Pathological and pharmacological functions of the metabolites of polyunsaturated fatty acids mediated by cyclooxygenases, lipoxygenases, and cytochrome P450s in cancers.

Oxylipins have garnered increasing attention because they were consistently shown to play pathological and/or pharmacological roles in the development of multiple cancers. Oxylipins are the metabolites of polyunsaturated fatty acids via both enzymatic and nonenzymatic pathways. The enzymes mediating the metabolism of PUFAs include but not limited to lipoxygenases (LOXs), cyclooxygenases (COXs), and cytochrome P450s (CYPs) pathways, as well as the down-stream enzymes. Here, we systematically summarized the pleiotropic effects of oxylipins in different cancers through pathological and pharmacological aspects, with specific reference to the enzyme-mediated oxylipins. We discussed the specific roles of oxylipins on cancer onset, growth, invasion, and metastasis, as well as the expression changes in the associated metabolic enzymes and the associated underlying mechanisms. In addition, we also discussed the clinical application and potential of oxylipins and related metabolic enzymes as the targets for cancer prevention and treatment. We found the specific function of most oxylipins in cancers, especially the underlying mechanisms and clinic applications, deserves and needs further investigation. We believe that research on oxylipins will provide not only more therapeutic targets for various cancers but also dietary guidance for both cancer patients and healthy humans.

Prostaglandin pathway activation in the diatom Skeletonema marinoi under grazer pressure.

Prostaglandins (Pgs) are eicosanoid lipid mediators detected in all vertebrates, in some marine invertebrates, macroalgae and in diatoms, a class of eukaryotic microalgae composing the phytoplankton. The enzymes involved in the Pgs pathway were found to be differentially expressed in two strains of the diatom Skeletonema marinoi, named FE7 and FE60, already known to produce different levels of oxylipins, a class of secondary metabolites involved in the defence of diatoms against copepod predation, with FE7 being higher producer than FE60. In the present study we investigated the response of genes involved in the production of oxylipins and Pgs, evaluating their expression after the exposure to the copepod Temora stylifera. Our results highlighted a grazer feeding preference for FE60, the strain having low oxylipins content and reduced expression of Pgs enzymes, and an impact on the gene expression of the enzymes involved in oxylipins (i.e. lipoxygenase) and Pgs (i.e. cyclooxygenase) biosynthesis, especially in FE7. A time course evaluation of the gene expression over 24 h showed an upregulation of the essential enzyme in the Pgs pathway, the cyclooxygenase, in FE60 after 6 h of exposure to the grazer, differently from FE7 where no upregulation of gene expression in the presence of copepods was revealed. These results provide preliminary indications regarding the existence of a complex involvement of the Pgs pathway in the prey-predator interaction that requires further investigations.

Intrauterine devices influence prostaglandin secretion by equine uterus: in vitro and in vivo studies.

BACKGROUND: Intrauterine devices (IUD) are used in the veterinary practice as the non-pharmacological method of oestrus suppression in mares. When placed in the uterus, IUD create a physical contact with the endometrium that mimics the presence of an equine embryo. However, the mechanism of their action has not been fully elucidated. The objective of the present study was to examine the effect of mechanical stimulation of IUD on mare`s endometrium in both in vitro and in vivo study. For this purpose, we demonstrated the effect of IUD on prostaglandin (PG) F2α and PGE2 secretion, and mRNA transcription of genes involved in PG synthesis pathway in equine endometrial cells in vitro. In the in vivo study, we aimed to compare short-term effect of IUD inserted on day 0 (oestrus) with day 5-6 post-ovulation (the specific time when embryo reaches uterus after fertilization) on PG secretion from equine endometrium. To determine the long-term effect on PG synthase mRNA transcription, a single endometrial biopsy was taken only once within each group of mares at certain time points of the estrous cycle from mares placement with IUD on days 0 or 5-6 post-ovualtion. RESULTS: We showed for the first time that the incubation of the endometrial cells with the presence of IUD altered the pattern of PG synthase mRNA transcription in equine epithelial and stromal endometrial cells. In vivo, in mares placement with IUD on day 0, PGE2 concentrations in blood plasma were upregulated between 1 and 6, and at 10 h after the IUD insertion, compared with the control mares (P < 0.05). Moreover, the decrease of PTGFS mRNA transcription on day 16- 18, associated with an elevation in PTGES mRNA transcription on day 20 -21 of the estrous cycle in endometrial biopsies collected from mares placement with IUD on days 5-6 suggest an antiluteolytic action of IUD during the estrous cycle. CONCLUSION: We conclude that the application of IUD may mimic the equine conceptus presence through the physical contact with the endometrium altering PG synthase transcription, and act as a potent modulator of endometrial PG secretion both in vitro and in vivo.

