8-hydroxy-2--deoxyguanosine and Reperfusion-Injury

One of the important factors in the occurrence of acute kidney injury (AKI) among renal transplant patients (RTPs) is ischemia reperfusion injury (IRI). The current study aimed at determining the anti-inflammatory and anti-oxidative effects of melatonin on the complications of IRI and the level of Klotho expression in these patients.. A total of 40 renal transplant candidates were randomly assigned into placebo or melatonin group receiving the same dose of 3 mg/day. In order to measure serum melatonin levels, inflammatory and oxidative stress factors, renal function biomarkers, and Klotho gene/protein expression, venous blood samples were taken from patients over two different time points, i e, 24 h before the transplantation and at discharge from hospital.. Melatonin was associated with improvement in renal transplantation, since the serum level of neutrophil gelatinase-associated lipocalin, as a renal functional marker, significantly decreased (P < .001). The effect of melatonin as a suppressor of inflammation and oxidative stress was also evident in the melatonin group due to a significant reduction in the serum levels of MDA, CP, 8-OHdG, and TNF-α markers (P < .001).. Reduction in serum levels of renal function and oxidative stress/inflammatory markers in the melatonin group indicates that melatonin can inhibit IRI outcomes in RTPs through its anti-oxidant and anti-inflammatory properties. However, these properties do not appear as a result of influence on the level of Klotho gene/protein expression.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Adult; Anti-Inflammatory Agents; Antioxidants; Double-Blind Method; Female; Gene Expression Regulation; Glucuronidase; Humans; Kidney Transplantation; Klotho Proteins; Lipocalin-2; Male; Malondialdehyde; Melatonin; Middle Aged; Protein Carbonylation; Reperfusion Injury; Tumor Necrosis Factor-alpha

We previously demonstrated that prehabilitation by running on a treadmill leads to improved survival after gut ischemia reperfusion (I/R) in mice. The purpose of this research was to examine whether prehabilitation attenuates inflammatory responses after gut I/R in mice.. Male C57BL/6J mice (n = 92) were assigned to the sedentary (n = 46) or the exercise (n = 46) group. The exercise group ran on a treadmill for 4 wk, while the sedentary mice did not exercise. After the 4-week pretreatment, all mice underwent gut I/R and the blood, urine, small intestine, lung, liver, and gastrocnemius were harvested prior to ischemia or at 0, 3, 6, or 24 h after reperfusion. Histologically demonstrated organ damage, cytokine levels in the blood, gut and gastrocnemius, myeloperoxidase activity in the gut, 8-hydroxy-2'-deoxyguanosine levels in urine and the gut, and adenosine triphosphate (ATP) and ATP + ADP + adenosine monophosphate levels in the gut and gastrocnemius were evaluated.. The treadmill exercise reduced gut and lung injuries at 3 h and liver injury at 6 h after reperfusion. Running on the treadmill also decreased proinflammatory cytokine levels in the blood at 6 h, gut at 3 h and gastrocnemius at 6 h after reperfusion, myeloperoxidase activity in the gut prior to ischemia, and 6 h after reperfusion and the urinary 8-hydroxy-2'-deoxyguanosine level at 24 h after reperfusion, while ATP levels in exercised mice prior to ischemia and 3 h after reperfusion were increased in the intestine as compared to the levels in sedentary mice.. Prehabilitation with treadmill exercise reduces inflammatory responses after gut I/R and may exert protective actions against gut I/R.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Adenosine Triphosphate; Animals; Antioxidants; Cytokines; Ischemia; Male; Mice; Mice, Inbred C57BL; Peroxidase; Physical Conditioning, Animal; Preoperative Exercise; Reperfusion Injury

Oxidative damage is critical in the pathogenesis of ovarian ischaemia/reperfusion (I/R) injury, and statins have been reported to exert antioxidant activity. However, the role of VCAM-1 and xanthine oxidase (XO)/uric acid (UA) in ovarian I/R injury is not known. Also, whether or not atorvastatin exerts antioxidant activity like other statins is unclear.. This study investigated the involvement of VCAM-1 and XO/UA in ovarian I/R injury and the likely protective role of atorvastatin.. Forty female Wistar rats were randomized into sham-operated, ischaemia, ischaemia/reperfusion (I/R), ischaemia and atorvastatin, and I/R and atorvastatin.. In comparison with the sham-operated group, atorvastatin blunted ischaemia and I/R-induced distortion of ovarian histoarchitecture and follicular degeneration. Also, atorvastatin alleviated ischaemia and I/R-induced rise in XO, UA, and malondialdehyde, which was accompanied by inhibition of ischaemia and I/R-induced reductions in reduced glutathione level, enzymatic antioxidant activities and increase in myeloperoxidase activity and TNF-α and IL-6 levels by atorvastatin treatment. Additionally, atorvastatin blocked ischaemia and I/R-induced increase in VCAM-1 expression, caspase 3 activity, 8-hydroxydeoxyguanosine level and ovarian DNA fragmentation index.. For the first time, this study revealed that atorvastatin-mediated downregulation of VCAM-1 and XO/UA/caspase 3 signaling averts oxidative injury, inflammation, and apoptosis induced by ovarian ischaemia/reperfusion injury.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Antioxidants; Apoptosis; Atorvastatin; Caspase 3; Down-Regulation; Female; Glutathione; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Interleukin-6; Ischemia; Malondialdehyde; Oxidative Stress; Peroxidase; Rats; Rats, Wistar; Reperfusion Injury; Tumor Necrosis Factor-alpha; Uric Acid; Vascular Cell Adhesion Molecule-1; Xanthine Oxidase

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Apoptosis; Cell Line, Tumor; Cell Survival; Disease Models, Animal; Enzyme Inhibitors; Hydrogen Peroxide; Infarction, Middle Cerebral Artery; Janus Kinase 2; Mice; Neuroblastoma; Oxidants; Oxidative Stress; Rats; Reperfusion Injury; Signal Transduction; STAT3 Transcription Factor; Tyrphostins

Sodium salt of deoxyribonucleic acid (DNA), Derinat, isolated from the soft roes of Russian sturgeon, has been utilized as an immunomodulator for the treatment of reactive oxygen species (ROS)-associated diseases in clinics. Here we show that treatment with Derinat has an anti-inflammatory and anti-oxidative effects on cutaneous ischemia-reperfusion (IR) injury in pressure ulcer (PU) model mice. Dorsal skin damage and dermal edema in mild PU model mice were attenuated by treatment with Derinat. Immunohistochemical and biochemical analyses showed that Derinat suppressed IR-induced oxidative damage, i.e. accumulation of 8-hydroxy-2'-deoxyguanosine (8-OHdG), and related inflammatory factors such as cyclooxygenase 2 (COX-2) and IL-6 receptor (IL-6R) in dorsal skin from PU model mice. We also verified that phospholyated/non-phosphorylated ratio of extracellular signal-regulated kinase (Erk) and p38 mitogen-activated protein kinase (MAPK) increased after IR, which were attenuated by Derinat. We then compared the effect of Derinat with that of salmon DNA and other PU therapeutic agents, prostaglandin E1 (PGE1) and basic fibroblast growth factor (bFGF), by using severe PU model mice. The effects of Derinat and salmon-DNA were compatible with those of PGE1 and bFGF. These results suggest that Derinat other fish-derived DNA formulation could be effective enough and become intriguing new therapeutic options.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Cyclooxygenase 2; Deoxyguanosine; Disease Models, Animal; DNA; Extracellular Signal-Regulated MAP Kinases; Fishes; Inflammation Mediators; Mice, Inbred BALB C; p38 Mitogen-Activated Protein Kinases; Pressure; Receptors, Interleukin-6; Reperfusion Injury; Skin; Skin Ulcer

This study evaluated the effect of mycophenolate mofetil (MMF) on uterine tissue preservation following ischaemia/reperfusion (I/R) injury. Uterine I/R injury was induced in rats by clamping the lower abdominal aorta and ovarian arteries for 30 min. Group I/R + V (n = 7) received vehicle alone while Group I/R + M (n = 7) received 20 mg/kg/day MMF. Control groups underwent sham surgery and received vehicle (Group C) or 20 mg/kg/day MMF (Group M) (n = 7 for both). Four hours after detorsion, uterine tissue 8-hydroxy-2'-deoxyguanosine (8-OHdG), glutathione, malondialdehyde (MDA), myeloperoxidase (MPO), superoxide dismutase (SOD) and serum ischaemia modified albumin (IMA) concentrations were measured. Histopathological analyses were performed. The I/R + M group showed significant reduction in serum IMA and uterine tissue 8-OHdG, MDA and MPO and significant increase in SOD concentrations compared with the I/R + V group, indicating a protective effect against I/R oxidative damage (P = 0.009, P = 0.006, P = 0.002, P = 0.003 and P = 0.009, respectively). Histopathological evaluation revealed MMF treatment resulted in significantly less tissue and cellular damage and apoptosis compared with the I/R + V group. These results indicate MMF is effective in attenuating uterine tissue damage and preventing apoptosis following uterine I/R injury, probably via anti-inflammatory and anti-oxidative action.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Albumins; Animals; Antibiotics, Antineoplastic; Antioxidants; Aorta, Abdominal; Arteries; Deoxyguanosine; Disease Models, Animal; Female; Glutathione; Immunosuppressive Agents; Mycophenolic Acid; Ovary; Peroxidase; Rats; Rats, Wistar; Reperfusion Injury; Superoxide Dismutase; Uterus

