Recombinant adenovirus-mediated shRNA silencing of midkine gene in BxPC-3 cells

Objective

To investigate the silencing effects of recombinant adenovirus Ad-shRNA-MK on midkine(MK) gene in pancreatic cancer cells.

Methods

Ad-shRNA-MK was used to infect pancreatic cancer BxPC-3 cells. Assays were conducted for knockdown of the MK gene on the day of infection and on the 1^st, 3^rd, 5^th, 7^th, and 9^th days post-infection by using immunocytochemistry, real-time RT-PCR, and Western blot analysis.

Results

The adenoviral Ad-shRNA-PTN was constructed successfully, and infection was confirmed by electron microscopic observation. By using real-time RT-PCR, the inhibition rates of MK mRNA expression in the BxPC-3 cells were 20%, 80%, 55%, and 23% on the 1^st, 3^rd, 5^th, and 7^th days post-infection. Immunocytochemistry and Western blot analysis confirmed this effect at the gene product level.

Conclusion

Efficient and specific knockdown of MK in pancreatic cancer cells by adenoviral Ad-shRNA-PTN is a potentially powerful tool for the study of gene therapy of pancreatic cancer nerve infiltration.