CREB, NF-κB, and NADPH oxidase coordinately upregulate indoxyl sulfate-induced angiotensinogen expression in proximal tubular cells

Am J Physiol Cell Physiol. 2013 Apr 1;304(7):C685-92. doi: 10.1152/ajpcell.00236.2012. Epub 2013 Feb 13.

Abstract

In chronic kidney disease (CKD), indoxyl sulfate, a uremic toxin, accumulates in serum, and the expression of angiotensinogen (AGT) is upregulated in renal proximal tubular cells. The present study aimed to determine the relationship between indoxyl sulfate and the upregulation of AGT expression in proximal tubular cells. Indoxyl sulfate induced expression of AGT in rat renal cortex and in cultured human proximal tubular cells (HK-2). In proximal tubular cells, indoxyl sulfate induced phosphorylation of cAMP response element-binding protein (CREB) on Ser-133, and small interfering RNA (siRNA) specific to CREB inhibited indoxyl sulfate-induced AGT expression. Our previous study demonstrated that indoxyl sulfate activated nuclear factor-κB (NF-κB) through reactive oxygen species (ROS) production. NF-κB inhibitors (pyrrolidine dithiocarbamate and isohelenin), NF-κB p65 siRNA, an antioxidant [N-acetylcysteine (NAC)], and a nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor [diphenyleneiodonium (DPI)] suppressed indoxyl sulfate-induced AGT expression. Both NAC and DPI suppressed indoxyl sulfate-induced expression of NF-κB p65 and CREB. CREB siRNA suppressed indoxyl sulfate-induced NF-κB p65 expression, whereas both NF-κB inhibitors and NF-κB p65 siRNA prevented indoxyl sulfate-induced CREB expression. Furthermore, we focused on the expression of NADPH oxidase 4 (NOX4), because indoxyl sulfate induced NOX4 expression in vascular smooth muscle cells and vascular endothelial cells. Indoxyl sulfate induced the expression of NOX4 in proximal tubular cells, which was suppressed by NAC, DPI, NF-κB inhibitors, NF-κB p65 siRNA, and CREB siRNA. Taken together, CREB, NF-κB, and NOX4 coordinately upregulate indoxyl sulfate-induced AGT expression in proximal tubular cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Angiotensinogen / genetics
  • Angiotensinogen / metabolism*
  • Animals
  • Cell Line
  • Cyclic AMP Response Element-Binding Protein / genetics
  • Cyclic AMP Response Element-Binding Protein / metabolism*
  • Gene Expression Regulation / physiology
  • Humans
  • Indican / pharmacology*
  • Kidney Cortex / metabolism
  • Kidney Tubules, Proximal / cytology*
  • NADPH Oxidase 4
  • NADPH Oxidases / genetics
  • NADPH Oxidases / metabolism*
  • NF-kappa B / genetics
  • NF-kappa B / metabolism*
  • Rats
  • Reactive Oxygen Species
  • Up-Regulation

Substances

  • Cyclic AMP Response Element-Binding Protein
  • NF-kappa B
  • Reactive Oxygen Species
  • Angiotensinogen
  • NADPH Oxidase 4
  • NADPH Oxidases
  • NOX4 protein, human
  • Indican