Glutathione loading prevents free radical injury in red blood cells after storage

We have previously demonstrated that the loss of glutathione (GSH) and GSH-peroxidase (GSH-PX) in banked red blood cells (RBCs) is accompanied by oxidative modifications of lipids, proteins and loss of membrane integrity^[1]. The objective of this study was to determine whether artificial increases in antioxidant (GSH) or antioxidant enzyme (catalase) content could protect membrane damage in the banked RBCs following an oxidant challenge. RBCs stored at 1-6°C for 0, 42 and 84 days in a conventional additive solution (Adsol®) were subjected to oxidative stress using ferric/ascorbic acid (Fe/ASC) before and after enriching them with GSH or catalase using a hypotonic lysis-isoosmotic resealing procedure. This lysis-resealing procedure in the presence of GSH/catalase raised intracellular GSH and catalase concentrations 4-6 fold, yet produced only a small reduction in mean cell volume (MCV), mean cell hemoglobin (MCH) and mean cell hemoglobin concentrations (MCHC). Indicators of oxidative stress and membrane integrity were measured, including acetylcholinesterase (AChE) activity, GSH concentration, phosphatidylserine (PS) externalization (prothrombin-converting activity) and transmembrane lipid movements (¹⁴C-lyso phosphatidylcholine flip-flop and PS transport). GSH-enrichment protected AChE activity in fresh (0 day) and stored (42 and 84 days) RBCs from Fe/ASC oxidation by 10, 23 and 26%, respectively, compared with not-enriched controls. Following oxidative stress, the rate of transbilayer lipid flip-flop did not increase in fresh cells, but increased 9.3% in 42-day stored cells. Phosphatidylserine exposure, as measured by prothrombinase activity, increased 2.4-fold in fresh and 5.2-fold in 42-day stored cells exposed to Fe/ASC. Previous studies have shown that 42-day storage causes a moderate decrease in PS transport (∼ 50 %), whereas transport rates declined by up to 75% in stored RBCs when challenged with Fe/ASC. GSH-enrichment prevented the increase in passive lipid flip-flop and the increase in prothrombinase activity, but offered no protection against oxidative damage of PS transport. In contrast to these effects, catalase-enrichment failed to protect GSH levels and AChE activity upon oxidative stress. Membrane protein thiol oxidation was assessed by labeling reactive protein thiols with 5-acetalamidofluorescein followed by immunoblotting with antifluorescein antibodies. Significant oxidation of membrane proteins was confirmed by a greater loss of thiols in stored RBCs than in fresh RBCs. These results demonstrate that it may be possible to prevent storage-mediated loss of AChE, increased lipid flip-flop, and increased PS exposure, by maintaining or increasing GSH levels of banked RBCs.     

冠状动脉介入治疗对冠心病患者Caspase-3,9水平的影响

目的：通过对冠心病患者经皮冠状动脉介入治疗（PCI）前后血清Caspase-3,9水平变化的观察,探讨冠状动脉介入治疗对冠心病患者血清Caspase-3,9水平的影响。方法：冠状动脉介入组（PCI组）30例和单纯造影组（CAG组）29例,在术前0.5h,术后3h,术后6h,术后12h,术后24h,术后48h分别测定血清Caspase-3,9水平。结果：CAG组术前与术后的血清Caspase-3,9水平差异均无统计学意义,PCI组术后3h、6h的Caspase-3水平较术前0.5h均有降低趋势,差异均有统计学意义（P〈0.05）,PCI组术后48h的Caspase-9水平与术后3h、6h相比均有升高趋势,差异均有统计学意义（P〈0.05）。结论：PCI术后狭窄或闭塞的冠状动脉血管重新恢复血供,血清中Caspase-3,9水平在PCI术后先降低后又逐渐升高,提示再灌注后可出现细胞的凋亡程度加重,血清中Caspase-3,9的水平可作为心肌再灌注损伤后细胞凋亡程度的评估指标并可作为诊断临床症状不典型的再灌注损伤的一个指标。PCI术可有效改善心肌血供,减少因术前缺血缺氧造成的细胞凋亡,但PCI术后再灌注损伤可进一步加重细胞凋亡。     

