血清氧化-还原态的监测及其与细胞损伤的关系

目的 :探讨如何评价体内的氧化还原状态及细胞的氧化还原态改变对细胞增殖、凋亡、损伤等的影响。方法 :测定正常人血清的NADPH/NADP+ 和GSH/GSSG比值 ,以了解该两项指标对机体氧化还原态的监测能力。并在培养的内皮细胞中加入各种浓度的氧化剂过氧化氢 (H2 O2 )和 /或还原剂N 乙酰 L 半胱氨酸(NAC) ,观察NADPH/NADP+ 和GSH/GSSG比值的变化与细胞活力及损伤之间的关系。以LDH释放率 ,细胞培养液中脂质过氧化物 (LPO)的浓度 ,细胞增殖活性、细胞凋亡率 ,观察细胞的增殖、损伤及其与氧化 还原态变化的关系。结果 :①在培养的内皮细胞中加入H2 O2 后 ,GSH/GSSG和NADPH/NADP+ 比值明显降低 ,细胞氧化还原状态偏向氧化应激方向 ,细胞出现氧化应激损伤 ;脂质过氧化物 (LPO)生成增多 ;LDH释放率增加 ;细胞增殖活力下降 ;凋亡率增高。氧化应激性损伤程度与GSH/GSSG和NADPH/NADP+ 比值呈负相关 ( p <0 .0 5 )。②在各浓度H2 O2 组中加入 0 .5mmol·L 1NAC ,GSH/GSSG和NADPH/NADP+ 比值恢复至接近对照组 ,上述氧化应激性损害也受到了不同程度的抑制 ,并在一定范围内呈量效关系。③单独加入NAC ,在高浓度NAC1mmol·L 1M和 2mmol·L 1时 ,GSH/GSSG和NADPH/NADP+ 比值明显升高 (VSControl,p <0 .0 1 ) ,细胞的氧化     

恶性肿瘤组织的氧化还原态及其抗氧化能力的改变

目的：探讨恶性肿瘤组织的氧化-还原态及其抗氧化能力的改变。 方法： 42例原发性消化道恶性肿瘤（食管癌13例、胃癌14例及结直肠癌15例），以其相对应的癌旁正常粘膜组织为对照，测定其谷胱甘肽氧化还原对（GSH、GSSG）的含量及辅酶Ⅱ氧化还原对（NADPH、NADP+）的含量，并据此计算出GSH/GSSG、NADPH/NADP+比值以及GSH/GSSG氧化还原电位值、NADPH/NADP+氧化还原电位值。 结果： 癌组织中的GSH、NADPH含量明显高于癌旁组织(P＜0.01)。GSH/GSSG、NADPH/NADP+的氧化-还原电位轻度还原偏移(P＜0.05)。而GSSG含量及NADP+含量则与对照组无明显差异(P＞0.05)。 结论： 消化道恶性肿瘤组织的GSH、NADPH含量明显升高，提示其抗氧化能力明显增强；而在此基础上，其氧化-还原电位仅较癌旁组织轻度偏向还原方向，提示癌组织内仍存有较明显的氧化应激态。     

HSPB1对心肌细胞氧化应激时氧化还原状态的影响

目的:本研究主要探讨HSPB1如何调控细胞的氧化还原状态从而发挥抗心肌损伤的作用。方法:以过氧化氢(H_2O_2)诱导的H9c2心肌细胞氧化应激损伤为模型,转染HSP25表达质粒或小分子干扰RNA,观察细胞存活率、LDH水平、细胞凋亡、活性氧水平、谷胱甘肽含量和GSH/GSSG比值的变化,非还原变性蛋白质印迹检测谷氧还蛋白1(Trx1)、谷胱甘肽过氧化物酶(GR)氧化还原状态的改变。结果:HSPB1过表达抑制H_2O_2所致的LDH释放以及caspase 3和PARP的断裂;HSPB1过表达不能减少过氧化氢引起的活性氧水平增加和不能增加细胞内GSH的含量,但可抑制H_2O_2所致的GSH/GSSG比值的降低。非还原性蛋白质印迹显示HSPB1过表达明显减少了H_2O_2所致的GR和Trx1的氧化。HSPB1表达下调促进H_2O_2所致的细胞死亡、LDH释放以及caspase 3和PARP的断裂,以及GSH/GSSG比值的下降;同时,非还原性蛋白质印迹显示HSPB1表达下调明显增加了H_2O_2所致的GR和Trx1的氧化。结论:HSPB1在心肌细胞氧化应激损伤中通过调控GR和Trx1的氧化还原状态起到抗细胞凋亡的作用。     

