Fig 2: A) Gating strategy for CD14+ cells. B) PD-L1 expression in CD14+ cells of healthy controls, Covid -19 positive and septic patients. Results are presented as both Mean Fluorescence Intensity and % of CD14+ cells. C) Gating strategy for CD3+ cells, CD4+, CD8+ and CD25+ CD127-cells (Tregs). D) PD-L1 expression in CD3+ cells (CD4+, CD8+ and Tregs) of healthy controls, Covid -19 positive and septic patients. Results are presented as both Mean Fluorescence Intensity and % of CD3+ cells.

Fig 3: Human ILC2 have fused and polarized mitochondria.a, ILC2 Gating strategy Live CD45+ CD7+ Lin- CD94- NKG2A- CD56- CD16- CD127+ CRTh2+ (ILC2) and Live CD45+ CD7+ Lin- CD94+ CD16+ (NKDim). b, Percentage of ILC2 and NKDim in live CD45+ cells. c, FACS measurement of TMRM and MitoTracker in healthy donors of fresh ILC2 and NKDim (n = 12). d, Confocal microscopy in fresh ILC2 and NKDim (n = 7). e, MitoTracker and TMRM intensity (SD/Mean). f, ATP and ADP intensities quantified by label-free mass spectrometry in freshly sorted ILC2 and NKDim cells (n = 3). a-b, Data representative of 10 independent experiments with 2 to 4 donors each. c, Data from 12 healthy donors from at least 5 independent experiments. d-e, Data representative of 2 independent experiments with at least 3 donors. Dots are single cells in the analyzed field. Scale bar 5µm. f, Each dot represents a donor. A minimum of 2 technical replicates were analyzed per run. b, e, f, Floating bars indicate the mean, minimum and maximum values within the dataset. Statistics were assessed by two tailed t-test (b,e,f), not significant (ns), p > 0.05; **p < 0.01; ****p < 0.0001. Source data

Fig 4: ILC2 have enhanced mitochondrial activity compared to naïve T cells.a, Gating strategy for CD4+ T cells subsets in the blood of healthy donors. FACS measurement of TMRM and MitoTracker gMFI in the indicated subsets (n = 4). b-f, Paired blood and tonsil samples were collected from pediatric donors. The percentage of ILC2 (b), GATA-3 gMFI (c), CRTH2 gMFI (d), representative plots of CD127, CD161, ST2 (e), TMRM and Mitotracker gMFI (f) were monitored by FACS (n = 3). a, Data representative of 2 experiments with at least 3 donors each. b-f, Data are representative of 2 independent experiments with 3 donors each. a, Floating bars indicate the mean, minimum and maximum values within the dataset. b-f, Bars are ± s.e.m. Statistics were assessed by one-way ANOVA with Dunnett correction (a) and two tailed t-test (b-f), not significant (ns), p > 0.05; *p < 0.05; **p < 0.01; ****p < 0.0001. Source data

Fig 5: Murine ILCPs yield group 1, 2, and 3 ILCs after co-culture with SIOs(A) Confocal image of SIOs with apical ZO-1 and Lyzozyme-1+ Paneth cells (scale bar: 50 µm).(B) Diagram of co-culture system and representative confocal image of SIO-ILCP co-culture (scale bar: 50 µm).(C) Schematic of experimental design.(D) Representative flow plot of DAPI-Lineage- (hematopoeitic stem cell [HSC] cocktail [CD3, CD45R, CD11b, TER-119] Ly-G6, CD5, CD19, and NK1.1), CD127+a4ß7+Flt3-, PD-1+ ILCP, and CD25+ ILC2P, gated based on PD-1 fluorescence minus one (FMO) controls (overlaid in red).(E) Quantification of ILCP count yielded per animal (N = 15 animals, five experiments).(F) Representative flow plots of DAPI-EpCAM+CD45- IECs and EpCAM-CD45+ ILCs from 7-day cultures of PD-1+ ILCPs only, ILCP + SIO, and SIO-only cultures. Arrows indicate corresponding lack of Lineage markers.(G) Fold change of CD45+Lineage- cells yielded after culture relative to the number of seeded precursors (N = 15 animals, five independent experiments).(H) Contour plot overlay representing expression of NKp46, ROR?t, NKp46, NK1.1, Klrg1, and Sca-1 in Lineage- populations derived from (F) in populations pregated as putative group 1 (magenta; Live, EpCAM-, CD45+, Lin-, ROR?t-, Klrg1-, NK1.1+, NKp46+), group 2 (green; Live, EpCAM-, CD45+, Lin-, ROR?t-, NKp46-, Klrg1+, Sca-1+), NKp46+ group 3 (lavender; Live, EpCAM-, CD45+, Lin-, Klrg1-, ROR?t+, NKp46+), NKp46- group 3 (blue; Live, EpCAM-, CD45+, Lin-, Klrg1-, ROR?t+, NKp46-), and a putative immature “other” population (gray; Live, EpCAM-, CD45+, Lin-, ROR?t-, Klrg1-, NK1.1-, NKp46-) in cultures derived from ILCPs only or ILCP co-culture with SIOs.(I) Quantification of pooled ILC groups depicted in (H) (ILCPs from N = 4 animals).Error bars represent SEM; p values are from unpaired Student’s t tests.