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|Storage Conditions||Store all components at 2°C to 8°C. Store the protein standard (component 90450) at -25° to -15°C within 2 weeks of arrival.|
|Material Size||200 assays|
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Referencias bibliográficas | 60 Disponible | Ver todas las referencias
|Visión general referencias||Especie||Pub Med ID|
|Aerobic exercise training upregulates skeletal muscle calpain and ubiquitin-proteasome systems in healthy mice. |
Telma F Cunha,Jose B N Moreira,Nathalie A Paixão,Juliane C Campos,Alex W A Monteiro,Aline V N Bacurau,Carlos R Bueno,Julio C B Ferreira,Patricia C Brum
Journal of applied physiology (Bethesda, Md. : 1985) 112 2012
Aerobic exercise training (AET) is an important mechanical stimulus that modulates skeletal muscle protein turnover, leading to structural rearrangement. Since the ubiquitin-proteasome system (UPS) and calpain system are major proteolytic pathways involved in protein turnover, we aimed to investigate the effects of intensity-controlled AET on the skeletal muscle UPS and calpain system and their association to training-induced structural adaptations. Long-lasting effects of AET were studied in C57BL/6J mice after 2 or 8 wk of AET. Plantaris cross-sectional area (CSA) and capillarization were assessed by myosin ATPase staining. mRNA and protein expression levels of main components of the UPS and calpain system were evaluated in plantaris by real-time PCR and Western immunoblotting, respectively. No proteolytic system activation was observed after 2 wk of AET. Eight weeks of AET resulted in improved running capacity, plantaris capillarization, and CSA. Muscle RING finger-1 mRNA expression was increased in 8-wk-trained mice. Accordingly, elevated 26S proteasome activity was observed in the 8-wk-trained group, without accumulation of ubiquitinated or carbonylated proteins. In addition, calpain abundance was increased by 8 wk of AET, whereas no difference was observed in its endogenous inhibitor calpastatin. Taken together, our findings indicate that skeletal muscle enhancements, as evidenced by increased running capacity, plantaris capillarization, and CSA, occurred in spite of the upregulated UPS and calpain system, suggesting that overactivation of skeletal muscle proteolytic systems is not restricted to atrophying states. Our data provide evidence for the contribution of the UPS and calpain system to metabolic turnover of myofibrillar proteins and skeletal muscle adaptations to AET.
|Chronic paracetamol treatment influences indices of reactive oxygen species accumulation in the aging Fischer 344 X Brown Norway rat aorta. |
Kevin M Rice,Sarath Meduru,Sunil K Kakarla,Anjaiah Katta,Sriram P Mupparaju,Brent Kidd,Lynne J Goebel,Eric R Blough
Annals of clinical and laboratory science 42 2012
Previous reports have demonstrated that increased levels of reactive oxygen species (ROS) and alterations in cell signaling characterize aging in the Fischer 344 X Brown Norway (FBN) rat aorta. Other work has suggested that increases in ROS may be related to vascular wall thickening and the development of hypertension. Paracetamol (acetaminophen) is a potent antioxidant that has been found to diminish free radicals in ischemia-reperfusion studies. However, it remains unclear whether chronic paracetamol administration influences signaling or ROS accumulation in the aging aorta. FBN rats (27 months old; n=8) were subjected to 6 months of treatment with a therapeutic dose of paracetamol (30 mg/kg/day) and compared to age-matched untreated FBN rat controls (n=8). Compared to measurements in the aortae of 6-month old animals, tunica media thickness, tissue superoxide levels, and protein oxidation levels were 38 ± 7%, 92 ± 31%, and 7 ± 2% higher in the aortae of 33-month control animals (p ?0.05). Chronic paracetamol treatment decreased tunica media thickness and the amount of oxidized protein by 13 ± 4% and 30 ± 1%, respectively (p ?0.05). This finding of diminished aortic thickening was associated with increased phosphorylation (activation) of the mitogen activated protein kinases and diminished levels of the anti-apoptotic protein Bcl-2. Taken together, these data suggest that chronic paracetamol treatment may decrease the deleterious effects of aging in the FBN rat aorta.
|Intra-coronary administration of tacrolimus markedly attenuates infarct size and preserves heart function in porcine myocardial infarction. |
Sarah Chua,Steve Leu,Jiunn-Jye Sheu,Yu-Chun Lin,Li-Teh Chang,Ying-Hsien Kao,Chia-Hung Yen,Tzu-Hsien Tsai,Yung-Lung Chen,Hsueh-Wen Chang,Cheuk-Kwan Sun,Hon-Kan Yip
Journal of inflammation (London, England) 9 2012
|Novel protective mechanisms for S-adenosyl-l-methionine against acetaminophen hepatotoxicity: Improvement of key antioxidant enzymatic function. |
James Michael Brown,John G Ball,Michael Scott Wright,Stephanie Van Meter,Monica A Valentovic
Toxicology letters 212 2012
Acetaminophen (APAP) overdose leads to severe hepatotoxicity, increased oxidative stress and mitochondrial dysfunction. S-adenosyl-l-methionine (SAMe) protects against APAP toxicity at a mmol/kg equivalent dose to N-acetylcysteine (NAC). SAMe acts as a principle biological methyl donor and participates in polyamine synthesis which increase cell growth and has a role in mitochondrial protection. The purpose of the current study tested the hypothesis that SAMe protects against APAP toxicity by maintaining critical antioxidant enzymes and markers of oxidative stress. Male C57Bl/6 mice were treated with vehicle (Veh; water 15ml/kg, ip), SAMe (1.25mmol/kg, ip), APAP (250mg/kg, ip), and SAMe+APAP (SAMe given 1h following APAP). Liver was collected 2 and 4h following APAP administration; mitochondrial swelling as well as hepatic catalase, glutathione peroxidase (GPx), glutathione reductase, and both Mn- and Cu/Zn-superoxide dismutase (SOD) enzyme activity were evaluated. Mitochondrial protein carbonyl, 3-nitrotyrosine cytochrome c leakage were analyzed by Western blot. SAMe significantly increased SOD, GPx, and glutathione reductase activity at 4h following APAP overdose. SAMe greatly reduced markers of oxidative stress and cytochrome C leakage following APAP overdose. Our studies also demonstrate that a 1.25mmol/kg dose of SAMe does not inhibit CYP 2E1 enzyme activity. The current study identifies a plausible mechanism for the decreased oxidative stress observed when SAMe is given following APAP.
|Mononuclear iron enzymes are primary targets of hydrogen peroxide stress. |
Adil Anjem,James A Imlay
The Journal of biological chemistry 287 2012
This study tested whether nonredox metalloenzymes are commonly charged with iron in vivo and are primary targets of oxidative stress because of it. Indeed, three sample mononuclear enzymes, peptide deformylase, threonine dehydrogenase, and cytosine deaminase, were rapidly damaged by micromolar hydrogen peroxide in vitro and in live Escherichia coli. The first two enzymes use a cysteine residue to coordinate the catalytic metal atom; it was quantitatively oxidized by the radical generated by the Fenton reaction. Because oxidized cysteine can be repaired by cellular reductants, the effect was to avoid irreversible damage to other active-site residues. Nevertheless, protracted H(2)O(2) exposure gradually inactivated these enzymes, consistent with the overoxidation of the cysteine residue to sulfinic or sulfonic forms. During H(2)O(2) stress, E. coli defended all three proteins by inducing MntH, a manganese importer, and Dps, an iron-sequestration protein. These proteins appeared to collaborate in replacing the iron atom with nonoxidizable manganese. The implication is that mononuclear metalloproteins are common targets of H(2)O(2) and that both structural and metabolic arrangements exist to protect them.
|Exercise training prevents oxidative stress and ubiquitin-proteasome system overactivity and reverse skeletal muscle atrophy in heart failure. |
Telma F Cunha,Aline V N Bacurau,Jose B N Moreira,Nathalie A Paixão,Juliane C Campos,Julio C B Ferreira,Marcelo L Leal,Carlos E Negrão,Anselmo S Moriscot,Ulrik Wisløff,Patricia C Brum
PloS one 7 2012
Heart failure (HF) is known to lead to skeletal muscle atrophy and dysfunction. However, intracellular mechanisms underlying HF-induced myopathy are not fully understood. We hypothesized that HF would increase oxidative stress and ubiquitin-proteasome system (UPS) activation in skeletal muscle of sympathetic hyperactivity mouse model. We also tested the hypothesis that aerobic exercise training (AET) would reestablish UPS activation in mice and human HF.
