Effect of intracellular Ca2+ concentration on endothelin-1 secretion

AbstractThe role of intracellular free Ca2+ concentration ([Ca2+]i in cellular regulation of endothelin-(ET-1) secretion was investigated in cultured porcine aortic endothelial cells of first passage. Intracellular Ca2+ concentration were adjusted between 50 nM and 1 μM using EGTA and thapsigargin, respectively. ET-1 secretion was maximal at [Ca2+]i of 190–470 nM, and reduced at low (50 and 110 nM) and high (470 nM) [Ca2+]i. The Ca2+ ionophores A23187 and ionomycin (each 1 μM), both of which raise [Ca2+]i above 1 μM, also potently inhibited ET-1 secretion under basal and stimulated conditions. The A23187-induced reduction in ET-1 secretion was not affected by NG-nitro-l-arginine (0.1 mM). Our results provide evidence that basal ET-1 secretion is regulated by Ca2+ and that Ca2+ ionophores reduce ET-1 secretion due to the inhibitory effect of high [Ca2+]i

Binding of endothelin to plasma proteins and tissue receptors: effects on endothelin determination, vasoactivity, and tissue kinetics

AbstractIn vitro binding of (3-[125I]Tyr)-endothelin-1 ([125I]ET-1) and (3-[125I]Tyr)-big ET-1(1–38) ([125I]big ET-1) to plasma proteins of healthy humans, cardiac patients and normotensive and hypertensive rats was investigated by equilibrium dialysis. Binding of both tracers was similar in plasma from healthy humans, patients with congestive heart failure, and following myocardial infarction (∼60%), and marginally higher in rat plasmas (∼70%). Binding of [125I]ET-1 to human plasma could be explained by binding to human serum albumin. Endogenous plasma ET-1 levels were ∼9 pg/ml in healthy humans, and ∼12–16 pg/ml in cardiac patients; big ET-1 concentrations were approximately two- to threefold higher. ET-1 bound to plasma protein was partly lost in column extraction. In rat isolated perfused hearts, the coronary dilator and constrictor potency of exogenous free and albumin-bound ET-1 was similar, whereas the kinetics of endogenous ET-1 was impeded by tight binding to ET receptors. The data indicate that binding of ET-1 to plasma proteins is without effect on peptide vasoactivity, but binding to tissue receptors greatly impedes its tissue kinetics

Effects of flavoring compounds used in electronic cigarette refill liquids on endothelial and vascular function.

Electronic cigarette refill liquids are commercially provided with a wide variety of flavoring agents. A recent study suggested that several common flavors may scavenge nitric oxide (NO) and cause endothelial dysfunction. It was the aim of the present study to investigate the effects of these flavors on NO/cyclic GMP-mediated signaling and vascular relaxation. We tested the flavoring agents for effects on Ca2+-induced cGMP accumulation and NO synthase activation in cultured endothelial cells. NO scavenging was studied with NO-activated soluble guanylate cyclase and as NO release from a NO donor, measured with a NO electrode. Blood vessel function was studied with precontracted rat aortic rings in the absence and presence of acetylcholine or a NO donor. Cinnamaldehyde inhibited Ca2+-stimulated endothelial cGMP accumulation and NO synthase activation at ≥0.3 mM. Cinnamaldehyde and diacetyl inhibited NO-activated soluble guanylate cyclase with IC50 values of 0.56 (0.54-0.58) and 0.29 (0.24-0.36) mM, respectively, and caused moderate NO scavenging at 1 mM that was not mediated by superoxide anions. The other compounds did not scavenge NO at 1 mM. None of the flavorings interfered with acetylcholine-induced vascular relaxation, but they caused relaxation of pre-contracted aortas. The most potent compounds were eugenol and cinnamaldehyde with EC50 values of ~0.5 mM. Since the flavors did not affect endothelium-dependent vascular relaxation, NO scavenging by cinnamaldehyde and diacetyl does not result in impaired blood vessel function. Although not studied in vivo, the low potency of the compounds renders it unlikely that the observed effects are relevant to humans inhaling flavored vapor from electronic cigarettes

Endothelial dysfunction in adipose triglyceride lipase deficiency

AbstractSystemic knockout of adipose triglyceride lipase (ATGL), the pivotal enzyme of triglyceride lipolysis, results in a murine phenotype that is characterized by progredient cardiac steatosis and severe heart failure. Since cardiac and vascular dysfunction have been closely related in numerous studies we investigated endothelium-dependent and -independent vessel function of ATGL knockout mice. Aortic relaxation studies and Langendorff perfusion experiments of isolated hearts showed that ATGL knockout mice suffer from pronounced micro- and macrovascular endothelial dysfunction. Experiments with agonists directly targeting vascular smooth muscle cells revealed the functional integrity of the smooth muscle cell layer. Loss of vascular reactivity was restored ~50% upon treatment of ATGL knockout mice with the PPARα agonist Wy14,643, indicating that this phenomenon is partly a consequence of impaired cardiac contractility. Biochemical analysis revealed that aortic endothelial NO synthase expression and activity were significantly reduced in ATGL deficiency. Enzyme activity was fully restored in ATGL mice treated with the PPARα agonist. Biochemical analysis of perivascular adipose tissue demonstrated that ATGL knockout mice suffer from perivascular inflammatory oxidative stress which occurs independent of cardiac dysfunction and might contribute to vascular defects. Our results reveal a hitherto unrecognized link between disturbed lipid metabolism, obesity and cardiovascular disease

Cardioprotective effects of atrasentan, an endothelin-A receptor antagonist, but not of nitric oxide in diabetic mice with myocyte-specific overexpression of endothelial nitric oxide synthase

1. We investigated the roles of nitric oxide (NO) and endothelin-1 (ET-1) in organ dysfunction in diabetic mice with normal genotype (wild-type, WT) or myocyte-specific overexpression of endothelial NO synthase (eNOS) (transgenic, TG) after chronic oral treatment with the endothelin-A (ET(A)) receptor antagonist atrasentan. 2. Mice were rendered diabetic by injection of 200 mg kg(−1) streptozotocin (STZ). Experimental groups were: untreated WT diabetic (n=9), untreated TG diabetic (n=9), atrasentan-treated WT diabetic (n=9), atrasentan-treated TG diabetic (n=8) and the four corresponding nondiabetic groups (n=5). Atrasentan was administered orally via drinking water at 3 mg kg(−1) per day over 28 days. All diabetic mice developed similar hyperglycaemia (27–30 mmol l(−1)). 3. Atrasentan treatment significantly improved left ventricular systolic and diastolic function in response to exogenous norepinephrine, but there were no differences between genotypes. 4. Atrasentan antagonized the diabetic impairments in endothelium-dependent coronary relaxation and thromboxane-receptor mediated aortic constriction. Further, it improved cardiac and renal oxidant status as evident from reduced tissue malondialdehyde levels. 5. Atrasentan reduced diabetic urine flow, proteinuria and plasma creatinine levels, but creatinine clearance was not significantly altered. 6. These results suggest that in experimental type 1 diabetes, blocking ET(A) receptors ameliorates myocardial, coronary and renal function and improves tissue oxidant status, whereas raising myocardial NO levels has neither beneficial nor deleterious effects on diabetic cardiomyopathy in this transgenic model