Nitric Oxide-dependent Mechanism Research Articles

Plant quiescence strategy and seed dormancy under hypoxia.

Plant quiescence and seed dormancy can be triggered by reduced oxygen availability. Under water, oxygen depletion caused by flooding can culminate in a quiescent state, which is a plant strategy for energy preservation and survival. In adult plants, a quiescent state can be activated by sugar starvation, leading to metabolic depression. In seeds, secondary dormancy can be activated by reduced oxygen availability, which creates an unfavourable state for germination. The physical dormancy of some seeds and buds includes barriers to external conditions, which indirectly results in hypoxia. The molecular processes that support seed dormancy and plant survival through quiescence under hypoxia include the N-degron pathway, which enables the modulation of ethylene-responsive factors of group VII and downstream targets. This oxygen- and nitric oxide-dependent mechanism interacts with phytohormone-related pathways to control growth.

In vivo mitochondria-targeted protection against uterine artery vascular dysfunction and remodelling in rodent hypoxic pregnancy.

Gestational hypoxia adversely affects uterine artery function, increasing complications. However, an effective therapy remains unidentified. Here, we show in rodent uterine arteries that hypoxic pregnancy promotes hypertrophic remodelling, increases constrictor reactivity via protein kinase C signalling, and triggers compensatory dilatation via nitric oxide-dependent mechanisms and stimulation of large conductance Ca2+ -activated K+ -channels. Maternal in vivo oral treatment with the mitochondria-targeted antioxidant MitoQ in hypoxic pregnancy normalises uterine artery reactivity and prevents vascular remodelling. From days 6-20 of gestation (term ∼22 days), female Wistar rats were randomly assigned to normoxic or hypoxic (13-14% O2 ) pregnancy±daily maternal MitoQ treatment (500 µm in drinking water). At 20 days of gestation, maternal, placental and fetal tissue was frozen to determine MitoQ uptake. The uterine arteries were harvested and, in one segment, constrictor and dilator reactivity was determined by wire myography. Another segment was fixed for unbiased stereological analysis of vessel morphology. Maternal administration of MitoQ in both normoxic and hypoxic pregnancy crossed the placenta and was present in all tissues analysed. Hypoxia increased uterine artery constrictor responses to norepinephrine, angiotensin II and the protein kinase C activator, phorbol 12,13-dibutyrate. Hypoxia enhanced dilator reactivity to sodium nitroprusside, the large conductance Ca2+ -activated K+ -channel activator NS1619 and ACh via increased nitric oxide-dependent mechanisms. Uterine arteries from hypoxic pregnancy showed increased wall thickness and MitoQ treatment in hypoxic pregnancy prevented all effects on uterine artery reactivity and remodelling. The data support mitochondria-targeted therapy against adverse changes in uterine artery structure and function in high-risk pregnancy. KEY POINTS: Dysfunction and remodelling of the uterine artery are strongly implicated in many pregnancy complications, including advanced maternal age, maternal hypertension of pregnancy, maternal obesity, gestational diabetes and pregnancy at high altitude. Such complications not only have immediate adverse effects on the growth of the fetus, but also they can also increase the risk of cardiovascular disease in the mother and offspring. Despite this, there is a significant unmet clinical need for therapeutics that treat uterine artery vascular dysfunction in adverse pregnancy. Here, we show in a rodent model of gestational hypoxia that in vivo oral treatment of the mitochondria-targeted antioxidant MitoQ protects against uterine artery vascular dysfunction and remodelling, supporting the use of mitochondria-targeted therapy against adverse changes in uterine artery structure and function in high-risk pregnancy.

