Decrease Of Nitric Oxide Content Research Articles

Pleiotropic vascular effects of ivabradine in streptozotocin-induced diabetes

AimsIvabradine (IVA) reduces heart rate (HR) by inhibiting hyperpolarization-activated cyclic nucleotide-gated (HCN) channels in sinoatrial node. Studies suggest that IVA has other beneficial effects on cardiovascular system that are not related to its effect on HR such as prevention of endothelial injury and the antioxidant effects. In addition to sinoatrial node, HCN channels exist in other tissues and their expression pattern differs in certain pathologies such as hypertension and hypertrophy. We investigated the mechanism of IVA effect in the setting of streptozotocin (STZ)-induced cardiovascular damage. Direct effects of IVA and their mechanism on thoracic aorta as well as possible prevention of vascular dysfunction in diabetes were investigated in this study. Methods and resultsThe effects of IVA on vascular function were investigated in control and STZ-diabetic rats. Some control and diabetic rats were treated with IVA. IVA treatment prevented diabetes-induced increase in plasma p-selectin and vascular cell adhesion molecule-1 levels and the decrease in nitric oxide content in the aortas of diabetic animals. When added to isolated organ bath, IVA induced concentration-dependent relaxations in thoracic aorta. Pre-incubation with Nω-Nitro- L -arginine methyl ester reduced IVA-induced relaxations. Expression patterns of all isoforms of HCN proteins were affected by both diabetes and IVA treatment. ConclusionIVA improves vascular function in diabetes and HCN channels support vascular activity against damaging effects of diabetes. IVA may be added to prevent diabetic cardiovascular dysfunction with these beneficial effects that are unrelated to its primary mechanism of action.

Disorders of microcirculation in a comorbid patient

Background. Diabetes mellitus (DM) is a global epidemic. According to the experts, by 2045 the number of patients with diabetes will rise by 48 %. Patients with diabetes are characterized by the high cardiovascular mortality and a significant reduction in life expectancy. Almost half of the deaths associated with diabetes are due to cardiovascular diseases (CVD). Patients with diabetes often have concomitant pathology (hypertension, dyslipidemia, obesity), which increases the cardiovascular risk.
 Objective. To describe microcirculation disorders in patients with comorbid conditions.
 Materials and methods. Analysis of literature sources on this topic.
 Results and discussion. Type 2 diabetes (DM2) is a multifaceted disease that has a number of cardiovascular, metabolic and renal complications. The links of the pathogenesis of cardiovascular complications of DM2 include dyslipidemia, systemic inflammation, insulin resistance, autonomic imbalance, and endothelial dysfunction (ED). ED leads to vasoconstriction, increased chronic inflammation, increased vascular permeability and hypercoagulation, which ultimately causes micro- and macroangiopathy. Neurological complications of diabetes are also mediated by ED and microangiopathies, which lead to nerve hypoxia with a decrease in conductivity velocity. In recent years, the literature has been actively discussing the syndrome of early vascular aging – premature and accelerated development of structural and functional age-related changes in blood vessels. Microcirculation disorders that accompany diabetes, CVD and their complications are mediated by nitric oxide (NO) imbalance. NO reduces the contractility and proliferation of smooth muscle cells, platelet aggregation, endothelin production, adhesion of monocytes and platelets, and oxidation of low-density lipoproteins. In humans, NO is produced from L-arginine. Increased L-arginase activity and decreased NO-synthase activity lead to a decrease in NO content and to the development of ED, atherosclerosis, and decreased insulin sensitivity. L-arginine as a substrate of NO improves the functional state of the endothelium, reduces the manifestations of oxidative stress, reduces the level of pro-inflammatory cytokines and adhesion molecules, inhibits platelet aggregation, reduces insulin resistance. It should be noted that the administration of 4.2 g of L-arginine in DM is not enough. In clinical studies, doses of 6.4-9 g per day were found to be effective in diabetes and obesity. Therefore, it is advisable to use Tivortin-200 (“Yuria-Pharm”), which contains 8.4 g of L-arginine in one vial.
 Conclusions. 1. Patients with diabetes are characterized by the high cardiovascular mortality and a significant reduction in life expectancy. 2. The links of the pathogenesis of cardiovascular complications of DM2 include dyslipidemia, systemic inflammation, insulin resistance, autonomic imbalance, and ED. 3. Disorders of microcirculation that accompany diabetes, CVD and their complications, are mediated by NO imbalance. 4. L-arginine as a substrate of NO improves the functional state of the endothelium, reduces the manifestations of oxidative stress, reduces the level of pro-inflammatory cytokines and adhesion molecules, inhibits platelet aggregation, reduces insulin resistance. 5. In diabetes and obesity, it is advisable to use Tivortin-200, which contains 8.4 g of L-arginine in one vial.

