Nitric Oxide Synthase Activity Research Articles

Endothelial insulin resistance induced by adrenomedullin mediates obesity-associated diabetes

Insulin resistance is a hallmark of obesity-associated type 2 diabetes. Insulin’s actions go beyond metabolic cells and also involve blood vessels, where insulin increases capillary blood flow and delivery of insulin and nutrients. We show that adrenomedullin, whose plasma levels are increased in obese humans and mice, inhibited insulin signaling in human endothelial cells through protein-tyrosine phosphatase 1B–mediated dephosphorylation of the insulin receptor. In obese mice lacking the endothelial adrenomedullin receptor, insulin-induced endothelial nitric oxide–synthase activation and skeletal muscle perfusion were increased. Treating mice with adrenomedullin mimicked the effect of obesity and induced endothelial and systemic insulin resistance. Endothelial loss or blockade of the adrenomedullin receptor improved obesity-induced insulin resistance. These findings identify a mechanism underlying obesity-induced systemic insulin resistance and suggest approaches to treat obesity-associated type 2 diabetes.

Efficacy of thiazole derivatives against colorectal cancer induced by dimethylhydrazine in male Wistar rats.

Identifying novel biological activities in hit compounds remains a significant challenge in cancer research. In this study, we evaluated the anticancer and safety profiles of two previously studied thiazole-based derivatives, Les-5303 and Les-6485, in a colorectal cancer (CRC) model induced by dimethylhydrazine (DMH) in male Wistar rats. CRC was induced through subcutaneous DMH administration at a 20mg/kg dose for 20 weeks. Les-5303 and Les-6485 were then administered intrarectally at a 6mg/kg dose for 5 days. The effects of these compounds on oxidative stress, the antioxidant system, inflammation, NO-synthase activity in the colonic mucosa and blood plasma, and hepatotoxicity markers, were thoroughly assessed. Both compounds demonstrated significant anticancer activity and antioxidant properties. Les-5303, however, exhibited increased hepatotoxicity, as evidenced by elevated AST activity and reduced urea concentration in the blood, indicating its potential for liver damage. In contrast, Les-6485 showed no significant hepatotoxic effects, maintaining normal hepatic enzyme activities and urea levels, suggesting a better safety profile. These findings highlight the distinct biological effects of Les-5303 and Les-6485, with Les-6485 showing promising anticancer activity coupled with minimal toxicity, making it a more favorable candidate for further development in CRC treatment.

Advances in the Regulation of Inflammatory Mediators in Nitric Oxide Synthase: Implications for Disease Modulation and Therapeutic Approaches

Nitric oxide synthases (NOS) are crucial enzymes responsible for the production of nitric oxide (NO), a signaling molecule with essential roles in vascular regulation, immune defense, and neurotransmission. The three NOS isoforms, endothelial NOS (eNOS), neuronal NOS (nNOS), and inducible NOS (iNOS), are tightly regulated by inflammatory mediators and cellular signaling pathways. While physiological NO production is vital for maintaining homeostasis, dysregulated NOS activity contributes to the pathogenesis of numerous diseases, including cardiovascular disorders, neurodegenerative conditions, and cancer. Recent advances in understanding the molecular mechanisms of NOS regulation have unveiled novel therapeutic opportunities, including isoform-specific modulators, upstream pathways, and nanotechnology-enhanced delivery systems. This review highlights these advancements, offering insights into how targeting NOS and its regulatory network can enable precise and effective therapeutic strategies for managing inflammation-driven pathologies.

Activation of APJ Receptors by CMF-019, But Not Apelin, Causes Endothelium-Dependent Relaxation of Spontaneously Hypertensive Rat Coronary Arteries.

