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Sample records for oxygenase ho-1 protects

  1. Epigallocatechin-3-gallate (EGCG) protects skin cells from ionizing radiation via heme oxygenase-1 (HO-1) overexpression

    Zhu Wei; Xu Jing; Ge Yangyang

    2014-01-01

    Epigallocatechin-3-gallate (EGCG), the major polyphenolic constituent of green tea, is a potent antioxidant and free radical scavenger that may have therapeutic applications for the treatment of many disorders. Radiation therapy is widely used for the treatment of various types of cancers; however, radiation-induced skin injury remains a serious concern. EGCG has not yet been reported as protecting skin cells against ionizing radiation. In the present study, we investigated whether EGCG confers cytoprotection against ionizing radiation. We found that, compared with the control, pretreatment with EGCG significantly enhanced the viability of human skin cells that were irradiated with X-rays, and decreased apoptosis induced by X-ray irradiation. Mito-Tracker assay showed that EGCG suppressed the damage to mitochondria induced by ionizing radiation via upregulation of SOD2. Reactive oxygen species (ROS) in HaCaT cells were significantly reduced when pretreated with EGCG before irradiation. Radiation-induced γH2AX foci, which are representative of DNA double-strand breaks, were decreased by pretreatment with EGCG. Furthermore, EGCG induced the expression of the cytoprotective molecule heme oxygenase-1 (HO-1) in a dose-dependent manner via transcriptional activation. HO-1 knockdown or treatment with the HO-1 inhibitor tin protoporphyrin (SnPPIX) reversed the protective role of EGCG, indicating an important role for HO-1. These results suggest that EGCG offers a new strategy for protecting skin against ionizing radiation. (author)

  2. Spatiotemporal expression of heme oxygenase-1 detected by in vivo bioluminescence after hepatic ischemia in HO-1/luc mice

    Su, Huawei; van Dam, Gooitzen M.; Buis, Carlijn I.; Visser, Dorien S.; Hesselink, Jan Willem; Schuurs, Theo A.; Leuvenink, Henri G. D.; Contag, Christopher H.; Porte, Robert J.

    Upregulation of heme oxygenase-1 (HO-1) has been proposed as a critical mechanism protecting against cellular stress during liver transplantation, providing a potential target for new therapeutic interventions. We investigated the feasibility of in vivo bioluminescence imaging (BLI) to noninvasively

  3. AN ELISA ASSAY FOR HEME OXYGENASE (HO-1)

    An ELISA assay for heme oxygenase (HO-l ) Abstract A double antibody capture ELISA for the HO-l protein has been developed to separately quantitate HO-I protein. The use of 2.5% NP40 detergent greatly assists in freeing HO-l protein from membranes and/or other cel...

  4. Heme oxygenase-1 (HO-1) inhibits postmyocardial infarct remodeling and restores ventricular function.

    Liu, Xiaoli; Pachori, Alok S; Ward, Christopher A; Davis, J Paul; Gnecchi, Massimiliano; Kong, Deling; Zhang, Lunan; Murduck, Jared; Yet, Shaw-Fang; Perrella, Mark A; Pratt, Richard E; Dzau, Victor J; Melo, Luis G

    2006-02-01

    We reported previously that predelivery of the anti-oxidant gene heme oxygenase-1 (HO-1) to the heart by adeno associated virus (AAV) markedly reduces injury after acute myocardial infarction (MI). However, the effect of HO-1 gene delivery on postinfarction recovery has not been investigated. In the current study, we assessed the effect of HO-1 gene delivery on post-MI left ventricle (LV) remodeling and function using echocardiographic imaging and histomorphometric approaches. Two groups of Sprague-Dawley rats were injected with 4 x 10(11) particles of AAV-LacZ (control) or AAV-hHO-1 in the LV wall. Eight wk after gene transfer, the animals were subjected to 30 min of ischemia by ligation of left anterior descending artery (LAD) followed by reperfusion. Echocardiographic measurements were obtained in a blinded fashion prior and at 1.5 and 3 months after I/R. Ejection fraction (EF) was reduced by 13% and 40% in the HO-1 and LacZ groups, respectively at 1.5 months after MI. Three months after MI, EF recovered fully in the HO-1, but only partially in the LacZ-treated animals. Post-MI LV dimensions were markedly increased and the anterior wall was markedly thinned in the LacZ-treated animals compared with the HO-1-treated animals. Significant myocardial scarring and fibrosis were observed in the LacZ-group in association with elevated levels of interstitial collagen I and III and MMP-2 activity. Post-MI myofibroblast accumulation was reduced in the HO-1-treated animals, and retroviral overexpression of HO-1 reduced proliferation of isolated cardiac fibroblasts. Our data indicate that rAAV-HO-1 gene transfer markedly reduces fibrosis and ventricular remodeling and restores LV function and chamber dimensions after myocardial infarction.

  5. Paracrine action of HO-1-modified mesenchymal stem cells mediates cardiac protection and functional improvement.

    Zeng, Bin; Ren, Xiaofeng; Lin, Guosheng; Zhu, Chengang; Chen, Honglei; Yin, Jiechao; Jiang, Hong; Yang, Bo; Ding, Danhua

    2008-10-01

    The aim has been to determine whether the supernatants of mesenchymal stem cells (MSCs) transfected with adenovirus carrying human heme oxygenase-1 (hHO-1) gene protect cardiomyocytes from ischemic injury. We have found that hHO-1 infected MSCs (hHO-1-MSCs) increased expression of hHO-1 protein. Apoptosis of cultured hHO-1-MSCs exposed to hypoxia was suppressed. Several cytokines, including HGF, bFGF, TGF-beta, VEGF and IL-1beta, were produced by hHO-1-MSCs, some being significantly enhanced under hypoxia stimulation. Meanwhile, those cytokines reduced caspase-3 level and activity in cultured adult rat ventricular cardiomyocytes (ARVCs) exposed to hypoxia. Supernatants obtained from hHO-1-MSCs improved left ventricular function, limited myocardial infarct size, increased microvessel density, and inhibited apoptosis of cardiomyocytes in rat myocardial infarction. It can be concluded hHO-1-modified MSCs prevent myocardial cell injury via secretion of paracrine-acting mediators.

  6. Expression and characterization of truncated human heme oxygenase (hHO-1) and a fusion protein of hHO-1 with human cytochrome P450 reductase.

    Wilks, A; Black, S M; Miller, W L; Ortiz de Montellano, P R

    1995-04-04

    A human heme oxygenase (hHO-1) gene without the sequence coding for the last 23 amino acids has been expressed in Escherichia coli behind the pho A promoter. The truncated enzyme is obtained in high yields as a soluble, catalytically-active protein, making it available for the first time for detailed mechanistic studies. The purified, truncated hHO-1/heme complex is spectroscopically indistinguishable from that of the rat enzyme and converts heme to biliverdin when reconstituted with rat liver cytochrome P450 reductase. A self-sufficient heme oxygenase system has been obtained by fusing the truncated hHO-1 gene to the gene for human cytochrome P450 reductase without the sequence coding for the 20 amino acid membrane binding domain. Expression of the fusion protein in pCWori+ yields a protein that only requires NADPH for catalytic turnover. The failure of exogenous cytochrome P450 reductase to stimulate turnover and the insensitivity of the catalytic rate toward changes in ionic strength establish that electrons are transferred intramolecularly between the reductase and heme oxygenase domains of the fusion protein. The Vmax for the fusion protein is 2.5 times higher than that for the reconstituted system. Therefore, either the covalent tether does not interfere with normal docking and electron transfer between the flavin and heme domains or alternative but equally efficient electron transfer pathways are available that do not require specific docking.

  7. Heme oxygenase-1 (HO-1 expression in prostate cancer cells modulates the oxidative response in bone cells.

    Mercedes Ferrando

    Full Text Available Prostate cancer (PCa is a leading cause of death among males. It is currently estimated that inflammatory responses are linked to 15-20% of all deaths from cancer worldwide. PCa is dominated by complications arising from metastasis to the bone where the tumor cells interact with the bone microenvironment impairing the balance between bone formation and degradation. However, the molecular nature of this interaction is not completely understood. Heme oxygenase-1 (HO-1 counteracts oxidative damage and inflammation. Previous studies from our laboratory showed that HO-1 is implicated in PCa, demonstrating that endogenous HO-1 inhibits bone derived-prostate cancer cells proliferation, invasion and migration and decreases tumor growth and angiogenesis in vivo. The aim of this work was to analyze the impact of HO-1 modulated PCa cells on osteoblasts proliferation in vitro and on bone remodeling in vivo. Using a co-culture system of PC3 cells with primary mice osteoblasts (PMOs, we demonstrated that HO-1 pharmacological induction (hemin treatment abrogated the diminution of PMOs proliferation induced by PCa cells and decreased the expression of osteoclast-modulating factors in osteoblasts. No changes were detected in the expression of genes involved in osteoblasts differentiation. However, co-culture of hemin pre-treated PC3 cells (PC3 Hem with PMOs provoked an oxidative status and activated FoxO signaling in osteoblasts. The percentage of active osteoblasts positive for HO-1 increased in calvarias explants co-cultured with PC3 Hem cells. Nuclear HO-1 expression was detected in tumors generated by in vivo bone injection of HO-1 stable transfected PC3 (PC3HO-1 cells in the femur of SCID mice. These results suggest that HO-1 has the potential to modify the bone microenvironment impacting on PCa bone metastasis.

  8. Relationship of Heme Oxygenase-1 (HO-1 Level with Onset and Severity in Normotensive Pregnancy and Severe Preeclampsia

    John Johannes Wantania

    2016-08-01

    Full Text Available Background: Preeclampsia still becomes a major problem in pregnancies. Various evidences showed that heme oxygenase-1 (HO-1 is very important in pregnancy. This study aims to understand the relationship of heme oxygenase-1 level with onset and severity in normotensive pregnancy and severe preeclampsia. Methods: This is a cross sectional analytic comparative study, the subjects consisted of 26 patients with normotensive pregnancies and 26 patients with severe preeclampsia. Blood samples from women with < 34 / ≥ 34 weeks’ normotensive pregnancies and women with severe preeclampsia were taken. HO-1 ELISA kit used to quantitate heme oxygenase-1 level in samples. Results: The level of heme oxygenase-1 in normotensive pregnant women < 34 weeks lower than severe preeclampsia pregnant women < 34 weeks (3.28 ± 0.46 ng/mL vs 4.20 ± 0.64 ng/mL, p=0.003, respectively. The median level of heme oxygenase-1 in normotensive pregnant women ≥ 34 weeks was 2.96 (2.41–4.39 ng/mL, while severe preeclampsia pregnant women ≥ 34 weeks was 3.52 (2.88–5.43 ng/mL, (p=0.040. The median level of heme oxygenase-1 in normotensive pregnant women was 3.04 (2.41–4.39 ng/mL, while severe preeclampsia pregnant women was 3.68 (2.88–5.67 ng/mL, (p=0.001. Conclusions: There is correlation between the incidence of severe preeclampsia with heme oxygenase-1 level in < 34 and ≥ 34 weeks of pregnancy. There is a significant difference between the level of heme oxygenase-1 in pregnant women with severe preeclampsia and in women with normotensive pregnancy. 

  9. Extratumoral Heme Oxygenase-1 (HO-1 Expressing Macrophages Likely Promote Primary and Metastatic Prostate Tumor Growth.

    Sofia Halin Bergström

    Full Text Available Aggressive tumors induce tumor-supporting changes in the benign parts of the prostate. One factor that has increased expression outside prostate tumors is hemoxygenase-1 (HO-1. To investigate HO-1 expression in more detail, we analyzed samples of tumor tissue and peritumoral normal prostate tissue from rats carrying cancers with different metastatic capacity, and human prostate cancer tissue samples from primary tumors and bone metastases. In rat prostate tumor samples, immunohistochemistry and quantitative RT-PCR showed that the main site of HO-1 synthesis was HO-1+ macrophages that accumulated in the tumor-bearing organ, and at the tumor-invasive front. Small metastatic tumors were considerably more effective in attracting HO-1+ macrophages than larger non-metastatic ones. In clinical samples, accumulation of HO-1+ macrophages was seen at the tumor invasive front, almost exclusively in high-grade tumors, and it correlated with the presence of bone metastases. HO-1+ macrophages, located at the tumor invasive front, were more abundant in bone metastases than in primary tumors. HO-1 expression in bone metastases was variable, and positively correlated with the expression of macrophage markers but negatively correlated with androgen receptor expression, suggesting that elevated HO-1 could be a marker for a subgroup of bone metastases. Together with another recent observation showing that selective knockout of HO-1 in macrophages reduced prostate tumor growth and metastatic capacity in animals, the results of this study suggest that extratumoral HO-1+ macrophages may have an important role in prostate cancer.

  10. Heme oxygenase and carbon monoxide protect from muscle dystrophy.

    Chan, Mun Chun; Ziegler, Olivia; Liu, Laura; Rowe, Glenn C; Das, Saumya; Otterbein, Leo E; Arany, Zoltan

    2016-11-28

    Duchenne muscle dystrophy (DMD) is one of the most common lethal genetic diseases of children worldwide and is 100% fatal. Steroids, the only therapy currently available, are marred by poor efficacy and a high side-effect profile. New therapeutic approaches are urgently needed. Here, we leverage PGC-1α, a powerful transcriptional coactivator known to protect against dystrophy in the mdx murine model of DMD, to search for novel mechanisms of protection against dystrophy. We identify heme oxygenase-1 (HO-1) as a potential novel target for the treatment of DMD. Expression of HO-1 is blunted in the muscles from the mdx murine model of DMD, and further reduction of HO-1 by genetic haploinsufficiency worsens muscle damage in mdx mice. Conversely, induction of HO-1 pharmacologically protects against muscle damage. Mechanistically, HO-1 degrades heme into biliverdin, releasing in the process ferrous iron and carbon monoxide (CO). We show that exposure to a safe low dose of CO protects against muscle damage in mdx mice, as does pharmacological treatment with CO-releasing molecules. These data identify HO-1 and CO as novel therapeutic agents for the treatment of DMD. Safety profiles and clinical testing of inhaled CO already exist, underscoring the translational potential of these observations.

  11. 50 Hz electric field effects on protein carbonyl (PCO), heme oxygenase-1 (HO-1) and hydroxyproline levels

    Ozgur, Elcin; Goknur, Guler; Seyhan, Nesrin

    2008-01-01

    Full text: Non-ionizing electromagnetic field (EMF) radiation sources, such as power lines and other Extremely Low Frequency (ELF) sources have become one of the most ubiquitous components of the spectrum of the human environment, and the possibility that they may have hazardous effects on human health is a major a public concern. Although it is well documented that EMFs have biological effects, the degree to which these exposures constitute a human health hazard is not clear yet. Today relation between production of oxidative stress resulted by reactive oxygen species and electrical stimulus, also the protective effects of antioxidant treatments are mentioned in many researches. In this study, it was aimed to determine both oxidation of proteins and protein collagen levels under 50 Hz 12 kV/m vertical Electric (E) Field exposure and the N-Acetylcysteine (NAC) administration which is a well-known antioxidant. To this end, protein carbonyl levels (PCO) as bio-markers of oxidative stress and Heme oxygenase-1 (HO-1), an enzyme that catalyzes the degradation of heme analyzed to figure out the protein oxidation. Hydroxyproline level, a major component of the protein collagen was measured in order to express the level of collagen in lung tissue. Guinea pigs, weighted 250-300 g, were used in the study. A total forty male guinea pigs were randomly divided into four groups which are composed of 10 guinea pigs each for groups: 1) Group I (Sham); 2) Group II (NAC-administrated group); 3) Group III (E Field Exposure group); 4) Group IV (NAC administrated + E Field exposed group). One week exposure period for 8 hours per daily was conducted for each exposure groups (Group III, Group IV ). The electric field exposure period was from 9 a.m. to 5 p.m. After the last exposure day, the guinea pigs were anesthetized by the injection of ketamine and xylazine. The guinea pigs were killed by decapitation. Statistical analyses were carried out using SPSS software (SPSS 11.5 for windows

  12. Heme oxygenase-1 expression protects the heart from acute injury caused by inducible Cre recombinase.

    Hull, Travis D; Bolisetty, Subhashini; DeAlmeida, Angela C; Litovsky, Silvio H; Prabhu, Sumanth D; Agarwal, Anupam; George, James F

    2013-08-01

    The protective effect of heme oxygenase-1 (HO-1) expression in cardiovascular disease has been previously demonstrated using transgenic animal models in which HO-1 is constitutively overexpressed in the heart. However, the temporal requirements for protection by HO-1 induction relative to injury have not been investigated, but are essential to employ HO-1 as a therapeutic strategy in human cardiovascular disease states. Therefore, we generated mice with cardiac-specific, tamoxifen (TAM)-inducible overexpression of a human HO-1 (hHO-1) transgene (myosin heavy chain (MHC)-HO-1 mice) by breeding mice with cardiac-specific expression of a TAM-inducible Cre recombinase (MHC-Cre mice), with mice containing an hHO-1 transgene preceded by a floxed-stop signal. MHC-HO-1 mice overexpress HO-1 mRNA and the enzymatically active protein following TAM administration (40 mg/kg body weight on 2 consecutive days). In MHC-Cre controls, TAM administration leads to severe, acute cardiac toxicity, cardiomyocyte necrosis, and 80% mortality by day 3. This cardiac toxicity is accompanied by a significant increase in inflammatory cells in the heart that are predominantly neutrophils. In MHC-HO-1 mice, HO-1 overexpression ameliorates the depression of cardiac function and high mortality rate observed in MHC-Cre mice following TAM administration and attenuates cardiomyocyte necrosis and neutrophil infiltration. These results highlight that HO-1 induction is sufficient to prevent the depression of cardiac function observed in mice with TAM-inducible Cre recombinase expression by protecting the heart from necrosis and neutrophil infiltration. These findings are important because MHC-Cre mice are widely used in cardiovascular research despite the limitations imposed by Cre-induced cardiac toxicity, and also because inflammation is an important pathological component of many human cardiovascular diseases.

  13. Heme oxygenase-1-mediated autophagy protects against pulmonary endothelial cell death and development of emphysema in cadmium-treated mice

    Surolia, Ranu; Karki, Suman; Kim, Hyunki; Yu, Zhihong; Kulkarni, Tejaswini; Mirov, Sergey B.; Carter, A. Brent; Rowe, Steven M.; Matalon, Sadis; Thannickal, Victor J.; Agarwal, Anupam

    2015-01-01

    Pulmonary exposure to cadmium, a major component of cigarette smoke, has a dramatic impact on lung function and the development of emphysema. Cigarette smoke exposure induces heme oxygenase-1 (HO-1), a cytoprotective enzyme. In this study, we employed a truncated mouse model of emphysema by intratracheal instillation of cadmium (CdCl2) solution (0.025% per 1 mg/kg body wt) in HO-1+/+, HO-1−/−, and overexpressing humanized HO-1 bacterial artificial chromosome (hHO-1BAC) mice. We evaluated the role of HO-1 in cadmium-induced emphysema in mice by analyzing histopathology, micro-computed tomography scans, and lung function tests. CdCl2-exposed HO-1−/− mice exhibited more severe emphysema compared with HO-1+/+ or hHO-1BAC mice. Loss of pulmonary endothelial cells (PECs) from the alveolar capillary membrane is recognized to be a target in emphysema. PECs from HO-1+/+, HO-1−/−, and hHO-1BAC were employed to define the underlying molecular mechanism for the protection from emphysema by HO-1. Electron microscopy, expression of autophagic markers (microtubule-associated protein 1B-light chain 3 II, autophagy protein 5, and Beclin1) and apoptotic marker (cleaved caspase 3) suggested induction of autophagy and apoptosis in PECs after CdCl2 treatment. CdCl2-treated HO-1−/− PECs exhibited downregulation of autophagic markers and significantly increased cleaved caspase 3 expression and activity (∼4-fold higher). Moreover, hHO-1BAC PECs demonstrated upregulated autophagy and absence of cleaved caspase 3 expression or activity. Pretreatment of HO-1+/+ PECs with rapamycin induced autophagy and resulted in reduced cell death upon cadmium treatment. Induction of autophagy following CdCl2 treatment was found to be protective from apoptotic cell death. HO-1 induced protective autophagy in PECs and mitigated cadmium-induced emphysema. PMID:26071551

  14. Heme oxygenase-1-mediated autophagy protects against pulmonary endothelial cell death and development of emphysema in cadmium-treated mice.

    Surolia, Ranu; Karki, Suman; Kim, Hyunki; Yu, Zhihong; Kulkarni, Tejaswini; Mirov, Sergey B; Carter, A Brent; Rowe, Steven M; Matalon, Sadis; Thannickal, Victor J; Agarwal, Anupam; Antony, Veena B

    2015-08-01

    Pulmonary exposure to cadmium, a major component of cigarette smoke, has a dramatic impact on lung function and the development of emphysema. Cigarette smoke exposure induces heme oxygenase-1 (HO-1), a cytoprotective enzyme. In this study, we employed a truncated mouse model of emphysema by intratracheal instillation of cadmium (CdCl2) solution (0.025% per 1 mg/kg body wt) in HO-1(+/+), HO-1(-/-), and overexpressing humanized HO-1 bacterial artificial chromosome (hHO-1BAC) mice. We evaluated the role of HO-1 in cadmium-induced emphysema in mice by analyzing histopathology, micro-computed tomography scans, and lung function tests. CdCl2-exposed HO-1(-/-) mice exhibited more severe emphysema compared with HO-1(+/+) or hHO-1BAC mice. Loss of pulmonary endothelial cells (PECs) from the alveolar capillary membrane is recognized to be a target in emphysema. PECs from HO-1(+/+), HO-1(-/-), and hHO-1BAC were employed to define the underlying molecular mechanism for the protection from emphysema by HO-1. Electron microscopy, expression of autophagic markers (microtubule-associated protein 1B-light chain 3 II, autophagy protein 5, and Beclin1) and apoptotic marker (cleaved caspase 3) suggested induction of autophagy and apoptosis in PECs after CdCl2 treatment. CdCl2-treated HO-1(-/-) PECs exhibited downregulation of autophagic markers and significantly increased cleaved caspase 3 expression and activity (∼4-fold higher). Moreover, hHO-1BAC PECs demonstrated upregulated autophagy and absence of cleaved caspase 3 expression or activity. Pretreatment of HO-1(+/+) PECs with rapamycin induced autophagy and resulted in reduced cell death upon cadmium treatment. Induction of autophagy following CdCl2 treatment was found to be protective from apoptotic cell death. HO-1 induced protective autophagy in PECs and mitigated cadmium-induced emphysema. Copyright © 2015 the American Physiological Society.

  15. Quercetin Protects Primary Human Osteoblasts Exposed to Cigarette Smoke through Activation of the Antioxidative Enzymes HO-1 and SOD-1

    Karl F. Braun

    2011-01-01

    Full Text Available Smokers frequently suffer from impaired fracture healing often due to poor bone quality and stability. Cigarette smoking harms bone cells and their homeostasis by increased formation of reactive oxygen species (ROS. The aim of this study was to investigate whether Quercetin, a naturally occurring antioxidant, can protect osteoblasts from the toxic effects of smoking. Human osteoblasts exposed to cigarette smoke medium (CSM rapidly produced ROS and their viability decreased concentration- and time-dependently. Co-, pre- and postincubation with Quercetin dose-dependently improved their viability. Quercetin increased the expression of the anti-oxidative enzymes heme-oxygenase- (HO- 1 and superoxide-dismutase- (SOD- 1. Inhibiting HO-1 activity abolished the protective effect of Quercetin. Our results demonstrate that CSM damages human osteoblasts by accumulation of ROS. Quercetin can diminish this damage by scavenging the radicals and by upregulating the expression of HO-1 and SOD-1. Thus, a dietary supplementation with Quercetin could improve bone matter, stability and even fracture healing in smokers.

  16. Protection from ischemic heart injury by a vigilant heme oxygenase-1 plasmid system.

    Tang, Yao Liang; Tang, Yi; Zhang, Y Clare; Qian, Keping; Shen, Leping; Phillips, M Ian

    2004-04-01

    Although human heme oxygenase-1 (hHO-1) could provide a useful approach for cellular protection in the ischemic heart, constitutive overexpression of hHO-1 may lead to unwanted side effects. To avoid this, we designed a hypoxia-regulated hHO-1 gene therapy system that can be switched on and off. This vigilant plasmid system is composed of myosin light chain-2v promoter and a gene switch that is based on an oxygen-dependent degradation domain from the hypoxia inducible factor-1-alpha. The vector can sense ischemia and switch on the hHO-1 gene system, specifically in the heart. In an in vivo experiment, the vigilant hHO-1 plasmid or saline was injected intramyocardially into myocardial infarction mice or sham operation mice. After gene transfer, expression of hHO-1 was only detected in the ischemic heart treated with vigilant hHO-1 plasmids. Masson trichrome staining showed significantly fewer fibrotic areas in vigilant hHO-1 plasmids-treated mice compared with saline control (43.0%+/-4.8% versus 62.5%+/-3.3%, PhHO-1 expression in peri-infarct border areas, concomitant with higher Bcl-2 levels and lower Bax, Bak, and caspase 3 levels in the ischemic myocardium compared with saline control. By use of a cardiac catheter, heart from vigilant hHO-1 plasmids-treated mice showed improved recovery of contractile and diastolic performance after myocardial infarction compared with saline control. This study documents the beneficial regulation and therapeutic potential of vigilant plasmid-mediated hHO-1 gene transfer. This novel gene transfer strategy can provide cardiac-specific protection from future repeated bouts of ischemic injury.

  17. Activation of the Nrf2/HO-1 Antioxidant Pathway Contributes to the Protective Effects of Lycium Barbarum Polysaccharides in the Rodent Retina after Ischemia-Reperfusion-Induced Damage

    Chang, Raymond Chuen-Chung; So, Kwok-Fai; Brecha, Nicholas C.; Pu, Mingliang

    2014-01-01

    Lycium barbarum polysaccharides (LBP), extracts from the wolfberries, are protective to retina after ischemia-reperfusion (I/R). The antioxidant response element (ARE)–mediated antioxidant pathway plays an important role in maintaining the redox status of the retina. Heme oxygenase-1 (HO-1), combined with potent AREs in its promoter, is a highly effective therapeutic target for the protection against neurodegenerative diseases, including I/R-induced retinal damage. The aim of our present study was to investigate whether the protective effect of LBP after I/R damage was mediated via activation of the Nrf2/HO-1-antioxidant pathway in the retina. Retinal I/R was induced by an increase in intraocular pressure to 130 mm Hg for 60 minutes. Prior to the induction of ischemia, rats were orally treated with either vehicle (PBS) or LBP (1 mg/kg) once a day for 1 week. For specific experiments, zinc protoporphyrin (ZnPP, 20 mg/kg), an HO-1 inhibitor, was intraperitoneally administered at 24 h prior to ischemia. The protective effects of LBP were evaluated by quantifying ganglion cell and amacrine cell survival, and by measuring cell apoptosis in the retinal layers. In addition, HO-1 expression was examined using Western blotting and immunofluorescence analyses. Cytosolic and nuclear Nrf2 was measured using immunofluorescent staining. LBP treatment significantly increased Nrf2 nuclear accumulation and HO-1 expression in the retina after I/R injury. Increased apoptosis and a decrease in the number of viable cells were observed in the ganglion cell layer (GCL) and inner nuclear layer (INL) in the I/R retina, which were reversed by LBP treatment. The HO-1 inhibitor, ZnPP, diminished the LBP treatment-induced protective effects in the retina after I/R. Taken together, these results suggested that LBP partially exerted its beneficial neuroprotective effects via the activation of Nrf2 and an increase in HO-1 protein expression. PMID:24400114

  18. Sulforaphane protects rodent retinas against ischemia-reperfusion injury through the activation of the Nrf2/HO-1 antioxidant pathway.

    Hong Pan

    Full Text Available Retinal ischemia-reperfusion (I/R injury induces oxidative stress, leukocyte infiltration, and neuronal cell death. Sulforaphane (SF, which can be obtained in cruciferous vegetables such as broccoli, exerts protective effects in response to oxidative stress in various tissues. These effects can be initiated through nuclear factor E2-related factor 2 (Nrf2-mediated induction of heme oxygenase-1 (HO-1. This investigation was designed to elucidate the neural protective mechanisms of SF in the retinal I/R rat model. Animals were intraperitoneally (i.p. injected with SF (12.5 mg/kg or vehicle (corn oil once a day for 7 consecutive days. Then, retinal I/R was made by elevating the intraocular pressure (IOP to 130 mmHg for 1 h. To determine if HO-1 was involved in the Nrf2 antioxidant pathway, rats were subjected to protoporphyrin IX zinc (II (ZnPP, 30 mg/kg, i.p. treatments at 24 h before retinal ischemia. The neuroprotective effects of SF were assessed by determining the morphology of the retina, counting the infiltrating inflammatory cells and the surviving retinal ganglion cells (RGCs and amacrine cells, and measuring apoptosis in the retinal layers. The expression of Nrf2 and HO-1 was studied by immunofluorescence analysis and western blotting. I/R induced a marked increase of ROS generation, caused pronounced inflammation, increased the apoptosis of RGCs and amacrine cells and caused the thinning of the inner retinal layer (IRL, and these effects were diminished or abolished by SF pretreatment. Meanwhile, SF pretreatment significantly elevated the nuclear accumulation of Nrf2 and the level of HO-1 expression in the I/R retinas; however, ZnPP reversed the protective effects of SF on I/R retinas. Together, we offer direct evidence that SF had protective effects on I/R retinas, which could be attributed, at least in part, to the activation of the Nrf2/HO-1 antioxidant pathway.

  19. Adenoviral transfer of the heme oxygenase-1 gene protects striatal astrocytes from heme-mediated oxidative injury.

    Teng, Zhi-Ping; Chen, Jing; Chau, Lee-Young; Galunic, Nicholas; Regan, Raymond F

    2004-11-01

    Heme oxygenase-1 (HO-1) is induced in the CNS after hemorrhage, and may have an effect on injury to surrounding tissue. Hemin, the preferred substrate of HO, is a neurotoxin that is present in intracranial hematomas. In a prior study, we observed that HO inhibitors increased the vulnerability of cultured cortical astrocytes to heme-mediated oxidative injury. To investigate the effect of HO more specifically, we used an adenoviral vector encoding the human HO-1 gene to specifically increase HO-1 expression. Incubation with 100 MOI of the HO-1 adenovirus (Adv-HHO-1) for 24 h increased both HO-1 protein and HO activity; a control adenovirus lacking the HO-1 gene had no effect. Using a DNA probe that was specific for human HO-1, 80.5 +/- 7.2% of astrocytes were observed to be infected by in situ hybridization. The cell death produced by 30-60 microM hemin was significantly reduced by pretreatment with 100 MOI Adv-HHO-1, as assessed by LDH release, propidium iodide exclusion, and MTT reduction assay. The threefold increase in cell protein oxidation produced by hemin was also attenuated in cultures pretreated with Adv-HHO-1. These results support the hypothesis that HO-1 protects astrocytes from heme-mediated oxidative injury. Specifically increasing astrocytic HO-1 by gene transfer may have a beneficial effect on hemorrhagic CNS injury.

  20. Heme oxygenase-1 protects endothelial cells from the toxicity of air pollutant chemicals

    Lawal, Akeem O.; Zhang, Min; Dittmar, Michael; Lulla, Aaron; Araujo, Jesus A.

    2015-01-01

    Diesel exhaust particles (DEPs) are a major component of diesel emissions, responsible for a large portion of their toxicity. In this study, we examined the toxic effects of DEPs on endothelial cells and the role of DEP-induced heme oxygenase-1 (HO-1) expression. Human microvascular endothelial cells (HMECs) were treated with an organic extract of DEPs from an automobile engine (A-DEP) or a forklift engine (F-DEP) for 1 and 4 h. ROS generation, cell viability, lactate dehydrogenase leakage, expression of HO-1, inflammatory genes, cell adhesion molecules and unfolded protein respone (UPR) gene were assessed. HO-1 expression and/or activity were inhibited by siRNA or tin protoporphyrin (Sn PPIX) and enhanced by an expression plasmid or cobalt protoporphyrin (CoPPIX). Exposure to 25 μg/ml of A-DEP and F-DEP significantly induced ROS production, cellular toxicity and greater levels of inflammatory and cellular adhesion molecules but to a different degree. Inhibition of HO-1 enzymatic activity with SnPPIX and silencing of the HO-1 gene by siRNA enhanced DEP-induced ROS production, further decreased cell viability and increased expression of inflammatory and cell adhesion molecules. On the other hand, overexpression of the HO-1 gene by a pcDNA 3.1D/V5-HO-1 plasmid significantly mitigated ROS production, increased cell survival and decreased the expression of inflammatory genes. HO-1 expression protected HMECs from DEP-induced prooxidative and proinflammatory effects. Modulation of HO-1 expression could potentially serve as a therapeutic target in an attempt to inhibit the cardiovascular effects of ambient PM. - Highlights: • We examined the role of HO-1 expression on diesel exhaust particle (DEP) in endothelial cells. • DEPs exert cytotoxic and inflammatory effects on human microvascular endothelial cells (HMECs). • DEPs induce HO-1 expression in HMECs. • HO-1 protects against the oxidative stress induced by DEps. • HO-1 attenuates the proinflammatory effects

  1. Heme oxygenase-1 protects endothelial cells from the toxicity of air pollutant chemicals

    Lawal, Akeem O.; Zhang, Min; Dittmar, Michael [Division of Cardiology, David Geffen School of Medicine, University of California, Los Angeles, 10833 Le Conte Avenue, CHS 43-264, Los Angeles, CA 90095 (United States); Lulla, Aaron [Division of Cardiology, David Geffen School of Medicine, University of California, Los Angeles, 10833 Le Conte Avenue, CHS 43-264, Los Angeles, CA 90095 (United States); Molecular Toxicology Interdepartmental Program, University of California, Los Angeles (United States); Araujo, Jesus A., E-mail: JAraujo@mednet.ucla.edu [Division of Cardiology, David Geffen School of Medicine, University of California, Los Angeles, 10833 Le Conte Avenue, CHS 43-264, Los Angeles, CA 90095 (United States); Molecular Toxicology Interdepartmental Program, University of California, Los Angeles (United States); Molecular Biology Institute, University of California, Los Angeles (United States)

    2015-05-01

    Diesel exhaust particles (DEPs) are a major component of diesel emissions, responsible for a large portion of their toxicity. In this study, we examined the toxic effects of DEPs on endothelial cells and the role of DEP-induced heme oxygenase-1 (HO-1) expression. Human microvascular endothelial cells (HMECs) were treated with an organic extract of DEPs from an automobile engine (A-DEP) or a forklift engine (F-DEP) for 1 and 4 h. ROS generation, cell viability, lactate dehydrogenase leakage, expression of HO-1, inflammatory genes, cell adhesion molecules and unfolded protein respone (UPR) gene were assessed. HO-1 expression and/or activity were inhibited by siRNA or tin protoporphyrin (Sn PPIX) and enhanced by an expression plasmid or cobalt protoporphyrin (CoPPIX). Exposure to 25 μg/ml of A-DEP and F-DEP significantly induced ROS production, cellular toxicity and greater levels of inflammatory and cellular adhesion molecules but to a different degree. Inhibition of HO-1 enzymatic activity with SnPPIX and silencing of the HO-1 gene by siRNA enhanced DEP-induced ROS production, further decreased cell viability and increased expression of inflammatory and cell adhesion molecules. On the other hand, overexpression of the HO-1 gene by a pcDNA 3.1D/V5-HO-1 plasmid significantly mitigated ROS production, increased cell survival and decreased the expression of inflammatory genes. HO-1 expression protected HMECs from DEP-induced prooxidative and proinflammatory effects. Modulation of HO-1 expression could potentially serve as a therapeutic target in an attempt to inhibit the cardiovascular effects of ambient PM. - Highlights: • We examined the role of HO-1 expression on diesel exhaust particle (DEP) in endothelial cells. • DEPs exert cytotoxic and inflammatory effects on human microvascular endothelial cells (HMECs). • DEPs induce HO-1 expression in HMECs. • HO-1 protects against the oxidative stress induced by DEps. • HO-1 attenuates the proinflammatory effects

  2. Heme oxygenase-1 enhances autophagy in podocytes as a protective mechanism against high glucose-induced apoptosis

    Dong, Chenglong [Department of Endocrinology, The Second Affiliated Hospital of Nanjing Medical University, Nanjing (China); Zheng, Haining [Department of Hyperbaric Oxygen, Nanjing General Hospital of Nanjing Military Command, Nanjing (China); Huang, Shanshan; You, Na; Xu, Jiarong; Ye, Xiaolong; Zhu, Qun; Feng, Yamin; You, Qiang; Miao, Heng [Department of Endocrinology, The Second Affiliated Hospital of Nanjing Medical University, Nanjing (China); Ding, Dafa, E-mail: dingdafa2004@aliyun.com [Department of Endocrinology, The Second Affiliated Hospital of Nanjing Medical University, Nanjing (China); Lu, Yibing, E-mail: luyibing2004@126.com [Department of Endocrinology, The Second Affiliated Hospital of Nanjing Medical University, Nanjing (China)

    2015-10-01

    Injury and loss of podocytes play vital roles in diabetic nephropathy progression. Emerging evidence suggests autophagy, which is induced by multiple stressors including hyperglycemia, plays a protective role. Meanwhile, heme oxygenase-1 (HO-1) possesses powerful anti-apoptotic properties. Therefore, we investigated the impact of autophagy on podocyte apoptosis under diabetic conditions and its association with HO-1. Mouse podocytes were cultured in vitro; apoptosis was detected by flow cytometry. Transmission electron microscopy and biochemical autophagic flux assays were used to measure the autophagy markers microtubule-associated protein 1 light chain 3-II (LC3-II) and beclin-1. LC3-II and beclin-1 expression peaked 12–24 h after exposing podocytes to high glucose. Inhibition of autophagy with 3-methyladenine or Beclin-1 siRNAs or Atg 5 siRNAs sensitized cells to apoptosis, suggesting autophagy is a survival mechanism. HO-1 inactivation inhibited autophagy, which aggravated podocyte injury in vitro. Hemin-induced autophagy also protected podocytes from hyperglycemia in vitro and was abrogated by HO-1 siRNA. Adenosine monophosphate-activated protein kinase phosphorylation was higher in hemin-treated and lower in HO-1 siRNA-treated podocytes. Suppression of AMPK activity reversed HO-1-mediated Beclin-1 upregulation and autophagy, indicating HO-1-mediated autophagy is AMPK dependent. These findings suggest HO-1 induction and regulation of autophagy are potential therapeutic targets for diabetic nephropathy. - Highlights: • High glucose leads to increased autophagy in podocytes at an early stage. • The early autophagic response protects against high glucose-induced apoptosis. • Heme oxygenase-1 enhances autophagy and decreases high glucose -mediated apoptosis. • Heme oxygenase-1 induces autophagy through the activation of AMPK.

  3. Heme oxygenase-1 enhances autophagy in podocytes as a protective mechanism against high glucose-induced apoptosis

    Dong, Chenglong; Zheng, Haining; Huang, Shanshan; You, Na; Xu, Jiarong; Ye, Xiaolong; Zhu, Qun; Feng, Yamin; You, Qiang; Miao, Heng; Ding, Dafa; Lu, Yibing

    2015-01-01

    Injury and loss of podocytes play vital roles in diabetic nephropathy progression. Emerging evidence suggests autophagy, which is induced by multiple stressors including hyperglycemia, plays a protective role. Meanwhile, heme oxygenase-1 (HO-1) possesses powerful anti-apoptotic properties. Therefore, we investigated the impact of autophagy on podocyte apoptosis under diabetic conditions and its association with HO-1. Mouse podocytes were cultured in vitro; apoptosis was detected by flow cytometry. Transmission electron microscopy and biochemical autophagic flux assays were used to measure the autophagy markers microtubule-associated protein 1 light chain 3-II (LC3-II) and beclin-1. LC3-II and beclin-1 expression peaked 12–24 h after exposing podocytes to high glucose. Inhibition of autophagy with 3-methyladenine or Beclin-1 siRNAs or Atg 5 siRNAs sensitized cells to apoptosis, suggesting autophagy is a survival mechanism. HO-1 inactivation inhibited autophagy, which aggravated podocyte injury in vitro. Hemin-induced autophagy also protected podocytes from hyperglycemia in vitro and was abrogated by HO-1 siRNA. Adenosine monophosphate-activated protein kinase phosphorylation was higher in hemin-treated and lower in HO-1 siRNA-treated podocytes. Suppression of AMPK activity reversed HO-1-mediated Beclin-1 upregulation and autophagy, indicating HO-1-mediated autophagy is AMPK dependent. These findings suggest HO-1 induction and regulation of autophagy are potential therapeutic targets for diabetic nephropathy. - Highlights: • High glucose leads to increased autophagy in podocytes at an early stage. • The early autophagic response protects against high glucose-induced apoptosis. • Heme oxygenase-1 enhances autophagy and decreases high glucose -mediated apoptosis. • Heme oxygenase-1 induces autophagy through the activation of AMPK

  4. Galantamine and carbon monoxide protect brain microvascular endothelial cells by heme oxygenase-1 induction

    Nakao, Atsunori; Kaczorowski, David J.; Zuckerbraun, Brian S.; Lei Jing; Faleo, Gaetano; Deguchi, Kentaro; McCurry, Kenneth R.; Billiar, Timothy R.; Kanno, Shinichi

    2008-01-01

    Galantamine, a reversible inhibitor of acetylcholine esterase (AChE), is a novel drug treatment for mild to moderate Alzheimer's disease and vascular dementia. Interestingly, it has been suggested that galantamine treatment is associated with more clinical benefit in patients with mild-to-moderate Alzheimer disease compared to other AChE inhibitors. We hypothesized that the protective effects of galantamine would involve induction of the protective gene, heme oxygenase-1 (HO-1), in addition to enhancement of the cholinergic system. Brain microvascular endothelial cells (mvECs) were isolated from spontaneous hypertensive rats. Galantamine significantly reduced H 2 O 2 -induced cell death of mvECs in association with HO-1 induction. These protective effects were completely reversed by nuclear factor-κB (NF-κB) inhibition or HO inhibition. Furthermore, galantamine failed to induce HO-1 in mvECs which lack inducible nitric oxide synthase (iNOS), supplementation of a nitric oxide (NO) donor or iNOS gene transfection on iNOS-deficient mvECs resulted in HO-1 induction with galantamine. These data suggest that the protective effects of galantamine require NF-κB activation and iNOS expression, in addition to HO-1. Likewise, carbon monoxide (CO), one of the byproducts of HO, up-regulated HO-1 and protected mvECs from oxidative stress in a similar manner. Our data demonstrate that galantamine mediates cytoprotective effects on mvECs through induction HO-1. This pharmacological action of galantamine may, at least in part, account for the superior clinical efficacy of galantamine in vascular dementia and Alzheimer disease

  5. Characterization of docosahexaenoic acid (DHA)-induced heme oxygenase-1 (HO-1) expression in human cancer cells: the importance of enhanced BTB and CNC homology 1 (Bach1) degradation.

    Wang, Shuai; Hannafon, Bethany N; Wolf, Roman F; Zhou, Jundong; Avery, Jori E; Wu, Jinchang; Lind, Stuart E; Ding, Wei-Qun

    2014-05-01

    The effect of docosahexaenoic acid (DHA) on heme oxygenase-1 (HO-1) expression in cancer cells has never been characterized. This study examines DHA-induced HO-1 expression in human cancer cell model systems. DHA enhanced HO-1 gene expression in a time- and concentration-dependent manner, with maximal induction at 21 h of treatment. This induction of HO-1 expression was confirmed in vivo using a xenograft nude mouse model fed a fish-oil-enriched diet. The increase in HO-1 gene transcription induced by DHA was significantly attenuated by the antioxidant N-acetyl cysteine, suggesting the involvement of oxidative stress. This was supported by direct measurement of lipid peroxide levels after DHA treatment. Using a human HO-1 gene promoter reporter construct, we identified two antioxidant response elements (AREs) that mediate the DHA-induced increase in HO-1 gene transcription. Knockdown of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) expression compromised the DHA-induced increase in HO-1 gene transcription, indicating the importance of the Nrf2 pathway in this event. However, the nuclear protein levels of Nrf2 remained unchanged upon DHA treatment. Further studies demonstrated that DHA reduces nuclear Bach1 protein expression by promoting its degradation and attenuates Bach1 binding to the AREs in the HO-1 gene promoter. In contrast, DHA enhanced Nrf2 binding to the AREs without affecting nuclear Nrf2 expression levels, indicating a new cellular mechanism that mediates DHA's induction of HO-1 gene transcription. To our knowledge, this is the first characterization of DHA-induced HO-1 expression in human malignant cells. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. Andrographolide protects liver cells from H2O2 induced cell death by upregulation of Nrf-2/HO-1 mediated via adenosine A2a receptor signalling.

    Mittal, Smriti P K; Khole, Swati; Jagadish, Nidhi; Ghosh, Debjani; Gadgil, Vijay; Sinkar, Vilas; Ghaskadbi, Saroj S

    2016-11-01

    Andrographolide, principle constituent of Andrographis paniculata Nees is used in traditional medicine in Southeast Asia and is known to exhibit various biological activities. Its antioxidant activity is due to its ability to activate one of the antioxidant enzymes, heme oxygenase-1 (HO-1) which is regulated transcriptionally through Nrf-2. However, molecular mechanism underlying activation of Nrf-2/HO-1 has not yet been clearly understood. Protective effect of andrographolide against H2O2 induced cell death, reactive oxygen species and lipid peroxidation was observed in HepG2 cells. Ability of andrographolide to modulate G-protein coupled receptor (GPCR) mediated signalling was determined using in silico docking and gene expression was analyzed by qRT-PCR, confocal microscopy and western blot analysis. We clearly show that andrographolide via adenosine A2A receptor signalling leads to activation of p38 MAP kinase, resulting in upregulation of Nrf-2, its translocation to nucleus and activation of HO-1. Additionally, it activates adenylate cyclase resulting in cAMP formation which in turn activates protein kinase A leading to inhibition of GSK-3β by phosphorylation. Inactivated GSK-3β leads to retention of Nrf-2 in the nucleus leading to sustained expression of HO-1 by binding to its antioxidant response element (ARE). Thus, andrographolide probably by binding to adenosine A2a receptor activates Nrf-2 transcription and also inhibits its exclusion from the nucleus by inactivating GSK-3β, together resulting in activation of HO-1. We speculate that andrographolide can be used as a therapeutic drug to combat oxidative stress implicated in pathogenesis of various diseases such as diabetes, osteoporosis, neurodegenerative diseases etc. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. Trimetazidine protects retinal ganglion cells from acute glaucoma via the Nrf2/Ho-1 pathway.

    Wan, Peixing; Su, Wenru; Zhang, Yingying; Li, Zhidong; Deng, Caibin; Zhuo, Yehong

    2017-09-15

    Acute glaucoma is one of the leading causes of irreversible vision impairment characterized by the rapid elevation of intraocular pressure (IOP) and consequent retinal ganglion cell (RGC) death. Oxidative stress and neuroinflammation have been considered critical for the pathogenesis of RGC death in acute glaucoma. Trimetazidine (TMZ), an anti-ischemic drug, possesses antioxidative and anti-inflammatory properties, contributing to its therapeutic potential in tissue damage. However, the role of TMZ in acute glaucoma and the underlying molecular mechanisms remain elusive. Here, we report that treatment with TMZ significantly attenuated retinal damage and RGC death in mice with acute glaucoma, with a significant decrease in reactive oxygen species (ROS) and inflammatory cytokine production in the retina. Furthermore, TMZ treatment directly decreased ROS production and rebalanced the intracellular redox state, thus contributing to the survival of RGCs in vitro TMZ treatment also reduced the production of inflammatory cytokines in vitro Mechanistically, the TMZ-mediated inhibition of apoptosis and inflammatory cytokine production in RGCs occurred via the regulation of the nuclear factor erythroid 2-related factor 2/heme oxygenase 1/caspase-8 pathway. Moreover, the TMZ-mediated neuroprotection in acute glaucoma was abrogated when an HO-1 inhibitor, SnPP, was used. Our findings identify potential mechanisms of RGC apoptosis and propose a novel therapeutic agent, TMZ, which exerts a precise neuroprotective effect against acute glaucoma. © 2017 The Author(s).

  8. EGR-1 regulates Ho-1 expression induced by cigarette smoke

    Chen, Huaqun; Wang, Lijuan; Gong, Tao; Yu, Yang; Zhu, Chunhua; Li, Fen; Wang, Li; Li, Chaojun

    2010-01-01

    As an anti-oxidant molecule, heme oxygenase-1 (HO-1) has been implicated in the protection of lung injury by cigarette smoke (CS). The mechanisms regulating its expression have not been defined. In this report, the role of early growth response 1 (EGR-1) in the regulation of Ho-1 expression was investigated. In C57BL/6 mice with CS exposure, HO-1 was greatly increased in bronchial epithelial cells and alveolar inflammatory cells. In primary cultured mouse lung fibroblasts and RAW264.7 cells exposed to cigarette smoke water extract (CSE), an increase in HO-1 protein level was detected. In addition, CSE induced HO-1 expression was decreased in Egr-1 deficient mouse embryo fibroblasts (Egr-1 -/- MEFs). Nuclear localization of EGR-1 was examined in mouse lung fibroblasts after exposure to CSE. Luciferase reporter activity assays showed that the enhancer region of the Ho-1 gene containing a proposed EGR-1 binding site was responsible for the induction of HO-1. A higher increase of alveolar mean linear intercept (Lm) was observed in lung tissues, and a larger increase in the number of total cells and monocytes/macrophages from bronchial alveolar lavage fluid was found in CS-exposed mice by loss of function of EGR-1 treatment. In summary, the present data demonstrate that EGR-1 plays a critical role in HO-1 production induced by CS.

  9. UVA-induced protection of skin through the induction of heme oxygenase-1.

    Xiang, Yuancai; Liu, Gang; Yang, Li; Zhong, Julia Li

    2011-12-01

    UVA (320-400 nm) and UVB (290-320 nm) are the major components of solar UV irradiation, which is associated with various pathological conditions. UVB causes direct damage to DNA of epidermal cells and is mainly responsible for erythema, immunosuppression, photoaging, and skin cancer. UVA has oxidizing properties that can cause damage or enhance UVB damaging effects on skin. On the other hand, UVA can also lead to high levels of heme oxygenase-1 (HO-1) expression of cells that can provide an antioxidant effect on skin as well as anti-inflammatory properties in mammals and rodents. Therefore, this review focuses on the potential protection of UVA wavebands for the skin immune response, instead of mechanisms that underlie UVA-induced damage. Also, the role of HO-1 in UVA-mediated protection against UVB-induced immunosuppression in skin will be summarized. Thus, this review facilitates further understanding of potential beneficial mechanisms of UVA irradiation, and using the longer UVA (UVA1, 340-400 nm) in combination with HO-1 for phototherapy and skin protection against sunlight exposure.

  10. Heme oxygenase-1 dependant pathway contributes to protection by tetramethylpyrazine against chronic hypoxic injury on medulla oblongata in rats.

    Ding, Yan; Hou, Xuefei; Chen, Li; Zhou, Hua; Gong, Yanju; Dai, Liqun; Zheng, Yu

    2016-02-15

    Tetramethylpyrazine (TMP), one of the active ingredients of the Chinese herb Lingusticum Wallichii (Chuan Xiong) has been proved to protect the medulla oblongata from chronic hypoxia injury. However, the underlying mechanism remains unclear. The purpose of this study was to determine whether the protective effects of TMP are associated with the heme oxygenase-1 (HO-1) dependant pathway in adult rats. The morphological changes of neurons in the hypoglossal nucleus (12N), the nucleus ambiguus (Amb), the nucleus tractus solitarius (NTS), and the pre-Bötzinger complex (pre-BötC) were investigated by Nissl staining; the malondialdehyde (MDA) content and superoxide dismutase (SOD) activity were measured to evaluate the anti-oxidant effect; some apoptosis parameters, Bax mRNA and Bcl-2 mRNA, were tested; and the double immunochemistry staining of active caspase-3/NeuN was performed. Meanwhile, the HO-1 protein expression and heme oxygenase (HO) activity were examined. Tin-protoporphyrin (SnPP), a potent inhibitor of HO, was used to further confirm the effect of HO-1. We found that TMP ameliorated the neuron loss in 12N, Amb and NTS, the decrease in SOD activity and the increase in MDA content, the decrease in Bcl-2 mRNA of medulla oblongata (Pmedulla oblongata in the rats. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. Eriodictyol Protects Endothelial Cells against Oxidative Stress-Induced Cell Death through Modulating ERK/Nrf2/ARE-Dependent Heme Oxygenase-1 Expression.

    Lee, Seung Eun; Yang, Hana; Son, Gun Woo; Park, Hye Rim; Park, Cheung-Seog; Jin, Young-Ho; Park, Yong Seek

    2015-06-26

    The pathophysiology of cardiovascular diseases is complex and may involve oxidative stress-related pathways. Eriodictyol is a flavonoid present in citrus fruits that demonstrates anti-inflammatory, anti-cancer, neurotrophic, and antioxidant effects in a range of pathophysiological conditions including vascular diseases. Because oxidative stress plays a key role in the pathogenesis of cardiovascular disease, the present study was designed to verify whether eriodictyol has therapeutic potential. Upregulation of heme oxygenase-1 (HO-1), a phase II detoxifying enzyme, in endothelial cells is considered to be helpful in cardiovascular disease. In this study, human umbilical vein endothelial cells (HUVECs) treated with eriodictyol showed the upregulation of HO-1 through extracellular-regulated kinase (ERK)/nuclear factor erythroid 2-related factor 2 (Nrf2)/antioxidant response element (ARE) signaling pathways. Further, eriodictyol treatment provided protection against hydrogen peroxide-provoked cell death. This protective effect was eliminated by treatment with a specific inhibitor of HO-1 and RNA interference-mediated knockdown of HO-1 expression. These data demonstrate that eriodictyol induces ERK/Nrf2/ARE-mediated HO-1 upregulation in human endothelial cells, which is directly associated with its vascular protection against oxidative stress-related endothelial injury, and propose that targeting the upregulation of HO-1 is a promising approach for therapeutic intervention in cardiovascular disease.

  12. Kidney injury and heme oxygenase-1

    Hai-xing MAI

    2012-02-01

    Full Text Available     Heme oxygenase-1 (HO-1 is one of the main pathways to degrade heme in mammals, and the main degradation products are free iron (Fe2+, carbon monoxide (CO, and bilirubin. Heme plays an important role in promoting cell survival, circulation of intracellular substrates, and immune regulation. Previous studies suggest that HO-1 pathway is an important internal factor in determining the susceptibility and severity of acute kidney injury (AKI. The induction of HO-1 expression can attenuate the severity of renal ischemia-reperfusion injury (IRI, and the inhibition of HO-1 expression will aggravate IRI. The present article summarizes the latest advances in research abroad and at home on protective mechanism by which HO-1 prevents AKI to further deepen our understanding of the role of HO-1 in the treatment of AKI.   

  13. Transduction of PEP-1-heme oxygenase-1 into insulin-producing INS-1 cells protects them against cytokine-induced cell death

    Lee, Su Jin; Kang, Hyung Kyung [Department of Physiology, College of Medicine, Hallym University, Chunchon 200-702 (Korea, Republic of); Song, Dong Keun [Department of Pharmacology, College of Medicine, Hallym University, Chunchon 200-702 (Korea, Republic of); Eum, Won Sik; Park, Jinseu [Department of Biomedical Science and Research Institute of Bioscience and Biotechnology, Hallym University, Chunchon 200-702 (Korea, Republic of); Choi, Soo Young, E-mail: sychoi@hallym.ac.kr [Department of Biomedical Science and Research Institute of Bioscience and Biotechnology, Hallym University, Chunchon 200-702 (Korea, Republic of); Kwon, Hyeok Yil, E-mail: hykwon@hallym.ac.kr [Department of Physiology, College of Medicine, Hallym University, Chunchon 200-702 (Korea, Republic of)

    2015-06-05

    Pro-inflammatory cytokines play a crucial role in the destruction of pancreatic β-cells, thereby triggering the development of autoimmune diabetes mellitus. We recently developed a cell-permeable fusion protein, PEP-1-heme oxygenase-1 (PEP-1-HO-1) and investigated the anti-inflammatory effects in macrophage cells. In this study, we transduced PEP-1-HO-1 into INS-1 insulinoma cells and examined its protective effect against cytokine-induced cell death. PEP-1-HO-1 was successfully delivered into INS-1 cells in time- and dose-dependent manner and was maintained within the cells for at least 48 h. Pre-treatment with PEP-1-HO-1 increased the survival of INS-1 cells exposed to cytokine mixture (IL-1β, IFN-γ, and TNF-α) in a dose-dependent manner. PEP-1-HO-1 markedly decreased cytokine-induced production of reactive oxygen species (ROS), nitric oxide (NO), and malondialdehyde (MDA). These protective effects of PEP-1-HO-1 against cytokines were correlated with the changes in the levels of signaling mediators of inflammation (iNOS and COX-2) and cell apoptosis/survival (Bcl-2, Bax, caspase-3, PARP, JNK, and Akt). These results showed that the transduced PEP-1-HO-1 efficiently prevented cytokine-induced cell death of INS-1 cells by alleviating oxidative/nitrosative stresses and inflammation. Further, these results suggested that PEP-1-mediated HO-1 transduction may be a potential therapeutic strategy to prevent β-cell destruction in patients with autoimmune diabetes mellitus. - Highlights: • We showed that PEP-1-HO-1 was efficiently delivered into INS-1 cells. • Transduced PEP-1-HO-1 exerted a protective effect against cytokine-induced cell death. • Transduced PEP-1-HO-1 inhibited cytokine-induced ROS and NO accumulation. • PEP-1-HO-1 suppressed cytokine-induced expression of iNOS, COX-2, and Bax. • PEP-1-HO-1 transduction may be an efficient tool to prevent β-cell destruction.

  14. Transduction of PEP-1-heme oxygenase-1 into insulin-producing INS-1 cells protects them against cytokine-induced cell death

    Lee, Su Jin; Kang, Hyung Kyung; Song, Dong Keun; Eum, Won Sik; Park, Jinseu; Choi, Soo Young; Kwon, Hyeok Yil

    2015-01-01

    Pro-inflammatory cytokines play a crucial role in the destruction of pancreatic β-cells, thereby triggering the development of autoimmune diabetes mellitus. We recently developed a cell-permeable fusion protein, PEP-1-heme oxygenase-1 (PEP-1-HO-1) and investigated the anti-inflammatory effects in macrophage cells. In this study, we transduced PEP-1-HO-1 into INS-1 insulinoma cells and examined its protective effect against cytokine-induced cell death. PEP-1-HO-1 was successfully delivered into INS-1 cells in time- and dose-dependent manner and was maintained within the cells for at least 48 h. Pre-treatment with PEP-1-HO-1 increased the survival of INS-1 cells exposed to cytokine mixture (IL-1β, IFN-γ, and TNF-α) in a dose-dependent manner. PEP-1-HO-1 markedly decreased cytokine-induced production of reactive oxygen species (ROS), nitric oxide (NO), and malondialdehyde (MDA). These protective effects of PEP-1-HO-1 against cytokines were correlated with the changes in the levels of signaling mediators of inflammation (iNOS and COX-2) and cell apoptosis/survival (Bcl-2, Bax, caspase-3, PARP, JNK, and Akt). These results showed that the transduced PEP-1-HO-1 efficiently prevented cytokine-induced cell death of INS-1 cells by alleviating oxidative/nitrosative stresses and inflammation. Further, these results suggested that PEP-1-mediated HO-1 transduction may be a potential therapeutic strategy to prevent β-cell destruction in patients with autoimmune diabetes mellitus. - Highlights: • We showed that PEP-1-HO-1 was efficiently delivered into INS-1 cells. • Transduced PEP-1-HO-1 exerted a protective effect against cytokine-induced cell death. • Transduced PEP-1-HO-1 inhibited cytokine-induced ROS and NO accumulation. • PEP-1-HO-1 suppressed cytokine-induced expression of iNOS, COX-2, and Bax. • PEP-1-HO-1 transduction may be an efficient tool to prevent β-cell destruction

  15. [Gene transfer-induced human heme oxygenase-1 over-expression protects kidney from ischemia-reperfusion injury in rats].

    Lü, Jin-xing; Yan, Chun-yin; Pu, Jin-xian; Hou, Jian-quan; Yuan, He-xing; Ping, Ji-gen

    2010-12-14

    To study the protection of gene transfer-induced human heme oxygenase-1 over-expression against renal ischemia reperfusion injury in rats. The model of kidney ischemia-reperfusion injury was established with Sprague-Dawley rats. In the therapy group (n=18), the left kidney was perfused and preserved with Ad-hHO-1 at 2.5×10(9) pfu/1.0 ml after flushed with 0-4°C HC-A organ storage solution via donor renal aorta. The rats in control groups were perfused with 0.9% saline solution (n=12) or the vector carrying no interest gene Ad-EGFP 2.5×10(9) pfu/1.0 ml (n=18) instead of Ad-hHO-1. BUN and Cr in serum were measured by slide chemical methods. The kidney samples of rats were harvested for assay of histology, immunohistochemistry and quantification of HO enzymatic activity. Apoptosis cells in the kidney were measured by TUNEL. Ad-hHO-1 via donor renal aorta could transfect renal cells of rats effectively, enzymatic activity of HO in treated group [(1.62±0.07) nmol×mg(-1)×min(-1)] is higher than in control groups treated with saline solution team [(1.27±0.07) nmol×mg(-1)×min(-1)] and vector EGFP team [(1.22±0.06) nmol×mg(-1)×min(-1)] (PhHO-1 expressed hHO-1 in kidneys at a high level. Corresponding to this, the level of BUN and Cr, as well as the number of apoptosis cells, were decreased, and the damage in histology by HE staining was ameliorated. Over-expression of human HO-1 can protect the kidney from ischemia/reperfusion injury in rats.

  16. Omega-3 fatty acids protect the brain against ischemic injury by activating Nrf2 and upregulating heme oxygenase 1.

    Zhang, Meijuan; Wang, Suping; Mao, Leilei; Leak, Rehana K; Shi, Yejie; Zhang, Wenting; Hu, Xiaoming; Sun, Baoliang; Cao, Guodong; Gao, Yanqin; Xu, Yun; Chen, Jun; Zhang, Feng

    2014-01-29

    Ischemic stroke is a debilitating clinical disorder that affects millions of people, yet lacks effective neuroprotective treatments. Fish oil is known to exert beneficial effects against cerebral ischemia. However, the underlying protective mechanisms are not fully understood. The present study tests the hypothesis that omega-3 polyunsaturated fatty acids (n-3 PUFAs) attenuate ischemic neuronal injury by activating nuclear factor E2-related factor 2 (Nrf2) and upregulating heme oxygenase-1 (HO-1) in both in vitro and in vivo models. We observed that pretreatment of rat primary neurons with docosahexaenoic acid (DHA) significantly reduced neuronal death following oxygen-glucose deprivation. This protection was associated with increased Nrf2 activation and HO-1 upregulation. Inhibition of HO-1 activity with tin protoporphyrin IX attenuated the protective effects of DHA. Further studies showed that 4-hydroxy-2E-hexenal (4-HHE), an end-product of peroxidation of n-3 PUFAs, was a more potent Nrf2 inducer than 4-hydroxy-2E-nonenal derived from n-6 PUFAs. In an in vivo setting, transgenic mice overexpressing fatty acid metabolism-1, an enzyme that converts n-6 PUFAs to n-3 PUFAs, were remarkably resistant to focal cerebral ischemia compared with their wild-type littermates. Regular mice fed with a fish oil-enhanced diet also demonstrated significant resistance to ischemia compared with mice fed with a regular diet. As expected, the protection was associated with HO-1 upregulation, Nrf2 activation, and 4-HHE generation. Together, our data demonstrate that n-3 PUFAs are highly effective in protecting the brain, and that the protective mechanisms involve Nrf2 activation and HO-1 upregulation by 4-HHE. Further investigation of n-3 PUFA neuroprotective mechanisms may accelerate the development of stroke therapies.

  17. Edaravone protected PC12 cells against MPP(+)-cytoxicity via inhibiting oxidative stress and up-regulating heme oxygenase-1 expression.

    Cheng, Baohua; Guo, Yunliang; Li, Chuangang; Ji, Bingyuan; Pan, Yanyou; Chen, Jing; Bai, Bo

    2014-08-15

    Oxidative stress is involved in the pathogenesis of Parkinson's disease (PD). Edaravone has been shown to have a neuroprotective effect. In the present work, we investigated the effect of edaravone on 1-methyl-4-phenylpyridinium (MPP(+))-treated PC12 cells. Edaravone inhibited the decrease of cell viability and apoptosis induced by MPP(+) in PC12 cells. In addition, edaravone alleviated intracellular reactive oxygen species (ROS) production. MPP(+) induced heme oxygenase-1 (HO-1) expression, which was further enhanced by edaravone. The inhibitor of HO-1 zinc protoporphyrin-IX attenuated the neuroprotection of edaravone. So edaravone protected PC12 cells against MPP(+)-cytoxicity via inhibiting oxidative stress and up-regulating HO-1 expression. The data showed that edaravone was neuroprotective and could be potentially therapeutics for PD in future. Copyright © 2014 Elsevier B.V. All rights reserved.

  18. The in vitro protection of human decay accelerating factor and hDAF/heme oxygenase-1 transgenes in porcine aortic endothelial cells against sera of Formosan macaques.

    Tu, C-F; Tai, H-C; Wu, C-P; Ho, L-L; Lin, Y-J; Hwang, C-S; Yang, T-S; Lee, J-M; Tseng, Y-L; Huang, C-C; Weng, C-N; Lee, P-H

    2010-01-01

    To mitigate hyperacute rejection, pigs have been generated with alpha-Gal transferase gene knockout and transgenic expression of human decay accelerating factor (hDAF), MCP, and CD59. Additionally, heme-oxygenase-1 (HO-1) has been suggested to defend endothelial cells. Sera (MS) (0%, 1%, 5%, 10%, and 15%) from Formosan macaques (Macaca cyclopis, MC), an Old World monkey wildly populated in Taiwan, was used to test the protective in vitro, effects of hDAF or hDAF/hHO-1 on porcine aortic endothelial cells (pAEC) derived from hDAF(+), hDAF(+)/hHO-1(+), and hDAF(+)/hHO-1(-) and 1 nontransgenic pAEC. Ten percent human serum (HS) served as a positive control. When MS addition increased to 10% or 15%, all transgenic pAEC exhibited a greater survival than nontransgenic pAEC. Noticeably, 15% MS reduced survived to 40% in nontransgenic and transgenic pAEC, respectively. These results revealed that hDAF exerted protective effects against MC complement activation. However, comparing with 10% MS and HS in pAEC of nontransgenic pigs, the survivability was higher in HS, suggesting that complement activation by MS was more toxic than that by HS. Furthermore, hDAF(+)/hHO-1(+) showed no further protection against effects of MS on transgenic pAEC. Copyright 2010 Elsevier Inc. All rights reserved.

  19. MAPK/JNK1 activation protects cells against cadmium-induced autophagic cell death via differential regulation of catalase and heme oxygenase-1 in oral cancer cells.

    So, Keum-Young; Kim, Sang-Hun; Jung, Ki-Tae; Lee, Hyun-Young; Oh, Seon-Hee

    2017-10-01

    Antioxidant enzymes are related to oral diseases. We investigated the roles of heme oxygenase-1 (HO-1) and catalase in cadmium (Cd)-induced oxidative stress and the underlying molecular mechanism in oral cancer cells. Exposing YD8 cells to Cd reduced the expression levels of catalase and superoxide dismutase 1/2 and induced the expression of HO-1 as well as autophagy and apoptosis, which were reversed by N-acetyl-l-cysteine (NAC). Cd-exposed YD10B cells exhibited milder effects than YD8 cells, indicating that Cd sensitivity is associated with antioxidant enzymes and autophagy. Autophagy inhibition via pharmacologic and genetic modulations enhanced Cd-induced HO-1 expression, caspase-3 cleavage, and the production of reactive oxygen species (ROS). Ho-1 knockdown increased autophagy and apoptosis. Hemin treatment partially suppressed Cd-induced ROS production and apoptosis, but enhanced autophagy and CHOP expression, indicating that autophagy induction is associated with cellular stress. Catalase inhibition by pharmacological and genetic modulations increased Cd-induced ROS production, autophagy, and apoptosis, but suppressed HO-1, indicating that catalase is required for HO-1 induction. p38 inhibition upregulated Cd-induced phospho-JNK and catalase, but suppressed HO-1, autophagy, apoptosis. JNK suppression exhibited contrary results, enhancing the expression of phospho-p38. Co-suppression of p38 and JNK1 failed to upregulate catalase and procaspase-3, which were upregulated by JNK1 overexpression. Overall, the balance between the responses of p38 and JNK activation to Cd appears to have an important role in maintaining cellular homeostasis via the regulation of antioxidant enzymes and autophagy induction. In addition, the upregulation of catalase by JNK1 activation can play a critical role in cell protection against Cd-induced oxidative stress. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. Inhibition of microRNA-153 protects neurons against ischemia/reperfusion injury in an oxygen-glucose deprivation and reoxygenation cellular model by regulating Nrf2/HO-1 signaling.

    Ji, Qiong; Gao, Jianbo; Zheng, Yan; Liu, Xueli; Zhou, Qiangqiang; Shi, Canxia; Yao, Meng; Chen, Xia

    2017-07-01

    MicroRNAs are emerging as critical regulators in cerebral ischemia/reperfusion injury; however, their exact roles remain poorly understood. miR-153 is reported to be a neuron-related miRNA involved in neuroprotection. In this study, we aimed to investigate the precise role of miR-153 in regulating neuron survival during cerebral ischemia/reperfusion injury using an oxygen-glucose deprivation and reoxygenation (OGD/R) cellular model. We found that miR-153 was significantly upregulated in neurons subjected to OGD/R treatment. Inhibition of miR-153 significantly attenuated OGD/R-induced injury and oxidative stress in neurons. Nuclear factor erythroid 2-related factor 2 (Nrf2) was identified as a target gene of miR-153. Inhibition of miR-153 significantly promoted the expression of Nrf2 and heme oxygenase-1 (HO-1). However, silencing of Nrf2 significantly blocked the protective effects of miR-153 inhibition. Our study indicates that the inhibition of miR-153 protects neurons against OGD/R-induced injury by regulating Nrf2/HO-1 signaling and suggests a potential therapeutic target for cerebral ischemia/reperfusion injury. © 2017 Wiley Periodicals, Inc.

  1. Andrographolide stimulates p38 mitogen-activated protein kinase-nuclear factor erythroid-2-related factor 2-heme oxygenase 1 signaling in primary cerebral endothelial cells for definite protection against ischemic stroke in rats.

    Yen, Ting-Lin; Chen, Ray-Jade; Jayakumar, Thanasekaran; Lu, Wan-Jung; Hsieh, Cheng-Ying; Hsu, Ming-Jen; Yang, Chih-Hao; Chang, Chao-Chien; Lin, Yen-Kuang; Lin, Kuan-Hung; Sheu, Joen-Rong

    2016-04-01

    Stroke pathogenesis involves complex oxidative stress-related pathways. The nuclear factor erythroid-2-related factor 2 (Nrf2) and heme oxygenase 1 (HO-1) pathways have been considered molecular targets in pharmacologic intervention for ischemic diseases. Andrographolide, a labdane diterpene, has received increasing attention in recent years because of its various pharmacologic activities. We determined that andrographolide modulates the mitogen-activated protein kinase (MAPK)-Nrf2-HO-1 signaling cascade in primary cerebral endothelial cells (CECs) to provide positive protection against middle cerebral artery occlusion (MCAO)-induced ischemic stroke in rats. In the present study, andrographolide (10 μM) increased HO-1 protein and messenger RNA expressions, Nrf2 phosphorylation, and nuclear translocation in CECs, and these activities were disrupted by a p38 MAPK inhibitor, SB203580, but not by the extracellular signal-regulated kinase inhibitor PD98059 or c-Jun amino-terminal kinase inhibitor SP600125. Similar results were observed in confocal microscopy analysis. Moreover, andrographolide-induced Nrf2 and HO-1 protein expressions were significantly inhibited by Nrf2 small interfering RNA. Moreover, HO-1 knockdown attenuated the protective effect of andrographolide against oxygen-glucose deprivation-induced CEC death. Andrographolide (0.1 mg/kg) significantly suppressed free radical formation, blood-brain barrier disruption, and brain infarction in MCAO-insulted rats, and these effects were reversed by the HO-1 inhibitor zinc protoporphyrin IX. The mechanism is attributable to HO-1 activation, as directly evidenced by andrographolide-induced pronounced HO-1 expression in brain tissues, which was highly localized in the cerebral capillary. In conclusion, andrographolide increased Nrf2-HO-1 expression through p38 MAPK regulation, confirming that it provides protection against MCAO-induced brain injury. These findings provide strong evidence that andrographolide could

  2. Taurine Protects Mouse Spermatocytes from Ionizing Radiation-Induced Damage Through Activation of Nrf2/HO-1 Signaling.

    Yang, Wenjun; Huang, Jinfeng; Xiao, Bang; Liu, Yan; Zhu, Yiqing; Wang, Fang; Sun, Shuhan

    2017-01-01

    The increasing prevalence of ionizing radiation exposure has inevitably raised public concern over the potential detrimental effects of ionizing radiation on male reproductive system function. The detection of drug candidates to prevent reproductive system from damage caused by ionizing radiation is urgent. We aimed to investigate the protective role of taurine on the injury of mouse spermatocyte-derived cells (GC-2) subjected to ionizing radiation. mouse spermatocytes (GC-2 cells) were exposed to ionizing radiation with or without treatment of Taurine. The effect of ionizing radiation and Taurine treatment on GC-2 cells were evaluated by cell viability assay (CCK8), cell cycle and apoptosis. The relative protein abundance change was determined by Western blotting. The siRNA was used to explore whether Nrf2 signaling was involved in the cytoprotection of Taurine. Taurine significantly inhibited the decrease of cell viability, percentage of apoptotic cells and cell cycle arrest induced by ionizing radiation. Western blot analysis showed that taurine significantly limited the ionizing radiation-induced down-regulation of CyclinB1 and CDK1, and suppressed activation of Fas/FasL system pathway. In addition, taurine treatment significantly increased the expression of Nrf2 and HO-1 in GC-2 cells exposed to ionizing radiation, two components in antioxidant pathway. The above cytoprotection of Taurine was blocked by siNrf2. Our results demonstrate that taurine has the potential to effectively protect GC-2 cells from ionizing radiation- triggered damage via upregulation of Nrf2/HO-1 signaling. © 2017 The Author(s). Published by S. Karger AG, Basel.

  3. Edaravone protects rats and human pulmonary alveolar epithelial cells against hyperoxia injury: heme oxygenase-1 and PI3K/Akt pathway may be involved.

    Cao, Huifang; Feng, Ying; Ning, Yunye; Zhang, Zinan; Li, Weihao; Li, Qiang

    2015-01-01

    Hyperoxic acute lung injury (HALI) is a clinical syndrome as a result of prolonged supplement of high concentrations of oxygen. As yet, no specific treatment is available for HALI. The present study aims to investigate the effects of edaravone on hyperoxia-induced oxidative injury and the underlying mechanism. We treated rats and human pulmonary alveolar epithelial cells with hyperoxia and different concentration of edaravone, then examined the effects of edaravone on cell viability, cell injury and two oxidative products. The roles of heme oxygenase-1 (HO-1) and PI3K/Akt pathway were explored using Western blot and corresponding inhibitors. The results showed that edaravone reduced lung biochemical alterations induced by hyperoxia and mortality of rats, dose-dependently alleviated cell mortality, cell injury, and peroxidation of cellular lipid and DNA oxidative damage. It upregulated cellular HO-1 expression and activity, which was reversed by PI3K/Akt pathway inhibition. The administration of zinc protoporphyrin-IX, a HO-1 inhibitor, and LY249002, a PI3K/Akt pathway inhibitor, abolished the protective effects of edaravone in cells. This study indicates that edaravone protects rats and human pulmonary alveolar epithelial cells against hyperoxia-induced injury and the antioxidant effect may be related to upregulation of HO-1, which is regulated by PI3K/Akt pathway.

  4. The cytoprotective enzyme heme oxygenase-1 suppresses Ebola virus replication.

    Hill-Batorski, Lindsay; Halfmann, Peter; Neumann, Gabriele; Kawaoka, Yoshihiro

    2013-12-01

    Ebola virus (EBOV) is the causative agent of a severe hemorrhagic fever in humans with reported case fatality rates as high as 90%. There are currently no licensed vaccines or antiviral therapeutics to combat EBOV infections. Heme oxygenase-1 (HO-1), an enzyme that catalyzes the rate-limiting step in heme degradation, has antioxidative properties and protects cells from various stresses. Activated HO-1 was recently shown to have antiviral activity, potently inhibiting the replication of viruses such as hepatitis C virus and human immunodeficiency virus. However, the effect of HO-1 activation on EBOV replication remains unknown. To determine whether the upregulation of HO-1 attenuates EBOV replication, we treated cells with cobalt protoporphyrin (CoPP), a selective HO-1 inducer, and assessed its effects on EBOV replication. We found that CoPP treatment, pre- and postinfection, significantly suppressed EBOV replication in a manner dependent upon HO-1 upregulation and activity. In addition, stable overexpression of HO-1 significantly attenuated EBOV growth. Although the exact mechanism behind the antiviral properties of HO-1 remains to be elucidated, our data show that HO-1 upregulation does not attenuate EBOV entry or budding but specifically targets EBOV transcription/replication. Therefore, modulation of the cellular enzyme HO-1 may represent a novel therapeutic strategy against EBOV infection.

  5. Heme oxygenase-1 regulates the progression of K/BxN serum transfer arthritis.

    Rita Brines

    Full Text Available Heme oxygenase-1 (HO-1 is induced in many cell types as a defense mechanism against stress. We have investigated the possible role of endogenous HO-1 in the effector phase of arthritis using the K/BxN serum transfer model of arthritis in HO-1 heterozygous and homozygous knock-out mice.Arthritis was induced in C57/Black-6 xFVB (HO-1(+/+, HO-1(+/- and HO-1(-/- mice by intraperitoneal injection of 150 µl serum from arthritic K/BxN mice at days 0 and 2. Blood was collected and animals were sacrificed at day 10. Histological analysis was performed in ankle sections. The levels of inflammatory mediators were measured in serum and paw homogenates by enzyme-linked immunosorbent assay or Multiplex technology. The incidence of arthritis was higher in HO-1(+/- and HO-1(-/- groups compared with HO-1(+/+. The inflammatory response was aggravated in HO-1(+/- mice as shown by arthritic score and the migration of inflammatory cells that could be related to the enhancement of CXCL-1 production. In addition, the HO-1(+/- group showed proteoglycan depletion significantly higher than HO-1(+/+ mice. Serum levels of matrix metalloproteinase-3, monocyte chemotactic protein-1, plasminogen activator inhibitor-1, E-selectin and intercellular adhesion molecule-1 were increased in arthritic HO-1(-/- mice, whereas vascular endothelial growth factor and some cytokines such as interferon-γ showed a reduction compared to HO-1(+/+ or HO-1(+/- mice. In addition, down-regulated gene expression of ferritin, glutathione S-reductase A1 and superoxide dismutase-2 was observed in the livers of arthritic HO-1(+/- animals.Endogenous HO-1 regulates the production of systemic and local inflammatory mediators and plays a protective role in K/BxN serum transfer arthritis.

  6. Heme oxygenase-1, oxidation, inflammation and atherosclerosis

    Jesus A Araujo

    2012-07-01

    Full Text Available Atherosclerosis is an inflammatory process of the vascular wall characterized by the infiltration of lipids and inflammatory cells. Oxidative modifications of infiltrating low density lipoproteins and induction of oxidative stress play a major role in lipid retention in the vascular wall, uptake by macrophages and generation of foam cells, a hallmark of this disorder. The vasculature has a plethora of protective resources against oxidation and inflammation, many of them regulated by the Nrf2 transcription factor. Heme oxygenase-1 (HO-1 is a Nrf2-regulated gene that plays a critical role in the prevention of vascular inflammation. It is the inducible isoform of heme oxygenase, responsible for the oxidative cleavage of heme groups leading to the generation of biliverdin, carbon monoxide and release of ferrous iron. HO-1 has important antioxidant, antiinflammatory, antiapoptotic, antiproliferative and immunomodulatory effects in vascular cells, most of which play a significant role in the protection against atherogenesis. HO-1 may also be an important feature in macrophage differentiation and polarization to certain subtypes. The biological effects of HO-1 are largely attributable to its enzymatic activity, which can be conceived as a system with three arms of action, corresponding to its three enzymatic byproducts. HO-1 mediated vascular protection may be due to a combination of systemic and vascular local effects. It is usually expressed at low levels but can be highly upregulated in the presence of several proatherogenic stimuli. The HO-1 system is amenable for use in the development of new therapies, some of them currently under experimental and clinical trials. Interestingly, in contrast to the HO-1 antiatherogenic actions, the expression of its transcriptional regulator Nrf2 leads to proatherogenic effects instead. This article reviews the evidence that supports the antiatherogenic role of HO-1, potential pathways and mechanisms mediating

  7. Ginsenoside Rb1 protects against 6-hydroxydopamine-induced oxidative stress by increasing heme oxygenase-1 expression through an estrogen receptor-related PI3K/Akt/Nrf2-dependent pathway in human dopaminergic cells

    Hwang, Yong Pil; Jeong, Hye Gwang

    2010-01-01

    Phytoestrogens are polyphenolic non-steroidal plant compounds with estrogen-like biological activity. Ginseng, the root of Panax ginseng C.A. Meyer (Araliaceae), is a popular traditional herbal medicine. Ginsenoside Rb1 (Rb1), an active component commonly found in ginseng root, is a phytoestrogen that exerts estrogen-like activity. In this study, we demonstrate that the phytoestrogen Rb1 inhibits 6-hydroxydopamine (6-OHDA)-induced oxidative injury via an ER-dependent Gβ1/PI3K/Akt and heme oxygenase-1 (HO-1) pathway. Pretreatment of SH-SY5Y cells with Rb1 significantly reduced 6-OHDA-induced caspase-3 activation and subsequent cell death. Rb1 also up-regulated HO-1 expression, which conferred cytoprotection against 6-OHDA-induced oxidative injury. Moreover, Rb1 induced both Nrf2 nuclear translocation, which is upstream of HO-1 expression and PI3K activation, a pathway that is involved in induced Nrf2 nuclear translocation, HO-1 expression and cytoprotection. Also, Rb1-mediated increases in PI3K activation and HO-1 induction were reversed by co-treatment with ICI 182,780 and pertussis toxin. Taken together, these results suggest that Rb1 augments the cellular antioxidant defenses through ER-dependent HO-1 induction via the Gβ1/PI3K/Akt-Nrf2 signaling pathway, thereby protecting cells from oxidative stress. Thus our study indicates that Rb1 has a partial cytoprotective role in dopaminergic cell culture systems.

  8. Heme oxygenase-1 mediates the protective effects of ischemic preconditioning on mitigating lung injury induced by lower limb ischemia-reperfusion in rats.

    Peng, Tsui-Chin; Jan, Woan-Ching; Tsai, Pei-Shan; Huang, Chun-Jen

    2011-05-15

    Lower limb ischemia-reperfusion (I/R) imposes oxidative stress, elicits inflammatory response, and subsequently induces acute lung injury. Ischemic preconditioning (IP), a process of transient I/R, mitigates the acute lung injury induced by I/R. We sought to elucidate whether the protective effects of IP involve heme oxygenase-1 (HO-1). Adult male rats were randomized to receive I/R, I/R plus IP, I/R plus IP plus the HO-1 inhibitor tin protoporphyrin (SnPP) (n = 12 in each group). Control groups were run simultaneously. I/R was induced by applying rubber band tourniquet high around each thigh for 3 h followed by reperfusion for 3 h. To achieve IP, three cycles of bilateral lower limb I/R (i.e., ischemia for 10 min followed by reperfusion for 10 min) were performed. IP was performed immediately before I/R. After sacrifice, degree of lung injury was determined. Histologic findings, together with assays of leukocyte infiltration (polymorphonuclear leukocytes/alveoli ratio and myeloperoxidase activity) and lung water content (wet/dry weight ratio), confirmed that I/R induced acute lung injury. I/R also caused significant inflammatory response (increases in chemokine, cytokine, and prostaglandin E(2) concentrations), imposed significant oxidative stress (increases in nitric oxide and malondialdehyde concentrations), and up-regulated HO-1 expression in lung tissues. IP significantly enhanced HO-1 up-regulation and, in turn, mitigated oxidative stress, inflammatory response, and acute lung injury induced by I/R. In addition, the protective effects of IP were counteracted by SnPP. The protective effects of IP on mitigating acute lung injury induced by lower limb I/R are mediated by HO-1. Copyright © 2011 Elsevier Inc. All rights reserved.

  9. Effect of heme oxygenase-1 on the protection of ischemia reperfusion injury of bile duct in rats after liver transplantation.

