Cyclic Guanosine Research Articles

A 2-week treatment with 5-azacytidine improved the hypercontractility state in prostate from obese mice: Role of the nitric oxide-cyclic guanosine monophosphate signalling pathway.

Benign prostatic hyperplasia (BPH) is characterised by increases in prostate volume and contraction. Downregulation of the nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) signalling pathway contributes to prostate dysfunctions. Previous studies in cancer cells or vessels have shown that the epigenetic mechanisms control the gene and protein expression of the enzymes involved in the production of NO and cGMP. This study is aimed to evaluate the effect of a 2-week treatment of 5-azacytidine (5-AZA), a DNA-methyltransferase inhibitor, in the prostate function of mice fed with a high-fat diet. Functional, histological, biochemical and molecular assays were carried out. Obese mice presented greater prostate weight, α-actin expression and contractile response induced by the α-1adrenoceptors agonist. The relaxation induced by the NO-donor and the protein expression of endothelial nitric oxide synthase (eNOS) and soluble guanylate cyclase (sGC) were significantly decreased in the prostate of obese mice. The treatment with 5-AZA reverted the higher expression of α-actin, reduced the hypercontractility state of the prostate and increased the expression of eNOS and sGC and intraprostatic levels of cGMP. When prostates from obese mice treated with 5-AZA were incubated in vitro with inhibitors of the NOS or sGC, the inhibitory effect of 5-AZA was reverted, therefore, showing the involvement of NO and cGMP. In conclusion, our study paves the way to develop or repurpose therapies that recover the expression of eNOS and sGC and, hence, to improve prostate function in BPH.

Role of mitochondrial dysfunction in kidney disease: Insights from the cGAS-STING signaling pathway.

Over the past decade, mitochondrial dysfunction has been investigated as a key contributor to acute and chronic kidney disease. However, the precise molecular mechanisms linking mitochondrial damage to kidney disease remain elusive. The recent insights into the cyclic guanosine monophosphate-adenosine monophosphate (GMP-AMP) synthetase (cGAS)-stimulator of interferon gene (STING) signaling pathway have revealed its involvement in many renal diseases. One of these findings is that mitochondrial DNA (mtDNA) induces inflammatory responses via the cGAS-STING pathway. Herein, we provide an overview of the mechanisms underlying mtDNA release following mitochondrial damage, focusing specifically on the association between mtDNA release-activated cGAS-STING signaling and the development of kidney diseases. Furthermore, we summarize the latest findings of cGAS-STING signaling pathway in cell, with a particular emphasis on its downstream signaling related to kidney diseases. This review intends to enhance our understanding of the intricate relationship among the cGAS-STING pathway, kidney diseases, and mitochondrial dysfunction.

Heme oxygenase-1 is an equid alphaherpesvirus 8 replication restriction host protein and suppresses viral replication via the PKCβ/ERK1/ERK2 and NO/cGMP/PKG pathway.

Equid alphaherpesvirus 8 (EqHV-8) is one of the most economically important viruses that is known to cause severe respiratory disease, abortion, and neurological syndromes in equines. However, no effective vaccines or therapeutic agents are available to control EqHV-8 infection. Heme oxygenase-1 (HO-1) is an antioxidant defense enzyme that displays significant cytoprotective effects against different viral infections. However, the literature on the function of HO-1 during EqHV-8 infection is little. We explored the effects of HO-1 on EqHV-8 infection and revealed its potential mechanisms. Our results demonstrated that HO-1 induced by cobalt-protoporphyrin (CoPP) or HO-1 overexpression inhibited EqHV-8 replication in susceptible cells. In contrast, HO-1 inhibitor (zinc protoporphyria) or siRNA targeting HO-1 reversed the anti-EqHV-8 activity. Furthermore, biliverdin, a metabolic product of HO-1, mediated the anti-EqHV-8 effect of HO-1 via both the protein kinase C (PKC)β/extracellular signal-regulated kinase (ERK)1/ERK2 and nitric oxide (NO)-dependent cyclic guanosine monophosphate (cGMP)-protein kinase G (PKG) signaling pathways. In addition, CoPP protected the mice by reducing the EqHV-8 infection in the lungs. Altogether, these results indicated that HO-1 can be developed as a promising therapeutic strategy to control EqHV-8 infection.IMPORTANCEEqHV-8 infections have threatened continuously donkey and horse industry worldwide, which induces huge economic losses every year. However, no effective vaccination strategies or drug against EqHV-8 infection until now. Our present study found that one host protien HO-1 restrict EqHV-8 replication in vitro and in vivo. Furthermore, we demonstrate that HO-1 and its metabolite biliverdin suppress EqHV-8 relication via the PKCβ/ERK1/ERK2 and NO/cGMP/PKG pathways. Hence, we believe that HO-1 can be developed as a promising therapeutic strategy to control EqHV-8 infection.