苯的代谢产物氢醌改变细胞氧化还原状态诱导TNF-α表达

目的:研究苯的代谢产物氢醌(HQ)对HL-60细胞TNF-α产生和细胞活性的影响,并探讨这种影响是否通过氧化还原调节来实现.方法:HL-60细胞用不同浓度HQ刺激24 h后,收集细胞及培养液上清,检测细胞内活性氧、氧化型/还原型谷胱甘肽含量,CCK8检测细胞增殖,流式细胞术(FCM)检测细胞凋亡,ELISA法检测TNF-α含量.结果:5 μmol/L HQ即可诱导GSH水平增高,抑制HL-60细胞增殖,但对GSH/GSSG比例,TNF-α表达及细胞凋亡无显著影响.50、 100 μmol/L HQ可破坏细胞内氧化还原状态,GSH/GSSG比例下降,TNF-α表达增高,显著抑制HL-60细胞增殖,诱导细胞凋亡,抗氧化剂N-乙酰半胱氨酸能拮抗HQ诱导的TNF-α表达、增殖抑制及细胞凋亡.结论:HQ可通过改变HL-60细胞内氧化还原状态诱导TNF-α表达,导致细胞凋亡.     

细胞内的氧化还原感受器蛋白与氧化还原信号转导

正1背景氧化还原反应是哺乳动物细胞存活所必需的,在细胞内的生化反应及代谢过程中氧化与还原占据着重要的位置。细胞内环境中氧化还原状态的轻微改变可能对细胞的生物活性产生重大影响。理论上,细胞内氧化还原状态的改变必须通过测定不同氧化还原电子对的还原电位来定量~([1-2])。但在实际的生物学研究当中,人们所说的氧化还原状态通常     

NAC通过调控胞浆抗氧化蛋白的氧化还原状态减轻心肌细胞氧化应激损伤

<正>目的:探讨N-乙酰半胱氨酸(NAC)是否通过影响心肌细胞内抗氧化通路蛋白的氧化还原状态从而在心肌细胞氧化应激中发挥保护作用。方法:用H2O2刺激H9c2心肌细胞导致氧化应激损伤。采用CM-H2DCFDA荧光染料检测细胞内ROS变化情况;谷胱甘肽试剂盒检测细胞内GSH和GSSG的水平;非还原Western blot技术检测Trx1、GR、Prx1和PTEN的氧化还原状     

慢性丙型肝炎患者氧化损伤的研究

目的 探讨慢性丙型肝炎(CHC)患者体内氧化损伤的情况.方法 52例CHC患者,按丙氨酸转氨酶(ALT)水平分为A组(ALT上升组)和B组(ALT正常组).正常对照组为20例健康志愿者.利用酶联免疫吸附法(ELISA)测定研究对象血清黄嘌呤氧化酶(XOD),丙二醛(MDA),氧化型谷胱甘肽(GSSG),谷胱甘肽(GSH),谷胱甘肽过氧化物酶(GSH-Px),谷胱甘肽巯基转移酶(GST),谷胱甘肽还原酶(GR)及维生素C(Vc)水平,并作出统计分析.结果 CHC患者血清XOD,MDA,GST和GR水平较正常对照组显著升高,而GSH,GSH-Px和Vc水平则明显降低.同时,A组患者血清XOD,MDA,GSSG,GST及GR水平较B组患者显著上调,而GSH,GSH-Px和Vc水平则显著下调.在CHC患者中,血清XOD,MDA,GSSG,GST水平与ALT水平呈正相关,血清GSH,GSH-Px,Vc与ALT水平呈负相关;血清XOD,MDA,GSSG,GR,GST水平与AST水平呈正相关,血清GSH-Px水平与AST水平呈负相关;血清GR水平与GGT水平呈正相关,血清GSH水平与GGT水平呈负相关;血清MDA,GR水平与AKP水平呈正相关.在CHC组中,仅血清XOD水平与血清HCV RNA水平间存在正相关关系.结论 CHC患者体内存在一定程度的氧化损伤,随血清ALT水平的升高,机体氧化损伤程度进一步加重.     

