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Detection of Inflammasome Activation and Pyroptotic Cell Death in Murine Bone Marrow-derived Macrophages
Published on: May 21, 2018
Quantitative investigations of apoptosis of murine mononuclear phagocytes during mild hyperthermia
J M Papadimitriou1, I van Bruggen
1Department of Pathology, University of Western Australia, Nedlands.
Abstract:
DNA fragmentation in murine monocytes and freshly harvested resident peritoneal macrophages (RPM) was assessed. Similar proportions of fragmented DNA were present in both cell types indicating that apoptosis is a significant occurrence in these cells and may provide a mechanism for controlling the numbers of mononuclear phagocytes in the tissues that house them. Culturing both of these cell types at 41 degrees C for 5 hr revealed a significant increase in the proportion of fragmented DNA only in cultures containing RPM. These latter preparations when cytophotometrically assessed also displayed reduced Feulgen DNA and cytoplasmic naphthol yellow S staining, again confirming the presence of apoptosis. It is suggested that the induction of apoptosis by mild hyperthermia may therefore be due to the relative absence of a temperature-sensitive inhibitor of apoptosis in a significant proportion of resident peritoneal macrophages. Lastly, electron microscope assessment indicated that the apoptotic macrophages were engulfed by their nonaffected neighbors. Programmed cell death in macrophage populations followed by phagocytosis of the apoptotic cells may therefore be a mechanism for controlling macrophage numbers in mammalian organisms.
Insights
Apoptosis, or programmed cell death, naturally occurs in monocytes and resident peritoneal macrophages (RPM). Mild hyperthermia significantly increases apoptosis in RPM, suggesting a mechanism for macrophage population control.
Area of Science:
- Immunology
- Cell Biology
- Histology
Background:
- Apoptosis is a crucial process for regulating cell populations.
- Mononuclear phagocytes, including macrophages, play vital roles in tissue homeostasis and immunity.
- Understanding the regulation of macrophage apoptosis is key to controlling inflammatory responses and tissue repair.
Purpose of the Study:
- To investigate the occurrence and regulation of apoptosis in murine monocytes and resident peritoneal macrophages (RPM).
- To determine the effect of mild hyperthermia on apoptosis in these cell types.
- To elucidate the mechanisms underlying macrophage population control through programmed cell death.
Main Methods:
- Assessment of DNA fragmentation in monocytes and RPM.
- Culturing cells at 41°C for 5 hours to induce hyperthermia.
- Cytophotometric analysis of Feulgen DNA and cytoplasmic naphthol yellow S staining.
- Electron microscopy to observe cellular interactions.
Main Results:
- Similar basal levels of DNA fragmentation (apoptosis) were observed in both monocytes and RPM.
- Mild hyperthermia (41°C) significantly increased DNA fragmentation specifically in RPM cultures.
- Cytophotometry confirmed increased apoptosis in RPM under hyperthermia, with reduced DNA and protein staining.
- Electron microscopy revealed phagocytosis of apoptotic macrophages by non-apoptotic neighbors.
Conclusions:
- Apoptosis is a significant mechanism for regulating mononuclear phagocyte numbers in tissues.
- Resident peritoneal macrophages are particularly susceptible to hyperthermia-induced apoptosis, potentially due to a lack of temperature-sensitive inhibitors.
- Phagocytosis of apoptotic macrophages by their healthy counterparts represents a key mechanism for controlling macrophage populations in mammals.
