AIM: To investigate the frequency of mutations in pre-core (pre-C) and basic core promoter (BCP) regions of hepatitis B virus (HBV) from Shanxi Province, and the association between mutations and disease related index...AIM: To investigate the frequency of mutations in pre-core (pre-C) and basic core promoter (BCP) regions of hepatitis B virus (HBV) from Shanxi Province, and the association between mutations and disease related indexes.METHODS: One hundred chronic hepatitis B patients treated at Shanxi Province Hospital of Traditional Chinese Medicine were included in this study. PCR-reverse dot blot hybridization and mismatch amplification mutation assay (MAMA)-PCR were used to detect the mutations in the HBV pre-C and BCP regions. HBV DNA content and liver function were compared between patients with mutant HBV pre-C and BCP loci and those with wild-type loci. The consistency between PCR-reverse dot blot hybridization and MAMA-PCR for detecting mutations in the HBV pre-C and BCP regions was assessed.RESULTS: Of the 100 serum samples detected, 9.38% had single mutations in the pre-C region, 29.17% had single mutations in the BCP region, 41.67% had mutations in both BCP and pre-C regions, and 19.79% had wild-type loci. The rates of BCP and pre-C mutations were 65.7% and 34.3%, respectively, in hepatitis B e antigen (HBeAg) positive patients, and 84.6% and 96.2%, respectively, in HBeAg negative patients. The rate of pre-C mutations was significantly higher in HBeAg negative patients than in HBeAg positive patients (χ<sup>2</sup> = 26.62, P = 0.00), but there was no significant difference in the distribution of mutations in the BCP region between HBeAg positive and negative patients (χ<sup>2</sup> = 2.43, P = 0.12). The presence of mutations in the pre-C (Wilcoxon W = 1802.5, P = 0.00) and BCP regions (Wilcoxon W = 2906.5, P = 0.00) was more common in patients with low HBV DNA content. Both AST and GGT were significantly higher in patients with mutant pre-C and BCP loci than in those with wild-type loci (P < 0.05). PCR-reverse dot blot hybridization and MAMA-PCR for detection of mutations in the BCP and pre-C regions had good consistency, and the Kappa values obtained were 0.91 and 0.58, respectively.CONCLUSION: HBeAg negative patients tend to have HBV pre-C mutations. However, these mutations do not cause increased DNA copies, but associate with damage of liver function.展开更多
Objectives: To investigate the mutation of the basic core promoter (BCP) of hepatitis B virus (HBV) and clarify the significance of HBV quasispecies groups in patients with chronic HBV infection. Methods: A set of spe...Objectives: To investigate the mutation of the basic core promoter (BCP) of hepatitis B virus (HBV) and clarify the significance of HBV quasispecies groups in patients with chronic HBV infection. Methods: A set of specific primers was synthesized according to the HBV DNA sequence of a Chinese strain. The BCP was amplified by PCR method from the serum of 40 patients with chronic HBV infection, and the PCR products of 2 patients were subcloned into pGEM Teasy vectors. Polyacrylamide gel elec- trophoresis (PAGE) was employed to display the de- letion mutations, and clones with differential length were selected to be sequenced. Sequence comparison was made to find the difference. Results: Two or three bands were displayed by PAGE in 60% patients. The results of sequence anal- ysis showed that there are some kinds of mutations in the BCP region. The substitution always occurs in TATA-like boxes, especially from T to C on 140 site. The deletion mutations were detected in TA1, TA2 and TA3. The 8bp, 20bp deletion mutations fre- quently happened. Conclusions: There is a hot deletion region in the BCP. The deletion and the substitution in the TATA- like box may influence the expression of preC/C pro- tein. The sequencing results indicate that there are HBV quasispecies groups in patients with chronic HBV infection.展开更多
Chronic hepatitis B virus(HBV)infection can lead to advanced liver pathology.Here,we establish a transgenic murine model expressing a basic core promoter(BCP)-mutated HBV genome.Unlike previous studies on the wild-typ...Chronic hepatitis B virus(HBV)infection can lead to advanced liver pathology.Here,we establish a transgenic murine model expressing a basic core promoter(BCP)-mutated HBV genome.Unlike previous studies on the wild-type virus,the BCP-mutated HBV transgenic mice manifest chronic liver injury that culminates in cirrhosis and tumor development with age.Notably,agonistic anti-Fas treatment induces fulminant hepatitis in these mice even at a negligible dose.As the BCP mutant exhibits a striking increase in HBV core protein(HBc)expression,we posit that HBc is actively involved in hepatocellular injury.Accordingly,HBc interferes with Fis1-stimulated mitochondrial recruitment of Tre-2/Bub2/Cdc16 domain family member 15(TBC1D15).HBc may also inhibit multiple Rab GTPase-activating proteins,including Rab7-specific TBC1D15 and TBC1D5,by binding to their conserved catalytic domain.In cells under mitochondrial stress,HBc thus perturbs mitochondrial dynamics and prevents the recycling of damaged mitochondria.Moreover,sustained HBc expression causes lysosomal consumption via Rab7 hyperactivation,which further hampers late-stage autophagy and substantially increases apoptotic cell death.Finally,we show that adenovirally expressed HBc in a mouse model is directly cytopathic and causes profound liver injury,independent of antigen-specific immune clearance.These findings reveal an unexpected cytopathic role of HBc,making it a pivotal target for HBV-associated liver disease treatment.The BCP-mutated HBV transgenic mice also provide a valuable model for understanding chronic hepatitis B progression and for the assessment of therapeutic strategies.展开更多
