Impaired Endothelium-dependent Vasodilation Research Articles

CD70 Modulates the Role of eNOS In Endothelial Cells

CD70 has traditionally been identified on antigen presenting cells and interacts with CD27 on T cells, promoting proliferation and memory cell formation. We have recently shown that CD70 is important in the formation of effector memory T cells in hypertension. However, the precise cell types that express CD70 and the role of CD70 on non‐immune cells has not been investigated. Here we describe a new role of CD70 in endothelial function. We found that human umbillical vein endothelial cells express CD70 mRNA and that this is increased by > 30 fold by laminar (15 dynes/cm2) compared to oscillatory shear. Studies of mesenteric vascular reactivity showed that CD70−/− mice have markedly impaired endothelium‐dependent vasodilation to acetylcholine compared to WT mice (45± 5 vs 25 ± 4 %) at baseline. In contrast, there were no differences in relaxation responses to sodium nitroprusside. Pre‐incubating vessels with the endothelial nitric oxide synthase (eNOS) inhibitor L‐NAME blocked endothelium‐dependent vasodilatation to acetylcholine in WT mice nearly 75%; in contrast, L‐NAME had no effect on endothelial vessel relaxation in CD70−/− mice. This suggests that vessels of CD70−/− mice lack the capacity to produce nitric oxide (NO). Western blot for the endothelial nitric oxide synthase (eNOS) showed a marked reduction of this enzyme in the aortas of CD70−/− as compared to wild type mice. We found that CD70 and eNOS interact by co‐immunoprecipitation, however this association is abolished by siRNA against CD70. In additional experiments, aortas from CD70−/− did demonstrate expression of both neuronal NOS (nNOS) and inducible NOS (iNOS). In order to assess functional effects of this expression, nNOS NO production was blocked with LVNIO, resulting in further impairment in vasodilation of CD70−/− mesenteric vessels (−logEC50 of 5.4 after treatment, compared to 6.2 before treatment); however blockade of iNOS with 1400 W resulted in apparent recovery of acetylcholine‐induced vasodilation. These data indicate that loss of CD70−/− leads to destabilization of the eNOS protein in endothelial cells resulting in endothelial dysfunction. In the absence of eNOS in CD70−/− vessels, iNOS contributes to endothelial dysfunction, while nNOS has a compensatory role. These studies have identified a new role of the immune co‐stimulatory protein CD70 in modulating endothelial eNOS protein levels and vascular function.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

The uremic toxin hippurate promotes endothelial dysfunction via the activation of Drp1-mediated mitochondrial fission

The accumulation of uremic toxins in chronic kidney disease (CKD) induces inflammation, oxidative stress and endothelial dysfunction, which is a key step in atherosclerosis. Accumulating evidence indicates increased mitochondrial fission is a contributing mechanism for impaired endothelial function. Hippurate, a uremic toxin, has been reported to be involved in cardiovascular diseases. Here, we assessed the endothelial toxicity of hippurate and the contribution of altered mitochondrial dynamics to hippurate-induced endothelial dysfunction.Treatment of human aortic endothelial cells with hippurate reduced the expression of endothelial nitric oxide synthase (eNOS) and increased the expression of intercellular cell adhesion molecule-1 (ICAM-1) and von Willebrand factor (vWF). The mechanisms of hippurate-induced endothelial dysfunction in vitro depended on the activation of Dynamin-related protein 1 (Drp1)-mediated mitochondrial fission and overproduction of mitochondrial reactive oxygen species (mitoROS). In a rat model in which CKD was induced by 5/6 nephrectomy (CKD rat), we observed increased oxidative stress, impaired endothelium-dependent vasodilation, and elevated soluble biomarkers of endothelial dysfunction (ICAM-1 and vWF). Similarly, endothelial dysfunction was identified in healthy rats treated with disease-relevant concentrations of hippurate. In aortas of CKD rats and hippurate-treated rats, we observed an increase in Drp1 protein levels and mitochondrial fission. Inhibition of Drp1 improved endothelial function in both rat models.These results indicate that hippurate, by itself, can cause endothelial dysfunction. Increased mitochondrial fission plays an active role in hippurate-induced endothelial dysfunction via an increase in mitoROS.

