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Sample records for neuronal apoptosis metallothionein

  1. Neuronal apoptosis, metallothionein expression and proinflammatory responses during cerebral malaria in mice

    DEFF Research Database (Denmark)

    Wiese, Lothar; Kurtzhals, Jørgen A L; Penkowa, Milena

    2006-01-01

    BACKGROUND: Cerebral malaria (CM) is an acute encephalopathy in humans due to the infection with Plasmodium falciparum. Neuro-cognitive impairment following CM occurs in about 10% of the treated survivors, while the precise pathophysiological mechanism remains unknown. Metallothionein I + II (MT-...

  2. Histological changes, apoptosis and metallothionein levels in Triturus carnifex (Amphibia, Urodela) exposed to environmental cadmium concentrations.

    Science.gov (United States)

    Capaldo, Anna; Gay, Flaminia; Scudiero, Rosaria; Trinchella, Francesca; Caputo, Ivana; Lepretti, Marilena; Marabotti, Anna; Esposito, Carla; Laforgia, Vincenza

    2016-04-01

    The aim of this study was to verify if the freshwater safety values established from the European Community (1998) and the Italian Ministry of Health (2001) for cadmium (44.5nM/L in drinking water and 178nM/L in sewage waters) were safe for amphibians, since at these same concentrations cadmium induced endocrine disruption in the newt Triturus carnifex. Adult male specimens of T. carnifex were exposed daily to cadmium (44.5nM/L and 178nM/L as CdCl2, nominal concentrations), respectively, during 3- and 9-months; at the same time, control newts were exposed to tap water only. The accumulation of cadmium in the skin, liver and kidney, the levels of metallothioneins in the skin and the liver, the expression of metallothionein mRNA in the liver, as well as the presence of histological alterations and of apoptosis in the target organs were evaluated. The 9-months exposure induced cadmium accumulation in all the tissues examined; moreover, histological changes were observed in all the tissues examined, irrespective of the dose or the time of exposure. Apoptosis was only detected in the kidney, whereas metallothioneins and metallothionein mRNA did not increase. This study demonstrates that the existing chronic water quality criterion established for cadmium induces in the newt T. carnifex cadmium accumulation and histological alterations in the target organs examined. Together with our previous results, showing that, at these same concentrations, cadmium induced endocrine disruption, the present results suggest that the existing chronic water quality criterion for cadmium appears to be not protective of amphibians.

  3. Histological changes, apoptosis and metallothionein levels in Triturus carnifex (Amphibia, Urodela) exposed to environmental cadmium concentrations

    Energy Technology Data Exchange (ETDEWEB)

    Capaldo, Anna, E-mail: anna.capaldo@unina.it [Department of Biology, University of Naples Federico II, Naples (Italy); Gay, Flaminia [Department of Chemistry and Biology, University of Salerno, Salerno (Italy); Scudiero, Rosaria; Trinchella, Francesca [Department of Biology, University of Naples Federico II, Naples (Italy); Caputo, Ivana; Lepretti, Marilena; Marabotti, Anna; Esposito, Carla [Department of Chemistry and Biology, University of Salerno, Salerno (Italy); Laforgia, Vincenza [Department of Biology, University of Naples Federico II, Naples (Italy)

    2016-04-15

    Highlights: • Specimens of the newt Triturus carnifex were exposed to environmental Cd doses. • Newts exposed to Cd during 9 months accumulated Cd in their tissues. • Cd induced histological alterations in the skin, liver and kidneys. • Cd induced apoptosis only in the kidneys. • Cd did not increase metallothionein levels in the skin and the liver, nor MTs mRNA. - Abstract: The aim of this study was to verify if the freshwater safety values established from the European Community (1998) and the Italian Ministry of Health (2001) for cadmium (44.5 nM/L in drinking water and 178 nM/L in sewage waters) were safe for amphibians, since at these same concentrations cadmium induced endocrine disruption in the newt Triturus carnifex. Adult male specimens of T. carnifex were exposed daily to cadmium (44.5 nM/L and 178 nM/L as CdCl{sub 2}, nominal concentrations), respectively, during 3- and 9-months; at the same time, control newts were exposed to tap water only. The accumulation of cadmium in the skin, liver and kidney, the levels of metallothioneins in the skin and the liver, the expression of metallothionein mRNA in the liver, as well as the presence of histological alterations and of apoptosis in the target organs were evaluated. The 9-months exposure induced cadmium accumulation in all the tissues examined; moreover, histological changes were observed in all the tissues examined, irrespective of the dose or the time of exposure. Apoptosis was only detected in the kidney, whereas metallothioneins and metallothionein mRNA did not increase. This study demonstrates that the existing chronic water quality criterion established for cadmium induces in the newt T. carnifex cadmium accumulation and histological alterations in the target organs examined. Together with our previous results, showing that, at these same concentrations, cadmium induced endocrine disruption, the present results suggest that the existing chronic water quality criterion for cadmium appears to

  4. The role of metallothionein II in neuronal differentiation and survival

    DEFF Research Database (Denmark)

    Køhler, Lene B; Berezin, Vladimir; Bock, Elisabeth

    2003-01-01

    -I+II can affect neurons directly. It is likely that MT isoforms could be beneficial also during neurodegenerative disorders. In this study, we have examined if MT-II affects survival and neurite extension of dopaminergic and hippocampal neurons. We show for the first time that MT-II treatment can...... significantly stimulate neurite extension from both dopaminergic and hippocampal neurons. Moreover, MT-II treatment significantly increases survival of dopaminergic neurons exposed to 6-hydroxydopamine (6-OHDA) and protects significantly hippocampal neurons from amyloid beta-peptide-induced neurotoxicity...

  5. Metallothionein and a peptide modeled after metallothionein, EmtinB, induce neuronal differentiation and survival through binding to receptors of the low-density lipoprotein receptor family

    DEFF Research Database (Denmark)

    Ambjørn, Malene; Asmussen, Johanne W; Lindstam, Mats;

    2007-01-01

    Accumulating evidence suggests that metallothionein (MT)-I and -II promote neuronal survival and regeneration in vivo. The present study investigated the molecular mechanisms underlying the differentiation and survival-promoting effects of MT and a peptide modeled after MT, EmtinB. Both MT...... and EmtinB directly stimulated neurite outgrowth and promoted survival in vitro using primary cultures of cerebellar granule neurons. In addition, expression and surface localization of megalin, a known MT receptor, and the related lipoprotein receptor-related protein-1 (LRP) are demonstrated in cerebellar...... granule neurons. By means of surface plasmon resonance MT and EmtinB were found to bind to both megalin and LRP. The bindings were abrogated in the presence of receptor-associated protein-1, an antagonist of the low-density lipoprotein receptor family, which also inhibited MT- and EmtinB-induced neurite...

  6. Protective role of metallothionein (Ⅰ/Ⅱ) against pathological damage and apoptosis induced by dimethylarsinic acid

    Institute of Scientific and Technical Information of China (English)

    Guang Jia; Yi-Qun Gu; Kung-Tung Chen; You-Yong Lu; Lei Yan; Jian-Ling Wang; Ya-Ping Su; J. C. Gaston Wu

    2004-01-01

    AIM: To better clarify the main target organs of dimethylarsinic acid toxicity and the role of metallothionein (MTs) in modifying dimethylarsinic acid (DMAA) toxicity.METHODS: MT-Ⅰ/Ⅱ null (MT-/-) mice and the corresponding wild-type mice (MT+/+), six in each group, were exposed to DMAA (0-750 mg/kg body weight) by a single oral injection.Twenty four hours later, the lungs, livers and kidneys were collected and undergone pathological analysis, induction of apoptotic cells as determined by TUNEL and MT concentration was detected by radio-immunoassay.RESULTS: Remarkable pathological lesions were observed at the doses ranging from 350 to 750 mg/kg body weight in the lungs, livers and kidneys and MT+/+ mice exhibited a relatively slight destruction when compared with that in dose matched MT-/- mice. The number of apoptotic cells was increased in a dose dependent manner in the lungs and livers in both types of mice. DMAA produced more necrotic cells rather than apoptotic cells at the highest dose of 750 mg/kg,however, no significant increase was observed in the kidney.Hepatic MT level in MT+/+ mice was significantly increased by DMAA in a dose-dependent manner and there was nodetectable amount of hepatic MT in untreated MT-/- mice.CONCLUSION: DMAA treatment can lead to the induction of apoptosis and pathological damage in both types of mice.MT exhibits a protective effect against DMAA toxicity.

  7. Isoflurane-induced neuronal apoptosis in developing hippocampal neurons

    Institute of Scientific and Technical Information of China (English)

    Hongliang Liu; Tijun Dai; Weitao Guo

    2013-01-01

    We hypothesized that the P2X7 receptor may be the target of isoflurane, so we investigated the roles of the P2X7 receptor and inositol triphosphate receptor in calcium overload and neuronal apoptosis induced by isoflurane in cultured embryonic rat hippocampal neurons. Results showed that isoflurane induced widespread neuronal apoptosis and significantly increased cytoplasmic Ca2+. Blockade of P2X7 receptors or removal of extracellular Ca2+ combined with blockade of inositol triphosphate receptors completely inhibited apoptosis or increase in cytoplasmic Ca2+. Removal of extracellular Ca2+ or blockade of inositol triphosphate receptor alone could partly inhibit these effects of isoflurane. Isoflurane could directly activate P2X7-gated channels and induce inward currents, but did not affect the expression of P2X7 receptor protein in neurons. These findings indicate that the mechanism by which isoflurane induced neuronal apoptosis in rat developing brain was mediated by intracellular calcium overload, which was caused by P2X7 receptor mediated calcium influx and inositol triphosphate receptor mediated calcium release.

  8. Hypoxia drives apoptosis independently of p53 and metallothionein transcript levels in hemocytes of the whiteleg shrimp Litopenaeus vannamei.

    Science.gov (United States)

    Felix-Portillo, Monserrath; Martínez-Quintana, José A; Arenas-Padilla, Marina; Mata-Haro, Verónica; Gómez-Jiménez, Silvia; Yepiz-Plascencia, Gloria

    2016-10-01

    The cellular mechanisms used by the shrimp Litopenaeus vannamei to respond to hypoxia have been studied from the energetic metabolism and antioxidant angles. We herein investigated the participation of p53 and metallothionein (MT) in the apoptotic process in response to hypoxia in shrimp hemocytes. The Lvp53 or LvMT genes were efficiently silenced by injection of double stranded RNA for p53 or MT. The effects of silencing on apoptosis were measured as caspase-3 activity and flow cytometry in hemocytes after 24 and 48 h of hypoxia (1.5 mg DO L(-1)). Hemocytes from unsilenced animals had significantly higher apoptosis levels upon both times of hypoxia. The apoptotic levels were diminished but not suppressed in dsp53-silenced but not dsMT-silenced hemocytes after 24 h of hypoxia, indicating a contribution of Lvp53 to apoptosis. Apoptosis in normoxia was significantly higher in dsp53-and dsMT-silenced animals compared to the unsilenced controls, pointing to a possible cytoprotective role of LvMT and Lvp53 during the basal apoptotic program in normoxia. Overall, these results indicate that hypoxia augments apoptosis in shrimp hemocytes and high mRNA levels of Lvp53 and LvMT are not necessary for this response.

  9. Neuronal apoptosis: signal and cell diversity

    Directory of Open Access Journals (Sweden)

    Lina Vanessa Becerra

    2009-12-01

    Full Text Available Programmed cell death occurs as a physiological process during development. In the brain and spinal cord this event determines the number and location of the different cell types. In adulthood, programmed cell death or apoptosis is more restricted but it may play a major role in different acute and chronic pathological entities. However, in contrast to other tissues where apoptosis has been widely documented from a morphological point of view, in the central nervous system complete anatomical evidence of apoptosis is scanty. In spite of this there is consensus about the activation of different signal systems associated to programmed cell death. In the present article we attempt to summarize the main apoptotic pathways so far identified in nervous tissue. Considering that apoptotic pathways are multiple, the neuronal cell types are highly diverse and specialized and that neuronal response to injury and survival depends upon tissue context, (i.e., preservation of connectivity, glial integrity and cell matrix, blood supply and trophic factors availability what is relevant for the apoptotic process in a sector of the brain may not be important in another.

  10. Nitric oxide damages neuronal mitochondria and induces apoptosis in neurons

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    The cytotoxic effect of nitric oxide on primarily cultured rat cerebellar granule cells was studied,and the mechanisms were discussed.The results showed that nitric oxide donor S-nitroso-N-acetyl-penicillamine (SNAP; 500 μmol/L) could induce apoptosis in immature cultures of cerebellar granule cells.Flow cytometry and HPLC analyses revealed that after treatment with SNAP,the mitochondrial transmembrane potential and the cellular ATP content decreased significantly.Nitric oxide scavenger hemoglobin could effectively prevent the neuronal mitochondria from dysfunction and attenuate apoptosis.The results suggested that nitric oxide activated the apoptotic program by inhibiting the activity of mitochondrial respiratory chain and thus decreasing the cellular ATP content.

  11. A metallothionein mimetic peptide protects neurons against kainic acid-induced excitotoxicity

    DEFF Research Database (Denmark)

    Sonn, Katrin; Pankratova, Stanislava; Korshunova, Irina

    2010-01-01

    Metallothioneins I and II (MTI/II) are metal-binding proteins overexpressed in response to brain injury. Recently, we have designed a peptide, termed EmtinB, which is modeled after the beta-domain of MT-II and mimics the biological effects of MTI/II in vitro. Here, we demonstrate the neuroprotect...

  12. Role of PUMA in methamphetamine-induced neuronal apoptosis.

    Science.gov (United States)

    Chen, Chuanxiang; Qincao, Litao; Xu, Jingtao; Du, Sihao; Huang, Enping; Liu, Chao; Lin, Zhoumeng; Xie, Wei-Bing; Wang, Huijun

    2016-01-05

    Exposure to methamphetamine (METH), a widely used illicit drug, has been shown to cause neuron apoptosis. p53 upregulated modulator of apoptosis (PUMA) is a key mediator in neuronal apoptosis. This study aimed to examine the effects of PUMA in METH-induced neuronal apoptosis. We determined PUMA protein expression in PC12 cells and SH-SY5Y cells after METH exposure using western blot. We also observed the effect of METH on neuronal apoptosis after silencing PUMA expression with siRNA using TUNEL staining and flow cytometry. Additionally, to investigate possible mechanisms of METH-induced PUMA-mediated neuronal apoptosis, we measured the protein expression of apoptotic markers, including cleaved caspase-3, cleaved PARP, Bax, B-cell leukemia/lymphoma-2 (Bcl-2) and cytochrome c (cyto c), after METH treatment with or without PUMA knockdown. Results showed that METH exposure induced cell apoptosis, increased PUMA protein levels, activated caspase-3 and PARP, elevated Bax and reduced Bcl-2 expression, as well as increased the release of cyto c from mitochondria to the cytoplasm in both PC12 and SH-SY5Y cells. All these effects were attenuated or reversed after silencing PUMA. A schematic depicting the role of PUMA in METH-induced mitochondrial apoptotic pathway was proposed. Our results suggest that PUMA plays an important role in METH-triggered apoptosis and it may be a potential target for ameliorating neuronal injury and apoptosis caused by METH.

  13. Redefining the role of metallothionein within the injured brain: extracellular metallothioneins play an important role in the astrocyte-neuron response to injury

    DEFF Research Database (Denmark)

    Chung, Roger S; Penkowa, Milena; Dittmann, Justin;

    2008-01-01

    A number of intracellular proteins that are protective after brain injury are classically thought to exert their effect within the expressing cell. The astrocytic metallothioneins (MT) are one example and are thought to act via intracellular free radical scavenging and heavy metal regulation, and...

  14. Cerebral ischemia—induced neuronal apoptosis mediated by nitric oxide

    Institute of Scientific and Technical Information of China (English)

    NomuY

    2002-01-01

    To elucidate the cellular and molecular mechanism of cerebral ischemia-induced neuronal apoptosis mediated by nitric oxide (NO) in the brain,we investigated:(1)cell death in hippocampal CA1 neurons of rats after a rransient four vessel occlusion (4VO)/reperfusion and (2) apoptosis induced by NOC18(NO releaser) using SHSY5Y cells,a human neuroblastoma cell line.We found that 4VO caused expression of inducible type of NO synthase (iNOS) in glial cells and neuronal apoptosis in CA1 region of rats.Next we examined in vitro apoptotic effects of NOC18 on SHSY5Y cells and suggest that NO decrease mitochondrial membrane potential,release cytochrome C from mitochondria,activates caspase-3,degrade inhibitor of caspase-activated DNase(Icad),and activated DNase translocate into nucleus and induce DNA fragmentation.Thus we conclude that the excess amount of NO produced by glial iNOS at cerebral ischemia could be involved in neuronal apoptosis in CA1 region.Regarding NO action on neurons,we further obtained that NO propects neuronal apoptosis in PC12 cells perhaps by nitrosylation of caspase,subsequent reduction of proteolytic activity.Taken together,we suggest that NO seem to exert dual effects(toxic and beneficial) on neuronal apoptosis,the one (toxic);apoptosis-induction throuth the decrease in mitochondrial membrane potentials and cytochrome C release and the othe (beneficial);protection against apoptosis through the inhibition of caspase activity.

  15. Mature neurons dynamically restrict apoptosis via redundant premitochondrial brakes.

    Science.gov (United States)

    Annis, Ryan P; Swahari, Vijay; Nakamura, Ayumi; Xie, Alison X; Hammond, Scott M; Deshmukh, Mohanish

    2016-12-01

    Apoptotic cell death is critical for the early development of the nervous system, but once the nervous system is established, the apoptotic pathway becomes highly restricted in mature neurons. However, the mechanisms underlying this increased resistance to apoptosis in these mature neurons are not completely understood. We have previously found that members of the miR-29 family of microRNAs (miRNAs) are induced with neuronal maturation and that overexpression of miR-29 was sufficient to restrict apoptosis in neurons. To determine whether endogenous miR-29 alone was responsible for the inhibition of cytochrome c release in mature neurons, we examined the status of the apoptotic pathway in sympathetic neurons deficient for all three miR-29 family members. Unexpectedly, we found that the apoptotic pathway remained largely restricted in miR-29-deficient mature neurons. We therefore probed for additional mechanisms by which mature neurons resist apoptosis. We identify miR-24 as another miRNA that is upregulated in the maturing cerebellum and sympathetic neurons that can act redundantly with miR-29 by targeting a similar repertoire of prodeath BH3-only genes. Overall, our results reveal that mature neurons engage multiple redundant brakes to restrict the apoptotic pathway and ensure their long-term survival. © 2016 Federation of European Biochemical Societies.

  16. Effect of Ca2EDTA on zinc mediated inflammation and neuronal apoptosis in hippocampus of an in vivo mouse model of hypobaric hypoxia.

    Directory of Open Access Journals (Sweden)

    Udayabanu Malairaman

    Full Text Available BACKGROUND: Calcium overload has been implicated as a critical event in glutamate excitotoxicity associated neurodegeneration. Recently, zinc accumulation and its neurotoxic role similar to calcium has been proposed. Earlier, we reported that free chelatable zinc released during hypobaric hypoxia mediates neuronal damage and memory impairment. The molecular mechanism behind hypobaric hypoxia mediated neuronal damage is obscure. The role of free zinc in such neuropathological condition has not been elucidated. In the present study, we investigated the underlying role of free chelatable zinc in hypobaric hypoxia-induced neuronal inflammation and apoptosis resulting in hippocampal damage. METHODS: Adult male Balb/c mice were exposed to hypobaric hypoxia and treated with saline or Ca2EDTA (1.25 mM/kg i.p daily for four days. The effects of Ca2EDTA on apoptosis (caspases activity and DNA fragmentation, pro-inflammatory markers (iNOS, TNF-α and COX-2, NADPH oxidase activity, poly(ADP ribose polymerase (PARP activity and expressions of Bax, Bcl-2, HIF-1α, metallothionein-3, ZnT-1 and ZIP-6 were examined in the hippocampal region of brain. RESULTS: Hypobaric hypoxia resulted in increased expression of metallothionein-3 and zinc transporters (ZnT-1 and ZIP-6. Hypobaric hypoxia elicited an oxidative stress and inflammatory response characterized by elevated NADPH oxidase activity and up-regulation of iNOS, COX-2 and TNF-α. Furthermore, hypobaric hypoxia induced HIF-1α protein expression, PARP activation and apoptosis in the hippocampus. Administration of Ca2EDTA significantly attenuated the hypobaric hypoxia induced oxidative stress, inflammation and apoptosis in the hippocampus. CONCLUSION: We propose that hypobaric hypoxia/reperfusion instigates free chelatable zinc imbalance in brain associated with neuroinflammation and neuronal apoptosis. Therefore, zinc chelating strategies which block zinc mediated neuronal damage linked with cerebral hypoxia

  17. Cold-inducible RNA-binding protein inhibits neuron apoptosis through the suppression of mitochondrial apoptosis.

    Science.gov (United States)

    Zhang, Hai-Tao; Xue, Jing-Hui; Zhang, Zhi-Wen; Kong, Hai-Bo; Liu, Ai-Jun; Li, Shou-Chun; Xu, Dong-Gang

    2015-10-01

    Cold-inducible RNA-binding protein (CIRP) is induced by mild hypothermia in several mammals, but the precise mechanism by which CIRP mediates hypothermia-induced neuroprotection remains unknown. We aimed to investigate the molecular mechanisms by which CIRP protects the nervous system during mild hypothermia. Rat cortical neurons were isolated and cultured in vitro under mild hypothermia (32°C). Apoptosis was measured by annexin V and propidium iodide staining, visualized by flow cytometry. Neuron ultrastructure was visualized by transmission electron microscopy. CIRP overexpression and knockdown were achieved via infection with pL/IRES/GFP-CIRP and pL/shRNA/F-CIRP-A lentivirus. RT(2) Profiler PCR Array Pathway Analysis and western blotting were used to evaluate the effects of CIRP overexpresion/knockdown on the neurons׳ transcriptome. Neuron late apoptosis was significantly reduced at day 7 of culture by 12h hypothermia, but neuron ultrastructure remained relatively intact. RT(2) Profiler PCR Array Pathway Analysis of 84 apoptosis pathway-associated factors revealed that mild hypothermia and CIRP overexpression induce similar gene expression profiles, specifically alterations of genes implicated in the mitochondrial apoptosis pathway. Mild hypothermia-treated neurons up-regulated 12 and down-regulated 38 apoptosis pathway-associated genes. CIRP-overexpressing neurons up-regulated 15 and down-regulated 46 genes. CIRP-knocked-down hypothermia-treated cells up-regulated 9 and down-regulated 40 genes. Similar results were obtained at the protein level. In conclusion, CIRP may inhibit neuron apoptosis through the suppression of the mitochondria apoptosis pathway during mild hypothermia.

  18. Acetaminophen induces apoptosis in rat cortical neurons.

    Directory of Open Access Journals (Sweden)

    Inmaculada Posadas

    Full Text Available BACKGROUND: Acetaminophen (AAP is widely prescribed for treatment of mild pain and fever in western countries. It is generally considered a safe drug and the most frequently reported adverse effect associated with acetaminophen is hepatotoxicity, which generally occurs after acute overdose. During AAP overdose, encephalopathy might develop and contribute to morbidity and mortality. Our hypothesis is that AAP causes direct neuronal toxicity contributing to the general AAP toxicity syndrome. METHODOLOGY/PRINCIPAL FINDINGS: We report that AAP causes direct toxicity on rat cortical neurons both in vitro and in vivo as measured by LDH release. We have found that AAP causes concentration-dependent neuronal death in vitro at concentrations (1 and 2 mM that are reached in human plasma during AAP overdose, and that are also reached in the cerebrospinal fluid of rats for 3 hours following i.p injection of AAP doses (250 and 500 mg/kg that are below those required to induce acute hepatic failure in rats. AAP also increases both neuronal cytochrome P450 isoform CYP2E1 enzymatic activity and protein levels as determined by Western blot, leading to neuronal death through mitochondrial-mediated mechanisms that involve cytochrome c release and caspase 3 activation. In addition, in vivo experiments show that i.p. AAP (250 and 500 mg/kg injection induces neuronal death in the rat cortex as measured by TUNEL, validating the in vitro data. CONCLUSIONS/SIGNIFICANCE: The data presented here establish, for the first time, a direct neurotoxic action by AAP both in vivo and in vitro in rats at doses below those required to produce hepatotoxicity and suggest that this neurotoxicity might be involved in the general toxic syndrome observed during patient APP overdose and, possibly, also when AAP doses in the upper dosing schedule are used, especially if other risk factors (moderate drinking, fasting, nutritional impairment are present.

  19. 5,7-Dimethoxycoumarin inhibits neuronal apoptosis by targeting ...

    African Journals Online (AJOL)

    However, those in the normal control and model groups received the same volume of ... cerebral infarction in animals the rate of neuronal apoptosis is ... Department of Health and Human Services, and guidelines ... The complex development was performed using an ECL .... and reproduction in any medium, provided the.

  20. Sulforaphane Prevents Neuronal Apoptosis and Memory Impairment in Diabetic Rats

    Directory of Open Access Journals (Sweden)

    Gengyin Wang

    2016-08-01

    Full Text Available Background/Aims: To explore the effects of sulforaphane (SFN on neuronal apoptosis in hippocampus and memory impairment in diabetic rats. Methods: Thirty male rats were randomly divided into normal control, diabetic model and SFN treatment groups (N = 10 in each group. Streptozotocin (STZ was applied to establish diabetic model. Water Morris maze task was applied to test learning and memory. Tunel assaying was used to detect apoptosis in hippocampus. The expressions of Caspase-3 and myeloid cell leukemia 1(MCL-1 were detected by western blotting. Neurotrophic factor levels and AKT/GSK3β pathway were also detected. Results: Compared with normal control, learning and memory were apparently impaired, with up-regulation of Caspase-3 and down-regulation of MCL-1 in diabetic rats. Apoptotic neurons were also found in CA1 region after diabetic modeling. By contrast, SFN treatment prevented the memory impairment, decreased the apoptosis of hippocampal neurons. SFN also attenuated the abnormal expression of Caspase-3 and MCL-1 in diabetic model. Mechanically, SFN treatment reversed diabetic modeling-induced decrease of p-Akt, p-GSK3β, NGF and BDNF expressions. Conclusion: SFN could prevent the memory impairment and apoptosis of hippocampal neurons in diabetic rat. The possible mechanism was related to the regulation of neurotropic factors and Akt/GSK3β pathway.

  1. Magnesium induces neuronal apoptosis by suppressing excitability

    OpenAIRE

    Dribben, W H; Eisenman, L N; Mennerick, S

    2010-01-01

    In clinical obstetrics, magnesium sulfate (MgSO4) use is widespread, but effects on brain development are unknown. Many agents that depress neuronal excitability increase developmental neuroapoptosis. In this study, we used dissociated cultures of rodent hippocampus to examine the effects of Mg++ on excitability and survival. Mg++-induced caspase-3-associated cell loss at clinically relevant concentrations. Whole-cell patch-clamp techniques measured Mg++ effects on action potential threshold,...

  2. Effects of calcium channel on 3-morpholinosydnonimine-induced rat hippocampal neuronal apoptosis

    Institute of Scientific and Technical Information of China (English)

    Quanzhong Chang; Shuling Zhang; Yuanyin Zheng; Lijuan Xu; Jinbao Yin; Shining Cai

    2011-01-01

    Previous studies have demonstrated that increased chloride channel activity plays a role in nitric oxide-induced neuronal apoptosis in the rat hippocampus.The present study investigated the effects of the broad-spectrum calcium channel blocker CdC12 on survival rate, percentage of apoptosis, and morphological changes in hippocampal neurons cultured in vitro, as well as the effects of calcium channels on neuronal apoptosis.The chloride channel blockers 4-acetamido-4'-isothiocyanatostilbene-2, 2'-disulfonic acid (SITS) or 4, 4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) increased the survival rate of 3-morpholinosydnonimine (SIN-1)-treated neurons and suppressed SIN-1-induced neuronal apoptosis.The calcium channel blocker CdC12 did not increase the survival rate of neurons and did not affect SIN-1-induced apoptosis or SITS- or DIDS-suppressed neuronal apoptosis.Results demonstrated that calcium channels did not significantly affect neuronal apoptosis.

  3. Mechanisms of Neuronal Apoptosis In Vivo

    Science.gov (United States)

    2004-02-01

    DAMD17-de Tapia , M., Lutz-Bucher, B., Gaiddon, C., and Loeffler, J. P. (2000) Alteration of the Bcl-x/Bax References ratio in a transgenic mouse model of... Ramirez P. 1999. Repair kinetics of gamma- hippocampal neurons in B27-supplemented NeurobasalTM, a new ray induced DNA damage determined by single...Biochemistry 15:4629-4637. 22. Ferrante, R. J., Browne, S. E., Shinobu, L. A., Bowling, A. C., 30. Mendiola-Cruz, M. T. and Morales- Ramirez , P. 1999

  4. Peptides modeled after the alpha-domain of metallothionein induce neurite outgrowth and promote survival of cerebellar granule neurons

    DEFF Research Database (Denmark)

    Asmussen, Johanne Wirenfeldt; Ambjørn, Malene; Bock, Elisabeth;

    2009-01-01

    Metallothionein (MT) is a metal-binding protein capable of preventing oxidative stress and apoptotic cell death in the central nervous system of mammals, and hence is of putative therapeutic value in the treatment of neurodegenerative disorders. Recently, we demonstrated that a peptide modeled...

  5. Metallothionein (MT)-III

    DEFF Research Database (Denmark)

    Carrasco, J; Giralt, M; Molinero, A

    1999-01-01

    Metallothionein-III is a low molecular weight, heavy-metal binding protein expressed mainly in the central nervous system. First identified as a growth inhibitory factor (GIF) of rat cortical neurons in vitro, it has subsequently been shown to be a member of the metallothionein (MT) gene family...... and renamed as MT-III. In this study we have raised polyclonal antibodies in rabbits against recombinant rat MT-III (rMT-III). The sera obtained reacted specifically against recombinant zinc-and cadmium-saturated rMT-III, and did not cross-react with native rat MT-I and MT-II purified from the liver of zinc...... injected rats. The specificity of the antibody was also demonstrated in immunocytochemical studies by the elimination of the immunostaining by preincubation of the antibody with brain (but not liver) extracts, and by the results obtained in MT-III null mice. The antibody was used to characterize...

  6. Thrombin-induced apoptosis in neurons through activation of c-Jun-N-terminal kinase.

    Science.gov (United States)

    Bao, Lei; Zu, Jie; He, Qianqian; Zhao, Hui; Zhou, Su; Ye, Xinchun; Yang, Xinxin; Zan, Kun; Zhang, Zuohui; Shi, Hongjuan; Cui, Guiyun

    2017-01-01

    Studies have shown that thrombin activation played a central role in cell injuries associated with intracerebral hemorrhage (ICH). Here, our study investigated the cytotoxicity of thrombin on neurons, and determined the involvement of JNK pathways in thrombin-induced neuronal apoptosis. Primary cultured neurons were treated with different doses of thrombin. Some neurons were given either SP600125 or vehicle. LDH release assay and flow cytometry were used to measure neuronal apoptosis caused by thrombin. The activation of JNK and capases-3 were measured by Western blot. Our results showed large doses of thrombin that increased the LDH release, the level of cleaved caspase-3 and apoptosis rate of neurons. JNK was activated by thrombin in a time-dependent manner. Administration of SP600125 protects neurons from thrombin-induced apoptosis. These data indicate that the activation of JNK is crucial for thrombin-induced neuronal apoptosis, and inhibition of JNK may be a potential therapeutic target for ICH.

  7. Acute upregulation of neuronal mitochondrial type-1 cannabinoid receptor and it's role in metabolic defects and neuronal apoptosis after TBI.

    Science.gov (United States)

    Xu, Zhen; Lv, Xiao-Ai; Dai, Qun; Ge, Yu-Qing; Xu, Jie

    2016-08-02

    Metabolic defects and neuronal apoptosis initiated by traumatic brain injury (TBI) contribute to subsequent neurodegeneration. They are all regulated by mechanisms centered around mitochondrion. Type-1 cannabinoid receptor (CB1) is a G-protein coupled receptor (GPCR) enriched on neuronal plasma membrane. Recent evidences point to the substantial presence of CB1 receptors on neuronal mitochondrial outer membranes (mtCB1) and the activation of mtCB1 influences aerobic respiration via inhibiting mitochondrial cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA)/complex I pathway. The expression and role of neuronal mtCB1 under TBI are unknown. Using TBI models of cultured neurons, wild type and CB1 knockout mice, we found mtCB1 quickly upregulated after TBI. Activation of mtCB1 promoted metabolic defects accompanied with ATP shortage but protected neurons from apoptosis. Selective activation of plasma membrane CB1 showed no effects on neuronal metabolism and apoptosis. Activation of mtCB1 receptors inhibited mitochondrial cAMP/PKA/complex I and resulted in exacerbated metabolic defects accompanied with a higher ratio of ATP reduction to oxygen consumption decrease as well as neuronal apoptosis. Further research found the remarkable accumulation of protein kinase B (AKT) on neuronal mitochondria following TBI and the activation of mtCB1 upregulated mitochondrial AKT/complex V activity. Upregulation of mitochondrial AKT/complex V activity showed anti-apoptosis effects and alleviated ATP shortage in metabolic defects. Taken together, we have identified mtCB1 quickly upregulate after TBI and a dual role the mtCB1 might play in metabolic defects and neuronal apoptosis initiated by TBI: the inhibition of mitochondrial cAMP/PKA/complex I aggravates metabolic defects, energy insufficiency as well as neuronal apoptosis, but the coactivation of mitochondrial AKT/complex V mitigates energy insufficiency and neuronal apoptosis.

  8. Amyloid β-42 induces neuronal apoptosis by targeting mitochondria.

    Science.gov (United States)

    Han, Xiao-Jian; Hu, Yang-Yang; Yang, Zhang-Jian; Jiang, Li-Ping; Shi, Sheng-Lan; Li, Ye-Ru; Guo, Miao-Yu; Wu, Hong-Li; Wan, Yu-Ying

    2017-10-01

    Alzheimer's disease (AD), with a typical pathological hallmark of amyloid‑beta (Aβ)‑containing plaques and neurofibrillary tangles, is one of the most common types of chronic neurodegenerative diseases. Aβ oligomers serve a crucial role in the pathogenesis of AD, and lead to neuronal loss. However, the precise mechanism of Aβ oligomers in AD remains to be elucidated. The present study demonstrated that 10 µM Aβ‑42 activated the caspase signaling pathway, and induced significant apoptosis in primary cultured mouse cerebral cortical neurons. The results of reverse transcription‑quantitative polymerase chain reaction and western blotting demonstrated that Aβ‑42 (10 µM) also significantly upregulated the transcription and expression of the mitochondrial fission protein dynamin‑related protein 1 (Drp1), and downregulated the transcription and expression of mitochondrial fusion proteins, including mitofusin 1/2 (Mfn1/2) and mitochondrial dynamin like GTPase (OPA‑1). Neurons were transfected with pDsRed2‑Mito for mitochondrial imaging, which revealed that 10 µM Aβ‑42 induced mitochondrial fission in cortical neurons. In addition, 2',7'‑dichlorodihydrofluorescein diacetate and tetramethylrhodamine ethyl ester staining indicated that Aβ‑42 increased the reactive oxygen species (ROS) level and reduced mitochondrial membrane potential in neurons. Inhibition of Drp1 activity by Mdivi‑1 efficiently prevented Aβ‑42‑induced ROS production and disruption of mitochondrial membrane potential. Loss of mitochondrial membrane potential may activate PTEN‑induced putative kinase 1 (Pink1), the prominent sensor for mitochondrial damage, and trigger the process of mitophagy to remove the damaged mitochondria. In the present study, western blotting revealed that the levels of autophagy marker microtubule‑associated proteins 1A/1B light chain 3B (LC3B) and Pink1 were upregulated after Aβ‑42 stimulation. In conclusion, these data indicated that

  9. Apoptosis of lumbar spinal cord neurons in cauda equina syndrome rats

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    Objective To explore the law of apoptosis of lumbar spinal cord neurons in cauda equina syndrome (CES). Methods Cauda equina of rats was compressed by a piece of silica gel stick. From day 1 to day 28,the lumbar spinal cord specimens were harvested and assessed by Nissl's staining and TUNEL staining. Results Compression of cauda equina caused lesion and apoptosis of neurons in lumbar spinal cord,and the extent of apoptosis reached the peak on 7th day after compression. Conclusion Apoptosis of neurons in lum...

  10. Aβ induces PUMA activation: a new mechanism for Aβ-mediated neuronal apoptosis.

    Science.gov (United States)

    Feng, Jie; Meng, Chengbo; Xing, Da

    2015-02-01

    p53 upregulated modulator of apoptosis (PUMA) is a promising tumor therapy target because it elicits apoptosis and profound sensitivity to radiation and chemotherapy. However, inhibition of PUMA may be beneficial for curbing excessive apoptosis associated with neurodegenerative disorders. Alzheimer's disease (AD) is a representative neurodegenerative disease in which amyloid-β (Aβ) deposition causes neurotoxicity. The regulation of PUMA during Aβ-induced neuronal apoptosis remains poorly understood. Here, we reported that PUMA expression was significantly increased in the hippocampus of transgenic mice models of AD and hippocampal neurons in response to Aβ. PUMA knockdown protected the neurons against Aβ-induced apoptosis. Furthermore, besides p53, PUMA transactivation was also regulated by forkhead box O3a through p53-independent manner following Aβ treatment. Notably, PUMA contributed to neuronal apoptosis through competitive binding of apoptosis repressor with caspase recruitment domain to activate caspase-8 that cleaved Bid into tBid to accelerate Bax mitochondrial translocation, revealing a novel pathway of Bax activation by PUMA to mediate Aβ-induced neuronal apoptosis. Together, we demonstrated that PUMA activation involved in Aβ-induced apoptosis, representing a drug target to antagonize AD progression.

  11. Beclin-1-mediated autophagy protects spinal cord neurons against mechanical injury-induced apoptosis.

    Science.gov (United States)

    Wang, Zhen-Yu; Lin, Jian-Hua; Muharram, Akram; Liu, Wen-Ge

    2014-06-01

    Apoptosis has been widely reported to be involved in the pathogenesis associated with spinal cord injury (SCI). Recently, autophagy has also been implicated in various neuronal damage models. However, the role of autophagy in SCI is still controversial and its interrelationship with apoptosis remains unclear. Here, we used an in vitro SCI model to observe a time-dependent induction of autophagy and apoptosis. Mechanical injury induced autophagy markers such as LC3 lipidation, LC3II/LC3I conversion, and Beclin-1 expression. Injured neurons showed decreased cell viability and increased apoptosis. To elucidate the effect of autophagy on apoptosis, the mechanically-injured neurons were treated with the mTOR inhibitor rapamycin and 3-methyl adenine (3-MA), which are known to regulate autophagy positively and negatively, respectively. Rapamycin-treated neurons showed the highest level of cell viability and lowest level of apoptosis among the injured neurons and those treated with 3-MA showed the reciprocal effect. Notably, rapamycin-treated neurons exhibited slightly reduced Bax expression and significantly increased Bcl-2 expression. Furthermore, by plasmid transfection, we showed that Beclin-1-overexpressing neuronal cells responded to mechanical injury with greater LC3II/LC3I conversion and cell viability, lower levels of apoptosis, higher Bcl-2 expression, and unaltered Bax expression as compared to vector control cells. Beclin-1-knockdown neurons showed almost the opposite effects. Taken together, our results suggest that autophagy may serve as a protection against apoptosis in mechanically-injured spinal cord neurons. Targeting mTOR and/or enhancing Beclin-1 expression might be alternative therapeutic strategies for SCI.

  12. Apoptosis during β-mercaptoethanol-induced differentiation of adult adipose-derived stromal cells into neurons

    Institute of Scientific and Technical Information of China (English)

    Yanan Cai; Xiaodong Yuan; Ya Ou; Yanhui Lu

    2011-01-01

    β-mercaptoethanol can induce adipose-derived stromal cells to rapidly and efficiently differentiate into neurons in vitro. However, because of the short survival time of the differentiated cells, clinical applications for this technique are limited. As such, we examined apoptosis of neurons differentiated from adipose-derived stromal cells induced with β-mercaptoethanol in vitro using terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling and transmission electron microscopy. The results revealed that the number of surviving cells decreased and apoptosis rate increased as induction time extended. Taken together, these results suggest that apoptosis occurring in the process of adipose-derived stromal cells differentiating into neurons is the main cause of cell death. However, the mechanism underlying cellular apoptosis should be researched further to develop methods of controlling apoptosis for clinical applications.

  13. Recombinant AAV-mediated Expression of Human BDNF Protects Neurons against Cell Apoptosis in Aβ-induced Neuronal Damage Model

    Institute of Scientific and Technical Information of China (English)

    LIU Zhaohui; MA Dongliang; FENG Gaifeng; MA Yanbing; HU Haitao

    2007-01-01

    The human brain-derived neurotrophic factor (hBDNF) gene was cloned by polymerase chain reaction and the recombinant adeno-associated viral vector inserted with hBDNF gene (AAV-hBDNF) was constructed. Cultured rat hippocampal neurons were treated with Aβ25-35 and serued as the experimental Aβ-induced neuronal damage model (AD model), and the AD model was infected with AAV-hBDNF to explore neuroprotective effects of expression of BDNF. Cell viability was assayed by MTT. The expression of bcl-2 anti-apoptosis protein was detected by immunocytochemical staining. The change of intracellular free Ca ion ([Ca2+]i) was measured by laser scanning confocal microscopy. The results showed that BDNF had protective effects against Aβ-induced neuronal damage. The expression of the bcl-2 anti-apoptosis protein was raised significantly and the balance of [Ca2+]i was maintained in the AAV-hBDNF treatment group as compared with AD model group. These data suggested that recombinant AAV mediated a stable expression of hBDNF in cultured hippocampal neurons and resulted in significant neuron protective effects in AD model. The BDNF may reduce neuron apoptosis through increasing the expression of the bcl-2 anti-apoptosis protein and inhibiting intracellular calcium overload. The viral vector-mediated gene expression of BDNF may pave the way of a novel therapeutic strategy for the treatment of neurodegenerative diseases such as Alzheimer's disease.

  14. Caspase-Mediated Apoptosis in Sensory Neurons of Cultured Dorsal Root Ganglia in Adult Mouse

    Directory of Open Access Journals (Sweden)

    Hamid Reza Momeni

    2013-01-01

    Full Text Available Objective: Sensory neurons in dorsal root ganglia (DRG undergo apoptosis after peripheral nerve injury. The aim of this study was to investigate sensory neuron death and the mechanism involved in the death of these neurons in cultured DRG.Materials and Methods: In this experimental study, L5 DRG from adult mouse were dissected and incubated in culture medium for 24, 48, 72 and 96 hours. Freshly dissected and cultured DRG were then fixed and sectioned using a cryostat. Morphological and biochemical features of apoptosis were investigated using fluorescent staining (Propidium iodide and Hoechst 33342 and the terminal Deoxynucleotide transferase dUTP nick end labeling (TUNEL method respectively. To study the role of caspases, general caspase inhibitor (Z-VAD.fmk, 100 μM and immunohistochemistry for activated caspase-3 were used.Results: After 24, 48, 72 and 96 hours in culture, sensory neurons not only displayed morphological features of apoptosis but also they appeared TUNEL positive. The application of Z-VAD.fmk inhibited apoptosis in these neurons over the same time period. In addition, intense activated caspase-3 immunoreactivity was found both in the cytoplasm and the nuclei of these neurons after 24 and 48 hours.Conclusion: Results of the present study show caspase-dependent apoptosis in the sensory neurons of cultured DRG from adult mouse.

