Modulatory Role of Nitric Oxide/cGMP System in Endothelin-1-Induced Signaling Responses in Vascular Smooth Muscle Cells

Nitric oxide (NO) is an important vasoprotective molecule that serves not only as a vasodilator but also exerts antihypertrophic and antiproliferative effects in vascular smooth muscle cells (VSMC). The precise mechanism by which the antihypertrophic and antiproliferative responses of NO are mediated remains obscure. However, recent studies have suggested that one of the mechanisms by which this may be achieved includes the attenuation of signal transduction pathways responsible for inducing the hypertrophic and proliferative program in VSMC. Endothelin-1 is a powerful vasoconstrictor peptide with mitogenic and growth stimulatory properties and exerts its effects by activating multiple signaling pathways which include ERK 1/2, PKB and Rho-ROCK. Both cGMP-dependent and independent events have been reported to mediate the effect of NO on these pathways leading to its vasoprotective response. This review briefly summarizes some key studies on the modulatory effect of NO on these signaling pathways and discusses the possible role of cGMP system in this process.     

神经元型钠通道在室性心律失常发生机制中的作用

室性心律失常是导致心脏性猝死最重要的原因,其发生机制与心室肌细胞离子电流紊乱有密切关系。近期研究发现,神经元型钠通道也表达于心脏,是介导晚钠电流的重要通道,可通过以下机制触发室性心律失常:(1)神经元型钠通道活性异常增高引起内向电流增大,破坏动作电位平台期电位平衡,直接引起细胞膜去极化,发生早后除极。(2)神经元型钠通道与钙调控蛋白在T管微域内共定位,其介导的Na+内流通过增强钙调控蛋白功能引起舒张期Ca2+释放,发生迟后除极。(3)晚钠电流具有速率依赖性和分布异质性,可增大复极离散,构成功能性折返的发生基质。现就神经元型钠通道在室性心律失常发生中的作用做一综述。     

Role of Neuronal Nitric Oxide in the Dopamine Deficit of HPRT-Deficient Mice

Lesch-Nyhan disease is a debilitating disorder caused by a lack of purine salvage activity. Basal ganglia dopamine deficits manifest in both patients and hypoxanthine phosphoribosyltransferase (HPRT) mutant mice. We previously reported decreased activity in an oxidant sensitive enzyme in the brain of HPRT-deficient mice. In the present study, we have investigated whether one source of free radicals, neuronal nitric oxide synthase (NOS1), contributes to the dopamine deficit associated with HPRT deficiency. HPRT knockout and wild-type mice were bred, either to lack, or to have the full complement of NOS1 alleles. Double mutant mice had striatal dopamine and dopamine metabolite levels indistinguishable from the HPRT single mutant counterparts. These results indicate that NOS1 produced nitric oxide does not contribute to the dopamine deficit seen in HPRT deficiency.     

Increased Monoester Lipase Activity of Red Blood Cells in Alcoholism

Monoester lipase activity was assayed by a radiochemical assay in the red blood cells (RBC) from 50 chronic alcoholic patients within 48 hr after discontinuation of chronic alcohol intake and from 40 nonalcoholic control subjects. The mean value of lipase activity was increased to 1213 +/- 229 milliunits/10(12) RBC in the alcoholics as compared with a value of 997 +/- 120 milliunits/10(12) RBC in the controls (p less than 0.001). The lipase increase was associated with increased values of the mean cellular volume of RBC. A subgroup (64%) of 32 alcoholic subjects with macrocytosis (mean cellular volume greater than 96 femtoliters) showed the highest mean lipase activity (1276 +/- 224 milliunits/10(12) RBC) as compared with a value of 1101 +/- 196 milliunits/10(12) RBC in the normocytic alcoholic subgroup (p less than 0.05). This latter subgroup had a mean value higher (p less than 0.05) than that in the control group. The relationship between values of mean cellular volume and lipase activity was not of predictive value in individual cases. The enzymatic increase was not related to a direct effect of alcohol on the intact RBC. It is postulated that this alteration might result from changes in the chemical and/or physical state of the plasma membrane induced by ethanol during RBC formation. In any event, the increased lipolytic activity of RBC represents a new biological characteristic of alcoholic subjects. Its determination might represent a noninvasive way of evaluating the influence of alcohol on a tissue parameter.     