    Zhan, Xi; Zhang, Zhiqing; Huang, Hanfei; Zhang, Yujun; Zeng, Zhong

    2018-06-01

    To investigate the effect of heme oxygenase-1 (HO-1) on the ischemic reperfusion injury (IRI) of bile duct in rat models after liver transplantation. 320 SD rats were equally and randomly divided into 5 groups, which were group A receiving injection of 3×10 8 /pfu/ml adenovirus (adv), group B with donor receiving Adv-HO-1 and recipient receiving Adv-HO-1-siRNA, group C with donor and recipient both receiving Adv-HO-1, group D with donor receiving Adv-HO-1-siRNA and recipient receiving Adv-HO-1, and group E with donor and recipient both receiving Adv-HO-1-siRNA at 24h before liver transplantation. Donor liver was stored in UW liquid at 4°C followed by measuring HO-1 level by western blot before transplantation. On d1, d3, d7 and d14, serum and liver was isolated for analysis of liver function, inflammatory cell infiltration by H&E staining, ultrastructure of liver by transmission electron microscopy as well as the expression of HO-1, Bsep, Mrp2 and Ntcp by western blot. Compared with group D and E, group B and C displayed improved liver function as demonstrated by lower level of ALT, AST, LDH, TBIL, ALP and GGT, increased secretion of TBA and PL as well as expression of transporter proteins (Bsep, Mrp2 and Ntcp), reduced inflammatory cells infiltration and liver injury. Our study demonstrated that overexpression of HO-1 in donor liver can ameliorate the damage to bile duct and liver, and improved liver function, suggesting HO-1 might be a new therapeutic target in the treatment of IRI after liver transplantation. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  10. Molecular mechanism and functional consequences of lansoprazole-mediated heme oxygenase-1 induction

    Schulz-Geske, Stephanie; Erdmann, Kati; Wong, Ronald J; Stevenson, David K; Schröder, Henning; Grosser, Nina

    2009-01-01

    AIM: To investigate the molecular mechanism and functional consequences of heme oxygenase-1 (HO-1) activation by lansoprazole in endothelial cells and macrophages. METHODS: Expression of HO-1 mRNA was analyzed by Northern blotting. Western blotting was used to determine the HO-1 and ferritin protein levels. NADPH-dependent reactive oxygen species (ROS) formation was measured with lucigenin-enhanced chemiluminescence. HO-1 promoter activity in mouse fibroblasts, stably transfected with a 15-kb HO-1 gene that drives expression of the reporter gene luciferase, was assessed using in vivo bioluminescence imaging. RESULTS: Lansoprazole increased HO-1 mRNA levels in endothelial cells and HO-1 protein levels in macrophages. In addition, lansoprazole-induced ferritin protein levels in both cell systems. Moreover, induction of the antioxidant proteins HO-1 and ferritin by lansoprazole was followed by a decrease in NADPH-mediated ROS formation. The radical scavenging properties of lansoprazole were diminished in the presence of the HO inhibitor, chromium mesoporphyrin IX. Induction of HO-1 gene expression by lansoprazole was not related to oxidative stress or to the activation of the mitogen-activated protein kinase pathway. However, the phosphatidylinositol 3-kinase inhibitor LY294002 showed a concentration-dependent inhibition of HO-1 mRNA and promoter activity. CONCLUSION: Activation of HO-1 and ferritin may account for the gastric protection of lansoprazole and is dependent on a pathway blocked by LY294002. PMID:19764090

  11. Gene therapy strategy for long-term myocardial protection using adeno-associated virus-mediated delivery of heme oxygenase gene.

    Melo, Luis G; Agrawal, Reitu; Zhang, Lunan; Rezvani, Mojgan; Mangi, Abeel A; Ehsan, Afshin; Griese, Daniel P; Dell'Acqua, Giorgio; Mann, Michael J; Oyama, Junichi; Yet, Shaw-Fang; Layne, Matthew D; Perrella, Mark A; Dzau, Victor J

    2002-02-05

    Ischemia and oxidative stress are the leading mechanisms for tissue injury. An ideal strategy for preventive/protective therapy would be to develop an approach that could confer long-term transgene expression and, consequently, tissue protection from repeated ischemia/reperfusion injury with a single administration of a therapeutic gene. In the present study, we used recombinant adeno-associated virus (rAAV) as a vector for direct delivery of the cytoprotective gene heme oxygenase-1 (HO-1) into the rat myocardium, with the purpose of evaluating this strategy as a therapeutic approach for long-term protection from ischemia-induced myocardial injury. Human HO-1 gene (hHO-1) was delivered to normal rat hearts by intramyocardial injection. AAV-mediated transfer of the hHO-1 gene 8 weeks before acute coronary artery ligation and release led to a dramatic reduction (>75%) in left ventricular myocardial infarction. The reduction in infarct size was accompanied by decreases in myocardial lipid peroxidation and in proapoptotic Bax and proinflammatory interleukin-1beta protein abundance, concomitant with an increase in antiapoptotic Bcl-2 protein level. This suggested that the transgene exerts its cardioprotective effects in part by reducing oxidative stress and associated inflammation and apoptotic cell death. This study documents the beneficial therapeutic effect of rAAV-mediated transfer, before myocardial injury, of a cytoprotective gene that confers long-term myocardial protection from ischemia/reperfusion injury. Our data suggest that this novel "pre-event" gene transfer approach may provide sustained tissue protection from future repeated episodes of injury and may be beneficial as preventive therapy for patients with or at risk of developing coronary ischemic events.

  12. Generation and characterization of human heme oxygenase-1 transgenic pigs.

    Hye-Jung Yeom

    Full Text Available Xenotransplantation using transgenic pigs as an organ source is a promising strategy to overcome shortage of human organ for transplantation. Various genetic modifications have been tried to ameliorate xenograft rejection. In the present study we assessed effect of transgenic expression of human heme oxygenase-1 (hHO-1, an inducible protein capable of cytoprotection by scavenging reactive oxygen species and preventing apoptosis caused by cellular stress during inflammatory processes, in neonatal porcine islet-like cluster cells (NPCCs. Transduction of NPCCs with adenovirus containing hHO-1 gene significantly reduced apoptosis compared with the GFP-expressing adenovirus control after treatment with either hydrogen peroxide or hTNF-α and cycloheximide. These protective effects were diminished by co-treatment of hHO-1 antagonist, Zinc protoporphyrin IX. We also generated transgenic pigs expressing hHO-1 and analyzed expression and function of the transgene. Human HO-1 was expressed in most tissues, including the heart, kidney, lung, pancreas, spleen and skin, however, expression levels and patterns of the hHO-1 gene are not consistent in each organ. We isolate fibroblast from transgenic pigs to analyze protective effect of the hHO-1. As expected, fibroblasts derived from the hHO-1 transgenic pigs were significantly resistant to both hydrogen peroxide damage and hTNF-α and cycloheximide-mediated apoptosis when compared with wild-type fibroblasts. Furthermore, induction of RANTES in response to hTNF-α or LPS was significantly decreased in fibroblasts obtained from the hHO-1 transgenic pigs. These findings suggest that transgenic expression of hHO-1 can protect xenografts when exposed to oxidative stresses, especially from ischemia/reperfusion injury, and/or acute rejection mediated by cytokines. Accordingly, hHO-1 could be an important candidate molecule in a multi-transgenic pig strategy for xenotransplantation.

  13. Generation and characterization of human heme oxygenase-1 transgenic pigs.

    Yeom, Hye-Jung; Koo, Ok Jae; Yang, Jaeseok; Cho, Bumrae; Hwang, Jong-Ik; Park, Sol Ji; Hurh, Sunghoon; Kim, Hwajung; Lee, Eun Mi; Ro, Han; Kang, Jung Taek; Kim, Su Jin; Won, Jae-Kyung; O'Connell, Philip J; Kim, Hyunil; Surh, Charles D; Lee, Byeong-Chun; Ahn, Curie

    2012-01-01

    Xenotransplantation using transgenic pigs as an organ source is a promising strategy to overcome shortage of human organ for transplantation. Various genetic modifications have been tried to ameliorate xenograft rejection. In the present study we assessed effect of transgenic expression of human heme oxygenase-1 (hHO-1), an inducible protein capable of cytoprotection by scavenging reactive oxygen species and preventing apoptosis caused by cellular stress during inflammatory processes, in neonatal porcine islet-like cluster cells (NPCCs). Transduction of NPCCs with adenovirus containing hHO-1 gene significantly reduced apoptosis compared with the GFP-expressing adenovirus control after treatment with either hydrogen peroxide or hTNF-α and cycloheximide. These protective effects were diminished by co-treatment of hHO-1 antagonist, Zinc protoporphyrin IX. We also generated transgenic pigs expressing hHO-1 and analyzed expression and function of the transgene. Human HO-1 was expressed in most tissues, including the heart, kidney, lung, pancreas, spleen and skin, however, expression levels and patterns of the hHO-1 gene are not consistent in each organ. We isolate fibroblast from transgenic pigs to analyze protective effect of the hHO-1. As expected, fibroblasts derived from the hHO-1 transgenic pigs were significantly resistant to both hydrogen peroxide damage and hTNF-α and cycloheximide-mediated apoptosis when compared with wild-type fibroblasts. Furthermore, induction of RANTES in response to hTNF-α or LPS was significantly decreased in fibroblasts obtained from the hHO-1 transgenic pigs. These findings suggest that transgenic expression of hHO-1 can protect xenografts when exposed to oxidative stresses, especially from ischemia/reperfusion injury, and/or acute rejection mediated by cytokines. Accordingly, hHO-1 could be an important candidate molecule in a multi-transgenic pig strategy for xenotransplantation.

  14. Cordycepin alleviates lipopolysaccharide-induced acute lung injury via Nrf2/HO-1 pathway.

    Qing, Rui; Huang, Zezhi; Tang, Yufei; Xiang, Qingke; Yang, Fan

    2018-04-24

    The present study is to investigate the protective effect of cordycepin on inflammatory reactions in rats with acute lung injury (ALI) induced by lipopolysaccharide (LPS), as well as the underlying mechanism. Wistar rat model of ALI was induced by intravenous injection of LPS (30 mg/kg body weight). One hour later, intravenous injection of cordycepin (1, 10 or 30 mg/kg body weight) was administered. The wet-to-dry weight ratio of lung tissues and myeloperoxidase activity in the lung tissues were measured. The contents of nitrite and nitrate were measured by reduction method, while chemiluminescence was used to determine the content of superoxide. Quantitative real-time polymerase chain reaction and Western blotting were used to determine the expression of mRNA and protein, respectively. Colorimetry was performed to determine the enzymatic activity of heme oxygenase-1 (HO-1). Nuclear translocation of Nrf2 was identified by Western blotting. The plasma contents of cytokines were measured by enzyme-linked immunosorbent assay. Cordycepin enhanced the expression and enzymatic activity of HO-1 in ALI rats, and activated Nrf2 by inducing the translocation of Nrf2 from cytoplasm to nucleus. In addition, cordycepin regulated the secretion of TNF-α, IL-6 and IL-10 via HO-1, and suppressed inflammation in lung tissues of ALI rats by inducing the expression of HO-1. HO-1 played important roles in the down-regulation of superoxide levels in lung tissues by cordycepin, and HO-1 expression induced by cordycepin affected nitrite and nitrate concentrations in plasma and iNOS protein expression in lung tissues. Cordycepin showed protective effect on injuries in lung tissues. The present study demonstrates that cordycepin alleviates inflammation induced by LPS via the activation of Nrf2 and up-regulation of HO-1 expression. Copyright © 2018. Published by Elsevier B.V.

  15. Association of Heme Oxygenase 1 with Lung Protection in Malaria-Associated ALI/ARDS.

    Pereira, Marcelo L M; Ortolan, Luana S; Sercundes, Michelle K; Debone, Daniela; Murillo, Oscar; Lima, Flávia A; Marinho, Claudio R F; Epiphanio, Sabrina

    2016-01-01

    Malaria is a serious disease, caused by the parasite of the genus Plasmodium , which was responsible for 440,000 deaths in 2015. Acute lung injury/acute respiratory distress syndrome (ALI/ARDS) is one of the main clinical complications in severe malaria. The murine model DBA/2 reproduces the clinical signs of ALI/ARDS in humans, when infected with Plasmodium berghei ANKA. High levels of HO-1 were reported in cases of severe malaria. Our data indicated that the HO-1 mRNA and protein expression are increased in mice that develop malaria-associated ALI/ARDS (MA-ALI/ARDS). Additionally, the hemin, a HO-1 inducing drug, prevented mice from developing MA-ALI/ARDS when administered prior to the development of MA-ALI/ARDS in this model. Also, hemin treatment showed an amelioration of respiratory parameters in mice, high VEGF levels in the sera, and a decrease in vascular permeability in the lung, which are signs of ALI/ARDS. Therefore, the induction of HO-1 before the development of MA-ALI/ARDS could be protective. However, the increased expression of HO-1 on the onset of MA-ALI/ARDS development may represent an effort to revert the phenotype of this syndrome by the host. We therefore confirm that HO-1 inducing drugs could be used for prevention of MA-ALI/ARDS in humans.

  16. Arctigenin Protects against Lipopolysaccharide-Induced Pulmonary Oxidative Stress and Inflammation in a Mouse Model via Suppression of MAPK, HO-1, and iNOS Signaling.

    Zhang, Wen-zhou; Jiang, Zheng-kui; He, Bao-xia; Liu, Xian-ben

    2015-08-01

    Arctigenin, a bioactive component of Arctium lappa (Nubang), has anti-inflammatory activity. Here, we investigated the effects of arctigenin on lipopolysaccharide (LPS)-induced acute lung injury. Mice were divided into four groups: control, LPS, LPS + DMSO, and LPS + Arctigenin. Mice in the LPS + Arctigenin group were injected intraperitoneally with 50 mg/kg of arctigenin 1 h before an intratracheal administration of LPS (5 mg/kg). Lung tissues and bronchoalveolar lavage fluids (BALFs) were collected. Histological changes of the lung were analyzed by hematoxylin and eosin staining. Arctigenin decreased LPS-induced acute lung inflammation, infiltration of inflammatory cells into BALF, and production of pro-inflammatory cytokines. Moreover, arctigenin pretreatment reduced the malondialdehyde level and increased superoxide dismutase and catalase activities and glutathione peroxidase/glutathione disulfide ratio in the lung. Mechanically, arctigenin significantly reduced the production of nitric oxygen and inducible nitric oxygen synthase (iNOS) expression, enhanced the expression of heme oxygenase-1, and decreased the phosphorylation of mitogen-activated protein kinases (MAPKs). Arctigenin has anti-inflammatory and antioxidative effects on LPS-induced acute lung injury, which are associated with modulation of MAPK, HO-1, and iNOS signaling.

  17. Beyond gastric acid reduction: Proton pump inhibitors induce heme oxygenase-1 in gastric and endothelial cells

    Becker, Jan C.; Grosser, Nina; Waltke, Christian; Schulz, Stephanie; Erdmann, Kati; Domschke, Wolfram; Schroeder, Henning; Pohle, Thorsten

    2006-01-01

    Proton pump inhibitors (PPIs) have been demonstrated to prevent gastric mucosal injury by mechanisms independent of acid inhibition. Here we demonstrate that both omeprazole and lansoprazole protect human gastric epithelial and endothelial cells against oxidative stress. This effect was abrogated in the presence of the heme oxygenase-1 (HO-1) inhibitor ZnBG. Exposure to either PPI resulted in a strong induction of HO-1 expression on mRNA and protein level, and led to an increased activity of this enzyme. Expression of cyclooxygenase isoforms 1 and 2 remained unaffected, and COX-inhibitors did not antagonize HO-1 induction by PPIs. Our results suggest that the antioxidant defense protein HO-1 is a target of PPIs in both endothelial and gastric epithelial cells. HO-1 induction might account for the gastroprotective effects of PPIs independently of acid inhibition, especially in NSAID gastropathy. Moreover, our findings provide additional perspectives for a possible but yet unexplored use of PPIs in vasoprotection

  18. Beyond gastric acid reduction: Proton pump inhibitors induce heme oxygenase-1 in gastric and endothelial cells

    Becker, Jan C [Department of Medicine B, University of Muenster, 48149 Muenster (Germany); Grosser, Nina [Department of Pharmacology and Toxicology, School of Pharmacy, Martin Luther University, Halle-Wittenberg, 06099 Halle (Saale) (Germany); Waltke, Christian [Department of Medicine B, University of Muenster, 48149 Muenster (Germany); Schulz, Stephanie [Department of Pharmacology and Toxicology, School of Pharmacy, Martin Luther University, Halle-Wittenberg, 06099 Halle (Saale) (Germany); Erdmann, Kati [Department of Pharmacology and Toxicology, School of Pharmacy, Martin Luther University, Halle-Wittenberg, 06099 Halle (Saale) (Germany); Domschke, Wolfram [Department of Medicine B, University of Muenster, 48149 Muenster (Germany); Schroeder, Henning [Department of Pharmacology and Toxicology, School of Pharmacy, Martin Luther University, Halle-Wittenberg, 06099 Halle (Saale) (Germany); Pohle, Thorsten [Department of Medicine B, University of Muenster, 48149 Muenster (Germany)

    2006-07-07

    Proton pump inhibitors (PPIs) have been demonstrated to prevent gastric mucosal injury by mechanisms independent of acid inhibition. Here we demonstrate that both omeprazole and lansoprazole protect human gastric epithelial and endothelial cells against oxidative stress. This effect was abrogated in the presence of the heme oxygenase-1 (HO-1) inhibitor ZnBG. Exposure to either PPI resulted in a strong induction of HO-1 expression on mRNA and protein level, and led to an increased activity of this enzyme. Expression of cyclooxygenase isoforms 1 and 2 remained unaffected, and COX-inhibitors did not antagonize HO-1 induction by PPIs. Our results suggest that the antioxidant defense protein HO-1 is a target of PPIs in both endothelial and gastric epithelial cells. HO-1 induction might account for the gastroprotective effects of PPIs independently of acid inhibition, especially in NSAID gastropathy. Moreover, our findings provide additional perspectives for a possible but yet unexplored use of PPIs in vasoprotection.

  19. Edaravone Improves Septic Cardiac Function by Inducing an HIF-1α/HO-1 Pathway

    Chao He

    2018-01-01

    Full Text Available Septic myocardial dysfunction remains prevalent and raises mortality rate in patients with sepsis. During sepsis, tissues undergo tremendous oxidative stress which contributes critically to organ dysfunction. Edaravone, a potent radical scavenger, has been proved beneficial in ischemic injuries involving hypoxia-inducible factor- (HIF- 1, a key regulator of a prominent antioxidative protein heme oxygenase- (HO- 1. However, its effect in septic myocardial dysfunction remains unclarified. We hypothesized that edaravone may prevent septic myocardial dysfunction by inducing the HIF-1/HO-1 pathway. Rats were subjected to cecal ligation and puncture (CLP with or without edaravone infusion at three doses (50, 100, or 200 mg/kg, resp. before CLP and intraperitoneal injection of the HIF-1α antagonist, ME (15 mg/kg, after CLP. After CLP, rats had cardiac dysfunction, which was associated with deformed myocardium, augmented lipid peroxidation, and increased myocardial apoptosis and inflammation, along with decreased activities of catalase, HIF-1α, and HO-1 in the myocardium. Edaravone pretreatment dose-dependently reversed the changes, of which high dose most effectively improved cardiac function and survival rate of septic rats. However, inhibition of HIF-1α by ME demolished the beneficial effects of edaravone at high dose, reducing the survival rate of the septic rats without treatments. Taken together, edaravone, by inducing the HIF-1α/HO-1 pathway, suppressed oxidative stress and protected the heart against septic myocardial injury and dysfunction.

  20. Therapeutic Roles of Heme Oxygenase-1 in Metabolic Diseases: Curcumin and Resveratrol Analogues as Possible Inducers of Heme Oxygenase-1

    Yong Son

    2013-01-01

    Full Text Available Metabolic diseases, such as insulin resistance, type II diabetes, and obesity, are associated with a low-grade chronic inflammation (inflammatory stress, oxidative stress, and endoplasmic reticulum (ER stress. Because the integration of these stresses is critical to the pathogenesis of metabolic diseases, agents and cellular molecules that can modulate these stress responses are emerging as potential targets for intervention and treatment of metabolic diseases. It has been recognized that heme oxygenase-1 (HO-1 plays an important role in cellular protection. Because HO-1 can reduce inflammatory stress, oxidative stress, and ER stress, in part by exerting antioxidant, anti-inflammatory, and antiapoptotic effects, HO-1 has been suggested to play important roles in pathogenesis of metabolic diseases. In the present review, we will explore our current understanding of the protective mechanisms of HO-1 in metabolic diseases and present some emerging therapeutic options for HO-1 expression in treating metabolic diseases, together with the therapeutic potential of curcumin and resveratrol analogues that have their ability to induce HO-1 expression.

  1. Induction of HO-1 by carbon monoxide releasing molecule-2 attenuates thrombin-induced COX-2 expression and hypertrophy in primary human cardiomyocytes

    Chien, Peter Tzu-Yu; Lin, Chih-Chung; Hsiao, Li-Der; Yang, Chuen-Mao

    2015-01-01

    Carbon monoxide (CO) is one of the cytoprotective byproducts of heme oxygenase (HO)-1 and exerts anti-inflammatory action in various models. However, the detailed mechanisms underlying CO-induced HO-1 expression in primary human cardiomyocytes remain largely unidentified. We used primary left ventricle myocytes as a model and applied CO releasing molecule (CORM)-2 to investigate the relationship of CO and HO-1 expression. We herein used Western blot, real-time PCR, promoter activity and EIA to investigate the role of HO-1 expression protecting against thrombin-mediated responses. We found that thrombin-induced COX-2 expression, PGE 2 release and cardiomyocyte hypertrophy markers (increase in ANF/BNP, α-actin expression and cell surface area) was attenuated by pretreatment with CORM-2 which was partially reversed by hemoglobin (Hb) or ZnPP (an inhibitor of HO-1 activity), suggesting that HO-1/CO system may be of clinical importance to ameliorate heart failure through inhibition of inflammatory responses. CORM-2-induced HO-1 protein expression, mRNA and promoter was attenuated by pretreatment with the inhibitors of Pyk2 (PF431396), PDGFR (AG1296), PI3K (LY294002), Akt (SH-5), p38 (SB202530), JNK1/2 (SP600125), FoxO1 (AS1842856) and Sp1 (mithramycin A). The involvement of these signaling components was further confirmed by transfection with respective siRNAs, consistent with those of pharmacological inhibitors. These results suggested that CORM-2-induced HO-1 expression is mediated through a Pyk2/PDGFR/PI3K/Akt/FoxO1/Sp1-dependent manner and exerts a cytoprotective effect in human cardiomyocytes. - Graphical abstract: In summary, CORM-2 treatment induces Pyk2 transactivated PDGFR, which induces PI3K/Akt/MAPK activation, and then recruits Sp1/Foxo1 transcriptional factors to regulate HO-1 gene expression in primary human cardiomyocytes. - Highlights: • CORM-2 induces HO-1 expression. • Pyk2-dependent PDGFR activates PI3K/Akt/MAPK pathway in CORM-2-induced HO-1

  2. Induction of HO-1 by carbon monoxide releasing molecule-2 attenuates thrombin-induced COX-2 expression and hypertrophy in primary human cardiomyocytes

    Chien, Peter Tzu-Yu [Department of Physiology and Pharmacology and Health Ageing Research Center, Chang Gung University, Kwei-Shan, Tao-Yuan, Taiwan (China); Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Kwei-Shan, Tao-Yuan, Taiwan (China); Lin, Chih-Chung; Hsiao, Li-Der [Department of Anesthetics, Chang Gung Memorial Hospital at Lin-Kou and College of Medicine, Chang Gung University, Kwei-San, Tao-Yuan, Taiwan (China); Yang, Chuen-Mao, E-mail: chuenmao@mail.cgu.edu.tw [Department of Physiology and Pharmacology and Health Ageing Research Center, Chang Gung University, Kwei-Shan, Tao-Yuan, Taiwan (China); Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Kwei-Shan, Tao-Yuan, Taiwan (China); Research Center for Industry of Human Ecology and Graduate Institute of Health Industry Technology, Chang Gung University of Science and Technology, Tao-Yuan, Taiwan (China)

    2015-12-01

    Carbon monoxide (CO) is one of the cytoprotective byproducts of heme oxygenase (HO)-1 and exerts anti-inflammatory action in various models. However, the detailed mechanisms underlying CO-induced HO-1 expression in primary human cardiomyocytes remain largely unidentified. We used primary left ventricle myocytes as a model and applied CO releasing molecule (CORM)-2 to investigate the relationship of CO and HO-1 expression. We herein used Western blot, real-time PCR, promoter activity and EIA to investigate the role of HO-1 expression protecting against thrombin-mediated responses. We found that thrombin-induced COX-2 expression, PGE{sub 2} release and cardiomyocyte hypertrophy markers (increase in ANF/BNP, α-actin expression and cell surface area) was attenuated by pretreatment with CORM-2 which was partially reversed by hemoglobin (Hb) or ZnPP (an inhibitor of HO-1 activity), suggesting that HO-1/CO system may be of clinical importance to ameliorate heart failure through inhibition of inflammatory responses. CORM-2-induced HO-1 protein expression, mRNA and promoter was attenuated by pretreatment with the inhibitors of Pyk2 (PF431396), PDGFR (AG1296), PI3K (LY294002), Akt (SH-5), p38 (SB202530), JNK1/2 (SP600125), FoxO1 (AS1842856) and Sp1 (mithramycin A). The involvement of these signaling components was further confirmed by transfection with respective siRNAs, consistent with those of pharmacological inhibitors. These results suggested that CORM-2-induced HO-1 expression is mediated through a Pyk2/PDGFR/PI3K/Akt/FoxO1/Sp1-dependent manner and exerts a cytoprotective effect in human cardiomyocytes. - Graphical abstract: In summary, CORM-2 treatment induces Pyk2 transactivated PDGFR, which induces PI3K/Akt/MAPK activation, and then recruits Sp1/Foxo1 transcriptional factors to regulate HO-1 gene expression in primary human cardiomyocytes. - Highlights: • CORM-2 induces HO-1 expression. • Pyk2-dependent PDGFR activates PI3K/Akt/MAPK pathway in CORM-2-induced HO

  3. Heme oxygenase-1 prevents non-alcoholic steatohepatitis through suppressing hepatocyte apoptosis in mice

    Fu Na

    2010-10-01

    Full Text Available Abstract Objective Heme oxygenase-1 (HO-1, the rate-limiting enzyme in heme catabolism, has been reported to have potential antioxidant properties. However, the role of HO-1 on hepatocyte apoptosis remains unclear. We aim to elucidate the effects of HO-1 on oxidative stress related hepatocellular apoptosis in nutritional steatohepatitis in mice. Methods C57BL/6J mice were fed with methionine-choline deficient (MCD diet for four weeks to induce hepatic steatohepatitis. HO-1 chemical inducer (hemin, HO-1 chemical inhibitor zinc protoporphyrin IX (ZnPP-IX and/or adenovirus carrying HO-1 gene (Ad-HO-1 were administered to mice, respectively. Hepatocyte apoptosis was evaluated by terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL assay, the mRNA and protein expression of apoptosis related genes were assayed by quantitative real-time PCR and Western blot. Results Hepatocyte signs of oxidative related apoptotic injury were presented in mice fed with MCD diet for 4 weeks. Induction of HO-1 by hemin or Ad-HO-1 significantly attenuated the severity of liver histology, which was associated with decreased hepatic lipid peroxidation content, reduced number of apoptotic cells by TUNEL staining, down-regulated expression of pro-apoptosis related genes including Fas/FasL, Bax, caspase-3 and caspase-9, reduced expression of cytochrome p4502E1 (CYP2E1, inhibited cytochrome c (Cyt-c release, and up-regulated expression of anti-apoptosis gene Bcl-2. Whereas, inhibition of HO-1 by ZnPP-IX caused oxidative stress related hepatic injury, which concomitant with increased number of TUNEL positive cells and up-regulated expression of pro-apoptosis related genes. Conclusions The present study provided evidences for the protective role of HO-1 in preventing nutritional steatohepatitis through suppressing hepatocyte apoptosis in mice.

  4. Role and molecular mechanism of HO-1-mediated NF-κB modulation in fibrosis progression of nonalcoholic steatohepatitis

    NAN Yuemin

    2013-05-01

    Full Text Available ObjectiveTo investigate the potential effects and molecular mechanisms of heme oxygenase-1 (HO-1-mediated modulation of nuclear factor-kappa B (NF-κB, and its downstream activation of intercellular adhesion molecule 1 (ICAM-1 and platelet derived growth factor (PDGF, in fibrosis progression of non-alcoholic steatohepatitis (NASH using the methionine and choline deficient (MCD mouse model of NASH. Methods Forty C57BL/6J male mice (18-20 g were randomly divided into four groups (n=10 each: NASH model group, administered the MCD diet; HO-1 agonist group, administered the MCD diet with intraperitoneal (ip injections of hemin (30 μmol/kg every other day; HO-1 inhibitor group, administered the MCD diet with ip injections of zinc protoporphyrin (ZnPP-IX; 20 μmol/kg every other day; and control group, administered a methionine and choline sufficient (MCS diet, without agonist nor inhibitor injections. After eight weeks, the mice were sacrificed and resected liver tissues used to assess successful model establishment by histological analysis (hematoxylin-eosin and Masson staining and the differential mRNA expression of HO-1, NF-κB, ICAM-1, and PDGF by real-time quantitative PCR (GAPDH normalized and protein expression of HO-1 and PDGF by western blotting ( β-actin normalized. Significance of an intergroup difference was assessed by single-factor analysis of variance test, and the Student-Newman-Keuls test was used for pairwise comparisons. ResultsThe NASH model group showed the appropriate histologic features of hepatic steatosis, necroinflammation and fibrogenesis, while the control group showed normal lobular architecture. In addition, the NASH model group showed significantly higher expression of HO-1, NF-κB, ICAM-1 and PDGF mRNA (all P<0.05, and concomitant increases in HO-1 and PDGF protein. The group treated with HO-1 agonist showed significant down-regulation of the NASH-induced NF-κB, ICAM-1 and PDGF expressions, while the opposite

  5. Heme oxygenase-1 expression protects melanocytes from stress-induced cell death: implications for vitiligo

    Elassiuty, Yasser E.; Klarquist, Jared; Speiser, Jodi; Yousef, Randa M.; El Refaee, Abdelaziz A.; Hunter, Nahla S.; Shaker, Olfat G.; Gundeti, Mohan; Nieuweboer-Krobotova, Ludmila; Caroline Le Poole, I.

    2011-01-01

    To study protection of melanocytes from stress-induced cell death by heme oxygenases during depigmentation and repigmentation in vitiligo, expression of isoforms 1 and 2 was studied in cultured control and patient melanocytes and normal skin explants exposed to UV or bleaching agent 4-TBP.

  6. Heme oxygenase-1 prevents hyperthyroidism induced hepatic damage via an antioxidant and antiapoptotic pathway.

    Giriş, Murat; Erbil, Yeşim; Depboylu, Bilge; Mete, Ozgür; Türkoğlu, Umit; Abbasoğlu, Semra Doğru; Uysal, Müjdat

    2010-12-01

    The exact pathogenesis of hepatic dysfunction in hyperthyroidism is still unknown. We aimed to investigate the pathogenesis of liver dysfunction caused by hyperthyroidism through inducing heme oxygenase-1 (HO-1) expression, which has antioxidant and anti-apoptotic properties. Rats were divided into six groups: untreated (group 1), treated with zinc protoporphyrin (ZnPP) (group 2), treated with hemin (group 3), treated with tri-iodothyronine (T3) (group 4), treated with T3 and ZnPP (group 5), and treated with T3 and hemin (group 6). After 22 d, oxidative stress and antioxidant enzymes and the expression of HO-1, mitochondrial permeability transition, cytochrome c, Bax, Bcl-2, caspase-3, caspase-8, and caspase-3 activity, and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay were examined. Hyperthyroidism induced oxidative stress of liver tissue was ameliorated by HO-1 induction. Administration of hemin (HO-1 inducer) increased Bcl-2 expression. Decreased expression of cytochrome c was accompanied by a decrease in caspase-3, caspase-8, Bax expression, and caspase-3 activity. The apoptotic activity and oxidative damage were found to be increased by the administration of ZnPP (HO-1 inhibitor). Immunohistochemistry findings supported these results. HO-1 induction plays a protective role in the pathogenesis of the liver dysfunction in hyperthyroidism. This effect is dependent on modulation of the antiapoptotic and antioxidative pathways by HO-1 expression. Copyright © 2010 Elsevier Inc. All rights reserved.

  7. L-ascorbate attenuates methamphetamine neurotoxicity through enhancing the induction of endogenous heme oxygenase-1.

    Huang, Ya-Ni; Wang, Jiz-Yuh; Lee, Ching-Tien; Lin, Chih-Hung; Lai, Chien-Cheng; Wang, Jia-Yi

    2012-12-01

    Methamphetamine (METH) is a drug of abuse which causes neurotoxicity and increased risk of developing neurodegenerative diseases. We previously found that METH induces heme oxygenase (HO)-1 expression in neurons and glial cells, and this offers partial protection against METH toxicity. In this study, we investigated the effects of l-ascorbate (vitamin C, Vit. C) on METH toxicity and HO-1 expression in neuronal/glial cocultures. Cell viability and damage were evaluated by 3-(4,5-dimethylthianol-2-yl)-2,5 diphenyl tetrazolium bromide (MTT) reduction and lactate dehydrogenase (LDH) release, respectively. Neuronal and glial localization of HO-1 were identified by double immunofluorescence staining. Reactive oxygen species (ROS) production was measured using the fluorochrome 2',7'-dichlorofluorescin diacetate. HO-1 mRNA and protein expression were examined by RT-qPCR and Western blotting, respectively. Results show that Vit. C induced HO-1 mRNA and protein expressions in time- and concentration-dependent manners. Inhibition of p38 mitogen-activated protein kinase (MAPK) but not extracellular signal-regulated kinase (ERK) significantly blocked induction of HO-1 by Vit. C. HO-1 mRNA and protein expressions were significantly elevated by a combination of Vit. C and METH, compared to either Vit. C or METH alone. Pretreatment with Vit. C enhanced METH-induced HO-1 expression and attenuated METH-induced ROS production and neurotoxicity. Pharmacological inhibition of HO activity abolished suppressive effects of Vit. C on METH-induced ROS production and attenuated neurotoxicity. We conclude that induction of HO-1 expression contributes to the attenuation of METH-induced ROS production and neurotoxicity by Vit. C. We suggest that HO-1 induction by Vit. C may serve as a strategy to alleviate METH neurotoxicity. Copyright © 2012 Elsevier Inc. All rights reserved.

  8. Genipin inhibits TNF-α-induced vascular smooth muscle cell proliferation and migration via induction of HO-1.

    Fengrong Jiang

    Full Text Available Vascular smooth muscle cell (VSMC proliferation and migration triggered by inflammatory stimuli contributes importantly to the pathogenesis of atherosclerosis and restenosis. On the other hand, genipin, an aglycon of geniposide, exhibits diverse pharmacological functions such as antitumor and anti-inflammatory effects. The protective effects of genipin on the cardiovascular system have also been reported. However, the molecular mechanism involved remains unknown. This study aimed to elucidate the precise function of genipin in VSMCs, focusing particularly on the role of heme oxygenase-1 (HO-1, a potent anti-inflammatory enzyme. We found that pretreatment of genipin induced HO-1 mRNA and protein levels, as well as its activity in VSMCs. Genipin inhibited TNF-α-induced VSMC proliferation and migration in a dose-dependent manner. At the molecular level, genipin prevented ERK/MAPK and Akt phosphorylation while left p38 MAPK and JNK unchanged. Genipin also blocked the increase of ROS generation induced by TNF-α. More importantly, the specific HO-1 siRNA partially abolished the beneficial effects of genipin on VSMCs. These results suggest that genipin may serve as a novel drug in the treatment of these pathologies by inducing HO-1 expression/activity and subsequently decreasing VSMC proliferation and migration.

  9. Heme oxygenase-1 induction alters chemokine regulation and ameliorates human immunodeficiency virus-type-1 infection in lipopolysaccharide-stimulated macrophages

    Zhou, Zhao-Hua; Kumari, Namita; Nekhai, Sergei; Clouse, Kathleen A.; Wahl, Larry M.; Yamada, Kenneth M.; Dhawan, Subhash

    2013-01-01

    Highlights: •Lipopolysaccharide stimulation of heme oxygenase-1 (HO-1) ameliorated HIV-1 infection of primary human macrophages. •The partial protection by HO-1 against HIV infection was associated with induction of chemokines such as MIP1α and MIP1β. •This mechanism explains lipopolysaccharide-stimulated HO-1-mediated inhibition of HIV-1 infection of macrophages. -- Abstract: We have elucidated a putative mechanism for the host resistance against HIV-1 infection of primary human monocyte-derived macrophages (MDM) stimulated with lipopolysaccharide (LPS). We show that LPS-activated MDM both inhibited HIV-1 entry into the cells and were refractory to post-entry productive viral replication. LPS-treated cells were virtually negative for mature virions as revealed by transmission electron microscopy. LPS activation of MDM markedly enhanced the expression of heme oxygenase-1 (HO-1), a potent inducible cytoprotective enzyme. Increased HO-1 expression was accompanied by elevated production of macrophage inflammatory chemokines (MIP1α and MIP1β) by LPS-activated MDM, significantly decreased surface chemokine receptor-5 (CCR-5) expression, and substantially reduced virus replication. Treatment of cells with HO-1 inhibitor SnPP IX (tin protoporphyrin IX) attenuated the LPS-mediated responses, HIV-1 replication and secretion of MIP1α, MIP1β, and LD78β chemokines with little change in surface CCR-5 expression. These results identify a novel role for HO-1 in the modulation of host immune response against HIV infection of MDM

  10. Red Yeast Rice Protects Circulating Bone Marrow-Derived Proangiogenic Cells against High-Glucose-Induced Senescence and Oxidative Stress: The Role of Heme Oxygenase-1

    Jung-Tung Liu

    2017-01-01

    Full Text Available The inflammation and oxidative stress of bone marrow-derived proangiogenic cells (PACs, also named endothelial progenitor cells, triggered by hyperglycemia contributes significantly to vascular dysfunction. There is supporting evidence that the consumption of red yeast rice (RYR; Monascus purpureus-fermented rice reduces the vascular complications of diabetes; however, the underlying mechanism remains unclear. This study aimed to elucidate the effects of RYR extract in PACs, focusing particularly on the role of a potent antioxidative enzyme, heme oxygenase-1 (HO-1. We found that treatment with RYR extract induced nuclear factor erythroid-2-related factor nuclear translocation and HO-1 mRNA and protein levels in PACs. RYR extract inhibited high-glucose-induced (30 mM PAC senescence and the development of reactive oxygen species (ROS in a dose-dependent manner. The HO-1 inducer cobalt protoporphyrin IX also decreased high-glucose-induced cell senescence and oxidative stress, whereas the HO-1 enzyme inhibitor zinc protoporphyrin IX and HO-1 small interfering RNA significantly reversed RYR extract-caused inhibition of senescence and reduction of oxidative stress in high-glucose-treated PACs. These results suggest that RYR extract serves as alternative and complementary medicine in the treatment of these diseases, by inducing HO-1, thereby decreasing the vascular complications of diabetes.

  11. Totarol prevents neuronal injury in vitro and ameliorates brain ischemic stroke: Potential roles of Akt activation and HO-1 induction

    Gao, Yuanxue; Xu, Xiaojun; Chang, Sai; Wang, Yunjie; Xu, Yazhou; Ran, Siqi [Jiangsu Key Laboratory of Drug Screening, State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009 (China); Huang, Zhangjian [Jiangsu Key Laboratory of Drug Discovery for Metabolic Diseases, China Pharmaceutical University, Nanjing 210009 (China); Li, Ping [Jiangsu Key Laboratory of Drug Screening, State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009 (China); Li, Jia [National Center for Drug Screening, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 189 Guo Shoujing Road, Shanghai 201203 (China); Zhang, Luyong [Jiangsu Key Laboratory of Drug Screening, State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009 (China); Saavedra, Juan M. [Department of Pharmacology and Physiology, Georgetown University Medical Center, Washington, DC 20057 (United States); Liao, Hong, E-mail: liaohong56@hotmail.com [Jiangsu Key Laboratory of Drug Screening, State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009 (China); Pang, Tao, E-mail: tpang@cpu.edu.cn [Jiangsu Key Laboratory of Drug Screening, State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009 (China); Department of Pharmacology and Physiology, Georgetown University Medical Center, Washington, DC 20057 (United States)

    2015-12-01

    The natural product totarol, a phenolic diterpenoid and a major constituent isolated from the sap of Podocarpus totara, has been reported to have a potent antimicrobial activity. In this study, we determined whether totarol possessed an additional neuroprotective activity in vitro and in vivo. We found that totarol prevented glutamate- and oxygen and glucose deprivation-induced neuronal death in primary rat cerebellar granule neuronal cells and cerebral cortical neurons. Totarol increased Akt and GSK-3β phosphorylation, Nrf2 and heme oxygenase-1 (HO-1) protein expressions and suppressed oxidative stress by increasing GSH and SOD activities. The PI3K/Akt inhibitor LY294002 prevented totarol neuroprotective effect by suppressing the totarol-induced changes in HO-1 expression and the activities of GSH and SOD. The HO-1 inhibitor ZnPPIX also prevented totarol-increased GSH and SOD activities. In a model of acute cerebral ischemic injury in Sprague–Dawley rats, produced by occlusion of the middle cerebral artery for 2 h followed by 22 h or 46 h of reperfusion, totarol significantly reduced infarct volume and improved the neurological deficit. In this model, totarol increased HO-1 expression and the activities of GSH and SOD. These observations suggest that totarol may be a novel activator of the Akt/HO-1 pathway protecting against ischemic stroke through reduction of oxidative stress. - Graphical abstract: It is unknown whether the natural product totarol has neuroprotective effects in vitro and in vivo. This study underscores that totarol prevents neuronal injury in vitro, not only by activating PI3K/Akt pathway, but also via induction of Nrf2, HO-1, GSH and SOD expressions. Totarol also ameliorated acute cerebral ischemic injury in a rat ischemic stroke model. The findings highlight that totarol may be exploited for protecting against ischemic stroke through Akt/HO-1 pathway. - Highlights: • Totarol protects glutamate- and OGD-induced neuronal injury in vitro.

  12. Totarol prevents neuronal injury in vitro and ameliorates brain ischemic stroke: Potential roles of Akt activation and HO-1 induction

    Gao, Yuanxue; Xu, Xiaojun; Chang, Sai; Wang, Yunjie; Xu, Yazhou; Ran, Siqi; Huang, Zhangjian; Li, Ping; Li, Jia; Zhang, Luyong; Saavedra, Juan M.; Liao, Hong; Pang, Tao

    2015-01-01

    The natural product totarol, a phenolic diterpenoid and a major constituent isolated from the sap of Podocarpus totara, has been reported to have a potent antimicrobial activity. In this study, we determined whether totarol possessed an additional neuroprotective activity in vitro and in vivo. We found that totarol prevented glutamate- and oxygen and glucose deprivation-induced neuronal death in primary rat cerebellar granule neuronal cells and cerebral cortical neurons. Totarol increased Akt and GSK-3β phosphorylation, Nrf2 and heme oxygenase-1 (HO-1) protein expressions and suppressed oxidative stress by increasing GSH and SOD activities. The PI3K/Akt inhibitor LY294002 prevented totarol neuroprotective effect by suppressing the totarol-induced changes in HO-1 expression and the activities of GSH and SOD. The HO-1 inhibitor ZnPPIX also prevented totarol-increased GSH and SOD activities. In a model of acute cerebral ischemic injury in Sprague–Dawley rats, produced by occlusion of the middle cerebral artery for 2 h followed by 22 h or 46 h of reperfusion, totarol significantly reduced infarct volume and improved the neurological deficit. In this model, totarol increased HO-1 expression and the activities of GSH and SOD. These observations suggest that totarol may be a novel activator of the Akt/HO-1 pathway protecting against ischemic stroke through reduction of oxidative stress. - Graphical abstract: It is unknown whether the natural product totarol has neuroprotective effects in vitro and in vivo. This study underscores that totarol prevents neuronal injury in vitro, not only by activating PI3K/Akt pathway, but also via induction of Nrf2, HO-1, GSH and SOD expressions. Totarol also ameliorated acute cerebral ischemic injury in a rat ischemic stroke model. The findings highlight that totarol may be exploited for protecting against ischemic stroke through Akt/HO-1 pathway. - Highlights: • Totarol protects glutamate- and OGD-induced neuronal injury in vitro.

  13. Involvement of heme oxygenase-1 in β-cyclodextrin-hemin complex-induced cucumber adventitious rooting process.

    Lin, Yuting; Li, Meiyue; Huang, Liqin; Shen, Wenbiao; Ren, Yong

    2012-09-01

    Our previous results showed that β-cyclodextrin-hemin complex (CDH) exhibited a vital protective role against cadmium-induced oxidative damage and toxicity in alfalfa seedling roots by the regulation of heme oxygenase-1 (HO-1) gene expression. In this report, we further test whether CDH exhibited the hormonal-like response. The application of CDH and an inducer of HO-1, hemin, were able to induce the up-regulation of cucumber HO-1 gene (CsHO1) expression and thereafter the promotion of adventitious rooting in cucumber explants. The effect is specific for HO-1 since the potent HO-1 inhibitor zinc protoporphyrin IX (ZnPP) blocked the above responses triggered by CDH, and the inhibitory effects were reversed further when 30% saturation of CO aqueous solution was added together. Further, molecular evidence showed that CDH triggered the increases of the HO-1-mediated target genes responsible for adventitious rooting, including one DnaJ-like gene (CsDNAJ-1) and two calcium-dependent protein kinase (CDPK) genes (CsCDPK1 and CsCDPK5), and were inhibited by ZnPP and reversed by CO. The calcium (Ca2+) chelator ethylene glycol-bis (2-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA) and the Ca2+ channel blocker lanthanum chloride (LaCl3) not only compromised the induction of adventitious rooting induced by CDH but also decreased the transcripts of above three target genes. However, the application of ascorbic acid (AsA), a well-known antioxidant in plants, failed to exhibit similar inducible effect on adventitious root formation. In short, above results illustrated that the response of CDH in the induction of cucumber adventitious rooting might be through HO-1-dependent mechanism and calcium signaling. Physiological, pharmacological and molecular evidence showed that β-cyclodextrin-hemin complex (CDH) was able to induce cucumber adventitious rooting through heme oxygenase-1 (HO-1)-dependent mechanism and calcium signaling.

  14. Cytoprotective effect exerted by geraniin in HepG2 cells is through microRNA mediated regulation of BACH-1 and HO-1.

    Aayadi, Hoda; Mittal, Smriti P K; Deshpande, Anjali; Gore, Makarand; Ghaskadbi, Saroj S

    2017-11-01

    Geraniin, a hydrolysable tannin, used in traditional medicine in Southeast Asia, is known to exhibit various biological activities. As an antioxidant it is known to up-regulate phase II enzyme Heme oxygenase-1 (HO-1). However its mechanism is not clearly understood. Nuclear factor erythroid-derived 2 related factor 2 (Nrf-2) is transcriptionally up-regulated by Extracellular signal-regulated kinase (ERK) 1/2 and retained in nucleus due to inactivated Glycogen synthase kinase 3 beta (GSK-3β). Geraniin additionally down-regulates expression of microRNA 217 and 377 (miR-217 and miR-377) which target HO-1 mRNA. Expression of BTB and CNC homolog 1 (BACH-1), another regulator of HO-1, is also down-regulated by up-regulating microRNA 98 (miR-98), a negative regulator of BACH-1. Thus, geraniin up-regulates HO-1 expression both through activating its positive regulator Nrf-2 and by down-regulating its negative regulator BACH-1. Up-regulation of HO-1 also confers protection to HepG2 cells from tertiary butyl hydroperoxide (TBH) induced cytotoxicity. [BMB Reports 2017; 50(11): 560-565].

  15. Targeted expression of heme oxygenase-1 prevents the pulmonary inflammatory and vascular responses to hypoxia

    Minamino, Tohru; Christou, Helen; Hsieh, Chung-Ming; Liu, Yuxiang; Dhawan, Vijender; Abraham, Nader G.; Perrella, Mark A.; Mitsialis, S. Alex; Kourembanas, Stella

    2001-07-01

    Chronic hypoxia causes pulmonary hypertension with smooth muscle cell proliferation and matrix deposition in the wall of the pulmonary arterioles. We demonstrate here that hypoxia also induces a pronounced inflammation in the lung before the structural changes of the vessel wall. The proinflammatory action of hypoxia is mediated by the induction of distinct cytokines and chemokines and is independent of tumor necrosis factor- signaling. We have previously proposed a crucial role for heme oxygenase-1 (HO-1) in protecting cardiomyocytes from hypoxic stress, and potent anti-inflammatory properties of HO-1 have been reported in models of tissue injury. We thus established transgenic mice that constitutively express HO-1 in the lung and exposed them to chronic hypoxia. HO-1 transgenic mice were protected from the development of both pulmonary inflammation as well as hypertension and vessel wall hypertrophy induced by hypoxia. Significantly, the hypoxic induction of proinflammatory cytokines and chemokines was suppressed in HO-1 transgenic mice. Our findings suggest an important protective function of enzymatic products of HO-1 activity as inhibitors of hypoxia-induced vasoconstrictive and proinflammatory pathways.

  16. Fibroblast growth factor 10 protects neuron against oxygen–glucose deprivation injury through inducing heme oxygenase-1

    Li, Yong-Hua; Yang, Li-Ye; Chen, Wei; Li, Ying-Ke; Yuan, Hong-Bin

    2015-01-01

    Highlights: • FGF10 attenuates OGD induced injury in cortical neuron. • FGF10 reduces OGD triggered ROS level in cortical neuron. • FGF10 induces HO-1 expression upon OGD stimuli in cortical neuron. • Knockdown of HO-1 impairs the neuroprotection of FGF10 in OGD model. - Abstract: Fibroblast growth factors (FGFs) are a family of structurally related heparin-binding proteins with diverse biological functions. FGFs participate in mitogenesis, angiogenesis, cell proliferation, development, differentiation and cell migration. Here, we investigated the potential effect of FGF10, a member of FGFs, on neuron survival in oxygen–glucose deprivation (OGD) model. In primary cultured mouse cortical neurons upon OGD, FGF10 treatment (100 and 1000 ng/ml) attenuated the decrease of cell viability and rescued the LDH release. Tuj-1 immunocytochemistry assay showed that FGF10 promoted neuronal survival. Apoptosis assay with Annexin V + PI by flow cytometry demonstrated that FGF10 treatment reduced apoptotic cell proportion. Moreover, immunoblotting showed that FGF10 alleviated the cleaved caspase-3 upregulation caused by OGD. FGF10 treatment also depressed the OGD-induced increase of caspase-3, -8 and -9 activities. At last, we found FGF10 triggered heme oxygenase-1 (HO-1) protein expression rather than hypoxia-inducible factor-1 (HIF-1), AMP-activated protein kinase (AMPK) signaling and extracellular signal-regulated kinases 1/2 (ERK1/2) signaling. Knockdown of HO-1 by siRNA partly abolished the neuroprotection of FGF10 in OGD model. In summary, our observations provide the first evidence for the neuroprotective function of FGF10 against ischemic neuronal injury and suggest that FGF10 may be a promising agent for treatment of ischemic stroke

  17. Fibroblast growth factor 10 protects neuron against oxygen–glucose deprivation injury through inducing heme oxygenase-1

    Li, Yong-Hua; Yang, Li-Ye; Chen, Wei; Li, Ying-Ke, E-mail: liyingke6f@126.com; Yuan, Hong-Bin, E-mail: yuanhongbin6f@126.com

    2015-01-02

    Highlights: • FGF10 attenuates OGD induced injury in cortical neuron. • FGF10 reduces OGD triggered ROS level in cortical neuron. • FGF10 induces HO-1 expression upon OGD stimuli in cortical neuron. • Knockdown of HO-1 impairs the neuroprotection of FGF10 in OGD model. - Abstract: Fibroblast growth factors (FGFs) are a family of structurally related heparin-binding proteins with diverse biological functions. FGFs participate in mitogenesis, angiogenesis, cell proliferation, development, differentiation and cell migration. Here, we investigated the potential effect of FGF10, a member of FGFs, on neuron survival in oxygen–glucose deprivation (OGD) model. In primary cultured mouse cortical neurons upon OGD, FGF10 treatment (100 and 1000 ng/ml) attenuated the decrease of cell viability and rescued the LDH release. Tuj-1 immunocytochemistry assay showed that FGF10 promoted neuronal survival. Apoptosis assay with Annexin V + PI by flow cytometry demonstrated that FGF10 treatment reduced apoptotic cell proportion. Moreover, immunoblotting showed that FGF10 alleviated the cleaved caspase-3 upregulation caused by OGD. FGF10 treatment also depressed the OGD-induced increase of caspase-3, -8 and -9 activities. At last, we found FGF10 triggered heme oxygenase-1 (HO-1) protein expression rather than hypoxia-inducible factor-1 (HIF-1), AMP-activated protein kinase (AMPK) signaling and extracellular signal-regulated kinases 1/2 (ERK1/2) signaling. Knockdown of HO-1 by siRNA partly abolished the neuroprotection of FGF10 in OGD model. In summary, our observations provide the first evidence for the neuroprotective function of FGF10 against ischemic neuronal injury and suggest that FGF10 may be a promising agent for treatment of ischemic stroke.

  18. Transgenic expression of human heme oxygenase-1 in pigs confers resistance against xenograft rejection during ex vivo perfusion of porcine kidneys.

    Petersen, Björn; Ramackers, Wolf; Lucas-Hahn, Andrea; Lemme, Erika; Hassel, Petra; Queisser, Anna-Lisa; Herrmann, Doris; Barg-Kues, Brigitte; Carnwath, Joseph W; Klose, Johannes; Tiede, Andreas; Friedrich, Lars; Baars, Wiebke; Schwinzer, Reinhard; Winkler, Michael; Niemann, Heiner

    2011-01-01

    The major immunological hurdle to successful porcine-to-human xenotransplantation is the acute vascular rejection (AVR), characterized by endothelial cell (EC) activation and perturbation of coagulation. Heme oxygenase-1 (HO-1) and its derivatives have anti-apoptotic, anti-inflammatory effects and protect against reactive oxygen species, rendering HO-1 a promising molecule to control AVR. Here, we report the production and characterization of pigs transgenic for human heme oxygenase-1 (hHO-1) and demonstrate significant protection in porcine kidneys against xenograft rejection in ex vivo perfusion with human blood and transgenic porcine aortic endothelial cells (PAEC) in a TNF-α-mediated apoptosis assay. Transgenic and non-transgenic PAEC were tested in a TNF-α-mediated apoptosis assay. Expression of adhesion molecules (ICAM-1, VCAM-1, and E-selectin) was measured by real-time PCR. hHO-1 transgenic porcine kidneys were perfused with pooled and diluted human AB blood in an ex vivo perfusion circuit. MHC class-II up-regulation after induction with IFN-γ was compared between wild-type and hHO-1 transgenic PAEC. Cloned hHO-1 transgenic pigs expressed hHO-1 in heart, kidney, liver, and in cultured ECs and fibroblasts. hHO-1 transgenic PAEC were protected against TNF-α-mediated apoptosis. Real-time PCR revealed reduced expression of adhesion molecules like ICAM-1, VCAM-1, and E-selectin. These effects could be abrogated by the incubation of transgenic PAECs with the specific HO-1 inhibitor zinc protoporphorine IX (Zn(II)PPIX, 20 μm). IFN-γ induced up-regulation of MHC class-II molecules was significantly reduced in PAECs from hHO-1 transgenic pigs. hHO-1 transgenic porcine kidneys could successfully be perfused with diluted human AB-pooled blood for a maximum of 240 min (with and without C1 inh), while in wild-type kidneys, blood flow ceased after ∼60 min. Elevated levels of d-Dimer and TAT were detected, but no significant consumption of fibrinogen and

  19. Heme-Oxygenase-1 Expression Contributes to the Immunoregulation Induced by Fasciola hepatica and Promotes Infection

    Paula Carasi

    2017-07-01

    Full Text Available Fasciola hepatica, also known as the liver fluke, is a trematode that infects livestock and humans causing fasciolosis, a zoonotic disease of increasing importance due to its worldwide distribution and high economic losses. This parasite immunoregulates the host immune system by inducing a strong Th2 and regulatory T immune response by immunomodulating dendritic cell (DC maturation and alternative activation of macrophages. In this paper, we show that F. hepatica infection in mice induces the upregulation of heme-oxygenase-1 (HO-1, the rate-limiting enzyme in the catabolism of free heme that regulates the host inflammatory response. We show and characterize two different populations of antigen presenting cells that express HO-1 during infection in the peritoneum of infected animals. Cells that expressed high levels of HO-1 expressed intermediate levels of F4/80 but high expression of CD11c, CD38, TGFβ, and IL-10 suggesting that they correspond to regulatory DCs. On the other hand, cells expressing intermediate levels of HO-1 expressed high levels of F4/80, CD68, Ly6C, and FIZZ-1, indicating that they might correspond to alternatively activated macrophages. Furthermore, the pharmacological induction of HO-1 with the synthetic metalloporphyrin CoPP promoted F. hepatica infection increasing the clinical signs associated with the disease. In contrast, treatment with the HO-1 inhibitor SnPP protected mice from parasite infection, indicating that HO-1 plays an essential role during F. hepatica infection. Finally, HO-1 expression during F. hepatica infection was associated with TGFβ and IL-10 levels in liver and peritoneum, suggesting that HO-1 controls the expression of these immunoregulatory cytokines during infection favoring parasite survival in the host. These results contribute to the elucidation of the immunoregulatory mechanisms induced by F. hepatica in the host and provide alternative checkpoints to control fasciolosis.

  20. Production and characterization of soluble human TNFRI-Fc and human HO-1(HMOX1) transgenic pigs by using the F2A peptide.

    Park, Sol Ji; Cho, Bumrae; Koo, Ok Jae; Kim, Hwajung; Kang, Jung Taek; Hurh, Sunghoon; Kim, Su Jin; Yeom, Hye Jung; Moon, Joonho; Lee, Eun Mi; Choi, Ji Yei; Hong, Ju Ho; Jang, Goo; Hwang, Joing-Ik; Yang, Jaeseok; Lee, Byeong Chun; Ahn, Curie

    2014-06-01

    Generation of transgenic pigs for xenotransplantation is one of the most promising technologies for resolving organ shortages. Human heme oxygenase-1 (hHO-1/HMOX1) can protect transplanted organs by its strong anti-oxidative, anti-apoptotic, and anti-inflammatory effects. Soluble human TNFRI-Fc (shTNFRI-Fc) can inhibit the binding of human TNF-α (hTNF-α) to TNF receptors on porcine cells, and thereby, prevent hTNF-α-mediated inflammation and apoptosis. Herein, we successfully generated shTNFRI-Fc-F2A-HA-hHO-1 transgenic (TG) pigs expressing both shTNFRI-Fc and hemagglutinin-tagged-human heme oxygenase-1 (HA-hHO-1) by using an F2A self-cleaving peptide. shTNFRI-Fc and HA-hHO-1 transgenes containing the F2A peptide were constructed under the control of the CAG promoter. Transgene insertion and copy number in the genome of transgenic pigs was confirmed by polymerase chain reaction (PCR) and Southern blot analysis. Expressions of shTNFRI-Fc and HA-hHO-1 in TG pigs were confirmed using PCR, RT-PCR, western blot, ELISA, and immunohistochemistry. shTNFRI-Fc and HA-hHO-1 were expressed in various organs, including the heart, lung, and spleen. ELISA assays detected shTNFRI-Fc in the sera of TG pigs. For functional analysis, fibroblasts isolated from a shTNFRI-Fc-F2A-HA-hHO-1 TG pig (i.e., #14; 1 × 10(5) cells) were cultured with hTNF-α (20 ng/mL) and cycloheximide (10 μg/mL). The viability of shTNFRI-Fc-F2A-HA-hHO-1 TG pig fibroblasts was significantly higher than that of the wild type (wild type vs. shTNFRI-Fc-F2A-HA-hHO-1 TG at 24 h, 31.6 ± 3.2 vs. 60.4 ± 8.3 %, respectively; p hHO-1 TG pig fibroblasts was lower than that of the wild type pig fibroblasts (wild type vs. shTNFRI-Fc-F2A-HA-hHO-1 TG at 12 h, 812,452 ± 113,078 RLU vs. 88,240 ± 10,438 RLU, respectively; p hHO-1 TG pigs generated by the F2A self-cleaving peptide express both shTNFRI-Fc and HA-hHO-1 molecules, which provides protection against oxidative and inflammatory injury

  1. [hHO-1 structure prediction and its mutant construct, expression, purification and activity analysis].

    Xia, Zhen Wei; Cui, Wen Jun; Zhou, Wen Pu; Zhang, Xue Hong; Shen, Qing Xiang; Li, Yun Zhu; Yu, Shan Chang

    2004-10-01

    Human Heme Oxygenase-1 (hHO-1) is the rate-limiting enzyme in the catabolism reaction of heme, which directly regulates the concentration of bilirubin in human body. The mutant structure was simulated by Swiss-pdbviewer procedure, which showed that the structure of active pocket was changed distinctly after Ala25 substituted for His25 in active domain, but the mutated enzyme still binded with heme. On the basis of the results, the expression vectors, pBHO-1 and pBHO-1(M), were constructed, induced by IPTG and expressed in E. coli DH5alpha strain. The expression products were purified with 30%-60% saturation (NH4)2SO4 and Q-Sepharose Fast Flow column chromatography. The concentration of hHO-1 in 30%-60% saturation (NH4)2SO4 components and in fractions through twice column chromatography was 3.6-fold and 30-fold higher than that in initial product, respectively. The activity of wild hHO-1 (whHO-1) and mutant hHO-1 (deltahHO-1) showed that the activity of deltahHO-1 was reduced 91.21% compared with that of whHO-1. The study shows that His25 is of importance for the mechanism of hHO-1, and provides the possibility for effectively regulating the activity to exert biological function.

  2. Methamphetamine induces heme oxygenase-1 expression in cortical neurons and glia to prevent its toxicity

    Huang, Y.-N.; Wu, C.-H.; Lin, T.-C.; Wang, J.-Y.

    2009-01-01

    The impairment of cognitive and motor functions in humans and animals caused by methamphetamine (METH) administration underscores the importance of METH toxicity in cortical neurons. The heme oxygenase-1 (HO-1) exerts a cytoprotective effect against various neuronal injures; however, it remains unclear whether HO-1 is involved in METH-induced toxicity. We used primary cortical neuron/glia cocultures to explore the role of HO-1 in METH-induced toxicity. Exposure of cultured cells to various concentrations of METH (0.1, 0.5, 1, 3, 5, and 10 mM) led to cytotoxicity in a concentration-dependent manner. A METH concentration of 5 mM, which caused 50% of neuronal death and glial activation, was chosen for subsequent experiments. RT-PCR and Western blot analysis revealed that METH significantly induced HO-1 mRNA and protein expression, both preceded cell death. Double and triple immunofluorescence staining further identified HO-1-positive cells as activated astrocytes, microglia, and viable neurons, but not dying neurons. Inhibition of the p38 mitogen-activated protein kinase pathway significantly blocked HO-1 induction by METH and aggravated METH neurotoxicity. Inhibition of HO activity using tin protoporphyrine IX significantly reduced HO activity and exacerbated METH neurotoxicity. However, prior induction of HO-1 using cobalt protoporphyrine IX partially protected neurons from METH toxicity. Taken together, our results suggest that induction of HO-1 by METH via the p38 signaling pathway may be protective, albeit insufficient to completely protect cortical neurons from METH toxicity.

  3. Isocyanides inhibit human heme oxygenases at the verdoheme stage.

    Evans, John P; Kandel, Sylvie; Ortiz de Montellano, Paul R

    2009-09-22

    Heme oxygenases (HO) catalyze the oxidative cleavage of heme to generate biliverdin, CO, and free iron. In humans, heme oxygenase-1 (hHO-1) is overexpressed in tumor tissues, where it helps to protect cancer cells from anticancer agents, while HOs in fungal pathogens, such as Candida albicans, function as the primary means of iron acquisition. Thus, HO can be considered a potential therapeutic target for certain diseases. In this study, we have examined the equilibrium binding of three isocyanides, isopropyl, n-butyl, and benzyl, to the two major human HO isoforms (hHO-1 and hHO-2), Candida albicans HO (CaHmx1), and human cytochrome P450 CYP3A4 using electronic absorption spectroscopy. Isocyanides coordinate to both ferric and ferrous HO-bound heme, with tighter binding by the more hydrophobic isocyanides and 200-300-fold tighter binding to the ferrous form. Benzyl isocyanide was the strongest ligand to ferrous heme in all the enzymes. Because the dissociation constants (KD) of the ligands for ferrous heme-hHO-1 were below the limit of accuracy for equilibrium titrations, stopped-flow kinetic experiments were used to measure the binding parameters of the isocyanides to ferrous hHO-1. Steady-state activity assays showed that benzyl isocyanide was the most potent uncompetitive inhibitor with respect to heme with a KI = 0.15 microM for hHO-1. Importantly, single turnover assays revealed that the reaction was completely stopped by coordination of the isocyanide to the verdoheme intermediate rather than to the ferric heme complex. Much tighter binding of the inhibitor to the verdoheme intermediate differentiates it from inhibition of, for example, CYP3A4 and offers a possible route to more selective inhibitor design.

  4. Isocyanides Inhibit Human Heme Oxygenases at the Verdoheme Stage†

    Evans, John P.; Kandel, Sylvie; Ortiz de Montellano, Paul R.

    2010-01-01

    Heme oxygenases (HO) catalyze the oxidative cleavage of heme to generate biliverdin, CO, and free iron. In humans, heme oxygenase-1 (hHO-1) is overexpressed in tumor tissues, where it helps to protect cancer cells from anticancer agents, while HOs in fungal pathogens, such as Candida albicans, function as the primary means of iron acquisition. Thus, HO can be considered a potential therapeutic target for certain diseases. In this study, we have examined the equilibrium binding of three isocyanides; isopropyl, n-butyl, and benzyl, to the two major human HO isoforms (hHO-1 and hHO-2), Candida albicans HO (CaHmx1), and human cytochrome P450 CYP3A4 using electronic absorption spectroscopy. Isocyanides coordinate to both ferric and ferrous HO-bound heme, with tighter binding by the more hydrophobic isocyanides, and 200-300-fold tighter binding to the ferrous form. Benzyl isocyanide was the strongest ligand to ferrous heme in all the enzymes. Because the dissociation constants (KD) of the ligands for ferrous heme-hHO-1 were below the limit of accuracy for equilibrium titrations, stopped-flow kinetic experiments were used to measure the binding parameters of the isocyanides to ferrous hHO-1. Steady-state activity assays showed that benzyl isocyanide was the most potent uncompetitive inhibitor with respect to heme with a KI = 0.15 μM for hHO-1. Importantly, single turnover assays revealed that the reaction was completely stopped by coordination of the isocyanide to the verdoheme intermediate rather than to the ferric heme complex. Much tighter binding of the inhibitor to the verdoheme intermediate differentiates it from inhibition of, for example, CYP3A4 and offers a possible route to more selective inhibitor design. PMID:19694439

  5. Adenovirus-mediated heme oxygenase-1 gene transfer into rabbit ocular tissues.

    Abraham, N G; da Silva, J L; Lavrovsky, Y; Stoltz, R A; Kappas, A; Dunn, M W; Schwartzman, M L

    1995-10-01

    Heme oxygenase-1 (HO-1) is a stress protein induced up to 100-fold within a few hours after exposure to oxidative stress, and it has been shown to counteract oxidative injury induced by ultraviolet light or free radicals. The current study was undertaken to determine whether the HO-1 gene can be introduced into adult rabbit ocular tissues by microinjection of a recombinant replication-deficient adenovirus human HO-1 cDNA (Adv-HHO). Human HO-1 gene was used for transfection studies to differentiate endogenous from transfected HO. The purified Adv-HHO construct (10(8) pfu/ml) was mixed with lipofectamine and microinjected into the anterior chamber, vitreous cavity, and subretinal space of New Zealand rabbit eyes. After 2 weeks, total RNA was extracted from different ocular tissues, reverse transcription-polymerase chain reaction was performed using specific human HO-1 primers, and amplification products were subjected to Southern hybridization. Transfection with the Adv-HHO construct into rabbit corneal epithelial cells in culture resulted in a functional expression of the human HO-1 gene; the human HO-1 mRNA was detected, and enzyme activity increased threefold. Human HO-1 mRNA was detected in the retina after microinjection of the Adv-HHO construct into the subretinal space. Microinjection into the vitreous resulted in HO-1 mRNA expression in the corneal endothelium, iris, lens, and retina; after intracameral injection of the Adv-HHO construct, human HO-1 mRNA was detected in corneal epithelium and endothelium, ciliary body, lens, and iris. Regardless of the injection site, transfected human HO-1 mRNA was undetectable in tissues outside the eye, that is, brain, liver, and kidney. These results demonstrated a tissue-selective functional transfer of the human HO-1 gene into rabbit ocular tissues in vivo. This technique may be a promising means for delivering HO-1 gene in vivo as a protective mechanism against oxidative stress that contributes to the pathogenesis of

  6. Heme oxygenase up-regulation under ultraviolet-B radiation is not epigenetically restricted and involves specific stress-related transcriptions factors

    Diego Santa-Cruz

    2017-08-01

    Full Text Available Heme oxygenase-1 (HO-1 plays a protective role against oxidative stress in plants. The mechanisms regulating its expression, however, remain unclear. Here we studied the methylation state of a GC rich HO-1 promoter region and the expression of several stress-related transcription factors (TFs in soybean plants subjected to ultraviolet-B (UV-B radiation. Genomic DNA and total RNA were isolated from leaves of plants irradiated with 7.5 and 15 kJ m-2 UV-B. A 304 bp HO-1 promoter region was amplified by PCR from sodium bisulfite-treated DNA, cloned into pGEMT plasmid vector and evaluated by DNA sequencing. Bisulfite sequencing analysis showed similar HO-1 promoter methylation levels in control and UV-B-treated plants (C: 3.4±1.3%; 7.5: 2.6±0.5%; 15: 3.1±1.1%. Interestingly, HO-1 promoter was strongly unmethylated in control plants. Quantitative RT-PCR analysis of TFs showed that GmMYB177, GmMYBJ6, GmWRKY21, GmNAC11, GmNAC20 and GmGT2A but not GmWRK13 and GmDREB were induced by UV-B radiation. The expression of several TFs was also enhanced by hemin, a potent and specific HO inducer, inferring that they may mediate HO-1 up-regulation. These results suggest that soybean HO-1 gene expression is not epigenetically regulated. Moreover, the low level of HO-1 promoter methylation suggests that this antioxidant enzyme can rapidly respond to environmental stress. Finally, this study has identified some stress-related TFs involved in HO-1 up-regulation under UV-B radiation. Keywords: Heme oxygenase, DNA methylation, Transcription factors, Ultraviolet-B radiation, Glycine max

  7. Role of the Nrf2-heme oxygenase-1 pathway in silver nanoparticle-mediated cytotoxicity

    Kang, Su Jin; Ryoo, In-geun; Lee, Young Joon; Kwak, Mi-Kyoung

    2012-01-01

    Silver nanoparticles (nano-Ag) have been widely used in various commercial products including textiles, electronic appliances and biomedical products. However, there remains insufficient information on the potential risk of nano-Ag to human health and environment. In the current study, we have investigated the role of NF-E2-related factor 2 (Nrf2) transcription factor in nano-Ag-induced cytotoxicity. When Nrf2 expression was blocked using interring RNA expression in ovarian carcinoma cell line, nano-Ag treatment showed a substantial decrease in cell viability with concomitant increases in apoptosis and DNA damage compared to the control cells. Target gene analysis revealed that the expression of heme oxygenase-1 (HO-1) was highly elevated by nano-Ag in nonspecific shRNA expressing cells, while Nrf2 knockdown cells (NRF2i) did not increase HO-1 expression. The role of HO-1 in cytoprotection against nano-Ag was reinforced by results using pharmacological inducer of HO-1: cobalt protoporphyrin-mediated HO-1 activation in the NRF2i cells prevented nano-Ag-mediated cell death. Similarly, pharmacological or genetic inhibition of HO-1 in nonspecific control cells exacerbated nano-Ag toxicity. As the upstream signaling mechanism, nano-Ag required the phosphoinositide 3-kinase (PI3K) and p38MAPK signaling cascades for HO-1 induction. The treatment with either PI3K inhibitor or p38MAPK inhibitor suppressed HO-1 induction and intensified nano-Ag-induced cell death. Taken together, these results suggest that Nrf2-dependent HO-1 up-regulation plays a protective role in nano-Ag-induced DNA damage and consequent cell death. In addition, nano-Ag-mediated HO-1 induction is associated with the PI3K and p38MAPK signaling pathways. -- Highlights: ► Role of Nrf2 signaling in silver nanoparticle toxicity. ► Silver nanoparticle toxicity is increased by Nrf2 blockade. ► Nrf2-dependent HO-1 induction protects cells from silver nanoparticle toxicity. ► PI3K and p38MAPK cascades are

  8. Hepatic expression of heme oxygenase-1 and antioxidant response element-mediated genes following administration of ethinyl estradiol to rats

    Morio, Lisa A.; Leone, Angelique; Sawant, Sharmilee P.; Nie, Alex Y.; Brandon Parker, J.; Taggart, Peter; Barron, Alfred M.; McMillian, Michael K.; Lord, Peter

    2006-01-01

    Heme oxygenase-1 (HO-1) is one of several enzymes induced by hepatotoxicants, and is thought to have an important protective role against cellular stress during liver inflammation and injury. The objective of the present study was to evaluate the role of HO-1 in estradiol-induced liver injury. A single dose of ethinyl estradiol (500 mg/kg, po) resulted in mild liver injury. Repeated administration of ethinyl estradiol (500 mg/kg/day for 4 days, po) resulted in no detectable liver injury or dysfunction. Using RT-PCR analysis, we demonstrate that HO-1 gene expression in whole liver tissue is elevated (> 20-fold) after the single dose of ethinyl estradiol. The number and intensity of HO-1 immunoreactive macrophages were increased after the single dose of ethinyl estradiol. HO-1 expression was undetectable in hepatic parenchymal cells from rats receiving Methocel control or a single dose of ethinyl estradiol, however cytosolic HO-1 immunoreactivity in these cells after repeated dosing of ethinyl estradiol was pronounced. The increases in HO-1 mRNA and HO-1 immunoreactivity following administration of a single dose of ethinyl estradiol suggested that this enzyme might be responsible for the observed protection of the liver during repeated dosing. To investigate the effect of HO-1 expression on ethinyl estradiol-induced hepatotoxicity, rats were pretreated with hemin (50 μmol/kg, ip, a substrate and inducer of HO-1), with tin protoporphyrin IX (60 μmol/kg, ip, an HO-1 inhibitor), or with gadolinium chloride (10 mg/kg, iv, an inhibitor/toxin of Kupffer cells) 24 h before ethinyl estradiol treatment. Pretreatment with modulators of HO-1 expression and activity had generally minimal effects on ethinyl estradiol-induced liver injury. These data suggest that HO-1 plays a limited role in antioxidant defense against ethinyl estradiol-induced oxidative stress and hepatotoxicity, and suggests that other coordinately induced enzymes are responsible for protection observed with

  9. Omega-3 polyunsaturated fatty acid has an anti-oxidant effect via the Nrf-2/HO-1 pathway in 3T3-L1 adipocytes

    Kusunoki, Chisato, E-mail: yosizaki@belle.shiga-med.ac.jp [Department of Medicine, Shiga University of Medical Science, Seta Tsukinowa-Cho, Otsu, Shiga 520-2192 (Japan); Yang, Liu; Yoshizaki, Takeshi; Nakagawa, Fumiyuki; Ishikado, Atsushi; Kondo, Motoyuki; Morino, Katsutaro; Sekine, Osamu; Ugi, Satoshi [Department of Medicine, Shiga University of Medical Science, Seta Tsukinowa-Cho, Otsu, Shiga 520-2192 (Japan); Nishio, Yoshihiko [Division of Diabetes, Metabolism and Endocrinology, Department of Graduate School of Medical and Dental Sciences, Kagoshima University, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan); Kashiwagi, Atsunori; Maegawa, Hiroshi [Department of Medicine, Shiga University of Medical Science, Seta Tsukinowa-Cho, Otsu, Shiga 520-2192 (Japan)

    2013-01-04

    Highlights: Black-Right-Pointing-Pointer Omega-3 PUFA has a direct anti-oxidant effect in adipocytes. Black-Right-Pointing-Pointer EPA and DHA induce HO-1 expression in 3T3-L1 adipocytes. Black-Right-Pointing-Pointer Omega-3 PUFA and its end-product, 4-HHE, activates the Nrf-2/HO-1 pathway. Black-Right-Pointing-Pointer Omega-3 PUFA protects against oxidative stress-induced cytotoxicity. -- Abstract: Oxidative stress is produced in adipose tissue of obese subjects and has been associated with obesity-related disorders. Recent studies have shown that omega-3 polyunsaturated fatty acid ({omega}3-PUFA) has beneficial effects in preventing atherosclerotic diseases and insulin resistance in adipose tissue. However, the role of {omega}3-PUFA on adipocytes has not been elucidated. In this study, 3T3-L1 adipocytes were treated with {omega}3-PUFA and its metabolites, eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), or 4-hydroxy hexenal (4-HHE). {omega}3-PUFA and its metabolites dose-dependently increased mRNA and protein levels of the anti-oxidative enzyme, heme oxygenase-1 (HO-1); whereas no changes in the well-known anti-oxidant molecules, superoxide dismutase, catalase, and glutathione peroxidase, were observed. Knockdown of nuclear factor erythroid 2-related factor 2 (Nrf-2) significantly reduced EPA, DHA or 4-HHE-induced HO-1 mRNA and protein expression. Also, pretreatment with {omega}3-PUFA prevented H{sub 2}O{sub 2}-induced cytotoxicity in a HO-1 dependent manner. In conclusion, treatment with EPA and DHA induced HO-1 through the activation of Nrf-2 and prevented oxidative stress in 3T3-L1 adipocytes. This anti-oxidant defense may be of high therapeutic value for clinical conditions associated with systemic oxidative stress.

  10. Omega-3 polyunsaturated fatty acid has an anti-oxidant effect via the Nrf-2/HO-1 pathway in 3T3-L1 adipocytes

    Kusunoki, Chisato; Yang, Liu; Yoshizaki, Takeshi; Nakagawa, Fumiyuki; Ishikado, Atsushi; Kondo, Motoyuki; Morino, Katsutaro; Sekine, Osamu; Ugi, Satoshi; Nishio, Yoshihiko; Kashiwagi, Atsunori; Maegawa, Hiroshi

    2013-01-01

    Highlights: ► Omega-3 PUFA has a direct anti-oxidant effect in adipocytes. ► EPA and DHA induce HO-1 expression in 3T3-L1 adipocytes. ► Omega-3 PUFA and its end-product, 4-HHE, activates the Nrf-2/HO-1 pathway. ► Omega-3 PUFA protects against oxidative stress-induced cytotoxicity. -- Abstract: Oxidative stress is produced in adipose tissue of obese subjects and has been associated with obesity-related disorders. Recent studies have shown that omega-3 polyunsaturated fatty acid (ω3-PUFA) has beneficial effects in preventing atherosclerotic diseases and insulin resistance in adipose tissue. However, the role of ω3-PUFA on adipocytes has not been elucidated. In this study, 3T3-L1 adipocytes were treated with ω3-PUFA and its metabolites, eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), or 4-hydroxy hexenal (4-HHE). ω3-PUFA and its metabolites dose-dependently increased mRNA and protein levels of the anti-oxidative enzyme, heme oxygenase-1 (HO-1); whereas no changes in the well-known anti-oxidant molecules, superoxide dismutase, catalase, and glutathione peroxidase, were observed. Knockdown of nuclear factor erythroid 2-related factor 2 (Nrf-2) significantly reduced EPA, DHA or 4-HHE-induced HO-1 mRNA and protein expression. Also, pretreatment with ω3-PUFA prevented H 2 O 2 -induced cytotoxicity in a HO-1 dependent manner. In conclusion, treatment with EPA and DHA induced HO-1 through the activation of Nrf-2 and prevented oxidative stress in 3T3-L1 adipocytes. This anti-oxidant defense may be of high therapeutic value for clinical conditions associated with systemic oxidative stress.

  11. HO-1-mediated macroautophagy: a mechanism for unregulated iron deposition in aging and degenerating neural tissues.

    Zukor, Hillel; Song, Wei; Liberman, Adrienne; Mui, Jeannie; Vali, Hojatollah; Fillebeen, Carine; Pantopoulos, Kostas; Wu, Ting-Di; Guerquin-Kern, Jean-Luc; Schipper, Hyman M

    2009-05-01

    Oxidative stress, deposition of non-transferrin iron, and mitochondrial insufficiency occur in the brains of patients with Alzheimer disease (AD) and Parkinson disease (PD). We previously demonstrated that heme oxygenase-1 (HO-1) is up-regulated in AD and PD brain and promotes the accumulation of non-transferrin iron in astroglial mitochondria. Herein, dynamic secondary ion mass spectrometry (SIMS) and other techniques were employed to ascertain (i) the impact of HO-1 over-expression on astroglial mitochondrial morphology in vitro, (ii) the topography of aberrant iron sequestration in astrocytes over-expressing HO-1, and (iii) the role of iron regulatory proteins (IRP) in HO-1-mediated iron deposition. Astroglial hHO-1 over-expression induced cytoplasmic vacuolation, mitochondrial membrane damage, and macroautophagy. HO-1 promoted trapping of redox-active iron and sulfur within many cytopathological profiles without impacting ferroportin, transferrin receptor, ferritin, and IRP2 protein levels or IRP1 activity. Thus, HO-1 activity promotes mitochondrial macroautophagy and sequestration of redox-active iron in astroglia independently of classical iron mobilization pathways. Glial HO-1 may be a rational therapeutic target in AD, PD, and other human CNS conditions characterized by the unregulated deposition of brain iron.

  12. Effect of a heme oxygenase-1 inducer on NADPH oxidase ...

    Effect of a heme oxygenase-1 inducer on NADPH oxidase expression in ... and immunohistochemistry of hepatic NOX1 and NOX4 were investigated in week 4. ... (HO-1 inhibitor) administration caused upregulation of NOX gene expression ...