细胞外半胱氨酸/胱氨酸氧化还原电位对NAFLD肝细胞线粒体功能的影响

目的:研究细胞外半胱氨酸/胱氨酸氧化还原电位(EhCys/Cy SS)对非酒精性脂肪肝(NAFLD)肝细胞线粒体功能的影响。方法:用EhCys/Cy SS分别为0 m V(氧化)、-80 m V(正常)和-150 m V(还原)的氧化还原培养基培养肝细胞株LO2并采用油酸诱导细胞建立NAFLD模型。荧光探针DCFH-DA和Mito SOX分别检测细胞整体水平和线粒体活性氧簇(ROS)生成,使用apocynin(NADPH氧化酶抑制剂)和Mito Q10(靶向线粒体抗氧化剂)、rotenone(线粒体呼吸链复合体I抑制剂)和antimycin A(线粒体呼吸链复合体Ⅲ抑制剂)作用细胞检测线粒体复合体活性从而探究ROS的来源,JC-1检测细胞线粒体膜电位。结果:油酸诱导的NAFLD细胞模型使肝细胞内ROS增多,线粒体膜电位下降;氧化的EhCys/Cy SS加剧了NAFLD肝细胞ROS的生成以及线粒体膜电位的下降,而还原的EhCys/Cy SS能清除ROS并逆转线粒体膜电位的下降。线粒体ROS清除剂Mito Q10能显著地减少氧化的EhCys/Cy SS所增加的ROS,而apocynin效果不明显。Rotenone作用于细胞后,ROS的增长率与细胞外EhCys/Cy SS有关,氧化状态下ROS增长率最小且复合体I活性减弱,即氧化的EhCys/Cy SS能通过抑制线粒体复合体I增加ROS的生成。结论:氧化的EhCys/Cy SS能通过抑制线粒体复合体I,从而加剧NAFLD肝细胞ROS的生成,并使线粒体膜电位进一步下降,而还原的EhCys/Cy SS能减少高脂所致ROS生成并减轻线粒体损伤。     

氧化低密度脂蛋白受体在人血管内皮细胞上的表达及其调控

采用放射配基结合及竞争抑制实验和反转录 聚合酶链反应检测人脐静脉内皮细胞 (HUVECs)上血凝素样的氧化低密度脂蛋白受体 (LOX1)的表达 ,表明HUVECs存在LOX1的基因表达和高亲和力位点Bmax (5 4 5±2 0 3)ng/ 10 6cell,Kd (2 0 1± 0 6 )× 10 -8mol/L ,氧化低密度脂蛋白对其自身受体LOX1的基因表达为正反馈调节 ,应用LOX1抑制剂可抑制此作用。     

血氧饱和度降低可促进红细胞氧化损伤

研究氧化应激条件下,血氧饱和度(SO2)对红细胞氧化损伤的影响。体外将红细胞孵育至SO2分别为0.3、0.5、0.7、0.9及0.98等状态,加入终浓度为0.15mmol/L叔丁基过氧化氢(BHP)使其发生氧化应激反应。测量胞内还原性谷胱甘肽(GSH)含量反映细胞抗氧化力,以胞内高铁血红蛋白(MetHb)、膜质过氧化(TBARS)及膜上沉淀的变性珠蛋白链水平体现细胞氧化损伤程度。结果显示,BHP存在的条件下,GSH含量随SO2上升而增加;MetHb、TBARS及膜上珠蛋白链水平则随SO2的上升而下降。上述结果表明,氧化应激条件下,红细胞抗氧化力和氧化损伤与SO2密切相关,SO2的降低可促进红细胞氧化损伤。     

