Gene expression variation analysis of M2c polarized human peripheral blood-derived macrophages stimulated with lysoPtdGlc by RNA-Seq.
Peripheral blood mononuclear cells were suspended in Dulbecco's Modified Eagle's Medium/Nutrient Mixture F-12 (Life technologies, Carlsbad, CA, USA), plated at a density of 1.5-2.0 Ã? 106 cells/ml, and incubated at 37Â°C for 3 hours. Adherent cells (monocytes) were cultured in RPMI 1640 supplemented with 10% fetal bovine serum (Biowest, Hiroshima, Japan) and 20 ng/ml M-CSF for 6 days to differentiate into M0 macrophages. 24 hours of culture in the presence of 100 nM Dexamethasone resulted in M2c macrophages were obtained when the medium was changed to 0.1% BSA and lysophosphatidylglucoside (lysoPtdGlc) 1.0 x 10-8M added 24 h after polarization of M2c macrophages. Cells were collected 2, 4, and 6 hours after addition, RNA was extracted, and submitted along with samples without lysoPtdGlc.