Notch signal interruption promotes CD4 and DP cell differentiation.

CB and adult HSCs were grown in OP9Deltapos co-culture, then Notch signalling was removed for 7 days by transfer to OP9Deltaneg co-culture or addition of DAPT. Cells were analysed by flow cytometry for CD4 and CD8 expression. (A) Representative dot plots for adult cells co-cultured for 70 days. (B–E) Adult (B, D) or CB (C, E) HSCs were grown in OP9Deltapos co-culture (▪) and then either transferred to OP9Deltaneg co-culture (○) or incubated with DAPT (▵) for 7 days. Data points show the percentage of CD4+ ISP (B, C) or CD4+ CD8+ DP (D, E) cells, n≥3, mean+SEM, * p<0.05 for DAPT vs OP9Deltapos (t-test), ** p<0.05 for OP9Deltaneg vs OP9Deltapos (t-test). (F) CD14 (▪) expression plotted against DP expression (▴) for adult cells in OP9Deltapos co-cultures after Notch signalling was removed for 7 days. (G) Cell numbers during DAPT treatment. Paired comparisons of six control and DAPT – treated cultures, showing all HSC-derived cells (left) and DP cells (right).