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The Effect of Picroside II on Ischemia-Reperfusion Injury in Myocardial Streptozotocin Induced Diabetic Rats
E. Chiaradia et al. / Free Radical Biology and Medicine 96 (2016) S32–S69
enzyme system, increasing concentrations of myeloperoxidase or H2O2 resulted in an increased extent of modification of the CBF and HBF domains on fibronectin. The presence of SCN- in this system decreased damage relative to that observed with MPO/ H2O2/Cl-. Conclusions: Overall, our data support the hypothesis that fibronectin, and particularly its functional CBF and HBF domains are susceptible to modification by HOCl or HOSCN, with this resulting in modification to its structure, function and biological properties. These changes may contribute to endothelial dysfunction and rupture of atherosclerotic lesions. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.092 P-30
The potent inflammatory oxidant, peroxynitrite, modifies the extracellular matrix of human atherosclerotic lesions
S45
due to diminished F-actin stress fibre formation, were also observed when HCAEC were exposed to peroxynitrite-modified HCAEC-ECM and FN. HCAEC exposed to peroxynitrite-modified HCAEC-ECM resulted in an up-regulation of inflammation-associated genes including matrix metalloproteinases (MMP) 7 and 13 as well as a down-regulation of integrins associated with cell adhesion to laminins and fibronectin. In advanced human atherosclerotic lesions, co-localization of perlecan, laminins and fibronectin with 3-nitrotyrosine was detected, while peroxynitrite-modified laminins was also detected in proteins extracted from human atherosclerotic lesions. Conclusion: Peroxynitrite modifies ECM proteins particularly to laminins and FN by inducing structural and functional changes, which are important in endothelial cell function. These data suggest a mechanism through which peroxynitrite can modify the arterial ECM basement membrane in atherosclerotic lesions and subsequently lead to ECM remodelling. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.093 P-31
Christine Y. Chuang 1, Georg Degendorfer 2,3, Astrid Hammer 4, John M. Whitelock 5, Ernst Malle 6, Michael J. Davies 1,2,3 1
Department of Biomedical Science, Panum Institute, University of Copenhagen, Copenhagen, Denmark 2 The Heart Research Institute, Sydney, Australia 3 Faculty of Medicine, University of Sydney, Sydney, Australia 4 The Institute of Cell Biology, Histology and Embryology, Centre for Molecular Medicine, Medical University of Graz, Graz, Austria 5 Graduate School of Biomedical Engineering, The University of New South Wales, Sydney, Australia 6 The Institute of Molecular Biology and Biochemistry, Centre for Molecular Medicine, Medical University of Graz, Graz, Austria
The extracellular matrix (ECM) of the vascular basement membrane is critical to the functional and mechanical properties of arteries, with matrix proteins interacting with growth factors and enzymes to regulate endothelial and smooth muscle cell activity. These interactions are perturbed in atherosclerosis, where activated monocytes and macrophages generate oxidants that can contribute to endothelial cell dysfunction, alter ECM composition, attribute to smooth muscle cell proliferation, resulting in alteration of ECM remodelling, arterial stiffening and plaque formation. Aims: To investigate the effects of the inflammatory oxidant, peroxynitrite, on ECM proteins synthesised by human coronary artery endothelial cells (HCAEC) and purified human plasma-derived fibronectin (FN). Results: Perlecan, laminins and fibronectin were detected in the native HCAEC-ECM in vitro. A loss of antibody reactivity against perlecan protein core and cell binding epitopes of laminin and fibronectin was detected upon exposure to peroxynitrite in native HCAEC-ECM and FN, accompanied by an increased reactivity against the peroxynitritegenerated product, 3-nitrotyrosine. Mass spectrometry data indicated that some of these modifications on FN are within critical sites such as heparin and collagen binding domains as well as the cell binding domains. In addition, a decrease of HCAEC adhesion and spreading,
The effect of Picroside II on ischemia-reperfusion injury in myocardial streptozotocin induced diabetic rats Yücel Polat 1, Ali Doğan Dursun 2, Ayşegül Küçük 3, Abdullah Özer 1, Dilek Erer 1, Mustafa Arslan 4 1
Department of Cardiovascular Surgery, Faculty of Medicine, Gazi University, Ankara, Turkey 2 Department of Physiology, Faculty of Medicine, Ankara University, Ankara, Turkey 3 Department of Physiology, Faculty of Medicine, Dumlupınar University, Kütahya, Turkey 4 Anaesthesiology and Reanimation, Faculty of Medicine, Gazi University, Ankara, Turkey Aim: Diabetes mellitus is one of the leading causes of morbidity and mortality which is one of the world's major health problems. That chronic metabolic disorder is accompanied by an increase in oxidative stress. Ischemia-reperfusion (I/R) injury is a cascade of events initiated by tissue ischemia. The cellular damage produced by reperfusion leads to an active infiammatory response. Picroside II extracted from Picrorhiza scrophulariiflora has a wide range of pharmacological effects, including antioxidant, anticarcinogenic, and immune-modulating activities. The aim of the study was to investigate the effect of picroside II on myocardial I/R injury in rats with streptozotocin-induced diabetes. Materials and Methods: Thirty Wistar rats were grouped into five groups (n ¼ 6). In Groups control (C), diabetes control (DC), and diabetes picroside II (DP) neither coronary artery occlusion nor reperfusion were performed. In diabetes I/R group (DIR), a left anterior descending artery branch was occluded for 60 minutes. This was followed by 120 minutes of reperfusion to produce I/R. In diabetes I/R picroside II group (DIRP), picroside II was administrated via 10 mg/kg intraperitoneal route 30 minutes before ligating the left anterior descending artery. At the end of the study, myocardial tissues were harvested to determine the total oxidant status and total antioxidant status levels.
