Discharge of solubilized and Dectin-1-reactive β-glucan from macrophage cells phagocytizing insoluble β-glucan particles: Involvement of reactive oxygen species (ROS)-driven degradation

Abstract

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 β-glucan-specific probes, we succeeded in detection of soluble and Dectin-1-reactive β-glucan discharged from mouse RAW 264.7 and human THP-1 macrophage cell lines as well as mouse peritoneal macrophages, which had phagocytized insoluble β-glucan particles. The RAW 264.7 cell culture-supernatant containing the discharged β-glucan stimulated naïve RAW 264.7 cells, resulting in the induction of cytokine expression. Such discharge of Dectin-1-reactive β-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 Cu(2+)/ascorbic acid system solubilized insoluble β-glucan particles in vitro, and a part of the solubilized β-glucan was Dectin-1 reactive and biologically active in macrophage activation. The soluble and biologically active β-glucan was degraded further during prolonged exposure to ROS. These results suggest that degraded but Dectin-1-reactive β-glucan is discharged from macrophage cells phagocytizing insoluble β-glucan particles and stimulates not only themselves again but also the other naïve phagocytes, leading to the effective elimination of infecting microbes and the ultimate breakdown and inactivation of metabolically resistant β-glucan.

The effect of Biolex-MB40 on the phagocytic activity and oxidative metabolism of peripheral blood granulocytes and monocytes in lambs

“…Hino et al (2012) demonstrated that macrophages released soluble glucans into the medium after phagocytizing insoluble β-glucan particles. They found that those particles were fragmented by macrophage-produced ROS, and that the released soluble β-glucan was dectin-1-reactive and biologically active in terms of macrophage activation.…”

Measurement of Barley β-Glucan Concentration in the Plasma by Sandwich ELISA Using Rat Dectin-1

“…In fact, our previous study has shown the discharge of -glucan from -glucan-phagocytized macrophages. 10) The present study has shown that the plasma BG concentrations were traceable by sandwich ELISA, using recombinant rat dectin-1. In other words, the present results show that BG had affinity to rat dectin-1, suggesting that BG potentially activated such immune cells as macrophages in rats, as has been shown in mice.…”

“…DEC-V5His expression and purification were performed as previously described. 10) The binding property of rat dectin-1 was assessed by competitive ELISA. Multi-well plate (96 wells, Nunc, Roskilde, Denmark) was coated overnight at 4 C with 125 mg/L of yeast -glucan (YG; BB-Glucan, Oriental Yeast Co., Tokyo, Japan) dissolved in 1 mol/L of NaOH.…”

“…9,10) Recombinant dectin-1 has another merit of being applied to histological staining. We prepared in the present study two types of recombinant rat dectin-1 proteins and measured the concentration of BG in the plasma by sandwich ELISA using the recombinant proteins.…”

The effect of Leiber Beta-S (1,3-1,6-β-D-glucan) on the phagocytic activity and oxidative metabolism of peripheral blood granulocytes and monocytes in calves

“…Kulicke et al (1997) suggested that immunostimulating effects of β-glucans are determined by the degree of branching, length and tertiary structure of polymers, their origin, synthesis, manner of administration, dose and period of application. An increase in the concentrations of strongly oxidizing compounds (superoxide anion O 2 -, hydrogen peroxide H 2 O 2 , hydroxyl radical •OH, singlet oxygen 1 O 2 , chloramine NH 2 Cl) generated by respiratory burst was also observed by Hino et al (2012) in an in vitro study after administration of curdlan, and by Eicher et al (2010) in an in vivo experiment on calves administered the EnergyPlus supplement (mannan and β-glucan). Surprisingly, in an in vitro study by Kim et al (2004), the production of O 2 -and H 2 O 2 by macrophages did not increase after the administration of an aqueous extract from the fruiting body and mycelia of Phellinus linteus.…”