绿茶提取物EGCG对视神经节细胞RGC-5的促氧化和抗氧化作用

目的探讨EGCG对视神经节细胞RGC-5的氧化和抗氧化作用。方法用中性红摄入法检测细胞增殖率,用碧云天生物技术研究所试剂检测过氧化氢和活性氧。结果在细胞培养条件下EGCG产生过氧化氢,细胞代谢EGCG产生的过氧化氢速率慢于直接加入的过氧化氢,在较高浓度下对RGC-5细胞表现抑制生长,具有浓度依赖性,半数致死量为367μmol。在25μmol浓度下24 h未见细胞毒作用,用25μmol的EGCG连续两次预处理RGC-5细胞,每次2 h,细胞表现抗过氧化氢的氧化损伤,耐受较高的过氧化氢的细胞毒作用;过氧化氢酶预处理EGCG(EGCG-E)细胞毒性弱于EGCG。结论 EGCG在较高浓度下产生过氧化氢对细胞表现细胞毒作用,在较低的浓度下诱导表现为保护细胞对过氧化氢的损伤;过氧化氢酶处理的EGCG细胞毒性弱于未处理的EGCG,EGCG的细胞毒性不仅是过氧化氢。     

神经细胞退行性病变机制研究进展

<正>神经退行性疾病(ND)是由神经元及(或)其髓鞘的丧失所致〔1,2〕,并随着时间的推移而恶化,以致功能障碍。ND病理改变有两种:一是细胞凋亡引起的大量神经细胞丢失;二是神经系统没有明显的神经元数量减少,但神经元出现结构和功能进行性退行性变性〔3〕。目前这些复杂疾病的病因仍未明确阐明。1氧化应激(OS)OS被认为是导致衰老和疾病的一个重要因素〔4〕。活性氧(ROS)包括超氧阴离子(·O2-)、羟自由基(·OH)和过氧化氢     

17β-雌二醇对H_2O_2致血管内皮损伤的保护作用及机制的研究

目的探讨17β-雌二醇对H2O2引起的血管内皮损伤的保护作用和可能的机制。方法人脐静脉内皮细胞株在37℃、5%CO2,条件下正常培养至细胞形成致密单层时,分别进行干预。实验分为5组,空白对照组:细胞正常培养,含10%胎牛血清DMEM培养基培养;过氧化氢损伤组:将培养融合状态达到80%~90%的HUVEC,每孔加入终浓度为0.75 mmol/L的H2O2;H2O2损伤+17β-雌二醇保护组:H2O2损伤浓度同过氧化氢损伤组,17β-雌二醇终浓度0.1 mmol/L;单纯17β-雌二醇组:17β-雌二醇终浓度0.1 mmol/L;H2O2损伤+17β-雌二醇保护+渥曼青霉素组H2O2损伤浓度同过氧化氢损伤组,17β-雌二醇终浓度0.1 mmol/L,渥曼青霉素终浓度100 nmol/L。收集干预后的各组细胞,Western-blot法检测细胞内PI3K/Akt蛋白表达情况,用CCK-8法比较细胞活性,流式细胞技术测定细胞凋亡率,荧光探针标记法测定细胞内活性氧(ROS)水平。结果 17β-雌二醇可以明显提高氧化应激损伤组内皮细胞中PI3K/Akt的表达,17β-雌二醇可以明显提高H2O2损伤后血管内皮细胞的存活率,降低早期和晚期凋亡率,并且减少H2O2损伤后血管内皮细胞内的ROS含量,差异有统计学意义(P0.001),在加入PI3K/Akt通路的特异性抑制剂渥曼青霉素以后,17β-雌二醇抗氧化损伤的作用明显减弱。结论 17β-雌二醇通过上调H2O2导致的血管内皮细胞损伤后胞内PI3K/Akt蛋白的表达,提高细胞存活率,发挥抗氧化损伤和抗细胞凋亡作用,这可能是雌激素发挥心血管保护效应的重要途径之一。     