Mycobacterium marinum mediates regulation of prostaglandin E2 expression on host immune response through cyclooxygenase pathway.

PURPOSE: Investigate the role of COX signaling in activating the PGE2-EP2 pathway. METHODS: Utilized a marine Mycobacterium infection model in zebrafish. Marine mycobacteria were stained with fluorescein isothiocyanate. The COX inhibitor indomethacin, EP2 receptor inhibitor AH6809, EP4 receptor inhibitor AH23848 and clodronate Liposomes were used to investigate the role of COX, EP2, EP4 and macrophage whether participating in combat marine mycobacterial infection. The expression level of the target gene was detected using real-time fluorescence quantitative PCR instrument. RESULTS: The findings revealed that larvae exposed to the COX inhibitor indomethacin or the EP2 receptor inhibitor AH6809 demonstrated a significantly higher mortality rate due to marine mycobacterium infection than those in the control group. Administration of exogenous prostaglandin E2 (PGE2) rescued the survival of zebrafish infected with marine mycobacteria and treated with indomethacin. Additionally, a significant reduction in survival rate was noted in macrophage-depleted zebrafish infected with marine mycobacteria. CONCLUSION: The host may combat marine mycobacterium infection via COX signaling, which activates the PGE2-EP2 pathway and mediates macrophage resistance.

The effect of aspirin and eicosapentaenoic acid on urinary biomarkers of prostaglandin E2 synthesis and platelet activation in participants of the seAFOod polyp prevention trial.

Urinary prostaglandin (PG) E metabolite (PGE-M) and 11-dehydro (d)-thromboxane (TX) B2 are biomarkers of cyclooxygenase-dependent prostanoid synthesis. We investigated (1) the effect of aspirin 300 mg daily and eicosapentaenoic acid (EPA) 2000 mg daily, alone and in combination, on urinary biomarker levels and, (2) whether urinary biomarker levels predicted colorectal polyp risk, during participation in the seAFOod polyp prevention trial. Urinary PGE-M and 11-d-TXB2 were measured by liquid chromatography-tandem mass spectrometry. The relationship between urinary biomarker levels and colorectal polyp outcomes was investigated using negative binomial (polyp number) and logistic (% with one or more polyps) regression models. Despite wide temporal variability in PGE-M and 11-d-TXB2 levels within individuals, both aspirin and, to a lesser extent, EPA decreased levels of both biomarkers (74% [P ≤ .001] and 8% [P ≤ .05] reduction in median 11-d-TXB2 values, respectively). In the placebo group, a high (quartile [Q] 2-4) baseline 11-d-TXB2 level predicted increased polyp number (incidence rate ratio [IRR] [95% CI] 2.26 [1.11,4.58]) and risk (odds ratio [95% CI] 3.56 [1.09,11.63]). A low (Q1) on-treatment 11-d-TXB2 level predicted reduced colorectal polyp number compared to placebo (IRR 0.34 [0.12,0.93] for combination aspirin and EPA treatment) compared to high on-treatment 11-d-TXB2 values (0.61 [0.34,1.11]). Aspirin and EPA both inhibit PGE-M and 11-d-TXB2 synthesis in keeping with shared in vivo cyclooxygenase inhibition. Colorectal polyp risk and treatment response prediction by 11-d-TXB2 is consistent with a role for platelet activation during early colorectal carcinogenesis. The use of urinary 11-d-TXB2 measurement for a precision approach to colorectal cancer risk prediction and chemoprevention requires prospective evaluation.