  15. Inhibitory effect of acupuncture on neuronal apoptosis in rats after cerebral ischemia

    Institute of Scientific and Technical Information of China (English)

    Bangyu Ju; Jing Zhang; Guohua Jiang

    2007-01-01

    BACKGROUND: Delayed neuronal death after total cerebral ischemia may accompany with apoptosis, but acupuncture may play a certain role in protecting nerve through inhibiting ischemic neuronal apoptosis.OBJECTIVE: To observe the effect of acupuncture on neuronal apoptosis in rats after cerebral ischemia and analyze its cerebral protective mechanism.DESIGN: Contrast observation among groups.SETTING: Heilongjiang University of Traditional Chinese Medicine.MATERIALS: A total of 30 male healthy Wistar rats of general grade and weighing (250±20) g were randomly divided into three groups, including sham operation group, cerebral ischemia group and acupuncture group with 10 rats in each group. Apoptosis in situ kit was provided by Baolingman Company,Germany.METHODS: The experiment was carried out in the Laboratory Center, Heilongjiang University of Traditional Chinese Medicine from May to November 2004. ① Rats in the cerebral ischemia group and the acupuncture group were used to establish total cerebral ischemic models with four vessels occlusion; in addition, models in the sham operation group were established with the same method as mentioned above.However, four vessels of rats in the sham operation were exposured and cerebral ischemia did not occur. Rats in the acupuncture group were given acupuncture treatment after operation. Needle of 40 mm in length was used to acupuncture bilateral Zusanli (St 36) and Quchi (LI 11) with the depth of 3 mm, and then bilateral acupoints were connected with KWD-808Ⅱ omnipotenc impulse electro-therapeutic apparatus (frequency: 1 Hz;thin waves; voltage: 2 V) once a day for totally 30 minutes. Meanwhile, needle of 25 mm in length was used to acupuncture Baihui (Du 20) with the depth of 3 mm, and then the needle was twirled once every 5 minutes for 30 minutes in total. The course was 7 days. ② Neuronal injuries in hippocampal CA1 area after cerebral ischemia were observed with Nissl body staining method at 7 days after treatment

  16. Changes in neuronal apoptosis and apoptosis modulatory factors in cerebral ischemia/reperfusion

    Institute of Scientific and Technical Information of China (English)

    Liying Qiu; Ying Li; Hongbin Fan; Bin Du; Zhiyong Yang; Jianqing Cheng

    2007-01-01

    BACKGROUND: The high concentration of glutamate release is the main cause for neuronal cell death. The relationship between glutamate level and apoptosis during ischemia/reperfusion injury is still unclear. OBJECTIVE: To observe the neuronal apoptosis at 24 and 72 hours following cerebral ischemia/reperfusion in rats, and analyze the possible influencing factors. DESIGN: A randomized controlled animal experiment. SETTING: School of Medicine, Southern Yangtze University.MATERIALS: Totally 30 male adult Sprague Dawley (SD) rats of clean grade, weighing 240 - 290 g, were obtained from Shanghai Experimental Animal Center, Chinese Academy of Sciences. The rats were randomly divided into sham-operated group (n=10) and model group (n=20). Each group was observed at 24 and 72 hours after ischemia/reperfusion, 5 rats at each time point in the sham-operated group, whereas 12 at 24 hours and 8 at 72 hours in the model group. Kits for determining apoptosis and Bcl-2 were bought from Wuhan Boster Biological Technology, Co., Ltd.; Kit for calcineurin from Nanjing Jiancheng Bioengineering Institute.METHODS: The experiment was carried out in the Functional Scientific Research Room of Southern Yangtze University from June to October in 2006.①Right middle cerebral artery was occluded by inserting a thread through internal carotid artery (ICA). The surgical process for the sham-operated rats was the same as that in the model group except a nylon suture inserted the ICA. According to Longa five-degree standard, the neurological deficit evaluation of rats was evaluated after surgery, and grades 1-3 were taken as successful model establishment. The blood was recirculated by withdrawing the nylon filament under anesthesia at 2 hours after ischemia in successful rat models.②After reperfusion, the brain tissue was quickly removed at 24 or 72 hours and the slices were obtained from optic chiasma to funnel manubrium. The changes of the number of apoptotic cells were observed using the

  17. Cesium chloride protects cerebellar granule neurons from apoptosis induced by low potassium.

    Science.gov (United States)

    Zhong, Jin; Yao, Weiguo; Lee, Weihua

    2007-10-01

    Neuronal apoptosis plays a critical role in the pathogenesis of neurodegenerative disorders, and neuroprotective agents targeting apoptotic signaling could have therapeutic use. Here we report that cesium chloride, an alternative medicine in treating radiological poison and cancer, has neuroprotective actions. Serum and potassium deprivation induced cerebellar granule neurons to undergo apoptosis, which correlated with the activation of caspase-3. Cesium prevented both the activation of caspase-3 and neuronal apoptosis in a dose-dependent manner. Cesium at 8 mM increased the survival of neurons from 45 +/- 3% to 91 +/- 5% of control. Cesium's neuroprotection was not mediated by PI3/Akt or MAPK signaling pathways, since it was unable to activate either Akt or MAPK by phosphorylation. In addition, specific inhibitors of PI3 kinase and MAP kinase did not block cesium's neuroprotective effects. On the other hand, cesium inactivated GSK3beta by phosphorylation of serine-9 and GSK3beta-specific inhibitor SB415286 prevented neuronal apoptosis. These data indicate that cesium's neuroprotection is likely via inactivating GSK3beta. Furthermore, cesium also prevented H(2)O(2)-induced neuronal death (increased the survival of neurons from 72 +/- 4% to 89 +/- 3% of control). Given its relative safety and good penetration of the brain blood barrier, our findings support the potential therapeutic use of cesium in neurodegenerative diseases.

  18. Effect of Batroxobin on Neuronal Apoptosis During Focal Cerebral Ischemia and Reperfusion in Rats

    Institute of Scientific and Technical Information of China (English)

    吴卫平; 匡培根; 李振洲

    2001-01-01

    We have found that Batroxobin plays a protactive role in ischemic brain injury, which attracted us to investigate the effect of Batroxobin on apoptosis of neurons during cerebral ischemia and reperfusion. The apoptotic cells in ischemic rat brains at different reperfusion intervals were tested with method of TdT-mediated dUTP-DIG nick end labeling (TUNEL) and the effect of Batroxobin on the apoptosis of neurons was studied in left middle cerebral artery (LMCA) occlusion and reperfusion in rat models (n=18). The results showed that few scattered apoptosis cells were observed in right cerebral hemispheres after LMCA occlusion and reperfusion, and that a lot of apoptosis cells were found in left ischemic cortex and caudoputamen at 12h reperfusion, and they reached peak at 24h~48h reperfusion. However, in the rats pretreated with Batroxobin, the number of apoptosis cells in left cerebral cortex and caudoputamen reduced significantly and the neuronal damage was much milder at 24h reperfusion than that of saline-treated rats. The results indicate that administration of Batroxobin may reduce the apoptosis of neurons induced by cerebral ischemia and reperfusion and afford significant cerebroprotection in the model of focal cerebral ischemia and reperfusion.

  19. Hydrogen sulfide prevents Abeta-induced neuronal apoptosis by attenuating mitochondrial translocation of PTEN.

    Science.gov (United States)

    Cui, Weigang; Zhang, Yinghua; Yang, Chenxi; Sun, Yiyuan; Zhang, Min; Wang, Songtao

    2016-06-14

    Neuronal cell apoptosis is an important pathological change in Alzheimer's disease (AD). Hydrogen sulfide (H(2)S) is known to be a novel gaseous signaling molecule and a cytoprotectant in many diseases including AD. However, the molecular mechanism of the antiapoptosis activity of H(2)S in AD is not yet fully understood. The aim of the present study is to evaluate the inhibitory effects of H(2)S on Abeta (Aβ)-induced apoptosis and the molecular mechanisms underlying primary neuron cells. Our results showed that sodium hydrosulfide (NaHS), a donor of H(2)S, significantly ameliorated Aβ-induced cell apoptosis. NaHS also reversed the Aβ-induced translocation of the phosphatase and tensin homologs deleted on chromosome 10 (PTEN) from the cytosol to the mitochondria. Furthermore, H(2)S increased the level of p-AKT/AKT significantly. Interestingly, the antiapoptosis effects of H(2)S were blocked down by specific PI3K/AKT inhibitor wortmannin. In conclusion, these data indicate that H(2)S inhibits Aβ-induced neuronal apoptosis by attenuating mitochondrial translocation of PTEN and that activation of PI3K/AKT signaling pathway plays a critical role in H(2)S-mediated neuronal protection. Our findings provide a novel route into the molecular mechanisms of neuronal apoptosis in AD.

  20. A viral vector expressing hypoxia-inducible factor 1 alpha inhibits hippocampal neuronal apoptosis

    Institute of Scientific and Technical Information of China (English)

    Xiqing Chai; Weina Kong; Lingyun Liu; Wenguo Yu; Zhenqing Zhang; Yimin Sun

    2014-01-01

    Hypoxia-inducible factor 1 (HIF-1) attenuates amyloid-beta protein neurotoxicity and decreases apoptosis induced by oxidative stress or hypoxia in cortical neurons. In this study, we construct-ed a recombinant adeno-associated virus (rAAV) vector expressing the human HIF-1αgene (rAAV-HIF-1α), and tested the assumption that rAAV-HIF-1αrepresses hippocampal neuronal apoptosis induced by amyloid-beta protein. Our results conifrmed that rAAV-HIF-1αsigniifcant-ly reduces apoptosis induced by amyloid-beta protein in primary cultured hippocampal neurons. Direct intracerebral rAAV-HIF-1αadministration also induced robust and prolonged HIF-1αproduction in rat hippocampus. Single rAAV-HIF-1αadministration resulted in decreased apoptosis of hippocampal neurons in an Alzheimer’s disease rat model established by intrace-rebroventricular injection of aggregated amyloid-beta protein (25-35). Our in vitro and in vivo ifndings demonstrate that HIF-1 has potential for attenuating hippocampal neuronal apoptosis induced by amyloid-beta protein, and provides experimental support for treatment of neurode-generative diseases using gene therapy.

  1. LINGO-1 receptor promotes neuronal apoptosis by inhibiting WNK3 kinase activity.

    Science.gov (United States)

    Zhang, Zhaohuan; Xu, Xiaohui; Xiang, Zhenghua; Yu, Zhongwang; Feng, Jifeng; He, Cheng

    2013-04-26

    LINGO-1 is a functional component of the Nogo receptor 1 · p75(NTR) · LINGO-1 and Nogo receptor 1 · TAJ (TNFRSF19/TROY)·LINGO-1 signaling complexes. It has recently been shown that LINGO-1 antagonists significantly improve neuronal survival after neural injury. However, the mechanism by which LINGO-1 signaling influences susceptibility to apoptosis remains unknown. In an effort to better understand how LINGO-1 regulates these signaling pathways, we used an established model of serum deprivation (SD) to induce neuronal apoptosis. We demonstrate that treatment either with a construct containing the intracellular domain of LINGO-1 or with Nogo66, a LINGO-1 receptor complex agonist, resulted in an enhanced rate of apoptosis in primary cultured cortical neurons under SD. Reducing the expression levels of the serine/threonine kinase WNK3 using shRNA or inhibiting its kinase activity had similar effects on the survival of serum-deprived neurons. Consistent with these observations, we found that LINGO-1 and WNK3 co-localized and co-precipitated in cultured cortical neurons and brain tissue. Significantly, this co-association was enhanced by Nogo66 treatment. Binding of WNK3 to the intracellular domain of LINGO-1 led to a reduction in WNK3 kinase activity, as did Nogo66 stimulation. Moreover, in vitro and in vivo evidence indicates that endogenous WNK3 suppresses SD-induced neuronal apoptosis in a kinase-dependent manner, as the expression of either a WNK3 RNAi construct or a kinase-dead N-terminal fragment of WNK3 led to increased apoptosis. Taken together, our results show that LINGO-1 potentiates neuronal apoptosis, likely by inhibiting WNK3 kinase activity.

  2. Intrinsic and extrinsic pathway signaling during neuronal apoptosis: lessons from the analysis of mutant mice.

    Science.gov (United States)

    Putcha, Girish V; Harris, Charles A; Moulder, Krista L; Easton, Rachael M; Thompson, Craig B; Johnson, Eugene M

    2002-04-29

    Trophic factor deprivation (TFD)-induced apoptosis in sympathetic neurons requires macromolecular synthesis-dependent BAX translocation, cytochrome c (cyt c) release, and caspase activation. Here, we report the contributions of other intrinsic and extrinsic pathway signals to these processes. Sympathetic neurons expressed all antiapoptotic BCL-2 proteins examined, yet expressed only certain BH3-only and multidomain proapoptotic BCL-2 family members. All coexpressed proapoptotic proteins did not, however, exhibit functional redundancy or compensatory expression, at least in the Bax-/-, Bak-/-, Bim-/-, Bid-/-, and Bad-/- neurons examined. Although the subcellular distribution or posttranslational modification of certain BCL-2 proteins changed with TFD, neither transcriptional nor posttranslational mechanisms regulated the expression or subcellular localization of BID, BAD, or BAK in this paradigm. Despite modest induction of Fas and FasL expression, Fas-mediated signaling did not contribute to TFD-induced apoptosis in sympathetic neurons. Similar findings were obtained with K+ withdrawal-induced apoptosis in cerebellar granule neurons, a model for activity-dependent neuronal survival in the CNS. Thus, expression alone does not guarantee functional redundancy (or compensation) among BCL-2 family members, and, at least in some cells, extrinsic pathway signaling and certain BH3-only proteins (i.e., BID and BAD) do not contribute to BAX-dependent cyt c release or apoptosis caused by TFD.

  3. Effect of morphine preconditioning on neuronal apoptosis following cerebral ischemia/reperfusion injury

    Institute of Scientific and Technical Information of China (English)

    He Dong; Xiangyu Ji; Dong Wang; Yueyi Ren; Shiduan Wang; Jianfang Song

    2010-01-01

    Apoptosis,a form of neuronal damage,takes place following cerebral ischemia/reperfusion injury,and caspase-3 plays an important role in apoptosis.Studies have shown that morphine preconditioning influences neuronal apoptosis and related protein expression following cerebral ischemia/reperfusion injury.In the present study,neuronal degeneration was attenuated,and the number of apoptotic cells and caspase-3 expression decreased following morphine preconditioning in a rat model of cerebral ischemia/reperfusion injury.Moreover,pathological changes were attenuated with increasing morphine doses,as well as the number of apoptotic cells and caspase-3 expression.Results from the present study revealed that morphine preconditioning reduced ischemic brain injury and improved cerebral ischemic tolerance in a dose-dependent manner.The anti-apoptotic mechanism of morphine is closely related to Caspase-3.

  4. Apoptosis in subicular neurons: A comparison between suicide and Addison's disease

    Science.gov (United States)

    Printha, K.; Hulathduwa, S. R.; Samarasinghe, K.; Suh, Y. H.; De Silva, K. R. D.

    2009-01-01

    Background: Stress and depression shows possible links to neuronal death in hippocampus. Subiculum plays a prominent role in limbic stress integration and direct effect of corticosteroids on subicular neurons needs to be defined to assess its subsequent impact on hippocampal plasticity. Aim: This study was intended to assess apoptosis in subicular neurons of a young depressed suicide victim, where presumably stress induced excess of corticosteroids and a case of young Addison's disease with low level of corticosteroids. Materials and Method: Both bilateral adrenal glands (Addison's) and subiculum (both cases) were initially stained with hematoxylin and eosin; subicular neurons of both cases were examined for the degree of apoptosis using ‘ApopTag Kit’. Apoptotic cell counts were expressed as average number of labeled cells/mm2 and the results were analysed statistically using a non-parametric Mann–Whitney U test. Result: Apoptotic neurons were detected in the subicular region of both suicide and Addison victims, and it is statistically significant in both right and left between the cases (P Addison disease where the number of neuronal cell death between right and left was statistically insignificant (P > 0.05). Conclusion: The present study confirms the vulnerability of the subicular neurons to apoptosis, possibly due to corticosteroids in both ends of spectrum. PMID:20048453

  5. Role of Caspase 3 in neuronal apoptosis after acute brain injury

    Institute of Scientific and Technical Information of China (English)

    杨新宇; 杨树源; 张建宁; 雪亮; 胡震

    2002-01-01

    To analyze the role of Caspase 3 in neuronal apoptosis after acute brain injury. Methods: Experiments were carried out with rat diffuse brain trauma model. The neuronal DNA injury in cortex and hippocampus was observed by TUNEL stain.The mRNA and protein expressions and enzyme activation of Caspase 3 were observed by Northern blot, in situ hybridization, immunohistochemistry stain and Western blot, respectively. Special Caspase 3 enzyme inhibitor was used to observe the therapeutic effect. Results: TUNEL positive neurons appeared 2 hours after severe trauma, peaked at 1 day and lasted for 7 days.Northern blot showed that the Caspase 3 mRNA expression was increased and peaked at 1 day, about twice higher than the control. In the area of cortex and hippocampus,positive mRNA staining neurons appeared most distinct on one day. With the antibody for Caspase 3 P20 subunit, the active Caspase 3 expression peaked at 1-3 days. The electrophoresis band of PARP degradation would be seen by Western blot. Caspase 3 enzyme inhibitor could reduce apoptotic neuronal death without any effect on Caspase 3 P20 subunit expression. Conclusions: After brain trauma, Caspase 3 mRNA and protein expressions and enzyme activation are enhanced in combination with neuronal apoptosis. Special Caspase 3 enzyme inhibitor can apparently decrease the neuronal apoptosis.

  6. Rhubarb extract has a protective role against radiation-induced brain injury and neuronal cell apoptosis.

    Science.gov (United States)

    Lu, Kui; Zhang, Cheng; Wu, Wenjun; Zhou, Min; Tang, Yamei; Peng, Ying

    2015-08-01

    Oxidative stress caused by ionizing radiation is involved in neuronal damage in a number of disorders, including trauma, stroke, Alzheimer's disease and amyotrophic lateral sclerosis. Ionizing radiation can lead to the formation of free radicals, which cause neuronal apoptosis and have important roles in the development of some types of chronic brain disease. The present study evaluated the effects of varying concentrations (2, 5 and 10 µg/ml) of ethanolic rhubarb extract on the neuronal damage caused by irradiation in primary neuronal cultures obtained from the cortices of rat embryos aged 20 days. Brain damage was induced with a single dose of γ-irradiation that induced DNA fragmentation, increased lactate dehydrogenase release in neuronal cells and acted as a trigger for microglial cell proliferation. Treatment with rhubarb extract significantly decreased radiation-induced lactate dehydrogenase release and DNA fragmentation, which are important in the process of cell apoptosis. The rhubarb extract exhibited dose-dependent inhibition of lactate dehydrogenase release and neuronal cell apoptosis that were induced by the administration of ionizing radiation. The effect of a 10 µg/ml dose of rhubarb extract on the generation of reactive oxygen species (ROS) induced by radiation was also investigated. This dose led to significant inhibition of ROS generation. In conclusion, the present study showed a protective role of rhubarb extract against irradiation-induced apoptotic neuronal cell death and ROS generation.

  7. Apoptosis of glutamatergic neurons fails to trigger a neurogenic response in the adult neocortex.

    Science.gov (United States)

    Diaz, Frank; McKeehan, Nicholas; Kang, Wenfei; Hébert, Jean M

    2013-04-10

    Adult neurogenesis is actively studied in part because of the potential to manipulate endogenous neural stem and progenitor cells for tissue repair. Although constitutive generation of neurons in the adult rodent olfactory bulb and hippocampal dentate gyrus is widely accepted and stroke-induced generation of striatal inhibitory neurons consistently observed, evidence supporting the generation of neurons in the neocortex after neuronal loss remains slim. Nevertheless, a few studies suggested that targeted apoptosis of neocortical glutamatergic neurons could trigger the generation of new ones in the adult brain. In light of such studies, we tested whether apoptosis of glutamatergic cortical neurons using two novel transgenic approaches in mice, an inducible Caspase-8 protein and an inducible diphtheria toxin gene, results in new neurons. After a thorough analysis, no new neurons were detected in the neocortex. Interestingly, an increase in the expression of the neuroblast marker DCX was observed in both models, in some cases in cells with morphologies previously associated with poststroke neuroblasts, but DCX(+) cells coexpressed the oligodendrocyte precursor marker Olig2, suggesting caution when using DCX as a marker for neuroblasts after injury. Given that the adult neocortex lacks an innate potential to regenerate lost glutamatergic neurons, future strategies should concentrate on manipulating the differentiation potential of endogenous or exogenous precursor cells.

  8. Bifunctional apoptosis inhibitor (BAR) protects neurons from diverse cell death pathways.

    Science.gov (United States)

    Roth, W; Kermer, P; Krajewska, M; Welsh, K; Davis, S; Krajewski, S; Reed, J C

    2003-10-01

    The bifunctional apoptosis regulator (BAR) is a multidomain protein that was originally identified as an inhibitor of Bax-induced apoptosis. Immunoblot analysis of normal human tissues demonstrated high BAR expression in the brain, compared to low or absent expression in other organs. Immunohistochemical staining of human adult tissues revealed that the BAR protein is predominantly expressed by neurons in the central nervous system. Immunofluorescence microscopy indicated that BAR localizes mainly to the endoplasmic reticulum (ER) of cells. Overexpression of BAR in CSM 14.1 neuronal cells resulted in significant protection from a broad range of cell death stimuli, including agents that activate apoptotic pathways involving mitochondria, TNF-family death receptors, and ER stress. Downregulation of BAR by antisense oligonucleotides sensitized neuronal cells to induction of apoptosis. Moreover, the search for novel interaction partners of BAR identified several candidate proteins that might contribute to the regulation of neuronal apoptosis (HIP1, Hippi, and Bap31). Taken together, the expression pattern and functional data suggest that the BAR protein is involved in the regulation of neuronal survival.

  9. Enhanced apoptosis during early neuronal differentiation in mouse ES cells with autosomal imbalance

    Institute of Scientific and Technical Information of China (English)

    Yoshiteru Kai; Teruhiko Wakayama; Mitsuo Oshimura; Chi Chiu Wang; Satoshi Kishigami; Yasuhiro Kazuki; Satoshi Abe; Masato Takiguchi; Yasuaki Shirayoshi; Toshiaki Inoue; Hisao Ito

    2009-01-01

    Although particular chromosomal syndromes are phenotypically and clinically distinct, the majority of individuals with autosomai imbalance, such as aneuploidy, manifest mental retardation. A common abnormal phenotype of Down syndrome (DS), the most prevalent autosomal aneuploidy, shows a reduction in both the number and the density of neurons in the brain. As a DS model, we have recently created chimeric mice from ES cells containing a single human chromosome 21. The mice mimicked the characteristic phenotypic features of DS, and ES cells showed a higher incidence of apoptosis during early neuronal differentiation in vitro. In this study, we examined the induction of anomalous early neural development by aneuploidy in mouse ES cells by transferring various human chromosomes or additional mouse chromosomes. Results showed an elevated incidence of apoptosis in all autosome-aneuploid clones examined during early neuronal differentiation in vitro. Further, cDNA microarray analysis revealed a common cluster of down-regulated genes, of which eight known genes are related to cell proliferation, neurite outgrowth and differentiation. Importantly, targeting of these genes by siRNA knockdown in normal mouse ES cells led to enhanced apoptosis during early neuronal differentiation. These findings strongly suggest that autosomal imbalance is associated with general neuronal loss through a common molecular mechanism for apoptosis.

  10. Sugammadex, a neuromuscular blockade reversal agent, causes neuronal apoptosis in primary cultures.

    Science.gov (United States)

    Palanca, José M; Aguirre-Rueda, Diana; Granell, Manuel V; Aldasoro, Martin; Garcia, Alma; Iradi, Antonio; Obrador, Elena; Mauricio, Maria Dolores; Vila, Jose; Gil-Bisquert, Anna; Valles, Soraya L

    2013-01-01

    Sugammadex, a γ-cyclodextrin that encapsulates selectively steroidal neuromuscular blocking agents, such as rocuronium or vecuronium, has changed the face of clinical neuromuscular pharmacology. Sugammadex allows a rapid reversal of muscle paralysis. Sugammadex appears to be safe and well tolerated. Its blood-brain barrier penetration is poor (Sugammadex in neurons in primary culture. Here we show that clinically relevant sugammadex concentrations cause apoptotic/necrosis neuron death in primary cultures. Studies on the underlying mechanism revealed that sugammadex-induced activation of mitochondria-dependent apoptosis associates with depletion of neuronal cholesterol levels. Furthermore SUG increase CytC, AIF, Smac/Diablo and CASP-3 protein expression in cells in culture. Potential association of SUG-induced alteration in cholesterol homeostasis with oxidative stress and apoptosis activation occurs. Furthermore, resistance/sensitivity to oxidative stress differs between neuronal cell types.

  11. Apoptosis and autophagy in nigral neurons of patients with Parkinson's disease.

    Science.gov (United States)

    Anglade, P; Vyas, S; Javoy-Agid, F; Herrero, M T; Michel, P P; Marquez, J; Mouatt-Prigent, A; Ruberg, M; Hirsch, E C; Agid, Y

    1997-01-01

    Parkinson's disease (PD) is a neurodegenerative disorder characterized by progressive cell loss confined mostly to dopaminergic neurons of the substantia nigra. Several factors, including oxidative stress, and decreased activity of complex I mitochondrial respiratory chain, are involved in the degenerative process. Yet, the underlying mechanisms leading to dopaminergic cell loss remain elusive. Morphological assessment for different modes of cell death: apoptosis, necrosis or autophagic degeneration, can contribute significantly to the understanding of this neuronal loss. Ultrastructural examination revealed characteristics of apoptosis and autophagic degeneration in melanized neurons of the substantia nigra in PD patients. The results suggest that even at the final stage of the disease, the dopaminergic neurons are undergoing active process of cell death.

  12. Metallothionein in Brain Disorders

    Directory of Open Access Journals (Sweden)

    Daniel Juárez-Rebollar

    2017-01-01

    Full Text Available Metallothioneins are a family of proteins which are able to bind metals intracellularly, so their main function is to regulate the cellular metabolism of essential metals. There are 4 major isoforms of MTs (I–IV, three of which have been localized in the central nervous system. MT-I and MT-II have been localized in the spinal cord and brain, mainly in astrocytes, whereas MT-III has been found mainly in neurons. MT-I and MT-II have been considered polyvalent proteins whose main function is to maintain cellular homeostasis of essential metals such as zinc and copper, but other functions have also been considered: detoxification of heavy metals, regulation of gene expression, processes of inflammation, and protection against free radicals generated by oxidative stress. On the other hand, the MT-III has been related in events of pathogenesis of neurodegenerative diseases such as Parkinson and Alzheimer. Likewise, the participation of MTs in other neurological disorders has also been reported. This review shows recent evidence about the role of MT in the central nervous system and its possible role in neurodegenerative diseases as well as in brain disorders.

  13. Chemotactic and mitogenic stimuli of neuronal apoptosis in patients with medically intractable temporal lobe epilepsy

    Science.gov (United States)

    Fiala, Milan; Avagyan, Hripsime; Merino, Jose Joaquin; Bernas, Michael; Valdivia, Juan; Espinosa-Jeffrey, Araceli; Witte, Marlys; Weinand, Martin

    2012-01-01

    To identify the upstream signals of neuronal apoptosis in patients with medically intractable temporal lobe epilepsy (TLE), we evaluated by immunohistochemistry and confocal microscopy brain tissues of 13 TLE patients and 5 control patients regarding expression of chemokines and cell-cycle proteins. The chemokine RANTES (CCR5) and other CC-chemokines and apoptotic markers (caspase-3, -8, -9) were expressed in lateral temporal cortical and hippocampal neurons of TLE patients, but not in neurons of control cases. The chemokine RANTES is usually found in cytoplasmic and extracellular locations. However, in TLE neurons, RANTES was displayed in an unusual location, the neuronal nuclei. In addition, the cell-cycle regulatory transcription factor E2F1 was found in an abnormal location in neuronal cytoplasm. The pro-inflammatory enzyme cyclooxygenase-2 and cytokine interleukin-1β were expressed both in neurons of patients suffering from temporal lobe epilepsy and from cerebral trauma. The vessels showed fibrin leakage, perivascular macrophages and expression of IL-6 on endothelial cells. In conclusion, the cytoplasmic effects of E2F1 and nuclear effects of RANTES might have novel roles in neuronal apoptosis of TLE neurons and indicate a need to develop new medical and/or surgical neuroprotective strategies against apoptotic signaling by these molecules. Both RANTES and E2F1 signaling are upstream from caspase activation, thus the antagonists of RANTES and/or E2F1 blockade might be neuroprotective for patients with medically intractable temporal lobe epilepsy. The results have implications for the development of new medical and surgical therapies based on inhibition of chemotactic and mitogenic stimuli of neuronal apoptosis in patients with medically intractable temporal lobe epilepsy. PMID:22444245

  14. Detecting the apoptosis of dopamine neurons with immunohistochemical staining and double-staining technique

    Institute of Scientific and Technical Information of China (English)

    Jiguo Zhang; Jing Zhang; Feng Zhang; Yunsheng Gao

    2006-01-01

    BACKGROUND: It is proved that the onset of Parkinson disease companies with neuronal apoptosis of dopamine in substantia nigra of midbrain. Previous researches on neuronal apoptosis of dopamine were analyzed on their consecutive tissue sections with immunohistochemical single-labeling method, immunofluorescence and electron microscope, and there are significant differences.OBJECTIVE: To observe the feasibility of neuronal apoptosis of dopamine with in situ end labeling and tyrosine-hydroxylase antibody immunohistochemical double-labeling staining technique.DESIGN: Controlled study.SETTING: College of Pharmacology of Taishan Medical College; College of Management of Taishan Medical College.MATERIALS: Wistar rats with 2 weeks old and of clean grade were provided by the Animal Center of Taishan Medical College. In situ end labeling kit (terminal deoxynucleotidyl transferase, mixed reactive solution of nucleotide, transfusion-POD), monoclonal antibody of rat antibody against tyrosine hydroxylase (Boehriuser).METHODS: The experiment was completed at the Pharmacological Laboratory of Taishan Medical College from February to December 2005. Tissue from midbrain of rats was taken out to make paraffin sections to observe the neuronal apoptosis of dopamine under microscope with in situ end labeling and tyrosine-hydroxylase antibody immunohistochemical double-labeling staining technique.MAIN OUTCOME MEASURES: Neuronal apoptosis of dopamine with in situ end labeling and tyrosine-hydroxylase antibody immunohistochemical double-labeling staining technique.RESULTS:① After double-labeling staining,two kinks of positive products were observed in neurons of dopamine which were suffered from apoptosis. One stained with tyrosine hydroxylase was hyacinthine, and the other stained with in situ end labeling was buffy. Cells of positive products stained with in situ end labeling shaped as strap and bend and was distributed in clustering.Cytoplasm was hyacinthine, staining was symmetrical

  15. Studies on the apoptosis of autonomic neuron in streptozotocin-induced diabetic rats

    Institute of Scientific and Technical Information of China (English)

    Qinglin Lou; Rongwen Bian; Tao Peng; Xiaojun Ouyang; Hui Xia; Liubao Gu; Yongzhen Mo

    2006-01-01

    Objective: The aim of the study was to investigate apoptosis of the autonomic neuron in streptozotocin-induced diabetic rats, and observe the effect of intervention with nerve growth factor (NGF) on the apoptosis. Methods: A total of 29 male Sprague-Dawley rats were divided into three groups, i.e. normal control (NC, n = 12), untreated diabetic (DM, n =9) and diabetic treated with NGF daily of 500 μg/kg for 30 days(DM+NGF, n = 8). The diabetic rat models were produced by intraperitoneal injection with streptozotocin(65 mg/kg). Over 3 months since the diabetes were setup, the superior cervical sympathetic ganglions(SCG) and the celiac ganglions(CG) were removed and fixed with 10% paraformaldchyde. Apoptosis was measured using terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling(TUNEL). Apoptotic index (AI) was calculated by computer image analysis system. Results: The AI of SCG and CG in DM and DM+NGF group were significant higher than those of NC group (P < 0.001) respectively. There was no difference of AI of SCG and CG between DM group and DM+NGF group (both P > 0.05). Conclusion: Neuron apoptosis may contribute to the pathophysiology of diabetic autonomic neuropathy and NGF can not prevent the apoptosis of autonomic neuron in diabetic rats.

  16. CaMKII inhibition promotes neuronal apoptosis by transcriptionally upregulating Bim expression.

    Science.gov (United States)

    Zhao, Yiwei; Zhu, Lin; Yu, Shaojun; Zhu, Jing; Wang, Chong

    2016-09-28

    The effects of Ca/calmodulin-dependent protein kinase II (CaMKII) on neuronal apoptosis are complex and contradictory, and the underlying mechanisms remain unclear. Bcl-2-interacting mediator of cell death (Bim) is an important proapoptotic protein under many physiological and pathophysiological conditions. However, there is no evidence that CaMKII and Bim are mechanistically linked in neuronal apoptosis. In this study, we showed that CaMKII inhibition by the inhibitors KN-62 and myristoylated autocamtide-2-related inhibitory peptide promoted apoptosis in cerebellar granule neurons in a dose-dependent manner. CaMKII inhibition increased Bim protein and messenger RNA levels. The expression of early growth response factor-1, a transcription factor of Bim, was also induced by CaMKII inhibitors. These data suggested that CaMKII repressed the transcriptional expression of Bim. Moreover, knockdown of Bim using small interfering RNAs attenuated the proapoptotic effects of CaMKII inhibition. Taken together, this is the first report to show that CaMKII inhibition transcriptionally upregulates Bim expression to promote neuronal apoptosis, providing new insights into the proapoptotic mechanism of CaMKII inhibition.

  17. Ischemic preconditioning reduces ischemic brain injury by suppressing nuclear factor kappa B expression and neuronal apoptosis

    Institute of Scientific and Technical Information of China (English)

    Songsheng Shi; Weizhong Yang; Xiankun Tu; Chunmei Chen; Chunhua Wang

    2013-01-01

    Ischemic stroke induces a series of complex pathophysiological events including blood-brain barrier disruption, inflammatory response and neuronal apoptosis. Previous studies demonstrate that ischemic preconditioning attenuates ischemic brain damage via inhibiting blood-brain barrier disruption and the inflammatory response. Rats underwent transient (15 minutes) occlusion of the bilateral common carotid artery with 48 hours of reperfusion, and were subjected to permanent middle cerebral artery occlusion. This study explored whether ischemic preconditioning could reduce ischemic brain injury and relevant molecular mechanisms by inhibiting neuronal apoptosis. Results found that at 72 hours following cerebral ischemia, myeloperoxidase activity was enhanced, malondialdehyde levels increased, and neurological function was obviously damaged. Simultaneously, neuronal apoptosis increased, and nuclear factor-κB and cleaved caspase-3 expression was significantly increased in ischemic brain tissues. Ischemic preconditioning reduced the cerebral ischemia-induced inflammatory response, lipid peroxidation, and neurological function injury. In addition, ischemic preconditioning decreased nuclear factor-κB p65 and cleaved caspase-3 expression. These results suggested that ischemic preconditioning plays a protective effect against ischemic brain injury by suppressing the inflammatory response, reducing lipid peroxidation, and neuronal apoptosis via inhibition of nuclear factor-κB and cleaved caspase-3 expression.

  18. MADP, a salidroside analog, protects hippocampal neurons from glutamate induced apoptosis.

    Science.gov (United States)

    Xian, Hua; Zhao, Jing; Zheng, Yuan; Wang, Meihong; Huang, Jun; Wu, Bingxin; Sun, Cheng; Yang, Yumin

    2014-05-08

    To investigate the anti-apoptotic effect of MADP, an analog of salidroside, against glutamate induced apoptosis in the cultured rat hippocampal neurons. Cytotoxicity was determined by the MTT method and lactate dehydrogenase release to the medium. Cell apoptosis was evaluated by Hoechst 33342 staining, TUNEL assay and flow cytometric analysis. Western blotting was applied for detecting protein levels of cellular signaling molecules. Our results showed that glutamate exposure significantly induces cell apoptosis, whereas the pretreatment of salidroside or MADP remarkably improves cell viability. Most importantly, the anti-apoptotic effect of MADP against glutamate insult is superior to salidroside. To explore the involved mechanisms, we measured some pro-apoptotic and anti-apoptotic protein levels, and several cell survival signaling pathways were analyzed as well. No visible alterations in Bcl-2 and Bax protein levels were observed by MADP or salidroside. Akt and JNK phosphorylation was robustly stimulated by MADP in the glutamate-treated neurons. Salidroside treatment results in a slight activation in Akt, while no significant alteration in JNK activity was observed. MADP exhibits higher capacity to attenuate glutamate induced cell apoptosis in the cultured rat hippocampal neurons, suggesting that MADP might be a better candidate than salidroside for developing novel drugs treating neuron loss associated disorders. Copyright © 2014 Elsevier Inc. All rights reserved.

  19. Effect of isoflurane on neuronal apoptosis in rats subjected to focal cerebral ischemia.

    Science.gov (United States)

    Kawaguchi, Masahiko; Drummond, John C; Cole, Daniel J; Kelly, Paul J; Spurlock, Mark P; Patel, Piyush M

    2004-03-01

    Although isoflurane can reduce ischemic neuronal injury after short postischemic recovery intervals, this neuroprotective efficacy is not sustained. Neuronal apoptosis can contribute to the gradual increase in infarct size after ischemia. This suggests that isoflurane, although capable of reducing early neuronal death, may not inhibit ischemia-induced apoptosis. We investigated the effects of isoflurane on markers of apoptosis in rats subjected to focal ischemia. Fasted Wistar-Kyoto rats were anesthetized with isoflurane and randomly allocated to awake (n = 40) or isoflurane (n = 40) groups. Animals in both groups were subjected to focal ischemia by filament occlusion of the middle cerebral artery for 70 min. Pericranial temperature was servo-controlled at 37 degrees C +/- 0.2 degrees C throughout the experiment. In the awake group, isoflurane was discontinued and the animals were allowed to awaken. In the isoflurane group, isoflurane anesthesia was maintained at 1.5 MAC (minimum alveolar anesthetic concentration). Animals were killed 7 h, 1 day, 4 days, or 7 days after reperfusion (n = 10/group/time point). The area of cerebral infarction was measured by image analysis in a hematoxylin and eosin stained section. In three adjacent sections, apoptotic neurons were identified by TUNEL staining and immunostaining for active caspase-9 and caspase-3. Infarct size was smaller in the isoflurane group than the awake group 7 h, 1 day, and 4 days after reperfusion (P awake versus isoflurane group. After a recovery period of 4 or 7 days, the number of apoptotic cells in the isoflurane group was more than in the awake group. After 7 days, the number of caspase-3 and -9 positive neurons was more in the isoflurane group (P < 0.05). The data indicate that isoflurane delays but does not prevent the development of cerebral infarction caused by ischemia. Isoflurane reduced the development of apoptosis early after ischemia but did not prevent it at later stages of postischemic

  20. Role of Mitochondria in Neuron Apoptosis during Ischemia-Reperfusion Injury

    Institute of Scientific and Technical Information of China (English)

    段秋红; 王西明; 王忠强; 卢涛; 韩义香; 何善述

    2004-01-01

    To investigate the role of mitochondria in neuronal apoptosis, ischemia-reperfusion mediated neuronal cell injury model was established by depriving of glucose, serum and oxygen in media.DNA fragmentation, cell viability, cytochrome C releasing, caspase3 activity and mitochondrial transmembrane potential were observed after N2a cells suffered the insults. The results showed that N2a cells in ischemic territory exhibited survival damage, classical cell apoptosis change, DNA ladder and activation of caspase3. Apoptosis-related alterations in mitochondrial functions, including release of cytochrome C and depression of mitochondrial transmembrane potential (△ψm)were testified in N2a cells after mimic ischemia-reperfusion. Moreover, activation of caspase3 occurred following the release of cytochrome C. However, the inhibitor of caspase3, Ac-DEVDinhibitor of mitochondria permeability transition pore only partly inhibited caspase3 activity and reduced DNA damage. Interestingly, treatment of Z-IETD-FMK, an inhibitor of caspase8 could comthat there were caspase3 dependent and independent cellular apoptosis pathways in N2a cells suffering ischemia-reperfusion insults. Mitochondria dysfunction may early trigger apoptosis and amplify apoptosis signal.

  1. Neuronal Death Following Soman Intoxication: Necrosis or Apoptosis?

    Science.gov (United States)

    2006-05-01

    progression of apoptotic cell death in the rat piriform cortex after soman intoxication. At various time intervals after seizure onset, animals were...We investigated the temporal progression of apoptotic cell death in the rat piriform cortex after soman intoxication. At various time intervals...and neuronal loss after acute soman exposure included the piriform cortex, hippocampus, septum, entorhinal cortex, dentate gyrus, amygdala, and

  2. Study of apoptosis pattern of dopaminergic neurons and neuroprotective effect of nicotine in MPTP mouse model

    Institute of Scientific and Technical Information of China (English)

    Dan Hu; Wei Cao; Shenggang Sun

    2007-01-01

    Objective:To investigate the apoptosis of dopaminergic neurons and the protective effect of nicotine in 1-methyl-4-phenyl-1, 2,3,6-tetrahydropyridine (MPTP)-induced mouse model of Parkinson's disease. Methods :The mouse model of Parkinson's disease were formed by MPTP (30 mg/kg/d×7, i.p.); and the loss and apoptosis of dopaminergic neurons was observed by Tyrosine Hydroxylase (TH) and TUNEL stains. In "Nicotime plus MPTP" group, mice were pretreated with nicotine before MPTP injection. The putative protective effect of nicotine was analyzed. Results:The number of TH-positive cells decreased during MPTP treatment. Apoptotic neurons began to appear after three injections of MPTP and peaked on the 8th day.In the MPTP-intoxicated mice treated with nicotine, the loss of TH-positive cells was significantly less than that of MPTP-treated group (30 mg/kg/d×7)(P < 0.05). Conclusion:The chronic treatment of MPTP can induce the apoptosis of dopaminergic neurons in substantia nigra, and nicotine might have a neuroprotecitve effect on dopaminergic neurons against MPTP toxicity.

  3. The pathways by which mild hypothermia inhibits neuronal apoptosis following ischemia/reperfusion injur y

    Institute of Scientific and Technical Information of China (English)

    Chun Luo; Su-yue Pan

    2015-01-01

    Several studies have demonstrated that mild hypothermia exhibits a neuroprotective role and it can inhibit endothelial cell apoptosis following ischemia/reperfusion injury by decreasing casp-ase-3 expression. It is hypothesized that mild hypothermia exhibits neuroprotective effects on neurons exposed to ischemia/reperfusion condition produced by oxygen-glucose deprivation. Mild hypothermia signiifcantly reduced the number of apoptotic neurons, decreased the expres-sion of pro-apoptotic protein Bax and increased mitochondrial membrane potential, with the peak of anti-apoptotic effect appearing between 6 and 12 hours after the injury. These ifndings indicate that mild hypothermia inhibits neuronal apoptosis following ischemia/reperfusion injury by protecting the mitochondria and that the effective time window is 6–12 hours after ischemia/reperfusion injury.

