Oxidative Stress Alters Physiological and Morphological Neuronal Properties

We investigated the effects of H₂O₂-induced oxidative stress on the delayed-rectifier current (IK_DR), neuronal physiological and morphological properties. Measurements were obtained from hippocampal CA1 neurons in control solution and from the same neurons after exposure to oxidative stress (short- and long-term H₂O₂ external applications at 0.1, 1, and 10 mM). With short-term (6 min) H₂O₂ (1 mM) treatment, IK_DR measured in the H₂O₂-containing solution (778 ± 23 pA, n = 20), was smaller than that measured in the control Ca²⁺-free Hepes solution (1,112 ± 38 pA, n = 20). Coenzyme Q₁₀ (0.1 mM) pretreatment prevented the H₂O₂-induced inhibition of IK_DR. With long-term (40, 80 min) H₂O₂ (0.1, 10 mM) treatment, the neuron lost its distinctive shape (rounded up) and the neurite almost disappeared. These results suggest that oxidative stress, which inhibits IK_DR, can alter neural activity. The morphological changes caused by H₂O₂ support the idea that oxidative stress causes intracellular damage and compromises neural function.     

Oxidative Stress Alters the Morphology and Toxicity of Aortic Medial Amyloid

The aggregation and fibril deposition of amyloid proteins have been implicated in a range of neurodegenerative and vascular diseases, and yet the underlying molecular mechanisms are poorly understood. Here, we use a combination of cell-based assays, biophysical analysis, and atomic force microscopy to investigate the potential involvement of oxidative stress in aortic medial amyloid (AMA) pathogenesis and deposition. We show that medin, the main constituent of AMA, can induce an environment rich in oxidative species, increasing superoxide and reducing bioavailable nitric oxide in human cells. We investigate the role that this oxidative environment may play in altering the aggregation process of medin and identify potential posttranslational modification sites where site-specific modification and interaction can be unambiguously demonstrated. In an oxidizing environment, medin is nitrated at tyrosine and tryptophan residues, with resultant effects on morphology that lead to longer fibrils with increased toxicity. This provides further motivation to investigate the role of oxidative stress in AMA pathogenicity.     

CpG-Oligodeoxynucleotide Treatment Protects against Ionizing Radiation-Induced Intestine Injury

Background

the bone marrow and the intestine are the major sites of ionizing radiation (IR)-induced injury. Our previous study demonstrated that CpG-oligodeoxynucleotide (ODN) treatment mitigated IR-induced bone marrow injury, but its effect on the intestine is not known. In this study, we sought to determine if CpG-ODN have protective effect on IR-induced intestine injury, and if so, to determine the mechanism of its effect.

Methods and Findings

Mice were treated with CpG-ODN after IR. The body weight and survival were daily monitored for 30 days consecutively after exposure. The number of surviving intestinal crypt was assessed by the microcolony survival assay. The number and the distribution of proliferating cell in crypt were evaluated by TUNEL assay and BrdU assay. The expression of Bcl-2, Bax and caspase-3 in crypt were analyzed by Immunohistochemistry assay. The findings showed that the treatment for irradiated mice with CpG-ODN diminished body weight loss, improved 30 days survival, enhanced intestinal crypts survival and maintained proliferating cell population and regeneration in crypt. The reason might involve that CpG-ODN up-regulated the expression of Bcl-2 protein and down-regulated the expression of Bax protein and caspase-3 protein.

Conclusion

CpG-ODN was effective in protection of IR-induced intestine injury by enhancing intestinal crypts survival and maintaining proliferating cell population and regeneration in crypt. The mechanism might be that CpG-ODN inhibits proliferating cell apoptosis through regulating the expression of apoptosis-related protein, such as Bax, Bcl-2 and caspase-3.     

