Figure 6: TREM-1 promotes foam cell formation of human monocytes in vitro.

(a,b) TREM-1 and DAP12-expressing U937-TD cells were incubated for 48 h in vitro in the presence of 5% serum from HFCD-fed mice and the indicated stimuli (plate-bound anti-TREM-1, or isotype control antibody +/−30 ng ml⁻¹ LPS). (a) Representative photomicrograph of ORO and DAPI-stained U937-TD cells. Scale bars indicate 100 μm. (b) Quantification of foam cell formation. The ratio of DAPI positive pixels versus ORO positive pixels was calculated using Image J software. Bars represent mean values+s.d. from 10 independent experiments. (c,d) U937-TD cells were incubated for 48 h with the indicated stimuli in the presence or absence of 5% HFCD serum. CD36 surface expression was determined by flow cytometry. (c) Representative histogram overlays showing CD36 surface expression (filled histograms represent isotype control-stained cells) (d) MFI values for CD36 surface expression. Bars show mean values+s.d. from three independent experiments. (e–g) CD14^hi monocytes were flow-sorted from human blood donors. (e,f) Foam cell formation capacity of human CD14^hi monocytes was determined as described for U937-TD cells. (e) Scale bars, 100 μm. (f) Bars show mean values+s.d. from three independent experiments with different blood donors. (g) After 20 h of culture with the indicated stimuli, CD14^hi monocytes were harvested for qRT-PCR-based analysis of genes involved in cholesterol metabolism. Symbols show expression levels for n=7 independent experiments with different blood donors. *P<0.05, **P<0.01 as determined by the one-way ANOVA test (b–f) and the paired t-test (g). Statistically not significant differences with P>0.05 are not indicated.