Immunological Study On ConA Combined With Mycobacterium Tuberculosis HSP65 DNA Vaccine

PurposerConA was combined with mycobacterium tuberculosis HSP65 DNA vaccine to enhance the function of APCs and cellular immunity induced by the DNA vaccine.Method:In this experiment recombinant plasmid DNA was got by basic solution from Escherichia coli ,that is mycobacterium tuberculosis HSP65 DNA vaccine we need,and,adjust its concentration to Img/ml with O.OIM PBS. Balb/c mice were divided into four groups, then injected respectively with PBS( group A),with ConA(group B);with DNA(group C) ; and DNA plus ConA( group D). Mice in group A were only injected O.OIM PBS,mice in group B were only received intramusclar injection with ConA 500μ g one times, mice of group C received mycobacterium tuberculosis HSP65 DNA vaccine 100 μ L three times once every two weeks ,and mice of group D with 500 μ g ConA plus DNA vaccine : ConA was applied only one times before two days of the first immunization of DNA vaccine,DNA vaccine was applied once every two weeks adding up to three times.Removed eyeballs of mice and collected blood ,determined HSP65-specific antibodies IgG by enzyme linked immenosorbent assay ( ELISA) at the 4 ,6,8 ,10 week after first immunization .At the 8 th week spleen of mice were used to detect mRNA expression of IL-4 and IFN- ?, peritoneal macrophages were collected from mice of different groups by washing peritoneal cavity,and cultured to measure NO level by commercially available kits . Results : HSP65-specific antibodies IgG were lower in the mice of group D than those of group C at the same dilution , at the 4th ,6th 8th,10th week the absorbency of HSP65-specific antibodies IgG of group C at 1:100 were 1.06 , 1.40, 1.83, 1.63,while in group D mice the corresponding absorbency were 0.93, 1.24, 1.71, 1.59 .At the 8th week the concentration of HSP65-specific antibodies IgG of group C and D respectivly was 729.38rng/L and 672.55mg/L (p<0.0l) .The mRNA expression of IFN- Y were detected in all groups ,but group D was the strongest among them ,group of A,B,C and D respectivly was 0.26 ,0.44 ,0.51 and 0.89 , the mRNA expression of IL-4were also detected in all groups ,group A was the strongest , group of A,B,C and D respectivly was 0.85 ,0.70 ,0.62 and 0.10 .the difference of mRNA expression of IL-4 and IFN- Y between group D and the other three groups are notable(P<001). NO of peritoneal macrophages was highest in group D(the concentration of NO in group A,BC and D were respectivly 323.42 umol/L ,448.20 umol/L ,439.03 jimol/L and 515.43 umol/L, (p<0.05 vs B C). Conclusion: At the level of moleculear we may draw a conclusion that ConA combined with mycobacterium tuberculosis HSP65 DNA vaccine have suppressed humoral immmune response in some degree, improved cellular immunity by prompting ThO cells to differentiate into Thl cells ,and have increased the level of NO, which demonstrated indirectly that ConA can enhance the function of APCs .