Prostaglandins as local regulators of ovarian physiology in ruminants.

Prostaglandins are synthesized from arachidonic acid through the catalytic activities of cyclooxygenase, while the production of different prostaglandin types, prostaglandin F2 alpha (PGF) and prostaglandin E2 (PGE), are regulated by specific prostaglandin synthases (PGFS and PGES). Prostaglandin ligands (PGF and PGE) bind to specific high-affinity receptors and initiate biologically distinct signalling pathways. In the ovaries, prostaglandins are known to be important endocrine regulators of female reproduction, in addition to maintaining local function through autocrine and/or paracrine effect. Many research groups in different animal species have already identified a variety of factors and molecular mechanisms that are responsible for the regulation of prostaglandin functions. In addition, prostaglandins stimulate their intrafollicular and intraluteal production via the pathway of prostaglandin self-regulation in the ovary. Therefore, the objective of the review article is to discuss recent findings about local regulation patterns of prostaglandin ligands PGF and PGE during different physiological stages of ovarian function in domestic ruminants, especially in bovine. In conclusion, the discussed local regulation mechanisms of prostaglandins in the ovary may stimulate further research activities in different methodological approaches, especially during final follicle maturation and ovulation, as well as corpus luteum formation and function.

The importance of cyclooxigenase in dentistry: a narrative review

Aim: Cyclooxygenase enzymes (COX) catalyze the conversion of arachidonic acid to prostaglandins and thromboxanes during pain and inflammation conditions. These enzymes have also been linked to several other conditions and diseases, and hence, in dentistry, it is crucial to identify the processes that increase the levels of these mediators. This paper aims to describe the significance of COX in dental practice through a narrative review. Methods: Articles relating to COX upregulation published in English and Spanish over the last 51 years in databases such as EBSCO, Google Scholar, Science Direct, PubMed, and Web of Science; were analyzed. Results: A total of 115 articles demonstrating the relationship between COX upregulation and multiple conditions and diseases of importance in prosthodontics, periodontics, oral pathology, orthodontics, and endodontics were included. Conclusions: COX upregulation is related to inflammatory and malignant diseases in oral tissues, such as periodontitis, pulpitis, and oral cancer, nevertheless, its expression is advantageous in other fields of study such as orthodontics. Additionally, is well documented that dental materials provoke an undesired increase in COX expression, which could be a significant factor that directly affects pulpal health

Exogenous oxygen is required for prostanoid induction under brain ischemia as evidence for a novel regulatory mechanism.

Previously, we and others reported a rapid and dramatic increase in brain prostanoids (PG), including prostaglandins, prostacyclins, and thromboxanes, under ischemia that is traditionally explained through the activation of esterified arachidonic acid (20:4n6) release by phospholipases as a substrate for cyclooxygenases (COX). However, the availability of another required COX substrate, oxygen, has not been considered in this mechanism. To address this mechanism for PG upregulation through oxygen availability, we analyzed mouse brain PG, free 20:4n6, and oxygen levels at different time points after ischemic onset using head-focused microwave irradiation (MW) to inactivate enzymes in situ before craniotomy. The oxygen half-life in the ischemic brain was 5.32 ± 0.45 s and dropped to undetectable levels within 12 s of ischemia onset, while there were no significant free 20:4n6 or PG changes at 30 s of ischemia. Furthermore, there was no significant PG increase at 2 and 10 min after ischemia onset compared to basal levels, while free 20:4n6 was increased ∼50 and ∼100 fold, respectively. However, PG increased ∼30-fold when ischemia was followed by craniotomy of nonMW tissue that provided oxygen for active enzymes. Moreover, craniotomy performed under anoxic conditions without MW did not result in PG induction, while exposure of these brains to atmospheric oxygen significantly induced PG. Our results indicate, for the first time, that oxygen availability is another important regulatory factor for PG production under ischemia. Further studies are required to investigate the physiological role of COX/PG regulation through tissue oxygen concentration.

