RESUMEN
The immunoregulatory cation channel TMEM176B plays a dual role in tumor immunity. On the one hand, TMEM176B promotes antigen cross-presentation to CD8+ T cells by regulating phagosomal pH in dendritic cells (DCs). On the other hand, it inhibits NLRP3 inflammasome activation through ionic mechanisms in DCs, monocytes and macrophages. We speculated that formulating BayK8644 in PEGylated chitosan nanoparticles (NP-PEG-BayK8644) should slowly release the compound and by that mean avoid cross-presentation inhibition (which happens with a fast 30 min kinetics) while still triggering inflammasome activation. Chitosan nanocarriers were successfully obtained, exhibiting a particle size within the range of 200 nm; they had a high positive surface charge and a 99 % encapsulation efficiency. In in vitro studies, NP-PEG-BayK8644 did not inhibit antigen cross-presentation by DCs, unlike the free compound. The NP-PEG-BayK8644 activated the inflammasome in a Tmem176b-dependent manner in DCs. We administered either empty (eNP-PEG) or NP-PEG-BayK8644 to mice with established tumors. NP-PEG-BayK8644 significantly controlled tumor growth and improved mice survival compared to both eNP-PEG and free BayK8644 in melanoma and lymphoma models. This effect was associated with enhanced inflammasome activation by DCs in the tumor-draining lymph node and infiltration of the tumor by CD8+ T cells. Thus, encapsulation of BayK8644 in chitosan NPs improves the anti-tumoral properties of the compound by avoiding inhibition of antigen cross-presentation.
Asunto(s)
Inmunidad Adaptativa , Quitosano , Células Dendríticas , Inmunidad Innata , Nanopartículas , Quitosano/química , Quitosano/farmacología , Animales , Nanopartículas/química , Ratones , Inmunidad Adaptativa/efectos de los fármacos , Células Dendríticas/inmunología , Células Dendríticas/efectos de los fármacos , Inmunidad Innata/efectos de los fármacos , Proteínas de la Membrana/inmunología , Inflamasomas/metabolismo , Línea Celular Tumoral , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/efectos de los fármacos , Ratones Endogámicos C57BL , Melanoma Experimental/inmunología , Melanoma Experimental/patología , Melanoma Experimental/tratamiento farmacológico , Polietilenglicoles/química , Polietilenglicoles/farmacologíaRESUMEN
The prevalence of fragrances in various hygiene products contributes to their sensorial allure. However, fragrances can induce sensitization in the skin or respiratory system, and the mechanisms involved in this process are incompletely understood. This study investigated the intricate mechanisms underlying the fragrance's effects on sensitization response, focusing on the interplay between CYP450 enzymes, a class of drug-metabolizing enzymes, and the adaptive immune system. Specifically, we assessed the expression of CYP450 enzymes and cytokine profiles in culture of BEAS-2B and mature dendritic cells (mDC) alone or in co-culture stimulated with 2 mM of a common fragrance, cinnamyl alcohol (CA) for 20 h. CYP1A1, CYP1A2, CYP1B1, CYP2A6, and CYP2A13 were analyzed by RT-PCR and IL-10, IL-12p70, IL-18, IL-33, and thymic stromal lymphopoietin (TSLP) by Cytometric Bead Array (CBA). Through RT-PCR analysis, we observed that CA increased CYP1A2 and CYP1B1 expression in BEAS-2B, with a further increased in BEAS-2B-mDC co-culture. Additionally, exposure to CA increased IL-12p70 levels in mDC rather than in BEAS-2B-mDC co-culture. In regards to IL-18, level was higher in BEAS-2B than in BEAS-2B-mDC co-culture. A positive correlation between the levels of IL-10 and CYP1B1 was found in mDC-CA-exposed and between IL-12p70 and CYP1A1 was found in BEAS-2B after CA exposure. However, IL-12p70 and CYP1A2 as well as IL-18, IL-33, and CYP1A1 levels were negative, correlated mainly in co-culture control. These correlations highlight potential immunomodulatory interactions and complex regulatory relationships. Overall, exposure to CA enhances CYP450 expression, suggesting that CA can influence immune responses by degrading ligands on xenosensitive transcription factors.
Asunto(s)
Técnicas de Cocultivo , Sistema Enzimático del Citocromo P-450 , Citocinas , Células Dendríticas , Propanoles , Humanos , Citocinas/metabolismo , Células Dendríticas/efectos de los fármacos , Células Dendríticas/metabolismo , Sistema Enzimático del Citocromo P-450/metabolismo , Sistema Enzimático del Citocromo P-450/genética , Propanoles/toxicidad , Propanoles/metabolismo , Línea Celular , Citocromo P-450 CYP1A1/metabolismo , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1B1/genética , Citocromo P-450 CYP1B1/metabolismo , Perfumes/toxicidad , Receptores de Hidrocarburo de Aril/metabolismo , Receptores de Hidrocarburo de Aril/genética , Citocromo P-450 CYP1A2/metabolismo , Citocromo P-450 CYP1A2/genéticaRESUMEN
SUMMARY: One of the reasons for acute kidney damage is renal ischemia. Nevertheless, there are limited protective and therapeutic approaches for this problem. Diacerein is an anti-inflammatory drug characterized by numerous biological activities. We aimed to determine the ameliorative impact of diacerein on renal ischemia/reperfusion injury (I/R) condition, exploring the underlying mechanisms. Twenty-four male rats were allotted into four groups (n= 6): sham group; Diacerein (DIA) group; I/R group, in which a non-crushing clamp occluded the left renal pedicle for 45 min, and the right kidney was nephrectomized for 5 min before the reperfusion process; I/R + diacerein group, injected intraperitoneally with 50 mg diacerein/kg i.m 30 minutes prior to I/R operation. Ischemia/ reperfusion was found to affect renal function and induce histopathological alterations. The flow cytometry analysis demonstrated an elevated expression of innate and mature dendritic cells in I/R renal tissues. Moreover, upregulation in the expression of the inflammatory genes (TLR4, Myd88, and NLRP3), and overexpression of the pro-inflammatory cytokines (IL-1β), apoptotic (caspase-3) and pyroptotic (caspase-1) markers were observed in I/R-experienced animals. The aforementioned deteriorations were mitigated by pre-I/R diacerein treatment. Diacerein alleviated I/R-induced inflammation and apoptosis. Thus, it could be a promising protective agent against I/R.
