RESUMEN
BACKGROUND: The interleukin (IL)-36 cytokines are a sub-family of the IL-1 family which are becoming increasingly implicated in the pathogenesis of inflammatory diseases and malignancies. Initial studies of IL-36 signalling in tumorigenesis identified an immune-mediated anti-tumorigenic function for these cytokines. However, more recent studies have shown IL-36 cytokines also contribute to the pathogenesis of lung and colorectal cancer (CRC). METHODS: The aim of this study was to investigate IL-36 expression in CRC using transcriptomic datasets and software such as several R packages, Cytoscape, GEO2R and AnalyzeR. Validation of results was completed by qRT-PCR on both cell lines and a patient cohort. Cellular proliferation was assessed by flow cytometry and resazurin reduction. RESULTS: We demonstrate that IL-36 gene expression increases with CRC development. Decreased tumoral IL-36 receptor expression was shown to be associated with improved patient outcome. Our differential gene expression analysis revealed a novel role for the IL-36/IL-17/IL-23 axis, with these findings validated using patient-derived samples and cell lines. IL-36γ, together with either IL-17a or IL-22, was able to synergistically induce different genes involved in the IL-17/IL-23 axis in CRC cells and additively induce colon cancer cell proliferation. CONCLUSIONS: Collectively, this data support a pro-tumorigenic role for IL-36 signalling in colon cancer, with the IL-17/IL-23 axis influential in IL-36-mediated colon tumorigenesis.
Asunto(s)
Neoplasias del Colon , Neoplasias Colorrectales , Humanos , Carcinogénesis , Neoplasias Colorrectales/patología , Citocinas/metabolismo , Perfilación de la Expresión Génica , Interleucina-17 , Interleucina-23/genética , TranscriptomaRESUMEN
The IL-36 cytokines are a recently described subset of the IL-1 family of cytokines, and have been shown to play a role in the pathogenesis of respiratory diseases such as asthma and COPD. Given the common aetiological links between COPD and lung cancer development, as well as the involvement of other IL-1 family members in lung tumorigenesis, the aim of this work was to investigate the role of IL-36 cytokines in the pathogenesis of lung cancer. In this study we demonstrate that expression of IL-36 cytokines and receptor mRNA and protein are significantly increased in lung cancer tissue compared to adjacent non-tumour tissue. In vitro assays showed that stimulation of two lung cancer cell lines, SKMES-1 human squamous cell and LLC murine lung cancer, with IL-36R agonists resulted in increased cellular migration and proliferation. All IL-36 cytokines induced the expression of pro-inflammatory chemokines in both lung cancer cell lines with synergistic effects identified upon co-stimulation of cells with IL-17, IL-22 and TNFα. Furthermore, we report that IL-36 cytokines induce protein expression of the immune checkpoint inhibitor protein PD-L1 on lung cancer cells. Taken together, this data indicates that targeting IL-36R signalling may be a useful targeted therapy for lung cancer patients with IL-36R+ cancer cells.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Enfermedad Pulmonar Obstructiva Crónica , Humanos , Ratones , Animales , Carcinoma de Pulmón de Células no Pequeñas/patología , Neoplasias Pulmonares/patología , Citocinas/metabolismo , Pulmón/metabolismo , Interleucina-1/metabolismo , CarcinogénesisRESUMEN
Environmental factors, including westernised diets and alterations to the gut microbiota, are considered risk factors for inflammatory bowel diseases (IBD). The mechanisms underpinning diet-microbiota-host interactions are poorly understood in IBD. We present evidence that feeding a lard-based high-fat (HF) diet can protect mice from developing DSS-induced acute and chronic colitis and colitis-associated cancer (CAC) by significantly reducing tumour burden/incidence, immune cell infiltration, cytokine profile, and cell proliferation. We show that HF protection was associated with increased gut microbial diversity and a significant reduction in Proteobacteria and an increase in Firmicutes and Clostridium cluster XIVa abundance. Microbial functionality was modulated in terms of signalling fatty acids and bile acids (BA). Faecal secondary BAs were significantly induced to include moieties that can activate the vitamin D receptor (VDR), a nuclear receptor richly represented in the intestine and colon. Indeed, colonic VDR downstream target genes were upregulated in HF-fed mice and in combinatorial lipid-BAs-treated intestinal HT29 epithelial cells. Collectively, our data indicate that HF diet protects against colitis and CAC risk through gut microbiota and BA metabolites modulating vitamin D targeting pathways. Our data highlights the complex relationship between dietary fat-induced alterations of microbiota-host interactions in IBD/CAC pathophysiology.
