羟自由基和脑缺血

羟自由基是一种活性高、毒性大的自由基,它在脑缺血及其再灌注的损伤中起着重要的作用,是自由基造成神经细胞死亡的关键环节之一。本文从以下几个方面加以综述,脑缺血中羟自由基产生的可能途径;脑缺血中羟自由基的变化;羟自由基在脑缺血性神经元损伤中的作用以及羟自由基清除剂对脑缺血的保护作用。     

褪黑激素对大鼠大脑皮层脑片在缺氧后羟自由基及乳酸脱氢酶生 …

目的 研究褪黑激素对大鼠大脑皮层脑片在缺氧后羟自由基及乳酸脱氢酶生成的影响。方法 通过水杨酸捕获法一观察缺氧再给氧中自由基含量的变化,脑片通以９１．６％,Ｎ２＋８．４％Ｏ２造成缺氧。ＬＤＨ用细胞毒检测盒测定。结果：ＤＨＢＡ水平在缺氧及再给氧后显著升高,缺氧时给予褪黑激素可以浓度依赖性地降低再给氧１５ｍｉｎ时ＤＨＢＡ的含量,但对缺氧３０ｍｉｎ时ＤＨＢＡ的含量没有显著作用,ＬＤＨ的含量在缺氧后１ｈ内持     

Melatonin decreases production of hydroxyl radical during cerebral ischemia-reperfusion

目的：研究褪黑激素（ｍｅｌａｔｏｎｉｎ）对大鼠脑缺血再灌注中羟自由基生成的影响．方法：采用栓线法阻塞左侧ＭＣＡ３０ｍｉｎ再灌注模型．通过水杨酸捕获法与微透析技术结合来观察缺血再灌中羟自由基含量的变化．结果：ＤＨＢＡ水平在缺血１５ｍｉｎ后显著升高，持续到再灌注后３０ｍｉｎ仍维持较高水平．缺血前３０ｍｉｎ给予ｍｅｌａｔｏｎｉｎ（４ｍｇ·ｋｇ－１，ｓｃ）显著降低缺血１６－３０ｍｉｎ及再灌注１－３０ｍｉｎ时ＤＨＢＡ的含量．结论：Ｍｅｌａｔｏｎｉｎ对大鼠缺血再灌中羟自由基的产生有抑制作用．     

5-羟甲基糠醛对脑缺血再灌注模型小鼠学习记忆及脑部自由基的影响

目的:探讨5-羟甲基糠醛(5-HMF)对脑缺血再灌注模型小鼠学习记忆及脑部自由基的影响。方法:小鼠分为假手术组、模型组、5-HMF低高剂量组和甲磺酸二氢麦角碱阳性对照组,各组给予相应药物后除假手术组外均建立脑缺血再灌注小鼠模型,以Morris水迷宫和跳台实验观测小鼠学习记忆能力,并检测其脑组织超氧化物歧化酶(SOD)活性和丙二醛(MDA)含量。结果:与假手术组比较,模型组Morris水迷宫实验中逃避潜伏期和游泳路径均明显延长,跳台实验的潜伏期缩短、错误次数增加;脑部SOD活性降低、MDA含量升高(P<0·01或P<0·05),与模型组比较,5-HMF组上述各指标均明显改善(P<0·01或0·05)。结论:5-HMF口服给药可改善脑缺血再灌注引起的学习记忆障碍,其机制可能与恢复脑组织清除自由基的酶活力抗自由基损伤有关。     

原花青素对脑缺血再灌注损伤羟自由基含量和肿瘤坏死因子-α表达影响

目的：观察葡萄籽原花青素（GSP）对大鼠脑缺血再灌注损伤中羟自由基含量和肿瘤坏死因子-α（TNF-α）表达的影响。方法 Wistar大鼠48只随机分为四组：假手术组、缺血再灌注模型组、GSP大剂量（40 mg/kg）组、GSP小剂量（10 mg/kg）组,每组12只,应用线栓法制作大鼠大脑中动脉栓塞模型（MCAO）,缺血2 h再灌注24 h。假手术组手术过程相同但不栓塞大脑中动脉。其中GSP大剂量组、小剂量组术前7 d分别腹腔注射2 ml/kg葡萄籽原花青素,模型组大鼠术前7 d腹腔注射2 ml/kg生理盐水,1次/d,再灌注前15 min加注1次。MCAO模型制作完成后首先进行神经功能评分,然后利用生物化学技术检测大鼠血清羟自由基含量和一氧化氮合酶活性,免疫组织化学染色检测半暗带区颞顶部脑皮质神经细胞TNF-α的表达。结果 GSP对大鼠脑缺血再灌注损伤有保护作用。其机制可能与降低NOS活性、脑组织羟自由基含量,降低脑组织TNF-α的表达,抑制神经细胞凋亡等有关。结论 GSP对脑缺血再灌注损伤具有一定的保护作用,具有临床研究价值。     

