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J. Biol. Chem., Vol. 278, Issue 11, 9142-9149, March 14, 2003
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From the Department of Pediatrics, University of Texas Southwestern
Medical Center, Dallas, Texas 75390 and the § Diabetes Unit,
Laboratory of Clinical Investigation, NCCAM, National Institutes of
Health, Bethesda, Maryland 20892-1755
High density lipoprotein (HDL) activates
endothelial nitric-oxide synthase (eNOS), leading to increased
production of the antiatherogenic molecule NO. A variety of stimuli
regulate eNOS activity through signaling pathways involving Akt kinase
and/or mitogen-activated protein (MAP) kinase. In the present study, we
investigated the role of kinase cascades in HDL-induced eNOS stimulation in cultured endothelial cells and COS M6 cells transfected with eNOS and the HDL receptor, scavenger receptor B-I. HDL
(10-50 µg/ml, 20 min) caused eNOS phosphorylation at Ser-1179, and
dominant negative Akt inhibited both HDL-mediated phosphorylation and
activation of the enzyme. Phosphoinositide 3-kinase (PI3 kinase)
inhibition or dominant negative PI3 kinase also blocked the
phosphorylation and activation of eNOS by HDL. Studies with genistein
and PP2 showed that the nonreceptor tyrosine kinase, Src, is an
upstream stimulator of the PI3 kinase-Akt pathway in this paradigm. In addition, HDL activated MAP kinase through PI3 kinase, and
mitogen-activated protein kinase/extracellular signal-regulated kinase
kinase inhibition fully attenuated eNOS stimulation by HDL without
affecting Akt or eNOS Ser-1179 phosphorylation. Conversely, dominant
negative Akt did not alter HDL-induced MAP kinase activation. These
results indicate that HDL stimulates eNOS through common upstream,
Src-mediated signaling, which leads to parallel activation of Akt and
MAP kinases and their resultant independent modulation of the enzyme.
High Density Lipoprotein-induced Endothelial Nitric-oxide
Synthase Activation Is Mediated by Akt and MAP Kinases*
,
*
This work was supported by the Scientist Development Program
of the American Heart Association (to C. M.) and National Institute of
Health Grants HL 58888, HL 53546, and HD 30276 (to P. W. S.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
To whom correspondence should be addressed: Dept. of Pediatrics,
University of Texas Southwestern Medical Center, 5323 Harry Hines
Blvd., Dallas, TX 75390. Tel.: 214-648-4574; Fax: 214-648-2481; E-mail:
chieko.mineo@utsouthwestern.edu.
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