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Colitis , Enfermedades Inflamatorias del Intestino , Neoplasias , Ratones , Animales , Vitamina D/metabolismo , Inflamación/metabolismo , Colitis/metabolismo , Enfermedades Inflamatorias del Intestino/metabolismo , Colon/patología , Dieta Alta en Grasa/efectos adversos , Bacterias , Ácidos y Sales Biliares/metabolismo , Ratones Endogámicos C57BL , Sulfato de Dextran/efectos adversos , Neoplasias/metabolismoRESUMEN
DNA sensor pathways can initiate inflammasome, cell death, and type I interferon (IFN) signaling in immune-mediated inflammatory diseases (IMIDs), including type I interferonopathies. We investigated the involvement of these pathways in the pathogenesis of ulcerative colitis (UC) by analyzing the expression of DNA sensor, inflammasome, and type I IFN biomarker genes in colonic mucosal biopsy tissue from control (n = 31), inactive UC (n = 31), active UC (n = 33), and a UC single-cell RNA-Seq dataset. The effects of type I IFN (IFN-ß), IFN-γ, and TNF-α on gene expression, cytokine production, and cell death were investigated in human colonic organoids. In organoids treated with cytokines alone, or in combination with NLR family pyrin domain-containing 3 (NLRP3), caspase, or JAK inhibitors, cell death was measured, and supernatants were assayed for IL-1ß/IL-18/CXCL10. The expression of DNA sensor pathway genes-PYHIN family members [absent in melanoma 2 (AIM2), IFI16, myeloid cell nuclear differentiation antigen (MNDA), and pyrin and HIN domain family member 1 (PYHIN1)- as well as Z-DNA-binding protein 1 (ZBP1), cyclic GMP-AMP synthase (cGAS), and DDX41 was increased in active UC and expressed in a cell type-restricted pattern. Inflammasome genes (CASP1, IL1B, and IL18), type I IFN inducers [stimulator of interferon response cGAMP interactor 1 (STING), TBK1, and IRF3), IFNB1, and type I IFN biomarker genes (OAS2, IFIT2, and MX2) were also increased in active UC. Cotreatment of organoids with IFN-ß or IFN-γ in combination with TNFα increased expression of IFI16, ZBP1, CASP1, cGAS, and STING induced cell death and IL-1ß/IL-18 secretion. This inflammatory cell death was blocked by the JAK inhibitor tofacitinib but not by inflammasome or caspase inhibitors. Increased type I IFN activity may drive elevated expression of DNA sensor genes and JAK-dependent but inflammasome-independent inflammatory cell death of colonic epithelial cells in UC.NEW & NOTEWORTHY This study found that patients with active UC have significantly increased colonic gene expression of cytosolic DNA sensor, inflammasome, STING, and type I IFN signaling pathways. The type I IFN, IFN-ß, in combination with TNF-α induced JAK-dependent but NLRP3 and inflammasome-independent inflammatory cell death of colonic organoids. This novel inflammatory cell death phenotype is relevant to UC immunopathology and may partially explain the efficacy of the JAKinibs tofacitinib and upadacitinib in patients with UC.
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Colitis Ulcerosa , Interferón Tipo I , Inhibidores de las Cinasas Janus , Humanos , Inflamasomas/metabolismo , Interleucina-18 , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Factor de Necrosis Tumoral alfa , Inhibidores de Caspasas , Organoides/metabolismo , Pirina , Caspasa 1/metabolismo , Nucleotidiltransferasas/metabolismo , ADN , Muerte Celular , Proteínas de Unión al ADN/metabolismo , Antígenos de DiferenciaciónRESUMEN
The IL-36 family of cytokines were first identified in 2000 based on their sequence homology to IL-1 cytokines. Over subsequent years, the ability of these cytokines to either agonise or antagonise an IL-1R homologue, now known as the IL-36 Receptor (IL-36R), was identified and these cytokines went through several cycles of renaming with the current nomenclature being proposed in 2010. Despite being identified over 20 years ago, it is only during the last decade that the function of these cytokines in health and disease has really begun to be appreciated, with both homeostatic functions in wound healing and response to infection, as well as pathological functions now ascribed. In the disease context, over activation of IL-36 has now been associated with many inflammatory diseases including Psoriasis and inflammatory bowel diseases, with roles in cancer also now being investigated. This review summarises the current knowledge of IL-36 biology, its role in inflammatory diseases and focuses on an emerging role for IL-36 in cancer.
