Vitamin C-induced activation of phospholipase D in lung microvascular endothelial cells: Regulation by MAP kinases

Saradhadevi Varadharaj, Emily Steinhour, Melissa G. Hunter, Tonya Watkins, Christopher P. Baran, Ulysses Magalang, Periannan Kuppusamy, Jay L. Zweier, Clay B. Marsh, Viswanathan Natarajan, Narasimham L. Parinandi

Research output: Contribution to journalArticlepeer-review

Abstract

Our earlier studies have shown that vitamin C at pharmacological doses (mM) induces loss of redox-dependent viability in bovine lung microvascular endothelial cells (BLMVECs) that is mediated by oxidative stress. Therefore, here, we investigated the vitamin C-induced activation of the lipid signaling enzyme, phospholipase D (PLD) in BLMVECs. Monolayer cultures of BLMVECs were treated with vitamin C (0-10 mM) for different time periods (0-2 h) and the activity of PLD was determined. Vitamin C induced activation of PLD in BLMVECs in a time- and dose-dependent fashion that was significantly attenuated by antioxidants, p38 mitogen-activated protein kinase (p38 MAPK)-specific inhibitor (SB203580), extracellular signal-regulated protein kinase (ERK)-specific inhibitor (PD98059), and transient transfection of cells with dominant-negative (DN)-p38 MAPK and DN-ERK1/ERK2. Vitamin C also induced phosphorylation and enhanced the activities of p38 MAPK and ERK in BLMVECs in a time-dependent fashion. It was also evident that vitamin C induced translocation of PLD1 and PLD2, association of p38 MAPK and ERK with PLD1 and PLD2, threonine phosphorylation of PLD1 and PLD2 and SB203580- and PD98059-inhibitable threonine phosphorylation of PLD1 in BLMVECs. Transient transfection of BLMVECs with DN-p38 MAPK and DN-ERK1/ERK2 resulted in marked attenuation of vitamin C-induced phosphorylation of threonine in PLD1 and PLD2. We, for the first time, showed that vitamin C at pharmacological doses, activated PLD in the lung microvascular ECs through oxidative stress and MAPK activation.

Original languageEnglish (US)
Pages (from-to)1396-1407
Number of pages12
JournalCellular Signalling
Volume18
Issue number9
DOIs
StatePublished - Sep 2006
Externally publishedYes

Keywords

  • Ascorbic acid
  • Endothelial cell
  • MAPK signaling
  • Oxidative stress
  • Phospholipase D

ASJC Scopus subject areas

  • Cell Biology

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