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WT vs. PPARd-/- Macrophages

ABSTRACT: Macrophages rapidly engulf apoptotic cells to limit the release of noxious cellular contents and to restrict autoimmune responses against self antigens. Although factors participating in recognition and engulfment of apoptotic cells have been identified, the transcriptional basis for the sensing and silently disposing of apoptotic cells is unknown. Here we show that peroxisome proliferator activated receptor delta (PPARdelta) is induced when macrophages engulf apoptotic cells and functions as a transcriptional sensor of dying cells. Genetic deletion of PPARdelta decreases expression of opsonins, such as C1qb, resulting in impairment of apoptotic cell clearance and reduction in anti-inflammatory cytokine production. This increases autoantibody production and predisposes global and macrophage-specific PPARdelta deficient mice to autoimmune kidney disease, a phenotype resembling the human disease systemic lupus erythematosus. Thus, PPARdelta plays a pivotal role in orchestrating the timely disposal of apoptotic cells by macrophages, ensuring that tolerance to self is maintained. HoxA9 immortalized ES cell derived myeloid precursors were created and differentiated as in Odegaard et al, 2007. After expansion, macrophages were differentiated for 8 days in the presence of MCSF in 20% L929 conditioned media with 2.5% bovine serum albumin in low glucose media. Macrophages were harvested for RNA using Qiagen's RNeasy Kit. A common reference was generated using e17.5 C57Bl/6 embryos. 30ug of either reference or macrophage RNA was labeled with Cy3 and Cy5 (Amersham) and hybridized to Stanford Functional Genomics Facility mouse cDNA microarrays for 16 hours. The arrays were washed and scanned on an Agilent G2505A ChipScan v. A.6.3.1. Florescence values were extracted using GenePix Pro v. 5.0.0.51. The data were normalized using regression correlation. Subsequently, missing values were imputed and significantly differentially expressed genes were found with Significance Analysis of Microarrays. For the Nature Medicine manuscript, significantly differentially expressed genes including genes involved in the process of uptake of apoptotic cells were extracted and clustered by complete linkage using Cluster and visualized using TreeView. Key to genotypes: "J1 untreated" are WT differentiated macrophages; "18 Untreated" are PPARd-/- differentiated macrophages Genotype: Wild type (WT) or PPAR delta minus (PPARd -/-) macrophages

ORGANISM(S): Mus musculus

PROVIDER: GSE17890 | GEO | 2009/10/18

SECONDARY ACCESSION(S): PRJNA120075

REPOSITORIES: GEO

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Project description:Macrophages rapidly engulf apoptotic cells to limit the release of noxious cellular contents and to restrict autoimmune responses against self antigens. Although factors participating in recognition and engulfment of apoptotic cells have been identified, the transcriptional basis for the sensing and silently disposing of apoptotic cells is unknown. Here we show that peroxisome proliferator activated receptor delta (PPARdelta) is induced when macrophages engulf apoptotic cells and functions as a transcriptional sensor of dying cells. Genetic deletion of PPARdelta decreases expression of opsonins, such as C1qb, resulting in impairment of apoptotic cell clearance and reduction in anti-inflammatory cytokine production. This increases autoantibody production and predisposes global and macrophage-specific PPARdelta deficient mice to autoimmune kidney disease, a phenotype resembling the human disease systemic lupus erythematosus. Thus, PPARdelta plays a pivotal role in orchestrating the timely disposal of apoptotic cells by macrophages, ensuring that tolerance to self is maintained. HoxA9 immortalized ES cell derived myeloid precursors were created and differentiated as in Odegaard et al, 2007. After expansion, macrophages were differentiated for 8 days in the presence of MCSF in 20% L929 conditioned media with 2.5% bovine serum albumin in low glucose media. Macrophages were harvested for RNA using Qiagen's RNeasy Kit. A common reference was generated using e17.5 C57Bl/6 embryos. 30ug of either reference or macrophage RNA was labeled with Cy3 and Cy5 (Amersham) and hybridized to Stanford Functional Genomics Facility mouse cDNA microarrays for 16 hours. The arrays were washed and scanned on an Agilent G2505A ChipScan v. A.6.3.1. Florescence values were extracted using GenePix Pro v. 5.0.0.51. The data were normalized using regression correlation. Subsequently, missing values were imputed and significantly differentially expressed genes were found with Significance Analysis of Microarrays. For the Nature Medicine manuscript, significantly differentially expressed genes including genes involved in the process of uptake of apoptotic cells were extracted and clustered by complete linkage using Cluster and visualized using TreeView. Key to genotypes: "J1 untreated" are WT differentiated macrophages; "18 Untreated" are PPARd-/- differentiated macrophages Genotype: Wild type (WT) or PPAR delta minus (PPARd -/-) macrophages genetic_modification_design

Project description:This a model from the article: Modelling the onset of Type 1 diabetes: can impaired macrophage phagocytosis make the difference between health and disease? Maree AF, Kublik R, Finegood DT, Edelstein-Keshet L.Philos Transact A Math Phys Eng Sci.2006 May 15;364(1842):1267-82. 16608707, Abstract: A wave of apoptosis (programmed cell death) occurs normally in pancreatic beta-cells of newborn mice. We previously showed that macrophages from non-obese diabetic (NOD) mice become activated more slowly and engulf apoptotic cells at a lower rate than macrophages from control (Balb/c) mice. It has been hypothesized that this low clearance could result in secondary necrosis, escalating inflammation and self-antigen presentation that later triggers autoimmune, Type 1 diabetes (T1D). We here investigate whether this hypothesis could offer a reasonable and parsimonious explanation for onset of T1D in NOD mice. We quantify variants of the Copenhagen model (Freiesleben De Blasio et al. 1999 Diabetes 48, 1677), based on parameters from NOD and Balb/c experimental data. We show that the original Copenhagen model fails to explain observed phenomena within a reasonable range of parameter values, predicting an unrealistic all-or-none disease occurrence for both strains. However, if we take into account that, in general, activated macrophages produce harmful cytokines only when engulfing necrotic (but not apoptotic) cells, then the revised model becomes qualitatively and quantitatively reasonable. Further, we show that known differences between NOD and Balb/c mouse macrophage kinetics are large enough to account for the fact that an apoptotic wave can trigger escalating inflammatory response in NOD, but not Balb/c mice. In Balb/c mice, macrophages clear the apoptotic wave so efficiently, that chronic inflammation is prevented. This model originates from BioModels Database: A Database of Annotated Published Models (http://www.ebi.ac.uk/biomodels/). It is copyright (c) 2005-2011 The BioModels.net Team. For more information see the terms of use. To cite BioModels Database, please use: Li C, Donizelli M, Rodriguez N, Dharuri H, Endler L, Chelliah V, Li L, He E, Henry A, Stefan MI, Snoep JL, Hucka M, Le Novère N, Laibe C (2010) BioModels Database: An enhanced, curated and annotated resource for published quantitative kinetic models. BMC Syst Biol., 4:92.

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WT vs. PPARd-/- Macrophages

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