8-11-14-eicosatrienoic-acid and Disease-Models--Animal

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Antihypertensive Agents; Disease Models, Animal; Disease Progression; Endothelium, Vascular; Fatty Acids; Humans; Hypertension; Kidney Diseases; Mice; Rats

To investigate the effect of 14,15-EET on the parthanatos in neurons induced by cerebral ischemia and reperfusion, middle cerebral artery occlusion and reperfusion (MCAO/R) and oxygen glucose deprivation/reoxygenation (OGD/R) were used to simulate cerebral ischemia reperfusion in vivo and in vitro, respectively. TTC staining and the Tunel method were used to detect cerebral infarct volume and neuronal apoptosis. Western blot and immunofluorescence were used to detect poly (ADP-ribose) polymerase-1 (PARP-1) activation and AIF nuclear translocation. The production of reactive oxygen species (ROS) and the expression of antioxidant genes were detected by Mito SOX, DCFH-DA and qPCR methods. MCAO/R increased cerebral infarct volume and neuronal apoptosis in mice, while 14,15-EET pretreatment increased cerebral infarct volume and neuronal apoptosis. OGD/R induced reactive oxygen species generation, PARP-1 cleavage, and AIF nuclear translocation in cortical neurons. 14,15-EET pretreatment could enhance the antioxidant gene expression of glutathione peroxidase (GSH-Px), heme oxygenase-1 (HO-1) and superoxide dismutase (SOD) in cortical neurons after ischemia and reperfusion. 14,15-EET inhibits the neuronal parthanatos induced by MCAO/R through upregulation of the expression of antioxidant genes and by reducing the generation of reactive oxygen species. This study advances the EET neuroprotection theory and provides a scientific basis for targeted clinical drugs that reduce neuronal parthanatos following cerebral ischemia and reperfusion.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Brain Injuries; Brain Ischemia; Disease Models, Animal; Glucose; Male; Mice; Models, Biological; Neurons; Neuroprotective Agents; Oxidative Stress; Parthanatos; Reactive Oxygen Species; Reperfusion Injury

Cytochrome P450 epoxygenase 2J2 (CYP2J2) metabolizes arachidonic acid to epoxyeicosatrienoic acids (EETs), which exert anti-inflammatory, anti-apoptotic, pro-proliferative, and antioxidant effects on the cardiovascular system. However, the role of CYP2J2 and EETs in pulmonary arterial hypertension (PAH) with lung ischemia-reperfusion injury (LIRI) remains unclear. In the present study, we investigated the effects of CYP2J2 overexpression and exogenous EETs on PAH with LIRI in vitro and in vivo.. CYP2J2 gene was transfected into rat lung tissue by recombinant adeno-associated virus (rAAV) to increase the levels of EETs in serum and lung tissue. A rat model of PAH with LIRI was constructed by intraperitoneal injection of monocrotaline (50 mg/kg) for 4 weeks, followed by clamping of the left pulmonary hilum for 1 h and reperfusion for 2 h. In addition, we established a cellular model of human pulmonary artery endothelial cells (HPAECs) with TNF-α combined with anoxia/reoxygenation (anoxia for 8 h and reoxygenation for 16 h) to determine the effect and mechanism of exogenous EETs.. CYP2J2 overexpression significantly reduced the inflammatory response, oxidative stress and apoptosis associated with lung injury in PAH with LIRI. In addition, exogenous EETs suppressed inflammatory response and reduced intracellular reactive oxygen species (ROS) production and inhibited apoptosis in a tumor necrosis factor alpha (TNF-α) combined hypoxia-reoxygenation model of HPAECs. Our further studies revealed that the anti-inflammatory effects of CYP2J2 overexpression and EETs might be mediated by the activation of PPARγ; the anti-apoptotic effects might be mediated by the PI3K/AKT pathway.. CYP2J2 overexpression and EETs protect against PAH with LIRI via anti-inflammation, anti-oxidative stress and anti-apoptosis, suggesting that increased levels of EETs may be a promising strategy for the prevention and treatment of PAH with LIRI.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Cells, Cultured; Cytochrome P-450 CYP2J2; Disease Models, Animal; Gene Expression Regulation; Humans; Hypertension, Pulmonary; Male; Rats; Reperfusion Injury; RNA

Epoxyeicosatrienoic acids (EETs) are metabolites of arachidonic acid that are rapidly metabolized into diols by soluble epoxide hydrolase (sEH). sEH inhibition has been shown to increase the biological activity of EETs, which are known to have anti-inflammatory properties. However, the role of EETs in pulmonary fibrosis remains unexplored. Liquid chromatography with tandem mass spectrometry (LC-MS/MS) was used to analyze EETs in the lung tissues of patients with idiopathic pulmonary fibrosis (IPF, n = 29) and controls (n = 15), and the function of 11,12-EET was evaluated in in vitro and in vivo in pulmonary fibrosis models. EET levels in IPF lung tissues, including those of 8,9-EET, 11,12-EET, and 14,15-EET, were significantly lower than those in control tissues. The 11,12-EET/11,12-DHET ratio in human lung tissues also differentiated IPF from control tissues. 11,12-EET significantly decreased transforming growth factor (TGF)-β1-induced expression of α-smooth muscle actin (SMA) and collagen type-I in MRC-5 cells and primary fibroblasts from IPF patients. sEH-specific siRNA and 1-trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea (TPPU; sEH inhibitor) also decreased TGF-β1-induced expression of α-SMA and collagen type-I in fibroblasts. Moreover, 11,12-EET and TPPU decreased TGF-β1-induced p-Smad2/3 and extracellular-signal-regulated kinase (ERK) expression in primary fibroblasts from patients with IPF and fibronectin expression in Beas-2B cells. TPPU decreased the levels of hydroxyproline in the lungs of bleomycin-induced mice. 11,12-EET or sEH inhibitors could inhibit pulmonary fibrosis by regulating TGF-β1-induced profibrotic signaling, suggesting that 11,12-EET and the regulation of EETs could serve as potential therapeutic targets for IPF treatment.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Arachidonic Acid; Biomarkers; Bleomycin; Cell Line; Disease Models, Animal; Disease Susceptibility; Female; Fibroblasts; Gene Expression Profiling; Humans; Idiopathic Pulmonary Fibrosis; Mice; Signal Transduction; Smad Proteins; Transforming Growth Factor beta1

Airway remodeling in asthma is difficult to treat because of its complex pathophysiology that involves proinflammatory cytokines, as well as the arachidonic acid cytochrome P-450 (CYP) pathway; however, it has received little attention. In this study, we assessed the efficacy of a soluble epoxide hydrolase (sEH) on airway remodeling in a mouse model of chronic asthma. The expression of sEH and CYP2J2 and the level of 14,15-epoxyeicosatrienoic acid (14,15-EET), airway remodeling and hyperresponsiveness (AHR) were analyzed to determine the level of sEH inhibition. AUDA, a sEH inhibitor, was given daily for 9 weeks orally, which significantly increased the level of 14,15-EET by inhibiting the expression of sEH and increasing the expression of CYP2J2 in lung tissues. The inhibition of sEH reduced the expression of remodeling-related molecular markers, such as interleukin (IL)-13, IL-17, matrix metalloproteinase 9, N-cadherin, α-smooth muscle actin (α-SMA), S100A4, Twist, epithelial goblet cell metaplasia, and collagen deposition in bronchoalveolar lavage fluid (BAL fluid) and lung tissues. Moreover, remodeling-related eosinophil accumulation in the BAL fluid and infiltration into the lung tissue were improved by AUDA. Finally, AUDA alleviated AHR, which is a functional indicator of airway remodeling. The effect of AUDA on airway remodeling was related to the downregulation of extracellular-regulated protein kinases (Erk1/2), c-Jun N-terminal kinases (JNK) and signal transducer and activator of transcription 3 (STAT3). To our knowledge, this is the first report to demonstrate that inhibition of sEH exerts significant protective effects on airway remodeling in asthma.

Topics: 8,11,14-Eicosatrienoic Acid; Adamantane; Airway Remodeling; Animals; Asthma; Bronchoalveolar Lavage Fluid; Cytochrome P-450 CYP2J2; Cytochrome P-450 Enzyme System; Disease Models, Animal; Epoxide Hydrolases; Female; Humans; Lauric Acids; Lung; MAP Kinase Signaling System; Mice; Ovalbumin; Signal Transduction; STAT3 Transcription Factor

Diabetes mellitus is a metabolic disorder that can lead to blood-brain barrier (BBB) disruption and cognitive decline. However, the mechanisms of BBB breakdown in diabetes are still unclear. Soluble epoxide hydrolase (sEH) is an enzyme that degrades epoxyeicosatrienoic acids (EETs), which have multiple protective effects on vascular structure and functions. In the current study, we showed increased vascular permeability of the BBB, which was accompanied by upregulation of sEH and downregulation of 14,15-EET. Moreover, the sEH inhibitor t-AUCB restored diabetic BBB integrity in vivo, and 14,15-EET prevented ROS accumulation and MEC injury in vitro. t-AUCB or 14,15-EET treatment provoked AMPK/HO-1 activation under diabetic conditions in vivo and in vitro. Thus, we suggest that decreased EET degradation by sEH inhibition might be a potential therapeutic approach to attenuate the progression of BBB injury in diabetic mice via AMPK/HO-1 pathway activation.

Topics: 8,11,14-Eicosatrienoic Acid; AMP-Activated Protein Kinases; Animals; Blood-Brain Barrier; Diabetes Mellitus, Type 2; Disease Models, Animal; Enzyme Inhibitors; Epoxide Hydrolases; Heme Oxygenase-1; Male; Membrane Proteins; Mice; Oxidative Stress; Protective Agents; Reactive Oxygen Species; Signal Transduction

Epoxyeicosatrienoic acids (EETs) are epoxy fatty acids that have biological actions that are essential for maintaining water and electrolyte homeostasis. An inability to increase EETs in response to a high-salt diet results in salt-sensitive hypertension. Vasodilation, inhibition of epithelial sodium channel, and inhibition of inflammation are the major EET actions that are beneficial to the heart, resistance arteries, and kidneys. Genetic and pharmacological means to elevate EETs demonstrated antihypertensive, anti-inflammatory, and organ protective actions. Therapeutic approaches to increase EETs were then developed for cardiovascular diseases. sEH (soluble epoxide hydrolase) inhibitors were developed and progressed to clinical trials for hypertension, diabetes mellitus, and other diseases. EET analogs were another therapeutic approach taken and these drugs are entering the early phases of clinical development. Even with the promise for these therapeutic approaches, there are still several challenges, unexplored areas, and opportunities for epoxy fatty acids.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Arachidonic Acid; Cardiovascular Diseases; Cytochrome P-450 Enzyme System; Disease Models, Animal; Epoxide Hydrolases; Forecasting; Humans; Hypertension; Kidney; Kidney Diseases; Mice; Natriuresis; Potassium; Rats; Rats, Inbred Dahl; Sodium Chloride; Sodium Chloride, Dietary; Vasodilation; Water-Electrolyte Balance; Water-Electrolyte Imbalance

NLRX1 weakens lipopolysaccharide (LPS)-induced NF-κB activation on immune cells. Cytochrome P450 epoxygenase 2J2 (CYP2J2) attenuates LPS-induced cardiac injury by inhibiting NF-κB activation. However, it is still unclear whether NLRX1 could reduce LPS-induced heart damage and whether it is involved in the anti-LPS cardioprotective effect of CYP2J2. In this study, we found that NLRX1 knockout further exacerbated LPS-induced heart injury and up-regulated the proinflammatory cytokines in serum and heart tissue, and weakened the inhibitory effect of CYP2J2 on the harmful effects caused by LPS. We also found that LPS treatment induced ubiquitination of NLRX1 and promoted its binding to IKKα/β in myocardial tissue, which should theoretically inhibit NF-κB activation. However, LPS eventually leads to activation of NF-κB and NLRP3 inflammasome. Under the action of LPS, CYP2J2 further promoted the ubiquitination of NLRX1 and its binding to IKKα/β, impaired NF-κB activation and NLRP3 inflammasome activation. NLRX1 knockout notably aggravated LPS-induced NF-κB activation and NLRP3 inflammasome activation, and attenuated the inhibitory effects of CYP2J2 on NF-κB signal and NLRP3 inflammasome. More, CYP2J2 reduced LPS-induced reactive oxygen species (ROS) production and mitochondrial depolarization in heart cells, thereby inhibiting NLRP3 inflammasome activation. NLRX1 knockdown aggravated mitochondrial depolarization induced by LPS and weakened the protective effect of CYP2J2 on mitochondrial potential, although it had no significant effect on reactive oxygen species production. Together, these findings demonstrated that NLRX1 knockout aggravated LPS-induced heart injury and weakened the anti-LPS cardioprotective effect of CYP2J2 by enhancing activation of NF-κB and NLRP3 inflammasome.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Cytochrome P-450 CYP2J2; Cytochrome P-450 Enzyme System; Cytokines; Disease Models, Animal; Heart Diseases; Inflammasomes; Inflammation Mediators; Lipopolysaccharides; Male; Mice, Inbred C57BL; Mice, Transgenic; Mitochondria, Heart; Mitochondrial Proteins; Myocytes, Cardiac; NF-kappa B; NLR Family, Pyrin Domain-Containing 3 Protein; Reactive Oxygen Species; Signal Transduction

