Changes of superoxide dismutase in cultured rat aortic smooth muscle cells (A7r5) by an incubation of vitamin E.

Supplementation of antioxidants such as vitamin E and vitamin C as health promotion food is popular recently. Epidemiological studies supported the beneficial effect of these antioxidants because oxygen free radicals have been linked to the process of diseases and aging. The present study evaluated the effect of alpha-tocopherol (vitamin E) on the changes of superoxide dismutase (SOD) in cultured rat aortic smooth muscle cells (A7r5) after a short-term (2 days) or long-term (7 days) incubation. Incubation of A7r5 cells with vitamin E at a concentration of 50 micromol/l for 2 days caused an increase of both the activity and mRNA level of SOD. At higher concentrations, such as 100 or 200 micromol/l, vitamin E failed to enhance SOD more effectively. However, after incubation for 7 days, vitamin E caused a decrease in both the activity and mRNA level of SOD in a concentration-dependent manner. Otherwise, the protein amount of SOD remained the same in these samples regardless of the concentration of vitamin E or the duration of incubation. The obtained results suggest that vitamin E can increase the effect of SOD to result in the beneficial influence of this antioxidant only at low concentration under a short-term supplementation because a down-regulation of SOD was observed in cells receiving a long-term incubation.     

Downregulation of superoxide dismutase activity and gene expression in cultured rat brain astrocytes after incubation with vitamin C

Reactive oxygen species have been linked with neuropathological changes in the central nervous system. Epidemiological studies supported the beneficial effect of supplementation of antioxidants. Superoxide dismutase (SOD) is an endogenous enzyme which can scavenge reactive oxygen species. This study investigated the effect of supplementation with ascorbic acid (vitamin C) on the changes of SOD in cultured neurological cells. Rat brain astrocytes (RBA-1 cells) were incubated with vitamin C and divided into four groups: a control group (without vitamin C) and three treatment groups with vitamin C at 40, 80, and 160 micromol/l. After short-term (2 days) and long-term (7 days) incubation, SOD activity, SOD mRNA level by Northern blotting, and SOD protein amounts by Western blotting were measured. After 2 days of incubation, vitamin C resulted in a decrease in the activity of SOD in a concentration-dependent manner (Mn-SOD from 14.8 +/- 1.2 to 13.2 +/- 0.5 U/mg protein and Cu/Zn-SOD from 64.8 +/- 1.2 to 51.7 +/- 0.9 U/mg protein; p < 0.05), and vitamin C also attenuated the Cu/Zn-SOD mRNA level from 100 to 86.3 +/- 6.7%; p < 0.01), whereas the protein amounts of these two SODs remained unchanged. After 7 days of incubation with vitamin C, the SOD activity of RBA-1 cells decreased significantly (Mn-SOD from 14.9 +/- 0.3 to 11.8 +/- 0.3 U/mg protein and Cu/Zn SOD from 61.8 +/- 1.8 to 54.6 +/- 0.9 U/mg protein; p < 0.01), and the mRNA level was also attenuated (Mn-SOD from 100 to 86.8 +/- 8.7% and Cu/Zn-SOD from 100 to 84.7 +/- 4.8%; p < 0.01). These results suggest that 2 and 7 days of incubation with relatively high concentrations of vitamin C may downregulate activity and gene expression of SOD in cultured RBA-1 cells.     

Superoxide dismutase: an emerging target for cancer therapeutics

Superoxide dismutase (SOD) is a critical enzyme responsible for the elimination of superoxide radicals and is considered to be a key anti-oxidant in aerobic cells. Cellular consumption of oxygen is essential for oxidative phosphorylation during ATP generation in the mitochondria, yet this cellular metabolism also leads to the production of reactive oxygen species (ROS), including the superoxide radical (O(2)(*)(-)) and hydrogen peroxide (H(2)O(2)). Accumulation of ROS results in cellular oxidative stress and, if not corrected, can lead to the damage of important biomolecules such as membrane lipids, proteins and DNA. Prolonged accumulation of high levels of free radicals in cells may cause irreversible cellular injury and ultimately result in cell death. Since SOD is the key enzyme in the first metabolic step of superoxide elimination, deficiency in SOD or inhibition of the enzyme activity may cause severe accumulation of O(2)(*)(-) in cells and lead to cell death. Thus, inhibition of SOD may provide a novel way to kill cancer cells. Due to dysfunction in the regulation of cell growth, cancer cells are active in energy metabolism, and thus produce high levels of O(2)(*)(-) and other ROS and are under constant oxidative stress. This may render the malignant cells more dependent on SOD to eliminate the toxic superoxide radicals and thus potentially more sensitive to SOD inhibitors. It is a plausible hypothesis that inhibition of SOD may preferentially kill malignant cells through a free radical-mediated mechanism. This article will review evidence that suggests SOD as an emerging therapeutic target for cancer treatment. The relevant clinical implications and potential risk will also be discussed.     