ENDOTHELIAL PROTECTIVE EFFECT OF LIRAGLUTIDE IN TYPE 2 DIABETES MELLITUS

Background . Endothelial dysfunction underlies the development of chronic complications in type 2 diabetes mellitus. Glucose-lowering drugs with additional protective effects on endothelium are required. Objective . To study endothelial protective action of liraglutide in patients with type 2 diabetes mellitus and to detect its relation with glycemic control. Design and methods . The study was held in patients with type 2 diabetes mellitus with glycated hemoglobin 7,5–9,0 % who received metformin monotherapy. Metformin dose had been titrated for 3 months. Patients who reached glycated hemoglobin level less than 7,5 % were included in the group 1 — they had received metformin monotherapy for next 6 months. Patients with glycated hemoglobin over 7,5 % after 3 months formed the group 2, and they received liraglutide in addition to metformin for the following 6 months. At baseline, at 3, 6 and 9 months, endothelium-dependent vasodilation was evaluated by forearm microcirculation dopplerography with 0,3 % acetylcholine solution ionophoresis. Results. At baseline, patients of groups 1 and 2 showed impaired endothelium-dependent vasodilation (decrease of the reaction to acetylcholine, curve form change). There was no significant change in the linear blood flow parameters in the group 1 when euglycemia was reached by metformin monotherapy. Additional administration of liraglutide in the group 2 led to the increase in the amplitude and favourable change of the curve form. These changes were more prominent after 6 months of combined therapy. Endothelium-dependent vasodilation amplitude reached values observed in healthy subjects after 9 months from the study beginning. Conclusions. The improvement of glycemia control (euglycemia) does not have endothelial protective properties on its own. Liraglutide has its own endothelial protective action independent of glycemia control.

An Update on Hypertension in Children With Type 1 Diabetes.

The prevalence of hypertension in children with type 1 diabetes is reported to be between 6% and 16%. This potentially modifiable cardiovascular risk factor may go undiagnosed and undertreated, particularly in children with type 1 diabetes. Recent updated Canadian clinical practice guidelines recommend blood pressure screening every 2 years in children with type 1 diabetes as well as routine use of ambulatory blood pressure monitoring. Risk factors for hypertension in type 1 diabetes include poor glycemic control, overweight and obesity and genetic predisposition for hypertension. In terms of pathophysiology, sustained hyperglycemia, angiotensin I and II and inflammatory cytokines have been implicated. Endothelial and vascular dysfunction, with impaired endothelial-dependent vasodilation and increased carotid artery intima-media thickness, are evident in preclinical and clinical studies of children and not just in adults with type 1 diabetes. Early targeted therapy is critical to the control of hypertension and the development of related morbidity. As with hypertension in adults with type 1 diabetes, lifestyle modifications remain first-line therapy, including diet and glycemic control. Initial antihypertensive therapy should be an angiotensin-converting enzyme inhibitor or an angiotensin receptor blocker because of their associated effects of reducing microalbuminuria and improving renovascular outcomes. Pediatric hypertension in type 1 diabetes is an area of evolving study and opinion; identification and appropriate treatment is critical for the prevention of micro- and macrovascular complications in adulthood.

Multiple common comorbidities produce left ventricular diastolic dysfunction associated with coronary microvascular dysfunction, oxidative stress, and myocardial stiffening

AimsMore than 50% of patients with heart failure have preserved ejection fraction characterized by diastolic dysfunction. The prevalance of diastolic dysfunction is higher in females and associates with multiple comorbidities such as hypertension (HT), obesity, hypercholesterolemia (HC), and diabetes mellitus (DM). Although its pathophysiology remains incompletely understood, it has been proposed that these comorbidities induce systemic inflammation, coronary microvascular dysfunction, and oxidative stress, leading to myocardial fibrosis, myocyte stiffening and, ultimately, diastolic dysfunction. Here, we tested this hypothesis in a swine model chronically exposed to three common comorbidities.Methods and resultsDM (induced by streptozotocin), HC (produced by high fat diet), and HT (resulting from renal artery embolization), were produced in 10 female swine, which were followed for 6 months. Eight female healthy swine on normal pig-chow served as controls. The DM + HC + HT group showed hyperglycemia, HC, hypertriglyceridemia, renal dysfunction and HT, which were associated with systemic inflammation. Myocardial superoxide production was markedly increased, due to increased NOX activity and eNOS uncoupling, and associated with reduced NO production, and impaired coronary small artery endothelium-dependent vasodilation. These abnormalities were accompanied by increased myocardial collagen content, reduced capillary/fiber ratio, and elevated passive cardiomyocyte stiffness, resulting in an increased left ventricular end-diastolic stiffness (measured by pressure–volume catheter) and a trend towards a reduced E/A ratio (measured by cardiac MRI), while ejection fraction was maintained.ConclusionsThe combination of three common comorbidities leads to systemic inflammation, myocardial oxidative stress, and coronary microvascular dysfunction, which associate with myocardial stiffening and LV diastolic dysfunction with preserved ejection fraction.