Water treatment with free-bubbling corrugated sheet MBR

Having previously introduced a new type corrugated-sheet membrane bioreactor (CS-MBR) with slug bubbling, this work investigates its water treatment performance and hydrodynamic optimization under a free bubbling regime. Experiments in parallel were conducted to compare CS-MBR with flat sheet membrane bioreactor (FS-MBR). The transmembrane pressure (TMP) and permeability results indicate that the corrugated sheet membrane has better anti-fouling performance with a final flux after 15 days of operation that was 45 % higher than that of the flat sheet membrane that had been tested in parallel. The analysis results of effluent quality and sludge extracellular polymeric substances (EPS) indicate that the membrane configuration has little impact on the effluent quality and sludge properties in the system. Scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) revealed the accumulation pattern of foulants on the surface of the corrugated sheet membrane and suggested that the corrugations were the cause of the improved anti-fouling performance. Bis-(3′-5′)-cyclic dimeric guanosine monophosphate (c-di-GMP) was extracted from fouling example at membrane surfaces, which was 50 % lower in the CS-MBR than in the FS-MBR. Computational Fluid Dynamics (CFD) simulations showed that channel gap distance of 2– 4 mm combined with 20 m/s nozzle velocity could produce an intense hydrodynamic effect. Based on the traditional free bubbling air usage, total air consumption of this CS-MBR optimal design is expected to save up to 70 % on energy.

The global regulation of c‐di‐GMP and cAMP in bacteria

AbstractNucleotide second messengers are highly versatile signaling molecules that regulate a variety of key biological processes in bacteria. The best‐studied examples are cyclic AMP (cAMP) and bis‐(3′–5′)‐cyclic dimeric guanosine monophosphate (c‐di‐GMP), which both act as global regulators. Global regulatory frameworks of c‐di‐GMP and cAMP in bacteria show several parallels but also significant variances. In this review, we illustrate the global regulatory models of the two nucleotide second messengers, compare the different regulatory frameworks between c‐di‐GMP and cAMP, and discuss the mechanisms and physiological significance of cross‐regulation between c‐di‐GMP and cAMP. c‐di‐GMP responds to numerous signals dependent on a great number of metabolic enzymes, and it regulates various signal transduction pathways through its huge number of effectors with varying activities. In contrast, due to the limited quantity, the cAMP metabolic enzymes and its major effector are regulated at different levels by diverse signals. cAMP performs its global regulatory function primarily by controlling the transcription of a large number of genes via cAMP receptor protein (CRP) in most bacteria. This review can help us understand how bacteria use the two typical nucleotide second messengers to effectively coordinate and integrate various physiological processes, providing theoretical guidelines for future research.