S46
E. Chiaradia et al. / Free Radical Biology and Medicine 96 (2016) S32–S69
Results: Total oxidant status levels were significantly higher in Group DIR, when compared with Groups C, DP, and DIRP (p ¼ .001, p ¼.019, and p ¼ .031, respectively). Total antioxidant status levels were significantly higher in Group DIR, when compared with groups C, DP, and DIRP (p ¼ .006, p ¼ .024, and p ¼ .007, respectively). Conclusion: These results indicate that administration of picroside II may have protective effects against I/R injury. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.094 P-32
Oxidative stress in brain during fetal-to-neonatal transition under an hypoxic atmosphere Maria Isabel Torres-Cuevas 1, J. Escobar 1, E. Cubells 1, R. Rodrigo 2, M. Vento 1,3 1
Neonatal Research Group, Health Research Institute La Fe, Valencia, Spain 2 Division of Genetics, University and Polytechnic Hospital La Fe, CIBERER, Valencia, Spain 3 Division of Neonatology, University and Polytechnic Hospital La Fe, Valencia, Spain
Background: Newborn asphyxia is one of the main complications during the perinatal period. The use of Oxygen for resuscitation has been broadly used in clinic for many years. However, it has been demonstrated that oxygen overexposure induces an oxidative stress (OS). Hypothesis: We speculate that delaying postnatal in the extrauterine oxygenation status would preserve reducing equivalents, enhance redox adaptation, and protect oxyregulator tissues (e.g.:brain). Objective: To assess oxidative stress and inflammation status, induced by Fetal to Neonatal Transition (FNT) under different FiO2 conditions, in brain of mice pups. Material and Methods: FiO2 in pregnant mice was reduced from 21% (room air) to 14% (Hx14) or not (Nx21) the night before of delivery (G18). 8 hours after birth both group were led to room air (Hx14/21 and Nx21/21 groups) or subjected to hyperoxia (FiO2 ¼100%) (Hx14/ 100 and Nx21/100 groups) and reset to 21% after 1 hour. At day 1 of life (P1) the pups were sacrificed and the brain tissues were snap frozen and kept at -80°C until the analysis. We set up a Mass Spectrometry (MS) method of four selected biomarkers: i) meta-tyrosine (m-tyr)/Phenylalanine (Phe), ii) orto-tyrosine (o-tyr)/Phe, iii) 3NO2tyrosine (3NO2-tyr)/para-tyrosine (tyr) and iv) 3Chloro-tyrosine (3Cltyr)/tyr ratios, to study cerebral OS and inflammation. Results: Hypoxia promotes significant changes in none of biomarkers. By contrast hyperoxia induces significant increases in m-tyr/phe and 3Cl-tyr/p-tyr. Remarkably, pre-conditioning hypoxia abrogates the rise of m-tyr/phe and 3Cl-tyr/p-tyr induced by hyperoxia as well as a decrease in the o-tyr/phe. Conclusions: Our results support the idea that FNT under hypoxic conditions could be protective to face a possible event of newborn resuscitation. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.095
P-33
Pharmacologic inhibition provides renoprotection nephropathy
of NADPH oxidase/Nox4 in contrast induced
Bo-Young Jeong 1, Hoi Young Lee 1, Doo-ri Choi 2, Seung-Yun Han 2, Moon Hyang Park 3, Kyung-Ho Lee 4, Dong-Il Kim 4, Won-Min Hwang 4, Sung-Ro Yun 4, Se-Hee Yoon 4 1
Pharmacology, Konyang University, Daejeon, Korea Anatomy, Konyang University, Daejeon, Korea 3 Pathology, Konyang University, Daejeon, Korea 4 Internal Medicine, Konyang University, Daejeon, Korea 2
