Donate Help Contact The AHA Sign In Home
American Heart Association
Circulation Research
Search: search_blue_button Advanced Search
Circulation Research. 2002;90:107-111
Published online before print November 26, 2001, doi: 10.1161/hh0102.102359
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
90/1/107    most recent
hh0102.102359v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Beckman, J. A.
Right arrow Articles by Creager, M. A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Beckman, J. A.
Right arrow Articles by Creager, M. A.
Right arrowPubmed/NCBI databases
*Compound via MeSH
*Substance via MeSH
Related Collections
Right arrow Other diabetes
Right arrow Endothelium/vascular type/nitric oxide
Right arrow Mechanism of atherosclerosis/growth factors
(Circulation Research. 2002;90:107.)
© 2002 American Heart Association, Inc.


Clinical Research

Inhibition of Protein Kinase Cß Prevents Impaired Endothelium-Dependent Vasodilation Caused by Hyperglycemia in Humans

Joshua A. Beckman, Allison B. Goldfine, Mary Beth Gordon, Leslie A. Garrett, Mark A. Creager

From the Cardiovascular Division (J.A.B., M.B.G., L.A.G., M.A.C.), Brigham and Women’s Hospital, and the Division of Cellular and Molecular Physiology (A.B.G.), Joslin Diabetes Center, Harvard Medical School, Boston, Mass.

Correspondence to Mark A. Creager, MD, Cardiovascular Division, Brigham and Women’s Hospital, 75 Francis St, Boston, MA 02115. E-mail mcreager{at}partners.org


*    Abstract
up arrowTop
*Abstract
down arrowIntroduction
down arrowMaterials and Methods
down arrowResults
down arrowDiscussion
down arrowReferences
 
The bioavailability of nitric oxide is decreased in animal models and humans with diabetes mellitus. Hyperglycemia, in particular, attenuates endothelium-dependent vasodilation in healthy subjects. In vitro and in vivo animal studies implicate activation of protein kinase Cß as an important mechanism whereby hyperglycemia decreases endothelium-derived nitric oxide. Accordingly, this study tested the hypothesis that inhibition of protein kinase Cß would prevent impairment of endothelium-dependent vasodilation in healthy humans exposed to hyperglycemia. This study was a randomized, double-blind, placebo-controlled, crossover trial. Healthy subjects were treated with an orally active, selective, protein kinase Cß inhibitor, LY333531, or matching placebo once a day for 7 days before vascular function testing. Forearm blood flow was measured using venous-occlusion, strain-gauge plethysmography. Endothelium-dependent vasodilation was measured via incremental brachial artery administration of methacholine chloride (0.3 to 10 µg/min) during euglycemia and after 6 hours of hyperglycemic clamp. The forearm blood flow dose-response curve to methacholine was significantly attenuated by hyperglycemia after placebo treatment (P=0.009 by ANOVA, euglycemia versus hyperglycemia) but not after treatment with LY333531. Inhibition of protein kinase Cß prevents the reduction in endothelium-dependent vasodilation induced by acute hyperglycemia in healthy humans in vivo. These findings suggest that hyperglycemia impairs endothelial function, in part, via protein kinase Cß activation.


Key Words: protein kinase C • nitric oxide • hyperglycemia • endothelium • diabetes


*    Introduction
up arrowTop
up arrowAbstract
*Introduction
down arrowMaterials and Methods
down arrowResults
down arrowDiscussion
down arrowReferences
 
Vascular disease is the principal cause of morbidity and mortality in patients with diabetes mellitus.1 Diabetes mellitus is associated with changes in endothelial cell function that augur the development of atherosclerosis. An important early change, decreased bioavailability of endothelium-derived nitric oxide, is linked to many of the pathological features of atherosclerosis including upregulation of leukocyte adhesion molecules, platelet activation, and an increased propensity for vasoconstriction.2,3 Previous studies have demonstrated decreased endothelium-dependent vasodilation, a physiological marker of decreased bioavailability of nitric oxide in both conduit arteries and resistance vessels in experimental models of diabetes and humans with type 1 and type 2 diabetes.46

The central importance of hyperglycemia to the development of cardiovascular disease in diabetes mellitus is becoming increasingly evident. Population studies have revealed that an incremental risk of cardiovascular disease is associated with higher levels of blood glucose, beginning in the upper normal range.7 Hyperglycemia, per se, impairs vasodilator function in animals and healthy humans, similar to that which occurs in patients with diabetes.8,9 This cannot be attributed to downregulation of endothelial nitric oxide synthase (eNOS), since glucose increases eNOS mRNA expression and protein levels in cultured endothelial cells and vessels from diabetic animals.10,11 Despite this upregulation, there is reduced endothelium-derived nitric oxide in hyperglycemic states. Of potential mechanisms by which hyperglycemia may decrease the bioavailability of nitric oxide, recent evidence implicates a prominent role for activation of protein kinase C.1214

Protein kinase C is a cytoplasmic family of enzymes with a wide variety of actions in intracellular signal transduction.15 The activation of protein kinase C decreases endothelium-derived nitric oxide synthesis, whereas its inhibition augments nitric oxide release.1618 Of the many types of protein kinase C in vascular tissue, the ß isoforms are activated to a greater magnitude than other isoforms in response to hyperglycemia.19,20 Thus, this subtype may be central to the vascular dysfunction seen with hyperglycemia.

