The Experimental Study Of Different Routes In Immunization Against Dental Caries With PVAX1-SG And PVAX1-SSG In Mice And Rats

Objective:To investigate the efficiency of pVAX1-SG and pVAX1-SSG immunized in BABL/c mice and SD rats by different routes,and to explore the best means immune.Methods:This study consists of three parts:Test one:identification and preparation of recombinant plasmid pVAX1-SG1, pVAX1-SSG and plasmid pVAX1.Firstly,the plasmid of pVAX1-SG,pVAX1-SSG and PVAX1 were prepared in a small quantity by E.Z.N.A.Plasmid Mini Kit I.Secondly,to identify those plasmids with restriction endonuclease digestion and gel electrophoresis. Thirdly,pVAX1-SG,pVAX1-SSG and pVAX1 were purified and measured in large quantity with QIAGEN Plasmid Giga Kit,which were up to the requirements of the animal immunition,on the basis of correct identification.Test two:To immunize BABL/c mice with the recombinant plasmids in order to observe the effect.Sixty-four 6-week-old male BALB/c mice were randomly divided into 8 groups of 8 mice.A:vaccinated with saline via intranasal route(negative control group); B:vaccinated with pVAX1 via intramuscular route(negative control group);C:vaccinated with pVAX1-SG via.subcutaneous injection in the submandibalar gland region route;D: vaccinated with pVAX1-SG via intramuscular route;E:vaccinated with pVAX1-SG via intranasal route;F:vaccinated with pVAX1-SSG via.subcutaneous injection in the submandibalar gland region route;G:vaccinated with pVAX1-SSG via intramuscular route H:vaccinated with pVAX1-SSG.via intranasal route;Every mouse was immuned a dose of 100μg in a week and for three times in all(saline group,100μl/only).The mice were weighted and killed after sampling.Then some important organs(heart,liver,spleen,lung, kidney) of the mice were separated for pathological examination.Using enzyme-linked immunosorbent assay(indirect ELISA) to test the level of specific antibodies S-IgA in saliva and IgG.in serum.Test three:To immunize SD rats with the recombinant plasmids in order to observe the effect.First,to establish animal model of dental caries with SD rats.Then methods of division,the immunity means,immunity time and specific antibody detection are the same as the mice.To collect samples from first immune to age 80 days of mice,once a week.The classic of Caries were assessed according to Keyes scoring method.Results:①pVAX1-SG,pVAX1-SSG and pVAX1 were isolated and purifiedy by large QIAGEN Plasmid Giga Kit in order to meet the requirements of the study;②The antibodies of S-IgA in saliva and IgG in serum were significantly increased and continued for several weeks after immunization in mice,at the same time nasal drip group and submandibalar gland group had induced higher than intramuscular group(P＜0.05).③The specific antibodies which pVAX1-SSG induced were more than pVAX1-SG induced under in the same immunition way,and with significant statistics(P＜0.05).④The level of caries are significantly lower in experimental groups than the negative control groups with significant statistics(P＜0.05).Conclusions:①Gene vaccine pVAX1-SG and pVAX1-SSG can induce to improve markedly the level of specific in BABL/c mice and SD rats;②Gene vaccine pVAX1-SG and pVAX1-SSG can reduce caries by three different immune ways in SD rats.③nasal mucosal immunity may be the most effective immune way of gene vaccine pVAX1-SG and pVAX1-SSG.④pVAX1-SSG can be a better selection for specific antibodies than pVAX1-SG.