You are on page 1of 17

Clinical Science (2005) 109, 143–159 (Printed in Great Britain









Vascular complications in diabetes mellitus: the role of endothelial dysfunction
Casper G. SCHALKWIJK∗ † and Coen D. A. STEHOUWER∗ †

Department of Internal Medicine, Academic Hospital Maastricht, 6202 AZ Maastricht, The Netherlands, and †Cardiovascular Research Institute Maastricht, University of Maastricht, 6200 MD Maastricht, The Netherlands









The endothelium is a complex organ with a multitude of properties essential for control of vascular functions. Dysfunction of the vascular endothelium is regarded as an important factor in the pathogenesis of diabetic micro- and macro-angiopathy. Endothelial dysfunction in Type I and II diabetes complicated by micro- or macro-albuminuria is generalized in that it affects many aspects of endothelial function and occurs not only in the kidney. The close linkage between microalbuminuria and endothelial dysfunction in diabetes is an attractive explanation for the fact that microalbuminuria is a risk marker for atherothrombosis. In Type I diabetes, endothelial dysfunction precedes and may cause diabetic microangiopathy, but it is not clear whether endothelial dysfunction is a feature of the diabetic state itself. In Type II diabetes, endothelial function is impaired from the onset of the disease and is strongly related to adverse outcomes. It is not clear whether impaired endothelial function is caused by hyperglycaemia or by other factors. Impaired endothelial function is closely associated with and may contribute to insulin resistance regardless of the presence of diabetes. Endothelial dysfunction in diabetes originates from three main sources. Hyperglycaemia and its immediate biochemical sequelae directly alter endothelial function or influence endothelial cell functioning indirectly by the synthesis of growth factors, cytokines and vasoactive agents in other cells. Finally, the components of the metabolic syndrome can impair endothelial function.

Diabetes mellitus is a common metabolic disease worldwide affecting approx. 150 million people in 2000, which is predicted to rise to 220 million in 2010 [1]. Diabetes and its associated complications have become a public health

problem of considerable magnitude. Cardiovascular disease causes most of the excess morbidity and mortality in diabetes mellitus. Adults with diabetes are at a 2- to 4-fold increased risk of cardiovascular events relative to those without diabetes [2]. Cardiovascular disease accounts for up to 80 % of premature excess mortality in diabetic

Key words: diabetes mellitus, endothelial dysfunction, hyperglycaemia, insulin resistance, metabolic syndrome, oxidative stress, vascular complication. Abbreviations: AGE, advanced glycosylation end-product; Ang II, angiotensin II; CRP, C-reactive protein; DAG, diacylglycerol; eNOS, endothelial NOS; ET-1, endothelin-1; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; GlcNAc, N-acetylglucosamine; HDL, high-density lipoprotein; IGF-1, insulin-like growth factor-1; IL, interleukin; LDL, low-density lipoprotein; LPL, lipoprotein lipase; MAPK, mitogen-activated protein kinase; MCP-1, monocyte chemotactic protein-1; NEFA, non-esterified fatty acid; NF-κB, nuclear factor κB; NO, nitric oxide; NOS, NO synthase; PAI-1, plasminogen activator inhibitor-1; PKC, protein kinase C; ROS, reactive oxygen species; TGF-β, transforming growth factor-β; TNF-α, tumour necrosis factor-α; t-PA, tissue-type plasminogen activator; VCAM-1, vascular cell adhesion molecule-1; VEGF, vascular endothelial growth factor; vWF, von Willebrand factor. Correspondence: Dr Casper G. Schalkwijk, Department of Internal Medicine, Maastricht University, Debeyelaan 25, 6202 AZ Maastricht, The Netherlands (email


2005 The Biochemical Society

smoking. dyslipidaemia. vWF (von Willebrand factor). they may differ according to the nature of the injury and may depend on the intrinsic properties of the endothelium (e. dyslipidaemia. hypercholesterolaemia. renal mesangial cells and large artery macrophages. hypertension. Risk factors such as hypercholesterolaemia. Furthermore. venous versus arterial versus microvascular endothelium). manifested by a low-grade inflammatory process and endothelial dysfunction [10].144 C. G.and macro-angiopathy. such as age. The endothelium actively regulates vascular tone and permeability. Macroangiopathy in diabetes consists mainly of an accelerated form of atherosclerosis and affects the coronary. the composition of the subendothelial matrix. Endothelial dysfunction is thought to play an important role not only in the initiation of atherosclerosis. retinal pericytes. hyperglycaemia. obesity and hyperhomocysteinaemia. limits activation of the coagulation cascade by the thrombomodulin/protein C. dyslipidaemia. the balance between coagulation and fibrinolysis. including NO (nitric oxide). Dysfunction of the vascular endothelium is regarded as an important factor in the pathogenesis of micro. such as vascular smooth muscle cells. have changed in a way that is inappropriate with regard to the preservation of organ function. and inflammatory activity in the vessel wall. Retinal and renal microangiopathy cause diabetic retinopathy and nephropathy and microangiopathy of the vasa nervorum is important in the pathogenesis of neuropathy. hypercholesterolaemia. stroke and diabetic foot disease [4–7]. Large clinical trials in both Type I and Type II diabetes have demonstrated that hyperglycaemia plays an important role in the pathogenesis of microvascular complications [8. A. Among important molecules synthesized by endothelial cells is NO. the endothelium actively decreases vascular tone. the adhesion and extravasation of leucocytes. D. Injury to the endothelium can cause endothelial dysfunction. adhesion molecules and cytokines. which serves as an important locus of control of vascular and thus organ functions [13]. however. obesity and hyperhomocysteinaemia are additional major causes of microangiopathy. inhibits platelet adhesion and aggregation by producing prostacyclin and NO. cause of clinically important microangiopathy. effects that are at least in part attributable to inhibition of the transcription factor NF-κB (nuclear factor κB) [17]. anti-proliferative. Normally. hyperinsulinaemia and impaired fibrinolysis cluster in the metabolic syndrome. insulin resistance. platelets. Vascular complications can be caused by micro. smoking and diabetes initiate atherosclerosis through endothelial activation and therefore endothelial dysfunction. On the other hand. PAI-1 (plasminogen activator inhibitor-1).12] and endothelial function has gained increasing attention in the study of vascular disease. these cells can affect endothelial cells. ENDOTHELIAL FUNCTION AND DYSFUNCTION The endothelium is the biological active inner layer of the blood vessels. NO inhibits leucocyte adhesion and rolling as well as cytokine-induced expression of VCAM-1 (vascular cell adhesion molecule-1) and MCP-1 (monocyte chemotactic protein-1) [16]. which is a particularly important endothelium-derived mediator. but also in its progression and clinical sequelae. but not a sufficient. leucocytes. Dysfunction of the endothelium can be considered present when its properties. The cardiovascular risk factors hypertension. Although diabetic patients with the most severe hyperglycaemia have the highest risk of microangiopathy. Endothelial activation designates one specific set of dysfunctions characterized by increased interactions with blood leucocytes in which adhesion molecules and chemoattractants such as MCP-1 and IL (interleukin)-8 are essential. smoking. because of its vasodilator. and regulates fibrinolysis by producing t-PA and its inhibitor PAI-1. ET-1 (endothelin-1). permeability-decreasing and anti-inflammatory properties [15]. Because of the huge premature morbidity and mortality associated with this disease. The production of these moieties is responsive to various stimuli [14]. but is related to general risk factors for atherothrombosis. All of the above-mentioned factors create a state of constant and progressive damage to the vascular wall. Schalkwijk and C. Ang II (angiotensin II). anti-platelet. maintains vascular permeability to nutrients.and macro-angiopathy [11. hormones. prevention of complications is a key issue. carotid and peripheral arteries. Such alterations do not necessarily occur simultaneously. These C . the endothelium produces components of the extracellular matrix such as collagen and a variety of regulatory mediators. This review will explore how endothelial dysfunction in diabetes relates to the pattern of disease occurrence described above and what current biochemical mechanisms have been proposed as an explanation for the development of endothelial dysfunction in diabetes. Hypertension. obesity. Therefore endothelial activation can be conceptualized as a transducer of atherogenic risk factors. dyslipidaemia.9]. is a necessary. limits leucocyte adhesion and thus inflammatory activity in the vessel wall. Stehouwer patients [3]. heparan sulphate/antithrombin and tissue factor/tissue factor pathway inhibitor interactions. It also affects the 2005 The Biochemical Society functions of other cell types. To carry out its above-mentioned functions. either in the basal state or after stimulation. thus increasing the risk of myocardial infarction. Risk of macroangiopathy does not appear to be strongly related to hyperglycaemia. prostanoids. t-PA (tissue-type plasminogen activator).g. other macromolecules and leucocytes.

high LDL (low-density lipoprotein)-cholesterol increases the production of ROS (reactive oxygen species) and thus NO scavenging. pre-eclampsia and vasculitis. and thus decreases NO availability. tests intended to estimate NOmediated endothelium-dependent vasodilation in part measure the effects of other endothelial vasodilators such as prostacyclin and EDHF (endothelium-derived hyperpolarizing factor). the assumptions are clearly invalid. MEASUREMENT OF ENDOTHELIAL DYSFUNCTION Endothelial function cannot be measured directly in humans. On the one hand. In this regard. Thirdly. Secondly. In some cases. such as atherothrombosis. possibly. some questions can be raised about the validity of the tests listed in Table 1. many crosssectional studies have shown impaired endothelium-dependent vasodilation and high levels of endotheliumderived regulatory proteins in patients with diseases that involve injury to the endothelium. but it is not known to what extent. as well as in individuals with risk factors for atherothrombosis [18– 20]. may provide additional information about the involvement of the endothelium in vascular permeability. which is involved in the control of cell–cell cohesion and thus endothelial permeability [27]. are valid estimates of endothelial dysfunction. high levels of endothelium-derived regulatory proteins or microalbuminuria have an adverse cardiovascular prognosis [21–23]. On the other hand. Measurement Impaired endothelium-dependent vasodilation Increased transcapillary escape rate of intravenously injected radiolabelled albumin. are probably in part endothelium-dependent. It is not entirely certain whether the measurements of endothelial function. with its very large surface area and synthetic capacity.Vascular complications in diabetes mellitus: the role of endothelial dysfunction 145 Table 1 Proposed estimates of endothelial dysfunction in humans ↑. microalbuminuria. PAI-1 can be produced not only by endothelial cells. estimates exist for assessing endothelial function in vivo in humans. For example. Moreover. adipocytes and vascular smooth muscle cells [24]. 2005 The Biochemical Society C . microalbuminuria and sCD146 ↑Endothelin ↑vWF ↑sThrombomodulin ↑t-PA and PAI-1 ↑sE-selectin and sVCAM-1 ↑sICAM-1 ↑Cellular fibronectin and Type IV collagen fragments Interpretation of altered endothelial function Decreased production of vasodilators and/or increased production of vasoconstrictors Increased permeability to macromolecules Increased production of vasoconstrictors Increased prothrombotic and procoagulant activity Decreased anticoagulant activity Decreased profibrinolytic activity Increased adhesion of and permeability to leucocytes Inflammatory activation Altered extracellular matrix synthesis atherogenic risk factors have in common that endothelial NO availability is decreased. whether through decreased production or through increased degradation. For example. which increases vascular NAD(P)H oxidases. but not perfect. such as arterial stiffness and intima/media thickness in the carotid artery. First. Thus reasonable. and (iii) endothelial function in the microvasculature parallels that in large arteries. but also by the biochemical and biophysical properties of the extracellular matrix [25] and by haemodynamic forces [26]. increased plasma or serum level. unless the latter should be endothelium dependent. novel data with soluble CD146. and may partly be confounded by impaired vascular smooth muscle function. Information on the validity of these assumptions is as yet scarce. plasma levels of endothelium-derived regulatory proteins and. it is likely that the transcapillary escape rate of albumin is determined not only by the endothelium. superoxide production and NO scavenging. These processes will lead to endothelial activation manifested by increased expression of adhesion molecules [16]. the concept that high plasma levels of endotheliumderived mediators reflect endothelial dysfunction in clinically relevant arteries (such as the coronary and carotid) requires that (i) other cell types are not an important source. s. which decreases NO production. but also increases the interaction between NOS (NO synthase) and caveolin-1. soluble. but also by hepatocytes. a member of the immunoglobulin superfamily. Estimates of different types of endothelial dysfunction may be obtained indirectly by measuring endothelium-dependent vasodilation. is the most important determinant of plasma levels of endotheliumderived mediators. the latter because of the fact that microvascular endothelium. prospective studies have shown that individuals with impaired endothelium-dependent vaso- dilation. as listed in Table 1. Some other vascular properties. (ii) synthesis is more important than clearance. Another important example is Ang II.