Recently, a selective inhibitor of protein kinase Cß, LY333531, has been described.21 Investigations using this compound in experimental models of diabetes indicate that inactivation of protein kinase Cß abrogates many of the pathophysiological vascular changes seen in hyperglycemia. Accordingly, the purpose of this investigation was to test the hypothesis that hyperglycemia-induced activation of protein kinase Cß decreases the bioavailability of endothelium-derived nitric oxide and impairs vasodilator function in healthy humans. Specifically, we sought to determine whether pretreatment with an inhibitor of protein kinase Cß, LY333531, would prevent decreases in endothelium-dependent vasodilation caused by experimental hyperglycemia in intact, healthy humans.


*    Materials and Methods
up arrowTop
up arrowAbstract
up arrowIntroduction
*Materials and Methods
down arrowResults
down arrowDiscussion
down arrowReferences
 
Subjects
Fifteen healthy volunteers were recruited via newspaper advertisement and provided written, informed consent. All subjects underwent screening, consisting of a medical history, physical examination, and laboratory studies to obtain values for complete blood cell count, serum electrolytes, fasting glucose, blood urea nitrogen, creatinine, transaminases, alkaline phosphatase, and lipid profile. Subjects with hypertension, history of tobacco use, LDL or total cholesterol greater than the 75th percentile for age and gender, cardiovascular disease, or other disease were excluded. The protocol was approved by the Human Research Committee of Brigham and Women’s Hospital.

Study Design
The effect of protein kinase Cß inhibition on endothelium-dependent vasodilation during hyperglycemia was studied in a randomized, double-blind, placebo-controlled, crossover design. All subjects were studied in the morning in the postabsorptive state, fasting after the previous midnight. Subjects were randomized to receive either LY333531 (Eli Lilly and Company), 32 mg orally once daily, or matching placebo for 7 days before and on the morning of each vascular function study. After a minimum 2-week washout period, subjects then crossed over and received the other medication for 7 days before the second study day. Female participants underwent vascular testing during the same menstrual phase each visit. Cyclooxygenase inhibitors, alcohol, and caffeine were prohibited for 12 hours before the study morning.

On the morning of each study, an indwelling antecubital venous catheter and brachial artery catheter were inserted. After a minimum of 30 minutes following catheter insertion, LY333531 levels and basal forearm blood flow were measured. Thereafter, endothelium-dependent vasodilation was assessed by measuring the forearm blood flow response to incremental intra-arterial doses of methacholine chloride (0.3, 1.0, 3.0, and 10.0 µg/min) infused at a flow rate of 0.388 mL/min. Each dose was administered for 6 minutes, and forearm blood flow measurements were made during the last 2 minutes of the infusion. After completion of euglycemic measurements, infusion of dextrose was initiated to maintain forearm hyperglycemia. After 6 hours of hyperglycemic clamp (see next section), basal forearm blood flow and the blood flow responses to methacholine were measured again. Endothelium-independent vasodilation was not tested because it has been demonstrated to be unimpaired in vitro, in animal models of hyperglycemia, and in intact humans with experimentally induced hyperglycemia.8,9,12,22,23 The vascular research laboratory was quiet, dimly lit, and temperature-controlled at 23°C.

Forearm Hyperglycemic Clamp
A forearm hyperglycemic clamp was used to raise and maintain forearm glucose concentration at 300 mg/dL (16.7 mmol/L) as previously described.9 Dextrose (50% solution) was infused via the brachial artery catheter into the forearm. Fifteen minutes after the infusion was started, the blood glucose level was determined from antecubital venous blood and the infusion rate adjusted to maintain the hyperglycemic clamp at 300 mg/dL. The infusion rate was adjusted every 10 to 15 minutes for the duration of the study, usually ranging between 0.1 and 0.3 mL/min. In addition, the somatostatin analogue, octreotide, was infused at 30 ng · kg-1 · min-1, to suppress pancreatic insulin release, since insulin is a known vasodilator whose vascular effects are mediated, at least in part, by endothelium-derived nitric oxide.24,25 The octreotide infusion was initiated at least 15 minutes before the first hemodynamic measurement and maintained throughout the protocol. No vasoactive effects have been identified in studies using a similar dose of octreotide.9,26,27

Hemodynamic Measurements
Bilateral forearm blood flow was measured by venous-occlusion, mercury-in-silastic, strain-gauge plethysmography (D.E. Hokanson, Issaquah, Wash) using established methods. The hand circulation was excluded during data acquisition using wrist cuffs inflated to 200 mm Hg. A venous occlusion pressure of 40 mm Hg was generated by cuffs placed on each arm above the elbow for each measurement of forearm blood flow. Forearm blood flow is reported as milliliters per 100 mL of tissue/min. Arterial blood pressure was measured via the brachial artery cannula. The cannula was attached to a pressure transducer contiguous with an amplifier on a Gould physiological recorder. Heart rate was determined by the RR interval of a continuous ECG monitor.

Laboratory Analyses
Whole-blood glucose concentration was measured at the bedside by the glucose oxidase method using a glucose reflectometer (Lifescan, Inc). Concentrations of LY333531 were determined by liquid chromatography with tandem mass spectrometry (Advanced BioAnalytical Services, Inc).

Statistical Analyses
Descriptive measures are reported as mean±SD. Experimental measures are reported as mean±SE. Basal forearm blood flows and glucose concentrations were compared by paired two-tailed t tests. Two-way, repeated-measures ANOVA was performed to compare the dose-response curves during euglycemia and after 6 hours of hyperglycemic clamp, using the absolute increase in blood flow from the basal flow rate. Statistical significance was accepted at the 95% confidence level (P<=0.05). Before the randomization code was broken, one subject was excluded from the analysis because of technical problems in acquiring forearm hemodynamic measurements. Data reported in the Results section do not include this subject. Additional statistical analysis was performed that included this subject’s data, and it did not change the significance of the interventions.