  4. The pathways by which mild hypothermia inhibits neuronal apoptosis following ischemia/reperfusion injury

    Directory of Open Access Journals (Sweden)

    Chun Luo

    2015-01-01

    Full Text Available Several studies have demonstrated that mild hypothermia exhibits a neuroprotective role and it can inhibit endothelial cell apoptosis following ischemia/reperfusion injury by decreasing casp-ase-3 expression. It is hypothesized that mild hypothermia exhibits neuroprotective effects on neurons exposed to ischemia/reperfusion condition produced by oxygen-glucose deprivation. Mild hypothermia significantly reduced the number of apoptotic neurons, decreased the expression of pro-apoptotic protein Bax and increased mitochondrial membrane potential, with the peak of anti-apoptotic effect appearing between 6 and 12 hours after the injury. These findings indicate that mild hypothermia inhibits neuronal apoptosis following ischemia/reperfusion injury by protecting the mitochondria and that the effective time window is 6-12 hours after ischemia/reperfusion injury

  5. Effect of cholecystokinin-8 on in vitro cultured rat cortical neurons against apoptosis

    Institute of Scientific and Technical Information of China (English)

    Ying Liu; Jiangbao Zhou

    2006-01-01

    BACKGROUND: Cholecystokinin (CCK-8) can regulate the synthesis of NO, release of amino acid substance and suppress Ca2+ inflow. It is unknown about neuroprotection of CCK-8 on neuronal apoptosis and its relationship with nerve growth factor (NGF).OBJECTryE: To investigate the protective effect of CCK-8 on in vitro cultured rat cortical neurons against apoptosis induced by glutamate, and explore its effect on expression of NGF in the neurons during apoptosis.DESIGN: Randomized controlled experiment on the basis of cells.SETTING: Children's Research Institute Affiliated to Children Hospital of Chongqing Medical University.MATERIALS: Eighty SD rats of 1-day old; DMEM/F12 culture medium (Biochrom Company, Germany);Fetal bovine serum (TBD Company, Tianjin); CCK-8 (Sigma Company, USA). Glutamate (Bioengineering Company, Shanghai); TUNEL kit and NGF- in situ hybridization kit (Boster Bioengineering Company,Wuhan); anti-NGF polyclonal antibody (Santa-Cluz Company); NGF immunocytochemistry kit (Zhongshan Company, Beijing).METHODS: The experiments were carried out in Children's Research Institute Affiliated to Children Hospital of Chongqing Medical University from December 2004 to September 2005. Primary cultured cortical neurons from SD rats of 1-day oldwere incubated for 7 days. The cultured cells were divided randomly into 3 groups:experimental group, model group and control group. Neurons in experimental groups were added CCK-8 of 1 ×10-6, 1 ×10-7, 1 ×10-8 μ mol/L respectively, and then added 50 μmol/L glutamate solution a hour later. Neurons in model groups were treated with 50 μ mol/L glutamate solution. In the control group, cells were treated with normal medium. Apoptosis of cultured cortical neurons were observed by fluorescent microscope, the expression of NGF protein and mRNA were determined respectively by immunocytochemistry and in situ hybridization, and apoptosis of cortical neurons was detected with terminal deoxynucleotidyl transferase-mediated nick

  6. Effects of lysophosphatidylcholine on β-amyloid-induced neuronal apoptosis

    Institute of Scientific and Technical Information of China (English)

    Zhen-xia QIN; Hui-yan ZHU; Ying-he HU

    2009-01-01

    Aim: We have investigated the effects of lysophosphatidylcholine (LPC), a product of lipid peroxidation, on Aβ1-42-induced SH-SY5Y cell apoptosis.Methods: The viability of cultured SH-SY5Y cells was measured using a CCK-8 kit. Apoptosis was determined by Chip-based flow cytometric assay. The mRNA transcription of Bcl-2, Bax, and caspase-3 were detected by using reverse transcrip-tion and real-time quantitative PCR and the protein levels of Bax and caspase-3 were analyzed by Western blotting. Thecytosolic calcium concentration of SH-SY5Y cells was tested by calcium influx assay. GZA expression in SH-SYSY cells wassilenced by small interfering RNA.Results: Long-term exposure of SH-SY5Y cells to LPC augmented the neurotoxicity of Aβ1-42. Furthermore, after LPC treatment, the Bax/Bcl-XL ratio and the expression levels, as well as the activity of caspase-3 were, elevated, whereas the expression level of TRAF1 was reduced. Because LPC was reported to be a specific ligand for the orphan G-protein coupled receptor, G2A, we investigated LPC-mediated changes in calcium levels in SH-SY5Y cells. Our results demonstrated that LPC can enhance the Aβ1-42-induced elevation of intracellular calcium. Interestingly, Aβ1-42 significantly increased the expression of G2A in SH-SY5Y cells, whereas knockdown of G2A using siRNA reduced the effects of LPC on Aβ1-42-induced neurotoxicity.Conclusion: The effects of LPC on Aβ1-42-induced apoptosis may occur through the signal pathways of the orphan G-protein coupled receptor.

  7. Metallothionein overexpression and its prognostic relevance in intrahepatic cholangiocarcinoma and extrahepatic hilar cholangiocarcinoma (Klatskin tumors).

    Science.gov (United States)

    Schmitz, Klaus Jürgen; Lang, Hauke; Kaiser, Gernot; Wohlschlaeger, Jeremias; Sotiropoulos, Georgios Charalambos; Baba, Hideo Andreas; Jasani, Bharat; Schmid, Kurt Werner

    2009-12-01

    Metallothionein is a group of small molecular weight cysteine-rich proteins with a broad variety of functions. Metallothionein has been shown to regulate apoptosis and proliferation. Overexpression of metallothionein frequently occurs in human tumors and is related to prognosis as well as therapy response. However, metallothionein expression and its clinical relevance in cholangiocarcinoma have not been investigated. The present study aimed to analyze metallothionein over-expression and its possible prognostic impact in intrahepatic cholangiocarcinoma and hilar extrahepatic cholangiocarcinoma (Klatskin tumors). We investigated the relationship of immunohistochemically demonstrated metallothionein expression with various clinicopathological parameters in a series of 56 intrahepatic and 56 extrahepatic cholangiocarcinoma. In noncancerous bile duct epithelia metallothionein was only occasionally weakly expressed; strong metallothionein overexpression (>50% metallothionein -positive tumor cells) was noted in 7 (12.5%) of 56 intrahepatic cholangiocarcinoma and 14 (25%) of 56 Klatskin tumors, which was associated with poor clinical outcome in univariate Kaplan-Meier testing in both intrahepatic cholangiocarcinoma (P = .002) and Klatskin tumors (P = .034). Moreover, strong metallothionein expression was identified as an independent prognostic parameter in multivariate Cox regression analysis in both intrahepatic cholangiocarcinoma (P = .005) and Klatskin tumors (P = .035). In contrast, cholangiocarcinoma with a papillary phenotype (8/112; 7.1%) exhibited a significant lack of strong metallothionein expression in all 8 of 8 cases. Strong metallothionein expression is identified as an independent poor prognostic parameter, and determination of the metallothionein expression may serve as an additional tool for the therapeutic management of patients with cholangiocarcinoma. In comparison, lack of metallothionein expression seems to be associated with cholangiocarcinoma with a

  8. Human neuronal apoptosis secondary to traumatic brain injury and the regulative role of apoptosis-related genes

    Institute of Scientific and Technical Information of China (English)

    杨树源; 雪亮

    2004-01-01

    Objective: To observe human neuronal apoptosis secondary to traumatic brain injury, and to elucidate its regulative mechanism and the change of expression of apoptosis-related genes.Methods: Specimens of brain were collected from cases of traumatic brain injury in humans. The histological and cellular morphology was examined by light and electron microscopy. The extent of DNA injury to cortical neurons was detected by using TUNEL. By in situ hybridisation and immunohistochemistry the mRNA changes and protein expression of Bcl-2, Bax, p53, and caspase 3 p20 subunit were observed.Results: Apoptotic neurons appeared following traumatic brain injury, peaked at 24 hours and lasted for 7 days. In normal brain tissue activated caspase 3 was rare,but a short time after trauma it became activated. The activity peaked at 20-28 hours and remained higher than normal for 5-7 days. There was no expression of Bcl-2 mRNA and Bcl-2 protein in normal brain tissue but 8 hours after injury their expression became evident and then increased, peaked at 2-3 days and remained higher than normal for 5-7 days. The primary expression of Bax-mRNA and Bax protein was high in normal brain tissue. At 20-28 hours they increased and remained high for 2-3 days; on the 7th days they returned to a normal level. In normal brain tissue, p53mRNA and P53 were minimally expressed.Increased expression was detected at the 8th hour, and decreased at 20-28 hours but still remained higher than normal on the 5th day.Conclusions: Following traumatic injury to the human brain, apoptotic neurons appear around the focus of trauma. The mRNA and protein expression of Bcl-2, Bax and p53 and the activity of caspase 3 enzyme are increased.

  9. Calreticulin Binds to Fas Ligand and Inhibits Neuronal Cell Apoptosis Induced by Ischemia-Reperfusion Injury

    Directory of Open Access Journals (Sweden)

    Beilei Chen

    2015-01-01

    Full Text Available Background. Calreticulin (CRT can bind to Fas ligand (FasL and inhibit Fas/FasL-mediated apoptosis of Jurkat T cells. However, its effect on neuronal cell apoptosis has not been investigated. Purpose. We aimed to evaluate the neuroprotective effect of CRT following ischemia-reperfusion injury (IRI. Methods. Mice underwent middle cerebral artery occlusion (MCAO and SH-SY5Y cells subjected to oxygen glucose deprivation (OGD were used as models for IRI. The CRT protein level was detected by Western blotting, and mRNA expression of CRT, caspase-3, and caspase-8 was measured by real-time PCR. Immunofluorescence was used to assess the localization of CRT and FasL. The interaction of CRT with FasL was verified by coimmunoprecipitation. SH-SY5Y cell viability was determined by MTT assay, and cell apoptosis was assessed by flow cytometry. The measurement of caspase-8 and caspase-3 activity was carried out using caspase activity assay kits. Results. After IRI, CRT was upregulated on the neuron surface and bound to FasL, leading to increased viability of OGD-exposed SH-SY5Y cells and decreased activity of caspase-8 and caspase-3. Conclusions. This study for the first time revealed that increased CRT inhibited Fas/FasL-mediated neuronal cell apoptosis during the early stage of ischemic stroke, suggesting it to be a potential protector activated soon after IRI.

  10. Human group IIA secretory phospholipase A2 induces neuronal cell death via apoptosis.

    Science.gov (United States)

    Yagami, Tatsurou; Ueda, Keiichi; Asakura, Kenji; Hata, Satoshi; Kuroda, Takayuki; Sakaeda, Toshiyuki; Takasu, Nobuo; Tanaka, Kazushige; Gemba, Takefumi; Hori, Yozo

    2002-01-01

    Expression of group IIA secretory phospholipase A2 (sPLA2-IIA) is documented in the cerebral cortex (CTX) after ischemia, suggesting that sPLA2-IIA is associated with neurodegeneration. However, how sPLA2-IIA is involved in the neurodegeneration remains obscure. To clarify the pathologic role of sPLA2-IIA, we examined its neurotoxicity in rats that had the middle cerebral artery occluded and in primary cultures of cortical neurons. After occlusion, sPLA2 activity was increased in the CTX. An sPLA2 inhibitor, indoxam, significantly ameliorated not only the elevated activity of the sPLA2 but also the neurodegeneration in the CTX. The neuroprotective effect of indoxam was observed even when it was administered after occlusion. In primary cultures, sPLA2-IIA caused marked neuronal cell death. Morphologic and ultrastructural characteristics of neuronal cell death by sPLA2-IIA were apoptotic, as evidenced by condensed chromatin and fragmented DNA. Before apoptosis, sPLA2-IIA liberated arachidonic acid (AA) and generated prostaglandin D2 (PGD2), an AA metabolite, from neurons. Indoxam significantly suppressed not only AA release, but also PGD2 generation. Indoxam prevented neurons from sPLA2-IIA-induced neuronal cell death. The neuroprotective effect of indoxam was observed even when it was administered after sPLA2-IIA treatment. Furthermore, a cyclooxygenase-2 inhibitor significantly prevented neurons from sPLA2-IIA-induced PGD2 generation and neuronal cell death. In conclusion, sPLA2-IIA induces neuronal cell death via apoptosis, which might be associated with AA metabolites, especially PGD2. Furthermore, sPLA2 contributes to neurodegeneration in the ischemic brain, highlighting the therapeutic potential of sPLA2-IIA inhibitors for stroke.

  11. Inhibition of apoptosis in neuronal cells infected with Chlamydophila (Chlamydia pneumoniae

    Directory of Open Access Journals (Sweden)

    Albert Elizabeth V

    2008-01-01

    Full Text Available Abstract Background Chlamydophila (Chlamydia pneumoniae is an intracellular bacterium that has been identified within cells in areas of neuropathology found in Alzheimer disease (AD, including endothelia, glia, and neurons. Depending on the cell type of the host, infection by C. pneumoniae has been shown to influence apoptotic pathways in both pro- and anti-apoptotic fashions. We have hypothesized that persistent chlamydial infection of neurons may be an important mediator of the characteristic neuropathology observed in AD brains. Chronic and/or persistent infection of neuronal cells with C. pneumoniae in the AD brain may affect apoptosis in cells containing chlamydial inclusions. Results SK-N-MC neuroblastoma cells were infected with the respiratory strain of C. pneumoniae, AR39 at an MOI of 1. Following infection, the cells were either untreated or treated with staurosporine and then examined for apoptosis by labeling for nuclear fragmentation, caspase activity, and membrane inversion as indicated by annexin V staining. C. pneumoniae infection was maintained through 10 days post-infection. At 3 and 10 days post-infection, the infected cell cultures appeared to inhibit or were resistant to the apoptotic process when induced by staurosporine. This inhibition was demonstrated quantitatively by nuclear profile counts and caspase 3/7 activity measurements. Conclusion These data suggest that C. pneumoniae can sustain a chronic infection in neuronal cells by interfering with apoptosis, which may contribute to chronic inflammation in the AD brain.

  12. Apoptotic neuron-secreted HN12 inhibits cell apoptosis in Hirschsprung’s disease

    Science.gov (United States)

    Du, Chunxia; Xie, Hua; Zang, Rujin; Shen, Ziyang; Li, Hongxing; Chen, Pingfa; Xu, Xiaoqun; Xia, Yankai; Tang, Weibing

    2016-01-01

    Perturbation in apoptosis can lead to Hirschsprung’s disease (HSCR), which is a genetic disorder of neural crest development. It is believed that long noncoding RNAs (lncRNAs) play a role in the progression of HSCR. This study shows that apoptotic neurons can suppress apoptosis of nonapoptotic cells by secreting exosomes that contain high levels of HN12 lncRNA. Elevated exogenous HN12 in nonapoptotic cells effectively inhibited cell apoptosis by maintaining the function of mitochondria, including the production of ATP and the release of cytochrome C. These results demonstrate that secreted lncRNAs may serve as signaling molecules mediating intercellular communication in HSCR. In addition, high HN12 levels in the circulation worked as a biomarker for predicting HSCR, providing a potential, novel, noninvasive diagnostic approach for early screening of HSCR. PMID:27853370

  13. Effects of endoplasmic reticulum stress and related apoptosis on selective death of dopaminergic neurons

    Institute of Scientific and Technical Information of China (English)

    Lan Wang; Shenggang Sun; Xuebing Cao; Zhentao Zhang; Li Xu

    2006-01-01

    Objective: To explore the mechanism of endoplasmic reticulum stress (ERS) response and related apoptosis in dopaminergic neurons death. Methods: Nerve growth factor (NGF)-treatedPC12 cells were treated with 6-OHDA, MPP+ and rotenone. MTr assay and flow cytometry were used to measure the cell viability and the rate of celluar apoptosis induced by those neurotoxins. The expression of ERS-related gene XBP1, Grp78, CHOP, caspase-12 in drug-treated group and reserpine preincubation group was determined with RT-polymerase chain reaction (RT-PCR) and immunohistochemistry. Results: After the exposure to different toxins, the viability of PC12 cells were decreased by 52%, 44%, 40% at 100μM6-OHDA, 75 μM MPP+, 20 nM rotenone for 24 h respectively. FCM assay confirmed time-dependent cell apoptosis (P < 0.01 ). The gene and protein expression of XBP1, Grp78 in drug-treated group were significantly increased and reached their peaks 8 h after the treatment(P < 0.05).The expression levels of CHOP and caspase-12 gene were increased 16-24 h after the treatment(P < 0.01 ), but the expression level of caspase-12 was inhibited by reserpine preincubayion (P < 0.05). Conclusion: The excessive ERS and relative activated cell apoptosis pathway may be associated with selective death of dopaminergic neurons.

  14. Apoptotic neuron-secreted HN12 inhibits cell apoptosis in Hirschsprung's disease

    Directory of Open Access Journals (Sweden)

    Du C

    2016-11-01

    Full Text Available Chunxia Du,1,2,* Hua Xie,1,2,* Rujin Zang,1,2,* Ziyang Shen,1,2 Hongxing Li,1,2 Pingfa Chen,1,2 Xiaoqun Xu,1,2 Yankai Xia,2,3 Weibing Tang1,2 1Department of Pediatric Surgery, Nanjing Children’s Hospital Affiliated to Nanjing Medical University, 2State Key Laboratory of Reproductive Medicine, Institute of Toxicology, School of Public Health, 3Key Laboratory of Modern Toxicology, Ministry of Education, Nanjing Medical University, Nanjing, People’s Republic of China *These authors contributed equally to this work Abstract: Perturbation in apoptosis can lead to Hirschsprung’s disease (HSCR, which is a genetic disorder of neural crest development. It is believed that long noncoding RNAs (lncRNAs play a role in the progression of HSCR. This study shows that apoptotic neurons can suppress apoptosis of nonapoptotic cells by secreting exosomes that contain high levels of HN12 lncRNA. Elevated exogenous HN12 in nonapoptotic cells effectively inhibited cell apoptosis by maintaining the function of mitochondria, including the production of ATP and the release of cytochrome C. These results demonstrate that secreted lncRNAs may serve as signaling molecules mediating intercellular communication in HSCR. In addition, high HN12 levels in the circulation worked as a biomarker for predicting HSCR, providing a potential, novel, noninvasive diagnostic approach for early screening of HSCR. Keywords: Hirschsprung’s disease, neuronal development, exosomal long noncoding RNA, intercellular communication, apoptosis, mitochondria

  15. HIF-1 antagonizes p53-mediated apoptosis through a secreted neuronal tyrosinase.

    Science.gov (United States)

    Sendoel, Ataman; Kohler, Ines; Fellmann, Christof; Lowe, Scott W; Hengartner, Michael O

    2010-06-03

    Hypoxia-inducible factor (HIF) is a transcription factor that regulates fundamental cellular processes in response to changes in oxygen concentration. HIFalpha protein levels are increased in most solid tumours and correlate with patient prognosis. The link between HIF and apoptosis, a major determinant of cancer progression and treatment outcome, is poorly understood. Here we show that Caenorhabditis elegans HIF-1 protects against DNA-damage-induced germ cell apoptosis by antagonizing the function of CEP-1, the homologue of the tumour suppressor p53. The antiapoptotic property of HIF-1 is mediated by means of transcriptional upregulation of the tyrosinase family member TYR-2 in the ASJ sensory neurons. TYR-2 is secreted by ASJ sensory neurons to antagonize CEP-1-dependent germline apoptosis. Knock down of the TYR-2 homologue TRP2 (also called DCT) in human melanoma cells similarly increases apoptosis, indicating an evolutionarily conserved function. Our findings identify a novel link between hypoxia and programmed cell death, and provide a paradigm for HIF-1 dictating apoptotic cell fate at a distance.

  16. Fractalkine is a "find-me" signal released by neurons undergoing ethanol-induced apoptosis

    Directory of Open Access Journals (Sweden)

    Jennifer D Sokolowski

    2014-11-01

    Full Text Available Apoptotic neurons generated during normal brain development or secondary to pathologic insults are efficiently cleared from the central nervous system. Several soluble factors, including nucleotides, cytokines, and chemokines are released from injured neurons, signaling microglia to find and clear debris. One such chemokine that serves as a neuronal-microglial communication factor is fractalkine, with roles demonstrated in several models of adult neurological disorders. Lacking, however, are studies investigating roles for fractalkine in perinatal brain injury, an important clinical problem with no effective therapies. We used a well-characterized mouse model of ethanol-induced apoptosis to assess the role of fractalkine in neuronal-microglial signaling. Quantification of apoptotic debris in fractalkine-knockout and CX3CR1-knockout mice following ethanol treatment revealed increased apoptotic bodies compared to wild type mice. Ethanol-induced injury led to release of soluble, extracellular fractalkine. The extracellular media harvested from apoptotic brains induces microglial migration in a fractalkine-dependent manner that is prevented by neutralization of fractalkine with a blocking antibody or by deficiency in the receptor, CX3CR1. This suggests fractalkine acts as a ‘find-me’ signal, recruiting microglial processes toward apoptotic cells to promote their clearance. Next, we aimed to determine whether there are downstream alterations in cytokine gene expression due to fractalkine signaling. We examined mRNA expression in fractalkine-knockout and CX3CR1-knockout mice after alcohol-induced apoptosis and found differences in cytokine production in the brains of these knockouts by 6 hours after ethanol treatment. Collectively, this suggests that fractalkine acts as a ‘find me’ signal released by apoptotic neurons, and subsequently plays a critical role in modulating both phagocytic clearance and inflammatory cytokine gene expression after

  17. Mesenchymal stem cells protect neurons against hypoxic-ischemic injury via inhibiting parthanatos, necroptosis, and apoptosis, but not autophagy.

    Science.gov (United States)

    Kong, Deyan; Zhu, Juehua; Liu, Qian; Jiang, Yongjun; Xu, Lily; Luo, Ning; Zhao, Zhenqiang; Zhai, Qijin; Zhang, Hao; Zhu, Mingyue; Liu, Xinfeng

    2017-03-01

    Cellular therapy with mesenchymal stem cells (MSCs) protects cortical neurons against hypoxic-ischemic injury of stroke. Although sorts of efforts have been made to confirm the neuroprotective effect of MSCs on neurons against hypoxic-ischemic injury, the mechanism is until now far away from clear. Here in this study, oxygen-glucose deprivation (OGD)-injured neuron model was applied to mimic the neuronal hypoxic-ischemic injury in vitro. Co-culturing with MSCs in a transwell co-culture system, the OGD injured neurons were rescued by 75.0 %. Further data demonstrated that co-culturing with MSCs protected the cortical neurons from the OGD-induced parthanatos by alleviating apoptosis-inducing factor (AIF) nuclear translocation; attenuated the neuronal necroptosis by down-regulating the expression of the two essential kinases in necroptosis, receptor interacting protein kinase1 (RIP1) and 3 (RIP3); rescued the neurons from apoptosis by deactivating caspase-3; whilst performed no significant influence on OGD-induced neuronal autophagy, according to its failed regulation on Beclin1. In conclusion, MSCs potentially protect the cortical neurons from OGD-injury in vitro, through rescuing neurons from the cell death of parthanatos, necroptosis, and apoptosis, but not autophagy, which could provide some evidence to the mechanism explanation on stem cell treatment for ischemic stroke.

  18. GnRH analogue attenuated apoptosis of rat hippocampal neuron after ischemia-reperfusion injury.

    Science.gov (United States)

    Chu, Chenyu; Xu, Bainan; Huang, Weiquan

    2010-12-01

    The expression and new functions of reproductive hormones in organs beyond hypothalamus-pituitary-gonad axis have been reported. So far, there is no report about the protective effects of GnRH analogue to hippocampal neurons suffering from ischemia-reperfusion injury. Middle cerebral artery occlusion model together with TUNEL staining were made in vivo and oxygen-glucose deprivation model together with double staining of Annexin V/PI with flow cytometer were made in vitro to observe the anti-apoptotic effects of GnRH analogue to hippocampal neurons after ischemia-reperfusion injury. The results found that the number of TUNEL positive pyramidal neurons in CA1 region in GnRH analogue experiment group was less than that in control group in vivo; the percentage of apoptotic neurons in GnRH analogue experiment group was less than that in control group in vitro. These findings suggested that pretreatment with certain concentration of GnRH analogue could attenuate apoptosis of hippocampal neurons. GnRH analogue has the protective effects to neurons.

  19. Investigation on apoptosis of neuronal cells induced by Amyloid beta-Protein

    Institute of Scientific and Technical Information of China (English)

    罗本燕; 徐增斌; 陈智; 陈峰; 唐敏

    2004-01-01

    Objective: To construct a PC12 cell strain with neuronal differentiation, and observe the apoptosis and pro-liferation activity effects induced these cells by Amyloid beta-Protein (Aβ3-43). Methods: 1) PC12 cells in logarithmic growth phase were subcultured for 24 h. After the culture fluid was changed, the cells were treated with Rat-β-NGF and cultured for 9 days. 2) Neuronal differentiation of PC 12 cells in logarithmic growth phase were divided into four groups:control group (0), experimental group (1), experimental group (2) and experimental group (3). The concentrations of Aβ in the four groups were 0 μmol/L, 1.25 μmol/L, 2.5 μmol/L and 5 μmol/L, respectively. The cells were harvested at 24, 48 and 72 h later and stained with AnnexinV-FITC/PI after centrifugation and washing. Then flow cytometry was conducted to examine the apoptosis percentage. 3) NGF-induced PC12 cells were selected and Aβ with different concentrations was added. The final concentrations of Aβ were 0 μmol/L, 1.25 μmol/L, 2.5 μmol/L and 5 μmol/L, respectively. After the cells were incubated in an atmosphere of 5% CO2 at 37 ℃ in an incubator for 72 h, the OD values were examined. Results: 1)Neuronal differentiated PC12 cell lines were successfully established. 2) Flow cytometric examination indicated that Aβ(1.25, 2.5, and 5.0 μmol/L) could effectively induce apoptosis of neuronal-differented cells at the 24 h, 48 h and 72 h time points. 3) Aβ (0-5.00 μmol/L) had no obvious effect on proliferation or restraining of the neuronal differentiation of the PC 12 cells after a 72 h interacting process. Conclusion: This investigation revealed successful neuronal differentiation of the PC12 cell strain. The induction of apoptosis of the neurocytes by various concentrations of Aβ was observed and the in-fluence of Aβ on induced proliferation of PC 12 cells by Rat-β-NGF was revealed. This study may provide basis for future research on the molecular cure of AD and interdiction of AD

  20. Metallothionein as a useful marker in Hodgkin lymphoma subclassification

    DEFF Research Database (Denmark)

    Penkowa, Milena; Sørensen, Brit Ladegaard; Nielsen, Signe Lidou

    2009-01-01

    Metallothionein (MT) expression is considered to be a prognostic factor that promotes tumor resistance to apoptosis. In non-Hodgkin lymphomas, MT is differentially expressed and constitutes a risk factor. We have characterised MT in lymph nodes of Hodgkin lymphoma (HL) [patients with nodular...

  1. Full Length Bid is sufficient to induce apoptosis of cultured rat hippocampal neurons

    Directory of Open Access Journals (Sweden)

    Ward Manus W

    2007-02-01

    Full Text Available Abstract Background Bcl-2 homology domain (BH 3-only proteins are pro-apoptotic proteins of the Bcl-2 family that couple stress signals to the mitochondrial cell death pathways. The BH3-only protein Bid can be activated in response to death receptor activation via caspase 8-mediated cleavage into a truncated protein (tBid, which subsequently translocates to mitochondria and induces the release of cytochrome-C. Using a single-cell imaging approach of Bid cleavage and translocation during apoptosis, we have recently demonstrated that, in contrast to death receptor-induced apoptosis, caspase-independent excitotoxic apoptosis involves a translocation of full length Bid (FL-Bid from the cytosol to mitochondria. We induced a delayed excitotoxic cell death in cultured rat hippocampal neurons by a 5-min exposure to the glutamate receptor agonist N-methyl-D-aspartate (NMDA; 300 μM. Results Western blot experiments confirmed a translocation of FL-Bid to the mitochondria during excitotoxic apoptosis that was associated with the release of cytochrome-C from mitochondria. These results were confirmed by immunofluorescence analysis of Bid translocation during excitotoxic cell death using an antibody raised against the amino acids 1–58 of mouse Bid that is not able to detect tBid. Finally, inducible overexpression of FL-Bid or a Bid mutant that can not be cleaved by caspase-8 was sufficient to induce apoptosis in the hippocampal neuron cultures. Conclusion Our data suggest that translocation of FL-Bid is sufficient for the activation of mitochondrial cell death pathways in response to glutamate receptor overactivation.

  2. Zinc Deficiency Induces Apoptosis via Mitochondrial p53- and Caspase-Dependent Pathways in Human Neuronal Precursor Cells

    Science.gov (United States)

    Seth, Rohit; Corniola, Rikki S.; Gower-Winter, Shannon D.; Morgan, Thomas J., Jr.; Bishop, Brian; Levenson, Cathy W.

    2015-01-01

    Previous studies have shown that zinc deficiency leads to apoptosis of neuronal precursor cells in vivo and in vitro. In addition to the role of p53 as a nuclear transcription factor in zinc deficient cultured human neuronal precursors (NT-2), we have now identified the translocation of phosphorylated p53 to the mitochondria and p53-dependent…

  3. Effects of melatonin on streptozotocin-induced retina neuronal apoptosis in high blood glucose rat.

    Science.gov (United States)

    Li, Xiaoyan; Zhang, Maonian; Tang, Weiqiang

    2013-03-01

    One of the main pathological symptoms of early diabetic retinal neuropathy is retina neuronal apoptosis. In the present work we investigated the effects of indoleamine hormone melatonin, a powerful free radical scavenger, on streptozotocin-induced retina neuronal cell apoptosis in high blood glucose rat. After melatonin treatment (10 mg/kg/day), tunel detection was used to monitor the apoptosis rate of neurons in the retinal ganglion cell layer; reversed quantitative PCR was used to measure the mRNA expression of retinal caspase-3, Mn superoxidase dismutase (SOD) and Cu-Zn SOD; and the activities of total SOD (T-SOD) and sub-type SOD was detected using xanthine oxidase enzymatic detection. Our data showed that melatonin treatment leads to a decrease of retinal cell apoptosis and the apoptotic index was (1.67 ± 0.54) % and (7.73 ± 0.95) % at 8 and 12 weeks after treatment. The relative quantitative (RQ) value for caspase-3 mRNA expression was (6.996 ± 1.192) and (7.267 ± 1.178) in melatonin group, which are much lower than the values of diabetic group (12.566 ± 2.272 and (14.297 ± 2.110) at 8 and 12 weeks, respectively) under the same condition. mRNA expression of Mn SOD and Cu-Zn SOD as well as their activities all decreased in the diabetic group compared with the control group. While melatonin treatment induced the expression of Mn SOD mRNA and a continual increase of Mn SOD activity as well as the activity and mRNA expression of Cu-Zn SOD at 12 weeks. Therefore, our results demonstrate that melatonin treatment prevented the decrease in mRNA expression of SOD and the increase in caspase-3 mRNA expression induced by diabetes thus exerts a beneficial effect on retina neuronal apoptosis.

  4. Aluminum—induced apoptosis in cultured cortical neurons and its effects on SAPK/JNK signal transduction pathway

    Institute of Scientific and Technical Information of China (English)

    FuHJ; DongSZ

    2002-01-01

    Aluminum (Al) exposure and apoptotic cell death have been implicated in several neurodegenerative diseases.the mechanisms by which Al interacts with the nervous system are only partly understood.In this study,we used cultured cortical neurons to investigate the ability of Al to induce the apoptosis of neurons and to explore the role of SAPK/JNK signal transduction pathway on the apoptosis induced by Al.It was found that Al-induced degeneration of cortical neurons involved the DNA fragmentation characteristic of apoptosis.The rate of apoptosis increased significantly,which was measured by TdT-mediated dUTKP nick end labeling.Westerm blot analysis showed that SAPK/JNK activities of cortical neurons varied when the dose and exposure time of AlCl3 were different.Our study demonstrates that Al can induce the apoptosis of cortical neurons and SAPK/JNK signal transduction pathway may play a great role in the apoptosis.

  5. The Fas/Fas ligand death receptor pathway contributes to phenylalanine-induced apoptosis in cortical neurons.

    Directory of Open Access Journals (Sweden)

    Xiaodong Huang

    Full Text Available Phenylketonuria (PKU, an autosomal recessive disorder of amino acid metabolism caused by mutations in the phenylalanine hydroxylase (PAH gene, leads to childhood mental retardation by exposing neurons to cytotoxic levels of phenylalanine (Phe. A recent study showed that the mitochondria-mediated (intrinsic apoptotic pathway is involved in Phe-induced apoptosis in cultured cortical neurons, but it is not known if the death receptor (extrinsic apoptotic pathway and endoplasmic reticulum (ER stress-associated apoptosis also contribute to neurodegeneration in PKU. To answer this question, we used specific inhibitors to block each apoptotic pathway in cortical neurons under neurotoxic levels of Phe. The caspase-8 inhibitor Z-IETD-FMK strongly attenuated apoptosis in Phe-treated neurons (0.9 mM, 18 h, suggesting involvement of the Fas receptor (FasR-mediated cell death receptor pathway in Phe toxicity. In addition, Phe significantly increased cell surface Fas expression and formation of the Fas/FasL complex. Blocking Fas/FasL signaling using an anti-Fas antibody markedly inhibited apoptosis caused by Phe. In contrast, blocking the ER stress-induced cell death pathway with salubrinal had no effect on apoptosis in Phe-treated cortical neurons. These experiments demonstrate that the Fas death receptor pathway contributes to Phe-induced apoptosis and suggest that inhibition of the death receptor pathway may be a novel target for neuroprotection in PKU patients.

  6. Apoptosis of motor neurons in the spinal cord after ischemia reperfusion injury delayed paraplegia in rabbits

    Institute of Scientific and Technical Information of China (English)

    Liu Bibo; Liu Miao; Ma Wei; Wang Duoning

    2007-01-01

    Objective To clarify the pathologic change of the motor neuron on spinal cord ischemia reperfusion injury delayed paraplegia. Methods The infrarenal aorta of White New Zealand rabbits (n=24) was occluded for 26 minutes using two bulldog clamps. Rabbits were killed after 8, 24, 72, or 168 hours (n=6 per group), respectively. The clamps was placed but never clamped in sham-operated rabbits (n=24). The lumbar segment of the spinal cord (L5 to L7) was used for morphological studies, including hematoxylin and eosin staining, the expression of bcl-2 and bax proteins in spinal cord was detected with immunohistochemistry. The apoptotic neurons in spinal cord were measured with terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end-labeling of DNA fragments (TUNEL) staining. Results Delayed paraplegia occurred in all rabbits of ischemia reperfusion group at 16-24 hours, but not in sham groups. Motor neurons were selectively lost at 7 days after transient ischemia. After ischemia, the positive expression of bcl-2 protein were in the sham controls but decreased significantly as compared with that of the IR group (P<0.01), especially in 72 hours reperfusion. The positive expression of bax protein were also in the sham controls, but increased in the IR group, especially in 72 hours reperfusion; In addition, TUNEL study demonstrated that no cells were positively labeled until 24 hours after ischemia, but nuclei of some motor neurons were positively labeled at peak after ischemia reperfusion at 72 hours. Conclusion Spinal cord ischemia in rabbits induces morphological and biochemical changes suggestive of apoptosis. These data raise the possibility that apoptosis contributes to neuronal cell death after spinal cord ischemia reperfusion.

  7. Ketamine induces toxicity in human neurons differentiated from embryonic stem cells via mitochondrial apoptosis pathway

    Science.gov (United States)

    Bosnjak, Zeljko J.; Yan, Yasheng; Canfield, Scott; Muravyeva, Maria Y.; Kikuchi, Chika; Wells, Clive; Corbett, John; Bai, Xiaowen

    2013-01-01

    Ketamine is widely used for anesthesia in pediatric patients. Growing evidence indicates that ketamine causes neurotoxicity in a variety of developing animal models. Our understanding of anesthesia neurotoxicity in humans is currently limited by difficulties in obtaining neurons and performing developmental toxicity studies in fetal and pediatric populations. It may be possible to overcome these challenges by obtaining neurons from human embryonic stem cells (hESCs) in vitro. hESCs are able to replicate indefinitely and differentiate into every cell type. In this study, we investigated the toxic effect of ketamine on neurons differentiated from hESCs. Two-week-old neurons were treated with different doses and durations of ketamine with or without the reactive oxygen species (ROS) scavenger, Trolox. Cell viability, ultrastructure, mitochondrial membrane potential (ΔΨm), cytochrome c distribution within cells, apoptosis, and ROS production were evaluated. Here we show that ketamine induced ultrastructural abnormalities and dose- and time-dependently caused cell death. In addition, ketamine decreased ΔΨm and increased cytochrome c release from mitochondria. Ketamine also increased ROS production and induced differential expression of oxidative stress-related genes. Specifically, abnormal ultrastructural and ΔΨm changes occurred earlier than cell death in the ketamine-induced toxicity process. Furthermore, Trolox significantly decreased ROS generation and attenuated cell death caused by ketamine in a dose-dependent manner. In conclusion, this study illustrates that ketamine time- and dose-dependently induces human neurotoxicity via ROS-mediated mitochondrial apoptosis pathway and that these side effects can be prevented by the antioxidant agent Trolox. Thus, hESC-derived neurons might provide a promising tool for studying anesthetic-induced developmental neurotoxicity and prevention strategies. PMID:22873495

  8. 5-Hydroxymethylfurfural from wine-processed Fructus corni inhibits hippocampal neuron apoptosis***

    Institute of Scientific and Technical Information of China (English)

    Hai Gu; Zequn Jiang; Mingyan Wang; Haiying Jiang; Fengming Zhao; Xia Ding; Baochang Cai; Zhen Zhan

    2013-01-01

    Previous studies have shown that 5-hydroxymethylfurfural, a compound extracted from wine- pro-cessed Fructus corni, has a protective effect on hippocampal neurons. The present study was de-signed to explore the related mechanisms. Our study revealed that high and medium doses (10, 1μmol/L) of 5-hydroxymethylfurfural could improve the morphology of H2O2-treated rat hippocampal neurons as revealed by inverted phase-contrast microscopy and transmission electron microscopy. MTT results showed that incubation with high and medium doses of 5-hydroxymethylfurfural caused a significant increase in the viability of neuronal cells injured by H2O2. Flow cytometry assays con-firmed that H2O2 could induce cellapoptosis, while high and medium doses of 5-hydroxymethylfurfural had a visible protective effect on apoptotic rat hippocampal neurons. Re-al-time PCR and western blot analysis showed that high and medium doses of 5-hydroxymethylfurfural prevented H2O2-induced up-regulation of p53, Bax and caspase-3 and antagonized the down-regulation of Bcl-2 induced by H2O2 treatment. These results suggested that 5-hydroxymethylfurfural could inhibit apoptosis of cultured rat hippocampal neurons injured by H2O2 via increase in Bcl-2 levels and decrease in p53, Bax and caspase-3 protein expression lev-els.

  9. Use of RNAi silencing to target preconditioned glial cell line-derived neurotrophic factor in neuronal apoptosis

    Institute of Scientific and Technical Information of China (English)

    Hongliang Guo; Zhongxin Xu; Xinhua Li; Jing Mang; Ying Xing; Jinting He; Guihua Xu; Shijun Yan; Lifeng Liu; Chunli Mei

    2011-01-01

    Several studies have suggested that exogenous glial cell line-derived neurotrophic factor may protect neurons from cerebral ischemic injury. However, the mechanisms underlying the neuroprotective effects of endogenous glial cell line-derived neurotrophic factor remain unclear. The present experiments sought to elucidate the influence of various conditioned media on neuronal apoptosis, using a normal culture medium for astrocytes, an astrocyte medium highly expressing glial cell line-derived neurotrophic factor, and an astrocyte medium in which glial cell line-derived neurotrophic factor expression was silenced using RNAi technology. The results confirmed that the use of RNAi silencing to target pretreated glial cell line-derived neurotrophic factor expression promoted neuronal apoptosis. In addition, oxygen and glucose deprivation preconditioning was found to upregulate glial cell line-derived neurotrophic factor expression, and significantly reduce neuronal apoptosis.

  10. Activin A prevents neuron-like PC12 cell apoptosis after oxygen-glucose deprivation

    Institute of Scientific and Technical Information of China (English)

    Guihua Xu; Zhongxin Xu; Jinting He; Hongliang Guo; Chunli Mei; Jiaoqi Wang; Zhongshu Li; Han Chen; Jing Mang; Hong Yang

    2013-01-01

    In this study, PC12 cells were induced to differentiate into neuron-like cells using nerve growth factor, and were subjected to oxygen-glucose deprivation. Cells were treated with 0, 10, 20, 30, 50, 100 ng/mL exogenous Activin A. The 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium bromide assay and Hoechst 33324 staining showed that the survival percentage of PC12 cells significantly decreased and the rate of apoptosis significantly increased after oxygen-glucose deprivation. Exogenous Activin A significantly increased the survival percentage of PC12 cells in a dose-dependent manner. Reverse transcription-PCR results revealed a significant increase in Activin receptor IIA, Smad3 and Smad4 mRNA levels, which are key sites in the Activin A/Smads signaling pathway, in neuron-like cells subjected to oxygen-glucose deprivation, while mRNA expression of the apoptosis-regulation gene caspase-3 decreased. Our experimental findings indicate that exogenous Activin A plays an anti-apoptotic role and protects neurons by means of activating the Activin A/Smads signaling pathway.

  11. Effect of Metallothionein on Cell Cycle, Apoptosis Rate and Subsets Distribution of Lymphocytes in Peripheral Blood of Dairy Cattle under Heat Stress

    Institute of Scientific and Technical Information of China (English)

    Jiajie LUO; Jun FANG; Lili LI; Bin ZHANG; LiZhuan WU; Zijun LI; Ying PENG; JueXin FAN; XinYi LAN; JinShun ZHAN

    2013-01-01

    [Objective] This study aimed to research the effect of metal othionein on cellcycle, apoptosis rate and subsets distribution of lymphocytes in peripheral blood of dairy cows under heat stress, so as to perfect the regulative mechanism re-searches of MT to anti-heat stress. [Method] Twenty lactating Chinese Holstein cows were randomly divided into four groups (A, B, C and D), and injected with 0, 4.0, 8.0 and 12.0 mg Zn-metal othionein, respectively by intravenous route. Blood sam-ples were col ected at 1st, 16th, 31st, 46th and 61st day, and the dynamic changes of cellcycle, apoptosis rate and subsets distribution of lymphocytes were determined. [Result] The apoptosis rate of cells in group B and C was lower than those in group A by 26.63% (P>0.05) and 24.84% (P>0.05) respectively. The number of cells in the G0/G1 phage in trial groups was increased and the number of cells in the S and G2/M phages tended to decrease, but there were no significant differences (P>0.05). The number of CD3+ T cellin three trial groups was greater than those in group A by 7.02% (P>0.05), 5.45% (P>0.05) and 3.85% (P>0.05) respectively, while the number of CD4+ T cellin trial groups was higher than those in control group by 31.04% (P<0.05), 35.68% (P<0.05) and 39.34% (P<0.05) respectively. The number of CD8+ T celland the levels of CD4+/CD8+ in trial groups were increased observ-ably, but significant difference (P<0.05) was observed in the levels of CD4+/CD8+between groups A and C only. It demonstrated that exogenous Zn-metal othionein can decrease apoptosis rate, improve cellcycle and regulate subsets distribution of lymphocytes in dairy cattle in a dose-dependent manner. [Conclusion] This study wil provide scientific basis for safe utilization of MT in dairy industry.