La isquemia renal es una de los motivos del daño renal agudo. Sin embargo, los enfoques protectores y terapéuticos para este problema son limitados. La diacereína es un fármaco antiinflamatorio caracterizado por numerosas actividades biológicas. Nuestro objetivo fue determinar el impacto de mejora de la diacereína en la condición de lesión por isquemia/ reperfusión renal (I/R), explorando los mecanismos subyacentes. Veinticuatro ratas macho se distribuyeron en cuatro grupos (n= 6): grupo simulado; grupo de diacereína (DIA); grupo I/R, en el que una pinza no aplastante ocluyó el pedículo renal izquierdo durante 45 min, y el riñón derecho fue nefrectomizado durante 5 min antes del proceso de reperfusión; Grupo I/R + diacereína, inyectado por vía intraperitoneal con 50 mg de diacereína/kg i.m. 30 min antes de la operación I/R. Se encontró que la isquemia/ reperfusión afecta la función renal e induce alteraciones histopatológicas. El análisis de citometría de flujo demostró una expresión elevada de células dendríticas innatas y maduras en tejidos renales I/R. Además, se observó una regulación positiva en la expresión de los genes inflamatorios (TLR4, Myd88 y NLRP3) y una sobreexpresión de las citoquinas proinflamatorias (IL-1β), marcadores apoptóticos (caspasa-3) y piroptóticos (caspasa-1) en animales con experiencia en I/R. Los deterioros antes mencionados fueron mitigados por el tratamiento previo a la diacereína I/R. La diacereína alivió la inflamación y la apoptosis inducidas por I/R. Por lo tanto, podría ser un agente protector prometedor contra I/R.
Asunto(s)
Animales , Ratas , Daño por Reperfusión/tratamiento farmacológico , Antraquinonas/administración & dosificación , Enfermedades Renales/tratamiento farmacológico , Antiinflamatorios/administración & dosificación , Células Dendríticas/efectos de los fármacos , Daño por Reperfusión/inmunología , Transducción de Señal , FN-kappa B/metabolismo , Antraquinonas/inmunología , Apoptosis/efectos de los fármacos , Estrés Oxidativo , Receptor Toll-Like 4/metabolismo , Interleucina-1beta/metabolismo , Citometría de Flujo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Inflamación , Inyecciones Intraperitoneales , Enfermedades Renales/inmunologíaRESUMEN
Dendritic cells are antigen-presenting cells, which identify and process pathogens to subsequently activate specific T lymphocytes. To regulate the immune responses, DCs have to mature by the recognition of TLR ligands, TNFα or IFNγ. These ligands have been used as adjuvants to activate DCs in situ or in vitro, with toxic effects. It has been shown that some molecules affect the immune system, e.g., Masticadienonic acid (MDA) and 3α-hydroxy masticadienoic acid (3α-OH MDA) triterpenes naturally occurring in several medicinal plants, since they activate the nitric oxide synthase in macrophages and induce T lymphocyte proliferation. The DCs maturation induced by MDA or 3a-OH MDA was determined by incubating these cells with MDA or 3α-OH MDA, and their phenotype was afterwards analyzed. The results showed that only 3α-OH MDA was able to induce DCs maturation. When mice with melanoma were inoculated with DCs/3α-OH MDA, a decreased tumor growth rate was observed along with an extended cell death area within tumors compared to mice treated with DCs incubated with MDA. In conclusion, it is proposed that 3α-OH MDA may be an immunostimulant molecule. Conversely, it is proposed that MDA may be a molecule with anti-inflammatory properties.
Asunto(s)
Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Factores Inmunológicos/química , Factores Inmunológicos/farmacología , Inmunomodulación/efectos de los fármacos , Triterpenos/química , Triterpenos/farmacología , Animales , Biomarcadores , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células Dendríticas/metabolismo , Modelos Animales de Enfermedad , Inmunofenotipificación , Ratones , Estructura Molecular , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Brucella lumazine synthase (BLS) is a homodecameric protein that activates dendritic cells via toll like receptor 4, inducing the secretion of pro-inflammatory cytokines and chemokines. We have previously shown that BLS has a therapeutic effect in B16 melanoma-bearing mice only when administered at early stages of tumor growth. In this work, we study the mechanisms underlying the therapeutic effect of BLS, by analyzing the tumor microenvironment. Administration of BLS at early stages of tumor growth induces high levels of serum IFN-γ, as well as an increment of hematopoietic immune cells within the tumor. Moreover, BLS-treatment increases the ratio of effector to regulatory cells. However, all treated mice eventually succumb to the tumors. Therefore, we combined BLS administration with anti-PD-1 treatment. Combined treatment increases the outcome of both monotherapies. In conclusion, we show that the absence of the therapeutic effect at late stages of tumor growth correlates with low levels of serum IFN-γ and lower infiltration of immune cells in the tumor, both of which are essential to delay tumor growth. Furthermore, the combined treatment of BLS and PD-1 blockade shows that BLS could be exploited as an essential immunomodulator in combination therapy with an immune checkpoint blockade to treat skin cancer.