It is of great importance to protect the brain against cerebral ischemia and reperfusion (I/R) injury, which leads to excitotoxicity, redox imbalance, inflammation and apoptosis; however, there is currently no effective treatment. The present study aimed to investigate the effect of H2S preconditioning on cerebral I/R injury and its underlying mechanism. The results demonstrated that H2S preconditioning significantly prevented the development of neurological function abnormality, inflammation and oxidative injury in mice as well as cognitive impairment caused by cerebral I/R. H2S preconditioning also suppressed the apoptosis caused by cerebral I/R. Moreover, the protective effect of H2S preconditioning was found to involve heat shock protein 70 (HSP70), in which the PI3K/Akt/Nrf2 pathway was involved. The data showed that H2S preconditioning could protect mice against cerebral I/R injury by the induction of HSP70 and the PI3K/Akt/Nrf2 pathway.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Administration, Inhalation; Animals; Brain Infarction; Deoxyguanosine; Disease Models, Animal; Enzyme Inhibitors; HSP70 Heat-Shock Proteins; Hydrogen Sulfide; Interleukin-6; Male; Malondialdehyde; Maze Learning; Mice; Mice, Inbred C57BL; Mice, Transgenic; Neurologic Examination; NF-E2-Related Factor 2; Oncogene Protein v-akt; Phosphatidylinositol 3-Kinases; Reperfusion Injury; Signal Transduction; Tumor Necrosis Factor-alpha

Liver ischemia-reperfusion (I/R) injury is a severe complication of liver surgery, and steatosis is a risk factor for liver damage. Reactive oxygen species generated by nicotinamide adenine dinucleotide phosphate oxidase (NOX) contribute to liver dysfunction. Here we examined the role of NOX in I/R injury of fatty livers.. Rats were fed a methionine and choline-deficient diet to induce a fatty liver. Rats then underwent surgically induced partial hepatic ischemia followed by reperfusion.. The overall survival rate after I/R was lower in rats with fatty livers than with normal livers (P < 0.01). Necrotic area and the concentrations of 8-hydroxy-2'-deoxyguanosine (8-OHdG), TNFα, and IL-6 were higher in fatty liver tissue than in normal liver tissue (P < 0.01). The number of p47phox-positive cells was significantly higher in fatty liver tissue than in normal liver tissue after reperfusion and peaked 24 hours after reperfusion. The number of TLR-4 positive cells was significantly higher in fatty liver tissue than in normal liver tissue after reperfusion and peaked 4 and 24 hours after reperfusion coupled with a decreased number of high-mobility group box 1-positive hepatocytes. Apocynin significantly improved the survival rate, necrotic area, and concentrations of 8-hydroxy-2'-deoxyguanosine, TNFα, and IL-6 (P < 0.01). The protective effect of apocynin on fatty livers was greater than on normal livers.. Ischemia-reperfusion injury was associated with increased high-mobility group box 1, TLR4, and NOX2. Inhibition of NOX activity improved oxidative stress and may prevent I/R injury in fatty liver.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Cells, Cultured; Choline Deficiency; Deoxyguanosine; Disease Models, Animal; Enzyme Inhibitors; Fatty Liver; HMGB1 Protein; Inflammation Mediators; Interleukin-6; Liver; Macrophages; Male; Membrane Glycoproteins; Methionine; NADPH Oxidase 2; NADPH Oxidases; Necrosis; Oxidative Stress; Rats, Sprague-Dawley; Reactive Oxygen Species; Reperfusion Injury; Signal Transduction; Time Factors; Toll-Like Receptor 4; Tumor Necrosis Factor-alpha

The present study investigates the effects of d-allose, a rare sugar, on the inflammatory response after transient forebrain ischemia in the gerbil and whether it reduces oxidative stress (8-hydroxyl-2'-deoxyguanosine levels) and behavioral deficits.. Transient forebrain ischemia was induced by occlusion of the bilateral common carotid arteries for 5 minutes. d-Allose was intraperitoneally injected immediately after ischemia (400 mg/kg). Inflammatory cytokines and oxidative damage in the hippocampus and behavioral deficits were examined 3 days after ischemia.. d-Allose administration reduced ischemia-induced cytokine production, oxidative stress, and behavioral deficits (motor and memory related).. The present results suggest that d-allose reduces brain injury after transient global ischemia by suppressing inflammation as well as by inhibiting oxidative stress.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Analysis of Variance; Animals; Blood Glucose; Blood Pressure; Cytokines; Deoxyguanosine; Disease Models, Animal; Dose-Response Relationship, Drug; Gene Expression Regulation; Gerbillinae; Glucose; Hippocampus; Male; Maze Learning; Movement Disorders; Reperfusion Injury; Sweetening Agents; Time Factors

Ischemia-reperfusion injury (IRI) has been considered as the major cause of acute kidney injury and can result in poor long-term graft function. Functional recovery after IRI is impaired in the elderly. In the present study, we aimed to compare kidney morphology, function, oxidative stress, inflammation, and development of renal fibrosis in young and aged rats after renal IRI.. Rat models of warm renal IRI were established by clamping left pedicles for 45 min after right nephrectomy, then the clamp was removed, and kidneys were reperfused for up to 12 wk. Biochemical and histologic renal damage were assessed at 12 wk after reperfusion. The immunohistochemical staining of monocyte macrophage antigen-1 (ED-1) and transforming growth factor beta 1 (TGF-β1) and messenger RNA level of TGF-β1 in the kidney were analyzed.. Renal IRI caused significant increases of malondialdehyde and 8-hydroxydeoxyguanosine levels and a decrease of superoxide dismutase activity in young and aged IRI rats; however, these changes were more obvious in the aged rats. IRI resulted in severe inflammation and tubulointerstitial fibrosis with decreased creatinine (Cr) clearance and increased histologic damage in aged rats compared with young rats. Moreover, we measured the ratio of Cr clearance between young and aged IRI rats. It demonstrated that aged IRI rats did have poor Cr clearance compared with the young IRI rats. ED-1 and TGF-β1 expression levels in the kidney were significantly higher in aged rats than in young rats after IRI.. Aged rats are more susceptible to IRI-induced renal failure, which may associate with the increased oxidative stress, increased histologic damage, and increased inflammation and tubulointerstitial fibrosis. Targeting oxidative stress and inflammatory response should improve the kidney recovery after IRI.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Acute Kidney Injury; Age Factors; Aging; Animals; Deoxyguanosine; Disease Models, Animal; Fibrosis; Inflammation; Malondialdehyde; Oxidative Stress; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Reperfusion Injury; Severity of Illness Index; Time

Liver transplant outcomes using grafts donated after cardiac death (DCD) remain poor.. We investigated the effects of ex vivo reconditioning of DCD grafts with venous systemic oxygen persufflation using nitric oxide gas (VSOP-NO) in rat liver transplants. Orthotopic liver transplants were performed in Lewis rats, using DCD grafts prepared using static cold storage alone (group-control) or reconditioning using VSOP-NO during cold storage (group-VSOP-NO). Experiment I: In a 30-min warm ischemia model, graft damage and hepatic expression of inflammatory cytokines, endothelial nitric oxide synthase (eNOS), inducible nitric oxide synthase (iNOS), and endothelin-1 (ET-1) were examined, and histologic analysis was performed 2, 6, 24, and 72 hr after transplantation. Experiment II: In a 60-min warm ischemia model, grafts were evaluated 2 hr after transplantation (6 rats/group), and survival was assessed (7 rats/group).. Experiment I: Group-VSOP-NO had lower alanine aminotransferase (ALT) (P<0.001), hyaluronic acid (P<0.05), and malondialdehyde (MDA) (P<0.001), hepatic interleukin-6 expression (IL-6) (P<0.05), and hepatic tumor necrosis factor-alpha (TNF-α) expression (P<0.001). Hepatic eNOS expression (P<0.001) was upregulated, whereas hepatic iNOS (P<0.01) and ET-1 (P<0.001) expressions were downregulated. The damage of hepatocyte and sinusoidal endothelial cells (SECs) were lower in group-VSOP-NO.Experiment II: VSOP-NO decreased ET-1 and 8-hydroxy-2'deoxyguanosine (8-OHdG) expression and improved survival after transplantation by 71.4% (P<0.01).. These results suggest that VSOP-NO effectively reconditions warm ischemia-damaged grafts, presumably by decreasing ET-1 upregulation and oxidative damage.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Alanine Transaminase; Animals; Cold Ischemia; Cytokines; Cytoprotection; Deoxyguanosine; Endothelin-1; Gases; Hyaluronic Acid; Inflammation Mediators; Liver; Liver Transplantation; Malondialdehyde; Models, Animal; Nitric Oxide; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Rats; Rats, Inbred Lew; Reperfusion Injury; RNA, Messenger; Time Factors; Warm Ischemia

Induced pluripotent stem cells (iPSC) are novel stem cell populations, but the role of iPSC in retinal ischemia and reperfusion (I/R) injury remains unknown. Since oncogene c-Myc is substantially contributed to tumor formation, in this study, we investigated the effects, mechanisms and safety of subretinal transplantation of iPSC without c-Myc (non-c-Myc iPSC) in a rat model of retinal I/R injury. Retinal I/R injury was induced by raising the intraocular pressure of Sprague-Dawley rats to 110 mmHg for 60 min. A subretinal injection of non-c-Myc iPSC or murine epidermal fibroblast was given 2 h after I/R injury. Electroretinograms (ERG) were performed to determine the functionality of the retinas. The surviving retinal ganglion cells (RGCs) and retinal apoptosis following I/R injury were determined by counting NeuN-positive cells in whole-mounted retinas and TUNEL staining, respectively. The generation of reactive oxygen species (ROS) and the activities of superoxide dismutase (SOD) and catalase (CAT) in the retinal tissues were determined by lucigenin- and luminol-enhanced chemiluminescence and enzyme-linked immunosorbent assay (ELISA). The degree of retinal oxidative damage was assessed by nitrotyrosine, acrolein, and 8-hydroxy-2'-deoxyguanosine (8-OHdG) staining. The expression of brain-derived neurotrophic factor (BDNF), ciliary neurotrophic factor (CNTF) and basic fibroblast growth factor (bFGF) in retinas was measured by immunohistochemistry and ELISA. We found that subretinal transplantation of non-c-Myc iPSC significantly suppressed the I/R-induced reduction in the ERG a- and b-wave ratio, attenuated I/R-induced loss of RGCs and remarkably ameliorated retinal morphological changes. Non-c-Myc iPSC potentially increased the activities of SOD and CAT, decreased the levels of ROS, which may account for preventing retinal cells from apoptotic cell death. In addition, the levels of BDNF and CNTF in retina were significantly elevated in non-c-Myc iPSC-treated eyes. Track the non-c-Myc iPSC after transplantation, most transplanted cells are remained in the subretinal space, with spare cells express neurofilament M markers at day 28. Six months after transplantation in I/R injured rats, no tumor formation was seen in non-c-Myc iPSC graft. In conclusion, non-c-Myc iPSC effectively rescued I/R-induced retinal damages and diminished tumorigenicity. Non-c-Myc iPSC transplantation attenuated retinal I/R injury, possibly via a mechanism involving the regulation o