Effects of long‐acting somatostatin analogues on redox systems in rat lens in experimental diabetes

The effects of long‐acting somatostatin analogues, angiopeptin (AGP) and Sandostatin (SMS), on the early decline in the lens content of glutathione (GSH), ATP and NADPH and increase in sorbitol were studied in STZ diabetic rats, and comparison was made with the effect of insulin. Three factors prompted this study: (i) the known increase in IGF‐1 in ocular tissue in diabetes and antagonistic effect of somatostatins, (ii) the known effect of IGF‐1 in increasing lens aldose reductase and (iii) the lack of effect of somatostatins on diabetic hyperglycaemia, the latter enabling a differentiation to be made between effects of hyperglycaemia per se and site(s) of IGF‐1/somatostatins. All four metabolites studied showed a significant restoration towards the normal control level after 7 days of treatment with AGP and SMS, and AGP was more effective on levels of GSH and ATP. A significant correlation was found between GSH and ATP across all groups at 7 days treatment. The redox state changes in diabetes include both NADP+/NADPH and NAD+/NADH in the conversion of glucose to sorbitol and via sorbitol dehydrogenase to fructose with a linked decrease in ATP formation via NAD+/NADH regulation of the glycolytic pathway. The interlinked network of change includes the requirement for ATP in the synthesis of GSH. The present study points to possible loci of action of somatostatins in improving metabolic parameters in the diabetic rat lens via effects on aldose reductase and/or glucose transport at GLUT 3.     

Modulation of inhibitory glycine receptors in cultured embryonic mouse hippocampal neurons by zinc, thiol containing redox agents and carnosine

Modulation of inhibitory glycine receptors by zinc (Zn(2+)) and endogenous redox agents such as glutathione may alter inhibition in the mammalian brain. Despite the abundance of Zn(2+) in the hippocampus and its ability to modulate glycine receptors, few studies have examined Zn(2+) modulation of hippocampal glycine receptors. Whether redox agents modulate hippocampal glycine receptors also remains unknown. This study examined Zn(2+) and redox modulation of glycine receptor-mediated currents in cultured embryonic mouse hippocampal neurons using whole-cell recordings. Zn(2+) concentrations below 10 microM potentiated currents elicited by low glycine, beta-alanine, and taurine concentrations by 300-400%. Zn(2+) concentrations above 300 microM produced nearly complete inhibition. Potentiating Zn(2+) concentrations shifted the dose-response curves for the three agonists to the left and decreased the Hill coefficient for glycine and beta-alanine but not taurine. Inhibiting Zn(2+) concentrations shifted the dose-response curves for glycine and beta-alanine to the right but reduced the maximum taurine response. Histidine residues may participate in potentiation because diethyl pyrocarbonate and pH 5.4 diminished Zn(2+) enhancement of glycine currents. pH 5.4 diminished Zn(2+) block of glycine currents, but diethyl pyrocarbonate did not. These findings indicate that separate sites mediate Zn(2+) potentiation and inhibition. The redox agents glutathione, dithiothreitol, tris(2-carboxyethyl)phosphine, and 5,5'-dithiobis(2-nitrobenzoic acid) did not alter glycine currents by a redox mechanism. However, glutathione and dithiothreitol interfered with the effects of Zn(2+) on glycine currents by chelating it. Carnosine had similar effects. Thus, Zn(2+) and thiol containing redox agents that chelate Zn(2+) modulate hippocampal glycine receptors with the mechanism of Zn(2+) modulation being agonist dependent.     