自由基与若干疾病的关系

近年来对自由基与疾病的关系进行深入探讨 ,发现一些有参考价值的规律。本文简述自由基与帕金森病、阿尔茨海默病、基因损伤、动脉粥样硬化的关系。1　自由基与帕金森病帕金森病 (PD)是以黑质致密层中多巴胺神经元为特征的疾病 ,这些神经元被认为对氧化应激特别敏感 ,这是由于反应活性氧通常是由多巴胺代谢产生。过氧化氢 (H2 O2 )产生于酪氨酸羟化酶 ,参与多巴胺的合成过程中及单胺氧化酶及非酶自动氧化的多巴胺分解代谢中。黑质神经元含有神经黑素 (neu-romelanin) ,它可以与金属离子络合 (如铁离子 ) ,提高了 H2 O2与还原性金属离子的…     

黄芩茎叶总黄酮对AD大鼠模型海马神经元超微结构的影响及抗氧化作用

目的 探讨黄芩茎叶总黄酮(SSTF)对AD大鼠模型学习记忆功能、海马神经元超微结构变化的影响及抗氧化作用.方法　采用Morris水迷宫实验观察大鼠学习记忆能力,电镜观察海马神经元的超微结构;检测血清中超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、谷胱甘肽过氧化物酶(GSH-Px)含量.结果 模型组大鼠2 min内穿越平台区域的次数较对照组明显减少(P<0.01),在平台象限停留的时间明显减少(P<0.01),给药组大鼠穿越平台区域的次数较模型组明显增多(P<0.05),在平台象限停留的时间较模型组明显延长(P<0.05).对照组海马神经元超微结构正常,模型组损伤严重,给药组较模型组损伤减轻.模型组SOD、CAT、GSH-Px含量较对照组明显降低(P<0.05),给药组较模型组明显升高(P<0.05).结论 SSTF对海马注射Aβ_(25-35)引起大鼠学习记忆能力降低及海马神经元超微结构损伤具有保护作用.其机制可能是增强大鼠体内抗氧化酶活性,从而减少活性氧对神经元的损伤.     

脑血管病性脑损伤后神经再生修复的策略

无论是何种原因造成的脑损伤后神经元缺乏自我再生和修复能力，这一直是长期困扰神经科学界的一大难题。由于脑损伤后中枢神经缺乏再生能力，特别是脑血管病造成的损伤，不能产生新的神经元或再生新的轴突，因而导致脑外伤后功能障碍难以恢复和无法恢复，如昏迷、瘫痪、失语、痴呆等。目前认为，脑损伤后神经元修复再生障碍主要原因有以下5方面：(1)神经元本身再生能力有限；(2)神经营养因子生成不足；(3)细胞外基质不适宜；(4)损伤产生了大量抑制神经元生长的因子；(5)损伤局部胶质细胞形成坚硬的瘢痕妨碍轴突生长穿过等。     

激活的ACM对鱼藤酮诱导的细胞氧化损伤的抑制作用

目的 观察激活的星形胶质细胞条件培养液(ACM)在鱼藤酮所致PC12细胞氧化损伤过程中的作用.方法 收集激活的ACM,加入到鱼藤酮染毒PC12细胞中,观察其对染毒神经元胞内丙二醛(MDA)、超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-Px)活性的影响,并检测细胞活性氧(ROS)水平和线粒体呼吸链复合物Ⅰ的活性变化.结果 ACM可明显降低染毒PC12细胞MDA和ROS水平;与完全培养基处理比较,ACM能有效增加染毒神经元SOD和GSH-Px的合成和释放,保护和提高抗氧化酶活性.结论 ACM能有效抑制鱼藤酮诱导的PC12细胞氧化损伤.     

自由基与心肌损伤

<正> 近年来,有关氧自由基在放射、炎症损伤和氧毒性等病理、生理改变过程中的作用的文章不断出现,本文的目的是验证活性氧物质在缺血后再灌注过程中加剧心肌损伤作用的假说。活性氧包括超氧阴离子、过氧化氢和羟自由基等。它们依靠其外层结构不成对电子,与多种细胞成份,尤其是含有-SH基的氨基酸和不饱和脂肪酸等物质产生反应。自由基的连续性的反应可致潜在的并发症,包括蛋白质变性、膜脂质过氧化、趋化因子产生和胶原合成的损害,结果是酶活性丢失、膜通透性紊乱及炎性细胞浸润增加。细胞内对抗氧自由基损害     