45]. that albumin excretion rate was significantly and indepen2005 The Biochemical Society dently correlated with levels of plasma vWF and soluble E-selectin. in Type II diabetes [32] and also in non-diabetic subjects [33].and macro-albuminuria are often associated with classic risk factors for microangiopathy and atherothrombosis. Prospective studies using markers such as plasma vWF have shown that high vWF concentrations are associated with an increased risk of developing microalbuminuria. have very high risk of developing severe complications [29–31]. by influencing glomerular mesangial and epithelial cell function in a paracrine fashion. by causing increased glomerular pressure and the synthesis of a leaky glomerular basement membrane. nephropathy and atherosclerosis in both Type I and Type II diabetes [12. dyslipidaemia and smoking [34].45]. patients who have had diabetes for more than 5–10 years and who have normal urinary C . Many studies have investigated this and have concluded that common risk factors explain at most a small part of the association between (micro)albuminuria and atherothrombosis [32.and macroalbuminuria are accompanied by a variety of markers of endothelial dysfunction.or macro-albuminuria is generalized in that it affects many aspects of endothelial function and occurs both in the kidney and elsewhere. obesity. which may include severe generalized endothelial dysfunction [37] and chronic low-grade inflammation [38]. progress with time. A. G. For example. macroalbuminuria. chronic low-grade inflammation can be both cause and consequence of endothelial dysfunction. we recently demonstrated [22]. endothelial function in diabetes has been investigated to answer several distinct questions. Because micro. Chronic low-grade inflammation is another candidate to explain the association between (micro)albuminuria and extrarenal complications [22.35. which has raised the question of what common mechanisms may be at work. regardless of the presence of diabetes. the occurrence of diabetic retinopathy and neuropathy and an increased risk of cardiovascular events and death [39–42]. i. However. We concluded that. In both types of diabetes. Indeed. together with more limited data showing that microalbuminuria is also associated with endothelial dysfunction in the absence of diabetes [37. chronic low-grade inflammation. The pattern of increased risk for vascular complications can be observed even in early nephropathy. Indeed.44. Other mechanisms must therefore be at work. an alternative or additional explanation is that there are other pathways that link (micro)albuminuria to extrarenal complications. such as (i) is the occurrence of microalbuminuria accompanied by severe endothelial dysfunction. such as autonomic neuropathy [47. microalbuminuria. In addition. Stehouwer ENDOTHELIAL DYSFUNCTION IN DIABETES General remarks A considerable body of evidence in humans indicates that endothelial dysfunction is closely associated with the development of diabetic retinopathy.32]. D. notably poor glycaemic control. increased urinary albumin excretion and endothelial dysfunction develop in parallel. and the two appear tightly linked [22. Endothelial dysfunction in Type I and II diabetes complicated by micro. and are strongly and independently associated with risk of death. an increased progression of microalbuminuria.43]. Such data. It is less clear how endothelial dysfunction would cause (micro)albuminuria. Importantly. (ii) is hyperglycaemia a sufficient cause of at least some types of endothelial dysfunction. a subgroup exists with a relatively normal compared with a very high risk of cardiovascular complications.36]. Thus. in a study of 328 patients with Type II diabetes who were followed for a mean of 9 years. an obvious possibility is that such risk factors cause both (micro)albuminuria and atherothrombosis and thus explain their association. (iii) what is the significance of the finding that insulin resistance is associated with endothelial dysfunction and (iv) what are the mechanisms that cause diabetes-associated endothelial dysfunction? Generalized endothelial dysfunction. One possibility is that such markers do not fully capture the processes they are meant to reflect. a substantial fraction will never develop severe vascular complications. the molecular pathways by which endothelial dysfunction causes (micro)albuminuria have yet to be worked out. recent data indicate that the association between (micro)albuminuria and atherothrombotic disease cannot be explained entirely by markers of endothelial dysfunction and chronic inflammation [22. have led to the concept that microalbuminuria itself is a marker of generalized renal and extrarenal endothelial dysfunction. endothelial dysfunction could contribute to the pathogenesis of albuminuria both directly.48] or prothrombotic mechanisms (Figure 1).146 C. i.e. microalbuminuria and atherothrombosis Although a majority of diabetic patients will develop vascular complications.e. In addition. and indirectly. in Type II diabetes. Nevertheless. Theoretically. chronic low-grade inflammation is associated with the occurrence and progression of (micro)albuminuria [44] and with risk of atherothrombotic disease [46]. At what stage of diabetes does endothelial dysfunction become manifest? Type I diabetes In Type I diabetes. the literature on endothelial function in diabetes is complex in part because endothelial function can be measured in many ways and in many vascular beds. patients with advanced nephropathy. within the group of diabetic patients.28].44. micro. in both types of diabetes. hypertension. Schalkwijk and C.

early uncom- plicated Type I diabetes is accompanied by dilation. It is at this stage that a general impairment of endothelial function. On the other hand. Taken together. hyperinsulinaemia. inflammation and urinary albumin excretion in Type II diabetes are progressive and closely interrelated [22]. Endothelial dysfunction.Vascular complications in diabetes mellitus: the role of endothelial dysfunction 147 Figure 1 Postulated pathways linking conventional risk factors. low HDL (high-density lipoprotein)-cholesterol levels. both in humans and in animal models [54]. as in Type I 2005 The Biochemical Society C . can usually be clearly observed.49].e. the (arbitrary) cut-off of 30 mg/24 h may be too high. but in and of itself is not a sufficient cause. However. is endothelial dysfunction a feature of Type I diabetes and is moderate hyperglycaemia sufficient to impair endothelial function? On the one hand. Impairment of endothelial function may be especially severe among diabetic women [58]. genetic or environmental. that patients with ‘normal’ urinary albumin excretion who show such cardiovascular abnormalities are those in whom urinary albumin excretion is in fact increased albeit within the normal range as conventionally defined. in time. Endothelial dysfunction thus occurs before microalbuminuria sets in. For example. does become impaired before the onset of microalbuminuria. endothelium-dependent and -independent vasodilation of resistance and conduit arteries are neither impaired nor enhanced [52–55]. studies that carefully stratified patients according to the absence or presence of a normal urinary albumin excretion rate have concluded that. although endothelial function may sometimes be normal even among microalbuminuric individuals [56]. Type II diabetes typically occurs in the context of a cluster of cardiovascular risk factors. suggesting normal endothelial function. Notably. including endothelial function. Secondly. Nevertheless. including impaired endothelium-dependent vasodilation.51].45. in conflict with these concepts [52–55]. in reasonably well-controlled Type I diabetes.36]. Microvascular dilation may cause capillary hypertension and. (micro)albuminuria and atherothrombosis in diabetes albumin excretion compared with non-diabetic individuals are characterized by subtle increases in blood pressure and large-artery stiffness and by autonomic dysfunction. according to the socalled haemodynamic hypothesis of the pathogenesis of microangiopathy. plasma vWF levels are not increased in patients with early uncomplicated and reasonably well-regulated Type I diabetes. It is not clear how they interact or whether impaired endothelial function is a common antecedent or a consequence. capillary hypertension will. but these data illustrate that cardiovascular function. abnormal LDL composition. Endothelial dysfunction in Type II diabetes appears to be independent of that induced by obesity [59]. such as microalbuminuria. This has a parallel in Type II diabetes and in non-diabetic individuals in whom any increase in microalbuminuria appears associated with increased risk of atherothrombosis [32. insulin resistance and chronic low-grade inflammation. notably obesity. hypertension. however. It is not clear to what extent this endothelial dysfunction is caused by hyperglycaemia. acutely impairs endothelium-dependent vasodilation in non-diabetic individuals [50. Other factors. the diabetic state predisposes to the development of endothelial dysfunction. Type II diabetes Endothelial dysfunction indicated by impaired endothelium-dependent vasodilation and increased plasma concentrations of markers of endothelial function is common in early and otherwise uncomplicated Type II diabetes. First. Several findings are. damage the microvascular endothelium and thus set the stage for more advanced stages of microangiopathy. the mediators responsible for the vasodilation typical of early Type I diabetes remain to be identified and it is not known whether the endothelium is involved. endothelial dysfunction. All these abnormalities are worse in the microalbuminuric stage [26. in the absence of other factors. of small and large blood vessels and an increase in microvascular blood flow. but the role of the other variables (compared with hyperglycaemia) remains to be established. vWF and soluble VCAM-1 levels are increased and have been found to be associated with an increased risk of cardiovascular mortality and development and progression of microalbuminuria [22. and some or perhaps most have impaired endothelial function and increased low-grade inflammation. inflammatory activity. however. all of which may impair endothelial function. An important contender is increased inflammatory activity. i. at least when the latter is defined as 30 mg/24 h. these data suggest that impaired endothelium-dependent vasodilation may occur early in a subset of Type I diabetic patients apparently dependent on other environmental or genetic factors such as the Ang II type I receptor gene polymorphism [57]. are likely to play a role in determining who among Type I diabetic patients go on to develop aggressive angiopathy and who do not. The possibility remains. not constriction. high triacylglycerol (triglyceride) levels. it has been argued that NO-mediated endothelium-dependent vasodilation is impaired in short-term uncomplicated diabetes and that hyperglycaemia.44]. In other words.

NEFAs and perhaps hyperinsulinaemia itself impair these actions of insulin. in vivo. The molecular pathways through which insulin increases NO synthesis. endothelial dysfunction C . however. How can endothelial dysfunction impair insulininduced glucose disposal? First. with vasodilation as the normal response and impaired vasodilation or even net vasoconstriction as abnormal responses [73]. and through active deformations. a potent vasoconstrictor. insulin can act on insulin receptors on endothelial cells to produce NO.e. Secondly. or how TNF-α. namely TNF-α (tumour necrosis factor-α). by impairing transendothelial insulin transport. INSULIN RESISTANCE AND THE METABOLIC SYNDROME Insulin resistance usually precedes the development of Type II diabetes and is often accompanied by a cluster of other risk factors (see above). A widely accepted theory states that insulin resistance is the primary abnormality that gives rise to Type II diabetes. endothelium-dependent vasodilation and capillary recruitment have not yet been fully elucidated. physical integrity and normal function of the arteriolar and capillary endothelium are prerequisites for normal metabolic insulin action. Nevertheless. LPL is the rate-limiting enzyme for triacylglycerol utilization and its physiological site of action is the capillary endothelial surface [72].g. microalbuminuria in Type II diabetes can occur in the absence of severe endothelial dysfunction [56. In addition to the regulation of peripheral resistance at the level of arterioles. by rarefaction (modelling indicates that for approx. notably Type II diabetes. reduced capillary density per volume of tissue). Stehouwer diabetes. In this way. Decreased capillary density and impaired capillary recruitment may in part explain why insulin resistance is associated with hypertension [69–71]. G. hypertension and dyslipidaemia. First. insulin may act at insulin receptors on vascular smooth cells to cause vasodilation and (or) on skeletal muscle to activate glucose metabolism to produce a metabolite (e. namely insulin resistance and endothelial dysfunction. D.60]. Schalkwijk and C. Indeed. ENDOTHELIAL DYSFUNCTION.e. A reduced capillary endothelial surface area may in turn result in reduced access of triacylglycerol-rich lipoprotein particles to LPL. there is a approx. contractility. obesity and hypertension. 20 % increase in peripheral vascular resistance). adenosine) that then acts on local endothelial and (or) smooth muscle cells. the endothelial dysfunction in large arteries that is an early and prominent event in atherothrombotic disease is paralleled by endothelial dysfunction in resistance vessels and metabolically important capillary beds that contributes to the development of the metabolic syndrome [61]. Therefore it is unlikely that a simple insulin-induced increase in total blood flow can increase glucose disposal. Secondly. insulin can redirect blood flow in skeletal muscle from non-nutritive capillaries (those that are not coupled to muscle cells) to nutritive capillaries (those that are) and thus increase glucose disposal even without increasing total blood flow [62–64]. capillaries can contribute by virtue of their narrow calibre and relative non-distensibility. in a process termed capillary recruitment. NEFAs (nonesterified fatty acids) and an impaired NO-dependent action [65–68]. i. but all elements of the cluster share two important pathophysiological features. and by impairing recruitment of previously underperfused muscle tissue. An alternative concept is that endothelial dysfunction is at the heart of the metabolic syndrome. insulin is a vasoactive hormone. and to be decreased by mediators closely associated with insulin resistance. 40 % rarefaction. Such a mechanism could explain why dyslipidaemia of the metabolic syndrome is confined to triacylglycerols and HDL-cholesterol. if this should be surface-area-dependent. chronic low-grade inflammation is closely associated with. but also ET-1. these data suggest that endothelial dysfunction and impaired capillary recruitment can cause insulin resistance with respect to glucose disposal both when the microvascular endothelium is 2005 The Biochemical Society otherwise healthy but cannot react properly to insulin (‘endothelial insulin resistance’) and when the microvascular endothelium is injured through other mechanisms. and then prognosis is relatively good [41]. regardless of the presence of diabetes. decreased capillary density and impaired capillary recruitment may decrease insulin-mediated glucose disposal by increasing the diffusion distance of glucose and insulin to glucose-metabolizing tissues. and that endothelial dysfunction merely represents the impact of hyperglycaemia and other features of the metabolic syndrome.148 C. i.. According to this concept. this does not exclude that. and may link. insulin’s vasodilator actions have been shown to be impaired in classic insulin-resistant states. The mechanisms underlying this clustering are still unclear. Endothelial insulin resistance can thus be conceptualized as a shift in the balance between vasodilators and vasoconstrictors produced by insulin. Taken together. insulininduced increases in glucose uptake and total blood flow have different concentration–effect curves as well as time kinetics. although insulin’s endothelial actions have been shown to occur in cell culture and in isolated vessels. such as age-related capillary drop-out (‘rarefaction’. Insulin increases muscle blood flow in a timeand concentration-dependent fashion through a mechanism that involves binding to the insulin receptor on the endothelial cell membrane. A. Thirdly. Specifically. Decreased capillary density and impaired capillary recruitment may also play a role in the development of atherogenic changes in lipoprotein concentrations through impaired action of endothelium-bound LPL (lipoprotein lipase).

