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Updated: Mar 24, 2026
Isolation, Characterization, and Purification of Macrophages from Tissues Affected by Obesity-related Inflammation
Published on: April 3, 2017
A novel CD14(high) CD16(high) subset of peritoneal macrophages from cirrhotic patients is associated to an increased
Antonio José Ruiz-Alcaraz1, Ana Tapia-Abellán1, María Dolores Fernández-Fernández1
1Departamento de Bioquímica, Biología Molecular (B) e Inmunología, Facultad de Medicina, IMIB and Regional Campus of International Excellence "Campus Mare Nostrum", Universidad de Murcia, 30100 Murcia, Spain.
Abstract:
The aim of this study was to characterize monocyte-derived macrophages (M-DM) from blood and ascites of cirrhotic patients comparatively with those obtained from blood of healthy controls. The phenotypic profile based on CD14/CD16 expression was analyzed by flow cytometry. Cells were isolated and stimulated in vitro with LPS and heat killed Candida albicans. Phosphorylation of ERK, c-Jun, p38 MAPK, and PKB/Akt was analyzed by Western blotting. A novel CD14(high)CD16(high) M-DM subpopulation is present in ascites (∼33%). The CD14(++)CD16(+) intermediate subset is increased in the blood of cirrhotic patients (∼from 4% to 11%) and is predominant in ascites (49%), while the classical CD14(++)CD16(-) subpopulation is notably reduced in ascites (18%). Basal hyperactivation of ERK and JNK/c-Jun pathways observed in ascites M-DM correlates with CD14/CD16 high expressing subsets, while PI3K/PKB does it with the CD16 low expressing cells. In vitro LPS treatment highly increases ERK1/2, PKB/Akt and c-Jun phosphorylation, while that of p38 MAPK is decreased in M-DM from ascites compared to control blood M-DM. Stimulation of healthy blood M-DM with LPS and C. albicans induced higher phosphorylation levels of p38 than those from ascites. Regarding cytokines secretion, in vitro activated M-DM from ascites of cirrhotic patients produced significantly higher amounts of IL-6, IL-10 and TNF-α, and lower levels of IL-1β and IL-12 than control blood M-DM. In conclusion, a new subpopulation of CD14(high)CD16(high) peritoneal M-DM has been identified in ascites of cirrhotic patients, which is very sensitive to LPS stimulation.
Insights
Researchers identified a new CD14(high)CD16(high) monocyte-derived macrophage (M-DM) subpopulation in cirrhotic ascites. These M-DM show altered signaling and cytokine production, indicating unique immune responses in liver cirrhosis.
Area of Science:
- Immunology
- Cell Biology
- Hepatology
Background:
- Cirrhosis is associated with altered immune cell function.
- Monocyte-derived macrophages (M-DM) play a critical role in immune responses.
- Understanding M-DM phenotypes in cirrhosis is crucial for immune modulation.
Purpose of the Study:
- To characterize M-DM from cirrhotic patients' blood and ascites.
- To compare M-DM phenotypes and functions with healthy controls.
- To investigate signaling pathway activation and cytokine secretion.
Main Methods:
- Flow cytometry for CD14/CD16 phenotyping.
- In vitro stimulation with LPS and Candida albicans.
- Western blotting for MAPK and Akt pathway phosphorylation.
- Cytokine level analysis (IL-6, IL-10, TNF-α, IL-1β, IL-12).
Main Results:
- A novel CD14(high)CD16(high) M-DM subpopulation (∼33%) was identified in ascites.
- Increased CD14(++)CD16(+) intermediate M-DM in cirrhotic blood and ascites.
- Reduced classical CD14(++)CD16(-) M-DM in ascites.
- Ascites M-DM showed basal hyperactivation of ERK/JNK pathways.
- LPS stimulation altered phosphorylation patterns and increased IL-6, IL-10, TNF-α production in ascites M-DM.
Conclusions:
- A novel CD14(high)CD16(high) peritoneal M-DM subpopulation exists in cirrhotic ascites.
- These M-DM exhibit distinct signaling and cytokine profiles compared to healthy controls.
- This subpopulation is highly sensitive to LPS stimulation, suggesting a role in cirrhotic inflammation.
