B-cell activation up-regulates LDL-R expression and promotes apoE enhancement of B cell–mediated NKT-cell activation. (A) Sorted B cells were cultured for 18 to 24 hours in media alone, media plus anti-Ig, or in media with a monolayer of CD40L-expressing cells. Representative histograms (from a total of 3 experiments) showing CD86 (top histograms) and LDL-R (bottom histograms) expression on unstimulated (media), anti-Ig activated, or CD40L-activated B cells. (B) ApoE-bound lipid antigen uptake by activated B cells promotes NKT-cell activation. Activated (anti-Ig or CD40L) and unstimulated (media) B cells were cocultured overnight with NKT cells and αGalCer ± apoE (2.5 μg/mL) as indicated. IFN-γ secretion was measured by ELISA. Results (mean ± SEM) obtained from 5 independent experiments using B cells purified from individual tonsils are shown. (C) CD40L-activated B cells were cocultured overnight with NKT cells and αGalGalCer with or without apoE (2.5 μg/mL) as indicated. Results (mean ± SEM) were obtained from independent experiments using B cells purified from 4 individual tonsils. (D) CD40L-activated B cells were pulsed for 4 hours in media, αGalCer (200 ng/mL), αGalGalCer (200 ng/mL) with or without apoE (2.5 μg/mL). Cells were washed and then cocultured overnight with NKT cells. Average (mean ± SEM) of B cells from 4 individual tonsils is shown. Alternatively, NKT cells were cultured overnight alone in media or αGalCer (200 ng/mL) ± apoE as indicated (NKT). *Significant differences (P < .05) versus control treatment without apoE (paired t test).