You are on page 1of 7

Nutrition, Metabolism & Cardiovascular Diseases (2006) 16, S39eS45

www.elsevier.com/locate/nmcd

Endothelial dysfunction in type 2


diabetes mellitus
Angelo Avogaro*, Gian Paolo Fadini, Alessandra Gallo,
Elisa Pagnin, Saula de Kreutzenberg

Department of Clinical and Experimental Medicine, Division of Metabolic Diseases,


University of Padova, School of Medicine, Via Giustiniani 2, 35128 Padova, Italy

Received 18 October 2005; received in revised form 20 October 2005; accepted 29 October 2005

KEYWORDS Abstract Aims: Vascular endothelial cells play a major role in maintaining cardio-
Endothelium; vascular homeostasis in health. Diabetes mellitus (DM) substantially impairs the
Diabetes; vasodilating properties of the endothelium and leads to endothelial dysfunction,
Protein kinase C; which can thus be considered the first step in the progression of cardiovascular dis-
Diabetic ease. The aim of the present study is to illustrate possible mechanisms responsible
complications; for endothelial dysfunction in DM.
Cardiovascular disease Data synthesis: We have shown that NADPH oxidase gene expression is increased
in circulating lymphomonocytes from patients with DM, and that this increased
gene expression is dependent upon metabolic control. Hyperglycemia can mediate
its adverse effects through the activation of protein kinase C. We have shown an
increase in membrane-associated PKC beta 2 activity in monocytes from patients
with DM. This activity was reduced by 40% in the euglycemic condition. Finally,
we show a reduction of the circulating endothelial progenitor cells, a subset of
bone marrow-derived endothelial-oriented stem cells, which can give rise to
mature endothelial cells.
Conclusion: Endothelial dysfunction, the initial step of the atherosclerotic process,
is reversible. Thus, major efforts should be made to control not only hyperglycemia
but also the other risk factors for cardiovascular disease, in order to prevent the
onset of all these processes that eventually leads the diabetic patient to premature
death.
ª 2005 Elsevier B.V. All rights reserved.

* Corresponding author. Tel.: þ39 049 8212178; fax: þ39 049 8754179.
E-mail address: angelo.avogaro@unipd.it (A. Avogaro).

0939-4753/$ - see front matter ª 2005 Elsevier B.V. All rights reserved.
doi:10.1016/j.numecd.2005.10.015
S40 A. Avogaro et al.

Introduction (NO-sensitive GC), which is the most important re-


ceptor for the signalling molecule NO. This leads to
Vascular endothelial cells play a major role in the conversion of GTP to cyclic guanosine mono-
maintaining cardiovascular homeostasis in health. phosphate (cGMP) [4]. The NO/cGMP signalling
In addition to providing a physical barrier between cascade is crucial in the cardiovascular system,
the vessel wall and lumen, the endothelium where it controls smooth muscle relaxation, and
secretes a number of mediators that regulate inhibition of platelet aggregation; cyclic nucleo-
platelet aggregation, coagulation, fibrinolysis and tide phosphodiesterases hydrolyze cGMP and thus
vascular tone. The term endothelial dysfunction terminate their action. NO has several important
refers to a condition in which the endothelium effects on the vasculature: first, it maintains basal
loses its physiological properties: the tendency to tone by relaxing vascular smooth muscle cells; it
promote vasodilation, fibrinolysis and antiaggrega- also inhibits platelet adhesion, activation, secre-
tion. Endothelial cells secrete several mediators tion, and aggregation and promotes platelet disag-
that can alternatively mediate either vasoconstric- gregation, in part through a cGMP-dependent
tion, such as endothelin-1 and thromboxane A2, or mechanism. In addition to these effects, endothe-
vasodilation such nitric oxide (NO), prostacyclin lial-derived NO inhibits leukocyte adhesion to the
and endothelium-derived hyperpolarizing factor endothelium and smooth muscle cell migration
(EDHF) [1]. NO is the major contributor to endo- and proliferation [2]: therefore NO is a powerful
thelium-dependent relaxation in conduit arteries inhibitor of the earlier phases of atherosclerosis
whereas the contribution of EDHF predominates and all the mechanisms that lead to neointimal
in smaller resistance vessels. Diabetes mellitus proliferation after vascular injury.
(DM) substantially impairs the vasodilating proper-
ties of the endothelium and leads to endothelial
dysfunction, which can thus be considered the first
Molecular mechanisms underlying
step in the progression of cardiovascular disease
(CVD) [2]. endothelial dysfunction in
diabetes mellitus

