鼠尾胶原在心肌细胞氧化损伤中的保护作用

背景：胶原蛋白具有抗氧化作用,但既往在实验室主要将鼠尾胶原用于促进细胞贴壁和支架构建,对于其抗氧化作用目前尚无相关研究。目的：探讨鼠尾胶原对过氧化氢导致离体心肌细胞氧化损伤的保护作用。方法：将原代培养的SD大鼠乳鼠心肌细胞随机接种到铺有鼠尾胶原的培养皿（实验组）和普通培养皿（对照组）中,并且均用0,10,100μmol/L H2O2诱导。24 h后,以电子显微镜观察各组乳鼠心肌细胞的形态,应用MTT 比色法检测心肌细胞存活率,TUNEL 法检测心肌细胞凋亡形态,流式细胞技术检测心肌细胞凋亡率,黄嘌呤氧化酶法检测超氧化物歧化酶活力,硫代巴比妥比色法检测丙二醛水平,Western-Blot 检测凋亡蛋白Bax、Bcl-2的表达。结果与结论：两组培养皿中,随着过氧化氢浓度的增高,均可见细胞凋亡状态明显加重,细胞存活率及细胞内过氧化氢活力明显下降,细胞凋亡率及细胞内丙二醛水平明显增高,Bcl-2/Bax比值显著降低,呈剂量依赖性。而在相同浓度过氧化氢诱导下,与对照组相比,实验组细胞凋亡状态明显减弱,细胞存活率、超氧化物歧化酶活力及Bcl-2/Bax比值增高,细胞凋亡率和丙二醛水平明显减低。表明鼠尾胶原对过氧化氢所致的心肌细胞氧化损伤具有保护作用,其机制可能与提高超氧化物歧化酶活力,减少丙二醛产生及提高Bcl-2的表达有关。     

白藜芦醇对中波紫外线照射致纤维细胞氧化损伤的影响

目的探讨白藜芦醇对中波紫外线（UVB）照射致体外培养人皮肤成纤维细胞氧化损伤的预防和治疗作用。 方法实验分为5组：正常对照组、白藜芦醇组、UVB照射组、UVB照射后白藜芦醇处理组及白藜芦醇预处理后UVB照射组。细胞处理后用甲基噻唑基四唑（MTT）法检测细胞增殖活性,酶生化法检测细胞超氧化物歧化酶（SOD）活性、丙二醛（MDA）含量。 结果白藜芦醇浓度&rt;100 μmol/L时可抑制成纤维细胞增殖,<100 μmol/L时可促进成纤维细胞增殖,最佳作用浓度为50 μmol/L;UVB照射后细胞活性下降31.7%,细胞SOD活性降低、MDA产生增加;与单纯UVB照射相比,UVB照射前或照射后用白藜芦醇处理均能使成纤维细胞存活率提高、SOD活性增强、MDA产生减少。 结论低浓度白藜芦醇可促进成纤维细胞增殖,从而对因UVB照射而损伤的体外培养成纤维细胞起到保护作用。     

Nitroxide stable radicals protect beating cardiomyocytes against oxidative damage.

The protective effect of stable nitroxide radicals against oxidative damage was studied using cardiomyocyte cultures obtained from newborn rats. Monolayered cardiomyocytes were exposed to H2O2 and the effect on spontaneous beating and leakage of LDH was determined. Hydrogen peroxide irreversibly blocked rhythmic beating and resulted in a significant membrane injury as shown by release of LDH. The injury was prevented by catalase which removes H2O2 and by cell-permeable, metal-chelating agents such as desferrioxamine or bipyridine. In contrast, reagents which are excluded from the cell such as superoxide dismutase or DTPA did not protect the cells against H2O2. Five- and six-membered ring, stable nitroxide radicals which have previously been shown to chemically act as low-molecular weight, membrane-permeable, SOD-mimetic compounds provided full protection. The nitroxides prevented leakage of LDH and preserved normal cardiomyocyte contractility, presumably by intercepting intracellular O2-radicals. Alternatively, protection may result through nitroxides reacting with reduced transition metal ions or by detoxifying secondary organic radicals.     

