Figure 5. Inhibition of system X_c⁻ is not required for HSPA5 expression or GPX4 degradation in ferroptosis.

(A) Indicated PDAC cells were treated with sulfasalazine (SAS) for six to 24 hours. The levels of GSH, GSSG, indicated protein, and GPX4 activity were assayed (n=3, *p < 0.05 versus untreated group). (B) Knockdown of SLC7A11 by shRNA did not affect erastin (20μM)-induced HSPA5 expression and GPX4 degradation at 24 hours in PANC1 cells (n=3, *p < 0.05). (C) SAS (400μM) inhibited erastin (20μM)-induced HSPA5 expression and GPX4 degradation at 24 hours in PANC1 cells (n=3, *p < 0.05). (D) Indicated PDAC cells were treated with erastin (20μM) with or without SAS (400μM) for three to 24 hours and cell death was assayed (n=3, *p < 0.05). (E) Indicated PDAC cells were treated with erastin (20 μM) /SAS (400 μM) with or without indicated inhibitors (ferrostatin-1, 1 μM; liprostatin-1, 1 μM; ZVAD-FMK,10 μM; necrosulfonamide, 0.5 μM) for 24 hours. Cell death was assayed using a CCK8 kit (n=3, *p < 0.05). (F, G) Indicated PANC1 cells were treated with erastin (20 μM) in combination with SAS (400 μM) for three to 24 hours. Cell death was assayed with a CCK8 kit (n=3, *p < 0.05).