Study Of Food Potential Allergenicity And Allergy Mechanism Using Animal Model

The increasing incidence make food allergy become an international research hotspot. In recent years, people have been exploring to ideal animal model of sensitization to evaluate many kinds of new food allergenic potential, and to study the related mechanism of food sensitization. However, at present, people are mostly research for single food sensitization protein (such as OVA, PNA) sensitization with animal, while ignoring the effects of proteins’ mutual interaction in food on allergenicity. In this research, we chose three kind of food protein extracts from peanut, milk and potato to sensitize BALB/c mice and studied an allergic murine model that can recognize potential allergenicity of different food. And on this basis, peanut protein extract were used to oral administrate C3H/HeJ mice whose immune system is more sensitive. The digital gene expression analysis method (DGE) was then used to investigate the sensitization mechanism and the influence of non-depletion anti-CD4monoclonal antibody on the allergy reaction. The main results and conclusions are as follows:(1) The BALB/c mice were orally sensitized with PPE, CME and PAPE respectively. Animals in PPE treatment group produced highest specific IgE, Thl-and Th2-cell, also the most severe aggregation and degranulation of mast cells in spleen. The levels of immune reaction of mice in CME group were lower than that in PPE group. The PAPE almost did not induce sensitization response in mice. This suggested that the chosen BALB/c mouse and sensitization method can be used to distinguish the different levels of potential allergenicity of food protein extracts.(2) DNAStar and NetMHCⅡ2.2online Server were used to analyse and predict the potential allergenicity of PPE, CME and PAPE. There are11kind of known allergic proteins in PPE, the Ara h1, Ara h2, Ara h3and Ara h5contain much epitopes. CME contains three known sensitization protein and epitopes only exist in the β-LG. On the contract, there is no epitopes in the four known sensitized protein in PAPE. The results of simulated gastric fluid digestion experiment suggest that the molecular weight of anti-digestion allergen in PPE were17KD and7KD,β-LG is the only anti-digested protein in CME, and the PAPE was completely digested. These results demonstrated that the potential allergenicity of the three protein extracts was PPE> CME> PAPE, this further validate the results obtained from BALB/c mice allergic experiment.(3) Orally sensitize C3H/HeJ mouse with PPE and explore the allergic related mechanism on molecular level using DGE method. The results appeared81differentially expressed genes were identified and contained30genes that are associated with immune system. These genes are mainly related to the inhibition of antigen-presenting function, immune cell migration and adhesion, promotion of T cell activation signal and enhancement of the B cell receptor (BCR) signal. This suggested that PPE processing activates the adoptive immune response of animal.(4) Non-depletion anti-CD4monoclonal antibody was used to i.v treated C3H/HeJ mice at the same time of sensitization. We fund it inhibited the allergy reactions on three aspects of humoral immunity, cellular immunity and systemic symptoms. DGE was also used to investigate the mechanisms of this affluence.131differentially expressed genes containing35related to immune system were identified. The35genes were primarily involve of macrophage activation and the innate immune response, T cells and B cells mature, inhibition of proliferation and antibody production, and to promotion of specific Treg cells. Anti-CD4antibodies not only inhibit the adoptive immune responses of animals to the PPE, also promoted the specific tolerance and moderation of innate immune response.