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Culture of Macrophage Colony-stimulating Factor Differentiated Human Monocyte-derived Macrophages
Published on: June 30, 2016
Inhibitory effects of blocking voltage-dependent potassium channel 1.3 on human monocyte-derived macrophage
Xin-jun Lei1, Ai-qun Ma, Yu-tao Xi
1Department of Cardiology, The First Affiliated Hospital of Medical School, Xi'an Jiaotong University, Xi'an 710061, China.
Objective:
To investigate the expression of voltage-dependent potassium channel 1.3 (Kv1.3) mRNA and protein during human monocyte-derived macrophage differentiation into foam cells and its function in foam cell formation.
Methods:
Human peripheral blood monocytes were isolated from healthy male volunteers by density gradient centrifugation and then by adherent method. The obtained monocytes were cultured for 5 days to differentiate into macrophages. Based on establishment of the human macrophage-derived foam cell model, the expression of Kv1.3 channel was investigated by immunocytochemical staining, reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. Furthermore, the effects of rMargatoxin, a Kv 1.3 channel-specific inhibitor, on cholesterol metabolism in macrophages incepting oxidized low density lipoprotein (OxLDL) were studied.
Results:
After the macrophages co-incubated with 30 mg/L OxLDL at 37 degrees C for 60 hours, the cellular volume obviously enlarged and many red lipid granules were deposited in cytoplasm. The total amount of cholesterol (TC), free cholesterol (FC) and cholesterol ester (CE) in cells markedly increased and the ratio of CE/TC rose from (14.4+/-6.8)% to (57.9+/-3.5)% (n=7, P<0.05). However, the expression of Kv1.3 channel had no significant change. rMargatoxin (0.1 nmol/L and 10 nmol/L) markedly reduced the contents of TC, FC and CE in macrophages and the ratios of CE/TC decreased to (42.8+/-11.6)% and (22.6+/-8.0)%, respectively (n=7, P<0.05). Meanwhile, the red lipid granules deposited in the cytoplasm of macrophages also decreased.
Conclusion:
These data clearly show that the expression of Kv1.3 channel does not change obviously during human monocyte-derived macrophage differentiation into foam cells and the blocking of it would prevent foam cell formation.
Insights
Voltage-dependent potassium channel 1.3 (Kv1.3) expression remains unchanged during human macrophage foam cell formation. Blocking Kv1.3 inhibits cholesterol accumulation and foam cell development.
Area of Science:
- Cell biology
- Immunology
- Cardiovascular research
Background:
- Macrophage-derived foam cells are key contributors to atherosclerosis.
- Voltage-dependent potassium channels play roles in cellular processes, including immune cell function.
Purpose of the Study:
- To investigate the role of voltage-dependent potassium channel 1.3 (Kv1.3) in human monocyte-derived macrophage differentiation into foam cells.
- To determine if Kv1.3 inhibition affects cholesterol metabolism and foam cell formation.
Main Methods:
- Human monocytes were differentiated into macrophages and then into foam cells using oxidized low-density lipoprotein (OxLDL).
- Kv1.3 expression was analyzed using immunocytochemistry, RT-PCR, and Western blot.
- The effect of the Kv1.3 inhibitor rMargatoxin on cholesterol content and foam cell formation was assessed.
Main Results:
- Macrophage foam cell formation led to increased intracellular cholesterol levels, with a significant rise in cholesterol ester ratio.
- Kv1.3 channel expression showed no significant change during foam cell differentiation.
- rMargatoxin treatment significantly reduced total cholesterol, free cholesterol, and cholesterol ester content, and decreased foam cell formation.
Conclusions:
- Kv1.3 channel expression is not significantly altered during human macrophage foam cell differentiation.
- Inhibition of Kv1.3 effectively prevents foam cell formation by modulating cholesterol metabolism.
