S-glutathiolation of p21ras by peroxynitrite mediates endothelial insulin resistance caused by oxidized low-density lipoprotein.
OBJECTIVE
To understand the mechanism by which oxidants are linked to insulin resistance, bovine aortic endothelial cells were exposed to oxidized low-density lipoproteins (oxLDL) or peroxynitrite.


RESULTS
OxLDL transiently increased phosphorylation of Erk and Akt within 5 minutes, but 60 minutes later, resulted in decreased insulin-induced Akt phosphorylation. OxLDL promoted a 2- to 5-fold increase in oxidant generation as measured by dihydrorhodamine or dihydroethidium oxidation that was ascribed to peroxynitrite. Exogenous peroxynitrite (25 to 100 micromol/L) or oxidized glutathione mimicked the effects of oxLDL. OxLDL increased the S-glutathiolation of p21ras, and adenoviral transfection with either a mutant p21ras (C118S) lacking the predominant site of S-glutathiolation or a dominant-negative mutant restored insulin-induced Akt phosphorylation. The requirement for oxidant-mediated S-glutathiolation and activation of p21ras in mediating insulin resistance was further implicated by showing that insulin signaling was restored by Mek inhibitors or by overexpression of glutaredoxin-1. Furthermore, oxLDL increased Erk-dependent phosphorylation of insulin receptor substrate-1 serine-616 that was prevented by inhibiting oxidant generation, Erk activation, or by the p21ras C118S mutant.


CONCLUSIONS
This study provides direct evidence for a novel molecular mechanism by which oxidants can induce insulin resistance via S-glutathiolation of p21ras and Erk-dependent inhibition of insulin signaling.
Clavreul
Nicolas
N
Director, Vascular Biology Unit, 650 Albany Street, X704, Boston, MA 02118. racohen@bu.edu.
Bachschmid
Markus M
MM
Hou
Xiuyun
X
Shi
Chaomei
C
Idrizovic
Azra
A
Ido
Yasuo
Y
Pimentel
David
D
Cohen
Richard A
RA
D000527:Alprostadil / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001011:Aorta / Q000166:cytology; D002417:Cattle; D015232:Dinoprostone / Q000819:agonists; D042783:Endothelial Cells / Q000187:drug effects / Q000502:physiology; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000502:physiology*; D004789:Enzyme Activation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism*; D019803:Glutathione Disulfide / Q000494:pharmacology; D007328:Insulin / Q000378:metabolism; D055504:Insulin Receptor Substrate Proteins; D007333:Insulin Resistance / Q000502:physiology*; D008077:Lipoproteins, LDL / Q000494:pharmacology*; D008244:Lysophosphatidylcholines / Q000494:pharmacology; D015689:Oncogene Protein p21(ras) / Q000187:drug effects / Q000378:metabolism*; D016877:Oxidants / Q000494:pharmacology; D030421:Peroxynitrous Acid / Q000494:pharmacology*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011459:Prostaglandins E, Synthetic / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology
D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D008077:Lipoproteins, LDL; D008244:Lysophosphatidylcholines; D016877:Oxidants; D010750:Phosphoproteins; D011459:Prostaglandins E, Synthetic; C099252:oxidized low density lipoprotein; D030421:Peroxynitrous Acid; D048049:Extracellular Signal-Regulated MAP Kinases; D015689:Oncogene Protein p21(ras); D000527:Alprostadil; D005978:Glutathione; C048491:butaprost; D015232:Dinoprostone; D019803:Glutathione Disulfide