Nitric oxide-generating system as an autocrine mechanism in human polymorphonuclear leucocytes.
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ABSTRACT:
SUBMITTER: Riesco A
PROVIDER: S-EPMC1134183 | biostudies-other | 1993 Jun
REPOSITORIES: biostudies-other
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Project description:The present study was aimed at determining the metabolic profile of PMNs in obese subjects, and to explore its potential relationship with insulin resistance (IR). To achieve this goal, a pilot clinical study was performed using PMNs from 17 patients with obesity and IR, and 17 lean controls without IR, which was validated in an additional smaller cohort (consisting of 10 patients and 10 controls). PMNs were isolated from peripheral blood and nuclear magnetic resonance was used to perform the metabolomic analysis. A total of 48 metabolites were quantified. The main metabolic change found in PMNs was a significant increase in 2-aminoisobutyric acid with a direct correlation with HOMA-IR (p<0.001), BMI (p<0.000001) and waist circumference (p<0.000001). By contrast, a decrease of 3-hydroxyisovalerate was observed with an inverse correlation with HOMA-IR (p = 0.001), BMI (p = 0.001) and waist circumference (p = 0.0001). Notably, the metabolic profile in plasma was different than that obtained in PMNs. In summary, our results suggest that the change in 3-hydroxyisovalerate and 2-aminoisobutyric is the key metabolic fingerprint in PMNs of obese subjects with IR. In addition, our methodology could be an easy and reliable tool for monitoring the effect of treatments in the setting of precision medicine.
| S-EPMC6044522 | biostudies-literature
Project description:ObjectivesPolymorphonuclear leucocytes (PMNs) constitute the first line of host defence and are crucial in maintaining periodontal health. Their survival and function are modulated by mesenchymal stromal cells (MSCs) from different origin. Gingival MSCs (GMSCs) play an important role in maintaining oral health and in the initial inflammatory response. The present study aimed to investigate the effects of GMSCs on PMNs apoptosis and reactive oxygen species (ROS) production.MethodsPMNs were either directly incubated with untreated, interleukin (IL)-1β- or tumour necrosis factor (TNF)-α-treated GMSCs or stimulated with their conditioned media. Resulting ROS production was evaluated by dichlorofluorescin diacetate staining, whereas PMNs apoptosis was assessed by Annexin V staining, followed by flow cytometry analysis.ResultsWhile conditioned media of untreated and TNF-α-treated GMSCs did not affect apoptosis of PMNs, it was significantly delayed by conditioned media of GMSCs treated with IL-1β. In direct co-culture, GMSCs exerted anti-apoptotic effects on PMNs independently of the previous stimulation. However, the strongest impact was observed by IL-1β-treated GMSCs. ROS production of PMNs was not influenced by GMSCs or their conditioned media.ConclusionThis study demonstrates for the first time the immunomodulatory properties of GMSCs towards PMNs, revealing that IL-1β enhances anti-apoptotic effects of GMSCs.
| S-EPMC9290793 | biostudies-literature
Project description:Symptomatic infection with Neisseria gonorrhoeae (Gc) is characterized by abundant neutrophil (PMN, polymorphonuclear leucocyte) influx, but PMNs cannot clear initial infection, indicating that Gc possess defences against PMN challenge. In this study, survival of liquid-grown Gc was monitored after synchronous infection of adherent, interleukin 8-treated human PMNs. 40-70% of FA1090 Gc survived 1 h of PMN exposure, after which bacterial numbers increased. Assays with bacterial viability dyes along with soybean lectin to detect extracellular Gc revealed that a subset of both intracellular and extracellular PMN-associated Gc were viable. Gc survival was unaffected in PMNs chemically or genetically deficient for producing reactive oxygen species (ROS). This result held true even for OpaB+ Gc, which stimulate neutrophil ROS production. Catalase- and RecA-deficient Gc, which are more sensitive to ROS in vitro, had no PMN survival defect. recN and ngo1686 mutant Gc also exhibit increased sensitivity to ROS and PMNs, but survival of these mutants was not rescued in ROS-deficient cells. The ngo1686 mutant showed increased sensitivity to extracellular but not intracellular PMN killing. We conclude that Gc are remarkably resistant to PMN killing, killing occurs independently of neutrophil ROS production and Ngo1686 and RecN defend Gc from non-oxidative PMN antimicrobial factors.
| S-EPMC2771623 | biostudies-literature
Project description:Nitric oxide (NO), a gaseous transmitter extensively present in the human body, regulates vascular relaxation, immune response, inflammation, neurotransmission, and other crucial functions. Nitrite donors have been used clinically to treat angina, heart failure, pulmonary hypertension, and erectile dysfunction. Based on NO's vast biological functions, it further can treat tumors, bacteria/biofilms and other infections, wound healing, eye diseases, and osteoporosis. However, delivering NO is challenging due to uncontrolled blood circulation release and a half-life of under five seconds. With advanced biotechnology and the development of nanomedicine, NO donors packaged with multifunctional nanocarriers by physically embedding or chemically conjugating have been reported to show improved therapeutic efficacy and reduced side effects. Herein, we review and discuss recent applications of NO nanomedicines, their therapeutic mechanisms, and the challenges of NO nanomedicines for future scientific studies and clinical applications. As NO enables the inhibition of the replication of DNA and RNA in infectious microbes, including COVID-19 coronaviruses and malaria parasites, we highlight the potential of NO nanomedicines for antipandemic efforts. This review aims to provide deep insights and practical hints into design strategies and applications of NO nanomedicines.
