Fucoidan suppresses excessive phagocytic capacity of porcine peripheral blood polymorphonuclear cells by modulating production of tumor necrosis factor-alpha by lipopolysaccharide-stimulated peripheral blood mononuclear cells

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Highlights

  • The clinical and pharmacological activities of fucoidan, a cell wall polysaccharide found in the brown seaweed, have been known to modulate the immune function. The objective of this study was to examine the effect of fucoidan on the phagocytic capacity of porcine peripheral blood polymorphonuclear cells (PMNs) in inflammation condition.

  • The phagocytic capacity of PMNs was enhanced by the culture supernatant from PBMCs treated with either fucoidan or LPS. In particular, the culture supernatant from PBMCs treated with LPS revealed excessive high phagocytosis of PMNs This excessive phagocytosis of PMNs was reduced by treatment of fucoidan.

  • In addition, the production of TNF-α was shown to increase upon the treatment of either fucoidan or LPS in PBMCs. However, the excessive production of TNF-α in PBMCs by LPS treatment was reduced by addition of fucoidan.

  • The level of TNF-α mRNA expression in PBMCs was also up-regulated by either the fucoidan or LPS treatment. However, the excessive expression of TNF-α mRNA in PBMCs by LPS treatment was decreased by fucoidan.

  • This ability of culture supernatant from PBMCs by LPS and/or fucoidan to increase the phagocytic capacity of PMNs was inhibited by addition of anti-rp TNF-α polyclonal antibody (pAb).

  • These results suggested that fucoidan can suppress excessive phagocytic capacity of PMNs through modulation of TNF-α production in inflammation condition.

Abstract

We examined the effect of fucoidan, an immune modulator, on the phagocytic capacity of porcine peripheral blood polymorphonuclear cells (PMNs) exposed to culture supernatant from lipopolysaccharide (LPS)-stimulated peripheral blood mononuclear cells (PBMCs). For this purpose, we evaluated the phagocytic capacity of porcine PMNs by flow cytometry and measured levels of tumor necrosis factor-alpha (TNF-α) protein and mRNA in porcine PBMCs by enzyme-linked immunosorbent assay (ELISA) and real time-polymerase chain reaction (PCR), respectively. Fucoidan or LPS alone did not affect the phagocytic capacity of PMNs, but phagocytosis by these cells was increased by exposure to culture supernatant from PBMCs treated with fucoidan or LPS. In particular, the culture supernatant from PBMCs treated with LPS revealed excessive phagocytosis of PMNs. This excessive phagocytic capacity was diminished by co-treatment LPS with fucoidan. Production of TNF-α mRNA and protein increased upon treatment of PBMCs with either fucoidan or LPS, but this effect was also diminished by co-treatment LPS with fucoidan. The ability of culture supernatant from PBMCs treated with LPS and/or fucoidan to increase the phagocytic capacity of PMNs was inhibited by anti-recombinant porcine TNF-α polyclonal antibody. These results suggested that fucoidan suppresses the phagocytic capacity of PMNs by modulating TNF-α production by LPS-stimulated PBMCs.

Introduction

Phagocytes, including polymorphonuclear cells (PMNs), monocytes, and macrophages, defend the body by phagocytizing foreign particles, harmful pathogens, and dead cells (Lee et al. 2003). The phagocytic capacities of these phagocytes are transduced by intracellular signal cascades that triggers several cellular processes, including activation of phagocytosis mechanisms and production of inflammation-related cytokines (Aderem and Underhill 1999). When mononuclear cells such as monocytes, tissue macrophages, and lymphocytes are activated, they produce pro-inflammatory cytokines, which induce activation of PMNs and upregulation of PMNs functions, including generation of reactive oxygen species (ROS) and phagocytosis (Pechkovsky et al. 1996).

Tumor necrosis factor-alpha (TNF-α) is a key regulator of the inflammatory response. It is known that high levels of TNF-α are correlated with pathological responses in a variety of inflammatory diseases and inflammatory conditions (Grimble 1998). Excess production of TNF-α is also related to a number of toxic manifestations of infection (Oliff et al., 1987; Waage et al. 1989).