Receptors for the vasoactive adipokine apelin, termed APJ receptors, are G-protein-coupled receptors and are widely expressed throughout the cardiovascular system. APJ receptors can also signal via G-protein-independent pathways, including G-protein-coupled-receptor kinase 2 (GRK2), which inhibits nitric oxide synthase (eNOS) activity and nitric oxide (NO) production in endothelial cells. Apelin causes endothelium-dependent, NO-mediated relaxation of coronary arteries from normotensive animals, but the effects of activating APJ receptor signaling pathways in hypertensive coronary arteries are largely unknown. We hypothesized that apelin-induced relaxation is impaired in coronary arteries from spontaneously hypertensive rats (SHR). Western blot and mRNA analysis revealed increased GRK2 expression in cultured SHR coronary endothelial cells. Apelin failed to cause relaxation in isolated SHR coronary arteries but, in the presence of apelin, relaxations to acetylcholine (ACh) were impaired. Apelin had no effect on relaxation to DEA NONOate. The GRK2 inhibitor, CMPD101, increased apelin-induced phosphorylation of Akt and eNOS in SHR endothelial cells and restored relaxation to apelin in SHR arteries. CMPD101 also blocked the inhibitory effect of apelin on ACh-induced relaxation. Relaxations to the APJ receptor-biased agonist, CMF-019, which preferentially activates the G-protein-dependent pathway with minimal effect on GRK2, were similar in SHR and WKY coronary arteries. Immunoblot analysis in SHR coronary endothelial cells demonstrated that CMF-019 increased Akt and eNOS phosphorylation whereas apelin had no effect. Thus, APJ receptor signaling via GRK2 impairs NO production or release from SHR endothelial cells. APJ receptor-biased agonists, such as CMF-019, may be more effective than apelin in causing vasodilation of SHR coronary arteries.

Deciphering the impact of NOS-derived NO on nitrogen metabolism and carbon flux in the heterocytous cyanobacterium Aphanizomenon flos-aquae 2012/KM1/D3.

Nitric oxide synthases (NOSs) are heme-based monooxygenases that catalyze the NADPH-dependent oxidation of L-arginine to produce NO and L-citrulline. Over the past five years, the identification and characterization of NOS homologs in cyanobacteria have significantly advanced our understanding of these enzymes. However, the precise mechanisms through which NOS-derived NO influences nitrogen metabolism remain incompletely elucidated. Therefore, the present study aims to investigates the impact of NOS-derived NO on nitrogen metabolism, heterocyte development, and carbon utilization dynamics in Aphanizomenon flos-aquae. Results demonstrate a three-fold increase in NOS-dependent NO production during the log to stationary growth phase in reponse to L-arginine availability. This increase in NOS activity substantially impacted critical cellular processes related to nitrogen metabolism. Specifically, the inhibition of NOS activity disrupted regulatory mechanisms involving ntcA and glnB genes, resulting in a failure to induce hetR, hep, dev and nif genes necessary for heterocyte differentiation and nitrogenase synthesis. NOS-derived NO also played a pivotal role in modulating the glutamine synthetase-glutamate synthase (GS-GOGAT) pathway, as evidenced by the sharp decline in glutamine and glutamate levels under NOS inhibition, which indicates impaired nitrogen assimilation. Besides, the observed alterations in succinate, fumarate, malate and pyruvate suggest regulatory roles of NOS in energy metabolism. NOS-inhibited cells redirected carbon flux towards glycogen/lipid biosynthesis, alongside protein degradation causing chlorosis, indicating nitrogen deficiency and compromised cellular viability. In contrast, NOS elicitation enhanced metabolic activity, supporting nitrogen assimilation and cellular growth. Overall, our results revealed the complex relationship among NOS-derived NO signaling, nitrogen metabolism, and carbon flux in cyanobacteria.

Fibrillin-1 deficiency perturbs aortic cholinergic relaxation and adrenergic contraction in a mouse model of early onset progressively severe Marfan syndrome.

The pathogenic role of nitric oxide (NO) signaling during development of thoracic aortic aneurysm (TAA) in Marfan syndrome (MFS) is currently unclear. We characterized vasomotor function and its relationship to the activity of the NO-generating enzymes in mice with early onset progressively severe MFS. Wire myography, immunoblotting, measurements of aortic NO and superoxide levels were used to compare vasomotor function, contractile-protein levels, and the activity of endothelial and inducible NO synthase (eNOS and iNOS, respectively) in ascending thoracic aortas of Fbn1mgR/mgR mice relative to wild type (WT) littermates. Isometric force measurements of aortic rings from 16-day-old male Fbn1mgR/mgR mice revealed a significant reduction in acetylcholine (ACh)-induced relaxation and increased phenylephrine (PE)-promoted contractility, associated with abnormally low eNOSSer1177 phosphorylation, decreased NO production and augmented superoxide levels. Greater aortic contractility was associated with α1-adrenoceptor upregulation and normal levels of contractile proteins. While iNOS inhibition had no effect on vasomotor functions, mutant aortic rings pre-incubated with a non-specific NOS inhibitor yielded a greater PE response, implying a significant contribution of endothelial dysfunction to aortic hypercontractility. Impaired eNOS signaling disrupts aortic cholinergic relaxation and adrenergic contraction in MFS mice with dissecting TAA.