| S-EPMC10395262 | biostudies-literature
Project description:Adenosine production in intact rat polymorphonuclear leucocytes was studied during 2-deoxyglucose-induced ATP catabolism. A cell-free system containing the cytosolic 5'-nucleotidase (EC 3.1.3.5) as the only phosphohydrolase was also studied. The rate of adenosine formation in both intact cells and the cell-free system showed a similar dependence on energy charge (([ATP] + 1/2 [ADP]/([ATP] + [ADP] + [AMP])), being maximal only at values close to 0.8. Sufficient cytosolic 5'-nucleotidase was present in intact cells to explain the observed rate of adenosine formation. We conclude that the cytosolic 5'-nucleotidase is responsible for adenosine production in rat polymorphonuclear leucocytes. This mechanism provides a direct biochemical link between the energy status of a cell and the rate of adenosine formation.
| S-EPMC1152251 | biostudies-other
Project description:We have previously observed that the phagocytosis of zymosan particles coated with complement by human polymorphonuclear leucocytes is accompanied by a time- and dose-dependent inhibition of phosphatidylcholine synthesis by transmethylation [García Gil, Alonso, Sánchez Crespo & Mato (1981) Biochem. Biophys. Res. Commun.101, 740-748]. The present studies show that phosphatidylcholine synthesis by a cholinephosphotransferase reaction is enhanced, up to 3-fold, during phagocytosis by polymorphonuclear cells. This effect was tested by both measuring the incorporation of radioactivity into phosphatidylcholine in cells labelled with [Me-(14)C]choline, and by assaying the activity of CDP-choline:diacylglycerol cholinephosphotransferase. The time course of CDP-choline:diacylglycerol cholinephosphotransferase activation by zymosan mirrors the inhibition of phospholipid methyltransferase activity previously reported. The extent of incorporation of radioactivity into phosphatidylcholine induced by various doses of zymosan correlates with the physiological response of the cells to this stimulus. This effect was specific for phosphatidylcholine, and phosphatidyl-ethanolamine turnover was not affected by zymosan. The purpose of this enhanced phosphatidylcholine synthesis is not to provide phospholipid molecules rich in arachidonic acid. The present studies show that about 80% of the arachidonic acid generated in response to zymosan derives from phosphatidylinositol. A transient accumulation of arachidonoyldiacylglycerol has also been observed, which indicates that a phospholipase C is responsible, at least in part, for the generation of arachidonic acid. Finally, isobutylmethylxanthine and quinacrine, inhibitors of phosphatidylinositol turnover, inhibit both arachidonic acid generation and phagocytosis, indicating a function for this pathway during this process.
| S-EPMC1158550 | biostudies-other
Project description:The subcellular distribution of nine glycosidases were studied in fractions of homogenized human polymorphonuclear leucocytes (neutrophils) obtained by isopycnic centrifugation through linear sucrose density gradients. The substrates were 4-methylumbelliferyl glycosides. All nine glycosides were hydrolysed by enzymes in neutrophil cytosol fractions, and by enzymes in at least one granule population. alpha-Glucosidase activity sedimented in sucrose density gradients to a point (p = 1.180 g/ml) just above the specific granules, possibly the 'tertiary' granule population. The peak corresponding to alpha-glucosidase did not co-sediment with, but considerably overlapped, the peak corresponding to lactoferrin, a marker for specific granules (p = 1.187 g/ml). alpha-Galactosidase activity was found primarily in heavy azurophil granules (p = 1.222 g/ml). alpha-Mannosidase activity was found primarily in light azurophil granules (p = 1.206 g/ml), following the distribution of myeloperoxidase, the commonly used azurophil granule marker. beta-Glucosidase activity was concentrated in mitochondrial fractions (p = 1.160 g/ml). All other glycosidases presented complex distributions, with activities not restricted to one granule class. Granule-associated glycosidase activities were increased 2--38 times when measured in the presence of 0.05% Triton X-100, indicating latency of the enzymes within granules.