HPLC检测在体大鼠脑缺血再灌期羟自由基的变化及维生素E的影响

选用水杨酸为捕捉剂,用HPLC—UV动态检测大鼠全脑缺血再灌期纹状体细胞外液羟自由基的变化。主要测定羟自由基与水杨酸生成稳定化合物2,5-二羟基苯甲酸(DHBA)的浓度。结果表明,2,5-DHBA在重灌15～25 min时达最高值,而在缺血期无明显变化。缺血前20 min分别ip Vit E30,60或120 mg·kg^-1均能抑制重灌期2,5-DHBA的升高。提示,本法可用于大鼠脑缺血再灌期羟自由基的测定,并可观察药物对在体脑缺血动物羟自由基反应的影响。     

黄芩素苷对脑缺血后海马羟自由基含量的影响

蛋白激酶Ｃ(ｐｒｏｔｅｉｎｋｉｎａｓｅｃ ,ＰＫＣ)在脑缺血再灌注损伤中占重要地位 ,脑缺血可引起ＰＫＣ移位激活 ,进而导致神经元损伤[1] 。黄芩素苷 (ｂｒｅｖｉｓｃａｐｉｎ ,Ｂｒｅ)是灯盏花注射液的有效成分 ,对ＰＫＣ有很强的抑制作用 ,其脑保护作用已得到国内学者的证实[2 ] 。此外 ,羟自由基 (ｈｙｄｒｏｘｙｌｒａｄｉｃａｌ,·ＯＨ)与脑缺血再灌注损伤关系密切 ,本实验旨在研究黄芩素苷的脑保护作用及其对·ＯＨ含量的影响 ,以进一步探讨其抗脑缺血再灌注损伤的可能机制。1　材料和方法1.1　动物与分组　蒙古沙土鼠 ,5 0～ 70 …     

依达拉奉对大鼠脑缺血再灌注后自由基含量的影响

目的通过观察雄性大鼠脑缺血再灌注后SOD活力、MDA、NO的表达,探讨依达拉奉的脑保护作用。方法选用雄性SD大鼠30只,随机分为3组:假手术(SO)组、缺血再灌注(I/R)组和依达拉奉(ED)组,每组10只。建立大鼠右侧大脑中动脉闭塞局灶性脑缺血再灌注模型,用生化法测定3组脑缺血再灌注后SOD活力、MDA、NO含量。结果 ED组及SO组脑组织SOD活力均高于I/R组(P<0.05),MDA及NO含量显著低于I/R组(P<0.05)。结论依达拉奉能增强SOD活力、减少MDA及NO含量,改善自由基代谢,有效减轻缺血再灌注损伤后的脑损伤。     

阿糖胞苷对羟自由基的清除作用

近年来,有关自由基与癌的研究已被广泛重视.羟自由基(·OH)是最强的氧化剂之一,甚至能引起连KMnO_4都不能引起的反应。·OH具有细胞毒素,因此·OH自由基抑制剂的研究,将会对癌的预防和治疗有一定作用。实验证明阿糖胞苷对羟自由基有一定的清除作用,其清除率随药物浓度的增加而加大,同时证明它对羟自由基的生成速率也有一定抑制作用。     

阿糖胞苷对羟自由基的清除作用

The cell damages caused by superoxide free radical (O₂^-) include a series of biochemical processes, including lipid peroxidation, DNA and chromosomal damage as well as deactivation of enzyme. It has been suggested that the intrinsic attacking agent is hydroxyl free radical (.OH), which is generated through Fenton reaction. Thus, any .OH scavenging agent may inhibit the relevant cell damage. Cytosine arabinoside was found to be able to scavenge .OH generated by Co~(60) radiolysis and Fe(11)-EDTA with H₂O₂ in aqueous solutions. Determination of the fluorescence of hydroxylated derivatives from benzoate was used to estimate the level of .OH. The scavenging percentage and inhibition ratio of .OH by cytosine arabinoside were measured.     

Effect of melatonin on production of hydroxyl radical and lactate dehydrogenase during hypoxia in rat cortical slices 1

目的：研究褪黑激素对大鼠大脑皮层脑片在缺氧后羟自由基及乳酸脱氢酶生成的影响．方法：通过水杨酸捕获法来观察缺氧再给氧中羟自由基含量的变化．脑片通以９１６％Ｎ２＋８４％Ｏ２造成缺氧．ＬＤＨ用细胞毒检测盒测定．结果：ＤＨＢＡ水平在缺氧及再给氧后显著升高;缺氧时给予褪黑激素可以浓度依赖性地降低再给氧１５ｍｉｎ时ＤＨＢＡ的含量,但对缺氧３０ｍｉｎ时ＤＨＢＡ的含量没有显著作用．ＬＤＨ的含量在缺氧后１ｈ内持续升高;褪黑激素可以显著降低缺氧后ＬＤＨ的释放．结论：褪黑激素降低大鼠缺氧后的损伤以及羟自由基的产生．     

Dietary glycine and renal denervation prevents cyclosporin A-induced hydroxyl radical production in rat kidney.

