Fig 2: HIOs promote proliferation and maturation of systemic human ILCPs(A) Gating strategy for PBMC-derived ILCPs, pregated on live, single, CD45+ cells (FMOs, blue and magenta).(B) Schematic of HIO + ILCP co-cultures, indicating the presence of mesenchymal cells and CD45+ ILCPs.(C) Representative plot overlays of EpCAM+ IECs, double-negative mesenchymal cells (M, magenta), and CD45+ ILCs (blue) after 14-day co-cultures, highlighting Matrigel debris in gray, and quantified in (D) as count of EpCAM-, CD45+, LINn- ILCs and (E) fold change expansion of ILCs after 14-day co-culture relative to the number of ILCPs seeded on day 1 (N = 3–15 across seven experiments).(F) Representative image of CD45+ ILCPs co-cultured with mesenchyme-depleted, EpCAM+ HIOs (scale bar: 25 µm). Error bars represent SEM; p values are from unpaired Student’s t tests.(G) Count of Live, EpCAM-CD45+LIN-ROR?t+ ILCs after 14-day co-culture with SD-HIOs or epithelial-depleted HIO-STROs expressing markers CCR6, NKp44, and/or T-bet (ILCPs from N = 3 PBMC donors).(H) Relative group 1 (Live, CD45+Lin-CRTH2-c-kit- [CD127+CD161+ in primary gut ILC only] and [EpCAM-ROR?t-GATA3- in SD-HIO and HIO-STRO only]), group 2 (Live, CD45+Lin-CRTH2+c-kit+/-, [CD127+CD161+/- in primary gut ILC only] and [EpCAM-ROR?t-GATA3+ in SD-HIO and HIO-STRO only]), group 3 (Live, CD45+Lin-CRTH2-c-kit+, NKp44+/- and [CD127+ CD161+ in primary gut ILC only] and [EpCAM-ROR?t+GATA3- in SD-HIO and HIO-STRO only]), and other LIN- ILCs (e.g., undifferentiated precursors; Live, CD45+, Lin-, CRTH2-, c-kit- and [CD127+ CD161+ in primary gut ILC only] and [EpCAM-, ROR?t-, GATA3- in SD-HIO and HIO-STRO only]) in unstimulated, primary human intestine (N = 13, adapted from Krämer et al., 2017), SD-HIOs (N = 13 from six experiments), and HIO-STROs (N = 7 from three experiments).