Related Experiment Video
Updated: May 18, 2026
Characterization of Human Monocyte Subsets by Whole Blood Flow Cytometry Analysis
Published on: October 17, 2018
Differential expression of TNFR1 (CD120a) and TNFR2 (CD120b) on subpopulations of human monocytes
Daniëlle Hijdra1, Adriane Dm Vorselaars, Jan C Grutters
1Department of Medical Microbiology and Immunology, St, Antonius Hospital, Nieuwegein, The Netherlands. g.rijkers@antoniusziekenhuis.nl.
Insights
Intermediate monocytes are more frequent in sarcoidosis patients and show higher TNFR1 expression. Nonclassical monocytes express more TNFR2, indicating distinct CD16+ monocyte functions in inflammation.
Area of Science:
- Immunology
- Cell Biology
Background:
- Human blood contains three monocyte subpopulations: classical (CD14++CD16-), intermediate (CD14++CD16+), and nonclassical (CD14+CD16++).
- CD16-expressing monocytes are elevated in sarcoidosis and other inflammatory conditions, but the specific subpopulation responsible is unclear.
- Tumor Necrosis Factor Receptor (TNFR) expression on these monocyte subsets has not been previously investigated.
Purpose of the Study:
- To determine the frequencies of monocyte subpopulations in sarcoidosis patients versus healthy controls.
- To analyze the expression levels of TNFR1 (CD120a) and TNFR2 (CD120b) on these monocyte subpopulations.
Main Methods:
- Peripheral blood cells from sarcoidosis patients and controls were stained for HLA-DR, CD14, CD16, TNFR1, and TNFR2.
- Flow cytometry (FACSCalibur) and FlowJo software were used for cell measurement and analysis.
- Statistical analysis included Student's t-test and One-way ANOVA.
Main Results:
- Sarcoidosis patients exhibited a significantly higher frequency of intermediate monocytes compared to healthy controls.
- Distinct TNFR1 and TNFR2 expression patterns were observed across monocyte subpopulations in both groups.
- Intermediate monocytes showed higher TNFR1 expression than classical and nonclassical monocytes, while nonclassical monocytes expressed more TNFR2 than intermediate monocytes.
Conclusions:
- Intermediate monocytes consistently display the highest TNFR1 expression, and nonclassical monocytes the highest TNFR2 expression, across both sarcoidosis patients and controls.
- The increased frequency of intermediate monocytes in sarcoidosis, coupled with differential TNFR expression, suggests functionally distinct roles for CD16-expressing monocyte subsets in the disease.
Background:
Three subpopulations of monocytes can be distinguished in human blood: classical (CD14++CD16-), intermediate (CD14++CD16+), and nonclassical (CD14+CD16++). CD16 expressing monocytes are expanded in patients with sarcoidosis and in various other inflammatory diseases. In sarcoidosis, it is unclear whether either intermediate, nonclassical or both CD16 expressing monocytes are responsible for this increase. Data relating to the monocyte subpopulations is receiving increasing attention, but the expression of TNF receptors on these subpopulations has not been studied thus far. The aim of this study was to determine frequencies of monocyte subpopulations and their expression of TNFR1 and TNFR2 in both sarcoidosis patients and healthy controls.
Methods:
Peripheral blood cells of sarcoidosis patients and healthy controls were stained for the markers HLA-DR, CD14, CD16, CD120a and CD120b. Cells were measured on a FACSCalibur and analyzed with FlowJo. We used Student's t-test and a parametric One-way ANOVA for statistical analysis.
Results:
Sarcoidosis patients had a significant higher frequency of intermediate monocytes than healthy controls. Significant differences in TNF receptor expression were found between the monocyte subpopulations, both in sarcoidosis patients as well as in healthy controls: intermediates expressed more TNFR1 than classicals and nonclassicals and nonclassicals expressed more TNFR2 than intermediates, whereas intermediates showed higher expression than classicals.
Conclusions:
In both sarcoidosis patients and healthy controls intermediate monocytes show the highest expression level of TNFR1 among monocyte subpopulations and nonclassical monocytes show the highest expression level of TNFR2. These findings, as wells as the higher frequency of intermediate monocytes in sarcoidosis patients, provide evidence for the existence of two functionally-distinct CD16 expressing monocyte subpopulations.
