Objective: To investigate the expression status of Mitogen-activated Protein Kinase Phosphatase-1 in oxygen-deprived gastric cancer cell line SGC7901 and its role in HIF-1 regulation.
Methods: The expression of MKP-1 in gastric cell line SGC7901 was detected with Western blot and semiquantity RT-PCR; Eukaryotic sense expression vector was constructed based on DNA recombination technology. Transfections of SGC7901 were performed using liposome; The luciferase activity was determined using Dual Luciferase Reporter System and the levels of VEGF in SGC7901cells under normoxia and hypoxia were measured by ELISA.
Results: (1) Semiquantity RT-PCR and Western blot suggested that the expression of MKP-1 was elevated in oxygen-deprived gastric cancer cell line SGC7901; (2) 48 hours after transfection, the phosphorylated form of HIF-1alpha in cell line transfected with recombinant plasmids was lower compared with that in cell line transfected with empty vectors after 12 hours of exposure to hypoxia; (3) There was very low luciferase activity under nomoxia while under hypoxia luciferase activity increased in a time-dependent manner and at all time points there was significant lower luciferase activity in SGC7901 cells than in cells transfected with empty plasmids. (4) At different points of time course, the expression of VEGF in SGC7901 was significantly higher under hypoxia than that in SGC7901 under nomorxia, while under hypoxia, the expression of VEGF in SGC7901 transfected with recombinant plasmids was significantly lower than that in SGC7901 transfected with empty vectors at all time points as indicated.
Conclusion: The expression of MKP-1 in SGC7901 was elevated under hypoxia, which could downregulate the HIF-1 trans-activition activity thereby repressing the expression of downstream target gene VEGF.