Effect of digoxin, sodium valproate, and celecoxib on the cerebral cyclooxygenase pathway and neuron-specific enolase under the pentylenetetrazole-induced kindling in mice.

Neuroinflammation plays an important role in the pathogenesis of epilepsy, so it is necessary to clarify the influence of standard antiepileptic drugs as well as adjuvant agents (e.g., cardiac glycoside digoxin, which previously showed a clear anticonvulsant potential) on cyclooxygenase pathway and neuron-specific enolase under the conditions of chronic epileptogenesis. The aim of the article is to determine the effect of digoxin, sodium valproate, and celecoxib per se, as well as the combination of digoxin with sodium valproate on the content of cyclooxygenase 1 and 2 types, prostaglandins E2, F2α, I2, thromboxane B2, 8-isoprostane and neuron-specific enolase in the brain of mice in the pentylenetetrazole-induced kindling model. It was found that only the combination of sodium valproate with digoxin provides a complete protective effect (absence of seizures) and shows the clearest influence on neuroinflammation markers and neuronal damage than monotherapy with each of these drugs and celecoxib, which appeared to be an ineffective anticonvulsant. The obtained results indicate that digoxin is a promising adjuvant drug to classical antiepileptic drugs (mostly sodium valproate) in epilepsy treatment.c.

Stress-mediated aggregation of disease-associated proteins in amyloid bodies.

The formation of protein aggregates is a hallmark of many neurodegenerative diseases and systemic amyloidoses. These disorders are associated with the fibrillation of a variety of proteins/peptides, which ultimately leads to cell toxicity and tissue damage. Understanding how amyloid aggregation occurs and developing compounds that impair this process is a major challenge in the health science community. Here, we demonstrate that pathogenic proteins associated with Alzheimer's disease, diabetes, AL/AA amyloidosis, and amyotrophic lateral sclerosis can aggregate within stress-inducible physiological amyloid-based structures, termed amyloid bodies (A-bodies). Using a limited collection of small molecule inhibitors, we found that diclofenac could repress amyloid aggregation of the ß-amyloid (1-42) in a cellular setting, despite having no effect in the classic Thioflavin T (ThT) in vitro fibrillation assay. Mapping the mechanism of the diclofenac-mediated repression indicated that dysregulation of cyclooxygenases and the prostaglandin synthesis pathway was potentially responsible for this effect. Together, this work suggests that the A-body machinery may be linked to a subset of pathological amyloidosis, and highlights the utility of this model system in the identification of new small molecules that could treat these debilitating diseases.

Impaired vascular ß-adrenergic relaxation in spontaneously hypertensive rats: The differences between conduit and resistance arteries.

It was suggested that impaired ß-adrenergic relaxation in spontaneously hypertensive rats (SHR) might contribute to their high blood pressure (BP). Our study was focused on isoprenaline-induced dilatation of conduit femoral or resistance mesenteric arteries and on isoprenaline-induced BP reduction in SHR and Wistar-Kyoto rats (WKY). We confirmed decreased ß-adrenergic relaxation of SHR femoral arteries due to the absence of its endothelium-independent component, whereas endothelium-dependent component of ß-adrenergic smooth muscle relaxation was similar in both strains. Conversely, isoprenaline-induced relaxation of resistance mesenteric arteries was similar in both strains and this was true for endothelium-dependent and endothelium-independent components. We observed moderately reduced sensitivity of SHR mesenteric arteries to salmeterol (ß2-adrenergic agonist) and this strain difference disappeared after endothelium removal. However, there was no difference in mesenteric arteries relaxation by dobutamine (ß1-adrenergic agonist) which was independent of endothelium. The increasing isoprenaline doses elicited similar BP decrease in both rat strains, although BP sensitivity to isoprenaline was slightly decreased in SHR. The blockade of cyclooxygenase (indomethacin) and NO synthase (L-NAME) further reduced BP sensitivity to isoprenaline in SHR. On the other hand, salmeterol elicited similar BP decrease in both strains and the blockade of cyclooxygenase and NO synthase increased BP sensitivity to salmeterol in SHR as compared to WKY. In conclusion, attenuated ß-adrenergic vasodilatation of conduit arteries of SHR but similar ß-adrenergic relaxation of resistance mesenteric arteries from WKY and SHR and their similar BP response to ß-adrenergic agonists do not support major role of altered ß-adrenergic vasodilatation for high BP in genetic hypertension.

