TGF-β affects the differentiation of human GM-CSF⁺ CD4⁺ T cells in an activation- and sodium-dependent manner

The cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF) is involved in the pathogenesis of chronic inflammatory diseases such as multiple sclerosis. However, the environmental cues promoting differentiation of GM-CSF producing T cells are unclear. Herein, we performed a broad experimental screening of cytokines and data-driven analysis assessing their ability to induce human GM-CSF⁺ CD4⁺ T cells and their subpopulations. TGF-β was discovered to induce GM-CSF production independently of proliferation and IL-2 signaling including STAT5. In contrast, IL-6 and IL-23 decreased GM-CSF production. On the population level, GM-CSF induction was highly correlated with expression of FOXP3 across cytokine stimulations but not with that of IL-17. However, on single-cell level GM-CSF and IFN-γ expression were most correlated, independently of the cytokine environment. Importantly, under low sodium conditions in the medium or upon stimulation with plate-bound instead of bead-bound anti-CD3 and anti-CD28 antibodies,the effects of TGF-β on GM-CSF, but not on FOXP3, were reversed. Our analysis indicates a novel role for TGF-β in generating GM-CSF⁺ subsets of human CD4⁺ T cells. These results are important for understanding of autoimmune disease and therapeutic considerations.