Some Aspects of Role of Nitric Oxide in the Mechanisms of Hypertension (Experimental Study).

BackgroundModulation of endothelial function is a therapeutic option to reduce some of the significant complications of hypertension. However, the relationship between endothelial dysfunction reduced nitric oxide (NO) production, and the development of hypertension is not fully understood. To establish a potential pathogenetic link between impaired NO synthesis and hypertension, we investigated the results of competitive interaction of the substrate of NO synthase, L-arginine, and its analog, an non-selective inhibitor of NO synthase, N-nitro-methyl ether-L-arginine (L-NAME), in experimental rats.MethodsArterial hypertension was induced in male Wistar rats by intraperitoneal administration of L-NAME (Sigma-Aldrich) for 4 - 7 weeks. During the last 3 weeks, to a separate group of animals simultaneously with L-NAME, L-arginine (Sigma-Aldrich) was administered. In animals monitored for systolic and diastolic pressure, the level of NO in blood samples was determined spectrophotometrically using a Griess reagent.ResultsAdministration of L-NAME for 4 - 7 weeks induced an irreversible decrease of NO content in blood, a reversible increase of systolic pressure (SP) and diastolic pressure (DP), and an irreversible increase in pulse pressure (PP). In rats against the background of 7 weeks of intraperitoneal administration of L-NAME, during the last 3 weeks, they were injected with L-arginine, the SP and DP indices returned to their initial values, PP decreased and the NO content in arterial blood increased.ConclusionsThe results of the study indicate the presence of residual endothelial dysfunction (characterized by insufficient NO) after the correction of hypertension. Therefore, in developing the new therapeutic approaches for the treatment of hypertension, it is necessary to include drugs that, in addition to correcting blood pressure, will support normalization, and potentiation of endothelial function and endogenous NO synthesis.

PL - 032 Physiological triggers involved in reduced slow myosin expression in disused postural muscle

PL - 032 Physiological triggers involved in reduced slow myosin expression in disused postural muscle

Brain Oxidative Stress and Neurodegeneration in the Ketamine Model of Schizophrenia during Antipsychotic Treatment: Effects of N-Acetylcysteine Treatment

We aimed to investigate the effect of N-acetylcysteine (NAC) on brain oxidative stress and on the response to therapy with clozapine or haloperidol in the ketamine model of schizophrenia. Mice received intraperitoneal (i.p.) injections of either saline, ketamine (30 mg/kg), NAC (50 mg/kg), ketamine + NAC, ketamine + clozapine (1.5 mg/kg), ketamine + clozapine + NAC, ketamine + haloperidol (1.5 mg/kg), or ketamine + haloperidol + NAC daily for one week. Malondialdehyde (MDA), reduced glutathione (GSH), nitric oxide, and paraoxonase-1 (PON-1) activity were determined in cerebral cortex and in the rest of the brain tissue. Results indicated that: (i) treatment with only NAC had no significant effects on MDA, GSH, or PON-1 activity, but resulted in a marked decrease in nitric oxide content in both the cortex and the rest of the brain compared to the saline control group; (ii) ketamine caused a significant increase in MDA, while decreasing nitric oxide, and GSH concentrations as well as PON-1activity in both the cerebral cortex and the rest of the brain. These biochemical alterations were alleviated by NAC; (iii) in ketamine-treated mice, either clozapine or haloperidol had no significant effect on MDA or GSH levels. Nitric oxide, however, showed a significant increase by either agent. Meanwhile, PON-1 activity showed a significant increase by clozapine in the cortex but was decreased by haloperidol in both the cortex and the rest of the brain compared with the ketamine only group; (iv) NAC resulted in a significant decrease in MDA in the cortex in ketamine + haloperidol-treated mice, and markedly alleviated the decline in GSH and PON-1 activity in both the cortex and the rest of the brain tissue in ketamine + clozapine- or ketamine + haloperidol-treated groups. NACshowed variable effects on brain nitric oxide in these groups; and (v) ketamine caused neurodegeneration in the cortex and striatum changes in the form of shrunken neurons, pyknotic and apoptotic nuclei, perineuronal vacuolations, and red neurons. These pathological changes were marked after treatment with either clozapine or haloperidol but ameliorated by NAC treatment. These data indicate increased brain oxidative stress in the schizophrenia model induced by ketamine in mice and that treatment with antipsychotics impairs brain antioxidants. The study suggests a potential therapeutic benefit for NACin alleviating brain oxidative stress, lipid peroxidation, and neurodegeneration during antipsychotic drug therapy in schizophrenia.

