Development of a novel in vitro assay to evaluate environmental water using an IL-8 reporter cell line

Yutaka Kimura, Chizu Fujimura, Toshifumi Imagawa, Socorro P. Lupisan, Mariko Saito-Obata, Mayuko Saito, Hitoshi Oshitani, Setsuya Aiba

Research output: Contribution to journalArticlepeer-review

Abstract

The IL-8 luciferase reporter cell line, THP-G8 cells, used in the in vitro sensitization test, OECD442E, can respond to a variety of stimuli other than haptens, such as lipopolysaccharide (LPS), other bacterial toxins, and detergents. Considering these characteristics, we examined the ability of the IL-8 luciferase assay using THP-G8 cells to evaluate water pollution. We first stimulated THP-G8 cell with various Toll-like receptor (TLR) agonists and nucleotide-binding oligomerization domain-like receptor (NLR) agonists, and found that TLR1, 2, 4, 5, 6 agonists and NOD 1, 2 agonists significantly augmented IL-8 luciferase activity (IL8LA). Then, we examined the detection threshold of LPS by THP-G8 cells, and found it 0.4 EU/ml. Next, we examined whether THP-G8 cells can differently respond to a variety of sources of environmental water around Sendai, Japan and Manila, Philippine and whether there is a correlation between the IL8LA of different sources of water and their level of endotoxin assessed by the LAL assay. There was a clear trend that the IL8LA was lower in the upper stream and higher in the downstream in both Japan and Philippine. Moreover, there was a strong correlation between the IL8LA of the environmental water and its endotoxin level. Finally, using N-acetyl-L-cysteine, an antioxidant/radical scavenger, and polymyxin B that neutralizes endotoxin, we demonstrated that there was a difference in the suppressive effects by them between the water from Japan and that from Philippine. These data suggest the potential of the IL-8 luciferase assay for evaluating environmental water pollution both quantitatively and qualitatively.

Original languageEnglish
Pages (from-to)1054-1063
Number of pages10
JournalEXCLI Journal
Volume19
DOIs
Publication statusPublished - 2020
Externally publishedYes

Keywords

  • Hapten
  • IL-8
  • Luciferase assay
  • Monocyte
  • NOD-like receptors
  • Toll-like receptors

ASJC Scopus subject areas

  • Molecular Medicine
  • Animal Science and Zoology
  • Pharmacology
  • Drug Discovery

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