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Apoptosis; Brain-Derived Neurotrophic Factor; Catalase; Cell Count; Ciliary Neurotrophic Factor; Deoxyguanosine; Disease Models, Animal; Electroretinography; Enzyme-Linked Immunosorbent Assay; Fibroblast Growth Factor 2; In Situ Nick-End Labeling; Induced Pluripotent Stem Cells; Mice; Nerve Growth Factors; Proto-Oncogene Proteins c-myc; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Reperfusion Injury; Retinal Diseases; Retinal Vessels; Stem Cell Transplantation; Superoxide Dismutase

Hematopoietic stem cells (HSCs) migrate to injury sites and aid in tissue repair. However, clinical success is poor and is partially due to limited HSC recruitment. We hypothesized that HSC pretreatment with H2O2 would enhance their recruitment to injured gut. As HSCs are rare cells, the number of primary cells obtained from donors is often inadequate for functional experiments. To circumvent this, in this study we utilized a functionally relevant cell line, HPC-7. Anesthetized mice were subjected to intestinal ischemia-reperfusion (IR) injury, and HPC-7 recruitment was examined intravitally. Adhesion to endothelial cells (ECs), injured gut sections, and ICAM-1/VCAM-1 protein were also quantitated in vitro. H2O2 pretreatment significantly enhanced HPC-7 recruitment to injured gut in vivo. A concomitant reduction in pulmonary adhesion was also observed. Enhanced adhesion was also observed in all in vitro models. Increased clustering of α4 and β2 integrins, F-actin polymerization, and filopodia formation were observed in pretreated HPC-7s. Importantly, H2O2 did not reduce HPC-7 viability or proliferative ability. HPC-7 recruitment to injured gut can be modulated by H2O2 pretreatment. This may be through increasing the affinity or avidity of surface integrins that mediate HPC-7 homing to injured sites or through stimulating the migratory apparatus. Strategies that enhance hematopoietic stem/progenitor cell recruitment may ultimately affect their therapeutic efficacy.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Apoptosis; CD18 Antigens; Cell Adhesion; Cell Line; Cellular Senescence; Culture Media, Conditioned; Deoxyguanosine; Endothelium; Frozen Sections; Gastrointestinal Tract; Hematopoietic Stem Cell Transplantation; Hematopoietic Stem Cells; Hydrogen Peroxide; Lung; Male; Mice; Mice, Inbred C57BL; Reperfusion Injury

The incidence of acute kidney injury (AKI) in the elderly population has steadily increased in recent years. Functional recovery after AKI is also impaired in the elderly; however, the mechanism underlying these age-related differences is not well understood. In the present study, we assessed kidney morphology, function, and oxidative stress in young and aged rats after renal ischemia and reperfusion.. Young (6- to 7-wk-old) and aged (60- to 65-wk-old) male Wistar rats were divided into four groups based on age and treatment: renal ischemia-reperfusion in young rats (young IR); renal ischemia-reperfusion in aged rats (aged IR); sham treatment in young rats (young control), and sham treatment in aged rats (aged control). Rats were sacrificed 24 h after treatment, serum blood urea nitrogen (BUN) and creatinine (Cre) concentrations were determined, and kidney tissue histology and 8-hydroxydeoxyguanosine (8-OHdG) levels were evaluated.. After ischemia-reperfusion, serum BUN, and Cre levels were higher in aged rats than in young rats. Reperfusion-induced kidney damage and kidney tissue 8-OHdG levels were also more severe in the aged IR group. Moreover, plasma antioxidant potential was lower in aged IR rats than in young IR rats.. Aged rats exhibited reduced antioxidant potential and increased oxidative stress after ischemia-reperfusion. Our findings demonstrate that aged rats experience more severe reperfusion-induced injuries compared with young rats.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Acute Kidney Injury; Aging; Animals; Blood Urea Nitrogen; Creatinine; Deoxyguanosine; Kidney; Male; Models, Animal; Oxidative Stress; Rats; Rats, Wistar; Reperfusion Injury

Acute kidney injury (AKI) is common in the intensive care unit, and one of its primary causes is renal ischemia-reperfusion (I/R) injury. Human atrial natriuretic peptide (hANP) exerts various pharmacologic effects, including renal protection. In the present study, we evaluated the renal protective effect of hANP in a rat model of renal I/R.. Male Wistar rats were divided into three groups that received the following treatments: induction of renal I/R (I/R group); continuous intravenous injection of hANP followed 30 min later by induction of renal I/R (hANP+I/R group); and sham treatment (control group). Rats were sacrificed after 60 min of ischemia and 24 h of reperfusion or sham treatment. To evaluate the renal protective effects if hANP, serum blood urea nitrogen (BUN) and creatinine (Cre) concentrations were determined, kidneys were histologically assessed, and serum biomarkers of oxidative stress were evaluated. In addition, antimycin A (AMA)-stimulated RAW264.7 cells were treated with hANP to assess its antioxidant effects.. Serum BUN and Cre levels were elevated in the I/R group; however, these increases were significantly inhibited in the hANP + I/R group. Similarly, kidney tissue damage observed in the I/R group was attenuated in the hANP + I/R group. In vitro, AMA-stimulated cells treated with hANP showed reduced reactive oxygen species activity compared to cells treated with AMA alone.. Our findings indicate that hANP may be effective in the treatment of various types of I/R injuries.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Acute Kidney Injury; Animals; Atrial Natriuretic Factor; Biomarkers; Blood Urea Nitrogen; Cell Line; Deoxyguanosine; Humans; Hydroxyl Radical; Kidney; Male; Oxidative Stress; Rats; Rats, Wistar; Reactive Oxygen Species; Reperfusion Injury

Activation of the NADPH oxidase subunit, NOX2, and increased oxidative stress are associated with neuronal death after cerebral ischemia and reperfusion. Inhibition of NOX2 by casein kinase 2 (CK2) leads to neuronal survival, but the mechanism is unknown. In this study, we show that in copper/zinc-superoxide dismutase transgenic (SOD1 Tg) mice, degradation of CK2α and CK2α' and dephosphorylation of CK2β against oxidative stress were markedly reduced compared with wild-type (WT) mice that underwent middle cerebral artery occlusion. Inhibition of CK2 pharmacologically or by ischemic reperfusion facilitated accumulation of poly(ADP-ribose) polymers, the translocation of apoptosis-inducing factor (AIF), and cytochrome c release from mitochondria after ischemic injury. The eventual enhancement of CK2 inhibition under ischemic injury strongly increased 8-hydroxy-2'-deoxyguanosine and phosphorylation of H2A.X. Furthermore, CK2 inhibition by tetrabromocinnamic acid (TBCA) in SOD1 Tg and gp91 knockout (KO) mice after ischemia reperfusion induced less release of AIF and cytochrome c than in TBCA-treated WT mice. Inhibition of CK2 in gp91 KO mice subjected to ischemia reperfusion did not increase brain infarction compared with TBCA-treated WT mice. These results strongly suggest that NOX2 activation releases reactive oxygen species after CK2 inhibition, triggering release of apoptogenic factors from mitochondria and inducing DNA damage after ischemic brain injury.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Apoptosis Inducing Factor; Brain Infarction; Casein Kinase II; Cell Death; Cells, Cultured; Cinnamates; Cytochromes c; Deoxyguanine Nucleotides; DNA Damage; Enzyme Activation; Female; Histones; Male; Membrane Glycoproteins; Mice; Mice, Knockout; Mitochondria; NADPH Oxidase 2; NADPH Oxidases; Nerve Tissue Proteins; Phosphorylation; Reperfusion Injury; Superoxide Dismutase; Superoxide Dismutase-1

Global cerebral ischemia and reperfusion (I/R) often result in high mortality. Free radicals have been reported to play an important role in global cerebral I/R, and therefore, reduction of these might improve the outcome. Here, we investigated the effect of hydrogen gas (H2) (a strong free radical scavenger) on the survival rate of mice following global cerebral I/R. We further examined the histopathological outcome and also the brain water content (as a possible determinant of mortality). Male C57BL/6J mice were subjected to global cerebral I/R by means of 45-min bilateral common carotid artery occlusion (BCCAO). A total of 160 mice were divided into three groups: sham surgery (sham group), BCCAO without H2 (BCCAO group), and BCCAO treated with 1.3% H2 (BCCAO + H2 group). We observed that H2 treatment significantly (P = 0.0232) improved the 7-day survival rate of mice, from 8.3% (BCCAO group, n = 12) to 50% (BCCAO + H2 group, n = 10). Histopathological analysis revealed that H2 treatment significantly attenuated neuronal injury and autophagy in the hippocampal cornu ammonis 1 sector and also brain edema, after 24 h of reperfusion. The beneficial effects of H2 treatment on brain injury were associated with significantly lower levels of oxidative stress markers (8-hydroxy-2'-deoxyguanosine and malondialdehyde) in the brain tissue. Thus, we believe that H2 may be an effective treatment for global cerebral I/R.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Administration, Inhalation; Animals; Body Water; Brain; Brain Ischemia; Capillary Permeability; Cerebrovascular Circulation; Deoxyguanosine; DNA Damage; Free Radical Scavengers; Hippocampus; Hydrogen; Lipid Peroxidation; Male; Malondialdehyde; Mice; Mice, Inbred C57BL; Microtubule-Associated Proteins; Reperfusion Injury; Serum Albumin; Survival Rate