English

The aim of this study was to investigate the efficiency of intrathecal glial cell line-derived neurotrophic factor (GDNF) on nitric oxide (NO) and nitric oxide synthase (NOS) activity following a spinal nerve ligation (SNL) of male Sprague Dawley rats. The rats were randomly divided into four groups: normal (control), sham-operated, SNL (SNL followed by a physiological saline injection into the subarachnoid space), and GDNF (SNL followed by a GDNF injection into the subarachnoid space). Each group was divided into three subgroups (n = 10). The rats in each subgroup were euthanised 3, 7, and 14 days after the operation. Rat behaviour was evaluated before euthanising, and the ipsilateral spinal cords were harvested after euthanising to determine the NO content and NOS activity. Compared with the control and sham-operated groups, the NO content and NOS activity in the SNL group increased significantly 3 days after the operation; this increase was maintained until 14 days after the operation (P < 0.01 or 0.05). A significant decrease was observed in the NO content and NOS activity in the GDNF group compared with the SNL group. The decrease continued until 14 days after the operation (P < 0.01 or 0.05). The results indicated that the NO and NOS activity in the rat spinal cord are associated with SNL-induced neuropathic pain. The decreased neuropathic pain from the intrathecal GDNF is correlated with the decrease in NO content and NOS activity in the spinal cord.   Key words: Glial cell line-derived neurotrophic factor, neuralgia, nitric oxide, nitric oxide synthase.

Protective effect of l-citrulline against ethanol-induced gastric ulcer in rats

We examined the protective effect of l-citrulline on ethanol-induced gastric ulcer in rats. Administration of l-citrulline at doses of 300, 600 and 900mg/kg body weight prior to ethanol ingestion protected the stomach from ulceration. The gastric lesions were significantly attenuated by all doses of l-citrulline as compared to the ethanol group. Pre-treatment with l-citrulline prevented the oxidative damage and the decrease of nitric oxide content as well as the increase of the myeloperoxidase activity. Consequently, significant changes observed included the attenuation in the elevation in total nitric oxide synthase activity and inducible nitric oxide synthase activity as well as the decrease in constitutive nitric oxide synthase activity in the gastric mucosa induced by ethanol. Analysis of serum cytokines of ethanol-induced rats showed a moderate decrease in interleukin-10 with considerable increase of interleukin-6 while l-citrulline inhibited the acute alteration of cytokines. These results suggested the gastroprotective effect of l-citrulline.

Effect of 2,3,5,4′-tetrahydroxystilbene-2-O-beta-d-glucoside on lipoprotein oxidation and proliferation of coronary arterial smooth cells

To investigate the effects of 2,3,5,4′-tetrahydroxystilbene-2-O-beta-d-glucoside (THSG), a compound extracted from the root of Polygonum multiflorum Thunb, on lipoprotein (LDL and VLDL) oxidation, proliferation and nitric oxide (NO) content of coronary arterial smooth cells (CASMCs) induced by LDL, VLDL, ox-LDL and ox-VLDL. The oxidation level of lipoprotein was determined by thiobabituric acid (TBA) method and agarose gel electrophoresis. The proliferative degree was determined by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) method. The NO content was assayed by nitrate reductase method. (1)THSG (0.1 ∼ 100 μmol·L− 1) could dose-dependently prevent lipoprotein from oxidation induced by Cu2+ and CASMCs. (2)THSG (0.1 ∼ 100 μmol·L− 1) inhibited porcine CASMCs proliferation elicited by LDL, VLDL, ox-LDL and ox-VLDL. (3)THSG (0.1 ∼ 100 μmol·L− 1) counterpoised the decrease of NO content in CASMCs evoked by LDL, VLDL, ox-LDL and ox-VLDL. As compared with control, it was found that the threshold concentration of THSG, which significantly exerted the actions mentioned above, were at the concentration of 1μmol·L− 1 (P < 0.01). In conclusion, THSG possesses the antagonistic effects on oxidation of lipoprotein, proliferation and decrease of NO content of CASMCs, which partially explain the mechanism of anti-atherosclerosis of Polygonum multiflorum Thunb.

