Evaluation of γ-Irradiated Peanut Extract as a Proper Immunogen for Immunotherapy in Murine Model of Peanut Allergy

BACKGROUND AND OBJECTIVES: Peanut allergy is one of the most serious forms of IgE-mediated food hypersensitivity, and gamma irradiation has been applied widely for the preservation of food. Previous studies indicated that the capacity of IgE to binds to ovalbumin (OVA) and ovamucoid (OM) was profoundly reduced after treatment with a combination of gamma irradiation and other processing treatments, and that this reduction in binding capacity was coupled with reduced OVA and OM skin reactions on skin prick tests. Therefore, this study was conducted to evaluate the feasibility of using the gamma irradiation technique to reduce peanut allergies by observing the changes that occur in the allergenecity and antigencity of gamma irradiated peanut proteins. MATERIALS & METHODS: 1) The physicochemical and imunochemical properties of gamma-irradiated peanuts - Peanut extract that had been gamma-irradiated at 5, 10, 20 and 50 kGy in aqueous solution were evaluated using SDS-PAGE, immunoblotting ELISA, and competitive indirect enzyme-linked immunosorbent assay (Ci-ELISA) with sera from peanut hypersensitive patients. 2) The effect of splenocyte stimulation with gamma-irradiated peanuts in mice that were allergic to peanuts - Mice were sensitized to peanuts by intragastric exposure to non-irradiated PN extracts at day 0, 1, 2, and 7 and then challenged at day 21. Four weeks later, we evaluated the cytokine production patterns and proliferation responses of splenocytes which had been stimulated with 0 kGy non-irradiated peanuts, 10 kGy irradiated peanuts, and 50 kGy irradiated peanuts. 3) The effect of intragastric administration of gamma-irradiated peanuts in mice – Mice were sensitized with non-irradiated PN extracts and irradiated PN extracts intragastricly at day 1, 2, 3, and 7 and then challenged them at day 21. Mice in groupⅠ(n=5) received crude PN (0 kGy), whereas mice in group Ⅱ (n=5) received 10 kGy irradiated PN, mice in group Ⅲ (n=5) received 50 kGy irradiated PN, and mice in group Ⅳ (n=5) were the naive control. We then analyzed the PN-specific serum IgE, IgG1, IgG2a antibody levels, PN-stimulated splenocyte cytokine production patterns and splenocyte proliferation responses. RESULTS: 1) The physicochemical and imunochemical properties of gamma-irradiated peanuts - The disappearance of the protein band after sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the structure of the peanut proteins changed after radiation treatment, and this change may have occurred as a result of fragmentation and/or aggregation of the proteins. In addition, the use of immunoglobulin E (IgE) from peanut-hypersensitive patients allowed the changes in the allergenicity and antigenicity of irradiated proteins to be observed by PN specific IgE immunoblotting and competitive indirect enzyme-linked immunosorbent assay (Ci-ELISA). The results of these analyses indicated that epitopes on the peanut allergens were structurally altered by gamma irradiation. 2) Effect of splenocyte stimulation with gamma-irradiated peanuts in mice that were allergic to peanuts – Gamma-irradiated PN-stimulated productions of IL-10 and IFN-γ were increased when compared to the non-irradiated PN-stimulated production. In addition, the Th1 / Th2 ratio increased in response to treatment with gamma-irradiated peanuts. 3) Effect of intragastric administration of gamma-irradiated peanuts in mice – The PN-specific IgE level was decreased in group Ⅲ (50 kGy irradiated peanut treatment) when compared to that of group Ⅰ(0 kGy non-irradiated peanut treatment). However, the PN-specific IgE level was increased at week 4 in group Ⅱ (10 kGy irradiated peanut treatment) when compared to group Ⅰ. Furthermore, the PN stimulated IL-4 and IL-10 levels in the 72 hour splenocyte culture supernatant were decreased in group Ⅲ, whereas the PN-stimulated production of IFN-γ was increased group Ⅱ and Ⅲ. CONCLUSION: The binding ability of patients’s IgE to irradiated peanut decreased depending on the dose. Also, SDS-PAGE and an immunoblotting assay revealed that fragmentation of the proteins had occurred, which shows that a structural change may reduce the binding capacity for the epitopes in ELISA. In murine experiment, that stimulation of the splenocytes of mice that were allergic to peanuts and mice themselves by gamma-irradiated peanuts was found to increase the Th1 / Th 2 ratio and IL-10 production. Therefore, this study showed the possibility of developing an immunogen capable of treatment and reduction of food allergy by using gamma-radiation technology. Future studies are required to further evaluate immunotherapy using irradiated peanuts in a murine model of peanut allergy.

Author dc:creator

• 오, 세조

Contributors dc:contributor

• 이, 수영
• 대학원 의학과
• 200424317

• Peanut allergy
• γ-irradiation
• Murine model
• Cytokine
• Immunotherapy
• 땅콩 알레르기
• 생쥐모델
• 감마선 조사
• 사이토카인
• 면역치료

Language dc:language

en

Identifier

http://dcoll.ajou.ac.kr:9080/dcollection/jsp/common/DcLoOrgPer.jsp?sItemId=000000005819
000000005819

OAI identifier oai:identifier

oai:repository.ajou.ac.kr:201003/1401