euk-134 and Disease-Models--Animal

Radiation-induced normal tissue toxicity is closely linked to endothelial cell (EC) damage and dysfunction (acute effects). However, the underlying mechanisms of radiation-induced adverse late effects with respect to the vascular compartment remain elusive, and no causative radioprotective treatment is available to date.. The importance of injury to EC for radiation-induced late toxicity in lungs after whole thorax irradiation (WTI) was investigated using a mouse model of radiation-induced pneumopathy. We show that WTI induces EC loss as long-term complication, which is accompanied by the development of fibrosis. Adoptive transfer of mesenchymal stem cells (MSCs) either derived from bone marrow or aorta (vascular wall-resident MSCs) in the early phase after irradiation limited the radiation-induced EC loss and fibrosis progression. Furthermore, MSC-derived culture supernatants rescued the radiation-induced reduction in viability and long-term survival of cultured lung EC. We further identified the antioxidant enzyme superoxide dismutase 1 (SOD1) as a MSC-secreted factor. Importantly, MSC treatment restored the radiation-induced reduction of SOD1 levels after WTI. A similar protective effect was achieved by using the SOD-mimetic EUK134, suggesting that MSC-derived SOD1 is involved in the protective action of MSC, presumably through paracrine signaling.. In this study, we explored the therapeutic potential of MSC therapy to prevent radiation-induced EC loss (late effect) and identified the protective mechanisms of MSC action.. Adoptive transfer of MSCs early after irradiation counteracts radiation-induced vascular damage and EC loss as late adverse effects. The high activity of vascular wall-derived MSCs for radioprotection may be due to their tissue-specific action. Antioxid. Redox Signal. 26, 563-582.

Topics: Animals; Disease Models, Animal; Endothelial Cells; Fibroblasts; Fibrosis; Gene Expression; Lung; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice; Organometallic Compounds; Phenotype; Radiation Injuries; Radiation Injuries, Experimental; Salicylates; Superoxide Dismutase-1

Patients with pulmonary hypertension (PH) suffer from inspiratory insufficiency, which has been associated with intrinsic contractile dysfunction in diaphragm muscle. Here, we examined the role of redox stress in PH-induced diaphragm weakness by using the novel antioxidant, EUK-134. Male Wistar rats were randomly divided into control (CNT), CNT + EUK-134 (CNT + EUK), monocrotaline-induced PH (PH), and PH + EUK groups. PH was induced by a single intraperitoneal injection of monocrotaline (60 mg/kg body weight). EUK-134 (3 mg/kg body weight/day), a cell permeable mimetic of superoxide dismutase (SOD) and catalase, was daily intraperitoneally administered starting one day after induction of PH. After four weeks, diaphragm muscles were excised for mechanical and biochemical analyses. There was a decrease in specific tetanic force in diaphragm bundles from the PH group, which was accompanied by increases in: protein expression of NADPH oxidase 2/gp91phox, SOD2, and catalase; 3-nitrotyrosine content and aggregation of actin; glutathione oxidation. Treatment with EUK-134 prevented the force decrease and the actin modifications in PH diaphragm bundles. These data show that redox stress plays a pivotal role in PH-induced diaphragm weakness. Thus, antioxidant treatment can be a promising strategy for PH patients with inspiratory failure.

Topics: Actins; Animals; Antioxidants; Biomimetic Materials; Catalase; Diaphragm; Disease Models, Animal; Glutathione; Hypertension, Pulmonary; Male; Monocrotaline; Muscle Contraction; Muscle Weakness; Organometallic Compounds; Oxidative Stress; Rats; Rats, Wistar; Salicylates; Superoxide Dismutase