Hronična niskostepena inflamacija u procesu starenja kao karika u lancu vaskularnih poremećaja koji su povezani sa gojaznošću

The pathogenesis of obesity-related vascular disorders has not been fully elucidated. The fundamental role of inflammation in aging process is now widely recognized, particularly for atherosclerotic disease which begins before birth. The number of obese individuals worldwide has reached two billion, leading to an explosion of obesity-related vascular disorders associated with increased morbidity and mortality. Obesity, as a chronic low grade inflammatory process, is important risk factor for metabolic and cardiovascular disease. Despite a well-known genetic component, this risk appears to originate from several abnormalities in adipose tissue function associated with a chronic inflammatory state. In particular, obesity as the most common nutritional disorder in industrialized countries, is closely related to impaired endothelial function, a well-known marker of preatherosclerotic disease. These conditions disrupt vascular homeostasis by causing an imbalance between the nitric oxide pathway and the endothelin-1 system, with impaired insulin-stimulated endothelium-dependent vasodilation. Having in mind the growing population of overweight and obese people worldwide, along with an increasingly aging population, understanding the pathophysiology of obesity on cardiovascular system is essential. The mechanisms linking obesity-related vascular disorders and low grade inflammation in aging process are the focus of this paper.

Comparative Study of Nitric Oxide Levels in Metabolic Syndrome and Diabetes Mellitus Patients

The metabolic syndrome is a clustering of risk factors of metabolic origin that increase the risk for cardiovascular disease and type 2 diabetes (DM). A proposed central event in metabolic syndrome is a decrease in the amount of bioavailable nitric oxide (NO) from endothelial NO synthase (eNOS). Patients with diabetes invariably show an impairment of endothelium-dependent vasodilation. Our aim is to evaluate the endothelial dysfunction in metabolic syndrome & Diabetes patients by estimation of nitric oxide levels. The study was conducted at SLIMS, Puducherry. The study included 200 diabetic patients, 200 metabolic syndrome patients and 200 Controls. When compared to controls, It was found a significant increase (p<0.001) in nitric oxide among metabolic syndrome and diabetes group and significant difference (p<0.001) between diabetes mellitus and metabolic syndrome groups being higher in metabolic syndrome group. Nitric oxide is reduced in the course of vascular disease in diabetes mellitus. Increased production of superioxide anion on oxidative stress to reduce plasma nitric oxide levels.

Arginine and aerobic training prevent endothelial and metabolic alterations in rats at high risk for the development of the metabolic syndrome - CORRIGENDUM.

Endothelial function is a key mechanism in the development of CVD. Arginine and exercise are important non-pharmacological strategies for mitigating the impact of metabolic changes in the metabolic syndrome, but the effect of their combined administration is unknown. Thus, the aim of this study was to investigate the isolated and combined effects of aerobic training and arginine supplementation on metabolic variables and vascular reactivity in rats at high risk for developing the metabolic syndrome. Wistar rats were divided into two groups: control and fructose (F - water with 10 % fructose). After 2 weeks, the F group was divided into four groups: F, fructose+arginine (FA, 880 mg/kg per d of l-arginine), fructose+training (FT) and fructose+arginine+training (FTA); treatments lasted for 8 weeks, and no difference was observed in body mass gain. Arginine did not improve the body protein content, and both the FA and FT groups show a reversal of the increase in adipose tissue. Insulin increase was prevented by training and arginine, without additive effect, and the increase in serum TAG was prevented only by training. The F group showed impaired endothelium-dependent vasodilation and hyperreactivity to phenylephrine, but arginine and training were capable of preventing these effects, even separately. Higher nitric oxide level was observed in the FA and FT groups, and no potentiating effect was detected. Thus, only training was able to prevent the increase in TAG and improve the protein mass, and training and arginine exert similar effects on fat content, insulin and endothelial function, but these effects are not additive.

Endothelial function is impaired in the cutaneous microcirculation of adults with psoriasis through reductions in nitric oxide-dependent vasodilation.

