Low-density miRNA microarray of induced mouse myeloid-derived suppressor cell
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE50914 | GEO | 2014/03/11
SECONDARY ACCESSION(S): PRJNA219401
REPOSITORIES: GEO
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Project description:Myeloid-derived suppressor cells (MDSCs) are one of the main cell populations that negatively regulate immune responses. However, the mechanism underlying the differentiation and expansion of MDSCs remains unclear. Employing a low-density miRNA microarray and a TaqMan probe-based qRT-PCR assay, we investigated the miRNAs expression profiles of MDSC. Compared to freshly isolated BM cells, BM-MDSCs had a significantly altered miRNA profile. There were 45 miRNAs that were upregulated and 17 miRNAs that were downregulated more than 2-fold. Among these miRNAs, miR-155 and miR-21 have the highest upregulation after cytokine induction. BM cells were planted into dishes using RPMI1640 medium supplemented with 2 mM L-glutamine, 10mM HEPES, 20M-BM-5M 2-mercaptoethanol, 150 U/ml streptomycin, 200 U/ml penicillin, 10% FBS and stimulated with combinations of GM-CSF (40ng/ml), and IL-6 (40ng/ml). Cells were cultured at 37M-BM-0C in 5% CO2-humidified atmosphere for 4 days.Total RNA were isolated, low-density miRNA microarray was performed
2014-03-11 | E-GEOD-50914 | biostudies-arrayexpress
Project description:Myeloid-derived suppressor cells (MDSCs) are immature myeloid cells that accumulate in the tumor microenvironment of most cancer patients. There MDSCs suppress both adaptive and innate immune responses, hindering immunotherapies. Moreover, many cancers are accompanied by inflammation, a processes that further intensifies MDSC suppressive activity, causing aggressive tumor progression and metastasis. MDSCs collected from tumor-bearing mice profusely release nano-scale membrane-bound extracellular vesicles, called exosomes, which carry biologically active proteins between cells and contribute directly to the immune suppressive functions of MDSC. Many studies on other cell types have shown that exosomes may also carry microRNAs (miRNAs) and messenger RNAs (mRNAs) which can also be transferred to surrounding and distant cells. However, to the best of our knowledge, the miRNA and mRNA cargo of MDSC-derived exosomes has not yet been interrogated. This study aims to identify and quantify the cargo of MDSC and their immunosuppressive exosomes to gather knowledge that can offer insights on the mechanisms by which MDSCs contribute to immune suppression, focusing on the role of exosomes as intercellular communication mediators in the tumor microenvironment. In order to achieve our objective a well-established mouse model based on a conventional mammary carcinoma (4T1 cells) and heightened inflammation (4T1 transduced to express the cytokine interleukin-1b) was used. We provide evidence that MDSC-derived exosomes carry proteins, mRNAs and miRNAs. Relative quantitation demonstrated quantitative differences between the exosome cargo and the cargo of their parental cells, supporting the hypothesis that selective loading into the exosomes is possible. Additionally, quantitative and functional analyses of the exosome cargo generated under conventional and heightened inflammation conditions are consistent with clinical observations that inflammation is linked to cancer development.
2017-11-17 | PXD006204 | Pride
Project description:Myeloid-derived suppressor cells (MDSCs), which accumulate in tumor bearers, are known to suppress anti-tumor immunity and thus promote tumor progression. MDSCs are considered a major cause of resistance against immune checkpoint inhibitors in patients with cancer. Therefore, MDSCs are potential targets in cancer immunotherapy. In this study, we modified an in vitro method of MDSC differentiation. Upon stimulating bone marrow (BM) cells with granulocyte-macrophage colony-stimulating factor in vitro, we obtained both lymphocyte antigen 6G positive (Ly-6G+) and negative (Ly-6G−) MDSCs (collectively, hereafter referred to as conventional MDSCs), which were non-immunosuppressive and immunosuppressive, respectively. We then found that MDSCs differentiated from Ly-6G− BM (hereafter called 6G− BM-MDSC) suppressed T-cell proliferation more strongly than conventional MDSCs, whereas the cells differentiated from Ly-6G+ BM (hereafter called 6G+ BM-MDSC) were non-immunosuppressive. In line with this, conventional MDSCs or 6G− BM-MDSC, but not 6G+ BM-MDSC, promoted tumor progression in tumor-bearing mice. Moreover, we identified that activated glutathione metabolism was responsible for the enhanced immunosuppressive ability of 6G− BM-MDSC. Finally, we showed that Ly-6G+ cells in 6G− BM-MDSC, which exhibited weak immunosuppression, expressed higher levels of Cybb mRNA, an immunosuppressive gene of MDSCs, than 6G+ BM-MDSC. Together, these data suggest that the depletion of Ly-6G+ cells from the BM cells leads to differentiation of immunosuppressive Ly-6G+ MDSCs. In summary, we propose a better method for MDSC differentiation in vitro. Moreover, our findings contribute to the understanding of MDSC subpopulations and provide a basis for further research on MDSCs.
