Abstract:

Abstract:Objective To explore the adjuvant effect of stimulating epitope  of mycobacterium tuberculosis heat shock protein 70(mtHSP70) on  fusion gene vaccine   of hepatocellular carcinoma associated antigens 661(HCA661), and to provide basis for improving immune effect of gene vaccine. Methods Eighteen BALB/c mice were randomly divided into three groups:pCI-neo group(n=6),pCI-HCA661-HSP70C group(n=6) and pCI-HCA661-HSP70E group(n=6).The  BALB/c mice were immunized intramuscularly with empty plasmid pCI-neo,recombinant plasmid pCI-HCA661-mtHSP70C and pCI-HCA661-mtHSP70E.The  changes of T cell subsets were analyzed by FCM. The antigen-stimulated production of IFNγ in the supernatants of  splenocytes in vitro and the specific  IgG antibody against HCA661 were detect by ELISA. The specific CTL cytotoxicity against tumor cells was used to detect the inhibitory effect on tumor cells. Results The percentagse of CD3+,CD4+T lymphocytes and the ratio of CD4+/CD8+ were significantly increased in  pCI-HCA661-mtHSP70C group and pCI-HCA661-mtHSP70E group compared with pCI-neo group（P<0.05）. The percentages of CD3+,CD4+ T lymphocytes and the ratio of CD4+/CD8+ were  significantly increased in pCI-HCA661-mtHSP70E group compared with pCI-HCA661-mtHSP70C group(P<0.05). The concentration of IFNγ in pCI-HCA661-mtHSP70E group was higher than that in pCI-HCA661-mtHSP70C group in supernatants from cultural splenocytes under HCA661 peptide stimulation（P<0.05）. The IFNγ levels in  pCI-HCA661-mtHSP70C group and pCI-HCA661-mtHSP70E group were higher than that in pCI-neo group(P<0.05).The levels of specific antibodies induced by fusion gene in pCI-HCA661-mtHSP70E group were increased significantly compared with pCI-HCA661-mtHSP70C group（P<0.05）.The anti-tumor effect of CTL was enhanced strongly in pCI-HCA661-mtHSP70E group. Conclusion Stimulating epitope of mtHSP70 can induce a strong specific immune response to HCA661 fusion gene vaccine in mice by its adjuvant effect.

Key words: fusion gene vaccine;hepatocellular carcinoma-associated antigens 661;mycobacterium tuberculosis heat protein 70;genetic adjuvant