CD36 cross-linking and PE phagocytosis do not induce TNF-α secretion.
(A) 5 × 105 monocytes were treated with F(ab′)2 fragments alone or cross-linking with 10-μg/mL CD36 (FA6-152, OKM5) or CD45 (monoclonal antibody 1214) followed by 10-μg/mL F(ab′)2 fragments, suspended in 500-μL RPMI–10% FCS–L-G, and exposed to the presence or absence of low-dose LPS (0.1 ng/mL). High-dose (100 ng/mL) LPS stimulation was used as a positive control. Cells were incubated for 4 hours at 37°C, 5% CO2; pelleted; and soluble TNF-α determined from the supernatants by enzyme-linked immunosorbent assay. CD36 cross-linking neither increased baseline TNF-α secretion nor primed for increased secretion in response to low-dose LPS, but CD45 cross-linking did prime for increased TNF-α following low-dose LPS. (B) 2.5 × 105 monocytes were allowed to adhere to plastic culture wells and incubated with medium alone, UEs, PEs (carefully synchronized to the trophozoite stage), or 1-μg/mL LPS for 4 hours at 37°C, 5% CO2. During this time, there was minimal PE rupture. At the end of 4 hours, the medium was collected, nonadherent cells pelleted, and soluble TNF-α determined. In panels A and B, n ≥ 3 per group and data are representative of results obtained in 3 separate experiments. Data = mean ± SEM. ***,P < .001 vs mock–cross-linked or unstimulated control cells (ANOVA with post hoc Tukey).