Protective effects of saturated hydrogen (H(2)) saline on cardiac ischaemia-reperfusion (I/R) injury have been demonstrated previously. This study was designed to show that hydrogen-rich saline is protective in preventing lung I/R injury in rats.. Adult male Sprague-Dawley rats underwent 45 min occlusion of the right lung roots and 120 min reperfusion. Rats were divided randomly into three groups: sham-operated control group, I/R plus saline treatment, and I/R plus hydrogen-rich saline treatment (0.6 mmol/L, 0.5 ml/kg/d). Three days of intraperitoneal injection of hydrogen-rich saline before the reperfusion combined with immediate administration of hydrogen-rich saline after the reperfusion were performed. Following reperfusion, the lung tissue and the pulmonary artery was immediately obtained and the W/D ratio, pulmonary artery contraction and relaxation ability, H-E staining, TUNEL staining, caspase-3, MDA, 8-OHdG content and measurement of such biomarkers as WBC, CRP were measured or carried out.. Hydrogen saline significantly protected vasoactivity of the pulmonary artery, reduced pulmonary oedema, decreased lung malondialdehyde (MDA), 8-OHdG concentration, alleviated lung epithelial cell apoptosis and lowered the level of such biomarkers as WBC, CRP, ALT and TBiL.. It is concluded that hydrogen-rich saline is a novel, simple, safe and effective method to attenuate pulmonary I/R injury.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Acute Lung Injury; Animals; Biomarkers; Deoxyguanosine; Hydrogen; Male; Malondialdehyde; Pulmonary Artery; Pulmonary Edema; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Respiratory Mucosa; Sodium Chloride

The present study investigates the anti-oxidative effects of D-allose on ischemic damage. Rats were subjected to transient middle cerebral artery occlusion (MCAO) for 1 h under pentobarbital anesthesia. D-allose was intravenously infused during occlusion and a further 1 h after reperfusion (400 mg/kg). The effects of D-allose on focal cerebral ischemia were examined by measuring brain damage (infarction and atrophy volume) and behavioral deficits 7 days after MCAO. In another set of rats, apurnic/apyrimidic abasic sites (AP-sites) and 8-hydroxy-2'-deoxyguanosine (8-OHdG), oxidative stress markers, were investigated 24 h after MCAO to examine the anti-oxidative effects of D-allose. Brain damage and behavioral deficits were significantly decreased by D-allose administration compared to vehicle. The number of AP-sites and 8-OHdG levels were also reduced by D-allose. Thus, the present study suggests that D-allose has anti-oxidative effects and induces neuroprotection in focal cerebral ischemia.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Antioxidants; Behavior, Animal; Brain Infarction; Deoxyguanosine; Disease Models, Animal; Glucose; Infarction, Middle Cerebral Artery; Male; Motor Activity; Neurologic Examination; Oxidative Stress; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Time Factors

Recently it has been demonstrated that hydrogen, as a novel antioxidant, can selectively reduce hydroxyl radicals (·OH) and peroxynitrite anion (ONOO-) in vitro and exert therapeutic antioxidant activity in many diseases. This study was designed to investigate the effect of hydrogen-rich saline on renal ischemia/reperfusion (I/R) injury in rats.. A rat model of renal I/R injury was induced by 45-min occlusion of the bilateral renal pedicles and 24-h reperfusion. Physiologic saline, hydrogen-rich saline, or nitrogen-rich saline (8 mL/kg) were administered intraperitoneally at 5 min before reperfusion, respectively.. After I/R injury, serum blood urea nitrogen (BUN), creatinine (Cr), tissue malondialdehyde (MDA), 8-hydroxydeoxyguanosine (8-OhdG), TNF-α, IL-1β, IL-6 levels, and myeloperoxidase (MPO) activity were all increased significantly, while tissue superoxide dismutase (SOD) and catalase (CAT) activities were all decreased significantly. Hydrogen-rich saline reversed these changes and relieved morphological renal injury and I/R-induced apoptosis, while no significant changes were observed in the nitrogen-rich saline-treated group compared with physiologic saline-treated group.. Hydrogen-rich saline is able to attenuate the renal I/R injury, which is possibly by reduction of oxidative stress and inflammation.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Cytokines; Deoxyguanosine; Disease Models, Animal; Hydrogen; Kidney; Male; Malondialdehyde; Neutrophil Infiltration; Oxidative Stress; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Sodium Chloride

Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a hypoxia-inducible neuroprotective protein that also stimulates proliferation of neuronal precursor cells. In this study, we investigated the possible role of HB-EGF in ischemia and reperfusion injury by measuring the changes in its mRNA expression following focal cerebral ischemia. We also examined neural damage after a middle cerebral artery occlusion (MCAO) and reperfusion in ventral forebrain specific HB-EGF knockout (KO) mice. The levels of HB-EGF mRNA in the cerebral cortex of wild-type (WT) mice were significantly increased 3-24 h after MCAO and reperfusion. Cerebral infraction in HB-EGF KO mice was aggravated at 1 day and 6 days after MCAO and reperfusion compared with WT mice. The number of terminal deoxynucleotidyl transferase (TdT)-mediated dNTP nick end labeling (TUNEL) and an oxidative stress marker, 8-hydroxy-2'-deoxyguanosine (8-OHdG) positive cells, were higher in HB-EGF KO mice than in WT mice. On the other hand, fewer bromodeoxyuridine (BrdU) positive cells were found in the subventricular zone in HB-EGF KO mice compared with WT mice. These results indicate that HB-EGF may play a pivotal role in ischemia and reperfusion injury and that endogenously synthesized HB-EGF is necessary for both the neuroprotective effect and for regulation of cell proliferation in the subventricular zone.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Adult Stem Cells; Analysis of Variance; Animals; Brain Infarction; Bromodeoxyuridine; Cerebral Ventricles; Deoxyguanosine; Disease Models, Animal; Epidermal Growth Factor; Gene Expression Regulation; Heparin-binding EGF-like Growth Factor; In Situ Nick-End Labeling; Infarction, Middle Cerebral Artery; Intercellular Signaling Peptides and Proteins; Mice; Mice, Knockout; Prosencephalon; Reperfusion Injury; RNA, Messenger; Transforming Growth Factor alpha

The angiotensin II type 2 (AT(2)) receptor is expressed in bone marrow cells and may affect cell differentiation. We previously reported a beneficial role of the AT(2) receptor in ischemic brain damage. Here, we investigated the effect of AT(2) receptor stimulation in hematopoietic cells on ischemic brain injury using chimeric mice. Chimeric mice were generated by bone marrow transplantation into wild-type mice after irradiation. Bone marrow cells were prepared from wild-type (Agtr2(+)) or AT(2) receptor-deficient mice (Agtr2(-)). Six weeks after bone marrow transplantation, these chimeric mice were subjected to ischemia/reperfusion injury. Both Agtr2(+) and Agtr2(-) chimeric mice did not show a significant change in systolic and diastolic blood pressures, whereas body weight decreased in Agtr2(-) chimera. Twenty-four hours after ischemia/reperfusion injury, ischemic brain damage in Agtr2(-) chimera was exaggerated compared with that in Agtr2(+) chimera. Moreover, cerebral blood flow in the peripheral region before and after ischemia/reperfusion injury was decreased in Agtr2(-) chimera. The inflammatory response in the ipsilateral hemisphere was not significantly different, whereas tumor necrosis factor-α and monocyte chemoattractant protein 1 expressions tended to increase in the Agtr2(-) chimeric brain. Expression of methylmethane sulfonate 2, which has a neuroprotective effect, was lower in the brain of Agtr2(-) chimera. These results indicate that deletion of AT(2) receptor in blood cells has a harmful effect on ischemic brain injury.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Blood Pressure; Body Weight; Bone Marrow Cells; Bone Marrow Transplantation; Brain; Brain Ischemia; Cerebrovascular Circulation; Chemokine CCL2; Deoxyguanosine; Enzyme-Linked Immunosorbent Assay; Female; Gene Expression; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Receptor, Angiotensin, Type 2; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; Transplantation Chimera; Tumor Necrosis Factor-alpha; Ubiquitin-Conjugating Enzymes

Retinal ischemia/reperfusion (I/R) injury causes profound tissue damage, especially retinal ganglion cell (RGC) death. The aims of the study were to investigate whether catalase (CAT) has a neuroprotective effect on RGC after I/R injury in rats, and to determine the possible antioxidant mechanism. Wistar female rats were randonmized into four groups: normal control group (Control group), retinal I/R with vehicle group (I/R with vehicle group), retinal I/R with AAV-CAT group (I/R with AAV-CAT group), and normal retina with AAV-CAT group (normal with AAV-CAT group). One eye of each rat was pretreated with recombinant adeno-associated virus containing catalase gene (I/R with AAV-CAT group or normal with AAV-CAT group) and recombinant adeno-associated virus containing GFP gene (I/R with vehicle group) by intravitreal injection 21 days before initiation of I/R injury. Retinal I/R injury was induced by elevating intraocular pressure to 100mmHg for 1h. The number of RGC and inner plexiform layer (IPL) thickness were measured by fluorogold retrograde labeling and hematoxylin and eosin staining at 6h, 24h, 72 h and 5d after injury. Hydrogen peroxide (H(2)O(2)), the number of RGC, IPL thickness, malondialdehyde(MDA), 8-hydroxy-2-deoxyguanosine (8-OHdG), CAT activity and nitrotyrosine were measured by fluorescence staining, immunohistochemistry and enzyme-linked immunosorbent assay analysis at 5 days after injury. Electroretinographic (ERG) evaluation was also used. Pretreatment of AAV-CAT significantly decreased the levels of H(2)O(2), MDA, 8-OHdG and nitrotyrosine, increased the catalase activity, and prevented the reduction of a- and b- waves in the I/R with AAV-CAT group compare with the I/R with vehicle group (p<0.01). Catalase attenuated the I/R-induced damage of RGC and IPL and retinal function. Therefore, catalase can protect the rat retina from I/R-induced injury by enhancing the antioxidative ability and reducing oxidative stress, which suggests that catalase may be relevant for the neuroprotection of inner retina from I/R-related diseases.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Antioxidants; Apoptosis; Catalase; Cell Count; Cell Survival; Deoxyguanosine; Dependovirus; Electroretinography; Enzyme-Linked Immunosorbent Assay; Female; Genetic Therapy; Genetic Vectors; Immunohistochemistry; Malondialdehyde; Neuroprotective Agents; Rats; Rats, Wistar; Reperfusion Injury; Retina; Retinal Diseases; Retinal Ganglion Cells; Tyrosine

