RNA-seq analysis identifies regulators of macrophage reprogramming from M2 to M1-like phenotype
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ABSTRACT: We analyzed expression profiles of thioglyolate elicted peritoneal exudate cells (peritoneal macrophages). Peritoneal macrophages were polarized into M1 and M2 macrophages by activation with IFNgamma+ LPS and Il4, respectively. We predicted a gene-regulatory network, which consists of four transcription factors (E2f1,Myc, Stat6, Pparg) regulating metabolic genes, M1 and M2-associated genes. The predicted regulators were all active in M2 macrophages. We hypothesized that these transcription factors are essential regulators to maintain M2 phenotype. To further validate our findings, we treated M2-polarized macrophages with siRNA-pools targeting E2f1, Myc, Stat6 and Pparg. In this study, we observed a switch towards an M1-like phenotype after transfection of siRNA-pools. In addition, Inflammatory pathways were upreguated while fatty acid metabolism was down regulated.
Project description:Monocytes mature to macrophages in the presence of the lineage determining cytokine M-CSF. They can be further polarized into M1 or M2 macrophages with distinct functional properties. We used microarrays to detail the global programme of gene expression underlying macrophage maturation and polarization and identified distinct classes of up-regulated genes during this process. Experiment Overall Design: Freshly isolated monocytes were cultured in the presence of M-CSF for 7 days, and then polarized to M1 or M2 cells. The study includes Monocytes at day 0, macrophages at day 3 and 7, M1 and m2 polarized macrophages.
Project description:Purpose: RNA-sequencing was performed to identify gene expression changes between bone marrow derived macrophages isolated from wildtype and mirn23a-/- (Mirc11-/-) mice that were either M1 or M2 polarized. Results: Diferential gene expression was examined between wildtype and mirn23a-/- M1 polarized macrophages and wildtype and mirn23a-/- M2 polarized macrophages. The number of genes with significant (p<0.05) 2-fold changes in our M1 dataset is 4-fold higher than the 2-fold changed genes in our M2 dataset. 43 unique genes were differentially expressed over 2-fold in M1 mutant macrophages compared to wildtype with 29 upregulated and 14 downregulated. 10 genes (8 downregulated/ 2 upregulated)were differentially expressed in mirn23a-/- M2 macrophages by at least 2-fold compared to wildtype.
Project description:We reported exosome-guided phenotype switches between M1- and M2-polarized BMDMs. M1- or M2-polarized BMDMs were successfully reprogrammed to M2- or M1-phenotype via the treatment of exosomes obtained from M2- or M1-polarized BMDMs. In this uploaded information, the exosomes from M1- and M2-polarized BMDMs were analyzed by high-throughput sequencing.
Project description:In this study, we used mass spectrometry and label-free quantification (LFQ) to characterize the global proteomics of polarized (M1, M2a, M2b, M2c, and M2d) and unpolarized (M0) phenotypes of macrophages from human THP-1 monocytes. The results described the biological functions of the four M2 macrophages subtypes and provided available references for identifying M2 macrophages or subtypes of M2 macrophages.
Project description:Lung cancer, a prevalent and aggressive disease, is characterized by recurrence and drug resistance. Understanding the underlying mechanisms and identifying new therapeutic targets are crucial for improving treatment outcomes. Our research indicates that methylation of cg09897064 and reduced expression of ZBP1 are associated with poor prognosis in lung cancer patients. Furthermore, these factors play a role in macrophage polarization, with ZBP1 upregulated in M1 macrophages compared to both M0 and M2 polarized macrophages. We observed cg09897064 methylation in M2 polarization, but not in M0 and M1 polarized macrophages. ATACseq analysis revealed closed chromatin accessibility of ZBP1 in M0 polarized macrophages, while open accessibility was observed in both M1 and M2 polarized macrophages. Our findings suggest that ZBP1 is downregulated in M0 polarized macrophages due to closed chromatin accessibility and downregulated in M2 polarized macrophages due to cg09897064 methylation. Further investigations manipulating cg09897064 methylation and ZBP1 expression through overexpression plasmids and shRNAs provided evidence for their role in modulating macrophage polarization and tumor growth. ZBP1 inhibits M2 polarization and suppresses tumor growth, while cg09897064 methylation promotes M2 polarization and macrophage-induced tumor growth. In mechanism investigations, we found that cg09897064 methylation impairs CEBPA binding to the ZBP1 promoter, leading to decreased ZBP1 expression. Targeting these factors may hold promise as a strategy for developing innovative checkpoint inhibitors in lung cancer treatment.