A c-di-GMP signaling module controls responses to iron in Pseudomonas aeruginosa

Cyclic dimeric guanosine monophosphate (c-di-GMP) serves as a bacterial second messenger that modulates various processes including biofilm formation, motility, and host-microbe symbiosis. Numerous studies have conducted comprehensive analysis of c-di-GMP. However, the mechanisms by which certain environmental signals such as iron control intracellular c-di-GMP levels are unclear. Here, we show that iron regulates c-di-GMP levels in Pseudomonas aeruginosa by modulating the interaction between an iron-sensing protein, IsmP, and a diguanylate cyclase, ImcA. Binding of iron to the CHASE4 domain of IsmP inhibits the IsmP-ImcA interaction, which leads to increased c-di-GMP synthesis by ImcA, thus promoting biofilm formation and reducing bacterial motility. Structural characterization of the apo-CHASE4 domain and its binding to iron allows us to pinpoint residues defining its specificity. In addition, the cryo-electron microscopy structure of ImcA in complex with a c-di-GMP analog (GMPCPP) suggests a unique conformation in which the compound binds to the catalytic pockets and to the membrane-proximal side located at the cytoplasm. Thus, our results indicate that a CHASE4 domain directly senses iron and modulates the crosstalk between c-di-GMP metabolic enzymes.

S-nitrosothiol homeostasis maintained by ADH5 facilitates STING-dependent host defense against pathogens

Oxidative (or respiratory) burst confers host defense against pathogens by generating reactive species, including reactive nitrogen species (RNS). The microbial infection-induced excessive RNS damages many biological molecules via S-nitrosothiol (SNO) accumulation. However, the mechanism by which the host enables innate immunity activation during oxidative burst remains largely unknown. Here, we demonstrate that S-nitrosoglutathione (GSNO), the main endogenous SNO, attenuates innate immune responses against herpes simplex virus-1 (HSV-1) and Listeria monocytogenes infections. Mechanistically, GSNO induces the S-nitrosylation of stimulator of interferon genes (STING) at Cys257, inhibiting its binding to the second messenger cyclic guanosine monophosphate-adenosine monophosphate (cGAMP). Alcohol dehydrogenase 5 (ADH5), the key enzyme that metabolizes GSNO to decrease cellular SNOs, facilitates STING activation by inhibiting S-nitrosylation. Concordantly, Adh5 deficiency show defective STING-dependent immune responses upon microbial challenge and facilitates viral replication. Thus, cellular oxidative burst-induced RNS attenuates the STING-mediated innate immune responses to microbial infection, while ADH5 licenses STING activation by maintaining cellular SNO homeostasis.

Discovery of Pyrido[2,3-d]pyrimidin-7-one Derivatives as Highly Potent and Efficacious Ectonucleotide Pyrophosphatase/Phosphodiesterase 1 (ENPP1) Inhibitors for Cancer Treatment.

Ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1) is an extracellular enzyme responsible for hydrolyzing cyclic guanosine monophosphate-adenosine monophosphate (cGAMP), the endogenous agonist for the stimulator of interferon genes (STING) pathway. Inhibition of ENPP1 can trigger STING and promote antitumor immunity, offering an attractive therapeutic target for cancer immunotherapy. Despite progress in the discovery of ENPP1 inhibitors, the diversity in chemical structures and the efficacy of the agents are far from desirable, emphasizing the demand for novel inhibitors. Herein, we describe the design, synthesis, and biological evaluation of a series of ENPP1 inhibitors based on the pyrido[2,3-d]pyrimidin-7-one scaffold. Optimization efforts led to compound 31 with significant potency in both ENPP1 inhibition and STING pathway stimulation in vitro. Notably, 31 demonstrated in vivo efficacy in a syngeneic 4T1 mouse triple negative breast cancer model. These findings provide a promising lead compound with a novel scaffold for further drug development in cancer immunotherapy.

Use of Genomics to Develop Novel Therapeutics and Personalize Hypertension Therapy.

Hypertension is a prevalent public health problem, contributing to >10 million deaths annually. Though multiple therapeutics exist, many patients suffer from treatment-resistant hypertension or try several medications before achieving blood pressure control. Genomic advances offer mechanistic understanding of blood pressure variability, therapeutic targets, therapeutic response, and promise a stratified approach to treatment of primary hypertension. Cyclic guanosine monophosphate augmentation, aldosterone synthase inhibitors, and angiotensinogen blockade with silencing RNA and antisense therapies are among the promising novel approaches. Pharmacogenomic studies have also been done to explore the genetic bases underpinning interindividual variability in response to existing therapeutics. A polygenic approach using risk scores is likely to be the next frontier in stratifying responses to existing therapeutics.

