PMID- 32339668 OWN - NLM STAT- MEDLINE DCOM- 20201215 LR - 20231213 IS - 1089-8611 (Electronic) IS - 1089-8603 (Print) IS - 1089-8603 (Linking) VI - 100-101 DP - 2020 Aug 1 TI - Isolation and culture of murine bone marrow-derived macrophages for nitric oxide and redox biology. PG - 17-29 LID - S1089-8603(19)30334-9 [pii] LID - 10.1016/j.niox.2020.04.005 [doi] AB - Macrophages are mononuclear phagocytes derived from haematopoietic progenitors that are widely distributed throughout the body. These cells participate in both innate and adaptive immune responses and lie central to the processes of inflammation, development, and homeostasis. Macrophage physiology varies depending on the environment in which they reside and they exhibit rapid functional adaption in response to external stimuli. To study macrophages in vitro, cells are typically cultured ex vivo from the peritoneum or alveoli, or differentiated from myeloid bone marrow progenitor cells to form bone marrow-derived macrophages (BMDMs). BMDMs represent an efficient and cost-effective means of studying macrophage biology. However, the inherent sensitivity of macrophages to biochemical stimuli (such as cytokines, metabolic intermediates, and RNS/ROS) makes it imperative to control experimental conditions rigorously. Therefore, the aim of this study was to establish an optimised and standardised method for the isolation and culture of BMDMs. We used classically activated macrophages isolated from WT and nitric oxide (NO)-deficient mice to develop a standardised culture method, whereby the constituents of the culture media are defined. We then methodically compared our standardised protocol to the most commonly used method of BMDM culture to establish an optimal protocol for the study of nitric oxide (NO)-redox biology and immunometabolism in vitro. CI - Crown Copyright (c) 2020. Published by Elsevier Inc. All rights reserved. FAU - Bailey, Jade D AU - Bailey JD AD - BHF Centre of Research Excellence, Division of Cardiovascular Medicine, Radcliffe Department of Medicine, John Radcliffe Hospital, University of Oxford, Oxford, OX3 9DU, UK; Wellcome Centre for Human Genetics, University of Oxford, Oxford, OX3 7BN, UK. FAU - Shaw, Andrew AU - Shaw A AD - BHF Centre of Research Excellence, Division of Cardiovascular Medicine, Radcliffe Department of Medicine, John Radcliffe Hospital, University of Oxford, Oxford, OX3 9DU, UK; School of Pharmacy & Biomedical Sciences, Faculty of Clinical & Biomedical Sciences, University of Central Lancashire, Preston, PR1 2HE, UK; Wellcome Centre for Human Genetics, University of Oxford, Oxford, OX3 7BN, UK. FAU - McNeill, Eileen AU - McNeill E AD - BHF Centre of Research Excellence, Division of Cardiovascular Medicine, Radcliffe Department of Medicine, John Radcliffe Hospital, University of Oxford, Oxford, OX3 9DU, UK; Wellcome Centre for Human Genetics, University of Oxford, Oxford, OX3 7BN, UK. FAU - Nicol, Thomas AU - Nicol T AD - BHF Centre of Research Excellence, Division of Cardiovascular Medicine, Radcliffe Department of Medicine, John Radcliffe Hospital, University of Oxford, Oxford, OX3 9DU, UK; Wellcome Centre for Human Genetics, University of Oxford, Oxford, OX3 7BN, UK. FAU - Diotallevi, Marina AU - Diotallevi M AD - BHF Centre of Research Excellence, Division of Cardiovascular Medicine, Radcliffe Department of Medicine, John Radcliffe Hospital, University of