HPLC and GC Characterization of Dicliptera bupleuroides Aerial Parts and Evaluation of Its Anti-Inflammatory Potential in Vitro, in silico and in Vivo Using Carrageenan and Formalin Induced Inflammation in Rat Models.

The current study aimed to evaluate the anti-inflammatory activity of Dicliptera bupleuroides Nees aerial parts methanol extract and its different fractions namely hexane, chloroform, ethyl acetate and butanol in vitro using cyclooxygenase inhibitory assay (COX-2). In vivo anti-inflammatory evaluation was performed using carrageenan and formalin induced inflammation in rat models followed by molecular docking. High performance liquid chromatography (HPLC) and gas chromatography coupled with mass chromatography (GC/MS) analyses were used for chemical analyses of the tested samples. The tested samples showed significant inhibition in COX-2 inhibitory assay where methanol extract (DBM) showed the highest inhibitory potential at 100 µg/mL estimated by 67.86 %. At a dose of 400 mg/kg, all of the examined samples showed pronounced results in carrageenan induced acute inflammation in rat model at 4th h interval with DBM showed the highest efficiency displaying 65.32 % inhibition as compared to the untreated rats. Formalin model was employed for seven days and DBM exhibited 65.33 % and 69.39 % inhibition at 200 and 400 mg/kg, respectively approaching that of the standard on the 7th day. HPLC revealed the presence of caffeic acid, gallic acid and sinapic acid, quercetin and myricetin in DBM. GC/MS analysis of its hexane fraction revealed the presence of 16 compounds belonging mainly to fatty acids and sterols that account for 85.26 % of the total detected compounds. Molecular docking showed that hexadecanoic acid followed by decanedioic acid and isopropyl myristate showed the best fitting within cyclooxygenase-II (COX-II) while nonacosane followed by hexatriacontane and isopropyl myristate revealed the most pronounced fitting within the 5-lipoxygenase (5-LOX) active sites. Absorption, metabolism, distribution and excretion and toxicity prediction (ADMET/ TOPKAT) concluded that most of the detected compounds showed reasonable pharmacokinetic, pharmacodynamic and toxicity properties that could be further modified to be more suitable for incorporation in pharmaceutical dosage forms combating inflammation and its undesirable consequences.

Production of glycerolamines based conjugated γ-aminobutyric acids using microbial COX and LOX as successor enzymes to GAD.

Gamma-aminobutyric acid (γ-ABA) can be produced by various microorganisms including bacteria, fungi and yeasts using enzymatic bioconversion, microbial fermentation or chemical hydrolysis. Regenerating conjugated glycerol-amines is valid by the intervention of microbial cyclooxygenase [COX] and lipooxygenase [LOX] enzymes produced via lactobacillus bacteria (LAB) as successor enzymes to glutamate decarboxylases (GAD). Therefore, the aim of this review is to provide an overview on γ-ABA production, and microbiological achievements used in producing this signal molecule based on those fermenting enzymes. The formation of aminoglycerides based conjugated γ-ABA is considered the key substances in controlling the host defense against pathogens and is aimed in increasing the neurotransmission effects and in suppressing further cardiovascular diseases.

Experimental validation for mechanisms of Qizhiweitong particles against Chronic Non-atrophic gastritis based on metabolomics and network pharmacology.