冠状动脉介入治疗对冠心病患者Caspase-3,9 水平的影响

目的:通过对冠心病患者经皮冠状动脉介入治疗(PCI)前后血清Caspase-3,9水平变化的观察,探讨冠状动脉介入治疗对冠心病患者血清Caspase-3,9水平的影响。方法:冠状动脉介入组(PCI组)30例和单纯造影组(CAG组)29例,在术前0.5h,术后3h,术后6h,术后12h,术后24h,术后48h分别测定血清Caspase-3,9水平。结果:CAG组术前与术后的血清Caspase-3,9水平差异均无统计学意义,PCI组术后3h、6h的Caspase-3水平较术前0.5h均有降低趋势,差异均有统计学意义(P<0.05),PCI组术后48h的Caspase-9水平与术后3h、6h相比均有升高趋势,差异均有统计学意义(P<0.05)。结论:PCI术后狭窄或闭塞的冠状动脉血管重新恢复血供,血清中Caspase-3,9水平在PCI术后先降低后又逐渐升高,提示再灌注后可出现细胞的凋亡程度加重,血清中Caspase-3,9的水平可作为心肌再灌注损伤后细胞凋亡程度的评估指标并可作为诊断临床症状不典型的再灌注损伤的一个指标。PCI术可有效改善心肌血供,减少因术前缺血缺氧造成的细胞凋亡,但PCI术后再灌注损伤可进一步加重细胞凋亡。     

Zinc protects HeLa-tat cells against free radical cytotoxicity induced by glucose

In the present study, we investigated the protective effect of zinc on the glucose-induced cytotoxicity in HeLa wild and HeLa-tat cells (30 and 20 mmol/l glucose, respectively). HeLa cells transfected with the protein Tat exhibit a lower antioxidant defense system. Incubation of HeLa wild and HeLa-tat cells with high glucose levels led to a rapid increase in generation of reactive oxygen species (ROS). As expected in the presence of high glucose concentrations, the viability was reduced for both cell lines. The redox status essentially regulated by thiol groups may play an important role in the apoptotic process. Thus, we developed a new method using the p-nitrophenyl disulfide to measure cytosolic thiol groups in intact cells. Cellular zinc was measured using inductively coupled plasma mass spectrometry. Intracellular thiol groups and intracellular zinc concentrations were significantly lower in HeLa cells cultured in hyperglycemic conditions, and their concentrations were significantly lower in HeLa-tat cells than in HeLa wild cells. However, the generation of ROS and the induction of apoptosis by a glucose specific mechanism were prevented by zinc (50 micromol/l) and the intracellular thiol groups and zinc concentrations significantly increased in both cell lines to become similar to the initial values. These results suggest that the glucose oxidation and its subsequent effects on the cells can be prevented by a biological antioxidant such as zinc.     

Primary antioxidant free radical scavenging and redox signaling pathways in higher plant cells

Antioxidants in plant cells mainly include glutathione, ascorbate, tocopherol, proline, betaine and others, which are also information-rich redox buffers and important redox signaling components that interact with cellular compartments. As an unfortunate consequence of aerobic life for higher plants, reactive oxygen species (ROS) are formed by partial reduction of molecular oxygen. The above enzymatic and non-enzymatic antioxidants in higher plant cells can protect their cells from oxidative damage by scavenging ROS. In addition to crucial roles in defense system and as enzyme cofactors, antioxidants influence higher plant growth and development by modifying processes from miotosis and cell elongation to senescence and death. Most importantly, they provide essential information on cellular redox state, and regulate gene expression associated with biotic and abiotic stress responses to optimize defense and survival. An overview of the literature is presented in terms of primary antioxidant free radical scavenging and redox signaling in plant cells. Special attention is given to ROS and ROS-anioxidant interaction as a metabolic interface for different types of signals derived from metabolisms and from the changing environment. This interaction regulates the appropriate induction of acclimation processes or execution of cell death programs, which are the two essential directions for higher plant cells.     

