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Culture of Macrophage Colony-stimulating Factor Differentiated Human Monocyte-derived Macrophages
Published on: June 30, 2016
[Inhibitory effect of blocking MaxiK channel on human monocyte-derived macrophages differentiation into foam cells]
Xin-jun Lei1, Ai-qun Ma, Yu-tao Xi
1Department of Cardiology, Key Laboratory of Environment and Genes Related to Diseases, Ministry of Education, Cardiovascular Ion Channelopathy Laboratory, the First Affiliated Hospital, Xi'an Jiaotong University, Xi'an 710061, China.
Aim:
To investigate the expression of high-conductance Ca2+-activated potassium channel (MaxiK channel) mRNA and protein during human monocyte-derived macrophage differentiation into foam cells and to study 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 human macrophage-derived foam cells model, the expression of MaxiK channel alpha-subunit was investigated by immunocytochemical staining, RT-PCR and Western blot. Furthermore, the effect of Paxilline, a MaxiK channel-specific inhibitor, on cholesterol metabolism in macrophages incepting oxidized low density lipoprotein (OxLDL) was studied.
Results:
After the macrophages were 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 (14.437+/-6.781)% to (57.946+/-3.507)% (n=7, P<0.05). However, the expression of MaxiK channel alpha-subunit had no significant change (P<0.05). Paxilline (5 micromol/L and 10 micromol/L) markedly reduced the content of TC, FC and CE in macrophages and the ratio of CE/TC decreased to (41.217+/-5.584)% (5 micromol/L Paxilline) and (18.017+/-11.559)% (10 micromol/L Paxilline), respectively (n=7, P<0.05). Meanwhile, the red lipid granules deposited in the cytoplasm of macrophages also decreased.
Conclusion:
Blocking MaxiK channel can inhibit human monocyte-derived macrophage to be differentiated into foam cells.
Insights
Blocking the MaxiK channel inhibits macrophage foam cell formation. This finding suggests MaxiK channel inhibition as a potential therapeutic strategy for atherosclerosis.
Area of Science:
- Cardiovascular Biology
- Ion Channel Physiology
- Atherosclerosis Research
Background:
- Macrophage foam cell formation is a key process in atherosclerosis.
- High-conductance Ca2+-activated potassium (MaxiK) channels play roles in cellular processes.
- The role of MaxiK channels in macrophage foam cell formation is not well understood.
Purpose of the Study:
- To investigate MaxiK channel expression during human monocyte-derived macrophage differentiation into foam cells.
- To determine the functional role of MaxiK channels in foam cell formation.
Main Methods:
- Human monocytes were differentiated into macrophages and then into foam cells using oxidized low-density lipoprotein (OxLDL).
- MaxiK channel alpha-subunit expression was assessed using immunocytochemistry, RT-PCR, and Western blot.
- The effect of Paxilline, a MaxiK channel inhibitor, on cholesterol metabolism in foam cells was evaluated.
Main Results:
- Macrophage foam cell formation led to increased intracellular cholesterol content and esterification.
- MaxiK channel alpha-subunit expression showed no significant change during foam cell differentiation.
- Paxilline treatment significantly reduced cholesterol accumulation and esterification in foam cells, decreasing the cholesterol ester to total cholesterol ratio.
Conclusions:
- MaxiK channel activity is involved in the process of human monocyte-derived macrophage foam cell formation.
- Inhibiting the MaxiK channel effectively reduces cholesterol accumulation in macrophages.
- MaxiK channel blockade presents a potential therapeutic target for mitigating foam cell formation in atherosclerosis.