Regulation of nitric oxide generation and consumption.

Nitric oxide (NO), originally discovered for its role in cardiovascular function, is a key molecule in physiological processes including metabolism, neurotransmission (including memory, learning, neuroprotection and synaptic plasticity), immunity, reproduction, and much more. NO can be synthesized by the catalytic activity of the enzyme nitric oxide synthase (NOS), which is found biologically in three isoforms, or nonenzymatically based on simple reduction of nitrate and nitrite or by the NO-donor S-nitrosothiol (R-SNO). Importantly, the deficiency of NO has been noted in a wide range of pathologies including cardiovascular disease, cancer, erectile dysfunction, male and female infertility, and mitochondrial disease. While there are several pathways that can lead to a reduction in the bioavailability of NO (i.e., consumption, inhibition, and substrate competition) it is the conclusion of the authors that multiple pathways co-exist in pathological states. This article outlines for the first time the major pathways of NO generation, the importance of NO in health, NO scavenging and enzyme inhibition, and the potential benefits of supplementation.

Luminal flow in the connecting tubule induces afferent arteriole vasodilation.

Renal autoregulatory mechanisms modulate renal blood flow. Connecting tubule glomerular feedback (CNTGF) is a vasodilator mechanism in the connecting tubule (CNT), triggered paracrinally when high sodium levels are detected via the epithelial sodium channel (ENaC). The primary activation factor of CNTGF-whether NaCl concentration, independent luminal flow, or the combined total sodium delivery-is still unclear. We hypothesized that increasing luminal flow in the CNT induces CNTGF via O2- generation and ENaC activation. Rabbit afferent arterioles (Af-Arts) with adjacent CNTs were microperfused ex-vivo with variable flow rates and sodium concentrations ranging from < 1 to 80mM and from 5 to 40 nL/min flow rates. Perfusion of the CNT with 5mM NaCl and increasing flow rates from 5 to 10, 20, and 40 nL/min caused a flow-rate-dependent dilation of the Af-Art (P < 0.001). Adding the ENaC blocker benzamil inhibited flow-induced Af-Art dilation, indicating a CNTGF response. In contrast, perfusion of the CNT with < 1mM NaCl did not result in flow-induced CNTGF vasodilation (P > 0.05). Multiple linear regression modeling (R2 = 0.51; P < 0.001) demonstrated that tubular flow (β = 0.163 ± 0.04; P < 0.001) and sodium concentration (β = 0.14 ± 0.03; P < 0.001) are independent variables that induce afferent arteriole vasodilation. Tempol reduced flow-induced CNTGF, and L-NAME did not influence this effect. Increased luminal flow in the CNT induces CNTGF activation via ENaC, partially due to flow-stimulated O2- production and independent of nitric oxide synthase (NOS) activity.

Pyrroloquinoline quinone-loaded coaxial nanofibers prevent oxidative stress after spinal cord injury.

Oxidative stress plays a major role in the secondary injury of the spinal cord tissue due to the high lipid content of nervous tissue. In the present study, coaxial nanofibers were loaded with the natural antioxidant pyrroloquinoline quinone (PQQ) and used as an implantable drug-delivery system and a scaffold post-SCI. The obtained data show that the concentration of NO and the activity of inducible nitric oxide synthase (iNOS) were significantly (P < 0.05) increased in the spinal cord injury (SCI) group. These levels were significantly decreased following treatment with nanofibers/PQQ. Implantation of nanofibers/PQQ resulted in a significant (P < 0.05) drop in the level of malondialdehyde (MDA) compared to the SCI group. The application of nanofibers loaded with PQQ after SCI caused a significant (P < 0.05) elevation of superoxide dismutase (SOD) and catalase (CAT) activity in the spinal cord tissue. The present work shows the protective role of coaxial nanofibers loaded with PQQ against oxidative stress in spinal cord injury. The reversal of oxidative stress with PQQ can lead to better outcomes following spinal cord injury.