Oxidative reactions are crucial in all the events


Nitric oxide synthesis and that lead to atherogenesis, including endothelial
biological effects dysfunction. In endothelium exposed to agents
that damage the vasculature there is stimulation
NO, which is the most powerful vasodilating of several enzymes that can produce reactive
compound, derives from L-arginine: the reaction oxygen species (ROS): the enzymes of the mito-
is catalyzed by a family of enzymes, called the chondrial electron transport chain, xanthine
NO synthases (NOS). Three NOS isoforms have oxidase, cyclooxygenases, lipoxygenases, myelo-
been identified: endothelial NOS (eNOS), neuronal peroxidases, cytochrome P450 monooxygenase,
NOS (nNOS) and inducible or inflammatory NOS uncoupled NOS, heme oxygenases, peroxidases,
(iNOS). In endothelial cells, eNOS generates NO and NAD(P)H oxidases. In turn, ROS can be pro-
only if one essential cofactor such as tetrahydro- duced intracellularly, extracellularly, or in specific
biopterin (BH4) is optimally present within the intracellular compartments. Among these en-
cells. Enzyme activity can be regulated by post- zymes, nicotinamide adenine dinucleotide/NADPH
translational modifications: these modifications oxidase is relevant because it is a major vascular
occur through the phosphorylation of Ser1179, source of ROS [5]. There is evidence that strong
which increases the activity of the enzyme [1]. correlations exist among NADPH oxidase activity,
Several kinases can phosphorylate this site, includ- atherosclerotic risk factors, and endothelial
ing protein kinase A, protein kinase C, and serine/ dysfunction. We have recently shown that NADPH
threonine kinase Akt/PKB. On the contrary eNOS oxidase gene expression is increased in circulating
activity can be downregulated by other proteins, lymphomonocytes from patients with diabetes
including caveolin-1, the major coating protein of mellitus (DM), and that this increased gene expres-
caveolae, vesicles found throughout endothelial sion is dependent upon metabolic control [6]. In-
cells, which are implicated not only in signal trans- terestingly, NADPH activity is increased not only
duction but also in endothelial barrier function and by factors that damage vascular endothelium,
mechanotransduction [3]. In order to induce but possibly by insulin itself [7,8]. ROS generation
vasodilation in the vascular smooth muscle cells, is enhanced in the blood vessels of hypertensive
NO must activate NO-sensitive guanylyl cyclase animal models and in atherosclerotic lesions in
Endothelium and diabetes S41