and increased oxidative stress by the regulation of several NADPH oxidases. the relatively low expression of aldose reductase in endothelial cells may not be sufficient to cause significant sorbitol accumulation. hyperglycaemia and its immediate biochemical sequelae directly alter endothelial function [77]. basement membrane thickening through TGF-β (transforming growth factor-β)-mediated increased synthesis of type IV collagen and fibronectin. increased sorbitol accumulation will increase osmotic stress. dysregulation of blood flow by decreasing endothelial NOS activity and (or) increasing ET-1 synthesis [94]. glucose and glucosederived dicarbonyl compounds react non-enzymatically with the basic amino acids lysine and arginine in proteins C . The sorbitol pathway In most cells. Figure 2 Postulated pathways linking cardiovascular risk factors. and of procoagulant proteins. and the activation of PKC (protein kinase C) via de novo synthesis of DAG (diacylglycerol). Thirdly. and increased apoptosis [81–86].Vascular complications in diabetes mellitus: the role of endothelial dysfunction 149 to form AGEs (advanced glycosylation end-products) both extra. MECHANISMS UNDERLYING ENDOTHELIAL DYSFUNCTION IN DIABETES General remarks Endothelial dysfunction in diabetes originates from three main sources [77–80]. in which an increase in blood glucose concentration will thus increase the intracellular accumulation of glucose and its metabolites. An increase in intracellular glucose will lead to an increase in the flux of glucose to sorbitol via the polyol pathway. Glucose transport is autoregulated by glucose in smooth muscle cells. and show decreased proliferation.and intra-cellularly. These postulated pathways are summarized in Figure 2. Thirdly. In contrast.90]. First. 2005 The Biochemical Society Hyperglycaemia and its immediate biochemical sequelae Various mechanisms have been proposed to explain how hyperglycaemia directly causes diabetic vascular complications. Figure 3 shows how. migration and fibrinolytic potential. insulin resistance and microvascular function and metabolic insulin resistance [74–76]. the components of the metabolic syndrome can affect endothelial function [88]. intracellularly. such as vWF and tissue factor. excess glucose can be metabolized to sorbitol and fructose by aldose reductase and sorbitol dehydrogenase. The full impact of the sorbitol pathway in vascular dysfunction is not completely understood and the role of inhibition of aldose reductase in the prevention and treatment of diabetic complications remains unclear. Glucose transport into endothelial and vascular smooth muscle cells occurs by facilitated diffusion and is thus insulin-independent. the redox imbalance favours the accumulation of triose phosphates which increases the formation of methylglyoxal and AGEs and enhances oxidative stress. Sorbitol accumulation decreases other osmolytes such as myo-inositol and taurine. these four biochemical mechanisms may all be the consequence of hyperglycaemia-induced overproduction of ROS in mitochondria. Aldose reductase activity in endothelial cells of different origin is low and it thus appears unlikely that the improved nerve conduction in diabetic neuropathy observed with aldose reductase inhibitors or myo-inositol supplementation is related to improved endothelial function. Secondly. the increase in the cytosolic NADH/ NAD+ ratio results in a redox imbalance that resembles that which occurs in tissue hypoxia and therefore is termed hyperglycaemic pseudohypoxia [91]. which can be exacerbated by NADPH-deficiency-induced depletion of reduced glutathione. The DAG/PKC pathway The cellular pathogenic consequences of hyperglycaemiainduced activation of PKC are multiple and include dysregulation of vascular permeability directly or indirectly {the latter through the induction of VEGF (vascular endothelial growth factor) in smooth muscle cells [93]}. These different pathways are interrelated and potentiate each other. In addition. an excess aldose reductase activity in human retinal endothelial cells can be a mechanism for human diabetic retinopathy [92]. Thus endothelial cells exposed to high glucose in vitro increase the production of extracellular matrix components. However. impaired fibrinolysis through increased expression of PAI-1. This pathway may impair endothelial function through three mechanisms. such as collagen and fibronectin. an increase in glucosamine-6phosphate via the hexosamine pathway. Secondly. First. but not in endothelial cells. which is accompanied by increased oxidation of NADPH to NADP+ and increased reduction of NAD+ to NADH [89. high glucose influences endothelial cell functioning indirectly by the synthesis of growth factor and vasoactive agents in other cells [87].

is synthesized de novo from the glycolytic intermediates dihydroxyacetone phosphate and glyceraldehyde-3-phosphate [95]. Hyperglycaemia-induced activation of PKC occurs through increased levels of the PKC activator DAG which. in which fructose 6-phosphate is converted into glucosamine 6phosphate by the enzyme glutamine:fructose-6-phosphate amidotransferase. G. Adapted from [77] with permission. in cells with a low aldose reductase activity such as endothelial cells. For example. Under normal conditions. Recent evidence indicates that these four biochemical and metabolic mechanisms are the consequence of a hyperglycaemia-induced overproduction of oxidative stress in the mitochondria. by the addition of GlcNAc to serine and threonine residues. This. into GlcNAc. In diabetic animals.150 C. the activation of PKC via de novo synthesis of DAG. subsequently. The mechanism responsible for the activation of PKC by hyperglycaemia is related to de novo synthesis of the PKC activator DAG from a stepwise acylation of glycerol 3-phosphate (GP) and phosphatidic acid (PA). excess glucose can be metabolized in the sorbitol pathway to sorbitol and fructose by aldose reductase (AR) and sorbitol dehydrogenase (SDH). and partly corrected urinary albumin excretion [99]. c (2001) Nature Publishing Group (http://nature. D. an increase in fructosamine 6-phosphate via the hexosamine pathway. type IV collagen and fibronectin. and the formation of AGEs. glucose is metabolized through the glycolytic pathway. hyperglycaemia may lead to increased ROS by activation of NAPDH oxidase. In addition to this pathway. Studies to evaluate the importance of the DAG/PKC pathway in humans are underway. A. the DAG/PKC pathway and the formation of AGEs) can be inhibited by the lipid-soluble thiamine derivative benfotiamine by activating the pentose phosphate pathway (PPP) enzyme transketolase (TK) [78. or uncoupling of eNOS.117]. Schalkwijk and C. vitamin E can inhibit PKCβ activity. Stehouwer Figure 3 Potential mechanisms by which hyperglycaemia and its immediate biochemical sequelae induce hyperglycaemic damage Various mechanisms have been proposed to explain how hyperglycaemia causes diabetic vascular dysfunction. 2005 The Biochemical Society The hexosamine pathway The vascular effects of the hexosamine such a modification of the Akt site of endothelial cell NOS has been shown to decrease enzyme activity [102]. Interestingly. indicating a link between oxidative stress and PKC activation [96]. such as PKC and endothelial cell NOS can be modified in a similar way. In aortic endothelial cells. An increase in intracellular glucose will lead to an increase in four pathways: the flux of glucose to sorbitol via the sorbitol pathway. the PKCβII isoform appears preferentially activated [98]. C . which decreased SP-1 phosphorylation and increased SP-1 activity. inactivation and reduced expression of the antioxidant enzymes catalase and superoxide dismutase (SOD). hyperglycaemia was shown to increase levels of hexosamine 6-phosphate and subsequently GlcNAc (Nacetylglucosamine). In turn this can increase transcription of PAI-1 and TGF-β1 [101]. The PKC family consists of at least eleven isoforms [97]. fructose 6-phosphate is converted into fructosamine 6-phosphate by the enzyme glutamine:fructose-6-phosphate amidotransferase (GFAT) and. are just beginning to be understood but may be profound [100]. increased O-linked glycosylation of the transcription factor SP-1. In the hexosamine pathway. an oral PKCβ inhibitor prevented diabetes-induced abnormalities in mRNA expression of TGF-β1. In several cell types. ameliorated increases in glomerular filtration rate and accelerated glomerular mesangial expansion. Three of the major biochemical pathways implicated in the pathogenesis of hyperglycaemia-induced vascular damage (the hexosamine pathway. In vascular cells. Other proteins.