Na+/Ca2+ exchanger inhibitor ameliorates impaired endothelium-dependent Na+ relaxation induced by high glucose in rat aorta

The present study was designed to investigate the effects of KB-R7943, an inhibitor of the Na+/Ca2+ exchanger, on impaired endothelium-dependent relaxation (EDR) induced by high glucose in rat isolated aorta. Both acetylcholine (ACh)-induced EDR and sodium nitroprusside (SNP)-induced endothelium-independent relaxation (EIR) were measured after aortic rings had been exposed to high glucose in the absence and presence of KB-R7943. Coincubation of aortic rings with high glucose (25 mmol/L) for 24 h resulted in a significant inhibition of EDR, but had no effect on EIR. After incubation of aortic rings in the presence of both KB-R7943 (0.1-10 micromol/L) and high glucose for 24 h, significantly attenuation of impaired EDR was observed. This protective effect of KB-R7943 (10 micromol/L) was abolished by superoxide dismutase (SOD; 200 U/mL) and l-arginine (3 mmol/L), whereas d-arginine (3 mmol/L) had no effect. Similarly, high glucose decreased SOD activity and the release of nitric oxide (NO) and increased superoxide anion (O2(-)) production in aortic tissue. KB-R7943 significantly decreased O2(-) production and increased SOD activity and NO release. These results suggest that KB-R7943 can restore impaired EDR induced by high glucose in rat isolated aorta, which may be related to the scavenging of oxygen free radicals and enhanced NO production.     

补虚化瘀祛痰饮对老年冠心病的抗氧化作用机制研究

目的:在论证老年冠心病中西医基本病机的基础上,研究补虚化瘀祛痰饮组方构成的抗氧化作用机制。方法:将66例老年冠心病患者随机分为中药干预组和对照组,2组常规治疗方法相同,中药干预组另加用补虚化瘀祛痰饮。采用生化及分子生物学手段测定治疗前、后超氧化物歧化酶(SOD)活性、血浆脂质过氧化物(LPO)含量以及外周血单个核细胞MnSOD基因mRNA的表达水平。结果:对照组患者治疗前、后血浆SOD活性及LPO含量无显著性差异(P>0.05),中药干预组患者治疗后血浆SOD活性显著升高,LPO含量显著降低(P<0.01);中药干预组MnSOD基因mRNA表达水平显著高于对照组(P<0.01)。结论:补虚化瘀祛痰饮能诱导单个核细胞MnSOD基因表达,干预氧自由基的产生,并增强消除氧自由基的功能。     

Superoxide dismutase: an emerging target for cancer therapeutics

Superoxide dismutase (SOD) is a critical enzyme responsible for the elimination of superoxide radicals and is considered to be a key anti-oxidant in aerobic cells. Cellular consumption of oxygen is essential for oxidative phosphorylation during ATP generation in the mitochondria, yet this cellular metabolism also leads to the production of reactive oxygen species (ROS), including the superoxide radical (O₂ ^{? -}) and hydrogen peroxide (H₂O₂). Accumulation of ROS results in cellular oxidative stress and, if not corrected, can lead to the damage of important biomolecules such as membrane lipids, proteins and DNA. Prolonged accumulation of high levels of free radicals in cells may cause irreversible cellular injury and ultimately result in cell death. Since SOD is the key enzyme in the first metabolic step of superoxide elimination, deficiency in SOD or inhibition of the enzyme activity may cause severe accumulation of O₂ ^{? -} in cells and lead to cell death. Thus, inhibition of SOD may provide a novel way to kill cancer cells. Due to dysfunction in the regulation of cell growth, cancer cells are active in energy metabolism, and thus produce high levels of O₂ ^{? -} and other ROS and are under constant oxidative stress. This may render the malignant cells more dependent on SOD to eliminate the toxic superoxide radicals and thus potentially more sensitive to SOD inhibitors. It is a plausible hypothesis that inhibition of SOD may preferentially kill malignant cells through a free radical-mediated mechanism. This article will review evidence that suggests SOD as an emerging therapeutic target for cancer treatment. The relevant clinical implications and potential risk will also be discussed.     

