Probucol promotes endogenous antioxidant reserve and confers protection against reperfusion injury

The present study examines whether a subchronic probucol treatment of rats offers protection against ischemia-reperfusion (IR) injury in isolated perfused hearts. Sprague-Dawley rats were treated every second day per week with probucol (cumulative dose 120 mg/kg body mass, i.p.) for 4 weeks. In the probucol group, baseline myocardial antioxidant enzyme, glutathione peroxidase (GSHPx), activity was increased (p<0.05), whereas superoxide dismutase (SOD) and catalase (CAT) activities were not changed. Baseline oxidative stress, as indicated by the myocardial lipid peroxidation, was less (p<0.05) in the probucol group. Isolated hearts were subjected to 60 min global I and 20 min R. Recovery of the contractile function in globally ischemic hearts upon reperfusion was 36% in untreated group and 74% in the probucol group. After IR, GSHPx and CAT activities were significantly (p<0.05) higher in the probucol group compared with the control group, whereas SOD did not change. Lipid peroxidation owing to IR was significantly less in the probocol group. These data suggest that probucol treatment improves endogenous antioxidant reserve and protects against increased oxidative stress following IR injury.     

急性低氧/复氧胁迫对克氏原螯虾抗氧化-能量代谢的影响

为研究低氧/复氧胁迫对克氏原螯虾(Procambarus clarkii)抗氧化及能量代谢的影响,将克氏原螯虾暴露于(1.0±0.1) mg/L急性低氧胁迫和后续(6.8±0.2) mg/L复氧环境中,于低氧胁迫1h、6h及复氧1h、12h分别采集肝胰腺、鳃和血淋巴,研究低氧/复氧胁迫下克氏原螯虾抗氧化-能量代谢酶的活力变化,分析鳃和肝胰腺组织的超微结构改变。在低氧胁迫下,肝胰腺和血淋巴中SOD酶活力显著下降(P<0.05);复氧以后,肝胰腺、血淋巴及鳃组织中SOD酶活力均出现了显著上升(P<0.05)。SOD酶活力变化可能与复氧过程中超氧阴离子自由基的过量产生有关。在复氧12h后,血淋巴和鳃组织中MDA含量均出现了显著性增加(P<0.01),提示机体细胞在复氧胁迫下产生了脂质过氧化。在低氧胁迫下,肝胰腺、鳃和血淋巴中ACP、AKP酶活力显著上升(P<0.05);在复氧12h后,肝胰腺和鳃组织中ACP酶活力显著降低(P<0.01)。显示低氧/复氧胁迫影响了机体的非特异性免疫应答。在急性低氧胁迫下,肝胰腺、血淋巴与鳃组织中的LDH含量和总ATPase活力均显...     

Hypoxia and recovery perturb free radical processes and antioxidant potential in common carp (Cyprinus carpio) tissues

The effects of hypoxia exposure and subsequent normoxic recovery on the levels of lipid peroxides (LOOH), thiobarbituric acid reactive substances (TBARS), carbonylproteins, total glutathione levels, and the activities of six antioxidant enzymes were measured in brain, liver, kidney and skeletal muscle of the common carp Cyprinus carpio. Hypoxia exposure (25% of normal oxygen level) for 5h generally decreased the levels of oxidative damage products, but in liver TBARS content were elevated. Hypoxia stimulated increases in the activities of catalase (by 1.7-fold) and glutathione peroxidase (GPx) (by 1.3-fold) in brain supporting the idea that anticipatory preparation takes place in order to deal with the oxidative stress that will occur during reoxygenation. In liver, only GPx activity was reduced under hypoxia and reoxygenation while other enzymes were unaffected. Kidney showed decreased activity of GPx under aerobic recovery but superoxide dismutase (SOD) and catalase responded with sharp increases in activities. Skeletal muscle showed minor changes with a reduction in GPx activity under hypoxia exposure and an increase in SOD activity under recovery. Responses by antioxidant defenses in carp organs appear to include preparatory increases during hypoxia by some antioxidant enzymes in brain but a more direct response to oxidative insult during recovery appears to trigger enzyme responses in kidney and skeletal muscle.     