Impairment of Endothelium-Dependent Relaxation of Rat Aortas by Homocysteine Thiolactone and Attenuation by Captopril

To explore the effects of angiotensin-converting enzyme (ACE) inhibitors on endothelial dysfunction induced by homocysteine thiolactone (HTL). Both endothelium-dependent relaxation and nondependent relaxation of thoracic aortic rings in rats induced by acetylcholine (Ach) or sodium nitroprusside (SNP) and biochemical parameters including malondialdehyde (MDA) and nitric oxide (NO) were measured in rat isolated aorta. Exposure of aortic rings to HTL (3 to 30 mM) for 90 minutes made a significant inhibition of endothelium-dependent relaxation induced by Ach, decreased contents of NO, and increased MDA concentration in aortic tissue. After incubation of aortic rings with captopril (0.003 to 0.03 mM) attenuated the inhibition of endothelium-dependent relaxation (EDR) and significantly resisted the decrease of NO content and elevation of MDA concentration caused by HTL (30 mmol/L) in aortic tissues, a similarly protective effect was observed when the aortic rings were incubated with both N-acetylcysteine (0.05 mM). Treatment with enalaprilat (0.003 to 0.01 mM) made no significant difference with the HTL (30 mM) group regarding EDR, but enalaprilat (0.03 mM) and losartan (0.03 mM) could partly restore the EDR in response to HTL (30 mM). Captopril was more effective than enalaprilat and losartan in attenuation of the inhibition of on acetylcholine-stimulated aortic relaxation by HTL in the same concentration. Moreover, superoxide dismutase (SOD, 200 U/mL), which is a scavenger of superoxide anions, apocynin (0.03 mM), which is an inhibitor of NADPH oxidase, and l-Arginine (3 mmol/L), a precursor of nitric oxide (NO), could reduce HTL (30 mM)-induced inhibition of EDR. After pretreatment with not only the NO synthase inhibitor Nomega-nitro-l-arginine methyl ester (L-NAME, 0.01 mM) but also the free sulfhydryl group blocking agent p-hydroxymercurybenzoate (PHMB, 0.05 mM) could abolish the protection of captopril and N-acetylcysteine, respectively. These results suggest that mechanisms of endothelial dysfunction induced by HTL may include the decrease of NO and the generation of oxygen free radicals and that captopril can restore the inhibition of EDR induced by HTL in isolated rat aorta, which may be related to scavenging oxygen free radicals and may be sulfhydryl-dependent.

Reactive species and apoptosis of neural precursor cells after γ-irradiation

This study addresses the participation of radiation-induced free radicals, mainly nitric oxide (NO), in modulating the apoptotic response in an in vitro model of neural cortical precursor cells exposed to γ-radiation. Cortical cells obtained from rats at 17 gestational day (GD) were irradiated with a dose of 2 Gy. The percentage of apoptotic cells was significantly increased 4 h post-irradiation (pi). NO content showed a significant increase after 30 min pi and the rate of generation reached a maximum 1 h pi. Luminol-dependent chemiluminescence (CL) was significantly higher in cells after 2 h pi as compared to control cells and this profile was maintained up to 4 h pi. Supplementation with l-NAME significantly increased light emission. Administration of superoxide dismutase (SOD) following l-NAME addition prevented the observed changes due to l-NAME administration. The caspase inhibitor zDEVD-fmk significantly reduced the radical generation. Moreover, the cellular decrease in NO content occurred coincidentally with the rise in oxygen radical generation and the activation of caspase-3. In vitro irradiation of neural precursor cells allowed us to suggest that an early radiation-induced generation of NO could exert a neuroprotective role. However, despite this NO initial protective effect and its role modulating the response against γ-radiation, NO generation was not able of fully preventing radiation-induced apoptosis.

