Reactive oxygen species and nuclear factor-kappa B pathway mediate high glucose-induced Pax-2 gene expression in mouse embryonic mesenchymal epithelial cells and kidney explants

Kidney Int. 2006 Nov;70(9):1607-15. doi: 10.1038/sj.ki.5001871. Epub 2006 Sep 20.

Abstract

Diabetic mellitus confers a major risk of congenital malformations, and is associated with diabetic embryopathy, affecting multiple organs including the kidney. The DNA paired box-2 (Pax-2) gene is essential in nephrogenesis. We investigated whether high glucose alters Pax-2 gene expression and aimed to delineate its underlying mechanism(s) of action using both in vitro (mouse embryonic mesenchymal epithelial cells (MK4) and ex vivo (kidney explant from Hoxb7-green florescent protein (GFP) mice) approaches. Pax-2 gene expression was determined by reverse transcriptase-polymerase chain reaction, Western blotting, and immunofluorescent staining. A fusion gene containing the full-length 5'-flanking region of the human Pax-2 promoter linked to a luciferase reporter gene, pGL-2/hPax-2, was transfected into MK4 cells with or without dominant negative IkappaBalpha (DN IkappaBalpha) cotransfection. Fusion gene expression level was quantified by cellular luciferase activity. Reactive oxygen species (ROS) generation was measured by lucigenin assay. Embryonic kidneys from Hoxb7-GFP mice were cultured ex vivo. High D(+) glucose (25 mM), compared to normal glucose (5 mM), specifically induced Pax-2 gene expression in MK4 cells and kidney explants. High glucose-induced Pax-2 gene expression is mediated, at least in part, via ROS generation and activation of the nuclear factor kappa B signaling pathway, but not via protein kinase C, p38 mitogen-activated protein kinase (MAPK), and p44/42 MAPK signaling.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line
  • Dose-Response Relationship, Drug
  • Electron Transport Complex I / antagonists & inhibitors
  • Electron Transport Complex I / genetics
  • Electron Transport Complex I / metabolism
  • Epithelium / drug effects
  • Epithelium / metabolism*
  • Gene Expression Regulation, Developmental / drug effects
  • Glucose / pharmacology*
  • Hyperglycemia / metabolism
  • Hyperglycemia / pathology
  • Kidney / drug effects
  • Kidney / embryology
  • Kidney / metabolism*
  • Mesoderm / cytology
  • Mesoderm / drug effects
  • Mesoderm / metabolism*
  • Mice
  • Mitogen-Activated Protein Kinase 1 / antagonists & inhibitors
  • Mitogen-Activated Protein Kinase 1 / genetics
  • Mitogen-Activated Protein Kinase 1 / metabolism
  • Mitogen-Activated Protein Kinase 3 / antagonists & inhibitors
  • Mitogen-Activated Protein Kinase 3 / genetics
  • Mitogen-Activated Protein Kinase 3 / metabolism
  • NADP / antagonists & inhibitors
  • NADP / genetics
  • NADP / metabolism
  • NF-kappa B / genetics
  • NF-kappa B / metabolism*
  • PAX2 Transcription Factor / genetics
  • PAX2 Transcription Factor / metabolism*
  • Protein Kinase C / antagonists & inhibitors
  • Protein Kinase C / genetics
  • Protein Kinase C / metabolism
  • Protein Kinase C / physiology
  • Reactive Oxygen Species / metabolism*
  • p38 Mitogen-Activated Protein Kinases / antagonists & inhibitors
  • p38 Mitogen-Activated Protein Kinases / genetics
  • p38 Mitogen-Activated Protein Kinases / metabolism

Substances

  • NF-kappa B
  • PAX2 Transcription Factor
  • Pax2 protein, mouse
  • Reactive Oxygen Species
  • NADP
  • Protein Kinase C
  • Mitogen-Activated Protein Kinase 1
  • Mitogen-Activated Protein Kinase 3
  • p38 Mitogen-Activated Protein Kinases
  • Electron Transport Complex I
  • Glucose