Blood Glutathione as an Index of Radiation-Induced Oxidative Stress in Mice and Humans

The effect of x-rays on GSH and GSSG levels in blood was studied in mice and humans. An HPLC method that we recently developed was applied to accurately determine GSSG levels in blood. The glutathione redox status (GSH/GSSG) decreases after irradiation. This effect is mainly due to an increase in GSSG levels. Mice received single fraction radiotherapy, at total doses of 1.0 to 7.0 Gy. Changes in GSSG in mouse blood can be detected 10 min after irradiation and last for 6 h within a range of 2.0–7.0 Gy. The highest levels of GSSG (20.1 ± 2.9 ), a 4.7-fold increase as compared with controls) in mouse blood are found 2 h after radiation exposure (5 Gy). Breast and lung cancer patients received fractionated radiotherapy at total doses of 50.0 or 60.0 Gy, respectively. GSH/GSSG also decreases in humans in a dose–response fashion. Two reasons may explain the radiation-induced increase in blood GSSG: (a) the reaction of GSH with radiation-induced free radicals resulting in the formation of thyl radicals that react to produce GSSG; and (b) an increase of GSSG release from different organs (e.g., the liver) into the blood. Our results indicate that the glutathione redox ratio in blood can be used as an index of radiation-induced oxidative stress. © 1997 Elsevier Science Inc.     

Elevated Expression of AKR1C3 Increases Resistance of Cancer Cells to Ionizing Radiation via Modulation of Oxidative Stress

With the aim to elucidate the etiology of radioresistance, we explored the genetic alterations in non-radioresistant vs. resistant esophageal cancer cells acquired by long-term fractionated radiation. We found AKR1C3, an aldo-keto reductase expressed seldom in most human tissues, expressed higher in radioresistance-acquired cells. Suppression of AKR1C3 via RNAi or its chemical inhibitors restored the sensitivity of the acquired tumor cells and xenograft BALB/c nude mice to ionizing radiation (IR). Cellular monitoring of the oxidative stress in the AKR1C3-elevated cells indicated that IR-induced ROS accumulation and the concomitant DNA damage was significantly alleviated, and such protective consequence disappeared upon AKR1C3 knockdown. These findings uncover the potential involvement of AKR1C3 in removal of cellular ROS and explain, at least partially, the acquired radioresistance by AKR1C3 overexpression. A retrospective analysis of esophageal carcinomas also indicated a significant expression of AKR1C3 in radio-resistant but not radio-sensitive surgical samples. Our study may provide a potential biomarker for predicting prognosis of radiotherapy and even direct a targeted therapy for esophageal cancer and other tumors.     