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Enfermedades Inflamatorias del Intestino/patología , Interleucina-1/metabolismo , Neoplasias/patología , Humanos , Inmunidad Innata , Enfermedades Inflamatorias del Intestino/metabolismo , Interleucina-1/química , Interleucina-1/genética , Interleucinas/genética , Interleucinas/metabolismo , Artropatías/metabolismo , Artropatías/patología , Neoplasias/metabolismo , Enfermedades Neurodegenerativas/metabolismo , Enfermedades Neurodegenerativas/patología , Psoriasis/metabolismo , Psoriasis/patología , Transducción de SeñalRESUMEN
Development of the human gut microbiota commences at birth, with certain bifidobacterial species representing dominant and early colonisers of the newborn gastrointestinal tract. The molecular basis of Bifidobacterium colonisation, persistence and presumed communication with the host has remained obscure. We previously identified tight adherence (Tad) pili from Bifidobacterium breve UCC2003 as an essential colonisation factor. Here, we demonstrate that bifidobacterial Tad pili also promote in vivo colonic epithelial proliferation. A significant increase in cell proliferation was detectable 5 days postadministration of B. breve UCC2003. Using advanced functional genomic approaches, bacterial strains either (a) producing the Tad2003 pili or (b) lacking the TadE or TadF pseudopilins were created. Analysis of the ability of these mutant strains to promote epithelial cell proliferation in vivo demonstrated that the pilin subunit, TadE, is the bifidobacterial molecule responsible for this proliferation response. These findings were confirmed in vitro using purified TadE protein. Our data imply that bifidobacterial Tad pili may contribute to the maturation of the naïve gut in early life through the production of a specific scaffold of extracellular protein structures, which stimulate growth of the neonatal mucosa.
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Bifidobacterium breve/fisiología , Proliferación Celular/efectos de los fármacos , Células Epiteliales/efectos de los fármacos , Células Epiteliales/fisiología , Proteínas Fimbrias/metabolismo , Fimbrias Bacterianas/fisiología , Mucosa Intestinal/microbiología , Bifidobacterium breve/genética , Línea Celular , Proteínas Fimbrias/genética , Eliminación de Gen , HumanosRESUMEN
It is widely accepted that the hypoxic nature of solid tumors contribute to their resistance to radiation therapy. There is increasing evidence that cyclooxygenase-2 (COX-2) contributes to increased resistance of tumors to radiation therapy. Several studies demonstrate that combination of COX-2 selective inhibitors with radiation therapy selectively enhances radio responsiveness of tumor cells. However, the majority of these studies utilised suprapharmacological concentrations under normoxic conditions only. Furthermore, the mechanism by which these agents act remain largely unclear. Therefore, the aim of this study was to determine the impact of COX-2 selective inhibitors on both normoxic and hypoxic radiosensitivity in vitro and the mechanisms underlying this. Because of the close, reciprocal relationship between COX-2 and p53 we investigated their contribution to radioresistance. To achieve this we exposed HeLa, MCF-7 and MeWo cells to the COX-2 selective inhibitor, NS398 (10µM). NS398 (10µM) selectively sensitized hypoxic HeLa and MCF-7 but not MeWo cells to ionising radiation (5 Gy). Furthermore, while knockdown of COX-2 with siRNA did not affect either normoxic radiosensitivity in HeLa cells, the radiosensitisation observed with NS398 was lost suggesting both COX-2 dependent and independent mechanisms. We also show that ionising radiation at 5 Gy results in phosphorylation of p53 at serine 15, a key phosphorylation site for p53-mediated apoptosis, and that hypoxia attenuates this phosphorylation. Attenuated phosphorylation of p53 under hypoxic conditions may therefore contribute to hypoxic radioresistance. We also show that NS398 selectively phosphorylates p53 under hypoxic conditions following irradiation at 5 Gy. p53 phosphorylation could be an underlying mechanism by which this agent and other COX-2 inhibitors sensitize tumors to radiation therapy.