Reduced vulnerability of the hypertrophied rat heart to oxygen-radical injury

Effects of xanthine--xanthine oxidase produced oxygen radicals were studied in hypertrophied rat hearts in a Langendorff preparation. Heart hypertrophy was produced by banding of the abdominal aorta for 6 weeks. This resulted in a 22% increase in ventricle/body weight ratio compared with that of sham-operated controls. Perfusion with xanthine--xanthine oxidase caused contractile failure and a significant rise in the resting tension. Complete contractile failure in hypertrophied hearts was seen at 25.5 +/- 3.2 min, whereas in control hearts it happened at 14.4 +/- 5.6 min. Contractile failure due to oxygen radicals in both groups was associated with a decline in high energy phosphates, increased lipid peroxidation, and extensive structural damage. Sarcolemma in both groups became permeable to the extracellular tracer lanthanum. As compared with control, in hypertrophied hearts the malondialdehyde content, indicative of lipid peroxidation, was less by 40%; whereas superoxide dismutase, a free radical scavenger, was higher by a similar amount. These data show a greater capacity of the 6-week hypertrophied heart to withstand a free radical induced contractile failure. This delay in oxygen radical effect can be partially explained by the reduced lipid peroxide content and increased superoxide dismutase activity in the hypertrophied hearts.     

The role aortic chemoreceptors during severe CO hypoxia

The importance of aortic chemoreceptors in the circulatory responses to severe carbon monoxide (CO) hypoxia was studied in anesthetized dogs. The aortic chemoreceptors were surgically denervated in eight dogs prior to the induction of CO hypoxia, with nine other dogs serving as intact controls. Values for both whole body and hindlimb blood flow, vascular resistance, and O2 uptake were determined prior to and at 30 min of CO hypoxia in the two groups. Arterial O2 content was reduced 65% using an in situ dialysis method to produce CO hypoxia. At 30 min of hypoxia, cardiac output increased but limb blood flow remained at prehypoxic levels in both groups. This indicated that aortic chemoreceptor input was not necessary for the increase in cardiac output during severe CO hypoxia, nor for the diversion of this increased flow to nonmuscle tissues. Limb O2 uptake decreased during CO hypoxia in the aortic-denervated group but remained at prehypoxic levels in the intact group. The lower resting values for limb blood flow in the aortic-denervated animals required a greater level of O2 extraction to maintain resting O2 uptake. When CO hypoxia was superimposed upon this compensation, an O2 supply limitation occurred because the limb failed to vasodilate even as maximal levels for O2 extraction were approached.     

超氧化物歧化酶对内皮细胞缺氧复氧损伤的防护作用

体外培养扔兔胸主动脉内皮细胞缺氧３０ｍｉｎ后复氧１０ｍｉｎ,可以发现缺氧后复氧可引起细胞乳酸脱氢酶释放量,细胞悬液丙二醛含量增加,谷胱甘肽过氧化酶活性降低,细胞合成释放一氧化氮减少,细胞内钙离子浓度明显升高;ＥＣ的这些损伤在缺氧期间即有表现,复氧后更为加剧。而在缺氧前预先加入终浓度为２００Ｕ／ｍｌ的超氧化物歧化酶可改善细胞的抗氧化能力,减轻缺氧复氧对ＥＣ的损伤。     

Silymarin modulates Cisplatin-induced oxidative stress and hepatotoxicity in rats

