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Characterization of murine macrophages from bone marrow, spleen and peritoneum.

ABSTRACT: Macrophages have heterogeneous phenotypes and complex functions within both innate and adaptive immune responses. To date, most experimental studies have been performed on macrophages derived from bone marrow, spleen and peritoneum. However, differences among macrophages from these particular sources remain unclear. In this study, the features of murine macrophages from bone marrow, spleen and peritoneum were compared.We found that peritoneal macrophages (PMs) appear to be more mature than bone marrow derived macrophages (BMs) and splenic macrophages (SPMs) based on their morphology and surface molecular characteristics. BMs showed the strongest capacity for both proliferation and phagocytosis among the three populations of macrophage. Under resting conditions, SPMs maintained high levels of pro-inflammatory cytokines expression (IL-6, IL-12 and TNF-?), whereas BMs produced high levels of suppressive cytokines (IL-10 and TGF-?). However, SPMs activated with LPS not only maintained higher levels of (IL-6, IL-12 and TNF-?) than BMs or PMs, but also maintained higher levels of IL-10 and TGF-?.Our results show that BMs, SPMs and PMs are distinct populations with different biological functions, providing clues to guide their further experimental or therapeutic use.

SUBMITTER: Wang C

PROVIDER: S-EPMC3574850 | biostudies-other | 2013 Feb

REPOSITORIES: biostudies-other

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Characterization of murine macrophages from bone marrow, spleen and peritoneum.

Wang Changqi C Yu Xiao X Cao Qi Q Wang Ya Y Zheng Guoping G Tan Thian Kui TK Zhao Hong H Zhao Ye Y Wang Yiping Y Harris David Ch DCh

BMC immunology 20130205

<h4>Background</h4>Macrophages have heterogeneous phenotypes and complex functions within both innate and adaptive immune responses. To date, most experimental studies have been performed on macrophages derived from bone marrow, spleen and peritoneum. However, differences among macrophages from these particular sources remain unclear. In this study, the features of murine macrophages from bone marrow, spleen and peritoneum were compared.<h4>Results</h4>We found that peritoneal macrophages (PMs) ...[more]

PMID: 23384230

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Project description:Osteoclasts are bone-resorbing cells differentiated from macrophage/monocyte precursors in response to macrophage colony-stimulating factor (M-CSF) and receptor activator of NF-κB ligand (RANKL). In vitro models are principally based on primary bone marrow macrophages (BMMs), but RAW 264.7 cells are frequently used because they are widely available, easy to culture, and more amenable to genetic manipulation than primary cells. Increasing evidence, however, has shown that the vastly different origins of these two cell types may have important effects on cell behavior. In particular, M-CSF is a prerequisite for the differentiation of BMMs, by promoting survival and proliferation and priming the cells for RANKL induction. RAW 264.7 cells readily form osteoclasts in the presence of RANKL, but M-CSF is not required. Based on these key differences, we sought to understand their functional implications and how it might affect osteoclast differentiation and related signaling pathways. Using a robust and high-throughput proteomics strategy, we quantified the global protein changes in osteoclasts derived from BMMs and RAW 264.7 cells at 1, 3, and 5 days of differentiation. Data are available via ProteomeXchange with the identifier PXD009610. Correlation analysis of the proteomes demonstrated low concordance between the two cell types (R2 ≈ 0.13). Bioinformatics analysis indicate that RANKL-dependent signaling was intact in RAW 264.7 cells, but biological processes known to be dependent on M-CSF were significantly different, including cell cycle control, cytoskeletal organization, and apoptosis. RAW 264.7 cells exhibited constitutive activation of Erk and Akt that was dependent on the activity of Abelson tyrosine kinase, and the timing of Erk and Akt activation was significantly different between BMMs and RAW 264.7 cells. Our findings provide the first evidence for major discrepancies between BMMs and RAW 264.7 cells, indicating that careful consideration is needed when using the RAW 264.7 cell line for studying M-CSF-dependent signaling and functions. © 2018 American Society for Bone and Mineral Research. © 2018 The Authors. JBMR Plus published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research.

Project description:Published data show that murine bone marrow-derived macrophages (BMDM) restrict growth of avirulent phase II, but not virulent phase I, Coxiella burnetii. Growth restriction of phase II bacteria is thought to result from potentiated recognition of pathogen-associated molecular patterns, which leads to production of inhibitory effector molecules. Past studies have used conditioned medium from L-929 murine fibroblasts as a source of macrophage-colony stimulating factor (M-CSF) to promote differentiation of bone marrow-derived myeloid precursors into macrophages. However, uncharacterized components of conditioned medium, such as variable amounts of type I interferons, can affect macrophage activation status and their permissiveness for infection. In the current study, we show that the C. burnetii Nine Mile phase II (NMII) strain grows robustly in primary macrophages from C57BL/6J mice when bone marrow cells are differentiated with recombinant murine M-CSF (rmM-CSF). Bacteria were readily internalized by BMDM, and replicated within degradative, LAMP1-positive vacuoles to achieve roughly 3 logs of growth over 6 days. Uninfected BMDM did not appreciably express CD38 or Egr2, markers of classically (M1) and alternatively (M2) activated macrophages, respectively, nor did infection change the lack of polarization. In accordance with an M0 phenotype, infected BMDM produced moderate amounts of TNF and nitric oxide. Similar NMII growth results were obtained using C57BL/6J myeloid progenitors immortalized with an estrogen-regulated Hoxb8 (ER-Hoxb8) oncogene. To demonstrate the utility of the ER-Hoxb8 system, myeloid progenitors from natural resistance-associated macrophage protein 1 (Nramp1) C57BL/6J knock-in mice were transduced with ER-Hoxb8, and macrophages were derived from immortalized progenitors using rmM-CSF and infected with NMII. No difference in growth was observed when compared to macrophages from wild type mice, indicating depletion of metal ions by the Nramp1 transporter does not negatively impact NMII growth. Results with NMII were recapitulated in primary macrophages where C57BL/6J Nramp1+ BMDM efficiently killed Salmonella enterica serovar Typhimurium. M-CSF differentiated murine macrophages from bone marrow and conditional ER-Hoxb8 myeloid progenitors will be useful ex vivo models for studying Coxiella-macrophage interactions.

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Characterization of murine macrophages from bone marrow, spleen and peritoneum.

Publications

Characterization of murine macrophages from bone marrow, spleen and peritoneum.

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