Quercetin protects mice kidney collagen against the damage induced by cadmium fluoride

Cadmium fluoride (CdF) is a well known environmental toxin. In the present study protective effect of quercetin against CdF induced changes in renal hydroxyproline (Hyp) fractions and collagen was investigated. Five experimental groups were studied viz., Group 1—normal (control) mice; Group 2—mice administered single intraperitoneal (ip) injection of CdF₂ at a dose of 2 mg/kg body weight (bw) and sacrificed after 24 h; Group 3—mice administered single ip injection of CdF at a dose of 2 mg/kg bw but sacrificed after 48 h; Group 4—mice injected with quercetin alone at a dose of 100 mg quercetin/kg bw ip and Group 5—mice injected ip with 100 mg quercetin/kg bw followed by 2 mg CdF₂/kg bw after 2 h. Mice in Groups 4 and 5 were sacrificed 24 h after the last treatment. 2 mg/kg bw CdF caused a significant decrease in free and protein bound Hyp fractions after 24 h. However after 48 h, significant decreases were observed in all the renal Hyp fractions viz., free, peptide bound, protein bound and total Hyp fractions. There was also a significant decrease in total renal collagen after 48 h of CdF treatment. Quercetin pretreatment followed by CdF injection restored the altered parameters to near normal levels. The authors conclude that quercetin may protect the kidney collagen against CdF assault.     

Diphenyl diselenide protects cultured MCF-7 cells against tamoxifen-induced oxidative DNA damage

Diphenyl diselenide (DPDS) is an electrophilic reagent used in the synthesis of a variety of pharmacologically active organoselenium compounds. Studies have shown its interesting pharmacodinamic properties, as antioxidant, antimutagenic and antitumoral effects. Here we report the antigenotoxic properties of DPDS against tamoxifen (TAM)-induced oxidative DNA damage in MCF-7 cultured cell line. We determined the cytotoxicity by lactate dehydrogenase (LDH) leakage assay and evaluated oxidative DNA damage by modified comet assay employing the enzymes formamidopyrimidine DNA-glycosylase (Fpg) and endonuclease III (Endo III). Our results demonstrate that the cellular effects of DPDS appear to be complex and concentration-dependent. The present findings show that DPDS is not genotoxic (at concentrations lower than 2.0 μmol/L) in MCF-7 cells, as observed in the modified comet assay. Moreover, DPDS protects against TAM-induced oxidative DNA damage, probably by its antioxidant activity, without interfering with its cytotoxicity. In this manner, the treatment with low concentrations of DPDS, a synthetic organoselenium compound, could be used as a potent antigenotoxic agent to prevent the risk of cancer induction triggered by tamoxifen hormone therapy. Thereby, more studies concerning the toxicity of DPDS and its structural derivatives are still necessary for future safe therapeutic application and development of novel chemopreventive compounds for combined therapy in breast cancer.     

Effect of sesame oil against acetaminophen-induced acute oxidative hepatic damage in rats

Acetaminophen (APAP) overdose causes acute liver injury or even death in both humans and experimental animals. We investigated the effect of sesame oil on APAP-induced acute liver injury. Male Wistar rats were given APAP (1,000 mg/kg; orally) to induce acute liver injury. Acetaminophen significantly increased aspartate transaminase, alanine transaminase, lipid peroxidation, and superoxide anion and hydroxyl radical generation levels; it also induced glutathione depletion. Sesame oil (8 mL/kg; orally) did not alter the gastric absorption of APAP, but it inhibited all the parameters altered by APAP and protected the rats against APAP-induced acute liver injury. We hypothesize that sesame oil maintained the intracellular glutathione levels, reduced reactive oxygen species levels, and inhibited lipid peroxidation in rats with APAP-induced acute liver injury.     

Antioxidative mechanisms protect resistant strains of Staphylococcus aureus against ciprofloxacin oxidative damage