基金Supported by Youth Foundation of Health and Family Planning Commission of Shanxi ProvinceNo.201301024
文摘AIM: To investigate the frequency of mutations in pre-core (pre-C) and basic core promoter (BCP) regions of hepatitis B virus (HBV) from Shanxi Province, and the association between mutations and disease related indexes.METHODS: One hundred chronic hepatitis B patients treated at Shanxi Province Hospital of Traditional Chinese Medicine were included in this study. PCR-reverse dot blot hybridization and mismatch amplification mutation assay (MAMA)-PCR were used to detect the mutations in the HBV pre-C and BCP regions. HBV DNA content and liver function were compared between patients with mutant HBV pre-C and BCP loci and those with wild-type loci. The consistency between PCR-reverse dot blot hybridization and MAMA-PCR for detecting mutations in the HBV pre-C and BCP regions was assessed.RESULTS: Of the 100 serum samples detected, 9.38% had single mutations in the pre-C region, 29.17% had single mutations in the BCP region, 41.67% had mutations in both BCP and pre-C regions, and 19.79% had wild-type loci. The rates of BCP and pre-C mutations were 65.7% and 34.3%, respectively, in hepatitis B e antigen (HBeAg) positive patients, and 84.6% and 96.2%, respectively, in HBeAg negative patients. The rate of pre-C mutations was significantly higher in HBeAg negative patients than in HBeAg positive patients (χ<sup>2</sup> = 26.62, P = 0.00), but there was no significant difference in the distribution of mutations in the BCP region between HBeAg positive and negative patients (χ<sup>2</sup> = 2.43, P = 0.12). The presence of mutations in the pre-C (Wilcoxon W = 1802.5, P = 0.00) and BCP regions (Wilcoxon W = 2906.5, P = 0.00) was more common in patients with low HBV DNA content. Both AST and GGT were significantly higher in patients with mutant pre-C and BCP loci than in those with wild-type loci (P < 0.05). PCR-reverse dot blot hybridization and MAMA-PCR for detection of mutations in the BCP and pre-C regions had good consistency, and the Kappa values obtained were 0.91 and 0.58, respectively.CONCLUSION: HBeAg negative patients tend to have HBV pre-C mutations. However, these mutations do not cause increased DNA copies, but associate with damage of liver function.
文摘Objectives: To investigate the mutation of the basic core promoter (BCP) of hepatitis B virus (HBV) and clarify the significance of HBV quasispecies groups in patients with chronic HBV infection. Methods: A set of specific primers was synthesized according to the HBV DNA sequence of a Chinese strain. The BCP was amplified by PCR method from the serum of 40 patients with chronic HBV infection, and the PCR products of 2 patients were subcloned into pGEM Teasy vectors. Polyacrylamide gel elec- trophoresis (PAGE) was employed to display the de- letion mutations, and clones with differential length were selected to be sequenced. Sequence comparison was made to find the difference. Results: Two or three bands were displayed by PAGE in 60% patients. The results of sequence anal- ysis showed that there are some kinds of mutations in the BCP region. The substitution always occurs in TATA-like boxes, especially from T to C on 140 site. The deletion mutations were detected in TA1, TA2 and TA3. The 8bp, 20bp deletion mutations fre- quently happened. Conclusions: There is a hot deletion region in the BCP. The deletion and the substitution in the TATA- like box may influence the expression of preC/C pro- tein. The sequencing results indicate that there are HBV quasispecies groups in patients with chronic HBV infection.
基金supported by the Shanghai Municipal Science and Technology Major Project(ZD2021CY001)the National Natural Science Foundation of China(82372233,82072279,81871647,and 82205055)the Innovation Fund for Medical Sciences from Chinese Academy of Medical Sciences(2019-I2M-5-040).
文摘Chronic hepatitis B virus(HBV)infection can lead to advanced liver pathology.Here,we establish a transgenic murine model expressing a basic core promoter(BCP)-mutated HBV genome.Unlike previous studies on the wild-type virus,the BCP-mutated HBV transgenic mice manifest chronic liver injury that culminates in cirrhosis and tumor development with age.Notably,agonistic anti-Fas treatment induces fulminant hepatitis in these mice even at a negligible dose.As the BCP mutant exhibits a striking increase in HBV core protein(HBc)expression,we posit that HBc is actively involved in hepatocellular injury.Accordingly,HBc interferes with Fis1-stimulated mitochondrial recruitment of Tre-2/Bub2/Cdc16 domain family member 15(TBC1D15).HBc may also inhibit multiple Rab GTPase-activating proteins,including Rab7-specific TBC1D15 and TBC1D5,by binding to their conserved catalytic domain.In cells under mitochondrial stress,HBc thus perturbs mitochondrial dynamics and prevents the recycling of damaged mitochondria.Moreover,sustained HBc expression causes lysosomal consumption via Rab7 hyperactivation,which further hampers late-stage autophagy and substantially increases apoptotic cell death.Finally,we show that adenovirally expressed HBc in a mouse model is directly cytopathic and causes profound liver injury,independent of antigen-specific immune clearance.These findings reveal an unexpected cytopathic role of HBc,making it a pivotal target for HBV-associated liver disease treatment.The BCP-mutated HBV transgenic mice also provide a valuable model for understanding chronic hepatitis B progression and for the assessment of therapeutic strategies.