We recently reported that siRNA-knockdown of delta-5-desaturase (D5D), the rate-limiting enzyme converting upstream ω - 6 dihomo-γ-linolenic acid (DGLA) to arachidonic acid, promoted formation of the anti-cancer byproduct 8-hydroxyoctanoic acid (8-HOA) from COX-2-catalyzed DGLA peroxidation, consequently suppressing pancreatic cancer cell growth, migration and invasion. In this study, we have further investigated the anti-tumor effects of D5D-knockdown and the resulting intensified COX-2-catalyzed DGLA peroxidation in subcutaneous xenograft tumors. Four-week old female nude mice (Jackson Laboratory, J:Nu-007850) were injected with human pancreatic cancer cell line BxPC-3 or its D5D knockdown counterpart (via shRNA), followed by 4-week treatments of: vehicle control, DGLA supplementation (8 mg/mouse, twice a week), gemcitabine (30 mg/kg, twice a week), and a combination of DGLA and gemcitabine. In D5D-knockdown tumors, DGLA supplementation promoted 8-HOA formation to a threshold level (> 0.3 µg/g) and resulted in significant tumor reduction (30% vs. control). The promoted 8-HOA not only induced apoptosis associated with altered expression of Bcl-2, cleaved PARP, procaspase 3 and procaspase 9, but also suppressed the tumor metastatic potential via altering MMP-2 and E-cadherin expression. DGLA supplementation resulted in similar anti-tumor effects to those of gemcitabine in our experiments, while the combined treatment led to most significant inhibitory effect on D5D-knockdown tumor growth (70% reduction vs. control). Compared to conventional COX-2 inhibition in cancer treatment, our new strategy that takes advantage of overexpressed COX-2 in cancer cells and tumors, and of abundant ω - 6 fatty acids in the daily diet, should lead us to develop a better and safer anti-pancreatic cancer therapy for patients.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Antineoplastic Agents; Apoptosis; Biomarkers; Cell Line, Tumor; Cell Proliferation; Delta-5 Fatty Acid Desaturase; Deoxycytidine; Disease Models, Animal; Drug Synergism; Fatty Acid Desaturases; Female; Gemcitabine; Gene Expression Regulation, Neoplastic; Humans; Mice; Neoplasm Metastasis; Neoplasm Staging; Pancreatic Neoplasms; RNA Interference; RNA, Small Interfering; Xenograft Model Antitumor Assays

Central poststroke pain (CPSP) is a neuropathic pain syndrome arising after a lesion of the central nervous system owing to cerebrovascular insult. Impaired daily activities and reduced quality of life in people suffering from CPSP justify the need for improved treatment. The detailed mechanism of CPSP is not well understood, but central disinhibition has been suggested. Recent reports indicated that epoxyeicosatrienoic acids (EETs), the cytochrome P450 metabolites of arachidonic acid, promoted neuronal survival after stroke, displayed antinociception in peripheral inflammatory pain, and reduced neuronal excitability in seizure model. Here, we tested the hypothesis that 14,15-EET may attenuate CPSP by suppressing thalamic disinhibition through neurosteroids-δ-subunit-containing gamma-aminobutyric acid A receptors (δGABA

Topics: 8,11,14-Eicosatrienoic Acid; Analgesics; Animals; Cerebral Hemorrhage; Disease Models, Animal; Gabapentin; Hyperalgesia; Male; Pregnanolone; Proof of Concept Study; Random Allocation; Rats, Sprague-Dawley; Receptors, GABA-A; Stroke; Thalamus

Coconut oil is used as a dietary oil worldwide, and its healthy effects are recognized by the fact that coconut oil is easy to digest, helps in weight management, increases healthy cholesterol, and provides instant energy. Although topical application of coconut oil is known to reduce skin infection and inflammation, whether dietary coconut oil has any role in decreasing skin inflammation is unknown. In this study, we showed the impact of dietary coconut oil in allergic skin inflammation by using a mouse model of contact hypersensitivity (CHS). Mice maintained on coconut oil showed amelioration of skin inflammation and increased levels of cis-5, 8, 11-eicosatrienoic acid (mead acid) in serum. Intraperitoneal injection of mead acid inhibited CHS and reduced the number of neutrophils infiltrating to the skin. Detailed mechanistic studies unveiled that mead acid inhibited the directional migration of neutrophils by inhibiting the filamentous actin polymerization and leukotriene B

Topics: 8,11,14-Eicosatrienoic Acid; Actins; Animals; Biomarkers; Capillary Permeability; Chemotaxis; Coconut Oil; Dermatitis, Atopic; Dermatitis, Contact; Dietary Fats, Unsaturated; Disease Models, Animal; Female; Immunohistochemistry; Immunophenotyping; Leukotriene B4; Lipid Metabolism; Mice; Neutrophils; Skin

Non-alcoholic fatty liver disease (NAFLD) is associated with obesity and is considered to be an inflammatory disorder characterized by fatty acid accumulation, oxidative stress, and lipotoxicity. We have previously reported that epoxyeicosatrienoic acid-agonist (EET-A) has multiple beneficial effects on cardiac, renal and adipose tissue function while exhibiting both anti-inflammatory and anti-oxidant activities. We hypothesized that EET-A intervention would play a central role in attenuation of obesity-induced steatosis and hepatic fibrosis that leads to NAFLD.. We studied the effect of EET-A on fatty liver using db/db mice as a model of obesity. Mice were fed a high fat diet (HFD) for 16 weeks and administered EET-A twice weekly for the final 8 weeks.. db/db mice fed HFD significantly increased hepatic lipid accumulation as manifested by increases in NAS scores, hepatic fibrosis, insulin resistance, and inflammation, and decreases in mitochondrial mitofusin proteins (Mfn 1/2) and anti-obesity genes Fibroblast growth factor 21 (FGF21) and Cellular Repressor of E1A-Stimulated Genes 1 (CREG1). EET-A administration reversed the decrease in these genes and reduced liver fibrosis. Knockout of Peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) in EET-A treated mice resulted in a reversal of the beneficial effects of EET-A administration.. EET-A intervention diminishes fatty acid accumulation, fibrosis, and NFALD associated with an increase in HO-1-PGC1α and increased insulin receptor phosphorylation. A pharmacological strategy involving EETs may offer a potential therapeutic approach in preventing fibrosis, mitochondrial dysfunction, and the development of NAFLD.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Disease Models, Animal; Fatty Liver; Heme Oxygenase-1; Mice; Mitochondria; Non-alcoholic Fatty Liver Disease; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha; Receptors, Leptin; Signal Transduction

Exercise training (ET) is a safe and efficacious therapeutic approach for myocardial infarction (MI). Given the numerous benefits of exercise, exercise-induced mediators may be promising treatment targets for MI. C57BL/6 mice were fed 1-trifluoromethoxyphenyl-3-(1-propionylpiperidine-4-yl) urea (TPPU), a novel soluble epoxide hydrolase inhibitor (sEHI), to increase epoxyeicosatrienoic acid (EET) levels, for 1 week before undergoing MI surgery. After 1-week recovery, the mice followed a prescribed exercise programme. Bone marrow-derived endothelial progenitor cells (EPCs) were isolated from the mice after 4 weeks of exercise and cultured for 7 days. Angiogenesis around the ischaemic area, EPC functions, and the expression of microRNA-126 (miR-126) and its target gene Spred1 were measured. The results were confirmed in vitro by adding TPPU to EPC culture medium. ET significantly increased serum EET levels and promoted angiogenesis after MI. TPPU enhanced the effects of ET to reduce the infarct area and improve cardiac function after MI. ET increased EPC function and miR-126 expression, which were further enhanced by TPPU, while Spred1 expression was significantly down-regulated. Additionally, the protein kinase B/glycogen synthase kinase 3β (AKT/GSK3β) signalling pathway was activated after the administration of TPPU. EETs are a potential mediator of exercise-induced cardioprotection in mice after MI. TPPU enhances exercise-induced cardiac recovery in mice after MI by increasing EET levels and promoting angiogenesis around the ischaemic area.

Topics: 8,11,14-Eicosatrienoic Acid; Adaptor Proteins, Signal Transducing; Animals; Bone Marrow Cells; Cardiotonic Agents; Coronary Vessels; Disease Models, Animal; Endothelial Progenitor Cells; Enzyme Inhibitors; Epoxide Hydrolases; Gene Expression Regulation; Glycogen Synthase Kinase 3 beta; Male; Mice; Mice, Inbred C57BL; MicroRNAs; Myocardial Infarction; Neovascularization, Physiologic; Phenylurea Compounds; Physical Conditioning, Animal; Piperidines; Primary Cell Culture; Proto-Oncogene Proteins c-akt; Repressor Proteins; Signal Transduction

14,15-epoxyeicosatrienoic acid (14,15-EET) is an important lipid signaling molecule involved in the regulation of tumor metastasis, however, the role and molecular mechanisms of 14,15-EET activity in breast cancer cell epithelial-mesenchymal transition (EMT) and drug resistance remain enigmatic.. The 14, 15-EET level in serum and in tumor or non-cancerous tissue from breast cancer patients was measured by ELISA. qRT-PCR and western blot analyses were used to examine expression of integrin αvβ3. The role of 14, 15-EET in breast cancer cell adhesion, invasion was explored by adhesion and Transwell assays. The role of 14, 15-EET in breast cancer cell cisplatin resistance in vitro was determined by MTT assay. Western blot was conducted to detect the protein expressions of EMT-related markers and FAK/PI3K/AKT signaling. Xenograft models in nude mice were established to explore the roles of 14, 15-EET in breast cancer cells EMT and cisplatin resistance in vivo.. In the present study, we show that serum level of 14, 15-EET increases in breast cancer patients and 14, 15-EET level of tumor tissue is higher than that of non-cancerous tissue. Moreover, 14, 15-EET increases integrin αvβ3 expression, leading to FAK activation. 14, 15-EET induces breast cancer cell EMT via integrin αvβ3 and FAK/PI3K/AKT cascade activation in vitro. Furthermore, we find that 14, 15-EET induces breast cancer cells EMT and cisplatin resistance in vivo, αvβ3 integrin and the resulting FAK/PI3K/AKT signaling pathway are responsible for 14, 15-EET induced-breast cancer cells cisplatin resistance.. Our findings suggest that inhibition of 14, 15-EET or inactivation of integrin αvβ3/FAK/PI3K/AKT pathway could serve as a novel approach to reverse EMT and cisplatin resistance in breast cancer cells.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Breast Neoplasms; Cell Adhesion; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cisplatin; Disease Models, Animal; Drug Resistance, Neoplasm; Epithelial-Mesenchymal Transition; Female; Humans; Immunohistochemistry; Integrin alphaVbeta3; Mice; Phosphatidylinositol 3-Kinases; Phosphorylation; Proto-Oncogene Proteins c-akt; Signal Transduction; Xenograft Model Antitumor Assays

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Cell Line; Cell Movement; Disease Models, Animal; Epithelial Cells; Humans; Lung; Mice; Multidrug Resistance-Associated Protein 2; Multidrug Resistance-Associated Proteins; Neutrophils; Pneumonia, Pneumococcal; Respiratory Mucosa