Effect of GCP-02, a PPARalpha/gamma dual activator, on glucose and lipid metabolism in insulin-resistant mice

This paper reports on the effect of GCP-02, a dual activator of the peroxisome proliferator-activated receptors alpha/gamma (PPARalpha/gamma), on glucose and lipid metabolism in insulin-resistant obese mice induced by monosodium glutamate. The mice were divided into four groups on the basis of treatment: control group, rosiglitazone (positive control) (7 micromol/kg), and low- and high-dosage GCP-02 (7 micromol/kg and 3.5 micromol/kg, respectively). Drugs were given orally once a day for 19 days, and mice underwent testing for insulin tolerance, oral glucose tolerance and gluconeogenesis, and plasma cholesterol, triglyceride and free fatty acid levels. Mice were sacrificed, and body length and weight were measured; intraperitoneal adipose, heart and liver weighed; and plasma alanine aminotransferase (ALT) level and aspartate aminotransferase (AST) activity measured. Liver, soleus muscle and myocardium were assayed for glycogen, triglyceride and free fatty acid content and myocardia tested for superoxide dismutase (SOD) activity and malonaldehyde content. RT-PCR revealed expression of insulin receptor substrate 1 and 2 (IRS1, IRS2) and related genes in liver. GCP-02 had a more powerful effect than rosiglitazone on improving insulin sensitivity, ameliorating glucose tolerance, suppressing L-alanine-induced gluconeogenesis, and decreasing plasma levels of cholesterol, triglyceride and free fatty acid. It reduced body weight in control mice, significantly lowered hepatic content of glycogen, triglyceride and free fatty acid and myocardial content of triglyceride, and increased myocardial SOD activity. IRS2 mRNA was down-regulated in control mice but up-regulated by GCP-02. Thus, GCP-02 is a potential candidate for the prevention and therapy of diseases associated with insulin resistance such as type 2 diabetes mellitus and cardiovascular disease.     

Oxidative status and anti-oxidant enzyme activity during calcium paradox in the rat isolated heart

1. The effect of calcium paradox on oxidative status and the activity of anti-oxidant enzymes were studied in the rat isolated heart. Glutathione status, sulphydryl group contents and lipid peroxidation in the myocardium, as well as the release of oxidized and reduced glutathione from the heart, were taken as indices of oxidative events. 2. Reperfusion with calcium after calcium-free perfusion induced a significant decrease in the myocardial content of reduced and oxidized glutathione and non-protein sulphydryl groups. At the same time, a significant release of both forms of glutathione from the heart was observed. However, the ratio of oxidized to reduced glutathione remained unchanged and was not different from control. Increased lipid peroxidation was observed only after 30 min of reperfusion with calcium. 3. Increased anti-oxidant activity during the reperfusion period was observed. Mitochondrial Mn-superoxide dismutase (SOD) activity was increased throughout the reperfusion period, while cytoplasmic Cu,Zn-SOD and glutathione peroxidase activity showed a transient increase at 5 min reperfusion. 4. The results do not support an important role of oxygen free radicals in cell damage observed during calcium paradox in the rat isolated heart. Production of oxygen free radicals may occur during the reperfusion period, but the quantity produced is insufficient to exceed the anti-oxidant capacity of the heart.     