Asunto(s)
Interferón gamma/genética , Melanoma Experimental/tratamiento farmacológico , Receptor de Muerte Celular Programada 1/genética , Receptor Toll-Like 4/genética , Animales , Quimiocinas/genética , Citocinas/genética , Células Dendríticas/efectos de los fármacos , Modelos Animales de Enfermedad , Humanos , Inhibidores de Puntos de Control Inmunológico/farmacología , Factores Inmunológicos/farmacología , Melanoma Experimental/genética , Melanoma Experimental/patología , Ratones , Receptor Toll-Like 4/agonistas , Microambiente Tumoral/efectos de los fármacosRESUMEN
Hepatocellular carcinoma (HCC) arises in the setting of advanced liver fibrosis, a dynamic and complex inflammatory disease. The tumor microenvironment (TME) is a mixture of cellular components including cancer cells, cancer stem cells (CSCs), tumor-associated macrophages (TAM), and dendritic cells (DCs), which might drive to tumor progression and resistance to therapies. In this work, we study the effects of 4-methylumbelliferone (4Mu) on TME and how this change could be exploited to promote a potent immune response against HCC. First, we observed that 4Mu therapy induced a switch of hepatic macrophages (MÏ) towards an M1 type profile, and HCC cells (Hepa129 cells) exposed to conditioned medium (CM) derived from MÏ treated with 4Mu showed reduced expression of several CSCs markers and aggressiveness. HCC cells incubated with CM derived from MÏ treated with 4Mu grew in immunosuppressed mice while presented delayed tumor progression in immunocompetent mice. HCC cells treated with 4Mu were more susceptible to phagocytosis by DCs, and when DCs were pulsed with HCC cells previously treated with 4Mu displayed a potent antitumoral effect in therapeutic vaccination protocols. In conclusion, 4Mu has the ability to modulate TME into a less hostile milieu and to potentiate immunotherapeutic strategies against HCC.
Asunto(s)
Carcinoma Hepatocelular/tratamiento farmacológico , Himecromona/farmacología , Neoplasias Hepáticas/tratamiento farmacológico , Microambiente Tumoral/efectos de los fármacos , Animales , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patología , Línea Celular Tumoral , Células Dendríticas/efectos de los fármacos , Modelos Animales de Enfermedad , Resistencia a Antineoplásicos/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica , Humanos , Himecromona/efectos adversos , Inmunidad/efectos de los fármacos , Cirrosis Hepática/tratamiento farmacológico , Cirrosis Hepática/genética , Cirrosis Hepática/patología , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patología , Ratones , Células Madre Neoplásicas/efectos de los fármacos , Fagocitosis/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Macrófagos Asociados a Tumores/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Multiple sclerosis (MS) is a chronic demyelinating disease of the central nervous system (CNS). MS and its animal model called experimental autoimmune encephalomyelitis (EAE) immunopathogenesis involve a plethora of immune cells whose activation releases a variety of proinflammatory mediators and free radicals. Vitamin D3 (VitD) is endowed with immunomodulatory and antioxidant properties that we demonstrated to control EAE development. However, this protective effect triggered hypercalcemia. As such, we compared the therapeutic potential of VitD and paricalcitol (Pari), which is a non-hypercalcemic vitamin D analog, to control EAE. From the seventh day on after EAE induction, mice were injected with VitD or Pari every other day. VitD, but not Pari, displayed downmodulatory ability being able to reduce the recruitment of inflammatory cells, the mRNA expression of inflammatory parameters, and demyelination at the CNS. Lower production of proinflammatory cytokines by lymph node-derived cells and IL-17 by gut explants, and reduced intestinal inflammation were detected in the EAE/VitD group compared to the EAE untreated or Pari groups. Dendritic cells (DCs) differentiated in the presence of VitD developed a more tolerogenic phenotype than in the presence of Pari. These findings suggest that VitD, but not Pari, has the potential to be used as a preventive therapy to control MS severity.