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Project description:Tumor growth is associated with a profound alteration of myelopoiesis, leading to recruitment of immunosuppressive cells known as myeloid-derived suppressor cells (MDSCs). Immuno-regulatory activity of both tumor-induced and BM-derived MDSCs (by GM-CSF and IL-6 treatment) was entirely dependent on C/EBP transcription factor (TF), a key component of the emergency myelopoiesis triggered by stress and inflammation. We used miR expression analysis to identify miRs which could drive MDSC recruitment/generation/activity by modulating specific TFs and pathway. In particular, we identified a miR signature of 79 miR differentially expressed between not suppressive CD11b+ cells and CD11b+ isolated from tumor mass and spleen of tumor-bearing mice. Moreover on the same samples we profiled gene expression with Affymetrix microarrays to perform an integrated analysis of mirna and gene expression. Keywords: Expression profiling by array
2015-07-23 | GSE39807 | GEO
Project description:Myeloid-derived suppressor cells (MDSCs) potently suppress the anti-tumor immune responses and also orchestrate the tumor microenvironment that favors tumor angiogenesis and metastasis. The immunosuppressive activity of MDSCs has been extensively investigated, however the molecular networks regulating the non-immunological functions of tumor-expanded MDSCs, are largely unknown. In this study, we identified microRNA-494 (miR-494), whose expression was dramatically induced by tumor-derived factors (TDFs), as an essential player, in regulating the non-immunological activity of MDSCs by targeting PTEN and activating the Akt pathway. TGF-beta 1 was found to be a main tumor-derived factor responsible for the up-regulation of miR-494 in MDSCs. Expression of miR-494 not only enhanced CXCR4-mediated MDSC chemotaxis but also altered the intrinsic apoptotic/survival signal by targeting PTEN, thus contributing to the accumulation of MDSCs in tumor tissues. Consequently, down-regulation of PTEN resulted in increased activity of the Akt pathway and the subsequent up-regulation of matrix metalloproteinases (MMPs) for facilitating tumor cell invasion and metastasis. Knock down of miR-494 significantly reversed the activity of MDSCs and inhibited the tumor growth and metastasis of 4T1 murine breast cancer in vivo. Collectively, our findings reveal that TGF-beta 1-induced miR-494 expression in MDSCs plays a critical role in the molecular events governing the accumulation and non-immunological functions of tumor-expanded MDSCs, and might be identified as a potential target in cancer therapy. BALB/c mice (female, 6- to 8-wk-old) were injected subcutaneously in the mammary fat pad with 100,000 4T1 tumor cells. Three weeks later, tumor-bearing animals were used for the indicated studies. Gr-1+ CD11b+ cells were isolated by immunomagnetic selection from the bone marrow of 4T1 tumor-bearing mice (n=3) and tumor-free BALB/c mice (n=3), then the miRNA expression profile were analyzed using miRCURY LNAM-bM-^DM-" microRNA Arrays.
2012-03-20 | E-GEOD-36309 | biostudies-arrayexpress
Project description:Background and methods: Myeloid-derived suppressor cells (MDSCs) are immature myeloid cells which originate in the bone marrow (BM) and have immunoregulatory functions. MDSCs have been implicated in the pathogenesis of several autoimmune diseases but not in immune aplastic anemia (AA). We examined the roles of granulocytic-MDSCs (G-MDSCs) in murine models of human AA and bone marrow failure (BMF). To perform Totalseq, bone marrow mononuclear cells were FACS sorted to obtain alive cells based on FSC and SSC from five bone marrow failure control mice and five G-MDSC-treated mice, mRNA profiles of single cells were generated and sequenced on an Illumina Novaseq System. Results: As both prophylaxis and therapy, BM-derived G-MDSCs improved pancytopenia and BM cellularity and decreased BM T cell infiltration in major histocompatibility (MHC)-matched CB10 BMF mice. Single cell RNA sequencing demonstrated that G-MDSCs downregulated cell cycle related pathways in BM infiltrated T cells, consistent with suppression of T cell proliferation by G-MDSCs. Conclusion: Our results demonstrate that BM-derived G-MDSCs improved pancytopenia and BM cellularity and decreased BM T cell infiltration in major histocompatibility (MHC)-matched CB10 BMF mice, but not in MHC-mismatched CByB6F1 BMF model. Therapeutic efficacy of G-MDSCs are immune context-dependent. Bone marrow mononuclear cells were FACS sorted to obtain alive cells based on FSC and SSC from five bone marrow failure control mice and five G-MDSC-treated mice.
