Analysis of amphotericin B-induced cell signaling with chemical inhibitors of signaling molecules

Kenichiro Matsuo, Hitoshi Hotokezaka, Naoya Ohara, Yuji Fujimura, Atsutoshi Yoshimura, Yukio Okada, Yoshitaka Hara, Noriaki Yoshida, Koji Nakayama

Research output: Contribution to journalArticlepeer-review

21 Citations (Scopus)

Abstract

Although amphotericin B (AmB) is a major polyene antibiotic against invasive fungal infection, administration to patients sometimes causes inflammatory side effects, which limits the usage of the antibiotic. We studied the intracellular signaling that was induced by AmB. p65 (RelA) of nuclear factor-κB (NF-κB), a well-known signaling molecule as an inducer of proinflammatory cytokines, was phosphorylated by AmB in RAW264.7 cells, a monocyte-like cell line. Among chemical inhibitors of signaling molecules, U-73122 (phospholipase C (PLC) inhibitor), Gö6976 (protein kinase C (PKC) inhibitor), BAPTA-AM (calcium chelator), LFM-A13 (Bruton's tyrosine kinase (Btk)-specific inhibitor), and PP2 (c-Src kinase inhibitor) suppressed AmB-induced phosphorylation of p65 and translocation of p65 into the nucleus. U-73122 and Gö6976 reduced AmB-mediated induction of proinflammatory cytokines (tumor necrosis factor (TNF)-α and interleukin (IL)-6) in RAW264.7 cells. Furthermore, AmB-induced activation of NF-κB was observed in toll-like receptor (TLR) 2-expressed cells, and the activation of NF-κB was inhibited by U-73122, whereas peptidoglycan-induced NF-κB activation, which was also dependent on TLR2, was not inhibited by U-73122. Finally, U-73122 partially suppressed in vivo production of TNF-α and IL-6 induced by AmB administration in BALB/c mice. These results suggested that the signaling from AmB stimulation to proinflammatory cytokine production is mediated by TLR2, Btk, PLC, PKC, c-Src and NF-κB. These signaling molecules may become a target for chemotherapy suppressing AmB-induced proinflammatory cytokine production.

Original languageEnglish
Pages (from-to)337-347
Number of pages11
JournalMICROBIOLOGY and IMMUNOLOGY
Volume50
Issue number4
DOIs
Publication statusPublished - 2006
Externally publishedYes

Keywords

  • Amphotericin B
  • Cell signaling
  • Phospholipase C
  • Toll-like receptor

ASJC Scopus subject areas

  • Microbiology
  • Immunology
  • Virology

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