Oxidative Stress Alters Syndecan-1 Distribution in Lungs with Pulmonary Fibrosis

Idiopathic pulmonary fibrosis (IPF) is an interstitial lung disease characterized by severe, progressive fibrosis. Roles for inflammation and oxidative stress have recently been demonstrated, but despite advances in understanding the pathogenesis, there are still no effective therapies for IPF. This study investigates how extracellular superoxide dismutase (EC-SOD), a syndecan-binding antioxidant enzyme, inhibits inflammation and lung fibrosis. We hypothesize that EC-SOD protects the lung from oxidant damage by preventing syndecan fragmentation/shedding. Wild-type or EC-SOD-null mice were exposed to an intratracheal instillation of asbestos or bleomycin. Western blot was used to detect syndecans in the bronchoalveolar lavage fluid and lung. Human lung samples (normal and IPF) were also analyzed. Immunohistochemistry for syndecan-1 and EC-SOD was performed on human and mouse lungs. In vitro, alveolar epithelial cells were exposed to oxidative stress and EC-SOD. Cell supernatants were analyzed for shed syndecan-1 by Western blot. Syndecan-1 ectodomain was assessed in wound healing and neutrophil chemotaxis. Increases in human syndecan-1 are detected in lung homogenates and lavage fluid of IPF lungs. Syndecan-1 is also significantly elevated in the lavage fluid of EC-SOD-null mice after asbestos and bleomycin exposure. On IHC, syndecan-1 staining increases within fibrotic areas of human and mouse lungs. In vitro, EC-SOD inhibits oxidant-induced loss of syndecan-1 from A549 cells. Shed and exogenous syndecan-1 ectodomain induce neutrophil chemotaxis, inhibit alveolar epithelial wound healing, and promote fibrogenesis. Oxidative shedding of syndecan-1 is an underlying cause of neutrophil chemotaxis and aberrant wound healing that may contribute to pulmonary fibrosis.Idiopathic pulmonary fibrosis (IPF)² is an interstitial lung disease characterized by severe and progressive fibrosis. IPF patients have a mean survival of 3–5 years (1, 2) and no effective therapies (3, 4), other than orthotopic lung transplantation, have proven to improve survival. The pathogenesis of IPF is poorly understood; however, inflammation and oxidant/antioxidant imbalances in the lung are thought to play important roles (5–7). A better understanding of the molecular mechanisms involved in oxidative injury and fibrosis could lead to the development of novel therapeutic targets.Extracellular superoxide dismutase (EC-SOD) is an antioxidant enzyme bound to heparan sulfate in the lung extracellular matrix (8–10), which can inhibit inflammation (11, 12) and prevent subsequent development of fibrosis (13–16). Despite its beneficial role, the mechanisms through which EC-SOD protects the lung remain unknown.The extracellular matrix (ECM) is essential for tissue homeostasis and changes in the ECM microenvironment can be detrimental to cell function during inflammation and wound healing. Heparan sulfate proteoglycans (HSPG) contain a membrane-bound core protein and extracellular carbohydrate side chains. Syndecans are the most abundant HSPG in humans; there are 4 isoforms with variable cell expression (17, 18). Both syndecan-1 and -4 are expressed in the lung, with epithelial cell and ubiquitous expression, respectively (19). Syndecans are essential for ECM homeostasis by binding cytokines and growth factors, acting as co-receptors and soluble effectors. They also have potential roles in inflammation (18, 20, 21), fibrosis (22, 23), and wound healing (24–26). Syndecans are shed under physiological and pathological conditions but the function of shed syndecans is poorly understood (22). Reactive oxygen species (ROS) are capable of fragmenting HSPG (27) and other ECM components. Notably, EC-SOD has been shown to prevent oxidative damage to many ECM components (23, 28, 29). Within the lung, EC-SOD binds to syndecan-1 on the cell surface via a heparin-binding domain (8, 30). Because of the known functions of syndecans and its close interaction with EC-SOD, syndecan-1 is a key target that may contribute to the anti-inflammatory and anti-fibrotic effects of EC-SOD in the lung and in the pulmonary fibrosis.This study was conducted to determine the role of EC-SOD in protecting the ECM from oxidative stress and to investigate our hypothesis that EC-SOD protects the lung from inflammation and fibrosis by inhibiting oxidant-induced shedding of syndecan-1. Our findings suggest that a loss of EC-SOD in the lung leaves syndecan-1 vulnerable to oxidative stress and that oxidatively shed syndecan-1 ectodomain induces neutrophil chemotaxis, impairs epithelial wound healing, and promotes fibrogenesis. The discovery that oxidative stress alters the distribution of syndecan-1 in the lung microenvironment is a novel finding in the context of pulmonary fibrosis. These findings advance the understanding of the pathogenesis of idiopathic pulmonary fibrosis and provide a potential new therapeutic target for intervention in IPF.     

Human Surfactant Protein D Alters Oxidative Stress and HMGA1 Expression to Induce p53 Apoptotic Pathway in Eosinophil Leukemic Cell Line

Surfactant protein D (SP-D), an innate immune molecule, has an indispensable role in host defense and regulation of inflammation. Immune related functions regulated by SP-D include agglutination of pathogens, phagocytosis, oxidative burst, antigen presentation, T lymphocyte proliferation, cytokine secretion, induction of apoptosis and clearance of apoptotic cells. The present study unravels a novel ability of SP-D to reduce the viability of leukemic cells (eosinophilic leukemic cell line, AML14.3D10; acute myeloid leukemia cell line, THP-1; acute lymphoid leukemia cell lines, Jurkat, Raji; and human breast epithelial cell line, MCF-7), and explains the underlying mechanisms. SP-D and a recombinant fragment of human SP-D (rhSP-D) induced G2/M phase cell cycle arrest, and dose and time-dependent apoptosis in the AML14.3D10 eosinophilic leukemia cell line. Levels of various apoptotic markers viz. activated p53, cleaved caspase-9 and PARP, along with G2/M checkpoints (p21 and Tyr15 phosphorylation of cdc2) showed significant increase in these cells. We further attempted to elucidate the underlying mechanisms of rhSP-D induced apoptosis using proteomic analysis. This approach identified large scale molecular changes initiated by SP-D in a human cell for the first time. Among others, the proteomics analysis highlighted a decreased expression of survival related proteins such as HMGA1, overexpression of proteins to protect the cells from oxidative burst, while a drastic decrease in mitochondrial antioxidant defense system. rhSP-D mediated enhanced oxidative burst in AML14.3D10 cells was confirmed, while antioxidant, N-acetyl-L-cysteine, abrogated the rhSP-D induced apoptosis. The rhSP-D mediated reduced viability was specific to the cancer cell lines and viability of human PBMCs from healthy controls was not affected. The study suggests involvement of SP-D in host’s immunosurveillance and therapeutic potential of rhSP-D in the eosinophilic leukemia and cancers of other origins.     