The use of non-heart-beating donors (NHBD) has come into practice to resolve the shortage of donor lungs. This study investigated whether hypothermic machine perfusion (HMP) can improve the quality of NHBD lungs.. An uncontrolled NHBD model was achieved in male Lewis rats. Ninety minutes after cardiac arrest, HMP was performed for 60 min at 6°C to 10°C. The first study investigated the physiological lung functions during HMP and the lung tissue energy levels before and after HMP. The second study divided the rats into three groups (n=6 each): no ischemia group; 90-min warm ischemia+60-min HMP+120-min static cold storage (SCS) (HMP group); and 90-min warm ischemia+180-min SCS group. All lungs were reperfused for 60 min at 37°C. Lung functions were evaluated at given timings throughout the experiments. Oxidative damage during reperfusion was evaluated immunohistochemically with a monoclonal antibody against 8-hydroxy-2'-deoxyguanosine.. The first study revealed that lung functions were stable during HMP. Lung tissue energy levels decreased during warm ischemia but were significantly increased by HMP (P<0.05). The second study confirmed that HMP significantly decreased pulmonary vascular resistance, increased pulmonary compliance, and improved pulmonary oxygenation. The ratio of 8-hydroxy-2'-deoxyguanosine positive cells to total cells significantly increased in the SCS group (P<0.01).. Short-term HMP improved lung tissue energy levels that decreased during warm ischemia and ameliorated ischemia-reperfusion injury with decreased production of reactive oxygen species.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Deoxyguanosine; Hypothermia, Induced; Lung; Lung Transplantation; Male; Nitroglycerin; Organ Preservation; Rats; Rats, Inbred Lew; Reperfusion Injury; Tissue Donors

Retinal ischemia-reperfusion (I/R) injury by transient elevation of intraocular pressure (IOP) is known to induce neuronal damage through the generation of reactive oxygen species. Study results have indicated that molecular hydrogen (H(2)) is an efficient antioxidant gas that selectively reduces the hydroxyl radical (*OH) and suppresses oxidative stress-induced injury in several organs. This study was conducted to explore the neuroprotective effect of H(2)-loaded eye drops on retinal I/R injury.. Retinal ischemia was induced in rats by raising IOP for 60 minutes. H(2)-loaded eye drops were prepared by dissolving H(2) gas into a saline to saturated level and administered to the ocular surface continuously during the ischemia and/or reperfusion periods. One day after I/R injury, apoptotic cells in the retina were quantified, and oxidative stress was evaluated by markers such as 4-hydroxynonenal and 8-hydroxy-2-deoxyguanosine. Seven days after I/R injury, retinal damage was quantified by measuring the thickness of the retina.. When H(2)-loaded eye drops were continuously administered, H(2) concentration in the vitreous body immediately increased and I/R-induced *OH level decreased. The drops reduced the number of retinal apoptotic and oxidative stress marker-positive cells and prevented retinal thinning with an accompanying activation of Müller glia, astrocytes, and microglia. The drops improved the recovery of retinal thickness by >70%.. H(2) has no known toxic effects on the human body. Thus, the results suggest that H(2)-loaded eye drops are a highly useful neuroprotective and antioxidative therapeutic treatment for acute retinal I/R injury.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Aldehydes; Animals; Apoptosis; Biomarkers; Deoxyguanosine; Diffusion; Disease Models, Animal; Hydrogen; Hydroxyl Radical; Immunoenzyme Techniques; In Situ Nick-End Labeling; Male; Microscopy, Confocal; Neuroglia; Ophthalmic Solutions; Oxidative Stress; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Retinal Diseases; Retinal Neurons; Vitreous Body

Renal ischemia-reperfusion (I/R), an important cause of acute kidney injury, is unavoidable during various types of operations, including renal transplantation, surgical revascularization of the renal artery, partial nephrectomy, and treatment of suprarenal aortic aneurysms. Exacerbation of I/R injury is mediated by reactive oxygen species (ROS). A recent study has shown that hydrogen has antioxidant properties. In this study, we tested the hypothesis that a hydrogen-rich saline solution (HRSS) attenuates renal I/R injury in a rodent model.. Rats were treated with an intravenous injection of HRSS or control saline solution followed by renal I/R. After 24 h of treatment, we performed a histological examination and transmission electron microscopy, and measured serum levels of 8-OHdG.. Histological analysis revealed a marked reduction of interstitial congestion, edema, inflammation, and hemorrhage in renal tissue harvested 24 h after HRSS treatment compared to saline administration. Renal I/R injury, which led to altered mitochondrial morphology, was also inhibited by HRSS. Furthermore, serum 8-OHdG levels were significantly lower in rats treated with HRSS and subjected to renal I/R.. These protective effects were likely due to the antioxidant properties of HRSS. These results suggest that HRSS is a potential therapeutic candidate for treating various I/R diseases.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Deoxyguanosine; Hydrogen; Kidney; Male; Rats; Rats, Wistar; Reperfusion Injury; Sodium Chloride

Renal ischemia-reperfusion injury (IRI), leading to acute kidney injury, is a frequent complication with renal transplantation and it is associated with graft function. Its pathogenesis involves ischemia, vascular congestion and reactive oxygen metabolites. Carvedilol is an antihypertensive drug with potent anti-oxidant properties. In this study we investigated the protective effects of carvedilol in a rat renal IRI model.. Twenty-four rats were randomized into sham, untreated control and carvedilol (2 mg/kg 30 min before surgery and 12 hr after reperfusion) treatment groups and were subjected to 60 min of left renal ischemia followed by reperfusion at 24, 48, 96 and 168 hr.. Treatment with carvedilol significantly decreased plasma creatinine levels after IRI (up to 168 hr) compared to controls (P < 0.001), suggesting an improvement in renal function. Histopathological analysis revealed decreased IRI-induced damage in kidneys from carvedilol-treated rats. A significant increase in the expression levels of Cu/Zn superoxide dismutase and reduction of 8-hydroxydeoxyguanosine and apoptosis levels (P < 0.005) suggested a protective effect after treatment with carvedilol.. Our findings suggest that carvedilol ameliorates IRI resulting in improved renal function.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Antihypertensive Agents; Apoptosis; Carbazoles; Carvedilol; Creatinine; Deoxyguanosine; Disease Models, Animal; Epithelial Cells; Kidney Tubules; Male; Oxidative Stress; Propanolamines; Rats; Rats, Sprague-Dawley; Renal Insufficiency; Reperfusion Injury; Superoxide Dismutase

Metallothioneins (MTs) are metal-binding proteins and have four isoforms. MT-III was, at first, found in the brains of patients with Alzheimer's disease. MT-III exists mainly in the central nervous system, and the main effects are thought to be anti-oxidative and regulate zinc levels. In some previous reports, MT-III exhibited neuroprotective effects in various pathological situations, but its detailed effects are still unclear. In the present study, we examined neuronal damage after a middle cerebral artery occlusion (MCAO) in MT-III knockout (KO) mice to elucidate the relationship between MT-III and cerebral infarction. There was no significant difference in cerebral infarction after 24-h permanent MCAO between the wild-type and MT-III KO mice. On the other hand, after 2-h MCAO and 22-h reperfusion, cerebral infarction in the MT-III KO mice was aggravated compared with the wild-type mice. Furthermore, fatal rate of MT-III KO mice increased from 3 days after MCAO, and neurological deficits at 5 and 7 days after MCAO of MT-III KO mice were worse than those of wild-type. We examined terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining and the immunostaining of an oxidative stress marker, 8-hydroxy-2'-deoxyguanosine (8-OHdG), at 24 h after transient MCAO. In the penumbra lesion, the positive cell numbers in both staining assays were higher in the MT-III KO mice than those of the wild-type mice. These findings indicate that neuronal damage was aggravated by reperfusion injury in the MT-III KO mice compared with the wild-type mice, suggesting that MT-III plays anti-oxidative and neuroprotective roles in transient cerebral ischemia.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Brain; Cell Count; Deoxyguanosine; Female; In Situ Nick-End Labeling; Infarction, Middle Cerebral Artery; Male; Metallothionein 3; Mice; Mice, Inbred Strains; Mice, Knockout; Nerve Tissue Proteins; Neurons; Random Allocation; Reperfusion Injury; Time Factors

We have previously demonstrated that melatonin protects against ischemia/reperfusion-induced oxidative damage to mitochondria in the fetal rat brain. The purpose of the present study was to evaluate the effects of maternally administered melatonin on ischemia/reperfusion-induced oxidative placental damage and fetal growth restriction in rats. The utero-ovarian arteries were occluded bilaterally for 30 min in rats on day 16 of pregnancy to induce fetal ischemia. Reperfusion was achieved by releasing the occlusion and restoring circulation. Melatonin solution (20 microg/mL) or the vehicle alone was administered orally during pregnancy. A sham operation was performed in control rats, which were treated with vehicle alone. Laparotomy was performed on day 20 of pregnancy and the number and weight of fetal rats and placentas were measured. Placental mitochondrial respiratory control index (RCI), a marker of mitochondrial respiratory activity, was also calculated for each group. Using immunohistochemistry, we investigated the degree of immunostaining of 8-hydroxy-2-deoxyguanosine (8-OHdG), a marker of oxidative DNA damage, and redox factor-1(ref-1), which repairs DNA damage and acts as a redox-modifying factor in rat placenta. Predictably, the ischemia/reperfusion operation significantly decreased the weight of fetal rats and placentas and the RCI. Melatonin prevented ischemia/reperfusion-induced changes in RCI (1.55 +/- 0.05 to 1.83 +/- 0.09, P < 0.05) and fetal growth (3.04 +/- 0.17 to 3.90 +/- 0.1, P < 0.0001). Immunohistochemistry revealed significant positive staining for 8-OHdG and ref-1 following ischemia/reperfusion; these effects were also reduced by melatonin treatment. Results indicated that ischemia/reperfusion-induced oxidative placental DNA and mitochondrial damage and fetal growth restriction can be prevented by maternally administered melatonin.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Deoxyguanosine; DNA Damage; DNA-(Apurinic or Apyrimidinic Site) Lyase; Female; Fetal Development; Fetal Growth Retardation; Immunohistochemistry; Melatonin; Mitochondria; Nitrosation; Oxidation-Reduction; Oxidative Stress; Placenta; Pregnancy; Rats; Reperfusion Injury