The Role of Antioxidant Micronutrients in the Prevention of Diabetic Complications

Diabetes mellitus is associated with an increased production of reactive oxygen species and a reduction in antioxidant defenses. This leads to oxidative stress, which is partly responsible for diabetic complications. Tight glycemic control is the most effective way of preventing or decreasing these complications. Nevertheless, antioxidant micronutrients can be proposed as adjunctive therapy in patients with diabetes. Indeed, some minerals and vitamins are able to indirectly participate in the reduction of oxidative stress in diabetic patients by improving glycemic control and/or are able to exert antioxidant activity. This article reviews the use of minerals (vanadium, chromium, magnesium, zinc, selenium, copper) and vitamins or cofactors (tocopherol [vitamin E], ascorbic acid [vitamin C], ubidecarenone [ubiquinone; coenzyme Q], nicotinamide, riboflavin, thioctic acid [lipoic acid], flavonoids) in diabetes, with a particular focus on the prevention of diabetic complications. Results show that dietary supplementation with micronutrients may be a complement to classical therapies for preventing and treating diabetic complications. Supplementation is expected to be more effective when a deficiency in these micronutrients exists. Nevertheless, many clinical studies have reported beneficial effects in individuals without deficiencies, although several of these studies were short term and had small sample sizes. However, a randomized, double-blind, placebo-controlled, multicenter trial showed that thioctic acid at an oral dosage of 800 mg/day for 4 months significantly improved cardiac autonomic neuropathy in type 2 diabetic patients. Above all, individuals with diabetes should be educated about the importance of consuming adequate amounts of vitamins and minerals from natural food sources, within the constraints of recommended sugar and carbohydrate intake.     

A Combined System for the Study of Glutathione Metabolism in Erythrocytes

A combined system for measuring giutathione stability, hexose monophosphate pathway activity, glutathione synthesis, glucose consumption and lactate formation in erythrocytes has been devised. By this method the results were obtained from the blood samples of six glucose 6-phosphate dehydrogenase deficiencies, one glucose phosphate isomerase deficiency and two pyruvate kinase deficiencies. These results indicate that one abnormal enzyme may exert its influence on the other several enzyme systems and that it is valuable to investigate the metabolism of erythrocytes using this system in different erythroenzymopathies as well as different variants of one enzyme deficiency.     

Vagal System Impairment in Human Immunodeficiency Virus-Positive Patients with Chronic Hepatitis C: Does Hepatic Glutathione Deficiency Have a Pathogenetic Role?

Background: Both an autonomic impairment and a systemic depletion of reduced glutathione (GSH) may be documented in patients with chronic liver diseases and in human immunodeficiency virus (HIV)-positive patients. Methods: The coefficients of electrocardiographic R-R interval variation (CVc) were assessed in 125 patients with chronic hepatitis C (CHC) (65 HIV-positive and 60 HIV-negative) and in 61 healthy controls. The CVc values were correlated with hepatic (H-GSH), plasmatic (P-GSH), lymphocyte (L-GSH), and erythrocyte (E-GSH) concentrations of GSH and with erythrocyte malonyldialdehyde (MDA) levels. Results: Compared with healthy controls, in CHC patients the concentrations of H-GSH, P-GSH, L-GSH, and E-GSH were reduced, whereas MDA levels were increased with a statistically significant difference (P < 0.001). CVc was significantly reduced in patients with CHC (especially in those who were HIV-positive) and correlated significantly with the values of H-GSH, P-GSH, L-GSH, E-GSH, and MDA (P < 0.001). Conclusions: A dysfunction of the cardiac vagal system may be detected in patients with CHC (especially in those who are HIV-positive); this abnormality may be related to a reduced response to oxidative stress because of a systemic depletion of GSH.     

High-Density Lipoprotein and Atherosclerosis: The Role of Antioxidant Activity

Levels of high-density lipoprotein (HDL) cholesterol are generally inversely associated with the risk for the development of atherosclerosis. The mechanism by which HDL imparts protection from the initiation and progression of occlusive vascular disease is complex and multifactorial. The major anti-atherosclerotic effect of HDL is felt to be reverse cholesterol transport. HDL has been demonstrated to scavenge cholesterol from the peripheral vasculature with transport to the liver, where is it excreted in the biliary system. However, HDL exhibits multiple other physiologic effects that may play a role in the reduced risk for atherosclerosis. HDL has been demonstrated to exhibit beneficial effects on platelet function, endothelial function, coagulation parameters, inflammation, and interactions with triglyceride-rich lipoproteins. Increasing amounts of clinical and experimental data have shown that HDL cholesterol has significant antioxidant effect that may significantly contribute to protection from atherosclerosis.     