  12. Effects of topiramate on hippocampal neuronal apoptosis in rats after kainic acid-evoked seizures

    Institute of Scientific and Technical Information of China (English)

    Yuan Wu; Jiarong Pang; Jinou Zheng; Xiaoqing Deng; Xiulin Liang; Jiaquan Li; Zhiying Chen

    2008-01-01

    BACKGROUND:Apoptosis plays an important role in brain injury after seizures and the formation of chronic epilepsy.It is important to investigate whether topiramate exhibits either antiepileptic and/or anti-apoptotic effects on hippocampal neurons.OBJECTIVE:To observe euronal apoptosis in hippocampus of rat seizure models,and to investigate the antagonizing effect of topiramate on neuronal apoptosis after seizures.DESIGN:An animal experiment of comparative observation.SETTING:First Affiliated Hospital of Guangxi Medical University.MATERIALS:Sixty healthy male Sprague Dawley(SD)rats,4-6 weeks old and weighing 160-220 g,were provided by the Experimental Animal Center of Guangxi Medical University.Main apparatus and reagents were as follows:Rat brain solid positioner(SR-6N,made in Japan); kainic acid by Sigma(USA);pathological image analyzer(DMR+550)by Leica(Germany); in situ apoptosis detection kit by Wuhan Boster Biological Technology Co.,Ltd; topiramate by Xi'an-Janssen Pharmaceutical,Ltd.The treatment on animals in the experiment was in accordance with the standards of animal ethics.METHODS:The experiments were performed at the Scientific Experimental Center of Guangxi Medical University from June to December 2006.The rats were randomly divided into a topiramate-treated group(n=30)and a model group(n=30).① After anesthesia,all rats were administered a kainic acid injection(0.2 μ L,2 g/L)into the right lateral ventricle.Grade Ⅲ and greater Racine standards were considered to be a successful model establishment.Thirty minutes after seizure ,rats in the topiramate-treated group were treated with an intraperitoneal(i.p.)injection of topiramate every day(40 mg/kg/d)for 2 weeks.The rats in the model group were treated with an equal volume of saline for 2 weeks.③Six rats in the topiramate-treated group were sacrificed at 1 day,and 1,2,3,and 4 weeks after treatment,respectively.The model group animals were sacrificed at corresponding time points.The brain

  13. Dysregulation of macrophage-secreted cathepsin B contributes to HIV-1-linked neuronal apoptosis.

    Directory of Open Access Journals (Sweden)

    Eillen J Rodriguez-Franco

    Full Text Available Chronic HIV infection leads to the development of cognitive impairments, designated as HIV-associated neurocognitive disorders (HAND. The secretion of soluble neurotoxic factors by HIV-infected macrophages plays a central role in the neuronal dysfunction and cell death associated with HAND. One potentially neurotoxic protein secreted by HIV-1 infected macrophages is cathepsin B. To explore the potential role of cathepsin B in neuronal cell death after HIV infection, we cultured HIV-1(ADA infected human monocyte-derived macrophages (MDM and assayed them for expression and activity of cathepsin B and its inhibitors, cystatins B and C. The neurotoxic activity of the secreted cathepsin B was determined by incubating cells from the neuronal cell line SK-N-SH with MDM conditioned media (MCM from HIV-1 infected cultures. We found that HIV-1 infected MDM secreted significantly higher levels of cathepsin B than did uninfected cells. Moreover, the activity of secreted cathepsin B was significantly increased in HIV-infected MDM at the peak of viral production. Incubation of neuronal cells with supernatants from HIV-infected MDM resulted in a significant increase in the numbers of apoptotic neurons, and this increase was reversed by the addition of either the cathepsin B inhibitor CA-074 or a monoclonal antibody to cathepsin B. In situ proximity ligation assays indicated that the increased neurotoxic activity of the cathepsin B secreted by HIV-infected MDM resulted from decreased interactions between the enzyme and its inhibitors, cystatins B and C. Furthermore, preliminary in vivo studies of human post-mortem brain tissue suggested an upregulation of cathepsin B immunoreactivity in the hippocampus and basal ganglia in individuals with HAND. Our results demonstrate that HIV-1 infection upregulates cathepsin B in macrophages, increases cathepsin B activity, and reduces cystatin-cathepsin interactions, contributing to neuronal apoptosis. These findings

  14. The effects of piracetam on heroin-induced CPP and neuronal apoptosis in rats.

    Science.gov (United States)

    Xu, Peng; Li, Min; Bai, Yanping; Lu, Wei; Ling, Xiaomei; Li, Weidong

    2015-05-01

    Piracetam is a positive allosteric modulator of the AMPA receptor that has been used in the treatment of cognitive disorders for decades. Recent surveys and drug analyses have demonstrated that a heroin mixture adulterated with piracetam has spread rapidly in heroin addicts in China, but its addictive properties and the damage it causes to the central neural system are currently unknown. The effect of piracetam on the reward properties of heroin was assessed by conditioned place preference (CPP). Electron microscopy and radioimmunoassay were used to compare the effects of heroin mixed with equivalent piracetam (HP) and heroin alone on neuronal apoptosis and the levels of beta-endorphin (β-EP) in different brain subregions within the corticolimbic system, respectively. Piracetam significantly enhanced heroin-induced CPP expression while piracetam itself didn't induce CPP. Morphological observations showed that HP-treated rats had less neuronal apoptosis than heroin-treated group. Interestingly, HP normalized the levels of β-EP in the medial prefrontal cortex (mPFC) and core of the nucleus accumbens (AcbC) subregions, in where heroin-treated rats showed decreased levels of β-EP. These results indicate that piracetam potentiate the heroin-induced CPP and protect neurons from heroin-induced apoptosis. The protective role of HP might be related to the restoration of β-EP levels by piracetam. Our findings may provide a potential interpretation for the growing trend of HP abuse in addicts in China. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  15. Phospho-Rb mediating cell cycle reentry induces early apoptosis following oxygen-glucose deprivation in rat cortical neurons.

    Science.gov (United States)

    Yu, Ying; Ren, Qing-Guo; Zhang, Zhao-Hui; Zhou, Ke; Yu, Zhi-Yuan; Luo, Xiang; Wang, Wei

    2012-03-01

    The aim of this study was to investigate the relationship between cell cycle reentry and apoptosis in cultured cortical neurons following oxygen-glucose deprivation (OGD). We found that the percentage of neurons with BrdU uptake, TUNEL staining, and colocalized BrdU uptake and TUNEL staining was increased relative to control 6, 12 and 24 h after 1 h of OGD. The number of neurons with colocalized BrdU and TUNEL staining was decreased relative to the number of TUNEL-positive neurons at 24 h. The expression of phosphorylated retinoblastoma protein (phospho-Rb) was significantly increased 6, 12 and 24 h after OGD, parallel with the changes in BrdU uptake. Phospho-Rb and TUNEL staining were colocalized in neurons 6 and 12 h after OGD. This colocalization was strikingly decreased 24 h after OGD. Treatment with the cyclin-dependent kinase inhibitor roscovitine (100 μM) decreased the expression of phospho-Rb and reduced neuronal apoptosis in vitro. These results demonstrated that attempted cell cycle reentry with phosphorylation of Rb induce early apoptosis in neurons after OGD and there must be other mechanisms involved in the later stages of neuronal apoptosis besides cell cycle reentry. Phosphoralated Rb may be an important factor which closely associates aberrant cell cycle reentry with the early stages of neuronal apoptosis following ischemia/hypoxia in vitro, and pharmacological interventions for neuroprotection may be useful directed at this keypoint.

  16. Oxidative Stress Associated with Neuronal Apoptosis in Experimental Models of Epilepsy

    Directory of Open Access Journals (Sweden)

    Marisela Méndez-Armenta

    2014-01-01

    Full Text Available Epilepsy is considered one of the most common neurological disorders worldwide. Oxidative stress produced by free radicals may play a role in the initiation and progression of epilepsy; the changes in the mitochondrial and the oxidative stress state can lead mechanism associated with neuronal death pathway. Bioenergetics state failure and impaired mitochondrial function include excessive free radical production with impaired synthesis of antioxidants. This review summarizes evidence that suggest what is the role of oxidative stress on induction of apoptosis in experimental models of epilepsy.

  17. Recombinant human erythropoietin increases survival and reduces neuronal apoptosis in a murine model of cerebral malaria

    DEFF Research Database (Denmark)

    Wiese, Lothar; Hempel, Casper; Penkowa, Milena;

    2008-01-01

    with recombinant human Epo (rhEpo; 50-5000 U/kg/OD, i.p.) at different time points. The effect on survival was measured. Brain pathology was investigated by TUNEL (Terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP)-digoxigenin nick end labelling), as a marker of apoptosis. Gene...... expression in brain tissue was measured by real time PCR. RESULTS: Treatment with rhEpo increased survival in mice with CM in a dose- and time-dependent manner and reduced apoptotic cell death of neurons as well as the expression of pro-inflammatory cytokines in the brain. This neuroprotective effect...

  18. Neuroprotective Effects of Kukoamine a against Radiation-induced Rat Brain Injury through Inhibition of Oxidative Stress and Neuronal Apoptosis.

    Science.gov (United States)

    Zhang, Yaqiong; Cheng, Zhihua; Wang, Changli; Ma, Hongda; Meng, Weihong; Zhao, Qingchun

    2016-10-01

    Radiation-induced brain injury (RIBI) is a prominent side effect of radiotherapy for cranial tumors. Kukoamine A (KuA) has the ability of anti-oxidative stress and anti-apoptosis in vitro. The aim of this study was to investigate whether KuA would prevent the detrimental effect of ionizing radiation on hippocampal neurons. For this study, male Wistar rats were received either sham irradiation or whole brain irradiation (30 Gy single dose of X-rays) followed by the immediate injection of either KuA or vehicle intravenously. The dose of KuA was 5, 10 and 20 mg/kg respectively. The protective effects of KuA were assessed by Nissl staining. The levels of oxidative stress marker and antioxidants activities were assayed by kits. TUNEL staining was performed to detect the level of apoptosis in hippocampal neurons. The expression of apoptosis-related proteins as well as the brain-derived neurophic factor (BDNF) was evaluated by western blot. Whole brain irradiation led to the neuronal abnormality and it was alleviated by KuA. KuA decreased malondialdehyde (MDA) level, increased glutathione (GSH) level, superoxide dismutase (SOD) and catalase (CAT) activities, as well as alleviated neuronal apoptosis by regulating the expression of cleaved caspase-3, cytochrome C, Bax and Bcl-2. Additionally, KuA increased the expression of BDNF. These data indicate that KuA has neuroprotective effects against RIBI through inhibiting neuronal oxidative stress and apoptosis.

  19. Influence of acute ethanol intoxication on neuronal apoptosis and Bcl-2 protein expression after severe traumatic brain injury in rats

    Institute of Scientific and Technical Information of China (English)

    HE Min; LIU Wei-guo; WEN Liang; DU Hang-gen; YIN Li-chun; CHEN Li

    2013-01-01

    To study the influence and mechanism of acute ethanol intoxication (AEI) on rat neuronal apoptosis after severe traumatic brain injury (TBI).Methods:Ninety-six Sprague-Dawley rats were randomly divided into four groups:normal control,AEI-only,TBI-only and TBI+AEI (n=24 for each).Severe TBI model was developed according to Feeney's method.Rats in TBI+AEI group were firstly subjected to AEI,and then suffered head trauma.In each group,animals were sacrificed at 6 h,24 h,72 h,and 168 h after TBI.The level of neuronal apoptosis and the expression of Bcl-2 protein were determined by TUNEL assay and immunohistochemical method,respectively.Results:Apoptotic cells mainly distributed in the cortex and white matter around the damaged area.Neuronal apoptosis significantly increased at 6 h after trauma and peaked at 72 h.Both the level of neuronal apoptosis and expression of Bcl-2 protein in TBI-only group and TBI+AEI group were higher than those in control group (P<0.05).Compared with TBI-only group,the two indexes were much higher in TBI+AEI group at all time points (P<0.05).Conclusion:Our findings suggest that AEI can increase neuronal apoptosis after severe TBI.

  20. Apoptosis of supraoptic AVP neurons is involved in the development of central diabetes insipidus after hypophysectomy in rats

    Directory of Open Access Journals (Sweden)

    Huang Lijin

    2008-06-01

    Full Text Available Abstract Background It has been reported that various types of axonal injury of hypothalamo-neurohypophyseal tract can result in degeneration of the magnocellular neurons (MCNs in hypothalamus and development of central diabetes insipidus (CDI. However, the mechanism of the degeneration and death of MCNs after hypophysectomy in vivo is still unclear. This present study was aimed to disclose it and to figure out the dynamic change of central diabetes insipidus after hypophysectomy. Results The analysis on the dynamic change of daily water consumption (DWC, daily urine volume(DUV, specific gravity of urine(USG and plasma vasopressin concentration showed that the change pattern of them was triphasic and neuron counting showed that the degeneration of vasopressin neurons began at 10 d, aggravated at 20 d and then stabilized at 30 d after hypophysectomy. There was marked upregulation of cleaved Caspase-3 expression of vasopressin neurons in hypophysectomy rats. A "ladder" pattern of migration of DNA internucleosomal fragments was detected and apoptotic ultrastructure was found in these neurons. There was time correlation among the occurrence of diabetes insipidus, the changes of plasma vasopressin concentration and the degeneration of vasopressin neurons after hypophysectomy. Conclusion This study firstly demonstrated that apoptosis was involved in degeneration of supraoptic vasopressin neurons after hypophysectomy in vivo and development of CDI. Our study on time course and correlations among water metabolism, degeneration and apoptosis of vasopressin neurons suggested that there should be an efficient therapeutic window in which irreversible CDI might be prevented by anti-apoptosis.

  1. Neurosteroid dehydroepiandrosterone interacts with nerve growth factor (NGF receptors, preventing neuronal apoptosis.

    Directory of Open Access Journals (Sweden)

    Iakovos Lazaridis

    2011-04-01

    Full Text Available The neurosteroid dehydroepiandrosterone (DHEA, produced by neurons and glia, affects multiple processes in the brain, including neuronal survival and neurogenesis during development and in aging. We provide evidence that DHEA interacts with pro-survival TrkA and pro-death p75(NTR membrane receptors of neurotrophin nerve growth factor (NGF, acting as a neurotrophic factor: (1 the anti-apoptotic effects of DHEA were reversed by siRNA against TrkA or by a specific TrkA inhibitor; (2 [(3H]-DHEA binding assays showed that it bound to membranes isolated from HEK293 cells transfected with the cDNAs of TrkA and p75(NTR receptors (K(D: 7.4 ± 1.75 nM and 5.6 ± 0.55 nM, respectively; (3 immobilized DHEA pulled down recombinant and naturally expressed TrkA and p75(NTR receptors; (4 DHEA induced TrkA phosphorylation and NGF receptor-mediated signaling; Shc, Akt, and ERK1/2 kinases down-stream to TrkA receptors and TRAF6, RIP2, and RhoGDI interactors of p75(NTR receptors; and (5 DHEA rescued from apoptosis TrkA receptor positive sensory neurons of dorsal root ganglia in NGF null embryos and compensated NGF in rescuing from apoptosis NGF receptor positive sympathetic neurons of embryonic superior cervical ganglia. Phylogenetic findings on the evolution of neurotrophins, their receptors, and CYP17, the enzyme responsible for DHEA biosynthesis, combined with our data support the hypothesis that DHEA served as a phylogenetically ancient neurotrophic factor.

  2. Neuroprotective effect of melatonin against ischemia/reperfusion-induced neuronal apoptosis in mouse cerebellum

    Institute of Scientific and Technical Information of China (English)

    Qiuhong Duan; Tao Lu; Yixiang Han; Zhiqiang Lu; Ximing Wang

    2007-01-01

    BACKGROUND: Some experiments have demonstrated that melatonin (N-aceyl-5-methoxytryptamine, Mel) has antioxidation. However, whether it has neuroprotective effect in the ischemia/reperfusion injury of central nervous system is unclear.OBJECTIVE: To observe the protective effect of Mel on ischemia/reperfusion-induced cerebellar neuronal apoptosis of rats, and the action mechanism. DESIGN: Controlled observation experiment.SETTING: Department of Biochemistry and Molecular Biology, Tongji Medical College, Huazhong University of Science and Technology.MATERIALS: Eight Sprague-Dawley rats aged 7-8 days and weighing 10-12 g were provided by Medical Experimental Animal Center, Tongji Medical College, Huazhong University of Science and Technology. Anti-cytochrome C monoclonal antibody was purchased from R & D Company; 7-dichlorodihydrofluorescein diacetate(DCFH-DA), rhodamine 123 and Mel were purchased from Sigma Company (USA). Lactate dehydrogenase (LDH) kit was purchased from Nanjing Jiancheng Bioengineering Institute.METHODS: This experiment was carried out in the laboratory for Department of Biochemistry and Molecule Biology, Tongji Medical College between October 2002 and March 2004. Cerebellar neurons of rats were cultured in vitro. After oxygen-glucose deprivation (OGD) for 90 minutes, 1×10-4, 1×10-6, 1×10-9 mol/L Mel was added, respectively, namely high-, middle-, and low-concentration Mel groups. Cells, which were cultured by OGD, served as model group, and control group, in which OGD intervention was omitted, was set. ①Cytochrome C level of mitochondrial cells in each group was detected by ELISA method. ②LDH activity in the cell culture fluid was measured, and cell membrane permeability change was analyzed. The cells in the Mel group with the lowest LDH activity served as Mel treatment group, I.e. Cells were cultured with OGD, and then Mel was added; Meanwhile, Mel prevention group was set, I.e. Mel was added before OGD. Intervention was not changed in the

  3. Effect of Qingkailing injection on rat embryonic neuronal apoptosis and mitochondrial membrane potential

    Institute of Scientific and Technical Information of China (English)

    He Pang; Lingqun Zhu; Shuoren Wang; Fuing Niu; Wei Cui

    2006-01-01

    BACKGROUND:The decrease of mitochondrial membrane potential(MMP)is an irreversible marker of neuronal apoptosis during ischemla/reperfusion(I/R)injury of brain tissue.Qingkaiing injection is proved to have protective effect on neuronal ischemic injury.Whether inhibiting the decrease of MMP can inhibit apoptosis when I/R injury of brain tissue occurs is unclear.OBJECTIVE:To observe the effect of Qingkaiing injection on rat embryonic hippocampal neuronal apoptosis,MMP and mitochondroal activity after hypoxia/hypoglycamia and reoxygenation,and make a comparison of therapeutic effect on I/R injury between Oingkaiing injection and nimodipine.DESIGN:Observation and controlled trial.SETTING:Peropheral Vascular Center,Dongzhimen Hospital, Beijing University of Chinese Medicine;the Key Laboratory of Chinese Internal Medicine of Ministry of Education and Beijing Key Laboratory.Dongzhimen Hospital,Beijing University of Chinese Medicine.MATERIALS:Eight Wistar rats at embryonic 18 days,provided by Breeding Farm of Experimental Animals,Chinese Academy of Medical Sciences(Permission No.SCXK-11-00-0006) were employed in this trial.Qingkaiing injection (Pharmaceutical Factory of Beijing University of Chinese Medicine,Batch No.213710A,10 Ml each,baicalin 50 g and total nitrogen 25 mg included)and nimodipine(ICN company,USA)were also used.METHODS:This experiment was carried out in the Key Laboratory of Chinese Internal Medicine of Ministry of Education,Dongzhimen Hospital,Beijing University of Chinese Medicine and Beijing Key Laboratory from January 2003 to December 2005.①The pregnant rats were anesthetized and fetal rats were isolated for culturong fetal rat hippocampal neurons.The neurons cultured for 10 days were used for expedment.The neurons were divided into 5 groups:model group,control group,nimodipine group.Qingkailing high-dose group and Oingkailing low-dose group.Hypoxia/hypoglycemia and reoxygenation models served as model group,and they were used to simulate reperfusion

  4. Calcium-sensing receptor antagonist NPS2390 attenuates neuronal apoptosis though intrinsic pathway following traumatic brain injury in rats.

    Science.gov (United States)

    Xue, Zhaoliang; Song, Zhengfei; Wan, Yingfeng; Wang, Kun; Mo, Lianjie; Wang, Yirong

    2017-04-29

    Traumatic brain injury (TBI) initiates a complex cascade of neurochemical and signaling changes that leads to neuronal apoptosis, which contributes to poor outcomes for patients with TBI. Previous study indicates that calcium-sensing receptor (CaSR) activation contributes to neuron death in focal cerebral ischemia-reperfusion mice, however, its role in neuronal apoptosis after TBI is not well-established. Using a controlled cortical impact model in rats, the present study was designed to determine the effect of CaSR inhibitor NPS2390 upon neuronal apoptosis after TBI. Rats were randomly distributed into three groups undergoing the sham surgery or TBI procedure, and NPS2390 (1.5 mg/kg) was infused subcutaneously at 30 min and 120 min after TBI. All rats were sacrificed at 24 h after TBI. Our data indicated that NPS2390 significantly reduced the brain edema and improved the neurological function after TBI. In addition, NPS2390 decreased caspase-3 levels and the number of apoptotic neurons. Furthermore, NPS2390 up-regulated anti-apoptotic protein Bcl-2 expression and down-regulated pro-apoptotic protein Bax, and reduced subsequent release of cytochrome c into the cytosol. In summary, this study indicated that inhibition of CaSR by NPS2390 attenuates neuronal apoptosis after TBI, in part, through modulating intrinsic apoptotic pathway. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. 14,15-EET promotes mitochondrial biogenesis and protects cortical neurons against oxygen/glucose deprivation-induced apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Lai; Chen, Man; Yuan, Lin; Xiang, Yuting [Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Peking University Health Science Center, Beijing (China); Zheng, Ruimao, E-mail: rmzheng@pku.edu.cn [Department of Anatomy, Histology and Embryology, School of Basic Medical Sciences, Peking University Health Science Center, Beijing (China); Zhu, Shigong, E-mail: sgzhu@bjmu.edu.cn [Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Peking University Health Science Center, Beijing (China)

    2014-07-18

    Highlights: • 14,15-EET inhibits OGD-induced apoptosis in cortical neurons. • Mitochondrial biogenesis of cortical neurons is promoted by 14,15-EET. • 14,15-EET preserves mitochondrial function of cortical neurons under OGD. • CREB mediates effect of 14,15-EET on mitochondrial biogenesis and function. - Abstract: 14,15-Epoxyeicosatrienoic acid (14,15-EET), a metabolite of arachidonic acid, is enriched in the brain cortex and exerts protective effect against neuronal apoptosis induced by ischemia/reperfusion. Although apoptosis has been well recognized to be closely associated with mitochondrial biogenesis and function, it is still unclear whether the neuroprotective effect of 14,15-EET is mediated by promotion of mitochondrial biogenesis and function in cortical neurons under the condition of oxygen–glucose deprivation (OGD). In this study, we found that 14,15-EET improved cell viability and inhibited apoptosis of cortical neurons. 14,15-EET significantly increased the mitochondrial mass and the ratio of mitochondrial DNA to nuclear DNA. Key makers of mitochondrial biogenesis, peroxisome proliferator activator receptor gamma-coactivator 1 alpha (PGC-1α), nuclear respiratory factor 1 (NRF-1) and mitochondrial transcription factor A (TFAM), were elevated at both mRNA and protein levels in the cortical neurons treated with 14,15-EET. Moreover, 14,15-EET markedly attenuated the decline of mitochondrial membrane potential, reduced ROS, while increased ATP synthesis. Knockdown of cAMP-response element binding protein (CREB) by siRNA blunted the up-regulation of PGC-1α and NRF-1 stimulated by 14,15-EET, and consequently abolished the neuroprotective effect of 14,15-EET. Our results indicate that 14,15-EET protects neurons from OGD-induced apoptosis by promoting mitochondrial biogenesis and function through CREB mediated activation of PGC-1α and NRF-1.

  6. Effect of cholecystokinin on learning and memory, neuronal proliferation and apoptosis in the rat hippocampus

    Science.gov (United States)

    Reisi, Parham; Ghaedamini, Ali Reza; Golbidi, Mohammad; Shabrang, Moloud; Arabpoor, Zohreh; Rashidi, Bahman

    2015-01-01

    Background: Cholecystokinin (CCK) has roles in learning and memory, but the cellular mechanism is poorly understood. This study investigated the effect of CCK on spatial learning and memory, neuronal proliferation and apoptosis in the hippocampus in rats. Materials and Methods: Experimental groups were control and CCK. The rats received CKK octapeptide sulfated (CCK-8S, 1.6 μg/kg, i.p.) for 14 days. Spatial learning and memory were tested by Morris water maze and finally immunohistochemical study was performed; neurogenesis by Ki-67 method and apoptosis by Terminal deoxynucleotidyl transferase mediated dUTP Nick End Labeling (TUNEL) assay in hippocampal dentate gyrus (DG). Results: Cholecystokinin increased Ki-67 positive cells and reduced TUNEL positive cells in the granular layer of hippocampal DG. CCK failed to have a significant effect on spatial learning and memory. Conclusion: Results indicate neuroprotective and proliferative effects of CCK in the hippocampus; however, other factors are probably involved until the newly born neurons achieve necessary integrity for behavioral changes. PMID:26623402

  7. Protective effects of Purendan superfine powder on retinal neuron apoptosis in a rat model of type 2 diabetes mellitus

    Institute of Scientific and Technical Information of China (English)

    Zhijun Dong; Xiangyi Tao; Xiaoxiao Fu; Haibin Wang; Donghua Wang; Tiemin Zhang

    2012-01-01

    This study sought to investigate the effects of Purendan superfine powder comprised of Momordica charantia, Radix Ginseng, and Radix Salviae Miltiorrhiae on neuronal apoptosis and expression of bcl-2, bax, and caspase-3, which are retinal apoptosis-associated factors in rats with diabetes mellitus induced by continuous intraperitoneal injection of streptozotocin. The results showed that Purendan superfine powder could upregulate the expression of bcl-2 protein and mRNA, and downregulate the expression of bax and caspase-3 in the retina of diabetes mellitus rats. In addition, Purendan superfine powder was shown to reduce the number of apoptotic neurons. Our experimental findings indicate that Purendan superfine powder can inhibit neuronal apoptosis in the retina of diabetes mellitus rats and has protective effects on diabetic retinopathy.

  8. [The study on the relationship between hippocampus neuronal apoptosis and hippocampus synaptic plasticity in rats exposed to aluminum].

    Science.gov (United States)

    Nie, Xiaohan; Qin, Xiujun; Zhang, Huifang; Kang, Pan; Li, Zhaoyang; Niu, Qiao

    2015-07-01

    To investigate the effect of aluminum exposure on neuronal apoptosis of rats hippocampus and the correlation of and synaptic plasticity. There were 40 SPF grade SD rats which were randomly divided into four groups: the control group, the low dose group, the medium dose group and the high dose group, 10 rats in each group. The rats were daily gavaged with aluminum lactate for 30 days. The hippocampal fEPSPs in rat was measured by electrophysiological grapher and the neuronal apoptosis in hippocampus was detected by Flow cytometer. In addition, the relative expression of gene which includes caspase-3, 8, 9 was measured by Real-time PCR. Compared to the control group, the average of fEPSPs which after HFS 10, 20, 30, 40, 50, 60 min was decreased at different time point in the low dose group, the medium dose group and the high dose group (P neuronal apoptosis in rats, and then affect hippocampal synaptic plasticity.

  9. Sugammadex-Enhanced Neuronal Apoptosis following Neonatal Sevoflurane Exposure in Mice

    Science.gov (United States)

    Sun, Zhongliang; Adachi, Yushi U.; Makita, Koshi

    2016-01-01

    In rodents, neonatal sevoflurane exposure induces neonatal apoptosis in the brain and results in learning deficits. Sugammadex is a new selective neuromuscular blockade (NMB) binding agent that anesthesiologists can use to achieve immediate reversal of an NMB with few side effects. Given its molecular weight of 2178, sugammadex is thought to be unable to pass through the blood brain barrier (BBB). Volatile anesthetics can influence BBB opening and integrity. Therefore, we investigated whether the intraperitoneal administration of sugammadex could exacerbate neuronal damage following neonatal 2% sevoflurane exposure via changes in BBB integrity. Cleaved caspase-3 immunoblotting was used to detect apoptosis, and the ultrastructure of the BBB was examined by transmission electron microscopy. Exposure to 2% sevoflurane for 6 h resulted in BBB ultrastructural abnormalities in the hippocampus of neonatal mice. Sugammadex alone without sevoflurane did not induce apoptosis. The coadministration of sugammadex with sevoflurane to neonatal mice caused a significant increase (150%) in neuroapoptosis in the brain compared with 2% sevoflurane. In neonatal anesthesia, sugammadex could influence neurotoxicity together with sevoflurane. Exposure to 2% sevoflurane for 6 h resulted in BBB ultrastructural abnormalities in the hippocampus of neonatal mice. PMID:27895665

  10. Sugammadex-Enhanced Neuronal Apoptosis following Neonatal Sevoflurane Exposure in Mice

    Directory of Open Access Journals (Sweden)

    Maiko Satomoto

    2016-01-01

    Full Text Available In rodents, neonatal sevoflurane exposure induces neonatal apoptosis in the brain and results in learning deficits. Sugammadex is a new selective neuromuscular blockade (NMB binding agent that anesthesiologists can use to achieve immediate reversal of an NMB with few side effects. Given its molecular weight of 2178, sugammadex is thought to be unable to pass through the blood brain barrier (BBB. Volatile anesthetics can influence BBB opening and integrity. Therefore, we investigated whether the intraperitoneal administration of sugammadex could exacerbate neuronal damage following neonatal 2% sevoflurane exposure via changes in BBB integrity. Cleaved caspase-3 immunoblotting was used to detect apoptosis, and the ultrastructure of the BBB was examined by transmission electron microscopy. Exposure to 2% sevoflurane for 6 h resulted in BBB ultrastructural abnormalities in the hippocampus of neonatal mice. Sugammadex alone without sevoflurane did not induce apoptosis. The coadministration of sugammadex with sevoflurane to neonatal mice caused a significant increase (150% in neuroapoptosis in the brain compared with 2% sevoflurane. In neonatal anesthesia, sugammadex could influence neurotoxicity together with sevoflurane. Exposure to 2% sevoflurane for 6 h resulted in BBB ultrastructural abnormalities in the hippocampus of neonatal mice.

  11. HIV-1gp120 induces neuronal apoptosis through enhancement of 4-aminopyridine-senstive outward K+ currents.

    Directory of Open Access Journals (Sweden)

    Lina Chen

    Full Text Available Human immunodeficiency virus type 1 (HIV-1-associated dementia (HAD usually occurs late in the course of HIV-1 infection and the mechanisms underlying HAD pathogenesis are not well understood. Accumulating evidence indicates that neuronal voltage-gated potassium (Kv channels play an important role in memory processes and acquired neuronal channelopathies in HAD. To examine whether Kv channels are involved in HIV-1-associated neuronal injury, we studied the effects of HIV-1 glycoprotein 120 (gp120 on outward K+ currents in rat cortical neuronal cultures using whole-cell patch techniques. Exposure of cortical neurons to gp120 produced a dose-dependent enhancement of A-type transient outward K+ currents (IA. The gp120-induced increase of IA was attenuated by T140, a specific antagonist for chemokine receptor CXCR4, suggesting gp120 enhancement of neuronal IA via CXCR4. Pretreatment of neuronal cultures with a protein kinase C (PKC inhibitor, GF109203X, inhibited the gp120-induced increase of IA. Biological significance of gp120 enhancement of IA was demonstrated by experimental results showing that gp120-induced neuronal apoptosis, as detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL assay and caspase-3 staining, was attenuated by either an IA blocker 4-aminopyridine or a specific CXCR4 antagonist T140. Taken together, these results suggest that gp120 may induce caspase-3 dependent neuronal apoptosis by enhancing IA via CXCR4-PKC signaling.

  12. Ammonia prevents glutamate-induced but not low K(+)-induced apoptosis in cerebellar neurons in culture.

    Science.gov (United States)

    Llansola, M; Boscá, L; Felipo, V; Hortelano, S

    2003-01-01

    Cultured rat cerebellar granule neurons are widely used as a model system for studying neuronal apoptosis. Either low K(+) (5 mM) or low concentrations of glutamate (1-10 microM) induce apoptosis in cerebellar neurons in culture. However, the molecular mechanism(s) involved remain unclear. We show that long-term treatment with ammonia prevents glutamate-induced but not low K(+)-induced apoptosis in cerebellar neurons, as assessed by measuring DNA fragmentation and activation of caspase 3. Ammonia prevented glutamate-induced increase of intracellular calcium, depolarization of the inner mitochondrial membrane, release of cytochrome c to the cytosol, activation of caspase 3 and fragmentation of DNA. However, ammonia did not prevent low K(+)-induced activation of caspase 3 and fragmentation of DNA. These results indicate that the initial steps involved in the induction of apoptosis by low K(+) or by glutamate are different and that ammonia prevents glutamate-induced apoptosis by reducing glutamate-induced rise of intracellular Ca(2+), thus avoiding the activation of subsequent events of the apoptotic process.

  13. Sericin can reduce hippocampal neuronal apoptosis by activating the Akt signal transduction pathway in a rat model of diabetes mellitus

    Institute of Scientific and Technical Information of China (English)

    Zhihong Chen; Yaqiang He; Chengjun Song; Zhijun Dong; Zhejun Su; Jingfeng Xue

    2012-01-01

    In the present study, a rat model of type 2 diabetes mellitus was established by continuous peritoneal injection of streptozotocin. Following intragastric perfusion of sericin for 35 days, blood glucose levels significantly reduced, neuronal apoptosis in the hippocampal CA1 region decreased, hippocampal phosphorylated Akt and nuclear factor kappa B expression were enhanced, but Bcl-xL/Bcl-2 associated death promoter expression decreased. Results demonstrated that sericin can reduce hippocampal neuronal apoptosis in a rat model of diabetes mellitus by regulating abnormal changes in the Akt signal transduction pathway.

  14. Effect of clausenamide on hippocampal neuron apoptosis induced by sodium nitroprusside

    Institute of Scientific and Technical Information of China (English)

    Yongjun Liu; Qifeng Zhu

    2007-01-01

    BACKGROUND: Aggregation of β -amyloid peptide (A β ), excitatory intoxication, oxidation injury and inflammation reaction are generally regarded as the main pathogenesis for Alzheimer disease (AD). (-)clausenamide is characterized by promoting intelligent development, resisting oxidation, cleaning free radicals, resisting A β neurotoxicity and nerve cell apoptosis, inhibiting over phosphorylation of tau protein,and improving central cholinergic system. However, whether (-) clausenamide has an effect on hippocampal neuron apoptosis or not need further study.OBJECTIVE: To observe the effect of ( - ) clausenamide on survival rate of hippocampal neurons due to sodium nitroprusside (SNP) and analyze the possible pathways.DESIGN: Contrast observation.SETTING: Institute of Biochemistry and Molecular Biology, Guangdong Medical College.MATERIALS: A total of 12 male SD rats of 24 hours old were provided by the Experimental Animal Center of Guangdong Medical College. The primer was synthesized by Beijing Huada Genetic Engineering Company and (-) clausenamide (99.6%) was provided by Pharmacological Department of Chinese Academy of Medical Sciences. SNP was provided by Sigma Company.METHODS: Bilateral hippocampus was collected from newborn rats to establish single cell suspension. On the 12th day, hippocampal neurons were pretreated with 0.2, 0.4, 0.8 and 1.6 μ mol/L ( - ) clausenamide for 6 hours; the culture medium was gotten rid of and neurons were washed with non-serum DMEM solution for three times. In addition, non-serum DMEM solution was added with the above mentioned volume of ( - ) clausenamide and 50 μ mol/L SNP to culture neurons for 24 hours and the collected cells were prepared for the experiment. Neurons were equally divided into control group (culture medium control), model group (SNP treatment) and experimental group [( - ) clausenamide + SNP]. Experiment of each group was done for three times at least. Survival rate of cells was measured with MTT

  15. I(2)(PP2A) regulates p53 and Akt correlatively and leads the neurons to abort apoptosis.

    Science.gov (United States)

    Liu, Gong-Ping; Wei, Wei; Zhou, Xin; Zhang, Yao; Shi, Hai-Hong; Yin, Jun; Yao, Xiu-Qing; Peng, Cai-Xia; Hu, Juan; Wang, Qun; Li, Hong-Lian; Wang, Jian-Zhi

    2012-02-01

    A chronic neuron loss is the cardinal pathology in Alzheimer disease (AD), but it is still not understood why most neurons in AD brain do not accomplish apoptosis even though they are actually exposed to an environment with enriched proapoptotic factors. Protein phosphatase-2A inhibitor-2 (I(2)(PP2A)), an endogenous PP2A inhibitor, is significantly increased in AD brain, but the role of I(2)(PP2A) in AD-like neuron loss is elusive. Here, we show that I(2)(PP2A) regulates p53 and Akt correlatively. The mechanisms involve activated transcription and p38 MAPK activities. More importantly, we demonstrate that the simultaneous activation of Akt induced by I(2)(PP2A) counteracts the hyperactivated p53-induced cell apoptosis. Furthermore, I(2)(PP2A), p53 and Akt are all elevated in the brain of mouse model and AD patients. Our results suggest that the increased I(2)(PP2A) may trigger apoptosis by p53 upregulation, but due to simultaneous activation of Akt, the neurons are aborted from the apoptotic pathway. This finding contributes to the understanding of why most neurons in AD brain do not undergo apoptosis.

  16. Protective effect of tubuloside B on TNFα-induced apoptosis in neuronal cells

    Institute of Scientific and Technical Information of China (English)

    Min DENG; Jin-yuan ZHAO; Xiao-dong JU; Peng-fei TU; Yong JIANG; Zheng-bin LI

    2004-01-01

    AIM: To investigate the neuroprotective effect of tubuloside B, one of the phenylethanoids isolated from the stems of Cistanche salsa, on tumor necrosis factor-alpha (TNFα)-induced apoptosis in SH-SY5Y neuronal cells.METHODS: Cell viability was analyzed using MTT assay. Apoptotic cells were detected using Hoechst33342 staining, and confirmed by DNA fragmentation and flow cytometric analysis. The activity of caspase-3 was measured with special assay kit. The concentration of free intracellular calcium was determined with the probe Indo-1 by spectrometer. The level of intracellular reactive oxygen species and the potential of mitochondrial membrane were determined by laser scanning confocal microscopy (LSCM) combined with fluorescence probe H2DCFDA or JC-1 respectively. RESULTS: SH-SY5Y cells treated with TNFα 100 μg/L for 36 h showed typical morphological changes of apoptosis. DNA ladder could be observed by agarose gel electrophoresis. The highest percentage of apoptotic cells accumulated to 37.5 %. Following 36 h treatment with TNFα, accumulation of intracellular ROS and [Ca2+]i and decrease in mitochondrial membrane potential were observed, and caspase-3 activity increased by about five-fold compared with controls. However, pretreatment with tubuloside B (1, 10, or 100 mg/L) for 2 h attenuated the TNFα-mediated apoptosis. The antiapoptotic action of tubuloside B was partially dependent on an anti-oxidative stress effects, maintain of mitochondria function, decrease of concentration of free intracellular calcium and inhibition of caspase-3 activity. CONCLUSION: Tubuloside B has the neuroprotective capacity to antagonize TNFα-induced apoptosis in SH-SY5Y cells and may be useful in treating some neurodegenerative diseases.

  17. L-carnitine protects neurons from 1-methyl-4-phenylpyridinium-induced neuronal apoptosis in rat forebrain culture.

    Science.gov (United States)

    Wang, C; Sadovova, N; Ali, H K; Duhart, H M; Fu, X; Zou, X; Patterson, T A; Binienda, Z K; Virmani, A; Paule, M G; Slikker, W; Ali, S F

    2007-01-05

    1-Methyl-4-phenylpyridinium ion (MPP+), an inhibitor of mitochondrial complex I, has been widely used as a neurotoxin because it elicits a severe Parkinson's disease-like syndrome with an elevation of intracellular reactive oxygen species (ROS) and apoptosis. L-carnitine plays an integral role in attenuating the brain injury associated with mitochondrial neurodegenerative disorders. The present study investigates the effects of L-carnitine against the toxicity of MPP+ in rat forebrain primary cultures. Cells in culture were treated for 24 h with 100, 250, 500 and 1000 microM MPP+ alone or co-incubated with L-carnitine. MPP+ produced a dose-related increase in DNA fragmentation as measured by cell death ELISA (enzyme-linked immunosorbent assay), an increase in the number of TUNEL (terminal dUTP nick-end labeling)-positive cells and a reduction in the mitochondrial metabolism of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). No significant effect was observed with the release of lactate dehydrogenase (LDH), indicating that cell death presumably occurred via apoptotic mechanisms. Co-incubation of MPP+ with L-carnitine significantly reduced MPP+-induced apoptosis. Western blot analyses showed that neurotoxic concentrations of MPP+ decreased the ratio of BCL-X(L) to Bax and decreased the protein levels of polysialic acid neural cell adhesion molecules (PSA-NCAM), a neuron specific marker. L-carnitine blocked these effects of MPP+ suggesting its potential therapeutic utility in degenerative disorders such as Parkinson's disease, Alzheimer's disease, ornithine transcarbamylase deficiency and other mitochondrial diseases.

  18. Hyperbaric oxygenation alleviates chronic constriction injury (CCI)-induced neuropathic pain and inhibits GABAergic neuron apoptosis in the spinal cord.

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    Fu, Huiqun; Li, Fenghua; Thomas, Sebastian; Yang, Zhongjin

    2017-09-15

    Dysfunction of GABAergic inhibitory controls contributes to the development of neuropathic pain. We examined our hypotheses that (1) chronic constriction injury (CCI)-induced neuropathic pain is associated with increased spinal GABAergic neuron apoptosis, and (2) hyperbaric oxygen therapy (HBO) alleviates CCI-induced neuropathic pain by inhibiting GABAergic neuron apoptosis. Male rats were randomized into 3 groups: CCI, CCI+HBO and the control group (SHAM). Mechanical allodynia was tested daily following CCI procedure. HBO rats were treated at 2.4 atmospheres absolute (ATA) for 60min once per day. The rats were euthanized and the spinal cord harvested on day 8 and 14 post-CCI. Detection of GABAergic cells and apoptosis was performed. The percentages of double positive stained cells (NeuN/GABA), cleaved caspase-3 or Cytochrome C in total GABAergic cells or in total NeuN positive cells were calculated. HBO significantly alleviated mechanical allodynia. CCI-induced neuropathic pain was associated with significantly increased spinal apoptotic GABA-positive neurons. HBO considerably decreased these spinal apoptotic cells. Cytochrome-C-positive neurons and cleaved caspase-3-positive neurons were also significantly higher in CCI rats. HBO significantly decreased these positive cells. Caspase-3 mRNA was also significantly higher in CCI rats. HBO reduced mRNA expression of caspase-3. CCI-induced neuropathic pain was associated with increased apoptotic GABAergic neurons induced by activation of key proteins of mitochondrial apoptotic pathways in the dorsal horn of the spinal cord. HBO alleviated CCI-induced neuropathic pain and reduced GABAergic neuron apoptosis. The beneficial effect of HBO may be via its inhibitory role in CCI-induced GABAergic neuron apoptosis by suppressing mitochondrial apoptotic pathways in the spinal cord. Increased apoptotic GABAergic neurons induced by activation of key proteins of mitochondrial apoptotic pathways in the dorsal horn of the spinal

  19. Metallothionein mediates leukocyte chemotaxis

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    Lynes Michael A

    2005-09-01

    Full Text Available Abstract Background Metallothionein (MT is a cysteine-rich, metal-binding protein that can be induced by a variety of agents. Modulation of MT levels has also been shown to alter specific immune functions. We have noticed that the MT genes map close to the chemokines Ccl17 and Cx3cl1. Cysteine motifs that characterize these chemokines are also found in the MT sequence suggesting that MT might also act as a chemotactic factor. Results In the experiments reported here, we show that immune cells migrate chemotactically in the presence of a gradient of MT. This response can be specifically blocked by two different monoclonal anti-MT antibodies. Exposure of cells to MT also leads to a rapid increase in F-actin content. Incubation of Jurkat T cells with cholera toxin or pertussis toxin completely abrogates the chemotactic response to MT. Thus MT may act via G-protein coupled receptors and through the cyclic AMP signaling pathway to initiate chemotaxis. Conclusion These results suggest that, under inflammatory conditions, metallothionein in the extracellular environment may support the beneficial movement of leukocytes to the site of inflammation. MT may therefore represent a "danger signal"; modifying the character of the immune response when cells sense cellular stress. Elevated metallothionein produced in the context of exposure to environmental toxicants, or as a result of chronic inflammatory disease, may alter the normal chemotactic responses that regulate leukocyte trafficking. Thus, MT synthesis may represent an important factor in immunomodulation that is associated with autoimmune disease and toxicant exposure.

  20. Neuroprotective mechanisms of curcumin against cerebral ischemia-induced neuronal apoptosis and behavioral deficits.

    Science.gov (United States)

    Wang, Qun; Sun, Albert Y; Simonyi, Agnes; Jensen, Michael D; Shelat, Phullara B; Rottinghaus, George E; MacDonald, Ruth S; Miller, Dennis K; Lubahn, Dennis E; Weisman, Gary A; Sun, Grace Y

    2005-10-01

    Increased oxidative stress has been regarded as an important underlying cause for neuronal damage induced by cerebral ischemia/reperfusion (I/R) injury. In recent years, there has been increasing interest in investigating polyphenols from botanical source for possible neuroprotective effects against neurodegenerative diseases. In this study, we investigated the mechanisms underlying the neuroprotective effects of curcumin, a potent polyphenol antioxidant enriched in tumeric. Global cerebral ischemia was induced in Mongolian gerbils by transient occlusion of the common carotid arteries. Histochemical analysis indicated extensive neuronal death together with increased reactive astrocytes and microglial cells in the hippocampal CA1 area at 4 days after I/R. These ischemic changes were preceded by a rapid increase in lipid peroxidation and followed by decrease in mitochondrial membrane potential, increased cytochrome c release, and subsequently caspase-3 activation and apoptosis. Administration of curcumin by i.p. injections (30 mg/kg body wt) or by supplementation to the AIN76 diet (2.0 g/kg diet) for 2 months significantly attenuated ischemia-induced neuronal death as well as glial activation. Curcumin administration also decreased lipid peroxidation, mitochondrial dysfunction, and the apoptotic indices. The biochemical changes resulting from curcumin also correlated well with its ability to ameliorate the changes in locomotor activity induced by I/R. Bioavailability study indicated a rapid increase in curcumin in plasma and brain within 1 hr after treatment. Together, these findings attribute the neuroprotective effect of curcumin against I/R-induced neuronal damage to its antioxidant capacity in reducing oxidative stress and the signaling cascade leading to apoptotic cell death.

