Abstract

Objective: To investigate the effect of stable transfection with NAD+-linked 15-hydroxyprostaglandin dehydrogenase (15-PGDH) on the growth, proliferation and migration of gastric cancer cells.
Methods: The levels of 15-PGDH in four kinds of gastric cancer cells with different differentiation rates were compared by RT-PCR measurements, and the poorly differentiated gastric cancer cells SGC7901 were chose to perform follow-up experiments. SGC7901 cells were transfected with recombinant plasmid pcDNA3/15-PGDH and empty plasmid pcDNA3 as a control by using lipofectamine 2000. Cells with steady expression capability were sorted out by G418. RT-PCR and Western blot were used to confirm the transfection and expression of 15-PGDH in SGC7901 cells. MTT, cell scratch assay, soft-agar colony formation assays were used to determine the proliferation, migration and cell clone formation activities of SGC7901 with stable transfection of 15-PGDH. Flow cytometry assay was used to examine the effect of 15-PGDH on cell cycle and apoptosis. RT-PCR was used to analyze the levels of cell cycle-related genes (p53, p21 and p16) and apoptosis-related genes (Survivin, BCL-2 and Caspase3).
Results: Both recombinant plasmid pcDNA3/15-PGDH and empty vector plasmid pcDNA3 were successfully transfected into human gastric cancer SGC7901 cell line. Proliferation, migration and cell clone formation capabilities of pcDNA3/15-PGDH groups were significantly inhibited compared with other two control groups (P<0.05). Flow cytometry results demonstrated an increased fraction of sub-G1 phase and the increase of apoptotic cells for pcDNA3/15-PGDH groups compared with other two control groups (P<0.05). The levels of p16, p21 and p53 mRNA for pcDNA3/15-PGDH groups were higher than those in other two control groups (p<0.05). pcDNA3/15-PGDH groups exhibited a higher level of caspase3 mRNA but lower levels of survivin and BCL-2 mRNA in SGC7901 cells compared with other two control groups (P< 0.05).
Conclusion: The gene transfection of 15-PGDH has negative effects on the growth, proliferation and migration of gastric cancer SGC7901 cells by inducing apoptosis and cell cycle arrest.