Experimental study of mouse alveolar macrophages cultured in vitro under different conditions

Aim This work set out to compare the characteristics of alveolar macrophages (AMs) or AM-like cells in vitro cultured from different progenitors in mice of different ages in the presence of GM-CSF, TGF-β, and PPAR-γ agonist rosiglitazone (in brief, GTR), according to a previously reported optimized culture procedure.Methods After bronchoalveolar lavage fluid (BALF) and bone marrow were isolated from mice of different ages and fetal livers were isolated from embryonic-day-15-17 mice, the mononuclear cells were seeded into 12-well plates and cultured in the presence of GTR. The generated cells were subjected to flow cytometry analysis to analyze the expression of AM markers.Results After culture, almost all BALF-derived cells were round-shaped under the light microscope, about half bone marrow-derived cells appeared round-shaped, and about 85% fetal liver-derived cells showed round-shaped morphology. The frequency of BALF-AMs from 4-week-old mice was nearly 90%. The frequency and number of BALF-AMs increased with the age of mice. The number of BALF-AMs from 12-week-old mice was nearly 1 × 106. The frequency of bone marrow-derived AM-like cells from 4-week-old mice was lower than 20% and increased to about 40-45% for 8-week-old and 12-week-old mice. The number of bone marrow-derived AM-like cells increased with the age of mice and about 0.8 × 106 bone marrow-derived AM-like cells/well were produced from 12-week-old mice. The frequency of fetal liver-derived AM-like cells was around 84% and their number could reach 4 × 106 /well.Conclusions The purity and number of AMs or AM-like cells in vitro cultured from different progenitors in mice of different ages vary dramatically. The detailed condition should be chosen according to specific study requirements.