Zinc as a potential regulator of the BCR-ABL oncogene in chronic myelocytic leukemia cells

BackgroundGenerally, decreased zinc in the serum of tumor patients but increased zinc in tumor cells can be observed. However, the role of zinc homeostasis in myeloid leukemia remains elusive. BCR-ABL is essential for the initiation, maintenance, and progression of chronic myelocytic leukemia (CML). We are currently investigating the association between zinc homeostasis and CML. MethodsGenes involved in zinc homeostasis were examined using three GEO datasets. Western blotting and qPCR were used to investigate the effects of zinc depletion on BCR-ABL expression. Furthermore, the effect of TPEN on BCR-ABL promoter activity was determined using the dual-luciferase reporter assay. MRNA stability and protein stability of BCR-ABL were assessed using actinomycin D and cycloheximide. ResultsTranscriptome data mining revealed that zinc homeostasis-related genes were associated with CML progression and drug resistance. Several zinc homeostasis genes were affected by TPEN. Additionally, we found that zinc depletion by TPEN decreased BCR-ABL mRNA stability and transcriptional activity in K562 CML cells. Zinc supplementation and sodium nitroprusside treatment reversed BCR-ABL downregulation by TPEN, suggesting zinc- and nitric oxide-dependent mechanisms. ConclusionOur in vitro findings may help to understand the role of zinc homeostasis in BCR-ABL regulation and thus highlight the importance of zinc homeostasis in CML.

Methamphetamine-induced vaginal lubrication in rats.

Based on previous studies of vaginal lubrication as well as our own previously reported interview study of women who self-reported methamphetamine (meth)-induced vaginal lubrication, in the current study we sought to determine the potential dose-response relationship leading to meth-induced vaginal lubrication. We also developed an animal model to study the reported effects and examine potential mechanisms mediating this phenomenon. We sought to characterize the effects of meth on vaginal lubrication in an animal model with the aim of providing a potential framework for new mechanisms that incorporate novel therapeutic agents for the treatment of vaginal dryness. Vaginal lubrication was measured via insertion of a preweighed, cotton-tipped swab into the vaginal canal of anesthetized rats following treatment with various doses of intravenous (IV) meth, up to 0.96mg/kg, and after additional pharmacological manipulations, including administration of a nitric oxide synthase inhibitor and an estrogen receptor antagonist. Plasma signaling molecules, including estradiol, progesterone, testosterone, nitric oxide, and vasoactive intestinal polypeptide, were measured immediately before and at 9 time points after IV meth administration. Blood was collected via a previously implanted chronic indwelling jugular catheter and analyzed by use of commercially available kits per the manufacturer's instructions. Outcomes for this study include the measurement of vaginal lubrication in anesthetized rats following various pharmacological manipulations and plasma levels of various signaling molecules. Meth dose-dependently increased vaginal lubrication in anesthetized female rats. Meth significantly increased plasma levels compared to baseline of estradiol (2 and 15 minutes after meth infusion) as well as progesterone, testosterone, and nitric oxide (10 minutes after meth infusion). Also, vasoactive intestinal polypeptide decreased significantly compared to baseline for 45minutes following meth infusion. Our data further suggest that nitric oxide, but not estradiol, is critical in the production of vaginal secretions in response to meth. This study has far-reaching implications for women who are suffering from vaginal dryness and for whom estrogen therapy is unsuccessful, as the investigation has demonstrated that meth presents a novel mechanism for producing vaginal lubrication that can be targeted pharmacologically. This study is, to our knowledge, the first performed to measure the physiological sexual effects of meth in an animal model. Animals were anesthetized when they were administered meth. In an ideal situation, animals would be self-administering the drug to recapitulate better the contingent nature of drug taking; however, this method was not feasible for the study reported here. Methamphetamine increases vaginal lubrication in female rats through a nitric oxide-dependent mechanism.

Effect of the Non-steroidal Anti-inflammatory Drug Diclofenac on Ischemia-Reperfusion Injury in Rat Liver: A Nitric Oxide-Dependent Mechanism.