Electron spin resonance and spin trapping technique provide direct evidence that edaravone prevents acute ischemia-reperfusion injury of the liver by limiting free radical-mediated tissue damage

A novel free radical scavenger, 3-methyl-1-phenyl-2-pyrazolin-5-one (edaravone), is used for the treatment of acute ischemic stroke and is protective in several animal models of organ injury. We tested whether edaravone is protective against acute liver warm ischemia/reperfusion injury in the rat by acting as a radical scavenger. When edaravone was administered prior to ischemia and at the time of initiation of the reperfusion, liver injury was markedly reduced. Production of oxidants in the liver in this model was assessed in vivo by spin-trapping/electron spin resonance (ESR) spectroscopy. Ischemia/reperfusion caused an increase in free radical adducts rapidly, an effect markedly blocked by edaravone. Furthermore, edaravone treatment blunted ischemia/reperfusion-induced elevation in pro-inflammatory cytokines, infiltration of leukocytes and lipid peroxidation in the liver. These results demonstrate that edaravone is an effective blocker of free radicals in vivo in the liver after ischemia/reperfusion, leading to prevention of organ injury by limiting the deleterious effects of free radicals.     

Biomarkers in breath condensate: a promising new non-invasive technique in free radical research

Oxidative stress is associated with a range of inflammatory lung diseases including asthma, adult respiratory distress syndrome, idiopathic pulmonary fibrosis, pneumonia, lung transplantation, chronic obstructive pulmonary disease, cystic fibrosis, bronchiectasis and lung cancer. Increased concentrations of reactive oxygen species (ROS) in the airways of such patients are reflected by elevated concentrations of oxidative stress markers in the breath, airways, lung tissue and blood. Traditionally, the measurement of these biomarkers has involved invasive procedures to procure the samples, or examine the compartments. As a consequence, there is a need for less invasive approaches to measure oxidative stress. Analysis of breath hydrocarbons has partly fulfilled this need, however only gas phase volatile constituents can be assessed by this approach. The collection of exhaled breath condensate (EBC) is a simple, non-invasive approach, which comprehensively samples the lower respiratory tract. It is currently used as a research and diagnostic tool in the free radical field, yielding information on redox disturbance and the degree and type of inflammation in the lung. With further technical developments, such an approach may ultimately have a role in the clinic, in helping to diagnose specific lung diseases. EBC can be exploited to assess a spectrum of potential biomarkers, thus generating a “finger print” characteristic of the disease. By assessing the nature of oxidative stress in this manner, the most appropriate therapy can be selected and the response to treatment monitored.     

Postischemic administration of liposome-encapsulated luteolin prevents against ischemia-reperfusion injury in a rat middle cerebral artery occlusion model

Oxidative stress-induced neuronal cell death has been implicated in neurodegenerative diseases; one such disease is ischemic stroke. Using reactive oxygen species (ROS)-insulted primary neurons, we screened neuroprotectants with clinical potential and then, using ischemia/reperfusion (I/R) model, investigated the anti-ischemic potential of candidate neuroprotectants. Here, we showed that luteolin, isolated from the ripe fruit of Perilla frutescens (L.) Britt, exhibited a neuroprotective action upon the in vitro platform, thus serving as candidate for in vivo pharmacological evaluation. Liposome-encapsulated luteolin produced dramatic preventing effects on I/R-induced behavioral and histological injuries after a 13-day post-ischemic treatment. Furthermore, this phytochemical not only lowered the increased level of mitochondrial ROS but also substantially up-regulated the decreased activity of catalase and glutathione in I/R rat brains. Collectively, luteolin as a neuroprotectant acts by anti-ischemic activity likely through a rebalancing of pro-oxidant/antioxidant status. Its multitarget mechanisms implicate potential effectiveness for clinically treating ischemia stroke.     

Polyethylene glycol immediately repairs neuronal membranes and inhibits free radical production after acute spinal cord injury

Membrane disruption and the production of reactive oxygen species (ROS) are important factors causing immediate functional loss, progressive degeneration, and death in neurons and their processes after traumatic spinal cord injury. Using an in vitro guinea pig spinal cord injury model, we have shown that polyethylene glycol (PEG), a hydrophilic polymer, can significantly accelerate and enhance the membrane resealing process to restore membrane integrity following controlled compression. As a result of PEG treatment, injury-induced ROS elevation and lipid peroxidation (LPO) levels were significantly suppressed. We further show that PEG is not an effective free radical scavenger nor does it have the ability to suppress xanthine oxidase, a key enzyme in generating superoxide. These observations suggest that it is the PEG-mediated membrane repair that leads to ROS and LPO inhibition. Furthermore, our data also imply an important causal effect of membrane disruption in generating ROS in spinal cord injury, suggesting membrane repair to be an effective target in reducing ROS genesis.     