  1. Mkp1 is a c-Jun target gene that antagonizes JNK-dependent apoptosis in sympathetic neurons.

    Science.gov (United States)

    Kristiansen, Mark; Hughes, Rosie; Patel, Pritika; Jacques, Thomas S; Clark, Andrew R; Ham, Jonathan

    2010-08-11

    Developing sympathetic neurons depend on NGF for survival. When sympathetic neurons are deprived of NGF in vitro, a well documented series of events, including c-Jun N-terminal kinase (JNK) pathway activation, release of cytochrome c from the mitochondria, and caspase activation, culminates in the death of the neuron by apoptosis within 24-48 h. This process requires de novo gene expression, suggesting that increased expression of specific genes activates the cell death program. Using rat gene microarrays, we found that NGF withdrawal induces the expression of many genes, including mkp1, which encodes a MAPK phosphatase that can dephosphorylate JNKs. The increase in mkp1 mRNA level requires the MLK-JNK-c-Jun pathway, and we show that Mkp1 is an important regulator of JNK-dependent apoptosis in sympathetic neurons. In microinjection experiments, Mkp1 overexpression can inhibit JNK-mediated phosphorylation of c-Jun and protect sympathetic neurons from apoptosis, while Mkp1 knockdown accelerates NGF withdrawal-induced death. Accordingly, the number of superior cervical ganglion (SCG) neurons is reduced in mkp1-/- mice at P1 during the period of developmental sympathetic neuron death. We also show that c-Jun and ATF2 bind to two conserved ATF binding sites in the mkp1 promoter in vitro and in chromatin. Both of these ATF sites contribute to basal promoter activity and are required for mkp1 promoter induction after NGF withdrawal. These results demonstrate that Mkp1 is part of a negative feedback loop induced by the MLK-JNK-c-Jun signaling pathway that modulates JNK activity and the rate of neuronal death in rat sympathetic neurons following NGF withdrawal.

  2. Caspase 3 siRNA decreases apoptosis in cultured neuronal cells

    Institute of Scientific and Technical Information of China (English)

    Chunting Ye; Yaoxiong Huang; Xiaohong Yang; Honghui Chen

    2009-01-01

    BACKGROUND: Lentiviral vectors, a type of retroviral vector, are able to infect cells at all phases of cell cycle. They are able to express exogenous target genes in vivo over long periods of time with limited immunological reaction. OBJECTIVE: To inhibit neuronal apoptosis by blocking the apoptotic cascade reaction, gene silencing of Caspase 3, and transfection of Caspase 3 short hairpin ribonucleic acid (shRNA) into Neuro 2a cells using a lentiviral vector. DESIGN, TIME AND SETTING: An observational, genetic engineering cellular biology experiment was performed in Guangzhou Red Cross Hospital and Guangzhou institute of Traumatic Surgery between March 2007 and June 2008.MATERIALS: PLL3.7, PCMV-VSV-G, and PH'8.9△PR plasmids were provided by the CBR institute for Biomedical Research, Harvard Medical School, USA. Staurosporine was purchased from Sigma, USA. METHODS: Caspase 3 siRNA was synthesized and cloned into the PLL3.7 plasmid. The Caspase 3 shRNA-PLL3.7 lentivirus was generated in 293T cells using a calcium phosphate transfection kit. After the lentivirus was transfected into Neuro 2a cells, apoptosis was induced in both the transfected and untransfected cells by staurosporine. Cell apoptosis was assessed by flow cytometry. MAIN OUTCOME MEASURES: Caspase 3 mRNA expression was measured by RT-PCR and Caspase protein expression was assessed by Western blot. Cellular apoptosis was determined by flow cytometry using Annevin V-PE/7aad-Cy7.RESULTS: The transfection rate of caspase 3 shRNA was>98% using the ientiviral vector. RT-PCR and Western blot results demonstrated that significantly reduced Caspase 3 mRNA and protein expression in the transfected Neuro 2a. The control group exhibited 38.7% Annexin V/7aad-positive cells, which suggested apoptotic anaphase, while only 5.0% cells in the Caspase 3 gene silencing group entered apoptotic anaphase. CONCLUSION: Caspase 3 shRNA inhibited Caspase 3 expression in Neuro 2a cells and decreased drug-induced apoptosis of

  3. Curcumin protects microglia and primary rat cortical neurons against HIV-1 gp120-mediated inflammation and apoptosis.

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    Luyan Guo

    Full Text Available Curcumin is a molecule found in turmeric root that has anti-inflammatory, antioxidant, and anti-tumor properties and has been widely used as both an herbal drug and a food additive to treat or prevent neurodegenerative diseases. To explore whether curcumin is able to ameliorate HIV-1-associated neurotoxicity, we treated a murine microglial cell line (N9 and primary rat cortical neurons with curcumin in the presence or absence of neurotoxic HIV-1 gp120 (V3 loop protein. We found that HIV-1 gp120 profoundly induced N9 cells to produce reactive oxygen species (ROS, tumor necrosis factor-α (TNF-α and monocyte chemoattractant protein-1 (MCP-1. HIV-1 gp120 also induced apoptosis of primary rat cortical neurons. Curcumin exerted a powerful inhibitory effect against HIV-1 gp120-induced neuronal damage, reducing the production of ROS, TNF-α and MCP-1 by N9 cells and inhibiting apoptosis of primary rat cortical neurons. Curcumin may exert its biological activities through inhibition of the delayed rectification and transient outward potassium (K(+ current, as curcumin effectively reduced HIV-1 gp120-mediated elevation of the delayed rectification and transient outward K(+ channel current in neurons. We conclude that HIV-1 gp120 increases ROS, TNF-α and MCP-1 production in microglia, and induces cortical neuron apoptosis by affecting the delayed rectification and transient outward K(+ channel current. Curcumin reduces production of ROS and inflammatory mediators in HIV-1-gp120-stimulated microglia, and protects cortical neurons against HIV-1-mediated apoptosis, most likely through inhibition of HIV-1 gp120-induced elevation of the delayed rectification and transient outward K(+ current.

  4. Serine-threonine protein kinase activation may be an effective target for reducing neuronal apoptosis after spinal cord injury

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    Mu Jin

    2015-01-01

    Full Text Available The signaling mechanisms underlying ischemia-induced nerve cell apoptosis are poorly understood. We investigated the effects of apoptosis-related signal transduction pathways following ischemic spinal cord injury, including extracellular signal-regulated kinase (ERK, serine-threonine protein kinase (Akt and c-Jun N-terminal kinase (JNK signaling pathways. We established a rat model of acute spinal cord injury by inserting a catheter balloon in the left subclavian artery for 25 minutes. Rat models exhibited notable hindlimb dysfunction. Apoptotic cells were abundant in the anterior horn and central canal of the spinal cord. The number of apoptotic neurons was highest 48 hours post injury. The expression of phosphorylated Akt (p-Akt and phosphorylated ERK (p-ERK increased immediately after reperfusion, peaked at 4 hours (p-Akt or 2 hours (p-ERK, decreased at 12 hours, and then increased at 24 hours. Phosphorylated JNK expression reduced after reperfusion, increased at 12 hours to near normal levels, and then showed a downward trend at 24 hours. Pearson linear correlation analysis also demonstrated that the number of apoptotic cells negatively correlated with p-Akt expression. These findings suggest that activation of Akt may be a key contributing factor in the delay of neuronal apoptosis after spinal cord ischemia, particularly at the stage of reperfusion, and thus may be a target for neuronal protection and reduction of neuronal apoptosis after spinal cord injury.

  5. Gadd45b prevents autophagy and apoptosis against rat cerebral neuron oxygen-glucose deprivation/reperfusion injury.

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    He, Guoqian; Xu, Wenming; Tong, Linyan; Li, Shuaishuai; Su, Shiceng; Tan, Xiaodan; Li, Changqing

    2016-04-01

    Autophagic (type II) cell death has been suggested to play pathogenetic roles in cerebral ischemia. Growth arrest and DNA damage response 45b (Gadd45b) has been shown to protect against rat brain ischemia injury through inhibiting apoptosis. However, the relationship between Gadd45b and autophagy in cerebral ischemia/reperfusion (I/R) injury remains uncertain. The aim of this study is to investigate the effect of Gadd45b on autophagy. We adopt the oxygen-glucose deprivation and reperfusion (OGD/R) model of rat primary cortex neurons, and lentivirus interference used to silence Gadd45b expression. Cell viability and injury assay were performed using CCK-8 and LDH kit. Autophagy activation was monitored by expression of ATG5, LC3, Beclin-1, ATG7 and ATG3. Neuron apoptosis was monitored by expression of Bcl-2, Bax, cleaved caspase3, p53 and TUNEL assay. Neuron neurites were assayed by double immunofluorescent labeling with Tuj1 and LC3B. Here, we demonstrated that the expression of Gadd45b was strongly up-regulated at 24 h after 3 h OGD treatment. ShRNA-Gadd45b increased the expression of autophagy related proteins, aggravated OGD/R-induced neuron cell apoptosis and neurites injury. ShRNA-Gadd45b co-treatment with autophagy inhibitor 3-methyladenine (3-MA) or Wortmannin partly inhibited the ratio of LC3II/LC3I, and slightly ameliorated neuron cell apoptosis under OGD/R. Furthermore, shRNA-Gadd45b inhibited the p-p38 level involved in autophagy, but increased the p-JNK level involved in apoptosis. ShRNA-Gadd45b co-treatment with p38 inhibitor obviously induced autophagy. ShRNA-Gadd45b co-treatment with JNK inhibitor alleviated neuron cell apoptosis. In conclusion, our data suggested that Gadd45b inhibited autophagy and apoptosis under OGD/R. Gadd45b may be a common regulatory protein to control autophagy and apoptosis.

  6. Pivotal roles of p53 transcription-dependent and -independent pathways in manganese-induced mitochondrial dysfunction and neuronal apoptosis

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    Wan, Chunhua [Department of Nutrition and Food Hygiene, School of Public Health, Nantong University, Nantong 226019 Jiangsu (China); Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226019 Jiangsu (China); Ma, Xa; Shi, Shangshi [Department of Occupational Medicine and Environmental Toxicology, School of Public Health, Nantong University, Nantong 226019 Jiangsu (China); Zhao, Jianya; Nie, Xiaoke [Department of Nutrition and Food Hygiene, School of Public Health, Nantong University, Nantong 226019 Jiangsu (China); Han, Jingling; Xiao, Jing; Wang, Xiaoke [Department of Occupational Medicine and Environmental Toxicology, School of Public Health, Nantong University, Nantong 226019 Jiangsu (China); Jiang, Shengyang [Department of Nutrition and Food Hygiene, School of Public Health, Nantong University, Nantong 226019 Jiangsu (China); Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226019 Jiangsu (China); Jiang, Junkang, E-mail: Jiang_junkang@163.com [Department of Occupational Medicine and Environmental Toxicology, School of Public Health, Nantong University, Nantong 226019 Jiangsu (China); Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226019 Jiangsu (China)

    2014-12-15

    Chronic exposure to excessive manganese (Mn) has been known to lead to neuronal loss and a clinical syndrome resembling idiopathic Parkinson's disease (IPD). p53 plays an integral role in the development of various human diseases, including neurodegenerative disorders. However, the role of p53 in Mn-induced neuronal apoptosis and neurological deficits remains obscure. In the present study, we showed that p53 was critically involved in Mn-induced neuronal apoptosis in rat striatum through both transcription-dependent and -independent mechanisms. Western blot and immunohistochemistrical analyses revealed that p53 was remarkably upregulated in the striatum of rats following Mn exposure. Coincidentally, increased level of cleaved PARP, a hallmark of apoptosis, was observed. Furthermore, using nerve growth factor (NGF)-differentiated PC12 cells as a neuronal cell model, we showed that Mn exposure decreased cell viability and induced apparent apoptosis. Importantly, p53 was progressively upregulated, and accumulated in both the nucleus and the cytoplasm. The cytoplasmic p53 had a remarkable distribution in mitochondria, suggesting an involvement of p53 mitochondrial translocation in Mn-induced neuronal apoptosis. In addition, Mn-induced impairment of mitochondrial membrane potential (ΔΨm) could be partially rescued by pretreatment with inhibitors of p53 transcriptional activity and p53 mitochondrial translocation, Pifithrin-α (PFT-α) and Pifithrin-μ (PFT-μ), respectively. Moreover, blockage of p53 activities with PFT-α and PFT-μ significantly attenuated Mn-induced reactive oxidative stress (ROS) generation and mitochondrial H{sub 2}O{sub 2} production. Finally, we observed that pretreatment with PFT-α and PFT-μ ameliorated Mn-induced apoptosis in PC12 cells. Collectively, these findings implicate that p53 transcription-dependent and -independent pathways may play crucial roles in the regulation of Mn-induced neuronal death. - Highlights: • p53 is

  7. Histone H4 deacetylation plays a critical role in early gene silencing during neuronal apoptosis

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    Schlamp Cassandra L

    2010-05-01

    Full Text Available Abstract Background Silencing of normal gene expression occurs early in the apoptosis of neurons, well before the cell is committed to the death pathway, and has been extensively characterized in injured retinal ganglion cells. The causative mechanism of this widespread change in gene expression is unknown. We investigated whether an epigenetic change in active chromatin, specifically histone H4 deacetylation, was an underlying mechanism of gene silencing in apoptotic retinal ganglion cells (RGCs following an acute injury to the optic nerve. Results Histone deacetylase 3 (HDAC3 translocates to the nuclei of dying cells shortly after lesion of the optic nerve and is associated with an increase in nuclear HDAC activity and widespread histone deacetylation. H4 in promoters of representative genes was rapidly and indiscriminately deacetylated, regardless of the gene examined. As apoptosis progressed, H4 of silenced genes remained deacetylated, while H4 of newly activated genes regained, or even increased, its acetylated state. Inhibition of retinal HDAC activity with trichostatin A (TSA was able to both preserve the expression of a representative RGC-specific gene and attenuate cell loss in response to optic nerve damage. Conclusions These data indicate that histone deacetylation plays a central role in transcriptional dysregulation in dying RGCs. The data also suggests that HDAC3, in particular, may feature heavily in apoptotic gene silencing.

  8. Baicalein Promotes Neuronal and Behavioral Recovery After Intracerebral Hemorrhage Via Suppressing Apoptosis, Oxidative Stress and Neuroinflammation.

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    Wei, Ning; Wei, Yinghai; Li, Binru; Pang, Linlin

    2017-01-21

    Intracerebral hemorrhage (ICH) is an important public health problem in neurology, which is not only associated with high mortality but also leading to disability. Yet no satisfactory treatment has been developed. The secondary injury that resulted from a number of self-destructive processes such as neuroinflammation, apoptosis and oxidative stress, is the key factor contributing to ICH-induced brain damage. Baicalein has been proved to improve neuronal functional recovery in rat model of subarachnoid hemorrhage and ischemic brain damage. To investigate the effect of baicalein on ICH and its underlying mechanism, a collagenase-induced ICH rat model was performed. Baicalein treatment significantly decreased neurological severity score at day 1 and 3 after ICH injury. Our results showed that the lesion volume, the brain water content, the expression levels of four pro-inflammatory cytokines (IL-1β, IL-4 and IL-6 and TNF-α) and the numbers of apoptotic cells were reduced significantly in ICH rats receiving baicalein treatment, especially in 50 mg/kg baicalein-treated group. Moreover, baicalein increased SOD and GSH-Px activities and down-regulated MDA level of brain tissues in rats. These results suggested that the therapeutic efficacy of baicalein on repairing brain damage is probably caused by suppressing apoptosis, oxidative stress and neuroinflammation. Baicalein could be developed into a novel drug for clinical treatment of ICH and ICH-related brain injuries.

  9. Estrogen protects neuronal cells from amyloid beta-induced apoptosis via regulation of mitochondrial proteins and function

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    Iwamoto Sean

    2006-11-01

    Full Text Available Abstract Background Neurodegeneration in Alzheimer's disease is associated with increased apoptosis and parallels increased levels of amyloid beta, which can induce neuronal apoptosis. Estrogen exposure prior to neurotoxic insult of hippocampal neurons promotes neuronal defence and survival against neurodegenerative insults including amyloid beta. Although all underlying molecular mechanisms of amyloid beta neurotoxicity remain undetermined, mitochondrial dysfunction, including altered calcium homeostasis and Bcl-2 expression, are involved in neurodegenerative vulnerability. Results In this study, we investigated the mechanism of 17β-estradiol-induced prevention of amyloid beta-induced apoptosis of rat hippocampal neuronal cultures. Estradiol treatment prior to amyloid beta exposure significantly reduced the number of apoptotic neurons and the associated rise in resting intracellular calcium levels. Amyloid beta exposure provoked down regulation of a key antiapoptotic protein, Bcl-2, and resulted in mitochondrial translocation of Bax, a protein known to promote cell death, and subsequent release of cytochrome c. E2 pretreatment inhibited the amyloid beta-induced decrease in Bcl-2 expression, translocation of Bax to the mitochondria and subsequent release of cytochrome c. Further implicating the mitochondria as a target of estradiol action, in vivo estradiol treatment enhanced the respiratory function of whole brain mitochondria. In addition, estradiol pretreatment protected isolated mitochondria against calcium-induced loss of respiratory function. Conclusion Therefore, we propose that estradiol pretreatment protects against amyloid beta neurotoxicity by limiting mitochondrial dysfunction via activation of antiapoptotic mechanisms.

  10. Scutellaria baicalensis stem-leaf total flavonoid reduces neuronal apoptosis induced by amyloid beta-peptide (25-35)

    Institute of Scientific and Technical Information of China (English)

    Ruiting Wang; Xingbin Shen; Enhong Xing; Lihua Guan; Lisheng Xin

    2013-01-01

    Scutellaria baicalensis stem-leaf total flavonoid might attenuate learning/memory impairment and neuronal loss in rats induced by amyloid beta-peptide. This study aimed to explore the effects of Scutellaria baicalensis stem-leaf total flavonoid on amyloid beta-peptide-induced neuronal apoptosis and the expression of apoptosis-related proteins in the rat hippocampus. Male Wistar rats were given intragastric administration of Scutellaria baicalensis stem-leaf total flavonoid, 50 or 100 mg/kg, once per day. On day 8 after administration, 10 μg amyloid beta-peptide (25–35) was injected into the bilateral hippocampus of rats to induce neuronal apoptosis. On day 20, hippocampal tissue was harvested and probed with the terminal deoxyribonucleotidyl transferase-mediated biotin-16-dUTP nick-end labeling assay. Scutellaria baicalensis stem-leaf total flavonoid at 50 and 100 mg/kg reduced neuronal apoptosis induced by amyloid beta-peptide (25–35) in the rat hippocampus. Immunohistochemistry and western blot assay revealed that expression of the pro-apoptotic protein Bax, cytochrome c and caspase-3 was significantly diminished by 50 and 100 mg/kg Scutellaria baicalensis stem-leaf total flavonoid, while expression of the anti-apoptotic protein Bcl-2 was increased. Moreover, 100 mg/kg Scutellaria baicalensis stem-leaf total flavonoid had a more dramatic effect than the lower dosage. These experimental findings indicate that Scutellaria baicalensis stem-leaf total flavonoid dose-dependently attenuates neuronal apoptosis induced by amyloid beta-peptide in the hippocampus, and it might mediate this by regulating the expression of Bax, cytochrome c, caspase-3 and Bcl-2.

  11. Recombinant-activated factor Ⅶ and neuronal apoptosis in a rat model of intracerebral hemorrhage

    Institute of Scientific and Technical Information of China (English)

    Qiang Li; Wei Li; Suju Ding; Jianping Tang; Jing Fang; Benqiang Deng; Tao Wu

    2009-01-01

    BACKGROUND:Activated clotting factor Ⅶ has been demonstrated to exhibit obvious anti-apoptosis effects.OBJECTIVE:To observe the effect of activated clotting factor Ⅶ on neuronal apoptosis at different time points following rat intracerebral hemorrhage (ICH).DESIGN,TIME AND SETTING:A randomized,controlled,animal experiment was performed at the Neurobiological Laboratory of Second Military Medical University from October 2005 to April 2006.MATERIALS:Recombinant-activated clotting factor Vlla (rFⅧa) was purchased from Danish Novo Nordisk,Denmark.In situ cell death detection kit-POD kit was purchased from Roche,Switzerland.Caspase-3 activity determination kit from Biovision,USA.METHODS:A total of 72 healthy,male,Sprague Dawley rats,aged 5-8 months,were randomly assigned to three groups (n=24):sham-operated,ICH model,and rFⅧa.In the ICH model and rFⅧa groups,80.0 μL autologous non-clotting blood from rat tails was injected into the right caudate putamen to establish the ICH.The empty microinjector was inserted into the caudate putamen in the sham-operated group.The ICH model and rFⅧa groups were subdivided into four subsets separately:6,24,72 hours and 7 clays following ICH.The rats in the rFⅧa group were injected with 160 μg/kg rFⅧa via the dorsal vein of the penis.MAIN OUTCOME MEASURES:Apoptotic cells were detected in the right caudate putamen by TUNEL;caspase-3 activity by spectrophotometry;and rat neurological function was evaluated by neurological functional impairment scales.RESULTS:Rat neurological function was deteriorated at 24,72 hours,and 7 days following ICH.The TUNEL-positive cells and caspase-3 activity in the right caudate putamen was significantly increased in the ICH rats (P<0.05);rFVlla treatment reduced the number of TUNEL-positive cells and caspase-3 activity in the right caudate putamen (P<0.05),and neurological function was significantly improved (P<0.05).CONCLUSION:rFⅧa was applied within 72 hours after ICH,which reduced

  12. Oxidative DNA Damage Mediated by Intranuclear MMP Activity Is Associated with Neuronal Apoptosis in Ischemic Stroke

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    Shihoko Kimura-Ohba

    2016-01-01

    Full Text Available Evidence of the pathological roles of matrix metalloproteinases (MMPs in various neurological disorders has made them attractive therapeutic targets. MMPs disrupt the blood-brain barrier and cause neuronal death and neuroinflammation in acute cerebral ischemia and are critical for angiogenesis during recovery. However, some challenges have to be overcome before MMPs can be further validated as drug targets in stroke injury. Identifying in vivo substrates of MMPs should greatly improve our understanding of the mechanisms of ischemic injury and is critical for providing more precise drug targets. Recent works have uncovered nontraditional roles for MMPs in the cytosol and nucleus. These have shed light on intracellular targets and biological actions of MMPs, adding additional layers of complexity for therapeutic MMP inhibition. In this review, we discussed the recent advances made in understanding nuclear location of MMPs, their regulation of intranuclear sorting, and their intranuclear proteolytic activity and substrates. In particular, we highlighted the roles of intranuclear MMPs in oxidative DNA damage, neuronal apoptosis, and neuroinflammation at an early stage of stroke insult. These novel data point to new putative MMP-mediated intranuclear actions in stroke-induced pathological processes and may lead to novel approaches to treatment of stroke and other neurological diseases.

  13. Oxidative DNA Damage Mediated by Intranuclear MMP Activity Is Associated with Neuronal Apoptosis in Ischemic Stroke.

    Science.gov (United States)

    Kimura-Ohba, Shihoko; Yang, Yi

    2016-01-01

    Evidence of the pathological roles of matrix metalloproteinases (MMPs) in various neurological disorders has made them attractive therapeutic targets. MMPs disrupt the blood-brain barrier and cause neuronal death and neuroinflammation in acute cerebral ischemia and are critical for angiogenesis during recovery. However, some challenges have to be overcome before MMPs can be further validated as drug targets in stroke injury. Identifying in vivo substrates of MMPs should greatly improve our understanding of the mechanisms of ischemic injury and is critical for providing more precise drug targets. Recent works have uncovered nontraditional roles for MMPs in the cytosol and nucleus. These have shed light on intracellular targets and biological actions of MMPs, adding additional layers of complexity for therapeutic MMP inhibition. In this review, we discussed the recent advances made in understanding nuclear location of MMPs, their regulation of intranuclear sorting, and their intranuclear proteolytic activity and substrates. In particular, we highlighted the roles of intranuclear MMPs in oxidative DNA damage, neuronal apoptosis, and neuroinflammation at an early stage of stroke insult. These novel data point to new putative MMP-mediated intranuclear actions in stroke-induced pathological processes and may lead to novel approaches to treatment of stroke and other neurological diseases.

  14. Neuroprotective effect of ebselen against intracerebroventricular streptozotocin-induced neuronal apoptosis and oxidative stress in rats.

    Science.gov (United States)

    Unsal, Cuneyt; Oran, Mustafa; Albayrak, Yakup; Aktas, Cevat; Erboga, Mustafa; Topcu, Birol; Uygur, Ramazan; Tulubas, Feti; Yanartas, Omer; Ates, Ozkan; Ozen, Oguz Aslan

    2016-04-01

    The goal of this study was to examine the neuroprotective effect of ebselen against intracerebroventricular streptozotocin (ICV-STZ)-induced oxidative stress and neuronal apoptosis in rat brain. A total of 30 adult male Sprague-Dawley rats were randomly divided into 3 groups of 10 animals each: control, ICV-STZ, and ICV-STZ treated with ebselen. The ICV-STZ group rats were injected bilaterally with ICV-STZ (3 mg/kg) on days 1 and 3, and ebselen (10 mg/kg/day) was administered for 14 days starting from 1st day of ICV-STZ injection to day 14. Rats were killed at the end of the study and brain tissues were removed for biochemical and histopathological investigation. Our results demonstrated, for the first time, the neuroprotective effect of ebselen on Alzheimer's disease (AD) model in rats. Our present study, in ICV-STZ group, showed significant increase in tissue malondialdehyde levels and significant decrease in enzymatic antioxidants superoxide dismutase and glutathione peroxidase in the frontal cortex tissue. The histopathological studies in the brain of rats also supported that ebselen markedly reduced the ICV-STZ-induced histopathological changes and well preserved the normal histological architecture of the frontal cortex tissue. The number of apoptotic neurons was increased in frontal cortex tissue after ICV-STZ administration. Treatment of ebselen markedly reduced the number of degenerating apoptotic neurons. The study demonstrates the effectiveness of ebselen, as a powerful antioxidant, in preventing the oxidative damage and morphological changes caused by ICV-STZ in rats. Thus, ebselen may have a therapeutic value for the treatment of AD.

  15. Effect of nerve growth factor on neuronal apoptosis after spinal cord injury in rats

    Institute of Scientific and Technical Information of China (English)

    曹晓建; 汤长华; 罗永湘

    2002-01-01

    To explore the molecular mechanism of the protective effect of nerve growth factor (NGF) on injured spinal cord. Methods: The posterior T8 (the 8th thoracic segment) spinal cords of 60 Wistar rats were injured by impacts caused by objects (weighing 10 g) falling from a height of 2.5 cm with Allens way. Solution with nerve growth factors (NGF) was given to 30 rats (the NGF group) through a microtubule inserted into the subarachnoid cavity immediately, and at 2, 4, 8, 12 and 24 hours after spinal cord injury (SCI) respectively. Normal saline (NS) with same volume was given to the other 30 rats (the NS group) with the same method. And 5 normal rats were taken as the normal controls. The expression of bcl-2 and bax proteins in spinal cord was detected with immunohistochemistry. The apoptotic neurons in spinal cord were measured with terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling of DNA fragments (TUNEL) staining. Results: The positive expression of bcl-2 protein was strong in the normal controls, but decreased in the NS group, and increased significantly in the NGF group as compared with that of the NS group (P<0.01). The positive expression of bax protein was also strong in the normal controls, but increased in the NS group, and decreased significantly in the NGF group as compared with that of the NS group (P<0.01). Apoptotic neurons were found in the NS group, and they decreased significantly in the NGF group as compared with that of the NS group (P<0.01). Conclusions: NGF can protect the injured nerve tissues through stimulating the expression of bcl-2 protein, inhibiting the expression of bax protein and inhibiting the neuronal apoptosis after SCI.

  16. Clinical significance of metallothioneins in cell therapy and nanomedicine

    Directory of Open Access Journals (Sweden)

    Sharma S

    2013-04-01

    Full Text Available Sushil Sharma,1 Afsha Rais,1 Ranbir Sandhu,1 Wynand Nel,1 Manuchair Ebadi21Saint James School of Medicine, Bonaire, The Netherlands; 2Department of Pharmacology, Physiology, and Therapeutics, Center of Excellence in Neuroscience, University of North Dakota School of Medicine and Health Sciences, Grand Forks, ND, USAAbstract: Mammalian metallothioneins (MTs are low molecular weight (6–7 kDa cysteine-rich proteins that are specifically induced by metal nanoparticles (NPs. MT induction in cell therapy may provide better protection by serving as antioxidant, anti-inflammatory, antiapoptotic agents, and by augmenting zinc-mediated transcriptional regulation of genes involved in cell proliferation and differentiation. Liposome-encapsulated MT-1 promoter has been used extensively to induce growth hormone or other genes in culture and gene-manipulated animals. MTs are induced as a defensive mechanism in chronic inflammatory conditions including neurodegenerative diseases, cardiovascular diseases, cancer, and infections, hence can serve as early and sensitive biomarkers of environmental safety and effectiveness of newly developed NPs for clinical applications. Microarray analysis has indicated that MTs are significantly induced in drug resistant cancers and during radiation treatment. Nutritional stress and environmental toxins (eg, kainic acid and domoic acid induce MTs and aggregation of multilamellar electron-dense membrane stacks (Charnoly body due to mitochondrial degeneration. MTs enhance mitochondrial bioenergetics of reduced nicotinamide adenine dinucleotide–ubiquinone oxidoreductase (complex-1, a rate-limiting enzyme complex involved in the oxidative phosphorylation. Monoamine oxidase-B inhibitors (eg, selegiline inhibit α-synuclein nitration, implicated in Lewy body formation, and inhibit 1-methyl 4-phenylpyridinium and 3-morpholinosydnonimine-induced apoptosis in cultured human dopaminergic neurons and mesencephalic fetal stem cells. MTs

  17. Metallothionein:An overview

    Institute of Scientific and Technical Information of China (English)

    N Thirumoorthy; KT Manisenthil Kumar; A Shyam Sundar; L Panayappan; Malay Chatterjee

    2007-01-01

    Metallothioneins(MTs)were discovered in 1957 by Margoshes and Vallee and identified as low-molecular weight and sulphydryl rich proteins.It is not surprising that most mammalian tissues contain age related basal levels of MTs since they are involved in metalloregulatory processes that include cell growth and multiplication.In an effort to understand the biology of this intriguing tumor,various biomarkers such as oncogenes,p53 tumor suppressor gene,waf 1 protein,proliferating cell nuclear antigen,telomerase,microsatellite markers and cytogenetic changes have been examined.One biomarker which has recently shown to be expressed in various human tumors but still less reported in carcinoma is MT.Immunohistochemical detection of MT proteins in cold acetone-fixed paraffin embedded liver sections was performed by the streptavidin-avidin-biotin immunoperoxidase complex method.

  18. Exposure to cell phone radiation up-regulates apoptosis genes in primary cultures of neurons and astrocytes.

    Science.gov (United States)

    Zhao, Tian-Yong; Zou, Shi-Ping; Knapp, Pamela E

    2007-01-22

    The health effects of cell phone radiation exposure are a growing public concern. This study investigated whether expression of genes related to cell death pathways are dysregulated in primary cultured neurons and astrocytes by exposure to a working Global System for Mobile Communication (GSM) cell phone rated at a frequency of 1900MHz. Primary cultures were exposed to cell phone emissions for 2h. We used array analysis and real-time RT-PCR to show up-regulation of caspase-2, caspase-6 and Asc (apoptosis associated speck-like protein containing a card) gene expression in neurons and astrocytes. Up-regulation occurred in both "on" and "stand-by" modes in neurons, but only in "on" mode in astrocytes. Additionally, astrocytes showed up-regulation of the Bax gene. The effects are specific since up-regulation was not seen for other genes associated with apoptosis, such as caspase-9 in either neurons or astrocytes, or Bax in neurons. The results show that even relatively short-term exposure to cell phone radiofrequency emissions can up-regulate elements of apoptotic pathways in cells derived from the brain, and that neurons appear to be more sensitive to this effect than astrocytes.

  19. Recombinant human brain-derived neurotrophic factor prevents neuronal apoptosis in a novel in vitro model of subarachnoid hemorrhage.

    Science.gov (United States)

    Li, Mingchang; Wang, Yuefei; Wang, Wei; Zou, Changlin; Wang, Xin; Chen, Qianxue

    2017-01-01

    Subarachnoid hemorrhage (SAH) is a hemorrhagic stroke with high mortality and morbidity. An animal model for SAH was established by directly injecting a hemolysate into the subarachnoid space of rats or mice. However, the in vitro applications of the hemolysate SAH model have not been reported, and the mechanisms remain unclear. In this study, we established an in vitro SAH model by treating cortical pyramidal neurons with hemolysate. Using this model, we assessed the effects of recombinant human brain-derived neurotrophic factor (rhBDNF) on hemolysate-induced cell death and related mechanisms. Cortical neurons were treated with 10 ng/mL or 100 ng/mL rhBDNF prior to application of hemolysate. Hemolysate treatment markedly increased cell loss, triggered apoptosis, and promoted the expression of caspase-8, caspase-9, and cleaved caspase-3. rhBDNF significantly inhibited hemolysate-induced cell loss, neuronal apoptosis, and expression of caspase-8, caspase-9, and cleaved caspase-3. Our data revealed a previously unrecognized protective activity of rhBDNF against hemolysate-induced cell death, potentially via regulation of caspase-9-, caspase-8-, and cleaved caspase-3-related apoptosis. This study implicates that hemolysate-induced cortical neuron death represents an important in vitro model of SAH.

  20. Lychee Seed Saponins Improve Cognitive Function and Prevent Neuronal Injury via Inhibiting Neuronal Apoptosis in a Rat Model of Alzheimer’s Disease

    Science.gov (United States)

    Wang, Xiuling; Wu, Jianming; Yu, Chonglin; Tang, Yong; Liu, Jian; Chen, Haixia; Jin, Bingjin; Mei, Qibing; Cao, Shousong; Qin, Dalian

    2017-01-01

    Lychee seed is a traditional Chinese medicine and possesses many activities, including hypoglycemia, liver protection, antioxidation, antivirus, and antitumor. However, its effect on neuroprotection is still unclear. The present study investigated the effects of lychee seed saponins (LSS) on neuroprotection and associated mechanisms. We established a rat model of Alzheimer’s disease (AD) by injecting Aβ25–35 into the lateral ventricle of rats and evaluated the effect of LSS on spatial learning and memory ability via the Morris water maze. Neuronal apoptosis was analyzed by hematoxylin and eosin stain and terminal deoxynucleotidyl transferase (Tdt)-mediated dUTP nick-end labeling analysis, and mRNA expression of caspase-3 and protein expressions of Bax and Bcl-2 by reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting, respectively. The results showed that LSS remarkably improved cognitive function and alleviated neuronal injury by inhibiting apoptosis in the hippocampus of AD rats. Furthermore, the mRNA expression of caspase-3 and the protein expression of Bax were downregulated, while the protein expression of Bcl-2 and the ratio of Bcl-2/Bax were increased by LSS. We demonstrate that LSS significantly improves cognitive function and prevent neuronal injury in the AD rats via regulation of the apoptosis pathway. Therefore, LSS may be developed as a nutritional supplement and sold as a drug for AD prevention and/or treatment. PMID:28165366

  1. Lychee Seed Saponins Improve Cognitive Function and Prevent Neuronal Injury via Inhibiting Neuronal Apoptosis in a Rat Model of Alzheimer’s Disease

    Directory of Open Access Journals (Sweden)

    Xiuling Wang

    2017-02-01

    Full Text Available Lychee seed is a traditional Chinese medicine and possesses many activities, including hypoglycemia, liver protection, antioxidation, antivirus, and antitumor. However, its effect on neuroprotection is still unclear. The present study investigated the effects of lychee seed saponins (LSS on neuroprotection and associated mechanisms. We established a rat model of Alzheimer’s disease (AD by injecting Aβ25–35 into the lateral ventricle of rats and evaluated the effect of LSS on spatial learning and memory ability via the Morris water maze. Neuronal apoptosis was analyzed by hematoxylin and eosin stain and terminal deoxynucleotidyl transferase (Tdt-mediated dUTP nick-end labeling analysis, and mRNA expression of caspase-3 and protein expressions of Bax and Bcl-2 by reverse transcription-polymerase chain reaction (RT-PCR and Western blotting, respectively. The results showed that LSS remarkably improved cognitive function and alleviated neuronal injury by inhibiting apoptosis in the hippocampus of AD rats. Furthermore, the mRNA expression of caspase-3 and the protein expression of Bax were downregulated, while the protein expression of Bcl-2 and the ratio of Bcl-2/Bax were increased by LSS. We demonstrate that LSS significantly improves cognitive function and prevent neuronal injury in the AD rats via regulation of the apoptosis pathway. Therefore, LSS may be developed as a nutritional supplement and sold as a drug for AD prevention and/or treatment.

  2. 14,15-EET promotes mitochondrial biogenesis and protects cortical neurons against oxygen/glucose deprivation-induced apoptosis.

    Science.gov (United States)

    Wang, Lai; Chen, Man; Yuan, Lin; Xiang, Yuting; Zheng, Ruimao; Zhu, Shigong

    2014-07-18

    14,15-Epoxyeicosatrienoic acid (14,15-EET), a metabolite of arachidonic acid, is enriched in the brain cortex and exerts protective effect against neuronal apoptosis induced by ischemia/reperfusion. Although apoptosis has been well recognized to be closely associated with mitochondrial biogenesis and function, it is still unclear whether the neuroprotective effect of 14,15-EET is mediated by promotion of mitochondrial biogenesis and function in cortical neurons under the condition of oxygen-glucose deprivation (OGD). In this study, we found that 14,15-EET improved cell viability and inhibited apoptosis of cortical neurons. 14,15-EET significantly increased the mitochondrial mass and the ratio of mitochondrial DNA to nuclear DNA. Key makers of mitochondrial biogenesis, peroxisome proliferator activator receptor gamma-coactivator 1 alpha (PGC-1α), nuclear respiratory factor 1 (NRF-1) and mitochondrial transcription factor A (TFAM), were elevated at both mRNA and protein levels in the cortical neurons treated with 14,15-EET. Moreover, 14,15-EET markedly attenuated the decline of mitochondrial membrane potential, reduced ROS, while increased ATP synthesis. Knockdown of cAMP-response element binding protein (CREB) by siRNA blunted the up-regulation of PGC-1α and NRF-1 stimulated by 14,15-EET, and consequently abolished the neuroprotective effect of 14,15-EET. Our results indicate that 14,15-EET protects neurons from OGD-induced apoptosis by promoting mitochondrial biogenesis and function through CREB mediated activation of PGC-1α and NRF-1.

  3. Apoptosis neuronal: la diversidad de señales y de tipos celulares

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    Lina Vanessa Becerra

    2009-03-01

    Full Text Available La muerte celular programada es un evento fisiológico durante el desarrollo. En el encéfalo y la médula espinal, este proceso determina el número y la localización de los diferentes tipos celulares. En el sistema nervioso del adulto, la muerte celular programada o apoptosis está más restringida, pero puede jugar un papel determinante en enfermedades crónicas o agudas. Al contrario de otros tejidos en los cuales la apoptosis está documentada ampliamente desde el punto de vista morfológico, en el sistema nervioso central la evidencia en este sentido es escasa. A pesar de esto, existe consenso acerca de la activación de diferentes sistemas de señalización apoptótica. En el presente artículo se intenta resumir las principales vías de señalización apoptótica identificadas en el tejido nervioso. Considerando que las vías apoptóticas son múltiples, los tipos neuronales diversos y especializados y que la respuesta neuronal a la lesión y la supervivencia dependen del contexto de la célula en el tejido (preservación de la conectividad, integridad glial y matriz extracelular, flujo sanguíneo y disponibilidad de factores tróficos, lo que es relevante en el proceso apoptótico en un sector del cerebro puede no serlo en otro.

  4. Inhibition of apoptosis blocks human motor neuron cell death in a stem cell model of spinal muscular atrophy.

    Directory of Open Access Journals (Sweden)

    Dhruv Sareen

    Full Text Available Spinal muscular atrophy (SMA is a genetic disorder caused by a deletion of the survival motor neuron 1 gene leading to motor neuron loss, muscle atrophy, paralysis, and death. We show here that induced pluripotent stem cell (iPSC lines generated from two Type I SMA subjects-one produced with lentiviral constructs and the second using a virus-free plasmid-based approach-recapitulate the disease phenotype and generate significantly fewer motor neurons at later developmental time periods in culture compared to two separate control subject iPSC lines. During motor neuron development, both SMA lines showed an increase in Fas ligand-mediated apoptosis and increased caspase-8 and-3 activation. Importantly, this could be mitigated by addition of either a Fas blocking antibody or a caspase-3 inhibitor. Together, these data further validate this human stem cell model of SMA, suggesting that specific inhibitors of apoptotic pathways may be beneficial for patients.

  5. Osthole promotes neuronal differentiation and inhibits apoptosis via Wnt/β-catenin signaling in an Alzheimer's disease model.

    Science.gov (United States)

    Yao, Yingjia; Gao, Zhong; Liang, Wenbo; Kong, Liang; Jiao, Yanan; Li, Shaoheng; Tao, Zhenyu; Yan, Yuhui; Yang, Jingxian

    2015-12-15

    Neurogenesis is the process by which neural stem cells (NSCs) proliferate and differentiate into neurons. This is diminished in several neurodegenerative disorders such as Alzheimer's disease (AD), which is characterized by the deposition of amyloid (A)β peptides and neuronal loss. Stimulating NSCs to replace lost neurons is therefore a promising approach for AD treatment. Our previous study demonstrated that osthole modulates NSC proliferation and differentiation, and may reduce Aβ protein expression in nerve cells. Here we investigated the mechanism underlying the effects of osthole on NSCs. We found that osthole enhances NSC proliferation and neuronal differentiation while suppressing apoptosis, effects that were exerted via activation of Wnt/β-catenin signaling. These results provide evidence that osthole can potentially be used as a therapeutic agent in the treatment of AD and other neurodegenerative disorders.

  6. Mutant presenilin2 promotes apoptosis through the p53/miR-34a axis in neuronal cells.

    Science.gov (United States)

    Li, Liu-Hong; Tu, Qiu-Yun; Deng, Xiao-Hua; Xia, Jian; Hou, De-Ren; Guo, Ke; Zi, Xiao-Hong

    2017-02-08

    Neurodegenerative disorders have attracted attention in last decades due to their high incidence in the world. The p53/miR-34a axis triggers apoptosis and suppresses viability in multiple types of cells, but little is known about its role in neurodegenerative diseases. In this study, we showed that presenilin (PS)-2, a major gene associated with familial Alzheimer's disease (AD) could trigger the apoptosis through the p53/miR-34a axis in PC12 cells. First we found that PC12 cell viability was downregulated by PS-2 and mutant PS-2 overexpression, especially by mutant PS-2 overexpression. Then, we established a mutant PS-2-overexpressing PC12 cell line and confirmed that mutant PS-2 induced not only p53 but also miR-34a expression. The transfection of miR-34a inhibitor reversed PS-2-induced effects on cellular viability and apoptosis. Mutant PS-2 overexpression promoted caspase-3 expression, reduced Sirt1 and Bcl-2 expression, all of which were miR-34a downstream genes related with cell apoptosis. Moreover, mutant PS-2 also activated the p53/miR-34a axis and induced apoptosis in AD transgenic mice brain. These results implied that mutant PS-2 might promote the apoptosis of neuronal cells through triggering the p53/miR-34a axis. Altogether our results provide a novel insight into neurodegenerative disease and deepen our understandings of AD pathogenic processes.

  7. Role of metallothionein-III following central nervous system damage

    DEFF Research Database (Denmark)

    Carrasco, Javier; Penkowa, Milena; Giralt, Mercedes

    2003-01-01

    We evaluated the physiological relevance of metallothionein-III (MT-III) in the central nervous system following damage caused by a focal cryolesion onto the cortex by studying Mt3-null mice. In normal mice, dramatic astrogliosis and microgliosis and T-cell infiltration were observed in the area...... the inflammatory response elicited in the central nervous system by a cryoinjury, nor does it serve an important antioxidant role, but it may influence neuronal regeneration during the recovery process....