2022-07-27 | GSE193421 | GEO
Project description:Tumor progression is accompanied by an altered myelopoiesis that causes the accumulation of cells inhibiting anti-tumor T lymphocytes. We previously reported that immunosuppressive cells can be generated in vitro from bone marrow cells (BM) after four days GM-CSF and IL-6 treatment. Here, we describe that miR-142-3p down-regulation directs macrophage differentiation and determines the acquisition of their immunosuppressive function in cancer. Enforced miR over-expression impaired monocyte to macrophage transition both in vitro and in vivo. Conversely, forced miR down-regulation promoted the generation of immunosuppressive macrophages even during G-CSF-induced granulocytic differentiation. To identify how miR-142-3p regulates MDSC generation and activity, we analyze the gene expression of BM cultures transfected with either CTRL- or miR 142-3p mimic oligo -transfected before four days GM-CSF and IL-6 treatment. Keywords: Expression profiling by array BM cells were transfected either CTRL- or miR 142-3p mimic oligo before GM-CSF and IL-6 treatment to generate in vitro MDSCs during enforced miR over-expression. A triplicate of each sample was considered.Total RNA from obtained in vitro BM-differentiated MDSCs was isolated by Trizol reagent, and cRNA samples were hybridized to the Affymetrix GeneChip MOE430 2.0.
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2015-02-20 | E-GEOD-64484 | biostudies-arrayexpress
Project description:Tumor growth is associated with a profound alteration of myelopoiesis, leading to recruitment of immunosuppressive cells known as myeloid-derived suppressor cells (MDSCs). Analyzing the cytokines affecting myelo-monocytic differentiation produced by various experimental tumors, we found that GM-CSF, G-CSF, and IL-6 allowed a rapid generation of MDSCs from precursors present in mouse and human bone marrow (BM). BM-MDSCs induced by GM-CSF+IL-6 possessed the highest tolerogenic activity, as revealed by the ability to impair the priming of IFN- -producing CD8+ T cells upon in vivo adoptive transfer. Moreover, adoptive transfer of syngeneic, GM-CSF+IL-6-conditioned MDSCs to diabetic mice transplanted with allogeneic pancreatic islets resulted in long term acceptance of the allograft and correction of the diabetic status. Cytokines inducing MDSCs acted on a common molecular pathway. Immunoregulatory activity of both tumor-induced and BM-derived MDSCs was entirely dependent on C/EBP transcription factor, a key component of the emergency myelopoiesis triggered by stress and inflammation. Adoptive transfer of tumor antigen-specific CD8+ T lymphocytes resulted in therapy of established tumors only in mice lacking C/EBP in myeloid compartment. These data unveil another link between inflammation and cancer and identify a novel molecular target to control tumor-induced immune suppression. We used gene expression analysis to identify those factors, secreted by tumor-infiltrating MDSC, which could drive emathopoiesis. Moreover we compare gene expression profile of tumor-induced MDSC, obtained from either the spleen and the tumor infiltrate of tumor bearing mice, and in vitro bone marrow-derived MDSC. CD11b+ cells were immunomagnetically enriched from various murine tissue and experimental conditions, and cRNA samples were prepared accordingly to Expression Analysis: Technical Manual. 701021 Rev. 5. Santa Clara, CA, Affymetrix; 2004, and hybridized to the Affymetrix GeneChip MOE430 2.0 array which contains more than 45,000 probe sets, representing more than 34,000 genes. CD11b+ cells obtained from the spleen of healthy BALB/c and C57BL/6 mice were used as reference sample for tumor induced CD11b+ MDSC, enriched from either the spleen and the tumor infiltrate of tumor-bearing mice. Moreover CD11b+ cells enriched from fresh bone marrow were used as reference sample for in vitro bone marrow-differentiated MDSC, obtained with either GM-CSF+IL-6 and GM-CSF+G-CSF 4 days cytokine cocktail treatment.
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