Psoriasis is an independent risk factor for cardiovascular disease; however, the underlying mechanisms are not fully understood. Deficits in conduit arterial function are evident in patients with psoriasis, but potential impairments in microcirculatory endothelial function remain unclear. We hypothesized that cutaneous microvascular dysfunction would be detectable in otherwise healthy individuals with psoriasis. Two intradermal microdialysis fibers were placed in (nonlesional) forearm skin of nine patients (3 men and 6 women, 39 ± 5 yr) with moderate (16 ± 2% of body surface area) plaque psoriasis and nine healthy (nonpsoriatic) control subjects (3 men and 6 women, 38 ± 5 yr) for local delivery of 1) lactated Ringer solution (control) and 2) 10 mM l-ascorbate (a nonspecific antioxidant). An index of skin blood flow was measured using laser-Doppler flowmetry during local heating (42°C). Nitric oxide (NO)-dependent vasodilation was directly quantified after perfusion of the nonspecific NO synthase inhibitor NG-nitro-l-arginine methyl ester (15 mM). A third fiber was perfused with increasing concentrations (10-10 - 10-2 M) of norepinephrine to elicit adrenoreceptor-mediated cutaneous vasoconstriction. NO-dependent vasodilation was attenuated in patients with psoriasis (57 ± 5% and 39 ± 7% maximum cutaneous vascular conductance in control subjects and adults with psoriasis, respectively, P < 0.01). l-Ascorbate did not improve NO-dependent vasodilation ( P > 0.05). There was no group difference in maximal vasoconstriction or microvascular sensitivity to norepinephrine ( P > 0.05). These data suggest that NO bioavailability is reduced in otherwise healthy individuals with psoriasis, which contributes to systemic microvascular dysfunction. NEW & NOTEWORTHY In adults with psoriasis, reduced nitric oxide bioavailability mediates impaired endothelium-dependent vasodilation, independent of increases in oxidative stress. Furthermore, the degree of psoriatic symptomology is directly related to greater reductions in nitric oxide-dependent vasodilation.

Assessment of Human Adipose Tissue Microvascular Function Using Videomicroscopy.

While obesity is closely linked to the development of metabolic and cardiovascular disease, little is known about mechanisms that govern these processes. It is hypothesized that pro-atherogenic mediators released from fat tissues particularly in association with central/visceral adiposity may promote pathogenic vascular changes locally and systemically, and the notion that cardiovascular disease may be the consequence of adipose tissue dysfunction continues to evolve. Here, we describe a unique method of videomicroscopy that involves analysis of vasodilator and vasoconstrictor responses of intact small human arterioles removed from the adipose depot of living human subjects. Videomicroscopy is used to examine functional properties of isolated microvessels in response to pharmacological or physiological stimuli using a pressured system that mimics in vivo conditions. The technique is a useful approach to gain understanding of the pathophysiology and molecular mechanisms that contribute to vascular dysfunction locally within the adipose tissue milieu. Moreover, abnormalities in the adipose tissue microvasculature have also been linked with systemic diseases. We applied this technique to examine depot-specific vascular responses in obese subjects. We assessed endothelium-dependent vasodilation to both increased flow and acetylcholine in adipose arterioles (50 - 350 µm internal diameter, 2 - 3 mm in length) isolated from two different adipose depots during bariatric surgery from the same individual. We demonstrated that arterioles from visceral fat exhibit impaired endothelium-dependent vasodilation compared to vessels isolated from the subcutaneous depot. The findings suggest that the visceral microenvironment is associated with vascular endothelial dysfunction which may be relevant to clinical observation linking increased visceral adiposity to systemic disease mechanisms. The videomicroscopy technique can be used to examine vascular phenotypes from different fat depots as well as compare findings across individuals with different degrees of obesity and metabolic dysfunction. The method can also be used to examine vascular responses longitudinally in response to clinical interventions.

Abstract P498: Redox Regulation of Vascular Smooth Muscle cGMP Signaling and Blood Pressure by Cytochrome B5 Reductase 3

