Isolation of functional mature peritoneal macrophages from healthy humans

• Published in: Immunology and cell biology 2020-02, Vol.98 (2), p.114-126
• Main Authors: Ruiz‐Alcaraz, Antonio J, Martínez‐Banaclocha, Helios, Marín‐Sánchez, Pilar, Carmona‐Martínez, Violeta, Iniesta‐Albadalejo, Miguel Angel, Tristán‐Manzano, María, Tapia‐Abellán, Ana, García‐Peñarrubia, Pilar, Machado‐Linde, Francisco, Pelegrín, Pablo, Martínez‐Esparza, María
• Format: Article
• Language: English
• Subjects: Adult; Cell activation; Cell Culture Techniques - methods; Cytokines; Cytokines - metabolism; Female; Flow Cytometry; Homeostasis; human macrophages; Humans; Immunity, Innate; Inflammasomes; Inflammasomes - metabolism; Inflammation; innate immunity; Interleukin-1beta - metabolism; Interleukin-6 - metabolism; Intracellular signalling; Laparoscopy; Leukocytes - metabolism; Macrophages; Macrophages, Peritoneal - cytology; Macrophages, Peritoneal - metabolism; peritoneal leukocytes; Peritoneum; Phagocytes; Phagocytosis; Reactive Oxygen Species; Reproductive sterilization; Tumor Necrosis Factor-alpha - metabolism
• ISSN: 0818-9641; 1440-1711
• DOI: 10.1111/imcb.12305

Macrophages play an important role in the inflammatory response. Their various biological functions are induced by different membrane receptors, including Toll‐like receptors, which trigger several intracellular signaling cascades and activate the inflammasomes, which in turn elicit the release of inflammatory mediators such as cytokines. In this study, we present a novel method for the isolation of human mature peritoneal macrophages. This method can be easily implemented by gynecologists who routinely perform laparoscopy for sterilization by tubal ligation or surgically intervene in benign gynecological pathologies. Our method confirms that macrophages are the main peritoneal leukocyte subpopulation isolated from the human peritoneum in homeostasis. We showed that primary human peritoneal macrophages present phagocytic and oxidative activities, and respond to activation of the main proinflammatory pathways such as Toll‐like receptors and inflammasomes, resulting in the secretion of different proinflammatory cytokines. Therefore, this method provides a useful tool for characterizing primary human macrophages as control cells for studies of molecular inflammatory pathways in steady‐state conditions and for comparing them with those obtained from pathologies involving the peritoneal cavity. Furthermore, it will facilitate advances in the screening of anti‐inflammatory compounds in the human system. We have optimized a new method to isolate functional and viable mature peritoneal macrophages, the most abundant human peritoneal leukocyte subpopulation in healthy individuals. We also present strong data supporting that these macrophages are distinct from blood monocytes and, in fact, we provide several hints in our method to discriminate peripheral blood monocytes contaminating the peritoneal lavage sample. This methodology will facilitate the advances in the general knowledge of this cell type and in the screening of anti‐inflammatory compounds in the human system. This work also opens the field to study different diseases such as endometriosis, hepatic cirrhosis or obesity, where peritoneal macrophages could play an important role.