Improvement by N-acetylcysteine of acute respiratory distress syndrome through increasing intracellular glutathione, and extracellular thiol molecules and anti-oxidant power: evidence for underlying toxicological mechanisms

In acute respiratory distress syndrome (ARDS), there is extensive overproduction of free radicals to the extent that endogenous anti-oxidants are overwhelmed, permitting oxidative cell damage. The present study examined the benefit of the anti-oxidant compound N-acetylcysteine (NAC) in the management of ARDS by measuring patient's intracellular glutathione (inside red blood cells) and extracellular (plasma) anti-oxidant defense biomarkers and outcome. Twenty-seven ARDS patients were recruited from the intensive care unit of a teaching Hospital and randomly divided into two groups. Both groups were managed similarly by regular treatments but 17 patients received NAC 150 mg/kg at the first day that followed by 50 mg/kg/day for three days and 10 patients did not receive NAC. Treatment by NAC increased extracellular total anti-oxidant power and total thiol molecules and also improved intracellular glutathione and the outcome of the patients. In conclusion, patients with ARDS are in a deficient oxidant-anti-oxidant balance that can get a significant benefit if supplemented with NAC.     

Evaluation of free radical-scavenging and anti-oxidant properties of black berry against fluoride toxicity in rats

Oxidative damage to cellular components such as lipids and cell membranes by free radicals and other reactive oxygen species is believed to be associated with the development of degenerative diseases. Fluoride intoxication is associated with oxidative stress and altered anti-oxidant defense mechanism. So the present study was extended to investigate black berry anti-oxidant capacity towards superoxide anion radicals, hydroxyl radicals and nitrite in different organs of fluoride-intoxicated rats. The data indicated that sodium fluoride (10.3 mg/kg bw) administration induced oxidative stress as evidenced by elevated levels of lipid peroxidation and nitric oxide in red blood cells, kidney, testis and brain tissues. Moreover, significantly decreased glutathione level, total anti-oxidant capacity and superoxide dismutase activity were observed in the examined tissues. On the other hand, the induced oxidative stress and the alterations in anti-oxidant system were normalized by the oral administration of black berry juice (1.6 g/kg bw). Therefore it can be concluded that black berry administration could minimize the toxic effects of fluoride indicating its free radical-scavenging and potent anti-oxidant activities.     

Protective effects of exogenously administered or endogenously produced melatonin on hyperbaric oxygen-induced oxidative stress in the rat brain

1. Hyperbaric oxygen (HBO) is a widely used treatment modality in many diseases. A known side-effect of HBO is the production of reactive oxygen species (ROS). Many anti-oxidants, such as vitamins C and E, riboflavin and selenium, have been used successfully to scavenge the ROS produced by HBO administration. 2. The aim of the present study was to determine whether melatonin, a newly discovered anti-oxidant, has a protective effect against the overproduction of ROS produced by HBO in rat brain tissue. 3. Seventy male Sprague-Dawley rats were divided into seven groups as follows: 1, daytime control; 2, daytime HBO; 3, melatonin; 4, daytime HBO plus melatonin; 5, night-time control; 6, night-time HBO; and 7, night-time HBO under light exposure. 4. Hyperbaric oxygen was administered at 303 kPa for 120 min. Melatonin was injected at a dose of 10 mg/kg, i.p. Brain malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione peroxidase (GPx) levels were measured to elucidate oxidant status. 4. The MDA and SOD levels of groups 2 and 7 increased significantly. Exogenous (group 4) and endogenous (group 6) melatonin protected against HBO-induced lipid peroxidation. Exogenously administered melatonin (groups 3 and 4) had increased levels of the anti-oxidant enzymes SOD and GPx. 5. In conclusion, HBO caused oxidative stress and melatonin exhibited protective effects. Both endogenously produced and exogenously administered melatonin were found to be effective.     

Protective effects of cariporide on endothelial dysfunction induced by high glucose