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Ciclooxigenasa 2/química , Nitrobencenos/farmacología , Tolerancia a Radiación/efectos de los fármacos , Sulfonamidas/farmacología , Proteína p53 Supresora de Tumor/metabolismo , Neoplasias del Cuello Uterino/radioterapia , Antiinflamatorios no Esteroideos/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/efectos de la radiación , Ciclooxigenasa 2/metabolismo , Femenino , Células HeLa , Humanos , Hipoxia/fisiopatología , Fosforilación , Radiación Ionizante , Neoplasias del Cuello Uterino/metabolismo , Neoplasias del Cuello Uterino/patologíaRESUMEN
BACKGROUND: Despite the importance of inflammation in cancer, the role of the cytokine IL-33, and its receptor ST2, in colon cancer is unclear. The aim of this study was to investigate the role of IL-33, and its receptor isoforms (ST2 and ST2L), in colon cancer. METHODS: Serum levels of IL-33 and sST2 were determined with ELISA. ST2 and IL-33 expression was detected with quantitative real-time PCR (qRT-PCR), western blotting and immunohistochemistry. ST2 expression in CT26 cells was stably suppressed using ST2-specific shRNA. Cytokine and chemokine gene expression was detected with qRT-PCR. RESULTS: Human colon tumours showed lower expression of ST2L as compared with adjacent non-tumour tissue (P<0.01). Moreover, the higher the tumour grade, the lower the expression of ST2L (P=0.026). Colon cancer cells expressed ST2 and IL-33 in vitro. Functional analyses showed that stimulation of tumour cells with IL-33 induced the expression of chemokine (C-C motif) ligand 2 (CCL2). Knockdown of ST2 in murine colon cancer cells resulted in enhanced tumour growth (P<0.05) in BALB/c mice in vivo. This was associated with a decrease in macrophage infiltration, with IL-33-induced macrophage recruitment reduced by antagonising CCL2 in vitro. CONCLUSION: The IL-33/ST2 signalling axis may have a protective role in colon carcinogenesis.
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Neoplasias del Colon/prevención & control , Interleucina-33/fisiología , Receptores de Superficie Celular/fisiología , Animales , Movimiento Celular , Proliferación Celular , Neoplasias del Colon/patología , Femenino , Humanos , Proteína 1 Similar al Receptor de Interleucina-1 , Interleucina-33/análisis , Macrófagos/fisiología , Ratones , Ratones Endogámicos BALB C , Clasificación del Tumor , Invasividad Neoplásica , Receptores de Superficie Celular/análisisRESUMEN
TLRs play an important role in mediating intestinal inflammation and homeostasis. Fas is best studied in terms of its function in apoptosis, but recent studies demonstrate that Fas signaling may mediate additional functions such as inflammation. The role of Fas, and the Fas ligand (FasL), in the intestine is poorly understood. The aim of this study was to evaluate potential cross-talk between TLRs and Fas/FasL system in intestinal epithelial cells (IECs). IECs were stimulated with TLR ligands, and expression of Fas and FasL was investigated. Treatment with TLR4 and TLR5 ligands, but not TLR2 and 9 ligands, increased expression of Fas and FasL in IECs in vitro. Consistent with this finding, expression of intestinal Fas and FasL was reduced in vivo in the epithelium of TLR4 knockout (KO), 5KO, and germ-free mice, but not in TLR2KO mice. Modulating Fas signaling using agonistic anti-Fas augmented TLR4- and TLR5-mediated TNF-α and IL-8 production by IECs. In addition, suppression of Fas in IECs reduced the ability of TLR4 and TLR5 ligands and the intestinal pathogens Salmonella typhimurium and Listeria monocytogenes to induce the expression of IL-8. In conclusion, this study demonstrates that extensive cross-talk in IECs occurs between the Fas and TLR signaling pathways, with the FasL/Fas system playing a role in TLR-mediated inflammatory responses in the intestine.
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Proteína Ligando Fas/metabolismo , Inflamación/metabolismo , Mucosa Intestinal/metabolismo , Transducción de Señal , Receptor Toll-Like 4/metabolismo , Receptor Toll-Like 5/metabolismo , Receptor fas/metabolismo , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Línea Celular , Citocinas/biosíntesis , Modelos Animales de Enfermedad , Proteína Ligando Fas/genética , Regulación de la Expresión Génica , Células HT29 , Humanos , Inflamación/genética , Inflamación/inmunología , Inflamación/microbiología , Mucosa Intestinal/inmunología , Intestinos/inmunología , Intestinos/microbiología , Ligandos , Listeria monocytogenes , Ratones , Ratones Noqueados , FN-kappa B/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Salmonella typhimurium , Receptor Toll-Like 4/agonistas , Receptor Toll-Like 4/genética , Receptor Toll-Like 5/agonistas , Receptor Toll-Like 5/genética , Receptor fas/agonistas , Receptor fas/genéticaRESUMEN
Listeria monocytogenes is a Gram-positive bacterium that can cause septicemia and meningitis. TLRs are central receptors of the innate immune system that drive inflammatory responses to invading microbes such as L. monocytogenes. Although intestinal epithelial cells (IECs) represent the initial point of entry used by L. monocytogenes for infection, the innate immune response to L. monocytogenes in these cells has been poorly characterized to date. The aim of this study was to determine which TLRs are involved in mediating the immune response to L. monocytogenes in IECs. We performed an RNA interference screen of TLRs 1-10 in the HT-29 IEC cell line and observed the most significant reduction in chemokine output following silencing of TLR10. This effect was also observed in the macrophage cell line THP-1. The chemokines CCL20, CCL1, and IL-8 were reduced following knockdown of TLR10. Silencing of TLR10 resulted in increased viability of L. monocytogenes in both HT-29 and THP-1 cells. TLR10 was found to be predominantly expressed intracellularly in epithelia, and activation required viable L. monocytogenes. NF-κB activation was seen to require TLR2 in addition to TLR10. Taken together, these data indicate novel roles for TLR10 in sensing pathogenic infection in both the epithelium and macrophages and have identified L. monocytogenes as a source of ligand for the orphan receptor TLR10.