*    Results
up arrowTop
up arrowAbstract
up arrowIntroduction
up arrowMaterials and Methods
*Results
down arrowDiscussion
down arrowReferences
 
Baseline Characteristics
The 14 evaluable subjects, aged 26±6 years, included seven men and seven women. At the screening visit, mean arterial pressure was 86±10 mm Hg, blood glucose was 82±15 mg/dL, and total cholesterol was 141±19 mg/dL. Values obtained for fasting glucose, total and HDL cholesterol, triglycerides, and systolic and diastolic blood pressure were within normal limits in every subject.

Effect of Hyperglycemia and Protein Kinase Cß Inhibition
Forearm glucose concentrations averaged 317±45 mg/dL during 6 hours of hyperglycemia with placebo treatment and 336±83 mg/dL with LY333531 treatment (P=NS). LY333531 treatment resulted in plasma LY333531 concentrations of 16.3±4.3 nmol/L before euglycemia forearm blood flow measurements and 3.5±2.8 nmol/L after 6 hours of hyperglycemia just before data acquisition.

Baseline forearm blood flow was measured before the methacholine chloride infusion in each condition. Basal euglycemic forearm blood flow did not differ between placebo or LY333531 treatment periods, 1.7±0.1 versus 1.6±0.1 mL per 100 mL/min, respectively, P=NS (Figure 1). Six hours of hyperglycemic clamp increased basal forearm blood flow compared with the respective euglycemic baseline in both settings: to 2.4±0.2 mL per 100 mL/min with placebo treatment (P=0.008) and to 2.9±0.2 mL per 100 mL/min with LY333531 treatment (P=0.001) (Figure 1). The increase in basal forearm blood flow from euglycemia to hyperglycemia was significantly greater with LY333531 pretreatment than placebo (P=0.037).



View larger version (24K):
[in this window]
[in a new window]
 
Figure 1. Basal forearm blood flow at baseline euglycemia and during clamp, with and without PKCß inhibition. Resting forearm blood flow significantly increased from euglycemic baseline to 6-hour hyperglycemic clamp with and without LY333531 (P=0.008 and 0.001, respectively). The increase in resting flow was greater in the setting of LY333531 (P=0.037).

Incremental doses of methacholine chloride increased forearm blood flow during euglycemia and hyperglycemia. The forearm blood flow response to methacholine chloride during euglycemia was not significantly different with or without inhibition of protein kinase Cß (P=NS). During placebo treatment, 6 hours of hyperglycemic clamp significantly diminished the forearm blood flow response to methacholine compared with euglycemia (P=0.009) (Figure 2). In contrast, with protein kinase Cß inhibition, there was no significant difference in the forearm blood flow response to methacholine chloride between euglycemia and hyperglycemia (P=NS) (Figure 3).



View larger version (15K):
[in this window]
[in a new window]
 
Figure 2. Effect of hyperglycemia on endothelium-dependent vasodilation during placebo treatment. The increase in forearm blood flow from baseline induced by incremental methacholine at baseline and during hyperglycemic clamping is illustrated. Endothelium-dependent vasodilation was significantly attenuated during hyperglycemia (P=0.009).



View larger version (15K):
[in this window]
[in a new window]
 
Figure 3. Effect of inhibition of PKCß and hyperglycemia on endothelium-dependent vasodilation. The increase in forearm blood flow from baseline induced by methacholine at baseline and during hyperglycemic clamping with LY333531 treatment is shown. Endothelium-dependent vasodilation was not significantly changed during hyperglycemia (P=0.68).

Throughout the vascular function studies, there was no change in forearm blood flow in the contralateral forearm. Blood pressure and heart rate also remained stable and without significant variation during the course of the study.


*    Discussion
up arrowTop
up arrowAbstract
up arrowIntroduction
up arrowMaterials and Methods
up arrowResults
*Discussion
down arrowReferences
 
The novel finding of this investigation is that selective inhibition of protein kinase Cß prevents hyperglycemia-induced impairment of endothelium-dependent vasodilation in healthy, nondiabetic humans in vivo. These results suggest that activation of protein kinase Cß by hyperglycemia occurs within hours and importantly contributes to endothelial dysfunction. To our knowledge, this is the first report in intact humans that specific inhibition of protein kinase Cß preserves endothelium-dependent vasodilator function in the presence of hyperglycemia.

Hyperglycemia and Protein Kinase C Activation
Diabetes-related protein kinase C activation results in endothelial dysfunction manifested by decreased nitric oxide bioavailability, increased production of oxygen-derived free radicals, increased leukocyte adhesion molecule expression and leukocyte adhesion, increased albumin permeability, and impaired fibrinolysis.2832 Experimentally, hyperglycemia activates protein kinase C in endothelial and vascular smooth muscle cells and consistently produces similar findings of endothelial dysfunction including decreased endothelium-dependent vasodilation.9,31,33,34 The effect of hyperglycemia on vascular smooth muscle cell function is less clear. Experimental evidence demonstrates both increased and decreased vasoconstriction in response to hyperglycemia.35,36 Hypersensitivity to vasoconstriction does not appear to contribute importantly to the impairment of endothelium-dependent vasodilation in humans with diabetes or healthy humans exposed to hyperglycemia.3739

Of the many isoforms of protein kinase C activated by hyperglycemia, the ß isoforms play a prominent role in vivo in vascular dysfunction. Protein kinase Cß is preferentially activated in vascular endothelium in both diabetic rat and hyperglycemic dog models.19,20 Further, protein kinase Cß causes physiological abnormalities in vivo including increased retinal mean circulation time, glomerular filtration rate, and albumin excretion time in an in vivo rat model.14 Thus, protein kinase Cß contributes importantly to hyperglycemia-mediated vascular dysfunction.