Aim: To explore the effects of cariporide, a selective sodium-hydrogen antiporter inhibitor, on endothelial dysfunction induced by high glucose. Methods: Acetylcholine (ACh)-induced endothelium-dependent relaxation (EDR), sodium nitroprusside (SNP)-induced endothelium-independent relaxation and biochemical parameters including malondialdehyde (MDA), superoxide dismutase (SOD), and nitric oxide (NO) were measured in rat isolated aorta. Results: A 6-h incubation of aortic rings with high glucose (44 mmol/L) resulted in a significant inhibition of EDR, but had no effects on endothelium-independent relaxation. After the 6-h incubation of aortic rings in the co-presence of cariporide (0.01, 0.1, and 1μmol/L) with high glucose, cariporide prevented the inhibition of EDR caused by high glucose in concentration-dependent manners. Similarly, high glucose decreased SOD activity and contents of NO, and increased MDA concentration in aortic tissue. Cariporide (1 μmol/L) significantly resisted the decrease of NO content and SOD activity, and elevation of MDA concentration caused by high glucose in aortic tissues. Mannitol (44 mmol/L) or cariporide (1μmol/L) alone had no effect on EDR, endothelium-independent relaxation and biochemical parameters. Conclusion: Cariporide significantly prevented endothelial dysfunction induced by high glucose. The mechanisms of endothelial dysfunction induced by high glucose may involve the activation of sodium-hydrogen antiporter and the generation of oxygen-free radicals, but it is not related to the change of osmolarity.     

TRILINOLEIN INHIBITS THE ADHESION OF NEUTROPHILS TO ENDOTHELIAL CELLS

1. Trilinolein is a triacylglycerol with linoleic acid as the only type of fatty acid in all three esterified positions of glycerol. It was recently reported to have a myocardial protective effect in coronary ligated rats. We now study its effect on the adhesion of human neutrophils to cultured bovine endothelial cells. 2. Pretreatment of an endothelial monolayer with trilinolein at concentrations ranging from 10^-10 to 10^-6 mol/L significantly inhibited neutrophil adhesion to endothelial cells. Trilinolein was less potent than sodium nitroprusside in inhibiting neutrophil adhesion. 3. The inhibitory effect of trilinolein was antagonized by methylene blue and N^G-nitro-L-arginine methyl ester. The inhibitory effect of trilinolein was not mediated through linoleic acid because linoleic acid did not inhibit neutrophil adhesion. 4. Pretreatment of neutrophils with trilinolein did not reduce neutrophil adhesion. However, in neutrophils activated with N-formyl-methionyl-leucyl-phenylalanine, trilinolein inhibited the neutrophil adhesion to endothelial cells. 5. We conclude that trilinolein inhibits neutrophil adhesion to the endothelial monolayer by stimulating the nitric oxide and cyclic GMP pathways in endothelial cells. It may also inhibit neutrophil adhesion by scavenging free radicals. The inhibitory effect of trilinolein on neutrophil adhesion may play a role in its myocardial protective activity.     

Increase of anti-oxidation by exercise in the liver of obese Zucker rats

1. The effects of endurance training on the anti-oxidant status in diabetes were studied using obese Zucker rats. 2. We used a moderate exercise programme consisting of treadmill running at 20 m/min and 0% incline for 1 h/day, 7 days/week, for 8 weeks. At the end of the experimental period, changes in hepatic anti-oxidant enzymes in terms of protein content and mRNA levels were detected using western blotting analysis and northern blotting analysis, respectively. In addition, anti-oxidant enzyme activity was determined. 2. A significant reduction in mRNA levels and the protein content of hepatic Mn-superoxide dismutase (SOD) and glutathione peroxidase (GPx) were observed in non-exercise obese groups, but the mRNA and protein levels of these enzymes were markedly increased after exercise training. In addition, exercise training reversed the decreased enzyme activities of Mn-SOD and GPx in obese Zucker rats. 3. The diabetes-related lowering of the glutathione (GSH) concentration was elevated in exercised obese Zucker rats, indicating a marked effect of regular moderate exercise on the endogenous anti-oxidant system. 4. There were no marked changes in hepatic Cu/Zn-SOD in terms of mRNA levels, protein content and activity in sedentary obese Zucker rats compared with their lean littermates. Endurance training did not modify the gene expression and activity of hepatic Cu/Zn-SOD. 5. The results of the present study suggest that regular moderate exercise could improve the anti-oxidant defence function of Mn-SOD, GPx and GSH in obese Zucker rats.     