Contrast media (CM) induced nephropathy (CIN) is an acute deterioration of renal function following administration of CM in the absence of any other known reason. CIN remains a leading cause of iatrogenic acute kidney injury (AKI), despite adherence to protocols of risk assessment and prevention strategies. CIN may occur, in part, as a result of intrarenal oxidative stress. NADPH oxidases are important sources of reactive oxygen species (ROS). Among various types of NADPH oxidases, Nox4 is expressed predominantly in rodent kidney. Aims: The aim of the present study was to assess the effect of Nox4 inhibition on the prevention of CIN. To determine the role of Nox4 in CIN, the current study used GKT 137831, a recently described pyrazolopyridinedione derivative that inhibits Nox4 and Nox1. Results: The expression of Nox4 in HK-2 cells significantly increased by iohexol exposure. Pretreatment of GKT137831(most specific Nox1/4 inhibitor) resulted in reduced production of ROS, down regulation of proinflammatory marker (p38), that are implicated in CIN, and increased cellular survival in iohexol exposed HK-2 cells. Silencing of the Nox4 gene replicated theses effects by down regulation of proinflammatory markers. In CIN mice model, pretreatment with GKT137831 resulted in an attenuated vacuolar degeneration, tubular epithelial cell shedding, cellular cast formation and tubular dilatation. Collectively, these results identify Nox4 as a key source of ROS responsible for kidney injury in CIN and provide proof of principle for an innovative small molecule approach to prevent contrast induced nephropathy. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.096 P-34
Paraoxonase activity in children with suspected mitochondrial disease Muscurel Corina 1, Elena Laura Gaman 1, Torac Elena 1, Tarta-Arsene Oana 1, Iosif Liviu 2, Gilca Marilena 1, Cristea Bogdan 1, Stroica Laura 1, Petran Madalina 1, Atanasiu Valeriu 1, Stoian Irina 1 1
Biochemistry Department, University of Medicine and Pharmacy "Carol Davila ", Bucharest, Romania 2 R&D Irist Labmed, Bucharest, Romania
enzyme system, increasing concentrations of myeloperoxidase or H2O2 resulted in an increased extent of modification of the CBF and HBF domains on fibronectin. The presence of SCN- in this system decreased damage relative to that observed with MPO/ H2O2/Cl-. Conclusions: Overall, our data support the hypothesis that fibronectin, and particularly its functional CBF and HBF domains are susceptible to modification by HOCl or HOSCN, with this resulting in modification to its structure, function and biological properties. These changes may contribute to endothelial dysfunction and rupture of atherosclerotic lesions. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.092 P-30
The potent inflammatory oxidant, peroxynitrite, modifies the extracellular matrix of human atherosclerotic lesions
S45
due to diminished F-actin stress fibre formation, were also observed when HCAEC were exposed to peroxynitrite-modified HCAEC-ECM and FN. HCAEC exposed to peroxynitrite-modified HCAEC-ECM resulted in an up-regulation of inflammation-associated genes including matrix metalloproteinases (MMP) 7 and 13 as well as a down-regulation of integrins associated with cell adhesion to laminins and fibronectin. In advanced human atherosclerotic lesions, co-localization of perlecan, laminins and fibronectin with 3-nitrotyrosine was detected, while peroxynitrite-modified laminins was also detected in proteins extracted from human atherosclerotic lesions. Conclusion: Peroxynitrite modifies ECM proteins particularly to laminins and FN by inducing structural and functional changes, which are important in endothelial cell function. These data suggest a mechanism through which peroxynitrite can modify the arterial ECM basement membrane in atherosclerotic lesions and subsequently lead to ECM remodelling. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.093 P-31
Christine Y. Chuang 1, Georg Degendorfer 2,3, Astrid Hammer 4, John M. Whitelock 5, Ernst Malle 6, Michael J. Davies 1,2,3 1