Hyperglycemia-mediated protein kinase C activation may be caused by a number of mechanisms including elevated diacylglycerol concentration and mitochondrial superoxide anion overproduction. Diacylglycerol concentrations increase as a result of de novo synthesis from augmented glucose metabolism.40,41 Increased diacylglycerol concentrations cause membrane translocation and activation of protein kinase C.40 A recent study indicates that protein kinase C activation also may result from glucose-induced superoxide anion production. Increases in cytosolic glucose concentrations enhance intracellular superoxide anion production by generating an electrochemical gradient in the mitochondrial electron transport chain of the tricarboxylic acid pathway.42 Inhibitors of this pathway abrogate increases in protein kinase C activity.42 Mitochondrial superoxide overproduction may cause the activation of protein kinase C by inducing the de novo synthesis of diacylglycerol or phosphatidylcholine synthesis.

Protein Kinase C and Endothelium-Derived Nitric Oxide
Our study supports an important role for protein kinase Cß activation as a cause of hyperglycemia-induced, impaired endothelium-dependent vasodilation in human arterial resistance vessels. Indeed, we observed that LY333531, a specific inhibitor of protein kinase Cß, prevented the expected decrement in endothelium-dependent vasodilation during acute hyperglycemia.9,21,27 These findings cannot be explained by constitutive activity of protein kinase Cß or by effects of LY333531 in euglycemic conditions, because the euglycemic responses to methacholine chloride, with and without LY333531, were not different. Because the change in response to methacholine becomes evident with hyperglycemia, it is likely that in healthy humans, significant protein kinase Cß activation occurs as a consequence of hyperglycemia and decreases the bioavailability of endothelium-derived nitric oxide.

Experimental evidence suggests several mechanisms by which protein kinase C may decrease the bioavailability of nitric oxide. Activation of protein kinase C antagonizes phosphatidylinositol-3 kinase–mediated activation of endothelial nitric oxide synthase and decreases endothelium-derived perivascular nitric oxide concentration.43 In addition, activation of protein kinase C increases superoxide anion formation from several sources. Protein kinase C induces NAD(P)H oxidase to produce superoxide anion, which subsequently uncouples endothelial nitric oxide synthase and augments production of superoxide anion preferentially over nitric oxide.10,11,44,45 Superoxide anion further decreases the bioavailability of nitric oxide by reacting with it to form peroxynitrite. We have previously demonstrated that hyperglycemia-induced impairment of endothelium-dependent vasodilation can be reversed via infusion of an antioxidant in humans in vivo.27 Thus, our present observations lend support to the notion that hyperglycemic activation of protein kinase C in humans yields an oxidative stress that further decreases the bioavailability of nitric oxide.

It is conceivable that protein kinase C also may impair endothelium-independent vasodilation. Hyperglycemia activates protein kinase C in vascular smooth muscle cells,19,46 and the contractile response in vascular smooth muscle is abnormal in animal models of diabetes mellitus and in humans with type 2 but not type 1 diabetes.35,37,47 However, in experimental and human models of hyperglycemia, no abnormality in endothelium-independent vasodilation has been demonstrated.8,9,12,22,23 Our observations do not exclude the possibility that hyperglycemia-mediated protein kinase C activation alters vascular smooth muscle vasodilator function, but these previous investigations make this possibility less likely.

Basal Forearm Blood Flow
As we and others have demonstrated previously, hyperglycemia significantly increases resting forearm blood flow from baseline euglycemia to the 6-hour hyperglycemic clamp. This effect has been mimicked by infusion of mannitol and hypertonic saline, implicating increased osmolality as a cause for this phenomenon.9,48 Interestingly, in our subjects, the increase in resting forearm blood flow in response to hyperglycemia was greater with inhibition of protein kinase Cß. Our findings do not provide a precise mechanism for the change in resting forearm flow; however, unimpaired constitutive nitric oxide production, despite hyperglycemia, resulting from protein kinase Cß inhibition, may contribute to this finding.

Conclusion
Hyperglycemia increases the activity of protein kinase Cß, which decreases endothelium-dependent relaxation. This mechanism may contribute to vascular dysfunction in patients with hyperglycemia. Inhibition of protein kinase Cß by LY333531 may improve vascular function in patients with diabetes mellitus. Protein kinase Cß may therefore be an appropriate therapeutic target for patients with diabetes and vascular dysfunction.


*    Acknowledgments
 
This study was supported by grants from Eli Lilly & Company and the National Institutes of Health (HL-56607, HL-48743, and K23 HL-04169). Dr Beckman is the recipient of an American College of Cardiology/Merck Award. We would like to thank Dr George King for his critical appraisal of the manuscript and Dr James R. Woodworth for analysis of the pharmacokinetic data.

Received May 23, 2001; revision received September 24, 2001; accepted November 8, 2001.


*    References
up arrowTop
up arrowAbstract
up arrowIntroduction
up arrowMaterials and Methods
up arrowResults
up arrowDiscussion
*References
 