Cisplatin (CDDP) is a widely used anticancer drug, but at high dose, it can produce undesirable side effects such as hepatotoxicity. Because silymrin has been used to treat liver disorders, the protective effect of silymarin on CDDP-induced hepatotoxicity was evaluated in rats. Hepatotoxicity was determined by changes in serum alanine aminotransferase [ALT] and aspartate aminotransferase [AST], nitric oxide [NO] levels, albumin and calcium levels, and superoxide dismutase [SOD], glutathione peroxidase [GSHPx] activities, glutathione content, malondialdehyde [MDA] and nitric oxide [NO] levels in liver tissue of rats. Male albino rats were divided into four groups, 10 rats in each. In the control group, rats were injected i.p. with 0.2 ml of propylene glycol in saline 75/25 (v/v) for 5 consecutive days [Silymarin was dissolved in 0.2 ml of propylene glycol in saline 75/25 v/v]. The second group were injected with CDDP (7.5 mg /kg, I.P.), whereas animals in the third group were i.p. injected with silymarin at a dose of 100 mg/kg/day for 5 consecutive days. The Fourth group received a daily i.p. injection of silymarin (100 mg/kg/day for 5 days) 1 hr before a single i.p. injection of CDDP (7.5 mg/kg). CDDP hepatotoxicity was manifested biochemically by an increase in serum ALT and AST, elevation of MDA and NO in liver tissues as well as a decrease in GSH and the activities of antioxidant enzymes, including SOD, GSHPx in liver tissues. In addition, marked decrease in serum NO, albumin and calcium levels were observed. Serum ALT, AST, liver NO level, MDA was found to decreased in the combination group in comparison with the CDDP group. The activities of SOD, GSHPx, GSH and serum NO were lower in CDDP group than both the control and CDDP pretreated with silymarin groups. The results obtained suggested that silymarin significantly attenuated the hepatotoxicity as an indirect target of CDDP in an animal model of CDDP-induced nephrotoxicity.     

Mechanism of hypoxic preconditionin in guinea pig papillary muscles

Brief ischemia or hypoxia has been found to protect the heart against susbsequent long-lasting ischemia and to improve contractile dysfunction as well to reduce cell necrosis and the incidence of lethal arrhythmias. This phenomenon, termed preconditioning (PC) has been demonstrated in different species. However, little is known about PC in guinea pigs. Moreover, electrophysiological changes underlying protection have not been studied so far in conjuntion with force recovery in a setting of PC. The aim of the study was to study PC in a guinea pig papillary muscle, using recovery of contractility after long hypoxic challenge as the main end-point of protection, and to investigate concominant electrophysiological alterations. In guinea pig papillary muscle preparations contracting isometrically (paced at 2 Hz), transmembrane action potentials (AP) and developed force (DF) were recorded by conventional microelectrode technique and a force tranducer. In addition, effective refractory periods (ERP) were determined. Hypoxia was induced by superfusion with 100% N₂ (pO₂ < 5 kPa) and pacing at 3,3 Hz. In the control group, long hypoxia lasted for 45 min and was followed by 30 min reoxygenation. In the PC group, muscles were subjected to 5 min hypoxia followed by 10 min recovery prior to sustained hypoxia/reoxygenation. Results: Long hypoxia induced a similar depression of DF in both, PC and control groups. However, a loss of contractile activity occured earlier in the PC group. AP duration and ERP decreased faster and were significantly shorter after PC. Upon reoxygenation, preconditioned muscles showed significantly better recovery of function (DF 86% of prehypoxic value vs. 36% in controls; p < 0,05). AP and ERP were completely restored in both, PC and control groups. Guinea pig papillary muscle can be preconditioned with a brief hypoxic challenge against contractile dysfunction upon long-lasting hypoxia/reoxygenation. Shortening of AP and loss of contractility occured more quickly during hypoxia and may participate in the protective effect of preconditioning. Possible mechanisms might involve facilitated opening of K_ATP-dependent channels.     

