Adolescent ; Capillary Leak Syndrome ; chemically induced ; Female ; Granulocyte Colony-Stimulating Factor ; adverse effects ; Humans ; Male ; Middle Aged

OBJECTIVETo investigate the clinical significance of neutralizing anti-interferon-alpha antibodies (NA) in chronic hepatitis B patients treated with recombinant interferon-alpha(rIFN-alpha).

METHODSOne hundred and eighty-one patients (128 male and 53 female) with histological proven chronic hepatitis B were treated with 5 MU recombinant interferon-alpha 1b (rIFN-alpha 1b) subcutaneously thrice weekly for 6 to 37 (median 10) months. For each patient, Specific detection of serum HBV DNA level with fluorescent-quantitative PCR, HBeAg with enzymoimmunoassay and NA with an antiviral neutralizing biological assay were performed during therapy.

RESULTSNA was found in 61 (33.7%) of 181 patients. At the end of treatment, complete-response was achieved in 17 (27.9%) of 61 patients with NA and in 54 (45.0%) of 120 patients without NA, respectively (chi2=4.979). For NA positive patients, the complete-response rate was significantly lower in those who had not achieved partial-response prior to or at the same time as NA occurred than in those who did (3.8%, 1/26, vs. 45.7%, 16/35, chi2 = 7.457). Moreover, it was lower in patients who either had 20pg/ml of serum HBV DNA or above or HBV DNA had being reduced by less than 60% recent 3 months, but higher in those who had less than 20pg/ml of HBV DNA and HBV DNA had being reduced by 60% or above (20.0%, 9/45, vs. 56.3%, 9/16, chi2 = 11.009).

CONCLUSIONNA may negate the antiviral effects of rIFN-alpha in chronic hepatitis B patients treated with rIFN-alpha, especially if they appear before partial-response or at the occasion at which serum HBV DNA level was not below 20pg/ml or HBV DNA had being reduced by less than 60% recent 3 months.

Antibodies ; blood ; DNA, Viral ; blood ; Female ; Hepatitis B, Chronic ; drug therapy ; virology ; Humans ; Interferon-alpha ; immunology ; therapeutic use ; Male ; Recombinant Proteins ; therapeutic use

OBJECTIVETo investigate the relationship of virological breakthrough and production of neutralizing anti-interferon antibody (NAb) in chronic hepatitis B patients treated with recombinant interferon-alpha (rIFN-alpha).

METHODFour hundred eighty-five patients with histological proven chronic hepatitis B were treated with 5 MU recombinant interferon-alpha 1b (rIFN-alpha1b) thrice weekly for 6-37 months (median 10). Serum HBV DNA, HBeAg and NAb levels of the patients were detected by fluorescent-quantitative PCR, enzymoimmunoassay and antiviral neutralizing biological assay respectively during the therapy.

RESULTSVirological breakthrough occurred in 66 patients (13.6%), and NAb was found in 98 patients (20.2%) of the total 485 patients. The rate of NAb positivity was higher in patients with viral breakthrough than those without it (68.2%, 45/66, vs 12.6%, 53/419, chi(2)=109.06, P < 0.01), and viral breakthrough occurred more in patients with positive NAb than with negative NAb (45.9%, 45/98, vs 5.4%, 21/387, chi(2)=109.06, P < 0.01). The time of the viral breakthrough occurrence and the time of NAb production had a significant correlation (P < 0.01). The occurrence of viral breakthrough was also influenced by the age of patients (P < 0.05) and HBeAg status (P < 0.01) before they were treated.

CONCLUSIONViral breakthrough occurred in 13.6% of our 485 chronic hepatitis B patients treated with recombinant interferon-alpha. Their viral breakthrough and production of NAb production had a significant correlation.

Adult ; Antibodies, Neutralizing ; biosynthesis ; Female ; Hepatitis B Antibodies ; biosynthesis ; Hepatitis B virus ; immunology ; Hepatitis B, Chronic ; drug therapy ; virology ; Humans ; Interferon Type I ; therapeutic use ; Male ; Recombinant Proteins ; Young Adult

OBJECTIVETo study hepatitis B virus (HBV) expression in 3 hepatocytes infected with recombinant adenovirus containing 1.2-copy HBV DNA.a

METHODSA chicken hepatoma cell line and two human hepatocytes were infected by the recombinant adenovirus containing 1.2-copy HBV DNA at 25 pfu/cell. HBV-specific mRNA was detected by RT-PCR 3 days after the infection, and HBsAg and HBeAg were detected by ELISA and HBV DNA by real-time PCR daily after the infection.

RESULTSHBV mRNA expression was detected in all the 3 cells after recombinant adenovirus infection, and the quantities of HBV DNA and HBV antigens in the culture supernatant increased with the passage of time.a

CONCLUSIONInfection with the recombinant adenovirus containing 1.2-copy HBV DNA can mediate HBV infection in the 3 cells in vitro.