  1. Grundy SM, Benjamin IJ, Burke GL, Chait A, Eckel RH, Howard BV, Mitch W, Smith SC, Sowers JR. Diabetes and cardiovascular disease: a statement for healthcare professionals from the American Heart Association. Circulation. 1999; 100: 1134–1146.
  2. Anderson TJ, Gerhard MD, Meredith IT, Charbonneau F, Delagrange D, Creager MA, Selwyn AP, Ganz P. Systemic nature of endothelial dysfunction in atherosclerosis. Am J Cardiol. 1995; 75: 71B–74B.
  3. Busse R, Fleming I. Endothelial dysfunction in atherosclerosis. J Vasc Res. 1996; 33: 181–194.
  4. Meraji S, Jayakody L, Senaratne MP, Thomson AB, Kappagoda T. Endothelium-dependent relaxation in aorta of BB rat. Diabetes. 1987; 36: 978–981.
  5. Clarkson P, Celermajer DS, Donald AE, Sampson M, Sorensen KE, Adams M, Yue DK, Betteridge J, Deanfield JE. Impaired vascular reactivity in insulin-dependent diabetes mellitus is related to disease duration and low density lipoprotein cholesterol levels. J Am Coll Cardiol. 1996; 28: 573–579.
  6. Williams SB, Cusco JA, Roddy MA, Johnstone MT, Creager MA. Impaired nitric oxide-mediated vasodilation in patients with non-insulin-dependent diabetes mellitus. J Am Coll Cardiol. 1996; 27: 567–574.
  7. Coutinho M, Gerstein HC, Wang Y, Yusuf S. The relationship between glucose and incident cardiovascular events: a metaregression analysis of published data from 20 studies of 95,783 individuals followed for 12.4 years. Diabetes Care. 1999; 22: 233–240.
  8. Bohlen HG, Lash JM. Topical hyperglycemia rapidly suppresses EDRF-mediated vasodilation of normal rat arterioles. Am J Physiol. 1993; 265: H219–H225.
  9. Williams SB, Goldfine AB, Timimi FK, Ting HH, Roddy MA, Simonson DC, Creager MA. Acute hyperglycemia attenuates endothelium-dependent vasodilation in humans in vivo. Circulation. 1998; 97: 1695–1701.
  10. Cosentino F, Hishikawa K, Katusic ZS, Luscher TF. High glucose increases nitric oxide synthase expression and superoxide anion generation in human aortic endothelial cells. Circulation. 1997; 96: 25–28.
  11. Hink U, Li H, Mollnau H, Oelze M, Matheis E, Hartmann M, Skatchkov M, Thaiss F, Stahl RA, Warnholtz A, Meinertz T, Griendling K, Harrison DG, Forstermann U, Munzel T. Mechanisms underlying endothelial dysfunction in diabetes mellitus. Circ Res. 2001; 88: e14–e22.
  12. Tesfamariam B, Brown ML, Cohen RA. Elevated glucose impairs endothelium-dependent relaxation by activating protein kinase C. J Clin Invest. 1991; 87: 1643–1648.
  13. Nishio E, Watanabe Y. Glucose-induced down-regulation of NO production and inducible NOS expression in cultured rat aortic vascular smooth muscle cells: role of protein kinase C. Biochem Biophys Res Commun. 1996; 229: 857–863.
  14. Ishii H, Jirousek MR, Koya D, Takagi C, Xia P, Clermont A, Bursell SE, Kern TS, Ballas LM, Heath WF, Stramm LE, Feener EP, King GL. Amelioration of vascular dysfunctions in diabetic rats by an oral PKC ß inhibitor. Science. 1996; 272: 728–731.
  15. Mellor H, Parker PJ. The extended protein kinase C superfamily. Biochem J. 1998; 332: 281–292.
  16. Ohara Y, Sayegh HS, Yamin JJ, Harrison DG. Regulation of endothelial constitutive nitric oxide synthase by protein kinase C. Hypertension. 1995; 25: 415–420.
  17. Hirata K, Kuroda R, Sakoda T, Katayama M, Inoue N, Suematsu M, Kawashima S, Yokoyama M. Inhibition of endothelial nitric oxide synthase activity by protein kinase C. Hypertension. 1995; 25: 180–185.
  18. Ohara Y, Peterson TE, Zheng B, Kuo JF, Harrison DG. Lysophosphatidylcholine increases vascular superoxide anion production via protein kinase C activation. Arterioscler Thromb. 1994; 14: 1007–1013.
  19. Inoguchi T, Battan R, Handler E, Sportsman J, Heath W, King G. Preferential elevation of protein kinase C isoform ß II and diacylglycerol levels in the aorta and heart of diabetic rats: differential reversibility to glycemic control by islet cell transplantation. Proc Natl Acad Sci U S A. 1992; 89: 11059–11063.
  20. Kunisaki M, Fumio U, Nawata H, King GL. Vitamin E normalizes diacylglycerol-protein kinase C activation induced by hyperglycemia in rat vascular tissues. Diabetes. 1996; 45: S117–S119.
  21. Jirousek MR, Gillig JR, Gonzalez CM, Heath WF, McDonald JH3rd, Neel DA, Rito CJ, Singh U, Stramm LE, Melikian-Badalian A, Baevsky M, Ballas LM, Hall SE, Winneroski LL, Faul MM. (S)-13-[(dimethylamino)methyl]-10,11,14,15-tetrahydro-4,9:16, 21-dimetheno-1H, 13H-dibenzo[e,k]pyrrolo[3,4-h][1,4,13]oxadiazacyclohexadecene-1,3(2H)-d ione (LY333531) and related analogues: isozyme selective inhibitors of protein kinase C ß. J Med Chem. 1996; 39: 2664–2671.
  22. Pieper GM, Meier DA, Hager SR. Endothelial dysfunction in a model of hyperglycemia and hyperinsulinemia. Am J Physiol. 1995; 269: H845–H850.
  23. Kawano H, Motoyama T, Hirashima O, Hirai N, Miyao Y, Sakamoto T, Kugiyama K, Ogawa H, Yasue H. Hyperglycemia rapidly suppresses flow-mediated endothelium-dependent vasodilation of brachial artery. J Am Coll Cardiol. 1999; 34: 146–154.
  24. Scherrer U, Randin D, Vollenweider P, Vollenweider L, Nicod P. Nitric oxide release accounts for insulin’s vascular effects in humans. J Clin Invest. 1994; 94: 2511–2515.
  25. Steinberg HO, Brechtel G, Johnson A, Fineberg N, Baron AD. Insulin-mediated skeletal muscle vasodilation is nitric oxide dependent: a novel action of insulin to increase nitric oxide release. J Clin Invest. 1994; 94: 1172–1179.