Cardioprotection from ischemia and reperfusion injury by Withania somnifera: a hemodynamic, biochemical and histopathological assessment

The efficacy of Withania somnifera (Ws) to limit myocardial injury after ischemia and reperfusion was explored and compared to that of Vit E, a reference standard known to reduce mortality and infarct size due to myocardial infarction. Wistar rats (150-200 g) were divided into six groups and received orally saline (sham, control group), Ws-50/kg (Ws control and treated group) and Vit E-100 mg/kg (Vit E control and treated group) respectively for 1 month. On the 31st day, rats of the control, Vit E and Ws treated groups were anesthetized and subjected to 45 min occlusion of the LAD coronary artery followed by 60 min reperfusion. Hemodynamic parameters: systolic, diastolic and mean arterial pressure (SAP, DAP, MAP), heart rate (HR), left ventricular end diastolic pressure (LVEDP), left ventricular peak (+)LVdP/dt and (-)LVdP/dt were monitored. Hearts were removed and processed for histopathological and biochemical studies: Myocardial enzyme viz, creatin phosphokinase (CPK), and antioxidant parameters: malondialdehyde (MDA), glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSHPx) were estimated. Postischemic reperfusion produced significant cardiac necrosis, depression of left ventricular functions (MAP, LVEDP, (+) and (-)LVdP/dt) and a significant fall in GSH (p < 0.01), SOD, CAT (p < 0.05), LDH and CPK (p < 0.01) as well as an increase in MDA level (p < 0.05) in the control group rats as compared to sham group. The changes in levels of protein and GPx was however, not significant. Ws and Vit E favorably modulated most of the hemodynamic, biochemical and histopathological parameters though no significant restoration in GSH, MAP (with Vit E) were observed. Ws on chronic administration markedly augmented antioxidants (GSH, GSHPx, SOD, CAT) while Vit E did not stimulate the synthesis of endogenous antioxidants compared to sham. Results indicate that Ws significantly reduced myocardial injury and emphasize the beneficial action of Ws as a cardioprotective agent.     

Recombinant PTD-Cu/Zn SOD attenuates hypoxia–reoxygenation injury in cardiomyocytes

Abstract

Background. Oxidative stress plays a pivotal role in myocardial ischemia–reperfusion injury. Increasing the protein expression of intracellular Cu/Zn SOD, which is the major endogenous antioxidant enzyme, may attenuate or prevent hypoxia–reoxygenation injury (HRI) in cultured cardiomyocytes. However, ectogenic Cu/Zn-SOD can hardly be transferred into cells to exert biological effects. In this study, we constructed PTD-Cu/Zn SOD plasmid with a kind of translocation structure-Protein transduction domain (PTD) and detected its transmembrane ability and antioxidant effects in H9c2 rat cardiomyocytes subjected to hypoxia/reoxygenation injury (HRI). Methods. We constructed the pET-PTD-Cu/Zn SOD (CDs) prokaryotic expression vectors in plasmid that were inserted into E. coli BL21 to induce the protein expression of PTD-Cu/Zn SOD. H9c2 cardiomyocyte HRI was achieved by exposing cardiomyocytes to 12 h hypoxia followed by 2 h reoxygenation. Protein expression of PTD-Cu/Zn SOD in cardiomyocytes was assayed by Western blot and their enzyme activities were investigated by immunohistochemistry and flow cytometry. Results. In cultured cardiomyocytes hypoxia–reoxygenation injury model, exogenous PTD-Cu/Zn SOD could penetrate cell membrane to clear superoxide anion and decrease hydrogen peroxide level in H9c2 cardiomyocytes subjected to HRI. The level of mitochondrial membrane potential was restored to normal, and the cell apoptosis was reduced in cardiomyocytes with PTD-Cu/Zn SOD treatment during HRI. Conclusion. Recombinant PTD-Cu/Zn SOD could scavenge intracellular-free superoxide anion, protect mitochondria from damages, and attenuate the hypoxia–reoxygenation injury in cultured cardiomyocytes.     

