporally restricted lineage tracing

The developmental origins of lymphatic endothelial cells (LECs) have been under intense research after a century-long debate. Although previously thought to be of solely venous endothelial origin, additional sources of LECs were recently identified in multiple tissues in mice. Here, we investigated the regional differences in the origin(s) of the dermal lymphatic vasculature by lineage tracing using the pan-endothelial Cdh5-CreER T2 line. Tamoxifen-induced labeling of blood ECs at E9.5, prior to initiation of lymphatic development, traced the majority of dermal LECs but with lower efficiency in the lumbar compared with cervical skin. By contrast, when used at E9.5 but not at E11.5, 4-hydroxytamoxifen, the active metabolite of tamoxifen that provides a tighter window of Cre activity, revealed low labeling frequency of LECs, and lymphvasculogenic clusters in the lumbar skin in particular. Temporally restricted lineage tracing thus reveals contribution of LECs of Cdh5 -lineage independent origin to dermal lymphatic vasculature. Our results further highlight the Cre induction strategy as a critical parameter in defining the temporal window for stage-specific lineage tracing during early developmental stages of rapid tissue differentiation.