A STING agonist-loaded bispecific nanobioconjugate modulates macrophage immune responses to enhance antitumor immunotherapy

Tumor-associated macrophages (TAMs) have become attractive therapeutic targets because of their multifaceted cancer-promoting functions and their abundance in the tumor microenvironment (TME). Here, we construct a stimulator of interferon genes (STING) agonist-loaded bispecific nanobioconjugate for macrophage-mediated cancer immunotherapy. Calcium phosphate (CaP) is used as a carrier to load STING agonist, 2′,5′-3′,5′ cyclic guanosine monophosphate-adenosine monophosphate (cGAMP), to prepare cGAMP-loaded CaP nanoparticles (cGAMP@CaP). Two types of antibodies, anti-CD47 antibody (aCD47) and anti-SIRPα antibody (aSIRPα), are modified on the CaP nanoparticles (cGAMP@CaP@Abs). The resulting STING agonist-loaded bispecific nanobioconjugate not only enhances macrophage phagocytosis through blocking of the CD47-SIRPα axis and the physical bridge of macrophages and tumor cells but also potentiates the activation of the STING pathway by cGAMP in TAMs, accordingly effectively triggering a type I interferon-driven inflammatory program. The nanobioconjugate can target and accumulate at the tumor site, facilitate the infiltration of CD8+ T cells, and reprogram the immunosuppressive tumor microenvironment, achieving remarkable therapeutic efficacy in multiple murine tumor models. Overall, the STING agonist-loaded bispecific nanobioconjugate provides a novel platform for macrophage-mediated tumor immunotherapy.

Modelling Cross Talk in the Spatiotemporal System Dynamics of Calcium, IP3 and Nitric Oxide in Neuron Cells.

The bioenergetic system of calcium ([Ca2+]), inositol 1, 4, 5-trisphophate (IP3) and nitric oxide (NO) regulate the diverse mechanisms in neurons. The dysregulation in any or all of the calcium, IP3 and nitric oxide dynamics may cause neurotoxicity and cell death. Few studies are noted in the literature on the interactions of two systems like [Ca2+] with IP3 and [Ca2+] with nitric oxide in neuron cells, which gives limited insights into regulatory and dysregulatory processes in neuron cells. But, no study is available on the cross talk in dynamics of three systems [Ca2+], IP3 and NO in neurons. Thus, the cross talk in the system dynamics of [Ca2+], IP3 and NO regulation processes in neurons have been studied using mathematical model. The two-way feedback process between [Ca2+] and IP3 and two-way feedback process between [Ca2+] and NO through cyclic guanosine monophosphate (cGMP) with plasmalemmal [Ca2+]-ATPase (PMCA) have been incorporated in the proposed model. This coupling handles the indirect two-way feedback process between IP3 and nitric oxide in neuronal cells automatically. The numerical outcomes were acquired by employing the finite element method (FEM) with the Crank-Nicholson scheme (CNS). The present model incorporating the sodium-calcium exchanger (NCX) and voltage-gated calcium channel (VGCC) provides novel insights into the various regulatory and dysregulatory processes due to buffer, IP3-receptor, ryanodine receptor, cGMP kinetics through PMCA channel, etc. and their impacts on the interactive spatiotemporal system dynamics of [Ca2+], IP3 and NO in neurons. It is concluded that the behavior of different crucial mechanisms is quite different for interactions of two systems of [Ca2+] and NO and the interactions of three systems of [Ca2+], IP3 and nitric oxide in neuronal cell due to mutual regulatory adjustments. The association of several neurological disorders with the alterations in calcium, IP3 and NO has been explored in neurons.

Investigating the Evolution of Drosophila STING-Dependent Antiviral Innate Immunity by Multispecies Comparison of 2'3'-cGAMP Responses.

