Fungal stimuli induce ct-CD45 in human monocytes. (A) Human monocytes were stimulated with Pam3CSK4 (P3C), peptidoglycan (PG), zymosan (Zym), poly(I:C) (PI:C), LPS, CpG, or PMA for 90 minutes. Lysates were analyzed by Western blot with mAb 8-301. (B) Binding of MEM-28 to resting monocytes (dashed line) or monocytes treated for 90 minutes with PMA, zymosan, or LPS (black lines) were examined by flow cytometry. Isotype-matched control mAb VIAP (dotted line) was used as nonbinding control. (C) Monocytes treated with zymosan were harvested at different time points and analyzed by Western blot with mAb 8-301. (D) Different numbers of C albicans hyphae and yeast were heat-inactivated and cocultured with monocytes. Lysates were analyzed by Western blot with mAb 8-301 for generation of ct-CD45. (E) Stimulation of monocytes with the Dectin-1 ligand β-glucan (β-Glu) leads to ct-CD45 production. Blocking with laminarin (Lam) also inhibited zymosan-induced ct-CD45. (F) The syk inhibitor piceatannol (picea; 50 μmol/L) strongly inhibited ct-CD45 production by zymosan. NADPH-oxidase inhibitor apocynin (apo; 1 mmol/L) blocked ct-CD45 induced by PMA and zymosan. Data represent 3 independently performed experiments.