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Culture of Macrophage Colony-stimulating Factor Differentiated Human Monocyte-derived Macrophages
Published on: June 30, 2016
Expression of Kir2.1 channel during differentiation of human macrophages into foam cells
Xin-jun Lei1, Ai-qun Ma, Yu-tao Xi
1Department of Cardiology, Ion Channelopathy Laboratory, Key Laboratory of Environment and Genes Related to Diseases of Ministry of Education, Xi'an Jiaotong University, China.
Objective:
Detected in non-transformed bone marrow-derived macrophages (BMDM) and identified as one of the key channels in modulating macrophage proliferation, activation and apoptosis, Kir2.1 channel is also characterized to play a crucial role in cell differentiation. The purpose of this study was to investigate the expression of Kir2.1 channel mRNA and protein during human monocyte-derived macrophage differentiation into foam cells.
Methods:
Human peripheral blood monocytes were isolated from healthy male volunteers by density gradient centrifugation and then by adherence method. The macrophages identified as a homogeneous population of adherent cells were obtained after 5 days of culture. Expression of Kir2.1 channel during human macrophage differentiation into foam cells was investigated by RT-PCR, Western blotting and immunocytochemistry, respectively.
Results:
After incubation of the macrophages with 30 mg/L OxLDL at 37 degrees C for 60 h, the cells were obviously enlarged in size and numerous red lipid granules observed under optical microscope. The cellular contents of the total cholesterol (TC), free cholesterol (FC) and cholesterol ester (CE) were markedly increased from 54.79+/-28.304 mg/g, 47.968+/-26.787 mg/g and 6.822+/-3.437 mg/g to 229.775+/-57.453 mg/g, 96.241+/-24.003 mg/g and 133.535+/-36.292 mg/g, respectively; the CE/TC ratio rose from (14.437+/-6.781)% to (57.946+/-3.507)% (n=7, P<0.05), suggesting the phenotype of foam cells. However, there was no significant difference in the relative expression of Kir2.1 channel mRNA between the macrophages and foam cells [(59.074+/-10.566)% vs (46.98+/-12.527)%, n=5, P>0.05], nor was there significant difference in the relative expression of Kir2.1 channel protein between them [(60.527+/-18.621)% vs (50.243+/-11.583)%, n=6, P>0.05].
Conclusion:
Incubation of human monocyte-derived macrophages with 30 mg/L OxLDL for 60 h induces the differentiation of the cells into foam cells, but the expression of Kir2.1 channel does not change obviously.
