Induction of virus-neutralizing antibodies and T cell responses by dengue virus type 1 virus-like particles prepared from Pichia pastoris / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Dengue is currently a significant global health problem but no vaccines are available against the four dengue serotypes virus infections. The development of safe and effective vaccines has been hampered by the requirement of conferring complete protection against all four dengue serotypes and the lack of a convenient animal model. Virus-like particles (VLPs) have emerged as a promising subunit vaccine candidate. One strategy of vaccine development is to produce a tetravalent dengue subunit vaccine by mixing recombinant VLPs, corresponding to all four dengue virus serotypes. Towards this end, this study aimed to establish a Pichia pastoris (P. pastoris) expression system for production of dengue virus type 1 (DENV-1) VLPs and evaluate the humoral and cellular immune response of this particle in mice.</p><p><b>METHODS</b>A recombinant yeast P. pastoris clone containing prM and E genes of DENV-1 was constructed and DENV-1 VLPs expressed by this clone were analyzed by sucrose density gradient centrifugation, Western blotting, and transmission electron microscope. Groups of mice were immunized by these particles plus adjuvant formulations, then mice were tested by ELISA and neutralization assay for humoral immune response, and by lymphocyte proliferation and cytokine production assays for a cellular immune response.</p><p><b>RESULTS</b>Our data demonstrated that recombinant DENV-1 VLPs consisting of prM and E protein were successfully expressed in the yeast P. pastoris. Sera of VLPs immunized mice were shown to contain a high-titer of antibodies and the neutralization assay suggested that those antibodies neutralized virus infection in vitro. Data from the T lymphocyte proliferation assay showed proliferation of T cell, and ELISA found elevated secretion levels of interferon IFN-γ and IL-4.</p><p><b>CONCLUSIONS</b>P. pastoris-expressed DENV-1 VLPs can induce virus neutralizing antibodies and T cell responses in immunized mice. Using P. pastoris to produce VLPs offers a promising and economic strategy for dengue virus vaccine development.</p>

Inoculation of plasmid expressing the dengue 2 NS1 gene elicits immunity in mice / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To study cellular and humoral immune responses to NS1 protein in mice inoculated intramuscularly with recombinant plasmid expressing dengue 2 NS1 gene.</p><p><b>METHODS</b>The eukaryotic expressing plasmid pCNX2-NS1 was injected into tibialis anterior muscle in mice. The mice were subsequently boosted with the same dose and same method twice after the initial inoculation. The mice were killed at four-week intervals and their serum and spleen cells were harvested for further test.</p><p><b>RESULTS</b>Dengue 2 antibodies were detectable in the sera from inoculated animals four weeks after the last boost. The changes of CD4+ T lymphocyte and CD8+ T lymphocyte were also determined by flow cytometry.</p><p><b>CONCLUSION</b>The recombinant plasmid containing dengue 2 NS1 genes is immunogenic in intramuscularly inoculated mice. The vaccinated mice produced dengue-2 specific and long lasting immunity.</p>