Antioxidant Action of Carvedilol: A Potential Role in Treatment of Heart Failure

Chronic heart failure (CHF) is a progressive disease of diverse origins, multiple mechanisms and complex therapeutic strategies. Contemporary drugs aimed to alleviate the progression of CHF attempt to improve volume/salt loads (edema), correct circulatory aberrations (vascular constriction, low cardiac output) and protect the heart from direct toxic effects of neurohormonal systems such as the renin-angiotensin-aldosterone and the sympathetic nervous system. Most recently (1997) carvedilol (Coreg), a vasodilating beta-adrenergic blocking agent was approved for treatment of heart failure thereby introducing the first beta-blocking agent to heart failure medicine. Carvedilol is a multiple action, non-specific - and ₁-adrenergic receptor blocker and a potent antioxidant agent. Carvedilol has unique antioxidant properties confirmed in physico-chemical, biochemical, cellular and in vivo animal studies. In this paper, the role of oxidative stress in heart failure is reviewed in reference to the antioxidant actions of carvedilol and its possible contribution to the overall efficacy of the drug in reducing morbidity and mortality in heart failure patients.     

Role of Nitric Oxide in Anorectal Function of Normal and Neuronal Nitric Oxide Synthase Knockout Mice

PURPOSE: In vitro data suggest that nitric oxide is an important inhibitory neurotransmitter in the internal anal sphincter, and morphologic evidence implies that it mediates the rectoanal inhibitory reflex. This study examined the anatomy, physiology, and pharmacology of the internal sphincter in control and neuronal nitric oxide synthase knockout mice. METHODS: Neuronal nitric oxide synthase, nicotinamide adenosine triphosphate dinucleotide phosphate diaphorase histochemistry, and PGP 9.5 immunohistochemistry were compared between knockout and sibling control mice. Anorectal manometry was performed with a balloon-tipped water-perfused catheter. In vitro studies were performed on both whole internal anal sphincter rings and strips. RESULTS: Staining of the myenteric plexus and nerves traversing the internal anal sphincter in sibling control mice demonstrated the presence of neuronal nitric oxide synthase and nicotinamide adenine dinucleotide phosphate diaphorase at these sites. These markers were absent in knockout mice. Maximum anal resting pressure was similar in control and knockout mice (15.6 +/- 2.6 cm H(2)O (n = 4) vs. 14.0 +/- 2.3 cm H(2)O (n = 7)). The rectoanal inhibitory reflex was present in all control mice (n = 4) but in only four of seven knockout mice. Field stimulation with parameters designed to activate inhibitory nerves produced relaxation of internal sphincter tissue from both control and knockout mice, which was partially attenuated in control mice only by pretreatment with the nitric oxide synthase inhibitor N omega-nitro-L-arginine. Further inhibition of nerve-induced relaxation in control mice was achieved with antagonists of vasoactive intestinal peptide, adenosine triphosphate, and heme oxygenase. CONCLUSIONS: Although in the normal mouse, nitric oxide is an inhibitory neurotransmitter in the internal sphincter, other transmitters may play a role in the rectoanal inhibitory reflex. These other inhibitory neurotransmitters can apparently compensate for the absence of nitric oxide synthase in knockout mice to maintain approximately normal function.     

中枢胆碱能神经系统功能的研究进展

中枢胆碱能神经系统是中枢胆碱能神经元及其纤维投射分布的部位 ,与学习、记忆等高级神经活动密切相关。围产期缺氧缺血脑损伤及大脑的退行性改变常引起中枢胆碱能神经系统受损并导致学习和记忆等高级神经功能障碍 ,成为老年痴呆等疾病的重要发病机制之一。中枢胆碱能神经系统与神经营养因子和一些胆碱酯酶抑制剂之间有着密切的关系 ,具有许多新的功能。     

Monoester lipase activity in the red cells of patients with various blood disordrs

S ummary . Membrane-bound monoester lipase (MEL) activity was measured by a radiochemical assay in intact red blood cells (RBC) from 77 patients with various blood disorders. MEL activity levels in their RBC were compared with levels in normal subjects. The increase in MEL activity was greatest in RBC of regenerative anaemias (N=8) and blastic crisis of chronic granulocytic leukaemias (7) whereas a lesser, although significant, increase was found in acute non-lymphoblastic leukaemias (14). MEL activity was markedly decreased in RBC of erythroleukaemias (N=5) and decreased to a lesser extent in acute lymphoblastic leukaemias (10) and in chronic myeloproliferative disorders other than chronic granulocytic leukaemias (8). MEL activity levels were roughly correlated with reticulocyte counts, but the relationship was not of predictive value in individual cases. Further exploration of the possible diagnostic or prognostic implications of these variations in enzyme levels seems warranted.
A monoester lipase (MEL) activity has been recently identified and characterized as a membrane-bound enzyme in red blood cell (RBC) from rats (Arnaud et al , 1979) and human subjects (Boyer et al , 1981). No attempt has been made so far to search for abnormalities of this enzyme. We have therefore measured MEL activity in RBC of 77 patients with various blood disorders.     

