Temozolomide Drives Ferroptosis via a DMT1-Dependent Pathway in Glioblastoma Cells

PurposeTemozolomide is used in first-line treatment for glioblastoma. However, chemoresistance to temozolomide is common in glioma patients. In addition, mechanisms for the anti-tumor effects of temozolomide are largely unknown. Ferroptosis is a form of programmed cell death triggered by disturbed redox homeostasis, overloaded iron, and increased lipid peroxidation. The present study was performed to elucidate the involvement of ferroptosis in the anti-tumor mechanisms of temozolomide.Materials and MethodsWe utilized the CCK8 assay to evaluate cytotoxicity. Levels of lactate dehydrogenase (LDH), malondialdehyde (MDA), iron, and glutathione (GSH) were measured. Flow cytometry and fluorescence microscope were used to detect the production of reactive oxygen species (ROS). Western blotting, RT-PCR and siRNA transfection were used to investigate molecular mechanisms.ResultsTemozolomide increased the levels of LDH, MDA, and iron and reduced GSH levels in TG905 cells. Furthermore, we found that ROS levels and DMT1 expression were elevated in TG905 cells treated with temozolomide and were accompanied by a decrease in the expression of glutathione peroxidase 4, indicating an iron-dependent cell death, ferroptosis. Our results also showed that temozolomide-induced ferroptosis is associated with regulation of the Nrf2/HO-1 pathway. Conversely, DMT1 knockdown by siRNA evidently blocked temozolomide-induced ferroptosis in TG905 cells.ConclusionTaken together, our findings indicate that temozolomide may suppress cell growth partly by inducing ferroptosis by targeting DMT1 expression in glioblastoma cells.     

Differential sensitivity of inflammatory macrophages and alternatively activated macrophages to ferroptosis

Acumulation of oxidized membrane lipids ultimately results in ferroptotic cell death, which can be prevented by the selenoenzyme glutathione peroxidase 4 (Gpx4). In vivo conditions promoting ferroptosis and susceptible cell types are still poorly defined. In this study, we analyzed the conditional deletion of Gpx4 in mice specifically in the myeloid cell lineages. Surprisingly, development and maintenance of LysM⁺ macrophages and neutrophils, as well as CD11c⁺ monocyte-derived macrophages and dendritic cells were unaffected in the absence of Gpx4. Gpx4-deficient macrophages mounted an unaltered proinflammatory cytokine response including IL-1β production following stimulation with TLR ligands and activation of several inflammasomes. Accordingly, Gpx4^fl/flLysM-cre mice were protected from bacterial and protozoan infections. Despite having the capacity to differentiate to alternatively activated macrophages (AAM), these cells lacking Gpx4 triggered ferroptosis both in vitro and in vivo following IL-4 overexpression and nematode infection. Exposure to nitric oxide restored viability of Gpx4-deficient AAM, while inhibition of iNOS in proinflammatory macrophages had no effect. These data together suggest that activation cues of tissue macrophages determine sensitivity to lipid peroxidation and ferroptotic cell death.     

HIV-1 Tat蛋白对U87细胞活性及氧化应激的影响

目的:研究HAD和非HAD的艾滋病患者不同部位来源的Tat蛋白氨基酸位点变异及其对U87细胞氧化应激的影响。方法:采用BLAST和MEGA6对一例HAD患者(H)和一例非HAD患者(N)的基底节(BG)、脾脏(SPL)共4个部位来源的HIV-1 Tat蛋白进行序列分析,研究其氨基酸位点变异,并将 tat基因...     