are believed to contribute importantly to the formation of AGEs in vivo [104]. AGE-modified matrix stimulates interactions with mononuclear cells and macromolecules such as LDL. such as G-proteins. AGEmodified plasma proteins can bind to AGE receptors. AGEs are a mixture of different moieties. glyoxal and 3-deoxyglucosone. Clinical trials with aminoguanidine. including RAGE (receptor for AGE). methylglyoxal is probably the main AGE formed [105. an AGE formation inhibitor that had shown promise in animal experiments. TNF-α and TGF-β. in particular by mitochondria that have been uncoupled by the flux of NADH from the hyperglycaemia-enhanced glycolysis. galectin-3. oxidative stress may be the initial event in endothelial cell dysfunction [77]. blockade of RAGE inhibited the development of macrovascular disease and diabetic nephropathy. on different cell types such as endothelial cells [107].117]. causes inhibition of GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and subsequent accumulation of glycolysis intermediates. This so-called carbonyl stress has been implicated in the accelerated vascular damage in both diabetes and uraemia. the highly reactive dicarbonyl compounds methylglyoxal. Hyperglycaemia-induced oxidative stress as a common activator of the four biochemical pathways Recent evidence suggests that hyperglycaemia-induced mitochrondrial overproduction of superoxide anion radicals plays a key role in the activation of the above pathways [77] (Figure 3).Vascular complications in diabetes mellitus: the role of endothelial dysfunction 151 Non-enzymatic glycation Non-enzymatic glycation of proteins is the condensation reaction of the carbonyl group of sugar aldehydes with the N-terminus of free amino acids of proteins and initially leads to a Schiff’s base. AGE-modified type I and IV collagen inhibit normal matrix formation and cross-linking. activation of NADPH oxidases and uncoupling of eNOS (endothelial NOS) [119– 121]. In endothelial cells. Hyperglycaemia-induced GAPDH inhibition was found to be a consequence of poly(ADP-ribosyl)ation of GAPDH by PARP [poly(ADP-ribose) polymerase]. The overproduction of superoxide. and AGEs may act as oxidants. and decrease arterial elasticity. activation of NF-κB and interference with the availability of NO. thus leading to the generation of superoxide instead of NO. and prevented experimental diabetic microvascular retinopathy and nephropathy [78. which are formed from the degradation of glycolytic intermediates. have unfortunately been halted because of unforeseen side effects [114]. These include direct effects on the endothelium such as peroxidation of membrane lipids.124] and urinary excretion of 8-iso-PGF2α (8-iso-prostaglandin F2α ) [125]. PKC activation and AGE formation by methylglyoxal. the macrophage scavenger receptor A. In endothelial cells. namely auto-oxidation of glucose and nonenzymatic glycation [118]. In animal models. Oxidative stress as a final common pathway of hyperglycaemia-induced vascular dysfunction Enhanced oxidative stress in hyperglycaemia is indicated by increased levels of lipid hydroperoxides [123. which then undergoes rearrangement to early glycation Amadori-adducts such as fructosamine [103]. Initially. AGE-R1/p60 and AGE-R2/p90. and may modify endothelial function by a variety of mechanisms [126]. Further support for the involvement of accumulated glycolysis intermediates came from experiments demonstrating that activation of the enzyme transketolase by benfotiamine reduced these glycolysis intermediates and the various pathways of endothelial activation. high doses of vitamin C can improve some aspects of endothelial 2005 The Biochemical Society C . which was activated by DNA strand breaks produced by mitochondrial superoxide overproduction [116]. tissue factor and VCAM-1 genes is adversely affected. so-called glycoxidation products such as pentosidine and Nε (carboxymethyl)lysine result. In addition. The introduction of AGEs in the extracellular matrix can interfere with endothelial function in several ways. Although in short-term experiments. In addition. in macrophages and mesangial cells. Ligation of RAGE has been shown to mediate signal transduction via a receptor-mediated induction of ROS and activation of the transcription factors NF-κB and p21ras . several other mechanisms can contribute to superoxide production in diabetes. but trials with other AGE formation inhibitors and with AGE crosslink breakers are underway [115]. Many aspects of diabetic complications are thus potentially related to the effect of Amadori-adducts and AGEs [108–113]. ROS can affect many signalling pathways. ion channels and transcription factors. eNOS may become uncoupled in the presence of low levels of l-arginine or cofactors. In addition to mitochondrial uncoupling. and impaired antioxidant status [122]. which form AGEs at a much faster rate than glucose. because of the slow rate of reaction of glucose with proteins. AGEs were thought to form only on long-lived extracellular molecules. there is an increased expression of cytokines and growth factors such as IL-1. protein kinases. the expression of the thrombomodulin. It is not known whether oxidative stress causes endothelial dysfunction in human diabetes. Amadori-adducts are relatively stable and only a small fraction undergoes rearrangements to irreversible AGEs. Since inhibition of hyperglycaemia-induced ROS production prevented the hyperglycaemia-associated activation of the aldose reductase and hexosamine pathways. intracellular and short-lived molecules have now also been shown to be targets for AGE formation through reactions with other sugars such as glucose 6-phosphate and glyceraldehyde 3-phosphate. When oxidation is involved in their formation.106]. However.

by inhibiting anti-coagulant pathways and by impairing fibrinolysis via stimulation of PAI-1. inflammatory cytokines increase vascular permeability. TGFβ-1 exerts its effects via two transmembrane serine/threonine kinase receptors. Finally. which stimulates differentiation. 2005 The Biochemical Society TNF-α TNF-α is an inflammatory cytokine produced by neutrophils. G. VEGF VEGF is a multi-tasking cytokine. randomized clinical trials with antioxidants have failed to show a decrease in cardiovascular disease [128. The expression of VEGF can be induced by hypoxia through HIF-1 (hypoxia-inducible factor-1). laminin and proteoglycans. endothelial dysfunction and atherothrombosis are so closely related (see above). Enhanced glomerular expression of TGF-β1 in human diabetic nephropathy has been reported [135]. stretch and Ang II. increase leucocyte adhesion to endothelium and facilitate thrombus formation by inducing pro-coagulant activity. cell death or apoptosis and cellular differentiation. type I and II. The role of TGF-β in the thickening of capillary basement membranes elsewhere (e. These mediators alone or in combination can impair endothelial function and contribute to atherothrombosis. that are co-expressed on mesangial and endothelial cells [134]. have been best characterized [143]. Ang II. importantly. TGF-β TGF-β1 is secreted by different cell types as an inactive dimer consisting of a latency-associated peptide and mature TGF-β1. TGF-β1 regulates multiple cellular functions.152 C. expansion of the mesangial matrix [135]. recent studies have shown that TNF-α and inflammation in general can contribute to the pathogenesis of diabetic nephropathy. In addition. Activation of the transcription factor NF-κB is crucial in cytokine regulation of gene expression in endothelial cells [151]. The role of increased or decreased VEGF in other diabetic complications is the subject of much ongoing research [148–150]. Growth factors and cytokines There is solid evidence that TGF-β [with other growth factors such IGF-1 (insulin-like growth factor-1) and EGF (epidermal growth factor)] plays a major role in diabetic nephropathy [131]. TGF-β1 has been proposed as the major candidate to mediate the progression of diabetic nephropathy [136] by inducing glomerular and tubular changes resulting in progressive thickening of the glomerular basement membrane. treatment of diabetic mice with anti-TGF-β1 antibodies significantly attenuated the increase in TGF-β1 activity and extracellular matrix expression [138]. tissue factor. migration. -6 and -8. NF-κB is activated not only by TNF-α and IL-1. Because vitamin E-treated patients had a worsening in some vascular reactivity measurements when compared with control subjects. suggesting that it may be important both in atherothrombosis and in microangiopathy. but also by IGF-1 and TGF-β1 [142]. E-selectin. TGF-β1 also has potent anti-inflammatory effects on vascular cells. and cytokine activation of endothelial cells [133]. long-term treatment with high doses of vitamin E has no beneficial effects on endothelial or left ventricular function [130]. but also by hyperglycaemia.129]. adipocytes. and is activated by serine proteases such as plasmin. TNF-α may cause insulin resistance and impair endothelial function in large arteries and also in the microcirculation. AGEs. down-regulating cytokine-induced expression of VCAM-1 [139] and MCP-1 [140]. macrophages and. Stehouwer dysfunction in diabetes [127]. On the other hand. In diabetic patients. Briefly. in turn. demonstrating a role for TGF-β1 in diabetic nephropathy. Furthermore. tubulogenesis and vascular permeability in endothelial cells [141]. Schalkwijk and C. of which VEGF receptors 1 and 2. Amadori-albumin and AGEs. In diabetic retinopathy. in the retina) is less well established. regulates the expression of CRP (C-reactive protein). IL-1. the use of high dosages of vitamin E cannot be recommended. TNF-α can induce insulin resistance. Examples of NF-κB-regulated genes include VCAM-1. alteration of retinal microvascalature and increased vascopermeability are strongly stimulated by the interaction of VEGF with the endothelium [144]. proliferation. prevention of Ang II formation was associated with lower VEGF concentrations in the vitreous fluid of patients with proliferative diabetic retinopathy [147]. The spectrum of endothelial cell responses elicited by cytokines is varied. NF-κB C . including inhibition and stimulation of cell growth. It is also a potent inducer of extracellular matrix protein synthesis such as type I and IV collagen. NF-κB consists of a family of transcription factors that serve as important regulators of the inflammatory response and the regulation of vascular cell function. TGF-β1 is increased through hyperglycaemia-induced PKC activation. fibronectin. TNF-α can induce other powerful cytokines such as IL-6 which. a reduced glomerular filtration function and an increased excretion of proteins [137]. VEGF binds to several receptors. D. and antagonists of VEGF and its receptors have been shown to reduce retinopathy in animal models [146]. survival. ICAM-1 (intercellular celladhesion molecule-1). alter vasoregulatory responses. which may at least in part explain why insulin resistance.g. oxidized lipids and insulin. Several studies have shown increased vitreal VEGF levels in patients with proliferative diabetic retinopathy compared with patients without proliferative retinopathy [145]. also known as fms-like tyrosine kinase and fetal liver kinase-1. and VEGF is important in severe retinopathy [132]. Furthermore. A. PAI-1 and inducible NOS.

Interestingly. but also in monocytes/macrophages and vascular smooth muscle cells. cell adhesion molecules.159]. Also. decreased medullary blood flow and impaired pressure natriuresis. largely in endothelial cells. promotes monocyte chemotaxis and adhesion. dyslipidaemia and obesity Insulin resistance How metabolic and endothelial insulin resistance occur and why they are closely related is not fully understood (see above). which phosphorylates and activates eNOS. pro-insulin split products and C-peptide. resulting in increased ROS and vascular smooth muscle cell proliferation. The vascular effects of these peptides have not been extensively investigated. Impaired endothelial function in secondary hypertension is usually reversible upon reduction of blood pressure. For the vasculature. Future studies should be directed at delineating the molecular mechanisms for these important in vitro observations. and. insulin precursor molecules. Both TNF-α and NEFAs can cause metabolic and endothelial insulin resistance. Plasma levels of CRP are increased in both Type I and Type II diabetes [45. In contrast.Vascular complications in diabetes mellitus: the role of endothelial dysfunction 153 protects against apoptosis and activates the antioxidant enzyme SOD (superoxide dismutase). increases ROS and pro-inflammatory cytokine release. mainly because it is not clear whether effects such as increased vascular permeability to macromolecules and increased vascular smooth muscle cell proliferation occurs at physiological concentrations. can be regarded as a form of endothelial dysfunction and conceivably contributes to both atherothrombosis and microangiopathy. Hypertension causes endothelial activation as indicated by elevated levels of soluble adhesion molecules [161] and impaired NO availability (see above). whether insulin has atherogenic effects is controversial. and increases oxidized LDL uptake. notably intact pro-insulin. some patients with essential hypertension do not normalize endothelial function with blood-pressure lowering [162. CRP An important downstream marker of inflammation is CRP. 2005 The Biochemical Society C . have now suggested a role for CRP in atherogenesis [154. and increased ET-1. insulin.163]. MCP-1. Experimental data indicate that a decrease NO availability in the kidney may contribute to vasoconstriction and decreased glomerular filtration. How NEFAs impair insulin’s endothelial actions is not clear. whereas the vasoconstrictor effects of insulin are mainly mediated by the vasoconstrictor peptide ET-1. Numerous studies have shown that CRP levels predict cardiovascular disease [153]. Taken together.155]. CRP has been shown in vascular smooth muscle cells to increase inducible NO production. Insulin precursor molecules Type II diabetes is characterized by high levels of molecules that arise as a by-product of insulin synthesis and secretion. these data suggest that NF-κB pathway is an important contributor to the pathogenesis of vascular disease in diabetes mellitus. In conclusion. However. most importantly. The metabolic syndrome: insulin resistance. studies should be directed at confirming these findings in animal models and other systems as proof of concept. in contrast. although the molecular basis for these effects has not been fully elucidated [158. Salt sensitivity of blood pressure may denote an inability to increase NO availability in response to increased blood pressure [162]. An imbalance between the release of NO and ET-1 may be involved in the pathophysiology of hypertension and also atherosclerosis in insulin-resistant states associated with endothelial dysfunction. Endothelial insulin resistance. Hypertension Hypertension is a major determinant of microangiopathy and atherothrombosis in diabetes [160]. TNF-α may induce endothelial insulin resistance through its ability to impair intracellular signalling by inhibition of insulin-stimulated autophosphorylation and phosphorylation of IRS-1 (insulin receptor substrate-1). Data on this issue in diabetes are scarce. CRP induces tissue factor secretion. hypertension. Insulin precursors may increase plasma levels of PAI-1. levels of these peptides are abnormally low in Type I diabetes. whether the latter then contributes to increased blood pressure is not clear. In monocytes/macrophages. impaired tubuloglomerular feedback. increase NF-κB and MAPK (mitogen-activated protein kinase) activities.152]. both vasodilator and vasoconstrictor effects of insulin have been described [156]. and progressive proteinuria. IL-8 and PAI-1. CRP is a risk marker for cardiovascular disease and could emerge as a mediator in atherogenesis. Insulin Both Type I and Type II diabetes are usually accompanied by chronic hyperinsulinaemia. up-regulate Ang II type 1 receptor. whether primary or secondary (Figure 2). C-peptide may affect vascular tone and permeability. NO is the main mediator of the acute dilatory effects of insulin in human. Also. Insulin stimulates NO production in endothelial cells by subsequently activating the intracellular enzymes PI3K (phosphoinositide 3-kinase) and Akt [157]. whereas ET-1 production is MAPK dependent [73]. The pro-inflammatory pro-atherogenic effects of CRP that have been documented in endothelial cells include the following: decreased NO and prostacyclin. much in vitro data.