  8. Recombinant human erythropoietin increases survival and reduces neuronal apoptosis in a murine model of cerebral malaria

    Directory of Open Access Journals (Sweden)

    Hempel Casper

    2008-01-01

    Full Text Available Abstract Background Cerebral malaria (CM is an acute encephalopathy with increased pro-inflammatory cytokines, sequestration of parasitized erythrocytes and localized ischaemia. In children CM induces cognitive impairment in about 10% of the survivors. Erythropoietin (Epo has – besides of its well known haematopoietic properties – significant anti-inflammatory, antioxidant and anti-apoptotic effects in various brain disorders. The neurobiological responses to exogenously injected Epo during murine CM were examined. Methods Female C57BL/6j mice (4–6 weeks, infected with Plasmodium berghei ANKA, were treated with recombinant human Epo (rhEpo; 50–5000 U/kg/OD, i.p. at different time points. The effect on survival was measured. Brain pathology was investigated by TUNEL (Terminal deoxynucleotidyl transferase (TdT-mediated deoxyuridine triphosphate (dUTP-digoxigenin nick end labelling, as a marker of apoptosis. Gene expression in brain tissue was measured by real time PCR. Results Treatment with rhEpo increased survival in mice with CM in a dose- and time-dependent manner and reduced apoptotic cell death of neurons as well as the expression of pro-inflammatory cytokines in the brain. This neuroprotective effect appeared to be independent of the haematopoietic effect. Conclusion These results and its excellent safety profile in humans makes rhEpo a potential candidate for adjunct treatment of CM.

  9. Dietary flavonoid fisetin regulates aluminium chloride-induced neuronal apoptosis in cortex and hippocampus of mice brain.

    Science.gov (United States)

    Prakash, Dharmalingam; Sudhandiran, Ganapasam

    2015-12-01

    Dietary flavonoids have been suggested to promote brain health by protecting brain parenchymal cells. Recently, understanding the possible mechanism underlying neuroprotective efficacy of flavonoids is of great interest. Given that fisetin exerts neuroprotection, we have examined the mechanisms underlying fisetin in regulating Aβ aggregation and neuronal apoptosis induced by aluminium chloride (AlCl3) administration in vivo. Male Swiss albino mice were induced orally with AlCl3 (200 mg/kg. b.wt./day/8 weeks). Fisetin (15 mg/Kg. b.wt. orally) was administered for 4 weeks before AlCl3-induction and administered simultaneously for 8 weeks during AlCl3-induction. We found aggregation of Amyloid beta (Aβ 40-42), elevated expressions of Apoptosis stimulating kinase (ASK-1), p-JNK (c-Jun N-terminal Kinase), p53, cytochrome c, caspases-9 and 3, with altered Bax/Bcl-2 ratio in favour of apoptosis in cortex and hippocampus of AlCl3-administered mice. Furthermore, TUNEL and fluoro-jade C staining demonstrate neurodegeneration in cortex and hippocampus. Notably, treatment with fisetin significantly (Pfisetin treatment. We have identified the involvement of fisetin in regulating ASK-1 and p-JNK as possible mediator of Aβ aggregation and subsequent neuronal apoptosis during AlCl3-induced neurodegeneration. These findings define the possibility that fisetin may slow or prevent neurodegneration and can be utilised as neuroprotective agent against Alzheimer's and Parkinson's disease. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. TL-2 attenuates β-amyloid induced neuronal apoptosis through the AKT/GSK-3β/β-catenin pathway.

    Science.gov (United States)

    Zhu, Xiaolei; Wang, Sulei; Yu, Linjie; Yang, Hui; Tan, Renxiang; Yin, Kailin; Jin, Jiali; Zhao, Hui; Guan, Dening; Xu, Yun

    2014-09-01

    β-amyloid (Aβ)-mediated neuronal apoptosis contributes to the progression of Alzheimer's disease (AD), although the exact mechanism remains unclear. This study aimed to investigate whether Dalesconol B (TL-2), a potent immunosuppressive agent with an unusual carbon skeleton, could inhibit Aβ-induced apoptosis in vitro and in vivo and to explore the underlying mechanisms. Aβ(1-42) was injected to bilateral hippocampus of mice to make the AD models in vivo. TL-2 was able to cross the blood-brain barrier and attenuate memory deficits in the AD mice. TL-2 also inhibited Aβ(1-42)-induced neuronal apoptosis in vitro and in vivo. In addition, TL-2 could activate the AKT/GSK-3β pathway, and inhibition of AKT and activation of GSK-3β partially eliminated the neuroprotective effects of TL-2. Furthermore, TL-2 induced the nuclear translocation of β-catenin and enhanced its transcriptional activity through the AKT/GSK-3β pathway to promote neuronal survival. These results suggest that TL-2 might be a potential drug for AD treatment.

  11. Microglia are mediators of Borrelia burgdorferi-induced apoptosis in SH-SY5Y neuronal cells.

    Directory of Open Access Journals (Sweden)

    Tereance A Myers

    2009-11-01

    Full Text Available Inflammation has long been implicated as a contributor to pathogenesis in many CNS illnesses, including Lyme neuroborreliosis. Borrelia burgdorferi is the spirochete that causes Lyme disease and it is known to potently induce the production of inflammatory mediators in a variety of cells. In experiments where B. burgdorferi was co-cultured in vitro with primary microglia, we observed robust expression and release of IL-6 and IL-8, CCL2 (MCP-1, CCL3 (MIP-1alpha, CCL4 (MIP-1beta and CCL5 (RANTES, but we detected no induction of microglial apoptosis. In contrast, SH-SY5Y (SY neuroblastoma cells co-cultured with B. burgdorferi expressed negligible amounts of inflammatory mediators and also remained resistant to apoptosis. When SY cells were co-cultured with microglia and B. burgdorferi, significant neuronal apoptosis consistently occurred. Confocal microscopy imaging of these cell cultures stained for apoptosis and with cell type-specific markers confirmed that it was predominantly the SY cells that were dying. Microarray analysis demonstrated an intense microglia-mediated inflammatory response to B. burgdorferi including up-regulation in gene transcripts for TLR-2 and NFkappabeta. Surprisingly, a pathway that exhibited profound changes in regard to inflammatory signaling was triggering receptor expressed on myeloid cells-1 (TREM1. Significant transcript alterations in essential p53 pathway genes also occurred in SY cells cultured in the presence of microglia and B. burgdorferi, which indicated a shift from cell survival to preparation for apoptosis when compared to SY cells cultured in the presence of B. burgdorferi alone. Taken together, these findings indicate that B. burgdorferi is not directly toxic to SY cells; rather, these cells become distressed and die in the inflammatory surroundings generated by microglia through a bystander effect. If, as we hypothesized, neuronal apoptosis is the key pathogenic event in Lyme neuroborreliosis, then

  12. Inhibition of Mitochondrial Clearance and Cu/Zn-SOD Activity Enhance 6-Hydroxydopamine-Induced Neuronal Apoptosis.

    Science.gov (United States)

    In, Sua; Hong, Chang-Won; Choi, Boyoung; Jang, Bong-Geum; Kim, Min-Ju

    2016-01-01

    Parkinson's disease (PD) is a common movement disorder among neurodegenerative diseases, involving neuronal cell death in the substantia nigra of the midbrain. Although mechanisms of cell death in PD have been studied, the exact molecular pathogenesis is still unclear. Here, we explore the relationship between two types of cell death, autophagy and apoptosis, which have been studied separately in parkinsonian mimetic model of 6-hydroxydopamine (6-OHDA). 6-OHDA induced autophagy firstly and then later inhibition of autophagy flux occurred with apoptosis. The apoptosis was prevented by treatment of pan-caspase inhibitor, zVAD-fmk (benzyloxycarbonyl-VAD-fluoromethylketone (zVAD)), or early phase inhibitor of autophagy, 3-methyladenine (3-MA), indicating that autophagic induction was followed by the apoptosis. Interestingly, late step inhibitor of autophagy, bafilomycin A1 (BafA), aggravated 6-OHDA-induced apoptosis. This was associated with mitochondrial abnormality such as the inhibition of damaged mitochondrial clearance and aberrant increase of extracellular oxygen consumption. Furthermore, treatment of BafA did not inhibit 6-OHDA-mediated superoxide formation but strongly reduced the hydrogen peroxide production to below basal levels, indicating failure from superoxide to hydrogen peroxide. These results were accompanied by a lowered expression and activity of copper/zinc superoxide dismutase (Cu/Zn-SOD) but not of manganese SOD (MnSOD) and catalase. Thus, the present study suggests that crosstalk among apoptosis, autophagy, and oxidative stress is a causative factor of 6-OHDA-induced neuronal death and provides a mechanistic understanding of PD pathogenesis.

  13. THE PROTECTIVE EFFECTS OF THE TOTAL SAPONIN OF DIPSACUS ASPEROIDES ON THE APOPTOSIS OF HIPPOCAMPAL NEURONS INDUCED BY β-AMYLOID PROTEIN

    Institute of Scientific and Technical Information of China (English)

    钱亦华; 杨杰; 胡海涛; 刘勇; 杨广德; 曹云新; 任惠民

    2004-01-01

    Objective To investigate the effects of the total saponin of Dipsacus asperoides (tSDA) and ginsenoside Rb1 (GRb1) on the apoptosis of primary cultured hippocampal neurons induced by β-amyloid protein (Aβ). Methods Primary cultured hippocampal neurons, the cultures were pretreated with tSDA and GRb1 on 10d for 24 hours respectively. Then the cultures were treated with 35μmol·L-1 Aβ25-35 for 24 hours, observed the changing of survival rate of neurons and the apoptosis of neurons with biochemical analysis combining immunofluorescent cytochemical double-staining technique. Results Hippocampal neurons were treated with 35μmol*L-1 Aβ for 24 hours, and survival rate of neurons downed to 52.6%. When neurons were pretreated by tSDA and GRb1, survival rate of neurons increased 11% to 15%. The findings of immunofluorescent cytochemical double-staining indicated that apoptotic neurons were obviously more than that of the blank group, reaching 43.9%.When neurons were pretreated by tSDA and GRb1, apoptotic neurons were downed to 16.6%, 10.8% respectively. Conclusion tSDA had the same effects as GRb1, protecting the neurons, antagonizing neurotoxicity of Aβ, increasing survival rate of neurons, and reducing apoptotic neurons induced by Aβ.

  14. Protective effect of nicotinamide adenine dinucleotide (NAD(+)) against spinal cord ischemia-reperfusion injury via reducing oxidative stress-induced neuronal apoptosis.

    Science.gov (United States)

    Xie, Lei; Wang, Zhenfei; Li, Changwei; Yang, Kai; Liang, Yu

    2017-02-01

    As previous studies demonstrate that oxidative stress and apoptosis play crucial roles in ischemic pathogenesis and nicotinamide adenine dinucleotide (NAD(+)) treatment attenuates oxidative stress-induced cell death among primary neurons and astrocytes as well as significantly reduce cerebral ischemic injury in rats. We used a spinal cord ischemia injury (SCII) model in rats to verify our hypothesis that NAD(+) could ameliorate oxidative stress-induced neuronal apoptosis. Adult male rats were subjected to transient spinal cord ischemia for 60min, and different doses of NAD(+) were administered intraperitoneally immediately after the start of reperfusion. Neurological function was determined by Basso, Beattie, Bresnahan (BBB) scores. The oxidative stress level was assessed by superoxide dismutase (SOD) activity and malondialdehyde (MDA) content. The degree of apoptosis was analyzed by deoxyuridinetriphosphate nick-end labeling (TUNEL) staining and protein levels of cleaved caspase-3 and AIF (apoptosis inducing factor). The results showed that NAD(+) at 50 or 100mg/kg significantly decreased the oxidative stress level and neuronal apoptosis in the spinal cord of ischemia-reperfusion rats compared with saline, as accompanied with the decreased oxidative stress, NAD(+) administration significantly restrained the neuronal apoptosis after ischemia injury while improved the neurological and motor function. These findings suggested that NAD(+) might protect against spinal cord ischemia-reperfusion via reducing oxidative stress-induced neuronal apoptosis.

  15. ELDEPRYL AND RILUZOLE INHIBIT 1-METHYL-4-PHENYL -1,2,3,6-TETRAHYDROPYRIDINE (MPTP)-INDUCED NIGRAL NEURONAL APOPTOSIS IN MICE

    Institute of Scientific and Technical Information of China (English)

    陈生弟; 郭明; 刘振国; 陈红专

    2002-01-01

    Objective To investigate the possible role of apoptosis in the pathogenesis of Parkinsons disease. Methods C57BL mice were treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP), and TUNEL and flow cytometry were employed to detect neuronal apoptosis in the substantia nigra. ResultsThe results of animal experiment demonstrated that the administration of MPTP 30mg/kg for 7d could induce neuronal apoptosis in the substantia nigra. The MPTP-induced nigral neuronal apoptosis could be completely prevented by pre-treatment of Eldepryl, an inhibitor of B typed monoamine oxidase (MAO-B);and partially protected by pre-treatment of Riluzole, an antagonist of excitatory amino acid receptors. Data of cell culture experiment showed that 20mmol 1-methyl-4-phenylpyridinium ion(MPP+) induced the apoptosis of pheochromocytoma(PC12 cells), whereas 20mmol MPTP did not cause PC12 cells apoptosis. Conclusion It is concluded that the apoptotic effect of MPTP in vivo on the nigral neurons may be mediated by its intermediate metabolite MPP+. The dopaminergic neuronal apoptosis in the substantia nigra may be a common pathway of various causes that lead to the onset of Parkinson's disease.

  16. Zeptomole electrochemical detection of metallothioneins.

    Directory of Open Access Journals (Sweden)

    Vojtech Adam

    Full Text Available BACKGROUND: Thiol-rich peptides and proteins possess a large number of biological activities and may serve as markers for numerous health problems including cancer. Metallothionein (MT, a small molecular mass protein rich in cysteine, may be considered as one of the promising tumour markers. The aim of this paper was to employ chronopotentiometric stripping analysis (CPSA for highly sensitive detection of MT. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we used adsorptive transfer stripping technique coupled with CPSA for detection of cysteine, glutathione oxidized and reduced, phytochelatin, bovine serum albumin, and metallothionein. Under the optimal conditions, we were able to estimate detection limits down to tens of fg per ml. Further, this method was applied to detect metallothioneins in blood serum obtained from patients with breast cancer and in neuroblastoma cells resistant and sensitive to cisplatin in order to show the possible role of metallothioneins in carcinogenesis. It was found that MT level in blood serum was almost twice higher as compared to the level determined in healthy individuals. CONCLUSIONS/SIGNIFICANCE: This paper brings unique results on the application of ultra-sensitive electroanalytical method for metallothionein detection. The detection limit and other analytical parameters are the best among the parameters of other techniques. In spite of the fact that the paper is mainly focused on metallothionein, it is worth mentioning that successful detection of other biologically important molecules is possible by this method. Coupling of this method with simple isolation methods such as antibody-modified paramagnetic particles may be implemented to lab-on-chip instrument.

  17. Ceramides in Alzheimer’s Disease: Key Mediators of Neuronal Apoptosis Induced by Oxidative Stress and Aβ Accumulation

    Directory of Open Access Journals (Sweden)

    Maja Jazvinšćak Jembrek

    2015-01-01

    Full Text Available Alzheimer’s disease (AD, the most common chronic and progressive neurodegenerative disorder, is characterized by extracellular deposits of amyloid β-peptides (Aβ and intracellular deposits of hyperphosphorylated tau (phospho-tau protein. Ceramides, the major molecules of sphingolipid metabolism and lipid second messengers, have been associated with AD progression and pathology via Aβ generation. Enhanced levels of ceramides directly increase Aβ through stabilization of β-secretase, the key enzyme in the amyloidogenic processing of Aβ precursor protein (APP. As a positive feedback loop, the generated oligomeric and fibrillar Aβ induces a further increase in ceramide levels by activating sphingomyelinases that catalyze the catabolic breakdown of sphingomyelin to ceramide. Evidence also supports important role of ceramides in neuronal apoptosis. Ceramides may initiate a cascade of biochemical alterations, which ultimately leads to neuronal death by diverse mechanisms, including depolarization and permeabilization of mitochondria, increased production of reactive oxygen species (ROS, cytochrome c release, Bcl-2 depletion, and caspase-3 activation, mainly by modulating intracellular signalling, particularly along the pathways related to Akt/PKB kinase and mitogen-activated protein kinases (MAPKs. This review summarizes recent findings related to the role of ceramides in oxidative stress-driven neuronal apoptosis and interplay with Aβ in the cascade of events ending in neuronal degeneration.

  18. Neuroprotective effects of sevoflurane against electromagnetic pulse-induced brain injury through inhibition of neuronal oxidative stress and apoptosis.

    Directory of Open Access Journals (Sweden)

    Bin Deng

    Full Text Available Electromagnetic pulse (EMP causes central nervous system damage and neurobehavioral disorders, and sevoflurane protects the brain from ischemic injury. We investigated the effects of sevoflurane on EMP-induced brain injury. Rats were exposed to EMP and immediately treated with sevoflurane. The protective effects of sevoflurane were assessed by Nissl staining, Fluoro-Jade C staining and electron microscopy. The neurobehavioral effects were assessed using the open-field test and the Morris water maze. Finally, primary cerebral cortical neurons were exposed to EMP and incubated with different concentration of sevoflurane. The cellular viability, lactate dehydrogenase (LDH release, superoxide dismutase (SOD activity and malondialdehyde (MDA level were assayed. TUNEL staining was performed, and the expression of apoptotic markers was determined. The cerebral cortexes of EMP-exposed rats presented neuronal abnormalities. Sevoflurane alleviated these effects, as well as the learning and memory deficits caused by EMP exposure. In vitro, cell viability was reduced and LDH release was increased after EMP exposure; treatment with sevoflurane ameliorated these effects. Additionally, sevoflurane increased SOD activity, decreased MDA levels and alleviated neuronal apoptosis by regulating the expression of cleaved caspase-3, Bax and Bcl-2. These findings demonstrate that Sevoflurane conferred neuroprotective effects against EMP radiation-induced brain damage by inhibiting neuronal oxidative stress and apoptosis.

  19. Effects of prolonged abstinence from METH on the hippocampal BDNF levels, neuronal numbers and apoptosis in methamphetamine-sensitized rats.

    Science.gov (United States)

    Hajheidari, Samira; Sameni, Hamid Reza; Bandegi, Ahmad Reza; Miladi-Gorji, Hossein

    2017-04-03

    Methamphetamine (METH) use is associated with neuronal damage in various regions of brain, while effects of prolonged abstinence on METH-induced damage are not quite clear. This study evaluated serum and hippocampal BDNF levels, neuronal numbers and apoptosis in METH-sensitized and abstinent rats. Rats were sensitized to METH (2mg/kg, daily/18 days, s.c.). All rats were evaluated for neuron counting, the TUNEL test and serum and hippocampal BDNF levels after 30 days of forced abstinence from METH. The results showed that increased BDNF levels in the hippocampus and serum of METH-sensitized rats returned to control level after 30 days of abstinence. The number of neurons in the DG and CA1 of hippocampus and also, the total hippocampal perimeter and area in METH-sensitized rats were significantly lower than the saline rats. While, the number of neurons was not significantly increased in the hippocampus after prolonged abstinence from METH. Also, METH-sensitized rats showed a significant increase in TUNEL-positive cells, whereas METH-abstinent rats showed a slight but significant decrease in TUNEL-positive cells in the DG and CA3 of hippocampus. These results suggest that despite the reduction in BDNF levels, reducing the number of neurons, perimeter and area of the hippocampus were stable after abstinence. Thus, the degenerative effects of METH have been sustained even after prolonged abstinence in the hippocampus.

  20. Mutations in CRADD Result in Reduced Caspase-2-Mediated Neuronal Apoptosis and Cause Megalencephaly with a Rare Lissencephaly Variant.

    Science.gov (United States)

    Di Donato, Nataliya; Jean, Ying Y; Maga, A Murat; Krewson, Briana D; Shupp, Alison B; Avrutsky, Maria I; Roy, Achira; Collins, Sarah; Olds, Carissa; Willert, Rebecca A; Czaja, Agnieszka M; Johnson, Rachel; Stover, Jessi A; Gottlieb, Steven; Bartholdi, Deborah; Rauch, Anita; Goldstein, Amy; Boyd-Kyle, Victoria; Aldinger, Kimberly A; Mirzaa, Ghayda M; Nissen, Anke; Brigatti, Karlla W; Puffenberger, Erik G; Millen, Kathleen J; Strauss, Kevin A; Dobyns, William B; Troy, Carol M; Jinks, Robert N

    2016-11-03

    Lissencephaly is a malformation of cortical development typically caused by deficient neuronal migration resulting in cortical thickening and reduced gyration. Here we describe a "thin" lissencephaly (TLIS) variant characterized by megalencephaly, frontal predominant pachygyria, intellectual disability, and seizures. Trio-based whole-exome sequencing and targeted re-sequencing identified recessive mutations of CRADD in six individuals with TLIS from four unrelated families of diverse ethnic backgrounds. CRADD (also known as RAIDD) is a death-domain-containing adaptor protein that oligomerizes with PIDD and caspase-2 to initiate apoptosis. TLIS variants cluster in the CRADD death domain, a platform for interaction with other death-domain-containing proteins including PIDD. Although caspase-2 is expressed in the developing mammalian brain, little is known about its role in cortical development. CRADD/caspase-2 signaling is implicated in neurotrophic factor withdrawal- and amyloid-β-induced dendritic spine collapse and neuronal apoptosis, suggesting a role in cortical sculpting and plasticity. TLIS-associated CRADD variants do not disrupt interactions with caspase-2 or PIDD in co-immunoprecipitation assays, but still abolish CRADD's ability to activate caspase-2, resulting in reduced neuronal apoptosis in vitro. Homozygous Cradd knockout mice display megalencephaly and seizures without obvious defects in cortical lamination, supporting a role for CRADD/caspase-2 signaling in mammalian brain development. Megalencephaly and lissencephaly associated with defective programmed cell death from loss of CRADD function in humans implicate reduced apoptosis as an important pathophysiological mechanism of cortical malformation. Our data suggest that CRADD/caspase-2 signaling is critical for normal gyration of the developing human neocortex and for normal cognitive ability. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  1. Levodopa activates apoptosis signaling kinase 1 (ASK1) and promotes apoptosis in a neuronal model: implications for the treatment of Parkinson's disease.

    Science.gov (United States)

    Liedhegner, Elizabeth A Sabens; Steller, Kelly M; Mieyal, John J

    2011-10-17

    Oxidative stress is implicated in the etiology of Parkinson's disease (PD), the second most common neurodegenerative disease. PD is treated with chronic administration of l-3,4-dihydroxyphenylalanine (levodopa, L-DOPA), and typically, increasing doses are used during progression of the disease. Paradoxically, L-DOPA is a pro-oxidant and induces cell death in cellular models of PD through disruption of sulfhydryl homeostasis involving loss of the thiol-disulfide oxidoreductase functions of the glutaredoxin (Grx1) and thioredoxin (Trx1) enzyme systems [Sabens, E. A., Distler, A. M., and Mieyal, J. J. (2010) Biochemistry 49 (12), 2715-2724]. Considering this loss of both Grx1 and Trx1 activities upon L-DOPA treatment, we sought to elucidate the mechanism(s) of L-DOPA-induced apoptosis. In other contexts, both the NFκB (nuclear factor κB) pathway and the ASK1 (apoptosis signaling kinase 1) pathway have been shown to be regulated by both Grx1 and Trx1, and both pathways have been implicated in cell death signaling in model systems of PD. Moreover, mixed lineage kinase (MLK) has been considered as a potential therapeutic target for PD. Using SHSY5Y cells as model dopaminergic neurons, we found that NFκB activity was not altered by L-DOPA treatment, and the selective MLK inhibitor (CEP-1347) did not protect the cells from L-DOPA. In contrast, ASK1 was activated with L-DOPA treatment as indicated by phosphorylation of its downstream mitogen-activated protein kinases (MAPK), p38 and JNK. Chemical inhibition of either p38 or JNK provided protection from L-DOPA-induced apoptosis. Moreover, direct knockdown of ASK1 protected from L-DOPA-induced neuronal cell death. These results identify ASK1 as the main pro-apoptotic pathway activated in response to L-DOPA treatment, implicating it as a potential target for adjunct therapy in PD.

  2. Bovine Herpes Virus 1 (BHV-1) and Herpes Simplex Virus Type 1 (HSV-1) Promote Survival of Latently Infected Sensory Neurons, in Part by Inhibiting Apoptosis

    Science.gov (United States)

    Jones, Clinton

    2013-01-01

    α-Herpesvirinae subfamily members, including herpes simplex virus type 1 (HSV-1) and bovine herpes virus 1 (BHV-1), initiate infection in mucosal surfaces. BHV-1 and HSV-1 enter sensory neurons by cell-cell spread where a burst of viral gene expression occurs. When compared to non-neuronal cells, viral gene expression is quickly extinguished in sensory neurons resulting in neuronal survival and latency. The HSV-1 latency associated transcript (LAT), which is abundantly expressed in latently infected neurons, inhibits apoptosis, viral transcription, and productive infection, and directly or indirectly enhances reactivation from latency in small animal models. Three anti-apoptosis genes can be substituted for LAT, which will restore wild type levels of reactivation from latency to a LAT null mutant virus. Two small non-coding RNAs encoded by LAT possess anti-apoptosis functions in transfected cells. The BHV-1 latency related RNA (LR-RNA), like LAT, is abundantly expressed during latency. The LR-RNA encodes a protein (ORF2) and two microRNAs that are expressed in certain latently infected neurons. Wild-type expression of LR gene products is required for stress-induced reactivation from latency in cattle. ORF2 has anti-apoptosis functions and interacts with certain cellular transcription factors that stimulate viral transcription and productive infection. ORF2 is predicted to promote survival of infected neurons by inhibiting apoptosis and sequestering cellular transcription factors which stimulate productive infection. In addition, the LR encoded microRNAs inhibit viral transcription and apoptosis. In summary, the ability of BHV-1 and HSV-1 to interfere with apoptosis and productive infection in sensory neurons is crucial for the life-long latency-reactivation cycle in their respective hosts. PMID:25278776

  3. Hyperlipidemia exacerbates cerebral injury through oxidative stress, inflammation and neuronal apoptosis in MCAO/reperfusion rats.

    Science.gov (United States)

    Cao, Xiao-Lu; Du, Jing; Zhang, Ying; Yan, Jing-Ting; Hu, Xia-Min

    2015-10-01

    stress, inflammation and neuronal apoptosis by coexistence of hyperlipidemia and cerebral I/R.

  4. Up-regulation of GBP2 is Associated with Neuronal Apoptosis in Rat Brain Cortex Following Traumatic Brain Injury.

    Science.gov (United States)

    Miao, Qi; Ge, Meihong; Huang, Lili

    2017-02-27

    Guanylate binding protein 2 (GBP2) is one member of GBP family. Recently, GBP2 has been proposed to be a novel target of anti-cancer drugs. However, the role of GBP2 in the traumatic brain injury (TBI) is very limited. In this study, we sought to define GBP2's role in brain injury. GBP2 protein levels were significantly increased in the brain 3 days after injury, suggesting a functional role for GBP2 in TBI. Neuronal cells overexpressing GBP2 exhibited up-regulation of co-location of GBP2 and NeuN following TBI, suggesting that GBP2 potentiates the neuron apoptosis. To confirm the role of GBP2 in neuron apoptosis process, we employed a highly potent inhibitor of GBP2 (GBP2 RNAi). In H2O2-stimulated PC12 cells, in vitro blockade of GBP2 activity using GBP2 RNAi markedly attenuated the neuron apoptosis number. GBP2 RNAi also inhibited the expression levels of active caspase3 and p-Stat1. Furthermore, we found the expression of p-Stat1 in line with GBP2 and GBP2 interacted with p-Stat1 following TBI. The Jak2 inhibitor, AG490 inhibited this interaction and decreased the active caspase3 expression as well as promoted the functional recovery. Taken together, these data suggest that GBP2 RNAi has a protective effect in a rat TBI. This study demonstrates that GBP2 is an important positive regulator of TBI and is a promising therapeutic target for brain injury.

  5. Effect of ketamine on aquaporin-4 expression and neuronal apoptosis in brain tissues following brain injury in rats

    Institute of Scientific and Technical Information of China (English)

    Zangong Zhou; Xiangyu Ji; Li Song; Jianfang Song; Shiduan Wang; Yanwei Yin

    2006-01-01

    BACKGROUND: Aquaporin-4 (AQP-4) is closely related to the formation of brain edema. Neuronal apoptosis plays an important part in the conversion of swelled neuron following traumatic brain injury. At present, the studies on the protective effect of ketamine on brain have involved in its effect on aquaporin-4 expression and neuronal apoptosis in the brain tissues following brain injury in rats.OBJECTIVE: To observe the effect of ketamine on AQP-4 expression and neuronal apoptosis in the brain tissue following rat brain injury, and analyze the time-dependence of ketamine in the treatment of brain injury.DESIGN: Randomized grouping design, controlled animal trial.SETTING: Department of Anesthesiology, the Medical School Hospital of Qingdao University.MATERIALS: Totally 150 rats of clean grade, aged 3 months, were involved and randomized into control group and ketamine-treated group, with 75 rats in each. Each group was divided into 5 subgroups separately at 6,12, 24, 48 and 72 hours after injury, with 15 rats at each time point. Main instruments and reagents:homemade beat machine, ketamine hydrochloride (Hengrui Pharmaceutical Factory, Jiangsu), rabbit anti-rat AQP-4 polyclonal antibody, SABC immunohistochemical reagent kit and TUNEL reagent kit (Boster Co.,Ltd.,Wuhan).METHODS: This trial was carried out in the Institute of Cerebrovascular Disease, Medical College of Qingdao University during March 2005 to February 2006. A weight-dropping rat model of brain injury was created with Feeney method. The rats in the ketamine-treated group were intraperitoneally administered with 50 g/L ketamine (120 mg/kg) one hour after injury, but ketamine was replaced by normal saline in the control group. In each subgroup, the water content of cerebral hemisphere was measured in 5 rats chosen randomly. The left 10 rats in each subgroup were transcardiacally perfused with ketamine, then the brain tissue was made into paraffin sections and stained by haematoxylin and eosin. Neuronal

  6. Bone Morphogenetic Protein-7 Ameliorates Cerebral Ischemia and Reperfusion Injury via Inhibiting Oxidative Stress and Neuronal Apoptosis

    Directory of Open Access Journals (Sweden)

    Haitao Pei

    2013-11-01

    Full Text Available Previous studies have indicated that bone morphogenetic protein-7 (BMP-7 is neuroprotective against cerebral ischemia/reperfusion (IR injury. The present study was undertaken to determine the molecular mechanisms involved in this effect. Adult male Wistar rats were subjected to 2 h of transient middle cerebral artery occlusion (MCAO, followed by 24 h of reperfusion. BMP-7 (10−4 g/kg or vehicle was infused into rats at the onset of reperfusion via the tail vein. Neurological deficits, infarct volume, histopathological changes, oxidative stress-related biochemical parameters, neuronal apoptosis, and apoptosis-related proteins were assessed. BMP-7 significantly improved neurological and histological deficits, reduced the infarct volume, and decreased apoptotic cells after cerebral ischemia. BMP-7 also markedly enhanced the activities of antioxidant enzymes superoxide dismutase (SOD and glutathione peroxidase (GSH-PX, and reduced the level of malondialdehyde (MDA in IR rats. In addition, Western blot analysis indicated that BMP-7 prevented cytochrome c release, inhibited activation of caspase-3, caspase-9 and caspase-8. Our data suggested that BMP-7 has protective effects against cerebral IR injury in rats, and the neuroprotective effects may be attributed to attenuating oxidative stress and inhibiting neuronal apoptosis.

  7. Apoptosis of motor neurons in the spinal cord after ischemia reperfusion injury delayed paraplegia in rabbits

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Apoptosis is known to occur in the centralnervous system during development and in patho-logical settings such ischemia reperfusion(IR)inju-ry[1].Apoptosis requires an active commit ment ofthe cell to degrade its own DNA,according to aninternal programof self-destruction[2].Newproteinsynthesis is required for apoptosis,and protein syn-thesis inhibitors have been shown to reduce celldeath postischemically[3].Incontrast,necrosis is nota gene-facilitated process but results frominjuriouschanges in the environm...

  8. Loss of GCN5 leads to increased neuronal apoptosis by upregulating E2F1- and Egr-1-dependent BH3-only protein Bim.

    Science.gov (United States)

    Wu, Yanna; Ma, Shanshan; Xia, Yong; Lu, Yangpeng; Xiao, Shiyin; Cao, Yali; Zhuang, Sidian; Tan, Xiangpeng; Fu, Qiang; Xie, Longchang; Li, Zhiming; Yuan, Zhongmin

    2017-01-26

    Cellular acetylation homeostasis is a kinetic balance precisely controlled by histone acetyl-transferase (HAT) and histone deacetylase (HDAC) activities. The loss of the counterbalancing function of basal HAT activity alters the precious HAT:HDAC balance towards enhanced histone deacetylation, resulting in a loss of acetylation homeostasis, which is closely associated with neuronal apoptosis. However, the critical HAT member whose activity loss contributes to neuronal apoptosis remains to be identified. In this study, we found that inactivation of GCN5 by either pharmacological inhibitors, such as CPTH2 and MB-3, or by inactivation with siRNAs leads to a typical apoptosis in cultured cerebellar granule neurons. Mechanistically, the BH3-only protein Bim is transcriptionally upregulated by activated Egr-1 and E2F1 and mediates apoptosis following GCN5 inhibition. Furthermore, in the activity withdrawal- or glutamate-evoked neuronal apoptosis models, GCN5 loses its activity, in contrast to Bim induction. Adenovirus-mediated overexpression of GCN5 suppresses Bim induction and apoptosis. Interestingly, the loss of GCN5 activity and the induction of Egr-1, E2F1 and Bim are involved in the early brain injury (EBI) following subarachnoid haemorrhage (SAH) in rats. HDAC inhibition not only significantly rescues Bim expression and apoptosis induced by either potassium deprivation or GCN5 inactivation but also ameliorates these events and EBI in SAH rats. Taken together, our results highlight a new mechanism by which the loss of GCN5 activity promotes neuronal apoptosis through the transcriptional upregulation of Bim, which is probably a critical event in triggering neuronal death when cellular acetylation homeostasis is impaired.

  9. Total Flavone of Hawthorn Leaf inhibits neuronal apoptosis in brain tissue of rat models of chronic cerebral ischemia

    Institute of Scientific and Technical Information of China (English)

    Tan Rong-fang; Xia Ai-hua; Wu Xiao-guang; Cao Na-na; Li Meng-meng; Zhang Tian-ge; Wang Yi-ru; Yue Zhi-ling

    2014-01-01

    BACKGROUND: Cerebrovascular disease often causes dysfunction of the brain nerve, and nerve cel apoptosis is the important factor of cerebral nerve dysfunction. The excessive expression of c-fos can block the transduction of intracelular signal so that producing some apoptosis-promoting factors, which involve in nerve cel apoptosis process after ischemia injury of brain. Bcl-2 is an inhibited factor. It might to be the key to treat ischemic cerebrovascular disease by inhibiting or reducing the apoptosis of nerve cels after ischemia injury. OBJECTIVE: To investigate the therapeutic effect and mechanism of the Total Flavone of Hawthorn Leaf on chronic cerebral ischemia rats. METHODS: A total of 72 healthy male Sprague-Dawley rats were randomly divided into sham surgery group, model group, Total Flavone of Hawthorn Leaf group and ginkgo leaf group. Permanent bilateral carotid artery ligation was used to prepare chronic cerebral ischemia model in the model group, Total Flavone of Hawthorn Leaf group and ginkgo leaf group. Total Flavone of Hawthorn Leaf group and ginkgo leaf group respectively received 140 mg/kg Total Flavone of Hawthorn Leaf and 12.3 mg/kg ginkgo leaf intragastricaly for 36 days from 36 days after model induction. Model group and sham surgery group received 3.5 mL/kg physiological saline intragastricaly. RESULTS AND CONCLUSION: Compared with the model group, the expression of c-fos protein significantly deceased in the Total Flavone of Hawthorn Leaf group (P 0.05). These data indicated that the protective effect of Total Flavone of Hawthorn Leaf on chronic cerebral ischemia was associated with its inhibition of neuronal apoptosis. Its mechanism of anti-apoptosis might be associated with up-regulating expression of Bcl-2, down-regulating expression of c-fos and decreasing Ca2+ content in brain.

  10. Inhibition of caspases and intracellular free Ca2+ concentrations are involved in resveratrol protection against apoptosis in rat primary neuron cultures

    Institute of Scientific and Technical Information of China (English)

    Qi-hai GONG; Qian WANG; Jing-shan SHI; Xie-nan HUANG; Qiong LIU; Hu MA

    2007-01-01

    Aim:To investigate the influence of resveratrol (Res),a nutritional antioxidant,on the inhibition of apoptosis in rat primary neuron cultures. Methods:The cultured cortical neurons of neonatal Sprague-Dawley rats were pretreated with Res (0. 1,1.0,and 10.0μmol/L) and oxygen-glucose deprivation/reperfusion (OGD/RP) with oxygen and glucose were initiated at d 10 in vitro. Neuronal apoptosis was determined by flow cytometry,and morphological changes of neurons were observed by an electron microscope. For the mechanism studies,the intracellular free calcium concentration ([Ca2+]i) and the transcription of caspases-3 and -12 in neurons were detected by Fura 2/AM loading and real-time RT-PCR,respectively.Results:OGD/RP insult could induce an increase in the apoptotic rate of neurons (from 11.1% to 49.0%),and elicit an obvious morphological change in neurons;pretreatments with Res (0.1,1.0,and 10.0 μmol/L,respectively) significantly reduced the elevated rate of apoptosis to 41.7%,40.8%,and 37.4%,respectively,and ameliorated the neuronal morphological injury. Similarly,the OGD/RP insult obviously elicited the elevated levels of the [Ca2+]i and the expressions of caspases-3 and-12 mRNA in neurons. Res pretreatments markedly depressed the neuronal abnormal elevation of [Ca2+]i and the overexpression of caspases-3 and -12 mRNA in a concentration-dependent manner. Conclusion:Res can attenuate the rat cortical neuronal apoptosis induced by OGD/RP. The mechanisms are,at least partly,due to the inhibition of the calcium overload and the overexpression of caspases-3 and - 12 mRNA.

  11. Human recombinant factor VIIa may improve heat intolerance in mice by attenuating hypothalamic neuronal apoptosis and damage.

    Science.gov (United States)

    Hsu, Chuan-Chih; Chen, Sheng-Hsien; Lin, Cheng-Hsien; Yung, Ming-Chi

    2014-10-01

    Intolerance to heat exposure is believed to be associated with hypothalamo-pituitary-adrenocortical (HPA) axis impairment [reflected by decreases in blood concentrations of both adrenocorticotrophic-hormone (ACTH) and corticosterone]. The purpose of this study was to determine the effect of human recombinant factor VIIa (rfVIIa) on heat intolerance, HPA axis impairment, and hypothalamic inflammation, ischemic and oxidative damage, and apoptosis in mice under heat stress. Immediately after heat stress (41.2 °C for 1 h), mice were treated with vehicle (1 mL/kg of body weight) or rfVIIa (65-270 µg/kg of body weight) and then returned to room temperature (26 °C). Mice still alive on day 4 of heat exposure were considered survivors. Cellular ischemia markers (e.g., glutamate, lactate-to-pyruvate ratio), oxidative damage markers (e.g., nitric oxide metabolite, hydroxyl radials), and pro-inflammatory cytokines (e.g., interleukin-6, interleukin-1β, tumor necrosis factor-α) in hypothalamus were determined. In addition, blood concentrations of both ACTH and corticosterone were measured. Hypothalamic cell damage was assessed by determing the neuronal damage scores, whereas the hypothalamic cell apoptosis was determined by assessing the numbers of cells stained with terminal deoxynucleotidyl transferase-mediated αUTP nick-end labeling, caspase-3-positive cells, and platelet endothelial cell adhesion molecula-1-positive cells in hypothalamus. Compared with vehicle-treated heated mice, rfVIIa-treated heated mice had significantly higher fractional survival (8/10 vs 1/10), lesser thermoregulatory deficit (34.1 vs 24.8 °C), lesser extents of ischemic, oxidative, and inflammatory markers in hypothalamus, lesser neuronal damage scores and apoptosis in hypothalamus, and lesser HPA axis impairment. Human recombinant factor VIIa appears to exert a protective effect against heatstroke by attenuating hypothalamic cell apoptosis (due to ischemic, inflammatory, and oxidative damage

  12. Netrin-1 rescues neuron loss by attenuating secondary apoptosis in ipsilateral thalamic nucleus following focal cerebral infarction in hypertensive rats.

    Science.gov (United States)

    Liao, S-J; Gong, Q; Chen, X-R; Ye, L-X; Ding, Q; Zeng, J-S; Yu, J

    2013-02-12

    Neurological deficit following cerebral infarction correlates with not only primary injury, but also secondary neuronal apoptosis in remote loci connected to the infarction. Netrin-1 is crucial for axonal guidance by interacting with its receptors, deleted in colorectal cancer (DCC) and uncoordinated gene 5H (UNC5H). DCC and UNC5H are also dependence receptors inducing cell apoptosis when unbound by netrin-1. The present study is to investigate the role of netrin-1 and its receptors in ipsilateral ventroposterior thalamic nucleus (VPN) injury secondary to stroke in hypertensive rats. Renovascular hypertensive Sprague-Dawley rats underwent middle cerebral artery occlusion (MCAO). Continuous intracerebroventricular infusion of netrin-1 (600 ng/d for 7 days) or vehicle (IgG/Fc) was given 24h after MCAO. Neurological function was evaluated by postural reflex 8 and 14 days after MCAO. Then, immunoreactivity was determined in the ipsilateral VPN for NeuN, glial fibrillary acidic protein, netrin-1 and its receptors (DCC and UNC5H2), apoptosis was detected with Terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP-biotin nick-end labeling (TUNEL) assay, and the expressions of caspase-3, netrin-1, DCC, and UNC5H2 were quantified by western blot analysis. MCAO resulted in the impaired postural reflex after 8 and 14 days, with decreased NeuN marked neurons and increased TUNEL-positive cells, as well as an up-regulation in the levels of cleaved caspase-3 and UNC5H2 protein in the ipsilateral VPN, without significant change in DCC or netrin-1 expression. By exogenous netrin-1 infusion, the number of neurons was increased in the ipsilateral VPN, and both TUNEL-positive cell number and caspase-3 protein level were reduced, while UNC5H2 expression remained unaffected, simultaneously, the impairment of postural reflex was improved. Taken together, the present study indicates that exogenous netrin-1 could rescue neuron loss by attenuating secondary apoptosis in the

  13. Metallothionein-I+II in neuroprotection

    DEFF Research Database (Denmark)

    Pedersen, Mie Ø; Jensen, Rikke; Pedersen, Dan S

    2009-01-01

    -I+II decrease inflammation and secondary tissue damage (oxidative stress, neurodegeneration, and apoptosis) and promote post-injury repair and regeneration (angiogenesis, neurogenesis, neuronal sprouting and tissue remodelling). Intracellularly the molecular MT-I+II actions involve metal ion control...

  14. Motor neuron apoptosis and neuromuscular junction perturbation are prominent features in a Drosophila model of Fus-mediated ALS

    Directory of Open Access Journals (Sweden)

    Xia Ruohan

    2012-03-01

    Full Text Available Abstract Backgound Amyotrophic lateral sclerosis (ALS is progressive neurodegenerative disease characterized by the loss of motor function. Several ALS genes have been identified as their mutations can lead to familial ALS, including the recently reported RNA-binding protein fused in sarcoma (Fus. However, it is not clear how mutations of Fus lead to motor neuron degeneration in ALS. In this study, we present a Drosophila model to examine the toxicity of Fus, its Drosophila orthologue Cabeza (Caz, and the ALS-related Fus mutants. Results Our results show that the expression of wild-type Fus/Caz or FusR521G induced progressive toxicity in multiple tissues of the transgenic flies in a dose- and age-dependent manner. The expression of Fus, Caz, or FusR521G in motor neurons significantly impaired the locomotive ability of fly larvae and adults. The presynaptic structures in neuromuscular junctions were disrupted and motor neurons in the ventral nerve cord (VNC were disorganized and underwent apoptosis. Surprisingly, the interruption of Fus nuclear localization by either deleting its nuclear localization sequence (NLS or adding a nuclear export signal (NES blocked Fus toxicity. Moreover, we discovered that the loss of caz in Drosophila led to severe growth defects in the eyes and VNCs, caused locomotive disability and NMJ disruption, but did not induce apoptotic cell death. Conclusions These data demonstrate that the overexpression of Fus/Caz causes in vivo toxicity by disrupting neuromuscular junctions (NMJs and inducing apoptosis in motor neurons. In addition, the nuclear localization of Fus is essential for Fus to induce toxicity. Our findings also suggest that Fus overexpression and gene deletion can cause similar degenerative phenotypes but the underlying mechanisms are likely different.

