The regulation of inflammation by oxidized phospholipids.

During inflammation or under conditions of oxidative stress, the polyunsaturated fatty acid side chains of phospholipids in cellular membranes or lipoproteins can be oxidatively modified. This process generates a complex mixture of structurally diverse oxidized phospholipid (OxPL) species, each of which may exert distinct biological effects. The presence of OxPLs has been documented in acute and chronic microbial infections, metabolic disorders, and degenerative diseases. It is now well recognized that OxPLs actively influence biological processes and contribute to the induction and resolution of inflammation. While many pro- and anti-inflammatory effects have been documented for bulk OxPL preparations, we are only beginning to understand the exact molecular mechanisms and signaling events that mediate the individual proinflammatory or anti-inflammatory bioactivities of discrete isolated OxPL species. Here, we review the current knowledge on the regulation of inflammation by OxPLs and summarize recent studies that establish cyclopentenone-containing OxPLs as a category of potent anti-inflammatory lipid mediators.

A Bayesian order-restricted model for hormonal dynamics during menstrual cycles of healthy women.

We propose a Bayesian framework for analyzing multivariate linear mixed effect models with linear constraints on the fixed effect parameters. The procedure can incorporate both firm and soft restrictions on the parameters and Bayesian model selection for the random effects. The framework is used to analyze data from the BioCycle study. One of the main objectives of the BioCycle study is to investigate the association between markers of oxidative stress and hormone levels during menstrual cycles of healthy women. Contrary to the popular belief that ovarian hormones are negatively associated with level of F (2) -isoprostanes, a known marker for oxidative stress, our analysis finds a positive association between ovarian hormone levels and isoprostane levels. The positive association corroborates the findings from a previous analysis of the BioCycle data.

The fatty acid oxidation product 15-A3t-isoprostane is a potent inhibitor of NFκB transcription and macrophage transformation.

Fatty acids such as eicosapentaenoic acid (EPA) have been shown to be beneficial for neurological function and human health. It is widely thought that oxidation products of EPA are responsible for biological activity, although the specific EPA peroxidation product(s) which exert these responses have not yet been identified. In this work we provide the first evidence that the synthesized representative cyclopentenone IsoP, 15-A(3t)-IsoP, serves as a potent inhibitor of lipopolysaccharide-stimulated macrophage activation. The anti-inflammatory activities of 15-A(3t)-IsoP were observed in response not only to lipopolysaccharide, but also to tumor necrosis factor alpha and IL-1b stimulation. Subsequently, this response blocked the ability of these compounds to stimulate nuclear factor kappa b (NFκB) activation and production of proinflammatory cytokines. The bioactivity of 15-A(3t)-IsoP was shown to be dependent upon an unsaturated carbonyl residue which transiently adducts to free thiols. Site directed mutagenesis of the redox sensitive C179 site of the Ikappa kinase beta subunit, blocked the biological activity of 15-A(3t)-IsoP and NFκB activation. The vasoprotective potential of 15-A(3t)-IsoP was underscored by the ability of this compound to block oxidized lipid accumulation, a critical step in foam cell transformation and atherosclerotic plaque formation. Taken together, these are the first data identifying the biological activity of a specific product of EPA peroxidation, which is formed in abundance in vivo. The clear mechanism linking 15-A(3t)-IsoP to redox control of NFκB transcription, and the compound's ability to block foam cell transformation suggest that 15-A(3t)-IsoP provides a unique and potent tool to provide vaso- and cytoprotection under conditions of oxidative stress.

Isoprostanes and asthma.

Isoprostanes are prostaglandin (PG)-like compounds generated in vivo following oxidative stress by non-enzymatic peroxidation of polyunsaturated fatty acids, including arachidonic acid. They are named based on their prostane ring structure and by the localization of hydroxyl groups on the carbon side chain; these structural differences result in a broad array of isoprostane molecules with varying biological properties. Generation of specific isoprostanes is also regulated by host cell redox conditions; reducing conditions favor F2-isoprostane production while under conditions with deficient antioxidant capacity, D2- and E2-isoprostanes are formed. F2-isoprostanes (F2-isoP) are considered reliable markers of oxidative stress in pulmonary diseases including asthma. Importantly, F2-isoP and other isoprostanes function as ligands for PG receptors, and potentially other receptors that have not yet been identified. They have been reported to have important biological properties in many organs. In the lung, isoprostanes regulate cellular processes affecting airway smooth muscle tone, neural secretion, epithelial ion flux, endothelial cell adhesion and permeability, and macrophage adhesion and function. In this review, we will summarize the evidence that F2-isoP functions as a marker of oxidative stress in asthma, and that F2-isoP and other isoprostanes exert biological effects that contribute to the pathogenesis of asthma. This article is part of a Special Issue entitled Biochemistry of Asthma.

