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Immortalization of Murine Bone Marrow-Derived Macrophages

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1784))

Abstract

Macrophages are specialized phagocytes that display a variety of important functions for the host immune system. They are particularly important for the recognition of exogenous and endogenous danger signals, forming the defensive front line as part of innate immune response. As such, murine macrophages are commonly used for in vitro cell-based assays examining the mechanisms of innate immune activation, which can require the ongoing breeding and housing of a large number of genetically modified mouse strains. Here, we describe a robust protocol for the generation of immortalized bone marrow-derived macrophages (iBMDMs) from primary murine bone marrow cells. We further provide general protocols for harvesting, freezing, and thawing of bone marrow cells, maintaining iBMDMs in culture and generation of monoclonal iBMDM populations by single-cell cloning.

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References

  1. Murray PJ, Wynn TA (2011) Protective and pathogenic functions of macrophage subsets. Nat Rev Immunol 11:723–737

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Ginhoux F, Schultze JL, Murray PJ et al (2016) New insights into the multidimensional concept of macrophage ontogeny, activation and function. Nat Immunol 17:34–40

    Article  CAS  PubMed  Google Scholar 

  3. Okabe Y, Medzhitov R (2016) Tissue biology perspective on macrophages. Nat Immunol 17:9–17

    Article  CAS  PubMed  Google Scholar 

  4. De Nardo D (2017) Activation of the innate immune receptors: guardians of the micro galaxy. Adv Exp Med Biol 1024:1–35

    Article  PubMed  Google Scholar 

  5. De Nardo D (2015) Toll-like receptors: activation, signalling and transcriptional modulation. Cytokine 74:181–189

    Article  CAS  PubMed  Google Scholar 

  6. Latz E, Xiao TS, Stutz A (2013) Activation and regulation of the inflammasomes. Nat Rev Immunol 13:397–411

    Article  CAS  PubMed  Google Scholar 

  7. Blasi E, Mathieson BJ, Varesio L et al (1985) Selective immortalization of murine macrophages from fresh bone marrow by a raf/myc recombinant murine retrovirus. Nature 318:667–670

    Article  CAS  PubMed  Google Scholar 

  8. Blasi E, Barluzzi R, Bocchini V et al (1990) Immortalization of murine microglial cells by a v-raf/v-myc carrying retrovirus. J Neuroimmunol 27:229–237

    Article  CAS  PubMed  Google Scholar 

  9. Cox GW, Mathieson BJ, Gandino L et al (1989) Heterogeneity of hematopoietic cells immortalized by v-myc/v-raf recombinant retrovirus infection of bone marrow or fetal liver. J Natl Cancer Inst 81:1492–1496

    Article  CAS  PubMed  Google Scholar 

  10. Roberson SM, Walker WS (1988) Immortalization of cloned mouse splenic macrophages with a retrovirus containing the v-raf/mil and v-myc oncogenes. Cell Immunol 116:341–351

    Article  CAS  PubMed  Google Scholar 

  11. Bauernfeind FG, Horvath G, Stutz A et al (2009) Cutting edge: NF-kappaB activating pattern recognition and cytokine receptors license NLRP3 inflammasome activation by regulating NLRP3 expression. J Immunol 183:787–791

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Franklin BS, Bossaller L, De Nardo D et al (2014) The adaptor ASC has extracellular and ‘prionoid’ activities that propagate inflammation. Nat Immunol 15:727–737

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Halle A, Hornung V, Petzold GC et al (2008) The NALP3 inflammasome is involved in the innate immune response to amyloid-beta. Nat Immunol 9:857–865

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Hornung V, Bauernfeind F, Halle A et al (2008) Silica crystals and aluminum salts activate the NALP3 inflammasome through phagosomal destabilization. Nat Immunol 9:847–856

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Labzin LI, Schmidt SV, Masters SL et al (2015) ATF3 is a key regulator of macrophage IFN responses. J Immunol 195:4446–4455

    Article  CAS  PubMed  Google Scholar 

  16. Sun J, Li N, Oh KS et al (2016) Comprehensive RNAi-based screening of human and mouse TLR pathways identifies species-specific preferences in signaling protein use. Sci Signal 9:ra3

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Cardona Gloria Y, Latz E, De Nardo D (2018) Generation of Innate Immune Reporter Cells Using Retroviral Transduction. Methods Mol Biol 1714:97–117. https://doi.org/10.1007/978-1-4939-7519-8_7

    Article  PubMed  Google Scholar 

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Correspondence to Dominic De Nardo .

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De Nardo, D., Kalvakolanu, D.V., Latz, E. (2018). Immortalization of Murine Bone Marrow-Derived Macrophages. In: Rousselet, G. (eds) Macrophages. Methods in Molecular Biology, vol 1784. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7837-3_4

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  • DOI: https://doi.org/10.1007/978-1-4939-7837-3_4

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7836-6

  • Online ISBN: 978-1-4939-7837-3

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