Antioxidant parameters and ageing in some animal species

Connection between ageing and some tissue antioxidant parameters have been studied in four experiments on different animal species. Prenatal studies on the developing chick embryos showed discrepancies between the lipid-rich liver and brain antioxidant defence. In the liver, high levels of reduced glutathione (GSH), vitamins A and E and high activities of the antioxidant enzymes glutathione peroxidase (GPX) and superoxide dismutase (SOD) were found whereas brain expressed a high vitamin C concentration. In newborn healthy calves during the first two days of life, atmospheric oxygen tension did not cause either increased lipid peroxidation as reflected in a high malondialdehyde (MDA) level or any changes in GSII, GPX, SOD and catalase (CAT) activities in red blood cells (RBC). Plasma vitamin E and carotene concentrations also did not change. In growing healthy calves during two months after birth increasing MDA, decreasing GSH, GPX and CAT are leading features, whereas plasma vitamin E and carotene concentrations significantly increased. In young (1-year-old) and old (9-year-old) dogs RBC results showed significant differences with the highest MDA and lowest GSH levels in the old males. Activity of GPX and SOD was higher in old dogs than in the young ones, especially in the females.     

核因子E2相关因子2对人永生化角质形成细胞氧化损伤的保护作用

目的 探讨核因子E2相关因子2(Nrf2)对过氧化氢(H2O2)诱导的人永生化角质形成细胞(HaCaT)氧化损伤的保护作用.方法 用慢病毒转染方式使HaCaT过表达Nrf2(Nrf2/HaCaT),并用MTT法和抗Ki67免疫荧光染色法检测其增殖活性.实验分为Nrf2/HaCaT组和HaCaT组,每组5个样本.应用20...     

VCP relocalization limits mitochondrial activity,GSH depletion and ferroptosis during starvation in PC3 prostate cancer cells

During periods of crisis, cells must compensate to survive. To this end, cells may need to alter the subcellular localization of crucial proteins. Here, we show that during starvation, VCP, the most abundant soluble ATPase, relocalizes and forms aggregate-like structures at perinuclear regions in PC3 prostate cancer cells. This movement is associated with a lowered metabolic state, in which mitochondrial activity and ROS production are reduced. VCP appears to explicitly sense glutamine levels, as removal of glutamine from complete medium triggered VCP relocalization and its addition to starvation media blunted VCP relocalization. Cells cultured in Gln(+) starvation media exhibited uniformly distributed VCP in the cytoplasm (free VCP) and underwent ferroptotic cell death, which was associated with a decrease in GSH levels. Moreover, the addition of a VCP inhibitor, CB-5083, in starvation media prevented VCP relocalization and triggered ferroptotic cell death. Likewise, expression of GFP-fused VCP proteins, irrespective of ATPase activities, displayed free VCP and triggered cell death during starvation. These results indicate that free VCP is essential for the maintenance of mitochondrial function and that PC3 cells employ a strategy of VCP self-aggregation to suppress mitochondrial activity in order to escape cell death during starvation, a novel VCP-mediated survival mechanism.     

Gastrodin improves cognitive dysfunction and decreases oxidative stress in vascular dementia rats induced by chronic ischemia

Objective: To study the potential protective effects of gastrodin on reducing tissue oxidative stress and attenuating cognitive deficits in vascular dementia induced by cerebral chronic hyperfusion. To explore the detailed molecular mechanisms. Methods: 6 to 8 week old male Wistar rats were adopted as experimental animals. Animals were divided into the following groups: Group 1 (sham group with no occlusion), Group 2 (control group with 2VO procedure), Group 3 (sham group with gastrodin administration), Group 4 (2VO group with gastrodin administration). Morris water maze (MWM) test was adopted to test the learning and memory function of rats within different groups. MDA, glutathione peroxidase and total thiol assessment was done to reflect the oxidative stress in the brain tissue. Cell counting kit-8 (CCK8) and flow cytometry (FCM) were performed to examine the cell viability and apoptosis rate of SH-SY5Y cells induced by hydrogen peroxide and rescued by gastrodin treatments. Reactive oxygen species (ROS) generation was determined by the 2’, 7’-dichlorofluorescein diacetate (DCFH-DA) assay. qPCR and Western blot (WB) were adopted to detect the molecular mechanisms related to the anti-apoptosis and ROS scavenging effects of gastrodin. Results: Our results indicated an obvious protective effect of gastrodin on vascular dementia induced brain ischemia. Administration of gastrodin could improve the impaired learning and memory function induced by 2VO procedure in rats. The levels of MDA were partially decreased by the administration of gastrodin. The levels of glutathione peroxidase and total thiol were partially restored by the administration of gastrodin. Cell viability was improved by gastrodin in a dose-dependent pattern on SH-SY5Y cells induced by hydrogen peroxide (P < 0.05). Cell apoptosis rate was reduced by gastrodin in a dose-dependent pattern on SH-SY5Y cells induced by hydrogen peroxide (P < 0.05). Gastrodin could scavenge ROS generation induced by pre-treatment of hydrogen peroxide. Both qPCR and WB results showed significant enhancements on the expression levels of NFE2L2, ADH7, GPX2 and GPX3 (P < 0.05). Conclusion: Gastrodin administration is protective on the learning and memory functions that might be affected by vascular dementia induced oxidative stress due to brain ischemia. On the molecular level, NFE2L2, ADH7, GPX2 and GPX3 were up regulated by gastrodin.     