vascular reactivity and several markers of endothelial dysfunction have been shown to be associated with an adverse cardiovascular prognosis regardless of the presence of diabetes.187]. P. Taken together. This is characterized by low HDL-cholesterol and a preponderance of small-dense LDL. insulin resistance and dyslipidaemia. Recently. J. Even slight elevations of LDLcholesterol in Type II diabetic patients are associated with a substantial increase in cardiovascular risk. K. Clin. which can influence microvascular function. C. G. the close link between endothelial dysfunction and (micro)albuminuria is an attractive explanation for the fact that microalbuminuria is a risk marker for atherothrombosis. G. A. In diabetes. [174] have recently demonstrated that impaired microvascular function may contribute to the development of obesityassociated microangiopathy. such as myocardial infarction and stroke [170]. S. G. JAMA. N. S. The inverse relationship of adiponectin with CRP [188. In addition. TNF-α (see above) and adiponectin. endothelial damage and vascular diseases. D. 44. A common mechanism underlying endothelial dysfunction relates to an increase in oxidative stress.154 C. hypertension and insulin resistance. as well as in patients with some of the disease states frequently associated with obesity such as Type II diabetes mellitus [177. which can enhance oxidative stress and impair endothelial function directly and indirectly by increasing the production of small-dense LDL particles and by reducing HDL [165]. adiponectin has recently been shown in vitro to modulate a wide array of biological functions with anti-atherogenic properties. (2004) Epidemiology of diabetes. Nature (London) 414.. Schalkwijk and C. hypertension and microalbuminuria. Sorlie. these may be the consequences of obesity-associated hypertension. these risk factors cannot entirely explain the association of obesity with largeartery disease and microangiopathy. REFERENCES 1 Zimmet. (2004) Trends in cardiovascular complications of diabetes. In diabetes. and Sowers. This is the case not only for largeartery disease. Am. Type II diabetes. J. but also for disease entities that are caused wholly or in part by microangiopathy.. hyperglycaemia and components of the metabolic syndrome cause endothelial dysfunction directly or indirectly. de Jongh et al. New insights into mechanisms of endothelial dysfunction may lead to novel important strategies of treatment. Pharmacol. Assoc. In addition to important roles in the regulation of energy homoeostasis and insulin sensitivity [181]. is characterized by increased postprandial triacylglycerol-rich lipoproteins (chylomicrons and veryLDL particles). and Shaw.178] and also in patients with coronary artery disease [179]. Since microvascular endothelial dysfunction is closely associated with and may contribute to insulin resistance. S. including the inhibition of the expression of adhesion molecules [182] 2005 The Biochemical Society ACKNOWLEDGMENTS C. Obesity The current obesity epidemic implies that obesity is becoming an increasingly important risk factor for cardiovascular disease [169]. CONCLUSIONS Assessment of blood flow. 782–787 2 Fox. Alberti. R. Dyslipidaemia has been associated with increases in urinary albumin excretion in both Type I and Type II diabetes [167. 397–405 C . J. Med. In part. (2001) Global and societal implications of the diabetes epidemic. Adiponectin is highly expressed in adipose cells. notably retinopathy. especially when glycaemic control is poor.168]. How obesity causes large-artery disease and microangiopathy is poorly understood. improvement of microvascular function should be one of the first targets. Coady. is supported by a personal fellowship from the Diabetes Fonds Nederland. J. such as NEFAs [175]. However. these data strongly indicate that hypoadiponectinaemia is linked to inflammation.189] also suggests that decreased production of adiponectin contributes to vascular complications. D. nephropathy and heart failure [171–173]. In humans. and reduction of monocyte attachment to endothelial cells [183]. Plasma adiponectin levels are reduced in patients with obesity [176]. In contrast with studies in Type II diabetic patients. Stehouwer Dyslipidaemia One important cardiovascular risk factor in Type II diabetic people is dyslipidaemia [164]. 2495–2499 3 Winer. The pathophysiological mechanism behind the relationship between obesity and microvascular dysfunction is probably multifactorial and may include mediators directly secreted by adipocytes. small cross-sectional studies in Type I diabetic patients found increased plasma levels of adiponectin [180].. These changes contribute to atherothrombosis and may also play a role in nephropathy [166]. et al. Although endothelial dysfunction predicts the occurrence of microalbuminuria. stimulation of the production of NO [184] and suppression of TNF-α in macrophages [185]. hypoadiponectinaemia is associated with impaired vasoreactivity [186. causality needs to be determined. P. evidence has been provided that an increased adiponectin concentration is associated prospectively with a lower risk of coronary artery disease in Type I diabetes and that this effect was independent of conventional risk factors and markers of inflammation and insulin resistance [190]. 292.

. Am. 382–386 40 Stehouwer. et al. J.Vascular complications in diabetes mellitus: the role of endothelial dysfunction 155 4 Duby. A. V. J.. 12 (Suppl. et al. (2004) Diabetic neuropathy: an intensive review. risk indicators.. Twisk. D. Diabetologia 45. and Rasmussen. M. Relation between endothelial dysfunction and microalbuminuria. F. (2000) Putative pathophysiological role of growth factors and cytokines in experimental diabetic kidney disease. M. 1911–1916 27 Bardin. M. but impaired glucose metabolism is not: The Hoorn Study. Buck.. Cardiol. Emeis. C. Am. 28. Intern. I. 986–993 35 Gerstein.. et al. Clin. 971–976 41 Jager. 17. Res.. V. 4–17 32 Jager. S. C. 1899–1906 24 Shimomura. M. Kostense. R. Mulder. F. J. Hypertens. (1996) Enhanced expression of PAI-1 in visceral fat: possible contributor to vascular disease in obesity. B. 617–624 33 Yudkin. (2003) Cardiovascular disease in patients who have diabetes. A. Funahashi. 329. 16. and van Hinsbergh. E. S.. Heine. Dekker. E. M. C. J. A. 1317–1327 18 Ashworth. Med. Arterioscler. J. van Kraats. 977–986 9 United Kingdom Prospective Diabetes Study (UKPDS) (1995) 13: relative efficacy of randomly allocated diet. and Johnson. (2004) Diabetic renal disease: from recent studies to improved clinical practice. Opin. and Zeiher. J.. S79–S83 30 Parving.. J. A. G. 421–426 37 Stehouwer. Q. 31. B.. Transplant. Med. G. Thromb. Stroes. Hypertens. 399–413.. 1). H. 55–68 13 Cines. Franz. Curr.. interrelated. Circulation 101. (2004) Nephropathy in diabetes. S19–S21 7 Porta. 406–407 31 Mogensen. 11. W. H. G. P. D..A. I. 9–23 12 Stehouwer. B.. K. 32S–39S 15 Kawashima. H. Yi. (2000) Increased levels of soluble vascular cell adhesion molecule 1 are associated with risk of cardiovascular mortality in type 2 diabetes: the Hoorn study. M.. M. R. A. E. A. Basciani.. A. Smits. (2001) Identification of CD146 as a component of the endothelial junction involved in the control of cell-cell cohesion. (1999) Mechanisms of endothelium-dependent relaxation in myometrial resistance vessels and their alteration in preeclampsia. (2004) Microalbuminuria and cardiovascular risk. J. 800–803 25 van den Born. K. J. (2000) Recent advances towards understanding redox mechanisms in the activation of nuclear factor κB. Opin. P. P. R. M. (1999) Microalbuminuria and peripheral arterial disease are independent predictors of cardiovascular and all-cause mortality. and Baeuerle. J. van Hinsbergh. 485–491 22 Stehouwer. 160–173 5 Goldberg. A.. and den Ottolander. C. Cardiovasc. M. V.. Res. J. J. Diabetes 49. E. H. S. insulin. 66 (Suppl. (2003) Endothelial dysfunction in obesity and insulin resistance: a road to diabetes and heart disease. Harrison. A. Med. 34.S. F. Assoc. G. A. Britten. (2002) Diabetic retinopathy: a clinical update. Mann. Clin. 1278–1289 11 De Caterina. P. D. Pollak. Nephrol. Diabetologia 43.. Kidney Dis. Forrest. J.. 49–55 20 Henry. J.. J.. Pharm. Kostense. V. (1998) Endothelial function in health and disease: new insights into the genesis of cardiovascular disease. Y. H. and chronic low-grade inflammation in type 2 diabetes: progressive. Kostense. 99–107 16 Khan. R. R. et al. Pediatr. (1997) Vessel wall heparan sulfate and transcapillary passage of albumin in experimental diabetes in the rat. Proc.. and Cooper. A. sulphonylurea. 49–56 21 Jager. R. Hackeng. M. R. J. 286. Lancet ii. North Am.. endothelial dysfunction. G. 57–71 19 Bacon. (2003) von Willebrand factor and its propeptide in children with diabetes. F. J. Diabetes Care 27 (Suppl. (2002) Increased urinary albumin excretion. (1996) Nitric oxide regulates vascular cell adhesion molecule 1 gene expression and redox-sensitive transcriptional events in human vascular endothelial cells. A. M... A. J.. Lambert. Henry. Nephrol. Masse. A. M. 93.. J. N.. 9114–9119 17 Janssen-Heininger. A. C. F. et al. Berden. M. vii 6 Kikkawa. Lutterman. Baker. 61. R.. H. Nat.. Acad. flow-mediated vasodilation in elderly individuals without and with diabetes: further evidence for a link between microalbuminuria and endothelial dysfunction: the Hoorn Study. 3677–3684 28 Flyvbjerg. 2. P.. and Mogensen. The example of albuminuria and the risk of cardiovascular disease in diabetes. (2003) Progression of diabetic nephropathy. (2000) Endothelial dysfunctions: common denominators in vascular disease. M.. and Kostense. 27–31 26 Vervoort. Free Radicals Biol... W. B. Poynter. T.. Hill. P. or metformin in patients with newly diagnosed non-insulin dependent diabetes followed for three years. especially among hypertensive subjects: five-year follow-up of the Hoorn Study. and Bandello. 310. R. 43. Med. C. and Berden. (2000) Prognostic impact of coronary vasodilator dysfunction on adverse long-term outcome of coronary heart disease. Lipidol. Takahashi. et al. W. 530–533 34 Karalliedde. van Hinsbergh. Am. Gall. et al.. S42–S44 38 Nakamura. A. T. T. 537–551 36 Gerstein. 3527–3561 14 Quyyumi. A. Onoda. J. Am. J. Warren. Engl. R. D. G. Y. Pregnancy 18. D. Health Syst. M. E. Knudsen.. A. and causal factors. Anfosso. Nijpels. J. W. death. (1997) Endothelial dysfunction and pathogenesis of diabetic angiopathy. and Haneda. P. W. Biol. 1617–1634 8 The Diabetes Control and Complications Trial Research Group (1993) The effect of intensive treatment of diabetes on the development and progression of long-term complications in insulin-dependent diabetes mellitus. Blood 91. and Viberti.. and independently associated with risk of death. R. Islington Diabetes Survey. 11. (1999) Transcapillary escape rate of albumin is increased and related to haemodynamic changes in normo-albuminuric type 1 diabetic patients. Curr. Br. Med. Ferreira. (2002) Does microalbuminuria predict diabetic nephropathy? Diabetes Care 25. M. A. J. 17. 21. J.. Blood 98.. 17... W. G. Metab. (2001) Albuminuria and risk of cardiovascular events. Dial.. M.. E. 83–88 10 Caballero. M. Ruhe. V. and Bouter. G. A. J. R. H. (1991) von Willebrand factor and development of diabetic nephropathy in IDDM. (1988) Microalbuminuria as predictor of vascular disease in non-diabetic subjects. J. D. et al. and Medford. D. E. H. S. Natl. N. 529–536 C 2005 The Biochemical Society . (1998) Endothelial cells in physiology and in the pathophysiology of vascular disorders.. and heart failure in diabetic and nondiabetic individuals. J. 1157–1165 23 Schachinger.. S. C. 919–925 39 Verrotti. L. Atherosclerosis 174. N. and Chiarelli. Res. M. (2005) Endothelial cell dysfunction in systemic vasculitis: new developments and therapeutic prospects. F.. (2002) Epidemiologic analyses of risk factors. Itai. A. 92). U. JAMA. 19. Campbell. 105. K.. (2004) Microalbuminuria is associated with impaired brachial artery.. E. N. Viberti. Olbrych. R. (2004) The two faces of endothelial nitric oxide synthase in the pathophysiology of atherosclerosis. White.. P. S. 2). et al.. Morgese. Chaturvedi. flow-mediated dilation. Hypertens. Transplant.. Koya. Bakker. H. (2004) Type 2 diabetes is associated with impaired endotheliumdependent.. et al. Vasc. Setter. Sci. and Jackson. M. Rheumatol. C. Greco. C. E. (2004) Association between serum C-reactive protein levels and microalbuminuria: a population-based cross-sectional study in northern Iwate. J.. Dial. Endocrinol. G. 41. Med. 21. Kidney Int. Diabetes 51. M. risk markers. D. Am. 1205–1223 29 Molitch. R. Obes. 53. Diabetic Med. Alexander. J. C. and Wetzels. J.. J. J. H. Japan. Endothelium 11. I. Med. Diabetes 40. P.. DeFronzo. and Parving. Donker. R.. (2001) Prognostic implications of retinopathy and a high plasma von Willebrand factor concentration in type 2 diabetic subjects with microalbuminuria. A. J. J..