TNF-α inhibitor reportedly protects against myocardial ischemia/reperfusion (MI/R) injury. It can also increase Notch1 expression in inflammatory bowel disease, revealing the regulation of Notch1 signaling by TNF-α inhibitor. However, the interaction between TNF-α inhibitor and Notch1 signaling in MI/R remains unclear. This study aimed to determine the involvement of TNF-α inhibitor with Notch1 in MI/R and delineate the related mechanism. Notch1-specific small interfering RNA (20 μg) or Jagged1 (a Notch ligand, 12 μg) was delivered through intramyocardial injection. Forty-eight hours after injection, mice received 30 min of myocardial ischemia followed by 3 h (for cell apoptosis and oxidative/nitrative stress) or 24h (for infarct size and cardiac function) of reperfusion. Ten minutes before reperfusion, mice randomly received an intraperitoneal injection of vehicle, etanercept, diphenyleneiodonium, 1400W, or EUK134. Finally, downregulation of Notch1 significantly reversed the alleviation of MI/R injury induced by etanercept, as evidenced by enlarged myocardial infarct size, suppressed cardiac function, and increased myocardial apoptosis. Moreover, Notch1 blockade increased the expression of inducible NO synthase (iNOS) and gp(91)(phox), enhanced NO and superoxide production, and accelerated their cytotoxic reaction product, peroxynitrite. Furthermore, NADPH inhibition with diphenyleneiodonium or iNOS suppression with 1400W mitigated the aggravation of MI/R injury induced by Notch1 downregulation in mice treated with etanercept. Additionally, either Notch1 activation with Jagged1 or peroxynitrite decomposition with EUK134 reduced nitrotyrosine content and attenuated MI/R injury. These data indicate that MI/R injury can be attenuated by TNF-α inhibitor, partly via Notch1 signaling-mediated suppression of oxidative/nitrative stress.

Topics: Animals; Calcium-Binding Proteins; Disease Models, Animal; Down-Regulation; Enzyme Activation; Etanercept; Intercellular Signaling Peptides and Proteins; Jagged-1 Protein; Membrane Proteins; Mice; Mice, Inbred C57BL; Myocardial Infarction; Myocardial Reperfusion Injury; Myocardium; NADP; Nitric Oxide Synthase Type II; Onium Compounds; Organometallic Compounds; Oxidative Stress; Peroxynitrous Acid; Reactive Nitrogen Species; Reactive Oxygen Species; Receptor, Notch1; RNA Interference; RNA, Small Interfering; Salicylates; Serrate-Jagged Proteins; Tumor Necrosis Factor-alpha

Reduced mechanical loading during bedrest, spaceflight, and casting, causes rapid morphological changes in skeletal muscle: fiber atrophy and reduction of slow-twitch fibers. An emerging signaling event in response to unloading is the translocation of neuronal nitric oxide synthase (nNOSμ) from the sarcolemma to the cytosol. We used EUK-134, a cell-permeable mimetic of superoxide dismutase and catalase, to test the role of redox signaling in nNOSμ translocation and muscle fiber atrophy as a result of short-term (54 h) hindlimb unloading. Fischer-344 rats were divided into ambulatory control, hindlimb-unloaded (HU), and hindlimb-unloaded + EUK-134 (HU-EUK) groups. EUK-134 mitigated the unloading-induced phenotype, including muscle fiber atrophy and muscle fiber-type shift from slow to fast. nNOSμ immunolocalization at the sarcolemma of the soleus was reduced with HU, while nNOSμ protein content in the cytosol increased with unloading. Translocation of nNOS from the sarcolemma to cytosol was virtually abolished by EUK-134. EUK-134 also mitigated dephosphorylation at Thr-32 of FoxO3a during HU. Hindlimb unloading elevated oxidative stress (4-hydroxynonenal) and increased sarcolemmal localization of Nox2 subunits gp91phox (Nox2) and p47phox, effects normalized by EUK-134. Thus, our findings are consistent with the hypothesis that oxidative stress triggers nNOSμ translocation from the sarcolemma and FoxO3a dephosphorylation as an early event during mechanical unloading. Thus, redox signaling may serve as a biological switch for nNOS to initiate morphological changes in skeletal muscle fibers.

Topics: Aldehydes; Animals; Antioxidants; Cytosol; Disease Models, Animal; Forkhead Box Protein O3; Forkhead Transcription Factors; Hindlimb Suspension; Membrane Glycoproteins; Muscle Fibers, Fast-Twitch; Muscle Fibers, Skeletal; Muscle Fibers, Slow-Twitch; Muscular Atrophy; NADPH Oxidase 2; NADPH Oxidases; Nitric Oxide Synthase Type I; Organometallic Compounds; Oxidation-Reduction; Oxidative Stress; Phenotype; Phosphorylation; Protein Transport; Rats; Rats, Inbred F344; Salicylates; Sarcolemma; Signal Transduction; Time Factors