  13. L-Ascorbate attenuates methamphetamine neurotoxicity through enhancing the induction of endogenous heme oxygenase-1

    Huang, Ya-Ni [Department of Nursing, Hsin Sheng College of Medical Care and Management, Taoyuan, Taiwan (China); Wang, Jiz-Yuh [Department of Neurology, Faculty of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan (China); Lee, Ching-Tien [Department of Nursing, Hsin Sheng College of Medical Care and Management, Taoyuan, Taiwan (China); Lin, Chih-Hung [Graduate Institute of Medical Sciences and Department of Physiology, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Lai, Chien-Cheng [Far Eastern Memorial Hospital, Department of Surgery, Taipei, Taiwan (China); Wang, Jia-Yi, E-mail: jywang2010@tmu.edu.tw [Graduate Institute of Medical Sciences and Department of Physiology, College of Medicine, Taipei Medical University, Taipei, Taiwan (China)

    2012-12-01

    Methamphetamine (METH) is a drug of abuse which causes neurotoxicity and increased risk of developing neurodegenerative diseases. We previously found that METH induces heme oxygenase (HO)-1 expression in neurons and glial cells, and this offers partial protection against METH toxicity. In this study, we investigated the effects of L-ascorbate (vitamin C, Vit. C) on METH toxicity and HO-1 expression in neuronal/glial cocultures. Cell viability and damage were evaluated by 3-(4,5-dimethylthianol-2-yl)-2,5 diphenyl tetrazolium bromide (MTT) reduction and lactate dehydrogenase (LDH) release, respectively. Neuronal and glial localization of HO-1 were identified by double immunofluorescence staining. Reactive oxygen species (ROS) production was measured using the fluorochrome 2′,7′-dichlorofluorescin diacetate. HO-1 mRNA and protein expression were examined by RT-qPCR and Western blotting, respectively. Results show that Vit. C induced HO-1 mRNA and protein expressions in time- and concentration-dependent manners. Inhibition of p38 mitogen-activated protein kinase (MAPK) but not extracellular signal-regulated kinase (ERK) significantly blocked induction of HO-1 by Vit. C. HO-1 mRNA and protein expressions were significantly elevated by a combination of Vit. C and METH, compared to either Vit. C or METH alone. Pretreatment with Vit. C enhanced METH-induced HO-1 expression and attenuated METH-induced ROS production and neurotoxicity. Pharmacological inhibition of HO activity abolished suppressive effects of Vit. C on METH-induced ROS production and attenuated neurotoxicity. We conclude that induction of HO-1 expression contributes to the attenuation of METH-induced ROS production and neurotoxicity by Vit. C. We suggest that HO-1 induction by Vit. C may serve as a strategy to alleviate METH neurotoxicity. -- Highlights: ► Besides the anti-oxidant effect, Vit. C also induces HO-1 expression in brain cells. ► Vit. C reduces METH neurotoxicity and ROS production by

  14. L-Ascorbate attenuates methamphetamine neurotoxicity through enhancing the induction of endogenous heme oxygenase-1

    Huang, Ya-Ni; Wang, Jiz-Yuh; Lee, Ching-Tien; Lin, Chih-Hung; Lai, Chien-Cheng; Wang, Jia-Yi

    2012-01-01

    Methamphetamine (METH) is a drug of abuse which causes neurotoxicity and increased risk of developing neurodegenerative diseases. We previously found that METH induces heme oxygenase (HO)-1 expression in neurons and glial cells, and this offers partial protection against METH toxicity. In this study, we investigated the effects of L-ascorbate (vitamin C, Vit. C) on METH toxicity and HO-1 expression in neuronal/glial cocultures. Cell viability and damage were evaluated by 3-(4,5-dimethylthianol-2-yl)-2,5 diphenyl tetrazolium bromide (MTT) reduction and lactate dehydrogenase (LDH) release, respectively. Neuronal and glial localization of HO-1 were identified by double immunofluorescence staining. Reactive oxygen species (ROS) production was measured using the fluorochrome 2′,7′-dichlorofluorescin diacetate. HO-1 mRNA and protein expression were examined by RT-qPCR and Western blotting, respectively. Results show that Vit. C induced HO-1 mRNA and protein expressions in time- and concentration-dependent manners. Inhibition of p38 mitogen-activated protein kinase (MAPK) but not extracellular signal-regulated kinase (ERK) significantly blocked induction of HO-1 by Vit. C. HO-1 mRNA and protein expressions were significantly elevated by a combination of Vit. C and METH, compared to either Vit. C or METH alone. Pretreatment with Vit. C enhanced METH-induced HO-1 expression and attenuated METH-induced ROS production and neurotoxicity. Pharmacological inhibition of HO activity abolished suppressive effects of Vit. C on METH-induced ROS production and attenuated neurotoxicity. We conclude that induction of HO-1 expression contributes to the attenuation of METH-induced ROS production and neurotoxicity by Vit. C. We suggest that HO-1 induction by Vit. C may serve as a strategy to alleviate METH neurotoxicity. -- Highlights: ► Besides the anti-oxidant effect, Vit. C also induces HO-1 expression in brain cells. ► Vit. C reduces METH neurotoxicity and ROS production by

  15. Orthodontic Forces Induce the Cytoprotective Enzyme Heme Oxygenase-1 in Rats

    Suttorp, Christiaan M.; Xie, Rui; Lundvig, Ditte M. S.; Kuijpers-Jagtman, Anne Marie; Uijttenboogaart, Jasper Tom; Van Rheden, René; Maltha, Jaap C.; Wagener, Frank A. D. T. G.

    2016-01-01

    Orthodontic forces disturb the microenvironment of the periodontal ligament (PDL), and induce craniofacial bone remodeling which is necessary for tooth movement. Unfortunately, orthodontic tooth movement is often hampered by ischemic injury and cell death within the PDL (hyalinization) and root resorption. Large inter-individual differences in hyalinization and root resorption have been observed, and may be explained by differential protection against hyalinization. Heme oxygenase-1 (HO-1) forms an important protective mechanism by breaking down heme into the strong anti-oxidants biliverdin/bilirubin and the signaling molecule carbon monoxide. These versatile HO-1 products protect against ischemic and inflammatory injury. We postulate that orthodontic forces induce HO-1 expression in the PDL during experimental tooth movement. Twenty-five 6-week-old male Wistar rats were used in this study. The upper three molars at one side were moved mesially using a Nickel-Titanium coil spring, providing a continuous orthodontic force of 10 cN. The contralateral side served as control. After 6, 12, 72, 96, and 120 h groups of rats were killed. On parasagittal sections immunohistochemical staining was performed for analysis of HO-1 expression and quantification of osteoclasts. Orthodontic force induced a significant time-dependent HO-1 expression in mononuclear cells within the PDL at both the apposition- and resorption side. Shortly after placement of the orthodontic appliance HO-1 expression was highly induced in PDL cells but dropped to control levels within 72 h. Some osteoclasts were also HO-1 positive but this induction was shown to be independent of time- and mechanical stress. It is tempting to speculate that differential induction of tissue protecting- and osteoclast activating genes in the PDL determine the level of bone resorption and hyalinization and, subsequently, “fast” and “slow” tooth movers during orthodontic treatment. PMID:27486402

  16. Orthodontic Forces Induce the Cytoprotective Enzyme Heme Oxygenase-1 in Rats.

    Suttorp, Christiaan M; Xie, Rui; Lundvig, Ditte M S; Kuijpers-Jagtman, Anne Marie; Uijttenboogaart, Jasper Tom; Van Rheden, René; Maltha, Jaap C; Wagener, Frank A D T G

    2016-01-01

    Orthodontic forces disturb the microenvironment of the periodontal ligament (PDL), and induce craniofacial bone remodeling which is necessary for tooth movement. Unfortunately, orthodontic tooth movement is often hampered by ischemic injury and cell death within the PDL (hyalinization) and root resorption. Large inter-individual differences in hyalinization and root resorption have been observed, and may be explained by differential protection against hyalinization. Heme oxygenase-1 (HO-1) forms an important protective mechanism by breaking down heme into the strong anti-oxidants biliverdin/bilirubin and the signaling molecule carbon monoxide. These versatile HO-1 products protect against ischemic and inflammatory injury. We postulate that orthodontic forces induce HO-1 expression in the PDL during experimental tooth movement. Twenty-five 6-week-old male Wistar rats were used in this study. The upper three molars at one side were moved mesially using a Nickel-Titanium coil spring, providing a continuous orthodontic force of 10 cN. The contralateral side served as control. After 6, 12, 72, 96, and 120 h groups of rats were killed. On parasagittal sections immunohistochemical staining was performed for analysis of HO-1 expression and quantification of osteoclasts. Orthodontic force induced a significant time-dependent HO-1 expression in mononuclear cells within the PDL at both the apposition- and resorption side. Shortly after placement of the orthodontic appliance HO-1 expression was highly induced in PDL cells but dropped to control levels within 72 h. Some osteoclasts were also HO-1 positive but this induction was shown to be independent of time- and mechanical stress. It is tempting to speculate that differential induction of tissue protecting- and osteoclast activating genes in the PDL determine the level of bone resorption and hyalinization and, subsequently, "fast" and "slow" tooth movers during orthodontic treatment.

  17. Electroacupuncture Ameliorates Acute Renal Injury in Lipopolysaccharide-Stimulated Rabbits via Induction of HO-1 through the PI3K/Akt/Nrf2 Pathways.

    Yu, Jian-Bo; Shi, Jia; Zhang, Yuan; Gong, Li-Rong; Dong, Shu-An; Cao, Xin-Shun; Wu, Li-Li; Wu, Li-Na

    2015-01-01

    Electroacupuncture at select acupoints have been verified to protect against organ dysfunctions during endotoxic shock. And, heme oxygenase (HO)-1 as a phase II enzyme and antioxidant contributed to the protection of kidney in septic shock rats. The phosphatidylinositol 3-kinase (PI3K)-Akt pathway mediated the activation of NF-E2 related factor-2 (Nrf2), which was involved in HO-1 induction. To understand the efficacy of electroacupuncture stimulation in ameliorating acute kidney injury (AKI) through the PI3K/Akt/Nrf2 pathway and subsequent HO-1 upregulation, a dose of LPS 5mg/kg was administered intravenously to replicate the rabbit model of AKI induced by endotoxic shock. Electroacupuncture pretreatment was handled bilaterally at Zusanli and Neiguan acupoints for five consecutive days while sham electroacupuncture at non-acupoints as control. Results displayed that electroacupuncture stimulation significantly alleviated the morphologic renal damage, attenuated renal tubular apoptosis, suppressed the elevated biochemical indicators of AKI caused by LPS, enhanced the expressions of phospho-Akt, HO-1protein, Nrf2 total and nucleoprotein, and highlighted the proportions of Nrf2 nucleoprotein as a parallel. Furthermore, partial protective effects of elecroacupuncture were counteracted by preconditioning with wortmannin (the selective PI3K inhibitor), indicating a direct involvement of PI3K/Akt pathway. Inconsistently, wortmannin pretreatment made little difference to the expressions of HO-1, Nrf2 nucleoprotein and total protein, which indicated that PI3K/Akt may be not the only pathway responsible for electroacupuncture-afforded protection against LPS-induced AKI. These findings provide new insights into the potential future clinical applications of electroacupuncture for AKI induced by endotoxic shock instead of traditional remedies.

  18. Soybean-Derived Phytoalexins Improve Cognitive Function through Activation of Nrf2/HO-1 Signaling Pathway

    Ji Yeon Seo

    2018-01-01

    Full Text Available As soy-derived glyceollins are known to induce antioxidant enzymes in various types of cells and tissues, we hypothesized that the compounds could protect neurons from damage due to reactive oxygen species (ROS. In order to examine the neuroprotective effect of glyceollins, primary cortical neurons collected from mice and mouse hippocampal HT22 cells were challenged with glutamate. Glyceollins attenuated glutamate-induced cytotoxicity in primary cortical neuron isolated from mice carrying wild-type nuclear factor (erythroid-derived 2-like 2 (Nrf2, but the compounds were ineffective in those isolated from Nrf2 knockout mice, suggesting the involvement of the Nrf2 signaling pathway in glyceollin-mediated neuroprotection. Furthermore, the inhibition of heme oxygenase-1 (HO-1, a major downstream enzyme of Nrf2, abolished the suppressive effect of glyceollins against glutamate-induced ROS production and cytotoxicity, confirming that activation of HO-1 by glyceollins is responsible for the neuroprotection. To examine whether glyceollins also improve cognitive ability, mice pretreated with glyceollins were challenged with scopolamine and subjected to behavioral tests. Glyceollins attenuated scopolamine-induced cognitive impairment of mice, but failed to enhance memory in Nrf2 knockout mice, suggesting that the memory-enhancing effect is also mediated by the Nrf2 signaling pathway. Overall, glyceollins showed neuroprotection against glutamate-induced damage, and attenuated scopolamine-induced memory deficits in an Nrf2-dependent manner.

  19. Nrf2-dependent induction of innate host defense via heme oxygenase-1 inhibits Zika virus replication

    Huang, Hanxia; Falgout, Barry; Takeda, Kazuyo [Food and Drug Administration, Silver Spring, MD (United States); Yamada, Kenneth M. [National Institutes of Health, Bethesda, MD (United States); Dhawan, Subhash, E-mail: subhash.dhawan@fda.hhs.gov [Food and Drug Administration, Silver Spring, MD (United States)

    2017-03-15

    We identified primary human monocyte-derived macrophages (MDM) as vulnerable target cells for Zika virus (ZIKV) infection. We demonstrate dramatic effects of hemin, the natural inducer of the heme catabolic enzyme heme oxygenase-1 (HO-1), in the reduction of ZIKV replication in vitro. Both LLC-MK2 monkey kidney cells and primary MDM exhibited hemin-induced HO-1 expression with major reductions of >90% in ZIKV replication, with little toxicity to infected cells. Silencing expression of HO-1 or its upstream regulatory gene, nuclear factor erythroid-related factor 2 (Nrf2), attenuated hemin-induced suppression of ZIKV infection, suggesting an important role for induction of these intracellular mediators in retarding ZIKV replication. The inverse correlation between hemin-induced HO-1 levels and ZIKV replication provides a potentially useful therapeutic modality based on stimulation of an innate cellular response against Zika virus infection. - Highlights: •Hemin treatment protected monocyte-derived macrophages against Zika virus (ZIKV) infection. •Innate cellular protection against ZIKV infection correlated with Nrf2-dependent HO-1 expression. •Stimulation of innate cellular responses may provide a therapeutic strategy against ZIKV infection.

  20. Nrf2-dependent induction of innate host defense via heme oxygenase-1 inhibits Zika virus replication

    Huang, Hanxia; Falgout, Barry; Takeda, Kazuyo; Yamada, Kenneth M.; Dhawan, Subhash

    2017-01-01

    We identified primary human monocyte-derived macrophages (MDM) as vulnerable target cells for Zika virus (ZIKV) infection. We demonstrate dramatic effects of hemin, the natural inducer of the heme catabolic enzyme heme oxygenase-1 (HO-1), in the reduction of ZIKV replication in vitro. Both LLC-MK2 monkey kidney cells and primary MDM exhibited hemin-induced HO-1 expression with major reductions of >90% in ZIKV replication, with little toxicity to infected cells. Silencing expression of HO-1 or its upstream regulatory gene, nuclear factor erythroid-related factor 2 (Nrf2), attenuated hemin-induced suppression of ZIKV infection, suggesting an important role for induction of these intracellular mediators in retarding ZIKV replication. The inverse correlation between hemin-induced HO-1 levels and ZIKV replication provides a potentially useful therapeutic modality based on stimulation of an innate cellular response against Zika virus infection. - Highlights: •Hemin treatment protected monocyte-derived macrophages against Zika virus (ZIKV) infection. •Innate cellular protection against ZIKV infection correlated with Nrf2-dependent HO-1 expression. •Stimulation of innate cellular responses may provide a therapeutic strategy against ZIKV infection.

  1. Orthodontic forces induce the cytoprotective enzyme heme oxygenase-1 in rats

    Christiaan M. Suttorp

    2016-07-01

    Full Text Available Orthodontic forces disturb the microenvironment of the periodontal ligament (PDL, and induce craniofacial bone remodeling which is necessary for tooth movement. Unfortunately, orthodontic tooth movement is often hampered by ischemic injury and cell death within the PDL (hyalinization and root resorption. Large inter-individual differences in hyalinization and root resorption have been observed, and may be explained by differential protection against hyalization. Heme oxygenase-1 (HO-1 forms an important protective mechanism by breaking down heme into the strong anti-oxidants biliverdin/bilirubin and the signaling molecule carbon monoxide. These versatile HO-products protect against ischemic and inflammatory injury. We postulate that orthodontic forces induce HO-1 expression in the PDL during experimental tooth movement. Twenty-five 6-week-old male Wistar rats were used in this study. The upper three molars at one side were moved mesially using a Ni-Ti 10 cN coil spring. The contralateral side served as control. After 6, 12, 72, 96 and 120 hrs rats were killed. On parasagittal sections immunohistochemical staining was performed for analysis of HO-1 expression and quantification of multinuclear osteoclasts. Orthodontic force induced a significant time-dependent HO-1 expression in the mononuclear cell population within the PDL at both the apposition- and resorption side. Shortly after appliance placement HO-1 expression was highly induced in PDL cells but dropped to control levels within 72 hours. Some osteoclasts were HO-1 positive but this induction was shown to be independent of time- and mechanical stress. It is tempting to speculate that differential induction of cytoprotective enzymes as HO-1 in the PDL determines the level of hyalinization and, subsequently, fast and slow tooth movers during orthodontic treatment.

  2. Iron depletion in HCT116 cells diminishes the upregulatory effect of phenethyl isothiocyanate on heme oxygenase-1

    Bolloskis, Michael P.; Carvalho, Fabiana P.; Loo, George

    2016-01-01

    Some of the health-promoting properties of cruciferous vegetables are thought to be partly attributed to isothiocyanates. These phytochemicals can upregulate the expression of certain cytoprotective stress genes, but it is unknown if a particular nutrient is involved. Herein, the objective was to ascertain if adequate iron is needed for enabling HCT116 cells to optimally express heme oxygenase-1 (HO-1) when induced by phenethyl isothiocyanate (PEITC). PEITC increased HO-1 expression and also nuclear translocation of Nrf2, which is a transcription factor known to activate the HO-1 gene. However, in HCT116 cells that were made iron-deficient by depleting intracellular iron with deferoxamine (DFO), PEITC was less able to increase HO-1 expression and nuclear translocation of Nrf2. These suppressive effects of DFO were overcome by replenishing the iron-deficient cells with the missing iron. To elucidate these findings, it was found that PEITC-induced HO-1 upregulation can be inhibited with thiol antioxidants (glutathione and N-acetylcysteine). Furthermore, NADPH oxidase inhibitors (diphenyleneiodonium and apocynin) and a superoxide scavenger (Tiron) each inhibited PEITC-induced HO-1 upregulation. In doing so, diphenyleneiodonium was the most potent and also inhibited nuclear translocation of redox-sensitive Nrf2. Collectively, the results imply that the HO-1 upregulation by PEITC involves an iron-dependent, oxidant signaling pathway. Therefore, it is concluded that ample iron is required to enable PEITC to fully upregulate HO-1 expression in HCT116 cells. As such, it is conceivable that iron-deficient individuals may not reap the full health benefits of eating PEITC-containing cruciferous vegetables that via HO-1 may help protect against multiple chronic diseases. - Highlights: • PEITC increased HO-1 expression in HCT116 cells. • PEITC-induced HO-1 upregulation was impaired in iron-depleted HCT116 cells. • Impairment of PEITC-induced HO-1 upregulation was

  3. Pharmacological Inhibition of Host Heme Oxygenase-1 Suppresses Mycobacterium tuberculosis Infection In Vivo by a Mechanism Dependent on T Lymphocytes

    Diego L. Costa

    2016-10-01

    Full Text Available Heme oxygenase-1 (HO-1 is a stress response antioxidant enzyme which catalyzes the degradation of heme released during inflammation. HO-1 expression is upregulated in both experimental and human Mycobacterium tuberculosis infection, and in patients it is a biomarker of active disease. Whether the enzyme plays a protective versus pathogenic role in tuberculosis has been the subject of debate. To address this controversy, we administered tin protoporphyrin IX (SnPPIX, a well-characterized HO-1 enzymatic inhibitor, to mice during acute M. tuberculosis infection. These SnPPIX-treated animals displayed a substantial reduction in pulmonary bacterial loads comparable to that achieved following conventional antibiotic therapy. Moreover, when administered adjunctively with antimycobacterial drugs, the HO-1 inhibitor markedly enhanced and accelerated pathogen clearance. Interestingly, both the pulmonary induction of HO-1 expression and the efficacy of SnPPIX treatment in reducing bacterial burden were dependent on the presence of host T lymphocytes. Although M. tuberculosis expresses its own heme-degrading enzyme, SnPPIX failed to inhibit its enzymatic activity or significantly restrict bacterial growth in liquid culture. Together, the above findings reveal mammalian HO-1 as a potential target for host-directed monotherapy and adjunctive therapy of tuberculosis and identify the immune response as a critical regulator of this function.

  4. Heme oxygenase and the immune system in normal and pathological pregnancies

    Maide eOzen

    2015-04-01

    Full Text Available Normal pregnancy is an immunotolerant state. Many factors, including environmental, socioeconomic, genetic, and immunologic changes by infection and/or other causes of inflammation, may contribute to inter-individual differences resulting in a normal or pathologic pregnancy. In particular, imbalances in the immune system can cause many pregnancy-related diseases, such as infertility, abortions, pre-eclampsia, and preterm labor, which result in maternal/fetal death, prematurity, or small-for-gestational age newborns. New findings imply that myeloid regulatory cells and regulatory T cells (Tregs may mediate immunotolerance during normal pregnancy. Effector T cells (Teffs have, in contrast, been implicated to cause adverse pregnancy outcomes. Furthermore, feto-maternal tolerance affects the developing fetus. It has been shown that the Treg/Teff balance affects litter size and adoptive transfer of pregnancy-induced Tregs can prevent fetal rejection in the mouse. Heme oxygenase-1 (HO-1 has a protective role in many conditions through its anti-inflammatory, anti-apoptotic, antioxidative, and anti-proliferative actions. HO-1 is highly expressed in the placenta and plays a role in angiogenesis and placental vascular development and in regulating vascular tone in pregnancy. In addition, HO-1 is a major regulator of immune homeostasis by mediating crosstalk between innate and adaptive immune systems. Moreover, HO-1 can inhibit inflammation-induced phenotypic maturation of immune effector cells and pro-inflammatory cytokine secretion and promote anti-inflammatory cytokine production. HO-1 may also be associated with T-cell activation and can limit immune-based tissue injury by promoting Treg suppression of effector responses. Thus, HO-1 and its byproducts may protect against pregnancy complications by its immunomodulatory effects, and the regulation of HO-1 or its downstream effects has the potential to prevent or treat pregnancy complications and

  5. Hemin inhibits NO production by IL-1β-stimulated human astrocytes through induction of heme oxygenase-1 and reduction of p38 MAPK activation

    Sheng Wen S

    2010-09-01

    Full Text Available Abstract Background Heme oxygenase (HO-1 has been shown to attenuate oxidative injury and reduce apoptosis. HO-1 can be induced by various stimuli released during cellular injury, such as heme. Deleterious free heme is degraded by HO-1 to carbon monoxide, iron and biliverdin, which have potent anti-oxidant and anti-inflammatory properties. In this study, we tested the hypothesis that upregulation of HO-1 would inhibit production of the free radical (NO by interlukin (IL-1β-activated human astrocytes. Methods To measure NO production, inducible NO synthase (iNOS, HO-1 expression and mitogen-activated protein (MAP kinase activation we used hemin as an HO-1 inducer and tin protoporphyrin (SnPP IX as an inhibitor of HO-1 activity in human astrocyte cultures prior to IL-1β exposure. Transfection of astrocyte cultures was performed using a pLEX expression vector carrying the human HO-1 sequence prior to IL-1β treatment. Supernatants of astrocyte cultures pretreated with inhibitors of p38 MAPK or MEK1/2 prior to IL-1β exposure were collected for NO assay. Results IL-1β treatment of astrocytes alone induced undetectable amounts of HO-1 protein by western blot. However, HO-1 mRNA expression was modestly up-regulated in response to IL-1β stimulation. Pretreatment with hemin alone substantially induced both HO-1 mRNA and protein expression, and HO-1 mRNA expression was further enhanced when hemin was combined with IL-1β treatment. In contrast, IL-1β-induced iNOS mRNA expression and NO production were markedly inhibited by hemin treatment. When pretreated with SnPP, the inhibitory effect of hemin on IL-1β-induced NO production and iNOS expression was reversed, suggesting the involvement of HO-1. IL-1β-induced p38 MAPK activation, which is known to be required for NO production, was also down-regulated by hemin. Conclusion These findings support the hypothesis that up-regulation of HO-1 in astrocytes is associated with down-regulation of i

  6. Cadmium-induced heme-oxygenase-1 expression plays dual roles in autophagy and apoptosis and is regulated by both PKC-δ and PKB/Akt activation in NRK52E kidney cells

    So, Keum-Young; Oh, Seon-Hee

    2016-01-01

    Heme oxygenase-1 (HO-1) protects cells against cadmium (Cd)-induced oxidative stress. However, the mechanism underlying this protection is not well understood. In this study, we elucidated the role of HO-1 in Cd-induced cytotoxicity. Exposure of NRK52E cells to Cd induced protein kinase B (PKB)/Akt, protein kinase C (PKC)-δ, and glycogen synthase kinase (GSK) 3αb phosphorylation, and eukaryotic initiation factor (eIF) 2α dephosphorylation. Pharmacological inhibition of Akt resulted in HO-1 suppression and eIF2α activation, which partially suppressed CHOP and PARP-1 cleavage, but promoted autophagy and decreased cell viability. Pharmacological inactivation of PKC-δ markedly suppressed Cd-induced phospho-serine (p-Ser) GSK3αβ, and HO-1, and partially inhibited PARP-1 cleavage, but massively induced autophagy and decreased cell viability. Pharmacological upregulation of p-Ser GSK3αβ enhanced Cd-induced HO-1, CHOP, and PARP-1 cleavage, but decreased autophagy. Genetic deficiency of GSK3β suppressed HO-1 and PARP-1 cleavage and increased autophagy. Genetic suppression of HO-1 reduced Cd-induced PARP-1 cleavage, but increased LC3-II. Cd exposure led to accumulation of p-PKC-δ, p-Ser GSK3αβ, and HO-1 in the nucleus and particulate fractions, suggesting that they have dual functions in response to Cd. N-acetylcysteine treatment suppressed Cd-induced activation of PKC-δ and Akt. These results indicate that HO-1 induced by Cd exposure is regulated by PKC-δ, p-Ser GSK3αβ, and PKB/Akt, which restrain autophagic cell death, but mildly induce apoptosis in NRK52E cells. Together, the results suggest that HO-1 expression in response to Cd maintains cellular homeostasis during oxidative stress.

  7. Casein Glycomacropeptide Hydrolysates Exert Cytoprotective Effect against Cellular Oxidative Stress by Up-Regulating HO-1 Expression in HepG2 Cells

    Tiange Li

    2017-01-01

    Full Text Available Oxidative stress is considered as an important mediator in the progression of metabolic disorders. The objective of this study was to investigate the potential hepatoprotective effects and mechanisms of bovine casein glycomacropeptide hydrolysates (GHP on hydrogen peroxide (H2O2-induced oxidative damage in HepG2 cells. Results showed that GHP significantly blocked H2O2-induced intracellular reactive oxygen species (ROS generation and cell viability reduction in a dose-dependent manner. Further, GHP concentration-dependently induced heme oxygenase-1 (HO-1 expression and increased nuclear factor-erythroid 2-related factor 2 (Nrf2 nuclear translocation. Moreover, pretreatment of GHP increased the activation of p38 mitogen-activated protein kinase (p38 MAPK and extracellular signal-regulated protein kinase 1/2 (ERK1/2, which were shown to contribute to Nrf2-mediated HO-1 expression. Taken together, GHP protected HepG2 cells from oxidative stress by activation of Nrf2 and HO-1 via p38 MAPK and ERK1/2 signaling pathways. Our findings indicate that bovine casein glycomacropeptide hydrolysates might be a potential ingredient in the treatment of oxidative stress-related disorders and further studies are needed to investigate the protective effects in vivo.

  8. Glomerular Epithelial Cells-Targeted Heme Oxygenase-1 Over Expression in the Rat: Attenuation of Proteinuria in Secondary But Not Primary Injury.

    Atsaves, Vassilios; Makri, Panagiota; Detsika, Maria G; Tsirogianni, Alexandra; Lianos, Elias A

    2016-01-01

    Induction of heme oxygenase 1 (HO-1) in glomerular epithelial cells (GEC) in response to injury is poor and this may be a disadvantage. We, therefore, explored whether HO-1 overexpression in GEC can reduce proteinuria induced by puromycin aminonucleoside (PAN) or in anti-glomerular basement membrane (GBM) antibody (Ab)-mediated glomerulonephritis (GN). HO-1 overexpression in GEC (GECHO-1) of Sprague-Dawley rats was achieved by targeting a FLAG-human (h) HO-1 using transposon-mediated transgenesis. Direct GEC injury was induced by a single injection of PAN. GN was induced by administration of an anti-rat GBM Ab and macrophage infiltration in glomeruli was assessed by immunohistochemistry and western blot analysis, which was also used to assess glomerular nephrin expression. In GECHO-1 rats, FLAG-hHO-1 transprotein was co-immunolocalized with nephrin. Baseline glomerular HO-1 protein levels were higher in GECHO-1 compared to wild type (WT) rats. Administration of either PAN or anti-GBM Ab to WT rats increased glomerular HO-1 levels. Nephrin expression markedly decreased in glomeruli of WT or GECHO-1 rats treated with PAN. In anti-GBM Ab-treated WT rats, nephrin expression also decreased. In contrast, it was preserved in anti-GBM Ab-treated GECHO-1 rats. In these, macrophage infiltration in glomeruli and the ratio of urine albumin to urine creatinine (Ualb/Ucreat) were markedly reduced. There was no difference in Ualb/Ucreat between WT and GECHO-1 rats treated with PAN. Depending on the type of injury, HO-1 overexpression in GEC may or may not reduce proteinuria. Reduced macrophage infiltration and preservation of nephrin expression are putative mechanisms underlying the protective effect of HO-1 overexpression following immune injury. © 2016 S. Karger AG, Basel.

  9. The Cytoprotective Effects of E-α-(4-Methoxyphenyl-2',3,4,4'-Tetramethoxychalcone (E-α-p-OMe-C6H4-TMC--A Novel and Non-Cytotoxic HO-1 Inducer.

    Kai B Kaufmann

    Full Text Available Cell protection against different noxious stimuli like oxidative stress or chemical toxins plays a central role in the treatment of many diseases. The inducible heme oxygenase isoform, heme oxygenase-1 (HO-1, is known to protect cells against a variety of harmful conditions including apoptosis. Because a number of medium strong electrophiles from a series of α-X-substituted 2',3,4,4'-tetramethoxychalcones (α-X-TMCs, X = H, F, Cl, Br, I, CN, Me, p-NO2-C6H4, Ph, p-OMe-C6H4, NO2, CF3, COOEt, COOH had proven to activate Nrf2 resulting in HO-1 induction and inhibit NF-κB downstream target genes, their protective effect against staurosporine induced apoptosis and reactive oxygen species (ROS production was investigated. RAW264.7 macrophages treated with 19 different chalcones (15 α-X-TMCs, chalcone, 2'-hydroxychalcone, calythropsin and 2'-hydroxy-3,4,4'-trimethoxychalcone prior to staurosporine treatment were analyzed for apoptosis and ROS production, as well as HO-1 protein expression and enzyme activity. Additionally, Nrf2 and NF-κB activity was assessed. We found that amongst all tested chalcones only E-α-(4-methoxyphenyl-2',3,4,4'-tetramethoxychalcone (E-α-p-OMe-C6H4-TMC demonstrated a distinct, statistically significant antiapoptotic effect in a dose dependent manner, showing no toxic effects, while its double bond isomer Z-α-p-OMe-C6H4-TMC displayed no significant activity. Also, E-α-p-OMe-C6H4-TMC induced HO-1 protein expression and increased HO-1 activity, whilst inhibition of HO-1 by SnPP-IX abolished its antiapoptotic effect. The only weakly electrophilic chalcone E-α-p-OMe-C6H4-TMC reduced the staurosporine triggered formation of ROS, while inducing the translocation of Nrf2 into the nucleus. Furthermore, staurosporine induced NF-κB activity was attenuated following E-α-p-OMe-C6H4-TMC treatment. Overall, E-α-p-OMe-C6H4-TMC demonstrated its effective cytoprotective potential via a non-toxic induction of HO-1 in RAW264

  10. GEC-targeted HO-1 expression reduces proteinuria in glomerular immune injury.

    Duann, Pu; Lianos, Elias A

    2009-09-01

    Induction of heme oxygenase (HO)-1 is a key defense mechanism against oxidative stress. Compared with tubules, glomeruli are refractory to HO-1 upregulation in response to injury. This can be a disadvantage as it may be associated with insufficient production of cytoprotective heme-degradation metabolites. We, therefore, explored whether 1) targeted HO-1 expression can be achieved in glomeruli without altering their physiological integrity and 2) this expression reduces proteinuria in immune injury induced by an anti-glomerular basement membrane (GBM) antibody (Ab). We employed a 4.125-kb fragment of a mouse nephrin promoter downstream to which a FLAG-tagged hHO-1 cDNA sequence was inserted and subsequently generated transgenic mice from the FVB/N parental strain. There was a 16-fold higher transgene expression in the kidney than nonspecific background (liver) while the transprotein immunolocalized in glomerular epithelial cells (GEC). There was no change in urinary protein excretion, indicating that GEC-targeted HO-1 expression had no effect on glomerular protein permeability. Urinary protein excretion in transgenic mice with anti-GBM Ab injury (days 3 and 6) was significantly lower compared with wild-type controls. There was no significant change in renal expression levels of profibrotic (TGF-beta1) or anti-inflammatory (IL-10) cytokines in transgenic mice with anti-GBM Ab injury. These observations indicate that GEC-targeted HO-1 expression does not alter glomerular physiological integrity and reduces proteinuria in glomerular immune injury.

  11. Postneonatal Mortality and Liver Changes in Cloned Pigs Associated with Human Tumor Necrosis Factor Receptor I-Fc and Human Heme Oxygenase-1 Overexpression.

    Kim, Geon A; Jin, Jun-Xue; Lee, Sanghoon; Taweechaipaisankul, Anukul; Oh, Hyun Ju; Hwang, Joing-Ik; Ahn, Curie; Saadeldin, Islam M; Lee, Byeong Chun

    2017-01-01

    Soluble human tumor necrosis factor (shTNFRI-Fc) and human heme oxygenase 1 (hHO-1) are key regulators for protection against oxidative and inflammatory injury for xenotransplantation. Somatic cells with more than 10 copy numbers of shTNFRI-Fc and hHO-1 were employed in somatic cell nuclear transfer to generate cloned pigs, thereby resulting in seven cloned piglets. However, produced piglets were all dead within 24 hours after birth. Obviously, postnatal death with liver apoptosis was reported in the higher copy number of shTNFRI-Fc and hHO-1 piglets. In liver, the transcript levels of ferritin heavy chain, light chain, transferrin, and inducible nitric oxide synthase were significantly highly expressed compared to those of lower copy number of shTNFRI-Fc and hHO-1 piglets ( P hHO-1 piglets ( P hHO-1 overexpression may apparently induce free iron in the liver and exert oxidative stress by enhancing reactive oxygen species production and block normal postneonatal liver metabolism.

  12. Postneonatal Mortality and Liver Changes in Cloned Pigs Associated with Human Tumor Necrosis Factor Receptor I-Fc and Human Heme Oxygenase-1 Overexpression

    Geon A. Kim

    2017-01-01

    Full Text Available Soluble human tumor necrosis factor (shTNFRI-Fc and human heme oxygenase 1 (hHO-1 are key regulators for protection against oxidative and inflammatory injury for xenotransplantation. Somatic cells with more than 10 copy numbers of shTNFRI-Fc and hHO-1 were employed in somatic cell nuclear transfer to generate cloned pigs, thereby resulting in seven cloned piglets. However, produced piglets were all dead within 24 hours after birth. Obviously, postnatal death with liver apoptosis was reported in the higher copy number of shTNFRI-Fc and hHO-1 piglets. In liver, the transcript levels of ferritin heavy chain, light chain, transferrin, and inducible nitric oxide synthase were significantly highly expressed compared to those of lower copy number of shTNFRI-Fc and hHO-1 piglets (P<0.05. Also, H2O2 contents were increased, and superoxide dismutase was significantly lower in the higher copy number of shTNFRI-Fc and hHO-1 piglets (P<0.05. These results indicate that TNFRI-Fc and hHO-1 overexpression may apparently induce free iron in the liver and exert oxidative stress by enhancing reactive oxygen species production and block normal postneonatal liver metabolism.

  13. Beneficial effect of prolonged heme oxygenase 1 activation in a rat model of chronic heart failure

    Massimo Collino

    2013-07-01

    We and others have previously demonstrated that heme oxygenase 1 (HO-1 induction by acute hemin administration exerts cardioprotective effects. Here, we developed a rat model of heart failure to investigate whether a long-term induction of HO-1 by chronic hemin administration exerted protective effects. Sprague Dawley rats that underwent permanent ligation of the left coronary artery were closely monitored for survival rate analysis and sacrificed on day 28 post-operation. Administration of hemin (4 mg/kg body weight every other day for 4 weeks induced a massive increase in HO-1 expression and activity, as shown by the increased levels of the two main metabolic products of heme degradation, bilirubin and carbon monoxide (CO. These effects were associated with significant improvement in survival and reduced the extension of myocardial damage. The ischemic hearts of the hemin-treated animals displayed reduced oxidative stress and apoptosis in comparison with the non-treated rats, as shown by the decreased levels of lipid peroxidation, free-radical-induced DNA damage, caspase-3 activity and Bax expression. Besides, chronic HO-1 activation suppressed the elevated levels of myeloperoxidase (MPO activity, interleukin 1β (IL-1β production and tumor necrosis factor-α (TNFα production that were evoked by the ischemic injury, and increased the plasma level of the anti-inflammatory cytokine IL-10. Interestingly, HO-1 inhibitor zinc protoporphyrin IX (ZnPP-IX; 1 mg/kg lowered bilirubin and CO concentrations to control values, thus abolishing all the cardioprotective effects of hemin. In conclusion, the results demonstrate that chronic HO-1 activation by prolonged administration of hemin improves survival and exerts protective effects in a rat model of myocardial ischemia by exerting a potent antioxidant activity and disrupting multiple levels of the apoptotic and inflammatory cascade.

  14. Beneficial effect of prolonged heme oxygenase 1 activation in a rat model of chronic heart failure.

    Collino, Massimo; Pini, Alessandro; Mugelli, Niccolò; Mastroianni, Rosanna; Bani, Daniele; Fantozzi, Roberto; Papucci, Laura; Fazi, Marilena; Masini, Emanuela

    2013-07-01

    We and others have previously demonstrated that heme oxygenase 1 (HO-1) induction by acute hemin administration exerts cardioprotective effects. Here, we developed a rat model of heart failure to investigate whether a long-term induction of HO-1 by chronic hemin administration exerted protective effects. Sprague Dawley rats that underwent permanent ligation of the left coronary artery were closely monitored for survival rate analysis and sacrificed on day 28 post-operation. Administration of hemin (4 mg/kg body weight) every other day for 4 weeks induced a massive increase in HO-1 expression and activity, as shown by the increased levels of the two main metabolic products of heme degradation, bilirubin and carbon monoxide (CO). These effects were associated with significant improvement in survival and reduced the extension of myocardial damage. The ischemic hearts of the hemin-treated animals displayed reduced oxidative stress and apoptosis in comparison with the non-treated rats, as shown by the decreased levels of lipid peroxidation, free-radical-induced DNA damage, caspase-3 activity and Bax expression. Besides, chronic HO-1 activation suppressed the elevated levels of myeloperoxidase (MPO) activity, interleukin 1β (IL-1β) production and tumor necrosis factor-α (TNFα) production that were evoked by the ischemic injury, and increased the plasma level of the anti-inflammatory cytokine IL-10. Interestingly, HO-1 inhibitor zinc protoporphyrin IX (ZnPP-IX; 1 mg/kg) lowered bilirubin and CO concentrations to control values, thus abolishing all the cardioprotective effects of hemin. In conclusion, the results demonstrate that chronic HO-1 activation by prolonged administration of hemin improves survival and exerts protective effects in a rat model of myocardial ischemia by exerting a potent antioxidant activity and disrupting multiple levels of the apoptotic and inflammatory cascade.

  15. Prunella vulgaris Suppresses HG-Induced Vascular Inflammation via Nrf2/HO-1/eNOS Activation

    Ho Sub Lee

    2012-01-01

    Full Text Available Vascular inflammation is an important factor which can promote diabetic complications. In this study, the inhibitory effects of aqueous extract from Prunella vulgaris (APV on high glucose (HG-induced expression of cell adhesion molecules in human umbilical vein endothelial cells (HUVEC are reported. APV decreased HG-induced expression of intercellular adhesion molecule-1 (ICAM-1, vascular cell adhesion molecule-1 (VCAM-1, and E-selectin. APV also dose-dependently inhibited HG-induced adhesion of HL-60 monocytic cells. APV suppressed p65 NF-κB activation in HG-treated cells. APV significantly inhibited the formation of intracellular reactive oxygen species (ROS. HG-stimulated HUVEC secreted gelatinases, however, APV inhibited it. APV induced Akt phosphorylation as well as activation of heme oxygenase-1 (HO-1, eNOS, and nuclear factor E2-related factor 2 (Nrf2, which may protect vascular inflammation caused by HG. In conclusion, APV exerts anti-inflammatory effect via inhibition of ROS/NF-κB pathway by inducing HO-1 and eNOS expression mediated by Nrf2, thereby suggesting that Prunella vulgaris may be a possible therapeutic approach to the inhibition of diabetic vascular diseases.

  16. Heme Oxygenase-1 Induction by Carbon Monoxide Releasing Molecule-3 Suppresses Interleukin-1β-Mediated Neuroinflammation

    Chih-Chung Lin

    2017-11-01

    Full Text Available Neurodegenerative disorders and brain damage are initiated by excessive production of reactive oxygen species (ROS, which leads to tissue injury, cellular death and inflammation. In cellular anti-oxidant systems, heme oxygenase-1 (HO-1 is an oxidative-sensor protein induced by ROS generation or carbon monoxide (CO release. CO releasing molecules (CORMs, including CORM-3, exert anti-oxidant and anti-inflammatory effects. However, the molecular mechanisms of CORM-3-induced HO-1 expression and protection against interleukin (IL-1β-induced inflammatory responses have not been fully elucidated in rat brain astrocytes (RBA-1. To study the regulation of CORM-3-induced HO-1 expression, signaling pathways, promoter activity, mRNA and protein expression were assessed following treatment with pharmacological inhibitors and gene-specific siRNA knockdown. We found that CORM-3 mediated HO-1 induction via transcritional and translational processes. Furthermore, CORM-3-induced HO-1 expression was mediated by phosphorylation of several protein kinases, such as c-Src, Pyk2, protein kinase Cα (PKCα and p42/p44 mitogen-activated protein kinase (MAPK, which were inhibited by respective pharmacological inhibitors or by gene-specific knockdown with siRNA transfections. Next, we found that CORM-3 sequentially activated the c-Src/Pyk2/PKCα/p42/p44 MAPK pathway, thereby up-regulating mRNA for the activator protein (AP-1 components c-Jun and c-Fos; these effects were attenuated by an AP-1 inhibitor (Tanshinone IIA; TSIIA and other relevant inhibitors. Moreover, CORM-3-induced upregulation of HO-1 attenuated the IL-1β-induced cell migration and matrix metallopeptidase-9 mRNA expression in RBA-1 cells. These effects were reversed by an matrix metalloproteinase (MMP2/9 inhibitor or by transfection with HO-1 siRNA.

  17. Heme oxygenase-1 accelerates tumor angiogenesis of human pancreatic cancer.

    Sunamura, Makoto; Duda, Dan G; Ghattas, Maivel H; Lozonschi, Lucian; Motoi, Fuyuhiko; Yamauchi, Jun-Ichiro; Matsuno, Seiki; Shibahara, Shigeki; Abraham, Nader G

    2003-01-01

    Angiogenesis is necessary for the continued growth of solid tumors, invasion and metastasis. Several studies clearly showed that heme oxygenase-1 (HO-1) plays an important role in angiogenesis. In this study, we used the vital microscope system, transparent skinfold model, lung colonization model and transduced pancreatic cancer cell line (Panc-1)/human heme oxygenase-1 (hHO-1) cells, to precisely analyze, for the first time, the effect of hHO-1 gene on tumor growth, angiogenesis and metastasis. Our results revealed that HO-1 stimulates angiogenesis of pancreatic carcinoma in severe combined immune deficient mice. Overexpression of human hHO-1 after its retroviral transfer into Panc-1 cells did not interfere with tumor growth in vitro. While in vivo the development of tumors was accelerated upon transfection with hHO-1. On the other hand, inhibition of heme oxygenase (HO) activity by stannous mesoporphyrin was able transiently to delay tumor growth in a dose dependent manner. Tumor angiogenesis was markedly increased in Panc-1/hHO-1 compared to mock transfected and wild type. Lectin staining and Ki-67 proliferation index confirmed these results. In addition hHO-1 stimulated in vitro tumor angiogenesis and increased endothelial cell survival. In a lung colonization model, overexpression of hHO-1 increased the occurrence of metastasis, while inhibition of HO activity by stannous mesoporphyrin completely inhibited the occurrence of metastasis. In conclusion, overexpression of HO-1 genes potentiates pancreatic cancer aggressiveness, by increasing tumor growth, angiogenesis and metastasis and that the inhibition of the HO system may be of useful benefit for the future treatment of the disease.

  18. Metallothionein-III protects against 6-hydroxydopamine-induced oxidative stress by increasing expression of heme oxygenase-1 in a PI3K and ERK/Nrf2-dependent manner

    Hwang, Yong Pil; Kim, Hyung Gyun; Han, Eun Hee; Jeong, Hye Gwang

    2008-01-01

    The zinc-binding protein metallothionein-III (MT-III) is associated with resistance to neuronal injury. However, the underlying mechanism for its effects is unclear. In this study, we demonstrate that MT-III prevents the accumulation of reactive oxygen species (ROS) in dopaminergic SH-SY5Y cells challenged with the Parkinson's disease-related neurotoxin 6-hydroxydopamine (6-OHDA) by a mechanism that involves phosphatidylinositol 3-kinase (PI3K) and ERK kinase/NF-E2-related factor 2 (Nrf2) dependent induction of the stress response protein heme oxygenase-1 (HO-1). Pretreatment of SH-SY5Y cells with MT-III significantly reduced 6-OHDA-induced generation of ROS, caspase-3 activation, and subsequent cell death. Also, MT-III up-regulates HO-1 expression and this expression confers neuroprotection against oxidative injury induced by 6-OHDA. Moreover, MT-III induces Nrf2 nuclear translocation, which is upstream of MT-III-induced HO-1 expression, and PI3K and ERK1/2 activation, a pathway that is involved in induced Nrf2 nuclear translocation, HO-1 expression and neuroprotection. Taken together, these results suggest that the PI3K and ERK/Nrf2 signaling pathway controls the intracellular levels of ROS by regulating the expression of the antioxidant enzyme HO-1

  19. The Haptoglobin-CD163-Heme Oxygenase-1 Pathway for Hemoglobin Scavenging

    Jens Haugbølle Thomsen

    2013-01-01

    Full Text Available The haptoglobin- (Hp- CD163-heme oxygenase-1 (HO-1 pathway is an efficient captor-receptor-enzyme system to circumvent the hemoglobin (Hb/heme-induced toxicity during physiological and pathological hemolyses. In this pathway, Hb tightly binds to Hp leading to CD163-mediated uptake of the complex in macrophages followed by lysosomal Hp-Hb breakdown and HO-1-catalyzed conversion of heme into the metabolites carbon monoxide (CO, biliverdin, and iron. The plasma concentration of Hp is a limiting factor as evident during accelerated hemolysis, where the Hp depletion may cause serious Hb-induced toxicity and put pressure on backup protecting systems such as the hemopexin-CD91-HO pathway. The Hp-CD163-HO-1 pathway proteins are regulated by the acute phase mediator interleukin-6 (IL-6, but other regulatory factors indicate that this upregulation is a counteracting anti-inflammatory response during inflammation. The heme metabolites including bilirubin converted from biliverdin have overall an anti-inflammatory effect and thus reinforce the anti-inflammatory efficacy of the Hp-CD163-HO-1 pathway. Future studies of animal models of inflammation should further define the importance of the pathway in the anti-inflammatory response.

  20. Regulation of heme oxygenase-1 expression and MAPK pathways in response to kaempferol and rhamnocitrin in PC12 cells

    Hong, J.-T.; Yen, J.-H.; Wang Lisu; Lo, Y.-H.; Chen, Z.-T.; Wu, M.-J.

    2009-01-01

    Oxidative stress has been considered as a major cause of cellular injuries in a variety of clinical abnormalities, especially neural diseases. Our aim of research is to investigate the protective effects and mechanisms of kaempferol and rhamnocitrin (kaempferol-7-methyl ether) on oxidative damage in rat pheochromocytoma PC12 cells induced by a limited supply of serum and hydrogen peroxide (H 2 O 2 ). The current result demonstrated that kaempferol protected PC12 cells from serum deprivation-induced apoptosis. Pretreatment of cells with kaempferol also diminished intracellular generation of reactive oxygen species (ROS) in response to H 2 O 2 and strongly elevated cell viability. RT-Q-PCR and Western blotting revealed that kaempferol and rhamnocitrin significantly induced heme oxygenase (HO)-1 gene expression. Addition of zinc protoporphyrin (Znpp), a HO-1 competitive inhibitor, significantly attenuated their protective effects in H 2 O 2 -treated cells, indicating the vital role of HO-1 in cell resistance to oxidative injury. While investigating the signaling pathways responsible for HO-1 induction, we observed that kaempferol induced sustained extracellular signal-regulated protein kinase 1/2 (ERK1/2) in PC12 cells grown in low serum medium; while rhamnocitrin only stimulated transient ERK cascade. Addition of U0126, a highly selective inhibitor of MEK1/2, which is upstream of ERK1/2, had no effect on kaempferol- or rhamnocitrin-induced HO-1 mRNA expression, indicating no direct cross-talk between these two pathways. Furthermore, both kaempferol and rhamnocitrin were able to persistently attenuate p38 phosphorylation. Taking together, the above findings suggest that kaempferol and rhamnocitrin can augment cellular antioxidant defense capacity, at least in part, through regulation of HO-1 expression and MAPK signal transduction.

  1. In vitro studies on heme oxygenase-1 and P24 antigen HIV-1 level ...

    Background: Heme oxygenase-1 (HO-1) is a protein secreted by immune cells as a part of immune response mechanism.HO-1 can be induced by variety agents that causingoxidative stress, such as exposure to 100% oxygenat2,4 ATA pressure.It plays a vital role in maintaining cellular homeostasis.This study was ...

  2. Rapamycin Induces Heme Oxygenase-1 in Liver but Inhibits Bile Flow Recovery after Ischemia

    Kist, Alwine; Wakkie, Joris; Madu, Max; Versteeg, Ruth; ten Berge, Judith; Nikolic, Andrej; Nieuwenhuijs, Vincent B.; Porte, Robert J.; Padbury, Robert T. A.; Barritt, Greg J.

    Background/Aims. Rapamycin, which is employed in the management of patients undergoing liver surgery, induces the synthesis of heme oxygenase-1 (HO-1) in some non-liver cell types. The aim was to investigate whether rapamycin can induce HO-1 expression in the liver, and to test the effects of

  3. Modulation of cGMP by human HO-1 retrovirus gene transfer in pulmonary microvessel endothelial cells.

    Abraham, Nader G; Quan, Shuo; Mieyal, Paul A; Yang, Liming; Burke-Wolin, Theresa; Mingone, Christopher J; Goodman, Alvin I; Nasjletti, Alberto; Wolin, Michael S

    2002-11-01

    Carbon monoxide (CO) stimulates guanylate cyclase (GC) and increases guanosine 3',5'-cyclic monophosphate (cGMP) levels. We transfected rat-lung pulmonary endothelial cells with a retrovirus-mediated human heme oxygenase (hHO)-1 gene. Pulmonary cells that expressed hHO-1 exhibited a fourfold increase in HO activity associated with decreases in the steady-state levels of heme and cGMP without changes in soluble GC (sGC) and endothelial nitric oxide synthase (NOS) proteins or basal nitrite production. Heme elicited significant increases in CO production and intracellular cGMP levels in both pulmonary endothelial and pulmonary hHO-1-expressing cells. N(omega)-nitro-L-arginine methyl ester (L-NAME), an inhibitor of NOS, significantly decreased cGMP levels in heme-treated pulmonary endothelial cells but not heme-treated hHO-1-expressing cells. In the presence of exogenous heme, CO and cGMP levels in hHO-1-expressing cells exceeded the corresponding levels in pulmonary endothelial cells. Acute exposure of endothelial cells to SnCl2, which is an inducer of HO-1, increased cGMP levels, whereas chronic exposure decreased heme and cGMP levels. These results indicate that prolonged overexpression of HO-1 ultimately decreases sGC activity by limiting the availability of cellular heme. Heme activates sGC and enhances cGMP levels via a mechanism that is largely insensitive to NOS inhibition.

  4. Epigallocatechin Gallate Attenuates Proliferation and Oxidative Stress in Human Vascular Smooth Muscle Cells Induced by Interleukin-1β via Heme Oxygenase-1

    Po-Len Liu

    2014-01-01

    Full Text Available Proliferation of vascular smooth muscle cells (VSMCs triggered by inflammatory stimuli and oxidative stress contributes importantly to atherogenesis. The association of green tea consumption with cardiovascular protection has been well documented in epidemiological observations, however, the underlying mechanisms remain unclear. This study aimed to elucidate the effects of the most active green tea catechin derivative, (−-epigallocatechin-3-gallate (EGCG, in human aortic smooth muscle cells (HASMCs, focusing particularly on the role of a potent anti-inflammatory and antioxidative enzyme heme oxygenase-1 (HO-1. We found that pretreatment of EGCG dose- and time-dependently induced HO-1 protein levels in HASMCs. EGCG inhibited interleukin- (IL-1β-induced HASMC proliferation and oxidative stress in a dose-dependent manner. The HO-1 inducer CoPPIX decreased IL-1β-induced cell proliferation, whereas the HO-1 enzyme inhibitor ZnPPIX significantly reversed EGCG-caused growth inhibition in IL-1β-treated HASMCs. At the molecular level, EGCG treatment significantly activated nuclear factor erythroid-2-related factor (Nrf2 transcription activities. These results suggest that EGCG might serve as a complementary and alternative medicine in the treatment of these pathologies by inducing HO-1 expression and subsequently decreasing VSMC proliferation.

  5. Expression of nerve growth factor and heme oxygenase-1 predict poor survival of breast carcinoma patients

    Noh, Sang Jae; Chung, Myoung Ja; Moon, Woo Sung; Kang, Myoung Jae; Jang, Kyu Yun; Bae, Jun Sang; Jamiyandorj, Urangoo; Park, Ho Sung; Kwon, Keun Sang; Jung, Sung Hoo; Youn, Hyun Jo; Lee, Ho; Park, Byung-Hyun

    2013-01-01

    Nerve growth factor (NGF) is a neurotrophin and has been suggested to induce heme oxygenase-1 (HO1) expression. Although the role of HO1 in tumorigenesis remains controversial, recent evidence suggests NGF and HO1 as tumor-progressing factors. However, the correlative role of NGF and HO1 and their prognostic impact in breast carcinoma is unknown. We investigated the expression and prognostic significance of the expression of NGF and HO1 in 145 cases of breast carcinoma. Immunohistochemical expression of NGF and HO1 was observed in 31% and 49% of breast carcinoma, respectively. The expression of NGF and HO1 significantly associated with each other, and both have a significant association with histologic grade, HER2 expression, and latent distant metastasis. The expression of NGF and HO1 predicted shorter overall survival of breast carcinoma by univariate and multivariate analysis. NGF expression was an independent prognostic indicator for relapse-free survival by multivariate analysis. The combined expression pattern of NGF and HO1 was also an independent prognostic indicator of overall survival and relapse-free survival. The patients with tumors expressing NGF had the shortest survival and the patients with tumor, which did not express NGF or HO1 showed the longest survival time. This study has demonstrated that individual expression of NGF or HO1, and the combined NGF/HO1 expression pattern could be prognostic indicators for breast carcinoma patients

  6. Structure prediction and activity analysis of human heme oxygenase-1 and its mutant.

    Xia, Zhen-Wei; Zhou, Wen-Pu; Cui, Wen-Jun; Zhang, Xue-Hong; Shen, Qing-Xiang; Li, Yun-Zhu; Yu, Shan-Chang

    2004-08-15

    To predict wild human heme oxygenase-1 (whHO-1) and hHO-1 His25Ala mutant (delta hHO-1) structures, to clone and express them and analyze their activities. Swiss-PdbViewer and Antheprot 5.0 were used for the prediction of structure diversity and physical-chemical changes between wild and mutant hHO-1. hHO-1 His25Ala mutant cDNA was constructed by site-directed mutagenesis in two plasmids of E. coli DH5alpha. Expression products were purified by ammonium sulphate precipitation and Q-Sepharose Fast Flow column chromatography, and their activities were measured. rHO-1 had the structure of a helical fold with the heme sandwiched between heme-heme oxygenase-1 helices. Bond angle, dihedral angle and chemical bond in the active pocket changed after Ala25 was replaced by His25, but Ala25 was still contacting the surface and the electrostatic potential of the active pocket was negative. The mutated enzyme kept binding activity to heme. Two vectors pBHO-1 and pBHO-1(M) were constructed and expressed. Ammonium sulphate precipitation and column chromatography yielded 3.6-fold and 30-fold higher purities of whHO-1, respectively. The activity of delta hHO-1 was reduced 91.21% after mutation compared with whHO-1. Proximal His25 ligand is crucial for normal hHO-1 catalytic activity. delta hHO-1 is deactivated by mutation but keeps the same binding site as whHO-1. delta hHO-1 might be a potential inhibitor of whHO-1 for preventing neonatal hyperbilirubinemia.

  7. A novel compound derived from danshensu inhibits apoptosis via upregulation of heme oxygenase-1 expression in SH-SY5Y cells.

    Pan, Li-Long; Liu, Xin-Hua; Jia, Yao-Ling; Wu, Dan; Xiong, Qing-Hui; Gong, Qi-Hai; Wang, Yang; Zhu, Yi-Zhun

    2013-04-01

    Heme oxygenase-1 (HO-1) has potential anti-apoptotic properties. A novel compound [4-(2-acetoxy-3-((R)-3-(benzylthio)-1-methoxy-1-oxopropan-2- ylamino)-3-oxopropyl)-1,2-phenylene diacetate (DSC)] was synthesized by joining danshensu and cysteine through an appropriate linker. This study investigated if the cytoprotective properties of DSC involved the induction of HO-1. We evaluated the cytoprotective effects of DSC on H2O2-induced cell damage, apoptosis, intracellular and mitochondrial reactive oxygen species (ROS) production, mitochondrial membrane potential (ΔΨm) loss, and apoptosis-related proteins expression and its underlying mechanisms. DSC concentration-dependently attenuated cell death, lactate dehydrogenase release, intracellular and mitochondrial ROS production, and ΔΨm collapse, modulated apoptosis-related proteins (Bcl-2, Bax, caspase-3, p53, and cleaved PARP) expression, and inhibited phosphorylation of extracellular signal-regulated kinase 1/2 in SH-SY5Y cells induced by H2O2. In addition, DSC concentration-dependently induced HO-1 expression associated with nuclear translocation of nuclear factor-erythroid 2 related factor 2 (Nrf-2), while the effect of DSC was inhibited by a phosphoinositide 3-kinase (PI3K) inhibitor LY294002. Furthermore, the protective effect of DSC on H2O2-induced cell death was abolished by HO-1 inhibitor ZnPP, but was mimicked by carbon monoxide-releasing moiety CORM-3 or HO-1 by-product bilirubin. Finally, DSC inhibited H2O2-induced changes of Bcl-2, Bax, and caspase-3 expression, and all of these effects were reversed by HO-1 silencing. Induction of HO-1 may be, at least in part, responsible for the anti-apoptotic property of DSC, an effect that involved the activation of PI3K/Akt/Nrf-2 axis. DSC might have the potential for beneficial therapeutic interventions for neurodegenerative diseases. Copyright © 2013. Published by Elsevier B.V.

  8. A vigilant, hypoxia-regulated heme oxygenase-1 gene vector in the heart limits cardiac injury after ischemia-reperfusion in vivo.

    Tang, Yao Liang; Qian, Keping; Zhang, Y Clare; Shen, Leping; Phillips, M Ian

    2005-12-01

    The effect of a cardiac specific, hypoxia-regulated, human heme oxygenase-1 (hHO-1) vector to provide cardioprotection from ischemia-reperfusion injury was assessed. When myocardial ischemia and reperfusion is asymptomatic, the damaging effects are cumulative and patients miss timely treatment. A gene therapy approach that expresses therapeutic genes only when ischemia is experienced is a desirable strategy. We have developed a cardiac-specific, hypoxia-regulated gene therapy "vigilant vector'' system that amplifies cardioprotective gene expression. Vigilant hHO-1 plasmids, LacZ plasmids, or saline (n = 40 per group) were injected into mouse heart 2 days in advance of ischemia-reperfusion injury. Animals were exposed to 60 minutes of ischemia followed by 24 hours of reperfusion. For that term (24 hours) effects, the protein levels of HO-1, inflammatory responses, apoptosis, and infarct size were determined. For long-term (3 week) effects, the left ventricular remodeling and recovery of cardiac function were assessed. Ischemia-reperfusion resulted in a timely overexpression of HO-1 protein. Infarct size at 24 hours after ischemia-reperfusion was significantly reduced in the HO-1-treated animals compared with the LacZ-treated group or saline-treated group (P < .001). The reduction of infarct size was accompanied by a decrease in lipid peroxidant activity, inflammatory cell infiltration, and proapoptotic protein level in ischemia-reperfusion-injured myocardium. The long-term study demonstrated that timely, hypoxia-induced HO-1 overexpression is beneficial in conserving cardiac function and attenuating left ventricle remodelling. The vigilant HO-1 vector provides a protective therapy in the heart for reducing cellular damage during ischemia-reperfusion injury and preserving heart function.

  9. Alpha-7 nicotinic acetylcholine receptor agonist treatment in a rat model of Huntington's disease and involvement of heme oxygenase-1

    Laura Foucault-Fruchard

    2018-01-01

    Full Text Available Neuroinflammation is a common element involved in the pathophysiology of neurodegenerative diseases. We recently reported that repeated alpha-7 nicotinic acetylcholine receptor (α7nAChR activations by a potent agonist such as PHA 543613 in quinolinic acid-injured rats exhibited protective effects on neurons. To further investigate the underlying mechanism, we established rat models of early-stage Huntington's disease by injection of quinolinic acid into the right striatum and then intraperitoneally injected 12 mg/kg PHA 543613 or sterile water, twice a day during 4 days. Western blot assay results showed that the expression of heme oxygenase-1 (HO-1, the key component of the cholinergic anti-inflammatory pathway, in the right striatum of rat models of Huntington's disease subjected to intraperitoneal injection of PHA 543613 for 4 days was significantly increased compared to the control rats receiving intraperitoneal injection of sterile water, and that the increase in HO-1 expression was independent of change in α7nAChR expression. These findings suggest that HO-1 expression is unrelated to α7nAChR density and the increase in HO-1 expression likely contributes to α7nAChR activation-related neuroprotective effect in early-stage Huntington's disease.

  10. Oxidative stress suppression by luteolin-induced heme oxygenase-1 expression

    Sun, Gui-bo; Sun, Xiao; Wang, Min [Key Laboratory of Bioactive Substances and Resources Utilization of Chinese Herbal Medicine, Ministry of Education, Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100193 (China); Ye, Jing-xue [Jilin Agricultural University, No.2888, Xincheng Street, Changchun, 130021, Jilin (China); Si, Jian-yong [Key Laboratory of Bioactive Substances and Resources Utilization of Chinese Herbal Medicine, Ministry of Education, Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100193 (China); Xu, Hui-bo [Academy of Chinese Medical Sciences of Jilin Province, Gongnongda road 1745, Changchun, 130021, Jiblin (China); Meng, Xiang-bao; Qin, Meng; Sun, Jing [Key Laboratory of Bioactive Substances and Resources Utilization of Chinese Herbal Medicine, Ministry of Education, Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100193 (China); Wang, Hong-wei, E-mail: hwang@nju.edu.cn [Center for Translational Medicine and Jiangsu Key Laboratory of Molecular Medicine, Medical School of Nanjing University, Nanjing 210093 (China); Sun, Xiao-bo, E-mail: sunsubmit@163.com [Key Laboratory of Bioactive Substances and Resources Utilization of Chinese Herbal Medicine, Ministry of Education, Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100193 (China)

    2012-12-01

    Luteolin, a flavonoid that exhibits antioxidative properties, exerts myocardial protection effects. However, the underlying molecular mechanisms are not yet fully understood. To investigate the effects of luteolin on myocardial injury protection and its possible mechanisms, a myocardial injury model was established with intragastric administration of 4 mg/kg isoproterenol (ISO) to male Sprague–Dawley rats (200–220 g) daily for 2 days. We found that pretreatment of luteolin (160, 80 and 40 mg/kg, i.g., respectively) daily for 15 days can prevent ISO-induced myocardial damage, including decrease of serum cardiac enzymes, improvement electrocardiography and heart vacuolation. Luteolin also improved the free radical scavenging and antioxidant potential, suggesting one possible mechanism of luteolin-induced cardio-protection is mediated by blocking the oxidative stress. To clarify the mechanisms, we performed the in vitro study by hydrogen peroxide (H{sub 2}O{sub 2})-induced cytotoxicty model in H9c2 cells. We found that luteolin pretreatment prevented apoptosis, increased the expression of heme oxygenase-1 (HO-1), and enhanced the binding of Nrf2 to the antioxidant response element, providing an adaptive survival response against H{sub 2}O{sub 2}-derived oxidative cytotoxicity. The addition of Znpp, a selective HO-1 competitive inhibitor, reduced the cytoprotective ability of luteolin, indicating the vital role of HO-1 on these effects. Luteolin also activated Akt and ERK, whereas the addition of LY294002 and U0126, the pharmacologic inhibitors of PI3K and ERK, attenuated luteolin-induced HO-1 expression and cytoprotective effect. Taken together, the above findings suggest that luteolin protects against myocardial injury and enhances cellular antioxidant defense capacity through the activation of Akt and ERK signal pathways that leads to Nrf2 activation, and subsequently HO-1 induction. -- Highlights: ► Luteolin prevents isoproterenol-induced myocardial damage.

  11. Oxidative stress suppression by luteolin-induced heme oxygenase-1 expression

    Sun, Gui-bo; Sun, Xiao; Wang, Min; Ye, Jing-xue; Si, Jian-yong; Xu, Hui-bo; Meng, Xiang-bao; Qin, Meng; Sun, Jing; Wang, Hong-wei; Sun, Xiao-bo

    2012-01-01

    Luteolin, a flavonoid that exhibits antioxidative properties, exerts myocardial protection effects. However, the underlying molecular mechanisms are not yet fully understood. To investigate the effects of luteolin on myocardial injury protection and its possible mechanisms, a myocardial injury model was established with intragastric administration of 4 mg/kg isoproterenol (ISO) to male Sprague–Dawley rats (200–220 g) daily for 2 days. We found that pretreatment of luteolin (160, 80 and 40 mg/kg, i.g., respectively) daily for 15 days can prevent ISO-induced myocardial damage, including decrease of serum cardiac enzymes, improvement electrocardiography and heart vacuolation. Luteolin also improved the free radical scavenging and antioxidant potential, suggesting one possible mechanism of luteolin-induced cardio-protection is mediated by blocking the oxidative stress. To clarify the mechanisms, we performed the in vitro study by hydrogen peroxide (H 2 O 2 )-induced cytotoxicty model in H9c2 cells. We found that luteolin pretreatment prevented apoptosis, increased the expression of heme oxygenase-1 (HO-1), and enhanced the binding of Nrf2 to the antioxidant response element, providing an adaptive survival response against H 2 O 2 -derived oxidative cytotoxicity. The addition of Znpp, a selective HO-1 competitive inhibitor, reduced the cytoprotective ability of luteolin, indicating the vital role of HO-1 on these effects. Luteolin also activated Akt and ERK, whereas the addition of LY294002 and U0126, the pharmacologic inhibitors of PI3K and ERK, attenuated luteolin-induced HO-1 expression and cytoprotective effect. Taken together, the above findings suggest that luteolin protects against myocardial injury and enhances cellular antioxidant defense capacity through the activation of Akt and ERK signal pathways that leads to Nrf2 activation, and subsequently HO-1 induction. -- Highlights: ► Luteolin prevents isoproterenol-induced myocardial damage. ► Luteolin

  12. Protectin DX suppresses hepatic gluconeogenesis through AMPK-HO-1-mediated inhibition of ER stress.

    Jung, Tae Woo; Kim, Hyung-Chun; Abd El-Aty, A M; Jeong, Ji Hoon

    2017-06-01

    Several studies have shown that protectins, which are ω-3 fatty acid-derived proresolution mediators, may improve insulin resistance. Recently, protectin DX (PDX) was documented to attenuate insulin resistance by stimulating IL-6 expression in skeletal muscle, thereby regulating hepatic gluconeogenesis. These findings made us investigate the direct effects of PDX on hepatic glucose metabolism in the context of diabetes. In the current study, we show that PDX regulates hepatic gluconeogenesis in a manner distinct from its indirect glucoregulatory activity via IL-6. We found that PDX stimulated AMP-activated protein kinase (AMPK) phosphorylation, thereby inducing heme oxygenase 1 (HO-1) expression. This induction blocked hepatic gluconeogenesis by suppressing endoplasmic reticulum (ER) stress in hepatocytes under hyperlipidemic conditions. These effects were significantly dampened by silencing AMPK or HO-1 expression with small interfering RNA (siRNA). We also demonstrated that administration of PDX to high fat diet (HFD)-fed mice resulted in increased hepatic AMPK phosphorylation and HO-1 expression, whereas hepatic ER stress was substantially attenuated. Furthermore, PDX treatment suppressed the expression of gluconeogenic genes, thereby decreasing blood glucose levels in HFD-fed mice. In conclusion, our findings suggest that PDX inhibits hepatic gluconeogenesis via AMPK-HO-1-dependent suppression of ER stress. Thus, PDX may be an effective therapeutic target for the treatment of insulin resistance and type 2 diabetes through the regulation of hepatic gluconeogenesis. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. Mechanisms of HO-1 mediated attenuation of renal immune injury: a gene profiling study.

    Duann, Pu; Lianos, Elias A

    2011-10-01

    Using a mouse model of immune injury directed against the renal glomerular vasculature and resembling human forms of glomerulonephritis (GN), we assessed the effect of targeted expression of the cytoprotective enzyme heme oxygenase (HO)-1. A human (h) HO-1 complementary DNAN (cDNA) sequence was targeted to glomerular epithelial cells (GECs) using a GEC-specific murine nephrin promoter. Injury by administration of antibody against the glomerular basement membrane (anti-GBM) to transgenic (TG) mice with GEC-targeted hHO-1 was attenuated compared with wild-type (WT) controls. To explore changes in the expression of genes that could mediate this salutary effect, we performed gene expression profiling using a microarray analysis of RNA isolated from the renal cortex of WT or TG mice with or without anti-GBM antibody-induced injury. Significant increases in expression were detected in 9 major histocompatibility complex (MHC)-class II genes, 2 interferon-γ (IFN-γ)-inducible guanosine triphosphate (GTP)ases, and 3 genes of the ubiquitin-proteasome system. The increase in MHC-class II and proteasome gene expression in TG mice with injury was validated by real-time polymerase chain reaction (PCR) or Western blot analysis. The observations point to novel mechanisms underlying the cytoprotective effect of HO-1 in renal immune injury. Copyright © 2011. Published by Mosby, Inc.

  14. Kidneys From α1,3-Galactosyltransferase Knockout/Human Heme Oxygenase-1/Human A20 Transgenic Pigs Are Protected From Rejection During Ex Vivo Perfusion With Human Blood.

    Ahrens, Hellen E; Petersen, Björn; Ramackers, Wolf; Petkov, Stoyan; Herrmann, Doris; Hauschild-Quintern, Janet; Lucas-Hahn, Andrea; Hassel, Petra; Ziegler, Maren; Baars, Wiebke; Bergmann, Sabine; Schwinzer, Reinhard; Winkler, Michael; Niemann, Heiner

    2015-07-01

    Multiple modifications of the porcine genome are required to prevent rejection after pig-to-primate xenotransplantation. Here, we produced pigs with a knockout of the α1,3-galactosyltransferase gene (GGTA1-KO) combined with transgenic expression of the human anti-apoptotic/anti-inflammatory molecules heme oxygenase-1 and A20, and investigated their xenoprotective properties. The GGTA1-KO/human heme oxygenase-1 (hHO-1)/human A20 (hA20) transgenic pigs were produced in a stepwise approach using zinc finger nuclease vectors targeting the GGTA1 gene and a Sleeping Beauty vector coding for hA20. Two piglets were analyzed by quantitative reverse-transcription polymerase chain reaction, flow cytometry, and sequencing. The biological function of the genetic modifications was tested in a (51)Chromium release assay and by ex vivo kidney perfusions with human blood. Disruption of the GGTA1 gene by deletion of few basepairs was demonstrated in GGTA1-KO/hHO-1/hA20 transgenic pigs. The hHO-1 and hA20 mRNA expression was confirmed by quantitative reverse-transcription polymerase chain reaction. Ex vivo perfusion of 2 transgenic kidneys was feasible for the maximum experimental time of 240 minutes without symptoms of rejection. Results indicate that GGTA1-KO/hHO-1/hA20 transgenic pigs are a promising model to alleviate rejection and ischemia-reperfusion damage in porcine xenografts and could serve as a background for further genetic modifications toward the production of a donor pig that is clinically relevant for xenotransplantation.

  15. hHO-1 combined with GATA-4 transduction promotes myocardial transdifferentiation and anti- apoptosis of rat mesenchymal stem cells

    Ning-bo DENG

    2017-06-01

    Full Text Available Objectives To explore if the rat bone marrow mesenchymal stem cells (BMSCs modified by human heme oxygenase 1 (hHO-1 gene combined with GATA-4 gene may promote the ability of anti-apoptosis and myocardial transdifferentiation in vitro in hypoxia ischemic environment. Methods The rat BMSCs were isolated and cultured by whole bone marrow adherence and identified in vitro, and then were transfected with recombinant adenovirus; Western blotting was used to determinate the optimal time of gene expression; the genetically modified BMSCs were taken to hypoxia serum-free conditions simulating ischemia hypoxia microenvironment in vivo; CCK-8 kit and trypan blue staining were performed to detect the 12, 24, 48 and 72h survival rates in hypoxia ischemia respectively; flow cytometry was used to detect the apoptosis of BMSCs in hypoxia ischemia for 24h. The cardiomyocyte-specific cardiac troponin I (cTnI was detected by Western blotting and cellular immunofluorescence. Results The 12, 24, 48 and 72h survival rates were higher in hHO-1+GATA-4 group cultured in ischemia and hypoxia condition than in hHO-1 group (P<0.05 and GATA-4 group (P<0.05. After 24h cultivation in ischemia hypoxia condition, the apoptotic rates were lower in hHO-1+GATA-4 group than in hHO-1 group (P<0.05 and GATA-4 group (P<0.05. No significant difference existed in cTnI expressions between GATA-4 group and hHO-1+GATA-4 group. Conclusion Compared with transfection of hHO-1 or GATA-4 single gene, hHO-1 combined with GATA-4 transduction can significantly increase the survival rate of BMSCs in hypoxia ischemic condition, but myocardial transdifferentiation does not increase significantly. DOI: 10.11855/j.issn.0577-7402.2017.04.08

  16. Embryotoxicity Caused by DON-Induced Oxidative Stress Mediated by Nrf2/HO-1 Pathway

    Miao Yu

    2017-06-01

    Full Text Available Deoxynivalenol (DON belongs to the type B group of trichothecenes family, which is composed of sesquiterpenoid metabolites produced by Fusarium and other fungi in grain. DON may cause various toxicities, such as cytotoxicity, immunotoxicity, genotoxicity as well as teratogenicity and carcinogenicity. In the present study, we focus on a hypothesis that DON alters the expressions of Nrf2/HO-1 pathway by inducing embryotoxicity in C57BL/6 mouse (5.0, 2.5, 1.0, and 0 mg/kg/day and BeWo cell lines (0 and 50 nM; 3 h, 12 h and 24 h. Our results indicate that DON treatment in mice during pregnancy leads to ROS accumulation in the placenta, which results in embryotoxicity. At the same time Nrf2/HO-1 pathway is up-regulated by ROS to protect placenta cells from oxidative damage. In DON-treated BeWo cells, the level of ROS has time–effect and dose–effect relationships with HO-1 expression. Moderate increase in HO-1 protects the cell from oxidative damage, while excessive increase in HO-1 aggravates the oxidative damage, which is called in some studies the “threshold effect”. Therefore, oxidative stress may be the critical molecular mechanism for DON-induced embryotoxicity. Besides, Nrf2/HO-1 pathway accompanied by the “threshold effect” also plays an important role against DON-induced oxidative damage in this process.

  17. Potential Ameliorative Effects of Qing Ye Dan Against Cadmium Induced Prostatic Deficits via Regulating Nrf-2/HO-1 and TGF-β1/Smad Pathways.

    Du, Lifen; Lei, Yongfang; Chen, Jinglou; Song, Hongping; Wu, Xinying

    2017-01-01

    Cadmium (Cd) is an environmental pollutant with reproductive toxicity. Swertia mileensis is used in Chinese medicine for the treatment of prostatic deficits and named as Qing Ye Dan (QYD). This study was undertaken to investigate the potential protective effects of QYD against Cd-induced prostatic deficits. Rat model of prostatic deficits was induced by 0.2 mg/kg/d CdCl2 subcutaneous injection for 15 days. The prostatic oxidative stress was evaluated by detecting the levels of malondialdehyde, nitric oxide, reduced/ oxidized glutathione, total sulfhydryl groups and enzymatic antioxidant status. The prostatic inflammation was estimated by testing the levels of pro-inflammatory cytokines. The levels of epithelial-mesenchymal transition (EMT) markers E-cadherin, fibronectin, vimentin and α-smooth muscle actin were measured by qPCR analysis. Additionally, the prostatic expressions of transforming growth factor-β1 (TGF-β1), type I TGF-β receptor (TGF-βRI), Smad2, phosphorylation-Smad2 (p-Smad2), Smad3, p-Smad3, Smad7, nuclear related factor-2 (Nrf-2), heme oxygenase-1 (HO-1), B-cell CLL/lymphoma (Bcl)-2 and Bcl-2-associated X protein (Bax) were measured by western blot assay. It was found that QYD ameliorated the Cd-induced prostatic oxidative stress and inflammation, attenuated prostatic EMT, inhibited the TGF-β1/Smad pathway, increased Bcl-2/Bax ratio and enhanced the activity of Nrf-2/HO-1 pathway. These results showed that QYD could ameliorate Cd-induced prostatic deficits via modulating Nrf-2/HO-1 and TGF-β1/Smad pathways. © 2017 The Author(s). Published by S. Karger AG, Basel.

  18. BTB and CNC homolog 1 (Bach1) deficiency ameliorates TNBS colitis in mice: role of M2 macrophages and heme oxygenase-1.

    Harusato, Akihito; Naito, Yuji; Takagi, Tomohisa; Uchiyama, Kazuhiko; Mizushima, Katsura; Hirai, Yasuko; Higashimura, Yasuki; Katada, Kazuhiro; Handa, Osamu; Ishikawa, Takeshi; Yagi, Nobuaki; Kokura, Satoshi; Ichikawa, Hiroshi; Muto, Akihiko; Igarashi, Kazuhiko; Yoshikawa, Toshikazu

    2013-01-01

    BTB and CNC homolog 1 (Bach1) is a transcriptional repressor of heme oxygenase-1 (HO-1), which plays an important role in the protection of cells and tissues against acute and chronic inflammation. However, the role of Bach1 in the gastrointestinal mucosal defense system remains little understood. HO-1 supports the suppression of experimental colitis and localizes mainly in macrophages in colonic mucosa. This study was undertaken to elucidate the Bach1/HO-1 system's effects on the pathogenesis of experimental colitis. This study used C57BL/6 (wild-type) and homozygous Bach1-deficient C57BL/6 mice in which colonic damage was induced by the administration of an enema of 2,4,6-trinitrobenzene sulfonic acid (TNBS). Subsequently, they were evaluated macroscopically, histologically, and biochemically. Peritoneal macrophages from the respective mice were isolated and analyzed. Then, wild-type mice were injected with peritoneal macrophages from the respective mice. Acute colitis was induced similarly. TNBS-induced colitis was inhibited in Bach1-deficient mice. TNBS administration increased the expression of HO-1 messenger RNA and protein in colonic mucosa in Bach1-deficient mice. The expression of HO-1 mainly localized in F4/80-immunopositive and CD11b-immunopositive macrophages. Isolated peritoneal macrophages from Bach1-deficient mice highly expressed HO-1 and also manifested M2 macrophage markers, such as Arginase-1, Fizz-1, Ym1, and MRC1. Furthermore, TNBS-induced colitis was inhibited by the transfer of Bach1-deficient macrophages into wild-type mice. Deficiency of Bach1 ameliorated TNBS-induced colitis. Bach1-deficient macrophages played a key role in protection against colitis. Targeting of this mechanism is applicable to cell therapy for human inflammatory bowel disease.

  19. Baicalein inhibition of oxidative-stress-induced apoptosis via modulation of ERKs activation and induction of HO-1 gene expression in rat glioma cells C6

    Chen, Y.-C.; Chow, J.-M.; Lin, C.-W.; Wu, C.-Y.; Shen, S.-C.

    2006-01-01

    In the present study, we examined the protective mechanism of baicalein (BE) and its glycoside, baicalin (BI), on hydrogen-peroxide (H 2 O 2 )-induced cell death in rat glioma C6 cells. Results of the MTT assay, LDH release assay, and morphological observation showed that H 2 O 2 addition reduced the viability of C6 cells, and this was prevented by the addition of BE but not BI. Incubation of C6 cells with BE significantly decreased the intracellular peroxide level induced by H 2 O 2 according to flow cytometric analysis using DCHF-DA as a fluorescent substrate. Suppression of H 2 O 2 -induced apoptotic events including DNA ladders, hypodiploid cells, and activation of caspases 3, 8, and, 9 by BE but not BI was identified in C6 cells. The cytotoxicity and phosphorylation of ERK proteins induced by H 2 O 2 were blocked by the ERK inhibitor PD98059. Catalase addition prevented H 2 O 2 -induced ROS production, ERKs protein phosphorylation, and cell death, and BE dose-dependently inhibited H 2 O 2 -induced ERK protein phosphorylation in C6 cells. These data suggest that ROS-scavenging activity is involved in BE prevention of H 2 O 2 -induced cell death via blocking ERKs activation. Additionally, BE but not BI induced heat shock protein 32 (HSP32; HO-1) protein expression in both time- and dose-dependent manners, but not heme oxygenase 2 (HO-2), heat shock protein 70 (HSP70), or heat shock protein 90 (HSP90) protein expression. In the absence of H 2 O 2 , BE induces ERKs protein phosphorylation, and HO-1 protein expression induced by BE was blocked by the addition of cycloheximide, actinomycin D, and the ERK inhibitor PD98059. The addition of the HO inhibitor ZnPP inhibited the protective effect of BE against H 2 O 2 -induced cytotoxicity in C6 cells according to the MTT assay and apoptotic morphology under microscopic observation, accompanied by blocking the ROS-scavenging activity of BE in C6 cells. However, BE treatment was unable to protect C6 cells from C2-ceramide

  20. Effect of heme oxygenase-1 transduced bone marrow mesenchymal stem cells on damaged intestinal epithelial cells in vitro.

    Cao, Yi; Wu, Ben-Juan; Zheng, Wei-Ping; Yin, Ming-Li; Liu, Tao; Song, Hong-Li

    2017-07-01

    In this study, we explored the effects of mesenchymal stem cells (MSCs) from bone marrow overexpressing heme oxygenase-1 (HO-1) on the damaged human intestinal epithelial barrier in vitro. Rat MSCs were isolated from bone marrow and transduced with rat HO-1 recombinant adenovirus (HO-MSCs) for stable expression of HO-1. Colorectal adenocarinoma 2 (Caco2) cells were treated with tumor necrosis factor-α (TNF-α) to establish a damaged colon epithelial model. Damaged Caco2 were cocultured with MSCs, Ad-MSCs, Ad-HO + MSCs or HO-MSCs. mRNA and protein expression of Zona occludens-1 (ZO-1) and human HO-1 and the release of cytokines were measured. ZO-1 and human HO-1 in Caco2 were significantly decreased after treatment with TNF-α; and this effect was reduced when coculture with MSCs from bone marrow. Expression of ZO-1 was not significantly affected by Caco2 treatment with TNF-α, Ad-HO, and MSCs. In contrast, ZO-1 and human HO-1 increased significantly when the damaged Caco2 was treated with HO-MSCs. HO-MSCs showed the strongest effect on the expression of ZO-1 in colon epithelial cells. Coculture with HO-MSCs showed the most significant effects on reducing the expression of IL-2, IL-6, IFN-γ and increasing the expression of IL-10. HO-MSCs protected the intestinal epithelial barrier, in which endogenous HO-1 was involved. HO-MSCs play an important role in the repair process by reducing the release of inflammatory cytokines and increasing the release of anti-inflammatory factors. These results suggested that HO-MSCs from bone marrow were more effective in repairing the damaged intestinal epithelial barrier, and the effectiveness of MSCs was improved by HO-1 gene transduction, which provides favorable support for the application of stem cell therapy in the intestinal diseases. © 2017 The Authors. Cell Biology International Published by John Wiley & Sons Ltd on behalf of International Federation of Cell Biology.

  1. Selective activation of heme oxygenase-2 by menadione.

    Vukomanovic, Dragic; McLaughlin, Brian E; Rahman, Mona N; Szarek, Walter A; Brien, James F; Jia, Zongchao; Nakatsu, Kanji

    2011-11-01

    While substantial progress has been made in elucidating the roles of heme oxygenases-1 (HO-1) and -2 (HO-2) in mammals, our understanding of the functions of these enzymes in health and disease is still incomplete. A significant amount of our knowledge has been garnered through the use of nonselective inhibitors of HOs, and our laboratory has recently described more selective inhibitors for HO-1. In addition, our appreciation of HO-1 has benefitted from the availability of tools for increasing its activity through enzyme induction. By comparison, there is a paucity of information about HO-2 activation, with only a few reports appearing in the literature. This communication describes our observations of the up to 30-fold increase in the in-vitro activation of HO-2 by menadione. This activation was due to an increase in Vmax and was selective, in that menadione did not increase HO-1 activity.

