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The IL1?-IL1R signaling is involved in the stimulatory effects triggered by hypoxia in breast cancer cells and cancer-associated fibroblasts (CAFs).

ABSTRACT: BACKGROUND:Hypoxia plays a relevant role in tumor-related inflammation toward the metastatic spread and cancer aggressiveness. The pro-inflammatory cytokine interleukin-1? (IL-?) and its cognate receptor IL1R1 contribute to the initiation and progression of breast cancer determining pro-tumorigenic inflammatory responses. The transcriptional target of the hypoxia inducible factor-1? (HIF-1?) namely the G protein estrogen receptor (GPER) mediates a feedforward loop coupling IL-1? induction by breast cancer-associated fibroblasts (CAFs) to IL1R1 expression by breast cancer cells toward the regulation of target genes and relevant biological responses. METHODS:In order to ascertain the correlation of IL-? with HIF-1? and further hypoxia-related genes in triple-negative breast cancer (TNBC) patients, a bioinformatics analysis was performed using the information provided by The Invasive Breast Cancer Cohort of The Cancer Genome Atlas (TCGA) project and Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) datasets. Gene expression correlation, statistical analysis and gene set enrichment analysis (GSEA) were carried out with R studio packages. Pathway enrichment analysis was evaluated with Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway. TNBC cells and primary CAFs were used as model system. The molecular mechanisms implicated in the regulation of IL-1? by hypoxia toward a metastatic gene expression profile and invasive properties were assessed performing gene and protein expression studies, PCR arrays, gene silencing and immunofluorescence analysis, co-immunoprecipitation and ChiP assays, ELISA, cell spreading, invasion and spheroid formation. RESULTS:We first determined that IL-1? expression correlates with the levels of HIF-1? as well as with a hypoxia-related gene signature in TNBC patients. Next, we demonstrated that hypoxia triggers a functional liaison among HIF-1?, GPER and the IL-1?/IL1R1 signaling toward a metastatic gene signature and a feed-forward loop of IL-1? that leads to proliferative and invasive responses in TNBC cells. Furthermore, we found that the IL-1? released in the conditioned medium of TNBC cells exposed to hypoxic conditions promotes an invasive phenotype of CAFs. CONCLUSIONS:Our data shed new light on the role of hypoxia in the activation of the IL-1?/IL1R1 signaling, which in turn triggers aggressive features in both TNBC cells and CAFs. Hence, our findings provide novel evidence regarding the mechanisms through which the hypoxic tumor microenvironment may contribute to breast cancer progression and suggest further targets useful in more comprehensive therapeutic strategies.

SUBMITTER: Lappano R

PROVIDER: S-EPMC7418191 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature

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The IL1β-IL1R signaling is involved in the stimulatory effects triggered by hypoxia in breast cancer cells and cancer-associated fibroblasts (CAFs).

Lappano Rosamaria R Talia Marianna M Cirillo Francesca F Rigiracciolo Damiano Cosimo DC Scordamaglia Domenica D Guzzi Rita R Miglietta Anna Maria AM De Francesco Ernestina Marianna EM Belfiore Antonino A Sims Andrew H AH Maggiolini Marcello M

Journal of experimental & clinical cancer research : CR 20200810 1

<h4>Background</h4>Hypoxia plays a relevant role in tumor-related inflammation toward the metastatic spread and cancer aggressiveness. The pro-inflammatory cytokine interleukin-1β (IL-β) and its cognate receptor IL1R1 contribute to the initiation and progression of breast cancer determining pro-tumorigenic inflammatory responses. The transcriptional target of the hypoxia inducible factor-1α (HIF-1α) namely the G protein estrogen receptor (GPER) mediates a feedforward loop coupling IL-1β inductio ...[more]