The aim of this investigation was to determine whether the antioxidant defences protect resistant strains of Staphylococcus aureus against ciprofloxacin oxidative damage. Reactive oxygen species (ROS) were determined by chemiluminescence and nitric oxide (NO) was assayed by Griess reaction. The accumulation of ciprofloxacin was examined by fluorometry and oxidation of protein, catalase, ferrous reduction antioxidant potency (FRAP), carbonyls and advanced oxidation protein products (AOPP), studied by spectrophotometry. Ciprofloxacin stimulated higher production of ROS and NO in the susceptible strains than in the resistant ones. There was higher accumulation of antibiotic in sensitive strains than in resistant ones, except for the most resistant strain, which accumulated an elevated amount of antibiotic. The FRAP/ciprofloxacin accumulation ratio of the antibiotic was lower in sensitive than in resistant strains. The most resistant strain exhibited the highest FRAP and presented a high catalase activity. There was oxidation of proteins in the presence of ciprofloxacin, with the carbonyl residues increasing in sensitive and resistant S. aureus. The degradation of carbonyls to AOPP in oxidized proteins was higher in the resistant than in sensitive strains. In conclusion, an increase in antioxidant capacity and a rapid oxidation of carbonyls to AOPP contributed to resistance to ciprofloxacin.     

肿节风水提物抗放射诱导氧化损伤的临床观察

目的观察肿节风水提物抗氧化损伤的临床疗效。方法选择Ⅲ和Ⅳa期鼻咽癌（NPC）患者46例随机分为对照组和研究组,其中对照组25例,研究组21例。所有患者均采用放化综合治疗的方法,研究组配合单剂中药肿节风水提物20g／d口服。分,NN定治疗前后血浆总超氧化物歧化酶（SOD）活力和丙二醛（MDA）含量,并观察2组治疗后肿瘤消退情况和急性放化疗副反应。结果所有患者均完成治疗后与治疗前比较,对照组血浆总SOD活力降低,血浆MDA含量升高;而研究组二者均降低,治疗后研究组血浆总SOD活力高于对照组,MDA含量显著低于对照组,差异均有统计学意义（P〈0．05）。放疗后两组肿瘤局部控制率无差异（P〉0．05）。研究组的急性放化疗副反应出现的频率和严重度均低于对照组,差异有统计学意义（P〈0．05）。结论肿节风水提物有抗氧化损伤的作用,与放化疗结合治疗可使Ⅲ和Ⅳa期NPC肿瘤消退满意,对放化疗所致的副反应有一定消除作用。     

The protective effect of eugenol against gentamicin-induced nephrotoxicity and oxidative damage in rat kidney

Gentamicin (GM) is an effective aminoglycoside antibiotic against life-threatening Gram-negative bacteria. However, a major complication of therapeutic doses of GM is nephrotoxicity, which is believed to be related to the generation of reactive oxygen species. The present study was therefore aimed to investigate the protective effect of eugenol, a phenolic antioxidant, on GM-induced nephrotoxicity in Sprague-Dawley rats. Intramuscular injection of rats with GM (80 mg/kg body weight/day) for six consecutive days induced marked acute renal failure, manifested by a sharp significant increase in serum urea and creatinine levels, along with a significant depletion of serum potassium level, compared to normal controls. GM-induced renal dysfunction was attributable to enhanced oxidative stress, as revealed by decreased superoxide dismutase and catalase activities, glutathione depletion and increased lipid peroxidation. Furthermore, kidney lactate dehydrogenase activity, as an indicator of hypoxia, was significantly increased by GM administration. Eugenol (100 mg/kg body weight, per os) administered four days before and six days concurrently with GM (80 mg/kg body weight, i.m.) restored normal renal functions and suppressed GM-induced oxidative stress and hypoxia. Light microscopical examination of the renal tissues of GM-treated animals demonstrated severe tubular necrosis at the cortex and increased cellular inflammatory processes. However, these alterations were considerably reduced with eugenol coadministration. In conclusion, eugenol ameliorates GM-induced nephrotoxicity and oxidative damage by scavenging oxygen free radicals, decreasing lipid peroxidation and improving intracellular antioxidant defense.     

