Fig. 6: IL-18 stimulation of cytotoxic NK cells suppresses thymus regeneration.

a,b, Female 1- to 2-month-old C57BL/6 WT or Ifng-reporter mice were exposed to SL-TBI, and thymuses were collected on days 0 and 3 after irradiation. IL-18R⁺ and IL-18R^lo–neg CD45⁺NK1.1⁺TCRβ⁻CD49b⁺ NK cells, CD45⁺NK1.1⁺TCRβ⁻CD49a⁺ ILC1s and CD45⁺NK1.1⁺TCRβ⁺CD49b⁻ NKT cells were compared. a, NK, ILC1 and NKT cellularity on day 0 (n = 3) and day 3 (n = 7) after SL-TBI. b, Fold change in NK cells (n = 10), ILC1s (n = 10) and NKT cells (n = 7) between days 0 and 3 after SL-TBI. c, Female C57BL/6 CD45.1⁺ and CD45.2⁺ mice were surgically conjoined to establish parabiotic pairs when both members of the pair were subjected to SL-TBI. Thymuses were collected, and chimerism was calculated on day 0 (n = 8), day 1 (n = 6), day 4 (n = 4) or day 7 (n = 8) after SL-TBI. Congenic markers (CD45.1 and CD45.2) were used to determine the mouse of origin (that is, cells expressing the same CD45 isoform as the mouse-pair thymus were classified as ‘intrathymically’ derived, while cells expressing the alternate isoform were considered ‘extrathymically’ derived). The numbers of intrathymic or extrathymic CD45⁺NK1.1⁺CD3⁻ NK/ILC1 cells at the indicated time points are shown graphically. d, Concatenated flow cytometry plots showing Ifng-GFP, perforin and granzyme B expression within CD45⁺NK1.1⁺TCRβ⁻CD49b⁺ NK cells. The gMFI of Ifng-GFP, perforin and granzyme B expression in IL-18R^lo–neg and IL-18R⁺ NK cells on day 3 after SL-TBI (n = 8 per group) is shown. e, Female 1- to 2-month-old C57BL/6 WT or Il18r1^−/− mice were exposed to SL-TBI, and thymuses were isolated 3 days later. Concatenated flow cytometry plots and gMFI of perforin expression within CD45⁺NK1.1⁺TCRβ⁻CD49b⁺ NK cells are shown (n = 5 per group). f, Female 1- to 2-month-old Il18r1^fl/fl:Ncr1-Cre⁻ (Il18r1^WT, n = 8) and Il18r1^fl/fl:Ncr1-Cre⁺ (Il18r1^ΔNK/ILC1, n = 9) mice were exposed to SL-TBI, and the total thymic IFNγ and perforin levels were measured 3 days later. g, Ifngr1 and Ifngr2 expression at baseline, taken from the scRNAseq dataset described in Fig. 2a. h, Female 1- to 2-month-old C57BL/6 Ifngr^fl/fl:Foxn1-Cre⁻ (Ifngr^WT, n = 6) and Ifngr^fl/fl:Foxn1-Cre⁺ (Ifngr^ΔTEC, n = 9) mice were exposed to SL-TBI, and thymus cellularity was measured on day 7. i, Female 1- to 2-month-old C57BL/6 WT (Ifngr^+/+, n = 10) or Ifngr1^−/− (n = 7) mice were exposed to SL-TBI, and thymus cellularity was measured on day 7. j, Female 1- to 2-month-old C57BL/6 WT (Prf^+/+, n = 10 and Gzmb^+/+, n = 5), Prf^−/− (n = 7) and Gzmb^−/− (n = 4) mice were exposed to SL-TBI, and thymus cellularity was measured on day 7. Graphs represent mean ± s.e.m.; each dot represents an individual biological replicate. Statistics were generated for a, e, f and h–j using unpaired two-tailed t tests, for b and d using paired two-tailed t tests, and for c using one-way ANOVA with Dunnet’s correction for multiple comparisons. Panel c created with BioRender.com.

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