Endothelial cell heterogeneity in arteriolar annuli: Implications in retinal blood flow regulation

Aims/Purpose: To investigate the endothelial cell heterogeneity in retinal arterioles with special focus in arteriolar annuli.Methods: We compared structural and molecular characteristics between endothelial cells located in arteriolar annuli and endothelial cells located in other regions of retinal arterioles. Immunohistochemical, histochemical, and electron transmission microscopy assays were performed.Results: Molecular analysis showed that arteriolar annuli endothelial cells showed a decreased expression of PECAM‐1 together with an increased expression of NADPH‐diaphorase in comparison with other regions of retinal arterioles. PECAM‐1 is involved cell‐cell adhesion. Therefore, this result suggests a decrease in cellular adhesion at the arteriolar annuli. NADPH diaphorase reflects nitric oxide synthase activity. Thus, its increase indicates NO synthesis suggesting localized vasodilation and decrease of platelet aggregation at arteriolar annuli. An increased expression of von Willebrand factor in concomitance with a higher presence of Weibel‐Palade bodies was also observed in endothelial cells at the arteriolar annuli, suggesting that von Willebrand factor is stored and released in a regulated manner. Furthermore, transmission electron microscopy showed the presence of two endothelial cell subtypes in arteriolar annuli: activated endothelial cells, and shear stress‐adapted endothelial cells. Activated endothelial cells exhibited enlarged nuclei with chromatin condensed in the periphery, swollen perinuclear space, and an increased number of vesicles and caveolae. By contrast, shear stress‐adapted endothelial cells were thicker than the rest of endothelial cells and showed bilobulated nuclei, a well‐developed endoplasmic reticulum, as well as large increase in the number of mitochondria.Conclusions: All our results suggest that endothelium at arteriolar annuli, a structure suggested to regulate retinal blood flow by exerting a sphincter‐like activity, is formed by a subset of endothelial cells endowed with distinctive structural and molecular features which allow them to play a key role in the maintenance of blood flow through two mechanisms: the establishment of an antithrombotic milieu, and the vasodilation of arteriolar annuli.

Insights into the differences of caspase and apoptosis levels in pork longissimus thoracis muscles with different tenderness: A perspective on S-nitrosylation modification.

This study investigated the differences of caspase and apoptosis levels in pork with different tenderness from the perspective of S-nitrosylation and further explored their role during pork tenderization. Ten longissimus thoracis muscles selected from 36 individual carcasses based on shear force were divided into high (HT) and low (LT) tenderness groups (n=5), respectively. Results demonstrated that total nitric oxide synthase activity and protein S-nitrosylation levels of LT group were higher than HT group, while myocyte apoptosis levels were lower in LT group (p<0.05). Additionally, LT group possessed a lower caspase-3 activity while a higher abundance of intact caspase-3 and greater S-nitrosylation levels of cleaved caspase-3 (p<0.05). However, none of the above differences were found in caspase-9 (p>0.05). The lower tenderness in LT group might be associated with reduced caspase-3 activity resulted from increased S-nitrosylation levels of its active subunits, which delayed myocyte apoptosis and lowed the degradation of desmin and troponin T.

Impact of Soybean Bioactive Peptides on Growth, Lipid Metabolism, Antioxidant Ability, Molecular Responses, and Gut Microbiota of Oriental River Prawn (Macrobrachium nipponense) Fed with a Low-Fishmeal Diet.