Ischemia/reperfusion injury (IRI) is an inevitable complication of liver surgery and transplantation. The purpose of this study was to examine the beneficial effects of diclofenac on hepatic IRI and the mechanism behind it. Wistar rats' livers were subjected to warm ischemia for 60min followed by 24h of reperfusion. Diclofenac was administered intravenously 15min before ischemia at 10, 20, and 40mg/kg body weight. To determine the mechanism of diclofenac protection, the NOS inhibitor L-Nitro-arginine methyl ester (L-NAME) was administered intravenously 10min after diclofenac injection (40mg/kg). Liver injury was evaluated by aminotransferases (ALT and AST) activities and histopathological analysis. Oxidative stress parameters (SOD, GPX, MPO, GSH, MDA, and PSH) were also determined. Then, eNOS gene transcription and p-eNOS and iNOS protein expressions were evaluated. The transcription factors PPAR-γ and NF-κB in addition to the regulatory protein IκBα were also investigated. Finally, the gene expression levels of inflammatory (COX-2, IL-6, IL-1β, IL-18, TNF-α, HMGB-1, and TLR-4) and apoptosis (Bcl-2 and Bax) markers were measured. Diclofenac, at the optimal dose of 40mg/kg, decreased liver injury and maintained histological integrity. It also reduced oxidative stress, inflammation, and apoptosis. Its mechanism of action essentially depended on eNOS activation rather than COX-2 inhibition, since pre-treatment with L-NAME abolished all the protective effects of diclofenac. To our knowledge, this is the first study demonstrating that diclofenac protects rat liver against warm IRI through the induction of NO-dependent pathway. Diclofenac reduced oxidative balance, attenuated the activation of the subsequent pro-inflammatory response and decreased cellular and tissue damage. Therefore, diclofenac could be a promising molecule for the prevention of liver IRI.

Anti-atherogenic and cardio-protective properties of sweet melon (Cucumis melo. L. Inodorus) seed extract on high fat diet induced obesity in male wistar rats

BackgroundCucumis melon is a medicinal plant with multiple pharmacological properties such as anti-inflammatory, antioxidant, and diuretic effects. An increasing body of scientific evidence established the anti-diabetic/anti-obesity effects of Cucumis melo in humans, mice, and hamster models. However, there are no tangible reports on its ability to prevent cardiovascular complications following diet-induced obesity. The anti-atherogenic and cardioprotective effects of the Methanolic extract of Cucumis melo. L. Inodorus seeds on a high-fat diet (HFD)-induced obese rats was assessed in this study. MethodsForty male Wistar rats were randomly divided into five groups, (n = 8/group); i.e., Normal (N), HFD, HFD + 50 mg/kg b.w. of MCMs (Methanolic extract of Cucumis melon seeds), HFD + 100 mg/Kg b.w. of MCMs and HFD + 200 mg/kg b.w. of MCMs. The experimental animals were anaesthetized and sacrificed after 10 weeks, and blood samples and heart tissue were collected for further analysis. Using the Graph Pad Prism version 5.0, the results expressed as Mean ± SD was tested using the one-way ANOVA to show intergroup differences, followed by Bonferonni ‘s post hoc test. The level of significance was determined at P ≤ 0.05.ResultsMCMs significantly (P < 0.05) reduced body weight, adiposity index, total fat mass, low-density lipoprotein cholesterol (LDL-c), and total cholesterol (TC) compared with the HFD obese groups MCMs caused a significant reduction in the body weight, total fat mass, adiposity index, low-density lipoprotein cholesterol (LDL-c), and total cholesterol (TC) when compared to the animals in HFD obese groups. Also, the Atherogenic index of plasma (AIP), Castelli index and, malondialdehyde (MDA) significantly (P < 0.05) decreased in MCMs treated groups compared to the HFD obese group. The catalase, protein, and HDL levels were significantly increased in MCMs treated groups compared to HFD-obese animals. Expression of nitric oxide in the form of nitrite in the heart tissue significantly increased in the MCMs treated compared to the HFD-obese rats, with the majority of the positive results recorded at 100 mg/Kg b.w. of MCMs.ConclusionsMCMs have anti-atherogenic and Cardio-protective properties on High Fat Diet-Induced Obesity in Male rats via an antioxidant and nitric oxide-dependent mechanism. Further study is recommended to evaluate the molecular mechanisms to which these anti-atherogenic and cardio-protective actions can be attributed and exploit the GCMS result in the development of drug candidates.

