RESUMEN
Melatonin regulates vital physiological processes in animals, such as the circadian cycle, sleep, locomotion, body temperature, food intake, and sexual and immune responses. In plants, melatonin modulates seed germination, longevity, circadian cycle, photoperiodicity, flowering, leaf senescence, postharvest fruit storage, and resistance against biotic and abiotic stresses. In plants, the effect of melatonin is mediated by various regulatory elements of the redox network, including RNS and ROS. Similarly, the radical gas NO mediates various physiological processes, like seed germination, flowering, leaf senescence, and stress responses. The biosynthesis of both melatonin and NO takes place in mitochondria and chloroplasts. Hence, both melatonin and nitric oxide are key signaling molecules governing their biological pathways independently. However, there are instances when these pathways cross each other and the two molecules interact with each other, resulting in the formation of N-nitrosomelatonin or NOMela, which is a nitrosated form of melatonin, discovered recently and with promising roles in plant development. The interaction between NO and melatonin is highly complex, and, although a handful of studies reporting these interactions have been published, the exact molecular mechanisms governing them and the prospects of NOMela as a NO donor have just started to be unraveled. Here, we review NO and melatonin production as well as RNS-melatonin interaction under normal and stressful conditions. Furthermore, for the first time, we provide highly sensitive, ozone-chemiluminescence-based comparative measurements of the nitric oxide content, as well as NO-release kinetics between NOMela and the commonly used NO donors CySNO and GSNO.
Asunto(s)
Melatonina , Óxido Nítrico , Plantas , Melatonina/metabolismo , Óxido Nítrico/metabolismo , Plantas/metabolismo , Donantes de Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/farmacología , Transducción de Señal , Fenómenos Fisiológicos de las PlantasRESUMEN
Mitochondria play a crucial role in maintaining Ca2+ homeostasis in cells. Due to the critical regulatory role of the products of oxidative and non-oxidative metabolism of L-arginine, it is essential to clarify their effect on Ca2+ transport in smooth muscle mitochondria. Experiments were performed on the uterine myocytes of rats and isolated mitochondria. The possibility of NO synthesis by mitochondria was demonstrated by confocal microscopy and spectrofluorimetry methods using the NO-sensitive fluorescent probe DAF-FM and Mitotracker Orange CM-H2TMRos. It was shown that 50 µM L-arginine stimulates the energy-dependent accumulation of Ca2+ in mitochondria using the fluorescent probe Fluo-4 AM. A similar effect occurred when using nitric oxide donors 100 µM SNP, SNAP, and sodium nitrite (SN) directly. The stimulating effect was eliminated in the presence of the NO scavenger C-PTIO. Nitric oxide reduces the electrical potential in mitochondria without causing them to swell. The stimulatory effect of spermine on the accumulation of Ca2+ by mitochondria is attributed to the enhancement of NO synthesis, which was demonstrated with the use of C-PTIO, NO-synthase inhibitors (100 µM NA and L-NAME), as well as by direct monitoring of NO synthesis fluorescent probe DAF-FM. A conclusion was drawn about the potential regulatory effect of the product of the oxidative metabolism of L-arginine - NO on the transport of Ca2+ in the mitochondria of the myometrium, as well as the corresponding effect of the product of non-oxidative metabolism -spermine by increasing the synthesis of NO in these subcellular structures.
Asunto(s)
Arginina , Calcio , Óxido Nítrico , Femenino , Animales , Arginina/metabolismo , Calcio/metabolismo , Ratas , Óxido Nítrico/metabolismo , Oxidación-Reducción , Miometrio/metabolismo , Miometrio/efectos de los fármacos , Mitocondrias Musculares/metabolismo , Mitocondrias Musculares/efectos de los fármacos , Ratas Wistar , Mitocondrias/metabolismo , Mitocondrias/efectos de los fármacos , Útero/metabolismo , Útero/efectos de los fármacos , Espermina/metabolismo , Espermina/farmacología , Donantes de Óxido Nítrico/farmacología , Donantes de Óxido Nítrico/metabolismo , Músculo Liso/metabolismo , Músculo Liso/efectos de los fármacos , Transporte Biológico/efectos de los fármacosRESUMEN
When plants are exposed to water stress, photosynthesis is downregulated due to enhanced reactive oxygen species (ROS) and nitric oxide (NO). In contrast, photorespiratory metabolism protected photosynthesis and sustained yield. Modulation of photorespiration by ROS was established, but the effect of NO on photorespiratory metabolism was unclear. We, therefore, examined the impact of externally added NO by using S-nitrosoglutathione (GSNO), a natural NO donor, in leaf discs of pea (Pisum sativum) under dark or light: moderate or high light (HL). Maximum NO accumulation with GSNO was under high light. The presence of 2-4-carboxyphenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO), a NO scavenger, prevented the increase in NO, confirming the release of NO in leaves. The increase in S-nitrosothiols and tyrosine-nitrated proteins on exposure to GSNO confirmed the nitrosative stress in leaves. However, the changes by GSNO in the activities and transcripts of five photorespiratory enzymes: glycolate oxidase, hydroxypyruvate reductase, catalase, glycerate kinase, and phosphoglycolate phosphatase activities were marginal. The changes in photorespiratory enzymes caused by GSNO were much less than those with HL. Since GSNO caused only mild oxidative stress, we felt that the key modulator of photorespiration might be ROS, but not NO.
