关键词: 【关键词】 乙型肝炎病毒x基因, 人近端肾小管上皮细胞, 转染, Toll样受体4 , 凋亡

Abstract:

[Abstract] Objective: To investigate the effect of transfection of hepatitis B virus x (HBx) gene on apoptosis of cells and the secretion of cytokines. Methodology: Human proximal renal tubular epithelial cell line HK-2 was cultured in K-SFM medium. The eukaroytic vector pcDNA3.1/myc-HBx containing HBx gene was transiently transfected into HK-2 cells by lipofectamine mediation. The expression of HBx was confirmed by Real-time PCR (RT-PCR) and Western-blotting assay. Untransfected HK-2 cells and those trasfected with empty vector were used as control, the morphology of HK-2 cells were observed by microscopy and the level of TLR4 was measured by RT-PCR and Western-blotting assay, and the apoptosis was detected by flow cytometry, and the contents of IL-4 and IFN-γ in the supernatant were detected by ELISA. Results: RT-PCR and Western blotting results verified that HBx was expressed in HK-2 cells after transfection. After transfection of HBx gene, the shape of HK-2 cells became irregular. Compared with the control groups, the level of TLR4 was upregulated (P<0.05) and the number of apoptotic cells was significantly increased in the HBx transfection group (P<0.05). The level of IFN-γ in the supernatant of HBx transfection group was higher, but IL-4 level was lower than that of control groups. Conclusions HBx gene is successfully transfected into HK-2 cells. Overexpression of HBx gene may induce apoptosis of HK-2 cells by regulating TLR4. HBx can also induce imbalance of cytokines.

Key words: Key words  , HBx  , renal tubular epithelial cells   , transfection   , TLR4 apoptosis