Nitric oxide-dependent mechanism of endothelial dysfunction formation is a promising target link for pharmacological management

Nitric oxide-dependent mechanism of endothelial dysfunction formation is a promising target link for pharmacological management

Nitric Oxide Synthase Inhibition Protects Against RotenoneInduced Neurodegeneration in vivo

Neuropathologic and motor features of Parkinson’s disease can be induced in rodents by the pesticide rotenone. In this study, we aimed to determine whether nitric oxide synthase inhibition by NG-nitro-L-arginine methyl ester (L-NAME) could prevent neurodegeneration and biochemical changes induced by rotenone in rat brain. For this purpose, rats received subcutaneous injections of rotenone (1.5 mg/kg) every other day for two consecutive weeks and were treated at the same time with L-NAME at doses of 10 or 20 mg/kg, intraperitoneally. Oxidative stress indictors, malondialdehyde, reduced glutathione, and nitric oxide as well as paraoxonase-1 (PON-1) activity in brain, and the concentration of the antiapoptotic protein B cell/lymphoma-2 (Bcl2) in striatum were determined. Motor strength and coordination was assessed using wire hanging and stair tests. Neuronal degeneration was evaluated using hematoxylin &amp; eosin-stained brain sections. Results: L-NAME reduced, in a dose-dependent manner the increase in lipid peroxidation (malondialdehyde) and nitric oxide and restored reduced glutathione levels in brain of rotenone-treated rats. Additionally, L-NAME prevented the inhibition of paraoxonase-1 activity and the decrease in Bcl2 concentration induced by the pesticide. It also prevented motor impairments in a dose-dependent manner. Furthermore, L-NAME treatment ameliorated the neurodegenerative effects of rotenone, as shown by the decrease in degenerated neurons in striatum and cerebral cortex. The results of the present study suggest that rotenone mediates its neurotoxic effects, at least in part by nitric oxide-dependent mechanisms.

Nitric oxide-mediated effects of L-ornithine-L-aspartate in acute toxic liver injury

This study was aimed to investigate nitric oxide-dependent mechanisms of L-ornithine-L-aspartate (LOLA) action in acute toxic liver injury in rats. Acute hepatitis was induced in Wistar rats using 50% oil solution of tetrachloromethane (CCl4) intragastrically (2 g/kg) twice in a 24 hour interval. Intraperitoneal treatment with LOLA (200 mg/kg) was started 6 hours after the second CCl4 administration and maintained for 3 consecutive days. L-Nω-Nitroarginine Methyl Ester (L-NAME) was used intraperitoneally (10 mg/kg). In CCl4-induced hepatitis, LOLA restores the structure of hepatocytes and prevents aminotransferases, alkaline phosphatase and gamma-glutamyl transferase elevation. It decreases total bilirubin concentration but does not affect increased cholesterol level. LOLA augments urea concentration, total protein level in blood and liver as well as serum and liver content of nitrite anions. LOLA enhances activity of catalase, glutathione S-transferase, manganese superoxide dismutase, increases reduced glutathione level and total antioxidant capacity and decreases thiobarbituric acid reactive substances level. The concomitant use of L-NAME inhibits the action of LOLA to enhance nitrite anions synthesis both in serum and liver, to delay the recovery of hepatocytes, to counteract LOLA effect against blood total protein reduction, to prevent the decline in aminotransferases, alkaline phosphatase,, gamma-glutamyl transferase and glutathione S-transferase activity and to reduce catalase activity and reduced glutathione level. Therefore, in CCl4-induced hepatitis, LOLA effectively prevents cytolysis and cholestasis, improves liver metabolism and protects against oxidative stress. Partially, these changes occur in nitric oxide-mediated mechanism since the use of L-NAME declines most of LOLA effects.

In vitro antitumor activity of nano-pulse stimulation on human anaplastic thyroid cancer cells through nitric oxide-dependent mechanisms

Although most thyroid cancer patients have good clinical outcomes, there are still no effective therapies for patients with anaplastic thyroid cancers (ATC). Nano-pulse stimulation (NPS) is increasingly recognized as a non-thermal ablation technology that selectively targeting cancer cells. The aim of this study was to investigate the in vitro antitumor mechanism of NPS on human ATC cells. The cell line 8305C were treated with NPS and the CCK-8 assay, clonogenic assay, apoptosis assay and nitric oxide synthase (NOS) activity assay were used to test cell proliferation, apoptosis and NOS changes respectively. Our results showed that NPS can significantly (P < 0.05) inhibit ATC cell proliferation and induce apoptosis in a field strength-dependent manner. Moreover, the NOS activity in NPS treated groups obviously increased (P < 0.05). Further analysis indicated that NPS induced an inducible nitric oxide synthase (iNOS)-dependent killing in ATC cells. Finally, our research reveals for the first time that NPS is an effective non-thermal candidate for the treatment of ATC and supports a new possible calcium independent mechanism involvement in NPS treatment.