Pefloxacin-induced achilles tendon toxicity in rodents: biochemical changes in proteoglycan synthesis and oxidative damage to collagen

Despite a relatively low incidence of serious side effects, fluoroquinolones and the fluoroquinolone pefloxacin have been reported to occasionally promote tendinopathy that might result in the complication of spontaneous rupture of tendons. In the present study, we investigated in rodents the intrinsic deleterious effect of pefloxacin (400 mg/kg of body weight) on Achilles tendon proteoglycans and collagen. Proteoglycan synthesis was determined by measurement of in vivo and ex vivo radiosulfate incorporation in mice. Collagen oxidative modifications were measured by carbonyl derivative detection by Western blotting. An experimental model of tendinous ischemia (2 h) and reperfusion (3 days) was achieved in rats. Biphasic changes in proteoglycan synthesis were observed after a single administration of pefloxacin, consisting of an early inhibition followed by a repair-like phase. The depletion phase was accompanied by a marked decrease in the endogenous serum sulfate level and a concomitant increase in the level of sulfate excretion in urine. Studies of ex vivo proteoglycan synthesis confirmed the in vivo results that were obtained. The decrease in proteoglycan anabolism seemed to be a direct effect of pefloxacin on tissue metabolism rather than a consequence of the low concentration of sulfate. Pefloxacin treatment for several days induced oxidative damage of type I collagen, with the alterations being identical to those observed in the experimental tendinous ischemia and reperfusion model. Oxidative damage was prevented by coadministration of N-acetylcysteine (150 mg/kg) to the mice. These results provide the first experimental evidence of a pefloxacin-induced oxidative stress in the Achilles tendon that altered proteoglycan anabolism and oxidized collagen.     

Human plasma and tirilazad mesylate protect stored human erythrocytes against the oxidative damage of gamma-irradiation

Transfusion-associated graft-versus-host disease (TA-GVHD) is a serious condition that under certain circumstances can be lethal in immunosuppressed patients. The risk of TA-GVHD can be reduced in this population by gamma irradiation (gammaRad) of blood components. gammaRad results in production of reactive oxygen species which can damage red blood cells (RBC). Tirilazad mesylate (TM) is a member of the 21-aminosteroids (Lazaroids) family and is a powerful antioxidant. We investigated the ability of TM and human plasma (which contain powerful antioxidants) to protect stored human RBC against the oxidative damage of gammaRad. Fresh intact packed RBC obtained from the normal donors, with and without autologous plasma or TM (0.05 mg mL-1 RBC), were exposed to gammaRad (50 Gy) and stored for 28 days at 4 degrees C. Oxidative damage was assessed by osmotic fragility at 65 mM NaCl concentration (expressed by percentage haemolysis in 65 mM NaCl solution) and lipid peroxidation (measured by thiobarbituric acid reactive substances, TBARS). Our results showed that storage and irradiation of untreated intact RBC increased the osmotic fragility at 65 mM NaCl concentration (65.8 +/- 1.3 vs. 51.20 +/- 0.87% haemolysis; irradiated vs. controls, respectively; P = 0.002) and lipid peroxidation (TBARS = 4.47 +/- 0. 12 vs. 3.45 +/- 0.09 microM L-1 RBC; irradiated vs. controls, respectively; P = 0.001). TM protected the intact RBC against radiation-induced haemolysis (35.8 +/- 5.0 vs. 65.8 +/- 1.3% haemolysis; treated vs. untreated irradiated RBC, respectively; P = 0.02) and lipid peroxidation (TBARS = 2.91 +/- 0.2 vs. 4.47 +/- 0.12 microM L-1 RBC; treated vs. untreated irradiated RBC, respectively; P = 0.005). Addition of autologous plasma to packed RBC significantly reduced the extent of radiation-induced haemolysis by more than six-fold (12.45 +/- 0.26 vs. 65.8 +/- 2.2% haemolysis; irradiated RBC with versus without plasma, respectively; P = 0.0001). In conclusion, these results show that irradiation and storage of blood damages RBC via oxidative processes and addition of autologous plasma and/or TM protects RBC against such damage and possibly enhances their storage and survival.     

Pretreatment with melatonin protects against cyclophosphamide‐induced oxidative stress and renal damage in mice