Project description:Bone-marrow macrophages polarized to M2 phenotype are immunosuppressive. Interestingly, treatment with whole-glucan particles converts M2 macrophages to M1 phenotype with an anti-tumor phenotype. In this study, the effect of WGP treatment for 6 hours on the gene expression of M2 macrophages was assessed.
Project description:Analysis of the effects of CNI-1493 treatment on M1 and M2 polarized macrophages. The purpose of this microarray is to identify genes that may be differentially expressed in M1 or M2 macrophages after treatment with CNI-1493. CNI-1493 is a known inhibitor of M1 macrophages but details of its molecular mechanism are unknown. The effect of CNI-1493 on M2 macrophages has yet to be explored, but we hypothesize that CNI-1493 treatment will attenuate pro-tumor properties of M2 macrophages. We demonstrate with this array that known macrophage markers are unchanged after treatment with CNI-1493, indicating that CNI-1493 does not change the macrophage phenotype on a transcriptional level. Additionally, no candidate genes to suggest how CNI-1493 may attenuate the pro-tumor effects of M2 macrophages are readily identifiable. Total RNA extracted from M1 or M2 macrophages after polarization with GM-CSF (25ng/ml) or M-CSF (25ng/ml) for 7 days, followed by addition of IFN-γ (20ng/ml) and LPS (100ng/ml) or IL-4 (40ng/ml) for 18 hours, respectively, from CD14+ human PBMCs, and treated with CNI-1493 (200nM)
Project description:in the present study we investigate polarized canine macrophages using transcriptome sequencing, a larger panel of flow cytometry markers, and antimicrobial functional assays. Transcriptome analysis of primary canine monocyte-derived macrophages revealed unique, previously unreported signatures for polarized M1 and M2 macrophages.
Project description:The M2 phenotype is controlled by key transcription factors such as STAT6. We describe an engineered exosome therapeutic candidate delivering an antisense oligonucleotide (ASO) targeting STAT6 (exoASO-STAT6), which preferentially silences STAT6 expression in tumor-associated macrophages (TAMs). We demonstrate that administration of exoASO-STAT6 reprograms TAMs to an M1 phenotype, leading to the induction of nitric oxide synthase 2 (NOS2) and remodeling of the TAMs.
Project description:We showed that co-culture with TAMs triggered Bmi1 expression in gastrointestinal cancer cell lines. miRNAs have been found to target various oncogenes and tumor suppressors. We therefore hypothesized that the regulation of Bmi1 expression in gastrointestinal cancer cells may be mediated by miRNAs using miRNA microarray analysis. THP-1 cells were seeded in the transwell inserts (3540, Corning) for 6-well plates (1 M-CM-^W 106 cells/well). For preparation of M1-polarized THP-1 macrophages, 320 nM phorbol myristate acetate (PMA) was added to THP-1 cells for 6 h, followed by PMA plus 20 ng/ml interferon (IFN)-M-NM-3 and 100 ng/ml lipopolysaccharide for the following 18 h. For preparation of M2-polarized THP-1 macrophages, 320 nM PMA was added to THP-1 cells for 6 h, followed by PMA plus 20 ng/ml interleukin (IL)-4/IL-13 for the following 18 h. After three washes to remove cytokines, M1- or M2-polarized THP-1 macrophages were co-cultured in upper inserts with AGS cells in 6-well plates (1 M-CM-^W 105 cells/well) without direct contact, in each medium without 10% FBS as described above. After 24 h of co-culture, the upper inserts containing macrophages were discarded. AGS cells were collected and analyzed to identify downregulated microRNA in a gastric cancer cell line co-cultured with M1- or M2-polarized macrophage.