Melatonin treatment during chronic hypoxic gestation improves neonatal cerebrovascular function

BackgroundFetal chronic hypoxia is associated with blood flow redistribution and oxidative damage in the brain, leading to increased perinatal morbimortality. Melatonin reduces oxidative stress, improves vascular function, and has neuroprotective effects. ObjectivesThis study aimed to determine the effects of an oral melatonin treatment to pregnant ewes at high-altitude, on the cerebrovascular function of their neonates. Study designTen high-altitude pregnant sheep received either vehicle or melatonin (10 mg/d) during the last third of gestation until delivery. Postnatal daily hemodynamic measurements were recorded from lambs until 12 days old. In addition, lambs were submitted to a graded oxygenation protocol to assess cerebrovascular responses. Subsequently, lambs were euthanized, and middle cerebral arteries (MCA) were collected for vascular function, protein levels, and morphostructural analyses. ResultsAntenatal treatment doubled plasma levels of melatonin in pregnant ewes. Melatonin increased carotid flow and decreased carotid vascular resistance in the lambs by the end of the first week. Furthermore, melatonin increased MCA's maximal vasoconstrictor and vasodilator responses, associated with nitric oxide-dependent and independent mechanisms. ConclusionsAn oral treatment with melatonin during pregnancy promotes postnatal cerebral perfusion in chronically hypoxic neonates. Melatonin is a potential treatment for cerebrovascular dysfunction due to perinatal chronic hypoxia.

Supplemental nitrite increases choroidal neovascularization in mice

Low doses of nitrite, close to physiological levels, increase blood flow in normal and ischemic tissues through a nitric oxide (NO) dependent mechanism. Given that nitrite therapy and dietary supplementation with vegetables high in nitrate (e.g. beets) are gaining popularity we decided to determine if low doses of nitrite impact the development of choroidal neovascularization (CNV), a key feature of wet age related macular degeneration (AMD). Sodium nitrite (at 50 mg/L, 150 mg/L, and 300 mg/L), nitrate (1 g/L) or water alone were provided in the drinking water of C57BL/6 J mice aged 2 or 12 months. Mice were allowed to drink ad libitum for 1 week at which time laser-induced choroidal neovascularization (L-CNV) was induced. The mice continued to drink the supplemented water ad libitum for a further 14 days at which point optical coherence tomography (OCT) was performed to determine the volume of the CNV lesion. Blood was drawn to determine nitrite and nitrate levels and eyes taken for histology. CNV volume was 2.86 × 107 μm3 (±0.4 × 107) in young mice on water alone but CNV volume more than doubled to >6.9 × 107 μm3 (±0.8 × 107) in mice receiving 300 mg/L nitrite or 7.34 × 107 μm3 (±1.4 × 107) in 1 g/L nitrate (p < 0.01). A similar trend was observed in older mice. CNV volume was 5.3 × 107 μm3 (±0.5 × 107) in older mice on water alone but CNV volume almost doubled to approximately 9.3 × 107 μm3 (±1.1 × 107) in mice receiving 300 mg/L nitrite or 8.7 × 107 μm3 (±0.9 × 107) 1 g/L nitrate (p < 0.01). Plasma nitrite levels were highest in young mice receiving 150 mg/L in the drinking water with no changes in plasma nitrate observed. In older mice, drinking water nitrite did not significantly change plasma nitrite, but plasma nitrate was increased. Plasma nitrate was elevated in both young and old mice provided with nitrate supplemented drinking water. Our data demonstrate that the CNV lesion is larger in older mice compared to young and that therapeutic levels of oral nitrite increase the volume of CNV lesions in both young and older mice. Therapeutic nitrite or nitrate supplementation should be used with caution in the elderly population prone to CNV.

Nitrate Regulates Maize Root Transcriptome through Nitric Oxide Dependent and Independent Mechanisms.

