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Updated: Apr 30, 2026
"Liver-on-a-Chip" Cultures of Primary Hepatocytes and Kupffer Cells for Hepatitis B Virus Infection
Published on: February 19, 2019
[Hepatitis B virus X protein upregulates the expression of CD59 and Crry in mouse podocytes]
1Department of Pediatrics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Objective:
Different from primary membranous nephropathy, hepatitis B virus associated membranous nephropathy (HBV-MN) shows lower deposits of membrane attack complex (C5b-9) in glomerular subepithelium. The causes of relatively low complement activation in this disease remain unclear. The aim of this study was to investigate the influence of hepatitis B x protein (HBx) on the expression of CD59 and Crry in mouse podocytes.
Method:
Cultured mouse podocytes were divided into adenovirus vector hepatitis B virus X gene (Ad-HBx) transfected group (Ad-X group), blank podocytes group (B group) and adenovirus vector transfected group (Ad group). CD59 and Crry mRNA expression were assayed by semiquantitative RT-PCR. CD59 and Crry expression were tested by flow cytometry. The effect of HBx on complement activation was evaluated with MTT method. And then, the effects of P38MAPK, PI-3K and ERK1/2 pathway inhibitors (SB203580, LY294002, U0126) and DMSO on CD59 and Crry expression were respectively detected by flow cytometry.
Result:
Proteins CD59 and Crry expression rates (%) in group B, Ad group and Ad-X group were 17.71 ± 3.81, 18.29 ± 3.36 and 45.7 ± 9.01; 18 ± 2.31, 21.78 ± 2.01 and 47.45 ± 9.95, respectively. Compared with group B, CD59 and Crry expression in group Ad was not significantly different (P values for both > 0.05), but CD59 and Crry protein expression in Ad-X group was significantly higher than that in groups B and Ad (P values for both < 0.005); CD59 and Crry gene expression in group Ad was not significantly different from that in group B (P values for both > 0.05). However, CD59 and Crry gene expression of Ad-X group was significantly higher than that in groups B and Ad (P values for both < 0.05). Flow cytometry detected CD59 protein expression rates (%) were 17.35 ± 1.24, 46.19 ± 9.77, 43.03 ± 6.83 and 40.04 ± 6.39 and Crry protein expression rates (%) were 18.14 ± 3.56, 31.95 ± 1.68, 31.95 ± 1.69 and 37.14 ± 3.92 after SB203580, LY294002, U0126 and DMSO were added to Ad-X group respectively. P38 pathway inhibition resulted in significantly lower CD59 and Crry expression than Ad-X group (P values for all < 0.005), but PI-3K, ERK1/2 pathway inhibitors and DMSO had no significant effect on the expression of CD59 and Crry (P values for all > 0.05). The inhibition rates of cell lysis were significantly higher in Ad-X group than in groups B and Ad at each serum dilution point (P values for all < 0.05), while groups B and Ad had no significant difference in cell viability.
Conclusion:
HBx can up-regulate CD59 and Crry expression in podocytes through activating P38 pathway, resulting in decreased complement activation, which may facilitate latent HBV infection in podocytes and play a role in development of hepatitis B virus associated glomerulonephritis (HBV-GN).
Insights
Hepatitis B virus X protein (HBx) increases CD59 and Crry expression in podocytes via the P38 pathway. This reduces complement activation, potentially aiding latent HBV infection and hepatitis B virus-associated glomerulonephritis (HBV-GN) development.
Area of Science:
- Nephrology
- Virology
- Immunology
Background:
- Hepatitis B virus-associated membranous nephropathy (HBV-MN) exhibits lower complement C5b-9 deposits than primary MN.
- The mechanisms behind reduced complement activation in HBV-MN are not fully understood.
Purpose of the Study:
- To investigate the effect of hepatitis B virus X protein (HBx) on CD59 and Crry expression in mouse podocytes.
- To explore the role of HBx in complement regulation within podocytes.
Main Methods:
- Mouse podocytes were transfected with Ad-HBx.
- CD59 and Crry mRNA and protein expression were analyzed using RT-PCR and flow cytometry.
- Complement activation was assessed via MTT assay.
- The involvement of P38MAPK, PI-3K, and ERK1/2 pathways was examined using specific inhibitors.
Main Results:
- HBx significantly upregulated both CD59 and Crry mRNA and protein expression in podocytes.
- Inhibition of the P38 pathway, but not PI-3K or ERK1/2, reversed the HBx-induced increase in CD59 and Crry.
- HBx transfection led to significantly higher inhibition rates of cell lysis, indicating reduced complement activation.
Conclusions:
- HBx upregulates CD59 and Crry expression in podocytes by activating the P38 pathway.
- This upregulation leads to decreased complement activation.
- These findings suggest a role for HBx in facilitating latent HBV infection and the pathogenesis of HBV-GN.
