Investigation Of Immune Responses Induced By Recombinant VP1-VP4Protein Of Foot-and-mouth Disease Virus

Foot-and-mouth disease (FMD) is an acute, febribible and highly contagiousviral disease of cloven-hoofed animals caused by foot-and-mouth disease virus(FMDV). There is not a safe and effective control approach for this disease yet. Theclearance of FMDV needs activated CD8+T cells, Th1response and high level ofneutrilizing antibody. As for an exogenous antigen, the activation of CD8+T cellsneeds CD8α+DCs in terms of cross-presentaion. In vitro assay, lymph node T cellsreleased vigorously IFN-γ when incubated with SR-A inhibited dendritic cells pulsedwith recombinant FMDV VP1-VP4antigen, indicating SR-A plays a negtive role inrecognizing VP1-VP4and priming cellular immune response by DCs. For thisfunction of SR-A in vitro assay, we blocked the combination of SR-A with its ligandsby intraperitoneal injection of sennoside B and/or tannic acid. Given that CD8α+DCsis capable of cross-presenting soluble antigens to activate CD8+T cells, cytochrome cwas administed through intravenous injection to deplete CD8α+DCs. Theexperimental vaccine, which consists of FMDV VP1-VP4protein and IMS251cVGadjuvant, is subcutaneously injected into mice pre-treated above. Na ve micevaccinated with recombinant VP1-VP4or commertially inactivated FMDV vaccine ascontrols. By meaturing the contents of serum IFN-γ, IL-4, total antibody titer and theserum antibody isotype classes and/or subclasses the profile of immune responses ofVP1-VP4was assessed.The results indicate that both serum IFN-γ and IL-4levels of VP1-VP4vaccinated mice were lower than that of mice administed with curmercial inactivatedFMDV vaccine. Moreover, serum IFN-γ level significantly decreaseed（P＜0.05）,serum IL-4level decreased unsignificantly(P＞0.05). Compared to VP1-VP4vaccinated mice, the level of IL-4is significantly downregulated（P＜0.05）while theIFN-γ content is unsignificantly reduced(P＞0.05) in SR-A inhibited mice. Comparedto of VP1-VP4vaccinated mice, the level of IL-4is significantly reduced（P＜0.05）while the IFN-γ content is unsignificantly reduced(P＞0.05) under the circumstanceof CD8α+DCs depletion in vivo.The antibody levels of mice were assayed with two-dimensionalimmunodiffusion test. It was found that both mice serum total antibody titer of SR-Ainhibitor treatment groups and splenic CD8+DCs depletion groups were significantlylower（P＜0.05）, except tanic acid pretreatment group (P＞0.05) compared with theVP1-VP4vaccinated mice. The total serum antibody titer of mice induced byVP1-VP4protein was lower than that of mice induced by commercial inactivatedvaccine. However, there is no significant difference beween the two groups (P> 0.05).ELASA assay results showed that the serum IgG1, IgG2b, κ type and λ typelight chain levels decreased in SR-A inhibited mice than that of VP1-VP4vaccinatedmice, while the content of serum IgG2a, IgG3, IgA and IgM increased than that ofVP1-VP4vaccinated mice. Importantly, IgA levels in serum raised significantly（P＜0.05). However, there is no difference between any other groups and only VP1-VP4vaccinated group (P>0.05). Comparing to VP1-VP4protein injected mice, the serumIgG1, IgG2b, IgG3, IgM, κ type and λ type light chain contents of CD8α+DCsdepleted mice were lower，except κ type light chain is significantly lower（P＜0.05)and λ type light chain is very significantly lower（P＜0.01), but not significantly.Thecontens of IgG2a and IgA raised but with no significant difference among the groups.All results above indicate that FMDV VP1-VP4protein can be recognized byCD8α+DCs in way of SR-A. Herein CD8α+DCs activate CD4+T cells mainlythrough MHC-II molecules and initiate cellular immune response in feature ofproducing IL-4. Interestingly, the outcome of immune responses in mice pretreated bySR-A inhibitor is very similar to that of CD8α+DC depleted mice, indicating CD8α+DCs may be the cardinal SR-A expressed cells and also be the main sennoside B ortannic acid targeted cells. Given that IL-4levels of FMDV infected cattle is higherthan that of FMDV uninfected cattle and SR-A inhibition or CD8α+DCs depletion invivo lead to the significant downregulation of IL-4response and the upregulation ofserum IgA, It is reasonable to argue that sennoside B and/or tannic acid can beutilized as adjuvant elements of recombinant FMDV VP1-VP4vaccine.