[Effect of the immune strategy based on BCG priming and Ag85A/GM-CSF DNA vaccine boosting in mice].

AIM: To investigate immune effect in mice on the basis of BCG priming and DNA vaccine boosting, and to provide a new strategy for development of new type of anti-tuberculosis DNA vaccine further. METHODS: After mice were inoculated with BCG of 1x10(6) clone formation unit each three weeks, 100 microg of DNA vaccine was injected intramuscularly in mice two times at 3 week intervals. The proliferative responses of murine cytotoxic T lymphocytes (CTL), natural killer (NK) and spleen cell to antigen 85A(Ag85A) were measured by MTT method respectively. The antibody titers and IFN-gamma level from the immunized mice were detected by ELISA. RESULTS: The proliferative responses of CTL and spleen cell to Ag85A, as well as IFN-gamma level in mice immunized with prime-boost strategy were significantly increased respectively compared with the control mice immunized with blank plasmid or BCG only. Although NK activity was a little higher in mice immunized with prime-boost strategy than that of immunized with blank plasmid mice, it was still lower than that of mice immunized with BCG alone. The titer of the specific antibody against Ag85A in mice immunized with prime-boost strategy was also higher than that of mice immunized with DNA vaccine alone. CONCLUSION: The immune strategy of BCG-prime and Ag85A/GM-CSF DNA vaccine boost improve immune effect, especially the Th1 cellular immune response increase obviously. This study provides the possibility of further research for investigating protective function in immunized mice challenged by Mycobacterium tuberculosis.

[Anti-tumor mechanisms of Sp2/0 tumor vaccine transfected with mIL-21 gene in mice].

AIM: To explore the anti-tumor mechanism of Sp2/0-mIL-21 tumor vaccine in mice. METHODS: The molecules of MHC-I and CD80 on the surface of Sp2/0-mIL-21 tumor vaccine were detected by flow cytometry (FCM) respectively. A flow cytometric CFSE-7-AAD cytotoxicity assay was used to detect the cytotoxic activities of NK cells and CTLs. The expression of I-TAC in the tumor tissue was tested by RT-PCR. RESULTS: The expression of MHC-I molecule on the surface of tumor vaccine was up-regulated obviously. The cytotoxic activities of NK cells and CTLs were significantly enhanced in the mice inoculated with Sp2/0-mIL-21 tumor vaccine compared with the mice inoculated with Sp2/0 tumor cell in control group. The expression of I-TAC in the tumor tissue was up-regulated. The histopathologic section analysis showed more lymphocytes were infiltrated in the tumor tissue. CONCLUSION: Sp2/0-mIL-21 tumor vaccine can induce strong cell-mediated immune response to tumor cells after it was inoculated s.c into mice. The anti-tumor mechanisms induced by Sp2/0 tumor cell vaccine are associated with the proliferation and activation of T lymphocyte, the differentiation and maturity of NK cell, the infiltration of lymphocytes in tumor tissue, and the enharuement of cytotoxic activities of NK cells and CTLs.

Novel constructs of tuberculosis gene vaccine and its immune effect on mice.

A novel tuberculosis (TB) gene vaccine containing mouse granulocyte macrophage-colony stimulating factor (mGM-CSF) and a TB antigen (Ag85A) was developed in this study. The genes encoding Ag85A and mGM-CSF were amplified by PCR respectively from the Ag85A-containing pBSby5 and pC-mGM-CSF. The genes were then cloned into two different polylinker sites of plasmid pIRES, forming a novel TB gene vaccine construct pI85AGM. Following transfection of pI85AGM plasmid into 7721 cell line by Lipofectamine(TM), the expression of Ag85A and GM-CSF proteins was identified by Western blotting or RT-PCR. Then Balb/c mice were inoculated with the recombinant pI85AGM, pI85A, pIGM or plasmid alone, respectively. The activities of CTL, NK cells and the Ag85A-stimulated proliferation of spleen cells were measured by MTT method. The serum antibody against Ag85A was detected by ELISA. The results showed that the Ag85A and GM-CSF proteins could be expressed in 7721 cell line and the activity of CTLs and the proliferation of spleen cells were significantly increased in the pI85AGM-immunized mice, indicating that the pI85AGM-immunized mice could generate specific immune responses to Ag85A. This study might provide possibility for developing novel anti-TB gene vaccine.