肢体远程预处理通过上调机体抗氧化酶活性诱导兔脊髓缺血耐受

目的:证实抗氧化酶活性上调是肢体远程预处理(remote preconditioning,RPC)诱导兔脊髓缺血耐受效应的主要机制之一。方法:60只雄性新西兰大白兔随机分成对照组、远程预处理组、缺血组及RPC 缺血组(对照组n=6,余组n=18)。RPC组行双下肢短暂缺血2次(每次10min,间隔10min);缺血组仅行脊髓缺血模型;RPC 缺血组在远程预处理后1h行脊髓缺血;对照组为假手术组。对照组于脊髓缺血再灌注后48h行神经功能评分后取脊髓,作为对照。余三组分别于再灌注后6h、24h及48h评分后取材,各时间点各取6只。所有动物于缺血前、缺血20min、再灌注20min及再灌注6h采动脉血测血清抗氧化酶活性和丙二醛(MDA)含量;于取材后测定脊髓匀浆抗氧化酶活性和MDA含量。结果:再灌注后6h、24h及48h时对照组、远程预处理组及远程预处理 缺血组神经功能评分均明显高于缺血组(P<0.05)。血浆超氧化物歧化酶(SOD)活性和过氧化氢酶(CAT)活性在每个时间点RPC组均高于对照组,RPC 缺血组高于缺血组(P<0.05);其中缺血20min时,缺血组血浆SOD、CAT活性低于对照组,RPC 缺血组低于RPC组(P<0.05);而与缺血前相比,缺血20min时缺血组及RPC 缺血组SOD和CAT活性显著下降(P<0.05)再灌注24h和48h时,脊髓匀浆SOD、CAT活性对照组低于RPC组,缺血组低于RPC 缺血组(P<0.01);而MDA含量再灌注24h时对照组高于RPC组,缺血组高于RPC 缺血组(P<0.05)。脊髓匀浆SOD、CAT活性与神经功能评分具有显著相关性。结论:RPC诱导脊髓缺血耐受的机制可能为上调抗氧化酶活性,增强机体在缺血再灌注过程中清除氧自由基的能力,从而减少氧自由基介导的损伤,发挥脊髓保护作用。     

血必净注射液对缺氧/复氧大鼠心肌功能与结构的影响

目的：探讨不同剂量的血必净注射液对缺氧/复氧大鼠心肌功能的保护作用。方法：采用Langendorff方法制备大鼠离体心脏缺氧/复氧模型。130只雄性SD大鼠随机分为对照组（sham组）,缺氧/复氧组（H/R组）,低、中、高剂量血必净组（XBJ_L、XBJ_M、XBJ_H组）,除对照组外,其他四组按复氧不同时相（复氧0.5 h、1 h、2 h）又分别分为3个亚组（n=10）。对照组在平衡灌注20 min时纪录左室发展压（LVDP）、左心室发展压最大上升/下降速率（±dp/dt_max）、左心室内压（LVP）、心率（HR）的值, ELISA检测心肌中肌酸激酶同工酶（CK-MB）的浓度,光镜下观察心肌组织结构的改变;其余各组平衡灌注20 min后,灌注ThomasⅡ停搏液使心脏完全停搏30 min之后复灌K-H液使其心脏复跳,连续记录LVDP、±dp/dt_max、LVP、HR在复氧不同时间点的动态变化,ELISA检测各组复氧不同时间点心肌中CK-MB的浓度,光镜下观察各组复氧不同时间点心肌组织结构的改变。结果：与sham组相比,其余各组LVDP、±dp/dt_max、LVP值均降低（P<0.05）,心肌中CK-MB浓度上升（P<0.05）,心肌组织结构发生异常改变,随着复氧时间延长,以上指标异常变化逐渐加剧;在复氧0.5 h、1 h、2 h,各剂量血必净组LVDP、±dp/dt_max、LVP的值均高于H/R组对应时间点的值（P<0.05）,心肌中CK-MB浓度均低于H/R组,心肌组织结构异常变化减轻,以中剂量改善效果最佳（P<0.05）。结论：血必净注射液能够有效改善缺氧/复氧大鼠心肌的功能及形态学结构,以中剂量血必净（4 ml/100 ml）效果最佳。     

Allergy to drugs: antioxidant enzymic activities, lipid peroxidation and protein oxidative damage in human blood

