Fig. 1

Lactate regulates LCN2 expression in bladder cancer cells and its association with clinical outcomes. (a) The expression of LCN2 in normal bladder tissues (n = 28, GTEx-BLCA) and bladder cancer tissues (n = 404, TCGA-BLCA cohort), with a significance threshold of p < 0.05 (b) Representative images at magnification x100 were obtained and LCN2 protein expression in normal tissue and bladder tumor tissues were analyzed through the Human Protein Atlas database. Normal urinary bladder tissue (Male, age 55). Low grade urothelial carcinoma (Female, age 73; weak LCN2 staining). High grade urothelial carcinoma (Male, age 81; moderate LCN2 staining). (c) Correlation of LCN2 gene expression with clinical tumor staging of bladder cancer in GSE83586 cohort. Box plots represent LCN2 expression levels across different tumor stages: Ta (n = 19), T1 (n = 44), T2 (n = 241), and T3 (n = 2). Statistical significance and expression trends are indicated (p = 0.0334), highlighting potential associations between LCN2 expression and tumor progression. (d) Kaplan-Meier survival analysis of bladder cancer patients based on LCN2 gene expression using the GEPIA2 online application (http://gepia2.cancer-pku.cn/#index). The follow-up period for overall survival was over 150 months. The blue and red lines represent the survival curve of patients with low (n = 201) and high (n = 201) LCN2 expression, respectively, under a 95% confidence interval for each survival curve. p-value = 0.015 (e) Kaplan-Meier Plotter database (http://kmplot.com/analysis/) was used to assess the correlation between LCN2 expression and overall survival in patients with bladder cancer. The analysis showed that patients with high LCN2 expression (n = 204) had a significantly lower survival rate than those with low LCN2 expression (n = 200) (HR = 1.45, 95% CI, p = 0.015). With a follow-up period of 60 months, suggesting that higher LCN2 expression is associated with a reduced survival probability. (f) 5637 bladder cancer cells were treated with different lactic acid concentrations (10 and 20 mM) and sodium lactate (50 and 100 mM) for 24 h, respectively. Cell lysates were collected and subjected to western blot analysis using anti-LCN2 antibody. b-actin was used as a loading control. (g) The correlation between LCN2 and LDHA expression of the TCGA database were generated by cBioPortal website. (h) 5637 were treated with oxamate (50 and 100 mM for 24 h) to inhibit LDHA activity. Cell lysates were harvested and subjected to western blot analysis to assess LCN2 expression. (i) Cells were stably transduced with either scrambled shRNA control lentivirus or shRNA lentivirus targeting LDHA. Western blotting was performed to examine LDHA and LCN2 protein levels in 5637 cells.