Asunto(s)
Pisum sativum , S-Nitrosoglutatión , Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/metabolismo , Hojas de la Planta/metabolismo , Especies Reactivas de Oxígeno/metabolismo , S-Nitrosoglutatión/farmacología , S-Nitrosoglutatión/metabolismoRESUMEN
The MT-TL2 m.12315G>A pathogenic variant has previously been reported in five individuals with mild clinical phenotypes. Herein we report the case of a 5-year-old child with heteroplasmy for this variant who developed neurological regression and stroke-like episodes similar to those observed in mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes (MELAS). Biochemical evaluation revealed depletion of arginine on plasma amino acid analysis and low z-scores for citrulline on untargeted plasma metabolomics analysis. These findings suggested that decreased availability of nitric oxide may have contributed to the stroke-like episodes. The use of intravenous arginine during stroke-like episodes and daily enteral L-citrulline supplementation normalized her biochemical values of arginine and citrulline. Untargeted plasma metabolomics showed the absence of nicotinamide and 1-methylnicotinamide, and plasma total glutathione levels were low; thus, nicotinamide riboside and N-acetylcysteine therapies were initiated. This report expands the phenotype associated with the rare mitochondrial variant MT-TL2 m.12315G>A to include neurological regression and a MELAS-like phenotype. Individuals with this variant should undergo in-depth biochemical analysis to include untargeted plasma metabolomics, plasma amino acids, and glutathione levels to help guide a targeted approach to treatment.
Asunto(s)
Acidosis Láctica , Síndrome MELAS , Encefalomiopatías Mitocondriales , Accidente Cerebrovascular , Preescolar , Femenino , Humanos , Arginina/genética , Citrulina , Glutatión/metabolismo , Síndrome MELAS/diagnóstico , Síndrome MELAS/genética , Síndrome MELAS/complicaciones , Donantes de Óxido Nítrico/metabolismo , Accidente Cerebrovascular/complicaciones , Accidente Cerebrovascular/tratamiento farmacológicoRESUMEN
Physiologically, smooth muscle cells (SMC) and nitric oxide (NO) produced by endothelial cells strictly cooperate to maintain vasal homeostasis. In atherosclerosis, where this equilibrium is altered, molecules providing exogenous NO and able to inhibit SMC proliferation may represent valuable antiatherosclerotic agents. Searching for dual antiproliferative and NO-donor molecules, we found that furoxans significantly decreased SMC proliferation in vitro, albeit with different potencies. We therefore assessed whether this property is dependent on their thiol-induced ring opening. Indeed, while furazans (analogues unable to release NO) are not effective, furoxans' inhibitory potency parallels with the electron-attractor capacity of the group in 3 of the ring, making this effect tunable. To demonstrate whether their specific block on G1-S phase could be NO-dependent, we supplemented SMCs with furoxans and inhibitors of GMP- and/or of the polyamine pathway, which regulate NO-induced SMC proliferation, but they failed in preventing the antiproliferative effect. To find the real mechanism of this property, our proteomics studies revealed that eleven cellular proteins (with SUMO1 being central) and networks involved in cell homeostasis/proliferation are modulated by furoxans, probably by interaction with adducts generated after degradation. Altogether, thanks to their dual effect and pharmacological flexibility, furoxans may be evaluated in the future as antiatherosclerotic molecules.