humans and animals: this process appears to pro- interface of blood and tissue, which maintains
mote vascular proliferation, hypertrophy and a balance between these two compartments.
remodelling, and is involved in the progression of With disruption of this balance, mediated by in-
atherosclerosis. Insulin resistance, which most flammation, the vascular wall becomes susceptible
often precedes the onset of type 2 diabetes, and to atheroma formation [16]. In response to DM,
increased FFA levels cause oxidative stress due blood leukocytes attach to the endothelium where
to increased mitochondrial uncoupling. Moreover, there is an exaggerated production of ROS [17]. In
oxidative stress plays a key role in the pathogene- their turn, ROS induce several inflammatory cyto-
sis of late diabetic complications. Hyperglycemia kines such that systemic oxidative stress also
leads to mitochondrial dysfunction and activation means systemic inflammation. Accumulating evi-
of stress pathways. Both in vitro and in vivo studies dence suggests a major link between C reactive
reveal that oxidative stress due to hyperglycemia protein (CRP) and atherosclerosis. CRP, which is
occurs before late complications become clinically produced in the liver in response to pro-inflamma-
evident. This indicates that oxidative stress plays tory states, potently downregulates eNOS tran-
a crucial role in the pathogenesis of late diabetic scription and destabilizes eNOS mRNA, with
complications [9]. resultant decreases in both basal and stimulated
Hyperglycemia also leads to generation of ad- NO release [18]. CRP also stimulates endothelin-1
vanced glycation endproducts (AGE), the products and interleukin-6 release, upregulates adhesion
of non-enzymatic glycation of proteins and lipids molecules, and stimulates MCP-1. Thus, CRP is
that accumulate in the vessel wall. They are signal not only a marker of endothelial dysfunction/
transduction ligands for the receptor for AGE cardiovascular risk [19,20] but also takes part in
(RAGE) [10]. AGEs accumulate in the vessel wall its pathogenesis [21]. The inflammatory response
exposed to hyperglycemia and alter the structural involved in the initiation and progression of ath-
integrity of the vessel wall and underlying base- erosclerotic lesions involves many mediators that
ment membrane and are able to quench nitric activate NF-kB, which has specific target sequen-
oxide. This contributes substantially to negatively ces on many immune and inflammatory gene
affect endothelial function [11]. promoters [22].
Hyperglycemia is one of the major causal There is general agreement that hyperglycemia
factors in the development of diabetic vascular impairs endothelium-dependent vasodilation in
complications and can mediate their adverse vivo, in humans [23e27]. Interestingly, even in
effects through multiple pathways. One of those normoglycemic subjects who are prone to develop
mechanisms is the activation of PKC by hypergly- DM and insulin resistance syndrome, impaired en-
cemia-induced increases in diacylglycerol (DAG) dothelial function has been observed during an
level, partly due to de novo synthesis [12]. There is oral glucose tolerance test [28]. This has led to
increasing evidence that PKC activation is impor- the hypothesis that endothelial dysfunction may
tant in diabetes-related endothelial dysfunction; precede the development of overt DM, and that
impaired NO-vasodilation and increased ET-1 re- a prolonged and repeated exposure to postprandial
lease involved PKC-mediated inhibition of eNOS hyperglycemia may play an important role in the
[11]. We have shown an increase in membrane- development of atherosclerosis, even in those
associated PKC beta 2 activity in monocytes from who have normal fasting plasma glucose levels.
patients with DM; this activity was reduced by With endothelial function being defined as the
40% in the euglycemic condition [13]. PKC activa- blood flow response to acetylcholine, insulin resis-
tion also mediates the overexpression of adhesion tance itself is characterized by endothelial dys-
molecules such as ICAM, VCAM and E-selectin. PKC function [29]. In DM, endothelial dysfunction
also play a crucial role in mediating vascular appears a consistent although not widespread
smooth muscle cell contractility thus mediating finding; our previous finding of a normal vascular
vascular vasoconstriction in response to risk fac- response to acetylcholine in DM without other
tors such as hyperglycemia. known risk factors for CHD suggests that the endo-
Atherosclerosis is also regarded as a progressive thelial dysfunction in DM cannot be attributed to
disease arising from the combination of endothe- a single, but possibly, to a variety of factors such
lial dysfunction and inflammation. Indeed, inflam- as the diameter of LDL particles, total cholesterol,
mation has been recognized as a keypoint in the leukocyte count, von Willebrand factor and serum
cluster of the insulin resistance syndrome, includ- AGEs [30]. However, there is general agreement
ing diabetes, obesity, hypertension and hyperlipe- that hyperglycemia and DM lead to an impairment
mia [14,15]. Much of the inflammatory reactions of NO production and activity. We have recently
are mediated by the endothelium, located at the shown that the fraction of L-arginine converted
S42 A. Avogaro et al.

