Reversion of the anti-ER stress and anti-apoptotic effects of pioglitazone by a SCD-1 inhibitor.

RAW264.7 cells were pretreated with a SCD-1 inhibitor (1, 5, 10 nmol/L) for 1 hour and incubated with pioglitazone (10 μmol/L) for additional 6 hours. Then, cells were stimulated with palmitate (400 μmol/L) for 16 hours. Western blot analyses for phospho (P)-PERK (A), CHOP (B), Phospho(P)-JNK (C), cleaved caspase-3 (D) were performed. The bar graphs indicate the ratio of P-PERK to PERK, CHOP to α-tubulin, P-JNK to JNK, and cleaved caspase-3 to α-tubulin, respectively. *P<0.05 vs control (palmitate (-), pioglitazone (-) and SCD-1inhibitor (-)). #P<0.05 vs palmitate. †P<0.05 vs Palmitate+Pioglitazone. n = 4. (E) RAW264.7 cells were pretreated with pioglitazone (10 μmol/L) for 6 hours and stimulated with or without palmitate (400 μmol/L) for 16 hours. SCD-1 inhibitor was added 1 hour before pioglitazone treatment. The number of cells stained positively with Annexin V–FITC and negatively with PI was counted by flowcytometry. * P<0.05 vs control (palmitate (-), pioglitazone (-) and SCD-1inhibitor (-)), ##P<0.01 vs palmitate, ††P<0.01 vs Palmitate+Pioglitazone. n = 3. (F) RAW264.7 cells were pretreated with CAY10566, another SCD-1 inhibitor (1, 5, 10 nmol/L) for 1 hour and incubated with pioglitazone (10 μmol/L) for additional 6 hours. Then, cells were stimulated with palmitate (400 μmol/L) for 16 hours. Western blot analyses for phosphor (P)-PERK and PERK were performed. The bar graphs indicate the ratio of P-PERK to PERK. **P<0.01 vs control (palmitate (-), pioglitazone (-) and SCD-1inhibitor (-)). ##P<0.01 vs palmitate. ††P<0.01, †P<0.05 vs Palmitate+Pioglitazone. n = 4. RAW264.7 cells were treated with a SCD-1 inhibitor or CAY10566 (1, 5, 10 nmol/L) for 23 hours. Western blot analyses for phospho (P)-PERK (G), cleaved caspase-3 (H) were performed. The bar graphs indicate the ratio of P-PERK to PERK and cleaved caspase-3 to α-tubulin, respectively. *P<0.05 vs control. n = 4.