Isoprostanes and phytoprostanes: Bioactive lipids.

Polyunsaturated fatty acids (PUFA) are important constituents in all eukaryotic organisms, contributing to the structural integrity of biological membranes and serving as precursors for enzymatically-generated local hormones. In addition to these functions, PUFA can generate by a free radical-initiated mechanism, key products which participate in a variety of pathophysiological processes. In particular, free radical-catalyzed peroxidation of PUFA leads to in vivo formation of isoprostanes (IsoP), neuroprostanes (NeuroP), and phytoprostanes (PhytoP) which display a wide range of biological actions. IsoP are now the most reliable indicators of oxidative stress in humans. In this review, we will discuss some advances in our knowledge regarding two cyclic PUFA derivatives, IsoP and PhytoP, and how their biological roles may be clarified through new approaches based on analytical and synthetic organic chemistry.

The neurobiology of isoprostanes and Alzheimer's disease.

In its sporadic form Alzheimer's disease (AD) results from a combination of genetic and environmental risk factors with abnormal oxidative reactions, which result in free radical mediated injury of the brain. Isoprostanes are oxidized lipids formed by a free radical mediated mechanism, which in recent years have emerged as a reliable and sensitive marker of lipid peroxidation and oxidative stress. Consistent data show that they are increased in the brain of human AD as well as AD animal models. Besides their role as biomarkers, isoprostanes possess important biological effects, functioning as mediators of the cellular response to oxidative stress within the CNS. Recent evidence indicates that these lipid oxidation products, by activating the thromboxane receptor system, mediate the pro-amyloidotic neuronal response to oxidative stress in an experimental model of AD. This novel observation has important clinical implication, since pharmacologic modulation of the TP receptor system by selective antagonists, some of which are already available, could represent a novel therapeutic opportunity for AD as disease-modifying agents.

Isoprostanes in fetal and neonatal health and disease.

Isoprostanes are prostaglandin-like bioactive molecules generated via nonenzymatic peroxidation of lipid membrane-derived arachidonic acid by free radicals and reactive oxygen species. Their cognate receptors, biological actions, and signaling pathways are poorly understood. Aside from being sensitive and specific biomarkers of oxidative stress, E- and F-ring isoprostanes have important biological functions and likely mediate many of the disease-related pathological changes for which they are used as indicators. The biochemical pathways involved in isoprostane formation, their pathogenetic relevance to adult disease states, and their biological function are addressed. Developmentally, plasma and tissue content data show that isoprostane levels are highest during fetal and early neonatal life, when compared with adults. As such, the available data suggesting that isoprostanes play an important biological role, as well as possibly actively participate in the regulation of pulmonary vascular tone and the transition from fetal to postnatal life, are here reviewed. Lastly, the association between isoprostanes and certain neonatal clinical conditions is addressed. Although its existence has been recognized for almost 20 years, little is known about the critical importance of isoprostanes during fetal life and immediate neonatal period. This review is an attempt to bridge this knowledge gap.

Hypoxia-induced hyperreactivity of pulmonary arteries: role of cyclooxygenase-2, isoprostanes, and thromboxane receptors.