|Impact of obesity control on circulating level of endothelial progenitor cells and angiogenesis in response to ischemic stimulation. |
Yung-Lung Chen,Chia-Lo Chang,Cheuk-Kwan Sun,Chiung-Jen Wu,Tzu-Hsien Tsai,Sheng-Ying Chung,Sarah Chua,Kuo-Ho Yeh,Steve Leu,Jiunn-Jye Sheu,Fan-Yen Lee,Chia-Hung Yen,Hon-Kan Yip
Journal of translational medicine 10 2012
|Neuroglial alterations in rats submitted to the okadaic acid-induced model of dementia. |
Costa AP, Tramontina AC, Biasibetti R, Batassini C, Lopes MW, Wartchow KM, Bernardi C, Tortorelli LS, Leal RB, Gonçalves CA
Behavioural brain research 226 420-7. Epub 2011 Oct 1. 2012
Several types of animal models have been developed to investigate Alzheimer\'s disease (AD). Okadaic acid (OA), a potent inhibitor of phosphatases 1 and 2A, induces characteristics that resemble AD-like pathology. Memory impairment induced by intra-hippocampal injection of OA has been reported, accompanied by remarkable neuropathological changes including hippocampal neurodegeneration, a paired helical filament-like phosphorylation of tau protein, and formation of β-amyloid containing plaque-like structures. Rats were submitted to bilateral intrahippocampal okadaic acid-injection (100ng) and, 12 days after the surgery, behavioral and biochemical tests were performed. Using this model, we evaluated spatial cognitive deficit and neuroglial alterations, particularly astroglial protein markers such as glial fibrillary acidic protein (GFAP) and S100B, metabolism of glutamate, oxidative parameters and alterations in MAPKs. Our results indicate significant hippocampal changes, including increased GFAP, protein oxidation, and phosphorylation of p38(MAPK); and decreases in glutathione content, transporter EAAT2/GLT-1, and glutamine synthetase activity as well as a decrease in cerebrospinal fluid S100B. No alterations were observed in glutamate uptake activity and S100B content. In conclusion, the OA-induced model of dementia caused spatial cognitive deficit and oxidative stress in this model and, for the first time to our knowledge, specific astroglial alterations. Findings contribute to understanding diseases accompanied by cognitive deficits and the neural damage induced by AO administration.Copyright © 2011 Elsevier B.V. All rights reserved.
|Autophagy proteins LC3B, ATG5 and ATG12 participate in quality control after mitochondrial damage and influence lifespan. |
Sören Mai,Britta Muster,Jürgen Bereiter-Hahn,Marina Jendrach
Autophagy 8 2012
Mitochondrial health is maintained by the quality control mechanisms of mitochondrial dynamics (fission and fusion) and mitophagy. Decline of these processes is thought to contribute to aging and neurodegenerative diseases. To investigate the role of mitochondrial quality control in aging on the cellular level, human umbilical vein endothelial cells (HUVEC) were subjected to mitochondria-targeted damage by combining staining of mitochondria and irradiation. This treatment induced a short boost of reactive oxygen species, which resulted in transient fragmentation of mitochondria followed by mitophagy, while mitochondrial dynamics were impaired. Furthermore, targeted mitochondrial damage upregulated autophagy factors LC3B, ATG5 and ATG12. Consequently these proteins were overexpressed in HUVEC as an in vitro aging model, which significantly enhanced the replicative life span up to 150% and the number of population doublings up to 200%, whereas overexpression of LAMP-1 did not alter the life span. Overexpression of LC3B, ATG5 and ATG12 resulted in an improved mitochondrial membrane potential, enhanced ATP production and generated anti-apoptotic effects, while ROS levels remained unchanged and the amount of oxidized proteins increased. Taken together, these data relate LC3B, ATG5 and ATG12 to mitochondrial quality control after oxidative damage, and to cellular longevity.
|Enhancement of proteasome function by PA28α overexpression protects against oxidative stress. |
Li J, Powell SR, Wang X
The FASEB journal : official publication of the Federation of American Societies for Experimental Biology 25 883-93. Epub 2010 Nov 23. 2011
The principal function of the proteasome is targeted degradation of intracellular proteins. Proteasome dysfunction has been observed in experimental cardiomyopathies and implicated in human congestive heart failure. Measures to enhance proteasome proteolytic function are currently lacking but would be beneficial in testing the pathogenic role of proteasome dysfunction and could have significant therapeutic potential. The association of proteasome activator 28 (PA28) with the 20S proteasome may play a role in antigen processing. It is unclear, however, whether the PA28 plays any important role outside of antigen presentation, although up-regulation of PA28 has been observed in certain types of cardiomyopathy. Here, we show that PA28α overexpression (PA28αOE) stabilized PA28β, increased 11S proteasomes, and enhanced the degradation of a previously validated proteasome surrogate substrate (GFPu) in cultured neonatal rat cardiomyocytes. PA28αOE significantly attenuated H(2)O(2)-induced increases in the protein carbonyls and markedly suppressed apoptosis in cultured cardiomyocytes under basal conditions or when stressed by H(2)O(2). We conclude that PA28αOE is sufficient to up-regulate 11S proteasomes, enhance proteasome-mediated removal of misfolded and oxidized proteins, and protect against oxidative stress in cardiomyocytes, providing a highly sought means to increase proteasomal degradation of abnormal cellular proteins.
|Oxidative stress-associated shape transformation and Membrane proteome remodeling in erythrocytes of end stage renal disease patients on hemodialysis. |
Antonelou MH, Kriebardis AG, Velentzas AD, Kokkalis AC, Georgakopoulou SC, Papassideri IS
Journal of proteomics 2011
This study was designed to evaluate the oxidative stress status of erythrocytes and its association with cellular ultrastructure and membrane proteome modifications in patients with end stage renal disease (ESRD) on hemodialysis (HD). For that purpose, we studied red blood cells\' (RBCs) modifications in twelve non-diabetic ESRD patients that were responsive in erythropoietin therapy. Intracellular ROS levels were measured by fluorometry, RBCs ultra-structure was examined by electron microscopy, while the membrane proteome by electrophoresis and immunoblotting. Compared to the healthy subjects, the uremic RBCs exhibited significantly increased ROS accumulation. Dialysis partially ameliorated the basal ROS levels but triggered cellular sensitivity to exogenous oxidative stimuli. Common membrane modifications involved loss, aggregation, fragmentation and carbonylation of critical components as well as over-expression of stress markers. HD significantly contributed to membrane proteome remodeling, especially for aquaporin-1, peroxiredoxin-2 and ubiquitinated proteins. The intracellular redox status and the closely associated membrane modifications seemed to be related to membrane instability, loss of surface area through vesiculation, echinocytosis and stomatocytosis. Our data evinced a network of interactions among the uremic toxins, the RBCs membrane composition and the cellular shape modifications in ESRD, which is developed around a core of oxidative provocations and cellular responses.Copyright © 2011 Elsevier B.V. All rights reserved.
|Na-K-Cl cotransporter-1 in the mechanism of cell swelling in cultured astrocytes after fluid percussion injury. |
Jayakumar AR, Panickar KS, Curtis KM, Tong XY, Moriyama M, Norenberg MD
Journal of neurochemistry 117 437-48. doi 2011
Brain edema and associated increased intracranial pressure are major consequences of traumatic brain injury (TBI). An important early component of the edema associated with TBI is astrocyte swelling (cytotoxic edema). Mechanisms for such swelling, however, are poorly understood. Ion channels/transporters/exchangers play a major role in cell volume regulation, and a disturbance in one or more of these systems may result in cell swelling. To examine potential mechanisms in TBI-mediated brain edema, we employed a fluid percussion model of in vitro barotrauma and examined the role of the ion transporter Na(+)-K(+)-2Cl(-)-cotransporter 1 (NKCC1) in trauma-induced astrocyte swelling as this transporter has been strongly implicated in the mechanism of cell swelling in various neurological conditions. Cultures exposed to trauma (3, 4, 5 atm pressure) caused a significant increase in NKCC1 activity (21%, 42%, 110%, respectively) at 3 h. At 5 atm pressure, trauma significantly increased NKCC1 activity at 1 h and it remained increased for up to 3 h. Trauma also increased the phosphorylation (activation) of NKCC1 at 1 and 3 h. Inhibition of MAPKs and oxidative/nitrosative stress diminished the trauma-induced NKCC1 phosphorylation as well as its activity. Bumetanide, an inhibitor of NKCC1, significantly reduced the trauma-induced astrocyte swelling (61%). Silencing NKCC1 with siRNA led to a reduction in trauma-induced NKCC1 activity as well as in cell swelling. These findings demonstrate the critical involvement of NKCC1 in the astrocyte swelling following in vitro trauma, and suggest that blocking NKCC1 activity may represent a useful therapeutic strategy for the cytotoxic brain edema associated with the early phase of TBI.© 2011 The Authors. Journal of Neurochemistry © 2011 International Society for Neurochemistry.