Alterations in brain antioxidant enzymes and redox proteomic identification of oxidized brain proteins induced by the anti-cancer drug adriamycin: implications for oxidative stress-mediated chemobrain

Adriamycin (ADR) is a chemotherapeutic for the treatment of solid tumors. This quinone-containing anthracycline is well known to produce large amounts of reactive oxygen species (ROS) in vivo. A common complaint of patients undergoing long-term treatment with ADR is somnolence, often referred to as “chemobrain.” While ADR itself does not cross the blood brain barrier (BBB), we recently showed that ADR administration causes a peripheral increase in tumor necrosis factor α (TNF-α), which migrates across the BBB and leads to inflammation and oxidative stress in brain, most likely contributing to the observed decline in cognition. In the current study, we measured levels of the antioxidant glutathione (GSH) in brains of mice injected intraparitoneally (i.p.) with ADR, as well as the levels and activities of several enzymes involved in brain GSH metabolism. We observed significantly decreased GSH levels, as well as altered GSH/GSSG ratio in brains of ADR treated mice relative to saline-treated controls. Also observed in brains of ADR treated mice were increased levels of glutathione peroxidase (GPx), glutathione-S-transferase (GST), and glutathione reductase (GR). We also observed increased activity of GPx, but a significant reduction in GST and GR activity in mice brain, 72 h post i.p. injection of ADR (20 mg/kg body weight). Furthermore, we used redox proteomics to identify specific proteins that are oxidized and/or have differential levels in mice brains as a result of a single i.p. injection of ADR. Visinin like protein 1 (VLP1), peptidyl prolyl isomerase 1 (Pin1), and syntaxin 1 (SYNT1) showed differential levels in ADR treated mice relative to saline-treated controls. Triose phosphate isomerase (TPI), enolase, and peroxiredoxin 1 (PRX-1) showed significantly increased specific carbonylation in ADR treated mice brain. These results further support the notion ADR induces oxidative stress in brain despite not crossing the BBB, and that antioxidant intervention may prevent ADR-induced cognitive dysfunction.     

高血压患者血清SOD、LPO、GSH-PX含量的变化及其临床意义

目的 :探讨了高血压患者血清中SOD、LPO、GSH -PX含量的变化。方法 :分别应用放射免疫分析和生化法对 5 5例高血压患者进行了SOD、LPO、GSH -PX水平的检测 ,并与 38名正常健康人作比较。结果 :高血压患者血清中SOD、GSH -PX水平明显低于正常人组 (P <0 .0 1 ) ,而LPO水平则明显高于正常人组 (P <0 .0 1 )。结论 :血清SOD、LPO、GSH -PX水平的变化与高血压病的发生和发展密切相关     

Comparison between the effects of aging and hyperoxia on glutathione redox state and protein mixed disulfides in Drosophila melanogaster

The main purpose of this study was to determine whether experimental enhancement of oxidative stress by exposure to hyperoxia is an appropriate model for the acceleration of the normal aging process or for establishing a causal association between oxidative stress and aging. Insect tissues are directly exposed to ambient air via the tracheolar invaginations and are thus highly susceptible to oxidative stress under hyperoxic conditions. Amounts of glutathione (GSH), glutathione disulfide (GSSG) and protein mixed disulfides (PrSSG) were compared under normoxic and 100% ambient oxygen in males of two different strains of Drosophila melanogaster (Oregon R (WT) and y w strains). The reason for using two different strains was to preclude the effects of genetic background and to determine whether variations in longevity of the two strains are associated with resistance to oxidative stress. Amounts of GSSG and PrSSG increased, whereas GSH:GSSG ratios declined as a function of age in both strains. Under hyperoxia, y w flies did not exhibit an increase in GSSG amount or a decline in GSH:GSSG ratio, whereas WT flies showed a decline in GSH:GSSG ratio only during the later part of hyperoxic exposure. In neither strain there was a progressive increase in PrSSG amount under hyperoxia. Results indicate that hyperoxia (100% oxygen) neither reproduces nor accelerates the pattern of alterations in glutathione redox state and PrSSG content observed during aging under normoxic conditions, although some other indicators of oxidative stress may be affected.     