Cyclophosphamide (CP) is widely used in treatment of different cancers. Nephrotoxicity is one of the dose‐limiting side effects of CP. This study was carried out to investigate the effect of melatonin (MEL) on CP‐induced nephrotoxicity in mice. In this study, 50 Swiss albino mice (20–25 g) were randomly divided into five groups. Mice were pretreated with MEL intraperitoneally (i.p) in doses of 5, 10 and 20 mg/kg for five consecutive days, and CP (200 mg/kg, i.p) was administrated on the 5th day 1 h after the last dose of MEL. Then on day 6, blood samples were collected to determine serum creatinine (Cr) and blood urea nitrogen (BUN) levels. The kidneys were used for histological examination, biochemical assays and real‐time PCR studies. Malondialdehyde (MDA), glutathione (GSH), protein carbonyl (PC), nitric oxide (NO) level, catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx) and myeloperoxidase (MPO) activity were assessed in renal tissue. In addition, the expression of SOD2 and PGx1 was measured using real‐time PCR method in renal tissue. Results showed that CP administration significantly increases Cr, BUN, MDA, PC, NO level and MPO activity. It also decreases renal GSH level, SOD, GPx and CAT activity. Pretreatment with MEL (especially 20 mg/kg, i.p.) for 5 days prevented these changes; however, it did not affect the SOD activity. Our results revealed that MEL might be useful for prevention of the nephrotoxicity induced by CP through ameliorative effects on biochemical indices and oxidative stress parameters.     

Liquid ventilation attenuates pulmonary oxidative damage

PURPOSE: Liquid perfluorochemicals reduce the production of reaction oxygen species by alveolar macrophages. We sought to determine whether the use of liquid perfluorochemicals in vivo during liquid ventilation would attenuate oxidative damage to the lung. MATERIALS AND METHODS: Healthy infant piglets (n = 16) were instrumented for mechanical ventilation and received intravenous oleic acid to create an acute lung injury. The animals were assigned to a nontreatment group receiving conventional mechanical ventilation or a treatment group receiving partial liquid ventilation with a liquid perfluorochemical. Following sacrifice, the bronchoalveolar lavage and lung parenchyma were analyzed for evidence of oxidative damage to lipids and proteins by determination of TBARS and carbonylated protein residues, respectively. RESULTS: Mortality in the control group was 50% at the completion of the study compared with no deaths in the partial liquid ventilation group (P = .025). The alveolar-arterial oxygen difference was more favorable following injury in the partial liquid ventilation group. The liquid ventilation group demonstrated a 32% reduction in TBARS (P = .043) and a 14% reduction in carbonylated protein residues (P = .061). CONCLUSION: These data suggest that partial liquid ventilation supports gas exchange and reduces mortality in association with a reduction in the production of reactive oxygen species and the concomitant attenuation of tissue damage during the early phase of acute lung injury.     

Glutathione-dependent protection against oxidative injury

Functions of GSH in detoxication during radical-induced injury in specific pathological and toxicological conditions are discussed. GSH protects against oxidative damage in systems that scavenge radicals, eliminate lipid peroxidation products, preserve thiol-disulfide status of proteins, and repair oxidant damage. Several factors which affect cellular GSH homeostasis can affect these functions, including nutritional status, hypoxia and pharmacological intervention. Evidence from a variety of pathological and toxicological conditions, e.g. ischemia-reperfusion injury, chemically induced oxidative injury, radiation damage, aging, and degenerative diseases, indicate that GSH is a primary component of physiological systems to protect against oxidant and free-radical-mediated cell injury.     

Isotretinoin therapy induces DNA oxidative damage.

BACKGROUND: Isotretinoin (Iso) is currently indicated for the treatment of cystic acne (CA) and is related to marked teratogenicity. AIM: The aim of the study was to evaluate the relationship between total antioxidant status (TAS) and a serum marker of DNA oxidative damage, 8-hydroxy-2-desoxyguanosine (8-OHdG), in patients on Iso treatment. PATIENTS AND METHODS: Patients with CA (n=18) were evaluated before and 45 days after Iso (0.5 mg/kg per day) treatment and non-diseased controls (n=22) were tested only once. Plasma TAS levels and 8-OHdG were measured spectrophotometrically and with an immunoassay, respectively. Liver biochemical parameters and muscle enzymes were measured on a blood chemistry analyzer. RESULTS: TAS levels were significantly (p<0.0001) lower in patients before treatment (921+/-124 micromol/L) compared with those after treatment (1335+/-93 micromol/L) and in controls (1536+/-126 micromol/L). In contrast, 8-OHdG serum levels were two-fold higher in patients after treatment (0.21+/-0.03 ng/mL) than before treatment (0.11+/-0.02 ng/mL) and three-fold higher than in controls (0.07+/-0.01 ng/mL; p<0.0001). Negative correlations were found between TAS and 8-OHdG (r=-0.754, p<0.0001) in patients before therapy and positive correlations were found between creatine kinase (CK) and 8-OHdG (r=0.488, p<0.001) and liver enzymes after Iso treatment. CONCLUSIONS: High serum levels of 8-OHdG in patients on Iso therapy may be due to a direct effect of Iso on liver, muscle and skin epidermal cells. Regular evaluation of 8-OHdG in sera of patients, especially of women of reproductive age, on Iso treatment could be a sensitive follow-up biomarker of DNA oxidation.     

