Decorin inhibits the M-CSF–dependent proliferation of BMDMs.
(A) BMDMs were obtained after 7 days of culture in the presence of M-CSF. Then, 105 macrophages were incubated in the presence of the indicated amounts of M-CSF in 24-well plates precoated with BSA (10 μg/mL; control) or with decorin (10 μg/mL). Proliferation was determined as described in “Materials and methods.” (B) Decorin inhibits macrophage proliferation in a dose-dependent manner. The 105 macrophages were incubated in 24-well plates precoated with the indicated amounts of decorin in the presence of 1000 U/mL M-CSF. For both panels A and B, each determination was made in triplicate and the values represented correspond to the mean ± SD of one representative of 3 independent experiments. (C) Decorin also blocks proliferation of macrophages induced by recombinant M-CSF (░, 2 ng/mL), GM-CSF (▪, 10 ng/mL), or control (■). (D) Decorin blocks the cell cycle at G1 phase. The 106 macrophages were cultured in the presence of 1000 U/mL M-CSF in 35-mm Petri dishes precoated with BSA or with 100 μg/mL decorin for 24 hours. DNA content was measured by DAPI staining and flow cytometry. Cell cycle distribution was analyzed using the Multicycle program (Phoenix Flow Systems). (E) Counting of viable cells cultured in 100 μg/mL decorin-precoated plates for 24 to 48 hours. The cells were counted by trypan blue exclusion using a hemocytometer. Each point was performed in triplicate, and the results were represented as the mean ± SD.
