Figure 2.

Neutrophil ROS production is stimulated by F. pedrosoi conidia and blocked by F. pedrosoi hyphae. (A) WT neutrophils were purified using Ficoll density layer and then seeded in a 96-well plate in the presence of a luminol reagent and stimulated with F. pedrosoi conidia or hyphae. The ROS production was measured every 2 min to approximately 60 min. As unstimulated control, neutrophils were incubated in the absence of fungi to measure the ROS production during the steady state. The area under the curve was calculated to measure the total ROS production after 60 min. (B) To confirm that hyphae block ROS production, we stimulated the cells with PMA (highly stimulated ROS production) in the presence or absence of hyphae. (C) Using heated-killed (HK) hyphae, we demonstrate that live hyphae blocks while HK hyphae stimulates ROS production. Data are expressed as mean ± SEM; n = 3, one-way ANOVA with Bonferroni’s posttest. **p < 0.01 ***p < 0.001.