A. Tanju Ozcelikay

The influence of diabetes on cardiac β-adrenoceptor subtypes

Despite the significant developments in the treatment of diabetes mellitus, diabetic patients still continue to suffer from cardiac complications. The increase of cardiac adrenergic drive may ultimately contribute to the development and progression of diabetic cardiomyopathy. β-Adrenoceptors play an important role in the regulation of heart function. However, responsiveness of diabetic heart to β-adrenoceptor agonist stimulation is diminished. The chronotropic responses mediated by β1-subtype, which is mainly responsible for cardiac effects of catecholamines are decreased in the atria of diabetic rats. The expression of cardiac β1-subtype is significantly decreased in diabetic rats as well. β2-Adrenoceptors also increase cardiac function. Although the expression of this subtype is slightly decreased in diabetic rat hearts, β2-mediated chronotropic responses are preserved. On the other hand, functional β3-adrenoceptor subtype was characterized in human heart. Interestingly, stimulation of cardiac β3-adrenoceptors, on the contrary of β1- and β2-subtypes, mediates negative inotropic effect in human ventricular muscle. Cardiac β3-adrenoceptors are upregulated in experimental diabetes as well as in human heart failure. These findings suggest that each β-adrenoceptor subtype may play an important role in the pathophysiology of diabetes-induced heart disease. However, it is still not known whether the changes in the expression and/or responsiveness of β-adrenoceptors are adaptive or maladaptive. Therefore, this review outlines the potential roles of these receptor subtypes in cardiac pathologies of diabetes.

Effects of L-arginine on blood pressure and metabolic changes in fructose-hypertensive rats.

In the present study, we examined the effects of chronic L-arginine treatment on plasma insulin levels and systolic blood pressure (SBP) in fructose-fed (F) rats. Fructose feeding resulted in hyperinsulinemia and elevated blood pressure when compared with that in controls (plasma insulin, 311.3+/-11.4 v control 164.4+/-11.8 pmol/L, P < .05; SBP, 135.4+/-4.2 v control 105.5+/-1.3 mm Hg, P < .05). L-arginine treatment of fructose-hypertensive rats prevented the development of hyperinsulinemia and hypertension (plasma insulin, 200.1+/-7.5 pmol/L; P < .05 compared with that in F rats; SBP, 108.0+/-0.9 mm Hg; P < .05 compared with F rats). However, treatment with L-arginine did not influence any of these parameters in control rats. Statistical analysis of the data of plasma insulin level and SBP, revealed a significant correlation between these two variables. On the other hand, L-arginine treatment of F rats prevented the increased glucose and insulin concentrations in response to oral glucose challenge. L-arginine treatment also prevented the decrease in insulin sensitivity of F rats. These results indicate that L-arginine treatment is able to prevent fructose-induced hypertension and hyperinsulinemia. Our data also suggest a strong relationship between hyperinsulinemia and hypertension in this hypertensive rat model. Therefore, the antihypertensive effect of L-arginine could be, at least in part, the result of the restoration of plasma insulin levels by its vasodilator ability to increase blood flow to insulin sensitive tissues.

The effects of chronic L-arginine treatment on vascular responsiveness of streptozotocin-diabetic rats

In this study, the protective effects of L-arginine treatment in vivo on vascular reactivity of streptozotocin (STZ)-induced 12-week-old diabetic rats were examined. Loss of weight, polydipsia, polyphagia, hyperglycemia, hypoinsulinemia, and elevated levels of plasma cholesterol and triglyceride were observed in diabetic rats. L-arginine treatment (1 mg/mL in drinking water) did not significantly affect these metabolic and biochemical abnormalities. Plasma malondialdehyde (MDA) levels in untreated diabetic rats were also significantly higher than untreated controls. However, L-arginine treatment prevented the increase in MDA level of plasma of diabetic rats. Contractile responses, but not sensitivity to noradrenaline (NA), were significantly increased in diabetic rats compared to controls. Treatment of diabetic rats with L-arginine completely prevented the increase in NA responses. Relaxation response to acetylcholine (ACh), but not to sodium nitroprusside (SNP), in diabetic aorta has been found to be significantly decreased as compared with controls. However, there were no significant differences in pD2 values of acetylcholine in either of the groups. L-arginine treatment increased the ACh responses to the control level. All effects of L-arginine on vascular reactivity were found to be specific for diabetic rats and not controls. These results suggest that functional abnormalities occurred in aorta from diabetic rat might at least in part result from L-arginine deficiency, and the lipid peroxidation-lowering effect of L-arginine may account for its protective effect on vascular reactivity of diabetic rats.