  15. Quercetin and sesamin protect neuronal PC12 cells from high-glucose-induced oxidation, nitrosative stress, and apoptosis.

    Science.gov (United States)

    Bournival, Julie; Francoeur, Marc-André; Renaud, Justine; Martinoli, Maria-Grazia

    2012-06-01

    Complications of diabetes are now well-known to affect sensory, motor, and autonomic nerves. Diabetes is also thought to be involved in neurodegenerative processes characteristic of several neurodegenerative diseases. Indeed, it has been acknowledged recently that hyperglycemia-induced oxidative stress contributes to numerous cellular reactions typical of central nervous system deterioration. The goal of the present study was to evaluate the effects of the polyphenol quercetin and the lignan sesamin on high-glucose (HG)-induced oxidative damage in an in vitro model of dopaminergic neurons, neuronal PC12 cells. When incubated with HG (13.5 mg/mL), neuronal PC12 cells showed a significant increase of cellular death. Our results revealed that quercetin and sesamin defend neuronal PC12 cells from HG-induced cellular demise. An elevated level of reactive oxygen and nitrogen species is a consequence of improved oxidative stress after HG administration, and we demonstrated that this production diminishes with quercetin and sesamin treatment. We also found that quercetin and sesamin elicited an increment of superoxide dismutase activity. DNA fragmentation, Bax/Bcl-2 ratio, nuclear translocation of apoptosis-inducing factor, as well as poly(adenosine diphosphate [ADP]-ribose) polymerase cleavage were significantly reduced by quercetin and sesamin administration, affirming their antiapoptotic features. Also, HG treatment impacted caspase-3 cleavage, supporting caspase-3-dependent pathways as mechanisms of apoptotic death. Our results indicate a powerful role for these natural dietary compounds and emphasize preventive or complementary nutritional strategies for diabetes control.

  16. Neuronal apoptosis and neurofilament protein expression in the lateral geniculate body of cats following acute optic nerve injuries

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    BACKGROUND: The visual pathway have 6 parts, involving optic nerve, optic chiasm, optic tract, lateral geniculate body, optic radiation and cortical striatum area. Corresponding changes may be found in these 6 parts following optic nerve injury. At present, studies mainly focus on optic nerve and retina, but studies on lateral geniculate body are few.OBJECTIVE: To prepare models of acute optic nerve injury for observing the changes of neurons in lateral geniculate body, expression of neurofilament protein at different time after injury and cell apoptosis under the optical microscope, and for investigating the changes of neurons in lateral geniculate body following acute optic nerve injury.DESIGN: Completely randomized grouping design, controlled animal experiment.SETTING: Department of Neurosurgery, General Hospital of Ji'nan Military Area Command of Chinese PLA.MATERIALS: Twenty-eight adult healthy cats of either gender and common grade, weighing from 2.0 to 3.5 kg, were provided by the Animal Experimental Center of Fudan University. The involved cats were divided into 2 groups according to table of random digit: normal control group (n =3) and model group (n =25). Injury 6 hours, 1, 3, 7 and 14 days five time points were set in model group for later observation, 5 cats at each time point. TUNEL kit (Bohringer-Mannheim company)and NF200& Mr 68 000 mouse monoclonal antibody (NeoMarkers Company) were used in this experiment.METHODS: This experiment was carried out in the Department of Neurosurgery, General Hospital of Ji'nan Military Area Command of Chinese PLA between June 2004 and June 2005. ① The cats of model group were developed into cat models of acute intracranial optic nerve injury as follows: The anesthetized cats were placed in lateral position. By imitating operation to human, pterion approach was used. An incision was made at the joint line between outer canthus and tragus, and deepened along cranial base until white optic nerve via optic nerve pore

  17. Metallothionein expression in the central nervous system of multiple sclerosis patients

    DEFF Research Database (Denmark)

    Penkowa, M; Espejo, C; Ortega-Aznar, A;

    2003-01-01

    Multiple sclerosis (MS) is a major chronic demyelinating and inflammatory disease of the central nervous system (CNS) in which oxidative stress likely plays a pathogenic role in the development of myelin and neuronal damage. Metallothioneins (MTs) are antioxidant proteins induced in the CNS...

  18. Insulin-like growth factor-1 attenuates apoptosis and protects neurochemical phenotypes of dorsal root ganglion neurons with paclitaxel-induced neurotoxicity in vitro.

    Science.gov (United States)

    Chen, Cheng; Bai, Xue; Bi, Yanwen; Liu, Guixiang; Li, Hao; Liu, Zhen; Liu, Huaxiang

    2017-02-01

    Paclitaxel (PT)-induced neurotoxicity is a significant problem associated with successful treatment of cancers. Insulin-like growth factor-1 (IGF-1) is a neurotrophic factor and plays an important role in promoting axonal growth from dorsal root ganglion (DRG) neurons. Whether IGF-1 has protective effects on neurite growth, cell viability, neuronal apoptosis and neuronal phenotypes in DRG neurons with PT-induced neurotoxicity is still unclear. In this study, primary cultured rat DRG neurons were used to assess the effects of IGF-1 on DRG neurons with PT-induced neurotoxicity. The results showed that PT exposure caused neurite retraction in a dose-dependent manner. PT exposure caused a decrease of cell viability and an increase in the ratio of apoptotic cells which could be reversed by IGF-1. The percentage of calcitonin gene-related peptide immunoreactive (CGRP-IR) neurons and neurofilament (NF)-200-IR neurons, mRNA, and protein levels of CGRP and NF-200 decreased significantly after treatment with PT. IGF-1 administration had protective effects on CGRP-IR neurons, but not on NF-200-IR neurons. Either extracellular signal-regulated protein kinase (ERK1/2) inhibitor PD98059 or phosphatidylinositol 3-kinase (PI3 K) inhibitor LY294002 blocked the effect of IGF-1. The results imply that IGF-1 may attenuate apoptosis to improve neuronal cell viability and promote neurite growth of DRG neurons with PT-induced neurotoxicity. Moreover, these results support an important neuroprotective role of exogenous IGF-1 on distinct subpopulations of DRG neurons which is responsible for skin sensation. The effects of IGF-1 might be through ERK1/2 or PI3 K/Akt signaling pathways. These findings provide experimental evidence for IGF-1 administration to alleviate neurotoxicity of distinct subpopulations of DRG neurons induced by PT.

  19. Effects of D2 receptor antagonist haloperidol on hippocampal neuronal apoptosis in a rat model of temporal epilepsy

    Institute of Scientific and Technical Information of China (English)

    Songqing Wang; Aihua Zhang; Haitang Chen; Qianghua He; Peizeng Xie; Yiquan Ke; Xiaodan Jiang

    2008-01-01

    BACKGROUND: Dopamine receptors are divided into D1 and D2 subgroups. It has been reported that D2 receptors resist neural toxicity induced by excitatory amino acids and muscarine, and also alleviate epilepsy attacks following pilocarpine treatment. However, it has not yet been established whether D2 receptors regulate temporal epilepsy.OBJECTIVE: To observe the effects of the D2 antagonist halopefidol on hippocampal neuronal apoptosis and electrical brain activity in a rat model of kainic acid-induced temporal epilepsy.DESIGN, TIME AND SETTING: Randomized grouping and histopathological study were performed at the Neurology Medicine Institute of Zhujiang Hospital, Southern Medical University from August to December 2004.MATERIALS: Twenty-five adult, male, Sprague Dawley rats were selected for the present study. Kainic acid (Sigma, USA) was injected into the right lateral ventricle to establish models of temporal epilepsy. A PowerLab multiplying channel electrophysiolograph was provided by AD Instruments, Australia.METHODS: The rats were randomly divided into 5 groups (n = 5): control, model, haloperidol hippocampus, haloperidol striatum, and haloperidol substantia nigra. Temporal epilepsy was established in all rats except the control group. Haloperidol was slowly injected into the hippocampus, striatum and substantia nigra, respectively, in three different injection groups. Normal saline was injected into the fight lateral ventricle of the control rats.MAIN OUTCOME MEASURES: Hippocampal apoptosis was observed on the day 3 of treatment using TUNEL staining. Changes in electroencephalogram at 0, 0.5, 2, 6, and 12 hours following treatment onset were observed using a PowerLab multiplying channel electrophysiolograph. Animal behaviors were classified according to the Racine criteria.RESULTS: Twenty-five rats were included in the final analysis. Seizures did not occur in the control group. In the model group, 10 minutes alter kainic acid injection to the lateral

  20. LPS-induced microglial secretion of TNFα increases activity-dependent neuronal apoptosis in the neonatal cerebral cortex.

    Science.gov (United States)

    Nimmervoll, Birgit; White, Robin; Yang, Jenq-Wei; An, Shuming; Henn, Christopher; Sun, Jyh-Jang; Luhmann, Heiko J

    2013-07-01

    During the pre- and neonatal period, the cerebral cortex reveals distinct patterns of spontaneous synchronized activity, which is critically involved in the formation of early networks and in the regulation of neuronal survival and programmed cell death (apoptosis). During this period, the cortex is also highly vulnerable to inflammation and in humans prenatal infection may have a profound impact on neurodevelopment causing long-term neurological deficits. Using in vitro and in vivo multi-electrode array recordings and quantification of caspase-3 (casp-3)-dependent apoptosis, we demonstrate that lipopolysaccharide-induced inflammation causes rapid alterations in the pattern of spontaneous burst activities, which subsequently leads to an increase in apoptosis. We show that these inflammatory effects are specifically initiated by the microglia-derived pro-inflammatory cytokine tumor necrosis factor α and the chemokine macrophage inflammatory protein 2. Our data demonstrate that inflammation-induced modifications in spontaneous network activities influence casp-3-dependent cell death in the developing cerebral cortex.

  1. Cisplatin-induced apoptosis in auditory, renal, and neuronal cells is associated with nitration and downregulation of LMO4.

    Science.gov (United States)

    Rathinam, Rajamani; Ghosh, Samiran; Neumann, William L; Jamesdaniel, Samson

    Cytotoxic effects of cisplatin occur primarily through apoptosis. Though several pro- and anti-apoptotic signaling molecules have been identified to play an important role in mediating the ototoxic, nephrotoxic, and neurotoxic side-effects of cisplatin, the underlying mechanism is yet to be fully characterized. We reported that nitration of LIM domain only 4 (LMO4), a transcriptional regulator, facilitates cochlear apoptosis in cisplatin-induced ototoxicity. However, its role in cisplatin-mediated nephrotoxicity and neurotoxicity is poorly understood. Therefore, HK2, and SH-SY5Y cells were employed along with UBOC1 cells, to investigate the perturbations of LMO4 in cisplatin-induced cytotoxicity, in renal, neuronal, and auditory cells, respectively. Cisplatin induced an increase in the expression of active caspase-3, indicating cellular apoptosis, and increased the nitration of proteins, 24 h post-treatment. Immunostaining with anti-nitrotyrosine and anti-LMO4 indicated that nitrotyrosine co-localized with LMO4 protein in cisplatin treated cells. Immunoblotting with anti-LMO4 indicated that cisplatin induced a decrease in LMO4 protein levels. However, a corresponding decrease in LMO4 gene levels was not observed. Inhibition of protein nitration with SRI110, a peroxynitrite decomposition catalyst, attenuated cisplatin-induced downregulation of LMO4. More importantly, overexpression of LMO4 mitigated the cytotoxic effects of cisplatin in UBOC1 cells while a dose-dependent decrease in LMO4 protein strongly correlated with cell viability in UBOC1, HK2, and SH-SY5Y cells. Collectively, these findings suggested a potential role of LMO4 in facilitating the cytotoxic effects of cisplatin in auditory, renal, and neuronal cells.

  2. Safrole oxide induces neuronal apoptosis through inhibition of integrin beta4/SOD activity and elevation of ROS/NADPH oxidase activity.

    Science.gov (United States)

    Su, Le; Zhao, BaoXiang; Lv, Xin; Wang, Nan; Zhao, Jing; Zhang, ShangLi; Miao, JunYing

    2007-02-20

    Neuronal apoptosis is a very important event in the development of the central nervous system (CNS), but the underlying mechanisms remain to be elucidated. We have previously shown that safrole oxide, a small molecule, induces integrin beta4 expression and promotes apoptosis in vascular endothelial cells. In this study, the effects of safrole oxide on cell growth and apoptosis have been examined in primary cultures of mouse neurons. Safrole oxide was found to significantly inhibit neuronal cell growth and to induce apoptosis. The inhibitory and apoptotic activities of safrole oxide followed a dose- and time-dependent manner. Interestingly, the expression of integrin beta4 was significantly inhibited with safrole oxide treatment. Furthermore, safrole oxide dramatically increases the level of intracellular reactive oxygen species (ROS) and the activity of NADPH oxidase. Moreover, manganese-dependent superoxide dismutase (MnSOD) activity was decreased significantly with safrole oxide treatment. Our study thus demonstrates that safrole oxide induces neuronal apoptosis through integrin beta4, ROS, NADPH, and MnSOD.

  3. p38 Activation Is Required Upstream of Potassium Current Enhancement and Caspase Cleavage in Thiol Oxidant-Induced Neuronal Apoptosis

    Science.gov (United States)

    McLaughlin, BethAnn; Pal, Sumon; Tran, Minhnga P.; Parsons, Andrew A.; Barone, Frank C.; Erhardt, Joseph A.; Aizenman, Elias

    2013-01-01

    Oxidant-induced neuronal apoptosis has been shown to involve potassium and zinc dysregulation, energetic dysfunction, activation of stress-related kinases, and caspase cleavage. The temporal ordering and interdependence of these events was investigated in primary neuronal cultures exposed to the sulfhydryl oxidizing agent 2,2′-dithiodipyridine (DTDP), a compound that induces the intracellular release of zinc. We previously observed that tetraethylammonium (TEA), high extracellular potassium, or cysteine protease inhibitors block apoptosis induced by DTDP. We now report that both p38 and extracellular signal-regulated kinase phosphorylation are evident in neuronal cultures within 2 hr of a brief exposure to 100 μm DTDP. However, only p38 inhibition is capable of blocking oxidant-induced toxicity. Cyclohexamide or actinomycin D does not attenuate DTDP-induced cell death, suggesting that posttranslational modification of existing targets, rather than transcriptional activation, is responsible for the deleterious effects of p38. Indeed, an early robust increase in TEA-sensitive potassium channel currents induced by DTDP is attenuated by p38 inhibition but not by caspase inhibition. Moreover, we found that activation of p38 is required for caspase 3 and 9 cleavage, suggesting that potassium currents enhancement is required for caspase activation. Finally, we observed that DTDP toxicity could be blocked with niacinamide or benzamide, inhibitors of poly (ADP-ribose) synthetase. Based on these findings, we conclude that oxidation of sulfhydryl groups on intracellular targets results in intracellular zinc release, p38 phosphorylation, enhancement of potassium currents, caspase cleavage, energetic dysfunction, and translationally independent apoptotic cell death. PMID:11331359

  4. Involvement of the JNK/FOXO3a/Bim Pathway in Neuronal Apoptosis after Hypoxic-Ischemic Brain Damage in Neonatal Rats.

    Directory of Open Access Journals (Sweden)

    Deyuan Li

    Full Text Available c-Jun N-terminal kinase (JNK plays a key role in the regulation of neuronal apoptosis. Previous studies have revealed that forkhead transcription factor (FOXO3a is a critical effector of JNK-mediated tumor suppression. However, it is not clear whether the JNK/FOXO3a pathway is involved in neuronal apoptosis in the developing rat brain after hypoxia-ischemia (HI. In this study, we generated an HI model using postnatal day 7 rats. Fluorescence immunolabeling and Western blot assays were used to detect the distribution and expression of total and phosphorylated JNK and FOXO3a and the pro-apoptotic proteins Bim and CC3. We found that JNK phosphorylation was accompanied by FOXO3a dephosphorylation, which induced FOXO3a translocation into the nucleus, resulting in the upregulation of levels of Bim and CC3 proteins. Furthermore, we found that JNK inhibition by AS601245, a specific JNK inhibitor, significantly increased FOXO3a phosphorylation, which attenuated FOXO3a translocation into the nucleus after HI. Moreover, JNK inhibition downregulated levels of Bim and CC3 proteins, attenuated neuronal apoptosis and reduced brain infarct volume in the developing rat brain. Our findings suggest that the JNK/FOXO3a/Bim pathway is involved in neuronal apoptosis in the developing rat brain after HI. Agents targeting JNK may offer promise for rescuing neurons from HI-induced damage.

  5. Ischemia leads to apoptosis--and necrosis-like neuron death in the ischemic rat hippocampus

    DEFF Research Database (Denmark)

    Müller, Georg Johannes; Stadelmann, Christine; Bastholm, Lone

    2004-01-01

    pyramidal cells of the rat hippocampus. The earliest ischemic changes were found on day 2 and 3, reflected by an upregulation of phospho-c-Jun in a proportion of morphologically intact CA1 neurons, which matched the number of neurons that succumbed to ischemia at later time points. At day 3 and later 3...... and/or caspase-3 expression represented a minor fraction (neurons, while the vast majority followed a necrosis-like pathway. Our studies suggest that CA1 pyramidal cell death following transient forebrain ischemia may be initiated through c-Jun N-terminal kinase (JNK) pathway...

  6. Metallothioneins I and II: neuroprotective significance during CNS pathology

    DEFF Research Database (Denmark)

    Penkowa, Milena; Stankovic, Roger; Chung, Roger

    2006-01-01

    isoforms of the protein (MT-I and MT-II) are induced by numerous stimuli and pathogens but most importantly their induction by metals is closely linked to the physiological metabolism of zinc and protection from the toxic affects following heavy metal exposure. Although the preservation of their genetic......Metallothioneins (MTs) constitutes a superfamily of highly conserved, low molecular weight polypeptides, which are characterized by high contents of cysteine (sulphur) and metals. As intracellular metal-binding proteins they play a significant role in the regulation of essential metals. The major...... may provide neurotherapeutic targets offering protection against neuronal injury and degeneration....

  7. The microRNA miR-181c controls microglia-mediated neuronal apoptosis by suppressing tumor necrosis factor

    Directory of Open Access Journals (Sweden)

    Zhang Li

    2012-09-01

    Full Text Available Abstract Background Post-ischemic microglial activation may contribute to neuronal damage through the release of large amounts of pro-inflammatory cytokines and neurotoxic factors. The involvement of microRNAs (miRNAs in the pathogenesis of disorders related to the brain and central nervous system has been previously studied, but it remains unknown whether the production of pro-inflammatory cytokines is regulated by miRNAs. Methods BV-2 and primary rat microglial cells were activated by exposure to oxygen-glucose deprivation (OGD. Global cerebral ischemia was induced using the four-vessel occlusion (4-VO model in rats. Induction of pro-inflammatory and neurotoxic factors, such as tumor necrosis factor (TNF-α, interleukin (IL-1β, and nitric oxide (NO, were assessed by ELISA, immunofluorescence, and the Griess assay, respectively. The miRNA expression profiles of OGD-activated BV-2 cells were subsequently compared with the profiles of resting cells in a miRNA microarray. BV-2 and primary rat microglial cells were transfected with miR-181c to evaluate its effects on TNF-α production after OGD. In addition, a luciferase reporter assay was conducted to confirm whether TNF-α is a direct target of miR-181c. Results OGD induced BV-2 microglial activation in vitro, as indicated by the overproduction of TNF-α, IL-1β, and NO. Global cerebral ischemia/reperfusion injury induced microglial activation and the release of pro-inflammatory cytokines in the hippocampus. OGD also downregulated miR-181c expression and upregulated TNF-α expression. Overproduction of TNF-α after OGD-induced microglial activation provoked neuronal apoptosis, whereas the ectopic expression of miR-181c partially protected neurons from cell death caused by OGD-activated microglia. RNAinterference-mediated knockdown of TNF-α phenocopied the effect of miR-181c-mediated neuronal protection, whereas overexpression of TNF-α blocked the miR-181c-dependent suppression of apoptosis

  8. Metallothionein as an Anti-Inflammatory Mediator

    Directory of Open Access Journals (Sweden)

    Ken-ichiro Inoue

    2009-01-01

    Full Text Available The integration of knowledge concerning the regulation of MT, a highly conserved, low molecular weight, cystein-rich metalloprotein, on its proposed functions is necessary to clarify how MT affects cellular processes. MT expression is induced/enhanced in various tissues by a number of physiological mediators. The cellular accumulation of MT depends on the availability of cellular zinc derived from the diet. MT modulates the binding and exchange/transport of heavy metals such as zinc, cadmium, or copper under physiological conditions and cytoprotection from their toxicities, and the release of gaseous mediators such as hydroxyl radicals or nitric oxide. In addition, MT reportedly affects a number of cellular processes, such as gene expression, apoptosis, proliferation, and differentiation. Given the genetic approach, the apparently healthy status of MT-deficient mice argues against an essential biological role for MT; however, this molecule may be critical in cells/tissues/organs in times of stress, since MT expression is also evoked/enhanced by various stresses. In particular, because metallothionein (MT is induced by inflammatory stress, its roles in inflammation are implied. Also, MT expression in various organs/tissues can be enhanced by inflammatory stimuli, implicating in inflammatory diseases. In this paper, we review the role of MT of various inflammatory conditions.

  9. Effects of arsenic poisoning on neuronal cell apoptosis and mRNA and protein expression of calpain 1,calpain 2,and cdk5/p25

    Institute of Scientific and Technical Information of China (English)

    李新

    2014-01-01

    Objective To study the effect of arsenic on neuronal cell apoptosis and the mRNA and protein expression of calpain 1,calpain 2,and cyclin-dependent kinases 5(cdk5)/p25 and to provide a scientific basis for the research on neurotoxic mechanism of arsenic trioxide(As2O3).Methods Primary cultured rat neurons were divided into untreated control group,dimethyl sulfoxide

  10. Interleukin-1β enhances neuronal vulnerability to proNGF-mediated apoptosis by increasing surface expression of p75NTR and sortillin

    OpenAIRE

    Choi, Soyoung; Friedman, Wilma J.

    2013-01-01

    Many types of injury such as seizure, ischemia, and oxidative stress cause upregulation of the p75 neurotrophin receptor (p75NTR) in brain neurons, where it promotes apoptosis, however the mechanism by which p75NTR is regulated under these conditions is not well understood. Proinflammatory cytokines such as interleukin-1β (IL-1β) are highly produced under these injury conditions and, in particular, are expressed rapidly in the rat hippocampus after seizure. IL-1β is known to increase neuronal...

  11. Association of copy numbers of survival motor neuron gene 2 and neuronal apoptosis inhibitory protein gene with the natural history in a Chinese spinal muscular atrophy cohort.

    Science.gov (United States)

    Qu, Yu-jin; Ge, Xiu-shan; Bai, Jin-li; Wang, Li-wen; Cao, Yan-yan; Lu, Yan-yu; Jin, Yu-wei; Wang, Hong; Song, Fang

    2015-03-01

    We evaluated survival motor neuron 2 (SMN2) and neuronal apoptosis inhibitory protein (NAIP) gene copy distribution and the association of copy number with survival in 232 Chinese spinal muscular atrophy (SMA) patients. The SMN2 and NAIP copy numbers correlated positively with the median onset age (r = 0.72 and 0.377). The risk of death for patients with fewer copies of SMN2 or NAIP was much higher than for those with more copies (P < .01). The survival probabilities at 5 years were 5.1%, 90.7%, and 100% for 2, 3, and 4 SMN2 copies and 27.9%, 66.7%, and 87.2% for 0, 1, and 2 NAIP copies, respectively. Our results indicated that combined SMN1-SMN2-NAIP genotypes with fewer copies were associated with earlier onset age and poorer survival probability. Better survival status for Chinese type I SMA might due to a higher proportion of 3 SMN2 and a lower rate of zero NAIP.

  12. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) induces microglial nitric oxide production and subsequent rat primary cortical neuron apoptosis through p38/JNK MAPK pathway.

    Science.gov (United States)

    Li, Yuanye; Chen, Gang; Zhao, Jianya; Nie, Xiaoke; Wan, Chunhua; Liu, Jiao; Duan, Zhiqing; Xu, Guangfei

    2013-10-04

    It has been widely accepted that microglia, which are the innate immune cells in the brain, upon activation can cause neuronal damage. In the present study, we investigated the role of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in regulating microglial nitric oxide production and its role in causing neuronal damage. The study revealed that TCDD stimulates the expression of inducible nitric oxide synthase (iNOS) as well as the production of nitric oxide (NO) in a dose- and time-dependent manner. Further, a rapid activation of p38 and JNK MAPKs was found in HAPI microglia following TCDD treatment. Blockage of p38 and JNK kinases with their specific inhibitors, SB202190 and SP600125, significantly reduced TCDD-induced iNOS expression and NO production. In addition, it was demonstrated through treating rat primary cortical neurons with media conditioned with TCDD treated microglia that microglial iNOS activation mediates neuronal apoptosis. Lastly, it was also found that p38 and JNK MAPK inhibitors could attenuate the apoptosis of rat cortical neurons upon exposure to medium conditioned by TCDD-treated HAPI microglial cells. Based on these observations, we highlight that the p38/JNK MAPK pathways play an important role in TCDD-induced iNOS activation in rat HAPI microglia and in the subsequent induction of apoptosis in primary cortical neurons.

  13. The role of myocardin-related transcription factor-A in Aβ25-35 induced neuron apoptosis and synapse injury.

    Science.gov (United States)

    Zhang, Ying; Pan, Hong-Yan; Hu, Xia-Min; Cao, Xiao-Lu; Wang, Jun; Min, Zhen-Li; Xu, Shi-Qiang; Xiao, Wan; Yuan, Qiong; Li, Na; Cheng, Jing; Zhao, Shu-Qi; Hong, Xing

    2016-10-01

    Myocardin-related transcription factor-A (MRTF-A) highly expressed in brain has been demonstrated to promote neuronal survival via regulating the transcription of related target genes as a powerful co-activator of serum response factor (SRF). However, the role of MRTF-A in Alzheimer's disease (AD) is still unclear. Here, we showed that MRTF-A was significantly downregulated in cortex of the Aβ25-35-induced AD rats, which played a key role in Aβ25-35 induced cerebral neuronal degeneration in vitro. Bilateral intracerebroventricular injection of Aβ25-35 caused significantly MRTF-A expression decline in cortex of rats, along with significant neuron apoptosis and plasticity damage. In vitro, transfection of MRTF-A into primary cultured cortical neurons prevented Aβ25-35 induced neuronal apoptosis and synapses injury. And luciferase reporter assay determined that MRTF-A could bind to and enhance the transactivity of the Mcl-1 (Myeloid cell leukemia-1) and Arc (activity-regulated cytoskeletal-associated protein) promoters by activating the key CArG box element. These data demonstrated that the decreasing of endogenous MRTF-A expression might contribute to the development of AD, whereas the upregulation MRTF-A in neurons could effectively reduce Aβ25-35 induced synapse injury and cell apoptosis. And the underlying mechanism might be partially due to MRTF-A-mediated the transcription and expression of Mcl-1 and Arc by triggering the CArG box.

  14. The MLK family mediates c-Jun N-terminal kinase activation in neuronal apoptosis.

    Science.gov (United States)

    Xu, Z; Maroney, A C; Dobrzanski, P; Kukekov, N V; Greene, L A

    2001-07-01

    Neuronal apoptotic death induced by nerve growth factor (NGF) deprivation is reported to be in part mediated through a pathway that includes Rac1 and Cdc42, mitogen-activated protein kinase kinases 4 and 7 (MKK4 and -7), c-Jun N-terminal kinases (JNKs), and c-Jun. However, additional components of the pathway remain to be defined. We show here that members of the mixed-lineage kinase (MLK) family (including MLK1, MLK2, MLK3, and dual leucine zipper kinase [DLK]) are expressed in neuronal cells and are likely to act between Rac1/Cdc42 and MKK4 and -7 in death signaling. Overexpression of MLKs effectively induces apoptotic death of cultured neuronal PC12 cells and sympathetic neurons, while expression of dominant-negative forms of MLKs suppresses death evoked by NGF deprivation or expression of activated forms of Rac1 and Cdc42. CEP-1347 (KT7515), which blocks neuronal death caused by NGF deprivation and a variety of additional apoptotic stimuli and which selectively inhibits the activities of MLKs, effectively protects neuronal PC12 cells from death induced by overexpression of MLK family members. In addition, NGF deprivation or UV irradiation leads to an increase in both level and phosphorylation of endogenous DLK. These observations support a role for MLKs in the neuronal death mechanism. With respect to ordering the death pathway, dominant-negative forms of MKK4 and -7 and c-Jun are protective against death induced by MLK overexpression, placing MLKs upstream of these kinases. Additional findings place the MLKs upstream of mitochondrial cytochrome c release and caspase activation.

  15. Time-Dependent Increases in Protease Activities for Neuronal Apoptosis in Spinal Cords of Lewis Rats During Development of Acute Experimental Autoimmune Encephalomyelitis

    Science.gov (United States)

    Das, Arabinda; Guyton, M. Kelly; Matzelle, Denise D.; Ray, Swapan K.; Banik, Naren L.

    2008-01-01

    Multiple sclerosis (MS) is characterized by axonal demyelination and neurodegeneration, the latter having been inadequately explored in the MS animal model experimental autoimmune encephalomyelitis (EAE). The purpose of this study was to examine the time-dependent correlation between increased calpain and caspase activities and neurodegeneration in spinal cord tissues from Lewis rats with acute EAE. An increase in TUNEL-positive neurons and internucleosomal DNA fragmentation in EAE spinal cords suggested that neuronal death was a result of apoptosis on days 8–10 following induction of EAE. Increases in calpain expression in EAE correlated with activation of pro-apoptotic proteases, leading to apoptotic cell death beginning on day 8 of EAE, which occurred before the appearance of visible clinical symptoms. Increases in calcineurin expression and decreases in phospho-Bad (p-Bad) suggested Bad activation in apoptosis during acute EAE. Increases in the Bax:Bcl-2 ratio and activation of caspase-9 showed the involvement of mitochondria in apoptosis. Further, caspase-8 activation suggested induction of the death receptor–mediated pathway for apoptosis. Endoplasmic reticulum stress leading to caspase-3 activation was also observed, indicating that multiple apoptotic pathways were activated following EAE induction. In contrast, cell death was mostly a result of necrosis on the later day (day 11), when EAE entered a severe stage. From these findings, we conclude that increases in calpain and caspase activities play crucial roles in neuronal apoptosis during the development of acute EAE. PMID:18521931

  16. Expression of NF-кB in Schwann cells and its effect on motor neuron apoptosis in spinal cord following sciatic nerves injury in rats

    Institute of Scientific and Technical Information of China (English)

    WANG Yong-tang; LU Xiu-min; YU Ying; YANG Yan-hong; GAO Jie

    2007-01-01

    Objective:To explore the expression of nuclear factor-kappa B (NF-кB) in Schwann cells (SCs) and its effect on motor neuron apoptosis in spinal cord following sciatic nerves injury in adult rats. Methods:Thirty-six adult Sprague-Dawley (SD) rats were divided randomly into normal control group (n=6),and sciatic nerves crushing group (n=30),and the later was further equally randomized into 5 nerves were examined by immunohistochemistry staining,and the apoptosis of motor neurons in spinalcord of lumbar 4 to lumbar 6(L4-L6)was investigated by terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling (TUNEL) assay.Both were quantitated by image analysis.Results:In the normal control group (P<0.05,P<0.01).At 1 d after sciatic nerves crushing,the expression of served in the time-course on motor neuron apoptosis after sciatic nerves injury.Correlation analyses refollowing sciatic nerves injury(r=0.976 0,P<0.01).Conclusion:After injury of sciatic nerves,the presence and up-regulation of NF-κB in SCs may be involved in motor neuron apoptosis in L4-L6 spinal cord.

  17. Inhibitor of apoptosis-stimulating protein of p53 (iASPP is required for neuronal survival after axonal injury.

    Directory of Open Access Journals (Sweden)

    Ariel M Wilson

    Full Text Available The transcription factor p53 mediates the apoptosis of post-mitotic neurons exposed to a wide range of stress stimuli. The apoptotic activity of p53 is tightly regulated by the apoptosis-stimulating proteins of p53 (ASPP family members: ASPP1, ASPP2 and iASPP. We previously showed that the pro-apoptotic members ASPP1 and ASPP2 contribute to p53-dependent death of retinal ganglion cells (RGCs. However, the role of the p53 inhibitor iASPP in the central nervous system (CNS remains to be elucidated. To address this, we asked whether iASPP contributes to the survival of RGCs in an in vivo model of acute optic nerve damage. We demonstrate that iASPP is expressed by injured RGCs and that iASPP phosphorylation at serine residues, which increase iASPP affinity towards p53, is significantly reduced following axotomy. We show that short interference RNA (siRNA-induced iASPP knockdown exacerbates RGC death, whereas adeno-associated virus (AAV-mediated iASPP expression promotes RGC survival. Importantly, our data also demonstrate that increasing iASPP expression in RGCs downregulates p53 activity and blocks the expression of pro-apoptotic targets PUMA and Fas/CD95. This study demonstrates a novel role for iASPP in the survival of RGCs, and provides further evidence of the importance of the ASPP family in the regulation of neuronal loss after axonal injury.

  18. Melissa officinalis Acidic Fraction Protects Cultured Cerebellar Granule Neurons Against Beta Amyloid-Induced Apoptosis and Oxidative Stress.

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    Soodi, Maliheh; Dashti, Abolfazl; Hajimehdipoor, Homa; Akbari, Shole; Ataei, Nasim

    2017-01-01

    Extracellular deposition of the beta-amyloid (Aβ) peptide, which is the main finding in the pathophysiology of Alzheimer's disease (AD), leads to oxidative damage and apoptosis in neurons. Melissa officinalis (M. officinalis) is a medicinal plant from the Lamiaceae family that has neuroprotective activity. In the present study we have investigated the protective effect of the acidic fraction of M. officinalis on Aβ-induced oxidative stress and apoptosis in cultured cerebellar granule neurons (CGN). Additionally, we investigated a possible role of the nicotinic receptor. This study was an in vitro experimental study performed on mice cultured CGNs. CGNs were pre-incubated with different concentrations of the acidic fraction of M. officinalis for 24 hours, followed by incubation with Aβ for an additional 48 hours. CGNs were also pre-incubated with the acidic fraction of M. officinalis and mecamylamin, followed by incubation with Aβ. We used the 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) assay to measure cell viability. Acetylcholinesterase (AChE) activity, reactive oxygen species (ROS) production, lipidperoxidation, and caspase-3 activity were measured after incubation. Hochst/annexin Vfluorescein isothiocyanate (FITC)/propidium iodide (PI) staining was performed to detect apoptotic cells. The acidic fraction could protect CGNs from Aβ-induced cytotoxicity. Mecamylamine did not abolish the protective effect of the acidic fraction. AChE activity, ROS production, lipid peroxidation, and caspase-3 activity increased after Aβ incubation. Preincubation with the acidic fraction of M. officinalis ameliorated these factors and decreased the number of apoptotic cells. Our results indicated that the protective effect of the acidic fraction of M. officinalis was not mediated through nicotinic receptors. This fraction could protect CGNs through antioxidant and anti-apoptotic activities.

  19. Role of alpha-synuclein in neuronal apoptosis induced by rotenone

    Institute of Scientific and Technical Information of China (English)

    Yanying Liu; Hui Yang

    2006-01-01

    BACKGROUND: Aggregation of α-synuclein is the major component of Lewy bodies, which are the pathological hallmarks of Parkinson disease (PD). Although the mechanism of this protein aggregates is unclear,previous study showed that environmental toxins such as rotenone could induce the expression and aggregation of α-synuclein.OBJECTIVE: To observe the role of α-synuclein in PD.DESIGN: A randomized controlled trial.SETTING: Beijing Institute for Neuroscience, Capital University of Medical Sciences.MATERIALS: This study was performed from July 2005 to January 2006 at the Beijing Institute for Neuroscience, Capital University of Medical Sciences. Human dopaminergic neuroblastoma SH-SY5Y cells were provided by Beijing Institute for Neuroscience, Capital University of Medical Sciences.METHODS: Human dopaminergic neuroblastoma SH-SY5Y cells were treated to make α-synuclein over express. Rotenone was added into the medium of cultured both native SH-SY5Y cells and α-synuclein-overexpression SH-SY5Y cells. Lactate dehydrogenase (LDH) assay was used to detect with the cell viability. Flow cytometry and electrophoresis were adopted to measure the cell apoptosis.MAIN OUTCOME MEASURES: Cell viability, DNA fragmentation, and the number of cell apoptosis.RESULTS: After being treated with rotenone, LDH activity of α-synuclein overexpressed SH-SY5Y cells was (76.625±6.34) μ kat/L, which was significantly lower than that of control group (P < 0.05). As compared with normal SH-SY5Y cell, α-synuclein over-expressed SH-SY5Y cells had less DNA fragments and apoptotic cells. Α-synuclein might play a role in cell apoptosis induced by rotenone, which was also confirmed by using of antioxidant reagent.CONCLUSION: α-synuclein may partially protect against cell apoptosis induced by rotenone in SH-SY5Y cells.

  20. Effects of propofol on neuronal apoptosis and aquaporin-4 expression in a rat model of traumatic brain injury

    Institute of Scientific and Technical Information of China (English)

    Jianfang Song; Xiangyu Ji; Zangong Zhou

    2008-01-01

    BACKGROUND: Several studies have demonstrated that propofol exhibits protective effects in the central nervous system. OBJECTIVE: To observe the effects of propofol on neuronal apoptosis and aquaporin-4 (AQP-4) expression in a rat model of traumatic brain injury and to further investigate the mechanisms of action. DESIGN, TIME AND SETTING: The present neuronal, pathomorphological experiment was performed at the Institute of Cerebrovascular Disease, Qingdao University Medical College between April 2007 and March 2008. MATERIALS: Traumatic brain injury was induced by free falling objects in 150 healthy, male, Wistar rats. Propotol was produced by AstraZeaeca, China. Rabbit anti-rat AQP-4 polyclonal antibody, SABC inununohistochemistry kit, and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick-end labeling (TUNEL) kit were purchased from Wuhan Boster Bioengineering Co., Ltd., China. METHODS: All 150 rats were randomly and evenly divided into lesion-only and propofol-treated groups. One hour after traumatic brain injury, propofol-treated animals received 1% propofol (10 mg/kg) through the caudal vein, followed by a sustained perfusion of 30 mg/kg propofol per hour for 2 hours, while the lesion-only group received equal volumes of physiological saline in parallel. MAIN OUTCOME MEASURES: At 6, 12, 24, 48, and 72 hours after traumatic brain injury, morphological changes in the peritraumatic and adjacent brain areas were analyzed in all rats by hematoxylin-eosin (HE) staining. In addition, cellular apoptosis was detected by TUNEL assay and the number of AQP-4-positive cells was determined by immunohistochemistry techniques. Brain water content was calculated as the ratio of dry to wet tissue weight. RESULTS: HE staining results demonstrated that, in the lesion-only group, the peritraumatic area exhibited neuronal and glial cell necrosis and disintegration. The adjacent area displayed swollen neuronal perikarya and vascular endothelial cells, cellular edema

  1. YiQiFuMai Powder Injection Ameliorates Cerebral Ischemia by Inhibiting Endoplasmic Reticulum Stress-Mediated Neuronal Apoptosis

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    Guosheng Cao

    2016-01-01

    Full Text Available YiQiFuMai (YQFM powder injection as a modern preparation derived from Sheng Mai San, a traditional Chinese medicine, has been widely used in the treatment of cardiovascular and cerebrovascular diseases. However, its neuroprotective effect and underlying mechanism in cerebral ischemia remain to be explored. The present study was designed to investigate the neuroprotective effect of YQFM on endoplasmic reticulum (ER stress-mediated neuronal apoptosis in the permanent middle cerebral artery occlusion- (MCAO- injured mice and the oxygen-glucose deprivation- (OGD- induced pheochromocytoma (PC12 cells. The results showed that single administration of YQFM (1.342 g/kg, i.p. could reduce the brain infarction and improve the neurological deficits and the cerebral blood flow (CBF after MCAO for 24 h in mice. Moreover, incubation with YQFM (100, 200, and 400 μg/mL could increase the cell viability, decrease the caspase-3 activity, and inhibit the cell apoptosis in OGD-induced PC12 cells for 12 h. In addition, YQFM treatment could significantly modulate cleaved caspase-3 and Bcl-2 expressions and inhibit the expressions of ER stress-related marker proteins and signaling pathways in vivo and in vitro. In conclusion, our findings provide the first evidence that YQFM ameliorates cerebral ischemic injury linked with modulating ER stress-related signaling pathways, which provided some new insights for its prevention and treatment of cerebral ischemia diseases.

  2. Zika virus infection during the period of maximal brain growth causes microcephaly and corticospinal neuron apoptosis in wild type mice

    Science.gov (United States)

    Huang, Wen-Chin; Abraham, Rachy; Shim, Byoung-Shik; Choe, Hyeryun; Page, Damon T.

    2016-01-01

    Zika virus (ZIKV) infection in pregnant women has been established as a cause of microcephaly in newborns. Here we test the hypothesis that neurodevelopmental stages when the brain is undergoing rapid growth are particularly vulnerable to the effects of ZIKV infection. We injected ZIKV intracranially into wild type C57BL/6 mice at two different time points: early postnatal development, when the brain is growing at its maximal rate, and at weaning, when the brain has largely reached adult size. Both time points showed widespread immunoreactivity for ZIKV and cleaved caspase 3 (CC3, a marker of apoptosis) throughout the brain. However, in early postnatal ZIKV injected mice, some brain areas and cell types display particularly large increases in apoptosis that we did not observe in older animals. Corticospinal pyramidal neurons, a cell type implicated in human microcephaly associated with ZIKV infection, are an example of one such cell type. Proliferating cells in the ventricular zone stem cell compartment are also depleted. These findings are consistent with the hypothesis that periods of rapid brain growth are especially susceptible to neurodevelopmental effects of ZIKV infection, and establish a valuable model to investigate mechanisms underlying neurodevelopmental effects of ZIKV infection and explore candidate therapeutics. PMID:27713505

  3. Nrdp1 Increases Ischemia Induced Primary Rat Cerebral Cortical Neurons and Pheochromocytoma Cells Apoptosis Via Downregulation of HIF-1α Protein

    Directory of Open Access Journals (Sweden)

    Yuan Zhang

    2017-09-01

    Full Text Available Neuregulin receptor degradation protein-1 (Nrdp1 is an E3 ubiquitin ligase that targets proteins for degradation and regulates cell growth, apoptosis and oxidative stress in various cell types. We have previously shown that Nrdp1 is implicated in ischemic cardiomyocyte death. In this study, we investigated the change of Nrdp1 expression in ischemic neurons and its role in ischemic neuronal injury. Primary rat cerebral cortical neurons and pheochromocytoma (PC12 cells were infected with adenoviral constructs expressing Nrdp1 gene or its siRNA before exposing to oxygen-glucose deprivation (OGD treatment. Our data showed that Nrdp1 was upregulated in ischemic brain tissue 3 h after middle cerebral artery occlusion (MCAO and in OGD-treated neurons. Of note, Nrdp1 overexpression by Ad-Nrdp1 enhanced OGD-induced neuron apoptosis, while knockdown of Nrdp1 with siRNA attenuated this effect, implicating a role of Nrdp1 in ischemic neuron injury. Moreover, Nrdp1 upregulation is accompanied by increased protein ubiquitylation and decreased protein levels of ubiquitin-specific protease 8 (USP8 in OGD-treated neurons, which led to a suppressed interaction between USP8 and HIF-1α and subsequently a reduction in HIF-1α protein accumulation in neurons under OGD conditions. In conclusion, our data support an important role of Nrdp1 upregulation in ischemic neuronal death, and suppressing the interaction between USP8 and HIF-1α and consequently the hypoxic adaptive response of neurons may account for this detrimental effect.