Asunto(s)
Donantes de Óxido Nítrico , Óxido Nítrico , Donantes de Óxido Nítrico/farmacología , Donantes de Óxido Nítrico/metabolismo , Óxido Nítrico/metabolismo , Células Endoteliales/metabolismo , Músculo Liso Vascular , Proteómica , Proliferación Celular , Células Cultivadas , Miocitos del Músculo LisoRESUMEN
Background: Desipramine a representative of tricyclic antidepressants (TCAs) promotes recovery of depressed patients by inhibition of reuptake of neurotransmitters serotonin (SER) and norepinephrine (NE) in the presynaptic membrane by directly blocking their respective transporters SERT and NET.Aims: To study the effect of desipramine on programmed erythrocyte death (eryptosis) and explore the underlying mechanisms.Methods: Phosphatidylserine (PS) exposure on the cell surface as marker of cell death was estimated from annexin-V-binding, cell volume from forward scatter in flow cytometry. Hemolysis was determined photometrically, and intracellular glutathione [GSH]i from high performance liquid chromatography.Results: Desipramine dose-dependently significantly enhanced the percentage of annexin-V-binding cells and didn´t impact glutathione (GSH) synthesis. Desipramine-induced eryptosis was significantly reversed by pre-treatment of erythrocytes with either nitric oxide (NO) donor sodium nitroprusside (SNP) or N-acetyl-L-cysteine (NAC). The highest inhibitory effect was obtained by using both inhibitors together. Calcium (Ca2+) depletion aggravated desipramine-induced eryptosis. Changing the order of treatment, i.e. desipramine first followed by inhibitors, could not influence the inhibitory effect of SNP or NAC.Conclusion: Antidepressants-caused intoxication can be treated by SNP and NAC, respectively. B) Patients with chronic hypocalcemia should not be treated with tricyclic anti-depressants or their dose should be noticeably reduced.
Asunto(s)
Eriptosis , Donantes de Óxido Nítrico , Humanos , Donantes de Óxido Nítrico/farmacología , Donantes de Óxido Nítrico/metabolismo , Nitroprusiato/farmacología , Nitroprusiato/metabolismo , Calcio/metabolismo , Acetilcisteína/farmacología , Desipramina/farmacología , Desipramina/metabolismo , Eritrocitos/metabolismo , Glutatión/metabolismo , Glutatión/farmacología , Anexinas/metabolismo , Anexinas/farmacología , Fosfatidilserinas/metabolismo , Tamaño de la Célula , Ceramidas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Estrés OxidativoRESUMEN
As a toxic anthropogenic pollutant, lead (Pb) can be harmful to both plants and animals. Here, the effects of the application of nitric oxide (NO) donor, sodium nitroprusside (SNP, 0, 50, and 100 µM), on the morphological, biochemical, and molecular responses of rice plants under Pb (0, 150, and 300 µM) toxicity in hydroponic conditions were investigated. Pb stress decreased biomass, photosynthetic pigments, Fv/Fm value, and nitrogen (N) and increased the accumulation of hydrogen peroxide (H2O2), methylglyoxal (MG), malondialdehyde (MDA), and electrolyte leakage (EL) in rice seedlings. However, by improving the metabolism of chlorophyll and proline, SNP increased the content of chlorophyll and proline, restored the performance of the photosynthetic apparatus, and stimulated the growth of Pb-stressed rice seedlings. SNP by reducing the expression of HMA2 and increasing the expression of HMA3 and HMA4 caused the immobilization of Pb in the roots and reduced its transfer to the leaves. Adding SNP increased the activity of antioxidant enzymes and glyoxalase cycle and decreased H2O2, MG, MDA, and EL in the leaves of Pb-stressed rice seedlings. By upregulating the expression of genes GSH1, PCS, and ABCC1, SNP increased the accumulation of GSH and PCs in the roots and leaves and increased the plant's tolerance to Pb stress. By modulating the activity of enzymes involved in N metabolism, SNP increased the concentration of N and nitrate and decreased the concentration of ammonium in the leaves of Pb-stressed seedlings. Our study provides evidence that NO may become a promising tool for increasing the tolerance of rice plants to Pb toxicity.
Asunto(s)
Antioxidantes , Oryza , Antioxidantes/metabolismo , Nitroprusiato/metabolismo , Nitroprusiato/farmacología , Oryza/metabolismo , Plomo/metabolismo , Peróxido de Hidrógeno/metabolismo , Donantes de Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/farmacología , Estrés Oxidativo , Plantones , Clorofila/metabolismo , Prolina/metabolismoRESUMEN
Metabolism of nitric oxide (NO) donors: dinitrosyl iron complexes (DNIC), nitrosothiols (RSNO), and nitroprusside was studied on a chick embryo model. The obtained results give reason to assume that DNIC constituting the main pool of nitroso compounds in the vast majority of tissues are NO donors immediately interacting with the physiological target of NO, and other NO donors can perform this function after their transformation into DNIC. NO is released from DNIC not spontaneously, but under a joint influence of a factor destroying the complex and a target having chemical affinity for NO. A similar mechanism is apparently implicated in NO passage through the cell membrane.