PMID: 32778144

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Project description:IntroductionCarcinoma-associated fibroblasts (CAFs) play a pivotal role in cancer progression by contributing to invasion, metastasis and angiogenesis. Solid tumors possess a unique microenvironment characterized by local hypoxia, which induces gene expression changes and biological features leading to poor outcomes. Hypoxia Inducible Factor 1 (HIF-1) is the main transcription factor that mediates the cell response to hypoxia through different mechanisms that include the regulation of genes strongly associated with cancer aggressiveness. Among the HIF-1 target genes, the G-protein estrogen receptor (GPER) exerts a stimulatory role in diverse types of cancer cells and in CAFs.MethodsWe evaluated the regulation and function of the key angiogenic mediator vascular endothelial growth factor (VEGF) in CAFs exposed to hypoxia. Gene expression studies, Western blotting analysis and immunofluorescence experiments were performed in CAFs and breast cancer cells in the presence of cobalt chloride (CoCl₂) or cultured under low oxygen tension (2% O₂), in order to analyze the involvement of the HIF-1α/GPER signaling in the biological responses to hypoxia. We also explored the role of the HIF-1α/GPER transduction pathway in functional assays like tube formation in human umbilical vein endothelial cells (HUVECs) and cell migration in CAFs.ResultsWe first determined that hypoxia induces the expression of HIF-1α and GPER in CAFs, then we ascertained that the HIF-1α/GPER signaling is involved in the regulation of VEGF expression in breast cancer cells and in CAFs exposed to hypoxia. We also assessed by ChIP assay that HIF-1α and GPER are both recruited to the VEGF promoter sequence and required for VEGF promoter stimulation upon hypoxic condition. As a biological counterpart of these findings, conditioned medium from hypoxic CAFs promoted tube formation in HUVECs in a HIF-1α/GPER dependent manner. The functional cooperation between HIF-1α and GPER in CAFs was also evidenced in the hypoxia-induced cell migration, which involved a further target of the HIF-1α/GPER signaling like connective tissue growth factor (CTGF).ConclusionsThe present results provide novel insight into the role elicited by the HIF-1α/GPER transduction pathway in CAFs towards the hypoxia-dependent tumor angiogenesis. Our findings further extend the molecular mechanisms through which the tumor microenvironment may contribute to cancer progression.

Project description:BackgroundThe chemical carcinogen 3-methylcholanthrene (3MC) binds to the aryl hydrocarbon receptor (AHR) that regulates the expression of cytochrome P450 (CYP) enzymes as CYP1B1, which is involved in the oncogenic activation of environmental pollutants as well as in the estrogen biosynthesis and metabolism. 3MC was shown to induce estrogenic responses binding to the estrogen receptor (ER) ? and stimulating a functional interaction between AHR and ER?. Recently, the G protein estrogen receptor (GPER) has been reported to mediate certain biological responses induced by endogenous estrogens and environmental compounds eliciting an estrogen-like activity.MethodsMolecular dynamics and docking simulations were performed to evaluate the potential of 3MC to interact with GPER. SkBr3 breast cancer cells and cancer-associated fibroblasts (CAFs) derived from breast tumor patients were used as model system. Real-time PCR and western blotting analysis were performed in order to evaluate the activation of transduction mediators as well as the mRNA and protein levels of CYP1B1 and cyclin D1. Co-immunoprecipitation studies were performed in order to explore the potential of 3MC to trigger the association of GPER with AHR and EGFR. Luciferase assays were carried out to determine the activity of CYP1B1 promoter deletion constructs upon 3MC exposure, while the nuclear shuttle of AHR induced by 3MC was assessed through confocal microscopy. Cell proliferation stimulated by 3MC was determined as biological counterpart of the aforementioned experimental assays. The statistical analysis was performed by ANOVA.ResultsWe first ascertained by docking simulations the ability of 3MC to interact with GPER. Thereafter, we established that 3MC activates the EGFR/ERK/c-Fos transduction signaling through both AHR and GPER in SkBr3 cells and CAFs. Then, we found that these receptors are involved in the up-regulation of CYP1B1 and cyclin D1 as well as in the stimulation of growth responses induced by 3MC.ConclusionsIn the present study we have provided novel insights regarding the molecular mechanisms by which 3MC may trigger a physical and functional interaction between AHR and GPER, leading to the stimulation of both SkBr3 breast cancer cells and CAFs. Altogether, our results indicate that 3MC may engage both GPER and AHR transduction pathways toward breast cancer progression.

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The IL1?-IL1R signaling is involved in the stimulatory effects triggered by hypoxia in breast cancer cells and cancer-associated fibroblasts (CAFs).

Publications

The IL1β-IL1R signaling is involved in the stimulatory effects triggered by hypoxia in breast cancer cells and cancer-associated fibroblasts (CAFs).

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