RESUMEN
Cholera toxin B subunit fusion to autoantigens such as proinsulin (CTB-INS) down regulate dendritic cell (DC) activation and stimulate synthesis of DC immunosuppressive cytokines. Recent studies of CTB-INS induction of immune tolerance in human DCs indicate that increased biosynthesis of indoleamine 2,3-dioxygenase (IDO1) may play an important role in CTB-INS vaccine suppression of DC activation. Studies in murine models suggest a role for transforming growth factor beta (TGF-ß) in the stimulation of IDO1 biosynthesis, for the induction of tolerance in DCs. Here, we investigated the contribution of TGF-ß superfamily proteins to CTB-INS induction of IDO1 biosynthesis in human monocyte-derived DCs (moDCs). We show that CTB-INS upregulates the level of TGF-ß1, activin-A and the TGF-ß activator, integrin αvß8 in human DCs. However, inhibition of endogenous TGF-ß, activin-A or addition of biologically active TGF-ß1, and activin-A, did not inhibit or stimulate IDO1 biosynthesis in human DCs treated with CTB-INS. While inhibition with the kinase inhibitor, RepSox, blocked SMAD2/3 phosphorylation and diminished IDO1 biosynthesis in a concentration dependent manner. Specific blocking of the TGF-ß type 1 kinase receptor with SB-431542 did not arrest IDO1 biosynthesis, suggesting the involvement of a different kinase pathway other than TGF-ß type 1 receptor kinase in CTB-INS induction of IDO1 in human moDCs. Together, our experimental findings identify additional immunoregulatory proteins induced by the CTB-INS fusion protein, suggesting CTB-INS may utilize multiple mechanisms in the induction of tolerance in human moDCs.
Asunto(s)
Células Dendríticas/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Indolamina-Pirrol 2,3,-Dioxigenasa/biosíntesis , Proteínas Recombinantes de Fusión/farmacología , Factor de Crecimiento Transformador beta1/genética , Activinas/genética , Activinas/inmunología , Animales , Diferenciación Celular , Toxina del Cólera/genética , Toxina del Cólera/inmunología , Clonación Molecular , Células Dendríticas/citología , Células Dendríticas/inmunología , Escherichia coli/genética , Escherichia coli/metabolismo , Humanos , Indolamina-Pirrol 2,3,-Dioxigenasa/genética , Indolamina-Pirrol 2,3,-Dioxigenasa/inmunología , Integrinas/genética , Integrinas/inmunología , Ratones , Monocitos/citología , Monocitos/efectos de los fármacos , Monocitos/inmunología , Cultivo Primario de Células , Proinsulina/genética , Proinsulina/inmunología , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/inmunología , Pirazoles/farmacología , Piridinas/farmacología , Receptor Tipo I de Factor de Crecimiento Transformador beta , Receptores de Factores de Crecimiento Transformadores beta/genética , Receptores de Factores de Crecimiento Transformadores beta/inmunología , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/inmunología , Transducción de Señal , Proteína Smad2/genética , Proteína Smad2/inmunología , Proteína smad3/genética , Proteína smad3/inmunología , Factor de Crecimiento Transformador beta1/inmunologíaRESUMEN
Transforming growth factor beta (TGF-ß) is a pleiotropic cytokine present in vertebrate and invertebrate organisms that functions in numerous physiological and pathological processes. TGF-ß impacts all the cells of the immune system, and of the three known TGF-ß isoforms, TGF-ß1 is the predominant isoform expressed in immune cells. TGF-ß1 is known to play a pivotal role in the function of all immune cells especially in the regulation of T cell development and in the induction of immunological tolerance in dendritic cells (DCs). Based on the importance of DCs in regulation of the innate and adaptive arms of the immune system, in this review we explore the regulatory functions of TGF-ß required for establishment and maintenance of DC-mediated immune tolerance.
Asunto(s)
Células Dendríticas/inmunología , Isoformas de Proteínas/inmunología , Linfocitos T Reguladores/inmunología , Factor de Crecimiento Transformador beta/inmunología , Animales , Diferenciación Celular , Humanos , Tolerancia Inmunológica , Inmunidad , Transducción de SeñalRESUMEN
A chimeric protein vaccine composed of the cholera toxin B subunit fused to proinsulin (CTB-INS) was shown to suppress type 1 diabetes onset in NOD mice and upregulate biosynthesis of the tryptophan catabolic enzyme indoleamine 2, 3-dioxygenase (IDO1) in human dendritic cells (DCs). Here we demonstrate siRNA inhibition of the NF-κB-inducing kinase (NIK) suppresses vaccine-induced IDO1 biosynthesis as well as IKKα phosphorylation. Chromatin immunoprecipitation (ChIP) analysis of CTB-INS inoculated DCs showed that RelB bound to NF-κB consensus sequences in the IDO1 promoter, suggesting vaccine stimulation of the non-canonical NF-κB pathway activates IDO1 expression in vivo. The addition of Tumor Necrosis Factor Associated Factors (TRAF) TRAF 2, 3 and TRAF6 blocking peptides to vaccine inoculated DCs was shown to inhibit IDO1 biosynthesis. This experimental outcome suggests vaccine activation of the TNFR super-family receptor pathway leads to upregulation of IDO1 biosynthesis in CTB-INS inoculated dendritic cells. Together, our experimental data suggest the CTB-INS vaccine uses a TNFR-dependent signaling pathway of the non-canonical NF-κB signaling pathway resulting in suppression of dendritic cell mediated type 1 diabetes autoimmunity.
Asunto(s)
Células Dendríticas/efectos de los fármacos , Diabetes Mellitus Tipo 1/terapia , Indolamina-Pirrol 2,3,-Dioxigenasa/inmunología , FN-kappa B/inmunología , Vacunas/administración & dosificación , Secuencia de Aminoácidos , Animales , Autoinmunidad/efectos de los fármacos , Secuencia de Bases , Toxina del Cólera/biosíntesis , Toxina del Cólera/genética , Toxina del Cólera/inmunología , Células Dendríticas/inmunología , Células Dendríticas/patología , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/inmunología , Diabetes Mellitus Tipo 1/patología , Regulación de la Expresión Génica , Humanos , Quinasa I-kappa B/genética , Quinasa I-kappa B/inmunología , Quinasa I-kappa B/metabolismo , Indolamina-Pirrol 2,3,-Dioxigenasa/genética , Indolamina-Pirrol 2,3,-Dioxigenasa/metabolismo , Ratones , Ratones Endogámicos NOD , Datos de Secuencia Molecular , FN-kappa B/genética , FN-kappa B/metabolismo , Proinsulina/biosíntesis , Proinsulina/genética , Proinsulina/inmunología , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/inmunología , Proteínas Serina-Treonina Quinasas/metabolismo , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Proteínas Recombinantes de Fusión/biosíntesis , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/inmunología , Transducción de Señal , Factor 2 Asociado a Receptor de TNF/farmacología , Factor 3 Asociado a Receptor de TNF/farmacología , Factor 6 Asociado a Receptor de TNF/farmacología , Quinasa de Factor Nuclear kappa BRESUMEN
Survivin is one of the most important members of the inhibitors of apoptosis protein family, as it is expressed in most human cancers but is absent in normal, differentiated tissues. Lending to its importance, survivin has proven associations with apoptosis and cell cycle control, and has more recently been shown to modulate the tumor microenvironment and immune evasion as a result of its extracellular localization. Upregulation of survivin has been found in many cancers including breast, prostate, pancreatic, and hematological malignancies, and it may prove to be associated with the advanced presentation, poorer prognosis, and lower survival rates observed in ethnically diverse populations.