  26. Moller N, Bagger JP, Schmitz O, Jorgensen JO, Ovesen P, Moller J, Alberti KG, Orskov H. Somatostatin enhances insulin-stimulated glucose uptake in the perfused human forearm. J Clin Endocrinol Metab. 1995; 80: 1789–1793.
  27. Beckman JA, Goldfine AB, Gordon MB, Creager MA. Ascorbate restores endothelium-dependent vasodilation impaired by acute hyperglycemia in humans. Circulation. 2001; 103: 1618–1623.
  28. Morigi M, Angioletti S, Imberti B, Donadelli R, Micheletti G, Figliuzzi M, Remuzzi A, Zoja C, Remuzzi G. Leukocyte-endothelial interaction is augmented by high glucose concentrations and hyperglycemia in a NF-{kappa}B-dependent fashion. J Clin Invest. 1998; 101: 1905–1915.
  29. Park JY, Takahara N, Gabriele A, Chou E, Naruse K, Suzuma K, Yamauchi T, Ha SW, Meier M, Rhodes CJ, King GL. Induction of endothelin-1 expression by glucose: an effect of protein kinase C activation. Diabetes. 2000; 49: 1239–1248.
  30. Hempel A, Maasch C, Heintze U, Lindschau C, Dietz R, Luft FC, Haller H. High glucose concentrations increase endothelial cell permeability via activation of protein kinase C{alpha}. Circ Res. 1997; 81: 363–371.
  31. Koya D, King GL. Protein kinase C activation and the development of diabetic complications. Diabetes. 1998; 47: 859–866.
  32. Yasunari K, Kohno M, Kano H, Yokokawa K, Horio T, Yoshikawa J. Possible involvement of phospholipase D and protein kinase C in vascular growth induced by elevated glucose concentration. Hypertension. 1996; 28: 159–168.
  33. Graier WF, Posch K, Wascher TC, Kostner GM. Role of superoxide anions in changes of endothelial vasoactive response during acute hyperglycemia. Horm Metab Res. 1997; 29: 622–626.
  34. Tesfamariam B, Cohen RA. Free radicals mediate endothelial cell dysfunction caused by elevated glucose. Am J Physiol. 1992; 236: H321–H326.
  35. Ungvari Z, Pacher P, Kecskemeti V, Papp G, Szollar L, Koller A. Increased myogenic tone in skeletal muscle arterioles of diabetic rats: possible role of increased activity of smooth muscle Ca2+ channels and protein kinase C. Cardiovasc Res. 1999; 43: 1018–1028.
  36. Liu Y, Terata K, Rusch NJ, Gutterman DD. High glucose impairs voltage-gated K+ channel current in rat small coronary arteries. Circ Res. 2001; 89: 146–152.
  37. Johnstone MT, Creager SJ, Scales KM, Cusco JA, Lee BK, Creager MA. Impaired endothelium-dependent vasodilation in patients with insulin-dependent diabetes mellitus. Circulation. 1993; 88: 2510–2516.
  38. Nugent AG, McGurk C, Hayes JR, Johnston GD. Impaired vasoconstriction to endothelin 1 in patients with NIDDM. Diabetes. 1996; 45: 105–107.
  39. Houben AJ, Schaper NC, de Haan CH, Huvers FC, Slaaf DW, de Leeuw PW, Nieuwenhuijzen Kruseman AC. The effects of 7-hour local hyperglycaemia on forearm macro and microcirculatory blood flow and vascular reactivity in healthy man. Diabetologia. 1994; 37: 750–756.
  40. Lee TS, Saltsman KA, Ohashi H, King GL. Activation of protein kinase C by elevation of glucose concentration: proposal for a mechanism in the development of diabetic vascular complications [published erratum appears in Proc Natl Acad Sci U S A. 1991;88:9907]. Proc Natl Acad Sci U S A. 1989; 86: 5141–5145.
  41. Xia P, Inoguchi T, Kern TS, Engerman RL, Oates PJ, King GL. Characterization of the mechanism for the chronic activation of diacylglycerol-protein kinase C pathway in diabetes and hypergalactosemia. Diabetes. 1994; 43: 1122–1129.
  42. Nishikawa T, Edelstein D, Du XL, Yamagishi S, Matsumura T, Kaneda Y, Yorek MA, Beebe D, Oates PJ, Hammes HP, Giardino I, Brownlee M. Normalizing mitochondrial superoxide production blocks three pathways of hyperglycaemic damage. Nature. 2000; 404: 787–790.
  43. Kuboki K, Jiang ZY, Takahara N, Ha SW, Igarashi M, Yamauchi T, Feener EP, Herbert TP, Rhodes CJ, King GL. Regulation of endothelial constitutive nitric oxide synthase gene expression in endothelial cells and in vivo: a specific vascular action of insulin. Circulation. 2000; 101: 676–681.
  44. Inoguchi T, Li P, Umeda F, Yu HY, Kakimoto M, Imamura M, Aoki T, Etoh T, Hashimoto T, Naruse M, Sano H, Utsumi H, Nawata H. High glucose level and free fatty acid stimulate reactive oxygen species production through protein kinase C–dependent activation of NAD(P)H oxidase in cultured vascular cells. Diabetes. 2000; 49: 1939–1945.
  45. Laursen JB, Somers M, Kurz S, McCann L, Warnholtz A, Freeman BA, Tarpey M, Fukai T, Harrison DG. Endothelial regulation of vasomotion in apoE-deficient mice: implications for interactions between peroxynitrite and tetrahydrobiopterin. Circulation. 2001; 103: 1282–1288.
  46. Muniyappa R, Srinivas PR, Ram JL, Walsh MF, Sowers JR. Calcium and protein kinase C mediate high-glucose-induced inhibition of inducible nitric oxide synthase in vascular smooth muscle cells. Hypertension. 1998; 31: 289–295.
  47. McVeigh GE, Brennan GM, Johnston GD, McDermott BJ, McGrath LT, Henry WR, Andrews JW, Hayes JR. Impaired endothelium-dependent and independent vasodilation in patients with type 2 (non-insulin-dependent) diabetes mellitus. Diabetologia. 1992; 35: 771–776.
  48. Bank AJ, Rector TS, Burke MN, Tschumperlin LK, Kubo SH. Impaired forearm vasodilation to hyperosmolal stimuli in patients with congestive heart failure secondary to idiopathic dilated cardiomyopathy or to ischemic cardiomyopathy. Am J Cardiol. 1992; 70: 1315–1319.



