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Am J Physiol Heart Circ Physiol (March 28, 2002). doi:10.1152/ajpheart.00029.2002
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Articles in PresS, published online ahead of print March 28, 2002
Am J Physiol Heart Circ Physiol, 10.1152/ajpheart.00029.2002
Submitted on January 15, 2002
Accepted on March 18, 2002

Native LDL and Minimally Oxidized LDL Differentially Regulate Superoxide Anion in Vascular Endothelium In Situ

David W Stepp1, Jingsong Ou2, Allan W Ackerman2, Scott Welak2, David Klick1, and Kirkwood A Pritchard, Jr.2*

1 Physiology, Medical College of Wisconsin, Milwaukee, WI, USA
2 Surgery, Div Pediatric Surgery, Medical College of Wisconsin, Milwaukee, WI, USA

* To whom correspondence should be addressed. E-mail: kpritch{at}mcw.edu.

Low-density lipoprotein (LDL) and its oxidized derivatives are hypothesized to impair vascular function by increasing superoxide anion (O2.-). To investigate mechanisms in situ isolated carotid arteries were incubated with native LDL (n-LDL) or minimally oxidized LDL (mm-LDL). Using en face fluorescent confocal microscopy and hydroethidine (HEt), an oxidant sensitive fluorescent probe, we found that n-LDL increased O2.- in vascular endothelium >4-fold by a L-nitroargininemethylester (L-NAME)-inhibitable mechanism. In contrast, mm-LDL increased O2.- in vascular endothelium >8-fold by mechanisms that were partially inhibited by L-NAME and allopurinol (AL) and essentially ablated by diphenyleneiodium (DPI). These data indicate that both n-LDL and mm-LDL uncouple endothelial nitric oxide synthase (eNOS) activity and that mm-LDL also activates xanthine oxidase and NADPH oxidoreductase to induce greater increases in O2.- generation than n-LDL. Western analysis revealed that both lipoproteins inhibited A23187-stimulated association of heat shock protein 90 (Hsp90) with eNOS without inhibiting phosphorylation of eNOS at serine 1179 (phospho-eNOS), an immunological index of electron flow. As Hsp90 mediates the balance of .NO and O2.- generation by eNOS these data provide new insight into the mechanisms by which n-LDL and mm-LDL uncouple eNOS activity to increase endothelial O2.- generation to impair vascular function.




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