Excess Dietary Sodium Selenite Alters Apoptotic Population and Oxidative Stress Markers of Spleens in Broilers

Three hundred 1-day-old avian broilers were fed on a basic diet (0.2 mg/kg selenium) or the same diet amended to contain 1, 5, 10, and 15 mg/kg selenium supplied as sodium selenite (n = 60/group). In comparison with those of 0.2 mg/kg selenium group, the percentages of annexin V-positive splenocytes were increased in 5, 10, and 15 mg/kg selenium groups. TUNEL assay revealed that apoptotic cells with brown-stained nuclei distributed within the red pulp and white pulp of the spleens with increased frequency of occurrence in 10 and 15 mg/kg selenium groups in comparison with that of 0.2 mg/kg Se group. Sodium selenite-induced oxidative stress in spleens of chickens was evidenced by decrease in glutathione peroxidase, superoxide dismutase, and catalase activities and increase in malondialdehyde contents. The results indicate that excess dietary selenium in the range of 5–15 mg/kg of feed causes oxidative stress, which may be mainly responsible for the increased apoptosis of splenocytes in chickens.     

Ionizing Radiation Alters the Properties of Sodium Channels in Rat Brain Synaptosomes

The effect of ionizing radiation on neuronal membrane function was assessed by measurement of neurotoxin-stimulated 22Na+ uptake by rat brain synaptosomes. High-energy electrons and gamma photons were equally effective in reducing the maximal uptake of 22Na+ with no significant change in the affinity of veratridine for its binding site in the channel. Ionizing radiation reduced the veratridine-stimulated uptake at the earliest times measured (3 and 5 s), when the rate of uptake was greatest. Batrachotoxin-stimulated 22Na+ uptake was less sensitive to inhibition by radiation. The binding of [3H]saxitoxin to its receptor in the sodium channel was unaffected by exposure to ionizing radiation. The effect of ionizing radiation on the lipid order of rat brain synaptic plasma membranes was measured by the fluorescence polarization of the molecular probes 1,6-diphenyl-1,3,5-hexatriene and 1-[4-(trimethylammonium)phenyl]-6-phenyl-1,3,5-hexatriene. A dose of radiation that reduced the veratridine-stimulated uptake of 22Na+ had no effect on the fluorescence polarization of either probe. These results demonstrate an inhibitory effect of ionizing radiation on the voltage-sensitive sodium channels in rat brain synaptosomes. This effect of radiation is not dependent on changes in the order of membrane lipids.     

Ionizing Radiation-Induced Adaptive Response in Fibroblasts under Both Monolayer and 3-Dimensional Conditions

To observe the adaptive response (AR) induced by ionizing radiation in human fibroblasts under monolayer and 3-dimensional (3-D) condition. Three kinds of fibroblasts were cultured under both monolayer and 3-D condition. Immunofluorescent staining was used to detect the γ-H2AX foci and the morphological texture. Trypan blue staining was used to detect the cell death. Western blot was used to detect the expressions of γ-H2AX, p53 and CDKN1A/p21 (p21). We found that DNA damage increased in a dose-dependent and time-dependent manner after high doses of radiation. When cells were pretreated with a priming low dose of radiation followed by high dose radiation, DNA damage was attenuated under both monolayer and 3-D condition, and the adaptive response (AR) was induced. Additionally, the morphology of cells under monolayer and 3-D conditions were different, and radiation also induced AR according to morphological texture analysis. Priming low dose radiation induced AR both under monolayer and 3-D condition. Interestingly, 3-D microenvironment made cells more sensitive to radiation. The expression of p53 and p21 was changed and indicated that they might participate in the regulation of AR.     