Oxford, Oxford, OX3 9DU, UK; Wellcome Centre for Human Genetics, University of Oxford, Oxford, OX3 7BN, UK. FAU - Chuaiphichai, Surawee AU - Chuaiphichai S AD - BHF Centre of Research Excellence, Division of Cardiovascular Medicine, Radcliffe Department of Medicine, John Radcliffe Hospital, University of Oxford, Oxford, OX3 9DU, UK; Wellcome Centre for Human Genetics, University of Oxford, Oxford, OX3 7BN, UK. FAU - Patel, Jyoti AU - Patel J AD - BHF Centre of Research Excellence, Division of Cardiovascular Medicine, Radcliffe Department of Medicine, John Radcliffe Hospital, University of Oxford, Oxford, OX3 9DU, UK; Wellcome Centre for Human Genetics, University of Oxford, Oxford, OX3 7BN, UK. FAU - Hale, Ashley AU - Hale A AD - BHF Centre of Research Excellence, Division of Cardiovascular Medicine, Radcliffe Department of Medicine, John Radcliffe Hospital, University of Oxford, Oxford, OX3 9DU, UK; Wellcome Centre for Human Genetics, University of Oxford, Oxford, OX3 7BN, UK. FAU - Channon, Keith M AU - Channon KM AD - BHF Centre of Research Excellence, Division of Cardiovascular Medicine, Radcliffe Department of Medicine, John Radcliffe Hospital, University of Oxford, Oxford, OX3 9DU, UK; Wellcome Centre for Human Genetics, University of Oxford, Oxford, OX3 7BN, UK. FAU - Crabtree, Mark J AU - Crabtree MJ AD - BHF Centre of Research Excellence, Division of Cardiovascular Medicine, Radcliffe Department of Medicine, John Radcliffe Hospital, University of Oxford, Oxford, OX3 9DU, UK; Wellcome Centre for Human Genetics, University of Oxford, Oxford, OX3 7BN, UK. Electronic address: mark.crabtree@well.ox.ac.uk. LA - eng GR - RG/17/10/32859/BHF_/British Heart Foundation/United Kingdom GR - RG/12/5/29576/BHF_/British Heart Foundation/United Kingdom GR - DH_/Department of Health/United Kingdom GR - RG/12/5/29576/BHF_/British Heart Foundation/United Kingdom GR - 090532/Z/09/Z/WT_/Wellcome Trust/United Kingdom GR - FS/14/56/31049/BHF_/British Heart Foundation/United Kingdom GR - RG/07/003/23133/BHF_/British Heart Foundation/United Kingdom GR - RE/18/3/34214/BHF_/British Heart Foundation/United Kingdom GR - RE/13/1/30181/BHF_/British Heart Foundation/United Kingdom PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20200424 PL - United States TA - Nitric Oxide JT - Nitric oxide : biology and chemistry JID - 9709307 RN - 0 (Biopterins) RN - 31C4KY9ESH (Nitric Oxide) RN - 83869-56-1 (Granulocyte-Macrophage Colony-Stimulating Factor) SB - IM MH - Animals MH - Biopterins/metabolism MH - Cell Culture Techniques/methods MH - Cell Differentiation/drug effects MH - Female MH - Granulocyte-Macrophage Colony-Stimulating Factor/pharmacology MH - Macrophages/*cytology/drug effects/*metabolism MH - Male MH - Mice, Inbred C57BL MH - Mice, Transgenic MH - Nitric Oxide/*metabolism PMC - PMC7284309 OTO - NOTNLM OT - MCSF OT - Macrophage OT - Nitric oxide OT - Tetrahydrobiopterin EDAT- 2020/04/28 06:00 MHDA- 2020/12/16 06:00 PMCR- 2020/08/01 CRDT- 2020/04/28 06:00 PHST- 2019/11/19 00:00 [received] PHST- 2020/03/10 00:00 [revised] PHST- 2020/04/20 00:00 [accepted] PHST- 2020/04/28 06:00 [pubmed] PHST- 2020/12/16 06:00 [medline] PHST- 2020/04/28 06:00 [entrez] PHST- 2020/08/01 00:00 [pmc-release] AID - S1089-8603(19)30334-9 [pii] AID - 10.1016/j.niox.2020.04.005 [doi] PST - ppublish SO - Nitric Oxide. 2020 Aug 1;100-101:17-29. doi: 10.1016/j.niox.2020.04.005. Epub 2020 Apr 24.