Decreased β-adrenergic responses of rat small intestine due to non-insulin-dependent diabetes

Abstract Relaxing responses to salbutamol of duodenum, jejunum and ileum isolated from alloxan-induced non-insulin-dependent diabetic rats were investigated by means of pharmacodynamic analysis using pD 2 (apparent receptor affinity) and α E (intrinsic activity) values. β-Adrenergic responses of these three intestinal parts of diabetic rats were found to be significantly decreased when compared to controls. The decreased responses may be attributable to a decrease in the number of β-adrenergic receptors. The gastro-intestinal manifestations reported in diabetic patients are possibly linked to the occurrence of a decrease in β-adrenergic receptors. Furthermore, this model seems to be useful for the investigations of gastro-intestinal complications resulting from non-insulin-dependent diabetes.

The effect of vanadate on alloxan-diabetic rat atria

In this study, we examined the effect of vanadate treatment on cardiac changes recognized in diabetic rats. In addition, the possible contribution of thyroid hormones to vanadates effect on alloxan-diabetic atria was also investigated. Administration of alloxan to rats, as expected, resulted in hyperglycemia; hypoinsulinemia reduced thyroid hormone levels, decreased body weight and depressed cardiac function. Vanadate treatment of diabetic rats normalized blood glucose and serum thyroid hormone levels, neither were serum insulin levels of diabetic animals corrected after vanadate treatment. Vanadate treatment, however, did not affect the body weights of diabetic rats. Spontaneously-beating atria from diabetic rats were found to have decreased rates but increased forces of contractions compared with those from controls. On the other hand, the responsiveness of diabetic atria to both inotropic and chronotropic effects of isoprenaline was found to be decreased. Vanadate treatment resulted in the normalization of these alterations observed in diabetic atria. These results thus indicate that the normalizing effect of vanadate on diabetes-induced hypothyroidism may contribute to its effect in preventing cardiac changes observed at the early stage of diabetes.

In situ niosome forming maltodextrin proniosomes of candesartan cilexetil: In vitro and in vivo evaluations.

The aim of this study is to develop novel proniosomal tablets of candesartan cilexetil. Drug loaded proniosomes were prepared as dry powder by slurry method. The critical parameters of the production process were the type of the carrier (sorbitol, maltodextrin, and lactose), addition of charge inducers, rotation speed of the rotavapor and solvent evaporation temperature. The effects of these parameters on proniosome specifications such as the recovery, drug loading, drug release were investigated and also the influence on the particle size and surface charge of the niosomes derived by the hydration of proniosomes were evaluated. The mean particle size and drug loading of niosomes formed from the hydrated proniosomes were 204 ± 2 nm and 99.09 ± 0.04% respectively, with a negative charge -43.65 ± 0.54 mV. The proniosomes demonstrated good flowability, compressibility and consolidation properties both alone and together with the tableting agents (microcrystalline cellulose and cross-linked poly vinylpyrrolidone). The niosomes derived by the hydration of proniosomal tablets preserved their initial properties. Compatibly with the increased in vitro drug dissolution rate, the relative bioavailability of drug from proniosomal tablets increased 1.86-fold and 2.75-fold and higher candesartan plasma levels were achieved within shorter time compared to the pure drug.