Maize root responds to nitrate by modulating its development through the coordinated action of many interacting players. Nitric oxide is produced in primary root early after the nitrate provision, thus inducing root elongation. In this study, RNA sequencing was applied to discover the main molecular signatures distinguishing the response of maize root to nitrate according to their dependency on, or independency of, nitric oxide, thus discriminating the signaling pathways regulated by nitrate through nitric oxide from those regulated by nitrate itself of by further downstream factors. A set of subsequent detailed functional annotation tools (Gene Ontology enrichment, MapMan, KEGG reconstruction pathway, transcription factors detection) were used to gain further information and the lateral root density was measured both in the presence of nitrate and in the presence of nitrate plus cPTIO, a specific NO scavenger, and compared to that observed for N-depleted roots. Our results led us to identify six clusters of transcripts according to their responsiveness to nitric oxide and to their regulation by nitrate provision. In general, shared and specific features for the six clusters were identified, allowing us to determine the overall root response to nitrate according to its dependency on nitric oxide.

A 50 Hz magnetic field affects hemodynamics, ECG and vascular endothelial function in healthy adults: A pilot randomized controlled trial.

Application of exposure to 50/60 Hz magnetic fields (MFs) has been conducted in the treatment of muscle pain and fatigue mainly in Japan. However, whether MFs could increase blood flow leading to muscle fatigue recovery has not been sufficiently tested. We investigated the acute effects of a 50 Hz sinusoidal MF at Bmax 180 mT on hemodynamics, electrocardiogram, and vascular endothelial function in healthy young men. Three types of regional exposures to a 50 Hz MF, i.e., forearm, upper arm, or neck exposure to MF were performed. Participants who received three types of real MF exposures had significantly increased ulnar arterial blood flow velocity compared to the sham exposures. Furthermore, after muscle loading exercise, MF exposure recovered hemoglobin oxygenation index values faster and higher than sham exposure from the loading condition. Moreover, participants who received real MF exposure in the neck region had significantly increased parasympathetic high-frequency activity relative to the sham exposure. The MF exposure in the upper arm region significantly increased the brachial artery flow-mediated dilation compared to the sham exposure. Computer simulations of induced in situ electric fields indicated that the order-of-magnitude estimates of the peak values were 100-500 mV/m, depending on the exposure conditions. This study provides the first evidence that a 50 Hz MF can activate parasympathetic activity and thereby lead to increase vasodilation and blood flow via a nitric oxide-dependent mechanism. Trial registration: UMIN Clinical Trial Registry (CTR) UMIN000038834. The authors confirm that all ongoing and related trials for this drug/intervention are registered.

LIT01-196, a Metabolically Stable Apelin-17 Analog, Normalizes Blood Pressure in Hypertensive DOCA-Salt Rats via a NO Synthase-dependent Mechanism.

Apelin is a neuro-vasoactive peptide that plays a major role in the control of cardiovascular functions and water balance, but has an in-vivo half-life in the minute range, limiting its therapeutic use. We previously developed LIT01-196, a systemically active metabolically stable apelin-17 analog, produced by chemical addition of a fluorocarbon chain to the N-terminal part of apelin-17. LIT01-196 behaves as a potent full agonist for the apelin receptor and has an in vivo half-life in the bloodstream of 28 min after intravenous (i.v.) and 156 min after subcutaneous (s.c.) administrations in conscious normotensive rats. We aimed to investigate the effects of LIT01-196 following systemic administrations on arterial blood pressure, heart rate, fluid balance and electrolytes in conscious normotensive and hypertensive deoxycorticosterone acetate (DOCA)-salt rats. Acute i.v. LIT01-196 administration, in increasing doses, dose-dependently decreases arterial blood pressure with ED50 values of 9.8 and 3.1 nmol/kg in normotensive and hypertensive rats, respectively. This effect occurs for both via a nitric oxide-dependent mechanism. Moreover, acute s.c. LIT01-196 administration (90 nmol/kg) normalizes arterial blood pressure in conscious hypertensive DOCA-salt rats for more than 7 h. The LIT01-196-induced blood pressure decrease remains unchanged after 4 consecutive daily s.c. administrations of 90 nmol/kg, and does not induce any alteration of plasma sodium and potassium levels and kidney function as shown by the lack of change in plasma creatinine and urea nitrogen levels. Activating the apelin receptor with LIT01-196 may constitute a novel approach for the treatment of hypertension.

