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In high glucose protein kinase C-zeta activation is required for mesangial cell generation of reactive oxygen species.
- Source :
-
Kidney international [Kidney Int] 2005 Dec; Vol. 68 (6), pp. 2526-41. - Publication Year :
- 2005
-
Abstract
- Background: We postulated that in mesangial cells exposed to high glucose, protein kinase C-zeta (PKC-zeta) is necessary for the generation of reactive oxygen species (ROS) by nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and that the requirement of PKC-zeta for filamentous (F)-actin disassembly may involve ROS. To identify signaling mechanisms relevant to PKC-zeta activation and ROS generation, including phosphoinositide 3 kinase (PI3 kinase), we examined mesangial cell stimulation with platelet-derived growth factor (PDGF).<br />Methods: In primary rat mesangial cells cultured in 5.6 mmol/L or 30 mmol/L d-glucose, PKC-zeta expression was identified with immunoblotting and activity was analyzed in cell membrane immunoprecipitates and by confocal immunofluorescence imaging. ROS generation was measured by dichlorofluorescein fluorescence using confocal microscopy and was inhibited by transfection of antisense against NADPH subunits p22(phox) or p47(phox) or with Tempol. F-actin disassembly was observed by dual-channel confocal fluorescence imaging. PI3 kinase activity was detected by immunoblotting of phosphorylated Akt.<br />Results: In high glucose, generation of NADPH oxidase-dependent ROS was dependent on PKC-zeta. Conversely, sustained PKC-zeta activity was dependent on ROS generation, suggesting a positive feedback. PKC-zeta-dependent F-actin disassembly in high glucose required ROS generation. PDGF stimulated NADPH oxidase generation of ROS through a PKC-zeta mechanism that was independent of Akt phosphorylation and remained unchanged in high glucose.<br />Conclusion: In high glucose, mesangial cell PKC-zeta is required for ROS generation from NADPH oxidase similar to PDGF stimulation of PKC-zeta-dependent ROS generation through a pathway independent of PI3 kinase. F-actin disassembly in high glucose also requires ROS. A positive feedback loop occurs between ROS and the activation of PKC-zeta in high glucose.
- Subjects :
- Actins metabolism
Animals
Cells, Cultured
Male
Membrane Transport Proteins genetics
Mesangial Cells cytology
Mesangial Cells drug effects
NADPH Oxidases genetics
NADPH Oxidases metabolism
Oligonucleotides, Antisense
Phosphatidylinositol 3-Kinases metabolism
Phosphoproteins genetics
Platelet-Derived Growth Factor pharmacology
Rats
Rats, Sprague-Dawley
Signal Transduction drug effects
Signal Transduction physiology
Glucose pharmacology
Mesangial Cells enzymology
Protein Kinase C metabolism
Reactive Oxygen Species metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0085-2538
- Volume :
- 68
- Issue :
- 6
- Database :
- MEDLINE
- Journal :
- Kidney international
- Publication Type :
- Academic Journal
- Accession number :
- 16316329
- Full Text :
- https://doi.org/10.1111/j.1523-1755.2005.00660.x