Fig 2: FLRT2 overexpression promotes ferroptosis of human bladder cancer cells. (A) KEGG analysis of the top five pathways altered in FLRT2‐silenced T24 cells (n = 3 or 6). (B) LDH releases were measured to evaluate cell death (n = 6). (C, D) ROS generation and lipid peroxidation in FLRT2‐overexpressed or CTRL human bladder cancer cells (n = 6). (E, F) Levels of 12/15‐HETEs in cell medium (n = 6). (G) Levels of ferrous iron (n = 6). (H, I) Levels of GSH and GPX4 activity in FLRT2‐overexpressed or CTRL human bladder cancer cells (n = 6). All data are presented as the mean ± SD, p < 0.05 was considered statistically significant. NS indicates no significance.

Fig 3: FLRT2 triggers ferroptosis through elevating ACSL4. (A) Heatmaps of the differentially expressed genes (n = 3). (B, C) ACSL4 protein levels in T24 or UMUC‐3 cells with or without FLRT2 silence (n = 6). (D) Spearman correlation analysis was used for the analysis between FLRT2 and ACSL4. (E) ACSL4 protein levels in T24 or UMUC‐3 cells with or without FLRT2 overexpression (n = 6). (F) ACSL4 mRNA level in T24 or UMUC‐3 cells with or without ACSL4 silence (n = 6). (G, H) ROS generation and lipid peroxidation in FLRT2‐overexpressed or CTRL human bladder cancer cells with or without ACSL4 silence (n = 6). (I) Levels of 12/15‐HETEs in cell medium (n = 6). (J) LDH releases were measured to evaluate cell death (n = 6). All data are presented as the mean ± SD, P < 0.05 was considered statistically significant.

Fig 4: FLRT2 overexpression inhibits the malignant phenotypes of human bladder cancer cells. (A) FLRT2 protein levels in T24 or UMUC‐3 cells with or without FLRT2 overexpression (n = 6). (B) CCK‐8 assay was performed to detect cell survival in FLRT2‐overexpressed or CTRL human bladder cancer cells (n = 6). (C) LDH releases were measured to evaluate cell death (n = 6). (D, E) Cell migration and invasion in FLRT2‐overexpressed or CTRL human bladder cancer cells (n = 6). All data are presented as the mean ± SD, p < 0.05 was considered statistically significant.

Fig 5: FLRT2 silence facilitates the malignant phenotypes of human bladder cancer cells. (A) FLRT2 protein levels in T24 or UMUC‐3 cells with or without FLRT2 silence (n = 6). (B) CCK‐8 assay was performed to detect cell survival in FLRT2‐silenced or CTRL human bladder cancer cells (n = 6). (C) LDH releases were measured to evaluate cell death (n = 6). (D, E) Cell migration and invasion in FLRT2‐silenced or CTRL human bladder cancer cells (n = 6). All data are presented as the mean ± SD, p < 0.05 was considered statistically significant.