AIMS: This study investigates the role of the cyclooxygenase (COX)/prostanoid pathway in chronic hypoxia-induced hyperreactivity of pulmonary arteries. METHODS AND RESULTS: Pulmonary arteries were removed from normoxic or hypoxic (0.5 atm for 21 days) mice and studied for protein expression/localization of COX-1, COX-2, and thromboxane A2 (TXA2)-synthase, release of TXA2, prostacyclin (PGI2) and the isoprostane 8-iso-prostaglandin F2alpha (8-iso-PGF2alpha), and vasomotor responses. COX-2 expression was increased in all layers of pulmonary arteries from hypoxic mice. In contrast, COX-1 expression was not significantly modified following chronic hypoxia, whereas TXA2-synthase was decreased. Chronic hypoxia differentially affected prostanoid release from pulmonary arteries: TXA2 secretion was not significantly modified; PGI2 secretion was decreased, whereas 8-iso-PGF2alpha secretion was increased. A selective COX-2 inhibitor decreased 8-iso-PGF2alpha release. Arachidonic acid elicited an endothelium- and COX-1-dependent relaxation in pulmonary arteries from normoxic mice. In contrast, arachidonic acid induced an endothelium-independent contraction in pulmonary arteries from hypoxic mice that was partially reduced by catalase, COX-1, COX-2, or TXA2-synthase inhibitors and was totally abolished by blockade of the thromboxane (TP) receptor. Hyperresponsiveness to phenylephrine (PE) of pulmonary arteries from hypoxic mice was also decreased by COX-2 inhibitors, TP receptor antagonists or catalase, but not by TXA2-synthase inhibitors. Finally, 8-iso-PGF2alpha induced a TP receptor-dependent contraction in pulmonary arteries and markedly potentiated the contractile response to PE. CONCLUSION: Chronic hypoxia up-regulates COX-2 expression, increases 8-iso-PGF2alpha release, and shifts arachidonic acid-induced, endothelium-dependent relaxation to an endothelium-independent and TP receptor-dependent contraction in pulmonary arteries. COX-2-dependent production of 8-iso-PGF2alpha, by activating TP receptors, participates in hypoxia-induced hyperreactivity of pulmonary arteries.

New bioactive oxylipins formed by non-enzymatic free-radical-catalyzed pathways: the phytoprostanes.

In animals and plants, fatty acids with at least three double bonds can be oxidized to prostaglandin-like compounds via enzymatic and non-enzymatic pathways. The most common fatty acid precursor in mammals is arachidonic acid (C20:4) (AA) which can be converted through the cyclooxygenase pathway to a series of prostaglandins (PG). Non-enzymatic cyclization of arachidonate yields a series of isoprostanes (IsoP) which comprises all PG (minor compounds) as well as PG isomers that cannot be formed enzymatically. In contrast, in plants, alpha-linolenic acid (C18:3) (ALA) is the most common substrate for the allene oxide synthase pathway leading to the jasmonate (JA) family of lipid mediators. Non-enzymatic oxidation of linolenate leads to a series of C18-IsoPs termed dinor IsoP or phytoprostanes (PP). PP structurally resemble JA but cannot be formed enzymatically. We will give an overview of the biological activity of the different classes of PP and also discuss their analytical applications and the strategies developed so far for the total synthesis of PP, depending on the synthetic approaches according to the targets and which key steps serve to access the natural products.

Lipid peroxidation: physiological levels and dual biological effects.

Lipid peroxidation (LPO) has been shown to induce disturbance of membrane organization and functional loss and modification of proteins and DNA bases, and it has been implicated in the pathogenesis of various diseases. At the same time, LPO products have been shown to act as redox signaling mediators. Free and ester forms of both polyunsaturated fatty acids and cholesterol are important substrates for LPO in vivo and they are oxidized by both enzymatic and nonenzymatic mechanisms to give a variety of products. The results of numerous studies reported in the literatures show that the levels of LPO products in plasma of healthy human subjects are below 1 muM and that the molar ratios of LPO products to the respective parent lipids are below 1/1000, that is, below 0.1%. The levels of LPO products in human erythrocytes were found to be higher than those in plasma. Considerable levels of cholesterol oxidation products were observed. Although many LPO products exert cytotoxicity, sublethal concentrations of LPO products induce cellular adaptive responses and enhance tolerance against subsequent oxidative stress through upregulation of antioxidant compounds and enzymes. This adaptive response is observed not only for chemically reactive alpha,beta-unsaturated carbonyl compounds such as 4-hydroxy-2-nonenal and 15-deoxy-delta-12,14-prostaglandin J(2) but also for chemically stable compounds such as hydroxyoctadecadienoic acid, hydroxylcholesterol, and lysophosphatidylcholine. Such opposite dual functions of LPO products imply that LPO, and probably oxidative stress in general, may exert both deleterious and beneficial effects in vivo. LPO as well as reactive oxygen and nitrogen species has been shown to play an important role as a regulator of gene expression and cellular signaling messenger. In order to exert physiologically important functions as a regulator of gene expression and mediator of cellular signaling, the formation of LPO products must be strictly controlled and programmed. In contrast to LPO products by enzymatic oxidation, it appears difficult to regulate the formation of free radical-mediated LPO products. Even such unregulated LPO products may exert beneficial effects at low levels, but excessive unregulated LPO may lead to pathological disorders and diseases.