高糖与活性氧对人腹膜间皮细胞p21Waf1表达的影响

目的 :研究不同浓度葡萄糖、活性氧 (过氧化氢 )、抗氧化剂 (丙酮酸等 )对体外培养人腹膜间皮细胞(HPMC)细胞周期的影响及其机制。　 方法 :以体外培养的HPMC作为研究对象 ,分别给予不同浓度的葡萄糖、过氧化氢、丙酮酸作为刺激因素 ,观察细胞形态 ,用流式细胞仪检测细胞周期改变 ,测定G1期细胞比例。用半定量逆转录多聚酶链反应 (RT PCR)检测细胞周期调控蛋白p2 1Waf1mRNA水平 ,用细胞免疫组化方法检测p2 1Waf1蛋白水平 ,分析细胞周期改变机制。　 结果 :高糖、过氧化氢可以引起细胞形态呈肥大、衰老改变 ,细胞周期分析提示G1期细胞比例升高 ,即细胞停滞于G1期 ;加入抗氧化剂 (丙酮酸 )后 ,G1期细胞比例下降。高糖、外源性过氧化氢可以使p2 1表达增加 (基因与蛋白水平 ) ,在高糖培养液中加入抗氧化剂 ,可以使p2 1表达下降。　 结论 :高糖、外源性过氧化氢皆可使细胞周期发生停滞 ,且高糖增加外源性过氧化氢的毒性作用 ;高糖的这种作用与内源性活性氧致p2 1Waf1的表达增加有关 ,使用抗氧化剂可使之减弱     

茶多酚拮抗冈田酸诱导皮层神经元损伤的作用观察

目的探讨茶多酚（TP）拮抗冈田酸（OA）诱导的皮层神经元神经损伤的作用及机制。方法冈田酸作用于原代培养皮层神经元后,加入不同浓度的茶多酚,采用细胞毒性检测技术,CCK-8检测、Calcein-AM染色,观察茶多酚对冈田酸诱导的皮层神经元损伤的细胞活力的影响。结果 TP能够拮抗OA冈田酸诱导的皮层神经元损伤,与对照组比较,TP可以使OA诱导的皮层神经元的活力升高（P〈0.05）,活细胞的数目增多。结论茶多酚在一定浓度可有效拮抗OA引起的皮层神经元的损伤,发挥神经保护作用。     

Mitochondrial ATP-sensitive potassium channels enhance angiotensin-induced oxidative damage and dopaminergic neuron degeneration. Relevance for aging-associated susceptibility to Parkinson’s disease

Recent studies have shown that renin–angiotensin system overactivation is involved in the aging process in several tissues as well as in longevity and aging-related degenerative diseases by increasing oxidative damage and inflammation. We have recently shown that angiotensin II enhances dopaminergic degeneration by increasing levels of reactive oxygen species and neuroinflammation, and that there is an aging-related increase in angiotensin II activity in the substantia nigra in rats, which may constitute a major factor in the increased risk of Parkinson’s disease with aging. The mechanisms involved in the above mentioned effects and particularly a potential angiotensin–mitochondria interaction have not been clarified. The present study revealed that activation of mitochondrial ATP-sensitive potassium channels [mitoK(ATP)] may play a major role in the angiotensin II-induced effects on aging and neurodegeneration. Inhibition of mitoK(ATP) channels with 5-hydroxydecanoic acid inhibited the increase in dopaminergic cell death induced by angiotensin II, as well as the increase in superoxide/superoxide-derived reactive oxygen species levels and the angiotensin II-induced decrease in the mitochondrial inner membrane potential in cultured dopaminergic neurons. The present study provides data for considering brain renin–angiotensin system and mitoK(ATP) channels as potential targets for protective therapy in aging-associated diseases such as Parkinson’s disease.     