EMS患者红细胞免疫功能与脂质过氧化的关系

目的:探讨了子宫内膜异位症(EMS)患者红细胞免疫功能的变化及其与脂质过氧化的关系。方法:应用红细胞酵母花环法测定54例EMS患者红细胞免疫功能和化学比色法测定血浆丙二醛(MDA)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-PX)含量,并与30名正常健康人作比较。结果:EMS患者红细胞免疫复合物(RBC-IC)花环率和MDA水平明显升高(P<0.05～0.01),而红细胞C3b受体(RBCC3b-RR)、SOD、GSH-PX、SOD/MDA低于正常(P<0.01)。相关分析显示,RBCC3b花环与MDA呈显著负相关(r=-0.4428,P<0.05)、RBC-ICR与MDA呈正相关(r=0.5488,P<0.05)。结论:EMS患者红细胞免疫功能降低,与活性氧代谢紊乱密切相关。     

肾病综合征患者红细胞免疫功能与脂质过氧化关系的探讨

目的：探讨了肾病综合征患者红细胞免疫功能与脂质过氧化的关系。方法：应用红细胞酵母花环法测定了31例肾病综合征患者的红细胞免疫功能和化学法测定血清丙二醛（MDA）、超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GSH-PX）含量,并与35名正常健康人作比较。结果：肾病综合征患者RBC-ICR花环率和MDA水平明显升高（P〈0.01）,而RBC-C3bR,SOD,GSH-PX水平则〈正常（P〈0.01）,相关分析显示：RBC-C3b花环与MDA呈显著负相关（r=-0.4786,P〈0.01）,RBC-ICR与MDA呈正相关（r=0.6702,P〈0.01）。结论：肾病综合征患者红细胞免疫黏附功能的降低与活性代谢紊乱密切相关。     

支气管哮喘患儿红细胞免疫功能与脂质过氧化关系的探讨

目的 :探讨了支气管哮喘患儿红细胞免疫功能的变化及其与脂质过氧化的关系。方法 :应用红细胞酵母花环法测定了 32例支气管哮喘患儿红细胞免疫功能和化学比色法测定血浆丙二醛 (MDA) ,超氧化物歧化酶 (SOD) ,谷胱甘肽过氧化物酶 (GSH -PX)含量 ,并与 35名正常健康儿童作对照。结果 :支气管哮喘患儿RBC-IC花环率和MDA水平明显地升高 (P <0 .0 1 ) ,而RBC -C3bR、SOD、GSH -PX、SOD/MDA低于正常 (P <0 .0 1 ) ,相关分析显示 :RBC -C3b花环与MDA呈显著的负相关 (r=- 0 .4 30 7,P <0 .0 5 ) ,RBC -ICR与MDA呈正相关 (r=0 .6 384 ,P <0 .0 1 )。结论 :支气管哮喘患儿红细胞免疫粘附功能降低 ,与活性氧代谢紊乱密切相关     

缺氧缺血性脑病患儿NO、NOS水平与脂质过氧化关系的研究

目的:探讨了缺氧缺血性脑病患儿血清NO、NOS水平与脂质过氧化的关系.方法:应用生化法对30例缺氧缺血性脑病患儿进行了血清NO、NOS测定和化学比色法测定血清丙二醛(MDA)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-PX)含量,并以35名正常健康人作比较.结果:缺氧缺血性脑病患儿血清NO、SOD、GSH-PX水平显著低于正常人组(P＜0.01),而NOS、MDA水平则显著高于正常人组(P＜0.01),相关分析显示:血清NO、SOD、GSH-PX水平与MDA水平呈明显负相关(r=-0.4286、-0.4125、-0.4108,P＜0.05).结论:缺氧缺血性脑病患儿血清NO、SOD、GSH-PX水平的降低与活性代谢紊乱密切相关.