Department of Biomedical Science, Panum Institute, University of Copenhagen, Copenhagen, Denmark 2 The Heart Research Institute, Sydney, Australia 3 Faculty of Medicine, University of Sydney, Sydney, Australia 4 The Institute of Cell Biology, Histology and Embryology, Centre for Molecular Medicine, Medical University of Graz, Graz, Austria 5 Graduate School of Biomedical Engineering, The University of New South Wales, Sydney, Australia 6 The Institute of Molecular Biology and Biochemistry, Centre for Molecular Medicine, Medical University of Graz, Graz, Austria
The extracellular matrix (ECM) of the vascular basement membrane is critical to the functional and mechanical properties of arteries, with matrix proteins interacting with growth factors and enzymes to regulate endothelial and smooth muscle cell activity. These interactions are perturbed in atherosclerosis, where activated monocytes and macrophages generate oxidants that can contribute to endothelial cell dysfunction, alter ECM composition, attribute to smooth muscle cell proliferation, resulting in alteration of ECM remodelling, arterial stiffening and plaque formation. Aims: To investigate the effects of the inflammatory oxidant, peroxynitrite, on ECM proteins synthesised by human coronary artery endothelial cells (HCAEC) and purified human plasma-derived fibronectin (FN). Results: Perlecan, laminins and fibronectin were detected in the native HCAEC-ECM in vitro. A loss of antibody reactivity against perlecan protein core and cell binding epitopes of laminin and fibronectin was detected upon exposure to peroxynitrite in native HCAEC-ECM and FN, accompanied by an increased reactivity against the peroxynitritegenerated product, 3-nitrotyrosine. Mass spectrometry data indicated that some of these modifications on FN are within critical sites such as heparin and collagen binding domains as well as the cell binding domains. In addition, a decrease of HCAEC adhesion and spreading,
The effect of Picroside II on ischemia-reperfusion injury in myocardial streptozotocin induced diabetic rats Yücel Polat 1, Ali Doğan Dursun 2, Ayşegül Küçük 3, Abdullah Özer 1, Dilek Erer 1, Mustafa Arslan 4 1
Department of Cardiovascular Surgery, Faculty of Medicine, Gazi University, Ankara, Turkey 2 Department of Physiology, Faculty of Medicine, Ankara University, Ankara, Turkey 3 Department of Physiology, Faculty of Medicine, Dumlupınar University, Kütahya, Turkey 4 Anaesthesiology and Reanimation, Faculty of Medicine, Gazi University, Ankara, Turkey Aim: Diabetes mellitus is one of the leading causes of morbidity and mortality which is one of the world's major health problems. That chronic metabolic disorder is accompanied by an increase in oxidative stress. Ischemia-reperfusion (I/R) injury is a cascade of events initiated by tissue ischemia. The cellular damage produced by reperfusion leads to an active infiammatory response. Picroside II extracted from Picrorhiza scrophulariiflora has a wide range of pharmacological effects, including antioxidant, anticarcinogenic, and immune-modulating activities. The aim of the study was to investigate the effect of picroside II on myocardial I/R injury in rats with streptozotocin-induced diabetes. Materials and Methods: Thirty Wistar rats were grouped into five groups (n ¼ 6). In Groups control (C), diabetes control (DC), and diabetes picroside II (DP) neither coronary artery occlusion nor reperfusion were performed. In diabetes I/R group (DIR), a left anterior descending artery branch was occluded for 60 minutes. This was followed by 120 minutes of reperfusion to produce I/R. In diabetes I/R picroside II group (DIRP), picroside II was administrated via 10 mg/kg intraperitoneal route 30 minutes before ligating the left anterior descending artery. At the end of the study, myocardial tissues were harvested to determine the total oxidant status and total antioxidant status levels.