斑点追踪技术评价冠心病患者PCI术前后左室心肌力学改变

目的:探讨斑点追踪成像(speckle tracking imaging,STI)技术评价经皮冠状动脉介入(percutaneous coronary intervention,PCI)术对冠状动脉严重狭窄患者左室心肌力学的改变。方法:病变组(冠状动脉左前降支狭窄75%患者)30例,分别于PCI术前1天和术后3天、术后3个月接受超声心动图检查,测量的常规指标包括:左室射血分数(LVEF)、左室舒张末内径(LVDd)、左室舒张末容积(LVEd V),同时应用STI技术测量缺血心肌节段收缩期峰值应变参数:纵向、径向、圆周应变LS、RS、CS。体检健康者30例为对照组进行比较分析。结果:1与对照组比较,病变组PCI术前缺血心肌应变值(LS、RS、CS)均呈不同程度减低,差异均有统计学意义(P0.05);术后3天与术前比较均无统计学差异(P0.05);PCI术后3个月病变组LS、RS、CS较术前均不同程度提高,差异均有统计学意义(P0.05),与对照组比较,差异无统计学意义(P0.05);2病变组PCI术前后不同时间点与对照组比较,LVEF、LVDd、LVEd V均无统计学差异(P0.05)。结论:STI技术可定量敏感的评价冠状动脉严重狭窄患者缺血心肌力学改变,为评价PCI术对冠心病患者的疗效提供了客观依据。     

冠心病患者经皮冠状动脉介入治疗术后支架内再狭窄的相关危险因素分析

目的:探究冠心病(CHD)患者经皮冠状动脉介入治疗(PCI)术后支架内再狭窄(ISR)的相关危险因素。方法:选取2014年6月～2017年6月期间我院收治的行PCI的CHD患者200例为研究对象,术后随访一年再行冠脉造影检测,根据患者是否发生ISR分为观察组(38例,发生ISR)和对照组(162例,未发生ISR),收集并比较两组患者基线资料及生化指标,采用多因素logistic回归分析CHD患者PCI术后发生ISR的危险因素。结果:观察组吸烟、饮酒、高血压、糖尿病的人数占比、病程及支架直径均高于对照组,差异有统计学意义(P0.05)。观察组脂蛋白(a)[LP(a)]、纤维蛋白原(FIB)及尿酸(UA)水平显著高于对照组,总胆红素(TBIL)水平显著低于对照组,差异有统计学意义(P0.05)。多因素logistic回归分析显示,吸烟、糖尿病、支架直径(小)以及高水平LP(a)、低水平UA为CHD患者行PCI术后发生ISR的危险因素。结论:CHD患者行PCI术后发生ISR的危险因素有吸烟、糖尿病、支架直径以及高水平LP(a)、低水平UA,因此在PCI术中应尽可能选用较大的支架,同时戒烟、控制血糖有利于预防ISR的发生,定期检测血清LP(a)、UA水平变化,并采取有效的医疗与保健措施能够减少ISR的发生风险。     

Glutathione reductase plays an anti-apoptotic role against oxidative stress in human hepatoma cells

The cellular roles of glutathione reductase (GR) in the reactive oxygen species (ROS)-induced apoptosis were studied using the HepG2 cells transfected with GR. The overexpression of GR caused a marked enhancement in reduced and oxidized glutathione (GSH/GSSG) ratio, and significantly decreased ROS levels in the stable transfectants. Hydrogen peroxide (H₂O₂), under the optimal condition for apoptosis, significantly decreased cellular viability and total GSH content, and rather increased ROS level, apoptotic percentage and caspase-3 activity in the mock-transfected cells. However, hydrogen peroxide could not largely generate these apoptotic changes in cellular viability, ROS level, apoptotic percentage, caspase-3 activity and total GSH content in the cells overexpressing GR. Taken together, GR may play a protective role against oxidative stress.     