Epoxyeicosatrienoic acids (EETs) decrease cardiac ischemia-reperfusion injury; however, the mechanism of their protective effect remains elusive. Here, we investigated the cardioprotective action of a novel EET analog, EET-B, in reperfusion and the role of hypoxia-inducible factor (HIF)-1α in such action of EET-B. Adult male rats were subjected to 30 min of left coronary artery occlusion followed by 2 h of reperfusion. Administration of 14,15-EET (2.5 mg/kg) or EET-B (2.5 mg/kg) 5 min before reperfusion reduced infarct size expressed as a percentage of the area at risk from 64.3 ± 1.3% in control to 42.6 ± 1.9% and 46.0 ± 1.6%, respectively, and their coadministration did not provide any stronger effect. The 14,15-EET antagonist 14,15-epoxyeicosa-5( Z)-enoic acid (2.5 mg/kg) inhibited the infarct size-limiting effect of EET-B (62.5 ± 1.1%). Similarly, the HIF-1α inhibitors 2-methoxyestradiol (2.5 mg/kg) and acriflavine (2 mg/kg) completely abolished the cardioprotective effect of EET-B. In a separate set of experiments, the immunoreactivity of HIF-1α and its degrading enzyme prolyl hydroxylase domain protein 3 (PHD3) were analyzed in the ischemic areas and nonischemic septa. At the end of ischemia, the HIF-1α immunogenic signal markedly increased in the ischemic area compared with the septum (10.31 ± 0.78% vs. 0.34 ± 0.08%). After 20 min and 2 h of reperfusion, HIF-1α immunoreactivity decreased to 2.40 ± 0.48% and 1.85 ± 0.43%, respectively, in the controls. EET-B blunted the decrease of HIF-1α immunoreactivity (7.80 ± 0.69% and 6.44 ± 1.37%, respectively) and significantly reduced PHD3 immunogenic signal in ischemic tissue after reperfusion. In conclusion, EET-B provides an infarct size-limiting effect at reperfusion that is mediated by HIF-1α and downregulation of its degrading enzyme PHD3. NEW & NOTEWORTHY The present study shows that EET-B is an effective agonistic 14,15-epoxyeicosatrienoic acid analog, and its administration before reperfusion markedly reduced myocardial infarction in rats. Most importantly, we demonstrate that increased hypoxia-inducible factor-1α levels play a role in cardioprotection mediated by EET-B in reperfusion likely by mechanisms including downregulation of the hypoxia-inducible factor -1α-degrading enzyme prolyl hydroxylase domain protein 3.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Disease Models, Animal; Down-Regulation; Hypoxia-Inducible Factor 1, alpha Subunit; Hypoxia-Inducible Factor-Proline Dioxygenases; Male; Myocardial Infarction; Myocardial Reperfusion Injury; Myocardium; Proteolysis; Rats, Sprague-Dawley; Signal Transduction; Ventricular Function, Left; Ventricular Remodeling

Epoxyeicosatrienoic acids (EETs) are cytochrome P450 epoxygenase (CYP) metabolites of arachidonic acid and have multiple cardiovascular effects. Ophiopogonin D (OP-D) is an important effective monomeric component in Shenmai injection (SM-I). Both have been reported to have a variety of biological functions, including anti-inflammatory, anti-oxidant, and anti-apoptotic effects. We previously demonstrated that OP-D-mediated cardioprotection involves activation of CYP2J2/3 and enhancement of circulating EETs levels in vitro and can be developed as a novel drug for the therapy of myocardial ischemia-reperfusion (MI/R) injury. We therefore hypothesized that the protective effects of OP-D and SM-I against MI/R injury are associated with increased expression of CYP2J3 and enhanced circulating 11,12-EET levels in vivo.. A rat model of MI/R injury was generated by ligation of the left anterior descending coronary artery for 40 min, followed by reperfusion for 2 h to determine the protective effects and potential mechanisms of OP-D and SM-I. Electrocardiogram and ultrasonic cardiogram were used to evaluate cardiac function; 2,3,5-triphenyltetrazolium chloride was used to measure myocardial infarct size; hematoxylin and eosin staining and transmission electron microscopy were used to observe the morphology of myocardial tissue; and the expression of related proteins in the mechanistic study was observed by western blot analysis.. We found that OP-D and SM-I exert protective effects on MI/R injury, including regulation of cardiac function, reduction of lactate dehydrogenase and creatine kinase production, attenuation of myocardial infarct size, and improvement of the recovery of damaged myocardial structures. We found that OP-D and SM-I activate CYP2J3 expression and increase levels of circulating 11,12-EET in MI/R-injured rats.. We tested the hypothesis that the cardioprotective effects of OP-D and SM-I on MI/R injury are associated with increased expression of CYP2J3 and enhanced circulating 11,12-EET levels in rats. Taken together, our results show that the effects of OP-D and SM-I were also mediated by the activation of the PI3K/Akt/eNOS signaling pathway, while inhibition of the NF-κB signaling pathway and antioxidant and anti-apoptotic effects were involved in the cardioprotective effects of OP-D and SM-I.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Apoptosis; Cardiotonic Agents; Cytochrome P-450 Enzyme System; Disease Models, Animal; Male; Myocardial Reperfusion Injury; Myocardium; NF-kappa B; Nitric Oxide Synthase Type III; Oxidative Stress; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; Saponins; Signal Transduction; Spirostans; Superoxide Dismutase; Up-Regulation

Epoxyeicosatrienoic acid (EET) has wide applications due to the unique biological effects of anti‑hyperlipidemia, inhibition of platelet aggregation, anti‑inflammation, anti‑cancer, anti‑lipid oxidation and the promotion of brain tissue development. The present study investigated whether EET ameliorates cerebral ischemia‑reperfusion injury (CIRI) by inhibiting inflammatory factors and pannexin. Specific pathogen‑free 7‑week‑old male Sprague‑Dawley rats were randomly divided into three groups: Sham, CIRI and EET. Neurological deficit scores, cerebral infarct volume and cerebral edema were assessed in CIRI rats. Enzyme‑linked immunosorbent assays were performed to detect tumor necrosis factor‑α, interleukin‑6, nuclear factor‑κB and inducible nitric oxide synthase (iNOS) levels, and western blot analysis was performed also used to assess cleaved caspase‑3, phospholipase A2 (PLA2), cyclooxygenase‑2 and prostaglandin E2 (PGE2) protein expression levels. EET ameliorated cerebral injury and CIRI‑induced cleaved caspase‑3 protein expression levels in rats. EET additionally suppressed CIRI‑induced inflammation reactions and iNOS protein expression in rats. Furthermore, the protein expression levels of PLA2, PGE2 and pannexin‑1 in CIRI rats were inhibited by treatment with EET. These results indicated that EET reduces CIRI by inhibiting inflammation and levels of cleaved caspase‑3, PLA2, PGE2 and pannexin-1.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Brain Ischemia; Caspase 3; Connexins; Cyclooxygenase 2; Cytokines; Dinoprostone; Disease Models, Animal; Down-Regulation; Enzyme-Linked Immunosorbent Assay; Fatty Acids, Unsaturated; Interleukin-6; Male; Nerve Tissue Proteins; NF-kappa B; Nitric Oxide Synthase Type II; Phospholipases A2; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Tumor Necrosis Factor-alpha

Here we use a rat model to investigate the effects of epoxyeicosatrienoic acids (EETs) on retinal macular degeneration along with pathological and physiological mechanisms of the disease.. Six choroidal neovascularization (CNV) rats were created with a 532 nm laser and received intravitreal injections of EETs in both eyes. On day 1, 3, 7 and 14 after photocoagulation, the thickness and area of CNV were measured with HE staining and choroidal flat mounts. COX-2 and VEGF levels in CNV were detected by immunohistochemistry method. Protein and mRNA expression were studied by Western blotting and RT-PCR.. 14 days after photocoagulation, CNV thickness and area were significantly reduced (p<0.01) in the treatment group compared with the control group. COX-2 and VEGF had high expression in vascular endothelial cells and stromal cells of CNV. Peak expression of COX-2 and VEGF was significantly higher (p<0.01) in the treatment group than in the control group. 7 days after photocoagulation, VEGF protein and mRNA expression were significantly lower (p<0.05) in the treatment group than in the control group, whereas COX-2 mRNA showed no significant difference (p>0.05). FFA found that CNV fluorescein leakage area was significantly reduced (p<0.05) in the treatment group than in the control group. 14 days after photocoagulation, neovascularization area was significantly smaller (p<0.05) in the treatment group than in the control group. Vitreous EETs levels in the treatment group were significantly higher than in the control group. Compared with the control group, the celecoxib treatment group had significantly increased vitreous EETs (p<0.05).. Intravitreal injection of celecoxib could suppress the thickness and area of laser-induced macular degeneration CNV. It also improved the vitreous EETs levels in CNV model rats. COX-2 expression was upregulated in the early generation of laser-induced CNV, which may play an important role in regulating expression of VEGF.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Celecoxib; Choroidal Neovascularization; Cyclooxygenase 2 Inhibitors; Disease Models, Animal; Fluorescein Angiography; Intravitreal Injections; Macular Degeneration; Male; Rats; Rats, Inbred BN; Vascular Endothelial Growth Factor A

Intervertebral disc (IVD) degeneration is considered a common cause of low back pain. In the degenerating IVD, the production of pro-inflammatory cytokines, including IL-1 and TNF-α, progressively increases, contributing to the degenerative process. Epoxyeicosatrienoic acids (EETs), synthesized from arachidonic acid by cytochrome P450 enzymes, act as autocrine and paracrine effectors in regulating inflammation, cardiovascular functions, and angiogenesis. EETs were shown to be especially potent promoters of tissue regeneration. Considering their anti-inflammatory and anti-catabolic potential, we investigated whether EETs can influence IVD degeneration. We found that 14,15-EET protected rat nucleus pulposus (NP) cells against death induced by treatment with H2O2and TNF-α in vitro. At the molecular level, 14,15-EET significantly inhibited the NF-κB pathway, which plays essential roles in the degeneration and survival of NP cells. As a result, 14,15-EET efficiently prevented the matrix remodeling response of NP cells to TNF-α. Using a needle-punctured rat tail model, the influence of 14,15-EET on IVD degeneration in vivo was evaluated using radiographs, magnetic resonance images (MRI), and histological analysis. We observed that 14,15-EET prevented IVD degeneration. Our findings demonstrated that 14,15-EET can enhance the survival of NP cells and inhibit IVD degeneration. The EET pathway may be a novel therapeutic target against IVD degeneration.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Apoptosis; Cells, Cultured; Disease Models, Animal; Hydrogen Peroxide; Intervertebral Disc Degeneration; Male; NF-kappa B; Nucleus Pulposus; Rats; Signal Transduction; Tumor Necrosis Factor-alpha

The biological role of epoxyeicosatrienoic acids (EETs) in the regulation of pulmonary circulation is currently under debate. We hypothesized that EETs initiate increases in right ventricular systolic pressure (RVSP) via perhaps, pulmonary vasoconstriction.. Mice were anesthetized with isoflurane. Three catheters, inserted into the left jugular vein, the left carotid artery, and the right jugular vein, were used for infusing EETs, monitoring blood pressure (BP), and RVSP respectively. BP and RVSP were continuously recorded at basal conditions, in response to administration of 4 regioisomeric EETs (5,6-EET; 8,9-EET; 11,12-EET, and 14,15-EET; 1, 2, 5 and 10 ng/g body weight (BW) for each EET), and during exposure of mice to hypoxia.. All 4 EETs initiated dose-dependent increases in RVSP, though reduced BP. 11,12-EET elicited the greatest increment in RVSP among all EET isoforms. To clarify the direct elevation of RVSP in a systemic BP-independent manner, equivalent amounts of 14,15-EET were injected over 1 and 2 minutes respectively. One-minute injection of 14,15-EET elicited significantly faster and greater increases in RVSP than the 2-minute injection, whereas their BP changes were comparable. Additionally, direct injection of low doses of 14,15-EET (0.1, 0.2, 0.5, and 1 ng/g BW) into the right ventricle caused significant increases in RVSP without effects on BP, confirming that systemic vasodilation-induced increases in venous return are not the main cause for the increased RVSP. Acute exposure of mice to hypoxia significantly elevated RVSP, as well as 14,15-EET-induced increases in RVSP.. EETs directly elevate RVSP, a response that may play an important role in the development of hypoxia-induced pulmonary hypertension (PH).