Nonalcoholic steatohepatitis (NASH), a progressive stage of nonalcoholic fatty liver disease (NAFLD), is characterized by steatosis (accumulation of triacylglycerols within hepatocytes) along with inflammation and ballooning degeneration. It has been suggested that oxidative stress may play an important role in the progress of NAFLD to NASH. The aim of present study was to determine whether antioxidant supplementations using EUK-8, EUK-134 and vitamin C could improve the biochemical and histological abnormalities associated with diet-induced NASH in rats.. NASH was induced in male N-Mary rats by feeding a methionine - choline deficient (MCD) diet. The rats were fed either normal chow or MCD diet for 10 weeks. After NASH development, the MCD-fed rats were randomly divided into four groups of six: the NASH group that received MCD diet, the EUK-8 group which was fed MCD diet plus EUK-8, the EUK-134 group which was fed MCD diet plus EUK-134 and the vitamin C group which received MCD diet plus vitamin C. EUK-8, EUK-134 and vitamin C (30 mg/kg body weight/day) were administered by gavage for eight weeks.. Treatment of MCD-fed rats with salens reduced the sera aminotransferases, cholesterol, low density lipoprotein contents, the extent of lipid peroxidation and protein carbonylation whereas the HDL-C cholesterol levels were significantly increased. In addition, EUK-8 and EUK-134 improved steatosis, ballooning degeneration and inflammation in liver of MCD-fed rats.. Antioxidant (EUK-8, EUK-134 and vitamin C) supplementation reduces NASH-induced biochemical and histological abnormalities, pointing out that antioxidant strategy could be beneficial in treatment of NASH.

Topics: Animals; Ascorbic Acid; Choline Deficiency; Diet; Disease Models, Animal; Ethylenediamines; Fatty Liver; Humans; Lipid Peroxidation; Methionine; Organometallic Compounds; Oxidative Stress; Rats; Salicylates

Although there often is a clinical co-incidence of increased adiposity and obstructive sleep apnea, each factor is independently associated with elevated oxidative stress.. We hypothesized that overweight rats exposed to simulated sleep apnea would develop exacerbated oxidative stress leading to impaired endothelium-dependent vasodilation.. Rats were fed either a chow or high-fat diet (HFD; 60% kcal from fat) for 6 weeks. During the final 14 days of each diet, animals were exposed to either air or eucapnic intermittent hypoxia (E-IH) to simulate sleep apnea.. Rats exposed to either E-IH or HFD alone showed increases of 161 and 176%, respectively, in oxidative stress (measured as thiobarbituric acid-reactive substances) compared to chow + air controls. However, oxidative stress was lower following combined HFD + E-IH treatment (132% of chow + air controls) compared to each individual treatment. All three treatment groups, chow + E-IH, HFD + air and HFD + E-IH, had increased blood pressure (144.5 ± 4.4, 148.2 ± 5.6, and 136.2 ± 2.0 mm Hg, respectively, vs. chow + air: 123 ± 2.0 mm Hg) and attenuated acetylcholine (ACh)-mediated vasodilation (78.3, 72.7, and 78.2% of the chow + air response at the highest dose of ACh) compared to chow + air controls. Combined HFD and E-IH treatment did not further impair vasodilation compared to chow + E-IH alone. Vasodilatory responses were normalized by the antioxidant EUK-134 in each treatment group.. Increased adiposity and simulated sleep apnea impair endothelium- dependent vasodilation through enhanced generation of reactive oxygen species (ROS). However, the combined treatment does not exacerbate either ROS generation or vascular dysfunction observed with HFD or E-IH alone.

Topics: Acetylcholine; Adipose Tissue; Animals; Blood Pressure; Dietary Fats; Disease Models, Animal; Humans; Male; Organometallic Compounds; Overweight; Oxidative Stress; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Salicylates; Sleep Apnea, Obstructive; Vasodilation

Nitrous oxide (N2O, laughing gas) has been used as an anaesthetic and analgesic for almost two centuries, but its cellular targets remain unclear. Here, we present a molecular mechanism of nitrous oxide's selective inhibition of CaV3.2 low-voltage-activated (T-type) calcium channels in pain pathways. Using site-directed mutagenesis and metal chelators such as diethylenetriamine pentaacetic acid and deferoxamine, we reveal that a unique histidine at position 191 of CaV3.2 participates in a critical metal binding site, which may in turn interact with N2O to produce reactive oxygen species (ROS). These free radicals are then likely to oxidize H191 of CaV3.2 in a localized metal-catalysed oxidation reaction. Evidence of hydrogen peroxide and free radical intermediates is given in that N2O inhibition of CaV3.2 channels is attenuated when H2O2 is neutralized by catalase. We also use the adrenochrome test as an indicator of ROS in vitro in the presence of N2O and iron. Ensuing in vivo studies indicate that mice lacking CaV3.2 channels display decreased analgesia to N2O in response to formalin-induced inflammatory pain. Furthermore, a superoxide dismutase and catalase mimetic, EUK-134, diminished pain responses to formalin in wild-type mice, but EUK-134 and N2O analgesia were not additive. This suggests that reduced ROS levels led to decreased inflammation, but without the presence of ROS, N2O was not able to provide additional analgesia. These findings reveal a novel mechanism of interaction between N2O and ion channels, furthering our understanding of this widely used analgesic in pain processing.

