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Induction of indoleamine 2, 3-dioxygenase in human dendritic cells by a cholera toxin B subunit-proinsulin vaccine.

Mbongue JC, Nicholas DA, Zhang K, Kim NS, Hamilton BN, Larios M, Zhang G, Umezawa K, Firek AF, Langridge WH - PLoS ONE (2015)

Bottom Line: Vaccination did not interfere with monocytes differentiation into DC, suggesting the vaccine can function safely in the human immune system.Together, our experimental data indicate that CTB-INS vaccine induction of IDO1 biosynthesis in human DCs may result in the inhibition of DC maturation generating a durable state of immunological tolerance.Understanding how CTB-INS modulates IDO1 activity in human DCs will facilitate vaccine efficacy and safety, moving this immunosuppressive strategy closer to clinical applications for prevention of type 1 diabetes autoimmunity.

View Article: PubMed Central - PubMed

Affiliation: Center for Health Disparities and Molecular Medicine, Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, CA, United States of America; Loma Linda University School of Medicine, Department of Basic Sciences, Division of Physiology, Loma Linda, CA, United States of America.

ABSTRACT
Dendritic cells (DC) interact with naïve T cells to regulate the delicate balance between immunity and tolerance required to maintain immunological homeostasis. In this study, 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). Increased biosynthesis of the immunosuppressive enzyme was detected in DCs inoculated with the CTB-INS fusion protein but not in DCs inoculated with proinsulin, CTB, or an unlinked combination of the two proteins. Immunoblot and PCR analyses of vaccine treated DCs detected IDO1mRNA by 3 hours and IDO1 protein synthesis by 6 hours after vaccine inoculation. Determination of IDO1 activity in vaccinated DCs by measurement of tryptophan degradation products (kynurenines) showed increased tryptophan cleavage into N-formyl kynurenine. Vaccination did not interfere with monocytes differentiation into DC, suggesting the vaccine can function safely in the human immune system. Treatment of vaccinated DCs with pharmacological NF-κB inhibitors ACHP or DHMEQ significantly inhibited IDO1 biosynthesis, suggesting a role for NF-κB signaling in vaccine up-regulation of dendritic cell IDO1. Heat map analysis of the proteomic data revealed an overall down-regulation of vaccinated DC functions, suggesting vaccine suppression of DC maturation. Together, our experimental data indicate that CTB-INS vaccine induction of IDO1 biosynthesis in human DCs may result in the inhibition of DC maturation generating a durable state of immunological tolerance. Understanding how CTB-INS modulates IDO1 activity in human DCs will facilitate vaccine efficacy and safety, moving this immunosuppressive strategy closer to clinical applications for prevention of type 1 diabetes autoimmunity.

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Diagram of upstream signals that regulate IDO1 biosynthesis in CTB-INS vaccinated human DCs.Signaling pathways involved in vaccine up-regulation of IDO1 biosynthesis in human DCs were identified by Ingenuity Pathway analysis of proteins detected in vaccinated DCs by mass spectrometer analysis. (Red: up-regulation of the protein, Green: down-regulation of the protein, Grey: protein levels unchanged, White: Undetected). Based on calculations generated from the mass spectrometry data, the IPA program indicates the vaccine up-regulates the aryl hydrocarbon receptor (AhR) signal transduction pathways and down regulates TYROBP to increase IDO biosynthesis. NF-κB1: is the P105 precursor subunit of NF-κB which is cleaved to generate the P50 subunit of the canonical NF-κB transcriptional activator protein, located in the cytoplasm. NF-κB2: is the P100 subunit of non-canonical NF-κB, cleaved to generate the p52 subunit, located in the cytoplasm.
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pone.0118562.g005: Diagram of upstream signals that regulate IDO1 biosynthesis in CTB-INS vaccinated human DCs.Signaling pathways involved in vaccine up-regulation of IDO1 biosynthesis in human DCs were identified by Ingenuity Pathway analysis of proteins detected in vaccinated DCs by mass spectrometer analysis. (Red: up-regulation of the protein, Green: down-regulation of the protein, Grey: protein levels unchanged, White: Undetected). Based on calculations generated from the mass spectrometry data, the IPA program indicates the vaccine up-regulates the aryl hydrocarbon receptor (AhR) signal transduction pathways and down regulates TYROBP to increase IDO biosynthesis. NF-κB1: is the P105 precursor subunit of NF-κB which is cleaved to generate the P50 subunit of the canonical NF-κB transcriptional activator protein, located in the cytoplasm. NF-κB2: is the P100 subunit of non-canonical NF-κB, cleaved to generate the p52 subunit, located in the cytoplasm.

Mentions: The mass spectrometry proteome data from vaccine treated DCs was compared with a network of upstream regulators of IDO1 identified by IPA. The experimental results indicated that vaccine induced expression of IDO1 mRNA may be dependent not only on NF-κB, but also on AhR and STAT1 transcriptional activators as well as vaccine-induced down-regulation of the tyrosine kinase binding protein (TYROBP) [42–44] (Fig. 5).


