Mechanisms underlying endothelial dysfunction in diabetes mellitus.
Incubation of endothelial cells in vitro with high concentrations of glucose activates protein kinase C (PKC) and increases nitric oxide synthase (NOS III) gene expression as well as superoxide production. The underlying mechanisms remain unknown. To address this issue in an in vivo model, diabetes was induced with streptozotocin in rats. Streptozotocin treatment led to endothelial dysfunction and increased vascular superoxide production, as assessed by lucigenin- and coelenterazine-derived chemiluminescence. The bioavailability of vascular nitric oxide (as measured by electron spin resonance) was reduced in diabetic aortas, although expression of endothelial NOS III (mRNA and protein) was markedly increased. NOS inhibition with N:(G)-nitro-L-arginine increased superoxide levels in control vessels but reduced them in diabetic vessels, identifying NOS as a superoxide source. Similarly, we found an activation of the NADPH oxidase and a 7-fold increase in gp91(phox) mRNA in diabetic vessels. In vitro PKC inhibition with chelerythrine reduced vascular superoxide in diabetic vessels, whereas it had no effect on superoxide levels in normal vessels. In vivo PKC inhibition with N:-benzoyl-staurosporine did not affect glucose levels in diabetic rats but prevented NOS III gene upregulation and NOS-mediated superoxide production, thereby restoring vascular nitric oxide bioavailability and endothelial function. The reduction of superoxide in vitro by chelerythrine and the normalization of NOS III gene expression and reduction of superoxide in vivo by N:-benzoyl-staurosporine point to a decisive role of PKC in mediating these phenomena and suggest a therapeutic potential of PKC inhibitors in the prevention or treatment of vascular complications of diabetes mellitus. The full text of this article is available at http://www.circresaha.org.
Hink
U
U
Universitätskrankenhaus Eppendorf, Germany.
Li
H
H
Mollnau
H
H
Oelze
M
M
Matheis
E
E
Hartmann
M
M
Skatchkov
M
M
Thaiss
F
F
Stahl
R A
RA
Warnholtz
A
A
Meinertz
T
T
Griendling
K
K
Harrison
D G
DG
Forstermann
U
U
Munzel
T
T
D000818:Animals; D001011:Aorta; D001786:Blood Glucose / Q000187:drug effects; D003921:Diabetes Mellitus, Experimental / Q000139:chemically induced / Q000150:complications / Q000378:metabolism* / Q000503:physiopathology; D004195:Disease Models, Animal; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism* / Q000503:physiopathology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D066298:In Vitro Techniques; D008163:Luminescent Measurements; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III; D018384:Oxidative Stress / Q000187:drug effects; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013311:Streptozocin; D013481:Superoxides / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects; D014652:Vascular Diseases / Q000209:etiology / Q000378:metabolism*
D001786:Blood Glucose; D004791:Enzyme Inhibitors; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D013481:Superoxides; D009569:Nitric Oxide; D013311:Streptozocin; D019001:Nitric Oxide Synthase; D052250:Nitric Oxide Synthase Type III; C496316:Nos3 protein, rat; C056558:CYBB protein, human; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; D011493:Protein Kinase C