The effects of chronic trimetazidine treatment on mechanical function and fatty acid oxidation in diabetic rat hearts

Clinical and experimental evidence suggest that increased rates of fatty acid oxidation in the myocardium result in impaired contractile function in both normal and diabetic hearts. Glucose utilization is decreased in type 1 diabetes, and fatty acid oxidation dominates for energy production at the expense of an increase in oxygen requirement. The objective of this study was to examine the effect of chronic treatment with trimetazidine (TMZ) on cardiac mechanical function and fatty acid oxidation in streptozocin (STZ)-diabetic rats. Spontaneously beating hearts from male Sprague-Dawley rats were subjected to a 60-minute aerobic perfusion period with a recirculating Krebs-Henseleit solution containing 11 mmol/L glucose, 100 muU/mL insulin, and 0.8 mmol/L palmitate prebound to 3% bovine serum albumin (BSA). Mechanical function of the hearts, as cardiac output x heart rate (in (mL/min).(beats/min).10-2), was deteriorated in diabetic (73 +/- 4) and TMZ-treated diabetic (61 +/- 7) groups compared with control (119 +/- 3) and TMZ-treated controls (131 +/- 6). TMZ treatment increased coronary flow in TMZ-treated control (23 +/- 1 mL/min) hearts compared with untreated controls (18 +/- 1 mL/min). The mRNA expression of 3-ketoacyl-CoA thiolase (3-KAT) was increased in diabetic hearts. The inhibitory effect of TMZ on fatty acid oxidation was not detected at 0.8 mmol/L palmitate in the perfusate. Addition of 1 mumol/L TMZ 30 min into the perfusion did not affect fatty acid oxidation rates, cardiac work, or coronary flow. Our results suggest that higher expression of 3-KAT in diabetic rats might require increased concentrations of TMZ for the inhibitory effect on fatty acid oxidation. A detailed kinetic analysis of 3-KAT using different concentrations of fatty acid will determine the fatty acid inhibitory concentration of TMZ in diabetic state where plasma fatty acid levels are increased.

Nebivolol prevents desensitization of β-adrenoceptor signaling and induction of cardiac hypertrophy in response to isoprenaline beyond β1-adrenoceptor blockage.

The importance of chronic stimulation of β-adrenoceptors in the development of cardiac dysfunction is the rationale for the use of β-blockers in the treatment of heart failure. Nebivolol is a third-generation β-blocker, which has further properties including stimulation of endothelial nitric oxide synthase and/or β3-adrenoceptors. The aim of this study was to investigate whether nebivolol has additional effects on β-adrenoceptor-mediated functional responses along with morphologic and molecular determinants of cardiac hypertrophy compared with those of metoprolol, a selective β1-adrenoceptor blocker. Rats infused by isoprenaline (100 μg·kg(-1)·day(-1), 14 days) were randomized into three groups according to the treatment with metoprolol (30 mg·kg(-1)·day(-1)), nebivolol (10 mg·kg(-1)·day(-1)), or placebo for 13 days starting on day 1 after implantation of minipump. Both metoprolol and nebivolol caused a similar reduction on heart rate. Nebivolol mediated a significant improvement on cardiac mass, coronary flow, mRNA expression levels of sarcoplasmic reticulum Ca(2+) ATPase (SERCA2a) and atrial natriuretic peptide and phospholamban (PLN)/SERCA2a and phospho-PLN/PLN ratio compared with metoprolol and placebo. Nebivolol prevented the detrimental effects of isoprenaline infusion on isoprenaline (68% of control at 30 μM), BRL37344 (63% of control at 0.1 μM), and forskolin (64% of control at 1 μM) responses compared with metoprolol (isoprenaline, 34% of control; BRL37344, no response; forskolin, 26% of control) and placebo (isoprenaline, 33% of control; BRL37344, 28% of control; forskolin, 12% of control). Both β-blockers improved the changes in mRNA expressions of β1- and β3-adrenoceptors. Our results suggest that nebivolol partially protects the responsiveness of β-adrenoceptor signaling and the development of cardiac hypertrophy independent of its β1-adrenoceptor blocking effect.

