Fig. 4.
CLLV-1 suppresses fMLF-induced inflammatory responses in differentiated HL-60 (dHL-60) cells. (a) The HL-60 cells were exposed to 1.3% DMSO for 5 days. The differentiation of HL-60 cells by DMSO was examined by flow cytometry, using anti-FPR1 antibodies. (B–F) The dHL-60 cells were preincubated with DMSO or CLLV-1 (0.03–1 μM) and then activated with or without fMLF (0.1 μM)/CB (1 μg/mL). (b) Superoxide anion generation was detected by spectrophotometry at 550 nm, using cytochrome c reduction. (c) The intracellular ROS was monitored by flow cytometry, using cell-permeable DHR123. (d) Phosphorylation of p47phox was analyzed by immunoblotting, using antibodies against the phosphorylated (S304) and total p47phox. (e) F-actin levels were assayed by flow cytometry, using Alexa Fluor 594 Phalloidin. (f) Phosphorylation of AKT was analyzed by immunoblotting, using antibodies against the phosphorylated (S473 and T308) and total AKT. All data are expressed as mean values ± SEM (n = 3); *p < .05, **p < .01, and ***p < .001 compared with the DMSO + fMLF group (Student's t-test).