Viruses represent a major threat to all animals, which defend themselves through induction of a large set of virus-stimulated genes that collectively control the infection. In vertebrates, these genes include interferons that play a critical role in the amplification of the response to infection. Virus- and interferon-stimulated genes include restriction factors targeting the different steps of the viral replication cycle, in addition to molecules associated with inflammation and adaptive immunity. Predictably, antiviral genes evolve dynamically in response to viral pressure. As a result, each animal has a unique arsenal of antiviral genes. Here, we exploit the capacity to experimentally activate the evolutionarily conserved stimulator of IFN genes (STING) signaling pathway by injection of the cyclic dinucleotide 2'3'-cyclic guanosine monophosphate-adenosine monophosphate into flies to define the repertoire of STING-regulated genes in 10 Drosophila species, spanning 40 million years of evolution. Our data reveal a set of conserved STING-regulated factors, including STING itself, a cGAS-like-receptor, the restriction factor pastel, and the antiviral protein Vago, but also 2 key components of the antiviral RNA interference pathway, Dicer-2, and Argonaute2. In addition, we identify unknown species- or lineage-specific genes that have not been previously associated with resistance to viruses. Our data provide insight into the core antiviral response in Drosophila flies and pave the way for the characterization of previously unknown antiviral effectors.

Facile manganese ion-coordination assembly of nanogels for synergistic cancer chemo–chemodynamic-immunotherapy

Metal-ion therapy is an emerging technique for cancer treatment, where anticancer metals trigger an immune response against tumor cells due to immunogenic cell death while enabling chemokinetic therapy, and the integration of these activities holds considerable anticancer potential for achieving high specificity and efficiency. Herein, we designed acid-responsive manganese-based nanogels with high drug-loading capacity for combined chemotherapy, chemodynamic therapy, and metal-immune therapy. The chemotherapeutic drug mitoxantrone, metallic manganese ions, and polymeric polyacrylic acid self-assembled into stable nanogels that could be efficiently internalized by cancer cells efficiently, releasing Mn2+ to catalyze highly toxic of hydroxyl generation from endogenous H2O2 and induce tumor cell apoptosis. Furthermore, the Mn2+-mediated activation of cyclic guanosine monophosphate adenosine synthetase activated the stimulator of interferon genes (STING) pathway, thereby inducing dendritic cell maturation. Following intravenous injection into immunocompetent tumor-bearing mice, the nanogels effectively exerted a tumor-killing effect and enhanced the antitumor immune response in combination with an aCTLA-4 antibody. Manganese ions also enhanced T1-weighted magnetic resonance imaging for integrated diagnosis and treatment. Overall, the manganese-based nanogel demonstrates significant synergistic efficacy through combined chemotherapy, chemodynamic therapy and metal-immune therapy, providing a versatile therapeutic strategy for cancer treatment.

Role of the NO-GC/cGMP signaling pathway in platelet biomechanics

Cyclic guanosine monophosphate (cGMP) is a second messenger produced by the NO-sensitive guanylyl cyclase (NO-GC). The NO-GC/cGMP pathway in platelets has been extensively studied. However, its role in regulating the biomechanical properties of platelets has not yet been addressed and remains unknown. We therefore investigated the stiffness of living platelets after treatment with the NO-GC stimulator riociguat or the NO-GC activator cinaciguat using scanning ion conductance microscopy (SICM). Stimulation of human and murine platelets with cGMP-modulating drugs decreased cellular stiffness and downregulated P-selectin, a marker for platelet activation. We also quantified changes in platelet shape using deep learning-based platelet morphometry, finding that platelets become more circular upon treatment with cGMP-modulating drugs. To test for clinical applicability of NO-GC stimulators in the context of increased thrombogenicity risk, we investigated the effect of riociguat on platelets from human immunodeficiency virus (HIV)-positive patients taking abacavir sulfate (ABC)-containing regimens. Our results corroborate a functional role of the NO-GC/cGMP pathway in platelet biomechanics, indicating that biomechanical properties such as stiffness or shape could be used as novel biomarkers in clinical research.

