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.

中文翻译：

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用于一氧化氮和氧化还原生物学的鼠骨髓源性巨噬细胞的分离和培养。

巨噬细胞是源自广泛分布于全身的造血祖细胞的单核吞噬细胞。这些细胞参与先天性和适应性免疫反应，并且是炎症、发育和体内平衡过程的核心。巨噬细胞的生理机能因它们所处的环境而异，它们对外部刺激表现出快速的功能适应。为了在体外研究巨噬细胞，通常从腹膜或肺泡离体培养细胞，或从骨髓祖细胞分化形成骨髓衍生巨噬细胞 (BMDM)。BMDMs 代表了研究巨噬细胞生物学的一种高效且具有成本效益的手段。然而，巨噬细胞对生化刺激（如细胞因子、代谢中间体、和 RNS/ROS）使得必须严格控制实验条件。因此，本研究的目的是建立一种优化和标准化的 BMDMs 分离和培养方法。我们使用从 WT 和一氧化氮 (NO) 缺陷小鼠中分离的经典活化巨噬细胞来开发标准化的培养方法，从而定义培养基的成分。然后，我们有条不紊地将我们的标准化方案与最常用的 BMDM 培养方法进行了比较，以建立用于研究一氧化氮 (NO)-氧化还原生物学和体外免疫代谢的最佳方案。我们使用从 WT 和一氧化氮 (NO) 缺陷小鼠中分离的经典活化巨噬细胞来开发标准化的培养方法，从而定义培养基的成分。然后，我们有条不紊地将我们的标准化方案与最常用的 BMDM 培养方法进行了比较，以建立用于研究一氧化氮 (NO)-氧化还原生物学和体外免疫代谢的最佳方案。我们使用从 WT 和一氧化氮 (NO) 缺陷小鼠中分离的经典活化巨噬细胞来开发标准化的培养方法，从而定义培养基的成分。然后，我们有条不紊地将我们的标准化方案与最常用的 BMDM 培养方法进行了比较，以建立用于研究一氧化氮 (NO)-氧化还原生物学和体外免疫代谢的最佳方案。