Effect of incubation temperature after devitrification on quality parameters in human sperm cells

Sperm cryopreservation is common in assisted reproduction laboratories, providing a therapeutic option for several clinical conditions. This process has been optimized; however, the effect of post-thaw incubation temperature has been poorly studied. This work analyzed the effect of incubation temperature after devitrification on human sperm quality. Spermatozoa from normozoospermic donors were cryopreserved by vitrification. After devitrification, the spermatozoa were separated into two aliquots: (i) incubated at room temperature (RT, 22-25 °C) and (ii) incubated at 37 °C. Reactive oxygen species (ROS), viability, mitochondrial membrane potential (ΔΨM), phosphatidylserine externalization and motility were analyzed immediately after devitrification (control) and after 2, 4 and 6 h. Spermatozoa incubated at RT showed a conserved viability and ΔΨM compared to the control, while the incubation at 37 °C promoted a decrease in these parameters. The ROS levels were increased at both incubation conditions. The progressive motility was decreased in all experimental groups and the decrease was more pronounced under incubation at RT. No increase in phosphatidylserine externalization was observed. In conclusion, prior to use in assisted reproduction procedures, devitrified spermatozoa at RT conserve a better viability and ΔΨM than at 37 °C.     

Autophagy prevents irradiation injury and maintains stemness through decreasing ROS generation in mesenchymal stem cells

Stem cells were characterized by their stemness: self-renewal and pluripotency. Mesenchymal stem cells (MSCs) are a unique type of adult stem cells that have been proven to be involved in tissue repair, immunoloregulation and tumorigenesis. Irradiation is a well-known factor that leads to functional obstacle in stem cells. However, the mechanism of stemness maintenance in human MSCs exposed to irradiation remains unknown. We demonstrated that irradiation could induce reactive oxygen species (ROS) accumulation that resulted in DNA damage and stemness injury in MSCs. Autophagy induced by starvation or rapamycin can reduce ROS accumulation-associated DNA damage and maintain stemness in MSCs. Further, inhibition of autophagy leads to augment of ROS accumulation and DNA damage, which results in the loss of stemness in MSCs. Our results indicate that autophagy may have an important role in protecting stemness of MSCs from irradiation injury.     

Oxidized LDLs affect nitric oxide and radical generation in brain endothelial cells

There is an increase in the generation of reactive oxygen species and nitric oxide in the cerebral microcirculation in Alzheimer's disease. The factors that cause this increase in oxidative stress have not been identified. Increasing evidence suggests that there are common mechanisms in atherosclerosis and Alzheimer's disease. The objective of this study was to determine the effects of oxidized low density lipoproteins (LDLs) on brain endothelial cells. Cultured rat brain endothelial cells were treated with either native LDL (10 microg/ml) or LDL oxidized in vitro using 4-hydroxy-2-nonenal (HNE-LDL) (10 microg/ml), for 24h. The results showed that HNE-LDL significantly increased production of nitric oxide (p<0.01), decreased membrane fluidity (p<0.05), and increased reactive oxygen species generation (p<0.01). These data demonstrate that oxidized LDLs affect nitric oxide and radical generation in brain endothelial cells and could contribute to cerebrovascular dysfunction in Alzheimer's disease.     

Advances in percutaneous coronary intervention for chronic total occlusions: current antegrade dissection and reentry techniques and updated algorithm

Percutaneous coronary intervention (PCI) for chronic total occlusions (CTO) is considered relatively complex with low success rates and high complication rates. Treating a CTO with PCI using the hybrid algorithm increases success rates with acceptable complication rates. An essential part of the hybrid algorithm is antegrade dissection and reentry (ADR). In PCI of a non-CTO coronary lesion, the guidewire over which the stent is advanced and placed stays within the true lumen of the coronary artery. ADR techniques make it possible to cross the lesion through the wall of the coronary artery, the subintimal space, thus creating a small bypass within the architecture of the coronary artery and restoring antegrade blood flow. ADR increases success rates, especially in more difficult CTO procedures. In the last decade, new materials and techniques have been introduced in quick succession, which are summarised in this review. Consequently an updated ADR algorithm is presented, which can support the CTO operator during an ADR procedure.