Topics: Adrenochrome; Analgesics, Non-Narcotic; Animals; Calcium Channel Blockers; Calcium Channels, T-Type; Catalase; Chelating Agents; Deferoxamine; Disease Models, Animal; Female; Ganglia, Spinal; HEK293 Cells; Histidine; Humans; Hydrogen Peroxide; Male; Membrane Potentials; Mice; Mice, Inbred C57BL; Mice, Knockout; Mutagenesis, Site-Directed; Nitrous Oxide; Organometallic Compounds; Oxidation-Reduction; Pain; Pentetic Acid; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Salicylates; Signal Transduction; Time Factors; Transfection

ROS have been implicated in the development of pathological ventricular hypertrophy and the ensuing contractile dysfunction. Using the rat monocrotaline (MCT) model of pulmonary arterial hypertension (PAH), we recently reported oxidative stress in the failing right ventricle (RV) with no such stress in the left ventricle of the same hearts. We used the antioxidant EUK-134 to assess the role of ROS in the pathological remodeling and dysfunction of the RV. PAH was induced by an injection of MCT (80 mg/kg, day 0), treatment with EUK-134 (25 mg/kg, once every 2 days) of control and MCT-injected animals [congestive heart failure (CHF) group] was started on day 10, and animals were analyzed on day 22. EUK-134 treatment of the CHF group attenuated cardiomyocyte hypertrophy and associated changes in mRNA expression (myosin heavy chain-beta and deiodinase type 3). It also reduced RV oxidative stress and proapoptotic signaling and prevented interstitial fibrosis. Cardiac MRI showed that ROS scavenging did not affect the 37% increase in end-diastolic volume of the RV in the CHF relative to the control group, but the threefold increase in end-systolic volume was reduced by 42% in the EUK-134-treated CHF group. The improved systolic function was confirmed using echocardiography by an assessment of tricuspid annular plane systolic excursion. These data indicate an important role of ROS in RV cardiomyocyte hypertrophy and contractile dysfunction due to PAH and show the potential of EUK-class antioxidants as complementary therapeutics in the treatment of RV dysfunction in PAH.

Topics: Animals; Antioxidants; Disease Models, Animal; Heart Failure; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Male; Monocrotaline; Organometallic Compounds; Rats; Rats, Wistar; Reactive Oxygen Species; Salicylates; Ventricular Dysfunction, Right; Ventricular Remodeling

EUK-134 is one of the most promising of the superoxide dismutase (SOD)/catalase mimetic compounds. The antioxidant effects of EUK-134 were tested in a rat model of paraquat pneumotoxicity.. Male Wistar rats (n = 72) were divided into three groups: group 1, controls; group 2, paraquat alone; group 3, paraquat + EUK-134. Paraquat dichloride was administered per os at a dose of 80 mg/kg. EUK-134 was injected intraperitoneally at 10 mg/kg 2 h before the paraquat and again 4 h later at 5 mg/kg.. On days 1, 3 and 5 after treatment with paraquat alone the LDH activity increased (P = 0.0001, P = 0.00001 and P = 0.03, respectively), and the total protein content increased (P = 0.00002, P = 0.001 and P = 0.01, respectively). The levels of acid phosphatase (AcP) in BAL fluid increased on days 1 and 3 (P = 0.006 and P = 0.04). In lung homogenates paraquat alone increased SOD activity on day 1 and decreased it on days 3 and 5. Combined treatment with paraquat and EUK-134 elevated LDH activity on day 3 (significantly less than paraquat alone) and day 5, elevated the total protein content on day 5 only, and did not change AcP activity. The combination of both agents did not alter SOD activity and decreased catalase activity on day 5 significantly less than treatment with paraquat alone (P = 0.05).. EUK-134, a superoxide dismutase/catalase mimetic compound decreased the pneumotoxic effect of paraquat in rats.

Topics: Animals; Antioxidants; Bronchoalveolar Lavage Fluid; Catalase; Disease Models, Animal; Herbicides; Lung; Male; Organometallic Compounds; Oxidative Stress; Paraquat; Rats; Rats, Wistar; Salicylates; Superoxide Dismutase