BM-derivedBCL-6^-/- macrophages hyperproliferate due to accelerated G₁-to-S phase transition. (A) Morphology of macrophage colonies from liquid culture stained with methylene blue. A Zeiss Stemi microscope (Carl Zeiss Microimaging, Thornwood, NY) was used, and was equipped with a 4 ×/0.65 objective lens. The image was acquired with QImaging Retiga (QImaging, Burnaby, BC, Canada). Scale bar is equal to 1 mm. (B) Growth curves of wild-type (⋄) and BCL-6^-/- macrophages (▪) in the high CSF-1 medium. (Error bars representing SD are too small to be visible in this case.) (C) Cell cycle analysis based on PI staining and flow cytometry. Percentages of G₁ cells from 8 independent primary macrophage preparations are plotted for each genotype. Bars in the graph represent group means, shown with SD at the top and the P from a 2-tailed Student t test below. (D) Cell cycle progression of CSF-1-starved cells at 0, 8, 12, 16, and 20 hours after CSF-1 readdition was monitored by PI staining followed by flow cytometry analysis. Red peaks indicate G₁ and G₂/M phases and dashed areas represent S phase fractions. Arrows point to the first appearance of S phase cells. Results in panels A, B, and D are representative of 2 independent experiments.