|Redox cycling and increased oxygen utilization contribute to diquat-induced oxidative stress and cytotoxicity in Chinese hamster ovary cells overexpressing NADPH-cytochrome P450 reductase. |
Fussell KC, Udasin RG, Gray JP, Mishin V, Smith PJ, Heck DE, Laskin JD
Free Radic Biol Med 50 874-82. Epub 2011 Jan 4. 2011
Diquat and paraquat are nonspecific defoliants that induce toxicity in many organs including the lung, liver, kidney, and brain. This toxicity is thought to be due to the generation of reactive oxygen species (ROS). An important pathway leading to ROS production by these compounds is redox cycling. In this study, diquat and paraquat redox cycling was characterized using human recombinant NADPH-cytochrome P450 reductase, rat liver microsomes, and Chinese hamster ovary (CHO) cells constructed to overexpress cytochrome P450 reductase (CHO-OR) and wild-type control cells (CHO-WT). In redox cycling assays with recombinant cytochrome P450 reductase and microsomes, diquat was 10-40 times more effective at generating ROS compared to paraquat (K(M)=1.0 and 44.2μM, respectively, for H(2)O(2) generation by diquat and paraquat using recombinant enzyme, and 15.1 and 178.5μM, respectively for microsomes). In contrast, at saturating concentrations, these compounds showed similar redox cycling activity (V(max)≈6.0nmol H(2)O(2)/min/mg protein) for recombinant enzyme and microsomes. Diquat and paraquat also redox cycle in CHO cells. Significantly more activity was evident in CHO-OR cells than in CHO-WT cells. Diquat redox cycling in CHO cells was associated with marked increases in protein carbonyl formation, a marker of protein oxidation, as well as cellular oxygen consumption, measured using oxygen microsensors; greater activity was detected in CHO-OR cells than in CHO-WT cells. These data demonstrate that ROS formation during diquat redox cycling can generate oxidative stress. Enhanced oxygen utilization during redox cycling may reduce intracellular oxygen available for metabolic reactions and contribute to toxicity.Copyright © 2010 Elsevier Inc. All rights reserved.
|Pseudomonas aeruginosa is associated with increased lung cytokines and asymmetric dimethylarginine compared with methicillin-resistant Staphylococcus aureus. |
Sousse LE, Jonkam CC, Traber DL, Hawkins HK, Rehberg SW, Traber LD, Herndon DN, Enkhbaatar P
Shock (Augusta, Ga) 36 466-70. 2011
The objective of the study was to investigate pulmonary responses to Pseudomonas aeruginosa and methicillin-resistant Staphylococcus aureus (MRSA) using ovine and mice models of sepsis with emphasis on lung cytokine expression, asymmetric dimethylarginine (ADMA) concentration, and the arginase pathway. Sheep were instilled with either MRSA, P. aeruginosa, or saline under deep anesthesia; mechanically ventilated; resuscitated with fluid; and killed after 24 h. Mice were instilled with either MRSA, P. aeruginosa, or saline under deep anesthesia and killed after 8 h. Lungs were assessed for ADMA concentration, arginase activity, oxidative stress, and cytokine expression, and plasma was assessed for nitrate/nitrite concentrations. The severity of lung injury was more pronounced in P. aeruginosa sepsis compared with MRSA. The significant changes in sheep lung function after P. aeruginosa sepsis were associated with significantly increased ADMA concentrations and arginase activity compared with MRSA. However, the plasma concentration of nitrites and nitrates were significantly increased in MRSA sepsis compared with P. aeruginosa sepsis. In the mice model, P. aeruginosa significantly increased lung cytokine expression (IL-1 and IL-13), protein oxidation, and arginase activity compared with MRSA. Our data suggest that the greater expression of cytokines and ADMA concentrations may be responsible for severity of acute lung injury in P. aeruginosa sepsis. The lack of arginase activity may explain the greater nitric oxide production in MRSA sepsis.
|Apolipoprotein JClusterin in Human Erythrocytes Is Involved in the Molecular Process of Defected Material Disposal during Vesiculation. |
Antonelou MH, Kriebardis AG, Stamoulis KE, Trougakos IP, Papassideri IS.
PloS one 6 e26033 2011
We have showed that secretory Apolipoprotein J/Clusterin (sCLU) is down-regulated in senescent, stressed or diseased red blood cells (RBCs). It was hypothesized that sCLU loss relates to RBCs vesiculation, a mechanism that removes erythrocyte membrane patches containing defective or potentially harmful components.
|Decreased sensitivity associated with an altered formulation of a commercially available kit for detection of protein carbonyls. |
Wang P, Powell SR
Free radical biology & medicine 2010
Carbonylation is a commonly studied form of oxidative modification to proteins which can be conveniently detected using commercially available kits. The most common of these kits is the Oxyblot(TM) Protein Oxidation Detection Kit (Chemicon/Millipore). Over the past year we have observed severely diminished sensitivity of these kits which was shown to be a result of a change in the formulation of one of the components supplied in the kit. This component, the 10X 2,4-dinitrophenylhydrazine derivatization solution, which had previously been dissolved in 100% trifluoroacetic acid (TFA), was now dissolved in 2N hydrochloric acid, which according to our results is not acid enough. Further, we observed that upon storage even DNPH dissolved in TFA is subject to degradation. Based on these studies, we make recomendations that should improve the sensitivity and reproducibilty of this assay. Copyright Â© 2010 Elsevier Inc. All rights reserved.
|Cytoskeletal and cell adhesion defects in wounded and Pax6+/- corneal epithelia. |
Jingxing Ou,Christina Lowes,J Martin Collinson
Investigative ophthalmology & visual science 51 2010
PAX6 heterozygosity (PAX6(+/-)) causes aniridia and aniridia-related keratopathy (ARK) in humans, but the pathway from gene dosage deficiency to clinical disease has not been fully characterized. Recently, the authors suggested a model of a chronic wound state exacerbated by oxidative stress, showed the barrier function of Pax6(+/-) corneas is compromised and suggested Pax6(+/-) corneas show the molecular signature of a perpetual wound-healing state.
|Bone Marrow Cells Reduce Fibrogenesis and Enhance Regeneration in Fibrotic Rat Liver. |
Sun CK, Chen CH, Kao YH, Yuen CM, Sheu JJ, Lee FY, Chen YT, Kung CT, Yip HK
J Surg Res 2010
BACKGROUND: This study aimed at investigating the cellular and molecular impacts of bone marrow-derived mononuclear cells (BMCs) on regeneration of fibrotic liver in rats.
|Mitochondrial complex I activity and oxidative damage to mitochondrial proteins in the prefrontal cortex of patients with bipolar disorder. |
Andreazza AC, Shao L, Wang JF, Young LT
Arch Gen Psychiatry 67 360-8. 2010
CONTEXT: Accumulating evidence suggests that mitochondrial dysfunction and oxidative stress contribute to the pathogenesis of bipolar disorder and schizophrenia. It remains unclear whether mitochondrial dysfunction, specifically complex I impairment, is associated with increased oxidative damage and, if so, whether this relationship is specific to bipolar disorder. OBJECTIVE: To evaluate whether decreased levels of the electron transport chain complex I subunit NDUFS7 are associated with complex I activity and increased oxidative damage to mitochondrial proteins in the prefrontal cortex of patients with bipolar disorder, schizophrenia, or major depressive disorder. DESIGN: Postmortem prefrontal cortex from patients and controls were assessed using immunoblotting, spectrophotometric, competitive enzyme immunoassay to identify group differences in expression and activity of complex I, and in oxidative damage in mitochondria. SETTING: University of British Columbia, Vancouver, Canada. Patients Forty-five patients with a psychiatric disorder (15 each with bipolar disorder, schizophrenia, and major depressive disorder) and 15 nonpsychiatric control subjects were studied. MAIN OUTCOME MEASURES: Oxidative damage to proteins and mitochondrial complex I activity. RESULTS: Levels of NDUFS7 and complex I activity were decreased significantly in patients with bipolar disorder but were unchanged in those with depression and schizophrenia compared with controls. Protein oxidation, as measured by protein carbonylation, was increased significantly in the bipolar group but not in the depressed or schizophrenic groups compared with controls. We observed increased levels of 3-nitrotyrosine in the bipolar disorder and schizophrenia groups. CONCLUSIONS: Impairment of complex I may be associated with increased protein oxidation and nitration in the prefrontal cortex of patients with bipolar disorder. Therefore, complex I activity and mitochondrial dysfunction may be potential therapeutic targets for bipolar disorder.