to NO is lower in DM patients than in normal sub- amputation. Thus, emerging evidence indicating
jects using a stable isotope approach [31]. It has that bone marrow-derived EPCs participate in
also been shown that an inappropriate production postnatal neovascularization led to the hypothesis
of asymmetric dimethylarginine (ADMA) can alter that alterations in EPC number or function could
endothelial function in patients with DM: this be involved in the pathogenesis of vascular com-
endogenous competitor for NOS increases after plications in DM. Lambiase et al. have shown that
a fatty meal in DM and its circulating levels corre- poor coronary collateral development, which is
late with a reduction of vascular function [32]. typical of DM, is related to low levels of circulating
EPCs [37].
A possible role for EPCs in diabetic vascular
disease was first investigated in diabetic mice with
The role of circulating endothelial experimental hindlimb ischemia, in which infusion
progenitor cells in DM of human CD34-positive leukocytes was able to
accelerate blood flow restoration [38]. Tepper
Circulating endothelial progenitor cells (EPCs) are et al. showed that PBMC-derived EPCs isolated
considered a subset of bone marrow-derived en- from patients with DM displayed a proliferation
dothelial-oriented stem cells, expressing both rate in culture decreased by 48% compared to con-
hematopoietic immature surface markers, such trol subjects, a weaker adherence to activated
as CD34 and CD133, and endothelial lineage human umbilical vein endothelial cells (HUVEC)
markers [33]. EPCs give rise to mature endothelial and a 2.5-fold reduced incorporation into vascular
cells, as demonstrated in pioneer in vitro studies structures in vitro [39]. The rate of EPC prolifera-
and recently confirmed in vivo. EPCs originally tion from plated PBMCs in patients with DM was
reside in the bone marrow and are mobilized to inversely correlated with the levels of glycated
peripheral blood by a number of stimuli, including hemoglobin, suggesting a possible relation be-
tissue ischemia, physical exercise, cytokines and tween glucose control and EPC function. Almost
certain drugs provided with pleiotropic cardiovas- identical results have been reported by Loomans
cular effects [34]. Currently, it is suggested that in type 1 diabetic patients [40]. EPC reduction in
increase in EPCs may be one of the mechanisms diabetes has been hailed as a novel concept in
by which these stimuli would ameliorate vascular the pathogenesis of macrovascular complications.
function: once in the bloodstream, EPCs could We have recently demonstrated that patients
help to maintain endothelial integrity and stimu- with DM and peripheral arterial disease have a pro-
late angiogenesis in ischemic organs. found reduction of circulating EPCs in comparison
On the contrary, reduction in circulating EPCs with controls, with patients with vascular disease
has been demonstrated in the presence of risk but without DM and with patients with DM but
factors for atherosclerosis, endothelial dysfunction, without vascular disease, thus strengthening the
hypercholesterolemia, aging, smoking, chronic re- hypothesis that EPC reduction may have a role in
nal failure, coronary artery disease, cerebral the pathogenesis of vascular disease in DM. We
vascular disease and DM [35]. Low levels of circu- also confirmed the negative relation between
lating EPCs may be associated with both endothe- EPC levels and the number of risk factors and
lial dysfunction and poor collateral development. showed a strong positive correlation between
Indeed, the concept that EPCs are involved both EPCs from patients with limb ischemia and the
in early and advanced events of the development ankle-brachial index [41].
of cardiovascular diseases is of crucial importance
for diabetology since DM is widely associated with
endothelial dysfunction as well as with poor new
vessel generation [36]. Antidiabetic therapy improves
In diabetes, atherosclerotic vascular disease is endothelial function
characterized by high prevalence, early develop-
ment and rapid progression. The severity of macro- Glycemic control remains the major intervention
vascular complications in DM seems to be due to for prevention of both micro- and macrovascular
profound impaired collateralization of vascular disease. However, in the UKPDS study, the 12%
ischemic beds, but mechanisms that hinder reduction in all diabetes-related endpoints
ischemia-induced neovascularization remain elu- achieved by intensive insulin treatment was mainly
sive. In DM collateralization results are insufficient due to a 25% decrease in microvascular complica-
to overcome the loss of blood flow through occluded tions, while myocardial infarction showed a non-
arteries, leading to ischemia, and often to limb statistical 16% reduction [42]. Stronger predictors
Endothelium and diabetes S43

