Mechanism of Chimeric Vaccine Mediated Immune Suppression of Human Dendritic Cells
Type 1 diabetes mellitus is a chronic inflammatory disease in which insulin producing β-cells of the pancreatic islets are killed by autoreactive cells of the immune system in response to a loss of tolerance. Dendritic cells (DC) interact predominantly with naïve T cells to regulate the delicate balance between immunity and tolerance required to maintain immunological homeostasis. In this dissertation, immature human dendritic cells (iDC) were inoculated with a chimeric fusion protein vaccine containing the pancreatic β-cell auto-antigen proinsulin linked to a mucosal adjuvant the cholera toxin B subunit (CTB-INS). Proteomic analysis of vaccine inoculated DCs revealed strong up-regulation of the tryptophan catabolic enzyme indoleamine 2, 3-dioxygenase (IDO1) shown to be directly linked to CTB-INS-induced DC activation and maturation. Treatment of vaccinated DCs with the pharmacological NF-κB inhibitors ACHP or DHMEQ inhibited IDO1 biosynthesis, suggesting a role for NF-κB signaling in CTB-INS vaccine up-regulation of dendritic cell IDO1. Further, blocking the NF-κB-inducing kinase (NIK) phosphorylation of IKK-α dimers and translocation of p52/RelB complexes to the nucleus with anti-NIK siRNA, significantly inhibited IDO1 expression in human DCs. Chromatin immunoprecipitation (ChIP) analysis, showed that RelB-p52 dimers bound dendritic cell NF-κB consensus sequences in the IDO1 promoter region, demonstrating vaccine stimulation of NF-κB non-canonical pathway activation of IDO1 expression in human DCs in vivo. Addition of the Tumor Necrosis Factor Associated Factors (TRAF) TRAF6 and TRAF 2, 3 blocking peptides to vaccinated DCs dramatically reduced the level of IDO1 biosynthesis and stimulated DC suggesting the vaccine may function in the same signaling pathway as TNFR super-family for the up-regulation of IDO1 biosynthesis in vaccinated dendritic cells. Together, these data confirm CTB-INS vaccine activation of the non-canonical NF-κB signaling pathway TNFR receptor family up-regulation of dendritic cell IDO1 biosynthesis permitting vaccine inhibition of DC maturation leading to the suppression of type 1 diabetes development.
Cellular and Molecular Physiology, Medical Biochemistry, Medical Cell Biology, Medical Immunology, Medical Physiology, Physiology, Dendritic Cells; Immunity; Cellular; Cell Differentiation; Diabetes Mellitus; Type 1;, Immunological Homeostasis; Chimeric Fusion Protein Vaccine; Vaccine inhibitation;
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