Fms-like tyrosine kinase 3 ligand (Flt3L) administration leads to dramatic increases in dendritic cells (DC) in lymphoid and nonlymphoid tissues. Conversely, mice lacking Flt3L (Flt3L(-)/(-)) show severe reductions in both myeloid (CD11c(+)CD8alpha(-)) and lymphoid-related DC (CD11c(+)CD8alpha(+)) in the thymus and secondary lymphoid organs. In this study marked reductions in CD11c(+) interstitial cardiac DC and in dermal, but not epidermal, DC (Langerhans cells) were also observed. CD11c(+) cells that migrated from Flt3L(-/-) skin explants expressed lower surface MHC class II and costimulatory molecules and naive T cell allostimulatory activity than migratory wild-type (wt) C57BL/6 (B6) CD11c(+) cells. We examined the survival of Flt3L(-)/(-) heart or tail skin grafts (H2(b)) in allogeneic wt (BALB/c; H2(d)) recipients. The outcome of transplantation of BALB/c organs into Flt3L(-)/(-) recipients was also determined. Flt3L(-)/(-) mice rejected BALB/c heart or skin grafts with similar kinetics as B6 wt recipients. Trafficking of donor DC into host spleens or draining lymph nodes was markedly reduced after transplantation of Flt3L(-)/(-) heart, but not skin grafts, respectively. Compared with wt hearts, survival of Flt3L(-)/(-) hearts was markedly prolonged in BALB/c recipients (median survival time, 37 and 15 days, respectively; p < 0.001). Skin graft survival was unaffected. Rejection of Flt3L(-/-) hearts was precipitated by infusion of wt donor DC at the time of transplant. Thus, severe depletion of interstitial heart DC resulting from targeted gene disruption prolongs, but does not indefinitely extend, heart survival. Acute rejection of wt grafts in Flt3L(-/-) recipients reflects presumably an intact role of the direct pathway of allorecognition.