Protective effects of cariporide on endothelial dysfunction induced by high glucose1

To explore the effects of cariporide, a selective sodium-hydrogen antiporter inhibitor, on endothelial dysfunction induced by high glucose. Acetylcholine (ACh)-induced endothelium-dependent relaxation (EDR), sodium nitroprusside (SNP)-induced endothelium-independent relaxation and biochemical parameters including malondialdehyde (MDA), superoxide dismutase (SOD), and nitric oxide (NO) were measured in rat isolated aorta. A 6-h incubation of aortic rings with high glucose (44 mmol/L) resulted in a significant inhibition of EDR, but had no effects on endothelium-independent relaxation. After the 6-h incubation of aortic rings in the co-presence of cariporide (0.01, 0.1, and 1 micromol/L) with high glucose, cariporide prevented the inhibition of EDR caused by high glucose in concentration-dependent manners. Similarly, high glucose decreased SOD activity and contents of NO, and increased MDA concentration in aortic tissue. Cariporide (1 micromol/L) significantly resisted the decrease of NO content and SOD activity, and elevation of MDA concentration caused by high glucose in aortic tissues. Mannitol (44 mmol/L) or cariporide (1 micromol/L) alone had no effect on EDR, endothelium-independent relaxation and biochemical parameters. Cariporide significantly prevented endothelial dysfunction induced by high glucose. The mechanisms of endothelial dysfunction induced by high glucose may involve the activation of sodium-hydrogen antiporter and the generation of oxygen-free radicals, but it is not related to the change of osmolarity.

Delayed toe transplantation: experimental study and clinical application.

Delay of the operation in toe transplantation was performed with circulatory crisis, and 100 percent survival of the transplants was achieved. In 10 cases of toe transplantation in which circulatory crisis occurred, the operation was delayed. The vascular pedicle was divided, and the toe was transferred 17 to 21 hours after the donor toe was dissected. To investigate the mechanism of delayed toe transplantation, experiments in rabbits were carried out. In 36 rabbits, the hind limbs were severed except for the vascular pedicle. The endothelin content and nitric oxide content of the tissues in the hind limb and the arterial wall were tested in postoperative intervals of 4, 8, 16, 24, and 48 hours, respectively. The results showed that in 10 cases of delayed toe transplantation, all toes survived. As for the results of the experiment, there was significant increase of endothelin content and decrease of nitric oxide content in the local tissues and the arterial wall 4 to 8 hours after the operation. In postoperative 16 to 24 hours, endothelin content returned to normal level, and nitric oxide content increased remarkably. It is concluded that delayed pedicle division and toe transfer is an effective method in toe transplantation with vascular variations and circulatory crisis. The mechanism is related to the endothelin and nitric oxide content in the local tissue and the arterial wall. The indications for delayed toe transfer are also discussed.

The role of NO and B-50 in neurotoxicity of excitatory amino acids.

To investigate the direct evidence for the role which nitric oxide (NO) plays in the neurotoxicity of excitatory amino acids, we evaluated NO level by Greiss testing solution when glutamate (Glu) and kainate (KA) induced neuronal degeneration in primary cortical cultures. Glutamate-induced neurotoxicity was accompanied by a rise in NO. 5 mM hemoglobin (Hb) led to a decrease of NO content and prevented excitotoxicity induced by 1 mM glutamate. 1 mM L-arginine (L-Arg) reversed the effect of hemoglobin by raising the NO level. No change in NO content was found in KA-induced neurotoxicity, which was not affected by L-Arg, Hb or L-Arg + Hb. It is suggested that NO plays an important role in glutamate-, but not KA-induced neurotoxicity in primary cortical cultures. We also investigated the effects of glutamate on a growth-associated protein, B-50. The B-50 level declined significantly 24 h after exposure to 100 microM glutamate for 30 min and then recovered 2 days later. The effect of glutamate on B-50 was concentration-dependent. This indicates that B-50 might be involved in both glutamate neurotoxicity and the following neuronal repair process.