Hepato-gastroenterology · 2013
Inhibition of hepatitis B virus production in vitro by proteasome inhibitor MG132.
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Wang Yi, Li Xiao-Liang, Yu Yong-Sheng, Tang Zheng-Hao, Zang Guo-Qing
Аннотация
BACKGROUND/AIMS: To investigate the suppressive effects of proteasome inhibitor MG132 on hepatitis B virus production.
METHODOLOGY: HepG2 2.2.15 hepatoblastoma cells, which constitutively produce HBV particles, were used in the present study. MTT assay was used to evaluate the cytotoxicity of MG132. A Proteasome-Glo chymotrypsin-like cell-based assay was used to access the proteasome activity. Quantitative PCR were performed to analyze HBV-DNA. Secreted HBV antigens in the culture medium were measured by ELISA. Western blot and immunofluorescent staining of HBV antigen were also performed.
RESULTS: After 6 days of MG132 treatment, proteasome activity was greatly decreased to 64.3 ± 7.8% and 36.4 ± 7.7% of untreated cells by 0.1μM and 0.3μM of MG132, respectively. The levels of HBsAg and HBeAg, and the copy number of extracellular HBV-DNA, were decreased to nearly half of the control group by 0.1μM MG132. The HBV replicative intermediates were also suppressed by MG132. Western blot and immunofluorescent staining clearly showed the lower levels of the expression of HBV proteins induced by MG132.
CONCLUSIONS: MG132 could effectively inhibit the HBV replication in vitro. Ubiquitin-proteasome pathway plays an important role in HBV life cycle and could be a promising therapeutic target for anti-HBV drugs.
METHODOLOGY: HepG2 2.2.15 hepatoblastoma cells, which constitutively produce HBV particles, were used in the present study. MTT assay was used to evaluate the cytotoxicity of MG132. A Proteasome-Glo chymotrypsin-like cell-based assay was used to access the proteasome activity. Quantitative PCR were performed to analyze HBV-DNA. Secreted HBV antigens in the culture medium were measured by ELISA. Western blot and immunofluorescent staining of HBV antigen were also performed.
RESULTS: After 6 days of MG132 treatment, proteasome activity was greatly decreased to 64.3 ± 7.8% and 36.4 ± 7.7% of untreated cells by 0.1μM and 0.3μM of MG132, respectively. The levels of HBsAg and HBeAg, and the copy number of extracellular HBV-DNA, were decreased to nearly half of the control group by 0.1μM MG132. The HBV replicative intermediates were also suppressed by MG132. Western blot and immunofluorescent staining clearly showed the lower levels of the expression of HBV proteins induced by MG132.
CONCLUSIONS: MG132 could effectively inhibit the HBV replication in vitro. Ubiquitin-proteasome pathway plays an important role in HBV life cycle and could be a promising therapeutic target for anti-HBV drugs.