Reactive oxygen species lead to lipid peroxidation and specific oxidation of some specific enzymes, proteins and other macromolecules, thus affecting many intra- and intercellular systems. Recently, antioxidant functions have been linked to anti-inflammatory properties. Cell defences against toxic oxygen include antioxidant enzymes. We studied the enzymic antioxidant capacity in human blood of both erythrocytes and mononuclear cells from patients suffering from an allergic reaction to different drugs. We determined superoxide dismutases (SODs), glutathione peroxidase (GSHPx) and catalase (CAT) activities in each cell type. We also determined the extent of thiobarbituric acid reactive substances (TBARS) and the oxidative damage to proteins, in order to study the correlation between the cellular enzymic activities, the oxidative status and the allergic reaction. In mononuclear cells from allergic patients, SODs and CAT activities were enhanced compared with controls. Conversely, a decrease in GSHPx activity was found. In erythrocytes, higher values for CAT, GSHPx and SODs activities were found in allergic patients. TBARS were also enhanced in both types of cells, and the carbonyl content of serum was equally increased. The respective enzymic imbalances in mononuclear cells and erythrocytes, namely, GSHPx/SOD and CAT/SOD, and their consequences are discussed. To our knowledge, this is the first global study of antioxidant enzyme determinations, including TBARS level and carbonyl content, in patients suffering from allergies to drugs.     

Regulation of atrial contraction by PKA and PKC during development and regression of eccentric cardiac hypertrophy

ANG II plays a major role in development of cardiac hypertrophy through its AT1 receptor subtype, whereas angiotensin-converting enzyme (ACE) inhibitors are effective in reversing effects of ANG II on the heart. The objective of this study was to investigate the role of PKA and PKC in the contractile response of atrial tissue during development and ACE inhibitor-induced regression of eccentric hypertrophy induced by aortocaval shunt. At 1 wk after surgery, sham and shunt rats were divided into captopril-treated and untreated groups for 2 wk. Then isometric contraction was assessed by electrical stimulation of isolated rat left atrial preparations superfused with Tyrode solution in the presence or absence of specific inhibitors KT-5720 (for PKA) and Ro-32-0432 (for PKC) and high Ca2+. Peak tension developed was greater in shunt than in sham hearts. However, when expressed relative to tissue mass, hypertrophied muscle showed weaker contraction than muscle from sham rats. In sham rats, peak tension developed was more affected by PKC than by PKA inhibition, whereas this differential effect was reduced in the hypertrophied heart. Treatment of shunt rats with captopril regressed left atrial hypertrophy by 67% and restored PKC-PKA differential responsiveness toward sham levels. In the hypertrophied left atria, there was an increase in the velocity of contraction and relaxation that was not evident when expressed in specific relative terms. Treatment with ACE inhibitor increased the specific velocity of contraction, as well as its PKC sensitivity, in shunt rats. We conclude that ACE inhibition during eccentric cardiac hypertrophy produces a negative trophic and a positive inotropic effect, mainly through a PKC-dependent mechanism.     

Cardioprotection from ischemia and reperfusion injury by Withania somnifera: A hemodynamic,biochemical and histopathological assessment

The efficacy of Withania somnifera (Ws) to limit myocardial injury after ischemia and reperfusion was explored and compared to that of Vit E, a reference standard known to reduce mortality and infarct size due to myocardial infarction. Wistar rats (150–200 g) were divided into six groups and received orally saline (sham, control group), Ws-50/kg (Ws control and treated group) and Vit E-100 mg/kg (Vit E control and treated group) respectively for 1 month. On the 31st day, rats of the control, Vit E and Ws treated groups were anesthetized and subjected to 45 min occlusion of the LAD coronary artery followed by 60 min reperfusion. Hemodynamic parameters: systolic, diastolic and mean arterial pressure (SAP, DAP, MAP), heart rate (HR), left ventricular end diastolic pressure (LVEDP), left ventricular peak (+) LVdP/dt and (–) LVdP/dt were monitored. Hearts were removed and processed for histopathological and biochemical studies: Myocardial enzyme viz, creatin phosphokinase (CPK), and antioxidant parameters: malondialdehyde (MDA), glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSHPx) were estimated. Postischemic reperfusion produced significant cardiac necrosis, depression of left ventricular functions (MAP, LVEDP, (+) and (–) LVdP/dt) and a significant fall in GSH (p < 0.01), SOD, CAT(p < 0.05), LDH and CPK (p < 0.01) as well as an increase in MDA level (p < 0.05) in the control group rats as compared to sham group. The changes in levels of protein and GPx was however, not significant. Ws and Vit E favorably modulated most of the hemodynamic, biochemical and histopathological parameters though no significant restoration in GSH, MAP (with Vit E) were observed. Ws on chronic administration markedly augmented antioxidants (GSH, GSHPx, SOD, CAT) while Vit E did not stimulate the synthesis of endogenous antioxidants compared to sham. Results indicate that Ws significantly reduced myocardial injury and emphasize the beneficial action of Ws as a cardioprotective agent.     