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Arterial Pressure; Disease Models, Animal; Dose-Response Relationship, Drug; Hypertension, Pulmonary; Hypoxia; Infusions, Intravenous; Male; Mice, Inbred C57BL; Pulmonary Artery; Time Factors; Ventricular Function, Right; Ventricular Pressure

Accumulating data suggest that epoxyeicosatrienoic acids (EETs) and 20-hydroxyeicosatetraenoic acid, both cytochrome P450 (P450) enzyme metabolites of arachidonic acid (AA), play important roles in cardiovascular diseases. For many years, the cardiotonic pill (CP), an herbal preparation derived from Salviae Miltiorrhizae Radix et Rhizoma, Notoginseng Radix et Rhizoma, and Borneolum Syntheticum, has been widely used in China for the treatment of coronary artery disease. However, its pharmacological mechanism has not been well elucidated. The purpose of this study was to investigate the chronic effects of the CP on myocardial ischemia-reperfusion injury (MIRI) and AA P450 enzyme metabolism in rats (in vivo) and H9c2 cells (in vitro). The results showed that CP dose dependently (10, 20, and 40 mg/kg/d; 7 days) mitigated MIRI in rats. The plasma concentrations of EETs in CP-treated ischemia-reperfusion (I/R) rats (40 mg/kg/d; 7 days) were significantly higher (P < 0.05) than those in controls. Cardiac Cyp1b1, Cyp2b1, Cyp2e1, Cyp2j3, and Cyp4f6 were significantly induced (P < 0.05); CYP2J and CYP2C11 proteins were upregulated (P < 0.05); and AA-epoxygenases activity was significantly increased (P < 0.05) after CP (40 mg/kg/d; 7 days) administration in rats. In H9c2 cells, the CP also increased (P < 0.05) the EET concentrations and showed protection in hypoxia-reoxygenation (H/R) cells. However, an antagonist of EETs, 14,15-epoxyeicosa-5(Z)-enoic acid, displayed a dose-dependent depression of the CP's protective effects in H/R cells. In conclusion, upregulation of cardiac epoxygenases after multiple doses of the CP-leading to elevated concentrations of cardioprotective EETs after myocardial I/R-may be the underlying mechanism, at least in part, for the CP's cardioprotective effect in rats.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Cardiotonic Agents; Cell Line; Creatine Kinase, MB Form; Cytochrome P-450 Enzyme System; Cytoprotection; Disease Models, Animal; Dose-Response Relationship, Drug; Drugs, Chinese Herbal; Eicosanoids; Isoenzymes; L-Lactate Dehydrogenase; Male; Myocardial Reperfusion Injury; Myocytes, Cardiac; Rats, Sprague-Dawley; Up-Regulation

Dietary supplementation with polyunsaturated fatty acids has been widely used for primary and secondary prevention of cardiovascular disease in individuals at risk; however, the cardioprotective benefits of polyunsaturated fatty acids remain controversial because of lack of mechanistic and in vivo evidence. We present direct evidence that an omega-6 polyunsaturated fatty acid, dihomo-γ-linolenic acid (DGLA), exhibits in vivo cardioprotection through 12-lipoxygenase (12-LOX) oxidation of DGLA to its reduced oxidized lipid form, 12(S)-hydroxy-8Z,10E,14Z-eicosatrienoic acid (12(S)-HETrE), inhibiting platelet activation and thrombosis.. DGLA inhibited ex vivo platelet aggregation and Rap1 activation in wild-type mice, but not in mice lacking 12-LOX expression (12-LOX(-/-)). Similarly, wild-type mice treated with DGLA were able to reduce thrombus growth (platelet and fibrin accumulation) after laser-induced injury of the arteriole of the cremaster muscle, but not 12-LOX(-/-) mice, supporting a 12-LOX requirement for mediating the inhibitory effects of DGLA on platelet-mediated thrombus formation. Platelet activation and thrombus formation were also suppressed when directly treated with 12(S)-HETrE. Importantly, 2 hemostatic models, tail bleeding and arteriole rupture of the cremaster muscle, showed no alteration in hemostasis after 12(S)-HETrE treatment. Finally, the mechanism for 12(S)-HETrE protection was shown to be mediated via a Gαs-linked G-protein-coupled receptor pathway in human platelets.. This study provides the direct evidence that an omega-6 polyunsaturated fatty acid, DGLA, inhibits injury-induced thrombosis through its 12-LOX oxylipin, 12(S)-HETrE, which strongly supports the potential cardioprotective benefits of DGLA supplementation through its regulation of platelet function. Furthermore, this is the first evidence of a 12-LOX oxylipin regulating platelet function in a Gs α subunit-linked G-protein-coupled receptor-dependent manner.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Arachidonate 12-Lipoxygenase; Blood Platelets; Cell Adhesion Molecules; Chromogranins; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Disease Models, Animal; Fibrinolytic Agents; GTP-Binding Protein alpha Subunits, Gs; Humans; Mice, Inbred C57BL; Mice, Knockout; Microfilament Proteins; Oxidation-Reduction; Phosphoproteins; Phosphorylation; Platelet Activation; Platelet Aggregation; Platelet Aggregation Inhibitors; Shelterin Complex; Signal Transduction; Telomere-Binding Proteins; Thrombosis; Time Factors

The objective of the present study was to assess the effect of elevating epoxygenated fatty acids on retinal vascular inflammation. To stimulate inflammation we utilized TNFα, a potent pro-inflammatory mediator that is elevated in the serum and vitreous of diabetic patients. In TNFα-stimulated primary human retinal microvascular endothelial cells, total levels of epoxyeicosatrienoic acids (EETs), but not epoxydocosapentaenoic acids (EDPs), were significantly decreased. Exogenous addition of 11,12-EET or 19,20-EDP when combined with 12-(3-adamantane-1-yl-ureido)-dodecanoic acid (AUDA), an inhibitor of epoxide hydrolysis, inhibited VCAM-1 and ICAM-1 expression and protein levels; conversely the diol product of 19,20-EDP hydrolysis, 19,20-DHDP, induced VCAM1 and ICAM1 expression. 11,12-EET and 19,20-EDP also inhibited leukocyte adherence to human retinal microvascular endothelial cell monolayers and leukostasis in an acute mouse model of retinal inflammation. Our results indicate that this inhibition may be mediated through an indirect effect on NFκB activation. This is the first study demonstrating a direct comparison of EET and EDP on vascular inflammatory endpoints, and we have confirmed a comparable efficacy from each isomer, suggesting a similar mechanism of action. Taken together, these data establish that epoxygenated fatty acid elevation will inhibit early pathology related to TNFα-induced inflammation in retinal vascular diseases.

Topics: 8,11,14-Eicosatrienoic Acid; Adamantane; Animals; Cells, Cultured; Disease Models, Animal; Down-Regulation; Endothelial Cells; Epoxy Compounds; Fatty Acids, Unsaturated; Humans; Intercellular Adhesion Molecule-1; Lauric Acids; Male; Mice; Retinal Vasculitis; Retinal Vessels; Tumor Necrosis Factor-alpha; Vascular Cell Adhesion Molecule-1

Patients recovering from aneurysmal subarachnoid hemorrhage (SAH) are at risk for developing delayed cerebral ischemia (DCI). Experimental and human studies implicate the vasoconstrictor P450 eicosanoid 20-hydroxyeicosatetraenoic acid (20-HETE) in the pathogenesis of DCI. To date, no studies have evaluated the role of vasodilator epoxyeicosatrienoic acids (EETs) in DCI.. Using mass spectrometry, we measured P450 eicosanoids in cerebrospinal fluid (CSF) from 34 SAH patients from 1 to 14 days after admission. CSF eicosanoid levels were compared in patients who experienced DCI versus those who did not. We then studied the effect of EETs in a model of SAH using mice lacking the enzyme soluble epoxide hydrolase (sEH), which catabolizes EETs into their inactive diol. To assess changes in vessel morphology and cortical perfusion in the mouse brain, we used optical microangiography, a non-invasive coherence-based imaging technique.. Along with increases in 20-HETE, we found that CSF levels of 14,15-EET were elevated in SAH patients compared to control CSF, and levels were significantly higher in patients who experienced DCI compared to those who did not. Mice lacking sEH had elevated 14,15-EET and were protected from the delayed decrease in microvascular cortical perfusion after SAH, compared to wild type mice.. Our findings suggest that P450 eicosanoids play an important role in the pathogenesis of DCI. While 20-HETE may contribute to the development of DCI, 14,15-EET may afford protection against DCI. Strategies to enhance 14,15-EET, including sEH inhibition, should be considered as part of a comprehensive approach to prevent DCI.

Topics: 8,11,14-Eicosatrienoic Acid; Aged; Animals; Brain Ischemia; Disease Models, Animal; Female; Humans; Hydroxyeicosatetraenoic Acids; Male; Mice; Middle Aged; Neuroprotective Agents; Subarachnoid Hemorrhage

Cytochrome P-450 epoxygenase-derived epoxyeicosatrienoic acids (EETs) exert diverse biological activities, which include potent vasodilatory, anti-inflammatory, antiapoptotic, and antioxidatant effects, and cardiovascular protection. Liver has abundant epoxygenase expression and high levels of EET production; however, the roles of epoxygenases in liver diseases remain to be elucidated. In this study, we investigated the protection against high-fat diet-induced nonalcoholic fatty liver disease (NAFLD) in mice with endothelial-specific CYP2J2 overexpression (Tie2-CYP2J2-Tr). After 24 wk of high-fat diet, Tie2-CYP2J2-Tr mice displayed attenuated NAFLD compared with controls. Tie2-CYP2J2-Tr mice showed significantly decreased plasma triglyceride levels and liver lipid accumulation, improved liver function, reduced inflammatory responses, and less increase in hepatic oxidative stress than wild-type control mice. These effects were associated with inhibition of NF-κB/JNK signaling pathway activation and enhancement of the antioxidant defense system in Tie2-CYP2J2-Tr mice in vivo. We also demonstrated that 14,15-EET treatment protected HepG2 cells against palmitic acid-induced inflammation and oxidative stress. 14,15-EET attenuated palmitic acid-induced changes in NF-κB/JNK signaling pathways, malondialdehyde generation, glutathione levels, reactive oxygen species production, and NADPH oxidase and antioxidant enzyme expression in HepG2 cells in vitro. Together, these results highlight a new role for CYP epoxygenase-derived EETs in lipotoxicity-related inflammation and oxidative stress and reveal a new molecular mechanism underlying EETs-mediated anti-inflammatory and antioxidant effects that could aid in the design of new therapies for the prevention and treatment of NAFLD.

Topics: 8,11,14-Eicosatrienoic Acid; Alanine Transaminase; Animals; Aspartate Aminotransferases; Catalase; Cytochrome P-450 CYP2J2; Cytochrome P-450 Enzyme System; Cytokines; Diet, High-Fat; Disease Models, Animal; Glutathione Peroxidase; Hep G2 Cells; Humans; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; NF-kappa B; Non-alcoholic Fatty Liver Disease; Oxidative Stress; Palmitic Acid; Reverse Transcriptase Polymerase Chain Reaction; RNA; Signal Transduction; Superoxide Dismutase; Triglycerides

Streptococcus pneumoniae infections are an important cause of morbidity and mortality in older patients. Uncontrolled neutrophil-driven pulmonary inflammation exacerbates this disease. To test whether the α-tocopherol (α-Toc) form of vitamin E, a regulator of immunity, can modulate neutrophil responses as a preventive strategy to mitigate the age-associated decline in resistance to S. pneumoniae, young (4 mo) and old (22-24 mo) C57BL/6 mice were fed a diet containing 30-PPM (control) or 500-PPM (supplemented) α-Toc for 4 wk and intratracheally infected with S. pneumoniae. Aged mice fed a control diet were exquisitely more susceptible to S. pneumoniae than young mice. At 2 d postinfection, aged mice suffered 1000-fold higher pulmonary bacterial burden, 2.2-fold higher levels of neutrophil recruitment to the lung, and a 2.25-fold higher rate of lethal septicemia. Strikingly, α-Toc supplementation of aged mice resulted in a 1000-fold lower bacterial lung burden and full control of infection. This α-Toc-induced resistance to pneumococcal challenge was associated with a 2-fold fewer pulmonary neutrophils, a level comparable to S. pneumoniae-challenged, conventionally fed young mice. α-Toc directly inhibited neutrophil egress across epithelial cell monolayers in vitro in response to pneumococci or hepoxilin-A3, an eicosanoid required for pneumococcus-elicited neutrophil trans-epithelial migration. α-Toc altered expression of multiple epithelial and neutrophil adhesion molecules involved in migration, including CD55, CD47, CD18/CD11b, and ICAM-1. These findings suggest that α-Toc enhances resistance of aged mice to bacterial pneumonia by modulating the innate immune response, a finding that has potential clinical significance in combating infection in aged individuals through nutritional intervention.