Asunto(s)
Óxido Nítrico , Óxidos de Nitrógeno , Embrión de Pollo , Animales , Óxido Nítrico/metabolismo , Óxidos de Nitrógeno/metabolismo , Donantes de Óxido Nítrico/farmacología , Donantes de Óxido Nítrico/metabolismo , Hierro/químicaRESUMEN
Nitric oxide (NO) is a key vasodilatory signalling molecule and NO releasing molecules (NO donors) are being examined as potential treatments for many pathologies. The photoresponsive NO donor tert-dodecane S-nitrosothiol (tDodSNO) has been designed to be highly resistant to metabolism; in principle photoactivation of tDodSNO should therefore enable the controlled release of NO in situ via light modulation. To investigate the therapeutic utility of tDodSNO, we tested drug efficacy in Sprague Dawley rats to assess systemic and localised hemodynamic responses under photoactivation, and to confirm drug safety. For comparison, drug action was evaluated alongside the existing NO donors sodium nitroprusside (SNP) and S-nitrosoglutathione (GSNO). Across a dosing range (0.1-3.0 mg/kg) tDodSNO exerted markedly reduced systemic hypotensive action compared to these standard NO donors, inducing a slight decrease in mean arterial pressure (maximum 14.2 ± 3.0%) without affecting heart rate. Target limb photoactivation of tDodSNO resulted in a substantial localized vasodilatory response, with increases to mean (26.0 ± 7.3%) and maximum (53.2 ± 10.4%) blood flow and decreases to vascular resistance (27.1 ± 3.9%) that were restricted to light exposed tissue. In comparison GSNO and SNP showed variable peripheral effects and were not responsive to photoactivation. tDodSNO did not induce met-Hb formation in blood, or display any signs of toxicity, and was rapidly cleared from the systemic circulation, with no hemodynamic effects detectable 5 min post administration. These data are the first demonstration that drugs based upon a metabolically stable S-nitrosothiol group can be photoactivated in vivo to release NO, and that such agents cause less systemic side effects than existing NO donors. Our data support the use of S-nitrosothiols to enable the spatiotemporal control of NO for therapeutic applications.
Asunto(s)
Donantes de Óxido Nítrico , S-Nitrosotioles , Animales , Ratas , Donantes de Óxido Nítrico/farmacología , Donantes de Óxido Nítrico/metabolismo , Vasodilatación , Ratas Sprague-Dawley , S-Nitrosotioles/farmacología , S-Nitrosotioles/metabolismo , Nitroprusiato/farmacología , Óxido Nítrico/metabolismoRESUMEN
Platelets are the "guardians" of the blood circulatory system. At sites of vessel injury, they ensure hemostasis and promote immunity and vessel repair. However, their uncontrolled activation is one of the main drivers of thrombosis. To keep circulating platelets in a quiescent state, the endothelium releases platelet antagonists including nitric oxide (NO) that acts by stimulating the intracellular receptor guanylyl cyclase (GC). The latter produces the second messenger cyclic guanosine-3',5'-monophosphate (cGMP) that inhibits platelet activation by stimulating protein kinase G, which phosphorylates hundreds of intracellular targets. Intracellular cGMP pools are tightly regulated by a fine balance between GC and phosphodiesterases (PDEs) that are responsible for the hydrolysis of cyclic nucleotides. Phosphodiesterase type 5 (PDE5) is a cGMP-specific PDE, broadly expressed in most tissues in humans and rodents. In clinical practice, PDE5 inhibitors (PDE5i) are used as first-line therapy for erectile dysfunction, pulmonary artery hypertension, and lower urinary tract symptoms. However, several studies have shown that PDE5i may ameliorate the outcome of various other conditions, like heart failure and stroke. Interestingly, NO donors and cGMP analogs increase the capacity of anti-platelet drugs targeting the purinergic receptor type Y, subtype 12 (P2Y12) receptor to block platelet aggregation, and preclinical studies have shown that PDE5i inhibits platelet functions. This review summarizes the molecular mechanisms underlying the effect of PDE5i on platelet activation and aggregation focusing on the therapeutic potential of PDE5i in platelet disorders, and the outcomes of a combined therapy with PDE5i and NO donors to inhibit platelet activation.
