L- threo -C6-pyridinium-ceramide bromide, a novel cationic ceramide, induces NADPH oxidase activation, mitochondrial dysfunction and loss in cell viability in INS 832/13 β-cells

Cell Physiol Biochem. 2012;30(4):1051-8. doi: 10.1159/000341481. Epub 2012 Sep 24.

Abstract

Background: Emerging evidence indicates that exposure of isolated pancreatic β-cells to elevated glucose [glucotoxicity] or saturated fatty acids such as palmitate [lipotoxicity] or both [glucolipotoxicity] results in excessive intracellular oxidative stress mediated by phagocyte-like NADPH oxidase [Nox2]. Pharmacological evidence also implicates the intracellular generation of ceramide [CER] as one of the mediators of palmitate-induced cytotoxicity of the islet β- cell. Herein, we investigated the effects of L-threo-C6-pyridinium-ceramide bromide, a novel water soluble cationic ceramide [Ws-CER], on mitochondrial function and cell viability in insulin-secreting INS 832/13 cells.

Methods: Ws-CER, was synthesized as we reported earlier. Rac1 activation was quantitated by pull-down assay. Mitochondrial membrane potential was quantitated by JC-1 staining. Nox2 subunit expression and caspase-3 activity were determined by Western blotting.

Results: Our findings suggested a marked increase in the Nox2 activation [i.e., ROS generation and subunit expression and activation] in cells exposed to Ws-CER. We also noticed a significant reduction in mitochondrial membrane potential, increased in caspase-3 activity and associated loss in cell viability in Ws-CER-treated cells.

Conclusion: Based on these data we conclude that ceramide-induced Nox2-mediated oxidative stress couples mitochondrial dysfunction to loss in cell viability in the pancreatic β-cell.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Bromides / chemistry
  • Bromides / metabolism
  • Caspase 3 / metabolism
  • Cell Line
  • Cell Survival
  • Ceramides / chemistry
  • Ceramides / metabolism*
  • Enzyme Activation
  • Insulin-Secreting Cells / cytology*
  • Insulin-Secreting Cells / metabolism*
  • Membrane Glycoproteins / metabolism*
  • Membrane Potential, Mitochondrial
  • Mitochondria / metabolism*
  • NADPH Oxidase 2
  • NADPH Oxidases / metabolism*
  • Oxidative Stress
  • Pyridinium Compounds / chemistry
  • Pyridinium Compounds / metabolism*
  • Rats
  • Reactive Oxygen Species / metabolism

Substances

  • 2-N-(6'-(1''-pyridinium)hexanoyl)sphingosine
  • Bromides
  • Ceramides
  • Membrane Glycoproteins
  • Pyridinium Compounds
  • Reactive Oxygen Species
  • Cybb protein, rat
  • NADPH Oxidase 2
  • NADPH Oxidases
  • Caspase 3