of macrovascular complications were factors such On the contrary, insulin has also been shown to
as LDL cholesterol and blood pressure, that may exert pro-atherogenic effects. In fact, chronically
be related more to insulin resistance than to glyce- elevated insulin levels, such those reached in the
mic control. setting of insulin resistance and when the hormone
Nonetheless, the correction of both hypergly- is chronically administered, force insulin to act as
cemia and insulin resistance improves endothelial a growth factor through the MAP kinase pathway,
function. Although weight loss ameliorates insulin thus promoting the atherogenetic process [52].
sensitivity, a parallel sharp improvement in endo-
thelial function has not been observed. However,
weight loss decreases serum levels of inflammatory
markers and enhances vasodilatation in response References
to L-arginine [43].
Oral agents that improve insulin sensitivity can [1] Vallance P. Nitric oxide. Biologist (London) 2001;48(4):
153e8.
ameliorate vascular function: metformin has been [2] Anderson TJ. Nitric oxide, atherosclerosis and the clinical
reported to improve endothelial function in pa- relevance of endothelial dysfunction. Heart Fail Rev
tients with type DM in one placebo-controlled trial 2003;8(1):71e86.
[44], while, in another study, 8 weeks of treatment [3] Gratton JP, Bernatchez P, Sessa WC. Caveolae and caveo-
with glibenclamide, metformin and glimepiride all lins in the cardiovascular system. Circ Res 2004 Jun 11;
94(11):1408e17.
had similar effects on endothelial function [45]. [4] Munzel T, Feil R, Mulsch A, Lohmann SM, Hofmann F,
We have recently showed that this positive effect Walter U. Physiology and pathophysiology of vascular
of metformin on endothelial function may reside signaling controlled by guanosine 3#, 5#-cyclic mono-
in its capability to inhibit the PKC b2 activation phosphate-dependent protein kinase. Circulation 2003;
in endothelial cells exposed to high glucose con- 108(18):2172e83.
[5] Griendling KK, FitzGerald GA. Oxidative stress and cardio-
centration [46]. vascular injury: part I: basic mechanisms and in vivo mon-
The thiazolidinediones seem to be promising itoring of ROS. Circulation 2003;108(16):1912e6.
drugs to correct endothelial dysfunction: they are [6] Avogaro A, Pagnin E, Calo L. Monocyte NADPH oxidase
provided with pleiotropic actions on the cardio- subunit p22(phox) and inducible hemeoxygenase-1 gene
vascular system, such as anti-hypertensive, anti- expressions are increased in type II diabetic patients: rela-
tionship with oxidative stress. J Clin Endocrinol Metab
thrombotic and anti-inflammatory effects. The 2003;88(4):1753e9.
anti-inflammatory capacity derives from de- [7] Goldstein BJ, Kalyankar M, Wu X. Redox paradox. Insulin