This article has been cited by other articles:


Home page
CirculationHome page
E. Osto, A. Kouroedov, P. Mocharla, A. Akhmedov, C. Besler, L. Rohrer, A. von Eckardstein, S. Iliceto, M. Volpe, T. F. Luscher, et al.
Inhibition of Protein Kinase C{beta} Prevents Foam Cell Formation by Reducing Scavenger Receptor A Expression in Human Macrophages
Circulation, November 18, 2008; 118(21): 2174 - 2182.
[Abstract] [Full Text] [PDF]


Home page
Am. J. Physiol. Heart Circ. Physiol.Home page
A. Goel, D. Thor, L. Anderson, and R. Rahimian
Sexual dimorphism in rabbit aortic endothelial function under acute hyperglycemic conditions and gender-specific responses to acute 17{beta}-estradiol
Am J Physiol Heart Circ Physiol, June 1, 2008; 294(6): H2411 - H2420.
[Abstract] [Full Text] [PDF]


Home page
Cardiovasc ResHome page
A. Goel, Y. Zhang, L. Anderson, and R. Rahimian
Gender difference in rat aorta vasodilation after acute exposure to high glucose: Involvement of protein kinase C {beta} and superoxide but not of Rho Kinase
Cardiovasc Res, November 1, 2007; 76(2): 351 - 360.
[Abstract] [Full Text] [PDF]


Home page
Endocr. Rev.Home page
R. Muniyappa, M. Montagnani, K. K. Koh, and M. J. Quon
Cardiovascular Actions of Insulin
Endocr. Rev., August 1, 2007; 28(5): 463 - 491.
[Abstract] [Full Text] [PDF]


Home page
Am. J. Physiol. Heart Circ. Physiol.Home page
I. F. Benter, M. H. M. Yousif, C. Cojocel, M. Al-Maghrebi, and D. I. Diz
Angiotensin-(1-7) prevents diabetes-induced cardiovascular dysfunction
Am J Physiol Heart Circ Physiol, January 1, 2007; 292(1): H666 - H672.
[Abstract] [Full Text] [PDF]


Home page
Am. J. Physiol. Heart Circ. Physiol.Home page
M. E. J. Lott, C. Hogeman, M. Herr, R. Gabbay, and L. I. Sinoway
Effects of an oral glucose tolerance test on the myogenic response in healthy individuals
Am J Physiol Heart Circ Physiol, January 1, 2007; 292(1): H304 - H310.
[Abstract] [Full Text] [PDF]


Home page
Drug Metab. Dispos.Home page
J. L. Burkey, K. M. Campanale, R. Barbuch, D. O'Bannon, J. Rash, C. Benson, and D. Small
Disposition of [14C]Ruboxistaurin in Humans
Drug Metab. Dispos., November 1, 2006; 34(11): 1909 - 1917.
[Abstract] [Full Text] [PDF]


Home page
J Am Coll CardiolHome page
S.-F. Yan, E. Harja, M. Andrassy, T. Fujita, and A. M. Schmidt
Protein Kinase C {beta}/Early Growth Response-1 Pathway: A Key Player in Ischemia, Atherosclerosis, and Restenosis
J. Am. Coll. Cardiol., October 27, 2006; 48(9_Suppl_A): A47 - A55.
[Abstract] [Full Text] [PDF]


Home page
J. Pharmacol. Exp. Ther.Home page
W. Zhou, X.-L. Wang, K. G. Lamping, and H.-C. Lee
Inhibition of Protein Kinase Cbeta Protects against Diabetes-Induced Impairment in Arachidonic Acid Dilation of Small Coronary Arteries
J. Pharmacol. Exp. Ther., October 1, 2006; 319(1): 199 - 207.
[Abstract] [Full Text] [PDF]


Home page
Arterioscler. Thromb. Vasc. Bio.Home page
I. J. Goldberg and H. M. Dansky
Diabetic Vascular Disease: An Experimental Objective
Arterioscler. Thromb. Vasc. Biol., August 1, 2006; 26(8): 1693 - 1701.
[Abstract] [Full Text] [PDF]


Home page
Am. J. Physiol. Endocrinol. Metab.Home page
M. A. Vincent, L. H. Clerk, J. R. Lindner, W. J. Price, L. A. Jahn, H. Leong-Poi, and E. J. Barrett
Mixed meal and light exercise each recruit muscle capillaries in healthy humans
Am J Physiol Endocrinol Metab, June 1, 2006; 290(6): E1191 - E1197.
[Abstract] [Full Text] [PDF]


