OBJECTIVE: To investigate whether transcription factor-kappaB (NF-κ B) is involved in the modulation of P-glycoprotein (P-gp) by glucosylceramide synthase (GCS) in a multidrug resistance leukemia cell line K562/A02 and to explore the relationship between NF-κ B and extracelluar signal-regulated kinase (ERK). METHODS: K562/A02 cells were treated with GCSsiRNA, pyrrolidine dithiocarbamate (PDTC, a NF-κ B specific inhibitor) and U0126 (a MEK1/2 inhibitor), respectively. The expression of GCS and multidrug resistance protein 1 (MDR1) mRNA were analyzed with qRT-PCR. Various proteins of different groups were measured by Western blotting. RESULTS: After transfected with GCSsiRNA for 48 h, GCS mRNA were reduced by 62% (51%-73%) and MDR1 mRNA was reduced by 52% (43%-61%) in the K562/A02 cells. Compared with the negative control, relative expression of NF-κ B p65 in nuclear and P-ERK1/2 were both down-regulated, and P-gp was also inhibited significantly at 72 h after transfected with GCSsiRNA (P< 0.05). In addition, the expression of P-gp was decreased at 24 h with 80 μ mol/L PDTC and 48 h with 20 μ mol/L PDTC. P-ERK1/2 was inhibited significantly when the cells were treated with 20 μ mol/L U0126 for 48 h. The expression of NF-κ B p65 in nuclear and P-gp were also down-regulated. CONCLUSION: NF-κ B can modulate the effect of GCS on P-gp in K562/A02 cells. P-ERK1/2 can activate NF-κ B in above signal transduction pathway.
OBJECTIVE: To investigate whether transcription factor-kappaB (NF-κ B) is involved in the modulation of P-glycoprotein (P-gp) by glucosylceramide synthase (GCS) in a multidrug resistance leukemia cell line K562/A02 and to explore the relationship between NF-κ B and extracelluar signal-regulated kinase (ERK). METHODS: K562/A02 cells were treated with GCSsiRNA, pyrrolidine dithiocarbamate (PDTC, a NF-κ B specific inhibitor) and U0126 (a MEK1/2 inhibitor), respectively. The expression of GCS and multidrug resistance protein 1 (MDR1) mRNA were analyzed with qRT-PCR. Various proteins of different groups were measured by Western blotting. RESULTS: After transfected with GCSsiRNA for 48 h, GCS mRNA were reduced by 62% (51%-73%) and MDR1 mRNA was reduced by 52% (43%-61%) in the K562/A02 cells. Compared with the negative control, relative expression of NF-κ B p65 in nuclear and P-ERK1/2 were both down-regulated, and P-gp was also inhibited significantly at 72 h after transfected with GCSsiRNA (P< 0.05). In addition, the expression of P-gp was decreased at 24 h with 80 μ mol/L PDTC and 48 h with 20 μ mol/L PDTC. P-ERK1/2 was inhibited significantly when the cells were treated with 20 μ mol/L U0126 for 48 h. The expression of NF-κ B p65 in nuclear and P-gp were also down-regulated. CONCLUSION: NF-κ B can modulate the effect of GCS on P-gp in K562/A02 cells. P-ERK1/2 can activate NF-κ B in above signal transduction pathway.