Reactive oxygen species and biological aging: a mechanistic approach

The experimental evidence for an age-dependent increased generation of reactive oxygen species and a progressive accumulation of oxidized biomolecules is growing. However, despite such facts there is no detailed mechanistic information on how the higher availability of reactive oxygen species translates into the accumulation of oxidized biomolecules. For example, open questions are which reactive oxygen species are responsible for what types of oxidation products in vivo, under what specific reaction conditions can we expect which reaction products, and why specifically are modified biomolecules eliminated whereas others accumulate? Mitochondria appear to serve as the major source for reactive oxygen species in aging tissue. Genetic experiments have demonstrated an effect of Cu,ZnSOD on life span and in the prevention of age-related oxidative damage, suggesting that extramitochondrial superoxide promotes biological aging. However, as superoxide does not easily cross membranes, potential chemical pathways that convert mitochondrial reactive oxygen species into superoxide outside the mitochondria are displayed. The chemical reactivity of individual reactive oxygen species with the amino acid side chain of methionine is surveyed to obtain mechanistic details on the oxidation pathways potentially leading to the age-dependent methionine oxidation of the protein calmodulin in vivo. It will evolve that the in vivo accumulation of oxidized calmodulin cannot be the result of the reaction of an individual reactive oxygen species with calmodulin in homogenous solution alone. Complexation of calmodulin to calmodulin-binding proteins and protein turnover are additional parameters likely contributing to the accumulation of specifically modified calmodulin.     

Oxidative stress decreases in the trophocytes and fat cells of worker honeybees during aging

Trophocytes and fat cells of honeybees have been used for cellular senescence studies, but their oxidative stress and antioxidant enzyme activities with aging in workers is unknown. Here, we assayed reactive oxygen species and the activities of antioxidant enzymes in the trophocytes and fat cells of young and old workers. Young workers had higher reactive oxygen species levels, higher superoxide dismutase and thioredoxin reductase activities as well as lower catalase and glutathione peroxidase activities compared to old workers. Adding these results up, we propose that oxidative stress decreases with aging in the trophocytes and fat cells of workers.     

Free radicals: key to brain aging and heme oxygenase as a cellular response to oxidative stress

Aging is one of the unique features in all organisms. The impaired functional capacity of many systems characterizes aging. When such impairments occur in the brain, the susceptibility to neurodegenerative diseases amplifies considerably. The free radical theory of aging posits that the functional impairments in brains are due to the attack on critical cellular components by free radicals, reactive oxygen species, and reactive nitrogen species produced during normal metabolism. In this review, we examine this concept based on the parameters of oxidative stress in correlation to aging. The parameters for lipid peroxidation are phospholipid composition, reactive aldehydes, and isoprostanes. The parameters for protein oxidation are protein carbonyl levels, protein 3-nitrotyrosine levels, electron paramagnetic resonance, and oxidative stress-sensitive enzyme activities. We conclude that free radicals are, at least partially, responsible for the functional impairment in aged brains. The aging brain, under oxidative stress, responds by induction of various protective genes, among which is heme oxygenase. The products of the reaction catalyzed by heme oxygenase, carbon monoxide, iron, and biliverdin (later to bilirubin) each have profound effects on neurons. Although there may be other factors contributing to brain aging, free radicals are involved in the damaging processes associated with brain aging, and cellular stress response genes are induced under free radical oxidative stress. Therefore, this review supports the proposition that free radicals are, indeed, a key to brain aging.     

Reactive Oxygen Species and Autonomic Regulation of Cardiac Excitability

Sympathetic hyper-activity and diminished parasympathetic activity are a consequence of many primary cardiovascular disease states and can trigger arrhythmias. Emerging evidence suggests that reactive oxygen species (ROS) including nitric oxide, superoxide, and peroxynitrite may contribute to cardiac sympathovagal imbalance in the brainstem, peripheral neurons, and in cardiomyocytes since all experience increased oxidative stress as a result of cardiac disease processes and aging. This article reviews the roles of ROS in autonomic dysfunction and arrhythmia. In addition, novel research directed toward finding targets for modulating sympathovagal balance in cardiac disease is discussed.     

Protein oxidative damage is associated with life expectancy of houseflies.

The objective of this study was to test some of the predictions of the oxidative-stress hypothesis of aging, which postulates that aging is causally associated with the molecular damage inflicted by reactive oxygen species. Protein carbonyl content was used as an index of molecular oxidative modifications. The carbonyl content was found to be associated with the physiological age or life expectancy of flies rather than with their chronological age. Exposure of flies to sublethal hyperoxia (100% oxygen) irreversibly enhanced the carbonyl content of the flies and decreased their rate of oxygen consumption. Results of this study indicate that protein carbonyl content may be a biomarker of aging and support the general concept that oxidative stress may be a causal factor in the aging process.     