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Células Epiteliales/inmunología , Inmunidad Mucosa , Mucosa Intestinal/inmunología , Listeria monocytogenes/inmunología , Macrófagos/inmunología , Receptor Toll-Like 10/fisiología , Quimiocinas/biosíntesis , Quimiocinas/genética , Células Epiteliales/microbiología , Regulación de la Expresión Génica/inmunología , Células HCT116 , Células HEK293 , Células HT29 , Humanos , Inmunidad Innata , Técnicas In Vitro , Interleucinas/biosíntesis , Interleucinas/genética , Mucosa Intestinal/citología , Ligandos , Macrófagos/microbiología , FN-kappa B/metabolismo , Especificidad de Órganos , Interferencia de ARN , ARN Interferente Pequeño/farmacología , Receptor Toll-Like 10/antagonistas & inhibidores , Receptor Toll-Like 10/genética , Receptor Toll-Like 10/inmunología , Receptor Toll-Like 2/fisiología , Receptores Toll-Like/biosíntesis , Receptores Toll-Like/genéticaRESUMEN
Fas, also known as CD95 or APO-1, is a member of the tumor necrosis factor/nerve growth factor superfamily. Although best characterized in terms of its apoptotic function, recent studies have identified several other cellular responses emanating from Fas. These responses include migration, invasion, inflammation, and proliferation. In this review, we focus on the diverse cellular outcomes of Fas signaling and the molecular switches identified to date that regulate its pro- and anti-apoptotic functions. Such switches occur at different levels of signal transduction, ranging from the receptor through to cross-talk with other signaling pathways. Factors identified to date including other extracellular signals, proteins recruited to the death-inducing signaling complex, and the availability of different intracellular components of signal transduction pathways. The success of therapeutically targeting Fas will require a better understanding of these pathways, as well as the regulatory mechanisms that determine cellular outcome following receptor activation.
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Transducción de Señal , Receptor fas/metabolismo , Animales , Apoptosis , Movimiento Celular , Proliferación Celular , Proteína Ligando Fas/metabolismo , Regulación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Inflamación/metabolismo , Activación de Linfocitos , FN-kappa B/metabolismo , Linfocitos T/citologíaRESUMEN
Development of the human gut microbiota commences at birth, with bifidobacteria being among the first colonizers of the sterile newborn gastrointestinal tract. To date, the genetic basis of Bifidobacterium colonization and persistence remains poorly understood. Transcriptome analysis of the Bifidobacterium breve UCC2003 2.42-Mb genome in a murine colonization model revealed differential expression of a type IVb tight adherence (Tad) pilus-encoding gene cluster designated "tad(2003)." Mutational analysis demonstrated that the tad(2003) gene cluster is essential for efficient in vivo murine gut colonization, and immunogold transmission electron microscopy confirmed the presence of Tad pili at the poles of B. breve UCC2003 cells. Conservation of the Tad pilus-encoding locus among other B. breve strains and among sequenced Bifidobacterium genomes supports the notion of a ubiquitous pili-mediated host colonization and persistence mechanism for bifidobacteria.
