ABSTRACT: Nearly 350 million persons worldwide are chronically infected with hepatitis B virus (HBV). Ubiquitin (Ub) is a highly conserved small regulatory protein, ubiquitous in eukaryotes, that usually serves as a signal for the target protein that is recognised and degraded in proteasomes . The Ub-mediated processing of antigens is rapid and efficient and stimulates cell-mediated immune responses. Accordingly, Ub-mediated processing of antigens has been widely used in chronic-infection and cancer studies to improve immune response.
Many clinical trials have shown that DNA vaccine potency needs to be greatly enhanced. Here, we report a new strategy for designing an HBV DNA vaccine using the ubiquitin (Ub) sequence. The aim of this study was to investigate a novel DNA vaccination, based on the expression of HBV core antigen (HBcAg), fused to Ub to enhance DNA vaccine potency.
Mouse ubiquitin fused to the HBcAg gene and cloned into the eukaryotic vector pcDNA3.1 (-). BALB/c mice were immunized with recombinant pUb-HBcAg or pHBcAg DNA vaccine. Lymphocyte proliferation assay, intracellular IFN-γ assay, CTL cytotoxicity assay, and antibody assay were performed to analyze the cellular and humoral immune responses to our DNA constructs.
HBcAg was expressed effectively in the COS-7 cells that were transiently transfected with pUb-HBcAg. Strong anti-HBc IgG responses were elicited in mice that were immunized with pUb-HBcAg. The endpoint titers of anti-HBc peaked at 1:656100 on the 42nd day after the third immunization. pUb-HBcAg stimulated greater lymphocyte proliferation and induced higher levels of IL-2 and IFN-γ and a greater percentage of HBcAg-specific CD8+ T cells in mice than pHBcAg. In the CTL assay, the specific lysis rate reached 56.5% at an effector:target ratio of 50:1 in mice that were immunized with pUb-HBcAg.
pUb-HBcAg elicits specific anti-HBc responses and induces HBc-specific CTL responses in immunized BALB/c mice. Our results imply that Ub can be used as a molecular adjuvant that enhances the potency of DNA vaccines.
Hepatitis Monthly 08/2011; 11(8):620-8. · 2.19 Impact Factor
ABSTRACT: To define the expression of single-chain variable fragment (ScFv) against hepatitis B virus core protein (HBc) mediated by recombinant replication defective adenovirus carrying the anti-HBc ScFv gene in vitro and to define the activity of anti-HBc ScFv combining HBcAg.
The recombinant adenoviruses carrying anti-HBc ScFv gene generated by homologous recombination in bacteria and packaged in 293 cells were transfected into HepG2 cells, and the anti-HBc ScFv was detected using SDS-PAGE and Western blot.
Green fluorescent protein (GFP) was observed in HepG2 cells after the transfection. SDS-PAGE displayed a protein strap about 2.7 x 10(4), and the result of Western blot displayed a positive reactive strap.
Anti-HBc ScFv can be expressed in cells mediated by recombinant replication defective adenovirus carrying the anti-HBc ScFv gene.
Zhonghua gan zang bing za zhi = Zhonghua ganzangbing zazhi = Chinese journal of hepatology 09/2006; 14(8):587-9.
ABSTRACT: To explore the effects of interferon-alpha (IFN-alpha) application on peripheral circulating CD1alpha dendritic cells (DCs)in patients with chronic hepatitis B, and the expression of HLA-DR, CD80, and ICAM-1 on CD1alpha DCs in order to explore the mechanism of immune modulation of IFN-alpha.
By flow cytometry technique, changes of CD1alpha DCs were monitored in 22 patients with chronic hepatitis B treated with IFN-alpha and in 16 such patients not treated with IFN-alpha within three months. Meanwhile, the expression of HLA-DR, CD80, and ICAM-1 on CD1alpha DCs was detected.
In the group of IFN-alpha treatment, the percentage of CD1alpha DCs in peripheral blood mononuclear cells was increased after three months of therapy. In patients who became negative for HBV-DNA after IFN-alpha treatment, the increase of DCs was more prominent, while in control, these changes were not observed. Increased expression of HLA-DR, CD80,and ICAM-1 on CD1alpha DCs was also observed.
CD1alpha DCs can be induced by IFN-alpha in vivo, and the immune related molecules such as HLA-DR, CD80,and ICAM-1 are up-regulated to some degree. This might be an important immune related mechanism of IFN-alpha treatment for chronic hepatitis B.
World Journal of Gastroenterology 04/2006; 12(9):1447-51. · 2.47 Impact Factor
ABSTRACT: To investigate the association between curative effects of interferon-alpha and partial human leucocyte antigen (HLA) II alleles in chronic viral hepatitis B.
Sixty patients with chronic viral hepatitis B in Shanghai were treated with a standard course of treatment with interferon-alpha for 6 mo. HLA-DRB1, -DQA1, and -DQB1 alleles were detected by polymerase chain reaction-sequence specific primer (PCR-SSP) method.
Frequencies of HLA-DRB1*04 (P<0.025) and HLA-DQA1*0303 (P<0.01) in non-responders were significantly higher than those in partial and complete responders. Frequencies of HLA-DQA1*0505 (P<0.025) and HLA-DQB1*0301 (P<0.005) in partial and complete responders were significantly higher than those in non-responders.
Non-response to interferon-alpha therapy is positively correlated with HLA-DRB1*04 and HLA-DQA1*0303, and negatively correlated with HLA-DQA1*0505 and -DQB1*0301 in patient with chronic viral hepatitis B. HLA II genes of the identification alleles provide a method for evaluating outcome of interferon-alpha treatment.
World Journal of Gastroenterology 07/2004; 10(14):2116-8. · 2.47 Impact Factor