     

左室压力-应变环评估冠心病患者经皮冠状动脉介入术前后心肌做功的临床研究

摘要 目的：应用左室压力-应变环（LV-PSL）评估冠心病患者经皮冠状动脉介入术（PCI）前后心肌做功的改变。方法：选取在中国人民解放军南部战区总医院行PCI治疗的冠心病患者40例为病例组,同期健康体检者42例为对照组,应用LV-PSL评估整体做功效率（GWE）、整体做功指数（GWI）、整体无用功（GWW）、整体有用功（GCW）,比较对照组与病例组术前、术后1周、术后1个月各参数的差异,分析GWI、GCW、GWE、GWW与左室射血分数（LVEF）及左室整体纵向应变（GLS）的相关性。结果：与对照组相比,病例组术前、术后1周、术后1个月GWW升高,GCW、GWI、GWE降低（P<0.05）。与术前相比,术后1周GWI、GCW、GWE、GWW差异均无统计学意义（P>0.05）。与术前及术后1周相比,术后1个月GWW降低,GCW、GWI、GWE升高（P<0.05）。GWI、GCW、GWE、GWW与LVEF及GLS均显著相关（P<0.05）。结论：LV-PSL可定量评估冠心病患者PCI前后的心肌做功,为临床提供一种无创性评价PCI后心脏收缩功能的新方法。     

Dynamic thiol/disulphide homeostasis before and after radical prostatectomy in patients with prostate cancer

Abstract

Thiol groups are important anti-oxidants and essential molecules protecting organism against the harmful effects of reactive oxygen species (ROS). The aim of our study is to evaluate thiol–disulphide homeostasis with a novel recent automated method in patients with localized prostate cancer (PC) before and six months after radical prostatectomy (RP). 18 patients with PC and 17 healthy control subjects were enrolled into the study. Blood samples were collected from the controls subjects and patients before and six months after RP. Thiol–disulphide homeostasis was determined using a recently developed novel method. Prostate-specific antigen (PSA), albumin, total protein, total thiol, native thiol, disulphide and total antioxidant status (TAS) were measured and compared between the groups. Native thiol, total thiol and TAS levels were significantly higher in the control group than the patients before RP (p?<?.001). There was a non-significant increase in the native thiol, total thiol and TAS levels in the patients six months after RP in comparison to the levels before RP (p values .3, .3 and .09, respectively). We found a significant negative correlation between PSA and thiol levels. Our study demonstrated that the decreased thiol and TAS levels weakened anti-oxidant defence mechanism in the patients with PC as indicated. Increased oxidative stress in prostate cancer patients may cause metabolic disturbance and have a role in the aetiopathogenesis of prostate cancer.     

Genotoxicity of chloroquine in rat liver cells: Protective role of free radical scavengers

The genotoxic effect of chloroquine (CQ), a 4-aminoquinoline antimalarial drug was investigated in rat liver cells using the alkaline comet assay. Chloroquine (0–1000 μmol/L) significantly increased DNA strand breaks of rat liver cells dose-dependently. Rat liver cells exposed to CQ (100–500 μmol/L) and treated with endonuclease III and formamidopyrimidine-DNA glycosylase, the bacterial DNA repair enzymes that recognize oxidized pyrimidine and purine, respectively, showed greater DNA damage than those not treated with the enzymes, providing evidence that CQ induced oxidation of purines and pyrimidines. Treatment of cells with 5 mmol/L N-acetylcysteine, an intracellular reactive oxygen species (ROS) scavenger, and 100 μmol/L and 250 μmol/L deferoxamine, an established iron chelator, significantly decreased the CQ-induced strand breaks and base oxidation, respectively. Similarly, the formation of DNA strand breaks and oxidized bases was prevented by vitamin C (10 μmol/L) (a water-soluble antioxidant), quercetin (50 μmol/L) (an antioxidant flavonoid), and kolaviron (30 μmol/L and 90 μmol/L) (an antioxidant and a liver hepatoprotective phytochemical). The results indicate that the genotoxicity of CQ in rat liver cells might involve ROS and that free radical scavengers may elicit protective effects in these cells.