creased ROS generation by mononuclear cells and action is facilitated by insulin-stimulated reactive oxygen
to reduce lipid peroxidation [47]. These effects are species with multiple potential signaling targets. Diabetes
observed in obese individuals both with and with- 2005;54:311e21.
[8] Ceolotto G, Bevilacqua M, Papparella I, Baritono E,
out DM. Another mechanism that may mediate Franco L, Corvaja C, et al. Insulin generates free radicals
the improvement of vascular reactivity by TZDs is by an NAD(P)H, phosphatidylinositol 3#-kinase-dependent
suppression of FFAs through inhibition of lipolysis mechanism in human skin fibroblasts ex vivo. Diabetes
in adipose tissue. 2004;53(5):1344e51.
Finally, insulin therapy has been demonstrated [9] Evans JL, Goldfine ID, Maddux BA, Grodsky GM. Oxidative
stress and stress-activated signaling pathways: a unifying
to improve or reverse vascular dysfunction as hypothesis of type 2 diabetes. Endocr Rev 2002;23(5):
extensively shown by the group of Yki-Jarvinen 599e622.
[48,49]. [10] Bucala R, Tracey KJ, Cerami A. Advanced glycosylation
Both insulin deficiency and insulin resistance are products quench nitric oxide and mediate defective
associated with endothelial dysfunction. Indeed, endothelium-dependent vasodilatation in experimental
diabetes. J Clin Invest 1991;87(2):432e8.
insulin is a weak vasodilator and emerging data [11] Sheetz MJ, King GL. Molecular understanding of hypergly-
support the notion that metabolic-vascular cou- cemia’s adverse effects for diabetic complications. JAMA
pling is fundamental for physiological insulin 2002;288(20):2579e88.
action. Anti-atherogenic effects of insulin are [12] Cosentino F, Eto M, De Paolis P, van der Loo B,
believed to originate not only from correction of Bachschmid M, Ullrich V, et al. High glucose causes upregu-
lation of cyclooxygenase-2 and alters prostanoid profile in
hyperglycemia and its negative vascular conse- human endothelial cells: role of protein kinase C and reac-
quences, but also from direct anti-inflammatory tive oxygen species. Circulation 2003;107(7):1017e23.
effects. For example, insulin therapy suppresses [13] Ceolotto G, Gallo A, Miola M, Sartori M, Trevisan R, Del
NF-kB and reduces inflammatory markers in acute Prato S, et al. Protein kinase C activity is acutely regulated
myocardial infarction [50,51]. These favorable in- by plasma glucose concentration in human monocytes in
vivo. Diabetes 1999;48(6):1316e22.
sulin pathways, which are mediated by the activa- [14] Dandona P, Aljada A, Bandyopadhyay A. Inflammation: the
tion of PI3K/Akt, act on glucose, lipid and protein link between insulin resistance, obesity and diabetes.
metabolism as well as on vascular biology. Trends Immunol 2004;25(1):4e7.
S44 A. Avogaro et al.