Our previous studies have shown that ferulic acid (4-hydroxy-3-methoxycinnamic acid, FA) inhibits intercellular adhesion molecule-1 (ICAM-1) expression in the ischemic striatum after 2 h of reperfusion in a transient middle cerebral artery occlusion model in rats. The purpose of this study is to further investigate the neuroprotective effects of FA during reperfusion after cerebral ischemia. Rats were subjected to 90 min of ischemia; they were then sacrificed after 2, 10, 24 and 36 h of reperfusion. ICAM-1 and macrophage-1 antigen (Mac-1) mRNA were detected using semi-quantitative RT-PCR at 2 h of reperfusion. Mac-1, 4-hydroxy-2-nonenal (4-HNE), 8-hydroxy-2'-deoxyguanosine (8-OHdG), active caspase 3, neuronal nuclei (NeuN) and TUNEL positive cells were measured at 2, 10, 24 and 36 h of reperfusion. FA (100 mg/kg, i.v.) administered immediately after MCAo inhibited ICAM-1 and Mac-1 mRNA expression in the striatum at 2 h of reperfusion, and reduced the number of Mac-1, 4-HNE and 8-OHdG positive cells in the ischemic rim and core at 10, 24 and 36 h of reperfusion. FA decreased TUNEL positive cells in the penumbra at 10 h, and in the ischemic boundary and core at 24 and 36 h of reperfusion. FA curtailed active caspase 3 expression in the penumbra at 10 h and restored NeuN-labeled neurons in the penumbra and ischemic core at 36 h of reperfusion. FA decreased the level of ICAM-1 mRNA and the number of microglia/macrophages, and subsequently down-regulated inflammation-induced oxidative stress and oxidative stress-related apoptosis, suggesting that FA provides neuroprotection against oxidative stress-related apoptosis by inhibiting ICAM-1 mRNA expression after cerebral ischemia/reperfusion injury in rats.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Aldehydes; Animals; Apoptosis; Brain Ischemia; Caspase 3; Coumaric Acids; Deoxyguanosine; Disease Models, Animal; DNA-Binding Proteins; Encephalitis; Free Radical Scavengers; Gene Expression; In Situ Nick-End Labeling; Infarction, Middle Cerebral Artery; Intercellular Adhesion Molecule-1; Macrophage-1 Antigen; Male; Nerve Tissue Proteins; Neuroprotective Agents; Nuclear Proteins; Oxidative Stress; Rats; Rats, Sprague-Dawley; Reperfusion Injury; RNA, Messenger

Apoptosis resulting from disruption of the normal cell-matrix relationship (anoikis) during islet isolation, and the reactive oxygen and nitrogen species generated following hypoxia/reoxygenation (H/R) can lead to a loss of islet tissue in culture and the reduced survival of transplanted pancreatic islets. The aim of this study was to investigate the effect of (-)-epigallocatechin-3-gallate (EGCG), a well-known antiapoptotic agent, on inhibiting anoikis and H/R injury in an in vitro islet culture system.. Islets were isolated from F344 rats and cultured under normal or H/R condition with/without EGCG.. EGCG inhibited apoptosis and lactate-dehydrogenase leakage from anoikis and H/R in a dose-dependent manner. Further, EGCG prevent increases in 8-hydroxy-2'-deoxyguanosine content and inhibited the decline of insulin secretory function induced by H/R.. These results suggest that the addition of EGCG to an islet culture system may improve the survival rate of isolated islets and reduce the loss of functional islet mass that compromises the stable reversal of diabetes after islet transplantation.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Anoikis; Antioxidants; Catechin; Cell Hypoxia; Deoxyguanosine; Insulin; Islets of Langerhans; L-Lactate Dehydrogenase; Rats; Rats, Inbred F344; Reperfusion Injury; Tea

Oxidative stress contributes to ischemia/reperfusion neuronal damage in a consecutive 2-phase pattern: an immediate direct cytotoxic effect and subsequent redox-mediated inflammatory insult. The present study was designed to assess the neuroprotective mechanisms of edaravone, a novel free radical scavenger, through antioxidative and anti-inflammatory pathways, from the early period to up to 7 days after ischemia/reperfusion in mice.. Mice were subjected to 60-minute ischemia followed by reperfusion. They were divided into the edaravone group (n=72; with different schedules for first administration) and the vehicle (control) group (n=36). Infarct volume and neurological deficit scores were evaluated at several time points after ischemia. Immunohistochemical analysis for 4-hydroxy-2-nonenal (HNE), 8-hydroxy-deoxyguanosine (8-OHdG), ionized calcium-binding adapter molecule 1 (Iba-1), inducible NO synthase (iNOS), and nitrotyrosine were performed at 24 hours, 72 hours, or 7 days after reperfusion.. Edaravone, even when administrated 6 hours after onset of ischemia/reperfusion, significantly reduced the infarct volume (68.10+/-6.24%; P<0.05) and improved the neurological deficit scores (P<0.05) at 24 hours after reperfusion. Edaravone markedly suppressed the accumulation of HNE-modified protein and 8-OHdG at the penumbra area during the early period after reperfusion (P<0.05) and reduced microglial activation, iNOS expression, and nitrotyrosine formation at the late period.. Our results indicated that edaravone exerts an early neuroprotective effect through the early free radicals scavenging pathway and a late anti-inflammatory effect and suggested that edaravone is important for expansion of the therapeutic time window in stroke patients.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Aldehydes; Animals; Anti-Inflammatory Agents; Antioxidants; Antipyrine; Brain; Brain Ischemia; Deoxyguanosine; DNA Damage; Edaravone; Electrophoresis, Polyacrylamide Gel; Free Radical Scavengers; Immunoblotting; Immunohistochemistry; Inflammation; Lipid Peroxidation; Male; Mice; Mice, Inbred C57BL; Neuroprotective Agents; Oxidative Stress; Reperfusion Injury; Time Factors; Tyrosine

The role of glycine has not been investigated in lung ischemia-reperfusion injury after cold preservation. Furthermore, the role of apoptosis after reperfusion following cold preservation has not been fully understood.. Lewis rats were divided into three groups (n=6 each). In the GLY(-) and GLY(+) groups, isolated lungs were preserved for 15 hr at 4 degrees C after a pulmonary artery (PA) flush using our previously developed preservation solution (ET-K; extracellular-type trehalose containing Kyoto), with or without the addition of glycine (5 mM). In the Fresh group, isolated lungs were reperfused immediately after a PA flush with ET-K. They were reperfused for 60 min with an ex vivo perfusion model. Pulmonary function, oxidative stress, apoptosis, and tumor necrosis factor (TNF)-alpha expression were assessed after reperfusion.. Shunt fraction and peak inspiratory pressure after reperfusion in the GLY(-) group were significantly higher than those in the GLY(+) and Fresh groups. Oxidative damage and apoptosis in the alveolar epithelial cells of the GLY(-) group, assessed by immunohistochemical staining and quantification of 8-hydroxy-2'-deoxyguanosine and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling method, were significantly higher than those of the GLY(+) and Fresh groups. There were correlations among shunt fraction, oxidative damage, and apoptosis. There was no expression of TNF-alpha messenger RNA in all groups evaluated by the reverse transcription-polymerase chain reaction.. Glycine attenuates ischemia/reperfusion injury after cold preservation by reducing oxidative damage and suppressing apoptosis independent of TNF-alpha in this model. The suppression of apoptosis might ameliorate lung function after reperfusion.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Apoptosis; Blood Pressure; Cryopreservation; Cytoprotection; Deoxyguanosine; Glycine; Immunohistochemistry; In Situ Nick-End Labeling; Inhalation; Ischemia; Lung; Male; Oxidative Stress; Pressure; Pulmonary Artery; Pulmonary Circulation; Rats; Rats, Inbred Lew; Reperfusion Injury; Staining and Labeling

Heat shock preconditioning (HSPC) is a promising strategy for providing ischemic tolerance. The objective of this study is to investigate the effectiveness of HSPC in preventing oxidative damage of cellular proteins and DNA during ischemia-reperfusion of the liver. Male Wistar rats were divided into a heat shock group (group HS) and control (group C). Forty-eight hours prior to ischemia, rats in group HS received HSPC at 42 degrees C for 15 min. All rats received hepatic warm ischemia for 30min and subsequent reperfusion. The formation of 8-hydroxy-2'-deoxyguanosine (8-OHdG), 4-hydroxy-2-nonenal (HNE) modified proteins in liver tissue, survival rate of the animals, and changes in biochemical and histological parameters were compared between groups. Heat shock protein 72 was produced only in group HS. The 7-day survival of rats was significantly better in group HS (10/10) than in group C (5/10) (p < 0.01). The serum release of alanine aminotransferase (n = 10, p < 0.01) and the concentration of adenosine triphosphate in liver tissue (n = 10, p < 0.01) 40min after reperfusion was significantly better in group HS than in group C. The formation of 8-OHdG in liver tissue measured by high-performance liquid chromatography was suppressed in group HS (p < 0.01). The production of HNE-modified proteins as determined by Western-blot analysis was also decreased in group HS. These results were also confirmed by immunohistochemical analysis. As determined by levels of 8-OHdG and HNE-modified proteins produced during ischemia-reperfusion of the liver, HSPC reduced the oxidative injury of cellular proteins and DNA in the liver tissue.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Aldehydes; Animals; Body Temperature; Deoxyguanosine; DNA Damage; Heat-Shock Proteins; Heat-Shock Response; HSP72 Heat-Shock Proteins; Hyperthermia, Induced; Liver; Male; Protein Processing, Post-Translational; Rats; Rats, Wistar; Reperfusion Injury; Survival Rate