Topics: 8,11,14-Eicosatrienoic Acid; Age Factors; alpha-Tocopherol; Animals; Cell Adhesion Molecules; Dietary Supplements; Disease Models, Animal; Disease Resistance; Disease Susceptibility; Epithelium; Gene Expression; Male; Mice; Neutrophil Infiltration; Neutrophils; Pneumonia, Pneumococcal; Streptococcus pneumoniae; Transendothelial and Transepithelial Migration

Neutrophilic infiltration is a leading contributor to pathology in a number of pulmonary disease states, including cystic fibrosis. Hepoxilin A3 (HXA3) is a chemotactic eicosanoid shown to mediate the transepithelial passage of neutrophils in response to infection in several model systems and at multiple mucosal surfaces. Another well-known eicosanoid mediating general neutrophil chemotaxis is leukotriene B4 (LTB4). We sought to distinguish the roles of each eicosanoid in the context of infection of lung epithelial monolayers by Pseudomonas aeruginosa. Using human and mouse in vitro transwell model systems, we used a combination of biosynthetic inhibitors, receptor antagonists, as well as mutant sources of neutrophils to assess the contribution of each chemoattractant in driving neutrophil transepithelial migration. We found that following chemotaxis to epithelial-derived HXA3 signals, neutrophil-derived LTB4 is required to amplify the magnitude of neutrophil migration. LTB4 signaling is not required for migration to HXA3 signals, but LTB4 generation by migrated neutrophils plays a significant role in augmenting the initial HXA3-mediated migration. We conclude that HXA3 and LTB4 serve independent roles to collectively coordinate an effective neutrophilic transepithelial migratory response.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Bacteria; Bacterial Infections; Calcium Signaling; Cell Line; Chemotaxis, Leukocyte; Disease Models, Animal; Female; Gene Knockdown Techniques; Humans; Leukotriene B4; Mice; Neutrophil Infiltration; Neutrophils; Pseudomonas aeruginosa; Receptors, Leukotriene B4; Transendothelial and Transepithelial Migration

Cardiac remodelling is one of the key pathological changes that occur with cardiovascular disease. Previous studies have demonstrated the beneficial effects of CYP2J2 expression on cardiac injury. In the present study, we investigated the effects of cardiomyocyte-specific CYP2J2 expression and EET treatment on angiotensin II-induced cardiac remodelling and sought to determine the underlying molecular mechanisms involved in this process.. Eight-week-old mice with cardiomyocyte-specific CYP2J2 expression (αMHC-CYP2J2-Tr) and wild-type (WT) control mice were treated with Ang-II. Ang-II treatment of WT mice induced changes in heart morphology, cardiac hypertrophy and dysfunction, as well as collagen accumulation; however, cardiomyocyte-specific expression of CYP2J2 attenuated these effects. The cardioprotective effects observed in α-MHC-CYP2J2-Tr mice were associated with peroxisome proliferator-activated receptor (PPAR)-γ activation, reduced oxidative stress, reduced NF-κB p65 nuclear translocation, and inhibition of TGF-β1/smad pathway. The effects seen with cardiomyocyte-specific expression of CYP2J2 were partially blocked by treatment with PPAR-γ antagonist GW9662. In in vitro studies, 11,12-EET(1 μmol/L) treatment attenuated cardiomyocyte hypertrophy and remodelling-related protein (collagen I, TGF-β1, TIMP1) expression by inhibiting the oxidative stress-mediated NF-κB pathway via PPAR-γ activation. Furthermore, conditioned media from neonatal cardiomyocytes treated with 11,12-EET inhibited activation of cardiac fibroblasts and TGF-β1/smad pathway.. Cardiomyocyte-specific expression of CYP2J2 or treatment with EETs protects against cardiac remodelling by attenuating oxidative stress-mediated NF-κBp65 nuclear translocation via PPAR-γ activation.

Topics: 8,11,14-Eicosatrienoic Acid; Angiotensin II; Animals; Cells, Cultured; Coculture Techniques; Cytochrome P-450 CYP2J2; Cytochrome P-450 Enzyme System; Disease Models, Animal; Hypertrophy, Left Ventricular; Male; Mice, Inbred C57BL; Mice, Transgenic; Myocytes, Cardiac; Oxidative Stress; PPAR gamma; Rats; Signal Transduction; Transcription Factor RelA; Ventricular Function, Left; Ventricular Remodeling

The detailed mechanisms determining the course of congestive heart failure (CHF) and associated renal dysfunction remain unclear. In a volume overload model of CHF induced by creation of aorto-caval fistula (ACF) in Hannover Sprague-Dawley (HanSD) rats we explored the putative pathogenetic contribution of epoxyeicosatrienoic acids (EETs), active products of CYP-450 dependent epoxygenase pathway of arachidonic acid metabolism, and compared it with the role of the renin-angiotensin system (RAS). Chronic treatment with cis-4-[4-(3-adamantan-1-yl-ureido) cyclohexyloxy]benzoic acid (c-AUCB, 3 mg/l in drinking water), an inhibitor of soluble epoxide hydrolase (sEH) which normally degrades EETs, increased intrarenal and myocardial EETs to levels observed in sham-operated HanSD rats, but did not improve the survival or renal function impairment. In contrast, chronic angiotensin-converting enzyme inhibition (ACEi, trandolapril, 6 mg/l in drinking water) increased renal blood flow, fractional sodium excretion and markedly improved survival, without affecting left ventricular structure and performance. Hence, renal dysfunction rather than cardiac remodeling determines long-term mortality in advanced stage of CHF due to volume overload. Strong protective actions of ACEi were associated with suppression of the vasoconstrictor/sodium retaining axis and activation of vasodilatory/natriuretic axis of the renin-angiotensin system in the circulating blood and kidney tissue.

Topics: 8,11,14-Eicosatrienoic Acid; Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme Inhibitors; Animals; Benzoates; Disease Models, Animal; Drug Evaluation, Preclinical; Epoxide Hydrolases; Epoxy Compounds; Heart Failure; Kidney; Male; Myocardium; Peptide Fragments; Random Allocation; Rats; Renal Insufficiency; Renin-Angiotensin System; Ultrasonography; Urea

The study aims to investigate the effect of cytochrome P450 2J2 (CYP2J2) overexpression on hyperlipidemia in mice and further to explore its effect on fatty acid oxidation in vivo and in vitro.. The effects and mechanisms of endothelial-specific CYP2J2 transgene (Tie2-CYP2J2-Tr) on lipid and fatty acid metabolism were investigated in high-fat diet (HFD) -treated mice. HepG2, LO2 cells, and HUVECs were exposed to 0.4 mM free fatty acid (FFA) for 24 h and used as a model to investigate the roles of CYP2J2 overexpression and epoxyeicosatrienoic acids (EETs) on fatty acid β-oxidation in vitro.. Tie2-CYP2J2-Tr mice had significantly lower plasma and liver triglycerides, lower liver cholesterol and fatty acids, and reduced HFD-induced lipid accumulation. CYP2J2 overexpression resulted in activation of the hepatic and endothelial AMPKα, increased ACC phosphorylation, and increased expression of CPT-1 and PPARα, which were all reduced by HFD treatment. In FFA-treated HepG2, LO2, and HUVECs, both CYP2J2 overexpression and EETs significantly decreased lipid accumulation and increased fatty acid oxidation via activating the AMPK and PPARα pathways.. Endothelial-specific CYP2J2 overexpression alleviates HFD-induced hyperlipidemia in vivo. CYP2J2 ameliorates FFA-induced dyslipidemia via increased fatty acid oxidation mediated by the AMPK and PPARα pathways.

Topics: 8,11,14-Eicosatrienoic Acid; AMP-Activated Protein Kinases; Animals; Cholesterol; Diet, High-Fat; Disease Models, Animal; Fatty Acids; Fatty Acids, Nonesterified; Hyperlipidemias; Lipid Metabolism; Liver; Mice; Mice, Inbred C57BL; Mice, Transgenic; Oxidative Stress; Triglycerides

This study examined the effects of a novel orally active 14,15-epoxyeicosatrienoic acid analog (EET-A) on blood pressure (BP) and myocardial infarct size (IS) in two-kidney, one-clip (2K1C) Goldblatt hypertensive rats during sustained phase of hypertension. Between days 31 and 35 after clip placement the rats were treated with EET-A and BP was monitored by radiotelemetry; sham-operated normotensive rats were used as controls. Tissue concentrations of epoxyeicosatrienoic acids served as a marker of production of epoxygenase metabolites. The rats were subjected to acute myocardial ischemia/reperfusion (I/R) injury and IS was determined. We found that EET-A treatment did not lower BP in 2K1C rats and did not alter availability of biologically active epoxygenase metabolites in 2K1C or in sham-operated rats. The myocardial IS was significantly smaller in untreated 2K1C rats as compared with normotensive controls and EET-A reduced it in controls but not in 2K1C rats. Our findings suggest that during the phase of sustained hypertension 2K1C Goldblatt hypertensive rats exhibit increased cardiac tolerance to I/R injury as compared with normotensive controls, and that in this animal model of human renovascular hypertension short-term treatment with EET-A does not induce any antihypertensive and cardioprotective actions.

Topics: 8,11,14-Eicosatrienoic Acid; Administration, Oral; Animals; Blood Pressure; Blood Pressure Monitoring, Ambulatory; Disease Models, Animal; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Hydroxyeicosatetraenoic Acids; Hypertension, Renovascular; Kidney; Male; Myocardial Infarction; Myocardial Reperfusion Injury; Myocardium; Proto-Oncogene Proteins c-akt; Rats, Sprague-Dawley; Renin-Angiotensin System; Signal Transduction; Telemetry; Time Factors

Cardiac remodeling is one of the most common cardiac abnormalities and associated with a high mortality in chronic renal failure (CRF) patients. Apocynin, a nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase inhibitor, has been showed cardio-protective effects. However, whether apocynin can improve cardiac remodeling in CRF and what is the underlying mechanism are unclear. In the present study, we enrolled 94 participants. In addition, we used 5/6 nephrectomized rats to mimic cardiac remodeling in CRF. Serum levels of epoxyeicosatrienoic acids (EETs) and its mainly metabolic enzyme-soluble epoxide hydrolase (sEH) were measured. The results showed that the serum levels of EETs were significantly decreased in renocardiac syndrome participants (P < 0.05). In 5/6 nephrectomized CRF model, the ratio of left ventricular weight / body weight, left ventricular posterior wall thickness, and cardiac interstitial fibrosis were significantly increased while ejection fraction significantly decreased (P < 0.05). All these effects could partly be reversed by apocynin. Meanwhile, we found during the process of cardiac remodeling in CRF, apocynin significantly increased the reduced serum levels of EETs and decreased the mRNA and protein expressions of sEH in the heart (P < 0.05). Our findings indicated that the protective effect of apocynin on cardiac remodeling in CRF was associated with the up-regulation of EETs. EETs may be a new mediator for the injury of kidney-heart interactions.

Topics: 8,11,14-Eicosatrienoic Acid; Acetophenones; Aged; Angiotensin II; Animals; Cardio-Renal Syndrome; Cardiotonic Agents; Cell Line; Disease Models, Animal; Epoxide Hydrolases; Female; Fibrosis; Humans; Kidney Failure, Chronic; Male; Middle Aged; Myocytes, Cardiac; Rats, Sprague-Dawley; Stroke Volume; Up-Regulation; Ventricular Function, Left; Ventricular Remodeling

Polyunsaturated fatty acids have been attracting considerable interest because of their many biological activities and important roles in human health and nutrition. Dihomo-γ-linolenic acid (DGLA; C20: 3n-6) is known to have an anti-inflammatory activity, but its range of effects was not well studied because of its limited natural sources. Taking advantage of genetic tractability and increasing wealth of accessible data of Saccharomyces cerevisiae, we have previously constructed a DGLA-producing yeast strain by introducing two types of desaturase and one elongase genes to convert endogenous oleic acid (C18:1n-9) to DGLA. In this study, we investigated the efficacy of oral intake of heat-killed whole DGLA-producing yeast cells in the absence of lipid purification on cutaneous inflammation. Topical application of croton oil to mouse ears induces ear swelling in parallel with the increased production of chemokines and accumulation of infiltrating cells into the skin sites. These inflammatory reactions were significantly suppressed in a dose-dependent manner by oral intake of the DGLA-producing yeast cells for only 7 days. This suppression was not observed by the intake of the γ-linolenic acid-producing (C18:3n-6, an immediate precursor of DGLA) yeast, indicating DGLA itself suppressed the inflammation. Further analysis demonstrated that DGLA exerted an anti-inflammatory effect via prostaglandin E1 formation because naproxen, a cyclooxygenase inhibitor, attenuated the suppression. Since 25-fold of purified DGLA compared with that provided as a form of yeast was not effective, oral administration of the whole DGLA-producing yeast is considered to be a simple but efficient method to suppress inflammatory responses.

