Cellular crosstalk between TNF-α, NADPH oxidase, PKCβ2, and C2GNT in human leukocytes

Joanna M. Tarr, Ning Ding, Kirti Kaul, Anna Antonell, Luis A. Pérez-Jurado, Rakesh Chibber

Research output: Contribution to journalArticlepeer-review

10 Citations (Scopus)

Abstract

Increasing evidence suggests that chronic, sub-clinical inflammation plays an important role in the pathogenesis of diabetic retinopathy. We have established the potential role of the inflammatory enzyme, core 2 β-1, 6-N-acetylglucosaminyltransferase (C2GNT) in diabetic retinopathy. The present study was designed to explore the NADPH oxidase signaling pathway in the tumor necrosis factor-alpha (TNF-α)-induced activity of C2GNT in leukocytes. Human leukocytes (U937 cells) and an Epstein-Barr-transformed lymphoblastoid cell line deficient in p47phox (F10007 cells) were used for the study. Cells were exposed to TNF-α for 24. h in the presence and absence of 1) NADPH oxidase inhibitors (apocynin and scrambled and unscrambled gp91ds-tat), 2) LY379196 (specific protein kinase C β1/2 (PKCβ1/2) inhibitor), and 3) the antioxidant tiron. Subsequent C2GNT and NADPH activity was measured and the adhesion of U937 and F10007 cells to endothelial cells was assessed. TNF-α-induced C2GNT activity (1813 ± 326. pmol/h/mg protein) (mean ± SEM) in human leukocytes was significantly reversed with apocynin (153 ± 82. pmol/h/mg protein), unscrambled gp91ds-tat (244 ± 122. pmol/h/mg protein) and tiron (756 ± 87. pmol/h/mg protein). We further supported this C2GNT-NADPH oxidase link using p47phox-deficient leukocytes. The deficiency in p47phox prevented TNF-α-induced NADPH oxidase and C2GNT activity and adherence to endothelial cells. The response to TNF-α was restored by transfection with an expression plasmid containing a p47phox cDNA inserted in the sense direction. Our results demonstrate for the first time a novel signaling crosstalk between TNF-α, NADPH oxidase, PKCβ1/2 and C2GNT in leukocytes.

Original languageEnglish
Pages (from-to)873-878
Number of pages6
JournalCellular Signalling
Volume24
Issue number4
DOIs
Publication statusPublished - 1 Apr 2012
Externally publishedYes

Keywords

  • 6-N-acetylglucosaminyltransferase
  • Adhesion
  • Core 2 β-1
  • Diabetic retinopathy
  • Inflammation
  • Leukostasis
  • NADPH oxidase

ASJC Scopus subject areas

  • Cell Biology

Cite this