Study On Transdifferentiation Effect And Mechanism Of HBV On Human Renal Tubular Epithelial Cells

Objective:To observe the effect of PHY106-HBV recombinant plasmid（C genotype HBV forthe whole genome）on the human proximal tubular epithelial cell line(HK-2)in vitro, andits influence on cell transdifferentiation.To investigate the role of the TGF-β1/p38MAPKsignaling pathway in HBV-induced epithelial-mesenchymal transition of HK-2cells. Thepurpose of the study can explore the mechanisms of renal injury in patient with hepatitis Bassociated glomerulonephritis(HBV-GN).Methods:HK-2cells were cultured in vitro and we devided them into the HK-2group, theHK-2-PHY106group (HK-2cells transfected with PHY106control plasmid), theHK-2-PHY106-HBV group (HK-2cells transfected with PHY106-HBV plasmid), andSB203580group (HK-2cells pretreated with a chemical inhibitor of p38MAPK beforetransfected with PHY106-HBV plasmid). HK-2cells were transfected bylipofectamineTM2000. Supernatant of culture was collected for examining HBsAg andHBeAg by using electrochemiluminescence (ECLIA) method. The expression ofE-cadherin and α-smooth muscle actin （α-SMA） in HK-2cells were assayed byimmunocytochemistry after the cells were transfected for72h. TGF-β1mRNA wasdetected by RT-PCR after the cells were transfected for72h. The expression ofE-cadherin、α-SMA、phosphorylated p38MAPK and p38MAPK in HK-2cells wereassayed by Western blot after the cells were transfected for72h. Results:The HBsAg and HBeAg were detected with a higher expression in theHK-2-PHY106-HBV group. The E-cadherin detected by immunocytochemistry andWestern blot showed a significantly lower expression in the HK-2-PHY106-HBV group（P＜0.05）. However,the expression of α-SMA in the HK-2-PHY106-HBV group washigher than the other three groups（P＜0.05）. Additionally, we also found TGF-β1mRNA expression was upregulated in the HK-2-PHY106-HBV group showed by RT-PCR（P＜0.05）. The expression of p-P38MAPK in the HK-2-PHY106-HBV group detectedby Western blot was higher than the other three groups（P＜0.05）,and it decreased （P＜0.05）when dealt with SB202380(the specific chemistry inhibitor of P38MAPK) beforetransfection.Also,the decrease of the expression of p-P38MAPK can prevent theupregulation of α-SMA and the downregulation of E-cadherin.Conclusion:HBV can be replicated highly efficiently in HK-2cells and can make HK-2cells occurtransdifferentiation.The mechanism may be by activating TGF-β1/P38MAPK signalingpathway. SB202380(the specific chemistry inhibitor of P38MAPK) can prevent thetransdifferentiation induced by HBV on HK-2cells.