Diabetes Care 23. (1997) Vascular effects of acute hyperglycemia in humans are reversed by L-arginine. Rich. hypertension... C.. Thromb. B. H. Arterioscler. 287. van Dijk. G. 1515–1522 64 Coggins.. Hypertension 38.. H. Gansevoort. Lindner. (Suppl. Dial. W. L. ter Maaten.. Gerritsen. Res. and Baron.. 22. J. J. Lindner.. Wetzels. C. 137–147 53 Huvers. J. Vasc. Pratley. ter Maaten. H. Crowder. (2001) Endothelium-dependent and independent vasodilation studies at normoglycaemia in type I diabetes mellitus with and without microalbuminuria. Diabetologia 43. 233–242 C 2005 The Biochemical Society . P. Diabetologia 44. Engl. Coppack. K. H. Roberts. Fioretto. Circulation 95.. C.. and Colhoun. Vasc. J.. D.. J. (2000) Type II diabetes abrogates sex differences in endothelial function in premenopausal women. R. J. and de Jong. et al. C.. Diabetes 53... A.. (2001) Impaired autonomic function is associated with increased mortality. D. W. A. 3071–3078 43 Verhave. and Smits. 19. (2002) Blood pressure and insulin resistance: role for microvascular function? Cardiovasc. Galecki.. Physiol. D.. Diabetes 49. P. J. P. 49. De Leeuw. Stehouwer 42 Jager. H. J. R. Diabetes 46 (Suppl. K. V. Cardiovasc. V. Biol. R. A.. (2000) Free fatty acid elevation impairs insulin-mediated vasodilation and nitric oxide production. Stehouwer. van Hinsbergh. G. lipid uptake. 1300–1307 54 Vervoort.. Praet. 972–978 76 Weyer. Lutterman. Stehouwer.. (1998) How heterogeneous is microalbuminuria in diabetes mellitus? The case for ‘benign’ and ‘malignant’ microalbuminuria. ter Maaten. H. (1999) Atherosclerosis: an inflammatory disease. Obes. C. W. A. Thai. S. Vasc. (2004) Vasoconstrictor effects of insulin in skeletal muscle arterioles are mediated by ERK1/2 activation in endothelium. and endothelial dysfunction: a potential role for cytokines originating from adipose tissue? Arterioscler. J.. Hillege. et al. J. A.. M. I. 115–126 47 Smulders. M. J. 326–332 50 Dogra. R. (2000) Cardiovascular autonomic function is associated with (micro-)albuminuria in elderly Caucasian subjects with impaired glucose tolerance or type 2 diabetes: the Hoorn Study. Halter. S18–S21 44 Jager. Metab. D.. J. 19. Eckel. Circulation 99. TenVoorde.. and Yudkin.. (2002) C-reactive protein and soluble vascular cell adhesion molecule-1 are associated with elevated urinary albumin excretion but do not explain its link with cardiovascular risk. Stanton. (2004) Microvascular recruitment is an early insulin effect that regulates skeletal muscle glucose uptake in vivo. C. D.. Transplant... W. C. and 5-year mortality in diabetic and nondiabetic subjects: the Hoorn Study.. P. Spek. Nieuw Amerongen. ter Wee. G.. and Stehouwer. Diabetologia 45. O. 2682–2690 65 Steinberg. et al. J. J. Diabetes 51. R. J. K. A. (2000) Nitric oxide and vascular responses in Type I diabetes. IJzerman. C... O. T. S. Stehouwer. 1585–1589 58 Steinberg. J.. Schalkwijk. Am. P.. IJzerman. Diabetes 49. E.. Atherosclerosis 161. E... 896–902 72 Merkel. T. N. 3). J. G... G. R. 13. D. (1998) Specific impairment of endothelium-dependent vasodilation in subjects with type 2 diabetes independent of obesity. D. Emeis. M. and Tataranni. (2000) Angiotensin II type 1 receptor gene polymorphism and the response to hyperglycemia in early type 1 diabetes. J. et al. Diabetes Care 24. Circulation 101. Thromb. Nephrol.. Yudkin.. Paradisi. and Stehouwer. H.. D. Donker. (2002) Direct evidence for insulin-induced capillary recruitment in skin of healthy subjects during physiological hyperinsulinemia. 857–862 56 Stehouwer. De Zeeuw. J. 27 (Suppl. E. G. C-reactive protein. S. Diabetes 48. D. Kostense. A.. D. and Watts. S9–S13 62 Serne. Heart Circ. (2002) Lipid infusion impairs physiologic insulin-mediated capillary recruitment and muscle glucose uptake in vivo. (2002) Humoral markers of inflammation and endothelial dysfunction in relation to adiposity and in vivo insulin action in Pima Indians. 233–236 61 Pinkney.... Gans. J. M. 1997–2006 73 Eringa. 593–601 51 Giugliano. E. D. 340.. Med. P. C.. et al. G. N. E. plasma markers of endothelial function. Berden.. (2001) Impaired skin capillary recruitment in essential hypertension is caused by both functional and structural capillary rarefaction. et al. et al. A. O. J. C.. D. Diabetes 51. or a history of cardiovascular disease: the Hoorn Study. M. 593–598 45 Schalkwijk. P.156 C. 1783–1790 52 Chan. J. F. G. C.. Stehouwer. Houben. J. Stehouwer. G. D. (2003) Adipose tissue.. and Scholey. Tangelder. insulin resistance and fatty acids. (2000) Nitric oxide synthesis and isoprostane production in subjects with type 1 diabetes and normal urinary albumin excretion. 418–419 63 Serne. Forte. Int. R..... J. Biol. Mauer. L. O. L. J. (1999) Endothelium-dependent vasodilatation. and Baron. (1999) Plasma concentration of C-reactive protein is increased in type I diabetic patients without clinical macroangiopathy and correlates with markers of endothelial dysfunction: evidence for chronic inflammation. (2002) Vascular function. J. J. Res. J. Yudkin. A. Westerhof. van Hinsbergh. N. S25–S28 75 Yudkin. Cronin. Dekker. H. A. C. Evidence for reduced availability of nitric oxide during hyperglycemia. Clerk.. J. Arterioscler. D. W. S. Hypertension 34. 1369–1374 48 Gerritsen. insulin action and vascular disease: inflammatory signals. M. Dial.. Greene. S. Ouwehand. S. Poland. de Jongh. Pitt. L. Hook. Bakker. 1793–1798 49 van Ittersum. 1231–1238 67 Vincent. G. C.. Disord. van Doorn. and Stehouwer. Rattigan. D. G. Cronin. Gans. 92). H. A. P. A. Relat. 1418–1423 68 Clerk. M. B. Diabetes 50. Clin. Lipid Res. G. et al. O. et al. C. and Kostense. F.. J. 2751–2754 57 Miller. Diabetologia 41. C. E. Vallance. 83. (2001) Capillary recruitment is impaired in essential hypertension and relates to insulin’s metabolic and vascular actions. Gans.. J. Metab. A. 1138–1145 69 Serne. (1999) Microvascular function relates to insulin sensitivity and blood pressure in normal subjects. C. Y. Jager.. J. P. 55. R. J. (1998) Heterogeneous nature of microalbuminuria in NIDDM: studies of endothelial function and renal structure. J. I. and Coppack. H.. et al. 13. 351–357 46 Ross. and adrenergic vasoconstrictor responses in type 1 diabetes under near-normoglycemic conditions. 1946–1952 60 Fioretto. H. H. R... J.. H. P.... S.. et al. 238–242 70 Serne. B. J. G. H. and Sipkema. O. 2040–2046 59 Hogikyan. (2004). L. D. 2). (2002) Lipoprotein lipase: genetics. P.. R. et al. L.. Endocrinol. and regulation. Thromb. Kidney Int. S.. insulin resistance. W. (1999) von Willebrand factor. Transplant. et al.. H2043–H2048 74 Yudkin. C. M. J. (1999) Elevated skeletal muscle blood flow in noncomplicated type 1 diabetes mellitus: role of nitric oxide and sympathetic tone.. C. J.. 161–168 71 Serne. G. Stehouwer.. L. 1080–1085 55 O’Byrne. S. J. (1997) Endothelial dysfunction: cause of the insulin resistance syndrome. H. Paradisi. Rattigan. Donker. C.. E. Biol. 623–634 66 Steinberg. V.. J. R. (1999) C-reactive protein in healthy subjects: associations with obesity. Marfella. (2001) Physiologic hyperinsulinemia enhances human skeletal muscle perfusion by capillary recruitment. and Supiano. J. Nephrol.. M. J. An elevated urinary albumin excretion predicts de novo development of renal function impairment in the general population. W. J. P. D. Coppola.. J. F. R. especially in subjects with diabetes. F. and Nosadini. G. M. T. and Goldberg. and Clark. P. Schalkwijk and C. et al. R. S.. (1998) Ambulatory blood pressures and autonomic nervous function in normoalbuminuric type I diabetic patients. A. A. C.. S. Physiol. 43. et al. L. A. Diabetologia 42. Diabetes 49. E. R. N..

