Role of Notchl receptor on apoptosis of renal tubular epithelial cells transfected with HBx gene

Objective: To explore the expression of Notchl receptor in renal tubular epithelial cells transfected with HBx gene and elucidate its role in cell apoptosis. Methods: The eukaryotic vector pcDNA3.1/myc-HBx containing HBx gene or vector pcDNA3.1/myc-Notch1 containing Notchl gene was transiently transfected into HK-2 cells and shRNA technique employed for silencing Notchl. HK-2 cells were divided into 7 groups of normal culture, pcDNA3.1/myc, HBx, HBx + pcDNA3.1/myc, HBx + Notch1 gene, HBx + shRNA and HBx + Notch1 shRNA. Real-time polymerase chain reaction (PCR) and Western blotting were used to confirm the expression of HBx and detect the expression of Notch1 receptor. Methyl thiazolyl tetrazolium (MTT) assay was used to observe the proliferation rate of HK-2 cells and flow cytometry to detect the apoptosis of HK-2 cells. Results: HBx and Notch1 receptor were successfully expressed in HK-2 cells post-transfection. The transfection efficiency of shRNA was 70%. The expression of Notch1 receptor in pcDNA3.1/myc-HBx group was higher than that in the normal culture and pcDNA3.1/myc groups. And the apoptotic ratio was also higher than that of normal culture and pcDNA3.1/myc group. The difference was significant (14.94% ±0.32% vs 13.86% ±0.18%, 13.67% ±0.54%, both P < 0.05). The apoptotic ratio in the HBx + Notch1 gene group was lower than that in control group (10.10% ±0.26% vs 14.79% ±0.02%, P <0.05). And the growth inhibition ratio of cells was also lower. But the apoptotic ratio and growth inhibition ratio were both higher in HBx + Notch1 shRNA group than those in HBx + shRNA group (both P < 0.05). Conclusions: HBx gene is successfully transfected into HK-2 cells. And its over-expression may induce apoptosis of HK-2 cells and growth inhibition by regulating Notchl signal.