Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcriptional profiling of THP1-derived human macrophages with different stimuli

ABSTRACT: Macrophages are key cell types of the innate immune system regulating host defense, inflammation, tissue homeostasis and cancer. Within this functional spectrum diverse and often opposing phenotypes are displayed which are dictated by environmental clues and depend on highly plastic transcriptional programs. Among these the ‘classical’ (M1) and ‘alternative’ (M2) macrophage polarization phenotypes are the best characterized. Understanding macrophage polarization in humans may reveal novel therapeutic intervention possibilities for chronic inflammation, wound healing and cancer. Systematic loss of function screening in human primary macrophages is limited due to lack of robust gene delivery methods and limited sample availability. To overcome these hurdles we developed cell-autonomous assays using the THP-1 cell line allowing genetic screens for human macrophage phenotypes. To confirm the relevance of the THP1-based system we performed microarray studies on THP1 cells and primary human cells subjected to various conditions. THP1 cells were treated with PMA (phorbol 12-myristate 13-acetate) to derive macrophages which were then cultured 1, 3 or 6 days with different stimuli.

ORGANISM(S): Homo sapiens

SUBMITTER: Florian Nigsch

PROVIDER: E-MTAB-5917 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Multidimensional pooled shRNA screens in human THP-1 cells identify candidate modulators of macrophage polarization.

Surdziel Ewa E Clay Ieuan I Nigsch Florian F Thiemeyer Anke A Allard Cyril C Hoffman Gregory G Reece-Hoyes John S JS Phadke Tanushree T Gambert Romain R Keller Caroline Gubser CG Ludwig Marie-Gabrielle MG Baumgarten Birgit B Frederiksen Mathias M Schübeler Dirk D Seuwen Klaus K Bouwmeester Tewis T Fodor Barna D BD

PloS one 20170824 8

Macrophages are key cell types of the innate immune system regulating host defense, inflammation, tissue homeostasis and cancer. Within this functional spectrum diverse and often opposing phenotypes are displayed which are dictated by environmental clues and depend on highly plastic transcriptional programs. Among these the 'classical' (M1) and 'alternative' (M2) macrophage polarization phenotypes are the best characterized. Understanding macrophage polarization in humans may reveal novel therap ...[more]

PMID: 28837623

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Project description:OBJECTIVE: The liver X receptor M-NM-1 (LXRM-NM-1) is a ligand-dependent nuclear receptor and the major regulator of reverse cholesterol transport in macrophages. This makes it an interesting target for mechanistic study and treatment of atherosclerosis. METHODS AND RESULTS: We optimized a promising stilbenoid structure (STX4) in order to reach nanomolar effective concentrations in LXRM-NM-1 reporter-gene assays. STX4 displayed the unique property to activate LXRM-NM-1 effectively but not its subtype LXRM-NM-2. The potential of STX4 to increase transcriptional activity as an LXRM-NM-1 ligand was tested with gene expression analyses in THP1-derived human macrophages and oxLDL-loaded human foam cells. Only in foam cells but not in macrophage cells STX4 treatment showed athero-protective effects with similar potency as the synthetic LXR ligand T0901317 (T09). Surprisingly, combinatorial treatment with STX4 and T09 resulted in an additive effect on reporter-gene activation and target gene expression. In physiological tests the cellular content of total and esterified cholesterol was significantly reduced by STX4 without the undesirable increase in triglyceride levels as observed for T09. CONCLUSIONS: STX4 is a new LXRM-NM-1-ligand to study transcriptional regulation of anti-atherogenic processes in cell or ex vivo models, and provides a promising lead structure for pharmaceutical development. 2 treatment groups (STX4 vs. DMSO in foam cells), 3 biological replicates per treatment (6 samples in total), each representing one well from cell culture plate 2 treatment groups (STX4 vs. DMSO in macrophages), 4 biological replicates per treatment (8 samples in total), each representing one well from cell culture plate 2 treatment groups (T0901317 vs. DMSO in foam cells), 3-4 biological replicates per treatment (7 samples in total), each representing one well from cell culture plate 2 treatment groups (T0901317 vs. DMSO in macrophages), 4 biological replicates per treatment (8 samples in total), each representing one well from cell culture plate

Project description:Nasu-Hakola disease (NHD), also designated polycystic lipomembranous osteodysplasia with sclerosing leukoencephalopathy (PLOSL), is a rare autosomal recessive disorder characterized by progressive presenile dementia and formation of multifocal bone cysts, caused by a loss-of-function mutation of DAP12 or TREM2. TREM2 and DAP12 constitute a receptor/adaptor complex expressed on osteoclasts, dendritic cells, macrophages, monocytes, and microglia. At present, the precise molecular mechanisms underlying development of leukoencephalopathy and bone cysts in NHD remain largely unknown. We established THP-1 human monocyte clones that stably express small interfering RNA (siRNA) targeting DAP12 for serving as a cellular model of NHD. Genome-wide transcriptome analysis identified a set of 22 genes consistently downregulated in DAP12 knockdown cells. They constituted the molecular network closely related to the network defined by cell-to-cell signaling and interaction, hematological system development and function, and inflammatory response, where NF-kappaB acts as a central regulator. These results suggest that a molecular defect of DAP12 in human monocytes deregulates the gene network pivotal for maintenance of myeloid cell function in NHD. We found that both DAP12 knockdown and control clones were capable of equally responding to phorbol 12-myristate 13-acetate (PMA), a known inducer of morphological differentiation of THP-1 cells, by exhibiting almost similar gene expression profiles between both, following a 24-hour exposure to 50 nM PMA. The siRNA vector construct targeting the DAP12 sequence (SI) and the control vector construct targeting the scrambled sequence (SCR) were generated by using GeneClip U1 Hairpin cloning system (Promega). The vectors were transfected in THP-1 cells by using Lipofectamine LTX reagent. The stable cell lines were selected by incubating them for approximately two months in the feeding medium with inclusion of 200 microgram/ml Hygromycin B. Then, two SI clones named SI5 and SI17, in addition to two SCR clones named SCR1 and SCR4, were selected by limiting dilution of the cells in a manner of a single cell per well plated in a 96-well cell culture plate. Microarray data for clones SCR4 and SI17 submitted in this Series.

Dataset Information

Transcriptional profiling of THP1-derived human macrophages with different stimuli

Publications

Multidimensional pooled shRNA screens in human THP-1 cells identify candidate modulators of macrophage polarization.

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