支原体污染增加人肿瘤细胞外泌体生成并改变其小RNA表达模式

目的:研究支原体污染对肿瘤细胞外泌体生成的影响,探讨支原体导致的外泌体量和质的变化。方法:应用荧光显微镜及荧光共振能量转移观察支原体污染导致的胞内膜脂质向胞膜再分布及跨细胞传输变化,应用western blotting检测污染细胞上清中外泌体标记物CD81蛋白含量,应用芯片检测污染细胞上清外泌体中小RNA的表达变化模式。结果:支原体污染导致Rab11和胞膜脂质染料信号强度上升2倍以上,提示胞内膜脂质向胞膜再分布增强;支原体污染导致Di O和Di I双染细胞胞内膜泡的FRET指数上升4倍,提示膜脂质由支原体污染细胞向未污染细胞的跨细胞传输增强;污染细胞的上清中外泌体标志物CD81明显升高;污染导致外泌体内小RNA表达模式改变。结论:体外实验显示支原体污染可造成外泌体质和量的改变。     

Protective Effects of Zinc on 2.45 GHz Electromagnetic Radiation-Induced Oxidative Stress and Apoptosis in HEK293 Cells

Biological Trace Element Research - Several epidemiological studies have shown that exposure to electromagnetic radiation (EMR) can be harmful to human health. The purpose of this study was to...     

Melatonin Alters the miRNA Transcriptome of Inflammasome Activation in Murine Microglial Cells

Neurochemical Research - Systemic inflammation can have devastating effects on the central nervous system via its resident immune cells, the microglia. One of the primary mediators of this...     

氧化应激诱导的细胞衰老过程中细胞生长相关基因的表达

应激诱导的细胞早衰与复制性细胞衰老有相似的细胞表型,但其机制不尽相同.分析二者的衰老相关基因表达特点对了解应激因素诱导细胞衰老的机制有重要意义. 本文对过氧化氢诱导的HeLa细胞早衰过程中的关键衰老相关基因及其转录后调控因子的表达做了分析.结果发现,在复制性衰老过程中明显降低的cyclin A、cyclin B1、c-fos及HuR,在温和过氧化氢诱导的细胞早衰过程中并无明显改变;在氧化应激诱导的细胞早衰过程中,p21与p16表达升高,AUF1则降低,与复制性衰老过程一致;p21 mRNA半衰期在复制性衰老过程中无明显变化,但在氧化应激诱导的细胞早衰过程中则显著延长.上述结果提示,尽管氧化应激诱导的细胞早衰与复制性衰老存在相似基因表达变化,调控机制则不尽相同.     

Nontypeable Haemophilus influenzae Induces Sustained Lung Oxidative Stress and Protease Expression

Nontypeable Haemophilus influenzae (NTHi) is a prevalent bacterium found in a variety of chronic respiratory diseases. The role of this bacterium in the pathogenesis of lung inflammation is not well defined. In this study we examined the effect of NTHi on two important lung inflammatory processes 1), oxidative stress and 2), protease expression. Bronchoalveolar macrophages were obtained from 121 human subjects, blood neutrophils from 15 subjects, and human-lung fibroblast and epithelial cell lines from 16 subjects. Cells were stimulated with NTHi to measure the effect on reactive oxygen species (ROS) production and extracellular trap formation. We also measured the production of the oxidant, 3-nitrotyrosine (3-NT) in the lungs of mice infected with this bacterium. NTHi induced widespread production of 3-NT in mouse lungs. This bacterium induced significantly increased ROS production in human fibroblasts, epithelial cells, macrophages and neutrophils; with the highest levels in the phagocytic cells. In human macrophages NTHi caused a sustained, extracellular production of ROS that increased over time. The production of ROS was associated with the formation of macrophage extracellular trap-like structures which co-expressed the protease metalloproteinase-12. The formation of the macrophage extracellular trap-like structures was markedly inhibited by the addition of DNase. In this study we have demonstrated that NTHi induces lung oxidative stress with macrophage extracellular trap formation and associated protease expression. DNase inhibited the formation of extracellular traps.