Changes of cell-surface thiols and intracellular signaling in human monocytic cell line THP-1 treated with diphenylcyclopropenone

Morihiko Hirota, Akira Motoyama, Mie Suzuki, Masashi Yanagi, Masato Kitagaki, Hirokazu Kouzuki, Shigenobu Hagino, Hiroshi Itagaki, Hitoshi Sasa, Saori Kagatani, Setsuya Aiba

Research output: Contribution to journalArticlepeer-review

10 Citations (Scopus)

Abstract

Changes of cell-surface thiols induced by chemical treatment may affect the conformations of membrane proteins and intracellular signaling mechanisms. In our previous study, we found that a non-toxic dose of diphenylcyclopropene (DPCP), which is a potent skin sensitizer, induced an increase of cell-surface thiols in cells of a human monocytic cell line, THP-1. Here, we examined the influence of DPCP on intracellular signaling. First, we confirmed that DPCP induced an increase of cell-surface thiols not only in THP-1 cells, but also in primary monocytes. The intracellular reduced-form glutathione/oxidized-form glutathione ratio (GSH/GSSG ratio) was not affected by DPCP treatment. By means of labeling with a membrane-impermeable thiol-reactive compound, Alexa Fluor 488 C5 maleimide (AFM), followed by two-dimensional gel electrophoresis and analysis by liquid chromatography coupled with electrospray tandem mass spectrometry (LC/MS/MS), we identified several proteins whose thiol contents were modified in response to DPCP. These proteins included cell membrane components, such as actin and β-tubulin, molecular chaperones, such as heat shock protein 27A and 70, and endoplasmic reticulum (ER) stress-inducible proteins. Next, we confirmed the expression in DPCP-treated cells of spliced XBP1, a known marker of ER stress. We also detected the phosphorylation of SAPK/JNK and p38 MAPK, which are downstream signaling molecules in the IRE1α-ASK1 pathway, which is activated by ER stress. These data suggested that increase of cell-surface thiols might be associated with activation of ER stress-mediated signaling.

Original languageEnglish
Pages (from-to)871-879
Number of pages9
JournalJournal of Toxicological Sciences
Volume35
Issue number6
DOIs
Publication statusPublished - 2010 Dec

Keywords

  • 2DE
  • Cell-surface thiols
  • Diphenylcyclopropenone
  • ER stress
  • SH test
  • THP-1

ASJC Scopus subject areas

  • Toxicology

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