The effect of oxygen and light on the structure and function of the neonatal rat retina.

The neonatal rat is born with its eyes closed and an immature visual system, that some say is equivalent to that of a human fetus at 26 weeks of gestation. From birth, the visual system of the newborn rat will gradually mature, the first manifestation of that being the opening of the eye which usually take place at postnatal day 14. Complete maturation of the retina and visual pathways is normally reached at the end of the first month of life. The neonatal rat model thus represents a unique paradigm to study the normal and abnormal maturation of the primary visual pathways that normally occurs in utero in human subjects. Our laboratory has, over the past decade, developed two animal models of postnatally induced retinopathy, namely the Oxygen-Induced Retinopathy (OIR) that share several common features with the human Retinopathy of Prematurity (ROP) and the Light-Induced Retinopathy that is viewed by some as a valid model of some forms of Retinitis Pigmentosa (RP). The following pages review what is known of the pathophysiological processes taking place and suggest possible therapeutic avenues that could be explored in order to halt the degenerative process.

Isoprostanes and lung vascular pathology.

Isoprostanes are products of peroxidative attack of membrane lipids. As such, they accumulate to substantial levels in conditions of oxidative stress, including many pulmonary vascular diseases such as acute lung injury and pulmonary hypertension, and are increasingly being used as indicators of disease state and severity. However, our group and others have hypothesized that they are more than inert markers, but may also act as signal transduction molecules. As isomers of prostaglandins, they can exert powerful biological effects on many lung cell types through actions on prostanoid receptors. In this review, we collect many lines of evidence that point to causal roles for the isoprostanes in those disease states.

Regulation of neurotransmitter release from ocular tissues by isoprostanes.

Isoprostanes are prostaglandin-like compounds formed in vivo primarily by free radical-catalyzed peroxidation of arachidonic acid independent of the cyclooxygenase enzyme. In addition to being utilized as reliable indicators of oxidative stress, 8-isoprostanes exert pharmacological actions on smooth muscles from several tissues and organs, and they play a role in the release of neurotransmitters from the central and peripheral nervous systems. In the anterior uvea of the eye, 8-isoprostanes produce both excitatory and inhibitory effects on sympathetic neurotransmission in isolated mammalian iris ciliary bodies. Thromboxane (TP) receptors mediate the stimulatory action of isoprostanes on norepinephrine (NE) release from sympathetic nerves. In bovine retina, the 8-isoprostanes exhibit a biphasic regulatory effect on potassium-induced [3H]-D-aspartate release, with low concentrations being inhibitory and high concentrations causing an excitatory effect. Excitatory effects of 8-isoprostanes are mediated by TP receptors, while inhibitory responses are mediated by prostaglandin E (EP) receptors. The 8-isoprostanes produce pharmacological actions on sympathetic neurotransmission in mammalian anterior uvea, a response that is species-dependent. In the posterior segment of the eye, 8-isoprostanes elicit a complex response on the retina involving the activation of both prostanoid TP and EP receptors. An effect of isoprostanes on neurotransmitter pools provides new pharmacological target sites for the therapy of some ocular diseases.

Insights into oxidative stress: the isoprostanes.

Oxidative stress, characterized by an imbalance between increased exposure to free radicals and antioxidant defenses, is a prominent feature of many acute and chronic diseases and even the normal aging process. However, definitive evidence for this association has often been lacking due to recognized shortcomings with methods previously available to assess oxidant stress status in vivo in humans. Several in vitro markers of oxidative stress are available, but most are of limited value in vivo because thay lack sensitivity and/or specificity or require invasive methods. Isoprostanes (IsoPs) are prostaglandin (PG)-like compounds that are produced in vivo independently of cyclooxygenase enzymes, primarily by free radical-induced peroxidation of arachidonic acid. F(2)-IsoPs are a group of 64 compounds isomeric in structure to cyclooxygenase-derived PGF(2alpha). Other products of the IsoP pathway are also formed in vivo by rearrangement of labile PGH(2)-like IsoP intermediates including E(2)- and D(2)-IsoPs, cyclopentenone-A(2)- and J(2)-IsoPs, and highly reactive acyclic-ketoaldehydes (isoketals). Oxidation of docosahexaenoic acid, an abundant unsaturated fatty acid in the central nervous system, results in the formation of IsoP-like compounds, termed neuroprostanes. Measurement of F(2)-IsoPs is the most reliable approach to assess oxidative stress status in vivo, providing an important tool to explore the role of oxidative stress in the pathogenesis of human disease. Moreover, F(2)-IsoPs and other products of the IsoP pathway exert potent biological actions both via receptor-dependent and independent mechanisms and therefore may be pathophysiological mediators of disease. Measurement of F(2)-IsoPs may provide a uniquely valuable approach to understanding of the clinical pharmacology of antioxidants.