Adenoviridae ; genetics ; Animals ; Cell Line, Tumor ; Culture Media, Conditioned ; metabolism ; DNA, Recombinant ; genetics ; DNA, Viral ; genetics ; metabolism ; Gene Expression ; Hepatitis B Antigens ; metabolism ; Hepatitis B virus ; Hepatocytes ; metabolism ; virology ; Humans ; Reverse Transcriptase Polymerase Chain Reaction

Hepatitis B Core Antigens ; immunology ; Hepatitis B virus ; genetics ; immunology ; Hepatitis B, Chronic ; virology ; Humans ; T-Lymphocytes, Cytotoxic ; immunology

BACKGROUNDIt is still unclear whether viral genetic variability influences response to interferon (IFN)-alpha treatment. Recent reports suggest that IFN-alpha effects may be associated with hepatitis B virus (HBV) post-transcriptional regulation. This study was designed to explore the heterogeneity of HBV post-transcriptional regulatory elements (HPRE) and the relationship between the diversity of HPRE and the response to IFN-alpha treatment.

METHODSThe HPRE sequences from 31 Chinese patients infected with HBV were determined by directly sequencing of polymerase chain reaction (PCR) product, and comparing them to those from Caucasian patients. Subsequently, eukaryotic expression vectors containing HPRE at various points were constructed and transfected into HepG2 cells, which were then exposed to recombinant human cytokines.

RESULTSThe T to C point mutation at nt 1504 and the C to T (G) at nt 1508 in HPRE were found in 21 and 19 patients with chronic hepatitis B, respectively; the C to T point mutation at nt 1509 was found in 17 patients. These point mutations did not exist in the HPRE of the Caucasian patients. The activity of the CAT gene obviously increased in the case of T to C point mutation at nt 1504, but did not change in the case of the C to T (G) mutations at nt 1508 and 1509. The activity of the CAT gene at these point mutations of HPRE could be inhibited by IFN-alpha/gamma and tumor necrosis factor (TNF)-alpha except for the point mutations at nt 1508 of HPRE which may escape the suppression role of IFN-alpha on HPRE.

CONCLUSIONSThere are point mutations between the HPRE of Chinese and Caucasian HBV patients, which might be correlated with response to IFN-alpha. The variation of HPRE might affect the function of HPRE and influence the regulative function of IFN-alpha other than that of IFN-gamma or TNF-alpha on HPRE.

Chloramphenicol O-Acetyltransferase ; metabolism ; Genes, Regulator ; Hepatitis B virus ; drug effects ; genetics ; Hepatitis B, Chronic ; drug therapy ; virology ; Humans ; Interferon-alpha ; pharmacology ; Interferon-gamma ; pharmacology ; Plasmids ; Point Mutation ; Tumor Necrosis Factor-alpha ; pharmacology

Objective To study the biological significance of HBV gene mutation at nucleotide (nt) 1862. Methods The EB virus eukarotic expression vector for HBV pre-C/C gene was constructed using molecular biological method, HBV pre-C/C gene mutation at nt 1862 was induced by way of site-specific mutation technique, and identified by PCR-RFLP and sequencing analysis. The resulted recombinant plasmid containing the HBV variant was subsequently transfected into Cos7 cell line mediated by lipofectin, to observe the expression of HBeAg. The cells transfected with the recombinant plasmid con- taining wild HBV pre-C/C gene fragment served as control. Results HBeAg expression was detected in the cells transfected with wild recombinant plasmid but not in those with HBV variant transfection. Conclusion The success in the construction of eukarotic expression vector for HBV pre-C/C gene mutation at nt 1862 may pave the way for further studying a series of biological changes of HBV resulted from the mutation addressed in this study.

Objective To study the biological significance of HBV gene mutation at nucleotide (nt) 1862. Methods The EB virus eukarotic expression vector for HBV pre-C/C gene was constructed using molecular biological method, HBV pre-C/C gene mutation at nt 1862 was induced by way of site-specific mutation technique, and identified by PCR-RFLP and sequencing analysis. The resulted recombinant plasmid containing the HBV variant was subsequently transfected into Cos7 cell line mediated by lipofectin, to observe the expression of HBeAg. The cells transfected with the recombinant plasmid con- taining wild HBV pre-C/C gene fragment served as control. Results HBeAg expression was detected in the cells transfected with wild recombinant plasmid but not in those with HBV variant transfection. Conclusion The success in the construction of eukarotic expression vector for HBV pre-C/C gene mutation at nt 1862 may pave the way for further studying a series of biological changes of HBV resulted from the mutation addressed in this study.

OBJECTIVETo study the mechanism of hepatitis B virus infected patients who is negative for HbsAg.

METHODSDNA sequences of 46 patients were analyzed. In these patients, HBsAg was negative but HBV DNA was positive and six new HBsAg variants were identified. Four of the six variants were combined point mutants and two were insertion variants. These S genes were subcloned into eukaryotic expression vector EBO-plpp, and the recombinant eukaryotic expression plasmids were transfected into COS7 cells. Cell lines expressing mutant type HBsAg were obtained. The supernatants were detected by ELISA and RIA.

RESULTSOnly the two-amino acid-insertion variants could be detected and the others failed to react with polyclonal and monoclonal antibodies against HbsAg.

CONCLUSIONThe results indicated that the point mutations and insertions may result in a conformational change of the S gene, which affect HBsAg antigenicity, suggesting a possible relationship between the variants and the negative conversion of HBsAg of the patients.

Animals ; Antigenic Variation ; COS Cells ; Cercopithecus aethiops ; Hepatitis B Surface Antigens ; genetics ; immunology ; Hepatitis B virus ; genetics ; immunology ; Hepatitis B, Chronic ; immunology ; virology ; Humans ; Plasmids ; genetics ; Point Mutation ; Transfection