Cyclosporin A (CsA) nephrotoxicity is associated with renal hypoxia and increases in free radicals in the urine. This study was designed to elucidate the mechanism of radical production caused by CsA. Pretreatment of rats with CsA (25 mg/kg, i.g.) for 5 days decreased glomerular filtration rates by 65%, an effect largely prevented by both dietary glycine (5%) or renal denervation. CsA dissolved in olive oil produced a 6-line alpha-(4-pyridyl 1-oxide)-N-tert-butylnitrone (4-POBN)/free radical signal in the urine, which partitioned predominantly into the aqueous phase after chloroform extraction (i.e., it is water soluble). Dimethyl sulfoxide (DMSO) is attacked by the hydroxyl radical to produce a methyl radical; administration of CsA with [(12)C]DMSO produced two radical species in urine, one with hyperfine coupling constants similar to the 4-POBN/methyl radical adduct found in aqueous solution. CsA given with [(13)C]DMSO produced a 12-line spectrum, confirming the formation of hydroxyl radicals. The methyl radical produced by the hydroxyl radical represented 62% of radicals detected in urine but only 15% in bile. Therefore, hydroxyl radicals are produced largely in the kidney. Free radicals in urine were increased about 5-fold by CsA, an effect completely blocked by the inhibitory neurotransmitter, glycine, or by renal denervation. CsA infusion for 30 min increased efferent renal nerve activity 2-fold, and dietary glycine (5%) totally blocked this phenomenon. Taken together, these data are consistent with the hypothesis that CsA increases hydroxyl radical formation by increasing renal nerve activity resulting in vasoconstriction and hypoxia-reoxygenation. Glycine blunts the effect of CsA on the renal nerve, which explains, in part, prevention of nephrotoxicity.     

The effect of caffeine on MDMA-induced hydroxyl radical production in the mouse striatum

BackgroundThe psychostimulant 3,4-methylenedioxymethamphetamine (MDMA) with a strong addictive potential is widely used as a recreational drug. Neurotoxicity of MDMA is related with the generation of highly reactive free radicals.MethodsMDMA was given in doses of 20 and 40 mg/kg ip alone or in combination with caffeine (CAF) 10 mg/kg ip. Extracellular concentration of hydroxyl radical was measured using microdialysis in freely moving mice and was assayed by HPLC with electrochemical detection.ResultsMDMA dose-dependently increased production of hydroxyl radical in the mouse striatum and its effect was reversed by caffeine.ConclusionsThe data show that caffeine may have neuroprotective properties as it decreased oxidative stress induced by MDMA.     

The hydroxyl radical formation system in polymorphonuclear leukocytes.

We studied the hydroxyl radical (OH*)-generating system in polymorphonuclear leukocytes (PMNs). When phenylalanine was incubated with the alpha, beta and gamma fractions prepared from pig polymorphonuclear leukocytes (PMNs) and hydrogen peroxide (H2O2), significant levels of formation of m- and o-tyrosine were observed in the alpha and beta fractions, but not in the gamma fraction. The amount of tyrosine formation per milligram of protein was greater with the beta than with the alpha fraction. Further, when phenylalanine was incubated with alpha or beta fractions with similar myeloperoxidase (MPO) activities in the presence of H2O2, tyrosine formation by the beta fraction was also more effective. Using the beta fraction in which the MPO activity was destroyed by heat treatment, no significant amount of tyrosine was formed. However, with the heat-treated beta fraction and MPO preparations from human neutrophils in the presence of H2O2, the amount of tyrosine formation increased with the addition of increasing amounts of heat-treated beta fraction. Tyrosine formation by the beta fraction in the presence of H2O2 was significantly reduced by OH* scavengers. The above results suggest the existence of an OH*-generating system in which MPO and H2O2 participate in the granules of PMNs and, especially, in specific granules, there may exist some factors that cause more effective OH* generation.     

Prazosin attenuates hydroxyl radical generation in the rat myocardium.