Asunto(s)
Antioxidantes/administración & dosificación , Colecalciferol/administración & dosificación , Encefalomielitis Autoinmune Experimental/prevención & control , Ergocalciferoles/administración & dosificación , Factores Inmunológicos/administración & dosificación , Profilaxis Posexposición/métodos , Animales , Antioxidantes/farmacología , Células de la Médula Ósea/efectos de los fármacos , Células de la Médula Ósea/inmunología , Colecalciferol/farmacología , Citocinas/metabolismo , Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Encefalomielitis Autoinmune Experimental/sangre , Encefalomielitis Autoinmune Experimental/inmunología , Ergocalciferoles/farmacología , Femenino , Factores Inmunológicos/farmacología , Ratones , Ratones Endogámicos C57BL , Esclerosis Múltiple/inmunología , Esclerosis Múltiple/prevención & control , Índice de Severidad de la Enfermedad , Transducción de Señal/efectos de los fármacos , Resultado del TratamientoRESUMEN
PURPOSE: Chemotherapy for advanced pancreatic cancer has limited efficacy due to the difficultly of treating established tumours and the evolution of tumour resistance. Chemotherapies for pancreatic cancer are typically studied for their cytotoxic properties rather than for their ability to increase the immunogenicity of pancreatic tumour cells. In this study Gemcitabine in combination with immune modulatory chemotherapies Oxaliplatin, zoledronic acid and pomalidomide was studied to determine how combination therapy alters the immunogenicity of pancreatic tumour cell lines and subsequent T-cell responses. METHODS: Pancreatic tumour cell lines were stimulated with the chemotherapeutic agents and markers of immune recognition were assessed. The effect of chemotherapeutic agents on DC function was measured using uptake of CFSE-stained PANC-1 cells, changes in markers of maturation and their ability to activate CD8+ T-cells. The effect of chemotherapeutic agents on T-cell priming prior to activation using anti-CD3 and anti-CD28 antibodies was determined by measuring IFN-γ expression and Annexin V staining using flow cytometry. RESULTS: These agents demonstrate both additive and inhibitory properties on a range of markers of immunogenicity. Gemcitabine was notable for its ability to induce the upregulation of human leukocyte antigen and checkpoints on pancreatic tumour cell lines whilst inhibiting T-cell activation. Pomalidomide demonstrated immune modulatory properties on dendritic cells and T-cells, even in the presence of gemcitabine. DISCUSSION: These data highlight the complex interactions of different agents in the modulation of tumour immunogenicity and immune cell activation and emphasise the complexity in rationally designing chemo immunogenic combinations for use with immunotherapy.
Asunto(s)
Antimetabolitos Antineoplásicos/farmacología , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Desoxicitidina/análogos & derivados , Inmunomodulación/efectos de los fármacos , Neoplasias Pancreáticas/inmunología , Anexina A5/metabolismo , Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/inmunología , Línea Celular Tumoral , Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Desoxicitidina/farmacología , Interacciones Farmacológicas/inmunología , Antígenos de Histocompatibilidad Clase I/efectos de los fármacos , Antígenos de Histocompatibilidad Clase I/metabolismo , Humanos , Inmunomodulación/inmunología , Interferón gamma/metabolismo , Activación de Linfocitos/efectos de los fármacos , Activación de Linfocitos/inmunología , Oxaliplatino/farmacología , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/patología , Talidomida/análogos & derivados , Talidomida/farmacología , Ácido Zoledrónico/farmacología , GemcitabinaRESUMEN
S-layer (glyco)-proteins (SLPs) form a nanostructured envelope that covers the surface of different prokaryotes and show immunomodulatory activity. Previously, we have demonstrated that the S-layer glycoprotein from probiotic Lactobacillus kefiri CIDCA 8348 (SLP-8348) is recognized by Mincle (macrophage inducible C-type lectin receptor), and its adjuvanticity depends on the integrity of its glycans. However, the glycan's structure has not been described so far. Herein, we analyze the glycosylation pattern of three SLPs, SLP-8348, SLP-8321, and SLP-5818, and explore how these patterns impact their recognition by C-type lectin receptors and the immunomodulatory effect of the L. kefiri SLPs on antigen-presenting cells. High-performance anion-exchange chromatography-pulse amperometric detector performed after ß-elimination showed glucose as the major component in the O-glycans of the three SLPs; however, some differences in the length of hexose chains were observed. No N-glycosylation signals were detected in SLP-8348 and SLP-8321, but SLP-5818 was observed to have two sites carrying complex N-glycans based on a site-specific analysis and a glycomic workflow of the permethylated glycans. SLP-8348 was previously shown to enhance LPS-induced activation on both RAW264.7 macrophages and murine bone marrow-derived dendritic cells; we now show that SLP-8321 and SLP-5818 have a similar effect regardless of the differences in their glycosylation patterns. Studies performed with bone marrow-derived dendritic cells from C-type lectin receptor-deficient mice revealed that the immunostimulatory activity of SLP-8321 depends on its recognition by Mincle, whereas SLP-5818's effects are dependent on SignR3 (murine ortholog of human DC-SIGN). These findings encourage further investigation of both the potential application of these SLPs as new adjuvants and the protein glycosylation mechanisms in these bacteria.
Asunto(s)
Antígenos CD/metabolismo , Lactobacillus/metabolismo , Lectinas Tipo C/metabolismo , Glicoproteínas de Membrana/metabolismo , Secuencia de Aminoácidos , Animales , Antígenos CD/genética , Cromatografía Líquida de Alta Presión , Células Dendríticas/citología , Células Dendríticas/efectos de los fármacos , Células Dendríticas/metabolismo , Glicopéptidos/análisis , Glicopéptidos/química , Glicosilación , Inmunización , Interferón gamma/metabolismo , Lectinas Tipo C/deficiencia , Lectinas Tipo C/genética , Lipopolisacáridos/farmacología , Glicoproteínas de Membrana/química , Proteínas de la Membrana/deficiencia , Proteínas de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Polisacáridos/análisis , Polisacáridos/química , Células RAW 264.7 , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
Triple Negative Breast Cancer (TNBC) treatment is still challenging, and immunotherapy is a potential approach in this tumor subtype. Cetuximab is an IgG1 monoclonal antibody (mAb) directed against Epidermic Growth Factor Receptor (EGFR), a protein overexpressed in a subgroup of TNBC patients and associated with poor prognosis. Previously, we demonstrated in vitro that Cetuximab triggers Ab-dependent cell cytotoxicity against TNBC cells. In this study, using co-cultures including TNBC cells, and NK and Dendritic Cells (DCs) from healthy donors, we studied the effect of Cetuximab-activated NK cells on DC function. Given that we already demonstrated that TNBC has an immunosuppressive effect on NK cells, we also tested Cetuximab combination with IL-15. We determined that Cetuximab opsonization of TNBC cells increased IFN-γ and TNF-α production by NK cells co-cultured with DCs. Moreover, we showed that NK cells activated by TNBC cells opsonized with Cetuximab promoted tumor material uptake and maturation of DCs, as well as their ability to produce IL-12. Furthermore, the stimulation with IL-15 increased the activation of NK cells and the maturation of DCs. These results suggest that IL-15 may enhance the efficacy of Cetuximab in the treatment of TNBC by promoting activation of both NK cells and DCs.