Caloric restriction or catalase inactivation extends yeast chronological lifespan by inducing H2O2 and superoxide dismutase activity

The free radical theory of aging posits oxidative damage to macromolecules as a primary determinant of lifespan. Recent studies challenge this theory by demonstrating that in some cases, longevity is enhanced by inactivation of oxidative stress defenses or is correlated with increased, rather than decreased reactive oxygen species and oxidative damage. Here we show that, in Saccharomyces cerevisiae, caloric restriction or inactivation of catalases extends chronological lifespan by inducing elevated levels of the reactive oxygen species hydrogen peroxide, which activate superoxide dismutases that inhibit the accumulation of superoxide anions. Increased hydrogen peroxide in catalase-deficient cells extends chronological lifespan despite parallel increases in oxidative damage. These findings establish a role for hormesis effects of hydrogen peroxide in promoting longevity that have broad implications for understanding aging and age-related diseases.     

Sustained inhibition of oxidative phosphorylation impairs cell proliferation and induces premature senescence in human fibroblasts

The mitochondrial theory of aging predicts that functional alterations in mitochondria contribute to the aging process. Whereas this hypothesis implicates increased production of reactive oxygen species (ROS) as a driving force of the aging process, little is known about molecular mechanisms by which mitochondrial impairment might contribute to aging. Using cellular senescence as a model for human aging, we have recently reported partial uncoupling of the respiratory chain in senescent human fibroblasts. In the present communication, we address a potential cause-effect relationship between mitochondrial impairment and the appearance of a senescence-like phenotype in young cells. We found that treatment by antimycin A delays proliferation and induces premature senescence in a subset of the cells, associated with increased reactive oxygen species (ROS) production. Quenching of ROS by antioxidants did however not restore proliferation capacity nor prevent premature senescence. Premature senescence is also induced upon chronic exposure to oligomycin, irrespective of ROS production, and oligomycin treatment induced the up-regulation of the cdk inhibitors p16, p21 and p27, which are also up-regulated in replicative senescence. Thus, besides the well-established influence of ROS on proliferation and senescence, a reduction in the level of oxidative phosphorylation is causally related to reduced cell proliferation and the induction of premature senescence.     

Molecular pathology of aging and its implications for senescent coronary atherosclerosis

PURPOSE OF REVIEW: This review highlights common mechanisms of organismal aging and inflammatory coronary atherosclerosis. RECENT FINDINGS: A substantial body of evidence now indicates that aging is largely due to molecular damage inflicted by reactive oxygen species, electrophiles, and other reactive endobiotic and xenobiotic metabolites. Our understanding of genetic pathways regulating longevity began 12 years ago with the discovery that a developmental-arrest program in the nematode Caenorhabditis elegans also has marked effects on adult lifespan. This pathway, closely related to the insulin and insulinlike growth factor-signaling pathways of mammals, modulates longevity and stress resistance in several model organisms. Insulin-like signaling also has an impact on redox signaling, antioxidant defenses, and metabolic generation of oxidative stress. Recently, additional signaling pathways--involving Sirtuins, AMP kinase, Jun N-terminal kinase 1, and other master regulatory proteins--have been implicated in longevity and stress-resistance mechanisms. The inflammatory process involves acute production of reactive oxygen species by specialized cells responding to infection, exposure to toxins or allergens, cell damage, hypoxia, ischemia/reperfusion, and other factors, initiating signaling through several of these pathways. Free radical chain reactions arise from lipid oxidation and generate oxidized low-density lipoprotein, a powerful inflammatory signal and potentiator of atherosclerosis. Oxidized low-density lipoprotein accumulates in atherosclerotic arteries, particularly in rupture-prone regions. Inflammation involving oxidative stress, by way of the production of reactive oxygen species, is a hallmark of coronary atherosclerosis. SUMMARY: Common pathways underlie both organismal aging and tissue-autonomous senescent pathologic processes, such as coronary atherosclerosis. The mechanisms discovered in model organisms may lead to pharmacotherapeutic interventions.