Effect of Notch1 signal on hepatitis B virus X-mediated abnormal immune activity in renal tubular epithelial cells

Objective: To explore the expression of Notch1 receptor in renal tubular epithelial cells transfected with hepatitis B virus X (HBx) gene and its role in immunologic activity. Methods: The eukaryotic vector pcDNA3.1/myc-HBx containing HBx gene or vector pcDNA3.1/myc-Notch1 containing Notch1 gene was transiently transfected into HK-2 cells. And the shRNA technique was used to silence Notch1. HK-2 cells were divided into 7 groups, including normal culture, pcDNA3.1/myc, HBx, HBx + pcDNA3.1/myc, HBx + Notch1, HBx + shRNA and HBx + Notch1 shRNA Real-time PCR and Western blotting were used to detect the expression of Notch1. The expressions of MHC-II and CD40 were examined by flow cytometry. And the supernatant contents of IL-4 and IFN-γ were measured by enzyme-linked immunosorbent assay (ELISA). Results: The results of real-time PCR and Western blotting verified that HBx and Notch1 were successfully expressed in HK-2 cells after transfection. The transfection efficiency of shRNA was 70%. Compared with normal culture and pcDNA3.1/myc groups, the expression of Notch1 increased. The expressions of MHC-II and CD40 also significantly increased in the HBx + Notch1 group (9.69%±0.52% vs 4.90%±0.32%, 21.56%±0.71% vs 15.74%±0.20%, both P < 0.05). The supernatant level of IFN-γ was lower in HBx + Notch1 group (11.9±1.7 vs 18.8±0.8, P < 0.05) while the level of IL-4 was higher than control groups (50.2±0.6 vs 28.1±1.2, P < 0.05). And the HBx + Notch1 shRNA group had the opposite results. Conclusions: An over-expression of HBx gene may up-regulate the expression of Notch1. And Notch1 promotes the expression of immune molecules of renal tubular epithelial cell and regulates the secretion of cytokines so as to cause injury of cells and dysfunction of immunomicroenviroment.