Asunto(s)
Cetuximab/farmacología , Interleucina-15/farmacología , Línea Celular Tumoral , Células Cultivadas , Células Dendríticas/efectos de los fármacos , Células Dendríticas/metabolismo , Femenino , Citometría de Flujo , Humanos , Interferón gamma/metabolismo , Interleucina-12/metabolismo , Células Asesinas Naturales/efectos de los fármacos , Células Asesinas Naturales/metabolismo , Fagocitosis/efectos de los fármacos , Neoplasias de la Mama Triple Negativas , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
The interaction of dendritic cells and macrophages with a variety of rigid noncellular particles triggers activation of the NLRP3 inflammasome and consequent secretion of interleukin 1ß (IL-1ß). Noncellular particles can also be generated in the context of helminth infection, since these large pathogens often shed their outermost structures during growth and/or molting. One such structure is the massive, mucin-based, soft, flexible laminated layer (LL), which protects the larval stages of cestodes of the genus Echinococcus We show that particles from the Echinococcus granulosus LL (pLL) trigger NLRP3- and caspase-1-dependent IL-1ß in lipopolysaccharide (LPS)-primed mouse bone marrow-derived dendritic cells (BMDC). This response can be elicited by pLL too large for phagocytosis and nonetheless requires actin dynamics, Syk, and phosphatidylinositol 3-kinase (PI3K). These three requirements had already been observed in our previous study on the alteration by pLL of CD86, CD40, IL-10, and IL-12 responses to LPS in BMDC; however, we now show that these alterations are independent of NLRP3 and caspase-1. In other words, an initial interaction with particles requiring actin dynamics, Syk, and PI3K, but not phagocytosis, elicits both NLRP3-dependent and NLRP3-independent responses. Intraperitoneal injection of pLL induced IL-1ß, suggesting that contact with LL materials induces IL-1ß in the E. granulosus infection setting. Our results extend our understanding of NLRP3 inflammasome activation by noncellular particulate materials both to helminth-derived materials and to flexible/soft materials.
Asunto(s)
Micropartículas Derivadas de Células/química , Células Dendríticas/efectos de los fármacos , Echinococcus granulosus/química , Regulación de la Expresión Génica/efectos de los fármacos , Interacciones Huésped-Parásitos/inmunología , Proteína con Dominio Pirina 3 de la Familia NLR/inmunología , Clorometilcetonas de Aminoácidos/farmacología , Animales , Células de la Médula Ósea/efectos de los fármacos , Células de la Médula Ósea/inmunología , Caspasa 1/genética , Caspasa 1/inmunología , Micropartículas Derivadas de Células/inmunología , Células Dendríticas/inmunología , Echinococcus granulosus/inmunología , Femenino , Regulación de la Expresión Génica/inmunología , Interacciones Huésped-Parásitos/genética , Indazoles/farmacología , Inflamasomas/efectos de los fármacos , Inflamasomas/genética , Inflamasomas/inmunología , Interleucina-12/genética , Interleucina-12/inmunología , Interleucina-1beta/genética , Interleucina-1beta/inmunología , Lipopolisacáridos/farmacología , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Ratones , Ratones Endogámicos C57BL , Proteína con Dominio Pirina 3 de la Familia NLR/agonistas , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Fagocitosis/efectos de los fármacos , Fosfatidilinositol 3-Quinasa/genética , Fosfatidilinositol 3-Quinasa/inmunología , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/inmunología , Transducción de Señal , Estilbenos/farmacología , Sulfonamidas/farmacología , Wortmanina/farmacologíaRESUMEN
Schistosomiasis is a debilitating parasitic disease that affects more than 200 million people worldwide and causes approximately 280,000 deaths per year. Inside the definitive host, eggs released by Schistosoma mansoni lodge in the intestine and especially in the liver where they induce a granulomatous inflammatory process, which can lead to fibrosis. The molecular mechanisms initiating or promoting hepatic granuloma formation remain poorly understood. Inflammasome activation has been described as an important pathway to induce pathology mediated by NLRP3 receptor. Recently, other components of the inflammasome pathway, such as NLRP6, have been related to liver diseases and fibrotic processes. Nevertheless, the contribution of these components in schistosomiasis-associated pathology is still unknown. In the present study, using dendritic cells, we demonstrated that NLRP6 sensor is important for IL-1ß production and caspase-1 activation in response to soluble egg antigens (SEA). Furthermore, the lack of NLRP6 has been shown to significantly reduce periovular inflammation, collagen deposition in hepatic granulomas and mRNA levels of α-SMA and IL-13. Livers of Nlrp6-/- mice showed reduced levels of CXCL1/KC, CCL2, CCL3, IL-5, and IL-10 as well as Myeloperoxidase (MPO) and Eosinophilic Peroxidase (EPO) enzymatic activity. Consistently, the frequency of macrophage and neutrophil populations were lower in the liver of NLRP6 knockout mice, after 6 weeks of infection. Finally, it was further demonstrated that the onset of hepatic granuloma and collagen deposition were also compromised in Caspase-1-/- , IL-1R-/- and Gsdmd-/- mice. Our findings suggest that the NLRP6 inflammasome is an important component for schistosomiasis-associated pathology.