Effective AAV-mediated gene replacement therapy in retinal organoids modeling AIPL1-associated LCA4

Biallelic variations in the aryl hydrocarbon receptor interacting protein-like 1 (AIPL1) gene cause Leber congenital amaurosis subtype 4 (LCA4), an autosomal recessive early-onset severe retinal dystrophy that leads to the rapid degeneration of retinal photoreceptors and the severe impairment of sight within the first few years of life. Currently, there is no treatment or cure for AIPL1-associated LCA4. In this study, we investigated the potential of adeno-associated virus-mediated AIPL1 gene replacement therapy in two previously validated human retinal organoid (RO) models of LCA4. We report here that photoreceptor-specific AIPL1 gene replacement therapy, currently being tested in a first-in-human application, effectively rescued molecular features of AIPL1-associated LCA4 in these models. Notably, the loss of retinal phosphodiesterase 6 was rescued and elevated cyclic guanosine monophosphate (cGMP) levels were reduced following treatment. Transcriptomic analysis of untreated and AAV-transduced ROs revealed transcriptomic changes in response to elevated cGMP levels and viral infection, respectively. Overall, this study supports AIPL1 gene therapy as a promising therapeutic intervention for LCA4.

Soluble guanylate cyclase stimulators and activators: new horizons in the treatment of priapism associated with sickle cell disease.

Priapism, defined as a prolonged and often painful penile erection occurring without sexual stimulation or desire, is a common complication in sickle cell disease (SCD), affecting up to 48% of male patients. This condition presents significant clinical challenges and can lead to erectile dysfunction if not properly managed. Current pharmacological treatments for SCD-related priapism are primarily reactive rather than preventative, highlighting a gap in effective medical intervention strategies. A critical factor in developing priapism is the reduced basal bioavailability of nitric oxide (NO) and cyclic guanosine monophosphate (cGMP) in erectile tissues. New prevention strategies should ideally target the underlying pathophysiology of the disease. Compounds that stimulate and activate soluble guanylate cyclase (sGC) emerge as potential therapeutic candidates since these compounds have the property of inducing cGMP production by sGC. This review explores the potential of sGC stimulators and activators in treating priapism associated with SCD. We discuss the advantages of these agents in the face of the challenging pathophysiology of SCD. Additionally, the review underscores the impact of intravascular hemolysis and oxidative stress on priapism pathophysiology in SCD, areas in which sGC stimulators and activators may also have beneficial therapeutic effects.

FEATURES OF RELATIONS OF MELATONIN WITH THE STATE OF INTRACELLULAR REGULATORSOF THE FUNCTIONAL ACTIVITY OF WHOLE BLOOD MONONUCLEAR CELLS IN CORONARY HEART DISEASE

Introduction. Coronary heart disease (CHD), leading among the causes of death in adulthood and old age, is an urgent medical and social problem. The pathogenesis of most forms of coronary heart disease is based on stenosing atherosclerosis of the coronary arteries, which develops against the background of dyslipidemia and arterial hypertension and is accompanied by the activation of immunocompetent cells (ICCs) of the vascular wall with the development of a subclinical inflammatory reaction, as well as the production of pro-inflammatory factors such as interleukins, chemokines, growth factors and etc. In turn, ICC activity is determined by the state of their intracellular molecular cascades, which transmit signals into the cell and ensure its reactivity to various external stimuli, such as mitogens, cytokines, pathogen components, etcIt has been shown that the central nervous system plays an important role in the regulation of ICC activity due to the production of neurohumoral molecules, such as melatonin, endorphin, sero-tonin, etc., which ensure the coordination of immune responses and their control by the central nervous system. The aim of this study was to study the relationship between melatonin production and intracellular factors that regulate the pro-inflammatory activi-ty of whole blood mononuclear cells and their metabolism in patients with coronary artery disease. Material and methods. As part of the cohort study, 58 patients of both sexes with coronary artery disease aged 49 to 67 years and 20 practically healthy individuals of both sexes were examined. In accordance with the purpose of the study, the concentration of focal adhesion protein kinase (FAK), 5'AMP-activated protein kinase (AMPK), AKT1 protein kinase, signal transducers and transcription activators (STAT) was determined in nuclear cy-toplasmic lysates of whole blood mononuclear cells: STAT3, STAT5A and STAT6, c-Jun N-terminal protein kinase 1 and 2 isoforms (JNK), mitogen-activated protein kinase p38 (p38), extracellular growth kinase 1 and 2 isoforms (ERK), Janus kinase type 2 (JAK2), nuclear transcription factor NF -kB, caspase-1, cyclooxygenase-2 (COX-2), p70-S6K1 protein kinase, p53, p27, p21 proteins. In addition, the concentration of cyclic adenosine monophos-phate (cAMP) and cyclic guanosine monophosphate (cGMP) was determined in cell supernatants. Melatonin concentration was determined in blood se-rum. The material for the study was venous blood taken from the cubital vein in the morning from 6.00 to 6.15. Results. The analysis showed that in patients with coronary artery disease, in comparison with practically healthy individuals, in MNCs of whole blood, there was an increased level of protein kinases FAK, AKT, JNK, ERK, p70-S6K1, factor STAT6, protein p21, against which there was a decrease in the content of STAT3, STAT5A, JAK2, transcription factor NF-kB and caspase-1. These changes were accompanied by increased levels of cGMP and cAMP. Against this background, a decrease in the content of factors was revealed in the MNC. A high concentration of melatonin in patients with CAD was as-sociated with a decrease in the content of protein kinases AMPK, AKT, Jak2, ERK1, protein p21, caspase-1, and cAMP in MNCs, which was observed against the background of an increase in the level of protein p27 and nuclear factor NF-kB. The results of the correlation analysis indicate a different na-ture of the relationship between the level of melatonin and such factors as caspase-1, protein kinases ERK, JAK2, as well as the transcription factor NF-kB and p21 protein, depending on the characteristics of melatonin production in patients with coronary artery disease. Conclusions. In patients with coronary artery disease, melatonin exhibits a modulating effect on the energy balance of ICCs and their metabolism, helps limit pro-inflammatory activity by limiting the functional activity of MAPK/SAPK signaling pathways in MNCs.