Asunto(s)
Óxido Nítrico , Inhibidores de Fosfodiesterasa 5 , Humanos , Masculino , Plaquetas/metabolismo , GMP Cíclico/metabolismo , Proteínas Quinasas Dependientes de GMP Cíclico/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 5/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 5/farmacología , Guanosina/metabolismo , Guanosina/farmacología , Guanilato Ciclasa/metabolismo , Guanilato Ciclasa/farmacología , Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/metabolismo , Nucleótidos Cíclicos/metabolismo , Nucleótidos Cíclicos/farmacología , Inhibidores de Fosfodiesterasa 5/farmacología , Inhibidores de Fosfodiesterasa 5/uso terapéutico , Inhibidores de Fosfodiesterasa 5/metabolismo , Inhibidores de Agregación Plaquetaria/farmacología , Inhibidores de Agregación Plaquetaria/uso terapéutico , Proteínas Quinasas/metabolismoRESUMEN
Oxidative stress induced by well-known toxins including ferric nitrilotriacetate (Fe-NTA), carbon tetrachloride (CCl4) and thioacetamide (TAA) has been attributed to causing tissue injury in the liver and kidney. In this study, the effect of glyceryl trinitrate (GTN), a donor of nitric oxide and NG-nitroarginine methyl ester (l-NAME), a nitric oxide inhibitor on TAA-induced hepatic oxidative stress, GSH and GSH-dependent enzymes, serum transaminases and tumor promotion markers such as ornithine decarboxylase (ODC) activity and [3H]-thymidine incorporation in rats were examined. The animals were divided into seven groups consisting of six healthy rats per group. The six rats were injected intraperitoneally with TAA to evaluate its toxic effect, improvement in its toxic effect if any, or worsening in its toxic effect if any, when given in combination with GTN or l-NAME. The single necrogenic dose of TAA administration caused a significant change in the levels of both hepatic and serum enzymes such as glutathione S-transferase (GST), glutathione reductase (GR), glutathione peroxidase (GPx), γ-glutamyl transpeptidase (GGT), glucose 6-phosphate dehydrogenase (G6PD), alanine aminotransferase (AST) and aspartate aminotransferase (ALT). In addition, treatment with TAA also augmented malondialdehyde (MDA), ornithine decarboxylase (ODC) activity and [3H]-thymidine incorporation in rats liver. Concomitantly, TAA treatment depleted the levels of GSH. However, most of these changes were alleviated by the treatment of animals with GTN dose-dependently. The protective effect of GTN against TAA was also confirmed histopathologically. The present data confirmed our earlier findings with other oxidants including Fe-NTA and CCl4. The GTN showed no change whatsoever when administered alone, however when it was given along with TAA then it showed protection thereby contributing towards defending the role against oxidants-induced organ toxicity. Overall, GTN may contribute to protection against TAA-induced oxidative stress, toxicity, and proliferative response in the liver, according to our findings.
Asunto(s)
Donantes de Óxido Nítrico , Nitroglicerina , Animales , Glutatión/metabolismo , Hígado , NG-Nitroarginina Metil Éster/farmacología , Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/farmacología , Nitroglicerina/farmacología , Ornitina Descarboxilasa/metabolismo , Oxidantes/farmacología , Estrés Oxidativo , Ratas , Ratas Wistar , Tioacetamida/farmacología , Timidina/metabolismoRESUMEN
Wound healing pathologies are an increasing problem in ageing societies. Chronic, non-healing wounds, which cause high morbidity and severely reduce the quality of life of affected individuals, are frequently observed in aged individuals and people suffering from diseases affected by the Western lifestyle, such as diabetes. Causal treatments that support proper wound healing are still scarce. Here, we performed expression proteomics to study the effects of the small molecule TOP-N53 on primary human skin fibroblasts and keratinocytes. TOP-N53 is a dual-acting nitric oxide donor and phosphodiesterase-5 inhibitor increasing cGMP levels to support proper wound healing. In contrast to keratinocytes, which did not exhibit global proteome alterations, TOP-N53 had profound effects on the proteome of skin fibroblasts. In fibroblasts, TOP-N53 activated the cytoprotective, lysosomal degradation pathway autophagy and induced the expression of the selective autophagy receptor p62/SQSTM1. Thus, activation of autophagy might in part be responsible for beneficial effects of TOP-N53.
