Object As B7-H1 plays a critical role in immune escape of tumor cells, we studied the effect of etoposide on the expression of B7-H1 inhuman retinoblastoma cells and its related mechinism.
Methods The effect of etoposide on B7-H1 expression was determined by the reverse-transcription polymerase chain re-action (RT-PCR), real-time PCR and flow cytometry analysis in Y79 cells. Then the involvement of Mitogen-activated protein kinases (MAPKs) signal pathways were tested by Western blotting and signal transduction inhibitor assays. Furthmore, specific small interfering RNA (siRNA) targeting B7-H1 was transfected into Y79 retinoblastoma cells using liposome. Silencing of B7-H1 expression was measured by RT-PCR and Western blotting assays.
Results Etoposide increased B7-H1 mRNA and protein levels in Y79 cells. The effect of etoposide on B7-H1 expression peaked at the concentration of 5μg / ml , as confirmed by RT-PCR, real-time PCR and flow cytometry assays (P<0.01). The phosphorylation status of extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) were constitutively activated by etoposide, and MEK inhibitor simultaneously reduced the expression of B7-H1 induced by etoposide (P<0.01). B7-H1 siRNA significantly silenced B7-H1 expression in Y79 cells, as confirmed by RT-PCR and Western blotting assays (P<0.01).
Conclusion Our results demonstrated that etoposide promoted B7-H1 expression via activating phosphorylations of the ERK signal pathway in human retinoblastoma cells. These findings might open up the prospective of immunotherapy as an important treatment for retinoblastoma. |
