Fig. 6: IL-35-triggered autocrine TGF-β drives NK cell dysfunction and conversion into ILC1-like cells.

a TGFB1 expression in total NK cells cultured for 2 and 4 days in IL-2 with or without IL-35 (analyzed from our scRNA-seq dataset). b Supernatants from NK cells cultured for 24 h in the presence of medium versus IL-12 and IL-18 with or without IL-35 were collected to quantify active TGF-β1 by ELISA. Mean values ± S.D are shown (n = 7 individual donors). c Representative images of NK cells after 7 days of culture in IL-2 with or without IL-35 and a TGF-βR inhibitor (Galunisertib). d Representative flow cytometry plots (upper) and quantification of IFN-γ, T-BET, and EOMES expression (lower) in NK cells at 24 h of culture in IL-12 + IL-18 with or without IL-35, TGF-βR inhibitor (Galunisertib) or anti-TGF-β1/2/3 neutralizing antibody. Results are expressed as relative MFI for each marker compared to control condition without IL-35. Mean values ± S.D are shown (n = 4 to 5 individual donors). e Representative flow cytometry plots (left) for IFN-γ expression in NK cells after 5 days in culture with IL-2 with or without IL-35 and TGF-βR inhibitor (Galunisertib) and quantification (right) of % IFN-γ⁺ cells in the same culture conditions. Mean values ± S.D are shown (n = 4 individual donors). f Representative flow cytometry plots (left) for CD9 and CD103 expression in NK cells after 8 days in culture with IL-2 with or without IL-35, TGF-βR inhibitor (Galunisertib) or anti-TGF-β1/2/3 neutralizing antibody and quantification (right) of % CD9⁺ CD103⁺ cells in the same culture conditions. Mean values ± S.D are shown (n = 3 to 6 individual donors). Statistical significance was determined using paired T test. Source data are provided as a Source Data file.