Free radical-induced oxidative damages of macromolecules and cell death are important factors in the pathogenesis of ischemia/reperfusion brain injury. In the present study, an investigation as to whether green tea extract reduces ischemia/reperfusion-induced brain injury in Mongolian gerbils was conducted. The effect of green tea on the ischemia/reperfusion-induced production of hydrogen peroxide, lipid peroxidation and oxidative DNA damage (formation of 8-hydroxydeoxyguanosine), and cell death in addition to locomotor activity was studied. Two doses (0.5 or 2%) of green tea extract were added into the drinking water and to be accessed by animals ad libitum for 3 weeks prior to the induction of ischemia. A global ischemia was induced by the bilateral occlusion of the common carotid arteries for 5 min. Reperfusion was achieved by releasing the occlusion and restoring blood circulation for 48 h. The infarction volumes were 112+/-31 mm(3) and 76+/-11 mm(3) in the 0.5 and 2% green tea pretreated animals compared to 189+/-12 mm(3) in the ischemia/reperfusion animals. Green tea extract also reduced the levels of ischemia/reperfusion-induced hydrogen peroxide (from 1470+/-170 to 1034+/-46 and 555+/-30 nmole/mg protein), lipid peroxidation products (from 1410+/-210 to 930+/-40 and 330+/-20 nmole/mg protein) and 8-oxodG (from 3.9+/-0.1 to 2.8+/-0.3 and1.9+/-0.3 ng/microg DNA, x10(-2)) by pretreatment of 0.5 or 2% green tea for 3 weeks, respectively. Moreover, green tea also reduced the number of ischemia/reperfusion-induced apoptotic cells (from 59+/-12 to 37+/-8, 15+/-11 apoptotic cells/high power field in the striatum region) and locomotor activity (from 15140+/-2940 to 3900+/-600 and 4100+/-1200). This study therefore suggests that green tea may be a useful agent for the prevention of cerebral ischemia damage.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Aldehydes; Animals; Apoptosis; Beverages; Brain; Brain Infarction; Cerebrovascular Circulation; Cysteine Proteinase Inhibitors; Deoxyguanosine; DNA; Female; Gerbillinae; Hydrogen Peroxide; Ischemic Attack, Transient; Lipid Peroxidation; Malondialdehyde; Motor Activity; Neurons; Oxidative Stress; Plant Extracts; Reperfusion Injury

Although beneficial roles of hepatocyte growth factor (HGF) and its variants on several hepatic disorders have been reported, their effects on hepatic ischemia-reperfusion (IR) injury remain undetermined. We investigated the action of a deleted form of HGF (dHGF) on hepatic IR injury in rats.. dHGF or phosphate-buffered saline was continuously infused intravenously for 20 h prior to a 20-min occlusion of hepatic vessels. Samples were taken before and after IR, for measurement of serum dHGF and released enzymes, liver gamma-glutamylcysteinyl glycine (GSH) level, as well as histological and immunohistochemical examinations.. After reperfusion, histological injury, as well as increase in the serum activities of aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, and creatine kinase-BB were significantly attenuated in the dHGF-treated rats. dHGF maintained a high GSH level and suppressed oxidative stress and intercellular adhesion molecule-1 (ICAM-1) expression on sinusoidal endothelial cells (SECs), on which c-Met was not detected. IR caused activation of c-Met expression, which was milder in the dHGF-treated group, in hepatocytes at the pericentral region.. dHGF attenuated liver injury after IR. It also maintained a higher GSH level, depressed oxidative stress and inhibited ICAM-1 expression on c-Met negative SECs, suggesting a paracrine effect of dHGF.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Alternative Splicing; Animals; Deoxyguanosine; Glutathione; Hepatocyte Growth Factor; Humans; Immunohistochemistry; Infusions, Intravenous; Intercellular Adhesion Molecule-1; Liver; Male; Proto-Oncogene Proteins c-met; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury; Sequence Deletion

EPC-K1, L-ascorbic acid 2-[3,4-dihydro-2,5,7,8-tetramethyl-2-(4,8,12-trimethyltridecyl)-2H-1-benzopyran-6-yl-hydrogen phosphate] potassium salt, is a novel antioxidant. In this study, we investigated a reduction of oxidative neuronal cell damage with EPC-K1 by immunohistochemical analysis for 8-hydroxy-2'-deoxyguanosine (8-OHdG) in rat brain with 60 min transient middle cerebral artery occlusion, in association with terminal deoxynucleotidyl transferase-mediated dUTP-biotin in situ nick end labeling (TUNEL) and staining for total and active caspase-3. Treatment with EPC-K1 (20 mg kg(-1) i.v.) significantly reduced infarct size (p < 0.05) at 24 h of reperfusion. There were no positive cells for 8-OHdG and TUNEL in sham-operated brain, but numerous cells became positive for 8-OHdG, TUNEL and caspase-3 in the brains with ischemia. The number was markedly reduced in the EPC-K1 treated group. These reductions were particularly evident in the border zone of the infarct area, but the degree of reduction was less in caspase-3 staining than in 8-OHdG and TUNEL stainings. These results indicate EPC-K1 attenuates oxidative neuronal cell damage and prevents neuronal cell death.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Antioxidants; Apoptosis; Ascorbic Acid; Brain; Brain Infarction; Brain Ischemia; Caspase 3; Caspases; Deoxyguanosine; DNA Damage; Free Radicals; Immunohistochemistry; In Situ Nick-End Labeling; Infarction, Middle Cerebral Artery; Male; Nerve Degeneration; Neurons; Neuroprotective Agents; Oxidative Stress; Rats; Rats, Wistar; Reperfusion Injury; Vitamin E

The repair enzyme 8-oxoguanine glycosylase/ apyrimidinic/apurinic lyase (OGG) removes 8-hydroxy-2'deoxyguanosine (oh8dG) in human cells. Our goal was to examine oh8dG-removing activity in the cell nuclei of male C57BL/6 mouse brains treated with either forebrain ischemia-reperfusion (FblR) or sham operations. We found that the OGG activity in nuclear extracts, under the condition in which other nucleases did not destroy the oligodeoxynucleotide duplex, excised oh8dG with the greatest efficiency on the oligodeoxynucleotide duplex containing oh8dG/dC and with less efficiency on the heteroduplex containing oh8dG/dT, oh8dG/dG, or oh8dG/dA. This specificity was the same as for the recombinant type 1 OGG (OGG1) of humans. We observed that the OGG1 peptide and its activity in the mouse brain were significantly increased after 90 min of ischemia and 20-30 min of reperfusion. The increase in the protein level and in the activity of brain OGG1 correlated positively with the elevation of FblR-induced DNA lesions in an indicator gene (the c-fos gene) of the brain. The data suggest a possibility that the OGG1 protein may excise oh8dG in the mouse brain and that the activity of OGG1 may have a functional role in reducing oxidative gene damage in the brain after FblR.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Base Sequence; Brain; Brain Ischemia; Cell Nucleus; Deoxyguanosine; DNA; DNA Repair; DNA-Formamidopyrimidine Glycosylase; Male; Mice; Mice, Inbred C57BL; N-Glycosyl Hydrolases; Prosencephalon; Reperfusion Injury; RNA, Messenger; Up-Regulation

Eicosanoids accumulation and formation of oxygen free radicals have been implicated in the pathogenesis of ischemia/reperfusion brain injury. In the present study, we examined whether green tea extract protects against ischemia/reperfusion-induced brain injury by minimizing eicosanoid accumulation and oxygen radical-induced oxidative damage in the brain. Green tea extract (0.5%) was orally administered to Wistar rats for 3 weeks before induction of ischemia. Ischemia was induced by the occlusion of middle cerebral arteries for 60 min and reperfusion was achieved for 24 h. Infarction volume in the ipsilateral hemisphere of ischemia/reperfusion animals was 114 +/- 16 mm(3) in the 0.5% green tea pretreated animals compared to 180 +/- 54 mm(3) in left hemisphere of nontreated animals. Green tea extract (0.5%) also reduced ischemia/reperfusion-induced eicosanoid concentration: Leukotriene C(4) (from 245 +/- 51 to186 +/- 22), prostoglandin E(2) (from 306 +/- 71 to 212 +/- 43) and thromboxane A(2) (327 +/- 69 to 251 +/- 87 ng/mg protein). Ischemia/reperfusion-induced increases of hydrogen peroxide level (from 688 +/- 76 to 501 +/- 99 nmole/mg protein), lipid peroxidation products (from 1010 +/- 110 to 820 +/- 70 nmole/mg protein) and 8-oxodG formation (from 1.3 +/- 0.3 to 0.8 +/- 0.2 ng/microg DNA, x10(-2)) were also reduced. Moreover, 0.5% green tea extract also reduced the apoptotic cell number (from 44 +/- 11 to 29 +/- 1 in the striatum, and from 72 +/- 11 to 42 +/- 5 apoptotic cells/high power field in the cortex region). Green tea extract pretreatment also promoted recovery from the ischemia/reperfusion-induced inhibition of active avoidance. The present study shows that the minimizing effect of green tea extract on the eicosanoid accumulation and oxidative damage in addition to the reduction of neuronal cell death could eventually result in protective effect on the ischemia/reperfusion-induced brain injury and behavior deficit.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Antioxidants; Apoptosis; Avoidance Learning; Brain; Brain Infarction; Brain Ischemia; Deoxyguanosine; Eicosanoids; Hydrogen Peroxide; Lipid Peroxidation; Neurons; Neuroprotective Agents; Plant Extracts; Rats; Rats, Wistar; Reperfusion Injury; Tea