Oxidized soluble guanylate cyclase (sGC) heme iron (Fe 3+ ) is desensitized to nitric oxide (NO) and attenuates cGMP production needed for downstream activation of PKG-dependent signaling and vasodilation. While reactive oxygen species drive oxidation of sGC heme iron, the basic mechanism(s) governing sGC heme iron recycling to its NO-sensitive, reduced state (Fe 2+ ), are unknown. Here we report cytochrome b5 reductase 3 (Cyb5R3), also known as methemoglobin reductase, as a novel sGC heme iron reductase and regulator of cGMP production in vascular smooth muscle cells (VSMCs). Oxidant challenge studies demonstrate that VSMCs have an intrinsic ability to reduce oxidized sGC heme iron and form protein-protein complexes between Cyb5R3 and oxidized sGC. Genetic knockdown and pharmacological inhibition in VSMCs reveal Cyb5R3 expression and activity is critical for NO-stimulated cGMP production and vasodilation. Mechanistically, Cyb5R3 directly reduces oxidized sGC for NO sensitization assessed by biochemical, cellular, and ex vivo assays. Furthermore, generation of a smooth muscle specific Cyb5R3 knockout shows increased blood pressure and impaired endothelial dependent vasodilation. Together, these findings uncover new insights into NO-sGC-cGMP signaling and reveal Cyb5R3 as the first identified physiological sGC heme iron reductase in VSMC. The co-expression of Cyb5R3 and sGC in multiple cell types may unveil a fundamental mechanistic partnership that is critical in numerous physiological and pathophysiological processes.

The oxidized phospholipid POVPC impairs endothelial function and vasodilation via uncoupling endothelial nitric oxide synthase

Endothelial dysfunction is an early stage of atherosclerosis. We recently have shown that 25-hydroxycholesterol found in atherosclerotic lesions could impair endothelial function and vasodilation by uncoupling and inhibiting endothelial nitric oxide synthase (eNOS). 1-Palmitoyl-2-(5-oxovaleroyl)-sn-glycero-3-phosphocholine (POVPC), the oxidation product of oxidized low-density lipoprotein, is another proinflammatory lipid and has also been found in atherosclerotic lesions. However, whether POVPC promotes atherosclerosis like 25-hydroxycholesterol remains unclear. The purpose of this study was to explore the effects of POVPC on endothelial function and vasodilation. Human umbilical vein endothelial cells (HUVECs) were incubated with POVPC. Endothelial cell proliferation, migration and tube formation were measured. Nitric oxide (NO) production and superoxide anion generation (O2−) were determined. The expression and phosphorylation of endothelial nitric oxide synthase (eNOS), AKT, PKC-βII and P70S6K as well as the association of eNOS and heat shock protein 90 (HSP90) were detected by immunoblotting and immunoprecipitation. Endothelial cell apoptosis was monitored by TUNEL staining. The expression of Bcl-2, Bax, and Cleaved Caspase 3 were detected by immunoblotting. Finally, aortic ring from C57BL6 mice were isolated and treated with POVPC and the endothelium-dependent vasodilation was evaluated. POVPC significantly inhibited HUVECs proliferation, migration, tube formation, decreased NO production but increased O2− generation. POVPC inhibited the phosphorylation of Akt and eNOS at Ser1177, increased activation of PKC-βII, P70S6K and the phosphorylation of eNOS at Thr495, reduced the association of HSP90 with eNOS. Meanwhile, POVPC induced endothelial cell apoptosis by inhibiting Bcl-2 expression, increasing Bax and cleaved caspase-3 expressions as well as caspase-3 activity and impaired endothelium-dependent vasodilation. These data demonstrated that POVPC impaired endothelial function by uncoupling and inhibiting eNOS as well as by inducing endothelial cell apoptosis. Therefore, POVPC may play an important role in the development of atherosclerosis and may be considered as a potential therapeutic target for atherosclerosis.

Role of glutathione biosynthesis in endothelial dysfunction and fibrosis

Glutathione (GSH) biosynthesis is essential for cellular redox homeostasis and antioxidant defense. The rate-limiting step requires glutamate-cysteine ligase (GCL), which is composed of the catalytic (GCLc) and the modulatory (GCLm) subunits. To evaluate the contribution of GCLc to endothelial function we generated an endothelial-specific Gclc haplo-insufficient mouse model (Gclc e/+ mice). In murine lung endothelial cells (MLEC) derived from these mice we observed a 50% reduction in GCLc levels compared to lung fibroblasts from the same mice. MLEC obtained from haplo-insufficient mice showed significant reduction in GSH levels as well as increased basal and stimulated ROS levels, reduced phosphorylation of eNOS (Ser 1177) and increased eNOS S-glutathionylation, compared to MLEC from wild type (WT) mice. Studies in mesenteric arteries demonstrated impaired endothelium-dependent vasodilation in Gclc(e/+) male mice, which was corrected by pre-incubation with GSH-ethyl-ester and BH4. To study the contribution of endothelial GSH synthesis to renal fibrosis we employed the unilateral ureteral obstruction model in WT and Gclc(e/+) mice. We observed that obstructed kidneys from Gclc(e/+) mice exhibited increased deposition of fibrotic markers and reduced Nrf2 levels. We conclude that the preservation of endothelial GSH biosynthesis is not only critical for endothelial function but also in anti-fibrotic responses.

