Fig. 3. LCN2 acts as an effector gene of NUPR1 in blocking ferroptosis.

a Heatmap of relative mRNA levels of iron metabolism-associated genes in Nupr1^+/+ and Nupr1^−/− mPDAC cells following treatment with erastin (10 µM) or RSL3 (1 µM) for 24 h. b, c Analysis of LCN2 protein expression and Lcn2 promoter activity in Nupr1^+/+ and Nupr1^−/− mPDAC cells following treatment with erastin (10 µM) or RSL3 (1 µM) for 24 h (n = 3 well/group, one-tailed t test). d Binding of NUPR1 to Lcn2 promoter was analyzed using ChIP-qPCR in indicated mPDAC cells following treatment with erastin (10 µM) or RSL3 (1 µM) for 24 h (n = 3 well/group, one-tailed t test). e qPCR analysis of Lcn2 mRNA in indicated mPDAC cells following treatment with erastin (10 µM) or RSL3 (1 µM) for 24 h (n = 3 well/group, one-tailed t test). f Fe²⁺ levels in indicated mPDAC cells following treatment with erastin or RSL3 for 24 h (n = 3 well/group, two-way ANOVA with Tukey’s multiple comparisons test on all pairwise combinations). g–j Indicated mPDAC cells were treated with erastin (10 µM) or RSL3 (1 µM) in the absence or presence of DFO (100 µM) or liproxstatin-1 (1 µM) for 24 h, and then intracellular MDA (g), intracellular 8-OHdG (h), extracellular HMGB1 (i), and cell viability (j) were quantified (n = 3 well/group, two-way ANOVA with Tukey’s multiple comparisons test on all pairwise combinations). Data in b–j are presented as mean ± SD. The results in a–j are representative of those from 2 to 3 independent experiments with three technical replicates each.