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Hígado/inmunología , Hígado/patología , Receptores de Superficie Celular/metabolismo , Schistosoma mansoni/inmunología , Esquistosomiasis mansoni/inmunología , Animales , Antígenos Helmínticos/metabolismo , Antígenos Helmínticos/farmacología , Caspasa 1/genética , Caspasa 1/metabolismo , Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Fibrosis , Técnicas de Inactivación de Genes , Granuloma/inmunología , Granuloma/metabolismo , Inflamasomas/metabolismo , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Hepatopatías/inmunología , Hepatopatías/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Proteínas de Unión a Fosfato/genética , Proteínas de Unión a Fosfato/metabolismo , Receptores de Superficie Celular/genética , Receptores de Interleucina-1/genética , Receptores de Interleucina-1/metabolismo , Esquistosomiasis mansoni/parasitologíaRESUMEN
Immune and molecular profiling of CD8 T cells of patients receiving DC vaccines expressing three full-length melanoma antigens (MAs) was performed. Antigen expression levels in DCs had no significant impact on T cell or clinical responses. Patients who received checkpoint blockade before DC vaccination had higher baseline MA-specific CD8 T cell responses but no evidence for improved functional responses to the vaccine. Patients who showed the best clinical responses had low PD-1 expression on MA-specific T cells before and after DC vaccination; however, blockade of PD-1 during antigen presentation by DC had minimal functional impact on PD-1high MA-specific T cells. Gene and protein expression analyses in lymphocytes and tumor samples identified critical immunoregulatory pathways, including CTLA-4 and PD-1. High immune checkpoint gene expression networks correlated with inferior clinical outcomes. Soluble serum PD-L2 showed suggestive positive association with improved outcome. These findings show that checkpoint molecular pathways are critical for vaccine outcomes and suggest specific sequencing of vaccine combinations.
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Linfocitos T CD8-positivos/inmunología , Vacunas contra el Cáncer/inmunología , Inhibidores de Puntos de Control Inmunológico/farmacología , Activación de Linfocitos/inmunología , Antígenos de Neoplasias/metabolismo , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/efectos de los fármacos , Antígeno CTLA-4/metabolismo , Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Supervivencia sin Enfermedad , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Interferón gamma/metabolismo , Activación de Linfocitos/efectos de los fármacos , Linfocitos Infiltrantes de Tumor/efectos de los fármacos , Antígeno MART-1/metabolismo , Melanoma/sangre , Melanoma/genética , Melanoma/inmunología , Melanoma/patología , Receptor de Muerte Celular Programada 1/metabolismo , VacunaciónRESUMEN
CD8 T cells can kill malignant cells in an antigen-specific manner. However, anti-tumoral responses are usually limited by suppressive factors that curb the effector responses of tumor-infiltrating CD8 T cells. Therapeutic strategies to overcome intra-tumoral T cell suppression, for example immune checkpoint inhibition, have been clinically effective in patients with cancer. Here, we provide data that demonstrates that GK-1, a peptide derived from the parasite Taenia crassiceps, promotes an anti-melanoma CD8 T cell response with heightened effector characteristics that leads to an increased amount of tumor-infiltrating CD44+ IFN-γ-producing CD8 T cells. The response induced by GK-1 was associated with a reduction in the expression of PD-1 and PD-L1 on tumor-infiltrating CD8 and dendritic cells, respectively, effects that led to a dramatic decrease in tumor burden. Our results suggest that the immunomodulatory properties of GK-1 may promote a CD8 T cell response that may be therapeutically useful in the setting of cancer.
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Antígeno B7-H1/efectos de los fármacos , Linfocitos T CD8-positivos/efectos de los fármacos , Células Dendríticas/efectos de los fármacos , Linfocitos Infiltrantes de Tumor/efectos de los fármacos , Melanoma Experimental/inmunología , Péptidos Cíclicos/farmacología , Receptor de Muerte Celular Programada 1/efectos de los fármacos , Neoplasias Cutáneas/inmunología , Microambiente Tumoral/efectos de los fármacos , Traslado Adoptivo , Animales , Antígeno B7-H1/inmunología , Linfocitos T CD8-positivos/inmunología , Células Dendríticas/inmunología , Regulación hacia Abajo , Receptores de Hialuranos/inmunología , Interferón gamma/inmunología , Linfocitos Infiltrantes de Tumor/inmunología , Ratones , Ratones Endogámicos C57BL , Receptor de Muerte Celular Programada 1/inmunología , Linfocitos T/trasplante , Taenia , Microambiente Tumoral/inmunologíaRESUMEN
Current antifungal drugs present poor effectiveness and there is no available vaccine for fungal infections. Thus, novel strategies to treat or prevent invasive mycosis, such as cryptococcosis, are highly desirable. One strategy is the use of immunomodulators of polysaccharide nature isolated from mushrooms. The purpose of the present work was to evaluate the immunostimulatory activity of ß-(1,3)-glucan-containing exopolysaccharides (EPS) from the edible mushrooms Auricularia auricula in phagocytes and mice infected with Cryptococcus neoformans. EPS triggered macrophages and dendritic cell activation upon binding to Dectin-1, a pattern recognition receptor of the C-type lectin receptor family. Engagement of Dectin-1 culminated in pro-inflammatory cytokine production and cell maturation via its canonical Syk-dependent pathway signaling. Furthermore, upon EPS treatment, M2-like phenotype macrophages, known to support intracellular survival and replication of C. neoformans, repolarize to M1 macrophage pattern associated with enhanced production of the microbicidal molecule nitric oxide that results in efficient killing of C. neoformans. Treatment with EPS also upregulated transcript levels of genes encoding products associated with host protection against C. neoformans and Dectin-1 mediated signaling in macrophages. Finally, orally administrated ß-glucan-containing EPS from A. auricular enhanced the survival of mice infected with C. neoformans. In conclusion, the results demonstrate that EPS from A. auricula exert immunostimulatory activity in phagocytes and induce host protection against C. neoformans, suggesting that polysaccharides from this mushroom may be promising as an adjuvant for vaccines or antifungal therapy.
