Phagocytes engulf pathogenic microbes, kill them and degrade their cellular macromolecules by hydrolytic enzymes in phagolysosomes. However, such enzymes are unable to degrade some microbial polysaccharides, and fate of such indigestible polysaccharides in phagocytes remains uncertain. Using the extracellular domain of Dectin-1 as beta-glucan-specific probes, we succeeded in detection of soluble and Dectin-1-reactive beta-glucan discharged from mouse RAW 264.7 and human THP-1 macrophage cell lines as well as mouse peritoneal macrophages, which had phagocytized insoluble beta-glucan particles. The RAW 264.7 cell culture-supernatant containing the discharged beta-glucan stimulated naive RAW 264.7 cells, resulting in the induction of cytokine expression. Such discharge of Dectin-1-reactive beta-glucan from macrophage cells was inhibited by either NADPH oxidase inhibitors (apocynin and diphenylene iodonium) or radical scavengers (N-acetyl cysteine and MCI-186). Moreover, reactive oxygen species (ROS) produced by a Cu2+/ascorbic acid system solubilized insoluble beta-glucan particles in vitro, and a part of the solubilized beta-glucan was Dectin-1 reactive and biologically active in macrophage activation. The soluble and biologically active beta-glucan was degraded further during prolonged exposure to ROS. These results suggest that degraded but Dectin-1-reactive beta-glucan is discharged from macrophage cells phagocytizing insoluble beta-glucan particles and stimulates not only themselves again but also the other naive phagocytes, leading to the effective elimination of infecting microbes and the ultimate breakdown and inactivation of metabolically resistant beta-glucan. (C) 2012 Elsevier Inc. All rights reserved.