The substitution of fishmeal with high-level soybean meal in the diet of crustaceans usually induces lipid accumulation and oxidative stress in the hepatopancreas. Therefore, it is essential to alleviate these adverse effects. In the present study, SBPs were used to alleviate the negative effects of a fishmeal decrease on the growth performance, lipid metabolism, antioxidant capacity, and gut microbiota of oriental river prawn (Macrobrachium nipponense) in an 8-week feeding trial. Three isonitrogenic and isolipidic diets were prepared as follows: R (reference diet with 32% fishmeal), CT (control diet with 22% fishmeal), and SBP (22% fishmeal with 1.25 g/kg soybean bioactive peptides). The prawns (initial biomass per tank 17 g) were randomly divided into three groups with four replicates. The results showed that the low-fishmeal diet induced the following: (1) the inhibition of growth performance and survival of prawns; (2) an increase in triglyceride content in the hepatopancreas and hemolymph and downregulation of carnitine palmitoyl transferase 1 (cpt1) gene expression; (3) a reduction in antioxidant enzymes' activities and their genes expression levels and an increase malondialdehyde (MDA) content; and (4) an increase in the abundance of the conditional pathogen Pseudomonas in the gut. SBPs supplementation in the CT diet effectively alleviated most of the above adverse effects. SBPs enhanced inducible nitric oxide synthase (iNOS) activity to synthesize nitric oxide (NO) by activating the imd-relish pathway. Most importantly, SBPs increased the potential probiotic Rikenellaceae_RC9_gut_group abundance and decreased the abundance of the conditional pathogen Pseudomonas in the gut. In conclusion, SBPs supplementation can improve low-fishmeal-diet-induced growth inhibition by regulating the gut microbiota composition to ameliorate lipid deposition and oxidative stress and strengthen immune status in oriental river prawn.

Neuronal nitric oxide synthase activation by tadalafil protects neurological impairments in a zebrafish larva model of hyperammonemia.

Hyperammonaemia (HA) is a metabolic disorder characterized by increased ammonia levels in the blood and is associated with severe neurological impairments. Some previous findings have shown the involvement of the nitric oxide pathway in HA-induced neurological impairments. The current study explored the impact of tadalafil on neurological impairments induced by HA in a zebrafish larval model due to its reported indirect interactions with the nitric oxide pathway. HA was induced in zebrafish larvae by ammonium acetate exposure from 2 to 9 days post fertilization (dpf). Locomotor and cognitive functions were analysed following the treatment. The levels of gamma-aminobutyric acid (GABA), glutamate, and dopamine were measured in the larval head. The expression of genes associated with apoptosis (baxa and bcl2a), selected neurotransmitter receptors and bdnf was analysed. The protein levels of CREB and nNOS were also quantified. Tadalafil incubation reversed the HA-associated locomotor and cognitive impairments in larvae. The treatment attenuated GABA, dopamine, and glutamate levels. An upregulation in the expression of grin1a, gria2b, drd1b, drd2b, bdnf, and bcl2a, and downregulation of gabrz, gabrd, gabrg2 and baxa was observed following tadalafil treatment. The protein expression showed increased nNOS, p-CREB(Ser133), and decreased p-nNOS(Ser847) levels in the larvae incubated with tadalafil. The study concluded that tadalafil mitigates HA-induced neurological impairments by activating neuronal nitric oxide synthase. The study highlighted the possible application of tadalafil in the symptomatic management of neurological impairments in HA provided its efficacy and safety are further ensured in higher mammals.

Mechanisms of action of ethyl acetate fractions of Liparis nervosa (Thunb.) Lindl. as potential central anti-nociceptive agents.

Opioids/non-steroidal anti-inflammatory drugs are used to alleviate pain; however, they are expensive and can have adverse effects, especially when used over extended periods. Therefore, there is immense demand for innovative, non-addictive analgesics. Here, we report a novel plant-derived central anti-nociceptive agent, Liparis nervosa (Thunb.) Lindl. (LN), validated in animal pain models. Ethyl acetate fractions of L. nervosa (EALN) exhibited central anti-nociceptive activity in hot plate, tail immersion, formalin-induced paw oedema, and acetic acid-induced abdominal writhing tests. The chemical composition of the EALN was determined using ultra-high-performance liquid chromatography-mass spectrometry. Reserpine (monoamine transmitter-depleting agent) and naltrexone (opioid antagonist) partially suppressed the anti-nociceptive effect of EALN in both phases of the formalin test. Oral administration of EALN activated the endogenous opioid and central descending inhibitory systems by increasing β-endorphin, 5-hydroxytryptamine, and norepinephrine expression. EALN treatment increased the expression of γ-aminobutyric acid B; inhibited the expression of prostaglandin E2, substance P, calcitonin gene-related peptide, and c-Fos; and blocked the transmission of pain signals in the spinal cord. EALN treatment reduced the activity of nitric oxide and nitric oxide synthase in the central region and inhibited the nitric oxide-cyclic guanosine monophosphate signal transduction pathway, thereby attenuating the transmission of nociceptive information in the descending inhibitory pathways. The central anti-nociceptive effect of EALN was achieved by integrating these pathways. This study provides new insights into the pharmacologic action of LN and provide a therapeutic approach as a promising candidate for central anti-nociceptive agents.

