Fig. 4.

TLR2-dependent ROS production induced by metHb, LTA or metHb + LTA complex. Isolated neutrophils were pretreated with either TLR2 blocking Ab or isotype Ab for 1 h prior to the treatment with metHb (1 mg/ml), LTA (10 μg/ml) or metHb (1 mg/ml) + LTA (10 μg/ml) complex for 1 h. Mock was buffer only treatment. (a) Intracellular ROS determined using CM-H₂DCFDA dye and measured by flow cytometry. (b) IL-8 secretion determined by ELISA. (c) TNFα secretion determined by ELISA. Data are representative of at least 3 independent experiments with similar results. * and § are effects of metHb + LTA combination and TLR2 antibody, respectively. P < 0.05 by two-factor ANOVA.