Qizhiweitong particles (QZWT), a classic Chinese herbal prescription derived from the Sinisan decoction in Shang Han Za Bing Lun, has definitive clinical efficacy in treating Chronic Non-atrophic Gastritis (CNG) in China. However, its mechanism of action at the metabolic level remains unclear. The aim of this study was to explore the mechanisms of QZWT against CNG based on non-targeted metabolomics combined with network pharmacology and experimentally validated by enzyme linked immunosorbent assays (ELISA). First, CNG model rats were established by free drinking ammonia water combined with starvation and satiety disorder for 12 weeks. Taking gastric tissue as the object, ultra-high performance liquid chromatography tandem mass spectrometry based metabolomics and network pharmacology were conducted to identify the key compounds, core targets and pathways that mediate the effects of QZWT against CNG. Furthermore, the targets from network pharmacology and the metabolites from metabolomics were jointly analyzed to select crucial metabolism pathways by MetaScape. Finally, the key metabolic enzymes and metabolites were experimentally validated by ELISA. The results indicated that there were 29 differential metabolites were identified and considered to be metabolic biomarkers of QZWT in the treatment of CNG. Among them, 8 of the differential metabolites showed a significant reduction in the content of QZWT groups. Arachidonic acid (AA) metabolic and glycerophospholipid (GP) metabolic are the most crucial metabolic pathways for QZWT to treat CNG. QZWT regulated AA and GP metabolism by synergetic reducing the level of AA, Phospholipid acid and Lysophosphatidic acid and inhibiting the enzyme activity of prostaglandin endoperoxide synthase 1 and prostaglandin endoperoxide synthase 2. And a compound-reaction-enzyme-gene network of mechanism for QZWT against CNG was established. In conclusion, this study reveals the complicated mechanisms of QZWT against CNG. Our work presents a novel strategy to identify the potential mechanisms of pharmacological effects derived from a compound prescription of TCM.

A physiological concentration of anandamide promotes the migration of human endometrial fibroblast and the interaction with endothelial cells invitro.

INTRODUCTION: The mechanisms that govern fibroblast behavior during the vascular adaptations of the uterus at early pregnancy remain unknown. Anandamide, an endocannabinoid, binds to cannabinoid receptors (CBs), and regulates gestation and angiogenesis. Its tone is regulated by fatty acid amide hydrolase (FAAH) within the uterus. We investigated the role of anandamide in endometrial fibroblasts migration and whether anandamide modulates fibroblasts-endothelial crosstalk. METHODS: T-hESC and EA.hy926 cell lines were used as models of endometrial stromal and endothelial cells, respectively. T-hESC were incubated with anandamide plus different agents. Migration was tested (wound healing assay and phalloidin staining). Protein expression and localization were studied by Western blot and immunofluorescence. To test fibroblast-endothelial crosstalk, EA.hy926 cells were incubated with fibroblast conditioned media obtained after T-hESC migration. RESULTS: Anandamide 1 nM increased T-hESC migration via CB1 and CB2. Cyclooxygenase-2 participated in anandamide-stimulated fibroblast migration. Prostaglandin F2alpha, and not prostaglandin E2, increased fibroblast wound closure. CB1, CB2, cyclooxygenase-2 and FAAH were expressed in T-hESC. Anandamide did not alter cyclooxygenase-2 localization but induced its cytoplasmic and nuclear expression through CB1 and CB2. URB-597, a FAAH selective inhibitor, also increased T-hESC migration via both CBs, and augmented cyclooxygenase-2 expression. Conditioned media from anandamide-induced T-hESC wound healing closure stimulated endothelial migration and did not alter their proliferation. Soluble factors from cyclooxygenase-2 were secreted by T-hESC and participated in T-hESC-induced EA.hy926 migration. Although anandamide-conditioned media augmented in EA.hy926 the expression of γH2AX, a marker of DNA damage, cyclooxygenase-2 was not involved in this effect. DISCUSSION: Our results provide novel evidence about an active role of anandamide on endometrial fibroblast behavior as a mechanism regulating uterine vascular adaptations in early gestation.

Multi-action platinum(IV) prodrugs conjugated with COX-inhibiting NSAIDs.