  2. GT-repeat polymorphism in the heme oxygenase-1 gene promoter is associated with cardiovascular mortality risk in an arsenic-exposed population in northeastern Taiwan

    Wu, Meei-Maan; Chiou, Hung-Yi; Chen, Chi-Ling; Wang, Yuan-Hung; Hsieh, Yi-Chen; Lien, Li-Ming; Lee, Te-Chang; Chen, Chien-Jen

    2010-01-01

    Inorganic arsenic has been associated with increased risk of atherosclerotic vascular disease and mortality in humans. A functional GT-repeat polymorphism in the heme oxygenase-1 (HO-1) gene promoter is inversely correlated with the development of coronary artery disease and restenosis after clinical angioplasty. The relationship of HO-1 genotype with arsenic-associated cardiovascular disease has not been studied. In this study, we evaluated the relationship between the HO-1 GT-repeat polymorphism and cardiovascular mortality in an arsenic-exposed population. A total of 504 study participants were followed up for a median of 10.7 years for occurrence of cardiovascular deaths (coronary heart disease, cerebrovascular disease, and peripheral arterial disease). Cardiovascular risk factors and DNA samples for determination of HO-1 GT repeats were obtained at recruitment. GT repeats variants were grouped into the S (< 27 repeats) or L allele (≥ 27 repeats). Relative mortality risk was estimated using Cox regression analysis, adjusted for competing risk of cancer and other causes. For the L/L, L/S, and S/S genotype groups, the crude mortalities for cardiovascular disease were 8.42, 3.10, and 2.85 cases/1000 person-years, respectively. After adjusting for conventional cardiovascular risk factors and competing risk of cancer and other causes, carriers with class S allele (L/S or S/S genotypes) had a significantly reduced risk of cardiovascular mortality compared to non-carriers (L/L genotype) [OR, 0.38; 95% CI, 0.16-0.90]. In contrast, no significant association was observed between HO-1 genotype and cancer mortality or mortality from other causes. Shorter (GT)n repeats in the HO-1 gene promoter may confer protective effects against cardiovascular mortality related to arsenic exposure.

  3. Heme oxygenase-1 mediates BAY 11-7085 induced ferroptosis.

    Chang, Ling-Chu; Chiang, Shih-Kai; Chen, Shuen-Ei; Yu, Yung-Luen; Chou, Ruey-Hwang; Chang, Wei-Chao

    2018-03-01

    Ferroptosis is a form of oxidative cell death and has become a chemotherapeutic target for cancer treatment. BAY 11-7085 (BAY), which is a well-known IκBα inhibitor, suppressed viability in cancer cells via induction of ferroptotic death in an NF-κB-independent manner. Reactive oxygen species scavenging, relief of lipid peroxidation, replenishment of glutathione and thiol-containing agents, as well as iron chelation, rescued BAY-induced cell death. BAY upregulated a variety of Nrf2 target genes related to redox regulation, particularly heme oxygenase-1 (HO-1). Studies with specific inhibitors and shRNA interventions suggested that the hierarchy of induction is Nrf2-SLC7A11-HO-1. SLC7A11 inhibition by erastin, sulfasalazine, or shRNA interference sensitizes BAY-induced cell death. Overexperession of SLC7A11 attenuated BAY-inhibited cell viability. The ferroptotic process induced by hHO-1 overexpression further indicated that HO-1 is a key mediator of BAY-induced ferroptosis that operates through cellular redox regulation and iron accumulation. BAY causes compartmentalization of HO-1 into the nucleus and mitochondrion, and followed mitochondrial dysfunctions, leading to lysosome targeting for mitophagy. In this study, we first discovered that BAY induced ferroptosis via Nrf2-SLC7A11-HO-1 pathway and HO-1 is a key mediator by responding to the cellular redox status. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Heme oxygenase-1 comes back to endoplasmic reticulum

    Kim, Hong Pyo [School of Biological Sciences, Ulsan University (Korea, Republic of); Pae, Hyun-Ock [Department of Immunology, Wonkwang University School of Medicine (Korea, Republic of); Back, Sung Hun; Chung, Su Wol [School of Biological Sciences, Ulsan University (Korea, Republic of); Woo, Je Moon [Department of Opthalmology, Ulasn University Hospital (Korea, Republic of); Son, Yong [Department of Anesthesiology and Pain Medicine, Wonkwang University School of Medicine (Korea, Republic of); Chung, Hun-Taeg, E-mail: chung@ulsan.ac.kr [School of Biological Sciences, Ulsan University (Korea, Republic of)

    2011-01-07

    Research highlights: {yields} Although multiple compartmentalization of HO-1 has been documented, the functional implication of this enzyme at these subcellular organelles is only partially elucidated. {yields} HO-1 expression at ER is induced by a diverse set of conditions that cause ER stressors. {yields} CO may induce HO-1 expression in human ECs by activating Nrf2 through PERK phosphorylation in a positive-feedback manner. {yields} ER-residing HO-1 and its cytoprotective activity against ER stress is discussed. -- Abstract: Originally identified as a rate-limiting enzyme for heme catabolism, heme oxygenase-1 (HO-1) has expanded its roles in anti-inflammation, anti-apoptosis and anti-proliferation for the last decade. Regulation of protein activity by location is well appreciated. Even though multiple compartmentalization of HO-1 has been documented, the functional implication of this enzyme at these subcellular organelles is only partially elucidated. In this review we discuss the endoplasmic reticulum (ER)-residing HO-1 and its cytoprotective activity against ER stress.

  5. Increased Plasma Levels of Heme Oxygenase-1 in Human Brucellosis.

    Chen, Zhe; Zhang, Yu-Xue; Fu, Dong-Wei; Gao, Qing-Feng; Ge, Feng-Xia; Liu, Wei-Hua

    2016-08-01

    Brucellosis is associated with inflammation and the oxidative stress response. Heme oxygenase-1 (HO-1) is a cytoprotective stress-responsive enzyme that has anti-inflammatory and anti-oxidant effects. Nevertheless, the role of HO-1 in human brucellosis has not yet been studied. The aim of this study was to examine the plasma levels of HO-1 in patients with brucellosis and to evaluate the ability of plasma HO-1 levels as an auxiliary diagnosis, a severity predictor, and a monitor for brucellosis treatments. A total of 75 patients with brucellosis were divided into the acute, subacute, chronic active, and chronic stable groups. An additional 20 volunteers were included as the healthy control group. The plasma HO-1 levels and other laboratory parameters were measured in all groups. Furthermore, the plasma levels of HO-1 in the acute group were compared before and after treatment. The plasma HO-1 levels were considerably increased in the acute (4.97 ± 3.55), subacute (4.98 ± 3.23), and chronic active groups (4.43 ± 3.00) with brucellosis compared to the healthy control group (1.03 ± 0.63) (p brucellosis (r = 0.707, p brucellosis status and may be used as a supplementary plasma marker for diagnosing brucellosis and monitoring its treatment.

  6. The role of heme oxygenase-1 in systemic-onset juvenile idiopathic arthritis.

    Takahashi, Akitaka; Mori, Masaaki; Naruto, Takuya; Nakajima, Shoko; Miyamae, Takako; Imagawa, Tomoyuki; Yokota, Shumpei

    2009-01-01

    We have determined the serum levels of heme oxygenase-1 (HO-1) in 56 patients with systemic-onset juvenile idiopathic arthritis (s-JIA) and compared these with serum HO-1 levels in healthy controls and patients with other pediatric rheumatic diseases. Serum HO-1 levels were measured by the sandwich enzyme-linked immunosorbent assay. The mean serum HO-1 level in s-JIA patients during the active phase was 123.6 +/- 13.83 ng/ml, which was significantly higher than that in patients with polyarticular juvenile idiopathic arthritis (p-JIA), Kawasaki disease, systemic lupus erythematosus or mixed connective tissue disease (P < 0.0005). The serum levels of HO-1, cytokines and cytokine receptors in patients with s-JIA were also assessed at both the active and inactive phases. The serum HO-1 level in patients with s-JIA in the active phase was found to be significantly greater than that in patients with the disease in the inactive phase (P < 0.0001). An assessment of the relationships between serum HO-1 levels and other laboratory parameters or cytokines in patients with s-JIA did not reveal any strong correlations. These results suggest that the serum level of HO-1 may be a useful marker for the differential diagnosis of s-JIA. Further study will be necessary to elucidate the mechanism of HO-1 production and to clarify the role of HO-1 in the disease process.

  7. A role for heme oxygenase-1 in the antioxidant and antiapoptotic effects of erythropoietin: the start of a good news/bad news story?

    Calò, Lorenzo A; Davis, Paul A; Piccoli, Antonio; Pessina, Achille C

    2006-01-01

    Erythropoietin (EPO) is the major regulator of erythropoiesis. EPO's actions have been shown to be antiapoptotic and dependent on JAK2 signaling and Akt phosphorylation. These effects serve as link between EPO and heme oxygenase-1 (HO-1). HO-1 is an inducible enzyme with potent antioxidant and antiapoptotic activities which are regulated by Akt signaling. EPO's ability to alter cellular systems that involve apoptosis and oxidants suggests that EPO treatments are likely to have multiple and different effects which may start a good news/bad news story. Recombinant human EPO is the recognized treatment of choice to address anemia and to stimulate erythropoiesis in chronic renal failure patients, through its antiapoptotic action which likely involves HO-1. On the other hand, EPO treatment to address anemia in cancer patients, while providing significant improvements in cancer patients' quality of life, its effects on survival are equivocal, likely due to its linkage with HO-1. Two clinical trials of EPO in patients with solid tumors have, in fact, shown specific negative effects on survival. However, EPO's effect on tumor growth and survival is not uniformily pro growth and pro survival, as EPO may act synergistically with chemotherapy to induce apoptosis. Finally, compounds have been synthesized that do not trigger EPO receptor and thus may allow experimental distinction and, therefore, at least potentially affect at the clinical level the tissue-protective effects of EPO (e.g., antiapoptosis) without provoking its other potentially detrimental effects. Copyright 2006 S. Karger AG, Basel

  8. Heme oxygenase-1 modulates degeneration of the intervertebral disc after puncture in Bach 1 deficient mice.

    Ohta, Ryo; Tanaka, Nobuhiro; Nakanishi, Kazuyoshi; Kamei, Naosuke; Nakamae, Toshio; Izumi, Bunichiro; Fujioka, Yuki; Ochi, Mitsuo

    2012-09-01

    Intervertebral disc degeneration is considered to be a major feature of low back pain. Furthermore, oxidative stress has been shown to be an important factor in degenerative diseases such as osteoarthritis and is considered a cause of intervertebral disc degeneration. The purpose of this study was to clarify the correlation between oxidative stress and intervertebral disc degeneration using Broad complex-Tramtrack-Bric-a-brac and cap'n'collar homology 1 deficient (Bach 1-/-) mice which highly express heme oxygenase-1 (HO-1). HO-1 protects cells from oxidative stress. Caudal discs of 12-week-old and 1-year-old mice were evaluated as age-related models. Each group and period, 5 mice (a total of 20 mice, a total of 20 discs) were evaluated as age-related model. C9-C10 caudal discs in 12-week-old Bach 1-/- and wild-type mice were punctured using a 29-gauge needle as annulus puncture model. Each group and period, 5 mice (a total of 60 mice, a total of 60 discs) were evaluated. The progress of disc degeneration was evaluated at pre-puncture, 1, 2, 4, 8 and 12 weeks post-puncture. Radiographic, histologic and immunohistologic analysis were performed to compare between Bach 1-/- and wild-type mice. In the age-related model, there were no significant differences between Bach 1-/- and wild-type mice radiologically and histologically. However, in the annulus puncture model, histological scoring revealed significant difference at 8 and 12 weeks post-puncture. The number of HO-1 positive cells was significantly greater in Bach 1-/- mice at every period. The apoptosis rate was significantly lower at 1 and 2 weeks post-puncture in Bach 1-/- mice. Oxidative stress prevention may avoid the degenerative process of the intervertebral disc after puncture, reducing the number of apoptosis cells. High HO-1 expression may also inhibit oxidative stress and delay the process of intervertebral disc degeneration.

  9. Heme oxygenase-1: a metabolic nike.

    Wegiel, Barbara; Nemeth, Zsuzsanna; Correa-Costa, Matheus; Bulmer, Andrew C; Otterbein, Leo E

    2014-04-10

    Heme degradation, which was described more than 30 years ago, is still very actively explored with many novel discoveries on its role in various disease models every year. The heme oxygenases (HO) are metabolic enzymes that utilize NADPH and oxygen to break apart the heme moiety liberating biliverdin (BV), carbon monoxide (CO), and iron. Heme that is derived from hemoproteins can be toxic to the cells and if not removed immediately, it causes cell apoptosis and local inflammation. Elimination of heme from the milieu enables generation of three products that influences numerous metabolic changes in the cell. CO has profound effects on mitochondria and cellular respiration and other hemoproteins to which it can bind and affect their function, while BV and bilirubin (BR), the substrate and product of BV, reductase, respectively, are potent antioxidants. Sequestration of iron into ferritin and its recycling in the tissues is a part of the homeodynamic processes that control oxidation-reduction in cellular metabolism. Further, heme is an important component of a number of metabolic enzymes, and, therefore, HO-1 plays an important role in the modulation of cellular bioenergetics. In this review, we describe the cross-talk between heme oxygenase-1 (HO-1) and its products with other metabolic pathways. HO-1, which we have labeled Nike, the goddess who personified victory, dictates triumph over pathophysiologic conditions, including diabetes, ischemia, and cancer.

  10. Heme oxygenase-1 induction improves cardiac function following myocardial ischemia by reducing oxidative stress.

    Yossi Issan

    Full Text Available Oxidative stress plays a key role in exacerbating diabetes and cardiovascular disease. Heme oxygenase-1 (HO-1, a stress response protein, is cytoprotective, but its role in post myocardial infarction (MI and diabetes is not fully characterized. We aimed to investigate the protection and the mechanisms of HO-1 induction in cardiomyocytes subjected to hypoxia and in diabetic mice subjected to LAD ligation.In vitro: cultured cardiomyocytes were treated with cobalt-protoporphyrin (CoPP and tin protoporphyrin (SnPP prior to hypoxic stress. In vivo: CoPP treated streptozotocin-induced diabetic mice were subjected to LAD ligation for 2/24 h. Cardiac function, histology, biochemical damage markers and signaling pathways were measured.HO-1 induction lowered release of lactate dehydrogenase (LDH and creatine phospho kinase (CK, decreased propidium iodide staining, improved cell morphology and preserved mitochondrial membrane potential in cardiomyocytes. In diabetic mice, Fractional Shortening (FS was lower than non-diabetic mice (35±1%vs.41±2, respectively p<0.05. CoPP-treated diabetic animals improved cardiac function (43±2% p<0.01, reduced CK, Troponin T levels and infarct size compared to non-treated diabetic mice (P<0.01, P<0.001, P<0.01 respectively. CoPP-enhanced HO-1 protein levels and reduced oxidative stress in diabetic animals, as indicated by the decrease in superoxide levels in cardiac tissues and plasma TNFα levels (p<0.05. The increased levels of HO-1 by CoPP treatment after LAD ligation led to a shift of the Bcl-2/bax ratio towards the antiapoptotic process (p<0.05. CoPP significantly increased the expression levels of pAKT and pGSK3β (p<0.05 in cardiomyocytes and in diabetic mice with MI. SnPP abolished CoPP's cardioprotective effects.HO-1 induction plays a role in cardioprotection against hypoxic damage in cardiomyocytes and in reducing post ischemic cardiac damage in the diabetic heart as proved by the increased levels of pAKT with

  11. HO-1 gene overexpression enhances the beneficial effects of superparamagnetic iron oxide labeled bone marrow stromal cells transplantation in swine hearts underwent ischemia/reperfusion: an MRI study.

    Jiang, Yibo; Chen, Lijuan; Tang, Yaoliang; Ma, Genshan; Shen, Chengxing; Qi, Chunmei; Zhu, Qi; Yao, Yuyu; Liu, Naifeng

    2010-05-01

    To determine the effect of intracoronary transfer of superparamagnetic iron oxide (SPIO) labeled heme oxygenase-1 (HO-1) overexpressed bone marrow stromal cells (BMSCs) in a porcine myocardial ischemia/reperfusion model. Cell apoptosis was assayed and supernatant cytokine concentrations were measured in BMSCs that underwent hypoxia/reoxygen in vitro. Female mini-swines that underwent 1 h LAD occlusion followed by 1 h reperfusion were randomly allocated to receive intracoronary saline (control), 1 x 10(7) SPIO-labeled BMSCs transfected with pcDNA3.1-Lacz plasmid (Lacz-BMSCs), pcDNA3.1-human HO-1 (HO-1-BMSCs), pcDNA3.1-hHO-1 pretreated with a HO inhibitor, tin protoporphyrin (SnPP, n = 10 each). MRI and postmortem histological analysis were made at 1 week or 3 months thereafter. Post hypoxia/reoxygen in vitro, apoptosis was significantly reduced, supernatant VEGF significantly increased while TNF-alpha and IL-6 significantly reduced in HO-1-BMSCs group compared with Lacz-BMSCs group (all p < 0.05). Myocardial expression of VEGF was significantly higher in HO-1-BMSCs than in Lacz-BMSCs group at 1 week post transplantation (all p < 0.05). Signal voids induced by the SPIO were detected in the peri-infarction region in all BMSC groups at 1 week but not at 3 months post transplantation and the extent of the hypointense signal was the highest in HO-1-BMSCs group, and histological analysis showed that signal voids represented cardiac macrophages that engulfed the SPIO-labeled BMSCs. Pretreatment with SnPP significantly attenuated the beneficial effects of HO-1-BMSCs. Transplantation of HO-1-overexpressed BMSCs significantly enhanced the beneficial effects of BMSCs on improving cardiac function in this model.

  12. Small molecule activators of the Nrf2-HO-1 antioxidant axis modulate heme metabolism and inflammation in BV2 microglia cells.

    Foresti, Roberta; Bains, Sandip K; Pitchumony, Tamil Selvi; de Castro Brás, Lisandra E; Drago, Filippo; Dubois-Randé, Jean-Luc; Bucolo, Claudio; Motterlini, Roberto

    2013-10-01

    The nuclear factor erythroid derived 2-related factor 2 (Nrf2) and the antioxidant protein heme oxygenase-1 (HO-1) are crucial components of the cellular stress response. These two systems work together to combat oxidative stress and inflammation and are attractive drug targets for counteracting different pathologies, including neuroinflammation. We aimed to identify the most effective Nrf2/HO-1 activators that modulate the inflammatory response in microglia cells. In the present study, we searched the literature and selected 56 compounds reported to activate Nrf2 or HO-1 and analyzed them for HO-1 induction at 6 and 24h and cytotoxicity in BV2 microglial cells in vitro. Approximately 20 compounds up-regulated HO-1 at the concentrations tested (5-20 μM) with carnosol, supercurcumin, cobalt protoporphyrin-IX and dimethyl fumarate exhibiting the best induction/low cytotoxicity profile. Up-regulation of HO-1 by some compounds resulted in increased cellular bilirubin levels but did not augment the expression of proteins involved in heme synthesis (ALAS 1) or biliverdin reductase. Bilirubin production by HO-1 inducers correlated with their potency in inhibiting nitrite production after challenge with interferon-γ (INF-γ) or lipopolysaccharide (LPS). The compounds down-regulated the inflammatory response (TNF-α, PGE2 and nitrite) more strongly in cells challenged with INF-γ than LPS, and silencing HO-1 or Nrf2 with shRNA differentially affected the levels of inflammatory markers. These findings indicate that some small activators of Nrf2/HO-1 are effective modulators of microglia inflammation and highlight the chemical scaffolds that can serve for the synthesis of potent new derivatives to counteract neuroinflammation and neurodegeneration. Copyright © 2013 Elsevier Ltd. All rights reserved.

  13. Activation of the complement cascade enhances motility of leukemic cells by downregulating expression of HO-1.

    Abdelbaset-Ismail, A; Borkowska-Rzeszotek, S; Kubis, E; Bujko, K; Brzeźniakiewicz-Janus, K; Bolkun, L; Kloczko, J; Moniuszko, M; Basak, G W; Wiktor-Jedrzejczak, W; Ratajczak, M Z

    2017-02-01

    As a crucial arm of innate immunity, the complement cascade (ComC) is involved both in mobilization of normal hematopoietic stem/progenitor cells (HSPCs) from bone marrow (BM) into peripheral blood and in their homing to BM. Despite the fact that ComC cleavage fragments alone do not chemoattract normal HSPCs, we found that leukemia cell lines as well as clonogenic blasts from chronic myeloid leukemia and acute myeloid leukemia patients respond robustly to C3 and C5 cleavage fragments by chemotaxis and increased adhesion. This finding was supported by the detection of C3a and C5a receptors in cells from human malignant hematopoietic cell lines and patient blasts at the mRNA (reverse transcriptase-polymerase chain reaction) and protein level (fluorescence-activated cell sorting), and by the demonstration that these receptors respond to stimulation by C3a and C5a by phosphorylation of p42/44 and p38 mitogen-activated protein kinases (MAPK), and protein kinase B (PKB/AKT). We also found that inducible heme oxygenase 1 (HO-1) is a negative regulator of ComC-mediated trafficking of leukemic cells, and that stimulation of leukemic cells by C3 or C5 cleavage fragments activates p38 MAPK, which downregulates HO-1 expression, rendering cells more mobile. We conclude that activation of the ComC in leukemia/lymphoma patients (for example, as a result of accompanying infections) enhances the motility of malignant cells and contributes to their spread in a p38 MAPK-HO-1-dependent manner. Therefore, inhibition of p38 MAPK or upregulation of HO-1 by small-molecule modulators would have a beneficial effect on ameliorating cell migration-mediated expansion of leukemia/lymphoma cells when the ComC becomes activated.

  14. Activation of the complement cascade enhances motility of leukemic cells by downregulating expression of HO-1

    Abdelbaset-Ismail, A; Borkowska-Rzeszotek, S; Kubis, E; Bujko, K; Brzeźniakiewicz-Janus, K; Bolkun, L; Kloczko, J; Moniuszko, M; Basak, G W; Wiktor-Jedrzejczak, W; Ratajczak, M Z

    2017-01-01

    As a crucial arm of innate immunity, the complement cascade (ComC) is involved both in mobilization of normal hematopoietic stem/progenitor cells (HSPCs) from bone marrow (BM) into peripheral blood and in their homing to BM. Despite the fact that ComC cleavage fragments alone do not chemoattract normal HSPCs, we found that leukemia cell lines as well as clonogenic blasts from chronic myeloid leukemia and acute myeloid leukemia patients respond robustly to C3 and C5 cleavage fragments by chemotaxis and increased adhesion. This finding was supported by the detection of C3a and C5a receptors in cells from human malignant hematopoietic cell lines and patient blasts at the mRNA (reverse transcriptase-polymerase chain reaction) and protein level (fluorescence-activated cell sorting), and by the demonstration that these receptors respond to stimulation by C3a and C5a by phosphorylation of p42/44 and p38 mitogen-activated protein kinases (MAPK), and protein kinase B (PKB/AKT). We also found that inducible heme oxygenase 1 (HO-1) is a negative regulator of ComC-mediated trafficking of leukemic cells, and that stimulation of leukemic cells by C3 or C5 cleavage fragments activates p38 MAPK, which downregulates HO-1 expression, rendering cells more mobile. We conclude that activation of the ComC in leukemia/lymphoma patients (for example, as a result of accompanying infections) enhances the motility of malignant cells and contributes to their spread in a p38 MAPK–HO-1-dependent manner. Therefore, inhibition of p38 MAPK or upregulation of HO-1 by small-molecule modulators would have a beneficial effect on ameliorating cell migration-mediated expansion of leukemia/lymphoma cells when the ComC becomes activated. PMID:27451975

  15. Overexpressed human heme Oxygenase-1 decreases adipogenesis in pigs and porcine adipose-derived stem cells.

    Park, Eun Jung; Koo, Ok Jae; Lee, Byeong Chun

    2015-11-27

    Adipose-derived mesenchymal stem cells (ADSC) are multipotent, which means they are able to differentiate into several lineages in vivo and in vitro under proper conditions. This indicates it is possible to determine the direction of differentiation of ADSC by controlling the microenvironment. Heme oxygenase 1 (HO-1), a type of antioxidant enzyme, attenuates adipogenicity and obesity. We produced transgenic pigs overexpressing human HO-1 (hHO-1-Tg), and found that these animals have little fatty tissue when autopsied. To determine whether overexpressed human HO-1 suppresses adipogenesis in pigs, we analyzed body weight increases of hHO-1-Tg pigs and wild type (WT) pigs of the same strain, and induced adipogenic differentiation of ADSC derived from WT and hHO-1-Tg pigs. The hHO-1-Tg pigs had lower body weights than WT pigs from 16 weeks of age until they died. In addition, hHO-1-Tg ADSC showed reduced adipogenic differentiation and expression of adipogenic molecular markers such as PPARγ and C/EBPα compared to WT ADSC. These results suggest that HO-1 overexpression reduces adipogenesis both in vivo and in vitro, which could support identification of therapeutic targets of obesity and related metabolic diseases. Copyright © 2015 Elsevier Inc. All rights reserved.

  16. Effects of combined mesenchymal stem cells and heme oxygenase-1 therapy on cardiac performance.

    Zeng, Bin; Chen, Honglei; Zhu, Chengang; Ren, Xiaofeng; Lin, Guosheng; Cao, Feng

    2008-10-01

    Bone marrow mesenchymal stem cells (MSCs) have the potential to repair the infarcted myocardium and improve cardiac function. However, this approach is limited by its poor viability after transplantation, and controversy still exists over the mechanism by which MSCs contribute to the tissue repair. The human heme oxygenase-1 (hHO-1) was transfected into cultured MSCs using an adenoviral vector. 1 x 10(6) Ad-hHO-1-transfected MSCs (HO-1-MSCs) or Ad-Null-transfected MSCs (Null-MSCs) or PBS only (PBS group) were injected intramyocardially into rat hearts 1h after myocardial infarction. HO-1-MSCs survived in the infarcted myocardium, and expressed hHO-1 mRNA. The expression of basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) was significantly enhanced in HO-1-MSCs-treated hearts. At the same time, there were significant reduction of TNF-alpha, IL-1-beta and IL-6 mRNA, and marked increase of IL-10 mRNA in HO-1-MSCs-treated hearts. Moreover, a further downregulation of proapoptotic protein, Bax, and a marked increase in microvessel density were observed in HO-1-MSCs-treated hearts. The infarct size and cardiac performance were also significantly improved in HO-1-MSCs-treated hearts. The combined approach improves MSCs survival and is superior to MSCs injection alone.

  17. Hormonal fluctuations during the estrous cycle modulate Heme Oxygenase-1 expression in the uterus

    Maria Laura Zenclussen

    2014-03-01

    Full Text Available Deletion of the Heme Oxygenase-1 (Hmox1 locus in mice results in intrauterine lethality. The expression of the heme catabolyzing enzyme encoded by this gene, namely HO 1, is required to successfully support reproductive events. We have previously observed that HO-1 acts at several key events in reproduction ensuring pregnancy. HO-1 defines ovulation, positively influences implantation and placentation and ensures fetal growth and survival. Here, we embarked on a study aimed to determine whether hormonal changes during the estrous cycle in the mouse define HO-1 expression, thus influencing receptivity. We analyzed the serum levels of progesterone and estrogen by ELISA and HO-1 mRNA expression in uterus by real time RT-PCR at the metestrus, proestrus, estrus and diestrus phases of the estrous cycle. Further, we studied the HO-1 protein expression by Western Blot upon hormone addition to cultured uterine AN3 cells. We observed that HO-1 variations in uterine tissue correlated to changes in hormonal levels at different phases of the estrus cycle. In vitro, HO-1 protein levels in AN3 cells augmented after the addition of physiological concentrations of progesterone and estradiol, which confirmed our in vivo observations. Our data suggest an important role for hormones in HO-1 regulation in uterus that has a significant impact in receptivity and later on blastocyst implantation.

  18. Transfection of the Human Heme Oxygenase Gene Into Rabbit Coronary Microvessel Endothelial Cells: Protective Effect Against Heme and Hemoglobin Toxicity

    Abraham, N. G.; Lavrovsky, Y.; Schwartzman, M. L.; Stoltz, R. A.; Levere, R. D.; Gerritsen, M. E.

    1995-07-01

    Heme oxygenase (HO) is a stress protein and has been suggested to participate in defense mechanisms against agents that may induce oxidative injury such as metals, endotoxin, heme/hemoglobin, and various cytokines. Overexpression of HO in cells might therefore protect against oxidative stress produced by certain of these agents, specifically heme and hemoglobin, by catalyzing their degradation to bilirubin, which itself has antioxidant properties. We report here the successful in vitro transfection of rabbit coronary microvessel endothelial cells with a functioning gene encoding the human HO enzyme. A plasmid containing the cytomegalovirus promoter and the human HO cDNA complexed to cationic liposomes (Lipofectin) was used to transfect rabbit endothelial cells. Cells transfected with human HO exhibited an ≈3.0-fold increase in enzyme activity and expressed a severalfold induction of human HO mRNA as compared with endogenous rabbit HO mRNA. Transfected and nontransfected cells expressed factor VIII antigen and exhibited similar acetylated low-density lipoprotein uptake (two important features that characterize endothelial cells) with >85% of cells staining positive for each marker. Moreover, cells transfected with the human HO gene acquired substantial resistance to toxicity produced by exposure to recombinant hemoglobin and heme as compared with nontransfected cells. The protective effect of HO overexpression against heme/hemoglobin toxicity in endothelial cells shown in these studies provides direct evidence that the inductive response of human HO to such injurious stimuli represents an important tissue adaptive mechanism for moderating the severity of cell damage produced by these blood components.

  19. Reduction of bilirubin by targeting human heme oxygenase-1 through siRNA.

    Xia, Zhen-Wei; Li, Chun-E; Jin, You-Xin; Shi, Yi; Xu, Li-Qing; Zhong, Wen-Wei; Li, Yun-Zhu; Yu, Shan-Chang; Zhang, Zi-Li

    2007-04-01

    Neonatal hyperbilirubinemia is a common clinical condition caused mainly by the increased production and decreased excretion of bilirubin. Current treatment is aimed at reducing the serum levels of bilirubin. Heme oxygenase-1 (HO-1) is a rate-limiting enzyme that generates bilirubin. In this study we intended to suppress HO-1 using the RNA interference technique. Small interfering RNA (siRNA)-A, -B, and -C were designed based on human HO-1 (hHO-1) mRNA sequences. siRNA was transfected into a human hepatic cell line (HL-7702). hHO-1 transcription and protein levels were then determined. In addition, the inhibitory effect of siRNA on hHO-1 was assessed in cells treated with hemin or transfected with an hHO-1 plasmid. siRNA-C showed the most potent suppressive effect on hHO-1. This inhibition is dose and time dependent. Compared with control, both hemin and hHO-1 plasmids up-regulated hHO-1 expression in HL-7702 cells. However, the up-regulation was significantly attenuated by siRNA-C. Furthermore, the decrease in hHO-1 activity was coincident with the suppression of its transcription. Finally, siRNA-C was shown to reduce hHO-1 enzymatic activity and bilirubin levels. Thus, this study provides a novel therapeutic rationale by blocking bilirubin formation via siRNA for preventing and treating neonatal hyperbilirubinemia and bilirubin encephalopathy at an early clinical stage.

  20. Proteção funcional da enzima heme-oxigenase-1 na lesão renal aguda isquêmica e tóxica Protección funcional de la enzima heme-oxigenasa-1 en la lesión renal aguda isquémica y tóxica Functional protection of heme-oxygenase-1 enzyme in ischemic and toxic acute kidney injury

    Cassiane Dezoti

    2009-01-01

    estudio demuestran que la isquemia y la PmxB inducen AKL por la elevación de los peróxidos urinarios. El inductor de HO-1 atenuó la lesión en ambos modelos por atenuación del mecanismo redox.OBJECTIVE: To investigate the functional protection of heme-oxygenase-1 enzyme (HO-1 when using its inducer (Hemin and inhibitor (zinc protoporphyrin-ZnPP in ischemic and toxic acute kidney injury by Polymixin B in mice. MATERIALS: Adult male Wistar mice divided into 8 groups were used: SHAM (control, Ischemic (Isq, Isq+Hemin (Inducer of H0-1, Isq+ZnPP (inhibitor of H0-1, SALINA (control, Polimyxin B (PmxB, PmxB+Hemin, PmxB+ZnPP. METHOD: Analysis consists of Jaffé (creatinine clearance [crCl] and FOX-2 (urinary peroxides [UP]. RESULTS: Thirty minutes renal ischemia and its treatment with PmxB reduced the crCl and maintained urinary output. Urinary peroxide levels increased in both injuries. The administration of the inducer of H0-1 resulted in improvement in renal function and reduction in the levels of urinary peroxide. CONCLUSIONS: Findings indicated that ischemia and PmxB induce LAR (acute kidney injury [AKI] by elevating the levels of urinary peroxide. The HO-1 inducer ameliorated the injury in both animal models through redox mechanism.

  1. Association of heme oxygenase-1 GT-repeat polymorphism with blood pressure phenotypes and its relevance to future cardiovascular mortality risk: an observation based on arsenic-exposed individuals.

    Wu, Meei-Maan; Chiou, Hung-Yi; Chen, Chi-Ling; Hsu, Ling-I; Lien, Li-Ming; Wang, Chih-Hao; Hsieh, Yi-Chen; Wang, Yuan-Hung; Hsueh, Yu-Mei; Lee, Te-Chang; Cheng, Wen-Fang; Chen, Chien-Jen

    2011-12-01

    Heme oxygenase (HO)-1 is up-regulated as a cellular defense responding to stressful stimuli in experimental studies. A GT-repeat length polymorphism in the HO-1 gene promoter was inversely correlated to HO-1 induction. Here, we reported the association of GT-repeat polymorphism with blood pressure (BP) phenotypes, and their interaction on cardiovascular (CV) mortality risk in arsenic-exposed cohorts. Associations of GT-repeat polymorphism with BP phenotypes were investigated at baseline in a cross-sectional design. Effect of GT-repeat polymorphism on CV mortality was investigated in a longitudinal design stratified by hypertension. GT-repeat variants were grouped by S (accounting for CV covariates. Totally, 894 participants were recruited and analyzed. At baseline, carriers with HO-1 S alleles had lower diastolic BP (L/S genotypes, P = 0.014) and a lower possibility of being hypertensive (L/S genotypes, P = 0.048). After follow-up, HO-1 S allele was significantly associated with a reduced CV risk in hypertensive participants [relative mortality ratio (RMR) 0.27 (CI 0.11, 0.69), P = 0.007] but not in normotensive. Hypertensive participants without carrying the S allele had a 5.23-fold increased risk [RMR 5.23 (CI 1.99, 13.69), P = 0.0008] of CV mortality compared with normotensive carrying the S alleles. HO-1 short GT-repeat polymorphism may play a protective role in BP regulation and CV mortality risk in hypertensive individuals against environmental stressors. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  2. Heme oxygenase-1 deletion affects stress erythropoiesis.

    Yu-An Cao

    Full Text Available Homeostatic erythropoiesis leads to the formation of mature red blood cells under non-stress conditions, and the production of new erythrocytes occurs as the need arises. In response to environmental stimuli, such as bone marrow transplantation, myelosuppression, or anemia, erythroid progenitors proliferate rapidly in a process referred to as stress erythropoiesis. We have previously demonstrated that heme oxygenase-1 (HO-1 deficiency leads to disrupted stress hematopoiesis. Here, we describe the specific effects of HO-1 deficiency on stress erythropoiesis.We used a transplant model to induce stress conditions. In irradiated recipients that received hmox(+/- or hmox(+/+ bone marrow cells, we evaluated (i the erythrocyte parameters in the peripheral blood; (ii the staining intensity of CD71-, Ter119-, and CD49d-specific surface markers during erythroblast differentiation; (iii the patterns of histological iron staining; and (iv the number of Mac-1(+-cells expressing TNF-α. In the spleens of mice that received hmox(+/- cells, we show (i decreases in the proerythroblast, basophilic, and polychromatophilic erythroblast populations; (ii increases in the insoluble iron levels and decreases in the soluble iron levels; (iii increased numbers of Mac-1(+-cells expressing TNF-α; and (iv decreased levels of CD49d expression in the basophilic and polychromatophilic erythroblast populations.As reflected by effects on secreted and cell surface proteins, HO-1 deletion likely affects stress erythropoiesis through the retention of erythroblasts in the erythroblastic islands of the spleen. Thus, HO-1 may serve as a therapeutic target for controlling erythropoiesis, and the dysregulation of HO-1 may be a predisposing condition for hematologic diseases.

  3. HO-1 inhibits IL-13-induced goblet cell hyperplasia associated with CLCA1 suppression in normal human bronchial epithelial cells.

    Mishina, Kei; Shinkai, Masaharu; Shimokawaji, Tadasuke; Nagashima, Akimichi; Hashimoto, Yusuke; Inoue, Yoriko; Inayama, Yoshiaki; Rubin, Bruce K; Ishigatsubo, Yoshiaki; Kaneko, Takeshi

    2015-12-01

    Mucus hypersecretion and goblet cell hyperplasia are common features that characterize asthma. IL-13 increases mucin (MUC) 5AC, the major component of airway mucus, in airway epithelial cells. According to the literature, IL-13 receptor activation leads to STAT6 activation and consequent induction of chloride channel accessory 1 (CLCA1) gene expression, associated with the induction of MUC5AC. Heme oxygenase-1 (HO-1) is an enzyme that catalyzes oxidation of heme to biliverdin, and has anti-inflammatory and anti-oxidant properties. We examined the effects of HO-1 on mucin production and goblet cell hyperplasia induced by IL-13. Moreover, we assessed the cell signaling intermediates that appear to be responsible for mucin production. Normal human bronchial epithelial (NHBE) cells were grown at air liquid interface (ALI) in the presence or absence of IL-13 and hemin, a HO-1 inducer, for 14 days. Protein concentration was analyzed using ELISA, and mRNA expression was examined by real-time PCR. Histochemical analysis was performed using HE staining, andWestern blotting was performed to evaluate signaling transduction pathway. Hemin (4 μM) significantly increased HO-1 protein expression (p b 0.01) and HO-1 mRNA expression (p b 0.001). IL-13 significantly increased goblet cells, MUC5AC protein secretion (p b 0.01) and MUC5AC mRNA (p b 0.001), and these were decreased by hemin by way of HO-1. Tin protoporphyrin (SnPP)-IX, a HO-1 inhibitor, blocked the effect of hemin restoring MUC5AC protein secretion (p b 0.05) and goblet cell hyperplasia. Hemin decreased the expression of CLCA1 mRNA (p b 0.05) and it was reversed by SnPP-IX, but could not suppress IL-13-induced phosphorylation of STAT6 or SAM pointed domain-containing ETS transcription factor (SPDEF) and Forkhead box A2 (FOXA2) mRNA expression. In summary, HO-1 overexpression suppressed IL-13-induced goblet cell hyperplasia and MUC5AC production, and involvement of CLCA1 in the mechanism was suggested.

  4. NRF2 activation is involved in ozonated human serum upregulation of HO-1 in endothelial cells

    Pecorelli, Alessandra; Bocci, Velio; Acquaviva, Alessandra; Belmonte, Giuseppe; Gardi, Concetta; Virgili, Fabio; Ciccoli, Lucia; Valacchi, Giuseppe

    2013-01-01

    During the last decade, it has been shown that the activation of NRF2 and the binding to electrophile-responsive element (EpREs), stimulates the expression of a great number of genes responsible for the synthesis of phase I and phase II proteins, including antioxidants enzymes and heme oxygenase-1 (HO-1). This critical cell response occurs in cardiovascular, degenerative and chronic infective diseases aggravated by a chronic oxidative stress. In our previous reports we have shown that ozonated plasma is able to up-regulate HO-1 expression in endothelial cells. In the present work we investigated a candidate mechanism involved in this process. After treatment with increasing doses of ozonated serum (20, 40 and 80 μg/mL O 3 per mL of serum), a clear dose dependent activation of NRF2 and the subsequent induction of HO-1 and NAD(P)H quinone oxidoreductase 1(NQO1) was observed. This effect was also present when cells were treated with serum and hydrogen peroxide (H 2 O 2 ) or serum and 4-hydroxynonenal (4HNE). Moreover, the treatment with ozonated serum was associated with a dose-dependent activation of extracellular-signal-regulated kinases (ERK1/2) and p38 MAP kinases (p38), not directly involved in NRF2 activation. These data, provide a new insight on the mechanism responsible for the induction of HO-1 expression by ozonated serum in the endothelium, and have a practical importance as an expedient approach to the treatment of patients with both effective orthodox drugs and ozonated autohemotherapy, targeted to the restoration of redox homeostasis. - Highlights: ► Endothelial HO1 is upregulated by ozonated plasma ► This activation is induced by NRF2 and it is ERK independent. ► 4HNE and H 2 O 2 are the main molecules involved in this process. ► Ozonated plasma induced a hormetic effect ► Combination of orthodox medicine and ozonated plasma can be a useful treatment

  5. NRF2 activation is involved in ozonated human serum upregulation of HO-1 in endothelial cells

    Pecorelli, Alessandra [Department of Molecular and Developmental Medicine, University of Siena (Italy); Child Neuropsychiatry Unit, University Hospital, AOUS, Siena (Italy); Bocci, Velio [Department of Physiology, University of Siena (Italy); Acquaviva, Alessandra [Department of Molecular and Developmental Medicine, University of Siena (Italy); Belmonte, Giuseppe [Department of Biomedical Sciences, University of Siena (Italy); Gardi, Concetta [Department of Molecular and Developmental Medicine, University of Siena (Italy); Virgili, Fabio [INRAN, Rome (Italy); Ciccoli, Lucia [Department of Molecular and Developmental Medicine, University of Siena (Italy); Valacchi, Giuseppe, E-mail: giuseppe.valacchi@unife.it [Department of Life Sciences and Biotechnology, University of Ferrara (Italy); Department of Food and Nutrition, Kyung Hee University, Seoul (Korea, Republic of)

    2013-02-15

    During the last decade, it has been shown that the activation of NRF2 and the binding to electrophile-responsive element (EpREs), stimulates the expression of a great number of genes responsible for the synthesis of phase I and phase II proteins, including antioxidants enzymes and heme oxygenase-1 (HO-1). This critical cell response occurs in cardiovascular, degenerative and chronic infective diseases aggravated by a chronic oxidative stress. In our previous reports we have shown that ozonated plasma is able to up-regulate HO-1 expression in endothelial cells. In the present work we investigated a candidate mechanism involved in this process. After treatment with increasing doses of ozonated serum (20, 40 and 80 μg/mL O{sub 3} per mL of serum), a clear dose dependent activation of NRF2 and the subsequent induction of HO-1 and NAD(P)H quinone oxidoreductase 1(NQO1) was observed. This effect was also present when cells were treated with serum and hydrogen peroxide (H{sub 2}O{sub 2}) or serum and 4-hydroxynonenal (4HNE). Moreover, the treatment with ozonated serum was associated with a dose-dependent activation of extracellular-signal-regulated kinases (ERK1/2) and p38 MAP kinases (p38), not directly involved in NRF2 activation. These data, provide a new insight on the mechanism responsible for the induction of HO-1 expression by ozonated serum in the endothelium, and have a practical importance as an expedient approach to the treatment of patients with both effective orthodox drugs and ozonated autohemotherapy, targeted to the restoration of redox homeostasis. - Highlights: ► Endothelial HO1 is upregulated by ozonated plasma ► This activation is induced by NRF2 and it is ERK independent. ► 4HNE and H{sub 2}O{sub 2} are the main molecules involved in this process. ► Ozonated plasma induced a hormetic effect ► Combination of orthodox medicine and ozonated plasma can be a useful treatment.

  6. Antihepatocarcinoma Effect of Portulaca oleracea L. in Mice by PI3K/Akt/mTOR and Nrf2/HO-1/NF-κB Pathway

    Guoyin, Zheng; Hao, Peng; Min, Li; Wei, Gu; Zhe, Chen

    2017-01-01

    The purpose of the present study was to evaluate the pharmacological effects of Portulaca oleracea L. (Purslane) (PL) on N-nitrosodiethylamine- (NDEA-) induced hepatocellular carcinomas (HCC) and explore its potential mechanism. Mice were randomly assigned to four groups: control group, NDEA group, NDEA + Purslane (100 mg/kg) group, and NDEA + Purslane (200 mg/kg) group. The animal of each group was given NDEA (100 ppm) in drinking water. 1 h later, Purslane dissolved in PBS was intragastrically administered for continuous seven days. The results showed that Purslane reduced the activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in liver and serum. Purslane also reduced the contents of interleukin-6 (IL-6), IL-1β, tumor necrosis factor-α (TNF-α), and methane dicarboxylic aldehyde (MDA) and restored the activity of superoxygen dehydrogenises (SOD) in serum. Purslane could obviously attenuate the hepatic pathological alteration. Furthermore, treatment with Purslane effectively inhibited the phosphorylations of phosphatidylinositol 3 kinase (PI3K), protein kinase B (Akt), mammalian target of rapamycin (mTOR), nuclear factor-kappa B (NF-κB), and inhibitor of NF-κBα (IκBα) and upregulated the expressions of NF-E2-related factor 2 (Nrf2) and heme oxygenase- (HO-) 1. In conclusion, our research suggested that Purslane exhibited protective effects on NDEA-induced hepatocellular carcinomas by anti-inflammatory and antioxidative properties via the PI3K/Akt/mTOR and Nrf2/HO-1/NF-κB pathway. PMID:28659990

  7. Lycopene ameliorates atrazine-induced oxidative damage in adrenal cortex of male rats by activation of the Nrf2/HO-1 pathway.

    Abass, Marwa Ahmed; Elkhateeb, Shereen Ahmed; Abd El-Baset, Samia Adel; Kattaia, Asmaa Alhosiny; Mohamed, Eman Mosallam; Atteia, Hebatallah Husseini

    2016-08-01

    Atrazine (ATZ) is one of the most commonly used herbicides contaminating plants, soil and water resources. Several strategies have been used to counteract ATZ toxicity. Here, we tested the hypothesis that lycopene could ameliorate ATZ-induced toxicity in the adrenal cortex. For this purpose, 35 adult male albino rats were randomized into five equal groups: untreated control, vehicle control (received 0.5 mL corn oil/day), lycopene (treated with lycopene dissolved in 0.5 mL corn oil, 10 mg/kg b.w./day), ATZ (received ATZ dissolved in 0.5 mL corn oil 300 mg/kg b.w./day), and ATZ + lycopene (treated with ATZ and lycopene at the same previously mentioned doses). All treatments were given by oral gavage for 4 weeks. We found that ATZ exposure significantly increased relative adrenal weight, plasma ACTH levels, and adrenal oxidative stress as manifested by elevated malondialdehyde levels, decreased reduced glutathione content and depressed antioxidant enzyme activities in adrenal cortex tissues with respect to control groups. Furthermore, the transcription of adrenal cortex nuclear factor erythroid 2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), nuclear factor kappa B, and caspase-3 genes was increased significantly compared with the control groups. This was accompanied with DNA fragmentation and structural and ultrastructural changes in zona glomerulosa and zona fasiculata of the adrenal cortex. Notably, all these changes were partially ameliorated in rats treated concomitantly with ATZ and lycopene. Our results showed that lycopene exerts protective effects against ATZ-induced toxicity in rat adrenal cortex. These effects may be attributed to the antioxidative property of lycopene and its ability to activate the Nrf2/HO-1 pathway.

  8. Preconditioning with Gua Lou Gui Zhi decoction enhances H2O2-induced Nrf2/HO-1 activation in PC12 cells

    MAO, JINGJIE; LI, ZUANFANG; LIN, RUHUI; ZHU, XIAOQIN; LIN, JIUMAO; PENG, JUN; CHEN, LIDIAN

    2015-01-01

    Spasticity is common in various central neurological conditions, including after a stroke. Such spasticity may cause additional problems, and often becomes a primary concern for afflicted individuals. A number of studies have identified nuclear factor (erythroid-derived 2)-like 2 (Nrf2) as a key regulator in the adaptive survival response to oxidative stress. Elevated expression of Nrf2, combined with heme oxygenase 1 (HO-1) resistance, in the central nervous system is known to elicit key internal and external oxidation protection. Gua Lou Gui Zhi decoction (GLGZD) is a popular traditional Chinese formula with a long history of clinical use in China for the treatment of muscular spasticity following a stroke, epilepsy or a spinal cord injury. However, the mechanism underlying the efficacy of the medicine remains unclear. In the present study, the antioxidative effects of GLGZD were evaluated and the underlying molecular mechanisms were investigated, using hydrogen peroxide (H2O2)-induced rat pheochromocytoma cells (PC12 cells) as an in vitro oxidative stress model of neural cells. Upon application of different concentrations of GLGZD, a 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide (MTT) assay and ATP measurement were conducted to assess the impact on PC12 cell proliferation. In addition, inverted microscopy observations, and the MTT and ATP assessments, revealed that GLGZD attenuated H2O2-induced oxidative damage and signaling repression in PC12 cells. Furthermore, the mRNA and protein expression levels of Nrf2 and HO-1, which are associated with oxidative stress, were analyzed using reverse transcription quantitative polymerase chain reaction (PCR) and confocal microscopy. Confocal microscopy observations, as well as the quantitative PCR assay, revealed that GLGZD exerted a neuroprotective function against H2O2-induced oxidative damage in PC12 cells. Therefore, the results demonstrated that GLGZD protected PC12 cells injured by H2O2, which may be

  9. Effect of heme oxygenase-1 on radiation-induced skin injury

    Song Chuanjun; Meng Xingjun; Xie Ling; Chen Qing; Zhou Jundong; Zhang Shuyu; Wu Jinchang

    2012-01-01

    Objective: To investigate the effect of heme oxygenase-1 (HO-1) on the acute radiation-induced skin injury by gene transfer. Methods: Thirty-three male SD rats were randomly divided into three groups as PBS-injected group, Ad-EGFP-injected group and Ad-HO-1-injected group (n=11). In each group, three rats were used for determining the expression of target gene and the other rats were irradiated on the buttock skin with 40 Gy electron beam generated by a linear accelerator. Immediately after irradiation, rats were administered with a subcutaneous injection of PBS, Ad-EGFP or Ad-HO-1, respectively. Subsequently, the skin reactions were measured twice a week using the semi-quantitative skin injury scale. Results: The strong positive expression of HO-1 was observed in subcutaneous dermal tissue after injection of Ad-HO-1. Compared to the PBS-injected group or the Ad-EGFP-injected group, a significant mitigation of skin injury was observed in Ad-HO-1-injected mice 14 d after irradiation (q=0.000-0.030, P<0.05). Conclusions: HO-1 could significantly mitigate radiation-induced acute skin injury and Ad-HO-1 could be used to treat radiation-induced skin injury. (authors)

  10. Myeloid Heme Oxygenase-1 Regulates the Acute Inflammatory Response to Zymosan in the Mouse Air Pouch

    Rita Brines

    2018-01-01

    Full Text Available Heme oxygenase-1 (HO-1 is induced by many stimuli to modulate the activation and function of different cell types during innate immune responses. Although HO-1 has shown anti-inflammatory effects in different systems, there are few data on the contribution of myeloid HO-1 and its role in inflammatory processes is not well understood. To address this point, we have used HO-1M-KO mice with myeloid-restricted deletion of HO-1 to specifically investigate its influence on the acute inflammatory response to zymosan in vivo. In the mouse air pouch model, we have shown an exacerbated inflammation in HO-1M-KO mice with increased neutrophil infiltration accompanied by high levels of inflammatory mediators such as interleukin-1β, tumor necrosis factor-α, and prostaglandin E2. The expression of the degradative enzyme matrix metalloproteinase-3 (MMP-3 was also enhanced. In addition, we observed higher levels of serum MMP-3 in HO-1M-KO mice compared with control mice, suggesting the presence of systemic inflammation. Altogether, these findings demonstrate that myeloid HO-1 plays an anti-inflammatory role in the acute response to zymosan in vivo and suggest the interest of this target to regulate inflammatory processes.

  11. Anti-inflammatory effect of transduced PEP-1-heme oxygenase-1 in Raw 264.7 cells and a mouse edema model

    Kwon, Soon Won; Sohn, Eun Jeong; Kim, Dae Won; Jeong, Hoon Jae; Kim, Mi Jin; Ahn, Eun Hee; Kim, Young Nam; Dutta, Suman; Kim, Duk-Soo; Park, Jinseu; Eum, Won Sik; Hwang, Hyun Sook; Choi, Soo Young

    2011-01-01

    Highlights: → Recombinant PEP-1 heme oxygenase-1 expression vector was constructed and overexpressed. → We investigated transduction efficiency of PEP-1-HO-1 protein in Raw 264.7 cells. → PEP-1-HO-1 was efficiently transduced into Raw 264.7 cells in a dose and time dependent manner. → PEP-1-HO-1 exerted anti-inflammatory activity in Raw 264.7 cells and in a mice edema model. → PEP-1-HO-1 could be used as a therapeutic drug against inflammatory diseases. -- Abstract: Heme oxygenase-1 (HO-1), which catalyzes the degradation of free heme to biliverdin, carbon monoxide (CO), and free iron (Fe 2+ ), is up-regulated by several cellular stress and cell injuries, including inflammation, ischemia and hypoxia. In this study, we examined whether fusion of HO-1 with PEP-1, a protein transduction domain that is able to deliver exogenous molecules to living cells or tissues, would facilitate HO-1 delivery to target cells and tissues, and thereby effectively exert a therapeutically useful response against inflammation. Western blot analysis demonstrated that PEP-1-HO-1 fusion proteins were transduced into Raw 264.7 cells in time- and dose-dependent manners, and were stably maintained in the cells for about 60 h. In addition, fluorescence analysis revealed that only PEP-1-HO-1 fusion proteins were significantly transduced into the cytoplasm of cells, while HO-1 proteins failed to be transduced. In lipopolysaccharide (LPS)-stimulated Raw 264.7 cells and 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced mouse edema model, transduced PEP-1-HO-1 fusion proteins effectively inhibited the overexpression of pro-inflammatory mediators and cytokines. Also, histological analysis demonstrated that PEP-1-HO-1 remarkably suppressed ear edema. The results suggest that the PEP-1-HO-1 fusion protein can be used as a therapeutic molecule against reactive oxygen species-related inflammatory diseases.

  12. Involvement of heme oxygenase-1 expression in neuroprotection by piceatannol, a natural analog and a metabolite of resveratrol, against glutamate-mediated oxidative injury in HT22 neuronal cells.

    Son, Yong; Byun, Seung Jae; Pae, Hyun-Ock

    2013-08-01

    Neuronal cell death caused by oxidative stress is common in a variety of neural diseases and can be investigated in detail in cultured HT22 neuronal cells, where the amino acid glutamate at high concentrations causes glutathione depletion by inhibition of the glutamate/cystine antiporter system, intracellular accumulation of reactive oxygen species (ROS) and eventually oxidative stress-induced neuronal cell death. Using this paradigm, we have previously reported that resveratrol (3,5,4'-trans-trihydroxystilbene) protects HT22 neuronal cells from glutamate-induced oxidative stress by inducing heme oxygenase (HO)-1 expression. Piceatannol (3,5,4',3'-trans-trihydroxystilbene), which is a hydroxylated resveratrol analog and one of the resveratrol metabolites, is estimated to exert neuroprotective effect similar to that of resveratrol. The aim of this study, thus, is to determine whether piceatannol, similarly to resveratrol, would protect HT22 neuronal cells from glutamate-induced oxidative stress. Glutamate at high concentrations induced neuronal cell death and ROS formation. Piceatannol reduced glutamate-induced cell death and ROS formation. The observed cytoprotective effect was much higher when HT22 neuronal cells were pretreated with piceatannol for 6 or 12 h prior to glutamate treatment than when pretreated for 0.5 h. Piceatannol also increased HO-1 expression and HO activity via its activation of nuclear factor-E2-related factor 2 (Nrf2). Interestingly, neuroprotective effect of piceatannol was partly (but not completely) abolished by either down-regulation of HO-1 expression or blockage of HO-1 activity. Taken together, our results suggest that piceatannol, similar to resveratrol, is capable of protecting HT22 neuronal cells against glutamate-induced cell death, at least in part, by inducing Nrf2-dependent HO-1 expression.

  13. The HO-1/CO system regulates mitochondrial-capillary density relationships in human skeletal muscle.

    Pecorella, Shelly R H; Potter, Jennifer V F; Cherry, Anne D; Peacher, Dionne F; Welty-Wolf, Karen E; Moon, Richard E; Piantadosi, Claude A; Suliman, Hagir B

    2015-10-15

    The heme oxygenase-1 (HO-1)/carbon monoxide (CO) system induces mitochondrial biogenesis, but its biological impact in human skeletal muscle is uncertain. The enzyme system generates CO, which stimulates mitochondrial proliferation in normal muscle. Here we examined whether CO breathing can be used to produce a coordinated metabolic and vascular response in human skeletal muscle. In 19 healthy subjects, we performed vastus lateralis muscle biopsies and tested one-legged maximal O2 uptake (V̇o2max) before and after breathing air or CO (200 ppm) for 1 h daily for 5 days. In response to CO, there was robust HO-1 induction along with increased mRNA levels for nuclear-encoded mitochondrial transcription factor A (Tfam), cytochrome c, cytochrome oxidase subunit IV (COX IV), and mitochondrial-encoded COX I and NADH dehydrogenase subunit 1 (NDI). CO breathing did not increase V̇o2max (1.96 ± 0.51 pre-CO, 1.87 ± 0.50 post-CO l/min; P = not significant) but did increase muscle citrate synthase, mitochondrial density (139.0 ± 34.9 pre-CO, 219.0 ± 36.2 post-CO; no. of mitochondrial profiles/field), myoglobin content and glucose transporter (GLUT4) protein level and led to GLUT4 localization to the myocyte membrane, all consistent with expansion of the tissue O2 transport system. These responses were attended by increased cluster of differentiation 31 (CD31)-positive muscle capillaries (1.78 ± 0.16 pre-CO, 2.37 ± 0.59 post-CO; capillaries/muscle fiber), implying the enrichment of microvascular O2 reserve. The findings support that induction of the HO-1/CO system by CO not only improves muscle mitochondrial density, but regulates myoglobin content, GLUT4 localization, and capillarity in accordance with current concepts of skeletal muscle plasticity. Copyright © 2015 the American Physiological Society.

  14. Erythropoietin Attenuates Pulmonary Vascular Remodeling in Experimental Pulmonary Arterial Hypertension through Interplay between Endothelial Progenitor Cells and Heme Oxygenase

    van Loon, Rosa Laura E; Bartelds, Beatrijs; Wagener, Frank A D T G; Affara, Nada; Mohaupt, Saffloer; Wijnberg, Hans; Pennings, Sebastiaan W C; Takens, Janny; Berger, Rolf M F

    2015-01-01

    BACKGROUND: Pulmonary arterial hypertension (PAH) is a pulmonary vascular disease with a high mortality, characterized by typical angio-proliferative lesions. Erythropoietin (EPO) attenuates pulmonary vascular remodeling in PAH. We postulated that EPO acts through mobilization of endothelial progenitor cells (EPCs) and activation of the cytoprotective enzyme heme oxygenase-1 (HO-1). METHODS: Rats with flow-associated PAH, resembling pediatric PAH, were treated with HO-1 inducer EPO in the pre...

  15. Erythropoietin Attenuates Pulmonary Vascular Remodeling in Experimental Pulmonary Arterial Hypertension through Interplay between Endothelial Progenitor Cells and Heme Oxygenase

    van Loon, Rosa Laura E.; Bartelds, Beatrijs; Wagener, Frank A. D. T. G.; Affara, Nada; Mohaupt, Saffloer; Wijnberg, Hans; Pennings, Sebastiaan W. C.; Takens, Janny; Berger, Rolf M. F.

    2015-01-01

    Background Pulmonary arterial hypertension (PAH) is a pulmonary vascular disease with a high mortality, characterized by typical angio-proliferative lesions. Erythropoietin (EPO) attenuates pulmonary vascular remodeling in PAH. We postulated that EPO acts through mobilization of endothelial progenitor cells (EPCs) and activation of the cytoprotective enzyme heme oxygenase-1 (HO-1). Methods Rats with flow-associated PAH, resembling pediatric PAH, were treated with HO-1 inducer EPO i...

  16. Heme oxygenase-1 prevents cardiac dysfunction in streptozotocin-diabetic mice by reducing inflammation, oxidative stress, apoptosis and enhancing autophagy.

    Yanli Zhao

    Full Text Available Heme oxygenase-1 (HO-1 has been implicated in cardiac dysfunction, oxidative stress, inflammation, apoptosis and autophagy associated with heart failure, and atherosclerosis, in addition to its recognized role in metabolic syndrome and diabetes. Numerous studies have presented contradictory findings about the role of HO-1 in diabetic cardiomyopathy (DCM. In this study, we explored the role of HO-1 in myocardial dysfunction, myofibril structure, oxidative stress, inflammation, apoptosis and autophagy using a streptozotocin (STZ-induced diabetes model in mice systemically overexpressing HO-1 (Tg-HO-1 or mutant HO-1 (Tg-mutHO-1. The diabetic mouse model was induced by multiple peritoneal injections of STZ. Two months after injection, left ventricular (LV function was measured by echocardiography. In addition, molecular biomarkers related to oxidative stress, inflammation, apoptosis and autophagy were evaluated using classical molecular biological/biochemical techniques. Mice with DCM exhibited severe LV dysfunction, myofibril structure disarray, aberrant cardiac oxidative stress, inflammation, apoptosis, autophagy and increased levels of HO-1. In addition, we determined that systemic overexpression of HO-1 ameliorated left ventricular dysfunction, myofibril structure disarray, oxidative stress, inflammation, apoptosis and autophagy in DCM mice. Furthermore, serine/threonine-specific protein kinase (Akt and AMP-activated protein kinase (AMPK phosphorylation is normally inhibited in DCM, but overexpression of the HO-1 gene restored the phosphorylation of these kinases to normal levels. In contrast, the functions of HO-1 in DCM were significantly reversed by overexpression of mutant HO-1. This study underlines the unique roles of HO-1, including the inhibition of oxidative stress, inflammation and apoptosis and the enhancement of autophagy, in the pathogenesis of DCM.

  17. The haptoglobin-CD163-heme oxygenase-1 pathway for hemoglobin scavenging

    Thomsen, Jens Haugbølle; Etzerodt, Anders; Svendsen, Pia

    2013-01-01

    The haptoglobin- (Hp-) CD163-heme oxygenase-1 (HO-1) pathway is an efficient captor-receptor-enzyme system to circumvent the hemoglobin (Hb)/heme-induced toxicity during physiological and pathological hemolyses. In this pathway, Hb tightly binds to Hp leading to CD163-mediated uptake of the complex...

  18. Dry powder inhalation of hemin to induce heme oxygenase expression in the lung

    Zijlstra, G.S.; Brandsma, C.; Harpe, M.F.H.; Van Dam, G.M.; Slebos, D.J.; Kerstjens, H.A.M.; de Boer, Anne; Frijlink, H.W.

    2007-01-01

    The purpose of this study was to formulate hemin as a powder for inhalation and to show proof of concept of heme oxygenase 1 (HO-1) expression in the lungs of mice by inhalation of hemin. Hemin was spray dried from a neutralized sodium hydroxide solution. The particle size distribution of the powder

  19. Human heme oxygenase oxidation of 5- and 15-phenylhemes.

    Wang, Jinling; Niemevz, Fernando; Lad, Latesh; Huang, Liusheng; Alvarez, Diego E; Buldain, Graciela; Poulos, Thomas L; de Montellano, Paul R Ortiz

    2004-10-08

    Human heme oxygenase-1 (hHO-1) catalyzes the O2-dependent oxidation of heme to biliverdin, CO, and free iron. Previous work indicated that electrophilic addition of the terminal oxygen of the ferric hydroperoxo complex to the alpha-meso-carbon gives 5-hydroxyheme. Earlier efforts to block this reaction with a 5-methyl substituent failed, as the reaction still gave biliverdin IXalpha. Surprisingly, a 15-methyl substituent caused exclusive cleavage at the gamma-meso-rather than at the normal, unsubstituted alpha-meso-carbon. No CO was formed in these reactions, but the fragment cleaved from the porphyrin eluded identification. We report here that hHO-1 cleaves 5-phenylheme to biliverdin IXalpha and oxidizes 15-phenylheme at the alpha-meso position to give 10-phenylbiliverdin IXalpha. The fragment extruded in the oxidation of 5-phenylheme is benzoic acid, one oxygen of which comes from O2 and the other from water. The 2.29- and 2.11-A crystal structures of the hHO-1 complexes with 1- and 15-phenylheme, respectively, show clear electron density for both the 5- and 15-phenyl rings in both molecules of the asymmetric unit. The overall structure of 15-phenylheme-hHO-1 is similar to that of heme-hHO-1 except for small changes in distal residues 141-150 and in the proximal Lys18 and Lys22. In the 5-phenylheme-hHO-1 structure, the phenyl-substituted heme occupies the same position as heme in the heme-HO-1 complex but the 5-phenyl substituent disrupts the rigid hydrophobic wall of residues Met34, Phe214, and residues 26-42 near the alpha-meso carbon. The results provide independent support for an electrophilic oxidation mechanism and support a role for stereochemical control of the reaction regiospecificity.

  20. A Heme Oxygenase-1 Transducer Model of Degenerative and Developmental Brain Disorders

    Hyman M. Schipper

    2015-03-01

    Full Text Available Heme oxygenase-1 (HO-1 is a 32 kDa protein which catalyzes the breakdown of heme to free iron, carbon monoxide and biliverdin. The Hmox1 promoter contains numerous consensus sequences that render the gene exquisitely sensitive to induction by diverse pro-oxidant and inflammatory stimuli. In “stressed” astroglia, HO-1 hyperactivity promotes mitochondrial iron sequestration and macroautophagy and may thereby contribute to the pathological iron deposition and bioenergetic failure documented in Alzheimer disease, Parkinson disease and certain neurodevelopmental conditions. Glial HO-1 expression may also impact neuroplasticity and cell survival by modulating brain sterol metabolism and the proteasomal degradation of neurotoxic proteins. The glial HO-1 response may represent a pivotal transducer of noxious environmental and endogenous stressors into patterns of neural damage and repair characteristic of many human degenerative and developmental CNS disorders.

  1. Selenolate complexes of CYP101 and the heme-bound hHO-1/H25A proximal cavity mutant.

    Jiang, Yongying; Ortiz de Montellano, Paul R

    2008-05-05

    Thiolate and selenolate complexes of CYP101 (P450cam) and the H25A proximal cavity mutant of heme-bound human heme oxygenase-1 (hHO-1) have been examined by UV-vis spectroscopy. Both thiolate and selenolate ligands bound to the heme distal side in CYP101 and gave rise to characteristic hyperporphyrin spectra. Thiolate ligands also bound to the proximal side of the heme in the cavity created by the H25A mutation in hHO-1, giving a Soret absorption similar to that of the H25C hHO-1 mutant. Selenolate ligands also bound to this cavity mutant under anaerobic conditions but reduced the heme iron to the ferrous state, as shown by the formation of a ferrous CO complex. Under aerobic conditions, the selenolate ligand but not the thiolate ligand was rapidly oxidized. These results indicate that selenocysteine-coordinated heme proteins will not be stable species in the absence of a redox potential stabilizing effect.

  2. Identification of Heme Oxygenase-1 as a Novel Predictor of Hematopoietic Stem Cell Transplantation Outcomes in Acute Leukemia

    Yinghao Lu

    2016-09-01

    Full Text Available Objective: The main aim of this study was to determine the correlation between clinical outcome and heme oxygenase-1 (HO-1 expression before and after hematopoietic stem cell transplantation (HSCT in acute leukemia. Methods: HO-1 mRNA levels in 83 patients were measured using qRT-PCR. In a comparative analysis of HO-1 levels in relation to different post-transplant outcomes, the HO-1 threshold, determined via the receiver operating characteristic (ROC curve, was effectively used to predict clinical relapse and acute graft-versus-host disease (aGVHD. The correlations among clinical relapse, aGVHD and HO-1 expression were analyzed based on this threshold. Results: Leukemia risk stratification and relative expression of HO-1 before pretreatment had significant effects on clinical relapse. Leukemia risk stratification, relative expression of HO-1 after HSCT and the interval from diagnosis to transplantation had a significant influence on aGVHD. Both relapse and aGVHD appeared to be associated with relative HO-1 expression. The relative expression rate of HO-1 was 1.131-1.186 before pretreatment, and strongly associated with post-transplantation relapse. The relative expression rate of HO-1 was 1.102-1.144 after transplantation, and closely related to aGVHD. ROC curve analysis revealed high specificity and sensitivity of HO-1 expression in predicting relapse and aGVHD after allo-HSCT. Conclusions: HO-1 expression can be effectively used as a predictor of relapse as well as a diagnostic factor of aGVHD after transplantation for allo-HSCT patients with acute leukemia.

  3. The binding sites on human heme oxygenase-1 for cytochrome p450 reductase and biliverdin reductase.

    Wang, Jinling; de Montellano, Paul R Ortiz

    2003-05-30

    Human heme oxygenase-1 (hHO-1) catalyzes the NADPH-cytochrome P450 reductase-dependent oxidation of heme to biliverdin, CO, and free iron. The biliverdin is subsequently reduced to bilirubin by biliverdin reductase. Earlier kinetic studies suggested that biliverdin reductase facilitates the release of biliverdin from hHO-1 (Liu, Y., and Ortiz de Montellano, P. R. (2000) J. Biol. Chem. 275, 5297-5307). We have investigated the binding of P450 reductase and biliverdin reductase to truncated, soluble hHO-1 by fluorescence resonance energy transfer and site-specific mutagenesis. P450 reductase and biliverdin reductase bind to truncated hHO-1 with Kd = 0.4 +/- 0.1 and 0.2 +/- 0.1 microm, respectively. FRET experiments indicate that biliverdin reductase and P450 reductase compete for binding to truncated hHO-1. Mutation of surface ionic residues shows that hHO-1 residues Lys18, Lys22, Lys179, Arg183, Arg198, Glu19, Glu127, and Glu190 contribute to the binding of cytochrome P450 reductase. The mutagenesis results and a computational analysis of the protein surfaces partially define the binding site for P450 reductase. An overlapping binding site including Lys18, Lys22, Lys179, Arg183, and Arg185 is similarly defined for biliverdin reductase. These results confirm the binding of biliverdin reductase to hHO-1 and define binding sites of the two reductases.

  4. Involvement of PKA and HO-1 signaling in anti-inflammatory effects of surfactin in BV-2 microglial cells

    Park, Sun Young; Kim, Ji-Hee [Department of Molecular Biology, College of Natural Sciences, Pusan National University, Jangjeon-dong, Keumjeong-gu, Busan 609-735 (Korea, Republic of); Lee, Sang Joon [Department of Microbiology, College of Natural Sciences, Pusan National University, Jangjeon-dong, Keumjeong-gu, Busan 609-735 (Korea, Republic of); Kim, YoungHee, E-mail: yheekim@pusan.ac.kr [Department of Molecular Biology, College of Natural Sciences, Pusan National University, Jangjeon-dong, Keumjeong-gu, Busan 609-735 (Korea, Republic of)

    2013-04-01

    Surfactin, one of the most powerful biosurfactants, is a bacterial cyclic lipopeptide. Here, we investigated the anti-neuroinflammatory properties of surfactin in lipoteichoic acid (LTA)-stimulated BV-2 microglial cells. Surfactin significantly inhibited excessive production of the pro-inflammatory mediators TNF-α, IL-1β, IL-6, monocyte chemoattractant protein-1 (MCP-1), prostaglandin E{sub 2} (PGE{sub 2}), nitric oxide (NO) and reactive oxygen species (ROS), and suppressed the expression of matrix metalloproteinase-9 (MMP-9), inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2). Subsequent mechanistic studies revealed that surfactin inhibited LTA-induced nuclear factor-kappaB (NF-κB) and signal transducer and activator of transcription-1 (STAT-1) activation. However, surfactin increases the phosphorylation of the STAT-3, a component of the homeostatic mechanism causing anti-inflammatory events. We also demonstrated that surfactin induces heme oxygenase-1 (HO-1) expression and nuclear factor-regulated factor-2 (Nrf-2) activation, and that the anti-inflammatory effects of surfactin are abrogated by small interfering RNA-mediated knock-down of HO-1 or Nrf-2. Interestingly, we found that surfactin increased the level of cAMP and induced phosphorylation of cAMP responsive element binding protein (CREB) in microglial cells. Furthermore, treatment with the protein kinase A (PKA) inhibitor, H-89, blocked HO-1 induction by surfactin and abolished surfactin's suppressive effects on ROS and NO production. These results indicate that HO-1 and its upstream effector, PKA, play a pivotal role in the anti-neuroinflammatory response of surfactin in LTA-stimulated microglia. Therefore, surfactin might have therapeutic potential for neuroprotective agents to treat inflammatory and neurodegenerative diseases. - Highlights: ► Surfactin inhibits proinflammatory mediator synthesis in LTA-activated BV-2 cells. ► Surfactin suppresses NF-κB and STAT-1, but potentiates

  5. Involvement of PKA and HO-1 signaling in anti-inflammatory effects of surfactin in BV-2 microglial cells

    Park, Sun Young; Kim, Ji-Hee; Lee, Sang Joon; Kim, YoungHee

    2013-01-01

    Surfactin, one of the most powerful biosurfactants, is a bacterial cyclic lipopeptide. Here, we investigated the anti-neuroinflammatory properties of surfactin in lipoteichoic acid (LTA)-stimulated BV-2 microglial cells. Surfactin significantly inhibited excessive production of the pro-inflammatory mediators TNF-α, IL-1β, IL-6, monocyte chemoattractant protein-1 (MCP-1), prostaglandin E 2 (PGE 2 ), nitric oxide (NO) and reactive oxygen species (ROS), and suppressed the expression of matrix metalloproteinase-9 (MMP-9), inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2). Subsequent mechanistic studies revealed that surfactin inhibited LTA-induced nuclear factor-kappaB (NF-κB) and signal transducer and activator of transcription-1 (STAT-1) activation. However, surfactin increases the phosphorylation of the STAT-3, a component of the homeostatic mechanism causing anti-inflammatory events. We also demonstrated that surfactin induces heme oxygenase-1 (HO-1) expression and nuclear factor-regulated factor-2 (Nrf-2) activation, and that the anti-inflammatory effects of surfactin are abrogated by small interfering RNA-mediated knock-down of HO-1 or Nrf-2. Interestingly, we found that surfactin increased the level of cAMP and induced phosphorylation of cAMP responsive element binding protein (CREB) in microglial cells. Furthermore, treatment with the protein kinase A (PKA) inhibitor, H-89, blocked HO-1 induction by surfactin and abolished surfactin's suppressive effects on ROS and NO production. These results indicate that HO-1 and its upstream effector, PKA, play a pivotal role in the anti-neuroinflammatory response of surfactin in LTA-stimulated microglia. Therefore, surfactin might have therapeutic potential for neuroprotective agents to treat inflammatory and neurodegenerative diseases. - Highlights: ► Surfactin inhibits proinflammatory mediator synthesis in LTA-activated BV-2 cells. ► Surfactin suppresses NF-κB and STAT-1, but potentiates phosphorylation

  6. Interaction of nitric oxide with human heme oxygenase-1.

    Wang, Jinling; Lu, Shen; Moënne-Loccoz, Pierre; Ortiz de Montellano, Paul R

    2003-01-24

    NO and CO may complement each other as signaling molecules in some physiological situations. We have examined the binding of NO to human heme oxygenase-1 (hHO-1), an enzyme that oxidizes heme to biliverdin, CO, and free iron, to determine whether inhibition of hHO-1 by NO can contribute to the signaling interplay of NO and CO. An Fe(3+)-NO hHO-1-heme complex is formed with NO or the NO donors NOC9 or 2-(N,N-diethylamino)-diazenolate-2-oxide.sodium salt. Resonance Raman spectroscopy shows that ferric hHO-1-heme forms a 6-coordinated, low spin complex with NO. The nu(N-O) vibration of this complex detected by Fourier transform IR is only 4 cm(-1) lower than that of the corresponding metmyoglobin (met-Mb) complex but is broader, suggesting a greater degree of ligand conformational freedom. The Fe(3+)-NO complex of hHO-1 is much more stable than that of met-Mb. Stopped-flow studies indicate that k(on) for formation of the hHO-1-heme Fe(3+)-NO complex is approximately 50-times faster, and k(off) 10 times slower, than for met-Mb, resulting in K(d) = 1.4 microm for NO. NO thus binds 500-fold more tightly to ferric hHO-1-heme than to met-Mb. The hHO-1 mutations E29A, G139A, D140A, S142A, G143A, G143F, and K179A/R183A do not significantly diminish the tight binding of NO, indicating that NO binding is not highly sensitive to mutations of residues that normally stabilize the distal water ligand. As expected from the K(d) value, the enzyme is reversibly inhibited upon exposure to pathologically, and possibly physiologically, relevant concentrations of NO. Inhibition of hHO-1 by NO may contribute to the pleiotropic responses to NO and CO.

  7. Mechanism of estrogen-mediated attenuation of hepatic injury following trauma-hemorrhage: Akt-dependent HO-1 up-regulation.

    Hsu, Jun-Te; Kan, Wen-Hong; Hsieh, Chi-Hsun; Choudhry, Mashkoor A; Schwacha, Martin G; Bland, Kirby I; Chaudry, Irshad H

    2007-10-01

    Protein kinase B (Akt) is known to be involved in proinflammatory and chemotactic events in response to injury. Akt activation also leads to the induction of heme oxygenase (HO)-1. Up-regulation of HO-1 mediates potent, anti-inflammatory effects and attenuates organ injury. Although studies have shown that 17beta-estradiol (E2) prevents organ damage following trauma-hemorrhage, it remains unknown whether Akt/HO-1 plays any role in E2-mediated attenuation of hepatic injury following trauma-hemorrhage. To study this, male rats underwent trauma-hemorrhage (mean blood pressure, approximately 40 mmHg for 90 min), followed by fluid resuscitation. At the onset of resuscitation, rats were treated with vehicle, E2 (1 mg/kg body weight), E2 plus the PI-3K inhibitor (Wortmannin), or the estrogen receptor (ER) antagonist (ICI 182,780). At 2 h after sham operation or trauma-hemorrhage, plasma alpha-GST and hepatic tissue myeloperoxidase (MPO) activity, IL-6, TNF-alpha, ICAM-1, cytokine-induced neutrophil chemoattractant-1, and MIP-2 levels were measured. Hepatic Akt and HO-1 protein levels were also determined. Trauma-hemorrhage increased hepatic injury markers (alpha-GST and MPO activity), cytokines, ICAM-1, and chemokine levels. These parameters were markedly improved in the E2-treated rats following trauma-hemorrhage. E2 treatment also increased hepatic Akt activation and HO-1 expression compared with vehicle-treated, trauma-hemorrhage rats, which were abolished by coadministration of Wortmannin or ICI 182,780. These results suggest that the salutary effects of E2 on hepatic injury following trauma-hemorrhage are in part mediated via an ER-related, Akt-dependent up-regulation of HO-1.

  8. Antigenicity and protective efficacy of a Leishmania amastigote-specific protein, member of the super-oxygenase family, against visceral leishmaniasis.

    Vivian T Martins

    Full Text Available The present study aimed to evaluate a hypothetical Leishmania amastigote-specific protein (LiHyp1, previously identified by an immunoproteomic approach performed in Leishmania infantum, which showed homology to the super-oxygenase gene family, attempting to select a new candidate antigen for specific serodiagnosis, as well as to compose a vaccine against VL.The LiHyp1 DNA sequence was cloned; the recombinant protein (rLiHyp1 was purified and evaluated for its antigenicity and immunogenicity. The rLiHyp1 protein was recognized by antibodies from sera of asymptomatic and symptomatic animals with canine visceral leishmaniasis (CVL, but presented no cross-reactivity with sera of dogs vaccinated with Leish-Tec, a Brazilian commercial vaccine; with Chagas' disease or healthy animals. In addition, the immunogenicity and protective efficacy of rLiHyp1 plus saponin was evaluated in BALB/c mice challenged subcutaneously with virulent L. infantum promastigotes. rLiHyp1 plus saponin vaccinated mice showed a high and specific production of IFN-γ, IL-12, and GM-CSF after in vitro stimulation with the recombinant protein. Immunized and infected mice, as compared to the control groups (saline and saponin, showed significant reductions in the number of parasites found in the liver, spleen, bone marrow, and in the paws' draining lymph nodes. Protection was associated with an IL-12-dependent production of IFN-γ, produced mainly by CD4 T cells. In these mice, a decrease in the parasite-mediated IL-4 and IL-10 response could also be observed.The present study showed that this Leishmania oxygenase amastigote-specific protein can be used for a more sensitive and specific serodiagnosis of asymptomatic and symptomatic CVL and, when combined with a Th1-type adjuvant, can also be employ as a candidate antigen to develop vaccines against VL.

  9. Comparison of the crystal structure and function to wild-type and His25Ala mutant human heme oxygenase-1.

    Zhou, Wen-Pu; Zhong, Wen-Wei; Zhang, Xue-Hong; Ding, Jian-Ping; Zhang, Zi-Li; Xia, Zhen-Wei

    2009-03-01

    Human heme oxygenase-1 (hHO-1) is a rate-limiting enzyme in heme metabolism. It regulates serum bilirubin level. Site-directed mutagenesis studies indicate that the proximal residue histidine 25 (His25) plays a key role in hHO-1 activity. A highly purified hHO-1 His25Ala mutant was generated and crystallized with a new expression system. The crystal structure of the mutant was determined by X-ray diffraction technology and molecular replacement at the resolution of 2.8 A, and the model of hHO-1 His25Ala mutant was refined. The final crystallographic and free R factors were 0.245 and 0.283, respectively. The standard bond length deviation was 0.007 A, and the standard bond angle deviation was 1.3 degrees . The mutation of His25 to Ala led to an empty pocket underneath the ferric ion in the heme, leading to loss of binding iron ligand. Although this did not cause an overall structural change, the enzymatic activity of the mutant hHO-1 was reduced by 90%. By supplementing imidazole, the HO-1 activity was restored approximately 90% to its normal level. These data suggest that Ala25 remains unchanged in the structure compared to His25, but the important catalytic function of hHO-1 is lost. Thus, it appears that His25 is a crucial residue for proper hHO-1 catalysis.

  10. Non-coding RNAs and heme oxygenase-1 in vaccinia virus infection

    Meseda, Clement A.; Srinivasan, Kumar; Wise, Jasen; Catalano, Jennifer; Yamada, Kenneth M.; Dhawan, Subhash

    2014-01-01

    Highlights: • Heme oxygenase-1 (HO-1) induction inhibited vaccinia virus infection of macrophages. • Reduced infectivity inversely correlated with increased expression of non-coding RNAs. • The regulation of HO-1 and ncRNAs suggests a novel host defense response against vaccinia virus infection. - Abstract: Small nuclear RNAs (snRNAs) are <200 nucleotide non-coding uridylate-rich RNAs. Although the functions of many snRNAs remain undetermined, a population of snRNAs is produced during the early phase of infection of cells by vaccinia virus. In the present study, we demonstrate a direct correlation between expression of the cytoprotective enzyme heme oxygenase-1 (HO-1), suppression of selective snRNA expression, and inhibition of vaccinia virus infection of macrophages. Hemin induced HO-1 expression, completely reversed virus-induced host snRNA expression, and suppressed vaccinia virus infection. This involvement of specific virus-induced snRNAs and associated gene clusters suggests a novel HO-1-dependent host-defense pathway in poxvirus infection

  11. The Protective Role of Carbon Monoxide (CO Produced by Heme Oxygenases and Derived from the CO-Releasing Molecule CORM-2 in the Pathogenesis of Stress-Induced Gastric Lesions: Evidence for Non-Involvement of Nitric Oxide (NO

    Katarzyna Magierowska

    2016-03-01

    Full Text Available Carbon monoxide (CO produced by heme oxygenase (HO-1 and HO-2 or released from the CO-donor, tricarbonyldichlororuthenium (II dimer (CORM-2 causes vasodilation, with unknown efficacy against stress-induced gastric lesions. We studied whether pretreatment with CORM-2 (0.1–10 mg/kg oral gavage (i.g., RuCl3 (1 mg/kg i.g., zinc protoporphyrin IX (ZnPP (10 mg/kg intraperitoneally (i.p., hemin (1–10 mg/kg i.g. and CORM-2 (1 mg/kg i.g. combined with NG-nitro-l-arginine (l-NNA, 20 mg/kg i.p., 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 10 mg/kg i.p., indomethacin (5 mg/kg i.p., SC-560 (5 mg/kg i.g., and celecoxib (10 mg/kg i.g. affects gastric lesions following 3.5 h of water immersion and restraint stress (WRS. Gastric blood flow (GBF, the number of gastric lesions and gastric CO and nitric oxide (NO contents, blood carboxyhemoglobin (COHb level and the gastric expression of HO-1, HO-2, hypoxia inducible factor 1α (HIF-1α, tumor necrosis factor α (TNF-α, cyclooxygenase (COX-2 and inducible NO synthase (iNOS were determined. CORM-2 (1 mg/kg i.g. and hemin (10 mg/kg i.g. significantly decreased WRS lesions while increasing GBF, however, RuCl3 was ineffective. The impact of CORM-2 was reversed by ZnPP, ODQ, indomethacin, SC-560 and celecoxib, but not by l-NNA. CORM-2 decreased NO and increased HO-1 expression and CO and COHb content, downregulated HIF-1α, as well as WRS-elevated COX-2 and iNOS mRNAs. Gastroprotection by CORM-2 and HO depends upon CO’s hyperemic and anti-inflammatory properties, but is independent of NO.