Asunto(s)
Donantes de Óxido Nítrico , Inhibidores de Fosfodiesterasa 5 , Anciano , Autofagia , Fibroblastos/metabolismo , Humanos , Queratinocitos/metabolismo , Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/farmacología , Inhibidores de Fosfodiesterasa 5/farmacología , Proteoma/metabolismo , Calidad de Vida , Piel/metabolismoRESUMEN
Nitric oxide (NO) is a powerful vasodilator in different vascular beds and NO-donors are widely used in clinical practice. Early data suggested that NO and NO-donors activate vascular smooth muscle high-conductance, calcium-activated potassium channels (BK channels). There exist two hypotheses explaining the effect of NO and NO-donors on BK channels-one stating that protein kinase G (PKG) mediates the effect of NO, and the other one stating that NO acts directly on the channel. Thus, the degree of contribution of PKG to the NO-induced activation of the BK channel is still not completely clear. This study tested the hypothesis that the sodium nitroprusside (SNP)-induced activation of vascular smooth muscle BK channels is fully mediated by PKG. This hypothesis was investigated using the patch-clamp technique and freshly isolated smooth muscle cells from rat tail artery. In whole-cell experiments, SNP considerably increased the outward current compared with the addition of the bath solution. SNP did not alter the current in the presence of iberiotoxin, the specific blocker of BK channels, during co-application with hydroxocobalamin, an NO-scavenger, and in the presence of Rp-8-Br-PET-cGMPS, the specific PKG-inhibitor. In inside-out patches, the activity of BK channels was increased by SNP, SNAP, and DEA-NO. However, these effects did not differ from the effect of the application of drug-free bath solution. Furthermore, a similar increase in single BK channel activity was induced by Rp-8-Br-PET-cGMPS, Rp-8-Br-PET-cGMPS together with SNP, hydroxocobalamin, and hydroxocobalamin together with SNP or DEA-NO. Finally, the activity of excised BK channels did not change in the absence of any application but was considerably increased by PKG compared with the addition of drug-free bath solution. These results suggest that NO released from NO-donors stimulates the BK current only through activation of PKG.
Asunto(s)
Proteínas Quinasas Dependientes de GMP Cíclico , Canales de Potasio de Gran Conductancia Activados por el Calcio , Animales , Proteínas Quinasas Dependientes de GMP Cíclico/metabolismo , Hidroxocobalamina/metabolismo , Hidroxocobalamina/farmacología , Canales de Potasio de Gran Conductancia Activados por el Calcio/metabolismo , Músculo Liso Vascular/metabolismo , Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/farmacología , Nitroprusiato/farmacología , RatasRESUMEN
BACKGROUND/AIM: Nitric oxide (NO) has antitumor activity against various solid tumor cell types in addition to its vasodilatory effect. In the current study, we investigated the effect and mechanism of the synthetic nitrated form (NO2-NAT) of nateglinide, a hypoglycemic agent, on the induction of cell death in human pancreatic cancer cells. MATERIALS AND METHODS: NO production was evaluated by measuring nitrite (NO2) and nitrate (NO3) (NOx). Apoptotic cell numbers were determined using annexin V. RESULTS: NO2-NAT released nitrate and nitrite ions immediately upon dissolving in aqueous solution. NO2-NAT caused significant extracellular leakage of lactate dehydrogenase (LDH) in the human pancreatic cancer cell lines AsPC1 and BxPC3, and increased annexin-positive cells in a time- and concentration-dependent manner. NO2-NAT also significantly increased the activity of caspases 3 and 7. Exposure to Z-VAD-FMK, a caspase inhibitor, significantly suppressed NO2-NAT-induced cell death. CONCLUSION: These results indicated that NO2-NAT induces apoptosis in human pancreatic cancer cells and may serve as a new NO-based chemotherapeutic agent for the treatment of pancreatic cancer.
Asunto(s)
Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Caspasa 3/metabolismo , Caspasa 7/metabolismo , Nateglinida/farmacología , Donantes de Óxido Nítrico/farmacología , Neoplasias Pancreáticas/tratamiento farmacológico , Antineoplásicos/metabolismo , Línea Celular Tumoral , Activación Enzimática , Humanos , Nateglinida/análogos & derivados , Nateglinida/metabolismo , Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/metabolismo , Neoplasias Pancreáticas/inmunología , Neoplasias Pancreáticas/patología , Transducción de SeñalRESUMEN
Organic nitrates are widely used to restore endogenous nitric oxide (NO) levels reduced by endothelial nitric oxide synthase dysfunction. However, these drugs are associated with undesirable side effects, including tolerance. This study aims to investigate the cardiovascular effects of the new organic nitrate 1,3-diisobutoxypropan-2-yl nitrate (NDIBP). Specifically, we assessed its effects on blood pressure, vascular reactivity, acute toxicity, and the ability to induce tolerance. In vitro and ex vivo techniques showed that NDIBP released NO both in a cell-free system and in isolated mesenteric arteries preparations through a process catalyzed by xanthine oxidoreductase. NDIBP also evoked endothelium-independent vasorelaxation, which was significantly attenuated by 2-phenyl-4,4,5,5,-tetramethylimidazoline-1-oxyl 3-oxide (PTIO, 300 µM), a nitric oxide scavenger; 1-H-[1,2,4] oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ, 10 µM), a soluble guanylyl cyclase inhibitor; tetraethylammonium (TEA, 3 mM), a potassium channel blocker; febuxostat (500 nM), a xanthine oxidase inhibitor; and proadifen (10 µM), an inhibitor of cytochrome P450 enzyme. Furthermore, this organic nitrate did not induce tolerance in isolated vessels and presented low toxicity following acute oral administration. In vivo changes on cardiovascular parameters were assessed using normotensive and renovascular hypertensive rats. NDIBP evoked a reduction of blood pressure that was significantly higher in hypertensive animals. Our results suggest that NDIBP acts as a NO donor, inducing blood pressure reduction without having the undesirable effects of tolerance. Those effects seem to be mediated by activation of NO-sGC-cGMP pathway and positive modulation of K+ channels in vascular smooth muscle.