Role of Microglia in Ethanol’s Apoptotic Action on Hypothalamic Neuronal Cells in Primary Cultures

Background: Microglia are the major inflammatory cells in the central nervous system and play a role in brain injuries as well as brain diseases. In this study, we determined the role of microglia in ethanol’s apoptotic action on neuronal cells obtained from the mediobasal hypothalamus and maintained in primary cultures. We also tested the effect of cAMP, a signaling molecule critically involved in hypothalamic neuronal survival, on microglia‐mediated ethanol’s neurotoxic action. Methods: Ethanol’s neurotoxic action was determined on enriched fetal mediobasal hypothalamic neuronal cells with or without microglia cells or ethanol‐activated microglia‐conditioned media. Ethanol’s apoptotic action was determined using nucleosome assay. Microglia activation was determined using OX6 histochemistry and by measuring inflammatory cytokines secretion from microglia in cultures using enzyme‐linked immunosorbent assay (ELISA). An immunoneutralization study was conducted to identify the role of a cytokine involved in ethanol’s apoptotic action. Results: We show here that ethanol at a dose range of 50 and 100 mM induces neuronal death by an apoptotic process. Ethanol’s ability to induce an apoptotic death of neurons is increased by the presence of ethanol‐activated microglia‐conditioned media. In the presence of ethanol, microglia showed elevated secretion of various inflammatory cytokines, of which TNF‐α shows significant apoptotic action on mediobasal hypothalamic neuronal cells. Ethanol’s neurotoxic action was completely prevented by cAMP. The cell‐signaling molecule also prevented ethanol‐activated microglial production of TNF‐α. Immunoneutralization of TNF‐α prevented the microglia‐derived media’s ability to induce neuronal death. Conclusions: These results suggest that ethanol’s apoptotic action on hypothalamic neuronal cells might be mediated via microglia, possibly via increased production of TNF‐α. Furthermore, cAMP reduces TNF‐α production from microglia to prevent ethanol’s neurotoxic action.     

Reduced Glutathione, Glutathione Reductase, Glutathione Peroxidase, and Pyruvate Kinase in Erythrocytes of Human Newborns and Adults in Malaysia

S ummary . The mean level of glutathione, and activities of glutathione reductase, glutathione peroxidase and pyruvate kinase in human newborns showed highly significant differences from those in adults. Except for glutathione peroxidase, the mean activity of which was significantly lower in neonates, the values were significantly higher in neonates than in adults. Among neonates, Chinese had a higher mean glutathione reductase activity than Malays or Indians, but lower than that of Caucasians; these racial differences are probably due to a different dietary intake of riboflavin. Such racial differences were not seen in the activity of glutathione peroxidase and pyruvate kinase, but the mean level of reduced glutathione was significantly higher in Chinese than in Indian neonates although not than in Malayan neonates. The levels of glutathione, glutathione peroxidase and pyruvate kinase were significantly higher in female than in male newborns.     

LRRK2: An Emerging New Molecule in the Enteric Neuronal System That Quantitatively Regulates Neuronal Peptides and IgA in the Gut

Background

Leucine-rich repeat kinase 2 (LRRK2) is a recently discovered molecule associated with familial and sporadic Parkinson’s disease. It regulates many central neuronal functions such as cell proliferation, apoptosis, autophagy, and axonal extension. However, in contrast to the well-documented function of LRRK2 in central neurons, it is unclear whether LRRK2 is expressed in enteric neurons and affects the physiology of the gut.

Aims

By examining LRRK2-KO mice, this study investigated whether enteric neurons express LRRK2 and whether intestinal neuronal peptides and IgA are quantitatively changed.

Methods

Intestinal protein lysates and sections prepared from male C57BL/6 J mice were analyzed by Western blotting and immunostaining using anti-LRRK2 antibody, respectively. Intestinal neuronal peptide-mRNAs were quantified by real-time PCR in wild-type mice and LRRK2-KO mice. Intestinal IgA was quantified by ELISA. Lamina propria mononuclear cells (LPMCs) were analyzed by flow cytometry to evaluate the ratio of B1 to B2 B cells.

Results

Western analysis and immunostaining revealed that LRRK2 is expressed in enteric neurons. The amounts of mRNA for vasoactive intestinal peptide, neuropeptide Y, and substance P were increased in LRRK2-KO mice accompanied by an increment of IgA. However, the intestinal B cell subpopulations were not altered in LRRK2-KO mice.

Conclusions

For the first time, we have revealed that LRRK2 is expressed in enteric neurons and related to quantitative alterations of neuronal peptide and IgA. Our study highlights the importance of LRRK2 in enteric neurons as well as central neurons.