Fig. 5: TNF protects RA fibroblasts from ferroptosis while IL-6 and TGF-β sensitize fibroblasts to ferroptosis.

a, b Relative viability of fibroblasts derived from RA patients and primed with TNF, IL-6, or TGF-β (20 ng/ml) for 72 h, followed by treatment with different concentrations of IKE or RSL3. Cell viability was assayed by measuring cellular ATP levels at 26 h (IKE) (a) or 12 h (RSL3) (b) after treatment. n = 3 biologically independent samples per condition. IC₅₀ values were calculated using nonlinear regression analysis. c, d Cells were treated as indicated for 18 h (IKE, 1 μM) (c) or 4 h (RSL3, 0.125 μM) (d) in the presence of the ferroptosis inhibitor Ferrostatin-1 (Fer1) (1 μM) and lipid ROS accumulation was measured by BODIPY C11 staining coupled with flow cytometry. n = 2 biologically independent samples per condition. ns, P > 0.9999, ****P < 0.0001, ***P = 0.0007; one-way ANOVA followed by multiple comparisons. e, f The relative viability of fibroblasts primed with TNF in the presence of the anti-TNF blocking antibody adalimumab (Ada) for 72 h, followed by treatment with IKE (e) or RSL3 (f). n = 3 biologically independent samples per condition. *P = 0.0183, 0.0216 (left to right), ***P = 0.0003, 0.0002 (left to right); one-way ANOVA followed by multiple comparisons. g Relative viability of fibroblasts primed with IL-6 in the presence of the anti-IL-6 receptor monoclonal antibody tocilizumab (Toci), followed by treatment with IKE or RSL3. n = 3 biologically independent samples per condition. Left, **P = 0.0011, ***P = 0.0008; right, **P = 0.0011, *P = 0.0182; one-way ANOVA. h Relative viability of fibroblasts primed with TGF-β in the presence of the ALK receptor inhibitor SB431542 followed by treatment with IKE or RSL3. n = 3 biologically independent samples per condition. Left, *P = 0.0131, **P = 0.0042; right, **P = 0.0067, **P = 0.0040; one-way ANOVA followed by multiple comparisons. All bar graphs are represented as mean ± SD. Source data are provided as a Source data file.