In the last decades, inflammation has been recognized as being closely connected to cancer, and joint strategies encompassing chemotherapeutic and anti-inflammatory agents have been extensively studied. In this work, a series of novel cisplatin and oxaliplatin-based Pt(IV) complexes comprising non-steroidal anti-inflammatory drugs (NSAIDs) and their carboxyl ester analogues as axial moieties were synthesized. Several of the cisplatin-based Pt(IV) complexes 22-30 showed increased cytotoxicity in the human cancer cell lines CH1/PA-1, SW480 and A549 compared to the Pt(II) drug. For the most potent complex 26, comprising two aceclofenac (AFC) moieties, the formation of Pt(II)-9-methylguanine (9-MeG) adducts after activation with ascorbic acid (AsA) was proven. Additionally, a significant inhibition of cyclooxygenase (COX) activity and prostaglandin E2 (PGE2) production was observed, as well as increased cellular accumulation, depolarization of mitochondrial membranes, and strong proapoptotic potencies in SW480 cells. Overall, these systematic effects shown in vitro confer 26 as a potential anticancer agent combined with anti-inflammatory properties.

Association between serum prostacyclin and cerebrovascular reactivity in healthy young and older adults.

NEW FINDINGS: What is the central question of this study? What is the relationship between prostacyclin and cerebrovascular reactivity to hypercapnia before and after administration of a cyclooxygenase inhibitor, indomethacin, in healthy young and older adults? What is the main finding and importance? Serum prostacyclin was not related to cerebrovascular reactivity to hypercapnia before or after administration of indomethacin. However, in older adults, serum prostacyclin was related to the magnitude of change in cerebrovascular reactivity from before to after indomethacin administration. This suggests that older adults with higher serum prostacyclin may rely more on cyclooxygenase products to mediate cerebrovascular reactivity. ABSTRACT: Platelet activation may contribute to age-related cerebrovascular dysfunction by interacting with the endothelial cells that regulate the response to vasodilatory stimuli. This study evaluated the relationship between a platelet inhibitor, prostacyclin, and cerebrovascular reactivity (CVR) in healthy young (n = 35; 25 ± 4 years; 17 women, 18 men) and older (n = 12; 62 ± 2 years; 8 women, 4 men) adults, who were not daily aspirin users, before and after cyclooxygenase inhibition. Prostacyclin was determined by levels of 6-keto-prostaglandin F1α (6-keto PGF1α) in the blood. CVR was assessed by measuring the middle cerebral artery blood velocity response to hypercapnia using transcranial Doppler ultrasound before (CON) and 90 min after cyclooxygenase inhibition with indomethacin (INDO). In young adults, there were no associations between prostacyclin and middle cerebral artery CVR during CON (r = -0.14, P = 0.415) or INDO (r = 0.27, P = 0.118). In older adults, associations between prostacyclin and middle cerebral artery CVR during CON (r = 0.53, P = 0.075) or INDO (r = -0.45, P = 0.136) did not reach the threshold for significance. We also evaluated the relationship between prostacyclin and the change in CVR between conditions (ΔCVR). We found no association between ΔCVR and prostacyclin in young adults (r = 0.27, P = 0.110); however, in older adults, those with higher baseline prostacyclin levels demonstrated significantly greater ΔCVR (r = -0.74, P = 0.005). In conclusion, older adults with higher serum prostacyclin, a platelet inhibitor, may rely more on cyclooxygenase products for cerebrovascular reactivity to hypercapnia.

Pulegone Prevents Hypertension through Activation of Muscarinic Receptors and Cyclooxygenase Pathway in L-NAME-Induced Hypertensive Rats.

