Withdrawal of TNF-α after the fifth day of differentiation of CD34+ cord blood progenitors generates a homogeneous population of Langerhans cells and delays their maturation

: Human cord blood CD34+ progenitors cultured in the presence of granulocyte‐macrophage colony‐stimulating factor (GM‐CSF), tumor necrosis factor‐α (TNF‐α) and transforming growth factor‐β (TGF‐β) generate a heterogeneous population of dendritic cells (DC), including Langerhans cells (LC). This combination of cytokines has been shown to be crucial for differentiation into LC. After day 5 of culture, TNF‐α has been maintained in the medium in most studies despite the observation of spontaneous maturation of LC after day 12. Five‐day samples of in vitro differentiated LC were cultured in parallel with or without TNF‐α. The absence of TNF‐α was shown to: (1) slow down proliferation without triggering apoptotic cell death, (2) enhance the percentage of LC, (3) delay or abrogat the expression of CD83, CD86, HLA‐DR and CD208 molecules, and (4) maintain endocytosis by receptor and macropinocytosis. The withdrawal of TNF‐α abrogated the spontaneous synthesis of matrix metalloproteinases. At day 12, TNF‐α‐deprived LC were less efficient in allogeneic T cell activation than LC cultivated with TNF‐α. These data indicate that the suppression of TNF‐α after day 5 maintains cells in an immature state and provides a population with 80% of LC at day 12.