Proteomic Analysis and Immunoregulation Mechanism of Wheat Germ Globulin

Background: Wheat germ, one of the byproducts of flour milling, contains abundant physiologically active components. Globulins in wheat germ are a class of high-quality functional proteins and have received widespread attention. However, the composition of wheat germ globulin（ WGG）and the structure of the typical proteins have not yet been proved. The immunological activities and immune mechanisms of the WGG have not yet been revealed in vivo. Objectives: The proteomic analysis of WGG and the structure simulation of typical proteins were studied. The immunoregulatory effects of WGG on immunosuppressed mice induced by cyclophosphamide were investigated, and the immunological activities of WGG were explored. Methods: The main components, functions, and metabolic signaling pathways of WGG were analyzed through a combination of LC-MS method and bioinformatics. The structure of WGG was predicted via the Phyre2 tool. Immunosuppression in mice was induced by cyclophosphamide. After an intraperitoneal injection of WGG for 10 days, organ indexes and pathological changes of mice were detected. The T-cell subgroups in peripheral blood were analyzed via flow cytometry. Levels of IL-2, IL-4, TNF-α, and IFN-γ were evaluated through ELISA. The mRNA expression levels of T-Bet and GATA-3 were measured using real-time PCR. Results: The results indicated that the main functional components of WGG were wheat germ globulins, histones, heat shock proteins (HSPs), and other functional proteins. Wheat germ globulins and HSPs were the major immune components of WGG. WGG significantly reduced immunosuppression in the spleen and thymus indexes (P<0.01), and mitigated the damage caused by cyclophosphamide in the spleen and thymus. Moreover, WGG significantly increased the CD4+/CD8+ of the immunosuppressed mice (P<0.01), restored Th1/Th2 imbalance (P < 0.01), enhanced the content of IL-2 and IL-4 (P<0.01), and modified the abnormal secretion of cytokines. WGG also observably reduced the mRNA expression of T-Bet and GATA-3 (P<0.01). These results manifested that WGG components improved the immune system. The action mechanisms might be related to the variation of Th1/Th2 cells resulted from the control of the mRNA expression levels of T-Bet and GATA-3. Conclusion: The wheat germ histone family and the HSPs are the major immune components of WGG. It may be the immune mechanism of WGG that these globulins affect the differentiation of Th1/Th2 cells via controlling the mRNA expression levels of related genes. The results indicated the potential application of WGG or its further purified products as a superior plant-derived immunomodulator in the future. Keywords: Immune mechanism, T-Bet/GATA-3, Th1/Th2 cells, WGG, HSPs, mRNA expression.