Temporal changes in the expression of mRNA of NADPH oxidase subunits in renal epithelial cells exposed to oxalate or calcium oxalate crystals

Saeed R. Khan, Aslam Khan, Karen J. Byer

Research output: Contribution to journalArticle

Abstract

Background. Exposure of renal epithelial cells to oxalate (Ox) or calcium oxalate (CaOx) crystals leads to the production of reactive oxygen species and cell injury. We have hypothesized that Ox and CaOx crystals activate NADPH oxidase through upregulation of its various subunits.Methods. Human renal epithelial-derived cell line, HK-2, was exposed to 100 μmol Ox or 66.7 μg/cm2 CaOx monohydrate crystals for 6, 12, 24 or 48 h. After exposure, the cells and media were processed to determine activation of NADPH oxidase, production of superoxide and 8-isoprostane (8IP), and release of lactate dehydrogenase (LDH). RT-PCR was performed to determine mRNA expression of NADPH subunits p22phox, p40phox, p47phox, p67phox and gp91phox as well as Rac-GTPase.Results. Exposure to Ox and CaOx crystals resulted in increase in LDH release, production of 8-IP, NADPH oxidase activity and production of superoxide. Exposure to CaOx crystals resulted in significantly higher NADPH oxidase activity, production of superoxide and LDH release than Ox exposure. Exposure to Ox and CaOx crystals altered the expression of various subunits of NADPH oxidase. More consistent were increases in the expression of membrane-bound p22phox and cytosolic p47phox. Significant and strong correlations were seen between NADPH oxidase activity, the expression of p22phox and p47phox, production of superoxide and release of LDH when cells were exposed to CaOx crystals. The expressions of neither p22phox nor p47phox were significantly correlated with increased NADPH oxidase activity after the Ox exposure.Conclusions. As hypothesized, exposure to Ox or CaOx crystals leads to significant increases in the expression of p22 phox and p47phox, leading to activation of NADPH oxidase. Increased NADPH oxidase activity is associated with increased superoxide production and lipid peroxidation. Different pathways appear to be involved in the stimulation of renal epithelial cells by exposure to Ox and CaOx crystals.

Original languageEnglish (US)
Pages (from-to)1778-1785
Number of pages8
JournalNephrology Dialysis Transplantation
Volume26
Issue number6
DOIs
StatePublished - Jun 2011
Externally publishedYes

Keywords

  • calcium oxalate
  • kidney stones
  • NADPH oxidase
  • oxidative stress
  • p47

ASJC Scopus subject areas

  • Nephrology
  • Transplantation

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