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Mouse Bone Marrow Cell Isolation and Macrophage Differentiation

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Non-Alcoholic Steatohepatitis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2455))

Abstract

The rapid increase in the incidence of obesity contributes to a parallel increase in nonalcoholic steatohepatitis (NASH). Monocyte-derived macrophages, recruited from the bone marrow to the liver, promote NASH-related inflammation and fibrosis. In addition, adipose tissue macrophages (ATMs) release pro-inflammatory cytokines (PICs) which stimulate adipose tissue lipolysis liberating free fatty acids (FFAs) that can accumulate in the liver as triglycerides (TGs), thereby inducing steatosis. As such, bone marrow-derived macrophages (BMDMs) function as an essential tool to study the pathogenesis of NASH. BMDMs are primary bone marrow-derived cells which are differentiated into macrophages in vitro in the presence of growth factors. Macrophage colony-stimulating factor (M-CSF) is required for the proliferation and differentiation of committed myeloid progenitors into cells of the macrophage/monocyte lineage. Here, we describe a protocol for the isolation of mouse bone marrow cells and subsequent macrophage differentiation in which bone marrow cells are cultured in the presence of M-CSF, supplemented either by conditioned medium from L929 cells or in purified form. The efficiency of the differentiation is confirmed by immunofluorescent staining of macrophage surface antigen F4/80. The BMDMs serve as an excellent ex vivo model for a variety of studies, including hepatocyte-macrophage and adipocyte-macrophage cross-talk regulating NASH.

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Acknowledgments

This work was supported in part by the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) under Grant 1R01DK107451-01A1, the National Cancer Institute (NCI) under Grants 1R01CA230561-01A1, 1R01CA240004-01, and 1R01CA244993-01, and the Department of Defense (DOD) under Grant CA170048.

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Authors and Affiliations

  1. Department of Human and Molecular Genetics, Virginia Commonwealth University, Richmond, VA, USA

    Rachel Mendoza, Indranil Banerjee, Debashri Manna, Rajesh Yetirajam & Devanand Sarkar

  2. C. Kenneth and Dianne Wright Center for Clinical and Translational Research, Virginia Commonwealth University, Richmond, VA, USA

    Saranya Chidambaranathan Reghupaty

  3. Massey Cancer Center, Virginia Commonwealth University, Richmond, VA, USA

    Devanand Sarkar

  4. VCU Institute of Molecular Medicine, Virginia Commonwealth University, Richmond, VA, USA

    Devanand Sarkar

Authors
  1. Rachel Mendoza
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  2. Indranil Banerjee
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  3. Debashri Manna
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  4. Saranya Chidambaranathan Reghupaty
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  5. Rajesh Yetirajam
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  6. Devanand Sarkar
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Corresponding author

Correspondence to Devanand Sarkar .

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Editors and Affiliations

  1. Department of Human and Molecular Genetics, Virginia Commonwealth University, Richmond, VA, USA

    Devanand Sarkar

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© 2022 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Mendoza, R., Banerjee, I., Manna, D., Reghupaty, S.C., Yetirajam, R., Sarkar, D. (2022). Mouse Bone Marrow Cell Isolation and Macrophage Differentiation. In: Sarkar, D. (eds) Non-Alcoholic Steatohepatitis. Methods in Molecular Biology, vol 2455. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2128-8_8

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  • DOI: https://doi.org/10.1007/978-1-0716-2128-8_8

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2127-1

  • Online ISBN: 978-1-0716-2128-8

  • eBook Packages: Springer Protocols

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