Pyruvate modulates antioxidant status of cultured human lens epithelial cells under hypergalactosemic conditions

Lens epithelial cells are the metabolic unit of the lens and antioxidant enzymes are mainly concentrated here. The purpose of this study was to maintain human lens epithelial cells (HLEC) in culture and examine the status of antioxidant enzymes (glutathione peroxidase (GSHPx), catalase (CAT), glutathione-S-transferase (GST)), lipid peroxidation product malondialdehyde (MDA) and glutathione (GSH) levels in these cells under normal as well as hypergalactosemic (30 mM galactose) conditions. Further, effect of pyruvate, a physiological antioxidant has also been evaluated on these parameters. For conducting experiments, anterior capsule specimens obtained from fresh cadaver eyes from eye bank were cultured in Dulbecco's Modified Eagle's Medium (DMEM) supplemented with 20% fetal calf serum. Upon confluency, the cells were subcultured in three separate flasks containing DMEM alone (normal group), DMEM + 30 mM D-galactose (control group), DMEM + 30 mM D-galactose + 5 mM pyruvate (test group) and incubated for 24 or 72 h. These cells were observed under the phase contrast microscope for any morphological changes and harvested for the estimation of various antioxidant parameters. Our results show significant weakened antioxidant defense in HLEC when incubated in the presence of galactose as compared to normal. Addition of pyruvate significantly modulated levels of GSH, MDA, GSHPx, CAT and GST.     

Effects of methyl palmoxirate on hypoxic rat atria.

Isolated rat atria in hypoxia released lactate into the bathing medium and underwent a decline of the contraction frequency which, in some cases led to a complete cessation of the pacemaker activity. A pronounced fall in the peak developed tension and a rise in the resting tension also appeared. The atria from 24 h fasted rats, which oxidize faster their reserve lipids than those from fed rats, exhibited greater functional disturbances during hypoxia, a lower lactate output and a smaller recovery of peak tension upon reoxygenation. Methyl palmoxirate, which is a selective inhibitor of carnitine palmitoyltransferase I, attenuated the decline of the beating rate and the rise of the resting tension in both groups of rats and the incidence of atrial arrest in the fasted rat group. The fall in the peak tension, lactate output and recovery upon reoxygenation were not altered by the inhibitor. These data indicate that methyl palmoxirate alleviates some of the hypoxic functional derangements. Hence, it may be inferred that inhibiting the oxidation of the fatty acid derived from the endogenous triacylglycerol is beneficial during oxygen-limited conditions and that these effects could not be ascribed to changes in the glycolytic flux.     