Insufficient sleep is associated with impaired nitric oxide-mediated endothelium-dependent vasodilation

Background and aimsHabitual short nightly sleep duration is associated with increased atherosclerotic cardiovascular disease risk and morbidity. Vascular endothelial dysfunction represents an important mechanism that may underlie this heightened cardiovascular risk. Impaired endothelium-dependent vasodilation, particularly NO-mediated vasodilation, contributes to the development and progression of atherosclerotic vascular disease and acute vascular events. We tested the hypothesis that chronic insufficient sleep is associated with impaired NO-mediated endothelium-dependent vasodilation in middle-aged adults. MethodsThirty adult men were studied: 15 with normal nightly sleep duration (age: 58 ± 2 y; sleep duration: 7.7 ± 0.2 h/night) and 15 with short nightly sleep duration (55 ± 2 y; 6.1 ± 0.2 h/night). Forearm blood flow (FBF) responses to intra-arterial infusion of acetylcholine, in the absence and presence of the endothelial NO synthase inhibitor NG-monomethyl-L-arginine (L-NMMA), as well as responses to sodium nitroprusside, were determined by strain-gauge venous occlusion plethysmography. ResultsThe FBF response to acetylcholine was lower (∼20%; p<0.05) in the short sleep duration group (from 4.6 ± 0.3 to 11.7 ± 1.0 ml/100 ml tissue/min) compared with normal sleep duration group (from 4.4 ± 0.3 to 14.5 ± 0.5 ml/100 ml tissue/min). L-NMMA significantly reduced the FBF response to acetylcholine in the normal sleep duration group (∼40%), but not the short sleep duration group. There were no group differences in the vasodilator response to sodium nitroprusside. ConclusionsThese data indicate that short nightly sleep duration is associated with endothelial-dependent vasodilator dysfunction due, in part, to diminished NO bioavailability. Impaired NO-mediated endothelium-dependent vasodilation may contribute to the increased cardiovascular risk with insufficient sleep.

Pyrrolidine dithiocarbamate ameliorates endothelial dysfunction in thoracic aorta of diabetic rats by preserving vascular DDAH activity.

ObjectiveEndothelial dysfunction plays a pivotal role in the development of diabetic cardiovascular complications. Accumulation of endogenous nitric oxide synthase (NOS) inhibitor asymmetric dimethylarginine (ADMA) and inhibition of dimethylarginine dimethylaminohydrolase (DDAH) activity have been involved in diabetic endothelial dysfunction. This study was to investigate the effect of pyrrolidine dithiocarbamate (PDTC) on impairment of endothelium-dependent vasodilatation in diabetic rats and its potential mechanism.MethodsDiabetic rats were induced by a single intraperitoneal injection of streptozotocin (60mg/kg), and PDTC (10mg/kg) was given in drinking water for 8 weeks. Blood glucose and serum ADMA concentrations were measured in experimental rats. Recombinant adenovirus encoding human DDAH2 gene were constructed and ex vivo transferred to isolated rat aortas. The maximal relaxation (Emax) and half maximal effective concentration (EC50) of aortic rings response to accumulative concentrations of acetylcholine and vascular DDAH activity were examined before and after gene transfection.ResultsDiabetic rats displayed significant elevations of blood glucose and serum ADMA levels compared to control group (P<0.01). Vascular DDAH activity and endothelium-dependent relaxation of aortas were inhibited, as expressed by the decreased Emax and increased EC50 in diabetic rats compared to control rats (P<0.01). Treatment with PDTC not only decreased blood glucose and serum ADMA concentration (P<0.01) but also restored vascular DDAH activity and endothelium-dependent relaxation, evidenced by the higher Emax and lower EC50 in PDTC-treated diabetic rats compared to untreated diabetic rats (P<0.01). Similar restoration of Emax, EC50 and DDAH activity were observed in diabetic aortas after DDAH2-gene transfection.ConclusionsThese results indicate that PDTC could ameliorate impairment of endothelium-dependent relaxation in diabetic rats. The underlying mechanisms might be related to preservation of vascular DDAH activity and consequent reduction of endogenous ADMA in endothelium via its antioxidant action. This study highlights the therapeutic potential of PDTC in impaired vasodilation and provides a new strategy for treatment of diabetic cardiovascular complications.