IL-6预处理保护H2O2致心肌细胞损伤作用机制初探

目的初步探讨IL-6预处理对H2O2致心肌细胞氧化应激损伤的作用机制。方法采用心肌细胞原代培养方法,以H2O2刺激心肌细胞,建立细胞氧化应激模型;采用MTT法检测细胞活力;AnnexinV-FITC染色法和流式细胞术检测细胞凋亡率;检测心肌细胞内谷胱甘肽（GSH）、超氧化物歧化酶（SOD）、丙二醛（MDA）的表达情况。结果 H2O2可降低心肌细胞存活率并能增加其凋亡,IL-6预处理后能显著改善细胞活力及凋亡情况,与模型组相比差异有统计学意义（P〈0.05）。低浓度IL-6能明显增加细胞内GSH与SOD的水平,并降低MDA的含量,随着IL-6浓度的增加,这种效应逐渐消失。结论 IL-6预处理能保护H2O2致心肌细胞损伤作用,这可能与IL-6调节细胞GSH、SOD、MDA表达有关。     

A cellular model for Friedreich Ataxia reveals small-molecule glutathione peroxidase mimetics as novel treatment strategy

Friedreich Ataxia (FRDA), the most prevalent of the inherited ataxias, is a multi-systemic disease with loss of sensory neurons and life-threatening hypertrophic cardiomyopathy as its most severe manifestations. Reduced levels of the mitochondrial protein frataxin lead to cell-damaging oxidative stress and consequently FRDA is considered as a model for more common neurodegenerative disorders in which reactive radicals and oxidative stress are involved. We have developed a cellular assay system that discriminates between fibroblasts from FRDA patients and unaffected donors on the basis of their sensitivity to pharmacological inhibition of de novo synthesis of glutathione. With this assay we observed that supplementation with selenium effectively improved the viability of FRDA fibroblasts, indicating that basal selenium concentrations are not sufficient to allow an adequate increase in the activity of certain detoxification enzymes (such as GPX). Furthermore, we characterized potential drug candidates and found that idebenone, a mitochondrially localized antioxidant that ameliorates cardiomyopathy in FRDA patients, as well as other lipophilic antioxidants protected FRDA cells from cell death. Our results also demonstrate for the first time that small-molecule GPX mimetics have potential as a novel treatment strategy for Friedreich Ataxia and presumably also for other neurodegenerative diseases with mitochondrial impairment.     

Ammonium Ferric Citrate induced Ferroptosis in Non-Small-Cell Lung Carcinoma through the inhibition of GPX4-GSS/GSR-GGT axis activity