|Proteomic analysis in cerebrospinal fluid of patients with atypical nonketotic hyperglycinemia and pulmonary hypertension - A pilot study. |
Carlos E Rodríguez,José A Arranz,Nuria Colomé,Joan J Bech-Serra,Francesc Canals,Mireia Del Toro,Encarnació Riudor
Proteomics. Clinical applications 3 2009
A variant phenotype of nonketotic hyperglycinemia has been described by our group associated with pulmonary hypertension. The aim of this study is to investigate the cerebrospinal fluid proteomes to get an insight into this neurodegenerative process producing leukoencephalopathy with white matter spongiform degeneration. DIGE and MALDI-TOF-TOF analyses were performed to carry out the proteomic study of four patients against three normal controls and one additional control of a classical nonketotic hyperglycinemia. The differential proteomic analysis showed a displacement of some series of spots toward the acidic side. The shifted proteins showed a high degree of carbonylation and increased methionine sulfoxidation was found in cystatin C and in vitamin-D-binding protein. These findings in addition to the increase of serum malondialdehyde concentration provide evidence of an oxidative stress in the patients under study, which is probably systemic rather than mainly confined to the CNS. The similarities of our findings with those found in other neurodegenerative diseases suggest that oxidative damage is commonly involved in these pathologies. DIGE technology improves the 2-D PAGE differential analysis and it is suitable in proteomic studies with a small number of cases.
|A transient antioxidant stress response accompanies the onset of disuse atrophy in human skeletal muscle. |
Dalla Libera, Luciano, et al.
J. Appl. Physiol., 107: 549-57 (2009) 2009
It is presently unknown whether oxidative stress increases in disused skeletal muscle in humans. Markers of oxidative stress were investigated in biopsies from the vastus lateralis muscle, collected from healthy subjects before [time 0 (T0)], after 1 wk (T8), and after 5 wk (T35) of bed rest. An 18% decrease in fiber cross-sectional area was detected in T35 biopsies (P<0.05). Carbonylation of muscle proteins significantly increased about twofold at T35 (P<0.02) and correlated positively with the decrease in fiber cross-sectional area (P=0.04). Conversely, T8 biopsies showed a significant increase in protein levels of heme oxygenase-1 and glucose-regulated protein-75 (Grp75)/mitochondrial heat shock protein-70, two stress proteins involved in the antioxidant defense (P<0.05). Heme oxygenase-1 increase, which involved a larger proportion of slow fibers compared with T0, appeared blunted in T35 biopsies. Grp75 protein level increased threefold in T8 biopsies and localized especially in slow fibers (P<0.025), to decrease significantly in T35 biopsies (P<0.05). Percent change in Grp75 levels positively correlated with fiber cross-sectional area (P=0.01). Parallel investigations on rat soleus muscles, performed after 1-15 days of hindlimb suspension, showed that Grp75 protein levels significantly increased after 24 h of unloading (P = 0.02), i.e., before statistically significant evidence of muscle atrophy, to decrease thereafter in relation to the degree of muscle atrophy (P=0.03). Therefore, in humans as in rodents, disuse muscle atrophy is characterized by increased protein carbonylation and by the blunting of the antioxidant stress response evoked by disuse.
|Radiation resistance of Deinococcus radiodurans R1 with respect to growth phase. |
Sukhi SS, Shashidhar R, Kumar SA, Bandekar JR
FEMS Microbiol Lett 297 49-53. Epub 2009 Jun 1. 2009
Deinococcus species exhibit an extraordinary ability to withstand ionizing radiation (IR). Most of the studies on radiation resistance have been carried out with exponential phase cells. The studies on radiation resistance of Deinococcus radiodurans R1 with respect to different phases of growth showed that late stationary phase cells of D. radiodurans R1 were fourfold more sensitive to IR and heat as compared with exponential or early stationary phase cells. The increased sensitivity of D. radiodurans R1 to IR in the late stationary phase was not due to a decrease in the intracellular Mn/Fe ratio or an increase in the level of oxidative protein damage. The resistance to IR was restored when late stationary phase cells were incubated for 15 min in fresh medium before irradiation, indicating that replenishment of exhausted nutrients restored the metabolic capability of the cells to repair DNA damage. These observations suggest that stress tolerance mechanisms in D. radiodurans R1 differ from established paradigms.
|Decreased catalase expression and increased susceptibility to oxidative stress in primary cultured corneal fibroblasts from patients with granular corneal dystrophy type II. |
Seung-il Choi, Tae-im Kim, Kyu Seo Kim, Bong-Yoon Kim, So-yeon Ahn, Hyun-ju Cho, Hyung Keun Lee, Hyun-Soo Cho, Eung Kweon Kim
The American journal of pathology 175 248-61 2009
Granular corneal dystrophy type II (GCD II) is an autosomal dominant disorder characterized by age-dependent progressive accumulation of transforming growth factor-beta-induced protein (TGFBIp) deposits in the corneal stroma. Several studies have suggested that corneal fibroblasts may decline with age in response to oxidative stress. To investigate whether oxidative stress is involved in the pathogenesis of GCD II, we assayed antioxidant enzymes, oxidative damage, and susceptibility to reactive oxygen species-induced cell death in primary cultured corneal fibroblasts (PCFs) from GCD II patients and healthy subjects. We found elevated protein levels of Mn-superoxide dismutase, Cu/Zn-superoxide dismutase, glutathione peroxidase, and glutathione reductase, as well as increased CAT mRNA and decreased catalase protein in GCD II PCFs. Furthermore, catalase is down-regulated in normal PCFs transfected with transforming growth factor-beta-induced gene-h3. We also observed an increase in not only intracellular reactive oxygen species and H(2)O(2) levels, but also malondialdehyde, 4-hydroxynonenal, and protein carbonyls levels in GCD II PCFs. Greater immunoreactivity for malondialdehyde was observed in the corneal tissue of GCD II patients. In addition, we observed a decrease in Bcl-2 and Bcl-xL levels and an increase in Bax and Bok levels in GCD II PCFs. Finally, GCD II PCFs are more susceptible to H(2)O(2)-induced cell death. Together, these results suggest that oxidative damage induced by decreased catalase is involved in GCD II pathogenesis, and antioxidant agents represent a possible treatment strategy.Artículo Texto completo
|Acetaminophen prevents aging-associated hyperglycemia in aged rats: effect of aging-associated hyperactivation of p38-MAPK and ERK1/2. |
Wu, Miaozong, et al.
Diabetes Metab. Res. Rev., 25: 279-86 (2009) 2009
BACKGROUND: Aging-related hyperglycemia is associated with increased oxidative stress and diminished muscle glucose transporter-4 (Glut4) that may be regulated, at least in part, by the mitogen-activated protein kinases (MAPK). METHODS: To test the possibility that aging-related hyperglycemia can be prevented by pharmacological manipulation of MAPK hyperactivation, aged (27-month old) Fischer 344/NNiaHSD x Brown Norway/BiNia F1 (F344BN) rats were administered acetaminophen (30 mg/kg body weight/day) for 6 months in drinking water. RESULTS: Hepatic histopathology, serum aspartate aminotransferase and alanine aminotransferase analyses suggested that chronic acetaminophen did not cause hepatotoxicity. Compared with adult (6-month) and aged (27-month) rats, very aged rats (33-month) had higher levels of blood glucose, phosphorylation of soleus p38-MAPK and extracellular-regulated kinase 1/2 (ERK1/2), superoxide and oxidatively modified proteins (p<0.05), and these changes were associated with decreased soleus Glut4 protein abundance (p<0.05). Chronic acetaminophen treatment attenuated age-associated increase in blood glucose by 61.3% (p<0.05) and increased soleus Glut4 protein by 157.2% (p<0.05). These changes were accompanied by diminished superoxide levels, decrease in oxidatively modified proteins (-60.8%; p<0.05) and reduced p38-MAPK and ERK1/2 hyperactivation (-50.4% and -35.4%, respectively; p<0.05). CONCLUSIONS: These results suggest that acetaminophen may be useful for the treatment of age-associated hyperglycemia.