Arginine and aerobic training prevent endothelial and metabolic alterations in rats at high risk for the development of the metabolic syndrome.

Endothelial function is a key mechanism in the development of CVD. Arginine and exercise are important non-pharmacological strategies for mitigating the impact of metabolic changes in the metabolic syndrome, but the effect of their combined administration is unknown. Thus, the aim of this study was to investigate the isolated and combined effects of aerobic training and arginine supplementation on metabolic variables and vascular reactivity in rats at high risk for developing the metabolic syndrome. Wistar rats were divided into two groups: control and fructose (F - water with 10 % fructose). After 2 weeks, the F group was divided into four groups: F, fructose+arginine (FA, 880 mg/kg per d of l-arginine), fructose+training (FT) and fructose+arginine+training (FTA); treatments lasted for 8 weeks, and no difference was observed in body mass gain. Arginine did not improve the body protein content, and both the FA and FT groups show a reversal of the increase in adipose tissue. Insulin increase was prevented by training and arginine, without additive effect, and the increase in serum TAG was prevented only by training. The F group showed impaired endothelium-dependent vasodilation and hyperreactivity to phenylephrine, but arginine and training were capable of preventing these effects, even separately. Higher nitric oxide level was observed in the FA and FT groups, and no potentiating effect was detected. Thus, only training was able to prevent the increase in TAG and improve the protein mass, and training and arginine exert similar effects on fat content, insulin and endothelial function, but these effects are not additive.

OP-16 - Role of Glutathione Biosynthesis in Endothelial Dysfunction and Fibrosis

Glutathione (GSH) biosynthesis is essential for cellular redox homeostasis and antioxidant defence. The rate-limiting step requires glutathione cysteine ligase (GCL), which is composed of the catalytic (GCLc) and modulatory (GCLm) subunits. To evaluate the contribution of GCLc to endothelial function we generated endothelial-specific GCLc haploinsufficient (GCLc e/+ mice). In murine lung endothelial cells (MLEC) derived from these mice we corroborated a 50% reduction in GCLc levels compared to fibroblasts from the same mice. MLEC obtained from haploinsufficient mice showed significant reduction in GSH levels as well as increased basal and stimulated ROS levels, reduced phosphorylation of eNOS (Ser 1176) and increased eNOS S-glutathionylation, compared to MLEC from wild type (WT) mice. Studies in mesenteric arteries demonstrated impaired endothelium-dependent vasodilation in GCLc e/+ male mice, which was corrected by pre-incubation with ethyl-ester GSH and BH4. To study the contribution of endothelial GSH synthesis to renal fibrosis we employed the unilateral ureteral obstruction model in WT and GCLc e/+ mice. We observed that obstructed kidneys from GCLc e/+ mice exhibited increased deposition of fibrotic markers and reduced Nrf2 levels. We conclude that the preservation of endothelial GSH biosynthesis is critical for endothelial function and anti-fibrotic response.

Endothelial dysfunction in individuals born after fetal growth restriction: cardiovascular and renal consequences and preventive approaches.

Individuals born after intrauterine growth restriction (IUGR) have an increased risk of perinatal morbidity/mortality, and those who survive face long-term consequences such as cardiovascular-related diseases, including systemic hypertension, atherosclerosis, coronary heart disease and chronic kidney disease. In addition to the demonstrated long-term effects of decreased nephron endowment and hyperactivity of the hypothalamic-pituitary-adrenal axis, individuals born after IUGR also exhibit early alterations in vascular structure and function, which have been identified as key factors of the development of cardiovascular-related diseases. The endothelium plays a major role in maintaining vascular function and homeostasis. Therefore, it is not surprising that impaired endothelial function can lead to the long-term development of vascular-related diseases. Endothelial dysfunction, particularly impaired endothelium-dependent vasodilation and vascular remodeling, involves decreased nitric oxide (NO) bioavailability, impaired endothelial NO synthase functionality, increased oxidative stress, endothelial progenitor cells dysfunction and accelerated vascular senescence. Preventive approaches such as breastfeeding, supplementation with folate, vitamins, antioxidants, L-citrulline, L-arginine and treatment with NO modulators represent promising strategies for improving endothelial function, mitigating long-term outcomes and possibly preventing IUGR of vascular origin. Moreover, the identification of early biomarkers of endothelial dysfunction, especially epigenetic biomarkers, could allow early screening and follow-up of individuals at risk of developing cardiovascular and renal diseases, thus contributing to the development of preventive and therapeutic strategies to avert the long-term effects of endothelial dysfunction in infants born after IUGR.