The morbidity and mortality rates associated with non-small-cell lung carcinoma (NSCLC) are increasing every year, placing new demands on existing therapies and drugs. Ammonium ferric citrate (AFC) is often used as a food additive for iron supplementation; however, to our knowledge, no studies have investigated whether AFC can induce ferroptosis in NSCLC. In this study, we demonstrated that specific concentrations of AFC effectively inhibit the proliferation and invasion of lung cancer cell lines in vitro using a cell proliferation inhibition test, a transwell assay, and flow cytometry analysis of cell cycle and apoptosis. In addition, AFC significantly induced oxidative stress injury in lung cancer cell lines. A quantitative polymerase chain reaction assay showed that AFC markedly reduced the expression levels of cell growth factors, negative regulators of ferroptosis, and autophagy regulators. Lastly, a protein-protein interaction analysis revealed that glutathione peroxidase 4 (GPX4) exerted its biological role through the regulation of the GSS/GSR complex and downstream GGT family proteins. When the expression of GPX4 changes, its biological activities, such as the glutathione metabolic process, cellular biosynthetic process, cellular response to chemical stimulus, and antioxidant activity, change accordingly, thereby affecting the survival quality and physiological and biochemical activities of cells. Overall, this study verifies that AFC has the biological activity of activating oxidative stress injury in NSCLC cell lines, leading to a decrease in their autophagy and inducing ferroptosis. We also confirmed that the GPX4-GSS/GSR-GGT axis is a crucial target of AFC-induced ferroptosis.     

抗衰老Klotho蛋白对高糖作用下血管内皮细胞的保护作用

目的:研究抗衰老Klotho蛋白对高糖作用下血管内皮细胞的保护作用及其作用机制。方法:体外培养人脐静脉血管内皮细胞(HUVECs),设置PBS对照组、5.5 mmol/L葡萄糖组、33.3 mmol/L葡萄糖组、0.1μmol/L Klotho+33.3 mmol/L葡萄糖组、1μmol/L Klotho+33.3 mmol/L葡萄糖组和10μmol/L Klotho+33.3 mmol/L葡萄糖组。使用MTT法检测各组细胞活力;同时检测各组细胞培养上清中丙二醛(MDA)的含量以及乳酸脱氢酶(LDH)、超氧化物歧化酶(SOD)和还原型谷胱甘肽(GSH)的活性;流式细胞术检测各组细胞中活性氧(ROS)含量变化;ELISA检测各组细胞培养液中一氧化氮(NO)、内皮素1(ET-1)和细胞间黏附分子1(ICAM-1)的浓度变化;Western blot法检测各组细胞中核因子κB(NF-κB)蛋白的表达。结果:与PBS对照组相比,33.3 mmol/L葡萄糖能够显著降低HUVECs的细胞活力,增加细胞中ROS的含量,增加细胞培养上清中LDH活性和MDA含量,降低SOD和GSH的活性,同时降低NO分泌,诱导ET-1、ICAM-1分泌及细胞中NF-κB蛋白的表达(P0.05)。不同浓度Klotho蛋白和33.3 mmol/L高糖同时作用HUVECs时,细胞活力逐渐上升,ROS和MDA含量以及LDH活性逐渐下降,SOD和GSH活性则逐渐上升,同时NO分泌增加,ET-1、ICAM-1分泌及NF-κB蛋白表达显著下降(P0.05)。结论:抗衰老Klotho蛋白能够提升高糖作用下HUVECs的细胞活力,减少高糖诱导的ROS生成及氧化损伤,恢复HUVECs的正常分泌功能,并通过减少NF-κB蛋白表达发挥抗损伤作用。     

Ferroptosis: bug or feature?

Ferroptosis is an iron-dependent, oxidative form of non-apoptotic cell death. This form of cell death does not share morphological, biochemical, or genetic similarities with classic necrosis, necroptosis, parthanatos, or other forms of non-apoptotic cell death. Ferroptosis can be triggered by depleting the cell of the amino acid cysteine, or by inhibiting the phospholipid hydroperoxidase glutathione peroxidase 4 (GPX4). Why certain stimuli trigger ferroptosis instead of another form of cell death, and whether this process could be adaptive in vivo, are two major unanswered questions concerning this process. Emerging evidence and consideration of related non-apoptotic pathways suggest that ferroptosis could be an adaptive process, albeit one regulated and executed in a manner very different from apoptosis and other forms of cell death.     

