Effect of human CD5 molecule-like (CD5L) overexpression on PMA-differentiated THP1 human monocyte-like cells
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ABSTRACT:
OTHER RELATED OMICS DATASETS IN: GSE111315
PROVIDER: PRJNA436578 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:To assay the differential gene expression in human THP1-PMA differentiated macrophages expressing human CD5 molecule-like (CD5L) compared to THP1 empty vector control cells. Abstract from associated publication: Lucía Sanjurjo, Gemma Aran, Érica Téllez, Núria Amézaga, Carolina Armengol, Daniel López, Clara Prats and Maria-Rosa Sarrias. CD5L Promotes M2 Macrophage Polarization Through Autophagy-Mediated Upregulation of ID3. Frontiers in Immunology. 9:480. doi: 10.3389/fimmu.2018.00480 (2018). ABSTRACT: CD5L (CD5 molecule-like) is a secreted glycoprotein that controls key mechanisms in inflammatory responses, with involvement in processes such as infection, atherosclerosis, and cancer. In macrophages, CD5L promotes an anti-inflammatory cytokine profile in response to TLR activation. In the present study, we questioned whether CD5L is able to influence human macrophage plasticity, and drive its polarization towards any specific phenotype. We compared CD5L-induced phenotypic and functional changes to those caused by IFN/LPS, IL4 and IL10 in human monocytes. Phenotypic markers were quantified by RT-qPCR and flow cytometry, and a mathematical algorithm was built for their analysis. Moreover, we compared ROS production, phagocytic capacity and inflammatory responses to LPS. CD5L drove cells towards a polarization similar to that induced by IL10. Furthermore, IL10- and CD5L-treated macrophages showed increased LC3II content and colocalization with acidic compartments, thereby pointing to the enhancement of autophagy-dependent processes. Accordingly, siRNA targeting ATG7 in THP1 cells blocked CD5L-induced CD163 and MERTK mRNA and efferocytosis. In these cells, gene expression profiling and validation indicated the upregulation of the transcription factor ID3 by CD5L through ATG7. In agreement, ID3 silencing reversed polarization by CD5L. Our data point to a significant contribution of CD5L-mediated autophagy to the induction of ID3 and provide the first evidence that CD5L drives macrophage polarization.
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Project description:Macrophages are sentinels of the immune system and THP1 monocytic cells are one of the widely used models to study immune responses in macrophages. Several monocyte-to-macrophage differentiation protocols exist with phorbol 12-myristate-13-acetate (PMA) being widely used and accepted. However, the concentrations and durations of PMA treatment to induce differentiation varies widely in published literature. In this study, we determined the proteome expression dynamics of THP1 macrophages differentiated from monocytes by three commonly used PMA-based differentiation protocols using dimethyl labeling-based quantitative proteomics analysis. Our analysis shows that variations in PMA concentration and varying periods of rest post-stimulation result in downstream differences in proteome expression and cellular processes. We demonstrate that these differences result in altered expression of cytokines upon stimulation with different TLR ligands. Together, these findings provide a valuable resource that significantly expands the knowledge of protein expression dynamics in in vitro models of macrophages which in turn has profound impact on the immune responses being studied.
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Project description: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.
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Project description:SP110b is an interferon (IFN)-induced nuclear protein and may function as a transcriptional co-activator/repressor. IFNγ activates monocytes/macrophages thereby mediating inflammation. However, uncontrolled activation induces monocyte/macrophage cell death, which may cause immunopathology. We have demonstrated that SP110b expression prevented IFNγ-mediated monocyte/macrophage cell death. To explore the molecular mechanisms by which SP110b suppresses IFNγ-induced cell death, we performed a genome-wide microarray analysis to identify genetic determinants associated with IFNγ-induced cell death and regulated by SP110b. We sought to identify genetic determinants associated with IFNγ-induced cell death and regulated by SP110b. To that end, THP1 human monocyte-like cells that could be induced by doxycycline (Dox) to over-express SP110b (THP1-SP110b) were generated and 5 experimental groups of THP1-SP110b cells were harvested for RNA extraction and hybridization on Affymetrix microarrays. The 5 groups are as follows: untreated THP1-SP110b cells as control (CON), cells treated with IFNγ for 2 days (IFN_2D), cells treated with Dox plus IFNγ for 2 days (DoxIFN_2D), cells treated with IFNγ for 4 days (IFN_4D), and cells treated with Dox plus IFNγ for 4 days (DoxIFN_4D).
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