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Agaricales/química , Criptococosis/prevención & control , Polisacáridos Fúngicos/inmunología , Fagocitos/efectos de los fármacos , Fagocitos/inmunología , beta-Glucanos/inmunología , Animales , Criptococosis/inmunología , Cryptococcus neoformans/inmunología , Citocinas/inmunología , Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Células Dendríticas/microbiología , Factores Inmunológicos/farmacología , Lectinas Tipo C/inmunología , Enfermedades Pulmonares Fúngicas , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Macrófagos/microbiología , Ratones , Ratones Endogámicos C57BL , Fagocitos/microbiología , Transducción de Señal , beta-Glucanos/farmacologíaRESUMEN
PURPOSE: Dendritic cells (DCs) are the most potent antigen-presenting cells (APCs), and their capacity to activate the immune response has been widely used in immunotherapies against different diseases, predominantly cancer. However, they have not been so widely used in immunotherapies against infectious diseases. Leishmania mexicana is the causative agent of cutaneous leishmaniasis in Mexico, which can result in localized cutaneous leishmaniasis (LCL) and diffuse cutaneous leishmaniasis (DCL). DCL is characterized by the incapability of the immune response to control the parasite, which thus disseminates to all teguments. Treatments against DCL have shown low efficacy, which is a reason why alternative therapies such as immunotherapies are promising. One adjuvant that has proven its effectiveness in immunotherapies against some cancers and infections is GK1, a component of the SPVac vaccine against porcine cysticercosis. GK1 has the capacity to elicit proinflammatory cytokines and chemokines from DCs and macrophages. METHODS: We pulsed bone marrow-derived dendritic cells (BMDCs) with GK1 and a lysate obtained from L. mexicana promastigotes and tested the efficacy of this combination against the infection of susceptible mice with L. mexicana. RESULTS: We found that BMDCs stimulated with GK1 and a lysate of L. mexicana promastigotes secreted IFN-γ and IL-12, and when they were adoptively transferred to BALB/c mice which were then infected with L. mexicana promastigotes, there was a reduction in the size of the lesion and in the parasite load. CONCLUSIONS: The adjuvant properties of GK1 along with parasite antigens may have a protective effect against the infection of BALB/c mice with L. mexicana.
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Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Péptidos Cíclicos/inmunología , Péptidos Cíclicos/farmacología , Proteínas Protozoarias/inmunología , Adyuvantes Inmunológicos/farmacología , Traslado Adoptivo , Animales , Interferón gamma/inmunología , Interleucina-12/inmunología , Leishmania mexicana , Leishmaniasis Cutánea/inmunología , Leishmaniasis Cutánea Difusa/inmunología , Ratones , Ratones Endogámicos BALB C , Carga de Parásitos , Proteínas Protozoarias/farmacologíaRESUMEN
PURPOSE: Fatty acid synthase (FASN), the multifunctional enzyme responsible for endogenous fatty acid synthesis, is highly expressed and associated with poor prognosis in several human cancers, including melanoma. Our group has previously shown that pharmacological inhibition of FASN with orlistat decreases proliferation, promotes apoptosis, and reduces the metastatic spread of B16-F10 cells in experimental models of melanoma. While most of the orlistat antitumor properties seem to be closely related to direct effects on malignant cells, its impact on the host immune system is still unknown. METHODS: The effects of orlistat on the phenotype and activation status of infiltrating leukocytes in primary tumors and metastatic lymph nodes were assessed using a model of spontaneous melanoma metastasis (B16-F10 cells/C57BL/6 mice). Cells from the primary tumors and lymph nodes were mechanically dissociated and immune cells phenotyped by flow cytometry. The expression of IL-12p35, IL-12p40, and inducible nitric oxide synthase (iNOS) was analyzed by qRT-PCR and production of nitrite (NO2-) evaluated in serum samples with the Griess method. RESULTS: Orlistat-treated mice exhibited a 25% reduction in the number of mediastinal lymph node metastases (mean 3.96 ± 0.78, 95% CI 3.63-4.28) compared to the controls (mean 5.7 ± 1.72; 95% CI 5.01-6.43). The drug elicited an antitumor immune response against experimental melanomas by increasing maturation of intratumoral dendritic cells (DC), stimulating the expression of cytotoxicity markers in CD8 T lymphocytes and natural killer (NK) cells, as well as reducing regulatory T cells (Tregs). Moreover, the orlistat-treatment increased serum levels of nitric oxide (NO) concentrations. CONCLUSION: Taken together, these findings suggest that orlistat supports an antitumor response against experimental melanomas by increasing CD80/CD81-positive and IL-12-positive DC populations, granzyme b/NKG2D-positive NK populations, and perforin/granzyme b-positive CD8 T lymphocytes as well as reducing Tregs counts within experimental melanomas.
