Effects Of Cytokines On Immunocontraception Of DNA Vaccine Of Mouse Zona Pellucida 3

In the process of mammalian fertilization, the zona pellucida-3 (ZP3) glycoprotein on egg surface as the sperm receptor can bind to sperm and induce the sperm acrosome reaction, then complete the fertilization process. ZP3-specific antibody can block the binding site of ZP3 protein on oocyte surface, and inhibit sperm-egg interaction. ZP-based contraceptive vaccine is an ideal approach for human contraception and a humanistic control method for animals. To develop the ZP-based contraceptive vaccine for human application, we made an effort to improve the ZP3-specific antibodies immune response by selecting suitable immue adjuvants and proper vaccination route. DNA vaccine is considered as promising vaccine approach since its safe, cheap and ease to get and save, and it is broadly used for the development of anti-virus vaccine. In our previous study, either intramuscular or intranasal delivery of mZP3 DNA vaccine induced contraception in mice, but the immunogenicity is too weak and it needs to be enhanced by adjuvant.The efficacy of DNA vaccines may crucially depend upon their capacity to transduce dendritic cells (DCs). Granulocyte-macrophage colony-stimulating factor (GM-CSF) can influence the activation and maturation of antigen-presenting cells (APCs), particularly enhance the recruitment and function of DCs. In this study, mouse GM-CSF eukaryotic expression vector was constructed, then co-intramuscularly delivered with or 4 days prior to mouse zona pellucida-3 (mZP3) DNA vaccination. The results showed that GM-CSF improved the maturation of DCs, co-administration of GM-CSF and mZP3 enhanced the mRNA expressive levels of co-stimulation factors greatly. These data suggest that GM-CSF could promote the maturation of DCs and enhance its function on antigen presentation; GM-CSF could be used as the adjuvant of DNA vaccine to develop immunocontraceptive vaccine.Interleukin-5 (IL-5) is a cytokine secreted by T helper 2 (Th2) cells, that participates in the regulation of antibody secretion. In this study, we investigated whether GM-CSF and IL-5 can be used as molecular adjuvants to increase the humoral immune response generated by mZP3 DNA vaccine. Mice in experimental group were injected by GM-CSF 4 days before the co-immunization of IL-5 and mZP3 DNA vaccine. The contraception and the correlation with humoral and cellular immune responses were analyzed after immunization and mating. The effect of cytokine adjuvant on the maturation of DCs was evaluated. Results: Co-immunization of GM-CSF and IL-5 with mZP3 DNA vaccine induced the highest level of serum IgG and IL-4 expression in CD4+ T cells. Importantly, this strategy reduced mice fertility without disrupting normal ovarian morphology. GM-CSF enhanced the matura-tion of DCs evidenced by up-regulating the expression of MHC-Ⅱ and CD86. Conclusion:GM-CSF and IL-5 co-administration enhanced humoral immune responses to mZP3, and this may be a potential strategy for development of immunocontraceptive vaccine.Beside IL-5, interleukin-4(IL-4) can improve the immunogenicity of DNA vaccine as well. In this study, we investigated whether GM-CSF and IL-4 can be used as cytokine adjuvants to increase immunogenicity of mZP3 DNA vaccine. Mice in experimental group were injected by GM-CSF 4 days before the co-immunization of IL-4 and mZP3 DNA vaccine. Results: Co-immunization of GM-CSF and IL-4 with mZP3 DNA vaccine induced the highest level of serum IgG and reduced mice fertility. Conclusion:GM-CSF and IL-4 co-administration enhanced humoral immune responses to mZP3, and this may be a potential strategy for development of immunocontraceptive vaccine.Intranasal or oral delivery is an ideal route for ZP3 vaccination. To enhance the efficiency of mucosal delivery for DNA regimens, chitosan was used for the nasal delivery of DNA vaccines. In this study, we tested whether intranasal administration of GM-CSF can improve the immunocontraception of mZP3 DNA vaccine. The results showed that intranasal co-administration of GM-CSF and mZP3 DNA vaccine could effectively increased the levels of IgG antibodies in serum and secretory IgA (slgA) in vaginal washes, respectively. GM-CSF by promoting the maturation of DCs, enhanced Th2 type immune response induced by mZP3 DNA vaccine. More importantly, the intranasal co-administration of GM-CSF and mZP3 DNA vaccine reduced the mouse fertility rate and mean litter size without interfering the normal follicular development. These data suggest that GM-CSF could be used as molecular adjuvant to promote the immunocontraception.There is a common receptor shared between IL-5 and GM-CSF which makes a collaborative immunomodulatory of these two cytokines. By intranasal immunization of GM-CSF and IL-5, we tested whether GM-CSF and IL-5 as mucosal adjuvants can affect the immunocontraception of mouse zona pellucida 3 DNA vaccine. The results showed that co-administrated GM-CSF and IL-5 with mZP3 DNA vaccine increased the level of IgG antibodies serum, but not the secretory IgA (sIgA) in vaginal washes. Co-immunized of IL-5 and mZP3 induced the highest sIgA compare to other immunized groups. But such co-administration did not reduce the fertility rate significantly. These data suggest that IL-5 alone or co-immunized with GM-CSF can enhance mZP3 DNA induced antibody response, but the immunocontraception effect should be further studied.Conclusion:In our study, plasmid GM-CSF, IL-5 and IL-4 were used for improving the immunocontraception of mZP3 DNA vaccine. For system immunization, GM-CSF 4d prior to IL-5+mZP3 administration is a potential strategy for development of immunocontraceptive vaccine; for mucosal immunization, GM-CSF 4d prior to mZP3 DNA vaccination maybe a safe and effective contraceptive vaccine strategy.