Role of Traditional Cardiovascular Risk Factors for the Occurrence of Erectile Dysfunction in Patients with Coronary Artery Disease

Background: Erectile Dysfunction (ED) is defined as the inability to achieve or maintain an erection sufficient to permit satisfactory sexual intercourse. Erectile dysfunction affects more than 150 million men worldwide and impairs psychological well-being and personal relationships, hence quality of life. Recent studies have shown that ED is present in 42% to 76% of men with coronary artery disease (CAD). Epidemiological study showed clearly role of traditional cardiovascular risk factors such as diabetes, hypertension, dyslipidemia and smoking in CAD. Erectile dysfunction and vascular diseases share a similar risk factors and pathogenic involvement of nitric oxide (NO)-pathway leading to impairment of endothelium-dependent vasodilatation (early phase) and structural vascular abnormalities (late phase). This study was conducted to determine whether the stable CAD patients who have traditional cardiovascular risk factors has a higher risk for ED compared with stable CAD patients without traditional cardiovascular risk factors.Methods: We performed an age matched-paired case control study. Men with CAD documented by angiography were evaluated for ED. Erectile function was assessed by a 5-item version of the International Index of Erectile Function (IIEF-5). Traditional cardiovascular risk factors such as diabetes, hypertension, dyslipidemia and cigarette smoking were assesed. Depression and anxiety were screened using Indonesian version of Hospital Anxiety and Depression Scale (HADS). Basic demographic and other variables were also collected.Results: This study evaluated 127 men, 96.8% of them had traditional cardiovascular risk factors, 25.2% had diabetes mellitus, 77.2% had dyslipidemia, 55.9% had hypertension and 64.6% were smoker. Traditional cardiovascular risk factors was strongly associated with ED (OR=10.67 [1.25-232.83]). ED was independently associated with diabetes mellitus (OR=4.17 [1.14-15.24]), hypertension (OR=2.64 [1.07-6.49]) and cigarette smoking (OR=2.26 [1.01-5.75]).Conclusion: CAD patients with traditional cardiovascular risk factor had more risk for developing ED than those with no traditional cardiovascular risk factor.

Impaired Hydrogen Sulfide-Mediated Vasodilation Contributes to Microvascular Endothelial Dysfunction in Hypertensive Adults.

Reductions in hydrogen sulfide (H2S) production have been implicated in the pathogenesis of vascular dysfunction in animal models of hypertension; however, no studies have examined a functional role for H2S in contributing to microvascular dysfunction in hypertensive (HTN) adults. We hypothesized that endogenous production of H2S would be reduced, impaired endothelium-dependent vasodilation would be mediated by reductions in H2S-dependent vasodilation, and vascular responsiveness to exogenous H2S (sodium sulfide) would be attenuated in HTN compared to normotensive adults. Fifteen normotensive (51±2 years; blood pressure, 116±3/76±3 mm Hg) and 14 HTN adults (57±2 years; blood pressure 140±3/89±2 mm Hg) participated. H2S biosynthetic enzyme expression (Western blot) and substrate-dependent H2S production (amperometric probe) were measured in cutaneous tissue homogenates. Red cell flux (laser Doppler flowmetry) was measured during graded perfusions of acetylcholine (ACh; 10-6-10-1 mol/L) and sodium sulfide (10-5-101 mol/L) using intradermal microdialysis; the functional role of H2S was determined using pharmacological inhibition with aminooxyacetic acid (0.5 mmol/L). H2S biosynthetic enzyme expression and substrate-dependent H2S production were reduced in HTN adults (all P<0.05). ACh-induced endothelium-dependent vasodilation was blunted in HTN adults (P=0.012). Aminooxyacetic acid attenuated ACh-induced vasodilation in normotensive adults (ACh, 1.31±0.13 versus ACh+aminooxyacetic acid, 1.07±0.09 flux/mm Hg; P=0.025) but had no effect on vasodilation in HTN adults (ACh, 1.16±0.10 versus ACh+aminooxyacetic acid, 1.37±0.11 flux/mm Hg; P=0.47). Sodium sulfide-induced vasodilation was not different between groups. Collectively, these findings indicate that while the microvasculature maintains the ability to vasodilate in response to exogenous H2S, reductions in endogenous synthesis and H2S-dependent vasodilation contribute to endothelial dysfunction in human hypertension.