Fig. 1

The growth of clonogenic TECs in the explant niche is supported by TICs including thymic fibroblasts, hematopoietic cells, and lineage negative cells that express high amounts of Aire and IL25 (IL-17E). A Cells from Fraction 3, which include clonogenic TECs and TICs, derived from cleaned thymic explants. B Representative images of epithelial cell colonies derived from thymic explants cultured with DMEM/F-12 FBS 20% and with DMEM/F-12 KnockOut™ 10% at day 14. The control group with DMEM/F-12 KnockOut™ 10% only exhibited small colonies of epithelial cells. Scale bar: 10 µm D = 10 n = 5. C Gating strategy to characterize TICs derived from fraction 3, including thymic fibroblasts, cells from the hematopoietic lineage (Hem lin^POS) and lineage negative cells (lin^NEG). Both lin^NEG cells and fibroblasts highly express IL25 (IL17E) and Aire (n = 2). D Proliferation profile of TICs derived from thymic explants. Blue histograms represent the proliferation profile of thymic fibroblasts, red histograms represent the proliferation profile of Hem lin^POS cells and green histograms represent the proliferation profile of lin^NEG cells (n = 2). E Intracellular blockade of FcRs in organelles and vesicles of the secretory pathway with Trustain FcX™ decreases the percentage of lin^NEG IL25⁺ TICs by 15%, but there are still more than 75% of lin^NEG TICs with truly positive expression of IL25 (n = 2). F Thymic fibroblasts and Hem lin^POS cells, interact and support the growth of clonogenic TECs in explant niche. Scale bar: 20 µm D = 100 n = 2. DIC = Differential interference contrast, Fb = Fibroblasts, FMO = Fluorescence minus one, FcRs = Fragment Crystallizable Receptors, Hem lin^POS = Hematopoietic lineage positive cells, lin^NEG = Lineage negative cells, PDP n = Podoplanin, TICs = Thymic Interstitial Cells