| S-EPMC1185884 | biostudies-other
Project description:PurposeNitric oxide (NO) is a free radical synthesized mainly by nitric oxide synthases (NOSs). NO regulates many aspects in sperm physiology in different species. However, in vitro studies investigating NOS distribution, and how NO influences sperm capacitation and fertilization (IVF) in porcine, have been lacking. Therefore, our study aimed to clarify these aspects.MethodsTwo main experiments were conducted: (i) boar spermatozoa were capacitated in the presence/absence of S-nitrosoglutathione (GSNO), a NO donor, and two NOS inhibitors, NG-nitro-L-arginine methyl ester hydrochloride (L-NAME) and aminoguanidine hemisulfate salt (AG), and (ii) IVF was performed in the presence or not of these supplements, but neither the oocytes nor the sperm were previously incubated in the supplemented media.ResultsOur results suggest that NOS distribution could be connected to pathways which lead to capacitation. Treatments showed significant differences after 30 min of incubation, compared to time zero in almost all motility parameters (P < 0.05). When NOSs were inhibited, three protein kinase A (PKA) substrates (~ 75, ~ 55, and ~50 kDa) showed lower phosphorylation levels between treatments (P < 0.05). No differences were observed in total tyrosine phosphorylation levels evaluated by Western blotting nor in situ. The percentage of acrosome-reacted sperm and phosphatidylserine translocation was significantly lower with L-NAME. Both inhibitors reduced sperm intracellular calcium concentration and IVF parameters, but L-NAME impaired sperm ability to penetrate denuded oocytes.ConclusionsThese findings point out to the importance of both sperm and cumulus-oocyte-derived NO in the IVF outcome in porcine.
| S-EPMC6707978 | biostudies-literature
Project description:About 30% of patients with type 2 diabetes mellitus develop clinically overt nephropathy. Hyperglycemia is necessary, but not sufficient, to cause the renal damage that leads to kidney failure. Diabetic nephropathy (DN) is a multifactorial disorder that results from interaction between environmental and genetic factors. In the present article we will review the role of the nitric oxide synthase (NOS) in the pathogenesis of DN.Nitric oxide (NO) is a short-lived gaseous lipophilic molecule produced in almost all tissues, and it has three distinct genes that encode three NOS isoforms: neuronal (nNOS), inducible (iNOS) and endothelial (eNOS).The correct function of the endothelium depends on NO, participating in hemostasis control, vascular tone regulation, proliferation of vascular smooth muscle cells and blood pressure homeostasis, among other features. In the kidney, NO plays many different roles, including control of renal and glomerular hemodynamics. The net effect of NO in the kidney is to promote natriuresis and diuresis, along with renal adaptation to dietary salt intake.The eNOS gene has been considered a potential candidate gene for DN susceptibility. Three polymorphisms have been extensively researched: G894T missense mutation (rs1799983), a 27-bp repeat in intron 4, and the T786C single nucleotide polymorphism (SNP) in the promoter (rs2070744). However, the potential link between eNOS gene variants and the induction and progression of DN yielded contradictory results in the literature.In conclusion, NOS seems to be involve in the development and progression of DN. Despite the discrepant results of many studies, the eNOS gene is also a good candidate gene for DN.
| S-EPMC3928920 | biostudies-literature
Project description:We showed previously that rat cytochrome P450 CYP2B1 undergoes NO-dependent proteasomal degradation in response to inflammatory stimuli, and that the related human enzyme CYP2B6 is also down-regulated by NO in primary human hepatocytes. To investigate the mechanism of CYP2B6 down-regulation, we made several cell lines (HeLa and HuH7 cells) in which native CYP2B6 or CYP2B6 with a C-terminal V5 tag (CYP2B6V5) are expressed from a lentiviral vector with a cytomegalovirus promoter. Native CYP2B6 protein was rapidly down-regulated in HeLa cells within 3h of treatment with the NO donor (Z)-1-[2-(2-Aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate, while its mRNA level was not down-regulated. Treatment of the cells with the NO donor (Z)-1-[N-(3-aminopropyl)-N-(3-ammoniopropyl)amino]diazen-1-ium-1,2-diolate also resulted in rapid down-regulation of CYP2B6 activity, measured as the formation of 7-hydroxy-4-trifluoromethylcoumarin, as well as 2B6 protein in the CYP2B6 HeLa cell line. CYP2B6V5 was also down-regulated by NO donors in HuH7 cells. Down-regulation was observed in the presence of cycloheximide, demonstrating that this occurs via a post-translational mechanism. We generated a HeLa cell line expressing both CYP2B6V5 and human nitric oxide synthase 2 (NOS2), the latter under positive control by tetracycline. The cellular NO produced by doxycycline treatment also effectively down-regulated CYP2B6 protein, which was blocked by the co-treatment with the NOS2 competitive inhibitor L-NG-nitroarginine methyl ester (L-NAME). We next investigated the proteolytic enzymes responsible for NO-dependent CYP2B6 degradation. Neither calpain inhibitors (N-Acetyl-L-leucyl-L-leucyl-L-norleucinal, carbobenzoxy-valinyl-phenylalaninal), nor lysosomal protease inhibitors (3-methyladenine and chloroquine) inhibited the NO dependent CYP2B6V5 down-regulation. The proteasome inhibitors MG132 and bortezomib attenuated, but did not completely block the NO-induced down-regulation in the HuH7 cell line. However, when cells were co-treated with NO donor and proteasome inhibitors, high molecular mass species could be detected on native CYP2B6 as well as CYP2B6V5 Western blots. Further investigation demonstrated that CYP2B6 protein was polyubiquitinated and this was dramatically enhanced by co-treatment with NO donor and bortezomib. Taken together, our data demonstrate that CYP2B6 is down-regulated in an NO-dependent manner via ubiquitination and proteasomal degradation.
| S-EPMC5507215 | biostudies-literature