Penicillin G Induces H+, K+-ATPase via a Nitric Oxide-Dependent Mechanism in the Rat Colonic Crypt.

Background/Aims:The colonic H+, K+ ATPase (HKA2) is a heterodimeric membrane protein that exchanges luminal K+ for intracellular H+ and is involved in maintaining potassium homeostasis. Under homeostatic conditions, the colonic HKA2 remains inactive, since most of the potassium is absorbed by the small intestine. In diarrheal states, potassium is secreted and compensatory potassium absorption becomes necessary. This study proposes a novel mechanism whereby the addition of penicillin G sodium salt (penG) to colonic crypts stimulates potassium uptake in the presence of intracellular nitric oxide (NO), under sodium-free (0-Na+) conditions.Methods:Sprague Dawley rat colonic crypts were isolated and pHi changes were monitored through the ammonium prepulse technique. Increased proton extrusion in 0-Na+ conditions reflected heightened H+, K+ ATPase activity. Colonic crypts were exposed to penG, L-arginine (a NO precursor), and N-nitro l-arginine methyl ester (L-NAME, a NO synthase inhibitor).Results:Isolated administration of penG significantly increased H+, K+ ATPase activity from baseline, p 0.0067. Co-administration of arginine and penG in 0-Na+ conditions further upregulated H+, K+ ATPase activity, p <0.0001. Crypt perfusion with L-NAME and penG demonstrated a significant reduction in H+, K+ ATPase activity, p 0.0058.Conclusion:Overall, acute exposure of colonic crypts to penG activates the H+, K+ ATPase in the presence of NO. This study provides new insights into colonic potassium homeostasis.

Triple therapy with pravastatin, low molecular weight heparin and low dose aspirin improves placental haemodynamics and pregnancy outcomes in obstetric antiphospholipid syndrome in mice and women through a nitric oxide-dependent mechanism

ObjectivesA previous pilot study showed that pravastatin supplementation improved pregnancy outcomes in women with obstetric antiphospholipid syndrome (OAPS) that developed placental insufficiency despite standard of care treatment low molecular weight heparin plus low dose aspirin (LMWH + LDA). In this study we investigated the mechanism behind the beneficial effects of the triple therapy LMWH + LDA + pravastatin in improving uteroplacental vascular function and reducing pregnancy complications in OAPS. We hypothesized that nitric oxide (NO) is involved in the vasculoprotective effects of the triple therapy. A mouse model of OAPS that resembles the clinical scenario was used to test this hypothesis. MethodsEleven women with OAPS that developed preeclampsia (PE) and/or intrauterine growth restriction (IUGR) associated with uteroplacental vascular dysfunction despite treatment with LMWH + LDA participated in this study after given informed written consent. Seven women were supplemented with pravastatin at the time abnormal uterine artery Dopplers were detected and 4 remained on LMWH + LDA treatment only. Wire myography was used to identify the mechanisms underpinning the protective effects of the triple therapy in the mouse model of OAPS. ResultsThe triple therapy increased serum NO levels, diminished uteroplacental vessels resistance improving placental function and prolonged pregnancies compared to conventional treatment LMWH + LDA, leading to live births in women with OAPS. Comparable to the observations in women, the triple therapy protected pregnancies in OAPS-mice, increasing placental perfusion and pregnancy outcomes. A synergistic vasculoprotective effect of the triple therapy on uterine arteries and aorta was demonstrated in OAPS-mice. LMWH + LDA showed a partial protection on endothelial function. Addition of pravastatin increase eNOS synthesis, expression and activity/signaling leading to a significant increment in nitric oxide (NO) generation, resulting in improved placental vascular function and total protection of pregnancies. ConclusionLMWH + LDA + PRAV increased serum NO levels and significantly improved placental haemodynamics and maternal and neonatal outcomes in women and mice with OAPS. A role for eNOS/NO in mediating the placental vasculoprotective effects in OAPS-mice was demonstrated, strengthening the concept that impaired NO production is a crucial mediator in the pathogenesis of OAPS and a potential target for pharmacological interventions. The efficacy of pravastatin supplementation should be confirmed in a larger clinical trial.