Glutathione depletion induces differential apoptosis in cells of mouse retina, in vivo

Oxidative stress affects numerous intracellular macromolecules, and may result in cell death unless precisely regulated. Unregulated oxidative stress can be controlled by various cellular defense mechanisms such as glutathione (GSH) which can critically counteract the damaging effects of oxidative stress in mammalian cells. We determined the effects of unregulated oxidative stress induced by GSH depletion on cells in mouse retina. Mice were intraperitoneally injected with buthionine sulphoximine (BSO) at 1.5 g/kg. After 0, 1, 4, and 7 days of BSO administration, retinas were excised and sections were subjected to GSH assay and terminal uridine deoxynucleotidyl nick end labeling (TUNEL) analysis. After 4 days of BSO administration, the number of TUNEL positive cells was significantly increased. However, after 7 days, TUNEL positive cells returned to the basal level. The retinal region most affected by the BSO treatment appeared to be the outer nuclear layer where the photoreceptor cells reside. Different from cells in other regions, retinal cells in the inner nuclear layer increased in their apoptosis even after the first day of BSO injection, and the increase was further potentiated after 4 days. Taken together, our studies suggested that GSH depletion may cause unregulated oxidative stress to the cells in the retina and indeed increased cell death in the retina. The cells in the inner nuclear layer seemed to be affected earlier than the cells in other layers of the retina. The GSH level in the retina may be a crucial therapeutic target in preventing blindness.     

Protective effect of lycopene against ochratoxin A induced renal oxidative stress and apoptosis in rats

This study was designed to investigate the possible protective effect of lycopene against the renal toxic effects of OTA. Male Sprague-Dawley rats (<200 g, n = 6) were treated with OTA (0.5 mg/kg/day) and/or lycopene (5 mg/kg/day) by gavage for 14 days. Histopathological examinations were performed and apoptotic cell death in both cortex and medulla was evaluated by TUNEL assay. Besides, biochemical parameters and activities of renal antioxidant selenoenzymes [glutathione peroxidase 1 (GPx1), thioredoxin reductase (TrxR)], catalase (CAT), superoxide dismutase (SOD); concentrations of total glutathione (GSH), and malondialdehyde (MDA) levels were measured. OTA treatment was found to induce oxidative stress in rat kidney, as evidenced by marked decreases in CAT (35%) activity and GSH levels (44%) as well as increase in SOD activity (22%) vs control group. Furthermore, TUNEL analysis revealed a significant increase in the number of TUNEL-positive cells in cortex (49%) and medulla (75%) in OTA administrated group compared to control (p < 0.05). Lycopene supplementation with OTA increased GPx1 activity and GSH levels, and decreased apoptotic cell death in both cortex and medulla vs. control. The results of this study showed that at least one of the mechanisms underlying the renal toxicity of OTA is oxidative stress and apoptosis is the major form of cell death caused by OTA. Besides, our data indicate that the natural antioxidant lycopene might be partially protective against OTA-induced nephrotoxicity and oxidative stress in rat.     

Physical activity, aerobic capacity and selected markers of oxidative stress and the anti-oxidant defence system in healthy active elderly men.

The relationship of oxidative stress and the anti-oxidant defence system to maximal oxygen consumption (VO2max) and habitual physical activity was assessed in 26 elderly men (71.0 +/- 4.2 years) and compared to that of 12 young men (22.1 +/- 5.1 years). Physical activity was assessed by a questionnaire. Malondialdehyde (MDA), plasma total anti-oxidant status (TAS), the levels of red blood cell (RBC) superoxide dismutase (SOD) and glutathione peroxidase (GPX), as well as serum GPX activities were determined under resting conditions. The older and young men had similar TAS and RBC SOD, while MDA, RBC GPX and plasma GPX were higher, and RBC SOD/GPX ratio was significantly lower in the older men. Neither MDA nor anti-oxidants were associated with any of the physical activity/aerobic capacity measures in the elderly men. We conclude that in healthy elderly men with a good nutritional status, indicators of the anti-oxidant defence system are not lower in comparison with young men. Increased RBC and plasma GPX coupled with a high level of lipid peroxidation marker may indicate an adaptation of anti-oxidant defences to sustained oxidative stress. Furthermore, the results of the present study suggest that the level of habitual physical activity and aerobic capacity have no major influence on the resting balance between radical generation and blood anti-oxidant potential in healthy older men.     