|Decline in NRF2-regulated antioxidants in chronic obstructive pulmonary disease lungs due to loss of its positive regulator, DJ-1. |
Deepti Malhotra, Rajesh Thimmulappa, Ana Navas-Acien, Andrew Sandford, Mark Elliott, Anju Singh, Linan Chen, Xiaoxi Zhuang, James Hogg, Peter Pare, Rubin M Tuder, Shyam Biswal
American journal of respiratory and critical care medicine 178 592-604 2008
RATIONALE: Oxidative stress is a key contributor in chronic obstructive pulmonary disease (COPD) pathogenesis caused by cigarette smoking. NRF2, a redox-sensitive transcription factor, dissociates from its inhibitor, KEAP1, to induce antioxidant expression that inhibits oxidative stress. OBJECTIVES: To determine the link between severity of COPD, oxidative stress, and NRF2-dependent antioxidant levels in the peripheral lung tissue of patients with COPD. METHODS: We assessed the expression of NRF2, NRF2-dependent antioxidants, regulators of NRF2 activity, and oxidative damage in non-COPD (smokers and former smokers) and smoker COPD lungs (mild and advanced). Cigarette smoke-exposed human lung epithelial cells (Beas2B) and mice were used to understand the mechanisms. MEASUREMENTS AND MAIN RESULTS: When compared with non-COPD lungs, the COPD patient lungs showed (1) marked decline in NRF2-dependent antioxidants and glutathione levels, (2) increased oxidative stress markers, (3) significant decrease in NRF2 protein with no change in NRF2 mRNA levels, and (4) similar KEAP1 but significantly decreased DJ-1 levels (a protein that stabilizes NRF2 protein by impairing KEAP1-dependent proteasomal degradation of NRF2). Exposure of Bea2B cells to cigarette smoke caused oxidative modification and enhanced proteasomal degradation of DJ-1 protein. Disruption of DJ-1 in mouse lungs, mouse embryonic fibroblasts, and Beas2B cells lowered NRF2 protein stability and impaired antioxidant induction in response to cigarette smoke. Interestingly, targeting KEAP1 by siRNA or the small-molecule activator sulforaphane restored induction of NRF2-dependent antioxidants in DJ-1-disrupted cells in response to cigarette smoke. CONCLUSIONS: NRF2-dependent antioxidants and DJ-1 expression was negatively associated with severity of COPD. Therapy directed toward enhancing NRF2-regulated antioxidants may be a novel strategy for attenuating the effects of oxidative stress in the pathogenesis of COPD.Artículo Texto completo
|Expression of the counter-regulatory peptide intermedin is augmented in the presence of oxidative stress in hypertrophied cardiomyocytes. |
Bell, David, et al.
Cell. Physiol. Biochem., 21: 409-20 (2008) 2008
|Chronic wound state exacerbated by oxidative stress in Pax6+/- aniridia-related keratopathy. |
J Ou, P Walczysko, R Kucerova, A M Rajnicek, C D McCaig, M Zhao, J M Collinson
The Journal of pathology 215 421-30 2008
Heterozygosity for the transcription factor PAX6 causes eye disease in humans, characterized by corneal opacity. The molecular aetiology of such disease was investigated using a Pax6+/- mouse model. We found that the barrier function of uninjured Pax6+/- corneas was compromised and that Ca2+-PKC/PLC-ERK/p38 signalling pathways were abnormally activated, mimicking a 'wounded' epithelial state. Using proteomic analysis and direct assay for oxidized proteins, Pax6+/- corneas were found to be susceptible to oxidative stress and they exhibited a wound-healing delay which could be rescued by providing reducing agents such as glutathione. Pax6 protein was oxidized and excluded from the nucleus of stressed corneal epithelial cells, with concomitant loss of corneal epithelial markers and expression of fibroblast/myofibroblast markers. We suggest a chronic wound model for Pax6-related corneal diseases, in which oxidative stress underlies a positive feedback mechanism by depleting nuclear Pax6, delaying wound healing, and activating cell signalling pathways that lead to metaplasia of the corneal epithelium. The study mechanistically links a relatively minor dosage deficiency of a transcription factor with potentially catastrophic degenerative corneal disease.
|Manganese effects on haematopoietic cells and circulating coelomocytes of Asterias rubens (Linnaeus). |
Carolina Oweson, Helen Sköld, Annalisa Pinsino, Valeria Matranga, Bodil Hernroth
Aquatic toxicology (Amsterdam, Netherlands) 89 75-81 2008
Manganese (Mn) is a naturally abundant metal in marine sediments where it mainly occurs as MnO(2). During hypoxic conditions it is converted into a bioavailable state, Mn(2+), and can reach levels that previously have shown effects on immune competent cells of the crustacean, Nephrops norvegicus. Here we investigated if Mn also affects circulating coelomocytes and their renewal in the common sea star, Asterias rubens, when exposed to concentrations of Mn that can be found in nature. When the sea stars were exposed to Mn it accumulated in the coelomic fluid and the number of circulating coelomocytes, in contrast to what was recorded in Nephrops, increased significantly. By using the substitute nucleotide, 5-bromo-2'-deoxyuridine, BrdU, for tracing cell division and by recording mitotic index by nuclei staining, we found that Mn induced proliferation of cells from a putative haematopoietic tissue, the coelomic epithelium. In addition, the haematopoietic tissue and coelomocytes showed stress response in terms of changes in HSP70 levels and protein carbonyls, as judged by immunohistochemistry and Western blotting. Measurement of dehydrogenase activity, using MTS/PMS, revealed that Mn showed cytotoxic properties. We also found that the phagocytotic capacity of coelomocytes was significantly inhibited by Mn. It was concluded that the exposure of A. rubens to Mn induced renewal of coelomocytes and impaired their immune response.
|Endurance training without weight loss lowers systemic, but not muscle, oxidative stress with no effect on inflammation in lean and obese women. |
Michaela C Devries, Mazen J Hamadeh, Alexander W Glover, Sandeep Raha, Imtiaz A Samjoo, Mark A Tarnopolsky
Free radical biology medicine 45 503-11 2008
Obesity is associated with oxidative stress. Endurance training (ET) in healthy individuals increases antioxidant enzyme activity and decreases oxidative stress, whereas its effects on oxidative status in obese humans have yet to be determined. We investigated the effects of obesity and ET on markers of oxidative stress, antioxidant defense, and inflammation. Obese (n=12) and lean (n=12) women underwent 12 weeks of ET with blood, 24-h urine, and muscle biopsies collected prior to and following training for determination of oxidative stress (urinary 8-hydroxy-2-deoxyguanosine and 8-isoprostanes, muscle protein carbonyls, and 4-hydroxynonenal), antioxidant enzyme protein content (muscle CuZnSOD, MnSOD, and catalase), and inflammation (C-reactive protein, leptin, adiponectin, interleukin-6). Obese women had elevated urinary 8-hydroxy-2-deoxyguanosine (P=0.03), muscle protein carbonyls (P=0.03), and 4-hydroxynonenal (P0.001); serum C-reactive protein (P=0.01); and plasma leptin (P=0.0001) and interleukin-6 (P=0.03). ET decreased urinary 8-hydroxy-2-deoxyguanosine (P=0.006) and 8-isoprostanes (P=0.02) in all subjects and CuZnSOD protein content (P=0.04) in obese women, in the absence of changes in body weight or composition. ET without weight loss decreases systemic oxidative stress, but not markers of inflammation, in obese women.