Troglitazone has no effect on KATP channel opener induced-relaxations in rat aorta and in human saphenous veins from patients with type 2 diabetes

Troglitazone, a thiazolidinedione derivative, is an oral antidiabetic agent that enhances insulin sensitivity in insulin-resistant states. K(ATP) channels, on the other hand, have important roles protecting cardiovascular system in ischemic and/or hypoxic states. They are also important in the control of vascular tone, and therefore of blood pressure. We tested whether troglitazone can directly affect vascular K(ATP) channel opener-induced relaxations in vitro. 1, 10 or 100 microM troglitazone incubations for 30 min did not alter cromakalim (a K(ATP) channel opener)--induced relaxations in endothelium-denuded aortas from rat, saphenous veins from type 2 diabetic and nondiabetic patients. In addition, we compared the sensitivity to cromakalim in diabetic saphenous veins with that of nondiabetic veins. The concentration-response curve for cromakalim was shifted to the right in diabetic vein. pD2 values for cromakalim were 6.85+/-0.08 vs. 6.61+/-0.04 (p<0.05) in nondiabetic (n:10) and diabetic (n:7) veins respectively. % maximum response of cromakalim was also significantly decreased by 24+/-3% in diabetic veins. However, responsiveness of veins to phenylephrine or sodium nitroprusside were similar in both groups. The results obtained may be clinically useful 1. suggesting that in ischaemic and/or hypoxic insults troglitazone may not worsen vascular dilatation, through K(ATP) channel, in diabetic patients who are more prone to these conditions than healthy people, 2. providing an evidence that diabetes causes an impaired dilatation of human saphenous vein through K(ATP) channels. This may partly be related with diabetes-induced vascular complications, such as vasospasm and even hypertension. Accordingly, since saphenous veins are used as conduit vessels in coronary by-pass graft surgery, the results also suggest that the defective dilatation through K(ATP) channels may play a role on the performance of saphenous vein grafts in type 2 diabetes.

Role of Beta-3-Adrenoceptor in Catecholamine-Induced Relaxations in Gastric Fundus from Control and Diabetic Rats

The contribution of β-adrenoceptor subtypes to the catecholamine-mediated relaxations in gastric fundus from control and streptozotocin (STZ)-induced diabetic rats were investigated. Isolated organ bath studies and molecular techniques were used to characterize the β-adrenoceptor subtypes mediating relaxation of rat gastric fundus. Isoprenaline-mediated relaxation was not significantly changed by nadolol (β1-/β2-adrenoceptor antagonist; 1 µmol/l) but only shifted to the right by SR59230A (3-(2-ethylphenoxy)-1-[[(1S)-1,2,3,4-tetrahydronaphth-1-yl]amino]-(2S)-2-propanol oxalate salt, 0.1–1 µmol/l), a selective β3-adrenoceptor antagonist, in a competitive manner. Relaxant responses to noradrenaline were antagonized in a concentration-dependent manner by SR59230A (0.1–1 µmol/l), but not by metoprolol (selective β1-adrenoceptor antagonist; 0.1–1 µmol/l) and ICI-118551 (1-[2,3-(dihydro-7-methyl-1Hinden- 4-yl)oxy]-3-[(1-methylethyl)amino]-2-butanol hydrochloride, selective β2-adrenoceptor antagonist; 0.1–1 µmol/l). SR59230A (1 µmol/l) also caused a significant rightward shift in fenoterol-induced relaxation while ICI-118551 (1 µmol/l) did not have any effect. Selective β3-adrenoceptor agonist, BRL37344 ([4-[2-[[2-(3-chlorophenyl)-2-hydroxyethyl]amino]propyl]phenoxy]acetic acid), caused biphasic relaxation which was not affected by nadolol (1 µmol/l). SR59230A (1 µmol/l) abolished only the first phase of BRL37344 response. β1-, β2- and β3-adrenoceptor mRNA expressions have been detected in a similar intensity in gastric fundus from control rats. Experimental diabetes caused a significant decrease in Emax and pD2 values of isoprenaline and noradrenaline. Diabetes also reduced Emax but not pD2 value of the first component of BRL37344-induced relaxation response. The band intensity of mRNA transcript of β3-adrenoceptor was reduced in diabetics while no alteration has been found for β1- and β2-adrenoceptor mRNA transcripts between groups. These results show that functional β-adrenoceptor subtype involved in catecholamine-mediated relaxations is β3-adrenoceptor, and its function and mRNA expression are decreased in diabetes.