Both plasmids mediated expression and secretion of the recombinant proteins in vitro in eukaryotic cells, detected with anti-DENV2 antibodies and evaluated by immunofluorescence or metabolic labeling assay followed by imunopreciptation. The vaccines pE1D2 and pE2D2 were also tested in combination with the chimeric YF17D-D2 virus, in a prime and booster system or with simultaneous immunizations. The pE1D2 DNA vaccine combined with the YF17D-D2 chimera induced high levels of neutralizing antibodies in animals vaccinated with any of the different immunization schedules.
These DNA vaccines will be further tested animal models. Concerning the primary immune response, eventhough the inoculation route applied was not natural, peripheral effects caused by DENV infection, like T cell activation and migration to target organs, as well as the production of antibodies and cytokines were observed.
Transference experiments of T cells obtained from vaccinated animals and traced by CFSE showed evident T lymphocytes activation, proliferation and migration to different sites in the host, including hepatic and brain tissue, after DENV challenge. In conclusion, this study contributed for the elucidation of the immune response dynamics in animals infected with DENV by the i. Moreover, this work provided new insights in the comprehension of the mechanisms involved in the protection induced by anti-dengue DNA vaccines.
DENV has four distinct serotypes DENV , each with several genotypes and all can cause non-severe dengue, with and without warning signals, and severe dengue. As we are an endemic country for Flavivirus, the first step of the work was to prove that the ZIKV samples received by our research center were really this virus, and there was no coinfection with DENV.
The main objective of this dissertation was to compare the cellular immune response triggered by two distinct isolates of DENV 3, one neurotropic genotype I and one non-neurotropic genotype III. The supernatants were analyzed by Griess reaction, for quantification of the nitric oxide produced by the cells and by CBA for the quantification of the cytokines.
It was also evaluated whether these cells were susceptible to virus infection by titration.
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