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Updated: Jun 21, 2026
A Quantitative Assessment of the Phagocytosis of Allogeneic and Xenogeneic Erythrocytes by Rat Macrophages In Vitro
Published on: August 22, 2025
Flow cytometric isolation and phenotypic characterization of two subsets of ED2(+) (CD163) hepatic macrophages in
Yao He1, Tomohito Sadahiro, Sang-Ik Noh
1Comprehensive Transplant Center, Department of Surgery, Cedars-Sinai Medical Center, Los Angeles, California, USA.
Aims:
Macrophages in the liver are well known for their functional heterogeneity. However, subpopulations of the hepatic macrophages are not well defined.
Methods:
Two subsets of hepatic macrophages isolated from rats via FACS with immunolabeling of ED2 (anti-CD163) antibody were studied for phenotypic and functional characteristics.
Results:
A subset showed an ED2(high) and autofluorescence(high) (ED2(high)/AF(high)) phenotype, exhibiting characteristics consistent with the description of the Kupffer cells (KC). A second subset, displaying an ED2(dim)/AF(dim) phenotype, was smaller in size, monocyte-like and weak in phagocytosis. Transmission electron microscopy demonstrated that both subsets are phagocytes. Quantitative RT-PCR revealed that in addition to expression of macrophage-related surface markers such as CD14, ED1 (CD68), fucose receptor, and CD163, the ED2(dim)/ AF(dim) cells expressed mRNA encoding for myeloid lineage differentiation markers ERMP12 (PECAM) and ERMP20 (Ly-6C). These two subsets exhibited differential in gene expression of selected cytokines, extracellular matrix proteinases, and Toll-like receptor in normal livers, as well as significantly upregulated expression in cholestatic livers induced by bile duct ligation.
Conclusion:
The data suggest that the ED2(high)/AF(high) population of the liver cells represent the conventional Kupffer cells. The ED2(dim)/AF(dim) cells, however, are small hepatic resident macrophages characteristically different from the conventional Kupffer cells.
Insights
Researchers identified two distinct hepatic macrophage subsets in rats. One subset represents conventional Kupffer cells, while the other is a novel, smaller, monocyte-like hepatic resident macrophage population.
Area of Science:
- Immunology
- Hepatology
- Cell Biology
Background:
- Hepatic macrophages, primarily Kupffer cells (KC), exhibit functional heterogeneity.
- Subpopulations of hepatic macrophages remain poorly defined, hindering a complete understanding of liver immunity.
Purpose of the Study:
- To characterize distinct subpopulations of hepatic macrophages.
- To elucidate the phenotypic and functional differences between hepatic macrophage subsets.
Main Methods:
- Isolation of rat hepatic macrophages using fluorescence-activated cell sorting (FACS).
- Immunolabeling with ED2 (anti-CD163) antibody to distinguish subsets.
- Phenotypic analysis via flow cytometry and transmission electron microscopy.
- Gene expression profiling using quantitative RT-PCR.
Main Results:
- Two subsets were identified: ED2(high)/autofluorescence(high) (consistent with KC) and ED2(dim)/autofluorescence(dim) (smaller, monocyte-like, less phagocytic).
- Both subsets are phagocytic, with the ED2(dim)/AF(dim) cells expressing myeloid differentiation markers (PECAM, Ly-6C).
- Differential gene expression of cytokines, matrix proteinases, and Toll-like receptors was observed between subsets, with upregulation in cholestatic livers.
Conclusions:
- The ED2(high)/AF(high) population represents conventional Kupffer cells.
- The ED2(dim)/AF(dim) population constitutes a distinct subset of small, hepatic resident macrophages.
- These findings contribute to a refined understanding of liver macrophage heterogeneity and their roles in liver disease.