Renal and cardiac effects of the PDE9 inhibitor BAY 73-6691 in 5/6 nephrectomized rats.

It has been suggested that the novel selective phosphodiesterase 9 (PDE9) inhibitor may improve cardiac and renal function by blocking 3',5'-cyclic guanosine monophosphate (cGMP) degradation.5/6 nephrectomized (5/6Nx) rats were used to investigate the effects of the PDE9 inhibitor (BAY 73-6691) on the heart and kidney. Two doses of BAY 73-6691 (1mg/kg/day and 5mg/kg/day) were given for 95days.The 5/6Nx rats developed albuminuria, a decrease in serum creatinine clearance (Ccr), and elevated serum troponin T levels. Echocardiographic data showed that 5/6 nephrectomy resulted in increased fractional shortening (FS), stroke volume (SV), and left ventricular ejection fraction (EF). However, 95days of PDE9 inhibitor treatment did not improve any cardiac and renal functional parameter. Histopathologically, 5/6 nephrectomy resulted in severe kidney and heart damage, such as renal interstitial fibrosis, glomerulosclerosis, and enlarged cardiomyocytes. Telmisartan attenuated renal interstitial fibrosis and glomerulosclerosis as well as improved cardiomyocyte size. However, except for cardiomyocyte size and renal perivascular fibrosis, BAY 73-6691 had no effect on other cardiac and renal histologic parameters. Pathway enrichment analysis using RNA sequencing data of kidney and heart tissue identified chronic kidney disease pathways, such as phosphatidylinositol 3-kinase (PI3K)-protein kinase B (Akt) signaling pathway, complement and coagulation cascades, and nuclear factor kappa B (NF-κB) signaling pathway. PDE9i did not affect any of these disease-related pathways.Two dosages of the PDE9 inhibitor BAY 73-6691 known to be effective in other rat models have only limited cardio-renal protective effects in 5/6 nephrectomized rats.