Vitamins and other nutrients that support nitric oxide homeostasis and counteract the development of endotheliopathy

Regulation of blood nitric oxide (NO) levels is an essential component of maintaining cerebrovascular and cardiovascular blood flow. The production of NO in the vascular endothelium is influenced by numerous factors, including the supply of macro- and micronutrients. This article presents the results of systematization of data on the relationship between NO metabolism and nutrient factors. Particular attention is paid to the effect of group B vitamins (folates, vitamins B1, B2, B7 and B12) and the microbiome on vascular endothelial function, NO synthase activity and the prevention of oxidative-nitrosative stress. The correction of deficiencies in these vitamins (group B, A, C, E, D3), magnesium and calcium is an important part of the prevention of stress and a variety of diseases of the nervous system associated with endothelial dysfunction.

Subcellular Localization Guides eNOS Function.

Nitric oxide synthases (NOS) are enzymes responsible for the cellular production of nitric oxide (NO), a highly reactive signaling molecule involved in important physiological and pathological processes. Given its remarkable capacity to diffuse across membranes, NO cannot be stored inside cells and thus requires multiple controlling mechanisms to regulate its biological functions. In particular, the regulation of endothelial nitric oxide synthase (eNOS) activity has been shown to be crucial in vascular homeostasis, primarily affecting cardiovascular disease and other pathophysiological processes of importance for human health. Among other factors, the subcellular localization of eNOS plays an important role in regulating its enzymatic activity and the bioavailability of NO. The aim of this review is to summarize pioneering studies and more recent publications, unveiling some of the factors that influence the subcellular compartmentalization of eNOS and discussing their functional implications in health and disease.

Veratridine Induces Vasorelaxation in Mouse Cecocolic Mesenteric Arteries.

The vegetal alkaloid toxin veratridine (VTD) is a selective voltage-gated Na+ (NaV) channel activator, widely used as a pharmacological tool in vascular physiology. We have previously shown that NaV channels, expressed in arteries, contribute to vascular tone in mouse mesenteric arteries (MAs). Here, we aimed to better characterize the mechanisms of action of VTD using mouse cecocolic arteries (CAs), a model of resistance artery. Using wire myography, we found that VTD induced vasorelaxation in mouse CAs. This VTD-induced relaxation was insensitive to prazosin, an α1-adrenergic receptor antagonist, but abolished by atropine, a muscarinic receptor antagonist. Indeed, VTD-vasorelaxant effect was totally inhibited by the NaV channel blocker tetrodotoxin (0.3 µM), the NO synthase inhibitor L-NNA (20 µM), and low extracellular Na+ concentration (14.9 mM) and was partially blocked by the NCX1 antagonist SEA0400 (45.4% at 1 µM). Thus, we assumed that the VTD-induced vasorelaxation in CAs was due to acetylcholine release by parasympathetic neurons, which induced NO synthase activation mediated by the NCX1-Ca2+ entry mode in endothelial cells (ECs). We demonstrated NCX1 expression in ECs by RT-qPCR and immunohisto- and western immunolabelling. VTD did not induce an increase in intracellular Ca2+ ([Ca2+]i), while SEA0400 partially blocked acetylcholine-triggered [Ca2+]i elevations in Mile Sven 1 ECs. Altogether, these results illustrate that VTD activates NaV channels in parasympathetic neurons and then vasorelaxation in resistance arteries, which could explain arterial hypotension after VTD intoxication.