Involvement of endothelial thromboxane A2 in the vasoconstrictor response induced by 15-E2t-isoprostane in isolated human umbilical vein.

The present study was undertaken to evaluate the contractile response of several E- and F-ring isoprostanes (IsoP) in human umbilical vein (HUV) and to investigate the role of the endothelium on the effect of 15-E2t-IsoP, the most potent vasoconstrictor isoprostane, in human vessels. HUV rings with or without endothelium were suspended in an organ bath for recording the isometric tension in response to different agonists. The inhibitors to be evaluated were applied 30 min before the addition of the agonist. All of the compounds tested produced concentration-dependent contractions when tested on HUV rings with endothelium. Although these compounds were equieffective, significant differences were observed in their potency, with U46619 being the most potent followed by 15-E2t-IsoP > 15-E1t-IsoP = 15-F2t-IsoP > 15-F1t-IsoP = 9-epi-15-F2t-IsoP in descending rank order of potency. 15-E2t-IsoP was the most potent of the isoprostanes evaluated and, therefore, the one employed in the present study. When intact endothelium HUV rings were used, 15-E2t-IsoP-induced contraction was unaffected by the endothelin-converting enzyme inhibitor, phosphoramidon (10 microM), suggesting that short-term endothelin-1 release is not involved in this response. However, the non-selective cyclooxygenase (COX) inhibitor, indomethacin (10 and 30 microM), and the COX-2 selective inhibitor, NS-398 (3, 10 and 30 microM) produced inhibitory effects on 15-E2t-IsoP-induced contraction of HUV rings with endothelium. These results indicate that COX-derived contractile prostanoids are involved in this effect. Furthermore, the apparent pKb values estimated for indomethacin (5.5) and NS-398 (5.4) suggest that the prostanoids involved are derived from the COX-2 isoenzyme pathway. On HUV rings with endothelium, the phospholipase A2 inhibitor, oleyloxyethyl phosphorylcholine (30 and 100 microM), induced an inhibitory effect on 15-E2t-IsoP-induced contraction, suggesting that the phospholipase A2 pathway is also involved in this effect. In addition, the thromboxane A2 synthase inhibitor furegrelate (10 and 30 microM) also inhibited 15-E2t-IsoP-induced contraction of HUV rings with endothelium, indicating that thromboxane A2 is one of the contractile prostanoids involved in this response. Endothelium denudation clearly diminished the vasoconstrictor potency of 15-E2t-IsoP, demonstrating that the endothelium releases a vasoconstrictor factor in response to 15-E2t-IsoP. The absence of an inhibitory effect at the highest concentration of furegrelate (30 microM) on 15-E2t-IsoP-induced contraction of HUV rings without endothelium suggested that endothelium is the source of thromboxane A2. We conclude that prostanoids derived from the COX-2 isoenzyme pathway participate in 15-E2t-IsoP-induced vasoconstriction of isolated HUV rings. Our results also indicate that endothelial thromboxane A2 is one of the prostanoids involved in this effect.

Sex differences in susceptibility to oxidative injury and sleepiness from intermittent hypoxia.

