摘要
The epidemic of HIV/AIDS is sweeping across the world. It is of great importance to figure out new ways to curb this disease. Epitope-based vaccine is one of these solutions. In this study, a chimeric gene was obtained by combination of a designed HIV-1 multi-epitope gene (MEG) and HIV-1 p24 gene. A re- combinant plasmid pUTA2-MEGp24 was then constructed by inserting MEGp24 gene into the down- stream of the promoter (ATI-P7.5×20) of fowlpox virus (FPV) transfer vector pUTA2. The recombinant plasmid and wild-type FPV 282E4 strain were then co-transfected into CEF cells and homologous re- combination occurred. A recombinant virus expressing HIV-1 protein MEGp24 was screened by ge- nome PCR and Western blot assay. Large scale preparation and purification of the recombinant fowl- pox virus (rFPV) were then carried out. BALB/c mice were immunized intramuscularly with the rFPV for three times on day 0, 14 and 42. Mice were executed and sampled one week after the third inoculation. Anti-HIV-1 antibody in serum and Th1 cytokines in the supernatant of cultured spleen cells were as- sayed by ELISA. The count of T lymphocyte subsets and the CTL activity of spleen lymphocytes were analyzed by flow cytometry and lactate dehydrogenase (LDH) release assay, respectively. The results showed that HIV-1 specific antibody in serum and increased T lymphocyte subsets (CD4+ T, CD8+ T) were detected in the immunization group. CTL target-killing activity and higher secretion of Th1 cyto- kines (IFN-γ and IL-2) of spleen lymphocytes stimulated by H-2d-restricted CTL peptide were observed in immunized mice. We concluded that the rFPV may induce HIV-1 specific immunity especially cellular immunity in mice.
The epidemic of HIV/AIDS is sweeping across the world. It is of great importance to figure out new ways to curb this disease. Epitope-based vaccine is one of these solutions. In this study, a chimeric gene was obtained by combination of a designed HIV-1 multi-epitope gene (MEG) and HIV-1 p24 gene. A re- combinant plasmid pUTA2-MEGp24 was then constructed by inserting MEGp24 gene into the down- stream of the promoter (ATI-P7.5×20) of fowlpox virus (FPV) transfer vector pUTA2. The recombinant plasmid and wild-type FPV 282E4 strain were then co-transfected into CEF cells and homologous re- combination occurred. A recombinant virus expressing HIV-1 protein MEGp24 was screened by ge- nome PCR and Western blot assay. Large scale preparation and purification of the recombinant fowl- pox virus (rFPV) were then carried out. BALB/c mice were immunized intramuscularly with the rFPV for three times on day 0, 14 and 42. Mice were executed and sampled one week after the third inoculation. Anti-HIV-1 antibody in serum and Th1 cytokines in the supernatant of cultured spleen cells were as- sayed by ELISA. The count of T lymphocyte subsets and the CTL activity of spleen lymphocytes were analyzed by flow cytometry and lactate dehydrogenase (LDH) release assay, respectively. The results showed that HIV-1 specific antibody in serum and increased T lymphocyte subsets (CD4+ T, CD8+ T) were detected in the immunization group. CTL target-killing activity and higher secretion of Th1 cyto- kines (IFN-γ and IL-2) of spleen lymphocytes stimulated by H-2d-restricted CTL peptide were observed in immunized mice. We concluded that the rFPV may induce HIV-1 specific immunity especially cellular immunity in mice.
基金
the China"863"Program(Grant No.2003AA219051)