|Protein oxidation implicated as the primary determinant of bacterial radioresistance. |
Michael J Daly, Elena K Gaidamakova, Vera Y Matrosova, Alexander Vasilenko, Min Zhai, Richard D Leapman, Barry Lai, Bruce Ravel, Shu-Mei W Li, Kenneth M Kemner, James K Fredrickson
PLoS biology 5 e92 2007
In the hierarchy of cellular targets damaged by ionizing radiation (IR), classical models of radiation toxicity place DNA at the top. Yet, many prokaryotes are killed by doses of IR that cause little DNA damage. Here we have probed the nature of Mn-facilitated IR resistance in Deinococcus radiodurans, which together with other extremely IR-resistant bacteria have high intracellular Mn/Fe concentration ratios compared to IR-sensitive bacteria. For in vitro and in vivo irradiation, we demonstrate a mechanistic link between Mn(II) ions and protection of proteins from oxidative modifications that introduce carbonyl groups. Conditions that inhibited Mn accumulation or Mn redox cycling rendered D. radiodurans radiation sensitive and highly susceptible to protein oxidation. X-ray fluorescence microprobe analysis showed that Mn is globally distributed in D. radiodurans, but Fe is sequestered in a region between dividing cells. For a group of phylogenetically diverse IR-resistant and IR-sensitive wild-type bacteria, our findings support the idea that the degree of resistance is determined by the level of oxidative protein damage caused during irradiation. We present the case that protein, rather than DNA, is the principal target of the biological action of IR in sensitive bacteria, and extreme resistance in Mn-accumulating bacteria is based on protein protection.Artículo Texto completo
|The effect of training on the expression of mitochondrial biogenesis- and apoptosis-related proteins in skeletal muscle of patients with mtDNA defects. |
Peter J Adhihetty, Tanja Taivassalo, Ronald G Haller, Donald R Walkinshaw, David A Hood
American journal of physiology. Endocrinology and metabolism 293 E672-80 2007
Mitochondrial myopathy patients (MMPs) have impaired oxidative phosphorylation and exercise intolerance. Endurance training of MMPs improves exercise tolerance, but also increases mutational load. To assess the regulation of mitochondrial content in MMPs, we measured proteins involved in 1) biogenesis, 2) oxidative stress, and 3) apoptosis in MMPs and healthy controls (HCs) both before and after endurance training. Before training, MMPs had a greater mitochondrial content, along with a 1.4-fold (P 0.05) higher expression of the biogenesis regulator peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha). The DNA repair enzyme 8-oxoguanine DNA glycolase-1 (OGG-1), the antioxidant manganese superoxide dismutase (MnSOD), and the apoptotic proteins AIF and Bcl-2 were higher in MMPs compared with HCs. Aconitase, an enzyme sensitive to oxidative stress, was 52% lower (P 0.05) in MMPs when calculated based on an estimate of mitochondrial volume and oxidative stress-induced protein modifications tended to be higher in MMPs compared with HCs. Endurance training (ET) induced increases in mitochondrial content in both HC subjects and MMPs, but there was no effect of training on the regulatory proteins Tfam or PGC-1alpha. In MMPs, training induced a selective reduction of OGG-1, an increase in MnSOD, and a reduction in aconitase activity. Thus, before training, MMPs exhibited an adaptive response of nuclear proteins indicative of a compensatory increase in mitochondrial content. Following training, several parallel adaptations occurred in MMPs and HCs, which may contribute to previously observed functional improvements of exercise in MMPs. However, our results indicate that muscle from MMPs may be exposed to greater levels of oxidative stress during the course of training. Further investigation is required to evaluate the long-term benefits of endurance training as a therapeutic intervention for mitochondrial myopathy patients.
|Caloric restriction and gender modulate cardiac muscle mitochondrial H2O2 production and oxidative damage. |
Bartomeu Colom,Jordi Oliver,Pilar Roca,Francisco J Garcia-Palmer
Cardiovascular research 74 2007
Gender and diet have an important effect in cardiovascular disease and other aging-associated disorders, whose initiation and/or worsening seem to be delayed in females from different species and in animals subjected to caloric restriction (CR). The aim of the present study was to investigate whether cardiac muscle bioenergetic mitochondrial features could be responsible for these beneficial effects.
|Oxidative stress and atrial fibrillation after cardiac surgery: a case-control study. |
Ramlawi, Basel, et al.
Ann. Thorac. Surg., 84: 1166-72; discussion 1172-3 (2007) 2007
|Sugar-induced premature aging and altered differentiation in human epidermal keratinocytes. |
Ulrich Berge, Juliane Behrens, Suresh I S Rattan
Annals of the New York Academy of Sciences 1100 524-9 2007
Normal human epidermal keratinocytes (NHEK) show both the Hayflick phenomenon and differentiation in vitro. The aim of this study was to induce senescence in keratinocytes using two sugars, glucose and glyoxal. Induction of senescence in early-passage NHEK was characterized by monitoring cell morphology, short-term growth characteristics, cell proliferation, and viability assay. In addition, apoptosis, senescence-associated (SA) beta-gal activity, proteasomal activity and glycation, and glycoxidation of total proteins were determined. Our results show that a 3-day treatment with 100 mM glucose or 0.1 mM glyoxal induces in early-passage NHEK various cellular and biochemical characteristics comparable to those observed in serially subcultured late passage NHEK. Furthermore, sugar-treated prematurely aged NHEK showed impaired differentiation, as measured by the quantification of involucrin. There is preliminary evidence that a preexposure of NHEK to mild heat shock (41 degrees C, 1 h, 6 h in advance) can abrogate some of the sugar-induced negative effects, which is an example of mild stress-induced hormesis. This experimental model can be useful to study the effects of potential antiaging interventions.
|Super CitriMax (HCA-SX) attenuates increases in oxidative stress, inflammation, insulin resistance, and body weight in developing obese Zucker rats. |
Mohammad Asghar, Emmanuel Monjok, Ghislaine Kouamou, Sunny E Ohia, Debasis Bagchi, Mustafa F Lokhandwala
Molecular and cellular biochemistry 304 93-9 2007
Super CitriMax (HCA-SX) is a novel calcium/potassium salt of (-)-hydroxycitric acid extracted from the dried fruit rind of the plant Garcinia cambogia, and commonly consumed as weight loss dietary supplement. In the present study, we investigated the effect of HCA-SX on inflammation, oxidative stress and insulin resistance in developing obese Zucker rats, an animal model of type II diabetes associated with inflammation and oxidative stress. Male Zucker rats (5-week old) were supplemented with vehicle (control) and HCA-SX in drinking water for 7 weeks. Oxidative stress markers, including malondialdehyde (MDA), protein carbonyl (DNPH), and protein tyrosine nitration (tyr-NO(2)) were measured in the liver and kidney tissues using biochemical and immunoblotting techniques. Compared to controls, the levels of MDA, DNPH and tyr-NO(2) were lower in the liver and kidney of HCA-SX-treated animals. Furthermore, the levels of C-reactive protein and interleukin-6, markers of inflammation measured by ELISA, were lower in the plasma of HCA-SX-supplemented animals compared to controls, as were levels of fasting plasma insulin, glucose, and triglycerides. Interestingly, insulin resistance did not develop in HCA-SX-supplemented rats. Food-intake and body weight gain was also lower in rats supplemented with HCA-SX compared to their control counterparts. These results suggest that HCA-SX supplementation in obese Zucker rats reduces food-intake, body weight gain, and also attenuates the increases in inflammation, oxidative stress, and insulin resistance observed in untreated animals. Therefore, HCA-SX may be used as an intervention to overcome obesity-related complications, including inflammation, oxidative stress, and insulin resistance.
|Insulin-like growth factor I deficiency prolongs survival and antagonizes paraquat-induced cardiomyocyte dysfunction: role of oxidative stress. |
Qun Li, Xiaoping Yang, Nair Sreejayan, Jun Ren
Rejuvenation research 10 501-12 2007
Interruption of insulin-like growth factor I (IGF-1) signaling has been demonstrated to prolong life span although the underlying mechanism has not been elucidated. The aim of this study was to examine the influence of severe IGF-1 deficiency on survival rate, cardiomyocyte viability, contractile function, and intracellular Ca(2+) property in response to challenge with the pro-oxidant paraquat. C57 negative and liver IGF-1 deficient (LID) transgenic mice were administrated paraquat (75 mg/kg) and survival was monitored. LID mice displayed a significantly improved survival than did C57 mice evaluated by the Kaplan-Meier curve. MTT assay revealed that in vitro IGF-1 treatment significantly sensitized paraquat-induced cell death in both C57 and LID groups, with significantly better cell viability in LID cardiomyocytes. Compared to C57 mouse cardiomyocytes, LID myocytes displayed reduced peak shortening (PS), decreased maximal velocity of shortening/relengthening (+/- dL/dt), prolonged time-to-90% relengthening (TR(90)), and comparable tolerance to high stimulus frequency and intracellular Ca(2+) homeostasis. Paraquat treatment for 48 hours reduced PS, +/- dL/dt, tolerance to high stimulus frequency, resting and rise in intracellular Ca(2+), and prolonged TR(90), all of which were nullified or masked by IGF-1 deficiency. Paraquat increased reactive oxygen species and carbonyl production upregulated the Ca(2+) regulating protein SERCA2a, and downregulated Na(+) -Ca(2+) exchanger, the effects of which were nullified or masked by IGF-1 deficiency. Although LID mice displayed reduced whole body glucose clearance, cardiomyocytes from LID mice exhibited dramatically enhanced insulin-stimulated phosphorylation of insulin receptor and Akt. These data demonstrated that IGF-1 deficiency may antagonize or mask the paraquat-induced decrease in survival, cardiomyocyte dysfunction, oxidative stress, and change in Ca(2+) regulating proteins.
|Elimination of damaged proteins during differentiation of embryonic stem cells. |
Hernebring, Malin, et al.