[15] Haffner SM. Insulin resistance, inflammation, and the pre- a high-fat meal in patients with type 2 diabetes. Arterios-
diabetic state. Am J Cardiol 2003;92(4A):18Je26J. cler Thromb Vasc Biol 2000 Sep;20(9):2039e44.
[16] Hansson GK, Libby P, Schonbeck U, Yan ZQ. Innate and [33] Asahara T, Murohara T, Sullivan A, et al. Isolation of puta-
adaptive immunity in the pathogenesis of atherosclerosis. tive progenitor endothelial cells for angiogenesis. Science
Circ Res 2002;91(4):281e91. 1997;275:964e7.
[17] Libby P. Inflammation in atherosclerosis. Nature 2002 Dec [34] Aicher A, Zeiher AM, Dimmeler S. Mobilizing endothelial
19e26;420(6917):868e74. progenitor cells. Hypertension 2005;45(3):321e5.
[18] Ratnam S, Mookerjea S. The regulation of superoxide gen- [35] Urbich C, Dimmeler S. Endothelial progenitor cells: charac-
eration and nitric oxide synthesis by C-reactive protein. terization and role in vascular biology. Circ Res 2004;95(4):
Immunology 1998;94(4):560e8. 343e53.
[19] Ridker PM, Wilson PW, Grundy SM. Should C-reactive pro- [36] Fadini GP, Agostini C, Avogaro A. Endothelial progenitor
tein be added to metabolic syndrome and to assessment cells and vascular biology in diabetes mellitus. Current
of global cardiovascular risk? Circulation 2004;109(23): knowledge and future perspectives. Curr Diab Rev 2005;
2818e25. 1(1):41.
[20] Pradhan AD, Manson JE, Rifai N, Buring JE, Ridker PM. [37] Lambiase PD, Edwards RJ, Anthopoulos P, Rahman S,
C-reactive protein, interleukin 6, and risk of developing Meng G, Clifford AB, et al. Circulating humoral factors
type 2 diabetes mellitus. JAMA 2001 Aug;286(3):327e34. and endothelial progenitor cells in patients with differing
[21] Verma S, Yeh ET. C-reactive protein and atherothrombosis- coronary collateral support. Circulation 2004;109:2993e9.
beyond a biomarker: an actual partaker of lesion forma- [38] Schatteman GC, Hanlon HD, Jiao C, Dodds SG, Christy BA.
tion. Am J Physiol Regul Integr Comp Physiol 2003;285(5): Blood-derived angioblasts accelerate blood-flow restora-
R1253e6. tion in diabetic mice. J Clin Invest 2000;106(4):571e8.
[22] Valen G, Yan ZQ, Hansson GK. Nuclear factor kappa-B and [39] Tepper OM, Galiano RD, Capla JM, Kalka C, Gagne PJ,
the heart. J Am Coll Cardiol 2001 Aug;38(2):307e14. Jacobowitz GR, et al. Human endothelial progenitor cells
[23] McVeigh GE, Brennan GM, Johnston GD, McDermott BJ, from type II diabetics exhibit impaired proliferation, adhe-
McGrath LT, Henry WR, et al. Impaired endothelium- sion, and incorporation into vascular structures. Circula-
dependent and independent vasodilation in patients with tion 2002;106(22):2781e6.
type 2 (non-insulin-dependent) diabetes mellitus. Diabeto- [40] Loomans CJ, de Koning EJ, Staal FJ, Rookmaaker MB,
logia 1992 Aug;35(8):771e6. Verseyden C, de Boer HC, et al. Endothelial progenitor
[24] Williams SB, Cusco JA, Roddy MA, Johnstone MT, cell dysfunction: a novel concept in the pathogenesis of
Creager MA. Impaired nitric oxide-mediated vasodilation vascular complications of type 1 diabetes. Diabetes 2004
in patients with non-insulin-dependent diabetes mellitus. Jan;53(1):195e9.
J Am Coll Cardiol 1996 Mar 1;27(3):567e74. [41] Fadini GP, Miorin M, Facco M, Bonamico S, Baesso I,
[25] Makimattila S, Liu ML, Vakkilainen J, Schlenzka A, Grego F, et al. Circulating endothelial progenitor cells
Lahdenpera S, Syvanne M, et al. Impaired endothelium- are reduced in peripheral vascular complications of
dependent vasodilation in type 2 diabetes. Relation to type 2 diabetes mellitus. J Am Coll Cardiol 2005;45(9):
LDL size, oxidized LDL, and antioxidants. Diabetes Care 1449e57.
1999 Jun;22(6):973e81. [42] Intensive blood-glucose control with sulphonylureas or in-
[26] Hogikyan RV, Galecki AT, Pitt B, Halter JB, Greene DA, sulin compared with conventional treatment and risk of