The current study was designed to determine pulegone's antihypertensive and vasoprotective activity in L-NAME-induced hypertensive rats. Firstly, the hypotensive dose-response relationship of pulegone was evaluated in normotensive anesthetized rats using the invasive method. Secondly, the mechanism involved in hypotensive activity was determined in the presence of pharmacological drugs such as atropine/muscarinic receptor blocker (1 mg/kg), L-NAME/NOS inhibitor (20 mg/kg), and indomethacin/COX inhibitor (5 mg/kg) in anesthetized rats. Furthermore, studies were carried out to assess the preventive effect of pulegone in L-NAME-induced hypertensive rats. Hypertension was induced in rats by administering L-NAME (40 mg/kg) orally for 28 days. Rats were divided into six groups which were treated orally with tween 80 (placebo), captopril (10 mg/kg), and different doses of pulegone (20 mg/kg, 40 mg/kg, and 80 mg/kg). Blood pressure, urine volume, sodium, and body weight were monitored weekly. After 28 days, the effect of pulegone on lipid profile, hepatic markers, antioxidant enzymes, and nitric oxide was estimated from the serum of treated rats. Moreover, plasma mRNA expression of eNOS, ACE, ICAM1, and EDN1 was measured using real-time PCR. Results show that pulegone dose-dependently decreased blood pressure and heart rate in normotensive rats, with the highest effect at 30 mg/kg/i.v. The hypotensive effect of pulegone was reduced in the presence of atropine and indomethacin, whereas L-NAME did not change its hypotensive effect. Concurrent treatment with pulegone for four weeks in L-NAME-treated rats caused a reduction in both systolic blood pressure and heart rate, reversed the reduced levels of serum nitric oxide (NO), and ameliorated lipid profile and oxidative stress markers. Treatment with pulegone also improved the vascular response to acetylcholine. Plasma mRNA expression of eNOS was reduced, whereas ACE, ICAM1, and EDN1 levels were high in the L-NAME group, which was facilitated by pulegone treatment. To conclude, pulegone prevented L-NAME-induced hypertension by demonstrating a hypotensive effect through muscarinic receptors and cyclooxygenase pathway, indicating its use as a potential candidate in managing hypertension.

miR-665 overexpression inhibits the apoptosis of luteal cells in small ruminants suppressing HPGDS.

Evidence has shown that microRNA-665 (miR-665) is highly expressed in the mid-luteal phase compared with the early and end-luteal phase of the corpus luteum (CL) life cycle. However, whether miR-665 is a positive regulator of the life span of the CL is still unknown. The objective of this study is to explore the effect of miR-665 on the structural luteolysis in the ovarian CL. In this study, the targeting relationship between miR-665 and hematopoietic prostaglandin synthase (HPGDS) was firstly verified by dual luciferase reporter assay. Then, quantitative real-time PCR (qRT-PCR) was used to detect the expression of miR-665 and HPGDS in luteal cells. Following miR-665 overexpression, the apoptosis rate of the luteal cells was determined using flow cytometry; B-cell lymphoma-2 (BCL-2) and caspase-3 mRNA and protein were measured using qRT-PCR and Western blot (WB) analysis. Finally, the DP1 and CRTH2 receptors of PGD2, a synthetic product of HPGDS, were localized using immunofluorescence. Results confirmed that HPGDS was a direct target gene of miR-665, and miR-665 expression was negatively correlated with HPGDS mRNA expression in luteal cells. Meanwhile, after miR-665 was overexpressed, the apoptotic rate of the luteal cells showed a significant decrease (P < 0.05) and this was accompanied by elevated expression levels of anti-apoptotic factor BCL-2 mRNA and protein and decreased expression levels of apoptotic factor caspase-3 mRNA and protein (P < 0.01). Moreover, the immune fluorescence staining results showed that the DP1 receptor was also significantly decreased (P < 0.05), but the CRTH2 receptor was significantly increased (P < 0.05) in luteal cells. Overall, these results indicate that miR-665 reduces the apoptosis of luteal cells via inhibiting caspase-3 expression and promoting BCL-2 expression, and the biological function of miR-665 may be attributed to its target gene HPGDS which regulates the balance of DP1 and CRTH2 receptors expression in luteal cells. As a consequence, this study suggests that miR-665 might be a positive regulator of the life span of the CL rather than destroy the integrity of CL in small ruminants.