Background & objective: Ku80 is a key protein plays a role in repairing DNA double strand break (DSB) after irradiation. There are a few studies about other roles of Ku80 except DSB repair. This study was to inhibit Ku80 expression in cervical carcinoma cell line HeLa with small interfering RNA (siRNA), and explore its effect on cell proliferation.
Methods: Plasmids pKu80-siRNA and pNeg-siRNA (negative control) were constructed and transfected into HeLa cells. The expression of Ku80 in HeLa cells was detected by Western blot. The proliferation of HeLa cells in vitro and in vivo was determined by clone formation assay, MTT assay, and subcutaneous tumor formation in nude mice.
Results: Two cell clones were screened from pKu80-siRNA-and pNeg-siRNA-transfected HeLa cells. Ku80 expression in HeLa cells was suppressed markedly after transfection of pKu80-siRNA; this clone was named Hela/Ku80-siRNA. The clone formation efficiency was significantly lower in HeLa/Ku80-siRNA cells than in control cells (0.46+/-0.05 vs. 0.62+/-0.02, t=5.11, P<0.01). The proliferation rate was significantly lower in HeLa/Ku80-siRNA cells than in control cells at 48 h and 72 h after transfection (P<0.05). At the 25th day after subcutaneous transplantation in nude mice, the tumor volume was significantly smaller in HeLa/Ku80-siRNA group than in control group [(18.92+/-3.60) mm(3) vs. (194.88+/-30.61) mm(3), t=12.69, P<0.01].
Conclusions: We successfully established a cell model that Ku80 expression is suppressed almost completely by siRNA. Ku80 inhibition inhibits the proliferation of HeLa cells in vivo and in vitro.