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Antineoplásicos/farmacología , Metástasis Linfática/tratamiento farmacológico , Melanoma Experimental/tratamiento farmacológico , Orlistat/farmacología , Animales , Apoptosis/efectos de los fármacos , Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Células Dendríticas/efectos de los fármacos , Células Dendríticas/metabolismo , Ácido Graso Sintasas/metabolismo , Células Asesinas Naturales/efectos de los fármacos , Células Asesinas Naturales/metabolismo , Masculino , Melanoma Experimental/metabolismo , Ratones , Ratones Endogámicos C57BL , Óxido Nítrico/metabolismo , Linfocitos T Reguladores/efectos de los fármacos , Linfocitos T Reguladores/metabolismoRESUMEN
Abstract Hippocampus is a part of the brain that has a major role in spatial learning and memory which can be affected by herbal extracts. Incense resin (Styrax benzoin) has been used by local communities to improve intelligence. However, there is no scientific evidence of the functions of Styrax benzoin for regulating hippocampal function. The aim of this study was intended to analyze and investigate the effect of incense resin on learning, memory, and dendrite complexity of mice. Three months old male Deutch Democratic Yokohama (DDY) mice were injected orally with graded doses of 100, 150, and 200 mg/kg of incense resin aqueous extract daily for 30 days. Spatial learning and memory performance levels were tested with Y-maze alternation, novel object recognition, and Morris water maze. The branches and maximum dendritic span in the dentate gyrus were observed by the Golgi-Cox staining. Overall, our results showed that incense resin extract increased learning and memory ability, and the number of dendrite branching in the dentate gyrus.
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Animales , Masculino , Ratones , Células Dendríticas/efectos de los fármacos , Extractos Vegetales/farmacología , Styrax/química , Aprendizaje Espacial/efectos de los fármacos , Memoria/efectos de los fármacos , Administración Oral , Aprendizaje por Laberinto/efectos de los fármacosRESUMEN
Elevated levels of immunoglobulin E (IgE) are associated with allergies and other immunological disorders. Sensitization with alum adjuvant favours IgE production while CpG-ODN adjuvant, a synthetic toll-like receptor 9 (TLR9) agonist, inhibits it. The cellular mechanisms underlying in vivo TLR regulation of immunoglobulin production, specially IgE, are still controversial. Specifically, TLR-mediated IgE regulation in vivo is not yet known. In this study we showed that augmented levels of IgE induced by sensitizations to OVA with or without alum adjuvant or with OVA-pulsed dendritic cells (DCs) were inhibited by co-administration of CpG. Notably, CpG-mediated suppression of IgE production required MyD88-expression on DCs but not on B-cells. This finding contrasts with previous in vitro studies reporting regulation of IgE by a direct action of CpG on B cells via MyD88 pathway. In addition, we showed that CpG also inhibited IgE production in a MyD88-dependent manner when sensitization was performed with OVA-pulsed DCs. Finally, CpG signalling through MyD88 pathway was also necessary and sufficient to prevent anaphylactic antibody production involved in active cutaneous anaphylaxis.
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Adyuvantes Inmunológicos/administración & dosificación , Células Dendríticas/inmunología , Inmunoglobulina E/metabolismo , Factor 88 de Diferenciación Mieloide/metabolismo , Oligodesoxirribonucleótidos/administración & dosificación , Ovalbúmina/efectos adversos , Hipersensibilidad Respiratoria/tratamiento farmacológico , Adyuvantes Inmunológicos/farmacología , Animales , Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Linfocitos B/metabolismo , Células Dendríticas/efectos de los fármacos , Células Dendríticas/metabolismo , Modelos Animales de Enfermedad , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Inmunoglobulina E/efectos de los fármacos , Ratones , Factor 88 de Diferenciación Mieloide/genética , Oligodesoxirribonucleótidos/farmacología , Hipersensibilidad Respiratoria/inducido químicamente , Hipersensibilidad Respiratoria/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
Background Dendritic cells (DCs) play a crucial role during HIV-1 transmission due to their ability to transfer virions to susceptible CD4+ T cells, particularly in the lymph nodes during antigen presentation which favors the establishment of systemic infection. As mature dendritic cells (mDCs) exhibit a greater ability to transfer virions, compared to immature DCs (iDCs), maintenance of an iDC phenotype could decrease viral transmission. The immunomodulatory vitamin D (VitD) has been shown to reduce activation and maturation of DCs; hence, we hypothesized that it would reduce viral transference by DCs. Materials and methods We evaluated the effect of in vitro treatment with a precursor of VitD, cholecalciferol, on the activation/maturation phenotype of differentiated monocyte-derived DCs and their ability to transfer HIV-1 to autologous CD4+ T cells. Results Our findings show that although cholecalciferol decreases the activation of iDCs, it did not impact the maturation phenotype after LPS treatment nor iDCs' ability to transfer viral particles to target cells. Conclusion These findings suggest that despite cholecalciferol potentially modulates the phenotype of mucosal iDCs in vivo, such modulation might not impact the ability of these cells to transfer HIV-1 to target CD4+ T cells.