Separation And Investigation On Neospora Caninum Soluble Complexes Which Induce TH-1 Immune Response

Neospora caninum is an intracellular parasites, the main damage to the livestock industry is causing pregnant cows and calves in abortion. For few of the newborn calf, there are no obvious clinical symptoms, but they had been infected before they were born.In present, there is no drug able to resist or eliminate the parasite stayed in brain or muscle in cattle. The commercialized attenuated vaccines now used in Neospora caninum prevention, not only prevention effects was unsatisfactory, but also has a bio-safety issue. Therefore, the safe and efficient Neospora Caninum vaccines developing become increasingly prominent practical problems. So, how to select a vaccine candidate and how to present antigen effectively become a research pot. Studies have shown that Neospora caninum multiply in cells, so the cell-mediated immune response in the host protective immune response is very important. Cell-mediated host protective immune response mainly produced IFN-γ. IL-12 and TNF-α. In these cytokines, the IFN-y is to inhibit the propagation of intracellular parasites has played a crucial role. In theory, the antigen recognized by immune sera and the immune T cell can be considered effective vaccine candidate antigen. On the other side, the Neospora caninum components which can activate CD4+T cells to produce IFN-γcan be used as a vaccine candidate. Therefore, based on the above viewpoints, this study focused on searching for the components in soluble antigen which can develop host protective immunity to the type-1 immune response, and try to clarify its mechanism.The experiment is divided into three parts:the different treatment of Neospora and Neospora Caninum antigen stimulate immune cells in vitro; try to find the Neospora components in Neospora soluble antigen, which can develop host immune response to the type-1 immune response; the mechanism of DNA/RAN and macromolecular complexes induce the immune response during the stimulation. First, we verified the Neospora Caninum stimulation to immune cells by T lymphocytes and dendritic cells in vitro, all which can induce the immune response to type-1. and frozen-killed parasites can develop better than others. All Neospora Caninum antigen stimulation for immune cells has a dose-dependent. Second, the Neospora Caninum antigen was fractionated by HPLC-SEC and we found the fractionated NcAg 27-33 can induce the immune cells to produce the IFN-γstrongly. At the same time, we proved the Neospora Caninum DNA involved in the inducing of type-1 immune response. The results of the enzymatic digestion experiments, immune cells stimulating experiments and DNA transfer experiments proved that the other proteins in fractionated 27-33 were involved in the responding of immune cells for Neospora Caninum DNA stimulating. The singular Neospora Caninum DNA can not effectively induce immune responses; 293-TLR9 cells stimulating experiment confirmed that TLR-9 pathway involved in the immune response developed by Neospora Caninum DNA. The mechanism of the proteins in fr 27-33 induce the type-1 immune response still unclear. The structure and activity of those proteins need to be further analyzed and identified.