Biomarkers of oxidative damage in human disease

Oxidative/nitrosative stress, a pervasive condition of increased amounts of reactive oxygen/nitrogen species, is now recognized to be a prominent feature of many acute and chronic diseases and even of the normal aging process. However, definitive evidence for this association has often been lacking because of recognized shortcomings with biomarkers and/or methods available to assess oxidative stress status in humans. Emphasis is now being placed on biomarkers of oxidative stress, which are objectively measured and evaluated as indicators of normal biological processes, pathogenic processes, or pharmacologic responses to therapeutic intervention. To be a predictor of disease, a biomarker must be validated. Validation criteria include intrinsic qualities such as specificity, sensitivity, degree of inter- and intraindividual variability, and knowledge of the confounding and modifying factors. In addition, characteristics of the sampling and analytical procedures are of relevance, including constraints and noninvasiveness of sampling, stability of potential biomarkers, and the simplicity, sensitivity, specificity, and speed of the analytical method. Here we discuss some of the more commonly used biomarkers of oxidative/nitrosative damage and include selected examples of human studies.     

Protective effects of liposomes encapsulating ferulic acid against CCl4-induced oxidative liver damage in vivo rat model

Antioxidants are useful for the treatment of oxidative stress mediated liver damage. A naturally occurring antioxidant γ-oryzanol is rapidly hydrolyzed to its active hydrophobic metabolite, ferulic acid, inside the body. Limitations associated with the hydrophobicity of ferulic acid can be overcome by encapsulating in a liposomal formulation. As intravenously administered nanoparticles (including liposomes) can effectively reach the liver, such systems may be suitable drug delivery carriers to treat liver injury. In this study, we prepared a liposomal formulation of ferulic acid (ferulic-lipo) and examined its effects on liver damage induced by CCl₄. Ferulic-lipo were ~100 nm in size and drug encapsulation efficiency was about 92%. Ferulic-lipo showed potent scavenging efficacy against hydroxyl radical compared to α-tocopherol liposomes. Ferulic-lipo significantly prevented CCl₄-mediated cytotoxicity in human hepatocarcinoma cells. Furthermore, intravenous administration of ferulic-lipo significantly reduced alanine aminotransferase and aspartate amino transferase levels in a rat model of liver injury. CCl₄-mediated reactive oxygen species generation in liver was also reduced by intravenous administration of ferulic-lipo. Hepatoprotective effects of ferulic-lipo were demonstrated by histological observation of CCl₄-induced liver tissue damage. Therefore, ferulic-lipo exhibit potent antioxidative capacity and were suggested to be an effective formulation for prevention of oxidative damage of liver tissue.     

Antioxidant gene therapy against neuronal cell death

Oxidative stress is a common hallmark of neuronal cell death associated with neurodegenerative disorders such as Alzheimer's disease, Parkinson's disease, as well as brain stroke/ischemia and traumatic brain injury. Increased accumulation of reactive species of both oxygen (ROS) and nitrogen (RNS) has been implicated in mitochondrial dysfunction, energy impairment, alterations in metal homeostasis and accumulation of aggregated proteins observed in neurodegenerative disorders, which lead to the activation/modulation of cell death mechanisms that include apoptotic, necrotic and autophagic pathways. Thus, the design of novel antioxidant strategies to selectively target oxidative stress and redox imbalance might represent important therapeutic approaches against neurological disorders. This work reviews the evidence demonstrating the ability of genetically encoded antioxidant systems to selectively counteract neuronal cell loss in neurodegenerative diseases and ischemic brain damage. Because gene therapy approaches to treat inherited and acquired disorders offer many unique advantages over conventional therapeutic approaches, we discussed basic research/clinical evidence and the potential of virus-mediated gene delivery techniques for antioxidant gene therapy.