Fig. 8

p38 inhibition in FRG1 knockdown prostate cancer cells affects expression of various molecules: a. Representative Western blot panel showing reduction of phosphorylated p38 levels in the first lane after treatment with 0.5 μM SB203580 in DU145 FRG1KD cells (Si). Phosphorylated p38 levels were higher in second lane with 0.1 μM SB203580 and in third lane with untreated FRG1KD cells (Si). We could also observe lower p38 phosphorylation levels in DU145 pLKO.1sc (Sc) cells (lane 4) compared to FRG1KD (Si) (lane 3). Third row representing FRG1 panel shows knockdown of FRG1 in FRG1KD (Si) cells compared to pLKO.1 sc (Sc) cells. GAPDH levels were detected as loading control. b. Representative western blot panel showing reduction of phosphorylated p38 levels in the first lane after treatment with 0.5 μM SB203580 in PC3 FRG1KD cells (Si). Phosphorylation levels were higher in second lane with 0.1 μM SB203580 and in third lane with untreated FRG1KD cells (Si). We can also observe lower p38 phosphorylation levels in PC3 pLKO.1sc (Sc) cells compared to FRG1KD (Si). Third row representing FRG1 panel shows knockdown of FRG1 in FRG1KD (Si) cells compared to pLKO.1 sc (Sc) cells. GAPDH levels were detected as loading control. c. q-RT PCR based expression analysis of GM-CSF, MMP1 and PLGF in DU145 FRG1KD cells after 8 h of treatment with 0.5 μM SB203580 (p38 inhibitor). Fold change in FRG1KD and FRG1KD + SB (p38 inhibitor) groups was derived in comparison with DU145 pLKO.1sc. After 8 h of treatment significant reduction in expression was observed in GM-CSF (FRG1KD, FC = 1.94 vs. FRG1KD + SB, FC = 1.25) and PLGF (FRG1KD, FC = 2.59 vs. FRG1KD + SB, FC = 1.13) expression, no significant reduction was observed in case of MMP1 (FRG1KD, FC = 1.78 to FRG1KD + SB, FC = 1.49). d. q-RT PCR based expression analysis of GM-CSF, MMP1, PDGFA and CXCL1 in PC3 FRG1KD cells after 8 h of treatment with 0.5 μM SB203580 (p38 inhibitor). Fold change in FRG1KD and FRG1KD + SB groups was derived in comparison with PC3 pLKO.1sc. After 8 h of treatment significant reduction in GM-CSF (FRG1KD, FC = 4.26 vs. FRG1KD + SB, FC = 1.12), MMP1 (FRG1KD, FC = 4.69 vs. FRG1KD + SB, FC =1.17), PDGFA (FRG1KD, FC = 4.17 vs. FRG1KD + SB, FC = 1.21) and CXCL1 (FRG1KD, FC = 1.71 vs. FRG1KD + SB, FC = 1.05) expression was observed. In panel C and D, t-test (2 tailed, for unpaired samples) was used for comparison of fold change values between experimental and control group. Each experimental group had three replicates. # represents p value > 0.05, * represents p value < 0.05,, FC represents fold change