F. association with diabetic nephropathy. D.. 3). Devaraj. Invest. I.. Kidney Int. Med. J. H. J. (1996) Amelioration of vascular dysfunctions in diabetic rats by an oral PKCβ inhibitor. W. et al. Stehouwer. Diabetes 45 (Suppl.. 221–229 86 Maiello. V. K. S. J. 1. 19. polyol metabolism. Inoguchi.. and endothelial dysfunction in the streptozotocin-induced diabetic rats vasculature. 728–731 100 Nerlich. Gray. Y. 1400–1402 107 Stern. 80. Clin. Edelstein... J. I. P.. Eur. E708–E717 85 Graier. and Xia. and Beilin. M. U.. S. Biochem.. 9. M. 324. Edelstein. 1497–1503 94 Chen. M. L. M. Yagihashi. G. (1975) Hyperglycemia. J. Diabetes Care 25.. Asnaghi. T. H. Diabetes 41. T.. D. Biophys. 212–218 84 McGinn. 1341–1348 103 Thorpe. T. Wagner. G. A. Z. (2002) α-Tocopherol decreases superoxide anion release in human monocytes under hyperglycemic conditions via inhibition of protein kinase C-α. H. A. 97. Ligtvoet. J.. P. Ageing Res. 735–738 83 Boeri. X. M. L.. Wagner. A. et al.. Mauer. Metab. S. Patel. M. Pettermann. M. D.. J. Angulo. P. and Lorenzi. D. 1204–1209 89 Gabbay. (2002) Receptor for advanced glycation endproducts (RAGE) and the complications of diabetes. (2001) Advanced glycation end-products: a review. Neurobiol. S. Transplant. B. (1992) Increased expression of tissue plasminogen activator and its inhibitor and reduced fibrinolytic potential of human endothelial cells cultured in elevated glucose. (1998) Overexpression of glyoxalase-I in bovine endothelial cells inhibits intracellular advanced glycation endproduct formation and prevents hyperglycemia-induced increases in macromolecular endocytosis... Trans. S. 2404–2411 93 Williams. Cirina. et al. P.. S. F. Amino Acids 25. Cagliero. (2004) AMPK inhibits fatty acid-induced increases in NF-κB transactivation in cultured human umbilical vein endothelial cells. and Lorenzi. Diabetes 46. Biochem.. Diabetes 51.. 945–959 96 Venugopal. 31. Howell. Conti. Kolm-Litty. Biochem. Du. (2003) High glucose-mediated effects on endothelial cell proliferation occur via p38 MAP kinase. Boeri. (2003) Benfotiamine blocks three major pathways of hyperglycemic damage and prevents experimental diabetic retinopathy. D. Barden. Edelstein. Shiba. S. D.. Arch. W. Yan. (2001) Protein kinase C activation: isozyme-specific effects on metabolism and cardiovascular complications in diabetes. Trans. 77). 659–673 98 King. Keaney. Med... et al. D. M.. Sci. and Schmidt. Biophys. Clin. (2001) Protein kinase C and the development of diabetic vascular complications. M. B. (2003) α-Oxoaldehyde metabolism and diabetic complications.. Sci. and Baynes. P. M. and Ido. Invest. Int. et al. R. N. et al.. H. M. (2004) Studies of rat and human retinas predict a role for the polyol pathway in human diabetic retinopathy. Rev. J. M. and Jialal.. 294–299 79 Singh. (2000) Interaction of endothelin-1 with vasoactive factors in mediating glucose-induced increased permeability in endothelial cells. et al. Dittrich. L. S.. G. Wascher. M. P.. Soc. Thornalley.. 2446–2453 112 Chen. Sauer. Natl. T. T. M. 2320–2327 111 Schalkwijk. 106. (2004) Amadoriconfigurated albumin induces nitric oxide-dependent apoptosis of endothelial cells: a possible mechanism of diabetic vasculopathy. S. 813–820 78 Hammes. 53–60 109 Rodriguez-Manas. H... Grillari. (2000) Amadori-glycated albumin in diabetic nephropathy: pathophysiologic connections. C. Nephrol.. Clin. Diabetes 44. Saad. 58 (Suppl. Biochem. M. K. R. Biochem. S. L. Clin. S. and Waldhausl. J. and Szwergold. J. Ruderman. (1996) Biochemical and molecular mechanisms in the development of diabetic vascular complications. Dimmeler. Nishio. 419. 18. L.. M. J. E. H. D. 801–813 92 Dagher. J.. Amadori albumin.. Katai. Vallejo.... Clin. (2002) Circulating and urinary transforming growth factor β1. Nature (London) 414. 170–178 101 Du. 50. 1–15 108 Amore. Rossetti. G. S105–S108 99 Ishii.. R. 80–88 114 Thornalley. J. Invest. M. M. F. U. J. A. Chang. K. C.. Diabetes 53. N. et al. D. L. Diabetes 47. 108. M. Arch. S. 1311–1321 95 Way. Science 272. Hoehn. 26. J. N. 12222–12226 102 Du.. Maiello. Pharmacol. Koch. B.. Commun. Invest. 419. 101. 556–566 110 Chaturvedi. 453–458 81 Baumgartner-Parzer. G. 108. (1993) Hyperglycemic pseudohypoxia and diabetic complications. Y. S. H. H. J. I. and Pollock. Rao. (2001) Hyperglycemia inhibits endothelial nitric oxide synthase activity by posttranslational modification at the Akt site.. Biophys.. Twaalfhoven. 31. (2002) Polyol pathway and diabetic peripheral neuropathy. Diabetologia 44. Proc. G. D. Cherian. and complications of diabetes mellitus. C. M. I. Yang.. Roy. S... L. et al. Koya.. 275–281 104 Beisswenger. Endocrinol. J.. 31–40 C 2005 The Biochemical Society . S. 129–146 80 Ginsberg. Diabetes 42. N. oxidative stress. (1998) Expression of glutamine:fructose-6-phosphate amidotransferase in human tissues: evidence for high variability and distinct regulation in diabetes..Vascular complications in diabetes mellitus: the role of endothelial dysfunction 157 77 Brownlee. J. van Hinsbergh. and Stehouwer. W.. (2003) Use of aminoguanidine (Pimagedine) to prevent the formation of advanced glycation endproducts. Diabetologia 44.. (2003) Blockade of receptor for advanced glycation endproducts: a new target for therapeutic intervention in diabetic complications and inflammatory disorders. Annu. 1358–1363 105 Shinohara. I. (2003) Role of methylglyoxal adducts in the development of vascular complications in diabetes mellitus. Boeri.. C. (2000) Insulin resistance and cardiovascular disease. G. D. S. and Brownlee. Schalkwijk. Cohen. S. L. and Donnelly. S.. Diabetologia 46. K. Podesta. M. Mori.. B... 3049–3054 97 Idris. L. Am. V. Maiello. Yan.. C. Nat. F. M. and Schleicher. N. G. and Chakrabarti.. C. Jr. C.. M. C. Ju. N. Y. 294. R. Diabetic Med. E. E. R. L.. Fuller. Gallacher. 82. E... Gerhardinger. et al. S40–S44 113 Hudson. Diabetes 38. X. P.. and complications of type 1 diabetes: the EURODIAB prospective complications study. D. and Kostner. P... (2003) Maillard reaction products in tissue proteins: new products and new perspectives.. Rev.. 325–392 91 Williamson.. (1988) Increased expression of basement membrane components in human endothelial cells cultured in high glucose. Physiol. Bucciarelli. V.. G. C. Dial.. (1995) Intracellular mechanism of high d-glucose-induced modulation of vascular cell proliferation. 205–213 88 Cacicedo. 285. D. Q. N. (2000) Hyperglycemia-induced mitochondrial superoxide overproduction activates the hexosamine pathway and induces plasminogen activator inhibitor-1 expression by increasing Sp1 glycosylation. Apostolova.. Giardino. F. 521–536 90 Oates. Invest. G. C. Nickel. E. T. B. 1323–1327 82 Cagliero. P.. Abrahamian.. F. Frangos. (1997) Glucose-induced protein kinase C activation regulates vascular permeability factor mRNA expression and peptide production by human vascular smooth muscle cells in vitro. Gessl. Jirousek.. M. and Weis. G. T. Poronnik. 1142–1147 106 Bourajjaj. and Ziyadeh. (1989) Modification of tissue-factor mRNA and protein response to thrombin and interleukin 1 by high glucose in cultured human endothelial cells.S. Soc. A. Res. P. Sui. R. Almus. Park. S. Wendt. 1009–1015 87 Kofler. Grubenthal. (1999) Amadori albumin in type 1 diabetic patients: correlation with markers of endothelial function... A. V. and localization in retinal capillaries. Lab. (2003) Early and intermediate Amadori glycosylation adducts. and Schalkwijk. and King. Rev. K. and Orme. X. T. Diabetes 48.A. Kunisaki. and Lorenzi. (1995) High-glucose–triggered apoptosis in cultured endothelial cells. W. M.. Nelson. (2001) Biochemistry and molecular cell biology of diabetic complications. (2005) The role of cytokines in cardiovascular diseases: focus on endothelial response to inflammation. Acad.

R.. and Border. (2003) Diabetic nephropathy and transforming growth factor-β: transforming our view of glomerulosclerosis and fibrosis build-up. (2002) Effects of vitamin E on cardiovascular and microvascular outcomes in high-risk patients with diabetes: results of the HOPE study and MICRO-HOPE substudy. A.. et al. F.. and Ziyadeh.. Trends Biochem. et al. Clin. Physiol. T. T. W. 19. and Schleicher. Metab. E. S. (2004) Nuclear factor κB: a potential therapeutic target in atherosclerosis and thrombosis. Diabetes Care 27.. A. C.A. B. L.. Santilli. M. M. J. Trans. and Runge. H. 1919–1927 130 Economides. Shimizu. Khaodhiar. J. L. V. A. Vries-Knoppert. Guo. M. S. E. Regul. (2003) Inhibition of GAPDH activity by poly(ADP-ribose) polymerase activates three major pathways of hyperglycemic damage in endothelial cells. Gastaldi. Clin. 805–815 147 Hogeboom. Pathol.. Devaraj. Semin. P.. and Klein. Kidney Int. (2003) Primary prevention of cardiovascular events with low-dose aspirin and vitamin E in type 2 diabetic patients: results of the Primary Prevention Project (PPP) trial.. Nephrol. 23. R1014–R1030 121 Christ. S. Jin. (2004) C-reactive protein: risk marker or mediator in atherothrombosis? Hypertension 44. A. (2000) Molecular mechanism of diabetic nephropathy.. Cardiovasc. et al. (2003) Prevention of incipient diabetic nephropathy by high-dose thiamine and benfotiamine. (2003) Enhanced lipid peroxidation and platelet activation in the early phase of type 1 diabetes mellitus: role of interleukin-6 and disease duration. VEGF-A.. C.. F. Avery. Am.. 61. 532–543 134 Choi. J. Matsumoto. III. M. Collins. 22–28 128 Sacco. 204–211 131 Ziyadeh. and Shah. 29–38 127 Ting. N. et al.. Res. 1480–1487 146 Poulaki. Dixelius. Biol.. S55–S57 132 Frank. Liebetrau. Reichert-Thoen. Circulation 99. R. et al. N. Ferris. J.. Rev. and Thorpe. Chim. 43–53 123 Marra. Acta 297. (2003) Clinical potential of advanced glycation end-product inhibitors in diabetes mellitus. Festuccia.. Invest 97. M. M. F. C. M. Clin. Exp. T. M. 200. R. A. Sci. Med... 94. P. Natl. A. Stehouwer 115 Mene. N. 1–9 119 Guzik. Acad. P. 3264–3272 129 Lonn. R.. Circulation 105. and Venugopal. R. Diabetes Care 26. Engl. A. X.. (2001) The key role of the transforming growth factor-β system in the pathogenesis of diabetic nephropathy... et al. P. and Ridker. J. N. (1999) Role of oxidative stress in diabetic complications: a new perspective on an old paradigm. Arterioscler. S.. Sci. L. A. Nephrol. 2648–2652 122 Sindhu. N. 1171–1177 143 Cross. 442–455 150 Bates. (2005) The effect of vitamin E on endothelial function of microand macrocirculation and left ventricular function in type 1 and type 2 diabetic patients. (1994) Vascular endothelial growth factor in ocular fluid of patients with diabetic retinopathy and other retinal disorders. Cardiovasc.. G. and Wehling. M.. Matsumura.. et al. S. (2004) Diabetic retinopathy.. A. catalase and glutathione peroxidase in diabetes: response to insulin and antioxidant therapies. Cotroneo. 48–58 133 Chen. A. (1996) Vitamin C improves endothelium-dependent vasodilation in patients with non-insulin-dependent diabetes mellitus.. 581–597 151 Monaco. 522–530 139 Park. K. Ruoslahti. B. 1). Renal Failure 23. Hypertens. N. R. D. P. B. (2002) Acute intensive insulin therapy exacerbates diabetic bloodretinal barrier breakdown via hypoxia-inducible factor-1α and VEGF. 2540–2553 145 Aiello. M. 203–209 148 Eyries. R.. J. Transplant.. van Hinsbergh. Soc. I.. (2002) Regulation of microvascular permeability by vascular endothelial growth factors. W. Circulation 109. I. E. S. F. Physiol. Diabetes Care 25. T. S. S. J. (2005) Oxidative stress and vascular disease. and Thornalley.. F. F. 1049–1057 117 Babaei-Jadidi. Hillman. 1042–1046 153 Willerson.. Neal. et al. and Claesson-Welsh. S. (1996) Neutralization of TGF-β by anti-TGF-β antibody attenuates kidney hypertrophy and the enhanced extracellular matrix gene expression in STZ-induced diabetic mice. U. 601–608 141 Benjamin. W. J... Creager. 1814–1818 136 Chen. et al. Heck. O. G. V. Exp. J. K.. Diabetologia 45.. W. S. T. Pitocco. D. Boles. Diabetes 54.. D. Soc. 350. Joussen. F. B. R. 26. 224–229 126 Madamanchi. J. (2002) Mechanisms of increased vascular superoxide production in human diabetes mellitus: role of NAD(P)H oxidase and endothelial nitric oxide synthase. (2004) Essential role for Smad3 in regulating MCP-1 expression and vascular inflammation. E.. Ahmed. Invest 112. Engl. S. M.. Med. and Tangelder. Noble. P. 31. Diabetes 45. (2004) Diabetic retinopathy. N. Ciabattoni. W. et al. Pellegrini. L. 15. Vasc. Hong. and diabetic complications. F. and Creager. J. Casaretto. I. J. P.. (1989) Increased levels of acute-phase serum proteins in diabetes. Diabetes Care 25. A. M. J. G.. v. J. N. Battah. (2002) Angiotensin converting enzyme inhibiting therapy is associated with lower vitreous vascular endothelial growth factor concentrations in patients with proliferative diabetic retinopathy. Caselli. N. D. J. J. Edelstein. Am.. G. S53–S58 135 Yamamoto. Anat. R.. Proc. Jim. F. C. Nephrol. 90.. (2002) Early increase of oxidative stress and reduced antioxidant defenses in patients with uncomplicated type 1 diabetes: a case for gender difference. L. W. J. Diabetes 52..S. S.. K. Endothelium 11. Integr. N. P.. (2003) VEGF signalling: integration and multitasking in endothelial cell biology. Koo. F. Williams. Karachalias. T. L. 133–139 149 Caldwell.. Behzadian. and Paleolog. (2000) TGF-β 1 down-regulates inflammatory cytokine-induced VCAM-1 expression in cultured human glomerular endothelial cells. E. 3199–3203 125 Davi. (2004) Mediators of diabetic renal disease: the case for TGF-β as the major mediator.. 6–11 C 2005 The Biochemical Society . Aiello. T. K. F. Invest 109. Circulation 107. Diabetes 48. J. H.. Mussa. Res. C. 596–604 140 Feinberg. Roberts.. K.. 77). Clin. Lebedeva. et al. R. M. L. and Nicolucci. D.. J. S. (2000) Mechanism of transforming growth factor-β1 signaling. Iglesias-de la Cruz. Isono. K. M. Diabetes Metab. and Ziyadeh. A.158 C. Hoogwerf. 287. and Pugliese.. M. G... K. Tognoni. Yang. Comp. M. 488–494 144 Fong. Am. Y. N. S. (2004) Modulation of growth factor gene expression in vascular cells by oxidative stress. Clin. J. 1656–1662 120 Li.. Bartoli. (2004) Inflammation as a cardiovascular risk factor. J.. N.. 158. Res. and Khachigian. S. Thromb. Consoli. W. (2003) VEGF-receptor signal transduction. S. 25. Yusuf. Qin. F. et al. Nakamura. L. K. J. A. Gunther.. II2–II10 154 Jialal. A. D. Avanzini. M.. Circ. (2004) Dysregulation of hepatic superoxide dismutase. J. 28... Chiarelli. 15 (Suppl.. 1181–1184 142 Zachary. Clin. 331. Polak. 671–682 152 McMillan. 315–320 116 Du. Schalkwijk and C. Timimi. C. Ganz. T. R. 58 (Suppl. 471–481 137 Lehmann. C. Lee. D. (2001) Glucose. 2110–2120 118 Baynes. A.. E. (2004) Endothelial cell superoxide generation: regulation and relevance for cardiovascular pathophysiology. Am. Vendrov. 38. (2002) Glucose increases endothelial-dependent superoxide formation in coronary arteries by NAD(P)H oxidase activation: attenuation by the 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor atorvastatin. M. Arrigg. M. and Pocock. W. B. E. K. D. (1999) In vivo formation of 8-iso-prostaglandin F2α and platelet activation in diabetes mellitus: effects of improved metabolic control and vitamin E supplementation. N. G. 135–144 138 Sharma.. and Ziyadeh. M. W. 370–375 124 Davi. Bauersachs.. Drugs 3. (2003) Vascular endothelial growth factor and diabetic retinopathy: pathophysiological mechanisms and treatment perspectives. P. J. Diabetes 51.. Dial. Biochem. (1993) Expression of transforming growth factor β is elevated in human and experimental diabetic nephropathy. G. M. G. and Vaziri. D.. B. Roncaglioni. M.