Topics: 8,11,14-Eicosatrienoic Acid; Administration, Oral; Animals; Anti-Inflammatory Agents; Biological Therapy; Croton Oil; Dermatitis; Disease Models, Animal; Metabolic Engineering; Mice; Oleic Acid; Saccharomyces cerevisiae

Cytochrome P-450 (CYP) epoxygenases metabolize arachidonic acid into epoxyeicosatrienoic acids (EETs), which play important roles in regulating cardiovascular functions. The anti-inflammatory, antiapoptotic, proangiogenic, and antihypertensive properties of EETs suggest a beneficial role for EETs in diabetic nephropathy. Endogenous EET levels are maintained by a balance between synthesis by CYP epoxygenases and hydrolysis by epoxide hydrolases into physiologically less active dihydroxyeicosatrienoic acids. Genetic disruption of soluble epoxide hydrolase (sEH/EPHX2) results in increased EET levels through decreased hydrolysis. This study investigated the effects of sEH gene disruption on diabetic nephropathy in streptozotocin-induced diabetic mice. Streptozotocin-induced diabetic manifestations were attenuated in sEH-deficient mice relative to wild-type controls, with significantly decreased levels of Hb A(1c), creatinine, and blood urea nitrogen and urinary microalbumin excretion. The sEH-deficient diabetic mice also had decreased renal tubular apoptosis that coincided with increased levels of antiapoptotic Bcl-2 and Bcl-xl, and decreased levels of the proapoptotic Bax. These effects were associated with activation of the PI3K-Akt-NOS3 and AMPK signaling cascades. sEH gene inhibition and exogenous EETs significantly protected HK-2 cells from TNFα-induced apoptosis in vitro. These findings highlight the beneficial role of the CYP epoxygenase-EETs-sEH system in the pathogenesis of diabetic nephropathy and suggest that the sEH inhibitors available may be potential therapeutic agents for this condition.

Topics: 8,11,14-Eicosatrienoic Acid; Albuminuria; Animals; Apoptosis; Apoptosis Regulatory Proteins; Cell Line, Transformed; Cytoplasm; Diabetic Nephropathies; Disease Models, Animal; Epoxide Hydrolases; Gene Silencing; Humans; Hyperglycemia; Kidney Cortex; Kidney Tubules, Proximal; Mice; Molecular Targeted Therapy; RNA, Small Interfering; Signal Transduction; Streptozocin; Tumor Necrosis Factor-alpha

Epoxyeicosatrienoic acids (EETs) are vasodilator, natriuretic, and antiinflammatory lipid mediators. Both cis- and trans-EETs are stored in phospholipids and in red blood cells (RBCs) in the circulation; the maximal velocity (V(max)) of trans-EET hydrolysis by soluble epoxide hydrolase (sEH) is threefold that of cis-EETs. Because RBCs of the spontaneously hypertensive rat (SHR) exhibit increased sEH activity, a deficiency of trans-EETs in the SHR was hypothesized to increase blood pressure (BP). This prediction was fulfilled, since sEH inhibition with cis-4-[4-(3-adamantan-1-ylureido)cyclohexyloxy]benzoic acid (AUCB; 2 mg·kg(-1)·day(-1) for 7 days) in the SHR reduced mean BP from 176 ± 8 to 153 ± 5 mmHg (P < 0.05), whereas BP in the control Wistar-Kyoto rat (WKY) was unaffected. Plasma levels of EETs in the SHR were lower than in the age-matched control WKY (16.4 ± 1.6 vs. 26.1 ± 1.8 ng/ml; P < 0.05). The decrease in BP in the SHR treated with AUCB was associated with an increase in plasma EETs, which was mostly accounted for by increasing trans-EET from 4.1 ± 0.2 to 7.9 ± 1.5 ng/ml (P < 0.05). Consistent with the effect of increased plasma trans-EETs and reduced BP in the SHR, the 14,15-trans-EET was more potent (ED(50) 10(-10) M; maximum dilation 59 ± 15 μm) than the cis-isomer (ED(50) 10(-9) M; maximum dilation 30 ± 11 μm) in relaxing rat preconstricted arcuate arteries. The 11,12-EET cis- and trans-isomers were equipotent dilators as were the 8,9-EET isomers. In summary, inhibition of sEH resulted in a twofold increase in plasma trans-EETs and reduced mean BP in the SHR. The greater vasodilator potency of trans- vs. cis-EETs may contribute to the antihypertensive effects of sEH inhibitors.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Arachidonic Acids; Benzoic Acid; Blood Pressure; Disease Models, Animal; Epoxide Hydrolases; Erythrocytes; Hypertension; Male; Rats; Rats, Inbred SHR; Rats, Inbred WKY

Recently, a novel observation was made in which nonischemic trauma at a site remote from the heart produced by a transverse abdominal incision resulted in a marked reduction of infarct size (IS) in the mouse heart via activation of sensory nerve fibers in the skin and subsequent activation of bradykinin 2 receptors (BK2R). This phenomenon was termed remote preconditioning of trauma (RPCT). Since RPCT may have potential clinical implications we attempted to confirm these findings in a large animal model, the dog. The epoxyeicosatrienoic acids (EETs) have also recently been shown to be antinociceptive and have been shown to mimic ischemic preconditioning (IPC) and postconditioning (POC) in dogs, therefore, we tested the role of the EETs in RPCT.. Anesthetized adult mongrel dogs of either sex were subjected to 60 min of left anterior descending (LAD) coronary artery occlusion followed by 3 h of reperfusion. In all groups except the controls (no slit), a transverse slit (9 cm) was applied to the abdominal wall of the dog being careful to only slit the skin. Subsequently, 15 min after the slit the heart was subjected to the ischemia/reperfusion protocol.. In the control dogs, the IS as a percent of the area at risk (AAR) was 22.5 ± 2.4%, whereas in the dogs subjected to the slit alone the IS/AAR was reduced to 9.2 ± 1.2% (*P < 0.01). The BR2R blocker, HOE 140 (50 ug/kg, iv) given 10 min prior to the slit, completely abolished the protective effects of RCPT as did pretreatment with 14,15-EEZE, a putative EET receptor blocker or pretreatment with the selective EET synthesis inhibitor, MSPPOH.. These results suggest that BK and the EETs share cardioprotective properties in a large animal model of RPCT.

Topics: 8,11,14-Eicosatrienoic Acid; Abdomen; Animals; Bradykinin; Bradykinin B2 Receptor Antagonists; Coronary Circulation; Cytochrome P-450 Enzyme Inhibitors; Cytochrome P-450 Enzyme System; Disease Models, Animal; Dogs; Female; Hemodynamics; Ischemic Postconditioning; Ischemic Preconditioning, Myocardial; Male; Myocardial Infarction; Myocardial Reperfusion Injury; Myocardium; Receptor, Bradykinin B2

One of the prominent features of Alzheimer's disease is the excessive accumulation of the protein amyloid beta (Aβ) in certain areas of the brain leading to neurodegeneration. Aβ is cytotoxic and disrupts several cytoprotective pathways. Recent literature has demonstrated that certain cytochrome P450 (CYP) products are neuroprotective, including epoxide metabolites of arachidonic acid (AA), epoxyeicosatrienoic acids (EETs). The action of Aβ with respect to regionally produced EETs in the brain has yet to be defined. Epoxygenases metabolize AA into four regioisomers of EETs (14,15-, 11,12-, 8,9- and 5,6-EET). EETs are rapidly degraded into dihydroxyeicosatrienoic acids (DiHETEs) by soluble epoxide hydrolase (sEH). To determine the effect of Aβ on the epoxygenase activity in different regions of the brain, microsomes were prepared from the cerebrum and cerebellum of adult Sprague-Dawley rats and incubated with 1 and 10 μM Aβ for 30 min after which epoxygenase activity assay was performed. Mass spectrometry indicated that incubation with Aβ reduced 14,15-EET production by 30% as compared to vehicle in the cerebrum, but not in the cerebellum. When we separated the cerebrum into cortex and hippocampus, significant decrease in the production of total EETs and DiHETEs were seen in presence of Aβ (81% and 74%) in the cortex. Moreover, 11,12-EET production was decreased to ∼70% of vehicle in both cortex and hippocampus. Epoxygenase activity in the cultured astrocytes and neurons also showed reduction in total EET and DiHETE production (to 80% and ∼70% of vehicle respectively) in presence of Aβ. Altogether, our data suggest that Aβ reduces epoxygenase activity differentially in a region-specific and cell-specific manner. The reduction of cytoprotective EETs by Aβ in the cerebrum may make it more prone to degeneration than the cerebellum. Further understanding of these interactions will improve our ability to protect against the pathology of Alzheimer's disease.

Topics: 8,11,14-Eicosatrienoic Acid; Alzheimer Disease; Amyloid beta-Peptides; Animals; Animals, Newborn; Brain; Cytochrome P-450 CYP2J2; Cytochrome P-450 Enzyme Inhibitors; Cytochrome P-450 Enzyme System; Disease Models, Animal; Male; Neuroprotective Agents; Primary Cell Culture; Rats; Rats, Sprague-Dawley

Soluble epoxide hydrolase (sEH) metabolizes epoxyeicosatrienoic acids (EETs), primarily 14,15-EET. EETs are derived from arachidonic acid via P-450 epoxygenases and are cardioprotective. We tested the hypothesis that sEH deficiency and pharmacological inhibition elicit tolerance to ischemia via EET-mediated STAT3 signaling in vitro and in vivo. In addition, the relevance of single nucleotide polymorphisms (SNPs) of EPHX2 (the gene encoding sEH) on tolerance to oxygen and glucose deprivation and reoxygenation and glucose repletion (OGD/RGR) was assessed in male C57BL\\6J (WT) or sEH knockout (sEHKO) cardiomyocytes by using transactivator of transcription (TAT)-mediated transduction with sEH mutant proteins. Cell death and hydrolase activity was lower in Arg287Gln EPHX2 mutants vs. nontransduced controls. Excess 14,15-EET and SEH inhibition did not improve cell survival in Arg287Gln mutants. In WT cells, the putative EET receptor antagonist, 14,15-EEZE, abolished the effect of 14,15-EET and sEH inhibition. Cotreatment with 14,15-EET and SEH inhibition did not provide increased protection. In vitro, STAT3 inhibition blocked 14,15-EET cytoprotection, but not the effect of SEH inhibition. However, STAT3 small interfering RNA (siRNA) abolished cytoprotection by 14,15-EET and sEH inhibition, but cells pretreated with JAK2 siRNA remained protected. In vivo, STAT3 inhibition abolished 14,15-EET-mediated infarct size reduction. In summary, the Arg287Gln mutation is associated with improved tolerance against ischemia in vitro, and inhibition of sEH preserves cardiomyocyte viability following OGD/RGR via an EET-dependent mechanism. In vivo and in vitro, 14,15-EET-mediated protection is mediated in part by STAT3.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Cell Survival; Cells, Cultured; Disease Models, Animal; Epoxide Hydrolases; Janus Kinases; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Mutation; Myocardial Reperfusion Injury; Myocytes, Cardiac; Polymorphism, Single Nucleotide; Signal Transduction; STAT3 Transcription Factor

The widespread use of statins for hypercholesterolemia has uncovered pleiotropic anti-inflammatory properties that were unexpected based on the drugs' original design; yet, mechanisms for these protective actions remain uncertain. In this study lovastatin triggered biosynthesis of the anti-inflammatory and pro-resolving mediator 15-epi-lipoxin A(4) (15-epi-LXA(4)). During interactions between human neutrophils and airway epithelial cells, the statin-induced increase in 15-epi-LXA(4) was associated with increased 14,15-epoxyeicosatrienoic acid (14,15-EET) generation. When added to activated neutrophils, 14,15-EET enhanced 15-epi-LXA(4) biosynthesis. In a murine model of airway mucosal injury and inflammation, lovastatin increased 15-epi-LXA(4) formation in vivo and markedly decreased acute lung inflammation. Administration of 15-epi-LXA(4) also inhibited lung inflammation in an additive manner with lovastatin. Together, these results indicate that statin-triggered 15-epi-LXA(4) generation during human leukocyte-airway epithelial cell interactions is an endogenous mechanism for statin-mediated tissue protection at mucosal surfaces that may also be relevant in the statins' ability to stimulate the resolution of inflammation.