Proc. Natl. Acad. Sci. U.S.A., 103: 7700-5 (2006) 2006
|Dietary curcumin counteracts the outcome of traumatic brain injury on oxidative stress, synaptic plasticity, and cognition |
Wu, A. et al.
Exp. Neurol, 197(2):309-317 (2006) 2006
|Membrane protein carbonylation in non-leukodepleted CPDA-preserved red blood cells. |
Anastasios G Kriebardis, Marianna H Antonelou, Konstantinos E Stamoulis, Effrosini Economou-Petersen, Lukas H Margaritis, Issidora S Papassideri
Blood cells, molecules diseases 36 279-82 2006
Transfusion of allogeneic blood products is associated with adverse reactions and complications. Some of the negative effects of RBC transfusion are associated with the storage lesion. The importance of RBC oxidative damage in the storage lesion is not well documented. We monitored the storage-induced membrane protein oxidation in CPDA-preserved non-leukodepleted RBCs units from five blood donors in the course of the storage period, as assessed by protein carbonylation levels estimation. Carbonylated protein content was determined following 2,4-dinitrophenylhydrazine derivatization and SDS-polyacrylamide gel electrophoresis coupled with Western blotting. Immunoblotting with dinitrophenol-specific antibody revealed increased RBC membrane protein carbonyls with prolonged storage in CPDA units. This finding supports the idea of oxidation as a part of the storage lesion.
|Proteasome inhibition by MG-132 induces apoptotic cell death and mitochondrial dysfunction in cultured rat brain oligodendrocytes but not in astrocytes. |
Olaf Goldbaum, Grit Vollmer, Christiane Richter-Landsberg
Glia 53 891-901 2006
Proteasomal dysfunction has been implicated in neurodegenerative disorders and during aging processes. In frontotemporal dementias, corticobasal degeneration, and progressive supranuclear palsy, oligodendrocytes are specifically damaged. Application of proteasomal inhibitors to cultured oligodendrocytes is associated with apoptotic cell death. The present study was undertaken to investigate the death pathway activated in oligodendrocytes by proteasomal inhibition. Our data show that the proteasomal inhibitor MG-132 causes oxidative stress, as indicated by the upregulation of the small heat shock protein heme oxygenase-1 (HO-1) and the appearance of oxidized proteins. Activation of the mitochondrial pathway was involved in the apoptotic process. Mitochondrial membrane potential was disturbed, and cytochrome c was released from the mitochondria. Concomitantly, death-related caspases 3 and 9 were activated and poly(ADP-ribose)-polymerase cleavage occurred. MG-132-induced cell death, DNA-fragmentation, and caspase activation could be prevented by the broad caspase inhibitor zVAD-fmk. In contrast to oligodendrocytes, cultured astrocytes showed resistance to the treatment with proteasomal inhibitors and did not reveal cytotoxic responses. This was also observed in astrocytes differentiated in the presence of dibutyryl cyclic AMP. Hence, individual cells respond differently to proteasomal inhibition and the therapeutic use of proteasomal inhibitors, e.g. for the treatment of cancer or inflammatory diseases, needs to be carefully evaluated.
|Gamma-glutamylcysteine ethyl ester protection of proteins from Abeta(1-42)-mediated oxidative stress in neuronal cell culture: a proteomics approach |
Boyd-Kimball, D. et al.
J. Neurosci. Res., 79(5):707-713 (2005) 2005
|Specific inactivation of cysteine protease-type cathepsin by singlet oxygen generated from naphthalene endoperoxides. |
Nagaoka, Yuki, et al.
Biochem. Biophys. Res. Commun., 331: 215-23 (2005) 2005
Singlet oxygen is a causal factor in light-induced skin photoaging and the cytotoxic process of tumor cells in photodynamic chemotherapy. To develop a better understanding of the functional consequences of protein modification by singlet oxygen, the effects of naphthalene endoperoxide on lysosomal protease, cathepsin, were examined. When the soluble fraction of normal human fetal skin fibroblast cells was treated with the endoperoxide, the activities of cysteine proteases, cathepsins B and L/S, were inhibited, but that of aspartate protease, cathepsin D/E, was not. The reduction of the endoperoxide-treated soluble fractions by treatment with dithiothreitol barely recovered the activities. Cathepsin B, purified from normal human liver, exhibited similar profiles to that in cytosol. These data suggest that singlet oxygen oxidatively modifies an amino acid residue essential for catalysis and consequently results in the irreversible inactivation of cysteine protease-type cathepsin.
|Ketone bodies stimulate chaperone-mediated autophagy. |
Finn, Patrick F and Dice, J Fred
J. Biol. Chem., 280: 25864-70 (2005) 2005
Chaperone-mediated autophagy (CMA) is a selective lysosomal protein degradative process that is activated in higher organisms under conditions of prolonged starvation and in cell culture by the removal of serum. Ketone bodies are comprised of three compounds (beta-hydroxybutyrate, acetoacetate, and acetone) that circulate during starvation, especially during prolonged starvation. Here we have investigated the hypothesis that ketone bodies induce CMA. We found that physiological concentrations of beta-hydroxybutyrate (BOH) induced proteolysis in cells maintained in media with serum and without serum; however, acetoacetate only induced proteolysis in cells maintained in media with serum. Lysosomes isolated from BOH-treated cells displayed an increased ability to degrade both glyceraldehyde-3-phosphate dehydrogenase and ribonuclease A, substrates for CMA. Isolated lysosomes from cells maintained in media without serum also demonstrated an increased ability to degrade glyceraldehyde-3-phosphate dehydrogenase and ribonuclease A when the reaction was supplemented with BOH. Such treatment did not affect the levels of lysosome-associated membrane protein 2a or lysosomal heat shock cognate protein of 70 kDa, two rate-limiting proteins in CMA. However, pretreatment of glyceraldehyde-3-phosphate and ribonuclease A with BOH increased their rate of degradation by isolated lysosomes. Lysosomes pretreated with BOH showed no increase in proteolysis, suggesting that BOH acts on the substrates to increase their rates of proteolysis. Using OxyBlot analysis to detect carbonyl formation on proteins, one common marker of protein oxidation, we showed that treatment of substrates with BOH increased their oxidation. Neither glycerol, another compound that increases in circulation during prolonged starvation, nor butanol or butanone, compounds closely related to BOH, had an effect on CMA. The induction of CMA by ketone bodies may provide an important physiological mechanism for the activation of CMA during prolonged starvation.
|Cigarette smoke metal-catalyzed protein oxidation leads to vascular endothelial cell contraction by depolymerization of microtubules |
Bernhard, D. et al.
FASEB J. , 19(9):1096-1107 (2005) 2005
|Proteomic identification of oxidatively modified retinal proteins in a chronic pressure-induced rat model of glaucoma. |
Tezel, Gülgün, et al.
Invest. Ophthalmol. Vis. Sci., 46: 3177-87 (2005) 2005
PURPOSE: Based on the evidence of an amplified production of reactive oxygen species (ROS) during glaucomatous neurodegeneration, proteomic analysis was performed to determine oxidative modification of retinal proteins after experimental elevation of intraocular pressure (IOP). METHODS: IOP elevation was induced in rats by hypertonic saline injections into episcleral veins. Protein expression was determined by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) of retinal protein lysates obtained from eyes matched for the cumulative IOP exposure and axon loss. To determine protein oxidation levels, protein carbonyls were detected through 2D-oxyblot analysis of 2,4-dinitrophenylhydrazine (DNPH)-treated samples using an anti-DNP antibody. For identification of oxidized proteins, peptide masses were analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF/MS) and liquid chromatography-tandem mass spectrometry (LC/MS/MS). In addition to use of different engines in a bioinformatic database search and performance of peptide sequencing and 2D-Western blot analysis for confirmation of the identified proteins, immunohistochemistry was used for further validation of the proteomic findings. RESULTS: Comparison of 2D-oxyblots with Coomassie Blue-stained 2D-gels revealed that approximately 60 protein spots obtained with retinal protein lysates from ocular hypertensive eyes (of >400 spots) exhibited protein carbonyl immunoreactivity, which reflects oxidatively modified proteins. There was a significant increase in anti-carbonyl reactivity in individual protein spots obtained with retinal protein lysates from ocular hypertensive eyes compared with the control (P < 0.01). The identified proteins through peptide mass fingerprinting and peptide sequencing included glyceraldehyde-3-phosphate dehydrogenase, a glycolytic enzyme; HSP72, a stress protein; and glutamine synthetase, an excitotoxicity-related protein. Immunolabeling of retina sections with specific antibodies demonstrated cellular localization of these proteins as well as retinal distribution of the increased protein carbonyl immunoreactivity in ocular hypertensive eyes. CONCLUSIONS: The findings of this in vivo study provide novel evidence for oxidative modification of many retinal proteins in ocular hypertensive eyes and identify three specific targets of retinal protein oxidation in these eyes, thereby supporting the association of oxidative damage with neurodegeneration in glaucoma. By using a proteomic approach, this study also exemplifies that proteomics provide a very promising way to elucidate pathogenic mechanisms in glaucoma at the protein level.
|Proteomic analysis of parkin knockout mice: alterations in energy metabolism, protein handling and synaptic function |
Periquet, M. et al.