Fucoidan, which is found in the extracellular matrix of brown algae and brown seaweed, is polysaccharides containing fucose, mannose, galactose, and sulfate groups. These compounds of fucoidan possess diverse medicinal effect due to their ability to imitate patterns of sulfate substitution on glycosaminoglycans and sulfated glucans (Damonte et al. 2004; Li et al. 2008). Algal fucoidan has been widely examined in various biological contexts, including their anti-coagulant (Chandía and Matsuhiro, 2008), anti-angiogenic (Koyanagi et al. 2003), anti-proliferative (McCaffrey et al. 1992), anti-tumor (Alekseyenko et al. 2007), and anti-viral (Hayashi et al. 2008) properties.

The phagocytic capacities of macrophages are upregulated by fucoidan, and the activated cells produced markedly elevated levels of TNF-α (Choi et al. 2005). Although previous studies revealed that culture supernatant of fucoidan-stimulated canine peripheral blood mononuclear cells (PBMCs) boosts the phagocytic capacity of canine PMNs (Kim et al. 2011), no studies to date have evaluated the effect of fucoidan on the phagocytic capacity of PMNs under inflammatory conditions (Park et al. 2015; Park et al. 2017; Wang et al. 2018), which may be important in modulation of the inflammatory immune response.

In this study, we examined the modulating effect of fucoidan on the phagocytic capacity of porcine PMNs under inflammatory conditions. For this purpose, we administered fucoidan to porcine PMNs exposed to culture supernatant from lipopolysaccharide (LPS)-stimulated PBMCs.

Section snippets

Chemicals and reagents

Fucoidan purified from Focus vesiculosus, LPS from Escherichia coli 0127:B8, Percoll, RPMI 1640 medium, and carboxylate-modified polystyrene-latex beads were purchased from Sigma-Aldrich (St. Louis, MO, USA). Fetal bovine serum was purchased from Gibco (Grand Island, NY, USA), and anti-recombinant porcine (rp) TNF-α polyclonal antibody (pAb) was purchased from RayBiotech (Norcross, GA, USA). Stock solution of fucoidan was prepared by dissolving the compound in phosphate buffered saline (PBS) to

Effect of fucoidan on viability of porcine PBMCs and PMNs

PBMCs and PMNs were cultured for 24 h with fucoidan from 0 to 500 μg/ml and LPS from 0 to 1.0 μg/ml, and then their viability was evaluated by Trypan blue dye exclusion. As shown in Table 1, both PBMCs and PMNs exhibited high viability at concentrations from 0 to 200 μg/ml fucoidan and from 0 to 0.5 μg/ml LPS. However, viability was significantly decreased by fucoidan at a concentration of 500 μg/ml (p < 0.01) and LPS at a concentration of 1.0 μg/ml (p < 0.001). Subsequently, fucoidan and LPS

Discussion

Phagocytosis is upregulated in cultured macrophages treated with fucoidan or LPS for 48 h, and this effect is mediated in an autocrine fashion (Hayashi et al. 2008). Phagocytes such as neutrophils, monocytes, and macrophages express a broad spectrum of receptors that participate in particle recognition (Underhill and Ozinsky 2002). Differences in the complement of receptors between PMNs and macrophages might be responsible for differences in the effect of a compound on phagocytosis. However, in

Conflict of interest

There were no conflicts of interest for the authors.

Acknowledgements

This work was supported by a Global Research and Development Center (GRDC) Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (2017K1A4A3014959).

Authorship contributions

Hyeong-Mok Kim performed research, analyzed data, and wrote the manuscript; Changhwan Ahn, Byeong-Teck Kang, and Ji-Houn Kang performed research; Eui-Bae Jeung analyzed data; Mhan-Pyo Yang designed the research, analyzed data, wrote the manuscript, will be involved in drafting the manuscript, and will have an opportunity to approve subsequent revisions.

References (39)

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