Therapeutic potential of anti-oxidants and diabetic retinopathy

Retinopathy, a severely disabling complication of diabetes mellitus, is today the leading cause of acquired blindness among young adults in developed countries. Good glycaemic control can attenuate the development of diabetic retinopathy but such metabolic control is often difficult to achieve and maintain and additional therapies need to be identified by which retinopathy can be prevented or arrested. Hyperglycaemia plays a critical role in the development and progression of retinopathy, but the mechanism by which hyperglycaemia results in the development of retinopathy is not clear. Oxidative stress is increased in the retina in diabetes. The possible sources of increased oxidative stress might include increased generation of free radicals or impaired anti-oxidant defence system. Dietary supplementation with anti-oxidants in animal models of diabetic retinopathy inhibits retinal metabolic abnormalities and retinal histopathology, suggesting that oxidative stress is associated with the development of retinopathy. The mechanism by which anti-oxidants inhibit retinopathy in diabetes warrants further investigation, but animal studies show that increasing the diversity of anti-oxidants provides significantly more protection than using any single anti-oxidant. Thus, supplementation with anti-oxidants represents an achievable adjunct therapy to help preserve vision in diabetic patients.     

卡托普利保护大鼠缺血再灌注心肌与抗心肌脂质过氧化的关系(英文)

本实验分别用大鼠在体、离体心脏和培养心肌细胞观察了卡托普利(甲巯丙脯酸)抗心肌缺血再灌注损伤与抗脂质过氧化作用。离体心脏缺氧缺糖45 min后再给氧30 min以及在体心脏缺血3 h后再灌注1h,心肌超氧化物歧化酶(SOD)活性明显下降而丙二醛(MDA)含量显著升高。卡托普利能显著保护再灌注(或再给氧)时心肌SOD活性和降低MDA含量。培养心肌细胞缺氧缺糖6 h,细胞MDA含量和乳酸脱氢酶(LDH)释放显著增加。卡托普利显著降低MDA含量和LDH释放。该作用能被吲哚美辛所取消。IIoprost显示有卡托普利相似的保护作用。结果表明卡托普利的保护作用与抗氧自由基和抗脂质过氧化有关。其机理主要通过促进心肌前列环素释放而发挥作用。     

前胡丙素对牛主动脉平滑肌细胞增殖的抑制作用

目的:探讨前胡丙素(pra-C)对牛主动脉平滑肌细胞(SMC)增殖的影响 方法:细胞DNA的合成量由~3H-胸腺嘧啶核苷[~3H]TdR)掺入试验测定,应用流式细胞术测定细胞周期,用乳酸脱氢酶活性测定观察药物的毒性.结果:无论有无血管紧张素Ⅱ(AngⅡ),在0.01μmol/L到10μmol/L浓度范围内,pra-C均可呈浓度依赖性地抑制SMC的增殖,抑制作用主要与阻止细胞进入有丝分裂期有关,而与细胞毒性无关.结论:pra-C能够完全阻断Aug Ⅱ诱导SMC的增殖效应,部分阻止小牛血清诱导的细胞分裂,这对于血管增生性疾病的防治有重要意义.     

Effects of huperzine A on secretion of nerve growth factor in cultured rat cortical astrocytes and neurite outgrowth in rat PC12 cells

Aim: To study the effects of huperzine A (HupA) on neuritogenic activity and the expression of nerve growth factor (NGF). Methods: After being treated with 10μmol/L HupA, neurite outgrowth of PC 12 cells was observed and counted under phase-contrast microscopy. Mitogenic activity was assayed by [^3H]thymidine incorporation. Cell cytotoxicity was evaluated by lactate dehydrogenase (LDH) release. AChE activity, mRNA and protein expression were measured by the Ellman‘s method, RT-PCR, and Western blot, respectively. NGF mRNA and protein levels were determined by RT-PCR and ELISA assays. Results: Treatment of PC 12 cells with 10μmol/L HupA for 48 h markedly increased the number of neuritebearing cells, but caused no significant alteration in cell viability or other signs of cytotoxicity. In addition to inhibiting AChE activity, 10μmol/L HupA also increased the mRNA and protein levels of this enzyme. In addition, following 2h exposure of the astrocytes to 10μmol/L HupA, there was a significant up-regulation of mRNA for NGF and P75 low-affinity NGF receptor. The protein level of NGF was also increased after 24 h treatment with HupA. Conclusion: Our findings demonstrate for the first time that HupA has a direct or indirect neurotrophic activity, which might be beneficial in treatment of neurodegenerative disorders such as Alzheimer disease.