| Objective:After immunization of BALB/c mice by lavage method, the distribution and expression of oral anti-caries DNA vaccine p VAX1-SG in tissues and organs were observed in this paper, aiming at providing the theory on judging of anti-caries effect.Methods:Sixty-six 6-8 weeks old female BALB/c mice weighted between 18 g and 22 g,were randomly divided into 3 groups: 30 in experimental group, 18 in vector control group and 18 in blank control group. Recombinant plasmid p VAX1-SG and vector p VAX1 were identified and massive extracted. The temperature sensitive hydrogel encapsulated recombinant plasmid p VAX1-SG was applied to synthetize oral anti-caries DNA vaccine.The experimental group vaccinated the temperature sensitive hydrogel encapsulated recombinant plasmid p VAX1-SG by lavage method, the empty plasmid vector group vaccinated the temperature sensitive hydrogel encapsulated empty plasmid vector p VAX1 by lavage method, and the blank control group was without any treatment. All group were immunized twice at 2 weeks interval, and each immune dose was 100μg. The mice were weighed and killed after the second immunization in 1, 2, 7, 15, 30 and 60 days. The distribution and expression of p VAX1-SG in tissues and organs, including heart, liver,spleen, lung, kidney, brain, stomach, submandibular gland, small intestine and organs of generation, were checked by quantitative real-time PCR. Histopathological examination of the second immunized mice on the 60 th day was initiated in heart, liver, spleen, lung,kidney and small intestine. The target protein expression of immunized small intestine and submandibular gland of the second immunized mice on the 7th day was checked by Envision method.Results:1 The DNA of p VAX1-SG was detected in the immunized mice between1 and 60 days by quantitative real-time PCR, but there were differences of DNA c opies in tissues and organs. The most distributions of p VAX1-SG were organs of g eneration in the immunized mice on the 1st day, liver on the 2nd day, heart on 7t h and 15 th days, submandibular gland on 30 th day and spleen on 60 th day, respectively, while the values of them were(3.08E+08)±(0.92E+08) copies/ml,(3.49E+07)±(0.27E+07) copies/ml,(7.96E+07)±(0.54E+07) copies/ml,(9.32E+08)±(0.27E+08)copies/ml and(3.49E+07)±(0.27E+07) copies/ml respectively.2 p VAX1-SG of the immunized mice was expressed in tissues and organs, and relative quantitation of m RNA expression decreased with time went by. The main expressions of p VAX1-SG were spleen and small intestine in the immunized mice on the 1st day(p <0.05), liver, small intestine and organs of generation on the 2nd day(p < 0.05), liver and submandibular gland on the 7th day(p < 0.05) and small intestine on the 15 th day,respectively. The relative quantitation of m RNA in tissues and organs decreased on the30 th and 60 th days.3 The expression of recombinant protein was observed in the relative cell of small intestine and submandibular gland of immunized mice.4 Compared with the control group, there was no difference in weight of animals. There were no obvious pathological changes according to the observation of gliosis of the important organs and tissues of the experimental group.Conclusions:1.Distribution and expression of oral anti-caries DNA vaccine p VAX1-SG in tissues and organs were examined in immunized BALB/c mice by lavage method.However, distribution and expression of temperature sensitive hydrogel excapsulated recombinant plasmid p VAX1-SG did not synchronize, indicating that the expression of the recombinant plasmid p VAX1-SG had spatio-temporal characteristic.2. Expression of the target protein of oral anti-caries DNA vaccine p VAX1-SG in small intestine and submandibular gland of immunized mice by lavage method was examined.There was no significant pathological change in heart, liver, spleen, lung, kidney and immunized small intestine, indicating that recomdinant plasmid p VAX1-SG could be expressed in mice, and no toxic effect was observed in oral anti-caries DNA vaccine p VAX1-SG immunized mice. |
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