Asunto(s)
Antihipertensivos/uso terapéutico , Hipertensión/tratamiento farmacológico , Arterias Mesentéricas/efectos de los fármacos , Nitratos/uso terapéutico , Donantes de Óxido Nítrico/uso terapéutico , Vasodilatadores/uso terapéutico , Animales , Antihipertensivos/metabolismo , Sistema Enzimático del Citocromo P-450/metabolismo , Femenino , Hipertensión/metabolismo , Masculino , Nitratos/metabolismo , Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/metabolismo , Canales de Potasio/metabolismo , Ratas Wistar , Transducción de Señal/efectos de los fármacos , Guanilil Ciclasa Soluble/metabolismo , Vasodilatadores/metabolismo , Xantina Deshidrogenasa/metabolismoRESUMEN
In modern chemical and biochemical studies, special attention is paid to molecular systems capable of generating nitric oxide (NO), which is one of the most important signalling molecules in the body and can trigger a whole cascade of reactions. Despite the importance of this molecule, the mechanisms of its formation in living organisms remain a subject of debate. This review combines the most important methods of releasing NO from endogenous and exogenous sources. The history of endogenous NO donors dates back more than 150 years, since the synthesis of nitroglycerin, which remains the standard vasodilator today, even though it is known that it and many other similar compounds lead to the development of a nitrate tolerance. Particular awareness is devoted to the mechanisms of NO formation without the participation of enzymes, since these methods are most important for creating exogenous sources of NO as drugs. The study of NO formation methods is centred on both the creation of new NO donors and understanding the mechanisms of tolerance to them.
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Óxido Nítrico/biosíntesis , Arginina/química , Arginina/metabolismo , Humanos , Modelos Químicos , Donantes de Óxido Nítrico/química , Donantes de Óxido Nítrico/metabolismo , S-Nitrosotioles/química , S-Nitrosotioles/metabolismoRESUMEN
In a relatively isolated system of avian embryo, the metabolism of NO, a component of the dinitrosyl iron complexes (DNIC), the main NO donor in most tissues, depends on the ligands that make up the complex. This fact corroborates the earlier hypothesis that these ligands perform a regulatory function in NO metabolism. It is also shown that nitrite injected into the embryo is not oxidized to nitrate like NO in DNIC, but is accumulated outside the amniotic sac. Normally, nitrite is present in an embryo in trace amounts. These facts suggest that NO in the embryo is transferred from the donor molecule to a target in the embryo tissues further transformed with minimum oxidation to nitrite.
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Quelantes del Hierro/farmacología , Hierro/metabolismo , Hierro/farmacología , Óxidos de Nitrógeno/metabolismo , Óxidos de Nitrógeno/farmacología , Animales , Catalasa/antagonistas & inhibidores , Catalasa/efectos de los fármacos , Catalasa/metabolismo , Embrión de Pollo , Ácido Cítrico/farmacología , Desarrollo Embrionario/efectos de los fármacos , Glutatión , Hemoglobinas/química , Hemoglobinas/metabolismo , Hemoglobinas/farmacología , Hierro/química , Hierro/fisiología , Quelantes del Hierro/metabolismo , Ligandos , Nitratos/metabolismo , Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/química , Donantes de Óxido Nítrico/metabolismo , Nitritos/metabolismo , Óxidos de Nitrógeno/química , Oxidación-Reducción/efectos de los fármacos , Fenantrolinas/farmacologíaRESUMEN
Nitric oxide (NO) plays a prominent physiological role as a low-molecular-mass signal molecule involved in diverse biological functions. Great attention has been directed to pharmacologically modulating the release of NO for various therapeutic applications. We have focused on O2-(2,4-dinitrophenyl) 1-[(4-ethoxycarbonyl)piperazin-1-yl]diazen-1-ium-1,2-diolate (JS-K) as an example of diazeniumdiolate prodrugs with potential for cancer chemotherapy. JS-K is reportedly activated by glutathione conjugation by glutathione transferase (GST), but the scope of activities among the numerous members of the GSTome is unknown. We demonstrate that all human GSTs tested except GST T1-1 are active with JS-K as a substrate, but their specific activities are notably spanning a > 100-fold range. The most effective enzyme was the mu class member GST M2-2 with a specific activity of 273 ± 5 µmol min-1 mg-1 and the kinetic parameters Km 63 µM, kcat 353 s-1, kcat/Km 6 × 106 M-1 s-1. The abundance of the GSTs as an ensemble and their high catalytic efficiency indicate that release of NO occurs rapidly in normal tissues such that this influence must be considered in clarification of the tumor-killing effect of JS-K.