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Bifidobacterium/genética , Bifidobacterium/fisiología , Fimbrias Bacterianas/genética , Fimbrias Bacterianas/fisiología , Genoma Bacteriano , Secuencia de Aminoácidos , Animales , Proteínas Bacterianas/genética , Proteínas Bacterianas/fisiología , Secuencia de Bases , Bifidobacterium/crecimiento & desarrollo , Bifidobacterium/ultraestructura , Hibridación Genómica Comparativa , ADN Bacteriano/genética , Femenino , Fimbrias Bacterianas/ultraestructura , Tracto Gastrointestinal/microbiología , Regulación Bacteriana de la Expresión Génica , Vida Libre de Gérmenes , Humanos , Masculino , Metagenoma , Ratones , Ratones Endogámicos BALB C , Microscopía Electrónica de Transmisión , Microscopía Inmunoelectrónica , Datos de Secuencia Molecular , Familia de Multigenes , Mutación , Homología de Secuencia de AminoácidoRESUMEN
Although classically recognized as a neurotransmitter, gamma aminobutyric acid (GABA) has also been identified in colonic tumors. Moreover, the gut microbiome represents another potential source of GABA. Both GABAA and GABAB receptors have been implicated in contributing to the effects of GABA in colorectal cancer, with both pro- and anti-tumorigenic functions identified. However, their subunit composition is often overlooked. Studies to date have not addressed whether the GABA-producing potential of the microbiome changes over the course of colon tumor development or whether receptor subunit expression patterns are altered in colon cancer. Therefore, we investigated the clusters of orthologous group frequencies of glutamate decarboxylase (GAD) in feces from two murine models of colon cancer and found that the frequency of microbial GAD was significantly decreased early in the tumorigenic process. We also determined that microbial-derived GABA inhibited proliferation of colon cancer cells in vitro and that this effect of GABA on SW480 cells involved both GABAA and GABAB receptors. GABA also inhibited prostaglandin E2 (PGE2)-induced proliferation and interleukin-6 (IL-6) expression in these cells. Gene expression correlations were assessed using the "Cancer Exploration" suite of the TIMER2.0 web tool and identified that GABA receptor subunits were differentially expressed in human colon cancer. Moreover, GABAA receptor subunits were predominantly positively associated with PGE2 synthase, cyclooxygenase-2 and IL-6. Collectively, these data demonstrate decreased potential of the microbiome to produce GABA during tumorigenesis, a novel anti-tumorigenic pathway for GABA, and that GABA receptor subunit expression adds a further layer of complexity to GABAergic signaling in colon cancer.
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Proliferación Celular , Neoplasias del Colon , Microbioma Gastrointestinal , Receptores de GABA-A , Receptores de GABA-B , Transducción de Señal , Ácido gamma-Aminobutírico , Animales , Neoplasias del Colon/metabolismo , Neoplasias del Colon/microbiología , Neoplasias del Colon/patología , Ácido gamma-Aminobutírico/metabolismo , Humanos , Ratones , Línea Celular Tumoral , Receptores de GABA-A/metabolismo , Receptores de GABA-A/genética , Receptores de GABA-B/metabolismo , Dinoprostona/metabolismo , Glutamato Descarboxilasa/metabolismo , Interleucina-6/metabolismo , Ciclooxigenasa 2/metabolismo , Ciclooxigenasa 2/genética , Carcinogénesis , Heces/microbiología , Receptores de GABA/metabolismo , Receptores de GABA/genética , Masculino , Ratones Endogámicos C57BL , FemeninoRESUMEN
Despite studies demonstrating that inhibition of cyclooxygenase-2 (COX-2)-derived prostaglandin E2 (PGE2 ) has significant chemotherapeutic benefits in vitro and in vivo, inhibition of COX enzymes is associated with serious gastrointestinal and cardiovascular side effects, limiting the clinical utility of these drugs. PGE2 signals through four different receptors (EP1-EP4) and targeting individual receptor(s) may avoid these side effects, while retaining significant anticancer benefits. Here, we show that targeted inhibition of the EP1 receptor in the tumor cells and the tumor microenvironment resulted in the significant inhibition of tumor growth in vivo. Both dietary administration and direct injection of the EP1 receptor-specific antagonist, ONO-8713, effectively reduced the growth of established CT26 tumors in BALB/c mice, with suppression of the EP1 receptor in the tumor cells alone less effective in reducing tumor growth. This antitumor effect was associated with reduced Fas ligand expression and attenuated tumor-induced immune suppression. In particular, tumor infiltration by CD4(+) CD25(+) Foxp3(+) regulatory T cells was decreased, whereas the cytotoxic activity of isolated splenocytes against CT26 cells was increased. F4/80(+) macrophage infiltration was also decreased; however, there was no change in macrophage phenotype. These findings suggest that the EP1 receptor represents a potential target for the treatment of colon cancer.
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Neoplasias del Colon/inmunología , Modelos Animales de Enfermedad , Proteína Ligando Fas/metabolismo , Subtipo EP1 de Receptores de Prostaglandina E/efectos de los fármacos , Animales , Western Blotting , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Neoplasias del Colon/patología , Ensayo de Inmunoadsorción Enzimática , Femenino , Citometría de Flujo , Ratones , Ratones Endogámicos BALB C , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
Electrochemotherapy (ECT) is becoming an established therapy for melanoma and is under investigation for application in additional cancer types. One potential cancer type that may benefit from ECT is lung cancer as lung cancer treatments remain unable to deliver long-lasting treatment responses. Given the importance of the immune system in lung cancer, here we have also examined the impact of ECT on immune populations. The impact of electroporation and ECT on three human lung cancer cell lines (A549, H460, SK-MES 1), one murine cell line (LLC) and murine T cells, dendritic cells and macrophages was examined. The viability, metabolic activity and recovery potential post-treatment of all cell types was determined to evaluate the potential utility of ECT as a lung cancer treatment. Our findings demonstrate that cisplatin at 11 µM would be the suggested drug of choice when using ECT for lung cancer treatment. Our study also shows that T cells are not impacted by any tested condition, whilst dendritic cells and macrophages are significantly negatively impacted by electric field strengths surpassing 800 V/cm in vitro. Therefore, current ECT protocols (using 1000 V/cm in vivo) might need to adapted to improve viability of the immune population, thus improving therapy outcomes.
