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Updated: Aug 7, 2026
Isolation of Murine Peritoneal Macrophages to Carry Out Gene Expression Analysis Upon Toll-like Receptors Stimulation
Published on: April 29, 2015
Stimulation and release of interleukin-1 from peritoneal macrophages of the mouse
1Department of Pharmacology, University College London, UK.
Abstract:
Lipopolysaccharide (LPS) caused a concentration-dependent increase of released and cell-associated interleukin-1 (IL-1) in resident peritoneal macrophages from the mouse. LPS was about 30 times more potent at stimulating the level of cell-associated IL-1 than it was at stimulating the release of IL-1. Human recombinant tumour necrosis factor-alpha (TNF-alpha) and the calcium ionophores A23187 and ionomycin induced a concentration-dependent increase of cell-associated IL-1 but failed to cause release of IL-1 at concentrations producing maximal stimulation of cell-associated IL-1. The phorbol ester, 4 beta-phorbol dibutyrate, stimulated the release of IL-1 from mouse macrophages but failed to induce an increase in cell-associated IL-1. Substance P, neurokinin A, neurokinin B, calcitonin gene-related peptide and platelet-activating factor did not increase the released or cell-associated IL-1 in mouse macrophages. These agents also failed to alter released or cell-associated IL-1 stimulated by LPS, 1 microgram ml-1. It appears that a calcium signal is sufficient for the transcription and translation of IL-1 mRNA but does not result in the secretion of biologically active forms of IL-1. Our data also indicate that different intracellular signals may control the release and the cell accumulation of IL-1. We conclude that inflammatory mediators may independently increase either the release of, or the cell accumulation of IL-1.
Insights
Lipopolysaccharide (LPS) significantly boosts cell-associated interleukin-1 (IL-1) more than released IL-1 in mouse macrophages. Different signals control IL-1 release versus accumulation, impacting inflammation.
Area of Science:
- Immunology
- Cell Biology
- Molecular Biology
Background:
- Interleukin-1 (IL-1) is a key inflammatory cytokine.
- Understanding the regulation of IL-1 production and release is crucial for inflammatory disease research.
Purpose of the Study:
- To investigate the differential regulation of cell-associated and released IL-1 by various inflammatory mediators in mouse macrophages.
- To elucidate the intracellular signaling pathways controlling IL-1 transcription, translation, and secretion.
Main Methods:
- Primary resident peritoneal macrophages were isolated from mice.
- Cells were stimulated with lipopolysaccharide (LPS), tumor necrosis factor-alpha (TNF-alpha), calcium ionophores (A23187, ionomycin), and phorbol ester (4 beta-phorbol dibutyrate).
- Released and cell-associated IL-1 levels were quantified.
Main Results:
- LPS increased both released and cell-associated IL-1 in a concentration-dependent manner, being significantly more potent for cell-associated IL-1.
- Calcium ionophores and TNF-alpha increased cell-associated IL-1 but not released IL-1.
- Phorbol ester stimulated IL-1 release without increasing cell-associated IL-1.
- Substance P and other neuropeptides did not affect IL-1 levels, even when co-administered with LPS.
Conclusions:
- A calcium signal is sufficient for IL-1 mRNA production but not for secretion of biologically active IL-1.
- Distinct intracellular signaling pathways regulate IL-1 release and cell accumulation.
- Inflammatory mediators can independently modulate IL-1 release or cell accumulation, offering insights into inflammatory processes.