Hepatic GPx1 and GIT1 expression altered by ethanol exposure during third trimester-equivalent development

Background Ethanol (EtOH) exposure throughout gestation and breastfeeding leads to multiple adverse outcomes in the hepatic system. Under oxidative stress, alterations in the liver are related to the inhibition of induced nitric oxide synthase activity in sinusoidal cells as a consequence of low expression of G-protein-coupled receptor (GPCR)-kinase interacting (GIT1). Here, we hypothesized that both glutathione peroxidase 1 (GPx1) and GIT1 could be altered by EtOH exposure during the third trimester of human equivalent development. Methods We exposed rats during the third trimester equivalent [postnatal days (PD) 2-8] to moderate levels of maternal EtOH (20%). GPx1 and GIT1 expression was detected by western blotting, and the antioxidant activity of glutathione peroxidase GPx and the concentration of hepatic carbonyl groups (CG were determined by spectrophotometry. Serum biochemistry parameters such as alanine aminotransferase (ALT), glucose (gluc), cholesterol (chol), and triglycerides (TG) were also measured. Results We found that ethanol decreased both GIT1 and GPx1 selenoprotein expression, affecting GPx antioxidant activity and increasing protein oxidation. Conclusions These results demonstrate for the first time that the GPx antioxidant system altered by EtOH exposure during the third trimester of development is related to a parallel decrease in GIT1 expression [1].

Human airway epithelium controls Pseudomonas aeruginosa infection via inducible nitric oxide synthase.

Airway epithelial cells play a central role in the innate immune response to invading bacteria, yet adequate human infection models are lacking. We utilized mucociliary-differentiated human airway organoids with direct access to the apical side of epithelial cells to model the initial phase of Pseudomonas aeruginosa respiratory tract infection. Immunofluorescence of infected organoids revealed that Pseudomonas aeruginosa invades the epithelial barrier and subsequently proliferates within the epithelial space. RNA sequencing analysis demonstrated that Pseudomonas infection stimulated innate antimicrobial immune responses, but specifically enhanced the expression of genes of the nitric oxide metabolic pathway. We demonstrated that activation of inducible nitric oxide synthase (iNOS) in airway organoids exposed bacteria to nitrosative stress, effectively inhibiting intra-epithelial pathogen proliferation. Pharmacological inhibition of iNOS resulted in expansion of bacterial proliferation whereas a NO producing drug reduced bacterial numbers. iNOS expression was mainly localized to ciliated epithelial cells of infected airway organoids, which was confirmed in primary human lung tissue during Pseudomonas pneumonia. Our findings highlight the critical role of epithelial-derived iNOS in host defence against Pseudomonas aeruginosa infection. Furthermore, we describe a human tissue model that accurately mimics the airway epithelium, providing a valuable framework for systemically studying host-pathogen interactions in respiratory infections.

Pasteurized Akkermansia muciniphila Ameliorates Preeclampsia in Mice by Enhancing Gut Barrier Integrity, Improving Endothelial Function, and Modulating Gut Metabolic Dysregulation.

Preeclampsia (PE) is a serious complication of pregnancy linked to endothelial dysfunction and an imbalance in the gut microbiota. While Akkermansia muciniphila (AKK) has shown promise in alleviating PE symptoms, the use of live bacteria raises safety concerns. This study explored the potential of pasteurized A. muciniphila (pAKK) as a safer alternative for treating PE, focusing on its effects on endothelial function and metabolic regulation. A PE mouse model was induced via the nitric oxide synthase inhibitor L-NAME, followed by treatment with either pAKK or live AKK. Fecal metabolomic profiling was performed via liquid chromatography-tandem mass spectrometry (LC-MS/MS), and in vivo and in vitro experiments were used to assess the effects of pAKK on endothelial function and metabolic pathways. pAKK exhibited therapeutic effects comparable to those of live AKK in improving L-NAME-induced PE-like phenotypes in mice, including enhanced gut barrier function and reduced endotoxemia. pAKK also promoted placental angiogenesis by restoring endothelial nitric oxide synthase (eNOS) activity and nitric oxide (NO) production. The in vitro experiments further confirmed that pAKK alleviated L-NAME-induced NO reduction and endothelial dysfunction in human umbilical vein endothelial cells (HUVECs). Metabolomic analysis revealed that both pAKK and live AKK reversed metabolic disturbances in PE by modulating key metabolites and pathways related to unsaturated fatty acid biosynthesis, folate, and linoleic acid metabolism. As a postbiotic, pAKK may support existing treatments for preeclampsia by improving gut barrier function, restoring endothelial function, and regulating metabolic dysregulation, offering a safer alternative to live bacteria. These findings highlight the potential clinical value of pAKK as an adjunctive therapy in managing PE.