  12. Identification of danthron as an isoform-specific inhibitor of HEME OXYGENASE-1/cytochrome P450 reductase interaction with anti-tumor activity.

    Chou, Yi-Tai; Hsu, Fu-Fei; Hu, Dun-Yao; Chen, Ying-Chih; Hsu, Yuan-Hao; Hsu, John T-A; Chau, Lee-Young

    2018-01-23

    Heme oxygenase (HO) catalyzes NADPH-dependent degradation of heme to liberate iron, carbon monoxide and biliverdin. The interaction between HO and cytochrome P450 reductase (CPR), an electron donor, is essential for HO activity. HO-1 is a stress-inducible isoform whereas HO-2 is constitutively expressed. HO-1 induction is commonly seen in cancers and impacts disease progression, supporting the possibility of targeting HO-1 for cancer therapy. We employed a cell-based bioluminescence resonance energy transfer assay to screen compounds with ability to inhibit HO-1/CPR interaction. The effect of the identified compound on HO-1/CPR interaction was confirmed by pull down assay. Moreover, the anti-tumorigenic activity of the identified compound on HO-1-enhanced tumor growth and migration was assessed by trypan blue exclusion method and wound healing assay. Danthron was identified as an effective small molecule able to interfere with the interaction between HO-1 and CPR but not HO-2 and CPR. Additional experiments with structural analogues of danthron revealed that the positions of hydroxyl moieties significantly affected the potency of inhibition on HO-1/CPR interaction. Pull-down assay confirmed that danthron inhibited the interaction of CPR with HO-1 but not HO-2. Danthron suppressed growth and migration of HeLa cells with stable HO-1 overexpression but not mock cells. In contrast, anthrarufin, a structural analog with no ability to interfere HO-1/CPR interaction, exhibited no significant effect on HO-1-overexpressing HeLa cells. These findings demonstrate that danthron is an isoform-specific inhibitor for HO-1/CPR interaction and may serve as a lead compound for novel anticancer drug.

  13. Regulation of human heme oxygenase in endothelial cells by using sense and antisense retroviral constructs.

    Quan, S; Yang, L; Abraham, N G; Kappas, A

    2001-10-09

    Our objective was to determine whether overexpression and underexpression of human heme oxygenase (HHO)-1 could be controlled on a long-term basis by introduction of the HO-1 gene in sense (S) and antisense (AS) orientation with an appropriate vector into endothelial cells. Retroviral vector (LXSN) containing viral long terminal repeat promoter-driven human HO-1 S (LSN-HHO-1) and LXSN vectors containing HHO-1 promoter (HOP)-controlled HHO-1 S and AS (LSN-HOP-HHO-1 and LSN-HOP-HHO-1-AS) sequences were constructed and used to transfect rat lung microvessel endothelial cells (RLMV cells) and human dermal microvessel endothelial cells (HMEC-1 cells). RLMV cells transduced with HHO-1 S expressed human HO-1 mRNA and HO-1 protein associated with elevation in total HO activity compared with nontransduced cells. Vector-mediated expression of HHO-1 S or AS under control of HOP resulted in effective production of HO-1 or blocked induction of endogenous human HO-1 in HMEC-1 cells, respectively. Overexpression of HO-1 AS was associated with a long-term decrease (45%) of endogenous HO-1 protein and an increase (167%) in unmetabolized exogenous heme in HMEC-1 cells. Carbon monoxide (CO) production in HO-1 S- or AS-transduced HMEC-1 cells after heme treatment was increased (159%) or decreased (50%), respectively, compared with nontransduced cells. HO-2 protein levels did not change. These findings demonstrate that HHO-1 S and AS retroviral constructs are functional in enhancing and reducing HO activity, respectively, and thus can be used to regulate cellular heme levels, the activity of heme-dependent enzymes, and the rate of heme catabolism to CO and bilirubin.

  14. Isoporphyrin Intermediate in Heme Oxygenase Catalysis

    Evans, John P.; Niemevz, Fernando; Buldain, Graciela; de Montellano, Paul Ortiz

    2008-01-01

    Human heme oxygenase-1 (hHO-1) catalyzes the O2- and NADPH-dependent oxidation of heme to biliverdin, CO, and free iron. The first step involves regiospecific insertion of an oxygen atom at the α-meso carbon by a ferric hydroperoxide and is predicted to proceed via an isoporphyrin π-cation intermediate. Here we report spectroscopic detection of a transient intermediate during oxidation by hHO-1 of α-meso-phenylheme-IX, α-meso-(p-methylphenyl)-mesoheme-III, and α-meso-(p-trifluoromethylphenyl)-mesoheme-III. In agreement with previous experiments (Wang, J., Niemevz, F., Lad, L., Huang, L., Alvarez, D. E., Buldain, G., Poulos, T. L., and Ortiz de Montellano, P. R. (2004) J. Biol. Chem. 279, 42593–42604), only the α-biliverdin isomer is produced with concomitant formation of the corresponding benzoic acid. The transient intermediate observed in the NADPH-P450 reductase-catalyzed reaction accumulated when the reaction was supported by H2O2 and exhibited the absorption maxima at 435 and 930 nm characteristic of an isoporphyrin. Product analysis by reversed phase high performance liquid chromatography and liquid chromatography electrospray ionization mass spectrometry of the product generated with H2O2 identified it as an isoporphyrin that, on quenching, decayed to benzoylbiliverdin. In the presence of H218O2, one labeled oxygen atom was incorporated into these products. The hHO-1-isoporphyrin complexes were found to have half-lives of 1.7 and 2.4 h for the p-trifluoromethyl- and p-methyl-substituted phenylhemes, respectively. The addition of NADPH-P450 reductase to the H2O2-generated hHO-1-isoporphyrin complex produced α-biliverdin, confirming its role as a reaction intermediate. Identification of an isoporphyrin intermediate in the catalytic sequence of hHO-1, the first such intermediate observed in hemoprotein catalysis, completes our understanding of the critical first step of heme oxidation. PMID:18487208

  15. 4-Hydroxyestradiol induces mammary epithelial cell transformation through Nrf2-mediated heme oxygenase-1 overexpression

    Park, Sin-Aye; Lee, Mee-Hyun; Na, Hye-Kyung; Surh, Young-Joon

    2016-01-01

    Estrogen (17?-estradiol, E2) undergoes oxidative metabolism by CYP1B1 to form 4-hydroxyestradiol (4-OHE2), a putative carcinogenic metabolite of estrogen. Our previous study showed that 4-OHE2-induced production of reactive oxygen species contributed to neoplastic transformation of human breast epithelial (MCF-10A) cells. In this study, 4-OHE2, but not E2, increased the expression of heme oxygenase-1 (HO-1), a sensor and regulator of oxidative stress, in MCF-10A cells. Silencing the HO-1 gene...

  16. Impact of heme oxygenase-1 on cholesterol synthesis, cholesterol efflux and oxysterol formation in cultured astroglia.

    Hascalovici, Jacob R; Song, Wei; Vaya, Jacob; Khatib, Soliman; Fuhrman, Bianca; Aviram, Michael; Schipper, Hyman M

    2009-01-01

    Up-regulation of heme oxygenase-1 (HO-1) and altered cholesterol (CH) metabolism are characteristic of Alzheimer-diseased neural tissues. The liver X receptor (LXR) is a molecular sensor of CH homeostasis. In the current study, we determined the effects of HO-1 over-expression and its byproducts iron (Fe(2+)), carbon monoxide (CO) and bilirubin on CH biosynthesis, CH efflux and oxysterol formation in cultured astroglia. HO-1/LXR interactions were also investigated in the context of CH efflux. hHO-1 over-expression for 3 days ( approximately 2-3-fold increase) resulted in a 30% increase in CH biosynthesis and a two-fold rise in CH efflux. Both effects were abrogated by the competitive HO inhibitor, tin mesoporphyrin. CO, released from administered CORM-3, significantly enhanced CH biosynthesis; a combination of CO and iron stimulated CH efflux. Free iron increased oxysterol formation three-fold. Co-treatment with LXR antagonists implicated LXR activation in the modulation of CH homeostasis by heme degradation products. In Alzheimer's disease and other neuropathological states, glial HO-1 induction may transduce ambient noxious stimuli (e.g. beta-amyloid) into altered patterns of glial CH homeostasis. As the latter may impact synaptic plasticity and neuronal repair, modulation of glial HO-1 expression (by pharmacological or other means) may confer neuroprotection in patients with degenerative brain disorders.

  17. Carbon monoxide mediates heme oxygenase 1 induction via Nrf2 activation in hepatoma cells

    Lee, Bok-Soo; Heo, JungHee; Kim, Yong-Man; Shim, Sang Moo; Pae, Hyun-Ock; Kim, Young-Myeong; Chung, Hun-Taeg

    2006-01-01

    Carbon monoxide (CO) and nitric oxide (NO) are two gas molecules which have cytoprotective functions against oxidative stress and inflammatory responses in many cell types. Currently, it is known that NO produced by nitric oxide synthase (NOS) induces heme oxygenase 1 (HO1) expression and CO produced by the HO1 inhibits inducible NOS expression. Here, we first show CO-mediated HO1 induction and its possible mechanism in human hepatocytes. Exposure of HepG2 cells or primary hepatocytes to CO resulted in dramatic induction of HO1 in dose- and time-dependent manner. The CO-mediated HO1 induction was abolished by MAP kinase inhibitors (MAPKs) but not affected by inhibitors of PI3 kinase or NF-κB. In addition, CO induced the nuclear translocation and accumulation of Nrf2, which suppressed by MAPKs inhibitors. Taken together, we suggest that CO induces Nrf2 activation via MAPKs signaling pathways, thereby resulting in HO1 expression in HepG2 cells

  18. Fructose Mediated Non-Alcoholic Fatty Liver Is Attenuated by HO-1-SIRT1 Module in Murine Hepatocytes and Mice Fed a High Fructose Diet.

    Komal Sodhi

    Full Text Available Oxidative stress underlies the etiopathogenesis of nonalcoholic fatty liver disease (NAFLD, obesity and cardiovascular disease (CVD. Heme Oxygenase-1 (HO-1 is a potent endogenous antioxidant gene that plays a key role in decreasing oxidative stress. Sirtuin1 (SIRT1 belongs to the family of NAD-dependent de-acyetylases and is modulated by cellular redox.We hypothesize that fructose-induced obesity creates an inflammatory and oxidative environment conducive to the development of NAFLD and metabolic syndrome. The aim of this study is to determine whether HO-1 acts through SIRT1 to form a functional module within hepatocytes to attenuate steatohepatitis, hepatic fibrosis and cardiovascular dysfunction.We examined the effect of fructose, on hepatocyte lipid accumulation and fibrosis in murine hepatocytes and in mice fed a high fructose diet in the presence and absence of CoPP, an inducer of HO-1, and SnMP, an inhibitor of HO activity. Fructose increased oxidative stress markers and decreased HO-1 and SIRT1 levels in hepatocytes (p<0.05. Further fructose supplementation increased FAS, PPARα, pAMPK and triglycerides levels; CoPP negated this increase. Concurrent treatment with CoPP and SIRT1 siRNA in hepatocytes increased FAS, PPARα, pAMPK and triglycerides levels suggesting that HO-1 is upstream of SIRT1 and suppression of SIRT1 attenuates the beneficial effects of HO-1. A high fructose diet increased insulin resistance, blood pressure, markers of oxidative stress and lipogenesis along with fibrotic markers in mice (p<0.05. Increased levels of HO-1 increased SIRT1 levels and ameliorated fructose-mediated lipid accumulation and fibrosis in liver along with decreasing vascular dysfunction (p<0.05 vs. fructose. These beneficial effects of CoPP were reversed by SnMP.Taken together, our study demonstrates, for the first time, that HO-1 induction attenuates fructose-induced hepatic lipid deposition, prevents the development of hepatic fibrosis and abates

  19. Reduced caveolin-1 promotes hyper-inflammation due to abnormal heme oxygenase-1 localizationin LPS challenged macrophages with dysfunctional CFTR

    Zhang, Ping-Xia; Murray, Thomas S.; Villella, Valeria Rachela; Ferrari, Eleonora; Esposito, Speranza; D'Souza, Anthony; Raia, Valeria; Maiuri, Luigi; Krause, Diane S.; Egan, Marie E.; Bruscia, Emanuela M.

    2013-01-01

    We have previously reported that TLR4 signaling is increased in lipopolysaccharide (LPS) -stimulated Cystic Fibrosis (CF) macrophages (MΦs), contributing to the robust production of pro-inflammatory cytokines. The heme oxygenase (HO-1)/carbon monoxide (CO) pathway modulates cellular redox status, inflammatory responses, and cell survival. The HO-1 enzyme, together with the scaffold protein caveolin 1 (CAV-1), also acts as a negative regulator of TLR4 signaling in MΦs. Here, we demonstrate that in LPS-challenged CF MΦs, HO-1 does not compartmentalize normally to the cell surface and instead accumulates intracellularly. The abnormal HO-1 localization in CF MΦs in response to LPS is due to decreased CAV-1 expression, which is controlled by the cellular oxidative state, and is required for HO-1 delivery to the cell surface. Overexpression of HO-1 or stimulating the pathway with CO-releasing molecules (CORM2)enhancesCAV-1 expression in CF MΦs, suggesting a positive-feed forward loop between HO-1/CO induction and CAV-1 expression. These manipulations reestablished HO-1 and CAV-1 cell surface localization in CF MΦ's. Consistent with restoration of HO-1/CAV-1 negative regulation of TLR4 signaling, genetic or pharmacological (CORM2)-induced enhancement of this pathway decreased the inflammatory response of CF MΦs and CF mice treated with LPS. In conclusion, our results demonstrate that the counter-regulatory HO-1/CO pathway, which is critical in balancing and limiting the inflammatory response, is defective in CF MΦs through a CAV-1-dependent mechanism, exacerbating the CF MΦ's response to LPS. This pathway could be a potential target for therapeutic intervention for CF lung disease. PMID:23606537

  20. Functional imaging: monitoring heme oxygenase-1 gene expression in vivo

    Zhang, Weisheng; Reilly-Contag, Pamela; Stevenson, David K.; Contag, Christopher H.

    1999-07-01

    The regulation of genetic elements can be monitored in living animals using photoproteins as reporters. Heme oxygenase (HO) is the key catabolic enzyme in the heme degradation pathway. Here, HO expression serves as a model for in vivo functional imaging of transcriptional regulation of a clinically relevant gene. HO enzymatic activity is inhibited by heme analogs, metalloporphyrins, but many members of this family of compounds also activate transcription of the HO-1 promoter. The degree of transcriptional activation by twelve metalloporphyrins, differing at the central metal and porphyrin ring substituents, was evaluated in both NIH 3T3 stable lines and transgenic animals containing HO-1 promoter-luciferase gene fusions. In the correlative cell culture assays, the metalloporphyrins increased transcription form the full length HO promoter fusion to varying degrees, but none increased transcription from a truncated HO-1 promoter. These results suggested that one or both of the two distal enhancer elements located at -4 and -10 Kb upstream from transcriptional start are required for HO-1 induction by heme and its analogs. The full-length HO-1-luc fusion was then evaluated as a transgene in mice. It was possible to monitor the effects of the metalloporphyrins, SnMP and ZnPP, in living animals over time. This spatiotemporal analyses of gene expression in vivo implied that alterations in porphyrin ring substituents and the central metal may affect the extent of gene activation. These data further indicate that using photoprotein reporters, subtle differences in gene expression can be monitored in living animals.

  1. Heme oxygenase-1: A new druggable target in the management of chronic and acute myeloid leukemia.

    Salerno, Loredana; Romeo, Giuseppe; Modica, Maria N; Amata, Emanuele; Sorrenti, Valeria; Barbagallo, Ignazio; Pittalà, Valeria

    2017-12-15

    Heme oxygenase-1 (HO-1) is the enzyme catalyzing the rate-limiting oxidative degradation of cellular heme into free iron, carbon monoxide (CO), and biliverdin, which is then rapidly converted into bilirubin. By means of these catabolic end-products and by removal of pro-oxidant heme, HO-1 exerts antioxidant, antiapoptotic, and immune-modulating effects, leading to overall cytoprotective and beneficial functions in mammalian cells. Therefore, HO-1 is considered a survival molecule in various stress-related conditions. By contrast, growing evidence suggests that HO-1 is a survival-enhancing molecule also in various solid and blood cancers, such as various types of leukemia, promoting carcinogenesis, tumor progression, and chemo-resistance. Among leukemias, chronic myeloid leukemia (CML) is currently therapeutically well treated with tyrosine kinase inhibitors (TKIs) such as Imatinib (IM) and its congeners; nevertheless, resistance to all kinds of current drugs persist in a number of patients. Moreover, treatment outcomes for acute myeloid leukemia (AML) remain unsatisfactory, despite progress in chemotherapy and hematopoietic stem cell transplantation. Therefore, identification of new eligible targets that may improve leukemias therapy is of general interest. Several recent papers prove that inhibition of HO-1 through HO-1 inhibitors as well as modulation of other pathways involving HO-1 by a number of different new or known molecules, are critical for leukemia treatment. This review summarizes the current understanding of the pro-tumorigenic role of HO-1 and its potential as a molecular target for the treatment of leukemias. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  2. Effects of Metalloporphyrins on Heme Oxygenase-1 Transcription: Correlative Cell Culture Assays Guide in Vivo Imaging

    Monica Hajdena-Dawson; Weisheng Zhang; Pamela R. Contag; Ronald J. Wong; Hendrik J. Vreman; David K. Stevenson; Christopher H. Contag

    2003-01-01

    Heme oxygenase (HO) is the rate-limiting step in the heme degradation pathway and is a potential target for the control, or prevention, of pathologic jaundice in neonates. Metalloporphyrins (Mps), a diverse set of synthetic derivatives of heme, can competitively inhibit the HO enzymes. However, certain Mps are phototoxic and some increase transcription of HO-1, the inducible HO isozyme. Therefore, effective development of this class of compounds as therapeutics for treating pathologic jaundic...

  3. Tyrosine oxidation in heme oxygenase: examination of long-range proton-coupled electron transfer.

    Smirnov, Valeriy V; Roth, Justine P

    2014-10-01

    Heme oxygenase is responsible for the degradation of a histidine-ligated ferric protoporphyrin IX (Por) to biliverdin, CO, and the free ferrous ion. Described here are studies of tyrosyl radical formation reactions that occur after oxidizing Fe(III)(Por) to Fe(IV)=O(Por(·+)) in human heme oxygenase isoform-1 (hHO-1) and the structurally homologous protein from Corynebacterium diphtheriae (cdHO). Site-directed mutagenesis on hHO-1 probes the reduction of Fe(IV)=O(Por(·+)) by tyrosine residues within 11 Å of the prosthetic group. In hHO-1, Y58· is implicated as the most likely site of oxidation, based on the pH and pD dependent kinetics. The absence of solvent deuterium isotope effects in basic solutions of hHO-1 and cdHO contrasts with the behavior of these proteins in the acidic solution, suggesting that long-range proton-coupled electron transfer predominates over electron transfer.

  4. Management of oxidative stress by heme oxygenase-1 in cisplatin-induced toxicity in renal tubular cells.

    Schaaf, G J; Maas, R F M; de Groene, E M; Fink-Gremmels, J

    2002-08-01

    Induction of heme oxygenase-1 (HO-1) may serve as an immediate protective response during treatment with the cytostatic drug cisplatin (CDDP). Oxidative pathways participate in the characteristic nephrotoxicity of CDDP. In the present study, cultured tubular cells (LLC-PK1) were used to investigate whether induction of HO provided protection against CDDP by maintaining the cellular redox balance. The antioxidants, alpha-tocopherol (TOCO) and N-acetylcysteine (NAC), were used to demonstrate that elevation of ROS levels contribute to the development of CDDP-induced cytotoxicity. Chemical modulators of HO activity were used to investigate the role of HO herein. Hemin was used to specifically induce HO-1, while exposure of the cells to tin-protoporphyrin (SnPP) was shown to inhibit HO activity. Hemin treatment prior to CDDP-exposure significantly decreased the generation of ROS to control levels, while inhibition of HO increased the ROS levels beyond the levels measured in cells treated with CDDP alone. Furthermore, HO induction protected significantly against the cytotoxicity of CDDP, although this protection was limited. Similar results were obtained when the cells were preincubated with TOCO, suggesting that mechanisms other than impairment of the redox ratio are important in CDDP-induced loss of cell viability in vitro. In addition, SnPP treatment exacerbated the oxidative response and cytotoxicity of CDDP, especially at low CDDP concentrations. We therefore conclude that HO is able to directly limit the CDDP-induced oxidative stress response and thus serves as safeguard of the cellular redox balance.

  5. Heme oxygenase-1 delays gibberellin-induced programmed cell death of rice aleurone layers subjected to drought stress by interacting with nitric oxide

    Huangming eWu

    2016-01-01

    Full Text Available Cereal aleurone layers undergo a gibberellin (GA-regulated process of programmed cell death (PCD following germination. Heme oxygenase-1 (HO-1 is known as a rate-liming enzyme in the degradation of heme to biliverdin IXα (BV, carbon monoxide (CO, and free iron ions (Fe2+. It is a critical component in plant development and adaptation to environment stresses. Our previous studies confirmed that HO-1 inducer hematin (Ht promotes the germination of rice seeds in drought (20% polyethylene glycol-6000, PEG conditions, but the corresponding effects of HO-1 on the alleviation of germination-triggered PCD in GA-treated rice aleurone layers remain unknown. The present study has determined that GA co-treated with PEG results in lower HO-1 transcript levels and HO activity, which in turn results in the development of vacuoles in aleurone cells, followed by PCD. The pharmacology approach illustrated that up- or down-regulated HO-1 gene expression and HO activity delayed or accelerated GA-induced PCD. Furthermore, the application of the HO-1 inducer hematin and nitric oxide (NO donor sodium nitroprusside (SNP not only activated HO-1 gene expression, HO activity, and endogenous NO content, but also blocked GA-induced rapid vacuolation and accelerated aleurone layers PCD under drought stress. However, both HO-1 inhibitor zinc protoporphyrin IX (ZnPPIX and NO scavenger 2-(4-carboxyphenyl0-4, 4, 5, 5-tetramethylimidazoline-l-oxyl-3-oxide potassium salt (cPTIO reserved the effects of hematin and SNP on rice aleurone layer PCD under drought stress by down-regulating endogenous HO-1 and NO, respectively. The inducible effects of hematin and SNP on HO-1 gene expression, HO activity, and NO content were blocked by cPTIO. Together, these results clearly suggest that HO-1 is involved in the alleviation of GA-induced PCD of drought-triggered rice aleurone layers by associating with NO.

  6. Chinese herbal medicine compound Yi-Zhi-Hao pellet inhibits replication of influenza virus infection through activation of heme oxygenase-1

    Jinqiu Yin

    2017-11-01

    Full Text Available As a leading cause of respiratory disease, influenza A virus (IAV presents a pandemic threat in annual seasonal outbreaks. Given the limitation of existing anti-influenza therapies, there remains to be a requirement for new drugs. Compound Yi-Zhi-Hao pellet (CYZH is a famous traditional Chinese medicine (TCM used in the clinic, whose formula has been recorded in Complication of National Standard for Traditional Chinese Medicine to treat common cold. In this study, we found that CYZH exhibited a broad-spectrum anti-influenza activity and inhibited the expression of viral RNA and proteins in vitro. Mechanistically, CYZH had no inhibitory activities against viral protein hemagglutinin and IAV RNA-dependent RNA polymerase. Instead, it induced activation of erythroid 2-related factor 2 (Nrf2 and nuclear factor kappa B (NF-κB, which subsequently upregulated heme oxygenase-1 (HO-1 expression. Also, CYZH protected cells from oxidative damage induced by reactive oxygen series. In conclusions, CYZH inhibits IAV replication in vitro, at least partly by activating expression of the Nrf2/HO-1 pathway.

  7. Physalis peruviana L. inhibits airway inflammation induced by cigarette smoke and lipopolysaccharide through inhibition of extracellular signal-regulated kinase and induction of heme oxygenase-1.

    Park, Hyun Ah; Lee, Jae-Won; Kwon, Ok-Kyoung; Lee, Gilhye; Lim, Yourim; Kim, Jung Hee; Paik, Jin-Hyub; Choi, Sangho; Paryanto, Imam; Yuniato, Prasetyawan; Kim, Doo-Young; Ryu, Hyung Won; Oh, Sei-Ryang; Lee, Seung Jin; Ahn, Kyung-Seop

    2017-11-01

    Physalis peruviana L. (PP) is a medicinal herb that has been confirmed to have several biological activities, including anticancer, antioxidant and anti-inflammatory properties. The aim of the present study was to evaluate the protective effect of PP on cigarette smoke (CS)- and lipopolysaccharide (LPS)-induced pulmonary inflammation. Treatment with PP significantly reduced the influx of inflammatory cells in the bronchoalveolar lavage fluid (BALF) and lung of mice with CS- and LPS-induced pulmonary inflammation. PP also decreased the levels of reactive oxygen species (ROS) and pro-inflammatory cytokines, such as tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in the BALF. PP effectively attenuated the expression of monocyte chemoattractant protein-1 (MCP-1) and the activation of extracellular signal-regulated kinase (ERK) in the lung. In addition, nuclear factor erythroid 2-related factor 2 (Nrf2) activation and heme oxygenase-1 (HO-1) expression were increased by PP treatment. In an in vitro experiment, PP reduced the mRNA expression of TNF-α and MCP-1, and the activation of ERK in CS extract-stimulated A549 epithelial cells. Furthermore, PP increased the activation of Nrf2 and the expression of HO-1 in A549 cells. These findings suggest that PP has a therapeutic potential for the treatment of pulmonary inflammatory diseases, such as chronic obstructive pulmonary disease.

  8. Opposite effect of oxidative stress on inducible nitric oxide synthase and haem oxygenase-1 expression in intestinal inflammation: anti-inflammatory effect of carbon monoxide

    Dijkstra, Gerard; Blokzijl, Hans; Bok, Lisette; Homan, Manon; van Goor, Harry; Faber, Klaas Nico; Jansen, Peter L. M.; Moshage, Han

    2004-01-01

    Inducible nitric oxide synthase (iNOS) is expressed in intestinal epithelial cells (IEC) of patients with active inflammatory bowel disease (IBD) and in IEC of endotoxaemic rats. The induction of iNOS in IEC is an element of the NF-kappaB-mediated survival pathway. Haem oxygenase-1 (HO-1) is an

  9. Mycobacterium tuberculosis Induction of Heme Oxygenase-1 Expression Is Dependent on Oxidative Stress and Reflects Treatment Outcomes

    Neesha Rockwood

    2017-05-01

    Full Text Available The antioxidant enzyme heme oxygenase-1 (HO-1 is implicated in the pathogenesis of tuberculosis (TB and has been proposed as a biomarker of active disease. Nevertheless, the mechanisms by which Mycobacterium tuberculosis (Mtb induces HO-1 as well as how its expression is affected by HIV-1 coinfection and successful antitubercular therapy (ATT are poorly understood. We found that HO-1 expression is markedly increased in rabbits, mice, and non-human primates during experimental Mtb infection and gradually decreased during ATT. In addition, we examined circulating concentrations of HO-1 in a cohort of 130 HIV-1 coinfected and uninfected pulmonary TB patients undergoing ATT to investigate changes in expression of this biomarker in relation to HIV-1 status, radiological disease severity, and treatment outcome. We found that plasma levels of HO-1 were elevated in untreated HIV-1 coinfected TB patients and correlated positively with HIV-1 viral load and negatively with CD4+ T cell count. In both HIV-1 coinfected and Mtb monoinfected patients, HO-1 levels were substantially reduced during successful TB treatment but not in those who experienced treatment failure or subsequently relapsed. To further delineate the molecular mechanisms involved in induction of HO-1 by Mtb, we performed a series of in vitro experiments using mouse and human macrophages. We found that Mtb-induced HO-1 expression requires NADPH oxidase-dependent reactive oxygen species production induced by the early-secreted antigen ESAT-6, which in turn triggers nuclear translocation of the transcription factor NRF-2. These observations provide further insight into the utility of HO-1 as a biomarker of both disease and successful therapy in TB monoinfected and HIV-TB coinfected patients and reveal a previously undocumented pathway linking expression of the enzyme with oxidative stress.

  10. Heme oxygenase attenuates angiotensin II-mediated superoxide production in cultured mouse thick ascending loop of Henle cells.

    Kelsen, Silvia; Patel, Bijal J; Parker, Lawson B; Vera, Trinity; Rimoldi, John M; Gadepalli, Rama S V; Drummond, Heather A; Stec, David E

    2008-10-01

    Heme oxygenase (HO)-1 induction can attenuate the development of angiotensin II (ANG II)-dependent hypertension. However, the mechanism by which HO-1 lowers blood pressure is not clear. The goal of this study was to test the hypothesis that induction of HO-1 can reduce the ANG II-mediated increase in superoxide production in cultured thick ascending loop of Henle (TALH) cells. Studies were performed on an immortalized cell line of mouse TALH (mTALH) cells. HO-1 was induced in cultured mTALH cells by treatment with cobalt protoporphyrin (CoPP, 10 microM) or hemin (50 microM) or by transfection with a plasmid containing the human HO-1 isoform. Treatment of mTALH cells with 10(-9) M ANG II increased dihydroethidium (DHE) fluorescence (an index of superoxide levels) from 35.5+/-5 to 136+/-18 relative fluorescence units (RFU)/microm2. Induction of HO-1 via CoPP, hemin, or overexpression of the human HO-1 isoform significantly reduced ANG II-induced DHE fluorescence to 64+/-5, 64+/-8, and 41+/-4 RFU/microm2, respectively. To determine which metabolite of HO-1 is responsible for reducing ANG II-mediated increases in superoxide production in mTALH cells, cells were preincubated with bilirubin or carbon monoxide (CO)-releasing molecule (CORM)-A1 (each at 100 microM) before exposure to ANG II. DHE fluorescence averaged 80+/-7 RFU/microm2 after incubation with ANG II and was significantly decreased to 55+/-7 and 53+/-4 RFU/microm2 after pretreatment with bilirubin and CORM-A1. These results demonstrate that induction of HO-1 in mTALH cells reduces the levels of ANG II-mediated superoxide production through the production of both bilirubin and CO.

  11. Generation of CMAHKO/GTKO/shTNFRI-Fc/HO-1 quadruple gene modified pigs.

    Kim, Geon A; Lee, Eun Mi; Jin, Jun-Xue; Lee, Sanghoon; Taweechaipaisankul, Anukul; Hwang, Jong Ik; Alam, Zahid; Ahn, Curie; Lee, Byeong Chun

    2017-08-01

    As an alternative source of organs for transplantation into humans, attention has been directed to pigs due to their similarities in biological features and organ size. However, severe immune rejection has prevented successful xenotransplantation using pig organs and tissues. To overcome immune rejection, recently developed genetic engineering systems such as TALEN coupled with somatic cell nuclear transfer (SCNT) to make embryos could be used to produce pigs compatible with xenotransplantation. We used the TALEN system to target the non-Gal antigen cytidine monophosphate-N-acetylneuraminic acid hydroxylase (CMAH) gene in pigs that is naturally deleted in humans. Gal-deleted cells expressing both soluble human tumor necrosis factor receptor I IgG 1 -Fc (shTNFRI-Fc) and human hemagglutinin -tagged-human heme oxygenase-1 (hHO-1) were transfected with a TALEN target for CMAH. Cells lacking CMAH were negatively selected using N-glyconeuraminic acid (Neu5Gc)/magnetic beads and the level of Neu5Gc expression of isolated cells were analyzed by FACS and DNA sequencing. Cloned embryos using 3 different genetically modified cell clones were respectively transferred into 3 recipients, with 55.6% (5/9) becoming pregnant and three cloned pigs were produced. Successful genetic disruption of the CMAH gene was confirmed by sequencing, showing lack of expression of CMAH in tail-derived fibroblasts of the cloned piglets. Besides decreased expression of Neu5Gc in piglets produced by SCNT, antibody-mediated complement-dependent cytotoxicity assays and natural antibody binding for examining immuno-reactivity of the quadruple gene modified pigs derived from endothelial cells and fibroblasts were reduced significantly compared to those of wild type animals. We conclude that by combining the TALEN system and transgenic cells, targeting of multiple genes could be useful for generating organs for xenotransplantation. We produced miniature pigs with quadruple modified genes CMAHKO/GTKO/shTNFRI-Fc/hHO

  12. Andrographolide Activates Keap1/Nrf2/ARE/HO-1 Pathway in HT22 Cells and Suppresses Microglial Activation by Aβ42 through Nrf2-Related Inflammatory Response.

    Seo, Ji Yeon; Pyo, Euisun; An, Jin-Pyo; Kim, Jinwoong; Sung, Sang Hyun; Oh, Won Keun

    2017-01-01

    Therapeutic approach of Alzheimer's disease (AD) has been gradually diversified. We examined the therapeutic and preventive potential of andrographolide, which is a lactone diterpenoid from Andrographis paniculata , and focused on the Kelch-like ECH-associated protein 1 (Keap1)/nuclear factor (erythroid-derived 2)-like 2 (Nrf2)-mediated heme oxygenase (HO)-1-inducing effects and the inhibitory activity of amyloid beta (A β ) 42 -induced microglial activation related to Nrf2 and nuclear factor κ B (NF- κ B)-mediated inflammatory responses. Andrographolide induced the expression and translocation of Nrf2 from the cytoplasm to the nucleus, thereby activating antioxidant response element (ARE) gene transcription and HO-1 expression in murine hippocampal HT22 cells. Andrographolide eliminated intracellular A β 42 in BV-2 cells and decreased the production of interleukin (IL)-6, IL-1 β , prostaglandin (PG)E 2 , and nitric oxide (NO) because of artificial phagocytic A β 42 . It decreased pNF- κ B accumulation in the nucleus and the expression of inducible nitric oxide synthase (i-NOS) and cyclooxygenase II (COX-II) in the microglial BV-2 cell line. In summary, andrographolide activates Nrf2-mediated HO-1 expression and inhibits A β 42 -overexpressed microglial BV-2 cell activation. These results suggested that andrographolide might have the potential for further examination of the therapeutics of AD.

  13. Andrographolide Activates Keap1/Nrf2/ARE/HO-1 Pathway in HT22 Cells and Suppresses Microglial Activation by Aβ42 through Nrf2-Related Inflammatory Response

    Ji Yeon Seo

    2017-01-01

    Full Text Available Therapeutic approach of Alzheimer’s disease (AD has been gradually diversified. We examined the therapeutic and preventive potential of andrographolide, which is a lactone diterpenoid from Andrographis paniculata, and focused on the Kelch-like ECH-associated protein 1 (Keap1/nuclear factor (erythroid-derived 2-like 2 (Nrf2-mediated heme oxygenase (HO-1-inducing effects and the inhibitory activity of amyloid beta (Aβ42-induced microglial activation related to Nrf2 and nuclear factor κB (NF-κB-mediated inflammatory responses. Andrographolide induced the expression and translocation of Nrf2 from the cytoplasm to the nucleus, thereby activating antioxidant response element (ARE gene transcription and HO-1 expression in murine hippocampal HT22 cells. Andrographolide eliminated intracellular Aβ42 in BV-2 cells and decreased the production of interleukin (IL-6, IL-1β, prostaglandin (PGE2, and nitric oxide (NO because of artificial phagocytic Aβ42. It decreased pNF-κB accumulation in the nucleus and the expression of inducible nitric oxide synthase (i-NOS and cyclooxygenase II (COX-II in the microglial BV-2 cell line. In summary, andrographolide activates Nrf2-mediated HO-1 expression and inhibits Aβ42-overexpressed microglial BV-2 cell activation. These results suggested that andrographolide might have the potential for further examination of the therapeutics of AD.

  14. Hyperglycemia in Streptozotocin-Induced Diabetes Leads to Persistent Inflammation and Tissue Damage Following Uveitis Due to Reduced Levels of Ciliary Body Heme Oxygenase-1

    2006-01-01

    Full Text Available This study investigated the heme oxygenase-1 (HO-1 and the endotoxin-induced uveitis (EIU in diabetic streptozotocin (STZ-hyperglycemic rats. STZ-hyperglycemic rats had impaired levels of the enzyme HO-1 within the ciliary bodies if compared with the nondiabetic rats. STZ-hyperglycemic rats also predisposed the eye to produce high levels of both the cytokines IL-1 β and CXCL8. Subsequent EIU further and significantly P<.01 increased the cytokines production, an effect partly prevented by hemin treatment. Most importantly, hemin, an inducer of heme oxygenase expression and activity, recovered the huge number of infiltrated polymorphonuclear leukocytes PMN within the ciliary bodies associated with STZ-hyperglycemic state and EIU damage. Impairment of the stress-sensitive enzyme HO-1 in STZ-hyperglycemic rats increases and prolongs the inflammatory response to EIU.

  15. Differential induction of heme oxygenase and other stress proteins in cultured hippocampal astrocytes and neurons by inorganic lead

    Cabell, Leigh; Ferguson, Charles; Luginbill, Deana; Kern, Marcey; Weingart, Adam; Audesirk, Gerald

    2004-01-01

    We examined the effects of exposure to inorganic lead (Pb 2+ ) on the induction of stress proteins in cultured hippocampal neurons and astrocytes, with particular emphasis on the induction of heme oxygenase-1 (HO-1). In radiolabeled neuronal cultures, Pb 2+ exposure had no significant effect on the synthesis of any protein at any concentration (up to 250 μM) or duration of exposure (up to 4 days). In radiolabeled astrocyte cultures, however, Pb 2+ exposure (100 nM to 100 μM; 1-4 days) increased synthesis of proteins with approximate molecular weights of 23, 32, 45, 57, 72, and 90 kDa. Immunoblot experiments showed that Pb 2+ exposure (100 nM to 10 μM, 1-14 days) induces HO-1 synthesis in astrocytes, but not in neurons; this is probably the 32-kDa protein. The other heme oxygenase isoform, HO-2, is present in both neurons and astrocytes, but is not inducible by Pb 2+ at concentrations up to 100 μM. HO-1 can be induced by a variety of stimuli. We found that HO-1 induction in astrocytes is increased by combined exposure to Pb 2+ and many other stresses, including heat, nitric oxide, H 2 O 2 , and superoxide. One of the stimuli that may induce HO-1 is oxidative stress. Lead exposure causes oxidative stress in many cell types, including astrocytes. Induction of HO-1 by Pb 2+ is reduced by the hydroxyl radical scavengers dimethylthiourea (DMTU) and mannitol, but not by inhibitors of calmodulin, calmodulin-dependent protein kinases, protein kinase C, or extracellular signal-regulated kinases (ERK). Therefore, we conclude that oxidative stress is an important mechanism by which Pb 2+ induces HO-1 synthesis in astrocytes

  16. Critical role of heme oxygenase-1 in Foxp3-mediated immune suppression

    Choi, Byung-Min; Pae, Hyun-Ock; Jeong, Young-Ran; Kim, Young-Myeong; Chung, Hun-Taeg

    2005-01-01

    Foxp3, which encodes the transcription factor scurfin, is indispensable for the development and function of CD4 + CD25 + regulatory T cells (Treg). Recent data suggest conversion of peripheral CD4 + CD25 - naive T cells to CD4 + CD25 + Treg by acquisition of Foxp3 through costimulation with TCR and TGF-β or forced expression of the gene. One critical question is how Foxp3 causes T cells to become regulatory. In the present work, we demonstrate that Foxp3 can induce heme oxygenase-1 (HO-1) expression and subsequently such regulatory phenotypes as the suppression of nontransfected cells in a cell-cell contact-dependent manner as well as impaired proliferation and production of cytokines upon stimulation in Jurkat T cells. Moreover, we confirm the expression of both Foxp3 and HO-1 in peripheral CD4 + CD25 + Treg and suppressive function of the cells are relieved by the inhibition of HO-1 activity. In summary, we demonstrate that Foxp3 induces HO-1 expression and HO-1 engages in Foxp3-mediated immune suppression

  17. Heme Oxygenase-1 Gene Therapy Provides Cardioprotection Via Control of Post-Ischemic Inflammation: An Experimental Study in a Pre-Clinical Pig Model.

    Hinkel, Rabea; Lange, Philipp; Petersen, Björn; Gottlieb, Elena; Ng, Judy King Man; Finger, Stefanie; Horstkotte, Jan; Lee, Seungmin; Thormann, Michael; Knorr, Maike; El-Aouni, Chiraz; Boekstegers, Peter; Reichart, Bruno; Wenzel, Philip; Niemann, Heiner; Kupatt, Christian

    2015-07-14

    Heme oxygenase-1 (HO-1) is an inducible stress-responsive enzyme converting heme to bilirubin, carbon monoxide, and free iron, which exerts anti-inflammatory and antiapoptotic effects. Although efficient cardioprotection after HO-1 overexpression has been reported in rodents, its role in attenuating post-ischemic inflammation is unclear. This study assessed the efficacy of recombinant adenoassociated virus (rAAV)-encoding human heme oxygenase-1 (hHO-1) in attenuating post-ischemic inflammation in a murine and a porcine ischemia/reperfusion model. Murine ischemia was induced by 45 min of left anterior descending occlusion, followed by 24 h of reperfusion and functional as well as fluorescent-activated cell sorting analysis. Porcine hearts were subjected to 60 min of ischemia and 24h of reperfusion before hemodynamic and histologic analyses were performed. Human microvascular endothelial cells transfected with hHO-1 displayed an attenuated interleukin-6 and intercellular adhesion molecule 1 expression, resulting in reduced monocytic THP-1 cell recruitment in vitro. In murine left anterior descending occlusion and reperfusion, the post-ischemic influx of CD45(+) leukocytes, Ly-6G(+) neutrophils, and Ly-6C(high) monocytes was further exacerbated in HO-1-deficient hearts and reversed by rAAV.hHO-1 treatment. Conversely, in our porcine model of ischemia, the post-ischemic influx of myeloperoxidase-positive neutrophils and CD14(+) monocytes was reduced by 49% and 87% after rAAV.hHO-1 transduction, similar to hHO-1 transgenic pigs. Functionally, rAAV.hHO-1 and hHO-1 transgenic left ventricles displayed a smaller loss of ejection fraction than control animals. Whereas HO-1 deficiency exacerbates post-ischemic cardiac inflammation in mice, hHO-1 gene therapy attenuates inflammation after ischemia and reperfusion in murine and porcine hearts. Regional hHO-1 gene therapy provides cardioprotection in a pre-clinical porcine ischemia/reperfusion model. Copyright © 2015 American

  18. Induction of Protective Genes Leads to Islet Survival and Function

    Hongjun Wang

    2011-01-01

    Full Text Available Islet transplantation is the most valid approach to the treatment of type 1 diabetes. However, the function of transplanted islets is often compromised since a large number of β cells undergo apoptosis induced by stress and the immune rejection response elicited by the recipient after transplantation. Conventional treatment for islet transplantation is to administer immunosuppressive drugs to the recipient to suppress the immune rejection response mounted against transplanted islets. Induction of protective genes in the recipient (e.g., heme oxygenase-1 (HO-1, A20/tumor necrosis factor alpha inducible protein3 (tnfaip3, biliverdin reductase (BVR, Bcl2, and others or administration of one or more of the products of HO-1 to the donor, the islets themselves, and/or the recipient offers an alternative or synergistic approach to improve islet graft survival and function. In this perspective, we summarize studies describing the protective effects of these genes on islet survival and function in rodent allogeneic and xenogeneic transplantation models and the prevention of onset of diabetes, with emphasis on HO-1, A20, and BVR. Such approaches are also appealing to islet autotransplantation in patients with chronic pancreatitis after total pancreatectomy, a procedure that currently only leads to 1/3 of transplanted patients being diabetes-free.

  19. Pharmacological Induction of Heme Oxygenase-1 Impairs Nuclear Accumulation of Herpes Simplex Virus Capsids upon Infection

    Francisco J. Ibáñez

    2017-10-01

    Full Text Available Heme oxygenase-1 (HO-1 is an inducible enzyme that is expressed in response to physical and chemical stresses, such as ultraviolet radiation, hyperthermia, hypoxia, reactive oxygen species (ROS, as well as cytokines, among others. Its activity can be positively modulated by cobalt protoporphyrin (CoPP and negatively by tin protoporphirin (SnPP. Once induced, HO-1 degrades iron-containing heme into ferrous iron (Fe2+, carbon monoxide (CO and biliverdin. Importantly, numerous products of HO-1 are cytoprotective with anti-apoptotic, anti-oxidant, anti-inflammatory, and anti-cancer effects. The products of HO-1 also display antiviral properties against several viruses, such as the human immunodeficiency virus (HIV, influenza, hepatitis B, hepatitis C, and Ebola virus. Here, we sought to assess the effect of modulating HO-1 activity over herpes simplex virus type 2 (HSV-2 infection in epithelial cells and neurons. There are no vaccines against HSV-2 and treatment options are scarce in the immunosuppressed, in which drug-resistant variants emerge. By using HSV strains that encode structural and non-structural forms of the green fluorescent protein (GFP, we found that pharmacological induction of HO-1 activity with CoPP significantly decreases virus plaque formation and the expression of virus-encoded genes in epithelial cells as determined by flow cytometry and western blot assays. CoPP treatment did not affect virus binding to the cell surface or entry into the cytoplasm, but rather downstream events in the virus infection cycle. Furthermore, we observed that treating cells with a CO-releasing molecule (CORM-2 recapitulated some of the anti-HSV effects elicited by CoPP. Taken together, these findings indicate that HO-1 activity interferes with the replication cycle of HSV and that its antiviral effects can be recapitulated by CO.

  20. Plasma heme oxygenase-1 concentration is elevated in individuals with type 2 diabetes mellitus.

    Wei Bao

    Full Text Available BACKGROUND: Circulating concentrations of heme oxygenase-1 (HO-1 have been recently reported to be elevated in several chronic disorders. However, no study has ever examined the association between circulating HO-1 concentrations and type 2 diabetes mellitus (T2DM. METHODS AND FINDINGS: 581 cases with newly-diagnosed T2DM (New-T2DM and 611 comparison controls were recruited in this two-phase case-control study, comprising 420 cases and 429 controls collected in the first phase study and 161 cases and 182 controls in the second phase replication study. Analyses, using both separated data and combined data from the two-phase studies, show that plasma HO-1 concentrations were significantly increased in New-T2DM cases compared to controls (P<0.001. Plasma HO-1 concentrations were significantly correlated with plasma glucose concentrations, HOMA-beta and HOMA-IR (P<0.001. After adjustment for age, sex, BMI and family history of diabetes, the ORs for New-T2DM in the highest quartile of plasma HO-1 concentrations, compared with the lowest, was 8.23 (95% CI 5.55-12.21; P for trend <0.001. The trend remained significant after additional adjustment for fasting plasma glucose/insulin, HOMA-beta/HOMA-IR, TC/TG, smoking, drinking and history of hypertension, and even in further stratification analysis by age, sex, BMI, smoking, drinking and history of hypertension. CONCLUSIONS: Elevated plasma HO-1 concentrations are associated with higher ORs for New-T2DM, which add more knowledge regarding the important role of oxidative stress in T2DM. More consequent studies were warranted to confirm the clinical utility of plasma HO-1, especially in diagnosis and prognosis of T2DM and its complications.

  1. Heme oxygenase-1 gene expression modulates angiotensin II-induced increase in blood pressure.

    Yang, Liming; Quan, Shuo; Nasjletti, Alberto; Laniado-Schwartzman, Michal; Abraham, Nader G

    2004-06-01

    The heme-heme oxygenase (HO) system has been implicated in the regulation of vascular reactivity and blood pressure. This study examines the notion that overexpression of HO decreases pressor responsiveness to angiotensin II (Ang II). Five-day-old Sprague-Dawley rats received an intraleft ventricular injection of approximately 5x10(9) cfu/mL of retroviruses containing human HO-1 sense (LSN-HHO-1), rat HO-1 antisense (LSN-RHO-1-AS), or control retrovirus (LXSN). Three months later, rats were instrumented with femoral arterial and venous catheters for mean arterial pressure (MAP) determination and Ang II administration, respectively. Rats injected with LSN-HHO-1, but not with LXSN, expressed human HO-1 mRNA and protein in several tissues. BP increased with administration of Ang II in rats expressing and not expressing human HO-1. However, the Ang II-induced pressor response (mm Hg) in LSN-HHO-1 rats (16+/-3, 27+/-3, and 38+/-3 at 0.5, 2, and 10 ng) was surpassed (PHHO-1 rats with the HO inhibitor tin mesoporphyrin (SnMP) enhanced (P<0.05) the Ang II-induced pressor response to a level not different from that observed in LXSN rats. Rats injected with LSN-RHO-1-AS showed a decrease in renal HO-1 protein expression and HO activity relative to control LXSN rats. Administration of Ang II (0.1 to 2 ng) caused small (4 to 5 mm Hg) but significant increases in MAP in rats injected with LSN-RHO-1-AS (P<0.05) compared with rats injected with LXSN. These data demonstrate that overexpression of HO-1 brings about a reduction in pressor responsiveness to Ang II, which is most likely due to increased generation of an HO-1 product, presumably CO, with the ability to inhibit vascular reactivity to constrictor stimuli.

  2. Anti-inflammatory and heme oxygenase-1 inducing activities of lanostane triterpenes isolated from mushroom Ganoderma lucidum in RAW264.7 cells

    Choi, Solip [Department of Biochemistry, College of Natural Sciences, Kangwon National University, Chuncheon, Gangwon-Do 200-701 (Korea, Republic of); Nguyen, Van Thu [College of Pharmacy, Catholic University of Daegu, Gyeongsan 712-702 (Korea, Republic of); Tae, Nara; Lee, Suhyun [Department of Biochemistry, College of Natural Sciences, Kangwon National University, Chuncheon, Gangwon-Do 200-701 (Korea, Republic of); Ryoo, Sungwoo [Department of Biological Sciences, College of Natural Sciences, Kangwon National University, Chuncheon, Gangwon-Do 200-701 (Korea, Republic of); Min, Byung-Sun [College of Pharmacy, Catholic University of Daegu, Gyeongsan 712-702 (Korea, Republic of); Lee, Jeong-Hyung, E-mail: jhlee36@kangwon.ac.kr [Department of Biochemistry, College of Natural Sciences, Kangwon National University, Chuncheon, Gangwon-Do 200-701 (Korea, Republic of)

    2014-11-01

    Ganoderma lucidum is a popular medicinal mushroom used in traditional medicine for preventing or treating a variety of diseases. In the present study, we investigated the anti-inflammatory and heme oxygenase (HO)-1 inducing effects of 12 lanostane triterpenes from G. lucidum in RAW264.7 cells. Of these, seven triterpenes, butyl lucidenateE{sub 2}, butyl lucidenateD{sub 2} (GT-2), butyl lucidenate P, butyl lucidenateQ, Ganoderiol F, methyl ganodenate J and butyl lucidenate N induced HO-1 expression and suppressed lipopolysaccharide (LPS)-induced nitric oxide (NO) production. Inhibiting HO-1 activity abrogated the inhibitory effects of these triterpenes on the production of NO in LPS-stimulated RAW264.7 cells, suggesting the involvement of HO-1 in the anti-inflammatory effects of these triterpenes. We further studied the anti-inflammatory and HO-1 inducing effects of GT-2. Mitogen-activated protein kinase inhibitors or N-acetylcysteine, an antioxidant, did not suppress GT-2-mediated HO-1 induction; however, LY294002, a phosphoinositide 3-kinase (PI3K) inhibitor, blocked GT-2-induced HO-1 mRNA and protein expression. GT-2 increased nuclear translocation of nuclear factor-E2-related factor 2 (Nrf2) and knockdown of Nrf2 by small interfering RNA blocked GT-2-mediated HO-1 induction, suggesting that GT-2 induced HO-1 expression via the PI3K/AKT-Nrf2 pathway. Consistent with the notion that HO-1 has anti-inflammatory properties, GT-2 inhibited the production of tumor necrosis factor-α and interleukin-6, as well as inducible nitric oxide synthase and cyclooxygenase-2 expression. These findings suggest that HO-1 inducing activities of these lanostane triterpenes may be important in the understanding of a novel mechanism for the anti-inflammatory activity of G. lucidum. - Highlights: • The anti-inflammatory effects of selected triterpenes from Ganoderma lucidum are demonstrated. • Heme oxygenase-1 induction is attributable to the anti-inflammatory properties of these

  3. Anti-inflammatory and heme oxygenase-1 inducing activities of lanostane triterpenes isolated from mushroom Ganoderma lucidum in RAW264.7 cells

    Choi, Solip; Nguyen, Van Thu; Tae, Nara; Lee, Suhyun; Ryoo, Sungwoo; Min, Byung-Sun; Lee, Jeong-Hyung

    2014-01-01

    Ganoderma lucidum is a popular medicinal mushroom used in traditional medicine for preventing or treating a variety of diseases. In the present study, we investigated the anti-inflammatory and heme oxygenase (HO)-1 inducing effects of 12 lanostane triterpenes from G. lucidum in RAW264.7 cells. Of these, seven triterpenes, butyl lucidenateE 2 , butyl lucidenateD 2 (GT-2), butyl lucidenate P, butyl lucidenateQ, Ganoderiol F, methyl ganodenate J and butyl lucidenate N induced HO-1 expression and suppressed lipopolysaccharide (LPS)-induced nitric oxide (NO) production. Inhibiting HO-1 activity abrogated the inhibitory effects of these triterpenes on the production of NO in LPS-stimulated RAW264.7 cells, suggesting the involvement of HO-1 in the anti-inflammatory effects of these triterpenes. We further studied the anti-inflammatory and HO-1 inducing effects of GT-2. Mitogen-activated protein kinase inhibitors or N-acetylcysteine, an antioxidant, did not suppress GT-2-mediated HO-1 induction; however, LY294002, a phosphoinositide 3-kinase (PI3K) inhibitor, blocked GT-2-induced HO-1 mRNA and protein expression. GT-2 increased nuclear translocation of nuclear factor-E2-related factor 2 (Nrf2) and knockdown of Nrf2 by small interfering RNA blocked GT-2-mediated HO-1 induction, suggesting that GT-2 induced HO-1 expression via the PI3K/AKT-Nrf2 pathway. Consistent with the notion that HO-1 has anti-inflammatory properties, GT-2 inhibited the production of tumor necrosis factor-α and interleukin-6, as well as inducible nitric oxide synthase and cyclooxygenase-2 expression. These findings suggest that HO-1 inducing activities of these lanostane triterpenes may be important in the understanding of a novel mechanism for the anti-inflammatory activity of G. lucidum. - Highlights: • The anti-inflammatory effects of selected triterpenes from Ganoderma lucidum are demonstrated. • Heme oxygenase-1 induction is attributable to the anti-inflammatory properties of these triterpenes

  4. Antioxidant mechanism of heme oxygenase-1 involves an increase in superoxide dismutase and catalase in experimental diabetes.

    Turkseven, Saadet; Kruger, Adam; Mingone, Christopher J; Kaminski, Pawel; Inaba, Muneo; Rodella, Luigi F; Ikehara, Susumu; Wolin, Michael S; Abraham, Nader G

    2005-08-01

    Increased heme oxygenase (HO)-1 activity attenuates endothelial cell apoptosis and decreases superoxide anion (O2-) formation in experimental diabetes by unknown mechanisms. We examined the effect of HO-1 protein and HO activity on extracellular SOD (EC-SOD), catalase, O2-, inducible nitric oxide synthase (iNOS), and endothelial nitric oxide synthase (eNOS) levels and vascular responses to ACh in control and diabetic rats. Vascular EC-SOD and plasma catalase activities were significantly reduced in diabetic compared with nondiabetic rats (P inhibitor of HO-1 activity, decreased EC-SOD protein. Increased HO-1 activity in diabetic rats was associated with a decrease in iNOS but increases in eNOS and plasma catalase activity. On the other hand, aortic ring segments from diabetic rats exhibited a significant reduction in vascular relaxation to ACh, which was reversed with cobalt protoporphyrin treatment. These data demonstrate that an increase in HO-1 protein and activity, i.e., CO and bilirubin production, in diabetic rats brings about a robust increase in EC-SOD, catalase, and eNOS with a concomitant increase in endothelial relaxation and a decrease in O2-. These observations in experimental diabetes suggest that the vascular cytoprotective mechanism of HO-1 against oxidative stress requires an increase in EC-SOD and catalase.

  5. Omeprazole induces heme oxygenase-1 in fetal human pulmonary microvascular endothelial cells via hydrogen peroxide-independent Nrf2 signaling pathway

    Patel, Ananddeep; Zhang, Shaojie; Shrestha, Amrit Kumar; Maturu, Paramahamsa; Moorthy, Bhagavatula; Shivanna, Binoy

    2016-01-01

    Omeprazole (OM) is an aryl hydrocarbon receptor (AhR) agonist and a proton pump inhibitor that is used to treat humans with gastric acid related disorders. Recently, we showed that OM induces NAD (P) H quinone oxidoreductase-1 (NQO1) via nuclear factor erythroid 2-related factor 2 (Nrf2)-dependent mechanism. Heme oxygenase-1 (HO-1) is another cytoprotective and antioxidant enzyme that is regulated by Nrf2. Whether OM induces HO-1 in fetal human pulmonary microvascular endothelial cells (HPMEC) is unknown. Therefore, we tested the hypothesis that OM will induce HO-1 expression via Nrf2 in HPMEC. OM induced HO-1 mRNA and protein expression in a dose-dependent manner. siRNA-mediated knockdown of AhR failed to abrogate, whereas knockdown of Nrf2 abrogated HO-1 induction by OM. To identify the underlying molecular mechanisms, we determined the effects of OM on cellular hydrogen peroxide (H 2 O 2 ) levels since oxidative stress mediated by the latter is known to activate Nrf2. Interestingly, the concentration at which OM induced HO-1 also increased H 2 O 2 levels. Furthermore, H 2 O 2 independently augmented HO-1 expression. Although N-acetyl cysteine (NAC) significantly decreased H 2 O 2 levels in OM-treated cells, we observed that OM further increased HO-1 mRNA and protein expression in NAC-pretreated compared to vehicle-pretreated cells, suggesting that OM induces HO-1 via H 2 O 2 -independent mechanisms. In conclusion, we provide evidence that OM transcriptionally induces HO-1 via AhR - and H 2 O 2 - independent, but Nrf2 - dependent mechanisms. These results have important implications for human disorders where Nrf2 and HO-1 play a beneficial role. - Highlights: • Omeprazole induces HO-1 in human fetal lung cells. • AhR deficiency fails to abrogate omeprazole-mediated induction of HO-1. • Nrf2 knockdown abrogates omeprazole-mediated HO-1 induction in human lung cells. • Hydrogen peroxide depletion augments omeprazole-mediated induction of HO-1.

  6. Omeprazole induces heme oxygenase-1 in fetal human pulmonary microvascular endothelial cells via hydrogen peroxide-independent Nrf2 signaling pathway

    Patel, Ananddeep; Zhang, Shaojie; Shrestha, Amrit Kumar; Maturu, Paramahamsa; Moorthy, Bhagavatula; Shivanna, Binoy, E-mail: shivanna@bcm.edu

    2016-11-15

    Omeprazole (OM) is an aryl hydrocarbon receptor (AhR) agonist and a proton pump inhibitor that is used to treat humans with gastric acid related disorders. Recently, we showed that OM induces NAD (P) H quinone oxidoreductase-1 (NQO1) via nuclear factor erythroid 2-related factor 2 (Nrf2)-dependent mechanism. Heme oxygenase-1 (HO-1) is another cytoprotective and antioxidant enzyme that is regulated by Nrf2. Whether OM induces HO-1 in fetal human pulmonary microvascular endothelial cells (HPMEC) is unknown. Therefore, we tested the hypothesis that OM will induce HO-1 expression via Nrf2 in HPMEC. OM induced HO-1 mRNA and protein expression in a dose-dependent manner. siRNA-mediated knockdown of AhR failed to abrogate, whereas knockdown of Nrf2 abrogated HO-1 induction by OM. To identify the underlying molecular mechanisms, we determined the effects of OM on cellular hydrogen peroxide (H{sub 2}O{sub 2}) levels since oxidative stress mediated by the latter is known to activate Nrf2. Interestingly, the concentration at which OM induced HO-1 also increased H{sub 2}O{sub 2} levels. Furthermore, H{sub 2}O{sub 2} independently augmented HO-1 expression. Although N-acetyl cysteine (NAC) significantly decreased H{sub 2}O{sub 2} levels in OM-treated cells, we observed that OM further increased HO-1 mRNA and protein expression in NAC-pretreated compared to vehicle-pretreated cells, suggesting that OM induces HO-1 via H{sub 2}O{sub 2}-independent mechanisms. In conclusion, we provide evidence that OM transcriptionally induces HO-1 via AhR - and H{sub 2}O{sub 2} - independent, but Nrf2 - dependent mechanisms. These results have important implications for human disorders where Nrf2 and HO-1 play a beneficial role. - Highlights: • Omeprazole induces HO-1 in human fetal lung cells. • AhR deficiency fails to abrogate omeprazole-mediated induction of HO-1. • Nrf2 knockdown abrogates omeprazole-mediated HO-1 induction in human lung cells. • Hydrogen peroxide depletion augments

  7. Glaucocalyxin B Alleviates Lipopolysaccharide-Induced Parkinson’s Disease by Inhibiting TLR/NF-κB and Activating Nrf2/HO-1 Pathway

    Wei Xu

    2017-12-01

    Full Text Available Background/Aims: Parkinson’s disease (PD is a common neurodegenerative disease in the old population, characterized by dopaminergic neuron loss, inflammation and oxidative stress injury in the substantia nigra. Glaucocalyxin B (GLB, an ent-kauranoid diterpenoid isolated from Rabdosia japonica, has anti-inflammation and anti-tumor effects. However, its effects on PD remain unclear. Methods: PD was introduced in rats via injection of lipopolysaccharide (LPS into cerebral corpus striatum, and GLB was given intracerebroventricularly to these rats. Their walking, climbing and sensory states were detected by Stepping, Whisker and Cylinder Tests. The expression of tyrosine hydroxylase (TH, glial fibrillary acidic protein (GFAP, CD11b and ionized calcium binding adaptor molecule (IBA-1 were detected by immunohischemical staining. The levels of a series of inflammatory factors, oxidative stress-related factors and apoptosis-related factors were measured by real-time PCR, immunoblotting and ELISA. In addition, Toll-like receptor (TLR/nuclear factor kappa B (NF-κB and nuclear factor erythroid 2-related factor 2 (Nrf2/heme oxygenase (HO-1 pathways were investigated to illustrate the underlying mechanism. In vitro, microglial cells exposed to LPS were treated with GLB. Results: The injection of LPS caused walking, climbing and sensory disturbances in rats, induced inflammation, oxidative stress response and apoptosis, and activated TLR/NF-κB and Nrf2/ HO-1 pathways in the cerebral tissue. GLB administration attenuated LPS-induced alterations. The TLR/NF-κB pathway was deactivated and Nrf2/HO-1 was activated after application of GLB. In vitro, cytotoxic effects induced by the conditioned medium derived from microglial cells exposed to LPS in PC12 cells were attenuated by GLB. Conclusion: GLB suppresses LPS-induced PD symptoms by modification of TLR/NF-κB and Nrf2/HO-1 pathways in vivo and in vitro.

  8. Identification of heme oxygenase-1 stimulators by a convenient ELISA-based bilirubin quantification assay.

    Rücker, Hannelore; Amslinger, Sabine

    2015-01-01

    The upregulation of heme oxygenase-1 (HO-1) has proven to be a useful tool for fighting inflammation. In order to identify new HO-1 inducers, an efficient screening method was developed which can provide new lead structures for drug research. We designed a simple ELISA-based HO-1 enzyme activity assay, which allows for the screening of 12 compounds in parallel in the setting of a 96-well plate. The well-established murine macrophage cell line RAW264.7 is used and only about 26µg of protein from whole cell lysates is needed for the analysis of HO-1 activity. The quantification of HO-1 activity is based on an indirect ELISA using the specific anti-bilirubin antibody 24G7 to quantify directly bilirubin in the whole cell lysate, applying a horseradish peroxidase-tagged antibody together with ortho-phenylenediamine and H2O2 for detection. The bilirubin is produced on the action of HO enzymes by converting their substrate heme to biliverdin and additional recombinant biliverdin reductase together with NADPH at pH 7.4 in buffer. This sensitive assay allows for the detection of 0.57-82pmol bilirubin per sample in whole cell lysates. Twenty-three small molecules, mainly natural products with an α,β-unsaturated carbonyl unit such as polyphenols, including flavonoids and chalcones, terpenes, an isothiocyanate, and the drug oltipraz were tested at typically 6 or 24h incubation with RAW264.7 cells. The activity of known HO-1 inducers was confirmed, while the chalcones cardamonin, flavokawain A, calythropsin, 2',3,4'-trihydroxy-4-methoxychalcone (THMC), and 2',4'-dihydroxy-3,4-dimethoxychalcone (DHDMC) were identified as new potent HO-1 inducers. The highest inductive power after 6h incubation was found at 10µM for DHDMC (6.1-fold), carnosol (3.9-fold), butein (3.1-fold), THMC (2.9-fold), and zerumbone (2.5-fold). Moreover, the time dependence of HO-1 protein production for DHDMC was compared to its enzyme activity, which was further evaluated in the presence of

  9. High Glucose Promotes Tumor Invasion and Increases Metastasis-Associated Protein Expression in Human Lung Epithelial Cells by Upregulating Heme Oxygenase-1 via Reactive Oxygen Species or the TGF-β1/PI3K/Akt Signaling Pathway

    Xiaowen Kang

    2015-02-01

    Full Text Available Background: Growing evidence indicates that heme oxygenase-1 (HO-1 is up-regulated in malignancies and subsequently alters tumor aggressiveness and various cancer-related factors, such as high glucose (HG levels. HO-1 expression can be induced when glucose concentrations are above 25 mM; however, the role of HO-1 in lung cancer patients with diabetes remains unknown. Therefore, in this study we investigated the promotion of tumor cell invasion and the expression of metastasis-associated proteins by inducing the up-regulation of HO-1 expression by HG treatment in A549 human lung epithelial cells. Methods: The expression of HO-1and metastasis-associated protein expression was explored by western blot analysis. HO-1 enzymatic activity, reactive oxygen species (ROS production and TGF-β1 production were examined by ELISA. Invasiveness was analyzed using a Transwell chamber. Results: HG treatment of A549 cells induced an increase in HO-1 expression, which was mediated by the HG-induced generation of reactive oxygen species (ROS and transforming growth factor-β1 (TGF-β1 in a concentration- and time-dependent manner. Following the increase in HO-1 expression, the enzymatic activity of HO-1 also increased in HG-treated cells. Pretreatment with N-acetyl-L-cysteine (NAC or with phosphatidylinositol 3-kinase (PI3K/Akt inhibitors attenuated the HG-induced increase in HO-1 expression. HG treatment of A549 cells enhanced the invasion potential of these cells, as shown with a Transwell assay, and increased metastasis-associated protein expression. However, HO-1 siRNA transfection significantly decreased these capabilities. Conclusion: this study is the first to demonstrate that HG treatment of A549 human lung epithelial cells promotes tumor cell invasion and increases metastasis-associated protein expression by up-regulating HO-1 expression via ROS or the TGF-β1/PI3K/Akt signaling pathway.

  10. Impact of Bariatric Surgery on Heme Oxygenase-1, Inflammation, and Insulin Resistance in Morbid Obesity with Obstructive Sleep Apnea.

    Tirado, Raquel; Masdeu, Maria José; Vigil, Laura; Rigla, Mercedes; Luna, Alexis; Rebasa, Pere; Pareja, Rocío; Hurtado, Marta; Caixàs, Assumpta

    2017-09-01

    Morbid obesity and obstructive sleep apnea (OSA) interact at an inflammatory level. Bariatric surgery reduces inflammatory responses associated with obesity. Heme oxygenase-1 (HO-1) is an enzyme with anti-inflammatory properties, which might be increased in morbid obesity or OSA. We studied morbidly obese patients with OSA to determine: (a) HO-1 plasma concentrations according to OSA severity and their relationship with insulin resistance and inflammation and (b) the impact of bariatric surgery on HO-1 and parameters of insulin resistance and inflammation. We analyzed the homeostasis model insulin resistance index (HOMA) and plasma concentrations of HO-1, tumor necrosis factor alpha, interleukin-6, interleukin-1-beta, C reactive protein (CRP), and adiponectin according to polysomnography findings in 66 morbidly obese patients before bariatric surgery and 12 months after surgery. Before surgery, HO-1 plasma concentrations were similar in three groups of patients with mild, moderate, and severe OSA, and correlated with HOMA (r = 0.27, p = 0.02). Twelve months after surgery, low-grade inflammation and insulin resistance had decreased in all the groups, but HO-1 plasma concentration had decreased only in the severe OSA group (p = 0.02). In this group, the reduction in HO-1 correlated with a reduction in CRP concentrations (r = 0.43, p = 0.04) and with improved HOMA score (r = 0.37, p = 0.03). Bariatric surgery decreases HO-1 concentrations in morbid obesity with severe OSA, and this decrease is associated with decreases in insulin resistance and in inflammation.

  11. Atractylenolide I restores HO-1 expression and inhibits Ox-LDL-induced VSMCs proliferation, migration and inflammatory responses in vitro

    Li, Weifeng, E-mail: liwf@mail.xjtu.edu.cn; Zhi, Wenbing; Liu, Fang; He, Zehong; Wang, Xiuei; Niu, Xiaofeng, E-mail: niuxf@mail.xjtu.edu.cn

    2017-04-01

    Pathogenesis of atherosclerosis is characterized by the proliferation and migration of vascular smooth muscle cells (VSMCs) and inflammatory lesions. The aim of this study is to elucidate the effect of atractylenolide I (AO-I) on smooth muscle cell inflammation, proliferation and migration induced by oxidized modified low density lipoprotein (Ox-LDL). Here, We found that atractylenolide I inhibited Ox-LDL-induced VSMCs proliferation and migration in a dose-dependent manner, and decreased the production of inflammatory cytokines and the expression of monocyte chemoattractant protein-1 (MCP-1) in VSMCs. The study also identified that AO-I prominently inhibited p38-MAPK and NF-κB activation. More importantly, the specific heme oxygenase-1 (HO-1) inhibitor zinc protoporphyrin (ZnPP) IX partially abolished the beneficial effects of atractylenolide I on Ox-LDL-induced VSMCs. Furthermore, atractylenolide I blocked the foam cell formation in macrophages induced by Ox-LDL. In summary, inhibitory roles of AO-I in VSMCs proliferation and migration, lipid peroxidation and subsequent inflammatory responses might contribute to the anti-atherosclerotic property of AO-I. - Highlights: • AO-I inhibited Ox-LDL-induced VSMCs proliferation and migration. • AO-I alleviated inflammatory response via inhibiting TNF-α, IL-6 and NO production. • AO-I restored HO-1 expression and down-regulated PCNA expression. • MCP-1 overexpression is potentially regulated by NF-κB and p38 MAPK pathway. • AO-I possesses strong anti-lipid peroxidation effect.

  12. Methane-rich water induces cucumber adventitious rooting through heme oxygenase1/carbon monoxide and Ca(2+) pathways.

    Cui, Weiti; Qi, Fang; Zhang, Yihua; Cao, Hong; Zhang, Jing; Wang, Ren; Shen, Wenbiao

    2015-03-01

    Methane-rich water triggered adventitious rooting by regulating heme oxygenase1/carbon monoxide and calcium pathways in cucumber explants. Heme oxygenase1/carbon monoxide (HO1/CO) and calcium (Ca(2+)) were reported as the downstream signals in auxin-induced cucumber adventitious root (AR) formation. Here, we observed that application of methane-rich water (MRW; 80% saturation) obviously induced AR formation in IAA-depleted cucumber explants. To address the universality, we checked adventitious rooting in soybean and mung bean explants, and found that MRW (50 and 10% saturation, respectively) exhibited the similar inducing results. To further determine if the HO1/CO system participated in MRW-induced adventitious rooting, MRW, HO1 inducer hemin, its activity inhibitor zinc protoporphyrin IX (ZnPP), and its catalytic by-products CO, bilirubin, and Fe(2+) were used to detect their effects on cucumber adventitious rooting in IAA-depleted explants. Subsequent results showed that MRW-induced adventitious rooting was blocked by ZnPP and further reversed by 20% saturation CO aqueous solution. However, the other two by-products of HO1, bilirubin and Fe(2+), failed to induce AR formation. Above responses were consistent with the MRW-induced increases of HO1 transcript and corresponding protein level. Further molecular evidence indicted that expression of marker genes, including auxin signaling-related genes and cell cycle regulatory genes, were modulated by MRW alone but blocked by the cotreatment with ZnPP, the latter of which could be significantly rescued by the addition of CO. By using the Ca(2+)-channel blocker and Ca(2+) chelator, the involvement of Ca(2+) pathway in MRW-induced adventitious rooting was also suggested. Together, our results indicate that MRW might serve as a stimulator of adventitious rooting, which was partially mediated by HO1/CO and Ca(2+) pathways.

  13. Heme Oxygenase-1/Carbon Monoxide System and Embryonic Stem Cell Differentiation and Maturation into Cardiomyocytes

    Suliman, Hagir B.; Zobi, Fabio

    2016-01-01

    Abstract Aims: The differentiation of embryonic stem (ES) cells into energetically efficient cardiomyocytes contributes to functional cardiac repair and is envisioned to ameliorate progressive degenerative cardiac diseases. Advanced cell maturation strategies are therefore needed to create abundant mature cardiomyocytes. In this study, we tested whether the redox-sensitive heme oxygenase-1/carbon monoxide (HO-1/CO) system, operating through mitochondrial biogenesis, acts as a mechanism for ES cell differentiation and cardiomyocyte maturation. Results: Manipulation of HO-1/CO to enhance mitochondrial biogenesis demonstrates a direct pathway to ES cell differentiation and maturation into beating cardiomyocytes that express adult structural markers. Targeted HO-1/CO interventions up- and downregulate specific cardiogenic transcription factors, transcription factor Gata4, homeobox protein Nkx-2.5, heart- and neural crest derivatives-expressed protein 1, and MEF2C. HO-1/CO overexpression increases cardiac gene expression for myosin regulatory light chain 2, atrial isoform, MLC2v, ANP, MHC-β, and sarcomere α-actinin and the major mitochondrial fusion regulators, mitofusin 2 and MICOS complex subunit Mic60. This promotes structural mitochondrial network expansion and maturation, thereby supporting energy provision for beating embryoid bodies. These effects are prevented by silencing HO-1 and by mitochondrial reactive oxygen species scavenging, while disruption of mitochondrial biogenesis and mitochondrial DNA depletion by loss of mitochondrial transcription factor A compromise infrastructure. This leads to failure of cardiomyocyte differentiation and maturation and contractile dysfunction. Innovation: The capacity to augment cardiomyogenesis via a defined mitochondrial pathway has unique therapeutic potential for targeting ES cell maturation in cardiac disease. Conclusion: Our findings establish the HO-1/CO system and redox regulation of mitochondrial biogenesis as

  14. Astroglia overexpressing heme oxygenase-1 predispose co-cultured PC12 cells to oxidative injury.

    Song, Linyang; Song, Wei; Schipper, Hyman M

    2007-08-01

    The mechanisms responsible for the progressive degeneration of dopaminergic neurons and pathologic iron deposition in the substantia nigra pars compacta of patients with Parkinson's disease (PD) remain unclear. Heme oxygenase-1 (HO-1), the rate-limiting enzyme in the oxidative degradation of heme to ferrous iron, carbon monoxide, and biliverdin, is upregulated in affected PD astroglia and may contribute to abnormal mitochondrial iron sequestration in these cells. To determine whether glial HO-1 hyper-expression is toxic to neuronal compartments, we co-cultured dopaminergic PC12 cells atop monolayers of human (h) HO-1 transfected, sham-transfected, or non-transfected primary rat astroglia. We observed that PC12 cells grown atop hHO-1 transfected astrocytes, but not the astroglia themselves, were significantly more susceptible to dopamine (1 microM) + H(2)O(2) (1 microM)-induced death (assessed by nuclear ethidium monoazide bromide staining and anti-tyrosine hydroxylase immunofluorescence microscopy) relative to control preparations. In the experimental group, PC12 cell death was attenuated significantly by the administration of the HO inhibitor, SnMP (1.5 microM), the antioxidant, ascorbate (200 microM), or the iron chelators, deferoxamine (400 microM), and phenanthroline (100 microM). Exposure to conditioned media derived from HO-1 transfected astrocytes also augmented PC12 cell killing in response to dopamine (1 microM) + H(2)O(2) (1 microM) relative to control media. In PD brain, overexpression of HO-1 in nigral astroglia and accompanying iron liberation may facilitate the bioactivation of dopamine to neurotoxic free radical intermediates and predispose nearby neuronal constituents to oxidative damage. (c) 2007 Wiley-Liss, Inc.

  15. Over-expression of heme oxygenase-1 promotes oxidative mitochondrial damage in rat astroglia.

    Song, Wei; Su, Haixiang; Song, Sisi; Paudel, Hemant K; Schipper, Hyman M

    2006-03-01

    Glial heme oxygenase-1 is over-expressed in the CNS of subjects with Alzheimer disease (AD), Parkinson disease (PD) and multiple sclerosis (MS). Up-regulation of HO-1 in rat astroglia has been shown to facilitate iron sequestration by the mitochondrial compartment. To determine whether HO-1 induction promotes mitochondrial oxidative stress, assays for 8-epiPGF(2alpha) (ELISA), protein carbonyls (ELISA) and 8-OHdG (HPLC-EC) were used to quantify oxidative damage to lipids, proteins, and nucleic acids, respectively, in mitochondrial fractions and whole-cell compartments derived from cultured rat astroglia engineered to over-express human (h) HO-1 by transient transfection. Cell viability was assessed by trypan blue exclusion and the MTT assay, and cell proliferation was determined by [3H] thymidine incorporation and total cell counts. In rat astrocytes, hHO-1 over-expression (x 3 days) resulted in significant oxidative damage to mitochondrial lipids, proteins, and nucleic acids, partial growth arrest, and increased cell death. These effects were attenuated by incubation with 1 microM tin mesoporphyrin, a competitive HO inhibitor, or the iron chelator, deferoxamine. Up-regulation of HO-1 engenders oxidative mitochondrial injury in cultured rat astroglia. Heme-derived ferrous iron and carbon monoxide (CO) may mediate the oxidative modification of mitochondrial lipids, proteins and nucleic acids in these cells. Glial HO-1 hyperactivity may contribute to cellular oxidative stress, pathological iron deposition, and bioenergetic failure characteristic of degenerating and inflamed neural tissues and may constitute a rational target for therapeutic intervention in these conditions. Copyright 2005 Wiley-Liss, Inc.

  16. Heme Oxygenase-1 Inhibits HLA Class I Antibody-Dependent Endothelial Cell Activation.

    Eva Zilian

    Full Text Available Antibody-mediated rejection (AMR is a key limiting factor for long-term graft survival in solid organ transplantation. Human leukocyte antigen (HLA class I (HLA I antibodies (Abs play a major role in the pathogenesis of AMR via their interactions with HLA molecules on vascular endothelial cells (ECs. The antioxidant enzyme heme oxygenase (HO-1 has anti-inflammatory functions in the endothelium. As complement-independent effects of HLA I Abs can activate ECs, it was the goal of the current study to investigate the role of HO-1 on activation of human ECs by HLA I Abs. In cell cultures of various primary human macro- and microvascular ECs treatment with monoclonal pan- and allele-specific HLA I Abs up-regulated the expression of inducible proinflammatory adhesion molecules and chemokines (vascular cell adhesion molecule-1 [VCAM-1], intercellular cell adhesion molecule-1 [ICAM-1], interleukin-8 [IL-8] and monocyte chemotactic protein 1 [MCP-1]. Pharmacological induction of HO-1 with cobalt-protoporphyrin IX reduced, whereas inhibition of HO-1 with either zinc-protoporphyrin IX or siRNA-mediated knockdown increased HLA I Ab-dependent up-regulation of VCAM-1. Treatment with two carbon monoxide (CO-releasing molecules, which liberate the gaseous HO product CO, blocked HLA I Ab-dependent EC activation. Finally, in an in vitro adhesion assay exposure of ECs to HLA I Abs led to increased monocyte binding, which was counteracted by up-regulation of HO-1. In conclusion, HLA I Ab-dependent EC activation is modulated by endothelial HO-1 and targeted induction of this enzyme may be a novel therapeutic approach for the treatment of AMR in solid organ transplantation.

  17. Heme and HO-1 inhibition of HCV, HBV, and HIV

    Warren N Schmidt

    2012-10-01

    Full Text Available Hepatitis C virus, human immunodeficiency virus, and hepatitis B virus are chronic viral infections that cause considerable morbidity and mortality throughout the world. In the decades following the identification and sequencing of these viruses, in vitro experiments demonstrated that heme oxygenase-1, its oxidative products, and related compounds of the heme oxygenase system are virucidal for all three viruses. The purpose of this review is to critically evaluate and summarize the seminal studies that described and characterized this remarkable behavior. It will also discuss more recent work that discovered the antiviral mechanisms and target sites of these unique antiviral agents. In spite of the fact that these viruses are diverse pathogens with quite profound differences in structure and life cycle, it is significant that heme and related compounds show striking similarity for viral target sites across all three species. Collectively, these findings strongly indicate that we should move forward and develop heme and related tetrapyrroles into versatile antiviral agents that could be used therapeutically in patients with single or multiple viral infections.

  18. Upregulation of endothelial heme oxygenase-1 expression through the activation of the JNK pathway by sublethal concentrations of acrolein.

    Wu, C C; Hsieh, C W; Lai, P H; Lin, J B; Liu, Y C; Wung, B S

    2006-08-01

    Acrolein is a highly electrophilic alpha,beta-unsaturated aldehyde that is present in cigarette smoke. Heme oxygenase-1 (HO-1) is a cytoprotective enzyme activated by various such electrophilic compounds. In this study, the regulatory effects of acrolein upon the expression of HO-1 were investigated in endothelial cells (ECs). We demonstrate that acrolein induces the elevation of HO-1 protein levels, and subsequent enzyme activity, at non-cytotoxic concentrations. An additional alpha,beta-unsaturated aldehyde, cinnamaldehyde, was also found to increase HO-1 expression and have less cytotoxicity than acrolein. Moreover, acrolein-mediated HO-1 induction is abrogated in the presence of actinomycin D and cycloheximide. Nrf2 is a transcription factor involved in the induction of HO-1 through an antioxidant response element (ARE) in the promoter region of the HO-1 gene. We show that acrolein induces Nrf2 translocation and ARE-luciferase reporter activity. Acrolein was also found to induce the production of both superoxide and H2O2 at levels greater than 100 microM. However, with the exception of NAC, no antioxidant generated any effect upon acrolein-dependent HO-1 expression in ECs. Our present findings suggest that reactive oxygen species (ROS) may not be a major modulator for HO-1 induction. Using buthionine sulfoximine to deplete the intracellular GSH levels further enhanced the effects of acrolein. We also found that cellular GSH level was rapidly reduced after both 10 and 100 microM acrolein treatment. However, after 6 h of exposure to ECs, only 10 microM acrolein treatment increases GSH level. In addition, only the JNK inhibitor SP600125 and tyrosine kinase inhibitor genistein had any significant inhibitory impact upon the upregulation of HO-1 by acrolein. Pretreatment with a range of other PI3 kinase inhibitors, including wortmannin and LY294002, showed no effects. Hence, we show in our current experiments that a sublethal concentration of acrolein is in fact a

  19. Upregulation of endothelial heme oxygenase-1 expression through the activation of the JNK pathway by sublethal concentrations of acrolein

    Wu, C.C.; Hsieh, C.W.; Lai, P.H.; Lin, J.B.; Liu, Y.C.; Wung, B.S.