S46
E. Chiaradia et al. / Free Radical Biology and Medicine 96 (2016) S32–S69
Results: Total oxidant status levels were significantly higher in Group DIR, when compared with Groups C, DP, and DIRP (p ¼ .001, p ¼.019, and p ¼ .031, respectively). Total antioxidant status levels were significantly higher in Group DIR, when compared with groups C, DP, and DIRP (p ¼ .006, p ¼ .024, and p ¼ .007, respectively). Conclusion: These results indicate that administration of picroside II may have protective effects against I/R injury. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.094 P-32
Oxidative stress in brain during fetal-to-neonatal transition under an hypoxic atmosphere Maria Isabel Torres-Cuevas 1, J. Escobar 1, E. Cubells 1, R. Rodrigo 2, M. Vento 1,3 1
Neonatal Research Group, Health Research Institute La Fe, Valencia, Spain 2 Division of Genetics, University and Polytechnic Hospital La Fe, CIBERER, Valencia, Spain 3 Division of Neonatology, University and Polytechnic Hospital La Fe, Valencia, Spain
Background: Newborn asphyxia is one of the main complications during the perinatal period. The use of Oxygen for resuscitation has been broadly used in clinic for many years. However, it has been demonstrated that oxygen overexposure induces an oxidative stress (OS). Hypothesis: We speculate that delaying postnatal in the extrauterine oxygenation status would preserve reducing equivalents, enhance redox adaptation, and protect oxyregulator tissues (e.g.:brain). Objective: To assess oxidative stress and inflammation status, induced by Fetal to Neonatal Transition (FNT) under different FiO2 conditions, in brain of mice pups. Material and Methods: FiO2 in pregnant mice was reduced from 21% (room air) to 14% (Hx14) or not (Nx21) the night before of delivery (G18). 8 hours after birth both group were led to room air (Hx14/21 and Nx21/21 groups) or subjected to hyperoxia (FiO2 ¼100%) (Hx14/ 100 and Nx21/100 groups) and reset to 21% after 1 hour. At day 1 of life (P1) the pups were sacrificed and the brain tissues were snap frozen and kept at -80°C until the analysis. We set up a Mass Spectrometry (MS) method of four selected biomarkers: i) meta-tyrosine (m-tyr)/Phenylalanine (Phe), ii) orto-tyrosine (o-tyr)/Phe, iii) 3NO2tyrosine (3NO2-tyr)/para-tyrosine (tyr) and iv) 3Chloro-tyrosine (3Cltyr)/tyr ratios, to study cerebral OS and inflammation. Results: Hypoxia promotes significant changes in none of biomarkers. By contrast hyperoxia induces significant increases in m-tyr/phe and 3Cl-tyr/p-tyr. Remarkably, pre-conditioning hypoxia abrogates the rise of m-tyr/phe and 3Cl-tyr/p-tyr induced by hyperoxia as well as a decrease in the o-tyr/phe. Conclusions: Our results support the idea that FNT under hypoxic conditions could be protective to face a possible event of newborn resuscitation. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.095
P-33
Pharmacologic inhibition provides renoprotection nephropathy
of NADPH oxidase/Nox4 in contrast induced
Bo-Young Jeong 1, Hoi Young Lee 1, Doo-ri Choi 2, Seung-Yun Han 2, Moon Hyang Park 3, Kyung-Ho Lee 4, Dong-Il Kim 4, Won-Min Hwang 4, Sung-Ro Yun 4, Se-Hee Yoon 4 1
Pharmacology, Konyang University, Daejeon, Korea Anatomy, Konyang University, Daejeon, Korea 3 Pathology, Konyang University, Daejeon, Korea 4 Internal Medicine, Konyang University, Daejeon, Korea 2
Contrast media (CM) induced nephropathy (CIN) is an acute deterioration of renal function following administration of CM in the absence of any other known reason. CIN remains a leading cause of iatrogenic acute kidney injury (AKI), despite adherence to protocols of risk assessment and prevention strategies. CIN may occur, in part, as a result of intrarenal oxidative stress. NADPH oxidases are important sources of reactive oxygen species (ROS). Among various types of NADPH oxidases, Nox4 is expressed predominantly in rodent kidney. Aims: The aim of the present study was to assess the effect of Nox4 inhibition on the prevention of CIN. To determine the role of Nox4 in CIN, the current study used GKT 137831, a recently described pyrazolopyridinedione derivative that inhibits Nox4 and Nox1. Results: The expression of Nox4 in HK-2 cells significantly increased by iohexol exposure. Pretreatment of GKT137831(most specific Nox1/4 inhibitor) resulted in reduced production of ROS, down regulation of proinflammatory marker (p38), that are implicated in CIN, and increased cellular survival in iohexol exposed HK-2 cells. Silencing of the Nox4 gene replicated theses effects by down regulation of proinflammatory markers. In CIN mice model, pretreatment with GKT137831 resulted in an attenuated vacuolar degeneration, tubular epithelial cell shedding, cellular cast formation and tubular dilatation. Collectively, these results identify Nox4 as a key source of ROS responsible for kidney injury in CIN and provide proof of principle for an innovative small molecule approach to prevent contrast induced nephropathy. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.096 P-34
Paraoxonase activity in children with suspected mitochondrial disease Muscurel Corina 1, Elena Laura Gaman 1, Torac Elena 1, Tarta-Arsene Oana 1, Iosif Liviu 2, Gilca Marilena 1, Cristea Bogdan 1, Stroica Laura 1, Petran Madalina 1, Atanasiu Valeriu 1, Stoian Irina 1 1
Biochemistry Department, University of Medicine and Pharmacy "Carol Davila ", Bucharest, Romania 2 R&D Irist Labmed, Bucharest, Romania