Fig. 2

Lactate activates STAT3 in bladder cancer cells. (a) Quantitative PCR (qPCR) was performed to assess LCN2 mRNA levels in 5637 and MC-T2 cells treated with 10 mM and 20 mM for 24 h. Results are expressed as the ratio of LCN2 to GAPDH mRNA levels. Data represent the mean ± SEM from three independent experiments. Statistical significance was determined by an unpaired two-tailed t-test (*p < 0.05, **p < 0.01). (b) Exploring the potential mechanistic network between lactate, LCN2, and STAT3 in bladder tumors using Ingenuity Pathway Analysis (IPA). Gene Ontology (GO) analysis of the TCGA BLCA cohort (N = 411) was performed using the ShinyGO platform, comparing significantly differentially expressed genes between LCN2-high and LCN2-low expression groups (cutoff: median LCN2 expression). (d) A positive correlation of LCN2 and STAT3 mRNA expression in bladder tumors using cBioPortal website. (e) and (f) Western blot analysis of phosphorylated STAT3 (p-Y705 STAT3), total STAT3 and LCN2 expressions of 5637 cells under 10 mM and 20 mM lactic acid in (d), and 50 mM and 100 mM sodium lactate treatment in (e). (g) and (h) p-Y705 STAT3, STAT3 and LCN2 expression levels were examined in oxamate (50 and 100 mM) treatment and LDHA-silenced 5637 cells by western blot analysis.