    2006-01-01

    Acrolein is a highly electrophilic α,β-unsaturated aldehyde that is present in cigarette smoke. Heme oxygenase-1 (HO-1) is a cytoprotective enzyme activated by various such electrophilic compounds. In this study, the regulatory effects of acrolein upon the expression of HO-1 were investigated in endothelial cells (ECs). We demonstrate that acrolein induces the elevation of HO-1 protein levels, and subsequent enzyme activity, at non-cytotoxic concentrations. An additional α,β-unsaturated aldehyde, cinnamaldehyde, was also found to increase HO-1 expression and have less cytotoxicity than acrolein. Moreover, acrolein-mediated HO-1 induction is abrogated in the presence of actinomycin D and cycloheximide. Nrf2 is a transcription factor involved in the induction of HO-1 through an antioxidant response element (ARE) in the promoter region of the HO-1 gene. We show that acrolein induces Nrf2 translocation and ARE-luciferase reporter activity. Acrolein was also found to induce the production of both superoxide and H 2 O 2 at levels greater than 100 μM. However, with the exception of NAC, no antioxidant generated any effect upon acrolein-dependent HO-1 expression in ECs. Our present findings suggest that reactive oxygen species (ROS) may not be a major modulator for HO-1 induction. Using buthionine sulfoximine to deplete the intracellular GSH levels further enhanced the effects of acrolein. We also found that cellular GSH level was rapidly reduced after both 10 and 100 μM acrolein treatment. However, after 6 h of exposure to ECs, only 10 μM acrolein treatment increases GSH level. In addition, only the JNK inhibitor SP600125 and tyrosine kinase inhibitor genistein had any significant inhibitory impact upon the upregulation of HO-1 by acrolein. Pretreatment with a range of other PI3 kinase inhibitors, including wortmannin and LY294002, showed no effects. Hence, we show in our current experiments that a sublethal concentration of acrolein is in fact a novel HO-1 inducer

  20. Maresin 1 Ameliorates Lung Ischemia/Reperfusion Injury by Suppressing Oxidative Stress via Activation of the Nrf-2-Mediated HO-1 Signaling Pathway

    Quanchao Sun

    2017-01-01

    Full Text Available Lung ischemia/reperfusion (I/R injury occurs in various clinical conditions and heavily damaged lung function. Oxidative stress reaction and antioxidant enzymes play a pivotal role in the etiopathogenesis of lung I/R injury. In the current study, we investigated the impact of Maresin 1 on lung I/R injury and explored the possible mechanism involved in this process. MaR 1 ameliorated I/R-induced lung injury score, wet/dry weight ratio, myeloperoxidase, tumor necrosis factor, bronchoalveolar lavage fluid (BALF leukocyte count, BALF neutrophil ratio, and pulmonary permeability index levels in lung tissue. MaR 1 significantly reduced ROS, methane dicarboxylic aldehyde, and 15-F2t-isoprostane generation and restored antioxidative enzyme (superoxide dismutase, glutathione peroxidase, and catalase activities. Administration of MaR 1 improved the expression of nuclear Nrf-2 and cytosolic HO-1 in I/R-treated lung tissue. Furthermore, we also found that the protective effects of MaR 1 on lung tissue injury and oxidative stress were reversed by HO-1 activity inhibitor, Znpp-IX. Nrf-2 transcription factor inhibitor, brusatol, significantly decreased MaR 1-induced nuclear Nrf-2 and cytosolic HO-1 expression. In conclusion, these results indicate that MaR 1 protects against lung I/R injury through suppressing oxidative stress. The mechanism is partially explained by activation of the Nrf-2-mediated HO-1 signaling pathway.

  1. A Lactobacillus rhamnosus strain induces a heme oxygenase dependent increase in Foxp3+ regulatory T cells.

    Khalil Karimi

    Full Text Available We investigated the consequences of feeding with a Lactobacillus species on the immune environment in GALT, and the role of dendritic cells and heme oxygenase-1 in mediating these responses. Feeding with a specific strain of Lactobacillus rhamnosus induced a significant increase in CD4+CD25+Foxp3+ functional regulatory T cells in GALT. This increase was greatest in the mesenteric lymph nodes and associated with a marked decrease in TNF and IFNγ production. Dendritic cell regulatory function and HO-1 expression was also increased. The increase in Foxp3+ T cells could be prevented by treatment with a heme oxygenase inhibitor. However, neither inhibition of heme oxygenase nor blockade of IL-10 and TGFβ prevented the inhibition of inflammatory cytokine production. In conclusion Lactobacillus feeding induced a tolerogenic environment in GALT. HO-1 was critical to the enhancement of Foxp3+ regulatory T cells while additional, as yet unknown, pathways were involved in the down-regulation of inflammatory cytokine production by T cells.

  2. The induction of heme oxygenase-1 suppresses heat shock protein 90 and the proliferation of human breast cancer cells through its byproduct carbon monoxide

    Lee, Wen-Ying [Department of Pathology, Chi-Mei Hospital, Tainan, Taiwan (China); Department of Pathology, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Chen, Yen-Chou [Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Shih, Chwen-Ming; Lin, Chun-Mao; Cheng, Chia-Hsiung; Chen, Ku-Chung [Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Department of Biochemistry, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Lin, Cheng-Wei, E-mail: cwlin@tmu.edu.tw [Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Department of Biochemistry, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan (China)

    2014-01-01

    Heme oxygenase (HO)-1 is an oxidative stress-response enzyme which catalyzes the degradation of heme into bilirubin, ferric ion, and carbon monoxide (CO). Induction of HO-1 was reported to have antitumor activity; the inhibitory mechanism, however, is still unclear. In the present study, we found that treatment with [Ru(CO){sub 3}Cl{sub 2}]{sub 2} (RuCO), a CO-releasing compound, reduced the growth of human MCF7 and MDA-MB-231 breast cancer cells. Analysis of growth-related proteins showed that treatment with RuCO down-regulated cyclinD1, CDK4, and hTERT protein expressions. Interestingly, RuCO treatment resulted in opposite effects on wild-type and mutant p53 proteins. These results were similar to those of cells treated with geldanamycin (a heat shock protein (HSP)90 inhibitor), suggesting that RuCO might affect HSP90 activity. Moreover, RuCO induced mutant p53 protein destabilization accompanied by promotion of ubiquitination and proteasome degradation. The induction of HO-1 by cobalt protoporphyrin IX (CoPP) showed consistent results, while the addition of tin protoporphyrin IX (SnPP), an HO-1 enzymatic inhibitor, diminished the RuCO-mediated effect. RuCO induction of HO-1 expression was reduced by a p38 mitogen-activated protein kinase inhibitor (SB203580). Additionally, treatment with a chemopreventive compound, curcumin, induced HO-1 expression accompanied with reduction of HSP90 client protein expression. The induction of HO-1 by curcumin inhibited 12-O-tetradecanoyl-13-acetate (TPA)-elicited matrix metalloproteinase-9 expression and tumor invasion. In conclusion, we provide novel evidence underlying HO-1's antitumor mechanism. CO, a byproduct of HO-1, suppresses HSP90 protein activity, and the induction of HO-1 may possess potential as a cancer therapeutic. - Highlights: • CO and HO-1 inhibited the growth of human breast cancer cells. • CO and HO-1 attenuated HSP90 and its client proteins expression. • CO induced mutant p53 protein

  3. Intermittent hypoxia simulating obstructive sleep apnea causes pulmonary inflammation and activates the Nrf2/HO-1 pathway.

    Wang, Yeying; Chai, Yanling; He, Xiaojie; Ai, Li; Sun, Xia; Huang, Yiling; Li, Yongxia

    2017-10-01

    Obstructive sleep apnea (OSA) is a disorder with high morbidity in adults. OSA damages multiple organs and tissues, including the cardiovascular and cerebrovascular systems, the metabolism system, the lungs, liver and heart. OSA-induced damage is earliest and greatest to the pulmonary tissue. The present study established a rat OSA model of differing severity by inducing intermittent hypoxia with different concentrations of O 2 and it was determined that OSA caused a severe oxidative stress response and pulmonary inflammation in a dose-dependent manner. OSA increased serum levels of C-reactive protein and 8-isoprostane and elevated the expression of malondialdehyde, tumor necrosis factor α, interleukin (IL)-1β and IL-6 in the pulmonary tissue. Furthermore, the expression of two important antioxidants, superoxide dismutase and glutathione, was downregulated following intermittent hypoxia. By contrast, levels of cylooxygenase 2 and inducible nitric oxide synthase, which are crucial in the antioxidative response, increased. In addition, OSA activates the nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase (OH)-1 antioxidative signaling pathway. Finally, all increases and decreases in levels of inflammatory and antioxidative substances were dependent on oxygen concentrations. Therefore, the present study demonstrated that OSA, simulated by intermittent hypoxia, caused an oxidative stress response and pulmonary inflammation, and activated the canonical antioxidative Nrf2/HO-1 signaling pathway in a dose-dependent manner. These results may facilitate the development of clinical therapies to treat pulmonary diseases caused by OSA.

  4. Microsatellite polymorphism in the heme oxygenase-1 gene promoter and the risk of atrial fibrillation in Taiwanese.

    Lung-An Hsu

    Full Text Available Atrial fibrillation (AF is associated with increased oxidative stress. Emerging evidence suggests that heme oxygenase-1 (HO-1 is a potent antioxidant system against various oxidative stress-related diseases. The human HO-1 promoter has a GT-repeat length polymorphism that can determine the level of gene transcription.The aim of this study is to assess the role of the GT-repeat polymorphism in the promoter region of the HO-1 gene in Chinese-Taiwanese patients with AF.This study enrolled 200 AF patients and 240 controls, comparable for age and gender. In each subject, the length of GT-repeat polymorphism in the HO-1 promoter region was examined by polymerase chain reactions. The frequencies of long GT-repeat alleles (≧32 were significantly higher in AF patients than in controls. Multivariate analysis showed that the presence of long allele was significantly and independently associated with AF (odds ratio: 1.91, 95% CI 1.07-3.72; P = 0.028. Right atrial tissues from patients with chronic AF were investigated with immunoconfocal microscopy. Patients homozygous for shorter GT-repeat alleles exhibited greater HO-1 expression in their atria than those homozygous for longer alleles, which was reflected by less oxidative stress, myofibril degradation, and fibrosis in the atria of patients with shorter GT-repeat. In vitro, transient transfection assay in HL-1 atrial myocytes showed that the responsiveness of HO-1 transcriptional activity to tachypacing was inversely correlated with the length of the GT-repeats.Our results suggest that the HO-1 microsatellite polymorphism may contribute to the genetic background of AF in Chinese-Taiwanese patients.

  5. Fetal Microsatellite in the Heme Oxygenase 1 Promoter Is Associated With Severe and Early-Onset Preeclampsia.

    Kaartokallio, Tea; Utge, Siddheshwar; Klemetti, Miira M; Paananen, Jussi; Pulkki, Kari; Romppanen, Jarkko; Tikkanen, Ilkka; Heinonen, Seppo; Kajantie, Eero; Kere, Juha; Kivinen, Katja; Pouta, Anneli; Lakkisto, Päivi; Laivuori, Hannele

    2018-01-01

    Preeclampsia is a vascular pregnancy disorder that often involves impaired placental development. HO-1 (heme oxygenase 1, encoded by HMOX1 ) is a stress response enzyme crucial for endothelial and placental function. Long version of the guanine-thymine (GT n ) microsatellite in the HMOX1 promoter decreases HO-1 expression, and the long maternal repeat is associated with late-onset preeclampsia. Our aim was to study whether the length of fetal repeat is associated with mother's preeclampsia, whether the length of fetal and maternal repeats affect HO-1 levels in placenta and maternal serum, and whether HO-1 levels are altered in preeclampsia. We genotyped the repeat in the cord blood of 609 preeclamptic and 745 nonpreeclamptic neonates. HO-1 levels were measured in 36 placental samples, and in the first (222 cases/243 controls) and third (176 cases/53 controls) pregnancy trimester serum samples using enzyme-linked immunosorbent assay. The long fetal GT n repeat was associated with preeclampsia and its severe and early-onset subtypes. Interaction analysis suggested the maternal and fetal effects to be independent. Placental or serum HO-1 levels were not altered in preeclamptics, possibly reflecting heterogeneity of preeclampsia. Carriers of the long fetal and maternal repeats had lower placental and serum HO-1 levels, respectively, providing functional evidence for the association. We conclude that the long fetal GT n repeat may increase mother's risk for especially severe and early-onset preeclampsia. The fetal and maternal risk alleles likely predispose to different disease subtypes. © 2017 American Heart Association, Inc.

  6. Antrodia camphorata Potentiates Neuroprotection against Cerebral Ischemia in Rats via Downregulation of iNOS/HO-1/Bax and Activated Caspase-3 and Inhibition of Hydroxyl Radical Formation

    Po-Sheng Yang

    2015-01-01

    Full Text Available Antrodia camphorata (A. camphorata is a fungus generally used in Chinese folk medicine for treatment of viral hepatitis and cancer. Our previous study found A. camphorata has neuroprotective properties and could reduce stroke injury in cerebral ischemia animal models. In this study, we sought to investigate the molecular mechanisms of neuroprotective effects of A. camphorata in middle cerebral artery occlusion (MCAO rats. A selective occlusion of the middle cerebral artery (MCA with whole blood clots was used to induce ischemic stroke in rats and they were orally treated with A. camphorata (0.25 and 0.75 g/kg/day alone or combined with aspirin (5 mg/kg/day. To provide insight into the functions of A. camphorata mediated neuroprotection, the expression of Bax, inducible nitric oxide synthase (iNOS, haem oxygenase-1 (HO-1, and activated caspase-3 was determined by Western blot assay. Treatment of aspirin alone significantly reduced the expressions of HO-1 (P<0.001, iNOS (P<0.001, and Bax (P<0.01 in ischemic regions. The reduction of these expressions was more potentiated when rats treated by aspirin combined with A. camphorata (0.75 g/kg/day. Combination treatment also reduced apoptosis as measured by a significant reduction in active caspase-3 expression in the ischemic brain compared to MCAO group (P<0.01. Moreover, treatment of A. camphorata significantly (P<0.05 reduced fenton reaction-induced hydroxyl radical (OH• formation at a dose of 40 mg/mL. Taken together, A. camphorata has shown neuroprotective effects in embolic rats, and the molecular mechanisms may correlate with the downregulation of Bax, iNOS, HO-1, and activated caspase-3 and the inhibition of OH• signals.

  7. Antioxidant and Antifibrotic Effect of a Herbal Formulation In Vitro and in the Experimental Andropause via Nrf2/HO-1 Signaling Pathway

    Woong Jin Bae

    2017-01-01

    Full Text Available The Korean herbal formulation Ojayeonjonghwan is used for improving late-onset hypogonadism (LOH symptoms such as erectile dysfunction (ED. A previous research suggested that a modified Ojayeonjonghwan (KH-204 could be used as an alternative to the treatment for ED. The pharmacological effects were examined in different conditions, including in vitro and in vivo. We measured the survival rate of TM3 Leydig cells under the oxidative stress condition. The s.c. injection of leuprorelin was used to induce androgen deprivation. We measured serum testosterone levels, oxidative stress, and apoptosis. The results of the treatment by KH-204 (1 preserved TM3 cells from oxidative stress by improving the expression of nuclear factor erythroid 2-related factor 2 (Nrf2/heme oxygenase-1 (HO-1; (2 lowered the expression of transforming growth factor-beta (TGF-β 1/SMAD; (3 increased the average of serum testosterone in androgen-deprived male rats; (4 kept the activation of spermatogenesis; (5 upgraded the contents of 8-hydroxy-20-deoxyguanosine (8-OHdG and degraded the contents of superoxide dismutase (SOD; and (6 reduced apoptosis. We studied that KH-204 improved testicular dysfunction in LOH. It is likely, at least in part, to degrade oxidative stress through the Nrf2/HO-1 pathway. These findings may offer credible evidences for the use of new alternative therapies to treat LOH.

  8. Serum heme oxygenase-1 levels in patients with primary dysmenorrhea.

    Aksoy, Ayse Nur; Laloglu, Esra; Ozkaya, Alev Lazoglu; Yilmaz, Emsal Pınar Topdagi

    2017-04-01

    Primary dysmenorrhea effects the life-quality of women negatively. The aim of this study was to evaluate heme oxygenase-1 (HO1) activity together with malondialdehyde (MDA) and nitric oxide (NO) levels in patients with primary dysmenorrhea. A total of 28 nulliparous women with the diagnosis of primary dysmenorrhea and 26 healthy controls were included in this study. On the first day of menstruation, all patients underwent ultrasound examination to exclude pelvic pathology and the visual analogue scale was applied to patients. Patient's visual analogue scale (VAS) scores, age, body mass index (BMI), menstrual cycle length (day), length of bleeding (day) were recorded. In the same day, fasting blood samples were taken from each patient for biochemical analysis. Serum MDA, NO and HO1 levels were found to be higher in women with primary dysmenorrhea compared to healthy controls (p = 0.012, p = 0.009, p dysmenorrhea. Antioxidant support might be helpful to reduce pain severity in primary dysmenorrhea.

  9. HO-1 Upregulation Attenuates Adipocyte Dysfunction, Obesity, and Isoprostane Levels in Mice Fed High Fructose Diets

    Zeid Khitan

    2014-01-01

    Full Text Available Background. Fructose metabolism is an unregulated metabolic pathway and excessive fructose consumption is known to activate ROS. HO-1 is a potent antioxidant gene that plays a key role in decreasing ROS and isoprostanes. We examined whether the fructose-mediated increase in adipocyte dysfunction involves an increase in isoprostanes and that pharmacological induction of HO-1 would decrease both isoprostane levels and adipogenesis. Methods and Results. We examined the effect of fructose, on adipogenesis in human MSCs in the presence and absence of CoPP, an inducer of HO-1. Fructose increased adipogenesis and the number of large lipid droplets while decreasing the number of small lipid droplets (P<0.05. Levels of heme and isoprostane in fructose treated MSC-derived adipocytes were increased. CoPP reversed these effects and markedly increased HO-1 and the Wnt signaling pathway. The high fructose diet increased heme levels in adipose tissue and increased circulating isoprostane levels (P<0.05 versus control. Fructose diets decreased HO-1 and adiponectin levels in adipose tissue. Induction of HO-1 by CoPP decreased isoprostane synthesis (P<0.05 versus fructose. Conclusion. Fructose treatment resulted in increased isoprostane production and adipocyte dysfunction, which was reversed by the increased expression of HO-1.

  10. Protective Effects of Curcumin on Manganese-Induced BV-2 Microglial Cell Death.

    Park, Euteum; Chun, Hong Sung

    2017-08-01

    Curcumin, a bioactive component in tumeric, has been shown to exert antioxidant, anti-inflammatory, anticarcinogenic, hepatoprotective, and neuroprotective effects, but the effects of curcumin against manganese (Mn)-mediated neurotoxicity have not been studied. This study examined the protective effects of curcumin on Mn-induced cytotoxicity in BV-2 microglial cells. Curcumin (0.1-10 µM) dose-dependently prevented Mn (250 µM)-induced cell death. Mn-induced mitochondria-related apoptotic characteristics, such as caspase-3 and -9 activation, cytochrome c release, Bax increase, and Bcl-2 decrease, were significantly suppressed by curcumin. In addition, curcumin significantly increased intracellular glutathione (GSH) and moderately potentiated superoxide dismutase (SOD), both which were diminished by Mn treatment. Curcumin pretreatment effectively suppressed Mn-induced upregulation of malondialdehyde (MDA), total reactive oxygen species (ROS). Moreover, curcumin markedly inhibited the Mn-induced mitochondrial membrane potential (MMP) loss. Furthermore, curcumin was able to induce heme oxygenase (HO)-1 expression. Curcumin-mediated inhibition of ROS, down-regulation of caspases, restoration of MMP, and recovery of cell viability were partially reversed by HO-1 inhibitor (SnPP). These results suggest the first evidence that curcumin can prevent Mn-induced microglial cell death through the induction of HO-1 and regulation of oxidative stress, mitochondrial dysfunction, and apoptotic events.

  11. Cloning and characterization of a heme oxygenase-2 gene from alfalfa (Medicago sativa L.).

    Fu, Guang-Qing; Jin, Qi-Jiang; Lin, Yu-Ting; Feng, Jian-Fei; Nie, Li; Shen, Wen-Biao; Zheng, Tian-Qing

    2011-11-01

    Heme oxygenase (HO, EC 1.14.99.3) catalyzes the oxidation of heme and performs vital roles in plant development and stress responses. Two HO isozymes exist in plants. Between these, HO-1 is an oxidative stress-response protein, and HO-2 usually exhibited constitutive expression. Although alfalfa HO-1 gene (MsHO1) has been investigated previously, HO2 is still poorly understood. In this study, we report the cloning and characterization of HO2 gene, MsHO2, from alfalfa (Medica sativa L.). The full-length cDNA of MsHO2 contains an ORF of 870 bp and encodes for 290 amino acid residues with a predicted molecular mass of 33.3 kDa. Similar to MsHO1, MsHO2 also appears to have an N-terminal transit peptide sequence for chloroplast import. Many conserved residues in plant HO were also conserved in MsHO2. However, unlike HO-1, the conserved histidine (His) required for heme-iron binding and HO activity was replaced by tyrosine (Tyr) in MsHO2. Further biochemical activity analysis of purified mature MsHO2 showed no HO activity, suggesting that MsHO2 may not be a true HO in nature. Semi-quantitative RT-PCR confirmed its maximum expression in the germinating seeds. Importantly, the expression levels of MsHO2 were up-regulated under sodium nitroprusside (SNP) and H(2)O(2) (especially) treatment, respectively.

  12. Edaravone attenuates hippocampal damage in an infant mouse model of pneumococcal meningitis by reducing HMGB1 and iNOS expression via the Nrf2/HO-1 pathway.

    Li, Zheng; Ma, Qian-Qian; Yan, Yan; Xu, Feng-Dan; Zhang, Xiao-Ying; Zhou, Wei-Qin; Feng, Zhi-Chun

    2016-09-01

    Edaravone (3-methyl-1-phenyl-2-pyrazolin-5-one) is a free radical scavenger that has shown potent antioxidant, anti-inflammatory and neuroprotective effects in variety of disease models. In this study, we investigated whether edaravone produced neuroprotective actions in an infant mouse model of pneumococcal meningitis. C57BL/6 mice were infected on postnatal d 11 by intracisternal injection of a certain inoculum of Streptococcus pneumoniae. The mice received intracisternal injection of 10 μL of saline containing edaravone (3 mg/kg) once a day for 7 d. The severity of pneumococcal meningitis was assessed with a clinical score. In mice with severe meningitis, the survival rate from the time of infection to d 8 after infection was analyzed using Kaplan-Meier curves. In mice with mild meningitis, the CSF inflammation and cytokine levels in the hippocampus were analyzed d 7 after infection, and the clinical neurological deficit score was evaluated using a neurological scoring system d 14 after infection. The nuclear factor (erythroid-derived 2)-like 2 knockout (Nrf2 KO) mice and heme oxygenase-1 knockout (HO-1 KO) mice were used to confirm the involvement of Nrf2/HO-1 pathway in the neuroprotective actions of edaravone. In mice with severe meningitis, edaravone treatment significantly increased the survival rate (76.4%) compared with the meningitis model group (32.2%). In mice with mild meningitis, edaravone treatment significantly decreased the number of leukocytes and TNF- levels in CSF, as well as the neuronal apoptosis and protein levels of HMGB1 and iNOS in the hippocampus, but did not affect the high levels of IL-10 and IL-6 in the hippocampus. Moreover, edaravone treatment significantly improved the neurological function of mice with mild meningitis. In Nrf2 KO or HO-1 KO mice with the meningitis, edaravone treatment was no longer effective in improving the survival rate of the mice with severe meningitis (20.2% and 53.6%, respectively), nor it affected the

  13. Covalent heme attachment to the protein in human heme oxygenase-1 with selenocysteine replacing the His25 proximal iron ligand.

    Jiang, Yongying; Trnka, Michael J; Medzihradszky, Katalin F; Ouellet, Hugues; Wang, Yongqiang; Ortiz de Montellano, Paul R

    2009-03-01

    To characterize heme oxygenase with a selenocysteine (SeCys) as the proximal iron ligand, we have expressed truncated human heme oxygenase-1 (hHO-1) His25Cys, in which Cys-25 is the only cysteine, in the Escherichia coli cysteine auxotroph strain BL21(DE3)cys. Selenocysteine incorporation into the protein was demonstrated by both intact protein mass measurement and mass spectrometric identification of the selenocysteine-containing tryptic peptide. One selenocysteine was incorporated into approximately 95% of the expressed protein. Formation of an adduct with Ellman's reagent (DTNB) indicated that the selenocysteine in the expressed protein was in the reduced state. The heme-His25SeCys hHO-1 complex could be prepared by either (a) supplementing the overexpression medium with heme, or (b) reconstituting the purified apoprotein with heme. Under reducing conditions in the presence of imidazole, a covalent bond is formed by addition of the selenocysteine residue to one of the heme vinyl groups. No covalent bond is formed when the heme is replaced by mesoheme, in which the vinyls are replaced by ethyl groups. These results, together with our earlier demonstration that external selenolate ligands can transfer an electron to the iron [Y. Jiang, P.R. Ortiz de Montellano, Inorg. Chem. 47 (2008) 3480-3482 ], indicate that a selenyl radical is formed in the hHO-1 His25SeCys mutant that adds to a heme vinyl group.

  14. Curcumin Attenuates on Carbon Tetrachloride-Induced Acute Liver Injury in Mice via Modulation of the Nrf2/HO-1 and TGF-β1/Smad3 Pathway

    Xinyan Peng

    2018-01-01

    Full Text Available This study aimed to investigate the protective effect of curcumin against carbon tetrachloride (CCl4-induced acute liver injury in a mouse model, and to explain the underlying mechanism. Curcumin at doses of 50, 100 and 200 mg/kg/day were administered orally once daily for seven days prior to CCl4 exposure. At 24 h, curcumin-attenuated CCl4 induced elevated serum transaminase activities and histopathological damage in the mouse’s liver. Curcumin pre-treatment at 50, 100 and 200 mg/kg significantly ameliorated CCl4-induced oxidative stress, characterized by decreased malondialdehyde (MDA formations, and increased superoxide dismutase (SOD, catalase (CAT activities and glutathione (GSH content, followed by a decrease in caspase-9 and -3 activities. Curcumin pre-treatment significantly decreased CCl4-induced inflammation. Furthermore, curcumin pre-treatment significantly down-regulated the expression of TGF-β1 and Smad3 mRNAs (both p < 0.01, and up-regulated the expression of nuclear-factor erythroid 2-related factor 2 (Nrf2 and HO-1 mRNA (both p < 0.01 in the liver. Inhibition of HO-1 attenuated the protective effect of curcumin on CCl4-induced acute liver injury. Given these outcomes, curcumin could protect against CCl4-induced acute liver injury by inhibiting oxidative stress and inflammation, which may partly involve the activation of Nrf2/HO-1 and inhibition of TGF-β1/Smad3 pathways.

  15. (+)-Nootkatone and (+)-valencene from rhizomes of Cyperus rotundus increase survival rates in septic mice due to heme oxygenase-1 induction.

    Tsoyi, Konstantin; Jang, Hwa Jin; Lee, Young Soo; Kim, Young Min; Kim, Hye Jung; Seo, Han Geuk; Lee, Jae Heun; Kwak, Jong Hwan; Lee, Dong-Ung; Chang, Ki Churl

    2011-10-11

    The rhizomes of Cyperus rotundus have been used as traditional folk medicine for the treatment of inflammatory diseases. However, the mechanism by which extract of rhizomes of Cyperus rotundus (ECR) elicits anti-inflammation has not been extensively investigated so far. The aim of the present study was to test whether heme oxygenase (HO)-1 induction is involved in the anti-inflammatory action of ECR. Induction of HO-1 and inhibition of inducible nitric oxide synthase (iNOS)/NO production by ECR and its 12 constituents (3 monoterpenes, 5 sesquiterpenes, and 4 aromatic compounds) were investigated using RAW264.7 cells in vitro. In addition, anti-inflammatory action of ECR and its two active ingredients (nookkatone, valencene) were confirmed in sepsis animal model in vivo. ECR increased HO-1 expression in a concentration-dependent manner, which was correlated with significant inhibition of iNOS/NO production in LPS-activated RAW264.7 cells. Among 12 compounds isolated from ECR, mostly sesquiterpenes induced stronger HO-1 expression than monoterpenes in macrophage cells. Nootkatone and valencene (sesquiterpenes) significantly inhibited iNOS expression and NO production in LPS-simulated RAW264.7 cells. Inhibition of iNOS expression by nootkatone, valencene, and ECR were significantly reduced in siHO-1 RNA transfected cells. Furthermore, all three showed marked inhibition of high mobility group box-1 (HMGB1) in LPS-activated macrophages and increased survival rates in cecal ligation and puncture (CLP)-induced sepsis in mice. Taken together, we concluded that possible anti-inflammatory mechanism of ECR is, at least, due to HO-1 induction, in which sesquiterpenes such as nootkatone and valencene play a crucial role. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  16. Protective effects of total glucosides of paeony and the underlying mechanisms in carbon tetrachloride-induced experimental liver injury

    Qin, Ying; Tian, Ya-ping

    2011-01-01

    Introduction We explored the protective effects of total glucosides of paeony (TGP) and the underlying mechanisms in carbon tetrachloride (CCl4)-induced experimental liver injury in mice. Material and methods Chronic liver damage was induced by intraperitoneal injection of CCl4 (0.5 µl/g) three times per week for 8 weeks. Mice also received 25, 50 or 100 mg/kg TGP. Liver sections were stained with haematoxylin/eosin. Serum amino transferases, lipid peroxidation and tumour necrosis factor-α (TNF-α) levels were determined using commercial assays. Quantitative real-time polymerase chain reaction was used to determine the changes in hepatic TNF-α, COX-2, iNOS and HO-1 expression. Protein levels of nitric oxide synthase, cyclooxygenase-2, haem oxygenase-1 and cytochrome P450 2E1 were determined by western blotting. Results Histological results showed that TGP improved the CCl4-induced changes in liver structure and alleviated lobular necrosis. The increases in serum protein and hepatic mRNA expression of TNF-α induced by CCl4 treatment were suppressed by TGP. Total glucosides of paeony also attenuated the increase the expression in iNOS and CYP2E1 but augmented the increase in HO-1.The mRNA and protein expression levels of inducible HO-1 increased significantly after CCl4 treatment. Conclusions Total glucosides of paeony protects hepatocytes from oxidative damage induced by CCl4. Total glucosides of paeony may achieve these effects by enhancing HO-1 expression and inhibiting the expression of proinflammatory mediators. PMID:22291795

  17. Expression and characterization of full-length human heme oxygenase-1: the presence of intact membrane-binding region leads to increased binding affinity for NADPH cytochrome P450 reductase.

    Huber, Warren J; Backes, Wayne L

    2007-10-30

    Heme oxygenase-1 (HO-1) is the chief regulatory enzyme in the oxidative degradation of heme to biliverdin. In the process of heme degradation, HO-1 receives the electrons necessary for catalysis from the flavoprotein NADPH cytochrome P450 reductase (CPR), releasing free iron and carbon monoxide. Much of the recent research involving heme oxygenase has been done using a 30 kDa soluble form of the enzyme, which lacks the membrane binding region (C-terminal 23 amino acids). The goal of this study was to express and purify a full-length human HO-1 (hHO-1) protein; however, due to the lability of the full-length form, a rapid purification procedure was required. This was accomplished by use of a glutathione-s-transferase (GST)-tagged hHO-1 construct. Although the procedure permitted the generation of a full-length HO-1, this form was contaminated with a 30 kDa degradation product that could not be eliminated. Therefore, attempts were made to remove a putative secondary thrombin cleavage site by a conservative mutation of amino acid 254, which replaces arginine with lysine. This mutation allowed the expression and purification of a full-length hHO-1 protein. Unlike wild type (WT) HO-1, the R254K mutant could be purified to a single 32 kDa protein capable of degrading heme at the same rate as the WT enzyme. The R254K full-length form had a specific activity of approximately 200-225 nmol of bilirubin h-1 nmol-1 HO-1 as compared to approximately 140-150 nmol of bilirubin h-1 nmol-1 for the WT form, which contains the 30 kDa contaminant. This is a 2-3-fold increase from the previously reported soluble 30 kDa HO-1, suggesting that the C-terminal 23 amino acids are essential for maximal catalytic activity. Because the membrane-spanning domain is present, the full-length hHO-1 has the potential to incorporate into phospholipid membranes, which can be reconstituted at known concentrations, in combination with other endoplasmic reticulum resident enzymes.

  18. An improved method for purification of recombinant truncated heme oxygenase-1 by expanded bed adsorption and gel filtration.

    Hu, Hong-Bo; Wang, Wei; Han, Ling; Zhou, Wen-Pu; Zhang, Xue-Hong

    2007-03-01

    Recombinant truncated human heme oxygenase-1 (hHO-1) expressed in Escherichia coli was efficiently separated and purified from feedstock by DEAE-ion exchange expanded bed adsorption. Protocol optimization of hHO-1 on DEAE adsorbent resulted in adsorption in 0 M NaCl and elution in 150 mM NaCl at a pH of 8.5. The active enzyme fractions separated from the expanded bed column were further purified by a Superdex 75 gel filtration step. The specific hHO-1 activity increased from 0.82 +/- 0.05 to 24.8 +/- 1.8 U/mg during the whole purification steps. The recovery and purification factor of truncated hHO-1 of the whole purification were 72.7 +/- 4.7 and 30.2 +/- 2.3%, respectively. This purification process can decrease the demand on the preparation of feedstock and simplify the purification process.

  19. Fructose during pregnancy provokes fetal oxidative stress: The key role of the placental heme oxygenase-1.

    Rodrigo, Silvia; Rodríguez, Lourdes; Otero, Paola; Panadero, María I; García, Antonia; Barbas, Coral; Roglans, Núria; Ramos, Sonia; Goya, Luis; Laguna, Juan C; Álvarez-Millán, Juan J; Bocos, Carlos

    2016-12-01

    One of the features of metabolic syndrome caused by liquid fructose intake is an impairment of redox status. We have investigated whether maternal fructose ingestion modifies the redox status in pregnant rats and their fetuses. Fructose (10% wt/vol) in the drinking water of rats throughout gestation, leads to maternal hepatic oxidative stress. However, this change was also observed in glucose-fed rats and, in fact, both carbohydrates produced a decrease in antioxidant enzyme activity. Surprisingly, mothers fed carbohydrates displayed low plasma lipid oxidation. In contrast, fetuses from fructose-fed mothers showed elevated levels of plasma lipoperoxides versus fetuses from control or glucose-fed mothers. Interestingly, a clearly augmented oxidative stress was observed in placenta of fructose-fed mothers, accompanied by a lower expression of the transcription factor Nuclear factor-erythroid 2-related factor-2 (Nrf2) and its target gene, heme oxygenase-1 (HO-1), a potent antioxidant molecule. Moreover, histone deacetylase 3 (HDAC3) that has been proposed to upregulate HO-1 expression by stabilizing Nrf2, exhibited a diminished expression in placenta of fructose-supplemented mothers. Maternal fructose intake provoked an imbalanced redox status in placenta and a clear diminution of HO-1 expression, which could be responsible for the augmented oxidative stress found in their fetuses. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Structural characterization of human heme oxygenase-1 in complex with azole-based inhibitors.

    Rahman, Mona N; Vlahakis, Jason Z; Roman, Gheorghe; Vukomanovic, Dragic; Szarek, Walter A; Nakatsu, Kanji; Jia, Zongchao

    2010-03-01

    The development of inhibitors specific for heme oxygenases (HO) aims to provide powerful tools in understanding the HO system. Based on the lead structure (2S, 4S)-2-[2-(4-chlorophenyl)ethyl]-2-[(1H-imidazol-1-yl)methyl]-4-[((4-aminophenyl)thio)methyl]-1,3-dioxolane (azalanstat, QC-1) we have synthesized structural modifications to develop novel and selective HO inhibitors. The structural study of human HO-1 (hHO-1) in complex with a select group of the inhibitors was initiated using X-ray crystallographic techniques. Comparison of the structures of four such compounds each in complex with hHO-1 revealed a common binding mode, despite having different structural fragments. The compounds bind to the distal side of heme through an azole "anchor" which coordinates with the heme iron. An expansion of the distal pocket, mainly due to distal helix flexibility, allows accommodation of the compounds without displacing heme or the critical Asp140 residue. Rather, binding displaces a catalytically critical water molecule and disrupts an ordered hydrogen-bond network involving Asp140. The presence of a triazole "anchor" may provide further stability via a hydrogen bond with the protein. A hydrophobic pocket acts to stabilize the region occupied by the phenyl or adamantanyl moieties of these compounds. Further, a secondary hydrophobic pocket is formed via "induced fit" to accommodate bulky substituents at the 4-position of the dioxolane ring. Copyright 2009 Elsevier Inc. All rights reserved.

  1. Nitric oxide and iron modulate heme oxygenase activity as a long distance signaling response to salt stress in sunflower seedling cotyledons.

    Singh, Neha; Bhatla, Satish C

    2016-02-29

    Nitric oxide is a significant component of iron signaling in plants. Heme is one of the iron sensors in plants. Free heme is highly toxic and can cause cell damage as it catalyzes the formation of reactive oxygen species (ROS). Its catabolism is carried out by heme oxygenase (HOs; EC 1.14.99.3) which uses heme both as a prosthetic group and as a substrate. Two significant events, which accompany adaptation to salt stress in sunflower seedlings, are accumulation of ROS and enhanced production of nitric oxide (NO) in roots and cotyledons. Present investigations on the immunolocalization of heme oxygenase distribution in sunflower seedling cotyledons by confocal laser scanning microscopic (CLSM) imaging provide new information on the differential spatial distribution of the inducible form of HO (HO-1) as a long distance in response to NaCl stress. The enzyme is abundantly distributed in the specialized cells around the secretory canals (SCs) in seedling cotyledons. Abundance of tyrosine nitrated proteins has also been observed in the specialized cells around the secretory canals in cotyledons derived from salt stressed seedlings. The spatial distribution of tyrosine nitrated proteins and HO-1 expression further correlates with the abundance of mitochondria in these cells. Present findings, thus, highlight a link among distribution of HO-1 expression, abundance of tyrosine nitrated proteins and mitochondria in specialized cells around the secretory canal as a long distance mechanism of salt stress tolerance in sunflower seedlings. Enhanced spatial distribution of HO-1 in response to NaCl stress in seedling cotyledons is in congruence with the observed increase in specific activity of HO-1 in NaCl stressed conditions. The enzyme activity is further enhanced by hemin (HO-1 inducer) both in the absence or presence of NaCl stress and inhibited by zinc protoporphyrin. Western blot analysis of cotyledon homogenates using anti-HO-1 polyclonal antibody shows one major band (29

  2. Effects of Nuclear Factor-E2-related factor 2/Heme Oxygenase 1 on splanchnic hemodynamics in experimental cirrhosis with portal hypertension.

    Qin, Jun; He, Yue; Duan, Ming; Luo, Meng

    2017-05-01

    We explored the effects of Nuclear Factor-E2-related factor 2 (Nrf2) and Heme Oxygenase 1 (HO-1) on splanchnic hemodynamics in portal hypertensive rats. Experimental cirrhosis with portal hypertension was induced by intraperitoneal injection of carbon tetrachloride. The expression of proteins was examined by immunoblotting. Hemodynamic studies were performed by radioactive microspheres. The vascular perfusion system was used to measure the contractile response of mesentery arterioles in rats. Nrf2 expression in the nucleus and HO-1 expression in cytoplasm was significantly enhanced in portal hypertensive rats. Portal pressure, as well as regional blood flow, increased significantly in portal hypertension and can be blocked by tin protoporphyrin IX. The expression of endogenous nitric oxide synthase and vascular endothelial growth factors increased significantly compared to normal rats, while HO-1 inhibition decreased the expression of these proteins significantly. The contractile response of mesenteric arteries decreased in portal hypertension, but can be partially recovered through tin protoporphyrin IX treatment. The expression of Nrf2/HO-1 increased in mesenteric arteries of portal hypertensive rats, which was related to oxidative stress. HO-1was involved in increased portal pressure and anomaly splanchnic hemodynamics in portal hypertensive rats. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Crocin Suppresses LPS-Stimulated Expression of Inducible Nitric Oxide Synthase by Upregulation of Heme Oxygenase-1 via Calcium/Calmodulin-Dependent Protein Kinase 4

    Ji-Hee Kim

    2014-01-01

    Full Text Available Crocin is a water-soluble carotenoid pigment that is primarily used in various cuisines as a seasoning and coloring agent, as well as in traditional medicines for the treatment of edema, fever, and hepatic disorder. In this study, we demonstrated that crocin markedly induces the expression of heme oxygenase-1 (HO-1 which leads to an anti-inflammatory response. Crocin inhibited inducible nitric oxide synthase (iNOS expression and nitric oxide production via downregulation of nuclear factor kappa B activity in lipopolysaccharide- (LPS- stimulated RAW 264.7 macrophages. These effects were abrogated by blocking of HO-1 expression or activity. Crocin also induced Ca2+ mobilization from intracellular pools and phosphorylation of Ca2+/calmodulin-dependent protein kinase 4 (CAMK4. CAMK4 knockdown and kinase-dead mutant inhibited crocin-mediated HO-1 expression, Nrf2 activation, and phosphorylation of Akt, indicating that HO-1 expression is mediated by CAMK4 and that Akt is a downstream mediator of CAMK4 in crocin signaling. Moreover, crocin-mediated suppression of iNOS expression was blocked by CAMK4 inhibition. Overall, these results suggest that crocin suppresses LPS-stimulated expression of iNOS by inducing HO-1 expression via Ca2+/calmodulin-CAMK4-PI3K/Akt-Nrf2 signaling cascades. Our findings provide a novel molecular mechanism for the inhibitory effects of crocin against endotoxin-mediated inflammation.

  4. Upregulation of human heme oxygenase gene expression by Ets-family proteins.

    Deramaudt, B M; Remy, P; Abraham, N G

    1999-03-01

    Overexpression of human heme oxygenase-1 has been shown to have the potential to promote EC proliferation and angiogenesis. Since Ets-family proteins have been shown to play an important role in angiogenesis, we investigated the presence of ETS binding sites (EBS), GGAA/T, and ETS protein contributing to human HO-1 gene expression. Several chloramphenicol acetyltransferase constructs were examined in order to analyze the effect of ETS family proteins on the transduction of HO-1 in Xenopus oocytes and in microvessel endothelial cells. Heme oxygenase promoter activity was up-regulated by FLI-1ERGETS-1 protein(s). Chloramphenicol acetyltransferase (CAT) assays demonstrated that the promoter region (-1500 to +19) contains positive and negative control elements and that all three members of the ETS protein family were responsible for the up-regulation of HHO-1. Electrophoretic mobility shift assays (EMSA), performed with nuclear extracts from endothelial cells overexpressing HHO-1 gene, and specific HHO-1 oligonucleotides probes containing putative EBS resulted in a specific and marked bandshift. Synergistic binding was observed in EMSA between AP-1 on the one hand, FLI-1, ERG, and ETS-1 protein on the other. Moreover, 5'-deletion analysis demonstrated the existence of a negative control element of HHO-1 expression located between positions -1500 and -120 on the HHO-1 promoter. The presence of regulatory sequences for transcription factors such as ETS-1, FLI-1, or ERG, whose activity is associated with cell proliferation, endothelial cell differentiation, and matrix metalloproteinase transduction, may be an indication of the important role that HO-1 may play in coronary collateral circulation, tumor growth, angiogenesis, and hemoglobin-induced endothelial cell injuries.

  5. Transforming growth factor β1 enhances heme oxygenase 1 expression in human synovial fibroblasts by inhibiting microRNA 519b synthesis.

    Shu-Jui Kuo

    Full Text Available Osteoarthritis (OA is manifested by synovial inflammation and cartilage destruction that is directly linked to synovitis, joint swelling and pain. In the light of the role of synovium in the pathogenesis and the symptoms of OA, synovium-targeted therapy is a promising strategy to mitigate the symptoms and progression of OA. Transforming growth factor beta 1 (TGF-β1, a secreted homodimeric protein, possesses unique and potent anti-inflammatory and immune-regulatory properties in many cell types. Heme oxygenase 1 (HO-1 is an inducible anti-inflammatory and stress responsive enzyme that has been proven to prevent injuries caused by many diseases. Despite the similar anti-inflammatory profile and their involvement in the pathogenesis of arthritic diseases, no studies have as yet explored the possibility of any association between the expression of TGF-β1 and HO-1.TGF-β1-induced HO-1 expression was examined by HO-1 promoter assay, qPCR, and Western blotting. The siRNAs and enzyme inhibitors were utilized to determine the intermediate involved in the signal transduction pathway. We showed that TGF-β1 stimulated the synthesis of HO-1 in a concentration- and time-dependent manner, which can be mitigated by blockade of the phospholipase (PLCγ/protein kinase C alpha (PKCα pathway. We also showed that the expression of miRNA-519b, which blocks HO-1 transcription, is inhibited by TGF-β1, and the suppression of miRNA 519b could be reversed via blockade of the PLCγ/PKCα pathway.TGF-β1 stimulated the expression of HO-1 via activating the PLCγ/PKCα pathway and suppressing the downstream expression of miRNA-519b. These results may shed light on the pathogenesis and treatment of OA.

  6. Study on association between genetic polymorphisms of haem oxygenase-1, tumour necrosis factor, cadmium exposure and malaria pathogenicity and severity

    Ruangweerayut Ronnatrai

    2010-09-01

    Full Text Available Abstract Background Malaria is the most important public health problems in tropical and sub-tropical countries. Haem oxygenase (HO enzyme and the pro-inflammatory cytokine tumour necrosis factor (TNF have been proposed as one of the factors that may play significant role in pathogenicity/severity of malaria infection. HO is the enzyme of the microsomal haem degradation pathway that yields biliverdin, carbon monoxide, and iron. In this study, the association between malaria disease pathogenicity/severity and (GTn repeat polymorphism in the promoter region of the inducible HO-1 including the effect of cadmium exposure (potent inducer of HO-1 transcription as well as polymorphism of TNF were investigated. Methods Blood samples were collected from 329 cases non-severe malaria with acute uncomplicated Plasmodium falciparum malaria (UM and 80 cases with Plasmodium vivax malaria (VM, and 77 cases with severe or cerebral malaria (SM for analysis of genetic polymorphisms of HO-1 and TNF and cadmium levels. These patients consisted of 123 (25.3% Thai, 243 (50.0% Burmese and 120 (24.7% Karen who were present at Mae Sot General Hospital, Mae Sot, Tak Province, Thailand. Results The number of (GTn repeats of the HO-1 gene in all patients varied between 16 and 39 and categorized to short (S, medium (M and long (L GTn repeats. The genotype of (GTn repeat of HO-1 was found to be significantly different among the three ethnic groups of patients. Significantly higher frequency of S/L genotype was found in Burmese compared with Thai patients, while significantly lower frequencies of S/S and M/L but higher frequency of M/M genotype was observed in Burmese compared with Karen patients. No significant association between HO-1 and TNF polymorphisms including the inducing effect of cadmium and malaria pathogenicity/severity was observed. Conclusions Difference in the expression of HO-1 genotype in different ethnic groups may contribute to different severity of malaria

  7. The role of heme oxygenase-1 in drug metabolizing dysfunction in the alcoholic fatty liver exposed to ischemic injury

    Park, Sang Won [Department of Pharmacology, Institute of Health Sciences, Gyeongsang National University School of Medicine, Jinju 52727 (Korea, Republic of); Kang, Jung-Woo [School of Pharmacy, Sungkyunkwan University, Suwon, Gyeonggi-do 16419 (Korea, Republic of); Lee, Sun-Mee, E-mail: sunmee@skku.edu [School of Pharmacy, Sungkyunkwan University, Suwon, Gyeonggi-do 16419 (Korea, Republic of)

    2016-02-01

    This study was designed to investigate the role of heme oxygenase-1 (HO-1) in hepatic drug metabolizing dysfunction after ischemia/reperfusion (IR) in alcoholic fatty liver (AFL). Rats were fed a Lieber–DeCarli diet for five weeks to allow for development of AFL and were then subjected to 90 min of hepatic ischemia and 5 h of reperfusion. Rats were pretreated with hemin (HO-1 inducer) or ZnPP (HO-1 inhibitor) for 16 h and 3 h before hepatic ischemia. After hepatic IR, ethanol diet (ED)-fed rats had higher serum aminotransferase activities and more severe hepatic necrosis compared to the control diet (CD)-fed rats. These changes were attenuated by hemin and exacerbated by ZnPP. The activity and gene expression of HO-1 and its transcription factor (Nrf2) level increased significantly after 5 h of reperfusion in CD-fed rats but not in ED-fed rats. After reperfusion, cytochrome P450 (CYP) 1A1, 1A2, and 2B1 activities were reduced to levels lower than those observed in sham group, whereas CYP2E1 activity increased. The decrease in CYP2B1 activity and the increase in CYP2E1 activity were augmented after hepatic IR in ED-fed animals. These changes were significantly attenuated by hemin but aggravated by ZnPP. Finally, CHOP expression and PERK phosphorylation, microsomal lipid peroxidation, and levels of proinflammatory mediators increased in ED-fed rats compared to CD-fed rats after reperfusion. These increases were attenuated by hemin. Our results suggest that AFL exacerbates hepatic drug metabolizing dysfunction during hepatic IR via endoplasmic reticulum stress and lipid peroxidation and this is associated with impaired HO-1 induction. - Highlights: • Endogenous HO-1 is generated in insufficient quantities in steatotic ischemic injury. • Impaired HO-1 induction leads to excessive ER stress response and lipid peroxidation. • Alcoholic steatosis exacerbates IR-induced hepatic drug-metabolizing dysfunction. • HO-1 induction is required for appropriate medication

  8. Effects of Metalloporphyrins on Heme Oxygenase-1 Transcription: Correlative Cell Culture Assays Guide in Vivo Imaging

    Monica Hajdena-Dawson

    2003-07-01

    Full Text Available Heme oxygenase (HO is the rate-limiting step in the heme degradation pathway and is a potential target for the control, or prevention, of pathologic jaundice in neonates. Metalloporphyrins (Mps, a diverse set of synthetic derivatives of heme, can competitively inhibit the HO enzymes. However, certain Mps are phototoxic and some increase transcription of HO-1, the inducible HO isozyme. Therefore, effective development of this class of compounds as therapeutics for treating pathologic jaundice will require rapid and integrated biological screens to identify the most efficacious and safe Mps. To study the safety of these compounds, we assessed their cytotoxic effects and measured luciferase activity by bioluminescent imaging (BLI as an index of HO-1 transcription, first in live cell cultures and then in living transgenic reporter mice. A total of 12 Mps were first evaluated in the correlative cell culture assay. Based on results from this study, 2 Mps, zinc protoporphyrin (ZnPP and zinc bis glycol porphyrin (ZnBG, were selected for further studies in the live animal model. In vitro BLI showed ZnPP to be a strong inducer of HO-1 transcription in comparison to ZnBG, which showed minimal induction. Cytotoxicity studies revealed that ZnPP was phototoxic, whereas ZnBG had no effect on cell viability. In vivo BLI showed that both ZnPP and ZnBG had minimal effects on the levels of HO-1 transcription in the animals. Furthermore, serum enzyme assays indicated that neither caused detectable liver toxicity. These findings, and especially those with ZnBG, support the use of selected Mps as therapies for pathologic jaundice. Coupling the high throughput advantage of cell culture with the capability of imaging for whole-body temporal analyses could accelerate and refine the preclinical phases of drug development. Thus, this study serves as a model for understanding the effects of specific compounds in relation to defined targets using an integrated approach.

  9. Curcumin attenuates quinocetone induced apoptosis and inflammation via the opposite modulation of Nrf2/HO-1 and NF-kB pathway in human hepatocyte L02 cells.

    Dai, Chongshan; Li, Bin; Zhou, Yan; Li, Daowen; Zhang, Shen; Li, Hui; Xiao, Xilong; Tang, Shusheng

    2016-09-01

    The potential toxicity of quinocetone (QCT) has raised widely concern, but its mechanism is still unclear. This study aimed to investigate the protective effect of curcumin on QCT induced apoptosis and the underlying mechanism in human hepatocyte L02 cells. The results showed that QCT treatment significantly decreased the cell viability of L02 cell and increased the release of lactate dehydrogenase (LDH), which was attenuated by curcumin pre-treatment at 1.25, 2.5 and 5 μM. Compared to the QCT alone group, curcumin pre-treatment significantly attenuated QCT induced oxidative stress, mitochondrial dysfunction and apoptosis. In addition, curcumin pretreatment markedly attenuated QCT-induced increase of iNOS activity and NO production in a dose-dependent manner. Meanwhile, curcumin pretreatment markedly down-regulated the expression of nuclear factor -kB (NF-kB) and iNOS mRNAs, but up-regulated the expressions of Nrf2 and HO-1 mRNAs, compared to the QCT alone group. Zinc protoporphyrin IX, a HO-1 inhibitor, markedly partly abolished the cytoprotective effect of curcumin against QCT-induced caspase activation, NF-kB mRNA expression. These results indicate that curcumin could effectively inhibit QCT induced apoptosis and inflammatory response in L02 cells, which may involve the activation of Nrf2/HO-1 and inhibition of NF-kB pathway. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Neutron diffraction studies of Ho1-xYxNi2B2C compounds

    Chang, L.J.; Tomy, C.V.; Paul, D.M.K.

    1996-01-01

    Neutron diffraction measurements have been carried out to investigate the nature of magnetic ordering in Ho(1-x)Y(x)Ni(2)B(2)C (x = 0, 0.1 and 0.2) compounds. HoNi(2)B(2)C shows a complex type of magnetic ordering below the superconducting transition, with a commensurate antiferromagnetic ordering...

  11. The Protective Effect of Lavender Essential Oil and Its Main Component Linalool against the Cognitive Deficits Induced by D-Galactose and Aluminum Trichloride in Mice

    Pan Xu

    2017-01-01

    Full Text Available Lavender essential oil (LO is a traditional medicine used for the treatment of Alzheimer’s disease (AD. It was extracted from Lavandula angustifolia Mill. This study was designed to investigate the effects of lavender essential oil (LO and its active component, linalool (LI, against cognitive impairment induced by D-galactose (D-gal and AlCl3 in mice and to explore the related mechanisms. Our results revealed that LO (100 mg/kg or LI (100 mg/kg significantly protected the cognitive impairments as assessed by the Morris water maze test and step-though test. The mechanisms study demonstrated that LO and LI significantly protected the decreased activity of superoxide dismutase (SOD, glutathione peroxidase (GPX, and protected the increased activity of acetylcholinesterase (AChE and content of malondialdehyde (MDA. Besides, they protected the suppressed nuclear factor-erythroid 2-related factor 2 (Nrf2 and heme oxygenase-1 (HO-1 expression significantly. Moreover, the decreased expression of synapse plasticity-related proteins, calcium-calmodulin-dependent protein kinase II (CaMKII, p-CaMKII, brain-derived neurotrophic factor (BDNF, and TrkB in the hippocampus were increased with drug treatment. In conclusion, LO and its active component LI have protected the oxidative stress, activity of cholinergic function and expression of proteins of Nrf2/HO-1 pathway, and synaptic plasticity. It suggest that LO, especially LI, could be a potential agent for improving cognitive impairment in AD.

  12. Fisetin inhibits TNF-α-induced inflammatory action and hydrogen peroxide-induced oxidative damage in human keratinocyte HaCaT cells through PI3K/AKT/Nrf-2-mediated heme oxygenase-1 expression.

    Seo, Seung-Hee; Jeong, Gil-Saeng

    2015-12-01

    Oxidative skin damage and skin inflammation play key roles in the pathogenesis of skin-related diseases. Fisetin is a naturally occurring flavonoid abundantly found in several vegetables and fruits. Fisetin has been shown to exert various positive biological effects, such as anti-cancer, anti-proliferative, neuroprotective and anti-oxidative effects. In this study, we investigate the skin protective effects and anti-inflammatory properties of fisetin in hydrogen peroxide- and TNF-α-challenged human keratinocyte HaCaT cells. When HaCaT cells were treated with non-cytotoxic concentrations of fisetin (1-20μM), heme oxygenase (HO)-1 mRNA and protein expression increased in a dose-dependent manner. Furthermore, fisetin dose-dependently increased cell viability and reduced ROS production in hydrogen peroxide-treated HaCaT cells. Fisetin also inhibited the production of NO, PGE2 IL-1β, IL-6, expression of iNOS and COX-2, and activation of NF-κB in HaCaT cells treated with TNF-α. Fisetin induced Nrf2 translocation to the nuclei. HO-1 siRNA transient transfection reversed the effects of fisetin on cytoprotection, ROS reduction, NO, PGE2, IL-1β, IL-6, and TNF-α production, and NF-κB DNA-binding activity. Moreover, fisetin increased Akt phosphorylation and a PI3K pathway inhibitor (LY294002) abolished fisetin-induced cytoprotection and NO inhibition. Taken together, these results provide evidence for a beneficial role of fisetin in skin therapy. Copyright © 2015. Published by Elsevier B.V.

  13. Andrographolide exerts anti-hepatitis C virus activity by up-regulating haeme oxygenase-1 via the p38 MAPK/Nrf2 pathway in human hepatoma cells.

    Lee, Jin-Ching; Tseng, Chin-Kai; Young, Kung-Chia; Sun, Hung-Yu; Wang, Shainn-Wei; Chen, Wei-Chun; Lin, Chun-Kuang; Wu, Yu-Hsuan

    2014-01-01

    This study aimed to evaluate the anti-hepatitis C virus (HCV) activity of andrographolide, a diterpenoid lactone extracted from Andrographis paniculata, and to identify the signalling pathway involved in its antiviral action. Using HCV replicon and HCVcc infectious systems, we identified anti-HCV activity of andrographolide by measuring protein and RNA levels. A reporter activity assay was used to determine transcriptional regulation of anti-HCV agents. A specific inhibitor and short hairpin RNAs were used to investigate the mechanism responsible for the effect of andrographolide on HCV replication. In HCV replicon and HCVcc infectious systems, andrographolide time- and dose-dependently suppressed HCV replication. When combined with IFN-α, an inhibitor targeting HCV NS3/4A protease (telaprevir), or NS5B polymerase (PSI-7977), andrographolide exhibited a significant synergistic effect. Andrographolide up-regulated the expression of haeme oxygenase-1 (HO-1), leading to increased amounts of its metabolite biliverdin, which was found to suppress HCV replication by promoting the antiviral IFN responses and inhibiting NS3/4A protease activity. Significantly, these antiviral effects were attenuated by an HO-1-specific inhibitor or HO-1 gene knockdown, indicating that HO-1 contributed to the anti-HCV activity of andrographolide. Andrographolide activated p38 MAPK phosphorylation, which stimulated nuclear factor erythroid 2-related factor 2 (Nrf2)-mediated HO-1 expression, and this was found to be associated with its anti-HCV activity. Our results demonstrate that andrographolide has the potential to control HCV replication and suggest that targeting the Nrf2-HO-1 signalling pathway might be a promising strategy for drug development. © 2013 The British Pharmacological Society.

  14. IN VITRO STUDIES ON HEME OXYGENASE-1 AND P24 ANTIGEN HIV-1 LEVEL AFTERHYPERBARIC OXYGEN TREATMENTOFHIV-1 INFECTED ON PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMCS).

    Budiarti, Retno; Kuntaman; Nasronudin; Suryokusumo; Khairunisa, Siti Qamariyah

    2018-01-01

    Heme oxygenase-1 (HO-1) is a protein secreted by immune cells as a part of immune response mechanism.HO-1 can be induced by variety agents that causingoxidative stress, such as exposure to 100% oxygenat2,4 ATA pressure.It plays a vital role in maintaining cellular homeostasis.This study was conducted to identify the effect of hyperbaric oxygen exposure in cultured ofPBMCthat infected by HIV-1. Primary culture of PBMCs were isolated from 16 healthy volunteers and HIV-1 infected MT4 cell line by co-culture. The PBMCs were aliquoted into two wells as control group and treatment group. The 16 samples of HIV-1 infected PBMCwere exposed to oxygen at 2,4 ATA in animal hyperbaric chamber forthree times in 30 minutes periods with 5 minutes spacing period, that called 1 session.The Treatment done on 5 sessions within 5 days. 16 samples of HIV-1 infected PMBCs that have no hyperbaric treatment became control group.The supernatant were measured the HO-1 production by ELISA andmRNA expression of HO-1 by real time PCR and the number ofantigen p24 HIV-1by ELISA. The result showed that there was no increasing of HO-1 at both mRNA level and protein level, there was a decreasing number of antigen p24 HIV-1 at the treatment group. In addition, hyperbaric exposure could not increase the expression of HO-1, more over the viral replication might be reduced by other mechanism. Hyperbaric oxygen could increases cellular adaptive response of PBMCs infected HIV-1 through increased expression of proteins that can inhibit HIV viralreplication.

  15. EXPRESSION AND CHARACTERIZATION OF FULL-LENGTH HUMAN HEME OXYGENASE-1: PRESENCE OF INTACT MEMBRANE-BINDING REGION LEADS TO INCREASED BINDING AFFINITY FOR NADPH-CYTOCHROME P450 REDUCTASE

    Huber, Warren J.; Backes, Wayne L.

    2009-01-01

    Heme oxygenase (HO) is the chief regulatory enzyme in the oxidative degradation of heme to biliverdin. In the process of heme degradation, this NADPH and cytochrome P450 reductase (CPR)-dependent oxidation of heme also releases free iron and carbon monoxide. Much of the recent research involving heme oxygenase is done using a 30-kDa soluble form of the enzyme, which lacks the membrane binding region (C-terminal 23 amino acids). The goal of this study was to express and purify a full-length human HO-1 (hHO-1) protein; however, due to the lability of the full-length form, a rapid purification procedure was required. This was accomplished by use of a GST-tagged hHO-1 construct. Although the procedure permitted the generation of a full-length HO-1, this form was contaminated with a 30-kDa degradation product that could not be eliminated. Therefore, we attempted to remove a putative secondary thrombin cleavage site by a conservative mutation of amino acid 254, which replaces lysine with arginine. This mutation allowed the expression and purification of a full length hHO-1 protein. Unlike wild-type HO-1, the K254R mutant could be purified to a single 32-kDa protein capable of degrading heme at the same rate as the wild-type enzyme. The K254R full-length form had a specific activity of ~200–225 nmol bilirubin hr−1nmol−1 HO-1 as compared to ~140–150 nmol bilirubin hr−1nmol−1 for the WT form, which contains the 30-kDa contaminant. This is a 2–3-fold increase from the previously reported soluble 30-kDa HO-1, suggesting that the C-terminal 23 amino acids are essential for maximal catalytic activity. Because the membrane spanning domain is present, the full-length hHO-1 has the potential to incorporate into phospholipid membranes, which can be reconstituted at known concentrations, in combination with other ER-resident enzymes. PMID:17915953

  16. Induction of HO-1 in tissue macrophages and monocytes in fatal falciparum malaria and sepsis

    Liomba N

    2003-11-01

    Full Text Available Abstract Background As well as being inducible by haem, haemoxygenase -1 (HO-1 is also induced by interleukin-10 and an anti-inflammatory prostaglandin, 15d PGJ2, the carbon monoxide thus produced mediating the anti-inflammatory effects of these molecules. The cellular distribution of HO-1, by immunohistochemistry, in brain, lung and liver in fatal falciparum malaria, and in sepsis, is reported. Methods Wax sections were stained, at a 1:1000 dilution of primary antibody, for HO-1 in tissues collected during paediatric autopsies in Blantyre, Malawi. These comprised 37 acutely ill comatose patients, 32 of whom were diagnosed clinically as cerebral malaria and the other 5 as bacterial diseases with coma. Another 3 died unexpectedly from an alert state. Other control tissues were from Australian adults. Results Apart from its presence in splenic red pulp macrophages and microhaemorrhages, staining for HO-1 was confined to intravascular monocytes and certain tissue macrophages. Of the 32 clinically diagnosed cerebral malaria cases, 11 (category A cases had negligible histological change in the brain and absence of or scanty intravascular sequestration of parasitized erythrocytes. Of these 11 cases, eight proved at autopsy to have other pathological changes as well, and none of these eight showed HO-1 staining within the brain apart from isolated moderate staining in one case. Two of the three without another pathological diagnosis showed moderate staining of scattered monocytes in brain vessels. Six of these 11 (category A cases exhibited strong lung staining, and the Kupffer cells of nine of them were intensely stained. Of the seven (category B cases with no histological changes in the brain, but appreciable sequestered parasitised erythrocytes present, one was without staining, and the other six showed strongly staining, rare or scattered monocytes in cerebral vessels. All six lung sections not obscured by neutrophils showed strong staining of

  17. Isoporphyrin intermediate in heme oxygenase catalysis. Oxidation of alpha-meso-phenylheme.

    Evans, John P; Niemevz, Fernando; Buldain, Graciela; de Montellano, Paul Ortiz

    2008-07-11

    Human heme oxygenase-1 (hHO-1) catalyzes the O2- and NADPH-dependent oxidation of heme to biliverdin, CO, and free iron. The first step involves regiospecific insertion of an oxygen atom at the alpha-meso carbon by a ferric hydroperoxide and is predicted to proceed via an isoporphyrin pi-cation intermediate. Here we report spectroscopic detection of a transient intermediate during oxidation by hHO-1 of alpha-meso-phenylheme-IX, alpha-meso-(p-methylphenyl)-mesoheme-III, and alpha-meso-(p-trifluoromethylphenyl)-mesoheme-III. In agreement with previous experiments (Wang, J., Niemevz, F., Lad, L., Huang, L., Alvarez, D. E., Buldain, G., Poulos, T. L., and Ortiz de Montellano, P. R. (2004) J. Biol. Chem. 279, 42593-42604), only the alpha-biliverdin isomer is produced with concomitant formation of the corresponding benzoic acid. The transient intermediate observed in the NADPH-P450 reductase-catalyzed reaction accumulated when the reaction was supported by H2O2 and exhibited the absorption maxima at 435 and 930 nm characteristic of an isoporphyrin. Product analysis by reversed phase high performance liquid chromatography and liquid chromatography electrospray ionization mass spectrometry of the product generated with H2O2 identified it as an isoporphyrin that, on quenching, decayed to benzoylbiliverdin. In the presence of H218O2, one labeled oxygen atom was incorporated into these products. The hHO-1-isoporphyrin complexes were found to have half-lives of 1.7 and 2.4 h for the p-trifluoromethyl- and p-methyl-substituted phenylhemes, respectively. The addition of NADPH-P450 reductase to the H2O2-generated hHO-1-isoporphyrin complex produced alpha-biliverdin, confirming its role as a reaction intermediate. Identification of an isoporphyrin intermediate in the catalytic sequence of hHO-1, the first such intermediate observed in hemoprotein catalysis, completes our understanding of the critical first step of heme oxidation.

  18. Involvement of Nrf2-Mediated Upregulation of Heme Oxygenase-1 in Mollugin-Induced Growth Inhibition and Apoptosis in Human Oral Cancer Cells

    Young-Man Lee

    2013-01-01

    Full Text Available Although previous studies have shown that mollugin, a bioactive phytochemical isolated from Rubia cordifolia L. (Rubiaceae, exhibits antitumor effects, its biological activity in oral cancer has not been reported. We thus investigated the effects and putative mechanism of apoptosis induced by mollugin in human oral squamous cell carcinoma cells (OSCCs. Results show that mollugin induces cell death in a dose-dependent manner in primary and metastatic OSCCs. Mollugin-induced cell death involved apoptosis, characterized by the appearance of nuclear shrinkage, flow cytometric analysis of sub-G1 phase arrest, and annexin V-FITC and propidium iodide staining. Western blot analysis and RT-PCR revealed that mollugin suppressed activation of NF-κB and NF-κB-dependent gene products involved in antiapoptosis (Bcl-2 and Bcl-xl, invasion (MMP-9 and ICAM-1, and angiogenesis (FGF-2 and VEGF. Furthermore, mollugin induced the activation of p38, ERK, and JNK and the expression of heme oxygenase-1 (HO-1 and nuclear factor E2–related factor 2 (Nrf2. Mollugin-induced growth inhibition and apoptosis of HO-1 were reversed by an HO-1 inhibitor and Nrf2 siRNA. Collectively, this is the first report to demonstrate the effectiveness of mollugin as a candidate for a chemotherapeutic agent in OSCCs via the upregulation of the HO-1 and Nrf2 pathways and the downregulation of NF-κB.

  19. Andrographolide induces Nrf2 and heme oxygenase 1 in astrocytes by activating p38 MAPK and ERK.

    Wong, Siew Ying; Tan, Michelle G K; Wong, Peter T H; Herr, Deron R; Lai, Mitchell K P

    2016-09-23

    Andrographolide is the major labdane diterpenoid originally isolated from Andrographis paniculata and has been shown to have anti-inflammatory and antioxidative effects. However, there is a dearth of studies on the potential therapeutic utility of andrographolide in neuroinflammatory conditions. Here, we aimed to investigate the mechanisms underlying andrographolide's effect on the expression of anti-inflammatory and antioxidant heme oxygenase-1 (HO-1) in primary astrocytes. Measurements of the effects of andrograholide on antioxidant HO-1 and its transcription factor, Nrf2, include gene expression, protein turnover, and activation of putative signaling regulators. Andrographolide potently activated Nrf2 and also upregulated HO-1 expression in primary astrocytes. Andrographolide's effects on Nrf2 seemed to be biphasic, with acute (within 1 h) reductions in Nrf2 ubiquitination efficiency and turnover rate, followed by upregulation of Nrf2 mRNA between 8 and 24 h. The acute regulation of Nrf2 by andrographolide seemed to be independent of Keap1 and partly mediated by p38 MAPK and ERK signaling. These data provide further insights into the mechanisms underlying andrographolide's effects on astrocyte-mediated antioxidant, and anti-inflammatory responses and support the further assessment of andrographolide as a potential therapeutic for neurological conditions in which oxidative stress and neuroinflammation are implicated.

  20. DJ-1 Modulates Nuclear Erythroid 2-Related Factor-2-Mediated Protection in Human Primary Alveolar Type II Cells in Smokers.

    Bahmed, Karim; Messier, Elise M; Zhou, Wenbo; Tuder, Rubin M; Freed, Curt R; Chu, Hong Wei; Kelsen, Steven G; Bowler, Russell P; Mason, Robert J; Kosmider, Beata

    2016-09-01

    Cigarette smoke (CS) is a main source of oxidative stress and a key risk factor for emphysema, which consists of alveolar wall destruction. Alveolar type (AT) II cells are in the gas exchange regions of the lung. We isolated primary ATII cells from deidentified organ donors whose lungs were not suitable for transplantation. We analyzed the cell injury obtained from nonsmokers, moderate smokers, and heavy smokers. DJ-1 protects cells from oxidative stress and induces nuclear erythroid 2-related factor-2 (Nrf2) expression, which activates the antioxidant defense system. In ATII cells isolated from moderate smokers, we found DJ-1 expression by RT-PCR, and Nrf2 and heme oxygenase (HO)-1 translocation by Western blotting and immunocytofluorescence. In ATII cells isolated from heavy smokers, we detected Nrf2 and HO-1 cytoplasmic localization. Moreover, we found high oxidative stress, as detected by 4-hydroxynonenal (4-HNE) (immunoblotting), inflammation by IL-8 and IL-6 levels by ELISA, and apoptosis by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay in ATII cells obtained from heavy smokers. Furthermore, we detected early DJ-1 and late Nrf2 expression after ATII cell treatment with CS extract. We also overexpressed DJ-1 by adenovirus construct and found that this restored Nrf2 and HO-1 expression and induced nuclear translocation in heavy smokers. Moreover, DJ-1 overexpression also decreased ATII cell apoptosis caused by CS extract in vitro. Our results indicate that DJ-1 activates the Nrf2-mediated antioxidant defense system. Furthermore, DJ-1 overexpression can restore the impaired Nrf2 pathway, leading to ATII cell protection in heavy smokers. This suggests a potential therapeutic strategy for targeting DJ-1 in CS-related lung diseases.

  1. 4-Hydroxy hexenal derived from docosahexaenoic acid protects endothelial cells via Nrf2 activation.

    Atsushi Ishikado

    Full Text Available Recent studies have proposed that n-3 polyunsaturated fatty acids (n-3 PUFAs have direct antioxidant and anti-inflammatory effects in vascular tissue, explaining their cardioprotective effects. However, the molecular mechanisms are not yet fully understood. We tested whether n-3 PUFAs showed antioxidant activity through the activation of nuclear factor erythroid 2-related factor 2 (Nrf2, a master transcriptional factor for antioxidant genes. C57BL/6 or Nrf2(-/- mice were fed a fish-oil diet for 3 weeks. Fish-oil diet significantly increased the expression of heme oxygenase-1 (HO-1, and endothelium-dependent vasodilation in the aorta of C57BL/6 mice, but not in the Nrf2(-/- mice. Furthermore, we observed that 4-hydroxy hexenal (4-HHE, an end-product of n-3 PUFA peroxidation, was significantly increased in the aorta of C57BL/6 mice, accompanied by intra-aortic predominant increase in docosahexaenoic acid (DHA rather than that in eicosapentaenoic acid (EPA. Human umbilical vein endothelial cells were incubated with DHA or EPA. We found that DHA, but not EPA, markedly increased intracellular 4-HHE, and nuclear expression and DNA binding of Nrf2. Both DHA and 4-HHE also increased the expressions of Nrf2 target genes including HO-1, and the siRNA of Nrf2 abolished these effects. Furthermore, DHA prevented oxidant-induced cellular damage or reactive oxygen species production, and these effects were disappeared by an HO-1 inhibitor or the siRNA of Nrf2. Thus, we found protective effects of DHA through Nrf2 activation in vascular tissue, accompanied by intra-vascular increases in 4-HHE, which may explain the mechanism of the cardioprotective effects of DHA.

  2. The 1-Tosylpentan-3-one Protects against 6-Hydroxydopamine-Induced Neurotoxicity

    Chien-Jen Kao

    2017-05-01

    Full Text Available Previous studies have demonstrated that the marine compound austrasulfone, isolated from the soft coral Cladiella australis, exerts a neuroprotective effect. The intermediate product in the synthesis of austrasulfone, dihydroaustrasulfone alcohol, attenuates several inflammatory responses. The present study uses in vitro and in vivo methods to investigate the neuroprotective effect of dihydroaustrasulfone alcohol-modified 1-tosylpentan-3-one (1T3O. Results from in vitro experiments show that 1T3O effectively inhibits 6-hydroxydopamine-induced (6-OHDA-induced activation of both p38 mitogen-activated protein kinase (MAPK and caspase-3 in SH-SY5Y cells; and enhances nuclear factor erythroid 2–related factor 2 (Nrf2 and heme oxygenase-1 (HO-1 expression via phosphoinositide 3-kinase (PI3K/protein kinase B (Akt signaling. Hoechst staining and Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL staining results reveal that 1T3O significantly inhibits 6-OHDA-induced apoptosis. In addition, the addition of an Akt or HO-1 inhibitor decreases the protective effect of 1T3O. Thus, we hypothesize that the anti-apoptotic activity of 1T3O in neuronal cells is mediated through the regulation of the Akt and HO-1 signaling pathways. In vivo experiments show that 1T3O can reverse 6-OHDA-induced reduction in locomotor behavior ability in zebrafish larvae, and inhibit 6-OHDA-induced tumor necrosis factor-alpha (TNF-α increase at the same time. According to our in vitro and in vivo results, we consider that 1T3O exerts its anti-apoptotic activities at SH-SY5Y cells after 6-OHDA challenges, probably via the regulation of anti-oxidative signaling pathways. Therefore, this compound may be a promising therapeutic agent for neurodegenerations.

  3. Role of the heme oxygenases in abnormalities of the mesenteric circulation in cirrhotic rats.

    Sacerdoti, David; Abraham, Nader G; Oyekan, Adebayo O; Yang, Liming; Gatta, Angelo; McGiff, John C

    2004-02-01

    Carbon monoxide (CO), a product of heme metabolism by heme-oxygenase (HO), has biological actions similar to those of nitric oxide (NO). The role of CO in decreasing vascular responses to constrictor agents produced by experimental cirrhosis induced by carbon tetrachloride was evaluated before and after inhibition of HO with tin-mesoporphyrin (SnMP) in the perfused superior mesenteric vasculature (SMV) of cirrhotic and normal rats and in normal rats transfected with the human HO-1 (HHO-1) gene. Perfusion pressure and vasoconstrictor responses of the SMV to KCl, phenylephrine (PE), and endothelin-1 (ET-1) were decreased in cirrhotic rats. SnMP increased SMV perfusion pressure and restored the constrictor responses of the SMV to KCl, PE, and ET-1 in cirrhotic rats. The relative roles of NO and CO in producing hyporeactivity of the SMV to PE in cirrhotic rats were examined. Vasoconstrictor responses to PE were successively augmented by stepwise inhibition of CO and NO production, suggesting a complementary role for these gases in the regulation of reactivity of the SMV. Expression of constitutive but not of inducible HO (HO-1) was increased in the SMV of cirrhotic rats as was HO activity. Administration of adenovirus containing HHO-1 gene produced detection of HHO-1 RNA and increased HO activity in the SMV within 7 days. Rats transfected with HO-1 demonstrated reduction in both perfusion pressure and vasoconstrictor responses to PE in the SMV. We propose that HO is an essential component in mechanisms that modulate reactivity of the mesenteric circulation in experimental hepatic cirrhosis in rats.

  4. Heme Oxygenase Induction Suppresses Hepatic Hepcidin and Rescues Ferroportin and Ferritin Expression in Obese Mice

    Nitin Puri

    2017-01-01

    Full Text Available Hepcidin, a phase II reactant secreted by hepatocytes, regulates cellular iron levels by increasing internalization of ferroportin-a transmembrane protein facilitating egress of cellular iron. Chronic low-grade inflammatory states, such as obesity, have been shown to increase oxidative stress and enhance hepcidin secretion from hepatocytes and macrophages. Heme-heme oxygenase (HO is a stress response system which reduces oxidative stress. We investigated the effects of HO-1 induction on hepatic hepcidin levels and on iron homeostasis in hepatic tissues from lean and obese mice. Obese mice exhibited hyperglycemia (p<0.05; increased levels of proinflammatory cytokines (MCP-1, IL-6, p<0.05; oxidative stress (p<0.05; and increased hepatic hepcidin levels (p<0.05. Enhancement of hepcidin was reflected in the reduced expression of ferroportin in obese mice (p<0.05. However, this effect is accompanied by a significant decline in ferritin expression. Additionally, there are reduced insulin receptor phosphorylation and attenuation of metabolic regulators pAMPK, pAKT, and pLKB1. Cobalt protoporphyrin- (CoPP- induced HO-1 upregulation in obese mice reversed these alterations (p<0.05, while attenuating hepatic hepcidin levels. These effects of CoPP were prevented in obese mice concurrently exposed to an inhibitor of HO (SnMP (p<0.05. Our results highlight a modulatory effect of HO on iron homeostasis mediated through the suppression of hepatic hepcidin.

  5. Human heme oxygenase-1 gene transfer lowers blood pressure and promotes growth in spontaneously hypertensive rats.

    Sabaawy, H E; Zhang, F; Nguyen, X; ElHosseiny, A; Nasjletti, A; Schwartzman, M; Dennery, P; Kappas, A; Abraham, N G

    2001-08-01

    Heme oxygenase (HO) catalyzes the conversion of heme to biliverdin, with release of free iron and carbon monoxide. Both heme and carbon monoxide have been implicated in the regulation of vascular tone. A retroviral vector containing human HO-1 cDNA (LSN-HHO-1) was constructed and subjected to purification and concentration of the viral particles to achieve 5x10(9) to 1x10(10) colony-forming units per milliliter. The ability of concentrated infectious viral particles to express human HO-1 (HHO-1) in vivo was tested. A single intracardiac injection of the concentrated infectious viral particles (expressing HHO-1) to 5-day-old spontaneously hypertensive rats resulted in functional expression of the HHO-1 gene and attenuation of the development of hypertension. Rats expressing HHO-1 showed a significant decrease in urinary excretion of a vasoconstrictor arachidonic acid metabolite and a reduction in myogenic responses to increased intraluminal pressure in isolated arterioles. Unexpectedly, HHO-1 chimeric rats showed a simultaneous significant proportionate increase in somatic growth. Thus, delivery of HHO-1 gene by retroviral vector attenuates the development of hypertension and promotes body growth in spontaneously hypertensive rats.

  6. The nuclear factor-erythroid 2-related factor/heme oxygenase-1 axis is critical for the inflammatory features of type 2 diabetes-associated osteoarthritis.

    Vaamonde-Garcia, Carlos; Courties, Alice; Pigenet, Audrey; Laiguillon, Marie-Charlotte; Sautet, Alain; Houard, Xavier; Kerdine-Römer, Saadia; Meijide, Rosa; Berenbaum, Francis; Sellam, Jérémie

    2017-09-01

    Epidemiological findings support the hypothesis that type 2 diabetes mellitus (T2DM) is a risk factor for osteoarthritis (OA). Moreover, OA cartilage from patients with T2DM exhibits a greater response to inflammatory stress, but the molecular mechanism is unclear. To investigate whether the antioxidant defense system participates in this response, we examined here the expression of nuclear factor-erythroid 2-related factor (Nrf-2), a master antioxidant transcription factor, and of heme oxygenase-1 (HO-1), one of its main target genes, in OA cartilage from T2DM and non-T2DM patients as well as in murine chondrocytes exposed to high glucose (HG). Ex vivo experiments indicated that Nrf-2 and HO-1 expression is reduced in T2DM versus non-T2DM OA cartilage (0.57-fold Nrf-2 and 0.34-fold HO-1), and prostaglandin E 2 (PGE 2 ) release was increased in samples with low HO-1 expression. HG-exposed, IL-1β-stimulated chondrocytes had lower Nrf-2 levels in vitro , particularly in the nuclear fraction, than chondrocytes exposed to normal glucose (NG). Accordingly, HO-1 levels were also decreased (0.49-fold) in these cells. The HO-1 inducer cobalt protoporphyrin IX more efficiently attenuated PGE 2 and IL-6 release in HG+IL-1β-treated cells than in NG+IL-1β-treated cells. Greater reductions in HO-1 expression and increase in PGE 2 /IL-6 production were observed in HG+IL-1β-stimulated chondrocytes from Nrf-2 -/- mice than in chondrocytes from wild-type mice. We conclude that the Nrf-2/HO-1 axis is a critical pathway in the hyperglucidic-mediated dysregulation of chondrocytes. Impairments in this antioxidant system may explain the greater inflammatory responsiveness of OA cartilage from T2DM patients and may inform treatments of such patients. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  7. Metformin inhibits heme oxygenase-1 expression in cancer cells through inactivation of Raf-ERK-Nrf2 signaling and AMPK-independent pathways

    Do, Minh Truong; Kim, Hyung Gyun; Khanal, Tilak; Choi, Jae Ho; Kim, Dong Hee; Jeong, Tae Cheon; Jeong, Hye Gwang

    2013-01-01

    Resistance to therapy is the major obstacle to more effective cancer treatment. Heme oxygenase-1 (HO-1) is often highly up-regulated in tumor tissues, and its expression is further increased in response to therapies. It has been suggested that inhibition of HO-1 expression is a potential therapeutic approach to sensitize tumors to chemotherapy and radiotherapy. In this study, we tested the hypothesis that the anti-tumor effects of metformin are mediated by suppression of HO-1 expression in cancer cells. Our results indicate that metformin strongly suppresses HO-1 mRNA and protein expression in human hepatic carcinoma HepG2, cervical cancer HeLa, and non-small-cell lung cancer A549 cells. Metformin also markedly reduced Nrf2 mRNA and protein levels in whole cell lysates and suppressed tert-butylhydroquinone (tBHQ)-induced Nrf2 protein stability and antioxidant response element (ARE)-luciferase activity in HepG2 cells. We also found that metformin regulation of Nrf2 expression is mediated by a Keap1-independent mechanism and that metformin significantly attenuated Raf-ERK signaling to suppress Nrf2 expression in cancer cells. Inhibition of Raf-ERK signaling by PD98059 decreased Nrf2 mRNA expression in HepG2 cells, confirming that the inhibition of Nrf2 expression is mediated by an attenuation of Raf-ERK signaling in cancer cells. The inactivation of AMPK by siRNA, DN-AMPK or the pharmacological AMPK inhibitor compound C, revealed that metformin reduced HO-1 expression in an AMPK-independent manner. These results highlight the Raf-ERK-Nrf2 axis as a new molecular target in anticancer therapy in response to metformin treatment. - Highlights: • Metformin inhibits HO-1 expression in cancer cells. • Metformin attenuates Raf-ERK-Nrf2 signaling. • Suppression of HO-1 by metformin is independent of AMPK. • HO-1 inhibition contributes to anti-proliferative effects of metformin

  8. The protective effect of magnesium lithospermate B against glucose-induced intracellular oxidative damage

    Qu, Jian [Institute of Clinical Pharmacology, Hunan Key Laboratory of Pharmacogenetics, Central South University, Xiangya School of Medicine, Changsha 410078 (China); Ren, Xian [Shanghai Green Valley Pharmaceutical Co., Ltd., Shanghai 201304 (China); Hou, Rui-ying; Dai, Xing-ping [Institute of Clinical Pharmacology, Hunan Key Laboratory of Pharmacogenetics, Central South University, Xiangya School of Medicine, Changsha 410078 (China); Zhao, Ying-chun [Laboratories of Functional Genomics and Proteomics, Creighton University Medical Center, Omaha, NE 68131 (United States); Xu, Xiao-jing; Zhang, Wei; Zhou, Gan; Zhou, Hong-hao [Institute of Clinical Pharmacology, Hunan Key Laboratory of Pharmacogenetics, Central South University, Xiangya School of Medicine, Changsha 410078 (China); Liu, Zhao-qian, E-mail: liuzhaoqian63@126.com [Institute of Clinical Pharmacology, Hunan Key Laboratory of Pharmacogenetics, Central South University, Xiangya School of Medicine, Changsha 410078 (China)

    2011-07-22

    Highlights: {yields} LAB reduced the ROS production in HEK293T cells cultured under oxidative stress. High dose of glucose enhanced the expression of HO-1 mRNA and HO-1 protein in a time-dependent manner. {yields} LAB enhanced the expression of HO-1 mRNA and HO-1 protein in a dose-dependent manner treated with high dose of glucose. {yields} LAB plays an important role against glucose-induced intracellular oxidative damage. {yields} The enhanced expression of HO-1 mRNA and HO-1 protein caused by LAB is regulated via Nrf2 signal pathway. -- Abstract: Objectives: To investigate the effects of magnesium lithospermate B (LAB) on intracellular reactive oxygen species (ROS) production induced by high dose of glucose or H{sub 2}O{sub 2}, we explored the influences of LAB on the expression of heme oxygenase-1 (HO-1) and nuclear factor E2-related factor-2 (Nrf2) in HEK293T cells after treatment with high dose of glucose. Materials and methods: The total nuclear proteins in HEK293T cells were extracted with Cytoplasmic Protein Extraction Kit. The ROS level was determined by flow cytometry. The mRNA and protein expression of HO-1 and Nrf2 were determined by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and Western blot. Results: LAB reduced the ROS production in HEK293T cells cultured under oxidative stress. High dose of glucose enhanced the expression of HO-1 mRNA and HO-1 protein in a time-dependent manner. LAB enhanced the expression of HO-1 mRNA and HO-1 protein in a dose-dependent manner treated with high dose of glucose. The amount of Nrf2 translocation was enhanced after cells were pretreated with 50 {mu}mol/L or 100 {mu}mol/L LAB. Silencing of Nrf2 gene eliminated the enhanced expression of HO-1 protein induced by high dose of glucose plus LAB. Conclusions: LAB plays an important role against glucose-induced intracellular oxidative damage. The enhanced expression of HO-1 mRNA and HO-1 protein caused by LAB is regulated via Nrf2 signal pathway.

