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In vitro-differentiated embryonic stem cell macrophages: a model system for studying atherosclerosis-associated
K J Moore1, R P Fabunmi, L P Andersson
1Lipid Metabolism Unit, Massachusetts General Hospital, Boston, MA 02114, USA.
Abstract:
Monocytes/macrophages (Mo) appear to play a critical role in the initiation and progression of atherosclerotic lesions. In this study, we characterized in vitro-differentiated embryonic stem (ES) cell macrophages as a model system for studying atherosclerosis-associated Mo functions. Using immunofluorescence staining and Western analysis, we demonstrate that ES Mo express typical macrophage cell surface markers, as well as the known receptors for modified forms of low density lipoprotein (LDL), including the Mo scavenger receptors (SR-A type I and type II), CD36, and CD68. Differentiated ES Mo specifically bind and degrade 125I-labeled acetylated LDL with high affinity, and their incubation with acetylated LDL (15 microg/mL) for 48 hours produces characteristic "foamy" Mo, as visualized by oil red O staining. ES Mo also express matrix-degrading metalloproteinases (MMP-3, MMP-9), which have been implicated in collagen breakdown in the fibrous cap of atherosclerotic plaques, and secrete cytokines (tumor necrosis factor-alpha, interleukin-6) in response to inflammatory stimuli. Transfection experiments, using a green fluorescent protein reporter gene, driven by the myeloid-specific promoter, CD11b, demonstrated that ES Mo can also be used to study macrophage-restricted gene expression in vitro. Taken together, these data demonstrate that ES Mo exhibit many properties typical of arterial lesion macrophages. Its ease of genetic manipulation makes it an attractive system for investigations of macrophage functions in vitro.
Insights
Embryonic stem cell-derived macrophages (ES Mo) effectively model arterial lesion macrophages, showing key markers and functions relevant to atherosclerosis research. This system offers a versatile platform for studying macrophage behavior in vitro.
Area of Science:
- Cell Biology
- Immunology
- Cardiovascular Research
Background:
- Monocytes/macrophages (Mo) are crucial in initiating and advancing atherosclerotic lesions.
- Understanding Mo functions in atherosclerosis is vital for developing targeted therapies.
Purpose of the Study:
- To characterize in vitro-differentiated embryonic stem (ES) cell macrophages as a model for studying atherosclerosis-associated Mo functions.
- To evaluate the suitability of ES Mo for investigating macrophage-restricted gene expression.
Main Methods:
- Immunofluorescence staining and Western analysis to identify macrophage markers and LDL receptors.
- Incubation with acetylated low-density lipoprotein (LDL) to induce foam cell formation.
- Oil red O staining to visualize lipid accumulation.
- Assessing expression of matrix-degrading metalloproteinases (MMPs) and cytokine secretion.
- Transfection studies using a green fluorescent protein reporter gene driven by the CD11b promoter.
Main Results:
- ES Mo express typical macrophage cell surface markers and receptors for modified LDL (SR-A, CD36, CD68).
- Differentiated ES Mo efficiently bind and degrade acetylated LDL, forming characteristic "foamy" macrophages.
- ES Mo express MMP-3 and MMP-9 and secrete inflammatory cytokines (TNF-α, IL-6) upon stimulation.
- ES Mo support macrophage-restricted gene expression studies via transfection.
Conclusions:
- ES Mo exhibit key properties of arterial lesion macrophages, making them a valuable in vitro model.
- The genetic manipulability of ES Mo provides an attractive system for in vitro investigations of macrophage functions in atherosclerosis.