J. Neurochem., 95(5):1259-1276 (2005) 2005
|Plasma infusions into porcine cerebral white matter induce early edema, oxidative stress, pro-inflammatory cytokine gene expression and DNA fragmentation: implications for white matter injury with increased blood-brain-barrier permeability. |
Wagner, Kenneth R, et al.
Current neurovascular research, 2: 149-55 (2005) 2005
Plasma infused into porcine cerebral white matter induces both acute interstitial and delayed vasogenic edema. Edematous white matter contains extracellular plasma proteins and rapidly induces oxidative stress as evidenced by increased protein carbonyl formation and heme oxygenase-1 induction. We tested the hypothesis that edematous white matter would also upregulate pro-inflammatory cytokine gene expression and develop DNA damage. We infused autologous plasma into the frontal hemispheric white matter of pentobarbital-anesthetized pigs. We monitored and controlled physiological variables and froze brains in situ at 1, 4 or 24 hrs. We determined edema volumes by computer-assisted morphometry. We measured white matter protein carbonyl formation by immunoblotting, cytokine gene expression by standard RT-PCR methods and DNA fragmentation by agarose gel electrophoresis. White matter edema developed acutely (1 hr) after plasma infusion and increased significantly in volume between 4 and 24 hrs. Protein carbonyl formation also occurred rapidly in edematous white matter with significant elevations (3 to 4-fold) already present at 1 hr. This increase remained through 24 hrs. Pro-inflammatory cytokine gene expression was also rapidly increased at 1 hr post-infusion. Evidence for DNA fragmentation began at 2 to 4 hrs, and a pattern indicative of both ongoing necrosis and apoptosis was robust by 24 hrs. Plasma protein accumulation in white matter induces acute edema development and a cascade of patho-chemical events including oxidative stress, pro-inflammatory cytokine gene expression and DNA damage. These results suggest that in diseases with increased blood-brain barrier (BBB) permeability or following intracerebral hemorrhage or traumatic brain injury, interstitial plasma can rapidly damage white matter.
|Smoking, oxidative stress and inflammation: impact on resting energy expenditure in diabetic nephropathy. |
BMC nephrology, 6: 13 (2005) 2005
BACKGROUND: Inflammation is associated with increased resting energy expenditure (REE) in patients with chronic kidney disease. Oxidative stress, on the other hand, appears not to increase REE. Smoking is a common mechanism for generating oxidative stress and inflammation. Whether smokers have increased REE and if so, whether it is accounted for by the pro-oxidant and inflammatory state is not known. METHODS: A case control study of 11 smokers and 24 non-smokers with overt diabetic nephropathy was performed to evaluate the chronic effect of smoking on REE. REE (indirect calorimetry), glomerular filtration rate (iothalamate clearance), markers of oxidative stress (urinary and plasma malondialdehyde (MDA), and protein carbonyls) and inflammation (C-reactive protein, tumor necrosis factor-alpha, interleukin-6) were measured on two occasions four months apart. RESULTS: Biomarkers of inflammation (C-reactive protein) and oxidative stress (urinary and plasma MDA) were increased in smokers. REE was increased in smokers, 24.3 kcal/kg/day compared to 21 kcal/kg/day (p = 0.009) in non-smokers. After adjusting for age, GFR, MDA, C-reactive protein, and hemoglobin A1C the difference in REE between the two groups persisted (adjusted difference 3.51 kcal/kg/d, 95% confidence interval 0.59-6.45, p = 0.020). CONCLUSION: Patients with overt diabetic nephropathy who smoke have a higher REE, oxidative and inflammatory state. Elevated REE is not attributable to heightened oxidative stress and inflammatory state. Smoking is an independent risk factor for elevated REE in patients with diabetic nephropathy and provides an additional mechanism by which it may lead to poor outcomes.
|N-acetylcysteine increases manganese superoxide dismutase activity in septic rat diaphragms |
Barreiro, E. et al.
Eur. Respir. J., 26(6):1032-1039 (2005) 2005
|Dual role of vitamin C in an oxygen-sensitive system: discrepancy between DNA damage and cell death. |
Minyi Shi, Baohui Xu, Kazuko Azakami, Tomohiro Morikawa, Kunitomo Watanabe, Kanehisa Morimoto, Masaharu Komatsu, Kohji Aoyama, Toru Takeuchi
Free radical research 39 213-20 2005
Although vitamin C is considered to act both as pro-oxidant and antioxidant, the mechanisms underlying these actions are still unclear. Using the oxygen-sensitive system of a strict anaerobe, Prevotella melaninogenica, we investigated both the pro-oxidant and antioxidant mechanisms of vitamin C. In the presence of vitamin C, the 8-hydroxydeoxyguanosine (8OHdG) formation induced by oxygen exposure was enhanced, probably due to the action of vitamin C on hydrogen peroxide generated during oxygen exposure: while catalase almost completely suppressed the enhancing effect of vitamin C, 8OHdG formation induced by hydrogen peroxide was enhanced by vitamin C. By contrast, the presence of vitamin C inhibited bacterial cell death, membrane damage, and lipid peroxidation induced by oxygen exposure. Sodium azide showed similar effects to vitamin C, thus the antioxidant action of vitamin C may be due to its quenching of the singlet oxygen generated in this system. Both the pro-oxidant and antioxidant effects of vitamin C were observed only in acidic conditions.
|Effects of apolipoprotein E on the human immunodeficiency virus protein Tat in neuronal cultures and synaptosomes |
Pocernich, C. B. et al.
J. Neurosci. Res., 77(4):532-539 (2004) 2004
|Oxidative stress, beta-cell apoptosis, and decreased insulin secretory capacity in mouse models of hemochromatosis |
Cooksey, R. C. et al.
Endocrinology, 145(11):5305-5312 (2004) 2004
|Human endogenous retrovirus glycoprotein-mediated induction of redox reactants causes oligodendrocyte death and demyelination |
Antony, J. M. et al.
Nat. Neurosci., 7(10):1088-1095 (2004) 2004
|Evidence of myofibrillar protein oxidation induced by postischemic reperfusion in isolated rat hearts |
Canton, M. et al.
Am. J. Physiol. Heart Circ. Physiol., 286(3):H870-H877 (2004) 2004
|Oxidative stress, induced by 6-hydroxydopamine, reduces proteasome activities in PC12 cells: implications for the pathogenesis of Parkinson's disease |
Elkon, H. et al.
J. Mol. Neurosci., 24(3):387-400 (2004) 2004
|Influence of early postmortem protein oxidation on beef quality |
Rowe, L. J. et al.
J. Anim. Sci., 82(3):785-793 (2004) 2004
|Oxidative stress in HIV demented patients and protection ex vivo with novel antioxidants |
Turchan, J, et al
Neurology, 60:307-14 (2003) 2003
|Changes in rat liver mitochondria with aging. Lon protease-like reactivity and N(epsilon)-carboxymethyllysine accumulation in the matrix |
Bakala, H. et al.
Eur. J. Biochem., 270(10):2295-2302 (2003) 2003
|The mitochondrial DNA polymerase as a target of oxidative damage |
Graziewicz, M. A. et al.
Nucleic Acids Res., 30(13):2817-2824 (2002) 2002
|The curry spice curcumin reduces oxidative damage and amyloid pathology in an Alzheimer transgenic mouse |
Lim, G. P. et al.
J. Neurosci., 21(21):8370-8377 (2001) 2001
|Hallmarks of Aging|
|Poster: Methods For The Detection of Oxidative Stress|
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|OxyBlot Protein Oxidation Detection Kit|