Supiano MA. Specific impairment of endothelium-depen- complications in patients with type 2 diabetes (UKPDS 33).
dent vasodilation in subjects with type 2 diabetes indepen- UK Prospective Diabetes Study (UKPDS) Group. Lancet
dent of obesity. J Clin Endocrinol Metab 1998 Jun;83(6): 1998;352:837e53.
1946e52. [43] Heilbronn LK, Noakes M, Clifton PM. Energy restriction and
[27] Caballero AE, Arora S, Saouaf R, Lim SC, Smakowski P, weight loss on very-low-fat diets reduce C-reactive protein
Park JY, et al. Microvascular and macrovascular reactivity concentrations in obese, healthy women. Arterioscler
is reduced in subjects at risk for type 2 diabetes. Diabetes Thromb Vasc Biol 2001 Jun;21(6):968e70.
1999 Sep;48(9):1856e62. [44] Mather KJ, Verma S, Anderson TJ. Improved endothelial
[28] Kawano H, Motoyama T, Hirashima O, Hirai N, Miyao Y, function with metformin in type 2 diabetes mellitus.
Sakamoto T, et al. Hyperglycemia rapidly suppresses J Am Coll Cardiol 2001 Apr;37(5):1344e50.
flow-mediated endothelium-dependent vasodilation of [45] Dhindsa P, Davis KR, Donnelly R. Comparison of the micro-
brachial artery. J Am Coll Cardiol 1999 Jun;34(1):146e54. and macro-vascular effects of glimepiride and gliclazide
[29] Steinberg HO, Chaker H, Leaming R, Johnson A, in metformin-treated patients with type 2 diabetes: a
Brechtel G, Baron AD. Obesity/insulin resistance is associ- double-blind, crossover study. Br J Clin Pharmacol 2003
ated with endothelial dysfunction. Implications for the Jun;55(6):616e9.
syndrome of insulin resistance. J Clin Invest 1996 Jun 1; [46] Gallo A, Ceolotto G, Pinton P, Iori E, Murphy E, Rutter GA,
97(11):2601e10. et al. Metformin prevents glucose-induced protein kina-
[30] Avogaro A, Piarulli F, Valerio A, Miola M, Calveri M, Pavan P, seC-b2 activation in human umbilical vein endothelial cells
et al. Forearm nitric oxide balance, vascular relaxation, Through an antioxidant mechanism. Diabetes 2005;54(4):
and glucose metabolism in NIDDM patients. Diabetes 1997 1123e31.
Jun;46(6):1040e6. [47] Dandona P. Insulin resistance and endothelial dysfunction
[31] Avogaro A, Toffolo G, Kiwanuka E, de Kreutzenberg SV, in atherosclerosis: implications and interventions. Diabe-
Tessari P, Cobelli C. L-arginine-nitric oxide kinetics tes Technol Ther 2002;4(6):809e15.
in normal and type 2 diabetic subjects: a stable- [48] Vehkavaara S, Yki-Jarvinen H. 3.5 years of insulin therapy
labelled 15N arginine approach. Diabetes 2003 Mar;52(3): with insulin glargine improves in vivo endothelial function
795e802. in type 2 diabetes. Arterioscler Thromb Vasc Biol 2004 Feb;
[32] Fard A, Tuck CH, Donis JA, Sciacca R, Di Tullio MR, Wu HD, 24(2):325e30.
et al. Acute elevations of plasma asymmetric dimethylargi- [49] Vehkavaara S, Makimattila S, Schlenzka A, Vakkilainen J,
nine and impaired endothelial function in response to Westerbacka J, Yki-Jarvinen H. Insulin therapy improves
Endothelium and diabetes S45

endothelial function in type 2 diabetes. Arterioscler kappaB and stimulates IkappaB in mononuclear cells in
Thromb Vasc Biol 2000 Feb;20(2):545e50. obese subjects: evidence for an anti-inflammatory effect?
[50] Chaudhuri A, Janicke D, Wilson MF, Tripathy D, Garg R, J Clin Endocrinol Metab 2001;86:3257e65.
Bandyopadhyay A, et al. Anti-inflammatory and profibrino- [52] Cusi K, Maezono K, Osman A, Pendergrass M, Patti ME,
lytic effect of insulin in acute ST-segment-elevation myo- Pratipanawatr T, et al. Insulin resistance differentially
cardial infarction. Circulation 2004;109:849e54. affects the PI 3-kinase- and MAP kinase-mediated
[51] Dandona P, Aljada A, Mohanty P, Ghanim H, Hamouda W, signaling in human muscle. J Clin Invest 2000;105:
Assian E, et al. Insulin inhibits intranuclear nuclear factor 311e20.