Abstract 29: Neuroprotective Potential of Nitroglycerin (GTN) in Ischemic Stroke: Insights Into Glucose Metabolism and ER Stress Inhibition Through NO-CGMP-PKG Signaling

Background: While Nitroglycerin (GTN) is predominantly recognized as a vasodilator for ischemic heart disease, its potential neuroprotective properties in acute ischemic stroke (AIS) remain under exploration. This investigation sought to discover the therapeutic advantages of GTN post-recanalization in AIS and underlying mechanisms. Methods: Adult male Sprague Dawley rats were categorized into sham, MCAO with or without GTN treatment, and MCAO treated with both GTN and KT5823, an inhibitor of cyclic guanosine monophosphate (cGMP)-protein kinase G (PKG). AIS was induced by MCAO for 2 hours followed by 6 or 24 hours of reperfusion (tMCAO). A 28-hour MCAO without reperfusion was defined as the permanent MCAO group (pMCAO). The study assessed infarct volumes, neurological impairments, glucose metabolism metrics (lactate and ROS levels), nitric oxide (NO) and cGMP levels (via ELISA). mRNA and protein expressions of hyperglycolysis (GLUT1, GLUT3, PFK, LDH), gluconeogenesis (PCK1, PCK2), ER stress (BIP, PERK, EIF2α, ATF4, CHOP) as well as signaling molecules (PKG, AMPK), were measured with RT-PCR and Western blotting. Apoptotic cell death was evaluated using TUNEL. Results: GTN significantly reduced cerebral infarct volumes, neurological deficits, apoptotic cell death, lactate and ROS levels and decreased the expression of NO and cGMP levels and key glucose metabolism and ER stress-related genes and proteins, including GLUT1, GLUT3, PFK1, LDH, PCK1, PCK2, BIP, PERK, EIF2α, ATF4, CHOP and phosphorylated AMPK, while boosting PKG expression only in tMCAO. The coadministration of GTN and KT5823 reversed the observed GTN benefits. Conclusion: Our findings illuminate that GTN showcases neuroprotective properties in tMCAO (but not pMCAO) by enhancing glucose metabolism (hyperglycolysis and gluconeogenesis), controlling ER stress, and working through the NO-CGMP-PKG signaling cascade to inhibit AMPK phosphorylation.

Safety, tolerability, and pharmacokinetics of aildenafil citrate tablets, a novel oral PDE5 inhibitor, in healthy Chinese volunteers after multiple-dose administration.

Aildenafil citrate is a potent and selective inhibitor of cyclic guanosine monophosphate-specific phosphodiesterase type 5, developed for the treatment of erectile dysfunction (ED). This study aimed to assess the pharmacokinetics, safety, and tolerability of aildenafil citrate tablets after multiple doses in healthy Chinese males. Twenty participants were divided into 2 groups, 10 participants each. Participants were administered multiple doses of aildenafil citrate tablets at 30and 60mg. The safety evaluation was based on clinical symptoms and adverse events. Concentrations of aildenafil and its key metabolites (M1, M5, and M12) in human serum were measured by liquid chromatography-tandem mass spectrometry. Pharmacokinetic analysis showed rapid absorption and elimination of aildenafil, with a median time to maximum serum concentration of 1hour and mean terminal half-lives of 2.75and 3.26hours in the respective dose groups. The mean maximum concentration was proportional to the aildenafil dose in the range of 30to 60mg, although the area under the curve was not proportional for serum concentration vs time 0 to the last measurable time point (24 hours). Multiple doses of aildenafil were well tolerated, with 60.0% of men experiencing treatment-emergent adverse events, notably myalgia and fatigue, particularly in the 60-mg group. Aildenafil citrate tablets demonstrated favorable tolerability with once-daily administration over the clinical dose range. The occurrence of myalgia and fatigue was more prevalent in the 60-mg group. From a pharmacokinetic perspective, optimal administration of aildenafil citrate tablets appears to be 1hour before sexual intercourse in men with ED. This study presents robust safety and pharmacokinetic data at expected therapeutic doses, unaffected by clinical factors. The efficacy of aildenafil citrate tablets warrants further validation in individuals with ED. Aildenafil citrate tablets exhibited good tolerability in healthy Chinese males following multiple doses at 30and 60mg. The 60-mg group showed an increased incidence of myalgia and fatigue, suggesting the need for heightened clinical vigilance. The mean maximum concentration, but not the area under the curve, displayed dose proportionality within the 30- to 60-mg dose range, and no significant drug accumulation was observed with repeated daily administration. CTR20192473 (http://www.chinadrugtrials.org.cn).