Probucol treatment reverses antioxidant and functional deficit in diabetic cardiomyopathy

Earlier we reported that probucol treatment subsequent to the induction of diabetes can prevent diabetes-associated changes in myocardial antioxidants as well as function at 8 weeks. In this study, we examined the efficacy of probucol in the reversal of diabetes induced myocardial changes. Rats were made diabetic with a single injection of streptozotocin (65 mg/kg, i.v.). After 4 weeks of induction of diabetes, a group of animals was treated on alternate days with probucol (10 mg/kg i.p.), a known lipid lowering agent with antioxidant properties. At 8 weeks, there was a significant drop in the left ventricle (LVSP) and aortic systolic pressures (ASP) in the diabetic group. Hearts from these animals showed an increase in the thiobarbituric acid reacting substances (TBARS), indicating increased lipid peroxidation. This was accompanied by a decrease in the myocardial antioxidant enzymes activities, superoxide dismutase (SOD) and glutathione peroxidase (GSHPx). Myocardial catalase activity in the diabetic group was higher. In the diabetic + probucol group both LVSP and ASP showed significant recovery. This was also accompanied by an improvement in SOD and GSHPx activities and there was further increase in the catalase activity. Levels of the TBARS were decreased in this group. These data provide evidence that diabetic cardiomyopathy is associated with an antioxidant deficit which can be reversed with probucol treatment. Improved cardiac function with probucol may be due to the recovery of antioxidants in the heart.     

Antioxidant system in Down syndrome: a possible role in cataractogenesis

Recent studies show a relationship between oxidants, antioxidants, and degenerative disease of aging like cataract formation. Focal lens cortical changes and cortical liquification have been reported in patients with Down syndrome (DS) over 14 years. There is evidence supporting the hypothesis that trisomy 21 patients have an increase in free radical reactions. These changes in antioxidant system may play a role in cataractogenesis in Down syndrome. We screened serum samples from 12 patients with DS and cataract: and 12 healthy age and sex-matched persons. We evaluated the antioxidant enzyme activities of superoxide dismutase (SOD), glutathione peroxidase (GSHPx), glutathione-S-transferase (GST) and reduced glutathione (GSH) in erythrocytes. SOD and GSHPx levels of patients with DS were significantly higher than the control group. No significant changes were observed in GST and GSH levels between the DS and control groups. These findings suggest impairment in antioxidant system, which may be a possible mechanism for early cataract formation in DS.     

Exogenous GSH protection during hypoxia-reoxygenation of the isolated rat heart: impact of hypoxia duration

The objective of this study was to determine the interaction between duration of myocardial hypoxia and presence of exogenous glutathione (GSH) on functional recovery upon subsequent reoxygenation. Isolated perfused rat hearts were subjected to 20, 30, 40, or 50 min hypoxia (HYP), which resulted in a progressive decline in the amount of contractile recovery (% of normoxic rate-pressure product (RPP) and developed pressure) during 30 min reoxygenation. Supplementation with 5 mM GSH throughout normoxia, hypoxia, and reoxygenation significantly improved contractile recovery during reoxygenation after 20 and 30 min hypoxia (p < 0.05), but had no effect after longer durations of hypoxia when contractile recovery was typically below 40% of RPP and significant areas of no-reflow were observed. ECG analysis revealed that GSH shifted the bell-shaped curve for reperfusion ventricular fibrillation to the right resulting in attenuated fibrillation after 20 and 30 min hypoxia then increased incidences after 40 min when Control hearts were slow to resume electrical activity. ECG conduction velocity was well preserved in all hearts after 20 and 30 min hypoxia, but GSH administration significantly attenuated the decline that occurred with longer durations. GSH supplementation did not attenuate the 35% decline in intracellular thiols during 30 min of hypoxia. When 5 mM GSH was added only during 40 min of hypoxia, RPP recovery after reoxygenation was improved compared to unsupplemented Controls (73% vs. 55% of pre-hypoxia value, p < 0.05). Administration of GSH only during reoxygenation following 40 min of hypoxia did not alter RPP recovery compared to Control hearts. We conclude that cardioprotection by exogenous GSH is dependent on the duration of hypoxia and the functional parameter being evaluated. It is not due to an enhancement of intracellular GSH suggesting that exogenous GSH acts extracellularly to protect sarcolemmal proteins against thiol oxidation during the phase of hypoxia when oxidative stress is a major contributor to cardiac dysfunction. Furthermore, if enough damage accrues during oxygen deprivation, supplementing with GSH during reoxygenation will not impact recovery.