介孔二氧化硅纳米颗粒经氧化应激所致肾细胞毒性及GSK-3β在其中的作用

研究氧化应激及GSK-3β在介孔二氧化硅纳米颗粒(MSNs)诱导肾细胞毒性中的作用及机制,以及抗氧化剂N-乙酰半胱氨酸(NAC)在此毒性中的保护作用。选用NRK-52E大鼠的肾小管上皮细胞,将其直接或用1 μmol/L NAC预处理,然后暴露于400 μg/mL MSNs中。采用MTT法测定细胞活力,采用JC-1荧光染色法检测线粒体膜电势,采用western blot 法检测GSK-3β等蛋白水平,并测定抗氧化物SOD、GSH和CAT活性。NRK-52E细胞暴露于MSNs内24 h即出现严重的细胞毒性,其IC₅₀为(438.6±7.1) μg/mL。400 μg/mL MSNs作用24 h后,细胞的存活率下降到47.57%±2.03%,胞内SOD、GSH、CAT活性水平分别下降到39.74%±2.23%、51.42%±3.08%、46.05%±3.71%(P<0.001)。400 μg/mL MSNs还可显著活化GSK-3β,崩解线粒体ΔΨ_m,促进线粒体Cyt C漏出,并剪切激活Caspase-3,引起细胞死亡(P<0.001);1 μmol/L NAC干预后可显著缓解400 μg/mL MSNs引起的这些变化(P<0.01)。MSNs通过激活GSK-3β信号通路来诱导肾细胞氧化应激损伤,NAC可以改善线粒体功能,提高细胞抗氧化能力,减轻此损伤。     

Decrease of the inhibition of lipid peroxidation by glutathione-dependent system in erythrocytes of non-insulin dependent diabetics

Summary The inhibition of lipid peroxidation of erythrocyte membranes by glutathione-dependent protection was studied in patients with non-insulin dependent diabetes mellitus. Incubation of red cells from diabetics with 1.5 mM t-butyl hydroperoxide resulted in a lipid peroxidation increase greater than that of normal controls. Glutathione-dependent and glutathione-independent protection against oxidative damage was examined using an artificial system, in which erythrocyte ghosts were incubated with t-butyl peroxide and dialysed hemolysate in the presence or the absence of 2 mM glutathione. The glutathione-dependent protection of hemolysate from diabetics was approximately 70% of that from normal controls.The results suggest that decrease in glutathione-dependent protection against lipid peroxidation, along with decrease in glutathione levels, increases oxidative damage in erythrocyte membranes taken from diabetic patients.Abbreviations GSH reduced form of glutathione - GSSG glutathione disulfide - TCA trichloracetic acid - SOD superoxide dismutase - GPX glutathione peroxidase - MDA malondialdehyde     