  9. 3D nuclear organization of telomeres in the Hodgkin cell lines U-HO1 and U-HO1-PTPN1: PTPN1 expression prevents the formation of very short telomeres including "t-stumps"

    Lemieux Bruno

    2010-12-01

    Full Text Available Abstract Background In cancer cells the three-dimensional (3D telomere organization of interphase nuclei into a telomeric disk is heavily distorted and aggregates are found. In Hodgkin's lymphoma quantitative FISH (3D Q-FISH reveals a major impact of nuclear telomere dynamics during the transition form mononuclear Hodgkin (H to diagnostic multinuclear Reed-Sternberg (RS cells. In vitro and in vivo formation of RS-cells is associated with the increase of very short telomeres including "t-stumps", telomere loss, telomeric aggregate formation and the generation of "ghost nuclei". Results Here we analyze the 3D telomere dynamics by Q-FISH in the novel Hodgkin cell line U-HO1 and its non-receptor protein-tyrosine phosphatase N1 (PTPN1 stable transfectant U-HO1-PTPN1, derived from a primary refractory Hodgkin's lymphoma. Both cell lines show equally high telomerase activity but U-HO1-PTPN differs from U-HO1 by a three times longer doubling time, low STAT5A expression, accumulation of RS-cells (p As expected, multinuclear U-HO1-RS-cells and multinuclear U-HO1-PTPN1-RS-cells differ from their mononuclear H-precursors by their nuclear volume (p Conclusion Abundant RS-cells without additional very short telomeres including "t-stumps", high rate of apoptosis, but low STAT5A expression, are hallmarks of the U-HO1-PTPN1 cell line. These characteristics are independent of telomerase activity. Thus, PTPN1 induced dephosphorylation of STAT5 with consecutive lack of Akt/PKB activation and cellular arrest in G2, promoting induction of apoptosis, appears as a possible pathogenetic mechanism deserving further experimental investigation.

  10. Transition between acute and chronic hepatotoxicity in mice is associated with impaired energy metabolism and induction of mitochondrial heme oxygenase-1.

    Aniket Nikam

    Full Text Available The formation of protein inclusions is frequently associated with chronic metabolic diseases. In mice, short-term intoxication with 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC leads to hepatocellular damage indicated by elevated serum liver enzyme activities, whereas only minor morphological changes are observed. Conversely, chronic administration of DDC for several weeks results in severe morphological damage, characterized by hepatocellular ballooning, disruption of the intermediate filament cytoskeleton, and formation of Mallory-Denk bodies consisting predominantly of misfolded keratins, Sqstm1/p62, and heat shock proteins. To evaluate the mechanistic underpinnings for this dichotomy we dissected the time-course of DDC intoxication for up to 10 weeks. We determined body weight change, serum liver enzyme activities, morphologic alterations, induction of antioxidant response (heme oxygenase-1, HO-1, oxidative damage and ATP content in livers as well as respiration, oxidative damage and the presence and activity of HO-1 in endoplasmic reticulum and mitochondria (mtHO-1. Elevated serum liver enzyme activity and oxidative liver damage were already present at early intoxication stages without further subsequent increase. After 2 weeks of intoxication, mice had transiently lost 9% of their body weight, liver ATP-content was reduced to 58% of controls, succinate-driven respiration was uncoupled from ATP-production and antioxidant response was associated with the appearance of catalytically active mtHO-1. Oxidative damage was associated with both acute and chronic DDC toxicity whereas the onset of chronic intoxication was specifically associated with mitochondrial dysfunction which was maximal after 2 weeks of intoxication. At this transition stage, adaptive responses involving mtHO-1 were induced, indirectly leading to improved respiration and preventing further drop of ATP levels. Our observations clearly demonstrate principally different

  11. DJ-1 Modulates Nuclear Erythroid 2–Related Factor-2–Mediated Protection in Human Primary Alveolar Type II Cells in Smokers

    Bahmed, Karim; Messier, Elise M.; Zhou, Wenbo; Tuder, Rubin M.; Freed, Curt R.; Chu, Hong Wei; Kelsen, Steven G.; Bowler, Russell P.; Mason, Robert J.

    2016-01-01

    Cigarette smoke (CS) is a main source of oxidative stress and a key risk factor for emphysema, which consists of alveolar wall destruction. Alveolar type (AT) II cells are in the gas exchange regions of the lung. We isolated primary ATII cells from deidentified organ donors whose lungs were not suitable for transplantation. We analyzed the cell injury obtained from nonsmokers, moderate smokers, and heavy smokers. DJ-1 protects cells from oxidative stress and induces nuclear erythroid 2–related factor-2 (Nrf2) expression, which activates the antioxidant defense system. In ATII cells isolated from moderate smokers, we found DJ-1 expression by RT-PCR, and Nrf2 and heme oxygenase (HO)-1 translocation by Western blotting and immunocytofluorescence. In ATII cells isolated from heavy smokers, we detected Nrf2 and HO-1 cytoplasmic localization. Moreover, we found high oxidative stress, as detected by 4-hydroxynonenal (4-HNE) (immunoblotting), inflammation by IL-8 and IL-6 levels by ELISA, and apoptosis by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay in ATII cells obtained from heavy smokers. Furthermore, we detected early DJ-1 and late Nrf2 expression after ATII cell treatment with CS extract. We also overexpressed DJ-1 by adenovirus construct and found that this restored Nrf2 and HO-1 expression and induced nuclear translocation in heavy smokers. Moreover, DJ-1 overexpression also decreased ATII cell apoptosis caused by CS extract in vitro. Our results indicate that DJ-1 activates the Nrf2-mediated antioxidant defense system. Furthermore, DJ-1 overexpression can restore the impaired Nrf2 pathway, leading to ATII cell protection in heavy smokers. This suggests a potential therapeutic strategy for targeting DJ-1 in CS-related lung diseases. PMID:27093578

  12. A Novel, “Double-Clamp” Binding Mode for Human Heme Oxygenase-1 Inhibition

    Rahman, Mona N.; Vlahakis, Jason Z.; Vukomanovic, Dragic; Lee, Wallace; Szarek, Walter A.; Nakatsu, Kanji; Jia, Zongchao

    2012-01-01

    The development of heme oxygenase (HO) inhibitors is critical in dissecting and understanding the HO system and for potential therapeutic applications. We have established a program to design and optimize HO inhibitors using structure-activity relationships in conjunction with X-ray crystallographic analyses. One of our previous complex crystal structures revealed a putative secondary hydrophobic binding pocket which could be exploited for a new design strategy by introducing a functional group that would fit into this potential site. To test this hypothesis and gain further insights into the structural basis of inhibitor binding, we have synthesized and characterized 1-(1H-imidazol-1-yl)-4,4-diphenyl-2-butanone (QC-308). Using a carbon monoxide (CO) formation assay on rat spleen microsomes, the compound was found to be ∼15 times more potent (IC50 = 0.27±0.07 µM) than its monophenyl analogue, which is already a potent compound in its own right (QC-65; IC50 = 4.0±1.8 µM). The crystal structure of hHO-1 with QC-308 revealed that the second phenyl group in the western region of the compound is indeed accommodated by a definitive secondary proximal hydrophobic pocket. Thus, the two phenyl moieties are each stabilized by distinct hydrophobic pockets. This “double-clamp” binding offers additional inhibitor stabilization and provides a new route for improvement of human heme oxygenase inhibitors. PMID:22276118

  13. A novel, "double-clamp" binding mode for human heme oxygenase-1 inhibition.

    Mona N Rahman

    Full Text Available The development of heme oxygenase (HO inhibitors is critical in dissecting and understanding the HO system and for potential therapeutic applications. We have established a program to design and optimize HO inhibitors using structure-activity relationships in conjunction with X-ray crystallographic analyses. One of our previous complex crystal structures revealed a putative secondary hydrophobic binding pocket which could be exploited for a new design strategy by introducing a functional group that would fit into this potential site. To test this hypothesis and gain further insights into the structural basis of inhibitor binding, we have synthesized and characterized 1-(1H-imidazol-1-yl-4,4-diphenyl-2-butanone (QC-308. Using a carbon monoxide (CO formation assay on rat spleen microsomes, the compound was found to be ∼15 times more potent (IC(50 = 0.27±0.07 µM than its monophenyl analogue, which is already a potent compound in its own right (QC-65; IC(50 = 4.0±1.8 µM. The crystal structure of hHO-1 with QC-308 revealed that the second phenyl group in the western region of the compound is indeed accommodated by a definitive secondary proximal hydrophobic pocket. Thus, the two phenyl moieties are each stabilized by distinct hydrophobic pockets. This "double-clamp" binding offers additional inhibitor stabilization and provides a new route for improvement of human heme oxygenase inhibitors.

  14. A novel, "double-clamp" binding mode for human heme oxygenase-1 inhibition.

    Rahman, Mona N; Vlahakis, Jason Z; Vukomanovic, Dragic; Lee, Wallace; Szarek, Walter A; Nakatsu, Kanji; Jia, Zongchao

    2012-01-01

    The development of heme oxygenase (HO) inhibitors is critical in dissecting and understanding the HO system and for potential therapeutic applications. We have established a program to design and optimize HO inhibitors using structure-activity relationships in conjunction with X-ray crystallographic analyses. One of our previous complex crystal structures revealed a putative secondary hydrophobic binding pocket which could be exploited for a new design strategy by introducing a functional group that would fit into this potential site. To test this hypothesis and gain further insights into the structural basis of inhibitor binding, we have synthesized and characterized 1-(1H-imidazol-1-yl)-4,4-diphenyl-2-butanone (QC-308). Using a carbon monoxide (CO) formation assay on rat spleen microsomes, the compound was found to be ∼15 times more potent (IC(50) = 0.27±0.07 µM) than its monophenyl analogue, which is already a potent compound in its own right (QC-65; IC(50) = 4.0±1.8 µM). The crystal structure of hHO-1 with QC-308 revealed that the second phenyl group in the western region of the compound is indeed accommodated by a definitive secondary proximal hydrophobic pocket. Thus, the two phenyl moieties are each stabilized by distinct hydrophobic pockets. This "double-clamp" binding offers additional inhibitor stabilization and provides a new route for improvement of human heme oxygenase inhibitors.

  15. Structure-Activity Relationships of 1,2-Disubstituted Benzimidazoles: Selective Inhibition of Heme Oxygenase-2 Activity.

    Kong, Xianqi; Vukomanovic, Dragic; Nakatsu, Kanji; Szarek, Walter A

    2015-08-01

    Devising ways to up- or down-regulate heme oxygenase activity is attracting much interest as a strategy for the treatment of a variety of disorders. With a view of obtaining compounds that exhibit high potency and selectivity as inhibitors of the heme oxygenase-2 (HO-2) isozyme (constitutive) relative to the heme oxygenase-1 (HO-1) isozyme (inducible), several 1,2-disubstituted 1H-benzimidazoles were designed and synthesized. Specifically, analogues were synthesized in which the C2 substituent was the following: (1H-imidazol-1-yl)methyl, (N-morpholinyl)methyl, cyclopentylmethyl, cyclohexylmethyl, or (norborn-2-yl)methyl. Compounds with the cyclic system in the C2 substituent being a carbocyclic ring, especially cyclohexyl or norborn-2-yl, and the N1 substituent being a ring-substituted benzyl group, especially 4-chlorobenzyl or 4-bromobenzyl, best exhibited the target criteria of high potency and selectivity toward inhibition of HO-2. The new candidates should be useful pharmacological tools and may have therapeutic applications. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Andrographolide inhibits TNFα-induced ICAM-1 expression via suppression of NADPH oxidase activation and induction of HO-1 and GCLM expression through the PI3K/Akt/Nrf2 and PI3K/Akt/AP-1 pathways in human endothelial cells.

    Lu, Chia-Yang; Yang, Ya-Chen; Li, Chien-Chun; Liu, Kai-Li; Lii, Chong-Kuei; Chen, Haw-Wen

    2014-09-01

    Andrographolide, the major bioactive component of Andrographis paniculata, has been demonstrated to have various biological properties including anti-inflammation, antioxidation, and anti-hepatotoxicity. Oxidative stress is considered a major risk factor in aging, inflammation, cancer, atherosclerosis, and diabetes mellitus. NADPH oxidase is a major source of endogenous reactive oxygen species (ROS). In this study, we used EA.hy926 endothelial-like cells to explore the anti-inflammatory activity of andrographolide. Andrographolide attenuated TNFα-induced ROS generation, Src phosphorylation, membrane translocation of the NADPH oxidase subunits p47(phox) and p67(phox), and ICAM-1 gene expression. In the small hairpin RNA interference assay, shp47(phox) abolished TNFα-induced p65 nuclear translocation, ICAM-1 gene expression, and adhesion of HL-60 cells. Andrographolide induced the gene expression of heme oxygenase 1 (HO-1) and glutamate cysteine ligase modifier subunit (GCLM) in a time-dependent manner. Cellular glutathione (GSH) content was increased by andrographolide. shGCLM attenuated the andrographolide-induced increase in GSH content and reversed the andrographolide inhibition of HL-60 adhesion. shHO-1 showed a similar effect on andrographolide inhibition of HL-60 adhesion to shGCLM. The mechanism underlying the up-regulation of HO-1 and GCLM by andrographolide was dependent on the PI3K/Akt pathway, and both the Nrf2 and AP-1 transcriptional factors were involved. Our results suggest that andrographolide attenuates TNFα-induced ICAM-1 expression at least partially through suppression of NADPH oxidase activation and induction of HO-1 and GCLM expression, which is PI3K/Akt pathway-dependent. Copyright © 2014. Published by Elsevier Inc.

  17. Amelioration of Ischemia/Reperfusion Injury During Resuscitation from Hemorrhage by Induction of Heme Oxygenase-1 (HO-1) in a Conscious Mouse Model of Uncontrolled Hemorrhage

    2013-10-01

    against menadione -induced-oxidative stress, also induces HIF1a and this may explain their cytoprotective effect. KEY RESEARCH ACCOMPLISHMENTS: Due to...August 2010 - October 2011 : Compared CDDO-lm, CAPE, CAPA induced H0-1 mediated cytoprotection against menadione -induced-oxidative stress in HUVEC cells...Pharmaceutical Scientists (AAPS) 2013: Comparison of atmospheric oxygen versus physiological levels on cytotoxicity of menadione and cytoprotection by

  18. Rapid, convenient method for screening imidazole-containing compounds for heme oxygenase inhibition.

    Vlahakis, Jason Z; Rahman, Mona N; Roman, Gheorghe; Jia, Zongchao; Nakatsu, Kanji; Szarek, Walter A

    2011-01-01

    Sensitive assays for measuring heme oxygenase activity have been based on the gas-chromatographic detection of carbon monoxide using elaborate, expensive equipment. The present study describes a rapid and convenient method for screening imidazole-containing candidates for inhibitory activity against heme oxygenase using a plate reader, based on the spectroscopic evaluation of heme degradation. A PowerWave XS plate reader was used to monitor the absorbance (as a function of time) of heme bound to purified truncated human heme oxygenase-1 (hHO-1) in the individual wells of a standard 96-well plate (with or without the addition of a test compound). The degradation of heme by heme oxygenase-1 was initiated using l-ascorbic acid, and the collected relevant absorbance data were analyzed by three different methods to calculate the percent control activity occurring in wells containing test compounds relative to that occurring in control wells with no test compound present. In the cases of wells containing inhibitory compounds, significant shifts in λ(max) from 404 to near 412 nm were observed as well as a decrease in the rate of heme degradation relative to that of the control. Each of the three methods of data processing (overall percent drop in absorbance over 1.5h, initial rate of reaction determined over the first 5 min, and estimated pseudo first-order reaction rate constant determined over 1.5h) gave similar and reproducible results for percent control activity. The fastest and easiest method of data analysis was determined to be that using initial rates, involving data acquisition for only 5 min once reactions have been initiated using l-ascorbic acid. The results of the study demonstrate that this simple assay based on the spectroscopic detection of heme represents a rapid, convenient method to determine the relative inhibitory activity of candidate compounds, and is useful in quickly screening a series or library of compounds for heme oxygenase inhibition

  19. Curcuma longa (curcumin) decreases in vivo cisplatin-induced ototoxicity through heme oxygenase-1 induction.

    Fetoni, Anna R; Eramo, Sara L M; Paciello, Fabiola; Rolesi, Rolando; Podda, Maria Vittoria; Troiani, Diana; Paludetti, Gaetano

    2014-06-01

    To investigate whether curcumin may have in vivo protective effects against cisplatin ototoxicity by its direct scavenger activity and/or by curcumin-mediated upregulation of HO-1. Cisplatin-induced ototoxicity is a major dose-limiting side effect in anticancer chemotherapy. A protective approach to decrease cisplatin ototoxicity without compromising its therapeutic efficacy remains a critical goal for anticancer therapy. Recent evidences indicate that curcumin exhibits antioxidant, anti-inflammatory, and chemosensitizer activities. In male adult Wistar rats, a curcumin dose of 200 mg/kg, selected from a dose-response curve, was injected 1 hour before cisplatin administration and once daily for the following 3 days. A single dose of cisplatin (16 mg/kg) was administered intraperitoneally. Rats were divided as follows: 1) control, 2) curcumin control, 3) vehicle control, 4) cisplatin, 5) cisplatin+ vehicle, and 6) curcumin+cisplatin. ABRs were measured before and at Days 3 and 5 after cisplatin administration. Rhodamine-phalloidin staining, 4-hydroxy-2-nonenal and heme-oxigenase-1 immunostainings, and Western blot analyses were performed to assess and quantify OHC loss, lipid peroxidation, and the endogenous response to cisplatin-induced damage and to curcumin protection. Curcumin treatment attenuated hearing loss induced by cisplatin, increased OHC survival, decreased 4-HNE expression, and increased HO-1 expression. This preclinical study demonstrates that systemic curcumin attenuates ototoxicity and provides molecular evidence for a role of HO-1 as an additional mediator in attenuating cisplatin-induced damage.

  20. Ultraviolet-B Protective Effect of Flavonoids from Eugenia caryophylata on Human Dermal Fibroblast Cells.

    Patwardhan, Juilee; Bhatt, Purvi

    2015-10-01

    The exposure of skin to ultraviolet-B (UV-B) radiations leads to deoxyribonucleic acid (DNA) damage and can induce production of free radicals which imbalance the redox status of the cell and lead to increased oxidative stress. Clove has been traditionally used for its analgesic, anti-inflammatory, anti-microbial, anti-viral, and antiseptic effects. To evaluate the UV-B protective activity of flavonoids from Eugenia caryophylata (clove) buds on human dermal fibroblast cells. Protective ability of flavonoid-enriched (FE) fraction of clove was studied against UV-B induced cytotoxicity, anti-oxidant regulation, oxidative DNA damage, intracellular reactive oxygen species (ROS) generation, apoptotic morphological changes, and regulation of heme oxygenase-1 (HO-1) gene through nuclear factor E2-related factor 2 antioxidant response element (Nrf2 ARE) pathway. FE fraction showed a significant antioxidant potential. Pretreatment of cells with FE fraction (10-40 μg/ml) reversed the effects of UV-B induced cytotoxicity, depletion of endogenous enzymatic antioxidants, oxidative DNA damage, intracellular ROS production, apoptotic changes, and overexpression of Nrf2 and HO-1. The present study demonstrated for the first time that the FE fraction from clove could confer UV-B protection probably through the Nrf2-ARE pathway, which included the down-regulation of Nrf2 and HO-1. These findings suggested that the flavonoids from clove could potentially be considered as UV-B protectants and can be explored further for its topical application to the area of the skin requiring protection. Pretreatment of human dermal fibroblast with flavonoid-enriched fraction of Eugenia caryophylata attenuated effects of ultraviolet-B radiationsIt also conferred protection through nuclear factor E2-related factor 2-antioxidant response pathway and increased tolerance of cells against oxidative stressFlavonoid-enriched fraction can be explored further for topical application to the skin as a

  1. Heme Oxygenase-1 Activity as a Correlate to Exercise-Mediated Amelioration of Cognitive Decline and Neuropathological Alterations in an Aging Rat Model of Dementia

    Andrea Kurucz

    2018-01-01

    Full Text Available Alzheimer’s disease (AD is a neurodegenerative disorder with cognitive impairment. Physical exercise has long been proven to be beneficial in the disorder. The present study was designed to examine the effect of voluntary exercise on spatial memory, imaging, and pathological abnormalities. Particular focus has been given to the role of heme oxygenase-1 (HO-1—an important cellular cytoprotectant in preserving mental acuity—using an aging rat model of dementia. Male and female Wistar rats were segregated into six groups—namely, (i aged sedentary (control females (ASF, n=8; (ii aged sedentary (control males (ASM, n=8; (iii aged running females (ARF, n=8; (iv aged running males (ARM, n=8; (v young control females (YCF, n=8; and (vi young control males (YCM, n=8. Rats in the ARF and ARM groups had free access to a standardized inbuilt running wheel during the 3-month evaluation period. Spatial memory was investigated using the Morris Water Test, imaging and pathological alterations were assessed using positron emission tomography (PET imaging and histopathological examinations (H&E, Congo red staining, respectively, and HO-1 enzyme activity assays were also conducted. The outcomes suggest that voluntary physical exercise mitigates impaired spatial memory and neuropathological changes exhibited by the aging sedentary group, via elevated HO-1 activity, contributing to the antioxidant capacity in the aging brain.

  2. Amomum tsao-ko suppresses lipopolysaccharide-induced inflammatory responses in RAW264.7 macrophages via Nrf2-dependent heme oxygenase-1 expression.

    Li, Bin; Choi, Hee-Jin; Lee, Dong-Sung; Oh, Hyuncheol; Kim, Youn-Chul; Moon, Jin-Young; Park, Won-Hwan; Park, Sun-Dong; Kim, Jai-Eun

    2014-01-01

    Amomum tsao-ko Crevost et Lemaire, used as a spice in Asia, is an important source of Chinese cuisine and traditional Chinese medicines. A. tsao-ko is reported to exert a variety of biological and pharmacological activities, including anti-proliferative, anti-oxidative and neuroprotective effects. In this study, NNMBS227, consisting of the ethanol extract of A. tsao-ko, exhibited potent anti-inflammatory activities in RAW264.7 macrophages. We investigated the effect of NNMBS227 in the suppression of pro-inflammatory mediators, including pro-inflammatory enzymes (inducible nitric oxide synthase and cyclooxygenase-2) and cytokines (tumor necrosis factor-α and interleukin-1β) in LPS stimulated macrophages. NNMBS227 also inhibited the phosphorylation and degradation of IκB-α, as well as the nuclear translocation of nuclear factor kappa B (NF-κB) p65 caused by stimulation with LPS. In addition, NNMBS227 induced heme oxygenase (HO)-1 expression through the nuclear translocation of nuclear factor E2-related factor 2 (Nrf2) in macrophages. Using tin protoporphyrin (SnPP), an HO activity inhibitor, we confirmed an association between the anti-inflammatory effects of NNMBS227 and the up-regulation of HO-1. These findings suggest that Nrf2-dependent increases in the expression of HO-1 induced by NNMBS227 conferred anti-inflammatory activities in LPS stimulated RAW264.7 macrophages.

  3. Glutaredoxin 1 (GRX1) inhibits oxidative stress and apoptosis of chondrocytes by regulating CREB/HO-1 in osteoarthritis.

    Sun, Jie; Wei, Xuelei; Lu, Yandong; Cui, Meng; Li, Fangguo; Lu, Jie; Liu, Yunjiao; Zhang, Xi

    2017-10-01

    GRX1 (glutaredoxin1), a sulfhydryl disulfide oxidoreductase, is involved in many cellular processes, including anti-oxidation, anti-apoptosis, and regulation of cell differentiation. However, the role of GRX1 in the oxidative stress and apoptosis of osteoarthritis chondrocytes remains unclear, prompting the current study. Protein and mRNA expressions were measured by Western blot and RT-qPCR. Oxidative stress was detected by the measurement of MDA and SOD contents. Cells apoptosis were detected by Annexin V-FITC/PI and caspase-3 activity assays. We found that the mRNA and protein expressions of GRX1 were significantly down-regulated in osteoarthritis tissues and cells. GRX1 overexpression increased the mRNA and protein expression of CREB and HO-1. Meanwhile, GRX1 overexpression inhibited oxidative stress and apoptosis in osteoarthritis chondrocytes. Furthermore, we found that GRX1 overexpression regulated HO-1 by increasing CREB, and that HO-1 regulated oxidative stress and apoptosis in osteoarthritis chondrocytes. Thus, GRX1 overexpression constrains oxidative stress and apoptosis in osteoarthritis chondrocytes by regulating CREB/HO-1, providing a novel insight into the molecular mechanism and potential treatment of osteoarthritis. Copyright © 2017. Published by Elsevier Ltd.

  4. Regiospecificity determinants of human heme oxygenase: differential NADPH- and ascorbate-dependent heme cleavage by the R183E mutant.

    Wang, Jinling; Lad, Latesh; Poulos, Thomas L; Ortiz de Montellano, Paul R

    2005-01-28

    The ability of the human heme oxygenase-1 (hHO-1) R183E mutant to oxidize heme in reactions supported by either NADPH-cytochrome P450 reductase or ascorbic acid has been compared. The NADPH-dependent reaction, like that of wild-type hHO-1, yields exclusively biliverdin IXalpha. In contrast, the R183E mutant with ascorbic acid as the reductant produces biliverdin IXalpha (79 +/- 4%), IXdelta (19 +/- 3%), and a trace of IXbeta. In the presence of superoxide dismutase and catalase, the yield of biliverdin IXdelta is decreased to 8 +/- 1% with a corresponding increase in biliverdin IXalpha. Spectroscopic analysis of the NADPH-dependent reaction shows that the R183E ferric biliverdin complex accumulates, because reduction of the iron, which is required for sequential iron and biliverdin release, is impaired. Reversal of the charge at position 183 makes reduction of the iron more difficult. The crystal structure of the R183E mutant, determined in the ferric and ferrous-NO bound forms, shows that the heme primarily adopts the same orientation as in wild-type hHO-1. The structure of the Fe(II).NO complex suggests that an altered active site hydrogen bonding network supports catalysis in the R183E mutant. Furthermore, Arg-183 contributes to the regiospecificity of the wild-type enzyme, but its contribution is not critical. The results indicate that the ascorbate-dependent reaction is subject to a lower degree of regiochemical control than the NADPH-dependent reaction. Ascorbate may be able to reduce the R183E ferric and ferrous dioxygen complexes in active site conformations that cannot be reduced by NADPH-cytochrome P450 reductase.

  5. Involvement of Heme Oxygenase-1 Participates in Anti-Inflammatory and Analgesic Effects of Aqueous Extract of Hibiscus taiwanensis.

    Liu, Shu-Ling; Deng, Jeng-Shyan; Chiu, Chuan-Sung; Hou, Wen-Chi; Huang, Shyh-Shyun; Lin, Wang-Ching; Liao, Jung-Chun; Huang, Guan-Jhong

    2012-01-01

    Anti-inflammatory effects of the aqueous extract of Hibiscus taiwanensis (AHT) were used in lipopolysaccharide (LPS-)stimulated mouse macrophage RAW264.7 cells and carrageenan (Carr-)induced mouse paw edema model. When RAW264.7 macrophages were treated with AHT together with LPS, a concentration-dependent inhibition of nitric oxide (NO), tumor necrosis factor (TNF-α), and prostaglandin E2 (PGE(2)) levels productions were detected. Western blotting revealed that AHT blocked protein expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), and elevated heme oxygenase-1 (HO-1), significantly. In the animal test, AHT decreased the paw edema at the 4th and the 5th h after Carr administration, and it increased the activities of catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx) in the paw tissue. We also demonstrated AHT decreased the NO, TNF-α, and PGE2 levels on the serum level at the 5th h after the Carr injection. Western blotting revealed that AHT decreased Carr-induced iNOS, and COX-2, and increased HO-1 expressions at the 5th h in the edema paw. These findings demonstrated that AHT has excellent anti-inflammatory activities in vitro and in vivo and thus it has great potential to be used as a source for natural health products.

  6. Involvement of Heme Oxygenase-1 Participates in Anti-Inflammatory and Analgesic Effects of Aqueous Extract of Hibiscus taiwanensis

    Shu-Ling Liu

    2012-01-01

    Full Text Available Anti-inflammatory effects of the aqueous extract of Hibiscus taiwanensis (AHT were used in lipopolysaccharide (LPS-stimulated mouse macrophage RAW264.7 cells and carrageenan (Carr-induced mouse paw edema model. When RAW264.7 macrophages were treated with AHT together with LPS, a concentration-dependent inhibition of nitric oxide (NO, tumor necrosis factor (TNF-α, and prostaglandin E2 (PGE2 levels productions were detected. Western blotting revealed that AHT blocked protein expression of inducible nitric oxide synthase (iNOS and cyclooxygenase-2 (COX-2, and elevated heme oxygenase-1 (HO-1, significantly. In the animal test, AHT decreased the paw edema at the 4th and the 5th h after Carr administration, and it increased the activities of catalase (CAT, superoxide dismutase (SOD, and glutathione peroxidase (GPx in the paw tissue. We also demonstrated AHT decreased the NO, TNF-α, and PGE2 levels on the serum level at the 5th h after the Carr injection. Western blotting revealed that AHT decreased Carr-induced iNOS, and COX-2, and increased HO-1 expressions at the 5th h in the edema paw. These findings demonstrated that AHT has excellent anti-inflammatory activities in vitro and in vivo and thus it has great potential to be used as a source for natural health products.

  7. Effect of dimethyl fumarate on heme oxygenase-1 expression in experimental allergic encephalomyelitis in rats

    Kaja Kasarełło

    2017-12-01

    Full Text Available Multiple sclerosis (MS is an autoimmunological disease leading to neurodegeneration. The etiology of the disease remains unknown, which strongly impedes the development of effective therapy. Most MS treatments focus on modulating the activity of the immune system. Dimethyl fumarate (DMF exerts a broad spectrum of action, such as modulating immune cell differentiation towards anti-inflammatory subtypes, influencing cytokine production, regulating immune cell migration into the central nervous system, and activating intracellular antioxidant mechanisms. It is well established that activation of the nuclear factor E2 (Nrf2-dependent pathway, leading to expression of the second-phase antioxidant enzymes, is influenced by DMF. In our experiments we used female Lewis rats in an animal model of MS – experimental allergic encephalomyelitis (EAE. The rats were fed with dimethyl fumarate to test the expression of heme oxygenase-1 (HO-1, one of the second-phase antioxidant enzymes, at specific time points of the symptomatic phases of the disease: on the first day of the occurrence of clinical symptoms (10th day post immunization, DPI; at the peak of clinical symptoms (14th DPI; and at the end of the relapse (21st DPI. The results showed that HO-1 expression, at both the mRNA and protein level, is influenced by DMF administration only at the very beginning of the symptomatic phase of EAE, and not at the peak of clinical symptoms, nor at the end of the relapse. This indicates that the regulation of the Nrf2-dependent antioxidant pathway by DMF occurs at a certain time interval (early EAE/MS and strongly underlines the importance of the earliest introduction of the therapy to the patient.

  8. Lycopene inhibits NF-κB activation and adhesion molecule expression through Nrf2-mediated heme oxygenase-1 in endothelial cells.

    Yang, Po-Min; Chen, Huang-Zhi; Huang, Yu-Ting; Hsieh, Chia-Wen; Wung, Being-Sun

    2017-06-01

    The endothelial expression of cell adhesion molecules plays a leading role in atherosclerosis. Lycopene, a carotenoid with 11 conjugated double bonds, has been shown to have anti-inflammatory properties. In the present study, we demonstrate a putative mechanism for the anti-inflammatory effects of lycopene. We demonstrate that lycopene inhibits the adhesion of tumor necrosis factor α (TNFα)-stimulated monocytes to endothelial cells and suppresses the expression of intercellular cell adhesion molecule-1 (ICAM-1) at the transcriptional level. Moreover, lycopene was found to exert its inhibitory effects by blocking the degradation of the inhibitory protein, IκBα, following 6 h of pre-treatment. In TNFα-stimulated endothelial cells, nuclear factor-κB (NF-κB) nuclear translocation and transcriptional activity were abolished by up to 12 h of lycopene pre-treatment. We also found that lycopene increased the intracellular glutathione (GSH) level and glutamate-cysteine ligase expression. Subsequently, lycopene induced nuclear factor-erythroid 2 related factor 2 (Nrf2) activation, leading to the increased expression of downstream of heme oxygenase-1 (HO-1). The use of siRNA targeting HO-1 blocked the inhibitory effects of lycopene on IκB degradation and ICAM-1 expression. The inhibitory effects of lycopene thus appear to be mediated through its induction of Nrf2-mediated HO-1 expression. Therefore, the findings of the present study indicate that lycopene suppresses the activation of TNFα-induced signaling pathways through the upregulation of Nrf2-mediated HO-1 expression.

  9. The Anti-Inflammatory Effects of Dimethyl Fumarate in Astrocytes Involve Glutathione and Haem Oxygenase-1

    Shao Xia Lin

    2011-03-01

    Full Text Available DMF (dimethyl fumarate exerts anti-inflammatory and prometabolic effects in a variety of cell types, and a formulation (BG-12 is being evaluated for monotherapy in multiple sclerosis patients. DMF modifies glutathione (GSH levels that can induce expression of the anti-inflammatory protein HO-1 (haem oxygenase-1. In primary astrocytes and C6 glioma cells, BG-12 dose-dependently suppressed nitrite production induced by either LI [LPS (lipopolysaccharide at 1 μg/ml plus IFNγ (interferon γ at 20 units/ml] or a mixture of proinflammatory cytokines, with greater efficacy in C6 cells. BG-12 reduced NOS2 (nitric oxide synthase 2 mRNA levels and activation of a NOS2 promoter, reduced nuclear levels of NF-κB (nuclear factor κB p65 subunit and attenuated loss of |κBα (inhibitory κBα in both cell types, although with greater effects in astrocytes. In astrocytes, LI decreased mRNA levels for GSHr (GSH reductase and GCL (c-glutamylcysteine synthetase, and slightly suppressed GSHs (GSH synthetase mRNAs. Co-treatment with BG-12 prevented those decreased and increased levels above control values. In contrast, LI reduced GSHp (GSH peroxidase and GCL in C6 cells, and BG-12 had no effect on those levels. BG-12 increased nuclear levels of Nrf2 (nuclear factor-erythroid 2 p45 subunit-related factor 2, an inducer of GSH-related enzymes, in astrocytes but not C6 cells. In astrocytes, GSH was decreased by BG-12 at 2 h and increased at 24 h. Prior depletion of GSH using buthionine-sulfoximine increased the ability of BG-12 to reduce nitrites. In astrocytes, BG-12 increased HO-1 mRNA levels and effects on nitrite levels were blocked by an HO-1 inhibitor. These results demonstrate that BG-12 suppresses inflammatory activation in astrocytes and C6 glioma cells, but with distinct mechanisms, different dependence on GSH and different effects on transcription factor activation.

  10. Polymorphisms in the Haem Oxygenase-1 promoter are not associated with severity of Plasmodium falciparum malaria in Ghanaian children.

    Hansson, Helle H; Maretty, Lasse; Balle, Christina; Goka, Bamenla Q; Luzon, Elisa; Nkrumah, Francis N; Schousboe, Mette L; Rodrigues, Onike P; Bygbjerg, Ib Christian; Kurtzhals, Jørgen A L; Alifrangis, Michael; Hempel, Casper

    2015-04-11

    Haem oxygenase-1 (HO-1) catabolizes haem and has both cytotoxic and cytoprotective effects. Polymorphisms in the promoter of the Haem oxygenase-1 (HMOX1) gene encoding HO-1 have been associated with several diseases including severe malaria. The objective of this study was to determine the allele and genotype frequencies of two single nucleotide polymorphisms; A(-413)T and G(-1135)A, and a (GT)n repeat length polymorphism in the HMOX1 promoter in paediatric malaria patients and controls to determine possible associations with malaria disease severity. Study participants were Ghanaian children (n=296) admitted to the emergency room at the Department of Child Health, Korle-Bu Teaching Hospital, Accra, Ghana during the malaria season from June to August in 1995, 1996 and 1997, classified as having uncomplicated malaria (n=101) or severe malaria (n=195; defined as severe anaemia (n=63) or cerebral malaria (n=132)). Furthermore, 287 individuals without a detectable Plasmodium infection or asymptomatic carriers of the parasite were enrolled as controls. Blood samples from participants were extracted for DNA and allele and genotype frequencies were determined with allele-specific PCR, restriction fragment length analysis and microsatellite analysis. The number of (GT)n repeats in the study participants varied between 21 and 46 with the majority of alleles having lengths of 26 (8.1%), 29/30 (13.2/17.9%) and 39/40 (8.0/13.8%) repeats, and was categorized into short, medium and long repeats. The (-413)T allele was very common (69.8%), while the (-1135)A allele was present in only 17.4% of the Ghanaian population. The G(-1135)A locus was excluded from further analysis after failing the Hardy-Weinberg equilibrium test. No significant differences in allele or genotype distribution of the A(-413)T and (GT)n repeat polymorphisms were found between the controls and the malaria patients, or between the disease groups, for any of the analysed polymorphisms and no associations with

  11. Anti-neuroinflammatory effect of 6,8,1'-tri-O-methylaverantin, a metabolite from a marine-derived fungal strain Aspergillus sp., via upregulation of heme oxygenase-1 in lipopolysaccharide-activated microglia.

    Kim, Kwan-Woo; Kim, Hye Jin; Sohn, Jae Hak; Yim, Joung Han; Kim, Youn-Chul; Oh, Hyuncheol

    2018-02-01

    In the course of searching for anti-neuroinflammatory metabolites from marine-derived fungi, three fungal metabolites, 6,8,1'-tri-O-methylaverantin, 6,8-di-O-methylaverufin, and 5-methoxysterigmatocystin were isolated from a marine-derived fungal strain Aspergillus sp. SF-6796. Among these, 6,8,1'-tri-O-methylaverantin induced the expression of heme oxygenase (HO)-1 protein in BV2 microglial cells. The induction of HO-1 protein was mediated by the activation of nuclear transcription factor erythroid-2 related factor 2 (Nrf2), and was regulated by the p38 mitogen-activated protein kinase and phosphatidylinositol 3-kinase/protein kinase B signaling pathways. Furthermore, 6,8,1'-tri-O-methylaverantin suppressed the overproduction of pro-inflammatory mediators, such as nitric oxide, prostaglandin E 2 , inducible nitric oxide synthase, and cyclooxygenase-2 in lipopolysaccharide (LPS)-stimulated BV2 microglial cells. These anti-neuroinflammatory effects were mediated through the negative regulation of the nuclear factor kappa B pathway, repressing the phosphorylation and degradation of inhibitor kappa B-α, translocation into the nucleus of p65/p50 heterodimer, and DNA-binding activity of p65 subunit. The anti-neuroinflammatory effect of 6,8,1'-tri-O-methylaverantin was partially blocked by a selective HO-1 inhibitor, suggesting that its anti-neuroinflammatory effect is at least partly mediated by HO-1 induction. In this study, 6,8,1'-tri-O-methylaverantin also induced HO-1 protein expression in primary microglial cells, and this correlated with anti-neuroinflammatory effects observed in LPS-stimulated primary microglial cells. In conclusion, 6,8,1'-tri-O-methylaverantin represents a potential candidate for use in the development of therapeutic agents for the regulation of neuroinflammation in neurodegenerative diseases. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. The effects of lead exposure on the expression of HMGB1 and HO-1 in rats and PC12 cells.

    Yang, Meiyuan; Li, Yaobin; Wang, Ying; Cheng, Nuo; Zhang, Yi; Pang, Shimin; Shen, Qiwei; Zhao, Lijuan; Li, Guilin; Zhu, Gaochun

    2018-05-15

    Lead (Pb) is an environmental neurotoxic metal. Chronic exposure to Pb causes deficits of learning and memory in children and spatial learning deficits in developing rats. In this study we investigated the effects of Pb exposure on the expression of HMGB1 and HO-1 in rats and PC12 cells. The animals were randomly divided to three groups: control group; low lead exposure group; high lead exposure group; PC12 cells were divided into 3 groups: 0 μM (control group), 1 μM and 100 μM Pb acetate. The results showed that Pb levels in blood and brain of Pb exposed groups were significantly higher than that of the control group (p < 0.05). The expression of HMGB1 and HO-1 were increased in Pb exposed groups than that of the control group (p < 0.05). Moreover, we found that the up-regulation of HO-1 in Pb exposure environment inhibited the expression of HMGB1. Copyright © 2018 Elsevier B.V. All rights reserved.

  13. Bursopentin (BP5 protects dendritic cells from lipopolysaccharide-induced oxidative stress for immunosuppression.

    Tao Qin

    Full Text Available Dendritic cells (DCs play a vital role in the regulation of immune-mediated inflammatory diseases. Thus, DCs have been regarded as a major target for the development of immunomodulators. However, oxidative stress could disturb inflammatory regulation in DCs. Here, we examined the effect of bursopentine (BP5, a novel pentapeptide isolated from chicken bursa of fabricius, on the protection of DCs against oxidative stress for immunosuppression. BP5 showed potent protective effects against the lipopolysaccharide (LPS-induced oxidative stress in DCs, including nitric oxide, reactive oxygen species and lipid peroxidation. Furthermore, BP5 elevated the level of cellular reductive status through increasing the reduced glutathione (GSH and the GSH/GSSG ratio. Concomitant with these, the activities of several antioxidative redox enzymes, including glutathione peroxidase (GPx, catalase (CAT and superoxide dismutase (SOD, were obviously enhanced. BP5 also suppressed submucosal DC maturation in the LPS-stimulated intestinal epithelial cells (ECs/DCs coculture system. Finally, we found that heme oxygenase 1 (HO-1 was remarkably upregulated by BP5 in the LPS-induced DCs, and played an important role in the suppression of oxidative stress and DC maturation. These results suggested that BP5 could protect the LPS-activated DCs against oxidative stress and have potential applications in DC-related inflammatory responses.

  14. Daily exercise prevents diastolic dysfunction and oxidative stress in a female mouse model of western diet induced obesity by maintaining cardiac heme oxygenase-1 levels.

    Bostick, Brian; Aroor, Annayya R; Habibi, Javad; Durante, William; Ma, Lixin; DeMarco, Vincent G; Garro, Mona; Hayden, Melvin R; Booth, Frank W; Sowers, James R

    2017-01-01

    Obesity is a global epidemic with profound cardiovascular disease (CVD) complications. Obese women are particularly vulnerable to CVD, suffering higher rates of CVD compared to non-obese females. Diastolic dysfunction is the earliest manifestation of CVD in obese women but remains poorly understood with no evidence-based therapies. We have shown early diastolic dysfunction in obesity is associated with oxidative stress and myocardial fibrosis. Recent evidence suggests exercise may increase levels of the antioxidant heme oxygenase-1 (HO-1). Accordingly, we hypothesized that diastolic dysfunction in female mice consuming a western diet (WD) could be prevented by daily volitional exercise with reductions in oxidative stress, myocardial fibrosis and maintenance of myocardial HO-1 levels. Four-week-old female C57BL/6J mice were fed a high-fat/high-fructose WD for 16weeks (N=8) alongside control diet fed mice (N=8). A separate cohort of WD fed females was allowed a running wheel for the entire study (N=7). Cardiac function was assessed at 20weeks by high-resolution cardiac magnetic resonance imaging (MRI). Functional assessment was followed by immunohistochemistry, transmission electron microscopy (TEM) and Western blotting to identify pathologic mechanisms and assess HO-1 protein levels. There was no significant body weight decrease in exercising mice, normalized body weight 14.3g/mm, compared to sedentary mice, normalized body weight 13.6g/mm (p=0.38). Total body fat was also unchanged in exercising, fat mass of 6.6g, compared to sedentary mice, fat mass 7.4g (p=0.55). Exercise prevented diastolic dysfunction with a significant reduction in left ventricular relaxation time to 23.8ms for exercising group compared to 33.0ms in sedentary group (pstress and myocardial fibrosis with improved mitochondrial architecture. HO-1 protein levels were increased in the hearts of exercising mice compared to sedentary WD fed females. This study provides seminal evidence that exercise

  15. Lycopene and Apo-10′-lycopenoic Acid Have Differential Mechanisms of Protection against Hepatic Steatosis in β-Carotene-9′,10′-oxygenase Knockout Male Mice123

    Ip, Blanche C; Liu, Chun; Lichtenstein, Alice H; von Lintig, Johannes; Wang, Xiang-Dong

    2015-01-01

    Background: Nonalcoholic fatty liver disease is positively associated with obesity and cardiovascular disease risk. Apo-10′-lycopenoic acid (APO10LA), a potential oxidation product of apo-10′-lycopenal that is generated endogenously by β-carotene-9′,10′-oxygenase (BCO2) cleavage of lycopene, inhibited hepatic steatosis in BCO2-expressing mice. Objective: The present study evaluated lycopene and APO10LA effects on hepatic steatosis in mice without BCO2 expression. Methods: Male and female BCO2-knockout (BCO2-KO) mice were fed a high saturated fat diet (HSFD) with or without APO10LA (10 mg/kg diet) or lycopene (100 mg/kg diet) for 12 wk. Results: Lycopene or APO10LA supplementation reduced hepatic steatosis incidence (78% and 72%, respectively) and severity in BCO2-KO male mice. Female mice did not develop steatosis, had greater hepatic total cholesterol (3.06 vs. 2.31 mg/g tissue) and cholesteryl ester (1.58 vs. 0.86 mg/g tissue), but had lower plasma triglyceride (TG) (229 vs. 282 mg/dL) and cholesterol (97.1 vs. 119 mg/dL) than male mice. APO10LA-mitigated steatosis in males was associated with reduced hepatic total cholesterol (18%) and activated sirtuin 1 signaling, which resulted in reduced fatty acids (FAs) and TG synthesis markers [stearoyl-coenzyme A (CoA) desaturase protein, 71%; acetyl-CoA carboxylase phosphorylation, 79%; AMP-activated protein kinase phosphorylation, 67%], and elevated cholesterol efflux genes (cytochrome P450 family 7A1, 65%; ATP-binding cassette transporter G5/8, 11%). These APO10LA-mediated effects were not mimicked by lycopene supplementation. Intriguingly, steatosis inhibition by lycopene induced peroxisome proliferator–activated receptor (PPAR)α- and PPARγ-related genes in mesenteric adipose tissue (MAT) that increases mitochondrial uncoupling [cell death–inducing DNA fragmentation factor, α subunit-like effector a, 55%; PR domain-containing 16, 47%; uncoupling protein 3 (Ucp3), 55%], FA β-oxidation (PPARα, 53

  16. Andrographolide inhibits hypoxia-induced HIF-1α-driven endothelin 1 secretion by activating Nrf2/HO-1 and promoting the expression of prolyl hydroxylases 2/3 in human endothelial cells.

    Lin, Hung-Chih; Su, Shih-Li; Lu, Chia-Yang; Lin, Ai-Hsuan; Lin, Wan-Chun; Liu, Chin-San; Yang, Ya-Chen; Wang, Hsiu-Miao; Lii, Chong-Kuei; Chen, Haw-Wen

    2017-03-01

    Andrographolide, the main bioactive component of the medicinal plant Andrographis paniculata, has been shown to possess potent anti-inflammatory activity. Endothelin 1 (ET-1), a potent vasoconstrictor peptide produced by vascular endothelial cells, displays proinflammatory property. Hypoxia-inducible factor 1α (HIF-1α), the regulatory member of the transcription factor heterodimer HIF-1α/β, is one of the most important molecules that responds to hypoxia. Changes in cellular HIF-1α protein level are the result of altered gene transcription and protein stability, with the latter being dependent on prolyl hydroxylases (PHDs). In this study, inhibition of pro-inflammatory ET-1 expression and changes of HIF-1α gene transcription and protein stability under hypoxia by andrographolide in EA.hy926 endothelial-like cells were investigated. Hypoxic conditions were created using the hypoxia-mimetic agent CoCl 2. We found that hypoxia stimulated the production of reactive oxygen species (ROS), the expression of HIF-1α mRNA and protein, and the expression and secretion of ET-1. These effects, however, were attenuated by co-exposure to andrographolide, bilirubin, and RuCO. Silencing Nrf2 and heme oxygenase 1 (HO-1) reversed the inhibitory effects of andrographolide on hypxoia-induced HIF-1α mRNA and protein expression. Moreover, andrographolide increased the expression of prolyl hydroxylases (PHD) 2/3, which hydroxylate HIF-1α and promotes HIF-1α proteasome degradation, with an increase in HIF-1α hydroxylation was noted under hypoxia. Inhibition of p38 MAPK abrogated the hypoxia-induced increases in HIF-1α mRNA and protein expression as well as ET-1 mRNA expression and secretion. Taken together, these results suggest that andrographolide suppresses hypoxia-induced pro-inflammatory ET-1 expression by activating Nrf2/HO-1, inhibiting p38 MAPK signaling, and promoting PHD2/3 expression. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 918-930, 2017. © 2016 Wiley

  17. Psidium guajava extract inhibits thymus and activation-regulated chemokine (TARC/CCL17) production in human keratinocytes by inducing heme oxygenase-1 and blocking NF-κB and STAT1 activation.

    Han, Eun Hee; Hwang, Yong Pil; Choi, Jae Ho; Yang, Ji Hye; Seo, Jong Kwon; Chung, Young Chul; Jeong, Hye Gwang

    2011-09-01

    Psidium guajava (P. guajava) is a food and medicinal plant with antioxidant, anti-inflammatory, and anti-allergic activities that support its traditional uses. The aim of this study was to determine the effects of P. guajava ethyl acetate extract (PGEA) on atopic dermatitis and to investigate the possible mechanisms by which PGEA inhibits cytokine-induced Th2 chemokine expression in HaCaT human keratinocyte cells. We found that PGEA suppressed the IFN-γ/TNF-α-co-induced production of thymus and activation-regulated chemokine (TARC) protein and mRNA in HaCaT cells. Additionally, PGEA inhibited the TNF-α/IFN-γ-co-induced activation of NF-κB and STAT1 and increased the expression of heme oxygenase-1 (HO-1) protein and mRNA. HO-1 inhibitor enhanced the suppressive effects of PGEA on TNF-α/IFN-γ-co-induced TARC production and gene expression. Collectively, these data demonstrate that PGEA inhibits chemokine expression in keratinocytes by inducing HO-1 expression and it suggests a possible therapeutic application in atopic dermatitis and other inflammatory skin diseases. Copyright © 2011 Elsevier B.V. All rights reserved.

  18. Co-operation of the transcription factor hepatocyte nuclear factor-4 with Sp1 or Sp3 leads to transcriptional activation of the human haem oxygenase-1 gene promoter in a hepatoma cell line.

    Takahashi, Shigeru; Matsuura, Naomi; Kurokawa, Takako; Takahashi, Yuji; Miura, Takashi

    2002-11-01

    We reported previously that the 5'-flanking region (nucleotides -1976 to -1655) of the human haem oxygenase-1 ( hHO-1 ) gene enhances hHO-1 promoter activity in human hepatoma HepG2 cells, but not in HeLa cells [Takahashi, Takahashi, Ito, Nagano, Shibahara and Miura (1999) Biochim. Biophys. Acta 1447, 231-235]. To define more precisely the regulatory elements involved, in the present study we have functionally dissected this region and localized the enhancer to a 50 bp fragment (-1793 to -1744). Site-direct mutagenesis analysis revealed that two regions were responsible for this enhancer activity, i.e. a hepatocyte nuclear factor-4 (HNF-4) homologous region and a GC box motif homologous region. Mutation in either region alone moderately decreased enhancer activity. However, mutations in both regions reduced promoter activity to the basal level. Electrophoretic mobility-shift assays demonstrated that the P5-2 fragment (-1793 to -1744) interacted with at least two nuclear factors, i.e. HNF-4 and Sp1/Sp3. Co-transfection experiments using Drosophila SL2 cells revealed that HNF-4 and Sp1/Sp3 synergistically stimulated the enhancer activity of the P5-2 fragment. These results indicate that co-operation of HNF-4 with Sp1 or Sp3 leads to the activation of hHO-1 gene expression in hepatoma cells.

  19. Bioavailability of andrographolide and protection against carbon tetrachloride-induced oxidative damage in rats.

    Chen, Haw-Wen; Huang, Chin-Shiu; Li, Chien-Chun; Lin, Ai-Hsuan; Huang, Yu-Ju; Wang, Tsu-Shing; Yao, Hsien-Tsung; Lii, Chong-Kuei

    2014-10-01

    Andrographolide, a bioactive diterpenoid, is identified in Andrographis paniculata. In this study, we investigated the pharmacokinetics and bioavailability of andrographolide in rats and studied whether andrographolide enhances antioxidant defense in a variety of tissues and protects against carbon tetrachloride-induced oxidative damage. After a single 50-mg/kg administration, the maximum plasma concentration of andrographolide was 1μM which peaked at 30min. The bioavailability of andrographolide was 1.19%. In a hepatoprotection study, rats were intragastrically dosed with 30 or 50mg/kg andrographolide for 5 consecutive days. The results showed that andrographolide up-regulated glutamate cysteine ligase (GCL) catalytic and modifier subunits, superoxide dismutase (SOD)-1, heme oxygenase (HO)-1, and glutathione (GSH) S-transferase (GST) Ya/Yb protein and mRNA expression in the liver, heart, and kidneys. The activity of SOD, GST, and GSH reductase was also increased in rats dosed with andrographolide (pandrographolide increased nuclear Nrf2 contents and Nrf2 binding to DNA, respectively. After the 5-day andrographolide treatment, one group of animals was intraperitoneally injected with carbon tetrachloride (CCl4) at day 6. Andrographolide pretreatment suppressed CCl4-induced plasma aminotransferase activity and hepatic lipid peroxidation (pandrographolide is quickly absorbed in the intestinal tract in rats with a bioavailability of 1.19%. Andrographolide protects against chemical-induced oxidative damage by up-regulating the gene transcription and activity of antioxidant enzymes in various tissues. Copyright © 2014 Elsevier Inc. All rights reserved.

  20. Diabetes Impairs the Vascular Recruitment of Normal Stem Cells by Oxidant Damage, Reversed by Increases in pAMPK, Heme Oxygenase-1, and Adiponectin

    Sambuceti, Gianmario; Morbelli, Silvia; Vanella, Luca; Kusmic, Claudia; Marini, Cecilia; Massollo, Michela; Augeri, Carla; Corselli, Mirko; Ghersi, Chiara; Chiavarina, Barbara; Rodella, Luigi F; L'Abbate, Antonio; Drummond, George; Abraham, Nader G; Frassoni, Francesco

    2009-01-01

    Background Atherosclerosis progression is accelerated in diabetes mellitus (DM) by either direct endothelial damage or reduced availability and function of endothelial progenitor cells (EPCs). Both alterations are related to increased oxidant damage. Aim We examined if DM specifically impairs vascular signaling, thereby reducing the recruitment of normal EPCs, and if increases in antioxidant levels by induction of heme oxygenase-1 (HO-1) can reverse this condition. Methods Control and diabetic rats were treated with the HO-1 inducer cobalt protoporphyrin (CoPP) once a week for 3 weeks. Eight weeks after the development of diabetes, EPCs harvested from the aorta of syngenic inbred normal rats and labeled with technetium-99m-exametazime were infused via the femoral vein to estimate their blood clearance and aortic recruitment. Circulating endothelial cells (CECs) and the aortic expression of thrombomodulin (TM), CD31, and endothelial nitric oxide synthase (eNOS) were used to measure endothelial damage. Results DM reduced blood clearance and aortic recruitment of EPCs. Both parameters were returned to control levels by CoPP treatment without affecting EPC kinetics in normal animals. These abnormalities of EPCs in DM were paralleled by reduced serum adiponectin levels, increased numbers of CECs, reduced endothelial expression of phosphorylated eNOS, and reduced levels of TM, CD31, and phosphorylated AMP-activated protein kinase (pAMPK). CoPP treatment restored all of these parameters to normal levels. Conclusion Type II DM and its related oxidant damage hamper the interaction between the vascular wall and normal EPCs by mechanisms that are, at least partially, reversed by the induction of HO-1 gene expression, adiponectin, and pAMPK levels. PMID:19038792

  1. The expression, function and targeting of haem oxygenase-1 in cancer

    Hjortsø, Mads Duus; Andersen, Mads Hald

    2014-01-01

    , several anticancer strategies aim at targeting HO-1. The inhibition of HO-1 may cause tumour cells to become more sensitive to chemotherapy and radiation therapy. The water-soluble forms of the HO-1 inhibitor Zinc protoporphyrin (ZnPP) have seemed promising in different in-vivo models, in which it has......-inflammatory functions which play a very important role in the negative regulation of the immune system. Immunological targeting of HO-1 might represent an interesting approach, as epitopes derived from HO- 1 have been found exclusively on tumour tissue. Natural HO-1-specific T-cell responses have been identified...... the different results of the targeting of HO-1 in preclinical and clinical settings, and discuss future opportunities....

  2. Repeat polymorphisms in the Homo sapiens heme oxygenase-1 gene in diabetic and idiopathic gastroparesis

    Gibbons, Simon J.; Grover, Madhusudan; Choi, Kyoung Moo; Wadhwa, Akhilesh; Zubair, Adeel; Wilson, Laura A.; Wu, Yanhong; Abell, Thomas L.; Hasler, William L.; Koch, Kenneth L.; McCallum, Richard W.; Nguyen, Linda A. B.; Parkman, Henry P.; Sarosiek, Irene; Snape, William J.; Tonascia, James; Hamilton, Frank A.; Pasricha, Pankaj J.

    2017-01-01

    Background Idiopathic and diabetic gastroparesis in Homo sapiens cause significant morbidity. Etiology or risk factors have not been clearly identified. Failure to sustain elevated heme oxygenase-1 (HO1) expression is associated with delayed gastric emptying in diabetic mice and polymorphisms in the HO1 gene (HMOX1, NCBI Gene ID:3162) are associated with worse outcomes in other diseases. Aim Our hypothesis was that longer polyGT alleles are more common in the HMOX1 genes of individuals with gastroparesis than in controls without upper gastrointestinal motility disorders. Methods Repeat length was determined in genomic DNA. Controls with diabetes (84 type 1, 84 type 2) and without diabetes (n = 170) were compared to diabetic gastroparetics (99 type 1, 72 type 2) and idiopathic gastroparetics (n = 234). Correlations of repeat lengths with clinical symptom sub-scores on the gastroparesis cardinal symptom index (GCSI) were done. Statistical analyses of short (32) repeat alleles and differences in allele length were used to test for associations with gastroparesis. Results The distribution of allele lengths was different between groups (P = 0.016). Allele lengths were longest in type 2 diabetics with gastroparesis (29.18±0.35, mean ± SEM) and longer in gastroparetics compared to non-diabetic controls (28.50±0.14 vs 27.64±0.20 GT repeats/allele, P = 0.0008). Type 2 diabetic controls had longer alleles than non-diabetic controls. In all gastroparetic groups, allele lengths were longer in African Americans compared to other racial groups, differences in the proportion of African Americans in the groups accounted for the differences between gastroparetics and controls. Diabetic gastroparetics with 1 or 2 long alleles had worse GCSI nausea sub-scores (3.30±0.23) as compared to those with 0 long alleles (2.66±0.12), P = 0.022. Conclusions Longer poly-GT repeats in the HMOX1 gene are more common in African Americans with gastroparesis. Nausea symptoms are worse in

  3. Repeat polymorphisms in the Homo sapiens heme oxygenase-1 gene in diabetic and idiopathic gastroparesis.

    Gibbons, Simon J; Grover, Madhusudan; Choi, Kyoung Moo; Wadhwa, Akhilesh; Zubair, Adeel; Wilson, Laura A; Wu, Yanhong; Abell, Thomas L; Hasler, William L; Koch, Kenneth L; McCallum, Richard W; Nguyen, Linda A B; Parkman, Henry P; Sarosiek, Irene; Snape, William J; Tonascia, James; Hamilton, Frank A; Pasricha, Pankaj J; Farrugia, Gianrico

    2017-01-01

    Idiopathic and diabetic gastroparesis in Homo sapiens cause significant morbidity. Etiology or risk factors have not been clearly identified. Failure to sustain elevated heme oxygenase-1 (HO1) expression is associated with delayed gastric emptying in diabetic mice and polymorphisms in the HO1 gene (HMOX1, NCBI Gene ID:3162) are associated with worse outcomes in other diseases. Our hypothesis was that longer polyGT alleles are more common in the HMOX1 genes of individuals with gastroparesis than in controls without upper gastrointestinal motility disorders. Repeat length was determined in genomic DNA. Controls with diabetes (84 type 1, 84 type 2) and without diabetes (n = 170) were compared to diabetic gastroparetics (99 type 1, 72 type 2) and idiopathic gastroparetics (n = 234). Correlations of repeat lengths with clinical symptom sub-scores on the gastroparesis cardinal symptom index (GCSI) were done. Statistical analyses of short (32) repeat alleles and differences in allele length were used to test for associations with gastroparesis. The distribution of allele lengths was different between groups (P = 0.016). Allele lengths were longest in type 2 diabetics with gastroparesis (29.18±0.35, mean ± SEM) and longer in gastroparetics compared to non-diabetic controls (28.50±0.14 vs 27.64±0.20 GT repeats/allele, P = 0.0008). Type 2 diabetic controls had longer alleles than non-diabetic controls. In all gastroparetic groups, allele lengths were longer in African Americans compared to other racial groups, differences in the proportion of African Americans in the groups accounted for the differences between gastroparetics and controls. Diabetic gastroparetics with 1 or 2 long alleles had worse GCSI nausea sub-scores (3.30±0.23) as compared to those with 0 long alleles (2.66±0.12), P = 0.022. Longer poly-GT repeats in the HMOX1 gene are more common in African Americans with gastroparesis. Nausea symptoms are worse in subjects with longer alleles.

  4. Repeat polymorphisms in the Homo sapiens heme oxygenase-1 gene in diabetic and idiopathic gastroparesis.

    Simon J Gibbons

    Full Text Available Idiopathic and diabetic gastroparesis in Homo sapiens cause significant morbidity. Etiology or risk factors have not been clearly identified. Failure to sustain elevated heme oxygenase-1 (HO1 expression is associated with delayed gastric emptying in diabetic mice and polymorphisms in the HO1 gene (HMOX1, NCBI Gene ID:3162 are associated with worse outcomes in other diseases.Our hypothesis was that longer polyGT alleles are more common in the HMOX1 genes of individuals with gastroparesis than in controls without upper gastrointestinal motility disorders.Repeat length was determined in genomic DNA. Controls with diabetes (84 type 1, 84 type 2 and without diabetes (n = 170 were compared to diabetic gastroparetics (99 type 1, 72 type 2 and idiopathic gastroparetics (n = 234. Correlations of repeat lengths with clinical symptom sub-scores on the gastroparesis cardinal symptom index (GCSI were done. Statistical analyses of short (32 repeat alleles and differences in allele length were used to test for associations with gastroparesis.The distribution of allele lengths was different between groups (P = 0.016. Allele lengths were longest in type 2 diabetics with gastroparesis (29.18±0.35, mean ± SEM and longer in gastroparetics compared to non-diabetic controls (28.50±0.14 vs 27.64±0.20 GT repeats/allele, P = 0.0008. Type 2 diabetic controls had longer alleles than non-diabetic controls. In all gastroparetic groups, allele lengths were longer in African Americans compared to other racial groups, differences in the proportion of African Americans in the groups accounted for the differences between gastroparetics and controls. Diabetic gastroparetics with 1 or 2 long alleles had worse GCSI nausea sub-scores (3.30±0.23 as compared to those with 0 long alleles (2.66±0.12, P = 0.022.Longer poly-GT repeats in the HMOX1 gene are more common in African Americans with gastroparesis. Nausea symptoms are worse in subjects with longer alleles.

  5. Protective effect of metalloporphyrins against cisplatin-induced kidney injury in mice.

    Hao Pan

    Full Text Available Oxidative and nitrative stress is a well-known phenomenon in cisplatin-induced nephrotoxicity. The purpose of this work is to study the role of two metalloporphyrins (FeTMPyP and MnTBAP, water soluble complexes, in cisplatin-induced renal damage and their ability to scavenge peroxynitrite. In cisplatin-induced nephropathy study in mice, renal nitrative stress was evident by the increase in protein nitration. Cisplatin-induced nephrotoxicity was also evident by the histological damage from the loss of the proximal tubular brush border, blebbing of apical membranes, tubular epithelial cell detachment from the basement membrane, or intra-luminal aggregation of cells and proteins and by the increase in blood urea nitrogen and serum creatinine. Cisplatin-induced apoptosis and cell death as shown by Caspase 3 assessments, TUNEL staining and DNA fragmentation Cisplatin-induced nitrative stress, apoptosis and nephrotoxicity were attenuated by both metalloporphyrins. Heme oxygenase (HO-1 also plays a critical role in metalloporphyrin-mediated protection of cisplatin-induced nephrotoxicity. It is evident that nitrative stress plays a critical role in cisplatin-induced nephrotoxicity in mice. Our data suggest that peroxynitrite is involved, at least in part, in cisplatin-induced nephrotoxicity and protein nitration and cisplatin-induced nephrotoxicity can be prevented with the use of metalloporphyrins.

  6. Protective effects of hesperidin against oxidative stress of tert-butyl hydroperoxide in human hepatocytes.

    Chen, Mingcang; Gu, Honggang; Ye, Yiyi; Lin, Bing; Sun, Lijuan; Deng, Weiping; Zhang, Jingzhe; Liu, Jianwen

    2010-10-01

    Increasing evidence regarding free radical generating agents and the inflammatory process suggest that accumulation of reactive oxygen species (ROS) could involve hepatotoxicity. Hesperidin, a naturally occurring flavonoid presents in fruits and vegetables, has been reported to exert a wide range of pharmacological effects that include antioxidant, anti-inflammatory, antihypercholesterolemic, and anticarcinogenic actions. However, the cytoprotection and mechanism of hesperidin to neutralize oxidative stress in human hepatic L02 cells remain unclear. In this work, we assessed the capability of hesperidin to prevent tert-butyl hydroperoxide (t-BuOOH)-induced cell damage by augmenting cellular antioxidant defense. Hesperidin significantly protected hepatocytes against t-BuOOH-induced cell cytotoxicity, such as mitochondrial membrane potential (MMP) deplete and lactate dehydrogenase (LDH) release. Hesperidin also remarkably prevented indicators of oxidative stress, such as the ROS and lipid peroxidation level in a dose-dependent manner. Western blot showed that hesperidin facilitated ERK/MAPK phosphorylation which appeared to be responsible for nuclear translocation of Nrf2, thereby inducing cytoprotective heme oxygenase-1 (HO-1) expression. Based on the results described above, it suggested that hesperidin has potential as a therapeutic agent in the treatment of oxidative stress-related hepatocytes injury and liver dysfunctions. Copyright © 2010 Elsevier Ltd. All rights reserved.

  7. Supplemental dietary phytosterin protects against 4-nitrophenol-induced oxidative stress and apoptosis in rat testes

    Yonghui Zhang

    2015-01-01

    Full Text Available 4-Nitrophenol (PNP, is generally regarded as an environmental endocrine disruptor (EED. Phytosterin (PS, a new feed additive, possesses highly efficient antioxidant activities. The transcription factor, nuclear factor-erythroid 2-related factor 2 (Nrf2, is an important regulator of cellular oxidative stress. Using rats, this study examined PNP-induced testicular oxidative damage and PS-mediated protection against that damage. The generation of MDA and H2O2 upon PNP and PS treatment was milder than that upon treatment with PNP alone. This mitigation was accompanied by partially reversed changes in SOD, CAT, GSH and GSH-Px. Moreover, PNP significantly reduced the caudal epididymal sperm counts and serum testosterone levels. Typical morphological changes were also observed in the testes of PNP-treated animals. PNP reduced the transcriptional level of Nrf2, as evaluated by RT-PCR, but it promoted the dissociation from the Nrf2 complex, stabilization and translocation into the nucleus, as evaluated by immunohistochemistry and Western blotting. In addition, PNP enhanced the Nrf2-dependent gene expression of heme oxygenase-1 (HO-1 and glutamate–cysteine ligase catalytic subunit (GCLC. These results suggest that the Nrf2 pathway plays an important role in PNP-induced oxidative damage and that PS possesses modulatory effects on PNP-induced oxidative damage in rat testes.

  8. Protective Effect of Tempol on Acute Kidney Injury Through PI3K/Akt/Nrf2 Signaling Pathway

    Gensheng Zhang

    2016-02-01

    Full Text Available Background/Aims: Tempol is a protective antioxidant against ischemic injury in many animal models. The molecular mechanisms are not well understood. Nuclear factor erythroid 2-related factor (Nrf2 is a master transcription factor during oxidative stress, which is enhanced by activation of protein kinase C (PKC pathway. Another factor, tubular epithelial apoptosis, is mediated by activation of phosphoinositide 3-kinase (PI3K/protein kinase B (PKB, Akt signaling pathway during renal ischemic injury. We tested the hypothesis that tempol activates PKC or PI3K/Akt/Nrf2 pathways to transcribe many genes that coordinate endogenous antioxidant defense. Methods: The right renal pedicle was clamped for 45 minutes and the left kidney was removed to study renal ischemia/reperfusion (I/R injury in C57BL/6 mice. The response was assessed from serum parameters, renal morphology and renal expression of PKC, phosphorylated-PKC (p-PKC, Nrf2, heme oxygenase-1 (HO-1, Akt, phosphorylated-Akt (p-Akt, pro-caspase-3 and cleaved caspase-3 in groups of sham and I/R mice given vehicle, or tempol (50 or 100 mg/kg, intraperitoneal injection. Results: The serum malondialdehyde (MDA, marker of reactive oxygen species doubled and the BUN and creatinine increased 5- to 10-fold after I/R injury. Tempol (50 or 100 mg/kg prevented the increases in MDA but only tempol (50 mg/kg lessened the increases in BUN and creatinine and moderated the acute tubular necrosis. I/R did not change expression of PKC or p-PKC but reduced renal expression of Nrf2, p-Akt, HO-1 and pro-caspase-3 and increased cleaved caspase-3. Tempol (50 mg/kg prevented these changes produced by I/R whereas tempol (100 mg/kg had lesser or inconsistent effects. Conclusion: Tempol (50 mg/kg prevents lipid peroxidation and attenuates renal damage after I/R injury. The beneficial pathway apparently is not dependent on upregulation or phosphorylation of PKC, at lower tempol doses, does implicate upregulation of Akt with

  9. Anti-oxidative and anti-inflammatory effects of cinnamaldehyde on protecting high glucose-induced damage in cultured dorsal root ganglion neurons of rats.

    Yang, Dan; Liang, Xiao-Chun; Shi, Yue; Sun, Qing; Liu, Di; Liu, Wei; Zhang, Hong

    2016-01-01

    To examine the mechanism underlying the beneficial role of cinnamaldehyde on oxidative damage and apoptosis in high glucose (HG)-induced dorsal root ganglion (DRG) neurons in vitro. HG-treated DRG neurons were developed as an in vitro model of diabetic neuropathy. The neurons were randomly divided into five groups: the control group, the HG group and the HG groups treated with 25, 50 and 100 nmol/L cinnamaldehyde, respectively. Cell viability was examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and apoptosis rate was evaluated by the in situ TdT-mediated dUTP nick end labeling (TUNEL) assay. The intracellular level of reactive oxygen species (ROS) was measured with flow cytometry. Expression of nuclear factor-kappa B (NF-κB), inhibitor of κB (IκB), phosphorylated IκB (p-IκB), tumor necrosis factor (TNF)-α, interleukin-6 (IL-6) and caspase-3 were determined by western blotting and real-time quantitative reverse transcription polymerase chain reaction (RT-PCR). Expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and hemeoxygenase-1 (HO-1) were also measured by western blotting. Cinnamaldehyde reduced HG-induced loss of viability, apoptosis and intracellular generation of ROS in the DRG neurons via inhibiting NF-κB activity. The western blot assay results showed that the HG-induced elevated expressions of NF-κB, IκB and p-IκB were remarkably reduced by cinnamaldehyde treatment in a dose-dependent manner (P neurons, but also lowered the elevated IL-6, TNF-α, cyclo-oxygenase and inducible nitric oxide synthase levels, indicating a reduction in inflammatory damage. Cinnamaldehyde protected DRG neurons from the deleterious effects of HG through inactivation of NF-κB pathway but not through activation of Nrf2/HO-1. And thus cinnamaldehyde may have potential application as a treatment for DPN.

  10. Heme oxygenase is not involved in the anti-proliferative effects of statins on pancreatic cancer cells

    Vanova, K.; Boukalova, S.; Gbelcova, H.; Muchova, L.; Neuzil, J.; Gurlich, R.; Ruml, T.; Vitek, L.

    2016-01-01

    Pancreatic cancer is recognized as one of the most fatal tumors due to its aggressiveness and resistance to therapy. Statins were previously shown to inhibit the proliferation of cancer cells via various signaling pathways. In healthy tissues, statins activate the heme oxygenase pathway, nevertheless the role of heme oxygenase in pancreatic cancer is still controversial. The aim of this study was to evaluate, whether anti-proliferative effects of statins in pancreatic cancer cells are mediated via the heme oxygenase pathway. In vitro effects of various statins and hemin, a heme oxygenase inducer, on cell proliferation were evaluated in PA-TU-8902, MiaPaCa-2 and BxPC-3 human pancreatic cancer cell lines. The effect of statins on heme oxygenase activity was assessed and heme oxygenase-silenced cells were used for pancreatic cancer cell proliferation studies. Cell death rate and reactive oxygen species production were measured in PA-TU-8902 cells, followed by evaluation of the effect of cerivastatin on GFP-K-Ras trafficking and expression of markers of invasiveness, osteopontin (SPP1) and SOX2. While simvastatin and cerivastatin displayed major anti-proliferative properties in all cell lines tested, pravastatin did not affect the cell growth at all. Strong anti-proliferative effect was observed also for hemin. Co-treatment of cerivastatin and hemin increased anti-proliferative potential of these agents, via increased production of reactive oxygen species and cell death compared to individual treatment. Heme oxygenase silencing did not prevent pancreatic cancer cells from the tumor-suppressive effect of cerivastatin or hemin. Cerivastatin, but not pravastatin, protected Ras protein from trafficking to the cell membrane and significantly reduced expressions of SPP1 (p < 0.05) and SOX2 (p < 0.01). Anti-proliferative effects of statins and hemin on human pancreatic cancer cell lines do not seem to be related to the heme oxygenase pathway. While hemin triggers reactive

  11. Osmopriming-induced salt tolerance during seed germination of alfalfa most likely mediates through H2O2 signaling and upregulation of heme oxygenase.

    Amooaghaie, Rayhaneh; Tabatabaie, Fatemeh

    2017-07-01

    The present study showed that osmopriming or pretreatment with low H 2 O 2 doses (2 mM) for 6 h alleviated salt-reduced seed germination. The NADPH oxidase activity was the main source, and superoxide dismutase (SOD) activity might be a secondary source of H 2 O 2 generation during osmopriming or H 2 O 2 pretreatment. Hematin pretreatment similar to osmopriming improved salt-reduced seed germination that was coincident with the enhancement of heme oxygenase (HO) activity. The semi-quantitative RT-PCR confirmed that osmopriming or H 2 O 2 pretreatment was able to upregulate heme oxygenase HO-1 transcription, while the application of N,N-dimethyl thiourea (DMTU as trap of endogenous H 2 O 2 ) and diphenyleneiodonium (DPI as inhibitor of NADPHox) not only blocked the upregulation of HO but also reversed the osmopriming-induced salt attenuation. The addition of CO-saturated aqueous rescued the inhibitory effect of DMTU and DPI on seed germination and α-amylase activity during osmopriming or H 2 O 2 pretreatment, but H 2 O 2 could not reverse the inhibitory effect of ZnPPIX (as HO inhibitor) or Hb (as CO scavenger) that indicates that the CO acts downstream of H 2 O 2 in priming-driven salt acclimation. The antioxidant enzymes and proline synthesis were upregulated in roots of seedlings grown from primed seeds, and these responses were reversed by adding DMTU, ZnPPIX, and Hb during osmopriming. These findings for the first time suggest that H 2 O 2 signaling and upregulation of heme oxygenase play a crucial role in priming-driven salt tolerance.

  12. DNA Protecting Activities of Nymphaea nouchali (Burm. f Flower Extract Attenuate t-BHP-Induced Oxidative Stress Cell Death through Nrf2-Mediated Induction of Heme Oxygenase-1 Expression by Activating MAP-Kinases

    Md Badrul Alam

    2017-09-01

    Full Text Available This study was performed to investigate the antioxidant activities of Nymphaea nouchali flower (NNF extract and the underlying mechanism using RAW 264.7 cells. The presence of gallic acid, catechin, epicatechin, epigallocatechin, epicatechin gallate, caffeic acid, quercetin, and apigenin in the NNF was confirmed by high-performance liquid chromatography (HPLC. The extract had a very potent capacity to scavenge numerous free radicals. NNF extract was also able to prevent DNA damage and quench cellular reactive oxygen species (ROS generation induced by tert-Butyl hydroperoxide (t-BHP with no signs of toxicity. The NNF extract was able to augment the expression of both primary and phase II detoxifying enzyme, resulting in combat the oxidative stress. This is accomplished by phosphorylation of mitogen-activated protein kinase (MAP kinase (p38 kinase and extracellular signal-regulated kinase (ERK followed by enhancing the nuclear translocation of the nuclear factor erythroid 2-related factor 2 (Nrf2. This attenuates cellular ROS generation and confers protection from cell death. Altogether, the results of current study revealed that Nymphaea nouchali flower could be a source of natural phytochemicals that could lead to the development of new therapeutic agents for preventing oxidative stress associated diseases and attenuating disease progression.

  13. Role of Nrf2/ARE pathway in protective effect of electroacupuncture against endotoxic shock-induced acute lung injury in rabbits.

    Jian-bo Yu

    Full Text Available NF-E2 related factor 2 (Nrf2 is a major transcription factor and acts as a key regulator of antioxidant genes to exogenous stimulations. The aim of current study was to determine whether Nrf2/ARE pathway is involved in the protective effect of electroacupuncture on the injured lung in a rabbit model of endotoxic shock. A dose of lipopolysaccharide (LPS 5 mg/kg was administered intravenously to replicate the model of acute lung injury induced by endotoxic shock. Electroacupuncture pretreatment was handled bilaterally at Zusanli and Feishu acupoints for five consecutive days while sham electroacupuncture punctured at non-acupoints. Fourty anesthetized New England male rabbits were randomized into normal control group (group C, LPS group (group L, electroacupuncture + LPS group (group EL and sham electroacupuncture + LPS (group SEL. At 6 h after LPS administration, the animals were sacrificed and the blood samples were collected for biochemical measurements. The lungs were removed for calculation of wet-to-dry weight ratios (W/D, histopathologic examination, determination of heme oxygenase (HO-1 protein and mRNA, Nrf2 total and nucleoprotein, as well as Nrf2 mRNA expression, and evaluation of the intracellular distribution of Nrf2 nucleoprotein. LPS caused extensive morphologic lung damage, which was lessened by electroacupuncture treatment. Besides, lung W/D ratios were significantly decreased, the level of malondialdehyde was inhibited, plasma levels of TNF-α and interleukin-6 were decreased, while the activities of superoxide dismutase, glutathione peroxidase and catalase were enhanced in the electroacupucnture treated animals. In addition, electroacupuncture stimulation distinctly increased the expressions of HO-1 and Nrf2 protein including Nrf2 total protein and nucleoprotein as well as mRNA in lung tissue, while these effects were blunted in the sham electroacupuncture group. We concluded that electroacupuncture treatment at ST36 and BL13

  14. Role of Nrf2/ARE pathway in protective effect of electroacupuncture against endotoxic shock-induced acute lung injury in rabbits.

    Yu, Jian-bo; Shi, Jia; Gong, Li-rong; Dong, Shu-an; Xu, Yan; Zhang, Yuan; Cao, Xin-shun; Wu, Li-li

    2014-01-01

    NF-E2 related factor 2 (Nrf2) is a major transcription factor and acts as a key regulator of antioxidant genes to exogenous stimulations. The aim of current study was to determine whether Nrf2/ARE pathway is involved in the protective effect of electroacupuncture on the injured lung in a rabbit model of endotoxic shock. A dose of lipopolysaccharide (LPS) 5 mg/kg was administered intravenously to replicate the model of acute lung injury induced by endotoxic shock. Electroacupuncture pretreatment was handled bilaterally at Zusanli and Feishu acupoints for five consecutive days while sham electroacupuncture punctured at non-acupoints. Fourty anesthetized New England male rabbits were randomized into normal control group (group C), LPS group (group L), electroacupuncture + LPS group (group EL) and sham electroacupuncture + LPS (group SEL). At 6 h after LPS administration, the animals were sacrificed and the blood samples were collected for biochemical measurements. The lungs were removed for calculation of wet-to-dry weight ratios (W/D), histopathologic examination, determination of heme oxygenase (HO)-1 protein and mRNA, Nrf2 total and nucleoprotein, as well as Nrf2 mRNA expression, and evaluation of the intracellular distribution of Nrf2 nucleoprotein. LPS caused extensive morphologic lung damage, which was lessened by electroacupuncture treatment. Besides, lung W/D ratios were significantly decreased, the level of malondialdehyde was inhibited, plasma levels of TNF-α and interleukin-6 were decreased, while the activities of superoxide dismutase, glutathione peroxidase and catalase were enhanced in the electroacupucnture treated animals. In addition, electroacupuncture stimulation distinctly increased the expressions of HO-1 and Nrf2 protein including Nrf2 total protein and nucleoprotein as well as mRNA in lung tissue, while these effects were blunted in the sham electroacupuncture group. We concluded that electroacupuncture treatment at ST36 and BL13 effectively

  15. Immunolocalization of heme oxygenase-1 in periodontal diseases

    G Gayathri

    2014-01-01

    Conclusion: The results of our study is an increasing evidence of involvement of antioxidant enzymes like heme oxygenase-1 in periodontal inflammation and their implication for treatment of chronic periodontitis.

  16. Agmatine Reduces Lipopolysaccharide-Mediated Oxidant Response via Activating PI3K/Akt Pathway and Up-Regulating Nrf2 and HO-1 Expression in Macrophages.

    Jianshen Chai

    Full Text Available Macrophages are key responders of inflammation and are closely related with oxidative stress. Activated macrophages can enhance oxygen depletion, which causes an overproduction of reactive oxygen species (ROS and leads to further excessive inflammatory response and tissue damage. Agmatine, an endogenous metabolite of L-arginine, has recently been shown to have neuroprotective effects based on its antioxidant properties. However, the antioxidant effects of agmatine in peripheral tissues and cells, especially macrophages, remain unclear. In this study we explored the role of agmatine in mediating antioxidant effects in RAW 264.7 cells and studied its antioxidant mechanism. Our data demonstrate that agmatine is an activator of Nrf2 signaling that markedly enhances Nrf2 nuclear translocation, increases nuclear Nrf2 protein level, up-regulates the expression of the Nrf2 downstream effector HO-1, and attenuates ROS generation induced by Lipopolysaccharide (LPS. We further demonstrated that the agmatine-induced activation of Nrf2 is likely through the PI3K/Akt pathway. LY294002, a specific PI3K/Akt inhibitor, abolished agmatine-induced HO-1 up-regulation and ROS suppression significantly. Inhibiting HO-1 pathway significantly attenuated the antioxidant effect of agmatine which the products of HO-1 enzymatic activity contributed to. Furthermore, the common membrane receptors of agmatine were evaluated, revealing that α2-adrenoceptor, I1-imidazoline receptor or I2-imidazoline receptor are not required by the antioxidant properties of agmatine. Taken together, our findings revealed that agmatine has antioxidant activity against LPS-induced ROS accumulation in RAW 264.7 cells involving HO-1 expression induced by Nrf2 via PI3K/Akt pathway activation.