One Step Closer to a Cure for Preeclampsia?

One Step Closer to a Cure for Preeclampsia?

Involvement of Peroxisome Proliferator-Activated Receptor-a and g in Apelin Action on the Kidney in Type 2 Diabetic Rat Model

AbstractBackground: Apelin is an adipokine that revealed numerousrenoprotective actions; however, its effect in diabeticnephropathy is controversial.Aim of Study: The objectives of the current study wereto determine the action of apelin administration on nephropathyin type 2 diabetic rat model and to clarify its possible mechanismsand if PPARs activation is involved in these mechanisms.Material and Methods: Thirty adult male wistar rats weresubdivided into control, diabetic and diabetic-apelin treatedrats. Type 2 diabetes was induced via a high-fat diet togetherwith a single low-dose of streptozotocin. Apelin was injectedintraperitoneally for 10 weeks. Serum glucose, insulin andlipid profile were estimated. Homeostatic Model Assessmentfor Insulin Resistance (HOMA-IR) was calculated. Renalfunctions were evaluated by serum urea and creatinine, urinaryalbumin excretion, urinary N-acetyl-beta-D-glucosaminidase(NAG) activity and histopathological inspection. Renal tissuehomogenate was assessed for Superoxide Dismutase (SOD),Malondialdehyde (MDA), Nitric Oxide (NO) content andPPAR-a and PPAR-g gene expression.Results: Apelin administration to diabetic rats improvedhyperglycemia, insulin resistance, dyslipidemia, renal functionparameters and pathological lesions in the kidney that resultedfrom induction of diabetes. It elevates renal SOD and NO,decreased MDA and increased PPAR-a and g gene expressionin comparison to diabetic rats.Conclusions: Apelin administration to diabetic rats improvedinsulin resistance, hyperglycemia, dyslipidemia andrenal functions which may be partially via its antioxidantproperties and NO dependent mechanism that struggled theharmful properties of diabetes on the kidney. Besides, apelininduced upregulation of both PPARa/g genes expression thatcould be involved in the renoprotective effect of apelin. Moreinvestigations for the mechanism by which apelin acts onPPARa/g are recommended.

New Nutraceutical Combination Reduces Blood Pressure and Improves Exercise Capacity in Hypertensive Patients Via a Nitric Oxide-Dependent Mechanism.

BackgroundHigh blood pressure (BP) has long been recognized as a major health threat and, particularly, a major risk factor for stroke, cardiovascular disease, and end‐organ damage. However, the identification of a novel, alternative, integrative approach for the control of BP and cardiovascular protection is still needed.Methods and ResultsSixty‐nine uncontrolled hypertension patients, aged 40 to 68 years, on antihypertensive medication were enrolled in 2 double‐blind studies. Forty‐five were randomized to placebo or a new nutraceutical combination named AkP05, and BP, endothelial function, and circulating nitric oxide were assessed before and at the end of 4 weeks of treatment. Twenty‐four patients were randomized to diuretic or AkP05 for 4 weeks and underwent a cardiopulmonary exercise test to evaluate the effects of AkP05 on functional capacity of the cardiovascular, pulmonary, and muscular systems. Vascular and molecular studies were undertaken on mice to characterize the action of the single compounds contained in the AkP05 nutraceutical combination. AkP05 supplementation reduced BP, improved endothelial function, and increased nitric oxide release; cardiopulmonary exercise test revealed that AkP05 increased maximum O2 uptake, stress tolerance, and maximal power output. In mice, AkP05 reduced BP and improved endothelial function, evoking increased nitric oxide release through the PKCα/Akt/endothelial nitric oxide synthase pathway and reducing reactive oxygen species production via NADPH‐oxidase inhibition. These effects were mediated by synergism of the single compounds of AkP05.ConclusionsThis is the first study reporting positive effects of a nutraceutical combination on the vasculature and exercise tolerance in treated hypertensive patients. Our findings suggest that AkP05 may be used as an adjunct for the improvement of cardiovascular protection and to better control BP in uncontrolled hypertension.