Induction of indoleamine 2, 3-dioxygenase in human dendritic cells by a cholera toxin B subunit-proinsulin vaccine.

Mbongue JC, Nicholas DA, Zhang K, Kim NS, Hamilton BN, Larios M, Zhang G, Umezawa K, Firek AF, Langridge WH - PLoS ONE (2015)

Diagram of upstream signals that regulate IDO1 biosynthesis in CTB-INS vaccinated human DCs.Signaling pathways involved in vaccine up-regulation of IDO1 biosynthesis in human DCs were identified by Ingenuity Pathway analysis of proteins detected in vaccinated DCs by mass spectrometer analysis. (Red: up-regulation of the protein, Green: down-regulation of the protein, Grey: protein levels unchanged, White: Undetected). Based on calculations generated from the mass spectrometry data, the IPA program indicates the vaccine up-regulates the aryl hydrocarbon receptor (AhR) signal transduction pathways and down regulates TYROBP to increase IDO biosynthesis. NF-κB1: is the P105 precursor subunit of NF-κB which is cleaved to generate the P50 subunit of the canonical NF-κB transcriptional activator protein, located in the cytoplasm. NF-κB2: is the P100 subunit of non-canonical NF-κB, cleaved to generate the p52 subunit, located in the cytoplasm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4340906&req=5

pone.0118562.g005: Diagram of upstream signals that regulate IDO1 biosynthesis in CTB-INS vaccinated human DCs.Signaling pathways involved in vaccine up-regulation of IDO1 biosynthesis in human DCs were identified by Ingenuity Pathway analysis of proteins detected in vaccinated DCs by mass spectrometer analysis. (Red: up-regulation of the protein, Green: down-regulation of the protein, Grey: protein levels unchanged, White: Undetected). Based on calculations generated from the mass spectrometry data, the IPA program indicates the vaccine up-regulates the aryl hydrocarbon receptor (AhR) signal transduction pathways and down regulates TYROBP to increase IDO biosynthesis. NF-κB1: is the P105 precursor subunit of NF-κB which is cleaved to generate the P50 subunit of the canonical NF-κB transcriptional activator protein, located in the cytoplasm. NF-κB2: is the P100 subunit of non-canonical NF-κB, cleaved to generate the p52 subunit, located in the cytoplasm.
Mentions: The mass spectrometry proteome data from vaccine treated DCs was compared with a network of upstream regulators of IDO1 identified by IPA. The experimental results indicated that vaccine induced expression of IDO1 mRNA may be dependent not only on NF-κB, but also on AhR and STAT1 transcriptional activators as well as vaccine-induced down-regulation of the tyrosine kinase binding protein (TYROBP) [42–44] (Fig. 5).

Bottom Line: Vaccination did not interfere with monocytes differentiation into DC, suggesting the vaccine can function safely in the human immune system.Together, our experimental data indicate that CTB-INS vaccine induction of IDO1 biosynthesis in human DCs may result in the inhibition of DC maturation generating a durable state of immunological tolerance.Understanding how CTB-INS modulates IDO1 activity in human DCs will facilitate vaccine efficacy and safety, moving this immunosuppressive strategy closer to clinical applications for prevention of type 1 diabetes autoimmunity.

View Article: PubMed Central - PubMed

Affiliation: Center for Health Disparities and Molecular Medicine, Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, CA, United States of America; Loma Linda University School of Medicine, Department of Basic Sciences, Division of Physiology, Loma Linda, CA, United States of America.

ABSTRACT
Dendritic cells (DC) interact with naïve T cells to regulate the delicate balance between immunity and tolerance required to maintain immunological homeostasis. In this study, 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). Increased biosynthesis of the immunosuppressive enzyme was detected in DCs inoculated with the CTB-INS fusion protein but not in DCs inoculated with proinsulin, CTB, or an unlinked combination of the two proteins. Immunoblot and PCR analyses of vaccine treated DCs detected IDO1mRNA by 3 hours and IDO1 protein synthesis by 6 hours after vaccine inoculation. Determination of IDO1 activity in vaccinated DCs by measurement of tryptophan degradation products (kynurenines) showed increased tryptophan cleavage into N-formyl kynurenine. Vaccination did not interfere with monocytes differentiation into DC, suggesting the vaccine can function safely in the human immune system. Treatment of vaccinated DCs with pharmacological NF-κB inhibitors ACHP or DHMEQ significantly inhibited IDO1 biosynthesis, suggesting a role for NF-κB signaling in vaccine up-regulation of dendritic cell IDO1. Heat map analysis of the proteomic data revealed an overall down-regulation of vaccinated DC functions, suggesting vaccine suppression of DC maturation. Together, our experimental data indicate that CTB-INS vaccine induction of IDO1 biosynthesis in human DCs may result in the inhibition of DC maturation generating a durable state of immunological tolerance. Understanding how CTB-INS modulates IDO1 activity in human DCs will facilitate vaccine efficacy and safety, moving this immunosuppressive strategy closer to clinical applications for prevention of type 1 diabetes autoimmunity.

Show MeSH
Related in: MedlinePlus