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Electroquimioterapia , Neoplasias Pulmonares , Melanoma , Animales , Bleomicina/uso terapéutico , Línea Celular Tumoral , Cisplatino , Electroquimioterapia/métodos , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Melanoma/tratamiento farmacológico , RatonesRESUMEN
Electrochemotherapy (ECT), the application of an electric impulse to deliver chemotherapy drugs into cells, has been in clinical trials since the early 1990s and has been used for a variety of different malignancies including melanoma and sarcoma. A standard operating procedure for the use of ECT in clinical settings has been established since 2006. ECT is very effective in reducing the local tumour burden via T-cell dependent killing of the cancer cells; however abscopal effects are not consistently observed. Currently little is known or understood about how ECT affects the immune cell population within the treated tumour and how these changes could impact the immune response. In this manuscript, we will review the current knowledge on ECT in the context of its interactions with the immune system and discuss how the gained knowledge could be harnessed to develop a potent ECT-immune co-treatment combination (Electroimmunotherapy).
Asunto(s)
Electroquimioterapia , Melanoma , Neoplasias Cutáneas , Bleomicina/uso terapéutico , Electroquimioterapia/métodos , Humanos , Melanoma/tratamiento farmacológico , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/patología , Carga TumoralRESUMEN
The IL-36 cytokines are a recently described subset of the IL-1 family of cytokines, shown to play a role in the pathogenesis of intestinal diseases such as Inflammatory Bowel Disease (IBD). Given the link between IBD and colitis -associated cancer, as well as the involvement of other IL-1 family members in intestinal tumorigenesis, the aim of this work was to investigate whether IL-36 cytokines play a role in the pathogenesis of colon cancer. Whilst research to date has focused on the role of IL-36 family members in augmenting the immune response to induce tumour rejection, very little remains known about IL-36R signalling in tumour cells in this context. In this study we demonstrate that expression of IL-36 family member mRNA and protein are significantly increased in colorectal cancer tissue compared to adjacent non-tumour. In vitro assays showed stimulation of colon cancer cell lines with IL-36R agonists resulted in the activation of the pro-tumorigenic phenotypes of increased cellular migration, invasion and proliferation in both 2D and 3D models. In addition, the IL-36 cytokines induced strong expression of pro-inflammatory chemokines in both human and murine cell lines. Intraperitoneal injection of IL-36Ra significantly reduced tumour burden using the subcutaneous CT26 tumour model in syngeneic Balb/mice, and this was associated with a decrease in Ki-67 expression by tumour cells in the IL-36Ra- treated group relative to untreated, suggesting the inhibition of the pro-proliferative signalling of IL-36 agonists resulted in the decreased tumour size. Moreover, colon cancer cells lacking the IL-36R also showed reduced tumour growth and reduced Ki-67 expression in vivo. Taken together, this data suggests that targeting IL-36R signalling may be a useful targeted therapy for colorectal cancer patients with IL-36R+ tumour cells.
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Neoplasias del Colon , Enfermedades Inflamatorias del Intestino , Animales , Carcinogénesis/genética , Transformación Celular Neoplásica , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/genética , Citocinas/metabolismo , Humanos , Enfermedades Inflamatorias del Intestino/metabolismo , Interleucina-1/genética , Interleucina-1/metabolismo , Antígeno Ki-67 , Ratones , FenotipoRESUMEN
The IL-1 family of cytokines currently comprises of seven ligands with pro-inflammatory activity (IL-1α and IL-1ß, IL-18, IL-33, IL-36α, IL-36ß, IL-36γ) as well as two ligands with anti-inflammatory activity (IL-37, IL-38). These cytokines are known to play a key role in modulating both the innate and adaptive immunes response, with dysregulation linked to a variety of autoimmune and inflammatory diseases. Given the increasing appreciation of the link between inflammation and cancer, the role of several members of this family in the pathogenesis of cancer has been extensively investigated. In this review, we highlight both the pro- and anti-tumorigenic effects identified for almost all members of this family, and explore potential underlying mechanisms accounting for these divergent effects. Such dual functions need to be carefully assessed when developing therapeutic intervention strategies targeting these cytokines in cancer.