蜂胶醇提物通过抑制caspase-12减轻氧化低密度脂蛋白诱导的巨噬细胞凋亡

 目的: 研究蜂胶醇取物(ethanol extract of propolis,EEP)对氧化低密度脂蛋白(oxidized low-density lipoprotein,ox-LDL)诱导的巨噬细胞凋亡的抑制作用,并探讨可能的分子机制。 方法: 体外培养RAW264.7巨噬细胞,给予EEP(7.5、15和30 mg/L)、4-苯丁酸(4-phenylbutyric acid,PBA;5 mmol/L)或二亚苯基碘鎓(diphenyleneiodo-nium, DPI;5 μmol/L)预处理1 h,再加入ox-LDL(100 mg/L)或衣霉素(tunicamycin,TM;4 mg/L)继续培养24 h。分别采用MTT法和Annexin V-FITC双染法检测细胞活力和凋亡情况;试剂盒测定细胞内超氧化物歧化酶(superoxide dismutase,SOD)活性、活性氧簇(reactive oxygen species,ROS)和丙二醛(malondialdehyde,MDA)的水平。采用免疫印迹技术检测内质网应激(endoplasmic reticulum stress,ERS)凋亡途径关键蛋白caspase-12的表达变化。结果: 与ERS抑制剂PBA相似,EEP呈剂量依赖性减轻ox-LDL所致的巨噬细胞损伤,表现为细胞活力增加(P<0.01),凋亡率降低(P<0.05),且可抑制ERS诱导剂TM所引起的巨噬细胞活力下降和凋亡(P<0.05);与氧化应激抑制剂DPI相似,EEP可抑制ox-LDL诱导的氧化应激反应,表现为ROS和MDA生成减少(P<0.01),SOD活性增加(P<0.05);EEP显著抑制ox-LDL和TM所诱导的caspase-12活化(P<0.05);与ox-LDL组比较,PBA和DPI预处理组caspase-12活性也受到明显抑制(P<0.01)。结论: EEP可减轻 ox-LDL 所诱导的RAW264.7巨噬细胞凋亡,其机制可能与抑制氧化应激和caspase-12活化有关。     

Beneficial effects of chlorogenic acid on methotrexate-induced cerebellar Purkinje cell damage in rats

Several studies have well confirmed the contribution of oxidative stress in the pathogenesis of methotrexate (MTX)-induced damage in the various organs. Many agents have been tested experimentally to reduce or inhibit the oxidative stress. The aim of this study was to determine the possible protective effect of chlorogenic acid (CLG) on MTX-induced cerebellar damage in rats. The rats were randomly divided into three groups as follows: I: control group; II: MTX group; III: CLG+MTX group. In the MTX group; malondialdehyde (MDA) content was found to be increased, whereas superoxide dismutase (SOD), catalase (CAT) activities, and glutathione (GSH) content were decreased. On the other hand, CLG markedly attenuated the elevated MDA content and prevented the deleterious effects of MTX on oxidative stress markers. MTX caused severe loss of Purkinje cells and apoptotic cell death in the cerebellum. The CLG administration before MTX treatment significantly reduced Purkinje cell damage and the expression of apoptotic cells. In conclusion, our results demonstrate that chlorogenic acid treatment may protect the impairment of oxidative stress and ameliorate MTX-induced cerebellar damage at biochemical and histological levels.     

SSAO介导的氧化脱氨反应对3T3-L1脂肪细胞的氧化应激作用

目的:观察氨基脲敏感型胺氧化酶(SSAO)催化其底物甲胺(MA)和苯甲胺(BZA)的氧化脱氨反应对成熟3T3-L1脂肪细胞的氧化应激作用。方法:3T3-L1前脂肪细胞诱导分化为成熟脂肪细胞;高效液相色谱法检测不同分化天数下细胞SSAO活性的变化;MTT法检测不同浓度的MA或BZA对细胞活力的影响;荧光探针方法检测细胞在药物作用下产生的活性氧;以0.5 mmol/L MA或BZA处理成熟脂肪细胞或未经诱导分化的前脂肪细胞4 h,观察细胞产生的甲醛(FA)、苯甲醛(BZ)、脂质过氧化指标丙二醛(MDA)、总超氧化物歧化酶(T-SOD)和谷胱甘肽(GSH)的变化。结果:SSAO活性随细胞分化天数的增加而增加,并于第8天达到高峰;不同浓度MA或BZA作用细胞4 h对细胞活力无显著影响(P0.05);0.5 mmol/L MA或BZA孵育后,细胞产生的活性氧增高,约为阴性对照组的3~4倍,差异有统计学意义(P0.05);在成熟脂肪细胞中,MDA的含量增加,而T-SOD和GSH的活性和含量减少,与阴性对照组相比,差别有统计学意义(P0.05);MA和BZA作用未经诱导分化的前脂肪细胞,其MDA和SOD和GSH的变化不明显,差别无统计学意义(P0.05)。结论:SSAO可能通过其介导的氧化脱氨反应引起3T3-L1成熟脂肪细胞产生氧化应激。