Actin filament-associated protein 1 (AFAP-1) is a key mediator in inflammatory signaling-induced rapid attenuation of intrinsic P-gp function in human brain capillary endothelial cells

Yutaro Hoshi, Yasuo Uchida, Masanori Tachikawa, Sumio Ohtsuki, Tetsuya Terasaki

Research output: Contribution to journalArticlepeer-review

13 Citations (Scopus)

Abstract

The purpose of this study was to identify regulatory molecule(s) involved in the inflammatory signaling-induced decrease in P-glycoprotein (P-gp) efflux function at the blood–brain barrier (BBB) that may occur in brain diseases. We confirmed that in vivo P-gp efflux activity at the BBB was decreased without any change in P-gp protein expression level in a mouse model of acute inflammation induced by 3 mg/kg lipopolysaccharide. In a human BBB model cell line (human brain capillary endothelial cells; hCMEC/D3), 1-h treatment with 10 ng/mL tumor necrosis factor-α (TNF-α; an inflammatory mediator) rapidly reduced P-gp efflux activity, but had no effect on P-gp protein expression level. To clarify the non-transcriptional mechanism that causes the decrease in intrinsic efflux activity of P-gp in acute inflammation, we applied comprehensive quantitative phosphoproteomics to compare hCMEC/D3 cells treated with TNF-α and vehicle (control). Actin filament-associated protein-1 (AFAP-1), MAPK1, and transcription factor AP-1 (AP-1) were significantly phosphorylated in TNF-α-treated cells, and were selected as candidate proteins. In validation experiments, knockdown of AFAP-1 expression blocked the reduction in P-gp efflux activity by TNF-α treatment, whereas inhibition of MAPK function or knockdown of AP-1 expression did not. Quantitative targeted absolute proteomics revealed that the reduction in P-gp activity by TNF-α did not require any change in P-gp protein expression levels in the plasma membrane. Our results demonstrate that AFAP-1 is a key mediator in the inflammatory signaling-induced, translocation-independent rapid attenuation of P-gp efflux activity in human brain capillary endothelial cells. (Figure presented.).

Original languageEnglish
Pages (from-to)247-262
Number of pages16
JournalJournal of Neurochemistry
Volume141
Issue number2
DOIs
Publication statusPublished - 2017 Apr 1

Keywords

  • P-glycoprotein
  • actin filament-associated protein-1
  • blood–brain barrier
  • inflammation
  • phosphorylation
  • proteomics

ASJC Scopus subject areas

  • Biochemistry
  • Cellular and Molecular Neuroscience

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