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Interleucina-1/inmunología , Neoplasias/inmunología , Animales , HumanosRESUMEN
Fas (CD95) signalling is best known for its role in apoptosis, however, recent reports have shown it to be involved in other cellular responses as well, including inflammation. Fas and its adaptor protein FADD are known to negatively regulate LPS-induced proinflammatory responses, but their role in LPS-induced type I interferon production is unknown. Here, we demonstrate that Fas engagement on macrophages, using an agonistic Fas antibody CH11, augments LPS-induced NF-κB responses, causing increased production of TNFα, IL-8, IL-6 and IL-12. Conversely, costimulation with both LPS and CH11 causes a significant reduction in the level of interferon-beta (IFNß) production. This differential effect involves the Fas adaptor FADD because while LPS-induced IL-6 production increased in FADD-/- murine embryonic fibroblasts, LPS-induced IFNß production was significantly reduced in these cells. Overexpression of a dominant negative form of FADD (FADD-DD) inhibits LPS-induced IFNß luciferase but not LPS-induced NF-κB luciferase. In contrast, overexpression of full-length FADD inhibited LPS-induced NF-κB luciferase activation but was seen to augment LPS-induced IFNß luciferase. Moreover, FADD-DD inhibits TRIF-, TRAM-, IKKε-, TBK-1- and TRAF3-induced IFNß luciferase production, with coimmunoprecipitation experiments demonstrating an interaction between FADD and TRIF. These data identify FADD as a novel component of the noncanonical Toll-like receptor 4/IFNß signalling pathway and demonstrate that both Fas and its adaptor FADD can differentially regulate the production of LPS-induced proinflammatory cytokines and type I interferons.
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Proteína de Dominio de Muerte Asociada a Fas/genética , Interferón beta/genética , Lipopolisacáridos/farmacología , Macrófagos/efectos de los fármacos , Receptor Toll-Like 4/genética , Receptor fas/genética , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/inmunología , Proteínas Adaptadoras del Transporte Vesicular/genética , Proteínas Adaptadoras del Transporte Vesicular/inmunología , Animales , Anticuerpos/farmacología , Proteína de Dominio de Muerte Asociada a Fas/inmunología , Regulación de la Expresión Génica , Células HEK293 , Humanos , Quinasa I-kappa B/genética , Quinasa I-kappa B/inmunología , Interferón beta/inmunología , Interleucina-12/genética , Interleucina-12/inmunología , Interleucina-6/genética , Interleucina-6/inmunología , Interleucina-8/genética , Interleucina-8/inmunología , Células Jurkat , Macrófagos/citología , Macrófagos/inmunología , Ratones , FN-kappa B/genética , FN-kappa B/inmunología , Cultivo Primario de Células , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/inmunología , Células RAW 264.7 , Transducción de Señal , Células THP-1 , Factor 3 Asociado a Receptor de TNF/genética , Factor 3 Asociado a Receptor de TNF/inmunología , Receptor Toll-Like 4/inmunología , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/inmunología , Receptor fas/antagonistas & inhibidores , Receptor fas/inmunologíaRESUMEN
Microsatellite instability (MSI) is an alternative pathway of colorectal carcinogenesis. It is found in 10% to 15% of sporadic colorectal neoplasms and is characterized by failure of the DNA mismatch-repair system. High-level MSI (MSI-H) is associated with tumor-infiltrating lymphocytes (TILs) and a favorable prognosis. Expression of Fas ligand (FasL/CD95L) by cancer cells may mediate tumor immune privilege by inducing apoptosis of antitumor immune cells. The aim of this study was to investigate the relationship between FasL expression and MSI status in primary colon tumors. Using immunohistochemistry, we detected FasL expression in 91 colorectal carcinoma specimens, previously classified according to the level of MSI as MSI-H (n = 26), MSI-low (MSI-L) (n = 29), and microsatellite stable (n = 36). Tumor-infiltrating lymphocyte density was quantified by immunohistochemical staining for CD3. MSI-H tumors were significantly associated with reduced frequency (P = .04) and intensity (P = .066) of FasL expression relative to non-MSI-H (ie, microsatellite stable and MSI-L) tumors. Higher FasL staining intensity correlated with reduced TIL density (P = .059). Together, these findings suggest that the abundance of TILs found in MSI-H tumors may be due to the failure of these tumor cells to up-regulate FasL and may explain, in part, the improved prognosis associated with these tumors.