  17. Tanshinol ameliorates CCl4-induced liver fibrosis in rats through the regulation of Nrf2/HO-1 and NF-κB/IκBα signaling pathway

    Wang R

    2018-05-01

    Full Text Available Rong Wang,* Jing Wang,* Fuxing Song, Shengnan Li, Yongfang Yuan Department of Pharmacy, Shanghai 9th People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China *These authors contributed equally to this work Abstract: Tanshinol, a water-soluble component isolated from Salvia miltiorrhiza Bunge, has a variety of biological activities involving anti-fibrotic effect. However, the exact role and the underlying mechanisms remain largely unclear. This study mainly focused on the anti-hepatic fibrotic activities and mechanisms of tanshinol on carbon tetrachloride (CCl4-induced liver fibrosis in rats via anti-oxidative and anti-inflammation pathways. The rats were divided into 4 groups as follows: control, model, tanshinol 20 mg/kg, and tanshinol 40 mg/kg. Except for the control group, CCl4 was used to induce liver fibrosis processing for 8 weeks, meanwhile rats in tanshinol groups were intraperitoneally injected with additional tanshinol. Control group simultaneously received the same volumes of olive oil and saline. The potentially protective effect and mechanisms of tanshinol on liver fibrosis in rats were evaluated. The serum levels of alanine aminotransferase, aspartate aminotransferase, and total bilirubin were obviously lower in the tanshinol treatment groups related to model group. Compared with the model group, the levels of hyaluronic acid, type IV collagen, Laminin (LN, and procollagen III peptide (PIIIP in serum were significantly decreased after tanshinol treatment. Furthermore, tanshinol could regulate Nrf2/HO-1 signaling pathway and increase the level of superoxide dismutase (SOD and glutathione peroxidase (GSH-Px, and also decrease the level of malondialdehyde (MDA to against damage induced by oxidative stress. Simultaneously tanshinol could regulate nuclear factor kappa B signaling pathway to inhibit expression of inflammation factors, including transforming growth factor-β, tumor necrosis factor-α, Cox-2

  18. Polydatin Protects Rat Liver against Ethanol-Induced Injury: Involvement of CYP2E1/ROS/Nrf2 and TLR4/NF-κB p65 Pathway

    Qiong-Hui Huang

    2017-01-01

    Full Text Available Excessive alcohol consumption leads to serious liver injury, associating with oxidative stress and inflammatory response. Previous study has demonstrated that polydatin (PD exerted antioxidant and anti-inflammatory effects and attenuated ethanol-induced liver damage, but the research remained insufficient. Hence, this experiment aimed to evaluate the hepatoprotective effect and potential mechanisms of PD on ethanol-induced hepatotoxicity. Our results showed that PD pretreatment dramatically decreased the levels of alanine aminotransferase (ALT, aspartate aminotransferase (AST, alkaline phosphatase (ALP, and lactate dehydrogenase (LDH in the serum, suppressed the malonaldehyde (MDA and triglyceride (TG content and the production of reactive oxygen species (ROS, and enhanced the activities of superoxide dismutase (SOD, glutathione peroxidase (GSH-Px, catalase (CAT, andalcohol dehydrogenase (ADH, and aldehyde dehydrogenase (ALDH, paralleled by an improvement of histopathology alterations. The protective effect of PD against oxidative stress was probably associated with downregulation of cytochrome P450 2E1 (CYP2E1 and upregulation of nuclear factor erythroid 2-related factor 2 (Nrf2 and its target gene haem oxygenase-1 (HO-1. Moreover, PD inhibited the release of proinflammatory cytokines (TNF-α, IL-1β, and IL-6 via downregulating toll-like receptor 4 (TLR4 and nuclear factor kappa B (NF-κB p65. To conclude, PD pretreatment protects against ethanol-induced liver injury via suppressing oxidative stress and inflammation.

  19. Heme oxygenase behavior in ultraviolet-B irradiated soybean plants

    Yannarelli, G.G.; Noriega, G.O.; Tomaro, M.L.

    2005-01-01

    Full text: Ultraviolet-B (UV-B) radiation has a negative impact on plant cells, and leads to the generation of reactive oxygen species (ROS). Heme oxygenase (HO) plays a protective role against oxidative stress in mammals, but little is known about this issue in plants. Here, we report for the first time the response of HO in leaves of soybean plants subjected to UV-B radiation. HO activity, protein and gene expression, as well as stress markers were evaluated. Under lower UV-B doses (7.5 and 15 kJ m -2 ), the production of thiobarbituric acid reactive substances (TBARS) remained unaltered, while quantitative RT-PCR revealed that HO and catalase (CAT) transcripts were increased 40% and 20% after 8 h, respectively. Treatment with 30 kJ m -2 brought about a 90% enhancement in TBARS indicating that an oxidative burst occurred, and a downregulation in gene expression was observed. Immunoblot analysis showed a 4.3 and 3.7-fold increase in HO protein after irradiation with 75 and 15 kJ m -2 , respectively. HO and CAT enzymes activities were enhanced at these doses but diminished at 30 kJ m -2 UV-B. These results indicate that the up regulation of HO and CAT genes at the lower doses occurred as a signal of cell protection against oxidative damage. On the other hand, irradiation with 30 kJ m -2 overcome the cellular antioxidant capacity and repressed the response as a result of ROS overproduction. (author)

  20. OxDBase: a database of oxygenases involved in biodegradation

    Raghava Gajendra PS

    2009-04-01

    Full Text Available Abstract Background Oxygenases belong to the oxidoreductive group of enzymes (E.C. Class 1, which oxidize the substrates by transferring oxygen from molecular oxygen (O2 and utilize FAD/NADH/NADPH as the co-substrate. Oxygenases can further be grouped into two categories i.e. monooxygenases and dioxygenases on the basis of number of oxygen atoms used for oxidation. They play a key role in the metabolism of organic compounds by increasing their reactivity or water solubility or bringing about cleavage of the aromatic ring. Findings We compiled a database of biodegradative oxygenases (OxDBase which provides a compilation of the oxygenase data as sourced from primary literature in the form of web accessible database. There are two separate search engines for searching into the database i.e. mono and dioxygenases database respectively. Each enzyme entry contains its common name and synonym, reaction in which enzyme is involved, family and subfamily, structure and gene link and literature citation. The entries are also linked to several external database including BRENDA, KEGG, ENZYME and UM-BBD providing wide background information. At present the database contains information of over 235 oxygenases including both dioxygenases and monooxygenases. This database is freely available online at http://www.imtech.res.in/raghava/oxdbase/. Conclusion OxDBase is the first database that is dedicated only to oxygenases and provides comprehensive information about them. Due to the importance of the oxygenases in chemical synthesis of drug intermediates and oxidation of xenobiotic compounds, OxDBase database would be very useful tool in the field of synthetic chemistry as well as bioremediation.

  1. Co-treatment with conjugated linoleic acid and nitrite protects against myocardial infarction

    Natia Qipshidze-Kelm

    2014-01-01

    Full Text Available According to the CDC, the most common type of heart disease is coronary artery disease, which commonly leads to myocardial infarction (MI. Therapeutic approaches to lessen the resulting cardiovascular injury associated with MI are limited. Recently, MicroRNAs (miRNAs have been shown to act as negative regulators of gene expression by inhibiting mRNA translation and/or stimulating mRNA degradation. A single miRNA can modulate physiological or disease phenotypes by regulating whole functional systems. Importantly, miRNAs can regulate cardiac function, thereby modulating heart muscle contraction, heart growth and morphogenesis. MicroRNA-499 (miRNA-499 is a cardiac-specific miRNA that when elevated causes cardiomyocyte hypertrophy, in turn preventing cardiac dysfunction during MI. Previous studies revealed that combination treatment with conjugated linoleic acid (cLA and nitrite preserved cardiovascular function in mice. Therefore, it was hypothesized that cLA and nitrite may regulate miRNA-499, thus providing cardiac protection during MI. To test this hypothesis, 12-week old mice were treated with cLA (10 mg/kg/d-via osmotic mini-pump or cLA and nitrite (50 ppm-drinking water 3 days prior to MI (ligation of the left anterior descending artery. Echocardiography and pressure–volume (PV-loop analysis revealed that cLA and nitrite-treated MI mice had improved heart function (10 days following MI compared to untreated MI mice. Treatment with cLA and nitrite significantly induced levels of miRNA-499 compared to untreated MI mice. In addition, treatment with cLA and nitrite abolished MI-induced protein expression of p53 and dynamin-related protein-1 (DRP-1. Moreover, the antioxidant enzyme expression of heme oxygenase-1 (HO-1 was elevated in MI mice treated with cLA and nitrite compared to untreated MI mice. Confocal imaging on heart tissue confirmed expression the levels of HO-1 and p53. Taken together, these results suggest that therapeutic

  2. Inhibitory Effects of Benzaldehyde Derivatives from the Marine Fungus Eurotium sp. SF-5989 on Inflammatory Mediators via the Induction of Heme Oxygenase-1 in Lipopolysaccharide-Stimulated RAW264.7 Macrophages

    Kyoung-Su Kim

    2014-12-01

    Full Text Available Two benzaldehyde derivatives, flavoglaucin (1 and isotetrahydro-auroglaucin (2, were isolated from the marine fungus Eurotium sp. SF-5989 through bioassay- and 1H NMR-guided investigation. In this study, we evaluated the anti-inflammatory effects of these compounds in lipopolysaccharide (LPS-stimulated RAW264.7 macrophages. We demonstrated that compounds 1 and 2 markedly inhibited LPS-induced nitric oxide (NO and prostaglandin E2 (PGE2 production by suppressing inducible nitric oxide synthase (iNOS and cyclooxygenase-2 (COX-2 protein expression without affecting cell viability. We also demonstrated that the compounds reduced the secretion of pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α, interleukin-1β (IL-1β and interleukin-6 (IL-6. Furthermore, compounds 1 and 2 inhibited LPS-induced nuclear factor-κB (NF-κB activation by suppressing phosphorylation of IkappaB (IκB. These results indicated that the anti-inflammatory effects of these benzaldehyde derivatives in LPS-stimulated RAW264.7 macrophages were due to the inactivation of the NF-κB pathway. In addition, compounds 1 and 2 induced heme oxygenase-1 (HO-1 expression through the nuclear transcription factor-E2–related factor 2 (Nrf2 translocation. The inhibitory effects of compounds 1 and 2 on the production of pro-inflammatory mediators and on NF-κB binding activity were reversed by HO-1 inhibitor tin protoporphyrin (SnPP. Thus, the anti-inflammatory effects of compounds 1 and 2 also correlated with their ability of inducing HO-1 expression.

  3. Epalrestat increases glutathione, thioredoxin, and heme oxygenase-1 by stimulating Nrf2 pathway in endothelial cells

    Kaori Yama

    2015-04-01

    Full Text Available Epalrestat (EPS is the only aldose reductase inhibitor that is currently available for the treatment of diabetic neuropathy. Recently, we found that EPS at near-plasma concentration increases the intracellular levels of glutathione (GSH in rat Schwann cells. GSH plays a crucial role in protecting endothelial cells from oxidative stress, thereby preventing vascular diseases. Here we show that EPS increases GSH levels in not only Schwann cells but also endothelial cells. Treatment of bovine aortic endothelial cells (BAECs, an in vitro model of the vascular endothelium, with EPS caused a dramatic increase in intracellular GSH levels. This was concomitant with the up-regulation of glutamate cysteine ligase, an enzyme catalyzing the first and rate-limiting step in de novo GSH synthesis. Moreover, EPS stimulated the expression of thioredoxin and heme oxygenase-1, which have important redox regulatory functions in endothelial cells. Nuclear factor erythroid 2-related factor 2 (Nrf2 is a key transcription factor that regulates the expression of antioxidant genes. EPS increased nuclear Nrf2 levels in BAECs. Nrf2 knockdown by siRNA suppressed the EPS-induced glutamate cysteine ligase, thioredoxin-1, and heme oxygenase-1 expression. Interestingly, LY294002, an inhibitor of phosphatidylinositol 3-kinase, abolished the EPS-stimulated GSH synthesis, suggesting that the kinase is associated with Nrf2 activation induced by EPS. Furthermore, EPS reduced the cytotoxicity induced by H2O2 and tert-butylhydroperoxide, indicating that EPS plays a role in protecting cells from oxidative stress. Taken together, the results provide evidence that EPS exerts new beneficial effects on endothelial cells by increasing GSH, thioredoxin, and heme oxygenase-1 levels through the activation of Nrf2. We suggest that EPS has the potential to prevent several vascular diseases caused by oxidative stress.

  4. Protective effects of flavonoids isolated from Korean milk thistle Cirsium japonicum var. maackii (Maxim.) Matsum on tert-butyl hydroperoxide-induced hepatotoxicity in HepG2 cells.

    Jung, Hyun Ah; Abdul, Qudeer Ahmed; Byun, Jeong Su; Joung, Eun-Ji; Gwon, Wi-Gyeong; Lee, Min-Sup; Kim, Hyeung-Rak; Choi, Jae Sue

    2017-09-14

    Milk thistle leaves and flowers have been traditionally used as herbal remedy to alleviate liver diseases for decades. Korean milk thistle, Cirsium japonicum var. maackii (Maxim.) Matsum has been employed in traditional folk medicine as diuretic, antiphlogistic, hemostatic, and detoxifying agents. The aim of current investigation was to evaluate hepatoprotective properties of the MeOH extract of the roots, stems, leaves and flowers of Korean milk thistle as well as four isolated flavonoids, luteolin, luteolin 5-O-glucoside, apigenin and apigenin 7-O-glucuronide during t-BHP-induced oxidative stress in HepG2 cells. Hepatoprotective potential of the MeOH extracts and flavonoids derived from Korean milk thistle against t-BHP-induced oxidative stress in HepG2 cells were evaluated following MTT method. Incubating HepG2 cells with t-BHP markedly decreased the cell viability and increased the intracellular ROS generation accompanied by depleted GSH levels. Protein expression of heme oxygenase (HO-1) and nuclear factor-E2-related factor 2 (Nrf-2) was determined by Western blot. Our findings revealed that pretreating HepG2 cells with MeOH extracts and bioactive flavonoids significantly attenuated the t-BHP-induced oxidative damage, followed by increased cell viability in a dose-dependent manner. The results illustrate that excess ROS generation was reduced and GSH levels increased dose-dependently when HepG2 cells were pretreated with four flavonoids. Moreover, Western blotting analysis demonstrated that protein expressions of Nrf-2 and HO-1 were also up-regulated by flavonoids treatment. These results clearly demonstrate that the MeOH extracts and flavonoids from Korean milk thistle protected HepG2 cells against oxidative damage triggered by t-BHP principally by modulating ROS generation and restoring depleted GSH levels in addition to the increased Nrf-2/HO-1 signaling cascade. These flavonoids are potential natural antioxidative biomarkers against oxidative stress

  5. Anti-Inflammatory and Cytoprotective Effects of TMC-256C1 from Marine-Derived Fungus Aspergillus sp. SF-6354 via up-Regulation of Heme Oxygenase-1 in Murine Hippocampal and Microglial Cell Lines

    Dong-Cheol Kim

    2016-04-01

    Full Text Available In the course of searching for bioactive secondary metabolites from marine fungi, TMC-256C1 was isolated from an ethyl acetate extract of the marine-derived fungus Aspergillus sp. SF6354. TMC-256C1 displayed anti-neuroinflammatory effect in BV2 microglial cells induced by lipopolysaccharides (LPS as well as neuroprotective effect against glutamate-stimulated neurotoxicity in mouse hippocampal HT22 cells. TMC-256C1 was shown to develop a cellular resistance to oxidative damage caused by glutamate-induced cytotoxicity and reactive oxygen species (ROS generation in HT22 cells, and suppress the inflammation process in LPS-stimulated BV2 cells. Furthermore, the neuroprotective and anti-neuroinflammatory activities of TMC-256C1 were associated with upregulated expression of heme oxygenase (HO-1 and nuclear translocation of nuclear factor-E2-related factor 2 (Nrf2 in HT22 and BV2 cells. We also found that TMC-256C1 activated p38 mitogen-activated protein kinases (MAPK and phosphatidylinositol 3-kinase (PI3K/Akt signaling pathways in HT22 and BV2 cells. These results demonstrated that TMC-256C1 activates HO-1 protein expression, probably by increasing nuclear Nrf2 levels via the activation of the p38 MAPK and PI3K/Akt pathways.

  6. Short repeats in the heme oxygenase 1 gene promoter is associated with increased levels of inflammation, ferritin and higher risk of type-2 diabetes mellitus.

    Andrews, Mónica; Leiva, Elba; Arredondo-Olguín, Miguel

    2016-09-01

    We evaluated the relationship between the HO1 genotype, ferritin levels and the risk of type-2 diabetes and inflammation. Eight hundred thirty-five individuals were evaluated and classified according to their nutritional status and the presence of type-2 diabetes: 153 overweight (OW); 62 obese (OB); 55 type-2 diabetes mellitus (DM); 202 OWDM; 239 OBDM and 124 controls (C). We studied biochemical (glycemia, insulin, lipid profile, liver enzyme, creatinine, hsCRP), hematological (hemoglobin, free erythrocyte protoporphyrin, transferrin receptor and serum Fe and ferritin) and oxidative stress (SOD, GHS and TBARS) parameters. We determined heme oxygenase activity and the (GT)n polymorphism in its gene promoter. Individuals with diabetes, independent of nutritional status, showed high levels of ferritin and HO activity compared to control subjects. Allelic frequency was not different between the groups (Chi(2), NS) however, genotypes were different (Chi(2), P1). The SS (short-short) genotype was higher in all DM individuals compared to controls and MM was higher in controls. SM (short-medium) genotype was an independent risk factor for DM in logistic regression analysis. We observed high risk for type-2 diabetes mellitus in the presence of SM genotype and high levels of ferritin (OR adjusted: 2.7; 1.9-3.6; p1; compared to control group). It was also significantly related to inflammation. The SM genotype in HO1 gene promoter and ferritin levels were associated with higher risk for type-2 diabetes and for having a higher marker of inflammation, which is the main risk factor for the development of chronic diseases. Copyright © 2016 Elsevier GmbH. All rights reserved.

  7. Alteration of the Regiospecificity of Human Heme Oxygenase-1 by Unseating of the Heme but not Disruption of the Distal Hydrogen Bonding Network†

    Wang, Jinling; Evans, John P.; Ogura, Hiroshi; La Mar, Gerd N.; Ortiz de Montellano, Paul R.

    2008-01-01

    Heme oxygenase regiospecifically oxidizes heme at the α-meso position to give biliverdin IXα, CO, and iron. The heme orientation within the active site, which is thought to determine the oxidation regiospecificity, is shown here for the human enzyme (hHO1) to be largely determined by interactions between the heme carboxylic acid groups and residues Arg183 and Lys18 but not Tyr134. Mutation of either Arg183 or Lys18 individually does not significantly alter the NADPH-cytochrome P450 reductase-dependent reaction regiochemistry, but partially shifts the oxidation to the β/δ-meso positions in the reaction supported by ascorbic acid. Mutation of Glu29 to a lysine, which places a positive charge where it can interact with a heme carboxyl if the heme rotates by ~90°, causes a slight loss of regiospecificity, but combined with the R183E and K18E mutations results primarily in β/δ-meso oxidation of the heme under all conditions. NMR analysis of heme binding to the triple K18E/E29K/R183E mutant confirms rotation of the heme in the active site. Kinetic studies demonstrate that mutations of Arg183 greatly impair the rate of the P450 reductase-dependent reaction, in accord with the earlier finding that Arg183 is involved in binding of the reductase to hHO1, but have little effect on the ascorbate reaction. Mutations of Asp140 and Tyr58 that disrupt the active site hydrogen bonding network, impair catalytic rates but do not influence the oxidation regiochemistry. The results indicate both that the oxidation regiochemistry is largely controlled by ionic interactions of the heme propionic acid groups with the protein and that shifts in regiospecificity involve rotation of the heme about an axis perpendicular to the heme plane. PMID:16388581

  8. Differential expression patterns of Nqo1, AKR1B8 and Ho-1 in the liver and small intestine of C57BL/6 mice treated with sulforaphane

    Lin Luo

    2015-12-01

    Full Text Available This data article contains complementary figures and results related to the research article entitled “butylated hydroxyanisole induces distinct expression patterns of Nrf2 and detoxification enzymes in the liver and small intestine of C57BL/6 mice” (Luo et al., 2015 [1], which defined the basal and butylated hydroxyanisole (BHA-induced expression patterns of Phase II enzymes Nqo1, AKR1B8, and Ho-1 in the liver and small intestine of C57BL/6 mice. Sulforaphane [1-isothiocyanato-4-(methylsulfinylbutane] (SFN, a naturally occurring isothiocyanate derived from cruciferous vegetables, is a highly potent inducer of phase II cytoprotective enzymes. This dataset reports the histological changes of Nqo1, AKR1B8, and Ho-1 in wild-type (WT and Nrf2-/- mice induced by SFN. The mice were given a 25 mg/kg single oral dose of SFN for 24 h and 48 h. Immunohistochemistry revealed that, in the liver from WT mice, SFN increased Nqo1 staining in hepatocytes with slight higher staining in the pericentral region. The induction of AKR1B8 appeared mostly in hepatocytes in the periportal region. The basal and inducible Ho-1 was located predominately in Kupffer cells. In the small intestine from WT mice, the inducible expression of Nqo1 and AKR1B8 appeared more obvious in the villus than that in the crypt.

  9. Bioavailability of andrographolide and protection against carbon tetrachloride-induced oxidative damage in rats

    Chen, Haw-Wen [Department of Nutrition, China Medical University, Taichung, Taiwan (China); Huang, Chin-Shiu [Department of Health and Nutrition Biotechnology, Asia University, Taichung, Taiwan (China); Li, Chien-Chun [School of Nutrition, Chung Shan Medical University, Taichung, Taiwan (China); Department of Nutrition, Chung Shan Medical University Hospital, Taichung, Taiwan (China); Lin, Ai-Hsuan; Huang, Yu-Ju [Department of Nutrition, China Medical University, Taichung, Taiwan (China); Wang, Tsu-Shing [Department of Biomedical Science, Chung Shan Medical University, Taichung, Taiwan (China); Yao, Hsien-Tsung [Department of Nutrition, China Medical University, Taichung, Taiwan (China); Lii, Chong-Kuei [Department of Nutrition, China Medical University, Taichung, Taiwan (China); Department of Health and Nutrition Biotechnology, Asia University, Taichung, Taiwan (China)

    2014-10-01

    Andrographolide, a bioactive diterpenoid, is identified in Andrographis paniculata. In this study, we investigated the pharmacokinetics and bioavailability of andrographolide in rats and studied whether andrographolide enhances antioxidant defense in a variety of tissues and protects against carbon tetrachloride-induced oxidative damage. After a single 50-mg/kg administration, the maximum plasma concentration of andrographolide was 1 μM which peaked at 30 min. The bioavailability of andrographolide was 1.19%. In a hepatoprotection study, rats were intragastrically dosed with 30 or 50 mg/kg andrographolide for 5 consecutive days. The results showed that andrographolide up-regulated glutamate cysteine ligase (GCL) catalytic and modifier subunits, superoxide dismutase (SOD)-1, heme oxygenase (HO)-1, and glutathione (GSH) S-transferase (GST) Ya/Yb protein and mRNA expression in the liver, heart, and kidneys. The activity of SOD, GST, and GSH reductase was also increased in rats dosed with andrographolide (p < 0.05). Immunoblot analysis and EMSA revealed that andrographolide increased nuclear Nrf2 contents and Nrf2 binding to DNA, respectively. After the 5-day andrographolide treatment, one group of animals was intraperitoneally injected with carbon tetrachloride (CCl{sub 4}) at day 6. Andrographolide pretreatment suppressed CCl{sub 4}-induced plasma aminotransferase activity and hepatic lipid peroxidation (p < 0.05). These results suggest that andrographolide is quickly absorbed in the intestinal tract in rats with a bioavailability of 1.19%. Andrographolide protects against chemical-induced oxidative damage by up-regulating the gene transcription and activity of antioxidant enzymes in various tissues. - Highlights: • The bioavailability of andrographolide is 1.19% in rats. • Plasma concentration reaches 1 μM after giving 50 mg/kg andrographolide. • Andrographolide up-regulates Nrf2-dependent antioxidant genes. • Andrographolide increases antioxidant defense

  10. Bioavailability of andrographolide and protection against carbon tetrachloride-induced oxidative damage in rats

    Chen, Haw-Wen; Huang, Chin-Shiu; Li, Chien-Chun; Lin, Ai-Hsuan; Huang, Yu-Ju; Wang, Tsu-Shing; Yao, Hsien-Tsung; Lii, Chong-Kuei

    2014-01-01

    Andrographolide, a bioactive diterpenoid, is identified in Andrographis paniculata. In this study, we investigated the pharmacokinetics and bioavailability of andrographolide in rats and studied whether andrographolide enhances antioxidant defense in a variety of tissues and protects against carbon tetrachloride-induced oxidative damage. After a single 50-mg/kg administration, the maximum plasma concentration of andrographolide was 1 μM which peaked at 30 min. The bioavailability of andrographolide was 1.19%. In a hepatoprotection study, rats were intragastrically dosed with 30 or 50 mg/kg andrographolide for 5 consecutive days. The results showed that andrographolide up-regulated glutamate cysteine ligase (GCL) catalytic and modifier subunits, superoxide dismutase (SOD)-1, heme oxygenase (HO)-1, and glutathione (GSH) S-transferase (GST) Ya/Yb protein and mRNA expression in the liver, heart, and kidneys. The activity of SOD, GST, and GSH reductase was also increased in rats dosed with andrographolide (p < 0.05). Immunoblot analysis and EMSA revealed that andrographolide increased nuclear Nrf2 contents and Nrf2 binding to DNA, respectively. After the 5-day andrographolide treatment, one group of animals was intraperitoneally injected with carbon tetrachloride (CCl 4 ) at day 6. Andrographolide pretreatment suppressed CCl 4 -induced plasma aminotransferase activity and hepatic lipid peroxidation (p < 0.05). These results suggest that andrographolide is quickly absorbed in the intestinal tract in rats with a bioavailability of 1.19%. Andrographolide protects against chemical-induced oxidative damage by up-regulating the gene transcription and activity of antioxidant enzymes in various tissues. - Highlights: • The bioavailability of andrographolide is 1.19% in rats. • Plasma concentration reaches 1 μM after giving 50 mg/kg andrographolide. • Andrographolide up-regulates Nrf2-dependent antioxidant genes. • Andrographolide increases antioxidant defense in

  11. Isorhamnetin inhibits Prevotella intermedia lipopolysaccharide-induced production of interleukin-6 in murine macrophages via anti-inflammatory heme oxygenase-1 induction and inhibition of nuclear factor-κB and signal transducer and activator of transcription 1 activation.

    Jin, J Y; Choi, E Y; Park, H R; Choi, J I; Choi, I S; Kim, S J

    2013-12-01

    Interleukin-6 (IL-6) is a key proinflammatory cytokine that has been considered to be important in the pathogenesis of periodontal disease. Therefore, host-modulatory agents directed at inhibiting IL-6 appear to be beneficial in terms of attenuating periodontal disease progression and potentially improving disease susceptibility. In the current study, we investigated the effect of the flavonoid isorhamnetin on the production of IL-6 in murine macrophages stimulated with lipopolysaccharide (LPS) from Prevotella intermedia, a pathogen implicated in inflammatory periodontal disease, and its mechanisms of action. Lipopolysaccharide from P. intermedia ATCC 25611 was isolated using the standard hot phenol-water method. Culture supernatants were collected and assayed for IL-6. We used real-time PCR to quantify IL-6 and heme oxygenase-1 (HO-1) mRNA expression. The expression of HO-1 protein and the levels of signaling proteins were monitored using immunoblot analyses. The DNA-binding activity of nuclear factor-κB (NF-κB) was analyzed using ELISA-based assay kits. Isorhamnetin significantly down-regulated P. intermedia LPS-induced production of IL-6 as well as its mRNA expression in RAW264.7 cells. Isorhamnetin up-regulated the expression of HO-1 at both gene transcription and translation levels in cells stimulated with P. intermedia LPS. In addition, inhibition of HO-1 activity by tin protoporphyrin IX blocked the inhibitory effect of isorhamnetin on IL-6 production. Isorhamnetin failed to prevent LPS from activating either c-Jun N-terminal kinase or p38 pathways. Isorhamnetin did not inhibit NF-κB transcriptional activity at the level of inhibitory κB-α degradation. Isorhamnetin suppressed NF-κB signaling through inhibition of nuclear translocation and DNA binding activity of NF-κB p50 subunit and attenuated signal transducer and activator of transcription 1 signaling. Although further research is required to clarify the detailed mechanism of action, we propose

  12. Heme oxygenase activity increases after exercise in healthy volunteers

    AbstractHeme oxygenase (HO) is an essential, rate-limiting protein which participates in the catabolism of heme to iron, carbon monoxide (CO), and biliverdin. The alpha methene bridge carbon of the heme is eliminated as CO which can be measured as blood carboxyhemoglobin (COHb)....

  13. Punicalagin, a PTP1B inhibitor, induces M2c phenotype polarization via up-regulation of HO-1 in murine macrophages.

    Xu, Xiaolong; Guo, Yuhong; Zhao, Jingxia; He, Shasha; Wang, Yan; Lin, Yan; Wang, Ning; Liu, Qingquan

    2017-09-01

    Current data have shown that punicalagin (PUN), an ellagitannin isolated from pomegranate, possesses anti-inflammatory and anti-oxidant properties; however, its direct targets have not yet been reported. This is the first report that PTP1B serves as a direct target of PUN, with IC 50 value of 1.04μM. Results from NPOI further showed that the K on and K off of PUN-PTP1B complex were 3.38e2M -1 s -1 and 4.13e-3s -1 , respectively. The active site Arg24 of PTP1B was identified as a key binding site of PUN by computation simulation and point mutation. Moreover, inhibition of PTP1B by PUN promoted an M2c-like macrophage polarization and enhanced anti-inflammatory cytokines expression, including IL-10 and M-CSF. Based on gene expression profile, we elucidated that PUN treatment significantly up-regulated 275 genes and down-regulated 1059 genes. M1-like macrophage marker genes, such as Tlr4, Irf1/2, Hmgb1, and Stat1 were down-regulated, while M2 marker genes, including Tmem171, Gpr35, Csf1, Il1rn, Cebpb, Fos, Vegfα, Slc11a1, and Bhlhe40 were up-regulated in PUN-treated macrophages. Hmox-1, a gene encoding HO-1 protein, was preferentially expressed with 16-fold change. Inhibition of HO-1 obviously restored PUN-induced M2 polarization and IL-10 secretion. In addition, phosphorylation of both Akt and STAT3 contributed to PUN-induced HO-1 expression. This study provided new insights into the mechanisms of PUN-mediated anti-inflammatory and anti-oxidant activities and provided new therapeutic strategies for inflammatory diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Pinocembrin Suppresses H2O2-Induced Mitochondrial Dysfunction by a Mechanism Dependent on the Nrf2/HO-1 Axis in SH-SY5Y Cells.

    de Oliveira, Marcos Roberto; da Costa Ferreira, Gustavo; Brasil, Flávia Bittencourt; Peres, Alessandra

    2018-02-01

    Mitochondria are susceptible to redox impairment, which has been associated with neurodegeneration. These organelles are both a source and target of reactive species. In that context, there is increasing interest in finding natural compounds that modulate mitochondrial function and mitochondria-related signaling in order to prevent or to treat diseases involving mitochondrial impairment. Herein, we investigated whether and how pinocembrin (PB) would prevent mitochondrial dysfunction elicited by the exposure of human neuroblastoma SH-SY5Y cells to hydrogen peroxide (H 2 O 2 ). PB (25 μM) was administrated for 4 h before H 2 O 2 treatment (300 μM for 24 h). PB prevented H 2 O 2 -induced loss of cell viability mitochondrial depolarization in SH-SY5Y cells. PB also attenuated redox impairment in mitochondrial membranes. The production of superoxide anion radical (O 2 -• ) and nitric oxide (NO • ) was alleviated by PB in cells exposed to H 2 O 2 . PB suppressed the H 2 O 2 -induced inhibition of the tricarboxylic acid (TCA) cycle enzymes aconitase, α-ketoglutarate dehydrogenase, and succinate dehydrogenase. Furthermore, PB induced anti-inflammatory effects by abolishing the H 2 O 2 -dependent activation of the nuclear factor-κB (NF-κB) and upregulation of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α). The PB-induced antioxidant and anti-inflammatory effects are dependent on the heme oxygenate-1 (HO-1) enzyme and on the activation of the transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2), since HO-1 inhibition (with 0.5 μM ZnPP IX) or Nrf2 silencing (with small interfering RNA (siRNA)) abolished the effects of PB. Overall, PB afforded cytoprotection by the Nrf2/HO-1 axis in H 2 O 2 -treated SH-SY5Y cells.

  15. MAT, a Novel Polyherbal Aphrodisiac Formulation, Enhances Sexual Function and Nrf2/HO-1 Pathway While Reducing Oxidative Damage in Male Rats

    Kazim Sahin

    2018-01-01

    Full Text Available Mucuna pruriens, Ashwagandha, and Tribulus terrestris are known as the enhancers for sexual health, functional activities, vitality, and longevity. These herbs had been widely used in the Ayurveda medicine as aphrodisiacs through the ages, and their efficacy was also verified separately in our previous publication. Therefore, the aim of this study was to determine the effects of Mucuna, Ashwagandha, and Tribulus complexes on sexual function in rats. Twenty-eight male rats allocated to four groups as follows: (i negative control (C; (ii positive control or sildenafil citrate treated group (5 mg/kg (S; (iii MAT1 (combination of 10 mg Mucuna (M + 10 mg Ashwagandha (A + 10 mg Tribulus (T/kg BW; (iv MAT 2 (20 mg Mucuna + 20 mg Ashwagandha + 20 mg Tribulus/kg BW. There was no significant difference found between the MAT1 and MAT2 groups while they showed significantly increased testosterone, follicle-stimulating hormone (FSH, and luteinizing hormone (LH levels when compared to the negative control. Significant increases in Nrf2/HO1 levels and decreases in NF-κB were detected in MAT groups similar to the decrease in serum and testis malondialdehyde (MDA levels as compared to both controls. The sperm motility, count, and rate also significantly improved in both MAT groups, while ALT, AST, creatinine, ALP, and urea levels did not change in any of the groups. Oral consumption of MATs combination in male rats resulted in inhibition of NF-κB and MDA and also increased sex hormones with Nrf2-mediated HO-1 induction. MAT combinations may improve sexual functions by increasing levels of sexual hormones and regulation of NF-κB and Nrf2/HO-1 signaling pathways.

  16. MAT, a Novel Polyherbal Aphrodisiac Formulation, Enhances Sexual Function and Nrf2/HO-1 Pathway While Reducing Oxidative Damage in Male Rats

    Tuzcu, Mehmet; Orhan, Cemal; Sahin, Nurhan; Akdemir, Fatih; Yilmaz, Ismet

    2018-01-01

    Mucuna pruriens, Ashwagandha, and Tribulus terrestris are known as the enhancers for sexual health, functional activities, vitality, and longevity. These herbs had been widely used in the Ayurveda medicine as aphrodisiacs through the ages, and their efficacy was also verified separately in our previous publication. Therefore, the aim of this study was to determine the effects of Mucuna, Ashwagandha, and Tribulus complexes on sexual function in rats. Twenty-eight male rats allocated to four groups as follows: (i) negative control (C); (ii) positive control or sildenafil citrate treated group (5 mg/kg) (S); (iii) MAT1 (combination of 10 mg Mucuna (M) + 10 mg Ashwagandha (A) + 10 mg Tribulus (T)/kg BW); (iv) MAT 2 (20 mg Mucuna + 20 mg Ashwagandha + 20 mg Tribulus/kg BW). There was no significant difference found between the MAT1 and MAT2 groups while they showed significantly increased testosterone, follicle-stimulating hormone (FSH), and luteinizing hormone (LH) levels when compared to the negative control. Significant increases in Nrf2/HO1 levels and decreases in NF-κB were detected in MAT groups similar to the decrease in serum and testis malondialdehyde (MDA) levels as compared to both controls. The sperm motility, count, and rate also significantly improved in both MAT groups, while ALT, AST, creatinine, ALP, and urea levels did not change in any of the groups. Oral consumption of MATs combination in male rats resulted in inhibition of NF-κB and MDA and also increased sex hormones with Nrf2-mediated HO-1 induction. MAT combinations may improve sexual functions by increasing levels of sexual hormones and regulation of NF-κB and Nrf2/HO-1 signaling pathways. PMID:29853975

  17. Unique sex- and age-dependent effects in protective pathways in acute kidney injury.

    Boddu, Ravindra; Fan, Chunlan; Rangarajan, Sunil; Sunil, Bhuvana; Bolisetty, Subhashini; Curtis, Lisa M

    2017-09-01

    Sex and age influence susceptibility to acute kidney injury (AKI), with young females exhibiting lowest incidence. In these studies, we investigated mechanisms which may underlie the sex/age-based dissimilarities. Cisplatin (Cp)-induced AKI resulted in morphological evidence of injury in all groups. A minimal rise in plasma creatinine (PCr) was seen in Young Females, whereas in Aged Females, PCr rose precipitously. Relative to Young Males, Aged Males showed significantly, but temporally, comparably elevated PCr. Notably, Aged Females showed significantly greater mortality, whereas Young Females exhibited none. Tissue KIM-1 and plasma NGAL were significantly lower in Young Females than all others. IGFBP7 levels were modestly increased in both Young groups. IGFBP7 levels in Aged Females were significantly elevated at baseline relative to Aged Males, and increased linearly through day 3 , when these levels were comparable in both Aged groups. Plasma cytokine levels similarly showed a pattern of protective effects preferentially in Young Females. Expression of the drug transporter MATE2 did not explain the sex/age distinctions. Heme oxygenase-1 (HO-1) levels (~28-kDa species) showed elevation at day 1 in all groups with highest levels seen in Young Males. Exclusively in Young Females, these levels returned to baseline on day 3 , suggestive of a more efficient recovery. In aggregate, we demonstrate, for the first time, a distinctive pattern of response to AKI in Young Females relative to males which appears to be significantly altered in aging. These distinctions may offer novel targets to exploit therapeutically in both females and males in the treatment of AKI.

  18. Hepatoprotective effect of curcumin in lipopolysaccharide/D-galactosamine model of liver injury in rats: Relationship to HO-1/CO antioxidant system

    Černý, D.; Lekič, N.; Váňová, K.; Muchová, L.; Hořínek, A.; Kmoníčková, Eva; Zídek, Zdeněk; Kameníková, L.; Farghali, H.

    2011-01-01

    Roč. 82, č. 5 (2011), s. 786-791 ISSN 0367-326X R&D Projects: GA ČR GA305/07/0061 Grant - others:GA ČR(CZ) GA305/09/0004 Institutional research plan: CEZ:AV0Z50390512 Keywords : Curcumin * Heme oxygenase-1 * Carbon monoxide Subject RIV: FR - Pharmacology ; Medidal Chemistry Impact factor: 1.848, year: 2011

  19. EGFR mediates astragaloside IV-induced Nrf2 activation to protect cortical neurons against in vitro ischemia/reperfusion damages

    Gu, Da-min; Lu, Pei-Hua; Zhang, Ke; Wang, Xiang; Sun, Min; Chen, Guo-Qian; Wang, Qiong

    2015-01-01

    In this study, we tested the potential role of astragaloside IV (AS-IV) against oxygen and glucose deprivation/re-oxygenation (OGD/R)-induced damages in murine cortical neurons, and studied the associated signaling mechanisms. AS-IV exerted significant neuroprotective effects against OGD/R by reducing reactive oxygen species (ROS) accumulation, thereby attenuating oxidative stress and neuronal cell death. We found that AS-IV treatment in cortical neurons resulted in NF-E2-related factor 2 (Nrf2) signaling activation, evidenced by Nrf2 Ser-40 phosphorylation, and its nuclear localization, as well as transcription of antioxidant-responsive element (ARE)-regulated genes: heme oxygenase-1 (HO-1), NAD(P)H:quinone oxidoreductase 1 (NQO-1) and sulphiredoxin 1 (SRXN-1). Knockdown of Nrf2 through lentiviral shRNAs prevented AS-IV-induced ARE genes transcription, and abolished its anti-oxidant and neuroprotective activities. Further, we discovered that AS-IV stimulated heparin-binding-epidermal growth factor (HB-EGF) release to trans-activate epidermal growth factor receptor (EGFR) in cortical neurons. Blockage or silencing EGFR prevented Nrf2 activation by AS-IV, thus inhibiting AS-IV-mediated anti-oxidant and neuroprotective activities against OGD/R. In summary, AS-IV protects cortical neurons against OGD/R damages through activating of EGFR-Nrf2 signaling. - Highlights: • Pre-treatment of astragaloside IV (AS-IV) protects murine cortical neurons from OGD/R. • AS-IV activates Nrf2-ARE signaling in murine cortical neurons. • Nrf2 is required for AS-IV-mediated anti-oxidant and neuroprotective activities. • AS-IV stimulates HB-EGF release to trans-activate EGFR in murine cortical neurons. • EGFR mediates AS-IV-induced Nrf2 activation and neuroprotection against OGD/R

  20. EGFR mediates astragaloside IV-induced Nrf2 activation to protect cortical neurons against in vitro ischemia/reperfusion damages

    Gu, Da-min [Department of Anesthesiology, Affiliated Yixing People' s Hospital, Jiangsu University, Yixing (China); Lu, Pei-Hua, E-mail: lphty1_1@163.com [Department of Medical Oncology, Wuxi People' s Hospital Affiliated to Nanjing Medical University, Wuxi (China); Zhang, Ke; Wang, Xiang [Department of Anesthesiology, Affiliated Yixing People' s Hospital, Jiangsu University, Yixing (China); Sun, Min [Department of General Surgery, Affiliated Yixing People' s Hospital, Jiangsu University, Yixing (China); Chen, Guo-Qian [Department of Clinical Laboratory, Wuxi People' s Hospital Affiliated to Nanjing Medical University, Wuxi (China); Wang, Qiong, E-mail: WangQiongprof1@126.com [Department of Clinical Laboratory, Wuxi People' s Hospital Affiliated to Nanjing Medical University, Wuxi (China)

    2015-02-13

    In this study, we tested the potential role of astragaloside IV (AS-IV) against oxygen and glucose deprivation/re-oxygenation (OGD/R)-induced damages in murine cortical neurons, and studied the associated signaling mechanisms. AS-IV exerted significant neuroprotective effects against OGD/R by reducing reactive oxygen species (ROS) accumulation, thereby attenuating oxidative stress and neuronal cell death. We found that AS-IV treatment in cortical neurons resulted in NF-E2-related factor 2 (Nrf2) signaling activation, evidenced by Nrf2 Ser-40 phosphorylation, and its nuclear localization, as well as transcription of antioxidant-responsive element (ARE)-regulated genes: heme oxygenase-1 (HO-1), NAD(P)H:quinone oxidoreductase 1 (NQO-1) and sulphiredoxin 1 (SRXN-1). Knockdown of Nrf2 through lentiviral shRNAs prevented AS-IV-induced ARE genes transcription, and abolished its anti-oxidant and neuroprotective activities. Further, we discovered that AS-IV stimulated heparin-binding-epidermal growth factor (HB-EGF) release to trans-activate epidermal growth factor receptor (EGFR) in cortical neurons. Blockage or silencing EGFR prevented Nrf2 activation by AS-IV, thus inhibiting AS-IV-mediated anti-oxidant and neuroprotective activities against OGD/R. In summary, AS-IV protects cortical neurons against OGD/R damages through activating of EGFR-Nrf2 signaling. - Highlights: • Pre-treatment of astragaloside IV (AS-IV) protects murine cortical neurons from OGD/R. • AS-IV activates Nrf2-ARE signaling in murine cortical neurons. • Nrf2 is required for AS-IV-mediated anti-oxidant and neuroprotective activities. • AS-IV stimulates HB-EGF release to trans-activate EGFR in murine cortical neurons. • EGFR mediates AS-IV-induced Nrf2 activation and neuroprotection against OGD/R.

  1. Curcumin protects against cytotoxic and inflammatory effects of quartz particles but causes oxidative DNA damage in a rat lung epithelial cell line

    Li Hui; Berlo, Damien van; Shi Tingming; Speit, Guenter; Knaapen, Ad M.; Borm, Paul J.A.; Albrecht, Catrin; Schins, Roel P.F.

    2008-01-01

    Chronic inhalation of high concentrations of respirable quartz particles has been implicated in various lung diseases including lung fibrosis and cancer. Generation of reactive oxygen species (ROS) and oxidative stress is considered a major mechanism of quartz toxicity. Curcumin, a yellow pigment from Curcuma longa, has been considered as nutraceutical because of its strong anti-inflammatory, antitumour and antioxidant properties. The aim of our present study was to investigate whether curcumin can protect lung epithelial cells from the cytotoxic, genotoxic and inflammatory effects associated with quartz (DQ12) exposure. Electron paramagnetic resonance (EPR) measurements using the spin-trap DMPO demonstrated that curcumin reduces hydrogen peroxide-dependent hydroxyl-radical formation by quartz. Curcumin was also found to reduce quartz-induced cytotoxicity and cyclooxygenase 2 (COX-2) mRNA expression in RLE-6TN rat lung epithelial cells (RLE). Curcumin also inhibited the release of macrophage inflammatory protein-2 (MIP-2) from RLE cells as observed upon treatment with interleukin-1 beta (IL-1β) and tumour necrosis factor-alpha (TNFα). However, curcumin failed to protect the RLE cells from oxidative DNA damage induced by quartz, as shown by formamidopyrimidine glycosylase (FPG)-modified comet assay and by immunocytochemistry for 8-hydroxydeoxyguanosine. In contrast, curcumin was found to be a strong inducer of oxidative DNA damage itself at non-cytotoxic and anti-inflammatory concentrations. In line with this, curcumin also enhanced the mRNA expression of the oxidative stress response gene heme oxygenase-1 (ho-1). Curcumin also caused oxidative DNA damage in NR8383 rat alveolar macrophages and A549 human lung epithelial cells. Taken together, these observations indicate that one should be cautious in considering the potential use of curcumin in the prevention or treatment of lung diseases associated with quartz exposure

  2. The right choice of antihypertensives protects primary human hepatocytes from ethanol- and recombinant human TGF-β1-induced cellular damage

    Ehnert S

    2013-03-01

    damage, except for furosemide, which had no effect. As a common mechanism, all antihypertensives increased heme-oxygenase-1 (HO-1 expression, and inhibition of HO-1 activity reversed the protective effect of the drugs. Interestingly, Smad3/4 signaling was reduced by all compounds except furosemide, which even enhanced this profibrotic signaling. This effect was mediated by expressional changes of Smad3 and/or Smad4.Conclusions: Our results suggest that antihypertensives may both positively and negatively influence chronic liver disease progression. Therefore, we propose that in future patients with ALD and high blood pressure, they could benefit from an adjusted antihypertensive therapy with additional antifibrotic effects.Keywords: primary human hepatocytes, alcoholic liver disease, ethanol, TGF-β1, antihypertensives

  3. Effects of heme oxygenase-1 gene modulated mesenchymal stem cells on vasculogenesis in ischemic swine hearts.

    Jiang, Yi-Bo; Zhang, Xiao-Li; Tang, Yao-Liang; Ma, Gen-Shan; Shen, Cheng-Xing; Wei, Qin; Zhu, Qi; Yao, Yu-Yu; Liu, Nai-Feng

    2011-02-01

    Mesenchymal stem cells (MSCs) transplantation may partially restore heart function in the treatment of acute myocardial infarction (AMI). The aim of this study was to explore the beneficial effects of MSCs modified with heme xygenase-1 (HO-1) on post-infarct swine hearts to determine whether the induction of therapeutic angiogenesis is modified by the angiogenic cytokines released from the implanted cells. In vitro, MSCs were divided into four groups: (1) non-transfected MSCs (MSCs group), (2) MSCs transfected with the pcDNA3.1-Lacz plasmid (Lacz-MSCs group), (3) MSCs transfected with pcDNA3.1-hHO-1 (HO-1-MSCs group), and (4) MSCs transfected with pcDNA3.1-hHO-1 and pretreatment with an HO inhibitor, tin protoporphyrin (SnPP) (HO-1-MSCs + SnPP group). Cells were cultured in an airtight incubation bottle for 24 hours, in which the oxygen concentration was maintained at < 1%, followed by 12 hours of reoxygenation. After hypoxia/reoxygen treatment, ELISA was used to measure transforming growth factor (TGF-β) and fibroblast growth factor (FGF-2) in the supernatant. In vivo, 28 Chinese mini-pigs were randomly allocated to the following treatment groups: (1) control group (saline), (2) Lacz-MSCs group, (3) HO-1-MSCs group, and (4) HO-1-MSCs + SnPP group. About 1 × 10(7) of autologous stem cells or an identical volume of saline was injected intracoronary into porcine hearts 1 hour after MI. Magnetic resonance imaging (MRI) assay and postmortem analysis were assessed four weeks after stem cell transplantation. Post hypoxia/reoxygenation in vitro, TGF-β in the supernatant was significantly increased in the HO-1-MSCs ((874.88 ± 68.23) pg/ml) compared with Lacz-MSCs ((687.81 ± 57.64) pg/ml, P < 0.001). FGF-2 was also significantly increased in the HO-1-MSCs ((1106.48 ± 107.06) pg/ml) compared with the Lacz-MSCs ((853.85 ± 74.44) pg/ml, P < 0.001). In vivo, at four weeks after transplantation, HO-1 gene transfer increased the capillary density in the peri-infarct area

  4. Cysteine-independent activation/inhibition of heme oxygenase-2

    Dragic Vukomanovic

    2016-01-01

    Full Text Available Reactive thiols of cysteine (cys residues in proteins play a key role in transforming chemical reactivity into a biological response. The heme oxygenase-2 (HO-2 isozyme contains two cys residues that have been implicated in binding of heme and also the regulation of its activity. In this paper, we address the question of a role for cys residues for the HO-2 inhibitors or activators designed in our laboratory. We tested the activity of full length recombinant human heme oxygenase-2 (FL-hHO-2 and its analog in which cys265 and cys282 were both replaced by alanine to determine the effect on activation by menadione (MD and inhibition by QC-2350. Similar inhibition by QC-2350 and almost identical activation by MD was observed for both recombinant FL-hHO-2s. Our findings are interpreted to mean that thiols of FL-hHO-2s are not involved in HO-2 activation or inhibition by the compounds that have been designed and identified by us. Activation or inhibition of HO-2 by our compounds should be attributed to a mechanism other than altering binding affinity of HO-2 for heme through cys265 and cys282.

  5. Cysteine-independent activation/inhibition of heme oxygenase-2.

    Vukomanovic, Dragic; Rahman, Mona N; Maines, Mahin D; Ozolinš, Terence Rs; Szarek, Walter A; Jia, Zongchao; Nakatsu, Kanji

    2016-03-01

    Reactive thiols of cysteine (cys) residues in proteins play a key role in transforming chemical reactivity into a biological response. The heme oxygenase-2 (HO-2) isozyme contains two cys residues that have been implicated in binding of heme and also the regulation of its activity. In this paper, we address the question of a role for cys residues for the HO-2 inhibitors or activators designed in our laboratory. We tested the activity of full length recombinant human heme oxygenase-2 (FL-hHO-2) and its analog in which cys265 and cys282 were both replaced by alanine to determine the effect on activation by menadione (MD) and inhibition by QC-2350. Similar inhibition by QC-2350 and almost identical activation by MD was observed for both recombinant FL-hHO-2s. Our findings are interpreted to mean that thiols of FL-hHO-2s are not involved in HO-2 activation or inhibition by the compounds that have been designed and identified by us. Activation or inhibition of HO-2 by our compounds should be attributed to a mechanism other than altering binding affinity of HO-2 for heme through cys265 and cys282.

  6. Heme oxygenase-1 affects generation and spontaneous cardiac differentiation of induced pluripotent stem cells.

    Stepniewski, Jacek; Pacholczak, Tomasz; Skrzypczyk, Aniela; Ciesla, Maciej; Szade, Agata; Szade, Krzysztof; Bidanel, Romain; Langrzyk, Agnieszka; Grochowski, Radoslaw; Vandermeeren, Felix; Kachamakova-Trojanowska, Neli; Jez, Mateusz; Drabik, Grazyna; Nakanishi, Mahito; Jozkowicz, Alicja; Dulak, Jozef

    2018-02-01

    Cellular stress can influence efficiency of iPSCs generation and their differentiation. However, the role of intracellular cytoprotective factors in these processes is still not well known. Therefore, we investigated the effect of HO-1 (Hmox1) or Nrf2 (Nfe2l2), two major cytoprotective genes. Hmox1 -/- fibroblasts demonstrated decreased reprogramming efficiency in comparison to Hmox1 +/+ cells. Reversely, pharmacological enhancement of HO-1 resulted in higher number of iPSCs colonies. Importantly, elevated level of both p53 and p53-regulated miR-34a and 14-3-3σ was observed in HO-1-deficient fibroblasts whereas downregulation of p53 in these cells markedly increased their reprogramming efficiency. In human fibroblasts HO-1 silencing also induced p53 expression and affected reprogramming outcome. Hmox1 +/+ and Hmox1 -/- iPSCs similarly differentiated in vitro to cells originating from three germ layers, however, lower number of contracting cells was observed during this process in HO-1-deficient cells indicating attenuated cardiac differentiation. Importantly, silencing of Hmox1 in murine ESC using CRISPR/Cas-9 editing also impaired their spontaneous cardiac differentiation. Decreased reprogramming efficiency was also observed in Nrf2-lacking fibroblasts. Reversely, sulforaphane, a Nrf2 activator, increased the number of iPSCs colonies. However, both Nfe2l2 +/+ and Nfe2l2 -/- iPSCs showed similar pluripotency and differentiation capacity. These results indicate that regulation of HO-1 expression can further optimize generation and cardiac differentiation of iPSCs. © 2018 IUBMB Life, 70(2):129-142, 2018. © 2018 International Union of Biochemistry and Molecular Biology.

  7. Active site architecture of a sugar N-oxygenase.

    Thoden, James B; Branch, Megan C; Zimmer, Alex L; Bruender, Nathan A; Holden, Hazel M

    2013-05-14

    KijD3 is a flavin-dependent N-oxygenase implicated in the formation of the nitro-containing sugar d-kijanose, found attached to the antibiotic kijanimicin. For this investigation, the structure of KijD3 in complex with FMN and its dTDP-sugar substrate was solved to 2.1 Å resolution. In contrast to the apoenzyme structure, the C-terminus of the protein becomes ordered and projects into the active site cleft [Bruender, N. A., Thoden, J. B., and Holden, H. M. (2010) Biochemistry 49, 3517-3524]. The amino group of the dTDP-aminosugar that is oxidized is located 4.9 Å from C4a of the flavin ring. The model provides a molecular basis for understanding the manner in which KijD3 catalyzes its unusual chemical transformation.

  8. Retuning Rieske-type Oxygenases to Expand Substrate Range

    Mohammadi, Mahmood; Viger, Jean-François; Kumar, Pravindra; Barriault, Diane; Bolin, Jeffrey T.; Sylvestre, Michel (INRS); (Purdue)

    2012-09-17

    Rieske-type oxygenases are promising biocatalysts for the destruction of persistent pollutants or for the synthesis of fine chemicals. In this work, we explored pathways through which Rieske-type oxygenases evolve to expand their substrate range. BphAE{sub p4}, a variant biphenyl dioxygenase generated from Burkholderia xenovorans LB400 BphAE{sub LB400} by the double substitution T335A/F336M, and BphAE{sub RR41}, obtained by changing Asn{sup 338}, Ile{sup 341}, and Leu{sup 409} of BphAE{sub p4} to Gln{sup 338}, Val{sup 341}, and Phe{sup 409}, metabolize dibenzofuran two and three times faster than BphAE{sub LB400}, respectively. Steady-state kinetic measurements of single- and multiple-substitution mutants of BphAE{sub LB400} showed that the single T335A and the double N338Q/L409F substitutions contribute significantly to enhanced catalytic activity toward dibenzofuran. Analysis of crystal structures showed that the T335A substitution relieves constraints on a segment lining the catalytic cavity, allowing a significant displacement in response to dibenzofuran binding. The combined N338Q/L409F substitutions alter substrate-induced conformational changes of protein groups involved in subunit assembly and in the chemical steps of the reaction. This suggests a responsive induced fit mechanism that retunes the alignment of protein atoms involved in the chemical steps of the reaction. These enzymes can thus expand their substrate range through mutations that alter the constraints or plasticity of the catalytic cavity to accommodate new substrates or that alter the induced fit mechanism required to achieve proper alignment of reaction-critical atoms or groups.

  9. Heme oxygenase activity correlates with serum indices of iron homeostasis in healthy nonsmokers

    Heme oxygenase (HO) catalyzes the breakdown of heme to carbon monoxide, iron, and biliverdin. While the use of genetically altered animal models in investigation has established distinct associations between HO activity and systemic iron availability, studies have not yet confirm...

  10. In vitro Activation of heme oxygenase-2 by menadione and its analogs.

    Vukomanovic, Dragic; Rahman, Mona N; Bilokin, Yaroslav; Golub, Andriy G; Brien, James F; Szarek, Walter A; Jia, Zongchao; Nakatsu, Kanji

    2014-02-18

    Previously, we reported that menadione activated rat, native heme oxygenase-2 (HO-2) and human recombinant heme oxygenase-2 selectively; it did not activate spleen, microsomal heme oxygenase-1. The purpose of this study was to explore some structure-activity relationships of this activation and the idea that redox properties may be an important aspect of menadione efficacy. Heme oxygenase activity was determined in vitro using rat spleen and brain microsomes as the sources of heme oxygenase-1 and -2, respectively, as well as recombinant, human heme oxygenase-2. Menadione analogs with bulky aliphatic groups at position-3, namely vitamins K1 and K2, were not able to activate HO-2. In contrast, several compounds with similar bulky but less lipophilic moieties at position-2 (and -3) were able to activate HO-2 many fold; these compounds included polar, rigid, furan-containing naphthoquinones, furan-benzoxazine naphthoquinones, 2-(aminophenylphenyl)-3-piperidin-1-yl naphthoquinones. To explore the idea that redox properties might be involved in menadione efficacy, we tested analogs such as 1,4-dimethoxy-2-methylnaphthalene, pentafluoromenadione, monohalogenated naphthoquinones, α-tetralone and 1,4-naphthoquinone. All of these compounds were inactive except for 1,4-naphthoquinone. Menadione activated full-length recombinant human heme oxygenase-2 (FL-hHO-2) as effectively as rat brain enzyme, but it did not activate rat spleen heme oxygenase. These observations are consistent with the idea that naphthoquinones such as menadione bind to a receptor in HO-2 and activate the enzyme through a mechanism that may involve redox properties.

  11. Heme oxygenase-2 gene deletion attenuates oxidative stress in neurons exposed to extracellular hemin

    Benvenisti-Zarom Luna

    2004-09-01

    Full Text Available Abstract Background Hemin, the oxidized form of heme, accumulates in intracranial hematomas and is a potent oxidant. Growing evidence suggests that it contributes to delayed injury to surrounding tissue, and that this process is affected by the heme oxygenase enzymes. In a prior study, heme oxygenase-2 gene deletion increased the vulnerability of cultured cortical astrocytes to hemin. The present study tested the effect of HO-2 gene deletion on protein oxidation, reactive oxygen species formation, and cell viability after mixed cortical neuron/astrocyte cultures were incubated with neurotoxic concentrations of hemin. Results Continuous exposure of wild-type cultures to 1–10 μM hemin for 14 h produced concentration-dependent neuronal death, as detected by both LDH release and fluorescence intensity after propidium iodide staining, with an EC50 of 1–2 μM; astrocytes were not injured by these low hemin concentrations. Cell death was consistently reduced by at least 60% in knockout cultures. Exposure to hemin for 4 hours, a time point that preceded cell lysis, increased protein oxidation in wild-type cultures, as detected by staining of immunoblots for protein carbonyl groups. At 10 μM hemin, carbonylation was increased 2.3-fold compared with control sister cultures subjected to medium exchanges only; this effect was reduced by about two-thirds in knockout cultures. Cellular reactive oxygen species, detected by fluorescence intensity after dihydrorhodamine 123 (DHR staining, was markedly increased by hemin in wild-type cultures and was localized to neuronal cell bodies and processes. In contrast, DHR fluorescence intensity in knockout cultures did not differ from that of sham-washed controls. Neuronal death in wild-type cultures was almost completely prevented by the lipid-soluble iron chelator phenanthroline; deferoxamine had a weaker but significant effect. Conclusions These results suggest that HO-2 gene deletion protects neurons in mixed

  12. Implicación de las vías Nrf2/HO-1 y NADPH oxidasa en los modelos experimentales de artritis y osteoporosis postmenópausicas.

    Ibáñez Torres, Lidia

    2012-01-01

    Hemos puesto a punto y caracterizado el modelo de artritis postmenopáusica mediante ovariectomía (OVX) y artritis inducida por colágeno (CIA). Además, hemos estudiado la influencia de la vía HO-1 en este modelo animal mediante la administración de tin protoporfirina IX (SnPP) (inhibidor reversible de HO-1) y de CORM-3 (molécula liberadora de CO), así como la influencia de la vía NADPH oxidasa mediante el empleo de ratones con una modificación genética en Ncf1 de manera que no producen especie...

  13. Regulation of human heme oxygenase-1 gene expression under thermal stress.

    Okinaga, S; Takahashi, K; Takeda, K; Yoshizawa, M; Fujita, H; Sasaki, H; Shibahara, S

    1996-06-15

    Heme oxygenase-1 is an essential enzyme in heme catabolism, and its human gene promoter contains a putative heat shock element (HHO-HSE). This study was designed to analyze the regulation of human heme oxygenase-1 gene expression under thermal stress. The amounts of heme oxygenase-1 protein were not increased by heat shock (incubation at 42 degrees C) in human alveolar macrophages and in a human erythroblastic cell line, YN-1-0-A, whereas heat shock protein 70 (HSP70) was noticeably induced. However, heat shock factor does bind in vitro to HHO-HSE and the synthetic HHO-HSE by itself is sufficient to confer the increase in the transient expression of a reporter gene upon heat shock. The deletion of the sequence, located downstream from HHO-HSE, resulted in the activation of a reporter gene by heat shock. These results suggest that HHO-HSE is potentially functional but is repressed in vivo. Interestingly, heat shock abolished the remarkable increase in the levels of heme oxygenase-1 mRNA in YN-1-0-A cells treated with hemin or cadmium, in which HSP70 mRNA was noticeably induced. Furthermore, transient expression assays showed that heat shock inhibits the cadmium-mediated activation of the heme oxygenase-1 promoter, whereas the HSP70 gene promoter was activated upon heat shock. Such regulation of heme oxygenase-1 under thermal stress may be of physiologic significance in erythroid cells.

  14. Protective

    Wessam M. Abdel-Wahab

    2013-10-01

    Full Text Available Many active ingredients extracted from herbal and medicinal plants are extensively studied for their beneficial effects. Antioxidant activity and free radical scavenging properties of thymoquinone (TQ have been reported. The present study evaluated the possible protective effects of TQ against the toxicity and oxidative stress of sodium fluoride (NaF in the liver of rats. Rats were divided into four groups, the first group served as the control group and was administered distilled water whereas the NaF group received NaF orally at a dose of 10 mg/kg for 4 weeks, TQ group was administered TQ orally at a dose of 10 mg/kg for 5 weeks, and the NaF-TQ group was first given TQ for 1 week and was secondly administered 10 mg/kg/day NaF in association with 10 mg/kg TQ for 4 weeks. Rats intoxicated with NaF showed a significant increase in lipid peroxidation whereas the level of reduced glutathione (GSH and the activity of superoxide dismutase (SOD, catalase (CAT, glutathione S-transferase (GST and glutathione peroxidase (GPx were reduced in hepatic tissues. The proper functioning of the liver was also disrupted as indicated by alterations in the measured liver function indices and biochemical parameters. TQ supplementation counteracted the NaF-induced hepatotoxicity probably due to its strong antioxidant activity. In conclusion, the results obtained clearly indicated the role of oxidative stress in the induction of NaF toxicity and suggested hepatoprotective effects of TQ against the toxicity of fluoride compounds.

  15. Anti-Fibrotic Effect of Losartan, an Angiotensin II Receptor Blocker, Is Mediated through Inhibition of ER Stress via Up-Regulation of SIRT1, Followed by Induction of HO-1 and Thioredoxin

    Hyosang Kim

    2017-01-01

    Full Text Available Endoplasmic reticulum (ER stress is increasingly identified as modulator of fibrosis. Losartan, an angiotensin II receptor blocker, has been widely used as the first choice of treatment in chronic renal diseases. We postulated that anti-fibrotic effect of losartan is mediated through inhibition of ER stress via SIRT1 (silent mating type information regulation 2 homolog 1 hemeoxygenase-1 (HO-1/thioredoxin pathway. Renal tubular cells, tunicamycin (TM-induced ER stress, and unilateral ureteral obstruction (UUO mouse model were used. Expression of ER stress was assessed by Western blot analysis and immunohistochemical stain. ER stress was induced by chemical ER stress inducer, tunicamycin, and non-chemical inducers such as TGF-β, angiotensin II, high glucose, and albumin. Losartan suppressed the TM-induced ER stress, as shown by inhibition of TM-induced expression of GRP78 (glucose related protein 78 and p-eIF2α (phosphospecific-eukaryotic translation initiation factor-2α, through up-regulation of SIRT1 via HO-1 and thioredoxin. Losartan also suppressed the ER stress by non-chemical inducers. In both animal models, losartan reduced the tubular expression of GRP78, which were abolished by pretreatment with sirtinol (SIRT1 inhibitor. Sirtinol also blocked the inhibitory effect of losartan on the UUO-induced renal fibrosis. These findings provide new insights into renoprotective effects of losartan and suggest that SIRT1, HO-1, and thioredoxin may be potential pharmacological targets in kidney diseases under excessive ER stress condition.

  16. A newly synthesized macakurzin C-derivative attenuates acute and chronic skin inflammation: The Nrf2/heme oxygenase signaling as a potential target

    Akram, Muhammad [College of Pharmacy Institute of Pharmaceutical Science and Technology, Hanyang University, Ansan (Korea, Republic of); Shin, Iljin [College of Pharmacy and Research Institute of Pharmaceutical Science and Technology (RIPST), Ajou University, Suwon (Korea, Republic of); Kim, Kyeong-A; Noh, Dabi [College of Pharmacy Institute of Pharmaceutical Science and Technology, Hanyang University, Ansan (Korea, Republic of); Baek, Seung-Hoon; Chang, Sun-Young [College of Pharmacy and Research Institute of Pharmaceutical Science and Technology (RIPST), Ajou University, Suwon (Korea, Republic of); Kim, Hyoungsu, E-mail: hkimajou@ajou.ac.kr [College of Pharmacy and Research Institute of Pharmaceutical Science and Technology (RIPST), Ajou University, Suwon (Korea, Republic of); Bae, Ok-Nam, E-mail: onbae@hanyang.ac.kr [College of Pharmacy Institute of Pharmaceutical Science and Technology, Hanyang University, Ansan (Korea, Republic of)

    2016-09-15

    Impaired immune responses in skin play a pivotal role in the development and progression of chemical-associated inflammatory skin disorders. In this study, we synthesized new flavonoid derivatives from macakurzin C, and identified in vitro and in vivo efficacy of a potent anti-inflammatory flavonoid, Compound 14 (CPD 14), with its underlying mechanisms. In lipopolysaccharide (LPS)-stimulated murine macrophages and IFN-γ/TNF-α-stimulated human keratinocytes, CPD 14 significantly inhibited the release of inflammatory mediators including nitric oxide (NO), prostaglandins, and cytokines (IC{sub 50} for NO inhibition in macrophages: 4.61 μM). Attenuated NF-κB signaling and activated Nrf2/HO-1 pathway were responsible for the anti-inflammatory effects of CPD 14. The in vivo relevance was examined in phorbol 12-myristate 13-acetate (TPA)-induced acute skin inflammation and oxazolone-induced atopic dermatitis models. Topically applied CPD 14 significantly protected both irritation- and sensitization-associated skin inflammation by suppressing the expression of inflammatory mediators. In summary, we demonstrated that a newly synthesized flavonoid, CPD 14, has potent inhibitory effects on skin inflammation, suggesting it is a potential therapeutic candidate to treat skin disorders associated with excessive inflammation. - Highlights: • An anti-inflammatory flavonoid CPD 14 was newly synthesized from macakurzin C. • CPD 14 potently inhibited inflammatory reaction in keratinocytes and macrophages. • Dermal toxicity by irritation or sensitization in rats was protected by CPD 14. • Attenuated NF-κB and activated Nrf2/HO-1 were main mechanisms of CPD 14 action.

  17. Host heme oxygenase-1: Friend or foe in tackling pathogens?

    Singh, Nisha; Ahmad, Zeeshan; Baid, Navin; Kumar, Ashwani

    2018-05-14

    Infectious diseases are a major challenge in management of human health worldwide. Recent literature suggests that host immune system could be modulated to ameliorate the pathogenesis of infectious disease. Heme oxygenase (HMOX1) is a key regulator of cellular signaling and it could be modulated using pharmacological reagents. HMOX1 is a cytoprotective enzyme that degrades heme to generate carbon monoxide (CO), biliverdin, and molecular iron. CO and biliverdin (or bilirubin derived from it) can restrict the growth of a few pathogens. Both of these also induce antioxidant pathways and anti-inflammatory pathways. On the other hand, molecular iron can induce proinflammatory pathway besides making the cellular environment oxidative in nature. Since microbial infections often induce oxidative stress in host cells/tissues, role of HMOX1 has been analyzed in the pathogenesis of number of infections. In this review, we have described the role of HMOX1 in pathogenesis of bacterial infections caused by Mycobacterium species, Salmonella and in microbial sepsis. We have also provided a succinct overview of the role of HMOX1 in parasitic infections such as malaria and leishmaniasis. In the end, we have also elaborated the role of HMOX1 in viral infections such as AIDS, hepatitis, dengue, and influenza. © 2018 IUBMB Life, 2018. © 2018 International Union of Biochemistry and Molecular Biology.

  18. Loss of Kynurenine 3-Mono-oxygenase Causes Proteinuria.

    Korstanje, Ron; Deutsch, Konstantin; Bolanos-Palmieri, Patricia; Hanke, Nils; Schroder, Patricia; Staggs, Lynne; Bräsen, Jan H; Roberts, Ian S D; Sheehan, Susan; Savage, Holly; Haller, Hermann; Schiffer, Mario

    2016-11-01

    Changes in metabolite levels of the kynurenine pathway have been observed in patients with CKD, suggesting involvement of this pathway in disease pathogenesis. Our recent genetic analysis in the mouse identified the kynurenine 3-mono-oxygenase (KMO) gene (Kmo) as a candidate gene associated with albuminuria. This study investigated this association in more detail. We compared KMO abundance in the glomeruli of mice and humans under normal and diabetic conditions, observing a decrease in glomerular KMO expression with diabetes. Knockdown of kmo expression in zebrafish and genetic deletion of Kmo in mice each led to a proteinuria phenotype. We observed pronounced podocyte foot process effacement on long stretches of the filtration barrier in the zebrafish knockdown model and mild podocyte foot process effacement in the mouse model, whereas all other structures within the kidney remained unremarkable. These data establish the candidacy of KMO as a causal factor for changes in the kidney leading to proteinuria and indicate a functional role for KMO and metabolites of the tryptophan pathway in podocytes. Copyright © 2016 by the American Society of Nephrology.

  19. Heme oxygenase-1 and abscisic acid effects MAPK´s gene expression in soybean seeds

    Giacometti, R.; Santa Cruz, D.; Noriega, G.; Balestrasse, K.

    2012-01-01

    In soybean previous studies enabled the identification of MAPK3 and 6 whose activity is enhanced within the signaling pathway leading to defense reactions. In this study the effects of different compounds related to hemeoxygenase (HO-1) biosynthesis on mitogen-activated protein kinase (MAPK’s) genes expression in soybean seeds were tested. To this end, 20μM hemine, 22μM ZnPPIX, 0.5mM furidine or 100μM 8-bromoguanosine 3',5'-cyclic monophosphate (8Br) were added to pre-hydrated seeds for 5 days. MAPK’s genes expression was enhanced in seeds treated with hemine. This result indicates that heme catabolism could be involved in the signaling mediated by this cascade pathway. To confirm this hypothesis experiments were carried out in the precsence of ZnPPIX, a potent irreversible HO-1 inhibitor. In this case, no gene induction was observed. On the other hand, 8Br, a cGMP analog, induced HO-1 gene expression but did not modulate MAPK’s, indicating that this effect could not be mediated by cGMP. When the action of furidine, an abscisic acid inhibitor, was tested a diminution of HO-1 gene expression was observed. In this regard, MAPK’s showed a different response, being MAPK6 the only transcript that showed a diminished respect to controls, while MAPK3 mRNA as well as MAPKK1 was enhanced. These results were confirmed by western blotting and activity determinations. (authors)

  20. Discovery and industrial applications of lytic polysaccharide mono-oxygenases.

    Johansen, Katja S

    2016-02-01

    The recent discovery of copper-dependent lytic polysaccharide mono-oxygenases (LPMOs) has opened up a vast area of research covering several fields of application. The biotech company Novozymes A/S holds patents on the use of these enzymes for the conversion of steam-pre-treated plant residues such as straw to free sugars. These patents predate the correct classification of LPMOs and the striking synergistic effect of fungal LPMOs when combined with canonical cellulases was discovered when fractions of fungal secretomes were evaluated in industrially relevant enzyme performance assays. Today, LPMOs are a central component in the Cellic CTec enzyme products which are used in several large-scale plants for the industrial production of lignocellulosic ethanol. LPMOs are characterized by an N-terminal histidine residue which, together with an internal histidine and a tyrosine residue, co-ordinates a single copper atom in a so-called histidine brace. The mechanism by which oxygen binds to the reduced copper atom has been reported and the general mechanism of copper-oxygen-mediated activation of carbon is being investigated in the light of these discoveries. LPMOs are widespread in both the fungal and the bacterial kingdoms, although the range of action of these enzymes remains to be elucidated. However, based on the high abundance of LPMOs expressed by microbes involved in the decomposition of organic matter, the importance of LPMOs in the natural carbon-cycle is predicted to be significant. In addition, it has been suggested that LPMOs play a role in the pathology of infectious diseases such as cholera and to thus be relevant in the field of medicine. © 2016 Authors; published by Portland Press Limited.

  1. Identification of a novel oxygenase capable of regiospecific hydroxylation of D-limonene into (+)-trans-carveol

    Groeneveld, M.; van Beek, H. L.; Duetz, W. A.; Fraaije, M. W.

    2016-01-01

    By genome sequencing and analysis, we have been able to identify a gene cluster encoding a four component oxygenase which is able to oxidize D-limonene, L-limonene, cumene, and indole. Heterologous expression of the complete oxygenase system in Pseudomonas putida S12 allowed efficient and highly

  2. Characterisation of Anopheles gambiae heme oxygenase and metalloporphyrin feeding suggests a potential role in reproduction.

    Spencer, Christopher S; Yunta, Cristina; de Lima, Glauber Pacelli Gomes; Hemmings, Kay; Lian, Lu-Yun; Lycett, Gareth; Paine, Mark J I

    2018-05-03

    The mosquito Anopheles gambiae is the principal vector for malaria in sub-Saharan Africa. The ability of A. gambiae to transmit malaria is strictly related to blood feeding and digestion, which releases nutrients for oogenesis, as well as substantial amounts of highly toxic free heme. Heme degradation by heme oxygenase (HO) is a common protective mechanism, and a gene for HO exists in the An. gambiae genome HO (AgHO), although it has yet to be functionally examined. Here, we have cloned and expressed An. gambiae HO (AgHO) in E. coli. Purified recombinant AgHO bound hemin stoichiometrically to form a hemin-enzyme complex similar to other HOs, with a K D of 3.9 ± 0.6 μM; comparable to mammalian and bacterial HOs, but 7-fold lower than that of Drosophila melanogaster HO. AgHO also degraded hemin to biliverdin and released CO and iron in the presence of NADPH cytochrome P450 oxidoreductase (CPR). Optimal AgHO activity was observed at 27.5 °C and pH 7.5. To investigate effects of AgHO inhibition, adult female A. gambiae were fed heme analogues Sn- and Zn-protoporphyrins (SnPP and ZnPP), known to inhibit HO. These led to a dose dependent decrease in oviposition. Cu-protoporphyrin (CuPP), which does not inhibit HO had no effect. These results demonstrate that AgHO is a catalytically active HO and that it may play a key role in egg production in mosquitoes. It also presents a potential target for the development of compounds aimed at sterilising mosquitoes for vector control. Copyright © 2018 Elsevier Ltd. All rights reserved.

  3. The heme-heme oxygenase system: a molecular switch in wound healing.

    Wagener, F.A.D.T.G.; Beurden, H.E. van; Hoff, J.W. Von den; Adema, G.J.; Figdor, C.G.

    2003-01-01

    When cells are injured they release their contents, resulting in a local accumulation of free heme proteins and heme. Here, we investigated the involvement of heme and its degrading enzyme heme oxygenase (HO) in the inflammatory process during wound healing. We observed that heme directly

  4. Evidence-based prescription for cyclo-oxygenase-2 inhibitors in ...

    to the different metabolites generated by the cyclo-oxygenase-1. (COX-1) and ... have shown equivalent risk in non-NSAID users in developing congestive heart ... http://www.drgavinshang.co.za/wp-content/uploads/2012/11/NSAIDs-and-.

  5. RoxB Is a Novel Type of Rubber Oxygenase That Combines Properties of Rubber Oxygenase RoxA and Latex Clearing Protein (Lcp).

    Birke, Jakob; Röther, Wolf; Jendrossek, Dieter

    2017-07-15

    Only two types of rubber oxygenases, rubber oxygenase (RoxA) and latex clearing protein (Lcp), have been described so far. RoxA proteins (RoxAs) are c -type cytochromes of ≈70 kDa produced by Gram-negative rubber-degrading bacteria, and they cleave polyisoprene into 12-oxo-4,8-dimethyltrideca-4,8-diene-1-al (ODTD), a C 15 oligo-isoprenoid, as the major end product. Lcps are common among Gram-positive rubber degraders and do not share amino acid sequence similarities with RoxAs. Furthermore, Lcps have much smaller molecular masses (≈40 kDa), are b -type cytochromes, and cleave polyisoprene to a mixture of C 20 , C 25 , C 30 , and higher oligo-isoprenoids as end products. In this article, we purified a new type of rubber oxygenase, RoxB Xsp (RoxB of Xanthomonas sp. strain 35Y). RoxB Xsp is distantly related to RoxAs and resembles RoxAs with respect to molecular mass (70.3 kDa for mature protein) and cofactor content (2 c -type hemes). However, RoxB Xsp differs from all currently known RoxAs in having a distinctive product spectrum of C 20 , C 25 , C 30 , and higher oligo-isoprenoids that has been observed only for Lcps so far. Purified RoxB Xsp revealed the highest specific activity of 4.5 U/mg (at 23°C) of all currently known rubber oxygenases and exerts a synergistic effect on the efficiency of polyisoprene cleavage by RoxA Xsp RoxB homologs were identified in several other Gram-negative rubber-degrading species, pointing to a prominent function of RoxB for the biodegradation of rubber in Gram-negative bacteria. IMPORTANCE The enzymatic cleavage of rubber (polyisoprene) is of high environmental importance given that enormous amounts of rubber waste materials are permanently released (e.g., by abrasion of tires). Research from the last decade has discovered rubber oxygenase A, RoxA, and latex clearing protein (Lcp) as being responsible for the primary enzymatic attack on the hydrophobic and water-insoluble biopolymer poly( cis -1,4-isoprene) in Gram

  6. Heme Oxygenase-1 and breast cancer resistance protein protect against heme-induced toxicity

    Wagener, Frank A D T G; Dankers, Anita C A; van Summeren, Frank; Scharstuhl, Alwin; van den Heuvel, Jeroen J M W; Koenderink, Jan B; Pennings, Sebastiaan W C; Russel, Frans G M; Masereeuw, R.

    2013-01-01

    Heme is the functional group of diverse hemoproteins and crucial for many cellular processes. However, heme is increasingly recognized as a culprit for a wide variety of pathologies, including sepsis, malaria, and kidney failure. Excess of free heme can be detrimental to tissues by mediating

  7. Peracetylated hydroxytyrosol, a new hydroxytyrosol derivate, attenuates LPS-induced inflammatory response in murine peritoneal macrophages via regulation of non-canonical inflammasome, Nrf2/HO1 and JAK/STAT signaling pathways.

    Montoya, Tatiana; Aparicio-Soto, Marina; Castejón, María Luisa; Rosillo, María Ángeles; Sánchez-Hidalgo, Marina; Begines, Paloma; Fernández-Bolaños, José G; Alarcón-de-la-Lastra, Catalina

    2018-03-18

    The present study was designed to investigate the anti-inflammatory effects of a new derivative of hydroxytyrosol (HTy), peracetylated hydroxytyrosol (Per-HTy), compared with its parent, HTy, on lipopolysaccharide (LPS)-stimulated murine macrophages as well as potential signaling pathways involved. In particular, we attempted to characterize the role of the inflammasome underlying Per-HTy possible anti-inflammatory effects. Isolated murine peritoneal macrophages were treated with HTy or its derivative in the presence or absence of LPS (5 μg/ml) for 18 h. Cell viability was determined using sulforhodamine B (SRB) assay. Nitric oxide (NO) production was analyzed by Griess method. Production of pro-inflammatory cytokines was evaluated by enzyme-linked immunosorbent assay (ELISA) and inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX)-2, janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway (STAT3), haem oxigenase 1 (HO1), nuclear factor (erythroid-derived 2)-like 2 (Nrf2) expression and mitogen-activated protein kinases (MAPKs) activation was determined by Western blot. Per-HTy significantly reduced the levels of NO and pro-inflammatory cytokines as well as both COX-2 and iNOS expressions. Furthermore, Per-HTy treatment inhibited STAT3 and increased Nrf2 and HO1 protein levels in murine macrophages exposed to LPS. In addition, Per-HTy anti-inflammatory activity was related with an inhibition of non-canonical nucleotide binding domain (NOD)-like receptor (NLRP3) inflammasome pathways by decreasing pro-inflammatory interleukin (IL)-1β and IL-18 cytokine levels as consequence of regulation of cleaved caspase-11 enzyme. These results support that this new HTy derivative may offer a new promising nutraceutical therapeutic strategy in the management of inflammatory-related pathologies. Copyright © 2018. Published by Elsevier Inc.

  8. Novel bacterial sulfur oxygenase reductases from bioreactors treating gold-bearing concentrates

    Chen, Z-W; Liu, Y-Y; Wu, J-F

    2007-01-01

    The microbial community and sulfur oxygenase reductases of metagenomic DNA from bioreactors treating gold-bearing concentrates were studied by 16S rRNA library, real-time polymerase chain reaction (RT-PCR), conventional cultivation, and molecular cloning. Results indicated that major bacterial......) of bacteria and archaea were 4.59 x 10(9) and 6.68 x 10(5), respectively. Bacterial strains representing Acidithiobacillus, Leptospirillum, and Sulfobacillus were isolated from the bioreactors. To study sulfur oxidation in the reactors, pairs of new PCR primers were designed for the detection of sulfur...... oxygenase reductase (SOR) genes. Three sor-like genes, namely, sor (Fx), sor (SA), and sor (SB) were identified from metagenomic DNAs of the bioreactors. The sor (Fx) is an inactivated SOR gene and is identical to the pseudo-SOR gene of Ferroplasma acidarmanus. The sor (SA) and sor (SB) showed...

  9. Rieske non-heme iron-dependent oxygenases catalyse diverse reactions in natural product biosynthesis.

    Perry, Christopher; de Los Santos, Emmanuel L C; Alkhalaf, Lona M; Challis, Gregory L

    2018-04-13

    Covering: up to the end of 2017The roles played by Rieske non-heme iron-dependent oxygenases in natural product biosynthesis are reviewed, with particular focus on experimentally characterised examples. Enzymes belonging to this class are known to catalyse a range of transformations, including oxidative carbocyclisation, N-oxygenation, C-hydroxylation and C-C desaturation. Examples of such enzymes that have yet to be experimentally investigated are also briefly described and their likely functions are discussed.