Ischemia-reperfusion (IR) injury is an intractable process associated not only with therapeutic recanalization of vessels, but also with partial resection or transplantation of solid organs including liver. To develop methods for predicting the degree of hepatic IR injury and further to identify injured cells, we studied the formation of 8-hydroxy-2'-deoxy-guanosine (8-OHdG) and 4-hydroxy-2-nonenal (HNE)-modified proteins in the normothermic hepatic IR model of rats using immunohistochemistry, high-performance liquid chromatography (HPLC) determination and Western blot. The Pringle maneuver for either 15 or 30 min duration produced reversible or lethal damage, respectively. The levels of both products were significantly increased in proportion to ischemia duration 40 min after reperfusion, suggesting the involvement of hydroxyl radicals. Increased immunoreactivity of 8-OHdG was observed not only in the nuclei of hepatocytes but also in those of bile canalicular and endothelial cells. However, immunoreactivity of HNE-modified proteins was detected in the cytoplasm of hepatocytes, which was confirmed by Western blot, and in addition, in the nuclei of hepatocytes after severe injury. Thus, localization of the two oxidatively modified products was not identical. Our data suggest that these two products could be used for the assessment of hepatic IR injury in tissue, but that the biological significance of the two products might be different.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Aldehydes; Animals; Blotting, Western; Body Temperature; Chromatography, High Pressure Liquid; Deoxyguanosine; DNA Adducts; DNA Damage; Hepatocytes; Hydroxyl Radical; Immunoenzyme Techniques; Ischemia; Liver; Liver Function Tests; Male; Molecular Weight; Nuclear Proteins; Oxidation-Reduction; Oxidative Stress; Proteins; Rats; Rats, Wistar; Reperfusion Injury; Specific Pathogen-Free Organisms; Time Factors

To investigate whether melatonin reduces the susceptibility of the fetal rat brain to oxidative damage of lipids and DNA, we created a model of fetal ischemia/reperfusion using rats at day 19 of pregnancy. Fetal ischemia was induced by bilateral occlusion of the utero-ovarian artery for 20 min. Reperfusion was achieved by releasing the occlusion and restoring the circulation for 30 min. A sham operation was performed in control rats. Melatonin (10 mg/kg) or vehicle was injected intraperitoneally 60 min prior to the occlusion. We measured the concentration of thiobarbituric acid reactive substances (TBARS) in fetal brain homogenates, as well as levels of deoxyguanosine (dG) and 8-hydroxydeoxyguanosine (8-OHdG) in DNA extracted from those homogenates. Ischemia for 20 min did not significantly alter the levels of dG, 8-OHdG, and TBARS. Subsequent reperfusion, however, led to a significant reduction in the dG level (P < 0.05) and to significant increases in the levels of 8-OHdG (P < 0.05) and TBARS (P < 0.05), and in the 8-OHdG/dG ratio (P < 0.005). Melatonin administration prior to ischemia significantly reduced the ischemia/reperfusion-induced increases in the levels of 8-OHdG (14.33 +/- 6.52-5.15 +/- 3.28 pmol/mg of DNA, P < 0.001) and TBARS (11.61 +/- 3.85-4.73 +/- 3.80 nmol/mg of protein, P < 0.001) as well as in the 8-OHdG/dG ratio (7.19 +/- 2.49-1.61 +/- 0.98, P < 0.001). Furthermore, melatonin significantly increased the dG level (210.19 +/- 49.02-299.33 +/- 65.08 nmol/mg of DNA, P < 0.05). Results indicate that melatonin administration to the pregnant rat may prevent the ischemia/reperfusion-induced oxidative lipid and DNA damage in fetal rat brain.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Brain; Brain Ischemia; Deoxyguanosine; DNA; DNA Damage; Female; Fetus; Lipid Metabolism; Lipid Peroxidation; Melatonin; Oxidation-Reduction; Pregnancy; Rats; Rats, Wistar; Reperfusion Injury; Thiobarbituric Acid Reactive Substances

There is much evidence to suggest that ischemic injury occurs during the reperfusion phase of ischemia-reperfusion insults, and that the injury may be due to reactive-oxygen-species (ROS)-mediated oxidative events, including lipid peroxidation and DNA damage. However, oxidative DNA damage has until now not been examined in situ. In the present study, we report for the first time observation of cell type- and region-specific oxidative DNA damages in 5 min transient ischemic model by immunohistochemical methods, using monoclonal antibody against 8-hydroxy-2'-deoxyguanosine (8-OHdG), an oxidative DNA product. The cell types containing 8-OHdG immunoreactivity were neurons, glia and endothelial cells in the hippocampus. The 8-OHdG immunoreactivity was present in the nucleus but not the cytoplasm of these cells. The level of 8-OHdG in CA1 increased significantly (P<0.05) at the end of 30 min after ischemia, but there was no increase within CA2 and CA3 areas. The 8-OHdG levels in the hippocampus increased significantly (about fourfold) after 3 h of reperfusion and remained significantly (P<0.01) elevated for at least 12 h. At 4 days after ischemia, 8-OHdG levels in the CA2 and CA3 areas decreased to levels of the sham without neuronal loss, while disappearance of 8-OHdG immunoreactivity in the CA1 coincided with neuronal death in this area. These findings strongly suggest that ischemia-induced DNA damage evolves temporally and spatially, and that oxidative DNA damage may be involved in delayed neuronal death in the CA1 region.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Deoxyguanosine; DNA Damage; Gerbillinae; Hippocampus; Immunohistochemistry; Male; Oxidation-Reduction; Prosencephalon; Reactive Oxygen Species; Reperfusion Injury

We investigated oxidative damage to the c-fos gene and to its transcription in the brain of Long-Evans rats using a transient focal cerebral ischemia and reperfusion (FCIR) model. We observed a significant (p < 0.001) increase in the immunoreactivity to 8-hydroxy-2'-guanine (oh8G) and its deoxy form (oh8dG) in the ischemic cortex at 0-30 min of reperfusion in all 27 animals treated with 15-90 min of ischemia. Treatment with a neuronal nitric oxide synthase (nNOS) inhibitor, 3-bromo-7-nitroindazole (60 mg/kg, i.p.), abolished the majority but not all of the oh8G/oh8dG immunoreactivity. Treatment with RNase A reduced the oh8G immunoreactivity, suggesting that RNA may be targeted. This observation was further supported by decreased levels of mRNA transcripts of the c-fos and actin genes in the ischemic core within 30 min of reperfusion using in situ hybridization. The reduction in mRNA transcription occurred at a time when nuclear gene damage, detected as sensitive sites to Escherichia coli Fpg protein in the transcribed strand of the c-fos gene, was increased 13-fold (p < 0.01). Our results suggest that inhibiting nNOS partially attenuates FCIR-induced oxidative damage and that nNOS or other mechanisms induce nuclear gene damage that interferes with gene transcription in the brain.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Deoxyguanosine; DNA Repair; Gene Expression Regulation; Genes, fos; Immunohistochemistry; In Situ Hybridization; Ischemic Attack, Transient; Male; Oxidative Stress; Rats; Rats, Long-Evans; Reperfusion Injury; RNA, Messenger

Nitric oxide has a clearly defined place in normal renal homoeostasis while there is a continuing debate as to its role under pathophysiological conditions. This study investigated the role of nitric oxide in a model of renal warm ischaemia-reperfusion injury.. Groups of rats underwent bilateral renal warm ischaemia (for 15-60 min) followed by reperfusion (20 or 80 min) before unilateral nephrectomy for measurement of renal nitric oxide (as nitroxides) and oxidative damage. Renal function was measured on days 2 and 7 before killing and nephrectomy. A further group received the nitric oxide synthase inhibitor N(G)-nitro L-arginine methyl ester (L-NAME; 50 mg per kg body-weight) before induction of warm ischaemia.. In early reperfusion there was a correlation between the duration of warm ischaemia (15-45 min) and renal nitrate (r2=0.97) which increased from a mean(s.e.m.) baseline value of 95(5.9) to 208(17.3) nmol per mg protein following 45 min of warm ischaemia. Levels were further raised at 80 min and maintained through to day 7 (241(12.5) nmol per mg protein in 45-min group). This rise was attenuated by L-NAME (P< 0.01) as was the early rise in oxidative damage seen otherwise. By day 7, however, oxidative damage was increased (all P< or = 0.01).. Renal nitric oxide increased early in recoverable warm ischaemia-reperfusion injury and remained raised to day 7. Nitric oxide synthase inhibition ameliorated early but exacerbated late damage suggesting that the early burst of nitric oxide is cytotoxic but that overall nitric oxide may exert a cytoprotective effect.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Deoxyguanosine; DNA Damage; Glomerular Filtration Rate; Hot Temperature; Ischemia; Kidney; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Rats; Rats, Wistar; Reperfusion Injury

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Deoxyguanosine; DNA Damage; Myocardial Ischemia; Rats; Reperfusion Injury

Oxidative damage is thought to play an important role in ischemia/reperfusion injury, including the outcome of transplantation of the liver and intestine. We have investigated oxidative DNA damage after combined transplantation of the liver and small intestine in 5 pigs. DNA damage was estimated from the urinary excretion of the repair product 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG). In the first 1-3 hr after reperfusion of the grafts, 8-oxodG excretion was increased 2.9-fold (1.7-4.1; 95% confidence intervals; P < 0.05). A control experiment included sham surgery with clamping of the suprarenal inferior caval vein in 2 pigs during steady state infusion of 8-oxodG. While the caval vein was clamped, the urinary excretion of 8-oxodG was almost blocked, whereas after removal of the clamp, the excretion returned to and did not exceed the preclamp levels. In a separate experiment with 2 pigs, the elimination of injected 8-oxodG was shown to adhere to first-order kinetics with a clearance and a terminal elimination half-life of approximately 4 ml min-1 kg-1 and 2 1/2 hr, respectively. The injected dose was completely excreted into the urine within 4 hr. It is concluded that substantial oxidative damage to DNA results from reperfusion of transplanted small intestine and liver in pigs, as estimated from the readily excreted repair product 8-oxodG.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Deoxyguanosine; DNA Damage; Intestine, Small; Liver; Liver Transplantation; Oxidative Stress; Reperfusion Injury; Swine