Topics: 8,11,14-Eicosatrienoic Acid; Acute Disease; Animals; Anticholesteremic Agents; Cell Line; Disease Models, Animal; Female; Humans; Hypercholesterolemia; Inflammation; Inflammation Mediators; Lipoxins; Lovastatin; Male; Mice; Neutrophils; Pneumonia; Respiratory Mucosa

We previously demonstrated that several epoxyeicosatrienoic acids (EETs) produce reductions in myocardial infarct size in rats and dogs. Since a recent study demonstrated the release of opioids in mediating the antinociceptive effect of 14,15-EET, we hypothesized that endogenous opioids may also be involved in mediating the cardioprotective effect of the EETs. To test this hypothesis, we used an in vivo rat model of infarction and a rat Langendorff model. In the infarct model, hearts were subjected to 30 min occlusion of the left coronary artery and 2 h reperfusion. Animals were treated with 11,12-EET or 14,15-EET (2.5 mg/kg) alone 15 min before occlusion or with opioid antagonists [naloxone, naltrindole, nor-binaltorphimine (nor-BNI), and d-Phe-Cys-Tyr-d-Trp-Om-Thr-Pen-Thr-NH(2) (CTOP), a nonselective, a selective delta, a selective kappa, and a selective mu receptor antagonist, respectively] 10 min before EET administration. In four separate groups, antiserum to Met- and Leu-enkephalin and dynorphin-A-(1-17) was administered 50 min before the 11,12-EET administration. Infarct size expressed as a percent of the area at risk (IS/AAR) was 63.5 + or - 1.2, 45.3 + or - 1.0, and 40.9 + or - 1.2% for control, 11,12-EET, and 14,15-EET, respectively. The protective effects of 11,12-EET were abolished by pretreatment with either naloxone (60.5 + or - 1.8%), naltrindole (60.8 + or - 1.0%), nor-BNI (62.3 + or - 2.8%), or Met-enkephalin antiserum (63.2 + or - 1.7%) but not CTOP (42.0 + or - 3.0%). In isolated heart experiments, 11,12-EET was administered to the perfusate 15 min before 20 min global ischemia followed by 45 min reperfusion in control hearts or in those pretreated with pertussis toxin (48 h). 11,12-EET increased the recovery of left ventricular developed pressure from 33 + or - 1 to 45 + or - 6% (P < 0.05) and reduced IS/AAR from 37 + or - 4 to 20 + or - 3% (P < 0.05). Both pertussis toxin and naloxone abolished these beneficial effects of 11,12-EET. Taken together, these results suggest that the major cardioprotective effects of the EETs depend on activation of a G(i/o) protein-coupled delta- and/or kappa-opioid receptor.

Topics: 8,11,14-Eicosatrienoic Acid; Analgesics, Opioid; Animals; Disease Models, Animal; GTP-Binding Protein alpha Subunits, Gi-Go; Male; Myocardial Infarction; Myocardial Reperfusion Injury; Naloxone; Naltrexone; Narcotic Antagonists; Rats; Rats, Sprague-Dawley; Receptors, Opioid; Somatostatin

15-Lipoxygenase (15-LO-1) metabolizes arachidonic acid (AA) to 11,12,15-trihydroxyeicosatrienoic acids (THETAs) and 15-hydroxy-11,12-epoxyeicosatrienoic acids (HEETA) that dilate rabbit arteries. Increased endothelial 15-LO-1 expression enhances arterial relaxations to agonists. We tested the effect of hypoxia on 15-LO-1 expression, THETA and HEETA synthesis, and relaxations in rabbit arteries. The incubation of rabbit aortic endothelial cells and isolated aortas in 0.7% O(2) increased 15-LO-1 expression. Rabbits were housed in a hypoxic atmosphere of 12% O(2) for 5 days. 15-LO-1 expression increased in the endothelium of the arteries of rabbits in 12% O(2) compared with room air. THETA and HEETA synthesis was also enhanced in aortas and mesenteric arteries. AA hyperpolarized the smooth muscle cells in indomethacin- and phenylephrine-treated mesenteric arteries of hypoxic rabbits from -29.4 +/- 1 to -50.1 +/- 3 mV. The hyperpolarization to AA was less in arteries of normoxic rabbits (from -26.0 +/- 2 to -37 +/- 2 mV). This AA-induced hyperpolarization was inhibited by the 15-LO inhibitor BW-755C. Nitric oxide and prostaglandin-independent maximum relaxations to acetylcholine (79.7 +/- 2%) and AA (38.3 +/- 4%) were enhanced in mesenteric arteries from hypoxic rabbits compared with the normoxic rabbits (49.7 +/- 6% and 19.9 +/- 2%, respectively). These relaxations were inhibited by BW-755C and nordihydroguaiaretic acid. Therefore, hypoxia increased the relaxations to agonists in the rabbit mesenteric arteries by enhancing endothelial 15-LO-1 expression and synthesis of the hyperpolarizing factors THETA and HEETA.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Arachidonate 15-Lipoxygenase; Arachidonic Acid; Arteries; Biological Factors; Cyclooxygenase Inhibitors; Disease Models, Animal; Dose-Response Relationship, Drug; Endothelium, Vascular; Hypoxia; Lipoxygenase Inhibitors; Male; Membrane Potentials; Muscle, Smooth, Vascular; Nitric Oxide; Rabbits; RNA, Messenger; Time Factors; Tunica Intima; Vasoconstriction; Vasoconstrictor Agents; Vasodilation; Vasodilator Agents

20-Hydroxyeicosatetraenoic acid (20-HETE) has been implicated as a potential mediator in epithelial cell proliferation and cyst formation in polycystic kidney disease (PKD). In the present study, we studied the effects of chronic blockade of 20-HETE synthesis in an orthologous rodent model of autosomal recessive polycystic kidney disease (ARPKD), the PCK rat. RT-PCR analysis indicated that the expression of CYP4A1, CYP4A2, CYP4A3, and CYP4A8 mRNA was increased two- to fourfold in cystic PCK compared with noncystic Sprague-Dawley rat kidneys. Daily administration of a 20-HETE synthesis inhibitor, HET-0016 (10 mg x kg(-1) x day(-1) ip) for 4-7 wk significantly reduced kidney size by 24% from 4.95 +/- 0.19 g in vehicle-treated PCK rats to 3.76 +/- 0.15 g (n = 4). Collecting tubule morphometric cystic indices were reduced in HET-0016-treated PCK rats (2.1 +/- 0.2; n = 4) compared with vehicle-treated PCK rats (4.4 +/- 0.1; n = 4). The cellular mechanism by which 20-HETE may play a role in cyst formation has not been well characterized, but there was a significantly lower (P < 0.05) level of intracellular cAMP and decreased phosphorylation (activation) of ERK1/2 protein in PCK rat kidneys (n = 3) treated with HET-0016 . These studies indicate a potential role of 20-HETE in cyst formation in the orthologous rodent PCK model of ARPKD.

Topics: 8,11,14-Eicosatrienoic Acid; Amidines; Animals; Cyclic AMP; Cytochrome P-450 CYP4A; Disease Models, Animal; Enzyme Activation; Epoxy Compounds; Extracellular Signal-Regulated MAP Kinases; Hydroxyeicosatetraenoic Acids; Kidney; Male; Microsomes; Organ Size; Polycystic Kidney, Autosomal Recessive; Protein Isoforms; Rats; Rats, Sprague-Dawley; RNA, Messenger

Soluble epoxide hydrolase (sEH), a key enzyme in the metabolism of vasodilator eicosanoids called epoxyeicosatrienoic acids (EETs), is sexually dimorphic and suppressed by estrogen. We determined if the sex difference in blood flow during focal cerebral ischemia is linked to sEH. Soluble epoxide hydrolase expression in brain, hydrolase activity in cerebral vessels, and plasma 14,15-dihydroxyeicosatrienoic acid (14,15-DHET) were determined in male and female wild-type (WT) and sEH knockout (sEHKO) mice. Male, female, and ovariectomized female WT and sEHKO mice were subjected to 2-h middle cerebral artery occlusion (MCAO) and infarct size was measured at 24 h of reperfusion. Laser-Doppler cortical perfusion during MCAO was compared among groups and differences in cortical blood flow rates were confirmed using in vivo quantitative optical microangiography. Cerebrovascular expression and activity of sEH and plasma 14,15-DHET were lower in WT female than male mice, and blood flow during MCAO was higher and infarct size was smaller in WT female compared with male mice. Sex differences in cerebral blood flow and ischemic damage were abolished after ovariectomy and were absent in sEHKO mice. We conclude that sEH is an important mechanism underlying sex-linked differences in blood flow and brain damage after cerebral ischemia.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Blood Flow Velocity; Blotting, Western; Brain Ischemia; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Epoxide Hydrolases; Estrogens; Female; Infarction, Middle Cerebral Artery; Male; Mice; Mice, Knockout; Ovariectomy; Sex Characteristics; Solubility

Soluble epoxide hydrolase (sEH) metabolizes epoxyeicosatrienoic acids (EETs) to dihydroxyeicosatrienoic acids. EETs are formed from arachidonic acid during myocardial ischemia and play a protective role against ischemic cell death. Deletion of sEH has been shown to be protective against myocardial ischemia in the isolated heart preparation. We tested the hypothesis that sEH inactivation by targeted gene deletion or pharmacological inhibition reduces infarct size (I) after regional myocardial ischemia-reperfusion injury in vivo. Male C57BL\\6J wild-type or sEH knockout mice were subjected to 40 min of left coronary artery (LCA) occlusion and 2 h of reperfusion. Wild-type mice were injected intraperitoneally with 12-(3-adamantan-1-yl-ureido)-dodecanoic acid butyl ester (AUDA-BE), a sEH inhibitor, 30 min before LCA occlusion or during ischemia 10 min before reperfusion. 14,15-EET, the main substrate for sEH, was administered intravenously 15 min before LCA occlusion or during ischemia 5 min before reperfusion. The EET antagonist 14,15-epoxyeicosa-5(Z)-enoic acid (EEZE) was given intravenously 15 min before reperfusion. Area at risk (AAR) and I were assessed using fluorescent microspheres and triphenyltetrazolium chloride, and I was expressed as I/AAR. I was significantly reduced in animals treated with AUDA-BE or 14,15-EET, independent of the time of administration. The cardioprotective effect of AUDA-BE was abolished by the EET antagonist 14,15-EEZE. Immunohistochemistry revealed abundant sEH protein expression in left ventricular tissue. Strategies to increase 14,15-EET, including sEH inactivation, may represent a novel therapeutic approach for cardioprotection against myocardial ischemia-reperfusion injury.

Topics: 8,11,14-Eicosatrienoic Acid; Adamantane; Animals; Disease Models, Animal; Enzyme Inhibitors; Epoxide Hydrolases; Female; Gene Deletion; Heart Ventricles; Injections, Intraperitoneal; Injections, Intravenous; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Myocardial Infarction; Myocardial Reperfusion Injury; Myocytes, Cardiac; Time Factors; Urea

Disorders of the metabolism of essential fatty acids (EFAs) are related to atopic dermatitis (AD). Concentrations of dihomo-gamma-linolenic acid (DGLA), an EFA, in the serum of AD patients are lower than those in healthy volunteers. Recently we developed a fermented DGLA oil, and examined whether oral administration of DGLA prevents development of dermatitis in NC/Nga mice, which spontaneously develop human AD-like skin lesions. NC/Nga mice were fed a diet either containing or not containing DGLA for 8 weeks under in air-uncontrolled conventional circumstances. Clinical skin severity scores were significantly lower in mice fed DGLA than in mice not fed it. Scratching behavior and plasma total IgE levels were also reduced in the DGLA group, in association with histological improvement. DGLA suppressed clinical severity of skin lesions dose-dependently, with an increase in DGLA contents in phospholipids of skin, spleen, and plasma. Discontinuation of DGLA administration resulted in the onset of dermatitis and a decrease in DGLA contents in skin, spleen, and plasma. These findings indicate that oral administration of DGLA effectively prevents the development of AD in NC/Nga mice, and that DGLA in phospholipids is a compound of key importance in the development and prevention of dermatitis.

Topics: 8,11,14-Eicosatrienoic Acid; Administration, Oral; Animals; Dermatitis, Atopic; Diet; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Immunoglobulin E; Male; Mice; Mice, Inbred Strains; Severity of Illness Index; Skin; Skin Tests

Cytochrome P450-derived epoxyeicosatrienoic acids (EET) are biologically active metabolites of arachidonic acid that have potent effects on renal vascular reactivity and tubular ion transport and have been implicated in the control of blood pressure. EETs are hydrolyzed to their less active diols, dihydroxyeicosatrienoic acids (DHET), by the enzyme soluble epoxide hydrolase (sEH). 1,3-dicyclohexylurea (DCU), a potent sEH inhibitor, lowers systemic blood pressure in spontaneously hypertensive rats when dosed intraperitoneally. However, DCU has poor aqueous solubility, posing a challenge for in vivo oral delivery. To overcome this limitation, we formulated DCU in a nanosuspension using wet milling. Milling reduced particle size, increasing the total surface area by approximately 40-fold. In rats chronically infused with angiotensin II, the DCU nanosuspension administered orally twice daily for 4 days produced plasma exposures an order of magnitude greater than unmilled DCU and lowered blood pressure by nearly 30 mmHg. Consistent with the mechanism of sEH inhibition, DCU increased plasma 14,15-EET and decreased plasma 14,15-DHET levels. These data confirm the antihypertensive effect of sEH inhibition and demonstrate that greatly enhanced exposure of a low-solubility compound is achievable by oral delivery using a nanoparticle drug delivery system.