45021–45026 185 Yokota. et al. A. Endocrinol. S. Arita. 180–189 159 Chakrabarti. adiponectin. I.Vascular complications in diabetes mellitus: the role of endothelial dysfunction 159 155 Venugopal. Metab. Penno.. M.. R. Dekker. Cardiol. Evans. I. 1665–1666 181 Chandran. G.. Funahashi. et al.. and Henry. I. (1997) Determinants of progression of microalbuminuria in patients with NIDDM. S. Curr. et al.. Navis. H. 733–741 174 de Jongh. A.. Med. Khan.. et al. Biochem. Kidney Dis. 20. Y. J. 315. 1930–1935 178 Hotta. Nature (London) 399. 10.. B. L. et al. G. 323–329 167 Chaturvedi. Vasc. E.. 60. J. Fisslthaler. S. 997–1000 170 Manson. I. J. Gorzelniak. even in lean subjects. T. (2002) Hyperlipidemia in kidney disease: causes and consequences. IJzerman. 1320–1325 172 Kenchaiah. Commun. Makri. Arita. J.. in patients with type 1 diabetes.. J.. M. C.. Curr. and risk of type 2 diabetes in the Pima Indian. L. Colditz. lipids. (2001) Hypoadiponectinemia in obesity and type 2 diabetes: close association with insulin resistance and hyperinsulinemia. K. Zhang. in type 2 diabetic patients. 41. J. and Zeiher.. and the future. Kilcoyne. T. Bandinelli. Stehouwer. (2002) Hypertension. 98. Serne. G. (2003) Inflammatory markers. (2004) C-peptide: new findings and therapeutic implications in diabetes. J. Circulation 103. J.. A. Int. Med.. Y. in obesity. J. F. Moll. 2529–2535 175 de Jongh. 305–313 173 Pinto-Sietsma. (2004) The prospective association between adiponectin and coronary artery disease among individuals with type 1 diabetes. 14. et al.. G. 41–48 Received 19 January 2005/15 March 2005. T. S. 91–96 160 Stas. Diabetes Care 20.... (2002) Diabetic dyslipidemia. 999–1005 169 Wilding. Kihara. 820–825 157 Dimmeler. 346. proatherogenic role. M. M. Biol. S. Kihara. T. D. A. and obesity are associated with retinopathy: the Hoorn study. R. R. 387–391 166 Sahadevan.. 5. (1999) Activation of nitric oxide synthase in endothelial cells by Akt-dependent phosphorylation.. Weijers. (2002) Nitric oxide synthase and hypertension. S. 221–225 161 Boulbou. Zgibor. Diabetes Care 26. M. B. Hypertens.. 882–889 171 van Leiden. N. S.. and de Jong. S. C. (1999) Paradoxical decrease of an adipose-specific protein. Funahashi.. accepted 11 May 2005 Published on the Internet 25 July 2005. Slaats. A. 219–227 168 Smulders. N. 1999–2001 164 Taskinen. Koukoulis. D. Busse. K. R. J. Biophys. and Rees. 23.. E. A. Diabetes 52.. Engl. C. M. J. 33–37 156 Cardillo. Janssen. and Jialal. T. R. H. R. Endocr. Chem. D. Nephrol. Exp. S. (2004) Pathogenesis of hypertension in diabetes. Xu. S.1042/CS20050025 C 2005 The Biochemical Society . The Pittsburgh Epidemiology of Diabetes Complications Study. Evans. Funahashi. Montagnani.. A. 1595–1599 179 Kumada. N.. S. Serne. El Atat. DOI 10.. De Vries. et al. et al. Endocrinol. A. J. (2005) Circulating adhesion molecules levels in type 2 diabetes mellitus and hypertension. C. K.. J. 11. et al.. et al. S. 257. (2001) Microalbuminuria in type 1 diabetes: rates. S. De Zeeuw. M. adiponectin. Clin. 86. M. W. S. Funahashi. Ohishi. risk factors and glycemic threshold. Oritani. J. Y.. et al. P. Thromb. N.. W. and Kasiske. adiponectin. D. and microangiopathy. C. Med. 47–51 165 Evans. D. 231–234 187 Tan. A. E.. N. Disord. Nephrol.. 322. Kihara. Diabetes Care 25.. 347. E. Nambi.. Engl. Biol. Circulation 100. R. T. S.. M. Engl. W. Med.. and insulin resistance. 278. J. N. et al... R.. Fleming. Circulation 109. M.. R. Rakic. E. 2873–2882 176 Arita. (1990) A prospective study of obesity and risk of coronary heart disease in women. (2003) Association of hypoadiponectinemia with impaired vasoreactivity. 1057–1063 184 Chen.. adiponectin. 39–44 162 Leclercq.. I. N. (1999) Insulin stimulates both endothelin and nitric oxide activity in the human forearm. Diabetes 53.. Metab. Diabetes Care 25.. Gans. R. G. H.. Kihara.. Mangili.. et al. Ouchi.. R. D. N. 185–189 163 Sowers. and Germenis. C. Nephrol. A. Devaraj. Jaimes. J. Diabetologia 48. S. medicine. Opin. Rev. Hypertension 42.. Opin.. S. A. 3. Tanaka. Imaging 24. H. (2000) Adiponectin. Biol. M. 1745–1751 189 Engeli. (2004) Hypoadiponectinemia is associated with impaired endothelium-dependent vasodilation. and Silberbusch. 2473–2476 183 Ouchi. G. Takahashi. T. K. E.. Clin. Kidney Int. J.. C. N. Opin. T. Ijzerman. a new member of the family of soluble defense collagens. Opin. M. et al. Stehouwer. Cukiernik. Y. M. E. Blood 96. Y. Arterioscler.. B. A prospective study.. (2003) Adiponectin stimulates production of nitric oxide in vascular endothelial cells.. (1997) Science. H. C. N. et al. Gourgoulianis. H.. Thromb.. (2002) Blood pressure. 1723–1732 186 Ouchi. G. Diabesity Res. (2002) Elevated serum concentration of adipose-derived factor. Feldpausch. 11. Funahashi. (2003) Association between adiponectin and mediators of inflammation in obese women. and Stehouwer. S. (2004) Free fatty acid levels modulate microvascular function: relevance for obesity-associated insulin resistance. A. W. Res. T.. (2001) Adipocytederived plasma protein. Z. M. Clin. 601–605 158 Wahren. Khan. R. Levy. hypertension. E. Stampfer. (2003) Association of hypoadiponectinemia with coronary artery disease in men. N. suppresses lipid accumulation and class A scavenger receptor expression in human monocyte-derived macrophages. Funct. N. E. K. E.. and Quon. A. R. adipose-specific protein. 79–83 177 Weyer. 765–769 188 Krakoff. and Stehouwer. Phillips.. (2002) Obesity and the risk of heart failure. Vasc. 942–947 190 Costacou. C. Arterioscler. Chow. De Vries. A. J. 85–89 180 Imagawa. C. et al. Lipidol. T. Hypertens. hypertension. (1999) Novel modulator for endothelial adhesion molecules: adipocyte-derived plasma protein adiponectin. Am. (2003) Adiponectin: more than just another fat cell hormone? Diabetes Care 26. (2004) C-peptide and retinal microangiopathy in diabetes. 5. and oxidative stress. C. Arita. negatively regulates the growth of myelomonocytic progenitors and the functions of macrophages. Obesity treatment... Shimomura. Atherosclerosis Suppl. G. Circulation 100. et al. Petinaki. Hypertens. (2003) A central body fat distribution is related to renal function impairment.. T. K. Curr. R. adiponectin.. R. J. Kihara.. S. S.. J. and Sima.. (2000) Plasma concentrations of a novel. Metab 89. S.. (1999) Diabetic dyslipidaemia and coronary heart disease: new perspectives. and Sowers. 2442–2450 182 Ouchi. Nakamura. J. Physiol. M.. C. angiotensin II. and Raij. (2004) Impaired microvascular function in obesity: implications for obesity-associated microangiopathy. O. M. Hermann.. J. Curr. J. and Fuller. Br. (2005) Effect of C-reactive protein on vascular cells: evidence for a proinflammatory. Ciaraldi. M. M. A. Rottiers. D... Sumitsuji.