The present study examined whether tyramine-induced hydroxyl radical (*OH) generation via noradrenaline release was attenuated by prazosin. A flexibly mounted microdialysis technique was used to detect the generation of *OH in in vivo rat hearts. The microdialysis probe was implanted in the left ventricular myocardium of anaesthetized rats and Ringer's solution was used. To measure the level of *OH, sodium salicylate in Ringer's solution (0.5 nmol/microl/min) was infused directly through a microdialysis probe to detect the generation of *OH as reflected by the nonenzymatic formation of 2,3-dihydroxybenzoic acid (DHBA). Tyramine (0.1, 0.5 and 1.0 mM) increased the level of 2,3-DHBA in a concentration-dependent manner. However, in the presence of prazosin (10 microM), the effect of tyramine was abolished. To confirm the generation of *OH by a Fenton type reaction, iron (II) was infused through a microdialysis probe. A positive linear correlation between iron (II) and the formation of 2,3-DHBA (R2 = 0.982) was observed. To examine the effect of prazosin on ischemic/reperfused rat myocardium, the heart was subjected to myocardial ischemia for 15 min by occlusion of the left anterior descending coronary artery. When the heart was reperfused, a marked elevation of the level of 2,3-DHBA was observed. However, in the presence of prazosin (10 microM), the elevation of 2,3-DHBA was not observed in ischemic/reperfused rat heart. Prazosin was shown to have a *OH scavenging effect. These results suggest that tyramine-induced noradrenaline causes *OH generation, an effect which is inhibited by prazosin as Na+ channel blocker, but not through its alpha1-adrenoceptor antagonistic action of prazosin.     

蕨麻与脑缺血-再灌注损伤

脑缺血-再灌注损伤是一个复杂的级联反应过程,包括多个环节,近几年研究发现氧化应激与炎症反应与脑缺血-再灌注损伤有着密切关系。因此,抗氧化应激与抗炎症反应治疗在脑缺血-再灌注损伤中愈来愈受到重视。蕨麻作为我国传统藏药之一,药用历史悠久,其药用价值有待进一步发掘。现在学者们已经从蕨麻中提取各种活性成分,经过现代药理研究证实蕨麻具有抗氧化应激、抗炎、保护心肌、提高免疫力、抑制细菌活性及抑癌等作用,但蕨麻是否具有脑缺血-再灌注损伤保护作用尚有待进一步研究。本文就脑缺血-再灌注损伤研究进展与蕨麻的药理作用作一综述。     

Characterization of hard metal dusts from sintering and detonation coating processes and comparative hydroxyl radical production

Dust samples from sintering and detonation coating hard-metal processes were characterized, compared, and contrasted for morphology, composition, and generation of hydroxyl radicals. Inhalation of respirable hard-metal (sintered carbide) dusts from hard-metal processes is known to cause fibrotic and asthmatic lung disease. Scanning electron microscopy/energy-dispersive X-ray analysis was used for morphology, composition, and elemental distribution. An electron spin resonance (ESR) spin trapping technique was used to detect hydroxyl radical generation. Samples were incubated with air-saturated buffer solutions containing a spin trap and analyzed by ESR for the presence of *OH in solution. Postdetonation coating samples often had surface contamination of Co on the WC particles, as shown by elemental mapping of individual particles; this was not evident in predetonation samples or unsintered materials in this study. ESR measurements show that both detonation-gun materials were capable of generating *OH , while the WC, cobalt, and presintered mixture did not produce detectable amounts of *OH radicals. The DMPO/*OH adduct formation was apparently facilitated by Fe-mediated reactions for predetonation dusts, and by Fe-mediated site-specific reactions for postdetonation dusts. The overspray materials from the detonation-gun process produced 9-fold more *OH radicals than the predetonation coating mixture. Overall, this study indicates there are substantial differences between postdetonation materials and both predetonation and unsintered hard-metal process materials with respect to morphology, elemental distribution, and *OH radical generation reactions and that these differences may be important in the toxic potential of those materials.     

Synthesis of spin traps specific for hydroxyl radical

Two nitrones, 3,3-diethyl-5,5-dimethylpyrroline 1-oxide (DEDMPO) and 3,3,5,5-tetramethylpyrroline 1-oxide (M4PO), were synthesized by the zinc/ammonium chloride reduction of appropiately substituted gamma-nitrocarbonyl compounds, followed by addition of methylmagnesium bromide to the resulting intermediate nitrones. The lipophilicities of these nitrones were estimated by determining their partition coefficients in an 1-octanol/water system. They were found to be considerably more lipophilic than 5,5-dimethylpyrroline 1-oxide (DMPO). The spin trapping of hydroxyl and superoxide radicals with these nitrones was investigated, and the hyperfine coupling constants were determined. M4PO was found to spin trap both free radicals, while DEDMPO spin trapped only hydroxyl radical. DEDMPO was used to determine if hydroxyl radical was produced during the metabolism of menadione or nitrazepam by porcine thoracic aorta endothelial cells. Our results indicate, in conjunction with spin-trapping studies utilizing DMPO, that only superoxide is generated during cellular metabolism of quinones and aromatic nitro-containing compounds by endothelial cells.