Topics: 8,11,14-Eicosatrienoic Acid; Administration, Oral; Animals; Blood Pressure; Chromatography, Liquid; Disease Models, Animal; Epoxide Hydrolases; Hypertension; Male; Nanoparticles; Particle Size; Rats; Rats, Sprague-Dawley; Solubility; Suspensions; Tandem Mass Spectrometry; Urea

Previously, we demonstrated (17) that 11,12- and 14,15-epoxyeicosatrienoic acids (EETs) produce marked reductions in myocardial infarct size. Although it is assumed that this cardioprotective effect of the EETs is due to a specific interaction with a membrane-bound receptor, no evidence has indicated that novel EET antagonists selectively block the EET actions in dogs. Our goals were to investigate the effects of 11,12- and 14,15-EET, the soluble epoxide hydrolase inhibitor, 12-(3-adamantan-1-yl-ureido)-dodecanoic acid (AUDA), and the putative selective EET antagonist, 14,15-epoxyeicosa-5(Z)-enoic acid (14,15-EEZE), on infarct size of barbital anesthetized dogs subjected to 60 min of coronary artery occlusion and 3 h of reperfusion. Furthermore, the effect of 14,15-EEZE on the cardioprotective actions of the selective mitochondrial ATP-sensitive potassium channel opener diazoxide was investigated. Both 11,12- and 14,15-EET markedly reduced infarct size [expressed as a percentage of the area at risk (IS/AAR)] from 21.8 +/- 1.6% (vehicle) to 8.7 +/- 2.2 and 9.4 +/- 1.3%, respectively. Similarly, AUDA significantly reduced IS/AAR from 21.8 +/- 1.6 to 14.4 +/- 1.2% (low dose) and 9.4 +/- 1.8% (high dose), respectively. Interestingly, the combination of the low dose of AUDA with 14,15-EET reduced IS/AAR to 5.8 +/- 1.6% (P < 0.05), further than either drug alone. Diazoxide also reduced IS/AAR significantly (10.2 +/- 1.9%). In contrast, 14,15-EEZE had no effect on IS/AAR by itself (21.0 +/- 3.6%), but completely abolished the effect of 11,12-EET (17.8 +/- 1.4%) and 14,15-EET (19.2 +/- 2.4%) and AUDA (19.3 +/- 1.6%), but not that of diazoxide (10.4 +/- 1.4%). These results suggest that activation of the EET pathway, acting on a putative receptor, by exogenous EETs or indirectly by blocking EET metabolism, produced marked cardioprotection, and the combination of these two approaches resulted in a synergistic effect. These data also suggest that 14,15-EEZE is not blocking the mitochondrial ATP-sensitive potassium channel as a mechanism for antagonizing the cardioprotective effects of the EETs.

Topics: 8,11,14-Eicosatrienoic Acid; Adamantane; Animals; Blood Pressure; Cardiovascular Agents; Coronary Circulation; Diazoxide; Disease Models, Animal; Dogs; Dose-Response Relationship, Drug; Enzyme Inhibitors; Epoxide Hydrolases; Female; Heart Rate; Lauric Acids; Male; Mitochondria, Heart; Myocardial Infarction; Myocardium; Potassium Channels

We have chemically synthesized several stable analogs of the naturally occurring hepoxilins, 12-LO products derived from arachidonic acid, which we found to have promising actions in a variety of test systems of disease. The analogs, PBTs, afford chemical and biological stability to the hepoxilin molecule. This article reviews some of our latest observations with the PBTs in the areas of inflammation (inhibition of the bleomycin-evoked lung fibrosis in mice in vivo), platelet aggregation (antagonism of the thromboxane receptor in human platelets in vitro) and thrombosis (inhibitors in vivo), and cancer (apoptosis of the human leukemia cell line, K562 in vitro and in vivo). The demonstration that the PBTs are active in vivo suggests that they can serve as a platform for their further development as novel therapeutics in disease.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Anti-Inflammatory Agents; Antineoplastic Agents; Apoptosis; Bleomycin; Blood Platelets; Disease Models, Animal; Fibrinolytic Agents; Humans; K562 Cells; Leukemia, Experimental; Lung; Mice; Platelet Aggregation; Pulmonary Fibrosis

Shell-less culture of chick chorioallantoic membrane (CAM) of developing chicken embryos is a useful model to evaluate the effects of vascular agents. We assessed the response of CAM vessels to epoxyeicosatrienoic acids (EETs), derivatives of the essential fatty acid arachidonic acid, that have a number of important biological functions, including dilation of microvessels in the coronary, cerebral, renal, and mesenteric circulations. Three of four regioisomers of EETs, 14,15-, 11,12-, and 8,9-EET, induced a characteristic dose-dependent acute hyperemia within 4 min after application on 10-day-old CAMs. This response was marked in early stages of development (between days 8 and 10), but the frequency and intensity of the response were reduced after 11 days of development. Histological examination demonstrated that the hyperemia was not due to extravasation of erythrocytes. However, many capillaries were distended and contained densely packed erythrocytes as compared to uniformly arranged vessels and erythrocytes in untreated CAMs. Transmission electron microscopy showed the basal laminae surrounding capillaries remained intact, similar to those in vehicle-treated or untreated CAM tissue. The hyperemia was specific to EETs since we did not observe it to be induced by other vasodilators such as nitric oxide or prostacyclin. In conclusion, we report a novel vascular response to EETs using the CAM as an in vivo model. These lipids specifically distend a subset of capillaries in a dose- and development-dependent manner.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Capillaries; Chick Embryo; Chorioallantoic Membrane; Culture Techniques; Disease Models, Animal; Dose-Response Relationship, Drug; Hyperemia; Neovascularization, Physiologic; Nitric Oxide Donors; Time Factors; Vasodilator Agents; Vitelline Membrane

In chronic heart failure, the lung endothelial permeability response to angiotensin II or thapsigargin-induced store depletion is ablated, although the mechanisms are not understood.. To determine whether the ablated permeability response to store depletion during heart failure was due to impaired expression of store operated Ca2+ channels in lung endothelium.. Heart failure was induced by aortocaval fistula in rats. Permeability was measured in isolated lungs using the filtration coefficient and a low Ca2+/Ca2+ add-back strategy to identify the component of the permeability response dependent on Ca2+ entry.. In fistulas, right ventricular mass and left ventricular end diastolic pressure were increased and left ventricular shortening fraction decreased compared with shams. Thapsigargin-induced store depletion increased lung endothelial permeability in shams, but not in fistulas. Permeability increased in both groups after the Ca2+ ionophore A23187 or 14,15-epoxyeicosatrienoic acid, independent of store depletion. A diacylglycerol analog had no impact on permeability. Increased distance between the endoplasmic reticulum and the plasmalemmal membrane was ruled out as a mechanism for the loss of the permeability response to store depletion. Endothelial expression of the endoplasmic reticulum Ca2+ ATPase was not altered in fistulas compared with shams, whereas the store-operated canonical transient receptor potential channels 1, 3, and 4 were downregulated in extraalveolar vessel endothelium.. We conclude that the adaptive mechanism limiting store depletion-induced endothelial lung injury in the aortocaval model of heart failure involves downregulation of store-operated Ca2+ channels.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Calcimycin; Calcium Channels; Calcium-Transporting ATPases; Disease Models, Animal; Endothelium; Enzyme Inhibitors; Heart Failure; Ionophores; Lung; Permeability; Rats; Sarcoplasmic Reticulum Calcium-Transporting ATPases; Thapsigargin; Tissue Culture Techniques; Vasodilator Agents

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; 8,11,14-Eicosatrienoic Acid; Animals; Burns; Contact Lenses; Cytochrome P-450 Enzyme System; Disease Models, Animal; Eicosanoids; Eye Injuries; Inflammation; Neovascularization, Physiologic; Rabbits; Wound Healing

Bleomycin has been suggested to incite plasma extravasation and influx of inflammatory cells leading to pulmonary fibrosis. We hypothesized that stable analogs of the 12-lipoxygenase product, hepoxilin, may attenuate these effects. We initially investigated the effects of the four hepoxilin analogs (PBT-1 to -4) coadministered intradermally with bleomycin and found that PBT-1 and -2 significantly opposed the vascular permeability effects of bleomycin in rat skin. We subsequently tested the hepoxilin analogs for their actions in opposing the intratracheal bleomycin-evoked acute inflammatory phase of lung fibrosis in the mouse, characterized by a marked accumulation of macrophages and an increase in the rate of collagen synthesis and deposition. We found that the bleomycin-evoked effects on macrophage influx were inhibited by all the hepoxilin analogs (PBT-1, -3, and -4 > PBT-2) administered i.p. for 8 days. Increased total lung collagen was completely abrogated by PBT-1 and -2, whereas PBT-3 and -4 had little effect. A dose-response study with PBT-1 indicated that the effective dose for inhibition of bleomycin-induced inflammatory and histological changes was below 10 microg/day. These studies demonstrate an in vivo action of stable analogs of hepoxilin and support an effect on inflammation and vascular permeability from these novel compounds, especially for PBT-1.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Bleomycin; Disease Models, Animal; Male; Mice; Mice, Inbred CBA; Pulmonary Fibrosis; Rats; Rats, Wistar

This study was designed to examine the contribution of lipoxygenase products to mechanisms of vascular contraction and elevated blood pressure in rats with aortic coarctation-induced hypertension. In cytosolic fractions of aortae taken from hypertensive rats, 12-lipoxygenase protein was increased as compared to normotensive controls. Aortic rings from hypertensive, but not from normotensive rats, exhibited a basal tone which was reduced 74+/-12 and 71+/-22%, respectively, by the lipoxygenase inhibitors cinnamyl-3,4-dihydroxy-alpha-cyanocinnamate (CDC, 10(-5) mol/L) and 5,8,11-eicosatriynoic acid (ETI, 10(-5) mol/L). CDC (8 mg/kg s.c.) did not affect the blood pressure of normotensive rats but decreased that of hypertensive rats from 182+/-6 to 151+/-10 mm Hg. The blood pressure lowering effect of CDC was blunted in hypertensive rats pretreated with indomethacin or antibodies against 5,6-dihydro-prostaglandin I2. These data suggest contribution of lipoxygenase-derived products to mechanisms underlying aortic smooth muscle basal tone and elevated blood pressure in rats with aortic coarctation-induced hypertension. The vasodepressor effect of CDC depends on a mechanism involving vasodilatory prostaglandins.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; 8,11,14-Eicosatrienoic Acid; Animals; Aorta, Thoracic; Blood Pressure; Caffeic Acids; Cyclooxygenase Inhibitors; Disease Models, Animal; Epoprostenol; Hypertension; Indomethacin; Leukotrienes; Lipoxygenase; Lipoxygenase Inhibitors; Male; Muscle, Smooth, Vascular; Prostaglandins, Synthetic; Rats; Rats, Sprague-Dawley; Vasoconstriction; Vasoconstrictor Agents

The present studies have demonstrated that topical application of a low concentration of eicosa-5,8,11-trienoic acid (a 20:3,n9 fatty acid previously reported to inhibit competitively the activity of the sheep vesicular cyclooxygenase) to skin of normal fed hairless mice produced severe scaly dermatosis which is characterized by marked hyperplasia and acanthosis of the epidermal layer. The precise mechanism of this induction of scaly dermatosis is presently unclear. It is nonetheless interesting that the treatment of skin with similar concentrations of other unsaturated fatty acids produced no visible or histologic effects. Furthermore, endogenous levels of arachidonic acid in epidermal phospholipid and triglyceride fractions were shown to increase significantly (p < 0.01) in skin treated with the 20:3,n9 fatty acid while the endogenous level of PGE2 in the same tissue decreased markedly. This latter observation is consistent at least in part, with a previous report from this laboratory in which the 20:3,n9 fatty acid inhibited in vitro the activity of the sheep vesicular cyclooxygenase (the rate limiting enzyme in the transformation of arachidonic acid into the prostaglandin endoperoxides) although the increase in arachidonic acid may also reflect an increased incorporation of this fatty acid into the epidermal lipids by the hyperproliferative tissue. Evaluation of the proliferative status of 20:3,n9 fatty acid-treated skin showed a significant increase (p < 0.01) in labeling and mitotic indices. The use of this potentially endogenous fatty acid may be a useful tool for further investigations of hyperproliferative skin diseases where dietary deficiency of essential fatty acids does not exist.

Topics: 8,11,14-Eicosatrienoic Acid; Animals; Arachidonic Acids; Cell Division; Disease Models, Animal; Fatty Acids, Unsaturated; Female; Lipids; Mice; Mice, Nude; Prostaglandins E; Skin; Skin Diseases