STUDY OBJECTIVES: Adult male mice exposed to long-term intermittent hypoxia (LTIH), modeling sleep apnea oxygenation patterns, develop nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-dependent residual hypersomnolence and oxidative injury in select brain regions, including wake-active regions. Premenopausal females are less susceptible to selective oxidative brain injuries. We sought to determine whether female mice exposed to LTIH would confer resistance to LTIH-induced wake impairments and oxidative injuries. SUBJECTS AND SETTING: Young adult male and female C57BI/6J mice were studied in a university laboratory. INTERVENTIONS: Mice were randomly assigned to either LTIH or sham LTIH for 8 weeks. Total (24-h) wake time and mean sleep latency were measured under 2 conditions: rested and following 6 hours of enforced wakefulness. NADPH oxidase activation, carbonylation, and lipid peroxidation assays were also performed to assess sex differences in oxidative responses to LTIH. RESULTS: In contrast with the significant LTIH-induced wake impairments observed in male mice, females following LTIH showed normal wake times and sleep latencies. Female mice revealed less baseline carbonylation and less carbonylation following LTIH but showed robust NADPH oxidase activation and lipid peroxidation. In contrast with the female relative resistance to LTIH sleepiness, female mice showed more-pronounced sleepiness and delta response after enforced wakefulness. CONCLUSIONS: Despite a robust oxidative response to LTIH, age-matched female mice may be protected, at least temporarily, from LTIH wake impairments by lower basal carbonylation. In contrast, females show greater wake impairments after sleep deprivation. We hypothesize sex differences in polysomnographic predictors of sleepiness and residual sleepiness in humans with sleep apnea.

[Isoprostanes, biomarkers of lipid peroxidation in humans. Part 3: Biomarkers and mediators in vascular physiology and disease]. / Les isoprostanes, biomarqueurs de peroxydation lipidique chez l'homme. Partie 3: biomarqueurs et médiateurs en physiologie et pathologie vasculaire.

The 15-series F(2)-isoprostanes mediate vasoconstriction in different vascular beds and species. This contraction is mediated by the thromboxane receptors stimulation, and may be modulated by the endothelium. Furthermore, 15-F(2t)-IsoP induces smooth muscle cells mitogenesis and monocyte adhesion to endothelial cells. Some 15-series E(2)-isoprostanes are more potent than F(2)-isoprostanes. In clinical studies, 15-F(2t)-IsoP levels are increased in vascular disorders involving atherosclerosis, ischemia-reperfusion and inflammation. F(2)-isoprostane levels correlate to the severity of heart failure and pulmonary hypertension, raising the potential prognostic interest of these biomarkers. Whether the effects observed in vitro are observed consistently in vivo at physiological concentrations and whether these effects contribute to pathological states in vivo is still debated.

The pulmonary biology of isoprostanes.

Isoprostanes were first recognized as convenient markers of oxidative stress, but their powerful effects on a variety of cell functions are now also being increasingly appreciated. This is particularly true of the lung, which is comprised of a wide variety of different cell types (smooth muscle, innervation, epithelium, lymphatics, etc.), all of which have been shown to respond to exogenously applied isoprostanes. In this review, we summarize these biological responses in the lung, and also consider the roles that isoprostanes might play in a range of pulmonary clinical disorders.

The endothelium in health and disease: a discussion of the contribution of non-nitric oxide endothelium-derived vasoactive mediators to vascular homeostasis in normal vessels and in type II diabetes.

Endothelial dysfunction is considered as a major risk factor of cardiovascular complications of type I and types II diabetes. Impaired endothelium-dependent vasodilatation can be directly linked to a decreased synthesis of the endothelium-derived nitric oxide (NO) and/or an increase in the production of reactive oxygen species such as superoxide. Administration of tetrahydrobiopterin, an important co-factor for the enzyme nitric oxide synthase (NOS), has been demonstrated to enhance NO production in prehypertensive rats, restore endothelium-dependent vasodilatation in coronary arteries following reperfusion injury, aortae from streptozotocin-induced diabetic rats and in patients with hypercholesterolemia. Tetrahydrobiopterin supplementation has been shown to improve endothelium-dependent relaxation in normal individuals, patients with type II diabetes and in smokers. These findings from different animal models as well as in clinical trials lead to the hypothesis that tetrahydrobiopterin, or a precursor thereof, could be a new and an effective therapeutic approach for the improvement of endothelium function in pathophysiological conditions. In addition to NO, the endothelium also produces a variety of other vasoactive factors and a key question is: Is there also a link to changes in the synthesis/action of these other endothelium-derived factors to the cardiovascular complications associated with diabetes? Endothelium-derived hyperpolarizing factor, or EDHF, is thought to be an extremely important vasodilator substance notably in the resistance vasculature. Unfortunately, the nature and, indeed, the very existence of EDHF remains obscure. Potentially there are multiple EDHFs demonstrating vessel selectivity in their actions. However, until now, identity and properties of EDHF that determine the therapeutic potential of manipulating EDHF remains unknown. Here we briefly review the current status of EDHF and the link between EDHF and endothelial dysfunction associated with diabetes.