Home page
DiabetesHome page
K. Naruse, C. Rask-Madsen, N. Takahara, S.-w. Ha, K. Suzuma, K. J. Way, J. R.C. Jacobs, A. C. Clermont, K. Ueki, Y. Ohshiro, et al.
Activation of Vascular Protein Kinase C-{beta} Inhibits Akt-Dependent Endothelial Nitric Oxide Synthase Function in Obesity-Associated Insulin Resistance
Diabetes, March 1, 2006; 55(3): 691 - 698.
[Abstract] [Full Text] [PDF]


Home page
Circ. Res.Home page
C. Vecchione, A. Aretini, G. Marino, U. Bettarini, R. Poulet, A. Maffei, M. Sbroggio, L. Pastore, M. T. Gentile, A. Notte, et al.
Selective Rac-1 Inhibition Protects From Diabetes-Induced Vascular Injury
Circ. Res., February 3, 2006; 98(2): 218 - 225.
[Abstract] [Full Text] [PDF]


Home page
Diabetes CareHome page
Z. He and G. L. King
Can Protein Kinase C {beta}-Selective Inhibitor, Ruboxistaurin, Stop Vascular Complications in Diabetic Patients?
Diabetes Care, November 1, 2005; 28(11): 2803 - 2805.
[Full Text] [PDF]


Home page
Am. J. Physiol. Heart Circ. Physiol.Home page
A. Phillipson, E. E. Peterman, P. Taormina Jr., M. Harvey, R. J. Brue, N. Atkinson, D. Omiyi, U. Chukwu, and L. H. Young
Protein kinase C-{zeta} inhibition exerts cardioprotective effects in ischemia-reperfusion injury
Am J Physiol Heart Circ Physiol, August 1, 2005; 289(2): H898 - H907.
[Abstract] [Full Text] [PDF]


Home page
J. Am. Soc. Nephrol.Home page
D. J. Kelly, A. Chanty, R. M. Gow, Y. Zhang, and R. E. Gilbert
Protein Kinase C{beta} Inhibition Attenuates Osteopontin Expression, Macrophage Recruitment, and Tubulointerstitial Injury in Advanced Experimental Diabetic Nephropathy
J. Am. Soc. Nephrol., June 1, 2005; 16(6): 1654 - 1660.
[Abstract] [Full Text] [PDF]


Home page
Arterioscler. Thromb. Vasc. Bio.Home page
C. Rask-Madsen and G. L. King
Proatherosclerotic Mechanisms Involving Protein Kinase C in Diabetes and Insulin Resistance
Arterioscler. Thromb. Vasc. Biol., March 1, 2005; 25(3): 487 - 496.
[Abstract] [Full Text] [PDF]


Home page
Vasc MedHome page
D. J Collinson, R. Rea, and R. Donnelly
Masterclass series in peripheral arterial disease: Vascular risk: diabetes
Vascular Medicine, November 1, 2004; 9(4): 307 - 310.
[PDF]


Home page
Arterioscler. Thromb. Vasc. Bio.Home page
T. J. Guzik, J. Sadowski, B. Kapelak, A. Jopek, P. Rudzinski, R. Pillai, R. Korbut, and K. M. Channon
Systemic Regulation of Vascular NAD(P)H Oxidase Activity and Nox Isoform Expression in Human Arteries and Veins
Arterioscler. Thromb. Vasc. Biol., September 1, 2004; 24(9): 1614 - 1620.
[Abstract] [Full Text] [PDF]


Home page
Am. J. Physiol. Renal Physiol.Home page
S. Chu and H. G. Bohlen
High concentration of glucose inhibits glomerular endothelial eNOS through a PKC mechanism
Am J Physiol Renal Physiol, September 1, 2004; 287(3): F384 - F392.
[Abstract] [Full Text] [PDF]


Home page
DiabetesHome page
K. J. Mather, A. Lteif, H. O. Steinberg, and A. D. Baron
Interactions Between Endothelin and Nitric Oxide in the Regulation of Vascular Tone in Obesity and Diabetes
Diabetes, August 1, 2004; 53(8): 2060 - 2066.
[Abstract] [Full Text] [PDF]


Home page
CirculationHome page
Z. He and G. L. King
Protein Kinase C{beta} Isoform Inhibitors: A New Treatment for Diabetic Cardiovascular Diseases
Circulation, July 6, 2004; 110(1): 7 - 9.
[Full Text] [PDF]


Home page
CirculationHome page
A. Kouroedov, M. Eto, H. Joch, M. Volpe, T. F. Luscher, and F. Cosentino
Selective Inhibition of Protein Kinase C{beta}2 Prevents Acute Effects of High Glucose on Vascular Cell Adhesion Molecule-1 Expression in Human Endothelial Cells
Circulation, July 6, 2004; 110(1): 91 - 96.
[Abstract] [Full Text] [PDF]


Home page
Am. J. Physiol. Endocrinol. Metab.Home page
A. S. Reed, N. Charkoudian, A. Vella, P. Shah, R. A. Rizza, and M. J. Joyner
Forearm vascular control during acute hyperglycemia in healthy humans
Am J Physiol Endocrinol Metab, March 1, 2004; 286(3): E472 - E480.
[Abstract] [Full Text]


Home page
Am. J. Physiol. Cell Physiol.Home page
J. H. Tinsley, N. R. Teasdale, and S. Y. Yuan
Involvement of PKC{delta} and PKD in pulmonary microvascular endothelial cell hyperpermeability
Am J Physiol Cell Physiol, January 1, 2004; 286(1): C105 - C111.
[Abstract] [Full Text] [PDF]


Home page
Exp. Biol. Med.Home page
E. R. Werner, A. C.F. Gorren, R. Heller, G. Werner-Felmayer, and B. Mayer
Tetrahydrobiopterin and N