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Compuestos Azo/metabolismo , Glutatión Transferasa/metabolismo , Donantes de Óxido Nítrico/metabolismo , Piperazinas/metabolismo , Profármacos/metabolismo , Biotransformación , Humanos , Cinética , Óxido Nítrico/metabolismoRESUMEN
In the vertebrate cochlea, the reticular lamina seals the organ of Corti against the endolymph filled scala media. After noise exposure, fast alterations in the endothelial nitric oxide synthase (eNOS) expression level were identified in this cochlear structure. Minor amounts of nitric oxide (NO) produced by eNOS or applied by NO donors such as S-nitroso-N-acetyl-penicillamine (SNAP) might protect this vulnerable part of the organ of Corti, on the line of gap junctions of supporting cells and cochlear microcirculation. In n=5 anesthetized guinea pigs, SNAP was intravenously applied in two concentrations. Six untreated animals served as controls. The cochleae were removed and prepared for immunoelectron microscopy using specific gold-labeled anti-eNOS antibodies. The density of the gold particles was quantified for seven cellular regions in the reticular lamina at the ultrastructural level. Following SNAP application, a significant increase in eNOS expression (+176%) was detected compared with controls (p=0.012). The increase occurred mainly in actin-rich cuticular structures and the prominent microtubules bundles. Correlation analysis revealed three clear and five moderate cellular associations for controls, whereas only one clear and one moderate after SNAP application. Thus, application of the NO donor SNAP resulted in an increase in eNOS expression in distinct regions of the reticular lamina.
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Donantes de Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa de Tipo III/metabolismo , Órgano Espiral/metabolismo , Lámina Espiral/metabolismo , Animales , Cobayas , MasculinoRESUMEN
The mechanism behind the cytoprotective potential of cerium oxide nanoparticles (CeO2 NPs) against cytotoxic nitric oxide (NO) donors and H2O2 is still not clear. Synthesized and characterized CeO2 NPs significantly ameliorated the lipopolysaccharide (LPS)-induced cytokines IL-1ß and TNF-α. The main goal of this study was to determine the capacities of NPs regarding signaling effects that could have occurred due to reactive oxygen species (ROS) and/or NO, since NP-induced ROS/NO did not lead to toxicity in HUVE cells. Concentrations that induced 50% cell death (i.e., IC50s) of two NO donors (DETA-NO; 1250 ± 110 µM and sodium nitroprusside (SNP); 950 ± 89 µM) along with the IC50 of H2O2 (120 ± 7 µM) were utilized to evaluate cytoprotective potential and its underlying mechanism. We determined total ROS (as a collective marker of hydrogen peroxide, superoxide radical (O2â¢-), hydroxyl radical, etc.) by DCFH-DA and used a O2â¢- specific probe DHE to decipher prominent ROS. The findings revealed that signaling effects mediated mainly by O2â¢- and/or NO are responsible for the amelioration of toxicity by CeO2 NPs at 100 µg/mL. The unaltered effect on mitochondrial membrane potential (MMP) due to NP exposure and, again, CeO2 NPs-mediated recovery in the loss of MMP due to exogenous NO donors and H2O2 suggested that NP-mediated O2â¢- production might be extra-mitochondrial. Data on activated glutathione reductase (GR) and unaffected glutathione peroxidase (GPx) activities partially explain the mechanism behind the NP-induced gain in GSH and persistent cytoplasmic ROS. The promoted antioxidant capacity due to non-cytotoxic ROS and/or NO production, rather than inhibition, by CeO2 NP treatment may allow cells to develop the capacity to tolerate exogenously induced toxicity.