Establishment and primary application of a mouse model with hepatitis B virus replication

Abstract

Aim: To establish a rapid and convenient animal model with hepatitis B virus (HBV) replication.

Methods: A naked DNA solution of HBV-replication-competent plasmid was transferred to BALB/C mice via the tail vein, using a hydrodynamic in vivo transfection procedure. After injection, these mice were sacrificed on d 1, 3, 4, 5, 7 and 10. HBV DNA replication intermediates in the liver were analyzed by Southern blot hybridization. The expression of hepatitis B core antigen (HBcAg) and hepatitis B surface antigen (HBsAg) in the liver was checked by immunohistochemistry. Serum HBsAg and hepatitis B e antigen (HBeAg) was detected by enzyme-linked immunosorbent assay (ELISA). Inhibition of HBV replication was compared in HBV replication model mice treated intraperitoneally with polyinosinic-polytidylin acid (polyIC) or phosphate-buffered saline (PBS).

Results: After hydrodynamic in vivo transfection, HBV DNA replication intermediates in the mouse liver were detectable on d 1 and abundant on d 3 and 4, the levels were slightly decreased and remained relatively stable between d 5 and 7, and were almost undetectable on d 10. The expression patterns of HBcAg and HBsAg were similar to that of HBV replication intermediate DNA, except that they reached a peak on d 1 after injection. No obvious differences in HBV DNA replication intermediates were observed in the left, right and middle lobes of the liver. After treatment with polyIC, the level of HBV intermediate DNA in the liver was lower than that in the control mice injected with PBS.

Conclusion: A rapid and convenient mouse model with a high level of HBV replication was developed and used to investigate the inhibitory effect of polyIC on HBV replication, which provides a useful tool for future functional studies of the HBV genome.

Figure 1

DNA-dose-dependent HBV replication. Various amount of pHBV4.1 in 1.8 mL saline were injected into BALB/c mice within 5-8 s. Mice were sacrificed on d 4 after injection, and HBV DNA intermediates were detected by Southern blotting.

Figure 2

Time-dependent viral replication after hydrodynamic in vivo transfection with pHBV4.1. Mice were injected hydrodynamically with 10 μg pHBV4.1 and were sacrificed at different time points after injection. HBV DNA intermediates were detected by Southern blotting.

Figure 3

Detection of HBcAg expression in the liver by immunohistochemical staining (DAB, × 40). Mice were injected hydrodynamically with 10 μg pHBV4.1 and were sacrificed at different time points after injection and HBcAg in mouse liver was detected. A: 1 d after transfection; B: 4 d after transfection; C: 7 d after transfection; D: 10 d after transfection.

Figure 4

Detection of HBsAg expression in liver by immunohistochemical staining (DAB, × 40). Mice were injected hydrodynamically with 10 μg pHBV4.1 and were sacrificed at different time points after injection and HBsAg in mouse liver was detected. A: 1 d after transfection; B: 4 d after transfection; C: 7 d after transfection; D: 10 d after transfection.

Figure 5

Time-dependent HBV antigen expression in serum after hydrodynamic transfection. Mice were injected with 10 μg pHBV4.1. At different time points, HBsAg and HBeAg in the serum were measured by ELISA.

Figure 6

Viral replication in different parts of the mouse liver after hydrodynamic transfection. The results from two mice (A and B) are shown. L: left lobe; R: right lobe; M, middle lobe.

Figure 7

Effect of polyIC on HBV replication in mice. After matched by body weight, age and serum HBeAg, mice were treated with PBS or polyIC. HBV replication levels in the liver of the two groups of mice were compared.