  4. Galanin Protects from Caspase-8/12-initiated Neuronal Apoptosis in the Ischemic Mouse Brain via GalR1

    Science.gov (United States)

    Li, Yun; Mei, Zhu; Liu, Shuiqiao; Wang, Tong; Li, Hui; Li, Xiao-Xiao; Han, Song; Yang, Yutao; Li, Junfa; Xu, Zhi-Qing David

    2017-01-01

    Galanin (GAL) plays key role in many pathophysiological processes, but its role in ischemic stroke remains unclear. Here, the models of 1 h middle cerebral artery occlusion (MCAO)/1-7 d reperfusion (R)-induced ischemic stroke and in vitro cell ischemia of 1 h oxygen-glucose deprivation (OGD)/24 h reoxygenation in primary cultured cortical neurons were used to explore GAL’s effects and its underlying mechanisms. The results showed significant increases of GAL protein levels in the peri-infarct region (P) and infarct core (I) within 48 h R of MCAO mice (p<0.001). The RT-qPCR results also demonstrated significant increases of GAL mRNA during 24-48 h R (p<0.001), and GAL receptors GalR1-2 (but not 3) mRNA levels in the P region at 24 h R of MCAO mice (p<0.001). Furthermore, the significant decrease of infarct volume (p<0.05) and improved neurological outcome (p<0.001-0.05) were observed in MCAO mice following 1 h pre- or 6 h post-treatment of GAL during 1-7 d reperfusion. GalR1 was confirmed as the receptor responsible for GAL-induced neuroprotection by using GalR2/3 agonist AR-M1896 and Lentivirus-based RNAi knockdown of GalR1. GAL treatment inhibited Caspase-3 activation through the upstream initiators Capsases-8/-12 (not Caspase-9) in both P region and OGD-treated cortical neurons. Meanwhile, GAL’s neuroprotective effect was not observed in cortical neurons from conventional protein kinase C (cPKC) γ knockout mice. These results suggested that exogenous GAL protects the brain from ischemic injury by inhibiting Capsase-8/12-initiated apoptosis, possibly mediated by GalR1 via the cPKCγ signaling pathway. PMID:28203483

  5. Cholesterol secoaldehyde, an ozonation product of cholesterol, induces amyloid aggregation and apoptosis in murine GT1-7 hypothalamic neurons.

    Science.gov (United States)

    Sathishkumar, K; Xi, Xiaochun; Martin, Roy; Uppu, Rao M

    2007-06-01

    Aldehydic products from ozonation of cholesterol and peroxidation of phospholipids have been shown to accelerate aggregation of amyloid-beta (Abeta) in vitro. Here, we show that 3beta-hydroxy-5-oxo-5,6-secocholestan-6-al (ChSeco), an ozonation product of cholesterol, induces Abeta aggregation, generation of reactive oxygen species (ROS), and cytotoxicity in murine GT1-7 hypothalamic neurons. The formation of Abeta aggregates in situ was dose-dependent at ChSeco concentrations ranging from 1 to 20 microM. The increase in insoluble Abeta aggregates at increasing concentrations of ChSeco was accompanied by a decrease in soluble Abeta as evidenced by Western blot analysis. The formation of ROS in neuronal cells was found to be dose- and time-dependent with the magnitude being higher at 20 microM compared to 10 microM ChSeco or untreated controls. The increase in ROS was associated with depletion of GSH. The cytotoxicity induced by ChSeco involved changes in phosphatidylserine translocation, DNA fragmentation, and caspase 3/7 activity that are characteristic of apoptosis. Pretreatment of neuronal cells with Trolox, a water-soluble analog of alpha-tocopherol offered partial, but significant protection against ChSeco-induced cell death, whereas, N-acetyl-L-cysteine (NAC) completely prevented the cytotoxic effects of ChSeco. NAC and Trolox were without any effects on ChSeco-induced Abeta aggregation. Fibrillogenesis inhibitors, which inhibited Abeta aggregation, did not inhibit cell death induced by ChSeco, implying that ROS generation, and not Abeta aggregation, plays a major role in the observed cytotoxicity. However, since Alzheimer's and other neurodegenerative diseases are slow and progressive, the formation of Abeta aggregates in vivo by ChSeco may have long-term pathological consequences.

  6. Effects of minocycline on apoptosis and neuronal changes in retinal ganglion cells from experimental optic neuritis rats

    Institute of Scientific and Technical Information of China (English)

    Jing Zhang

    2008-01-01

    pathological changes in the optic nerve met the changes in demyelinated optic neuritis. Moreover, there was significant apoptosis of RGCs. The percentage area of optic nerve axons in the transverse section was significantly increased and the number of apoptotic RGCs was increased after treatment with methylprednisolone and minocycline. Compared with methylprednisolone, minocycline had better effects on reducing RGC apoptosis (P < 0.05).CONCLUSION: Minocycline has better inhibitory effects on RGC apoptosis than methylprednisolone. Minocycline can decrease the damage to axons of demyelinated optic neuritis rats, and has similar protective effects on neurons from demyelinated optic neuritis rats as methylprednisolone.

  7. Effect of polygonatum polysaccharide on the hypoxia-induced apoptosis and necrosis in in vitro cultured cerebral cortical neurons from neonatal rats

    Institute of Scientific and Technical Information of China (English)

    Guozhu Hu; Jin Zhang; Ning Tang; Zhu Wen; Rongqing Nie

    2006-01-01

    BACKGROUND: Cardiocerebrovascular diseases induced cerebral circulation insufficiency and senile vascular dementia can result in ischemic/hypoxic apoptosis of central neurons, which we should pay more attention to and prevent and treat as early as possible. Traditional Chinese medicine possesses the unique advantage in this field. Polygonatum, a Chinese herb for invigorating qi, may play a role against the hypoxic apoptosis of brain neurons.OBJECTIVE: To observe the protective effect of polygonatum polysaccharide on hypoxia-induced apoptosis and necrosis in cerebral cortical neurons cultured in vitro.DESIGN: A comparative experiment.SETTING: Laboratory of Cell Biology, Institute of Basic Medical Sciences, Jiangxi Provincial Academy of Traditional Chinese Medicine.MATERIALS: The experiment was carried out in the Laboratory of Cell Biology, Institute of Basic Medical Sciences, Jiangxi Provincial Academy of Traditional Chinese Medicine from November 2003 to April 2005.Totally 218 Wistar rats (male or female) of clean degree within 24 hours after birth were purchased from the animal center of Jiangxi Medical College (certification number was 021-97-03).METHODS: ① Preparation of cerebral cortical neurons of rats: The cerebral cortical tissues were isolated from the Wistar rats within 24 hours after birth, and prepared to single cell suspension, and the cerebral cortical neurons of neonatal rats were in vitro cultured in serum free medium with Neurobasal plus B27Supplement. ② Observation on the non-toxic dosage of polygonatum polysaccharide on neurons: After the neurons were cultured for 4 days, polygonatum polysaccharide of different dosages (1-20 g/L) was added for continuous culture for 48 hours, the toxicity and non-toxic dosage of polygonatum polysaccharide on neurons were observed and detected with trypan blue staining. ③ Grouping: After hypoxia/reoxygenation,the cultured neurons were divided into normal control group, positive apoptotic group and polygonatum

  8. Arsenic binding to Fucus vesiculosus metallothionein.

    Science.gov (United States)

    Merrifield, Maureen E; Ngu, Thanh; Stillman, Martin J

    2004-11-05

    The seaweed Fucus vesiculosus is a member of the brown algae family. Kille and co-workers [Biochem. J. 338 (1999) 553] reported that this species contains the gene for metallothionein. Metallothionein is a metalloprotein having low molecular weight, and high cysteine content, which binds a range of metals. F. vesiculosus bioaccumulates arsenic from the aquatic environment [Mar. Chem. 18 (1986) 321]. In this paper we describe arsenic binding to F. vesiculosus metallothionein, characterized by electrospray ionization mass spectrometry. Five arsenic-MT species were detected with increasing As to protein ratios. These results provide important information about the metal-chelation behaviour of this novel algal metallothionein which is a putative model for arsenic binding to F. vesiculosus in vivo.

  9. Rolipram stimulates angiogenesis and attenuates neuronal apoptosis through the cAMP/cAMP-responsive element binding protein pathway following ischemic stroke in rats.

    Science.gov (United States)

    Hu, Shouye; Cao, Qingwen; Xu, Peng; Ji, Wenchen; Wang, Gang; Zhang, Yuelin

    2016-03-01

    Rolipram, a phosphodiesterase-4 inhibitor, can activate the cyclic adenosine monophosphate (cAMP)/cAMP-responsive element binding protein (CREB) pathway to facilitate functional recovery following ischemic stroke. However, to date, the effects of rolipram on angiogenesis and cerebral ischemia-induced neuronal apoptosis are yet to be fully elucidated. In this study, the aim was to reveal the effect of rolipram on the angiogenesis and neuronal apoptosis following brain cerebral ischemia. Rat models of ischemic stroke were established following transient middle cerebral artery occlusion and rolipram was administered for three, seven and 14 days. The results were examined using behavioral tests, triphenyl tetrazolium chloride staining, immunostaining and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) to evaluate the effects of rolipram therapy on functional outcome, angiogenesis and apoptosis. Western blot analysis was used to show the phosphorylated- (p-)CREB protein level in the ischemic hemisphere. The rolipram treatment group exhibited a marked reduction in infarct size and modified neurological severity score compared with the vehicle group, and rolipram treatment significantly promoted the microvessel density in the ischemic boundary region and increased p-CREB protein levels in the ischemic hemisphere. Furthermore, a significant reduction in the number of TUNEL-positive cells was observed in the rolipram group compared with the vehicle group. These findings suggest that rolipram has the ability to attenuate cerebral ischemic injury, stimulate angiogenesis and reduce neuronal apoptosis though the cAMP/CREB pathway.

  10. Effects of acrous gramineus and its component,alpha-asarone, on apoptosis of hippocampal neurons after seizure in immature rats

    Institute of Scientific and Technical Information of China (English)

    Libin Yang; Shulei Li; Yanzhi Huang; Jianmin Liang; Yuhong Wang

    2008-01-01

    BACKGROUND: α-asarone and acrous gramineus have been shown to play a necessary function in enhancing the reactivity and convulsant threshold to electric stimulation of immature rats. They have also been shown to effectively suppress epileptic seizures induced by pentylenetetrazol in young rats. However, the mechanisms for these roles have been still unclear.OBJECTIVE: To observe the effects in immature rats of acrous gramineus and α-asarone on apoptosis of hippocampal neurons after epileptic seizure at the protein level, and to analyze the mechanism for these effects.DESIGN: A randomized controlled animal experiment.SETTINGS: Department of Pediatrics, First Hospital of Jilin University; Department of Histology and Embryology, Norman Bethune Medical School of Jilin University; Department of Internal Medicine, Children's Hospital of Changchun City; Department of Neurology, First Clinical Hospital affiliated to Harbin Medical University.MATERIALS: Fifty 3-week old Wistar rats, 34-40 g, irrespective of gender, were provided by Gaoxin Research Center of Medical Animal Experiment, Changchun. The animals were treated according to the animal ethical standards. The following chemicals were used for this study: acrous gramineus powders or infusion (Batch No. 0307113, Tianjiang Medicine Company Limited, Jiangyin), α-asarone tablets (Batch No. 030219, Tianwei Pharmaceutil Factory, Shenyang), and phenobarbital sodium tablets (Batch No. 020608, Xinya Medicine Company Limited, Shanghai). The animals were divided into five groups randomly. First, ten rats were chosen as the normal controls. The remaining rats were treated with I.p. Injections of pentylenetetrazol to stimulate an epileptic model. METHODS: The experiments were performed at the Neurological Laboratory of the First Hospital of Jilin University between October and December 2004. The rats were treated with I.p. Injections of pentylenetetrazol (60 mg/kg) to establish an epileptic model. According to Racine' s standard

  11. Bone marrow mesenchymal stem cells attenuate 2,5-hexanedione-induced neuronal apoptosis through a NGF/AKT-dependent pathway

    Science.gov (United States)

    Wang, Qingshan; Sun, Guohua; Gao, Chenxue; Feng, Lina; Zhang, Yan; Hao, Jie; Zuo, Enjun; Zhang, Cong; Li, Shuangyue; Piao, Fengyuan

    2016-01-01

    Growing evidence suggests that the increased neuronal apoptosis is involved in n-hexane-induced neuropathy. We have recently reported that bone marrow-mesenchymal stem cells-derived conditioned medium (BMSC-CM) attenuated 2,5-hexanedione (HD, the active metabolite of n-hexane)-induced apoptosis in PC12 cells. Here, we explored the anti-apoptotic efficacy of BMSC in vivo. HD-treated rats received BMSC by tail vein injection 5 weeks after HD intoxication. We found that in grafted rats, BMSC significantly attenuated HD-induced neuronal apoptosis in the spinal cord, which was associated with elevation of nerve growth factor (NGF). Neutralization of NGF in BMSC-CM blocked the protection against HD-induced apoptosis in VSC4.1 cells, suggesting that NGF is essential for BMSC-afforded anti-apoptosis. Mechanistically, we found that the decreased activation of Akt induced by HD was significantly recovered in the spinal cord by BMSC and in VSC4.1 cells by BMSC-CM in a TrkA-dependent manner, leading to dissociation of Bad/Bcl-xL complex in mitochondria and release of anti-apoptotic Bcl-xL. The importance of Akt was further corroborated by showing the reduced anti-apoptotic potency of BMSC in HD-intoxicated VSC4.1 cells in the presence of Akt inhibitor, MK-2206. Thus, our findings show that BMSC attenuated HD-induced neuronal apoptosis in vivo through a NGF/Akt-dependent manner, providing a novel solution against n-hexane-induced neurotoxicity. PMID:27703213

  12. Genistein inhibition of OGD-induced brain neuron death correlates with its modulation of apoptosis, voltage-gated potassium and sodium currents and glutamate signal pathway.

    Science.gov (United States)

    Ma, Xue-Ling; Zhang, Feng; Wang, Yu-Xiang; He, Cong-Cong; Tian, Kun; Wang, Hong-Gang; An, Di; Heng, Bin; Liu, Yan-Qiang

    2016-07-25

    In the present study, we established an in vitro model of hypoxic-ischemia via exposing primary neurons of newborn rats to oxygen-glucose deprivation (OGD) and observing the effects of genistein, a soybean isoflavone, on hypoxic-ischemic neuron viability, apoptosis, voltage-activated potassium (Kv) and sodium (Nav) currents, and glutamate receptor subunits. The results indicated that OGD exposure reduced the viability and increased the apoptosis of brain neurons. Meanwhile, OGD exposure caused changes in the current-voltage curves and current amplitude values of voltage-activated potassium and sodium currents; OGD exposure also decreased GluR2 expression and increased NR2 expression. However, genistein at least partially reversed the effects caused by OGD. The results suggest that hypoxic-ischemia-caused neuronal apoptosis/death is related to an increase in K(+) efflux, a decrease in Na(+) influx, a down-regulation of GluR2, and an up-regulation of NR2. Genistein may exert some neuroprotective effects via the modulation of Kv and Nav currents and the glutamate signal pathway, mediated by GluR2 and NR2.

  13. A physically-modified saline suppresses neuronal apoptosis, attenuates tau phosphorylation and protects memory in an animal model of Alzheimer's disease.

    Science.gov (United States)

    Modi, Khushbu K; Jana, Arundhati; Ghosh, Supurna; Watson, Richard; Pahan, Kalipada

    2014-01-01

    Alzheimer's disease (AD), the leading cause of dementia in the aging population, is characterized by the presence of neuritic plaques, neurofibrillary tangles and extensive neuronal apoptosis. Neuritic plaques are mainly composed of aggregates of amyloid-β (Aβ) protein while neurofibrillary tangles are composed of the hyperphosphorylated tau protein. Despite intense investigations, no effective therapy is currently available to halt the progression of this disease. Here, we have undertaken a novel approach to attenuate apoptosis and tau phosphorylation in cultured neuronal cells and in a transgenic animal model of AD. RNS60 is a 0.9% saline solution containing oxygenated nanobubbles that is generated by subjecting normal saline to Taylor-Couette-Poiseuille (TCP) flow under elevated oxygen pressure. In our experiments, fibrillar Aβ1-42, but not the reverse peptide Aβ42-1, induced apoptosis and cell death in human SHSY5Y neuronal cells. RNS60, but not NS (normal saline), RNS10.3 (TCP-modified saline without excess oxygen) or PNS60 (saline containing excess oxygen without TCP modification), attenuated Aβ(1-42)-induced cell death. RNS60 inhibited neuronal cell death via activation of the type 1A phosphatidylinositol-3 (PI-3) kinase-Akt-BAD pathway. Furthermore, RNS60 also decreased Aβ(1-42)-induced tau phosphorylation via (PI-3 kinase-Akt)-mediated inhibition of GSK-3β. Similarly, RNS60 treatment suppressed neuronal apoptosis, attenuated Tau phosphorylation, inhibited glial activation, and reduced the burden of Aβ in the hippocampus and protected memory and learning in 5XFAD transgenic mouse model of AD. Therefore, RNS60 may be a promising pharmaceutical candidate in halting or delaying the progression of AD.

  14. A physically-modified saline suppresses neuronal apoptosis, attenuates tau phosphorylation and protects memory in an animal model of Alzheimer's disease.

    Directory of Open Access Journals (Sweden)

    Khushbu K Modi

    Full Text Available Alzheimer's disease (AD, the leading cause of dementia in the aging population, is characterized by the presence of neuritic plaques, neurofibrillary tangles and extensive neuronal apoptosis. Neuritic plaques are mainly composed of aggregates of amyloid-β (Aβ protein while neurofibrillary tangles are composed of the hyperphosphorylated tau protein. Despite intense investigations, no effective therapy is currently available to halt the progression of this disease. Here, we have undertaken a novel approach to attenuate apoptosis and tau phosphorylation in cultured neuronal cells and in a transgenic animal model of AD. RNS60 is a 0.9% saline solution containing oxygenated nanobubbles that is generated by subjecting normal saline to Taylor-Couette-Poiseuille (TCP flow under elevated oxygen pressure. In our experiments, fibrillar Aβ1-42, but not the reverse peptide Aβ42-1, induced apoptosis and cell death in human SHSY5Y neuronal cells. RNS60, but not NS (normal saline, RNS10.3 (TCP-modified saline without excess oxygen or PNS60 (saline containing excess oxygen without TCP modification, attenuated Aβ(1-42-induced cell death. RNS60 inhibited neuronal cell death via activation of the type 1A phosphatidylinositol-3 (PI-3 kinase-Akt-BAD pathway. Furthermore, RNS60 also decreased Aβ(1-42-induced tau phosphorylation via (PI-3 kinase-Akt-mediated inhibition of GSK-3β. Similarly, RNS60 treatment suppressed neuronal apoptosis, attenuated Tau phosphorylation, inhibited glial activation, and reduced the burden of Aβ in the hippocampus and protected memory and learning in 5XFAD transgenic mouse model of AD. Therefore, RNS60 may be a promising pharmaceutical candidate in halting or delaying the progression of AD.

  15. The effect of silver nanoparticles on apoptosis and dark neuron production in rat hippocampus

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    Farzaneh Bagheri-abassi

    2015-07-01

    Full Text Available Objective(s:Silver nanoparticles (Ag-NPs are used widely in bedding, water purification, tooth paste and toys. These nanoparticles can enter into the body and move into the hippocampus. The aim of this study was to investigate the neurotoxicity of silver nanoparticles in the adult rat hippocampus. Materials and Methods:12 male Wistar rats were randomly divided into two experimental and control groups (6 rats in each group. Animals in the experimental group received Ag-NPs (30 mg/kg orally (gavage for 28 consecutive days. Control group in the same period was treated with distilled water via gavage. At the end of experiment, animals were deeply anesthetized, sacrificed, and their brains were collected from each group. Finally the brain sections were stained using toluidine blue and TUNEL. Then to compare the groups, dark neurons (DNs and apoptotic neurons were counted by morphometric method. Results: Results showed that the num­bers of DNs and apoptotic cells in the CA1, CA2, CA3, and dentate gyrus (DG of hippocampus significantly increased in the Ag-NPs group in comparison to the control group (P

  16. Interleukin-1β enhances neuronal vulnerability to proNGF-mediated apoptosis by increasing surface expression of p75(NTR) and sortillin.

    Science.gov (United States)

    Choi, S; Friedman, W J

    2014-01-17

    Many types of injury such as seizure, ischemia, and oxidative stress cause upregulation of the p75 neurotrophin receptor (p75(NTR)) in brain neurons, where it promotes apoptosis, however the mechanism by which p75(NTR) is regulated under these conditions is not well understood. Proinflammatory cytokines such as interleukin-1β (IL-1β) are highly produced under these injury conditions and, in particular, are expressed rapidly in the rat hippocampus after seizure. IL-1β is known to increase neuronal vulnerability under many conditions, although it does not directly induce neuronal death. Recently, we have shown that these cytokines regulate p75(NTR) induction both in neurons and astrocytes in vitro. Here, we show that IL-1β infusion into the brain induces p75(NTR) in neurons of the CA1 area of the hippocampus. While IL-1β induction of p75(NTR) is not sufficient to induce cell death, we demonstrate that IL-1β primes the neurons by recruiting p75(NTR) and its coreceptor sortilin to the cell surface, making the neurons more vulnerable to subsequent challenge by proNGF. These results suggest a mechanism by which IL-1β exacerbates neuronal death following injury.

  17. Aldehyde dehydrogenase 2 overexpression inhibits neuronal apoptosis after spinal cord ischemia/reperfusion injury

    Directory of Open Access Journals (Sweden)

    Xing-zhen Liu

    2017-01-01

    Full Text Available Aldehyde dehydrogenase 2 (ALDH2 is an important factor in inhibiting oxidative stress and has been shown to protect against renal ischemia/reperfusion injury. Therefore, we hypothesized that ALDH2 could reduce spinal cord ischemia/reperfusion injury. Spinal cord ischemia/reperfusion injury was induced in rats using the modified Zivin's method of clamping the abdominal aorta. After successful model establishment, the agonist group was administered a daily consumption of 2.5% alcohol. At 7 days post-surgery, the Basso, Beattie, and Bresnahan score significantly increased in the agonist group compared with the spinal cord ischemia/reperfusion injury group. ALDH2 expression also significantly increased and the number of apoptotic cells significantly decreased in the agonist group than in the spinal cord ischemia/reperfusion injury group. Correlation analysis revealed that ALDH2 expression negatively correlated with the percentage of TUNEL-positive cells (r = −0.485, P < 0.01. In summary, increased ALDH2 expression protected the rat spinal cord against ischemia/reperfusion injury by inhibiting apoptosis.

  18. Molecular functions of metallothionein and its role in hematological malignancies

    Directory of Open Access Journals (Sweden)

    Takahashi Shinichiro

    2012-07-01

    Full Text Available Abstract Metallothionein (MT was reported to be a potential negative regulator of apoptosis, and various reports have suggested that it may play roles in carcinogenesis and drug resistance, in at least a portion of cancer cells. The author summarizes the current understanding of the molecular functions of MT for tumor cell growth and drug resistance. These activities are regulated through intracellular metal ion modulation and free radical scavenging. Compared with analyses of solid tumors, few studies have analyzed the roles of MT in hematological malignancies. This review mainly describes the functions of MT in hematopoietic cells. Furthermore, through expression analyses of leukemias and lymphomas, the roles of MT in the biology of these diseases are particularly focused upon.

  19. Protective effects of quercetin on dieldrin-induced endoplasmic reticulum stress and apoptosis in dopaminergic neuronal cells.

    Science.gov (United States)

    Park, Euteum; Chun, Hong Sung

    2016-10-19

    Dieldrin, an organochlorine pesticide still used in several developing countries, has been proposed as a risk factor for Parkinson's disease. Quercetin is one of the potent bioactive flavonoids present in numerous plants. In this study, we investigated the protective effects of quercetin on neurotoxicity induced by dieldrin in cultured dopaminergic SN4741 cells. Our initial experiments showed that quercetin (10-40 μM) dose dependently prevented dieldrin (20 μM)-induced cytotoxicity in SN4741 cells. Pretreatment for 1 h with quercetin before dieldrin application could significantly suppress dieldrin-induced apoptotic characteristics, including nuclear condensation, DNA fragmentation, and caspase-3/7 activation. Results showed that dieldrin-induced markers of endoplasmic reticulum (ER) stress response such as chaperone GRP78, heme oxygenase-1, and phosphorylation of the α subunit of eukaryotic initiation factor 2. In addition, dieldrin reduced antiapoptotic Bcl-2 expression, but significantly elevated a proapoptotic transcription factor CHOP. Furthermore, RNA interference to CHOP almost completely repressed dieldrin-induced apoptotic cell death. Interestingly, quercetin prevented the changes in dieldrin-induced ER stress markers. These results suggest that quercetin may suppress the ER stress-CHOP pathway and dieldrin-induced apoptosis in dopaminergic neurons.

  20. Activation of c—Jun and suppression of phospho—p44/42 were involved in diphenylhydantoin—induced apoptosis of cultured rat cerebellar granule neurons

    Institute of Scientific and Technical Information of China (English)

    ZHAOLing-Zhi; SUXing-Wen; HUANGYi-Jun; QIUPeng-Xin; YANGGuang-Mei

    2003-01-01

    AIM:To investigate possible intracellular signal molecules involved in diphenylhydantoin (DPH)-mediated apoptosis of cerebellar granule neurons (CGN) and explore possible nolecular mechanisms of neurotoxicity of DPH.METHODS: Fluorescein diacetate (FDA) stain, hochest 33258 stain, and agar gel electrophoresis were used to test morphological and biological characters of primary CGN and cortical neurons (CN) in the presence or absence of 100μmol/L DPH; Western blot and RT-PCR were employed to further investigate apoptotic/survival signal moleculars involved in the neuronal apoptotic signal transdution. RESULTS:DPH 100μmol/L induced a typical apoptosis of CGN but had no toxicity on CN. Cerebellar granule neural apoptosis induced by 100μmol/L DPH was significantly inhibited by pre-treatment with SB203580(10μmol/L) or CEP-11004(1μmol/L) for 1h. DPH markedly upregulated the levels of phospho-c-Jun (active c-Jun), total c-Jun protein and c-jun mRNA in CGN. The levels of phospho-c-Jun dramatically elevated by DPH at 8 h were significantly inhibited by SB203580(10μmol/L) or CEP-11004 (1μmol/L). Moreover, the activities of p44/42 (ERK1/ERK2), other members of MAP kinases and generally believed to be important survival effetors in CGN, were markedly suppressed. However, the activities of both JNK and p38 were little affected in the process of apoptosis of CGN induced by 100μmol/L DPH. CONCLUSION: The selective toxicity of DPH on CGN is likely due to its ability to induce apoptosis of CGN, it is a process involved activation of c-Jun and suppression of the activity of p44/42.

  1. Interleukin-1 beta induction of neuron apoptosis depends on p38 mitogen-activated protein kinase activity after spinal cord injury

    Institute of Scientific and Technical Information of China (English)

    Xin-jia WANG; Kang-mei KONG; Wei-li QI; Wei-lian YE; Pei-song SONG

    2005-01-01

    Aim: Interleukin-1 beta (IL-1β) has been implicated as an extracellular signal in the initiation of apoptosis in neurons and oligodendrocytes after spinal cord injury (SCI). To further characterize the apoptotic cascade initiated by IL-1β after SCI, we examined the expression of IL-1 β, p38 mitogen-activated protein kinase (p38 MAPK) and caspase-3 after SCI, and further investigated whether p38 MAPK was involved in neuron apoptosis induced by IL-1 β. Methods: Adult rats were given contusion SCI at the T-10 vertebrae level with a weight-drop impactor (10 g weight dropped 25.0 mm). The expression levels of IL-1β, p38 MAPK and caspase-3after SCI were assessed with Western blots, immunohistochemistry staining, and real time reverse transcription polymerase chain reactions (RT-PCR). Neuron apoptosis was assessed with the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) method. Results:Increased levels of IL-1β and p38 MAPK were observed soon after injury, with a peak in expression levels within 6 h of injury. By 24 h after injury, caspase-3expression was markedly increased in the injured spinal cord. TUNEL-positive cells were first observed in the lesioned area 6 h after SCI. The largest number of TUNEL-positive cells was observed at 24 h post-SCI. Intrathecal injection of the IL-1 receptor antagonist IL-1Ra significantly reduced expression of p38 MAPK and caspase-3, and reduced the number of TUNEL-positive cells. Moreover,intrathecal injection of an inhibitor of p38 MAPK, SB203580, also significantly reduced the expression of caspase-3, and reduced the number of TUNEL-positive cells in the injured spinal cord. Conclusion: The p38MAPK signaling pathway plays an important role in IL-1β mediated induction of neuron apoptosis following SCI in rats.

  2. How do Chinese medicines that tonify the kidney inhibit dopaminergic neuron apoptosis?**

    Institute of Scientific and Technical Information of China (English)

    Shaogang Lin; Shuifen Ye; Jinmu Huang; Yun Tian; Yihui Xu; Mengqi Wu; Jingxia Wang; Songying Wu; Jing Cai

    2013-01-01

    Wistar rats were intragastrical y perfused with Chinese medicines used for tonifying the kidney. These included 0.180 g/mL of Herba Epimedi (Epimedium), Semen Cuscutae (Dodder Seed), or Herba Cistanches (Desertliving Cistanche), 0.04 mg/mL monoamine oxidase-B inhibitor selegiline, or distil ed water for 14 consecutive days to prepare drug-containing serum or blank serum. MES23.5 cells in the logarithmic phase were cultured in media supplemented with 15%drug-containing serum for 24 hours, fol owed by incubation in culture solution containing 100μmol/L H2O2 for 3 hours. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and flow tometry results showed that al drug-containing serums improved the survival rate of H 2 O 2-injured MES23.5 cells, inhibited pro-apoptotic FasL and caspase-3 expression, promoted anti-apoptotic Bcl-2 expression. However, drug-containing serums had little influence on Fas expression in H 2 O 2-injured MES23.5 cells. Enzyme-linked immunosorbent assay results showed that serum containing Herba Cistanches or Herba Epimedi increased the expression of nerve growth factor, brain-derived neurotrophic factor, and glial cellline-derived neurotrophic factor in injured MES23.5 cells;serum containing Semen Cuscutae only increased brain-derived neurotrophic factor expres-sion; while expression of the above neurotrophic factors remained the same in cells treated with serum containing selegiline. These findings indicate that Chinese medicines used to tonify the kid-ney can protect nerve cells by regulating the expression of apoptosis-related factors and neuro-trophic factors in MES23.5 cells.

  3. Arsenic Induction of Metallothionein and Metallothionein Induction Against Arsenic Cytotoxicity.

    Science.gov (United States)

    Rahman, Mohammad Tariqur; De Ley, Marc

    Human exposure to arsenic (As) can lead to oxidative stress that can become evident in organs such as the skin, liver, kidneys and lungs. Several intracellular antioxidant defense mechanisms including glutathione (GSH) and metallothionein (MT) have been shown to minimize As cytotoxicity. The current review summarizes the involvement of MT as an intracellular defense mechanism against As cytotoxicity, mostly in blood. Zinc (Zn) and selenium (Se) supplements are also proposed as a possible remediation of As cytotoxicity. In vivo and in vitro studies on As toxicity were reviewed to summarize cytotoxic mechanisms of As. Intracellular antioxidant defense mechanisms of MT are linked in relation to As cytotoxicity. Arsenic uses a different route, compared to major metal MT inducers such as Zn, to enter/exit blood cells. A number of in vivo and in vitro studies showed that upregulated MT biosynthesis in blood components are related to toxic levels of As. Despite the cysteine residues in MT that aid to bind As, MT is not the preferred binding protein for As. Nonetheless, intracellular oxidative stress due to As toxicity can be minimized, if not eliminated, by MT. Thus MT induction by essential metals such as Zn and Se supplementation could be beneficial to fight against As toxicity.

  4. Quantification of sPLA2-induced early and late apoptosis changes in neuronal cell cultures using combined TUNEL and DAPI staining.

    Science.gov (United States)

    Daniel, Bron; DeCoster, Mark A

    2004-08-01

    The terminal deoxynucleotidyl transferase (TdT) dUTP nick end labeling (TUNEL) stain is in wide use for measuring apoptosis in neurons, as well as in other cell types. TUNEL may give false positive results due to variations in labeling technique as well as staining of cells that have undergone non-apoptotic DNA strand breaks. Therefore, in isolation, TUNEL is not a certain indicator of apoptosis. Recently, we have demonstrated the potent apoptotic effect of secreted phospholipase A2 from group III (sPLA2-III) on primary cortical neurons from rat. Here we describe a computer-assisted method for quantifying TUNEL-positive neurons after sPLA2-III induced apoptosis. Extent of TUNEL is normalized to total nuclear content using 4',6-diamidino-2-phenylindole (DAPI) staining. Furthermore, DAPI counterstaining allows for determination of a nuclear morphology indicator, based on nuclear size and roundness, which we call the nuclear area factor. We found that the nuclear area factor is an early indicator of cell death (significant after 4 h post treatment), while TUNEL staining is significant at later times (26 h). Thus, the independent staining techniques using TUNEL and DAPI complement each other, and with commercially available image analysis software, may be used to indicate early as well as delayed cell injury processes.

  5. Mechanisms underlying attenuation of apoptosis of cortical neurons in the hypoxic brain by lfavonoids from the stems and leaves of Scutellaria baicalensis Georgi

    Institute of Scientific and Technical Information of China (English)

    Guangxin Miao; Hongxiang Zhao; Ke Guo; Jianjun Cheng; Shufeng Zhang; Xiaofeng Zhang; Zhenling Cai; Hong Miao; Yazhen Shang

    2014-01-01

    Flavonoids from the stems and leaves of Scutellaria baicalensis Georgi, an antioxidant, marked-ly improve memory impairments and neuronal injuries. In the present study, primary cortical neurons of rats were exposed to potassium cyanide to establish a model of in vitro neural cell apoptosis. Inhibition of apoptosis by lfavonoids from the stems and leaves of Scutellaria baical-ensis Georgi at concentrations of 18.98, 37.36, and 75.92μg/mL was detected using this model. These lfavonoids dramatically increased cell survival, inhibited cell apoptosis and excessive pro-duction of malondialdehyde, and increased the activities of superoxide dismutase, glutathione peroxidase, and Na+-K+-ATPase in primary cortical neurons exposed to potassium cyanide. The lfavonoids from the stems and leaves of Scutellaria baicalensis Georgi were originally found to have a polyhydric structure and to protect against cerebral hypoxia in in vitro and in vivo models, including hypoxia induced by potassium cyanide or cerebral ischemia. The present study suggests that lfavonoids from the stems and leaves of Scutellaria baicalensis Georgi exert neuroprotective effects via modulation of oxidative stress, such as malondialdehyde, superoxide dismutase, gluta-thione peroxidase and Na+-K+-ATPase disorders induced by potassium cyanide.

  6. Glucocorticoids increase impairments in learning and memory due to elevated amyloid precursor protein expression and neuronal apoptosis in 12-month old mice.

    Science.gov (United States)

    Li, Wei-Zu; Li, Wei-Ping; Yao, Yu-You; Zhang, Wen; Yin, Yan-Yan; Wu, Guo-Cui; Gong, Hui-Ling

    2010-02-25

    Alzheimer's disease is a chronic neurodegenerative disorder marked by a progressive loss of memory and cognitive function. Stress level glucocorticoids are correlated with dementia progression in patients with Alzheimer's disease. In this study, twelve month old male mice were chronically treated for 21 days with stress-level dexamethasone (5mg/kg). We investigated the pathological consequences of dexamethasone administration on learning and memory impairments, amyloid precursor protein processing and neuronal cell apoptosis in 12-month old male mice. Our results indicate that dexamethasone can induce learning and memory impairments, neuronal cell apoptosis, and mRNA levels of the amyloid precursor protein, beta-secretase and caspase-3 are selectively increased after dexamethasone administration. Immunohistochemistry demonstrated that amyloid precursor protein, caspase-3 and cytochrome c in the cortex and CA1, CA3 regions of the hippocampus are significantly increased in 12-month old male mice. Furthermore, dexamethasone treatment induced cortex and hippocampus neuron apoptosis as well as increasing the activity of caspase-9 and caspase-3. These findings suggest that high levels of glucocorticoids, found in Alzheimer's disease, are not merely a consequence of the disease process but rather play a central role in the development and progression of Alzheimer's disease. Stress management or pharmacological reduction of glucocorticoids warrant additional consideration of the regimen used in Alzheimer's disease therapies.

  7. Rapid eye movement sleep loss induces neuronal apoptosis in the rat brain by noradrenaline acting on alpha 1-adrenoceptor and by triggering mitochondrial intrinsic pathway

    Directory of Open Access Journals (Sweden)

    Bindu I Somarajan

    2016-03-01

    Full Text Available Many neurodegenerative disorders are associated with rapid eye movement sleep (REMS-loss, however the mechanism was unknown. As REMS-loss elevates noradrenaline (NA level in the brain as well as induces neuronal apoptosis and degeneration, in this study we have delineated the intracellular molecular pathway involved in REMS deprivation (REMSD associated NA-induced neuronal apoptosis. Rats were REMS deprived for 6 days by the classical flower-pot method, suitable controls were conducted and the effects on apoptosis markers evaluated. Further, the role of NA was studied by one, intraperitoneal (i.p. injection of NA-ergic alpha1-adrenoceptor antagonist prazosin (PRZ and two, by down-regulation of NA synthesis in locus coeruleus (LC neurons by local microinjection of tyrosine hydroxylase siRNA (TH-siRNA. Immunoblot estimates showed that the expressions of pro-apoptotic proteins viz. Bcl2-associated death promoter (BAD protein, apoptotic protease activating factor-1 (Apaf-1, cytochrome c, caspase9, caspase3 were elevated in the REMS-deprived rat brains, while caspase8 level remained unaffected; PRZ treatment did not allow elevation of these pro-apoptotic factors. Further, REMSD increased cytochrome c expression, which was prevented if the NA synthesis from the LC neurons was blocked by microinjection of TH-siRNA in vivo into the LC during REMSD in freely moving normal rats. Mitochondrial damage was re-confirmed by transmission electron microscopy (TEM, which showed distinctly swollen mitochondria with disintegrated cristae, chromosomal condensation and clumping along the nuclear membrane and all these changes were prevented in PRZ treated rats. Combining findings of this study along with earlier reports we propose that upon REMSD NA level increases in the brain as the LC NA-ergic REM-OFF neurons do not cease firing and TH is up-regulated in those neurons. This elevated NA acting on alpha1-adrenoceptors damages mitochondria causing release of

  8. 金属硫蛋白对热应激下体外培养奶牛淋巴细胞的影响%Effect of metallothionein on lymphocyte apoptosis of the dairy cow cultured in vitro under heat stress

    Institute of Scientific and Technical Information of China (English)

    占今舜; 吴力专; 李丽立; 张彬

    2014-01-01

    本研究的目的在于探讨外源性金属硫蛋白(MT)对热应激下体外培养奶牛淋巴细胞凋亡的影响。将屠宰获取的奶牛脾脏淋巴细胞随机分为4组,每组细胞培养液中MT的终浓度分别为0μg/mL(对照组)、70μg/mL(第Ⅰ组)、140μg/mL(第Ⅱ组)、210μg/mL(第Ⅲ组),同时置于细胞培养箱(37℃、5%CO2)中培养20 h,之后同时置于43℃水浴锅中进行热应激处理1 h,再回细胞培养箱中培养3 h,然后利用形态学观察、RT-PCR等方法进行各项指标的检测。结果如下,1)通过形态学观察和AO/EB染色观察,表明添加MT可以有效缓解热应激对奶牛淋巴细胞的损伤,但缓解效果随 MT浓度的增加而下降;2)第Ⅰ~Ⅲ组的CAT活性、SOD活性和 GSH-Px活性均高于对照组(P<0.01),MDA含量均低于对照组(P<0.01),表明 MT能够有效改善淋巴细胞的抗氧化能力;3)试验Ⅰ~Ⅲ组的Bax基因相对表达量均显著低于对照组(P<0.01),但试验Ⅰ~Ⅲ组Bcl-2基因的表达量和p50基因的表达量均显著低于对照组(P<0.01),表明MT可以调节与细胞凋亡相关基因的表达。综上所述,添加MT可以降低奶牛淋巴细胞死亡率,改善奶牛淋巴细胞抗氧化性能,抑制奶牛淋巴细胞凋亡,以添加终浓度为70μg/mL的 MT效果最好。%The effect of exogenous MT on lymphocyte apoptosis of the dairy cow was studied invitro under heat stress.The splenic dairy cow lymphocytes were divided into 4 groups which had final concentrations of MT 0 (control group),70 (groupⅠ),140 (groupⅡ)and 210μg/mL (groupⅢ).The cells of each group were cul-tured in a cell incubator (37℃,5%CO2 ).After 20 h,the cells of each group were put into water bath pot (43℃)for 1 h,and then recultured in a cell incubator (37℃,5%CO2)for 3 h.Finally,various indexes were tested by observation of morphology and RT-PCR.1)MT added in cell

  9. De-repression of FOXO3a death axis by microRNA-132 and -212 causes neuronal apoptosis in Alzheimer's disease.

    Science.gov (United States)

    Wong, Hon-Kit Andus; Veremeyko, Tatiana; Patel, Nehal; Lemere, Cynthia A; Walsh, Dominic M; Esau, Christine; Vanderburg, Charles; Krichevsky, Anna M

    2013-08-01

    Alzheimer's disease (AD) is a multifactorial and fatal neurodegenerative disorder for which the mechanisms leading to profound neuronal loss are incompletely recognized. MicroRNAs (miRNAs) are recently discovered small regulatory RNA molecules that repress gene expression and are increasingly acknowledged as prime regulators involved in human brain pathologies. Here we identified two homologous miRNAs, miR-132 and miR-212, downregulated in temporal cortical areas and CA1 hippocampal neurons of human AD brains. Sequence-specific inhibition of miR-132 and miR-212 induces apoptosis in cultured primary neurons, whereas their overexpression is neuroprotective against oxidative stress. Using primary neurons and PC12 cells, we demonstrate that miR-132/212 controls cell survival by direct regulation of PTEN, FOXO3a and P300, which are all key elements of AKT signaling pathway. Silencing of these three target genes by RNAi abrogates apoptosis caused by the miR-132/212 inhibition. We further demonstrate that mRNA and protein levels of PTEN, FOXO3a, P300 and most of the direct pro-apoptotic transcriptional targets of FOXO3a are significantly elevated in human AD brains. These results indicate that the miR-132/miR-212/PTEN/FOXO3a signaling pathway contributes to AD neurodegeneration.

  10. Genome-wide microRNA profiling of rat hippocampus after status epilepticus induced by amygdala stimulation identifies modulators of neuronal apoptosis.

    Directory of Open Access Journals (Sweden)

    Zhen Sun

    Full Text Available MicroRNAs (miRNAs are small and endogenously expressed non-coding RNAs that negatively regulate the expression of protein-coding genes at the translational level. Emerging evidence suggests that miRNAs play critical roles in central nervous system under physiological and pathological conditions. However, their expression and functions in status epilepticus (SE have not been well characterized thus far. Here, by using high-throughput sequencing, we characterized miRNA expression profile in rat hippocampus at 24 hours following SE induced by amygdala stimulation. After confirmation by qRT-PCR, six miRNAs were found to be differentially expressed in brain after SE. Subsequent Kyoto Encyclopedia of Genes and Genomes pathway analysis indicated that most of the predicted target genes for these six miRNAs were related to neuronal apoptosis. We then investigated the dynamic changes of these six miRNAs at different time-point (4 hours, 24 hours, 1 week and 3 weeks after SE. Meanwhile, neuronal survival and apoptosis in the hippocampus after SE were evaluated by Nissl staining and terminal deoxynucleotidyl transferase-mediated dUTP end-labeling assay. We found that the expression of miR-874-3p, miR-20a-5p, miR-345-3p, miR-365-5p, and miR-764-3p were significantly increased from 24 hours to 1 week, whereas miR-99b-3p level was markedly decreased from 24 hours to 3 weeks after SE. Further analysis revealed that the levels of miR-365-5p and miR-99b-3p were significantly correlated with neuronal apoptosis after SE. Taken together, our data suggest that miRNAs are important modulators of SE-induced neuronal apoptosis. These findings also open new avenues for future studies aimed at developing strategies against neuronal apoptosis after SE.

  11. Third-Degree Hindpaw Burn Injury Induced Apoptosis of Lumbar Spinal Cord Ventral Horn Motor Neurons and Sciatic Nerve and Muscle Atrophy in Rats

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    Sheng-Hua Wu

    2015-01-01

    Full Text Available Background. Severe burns result in hypercatabolic state and concomitant muscle atrophy that persists for several months, thereby limiting patient recovery. However, the effects of burns on the corresponding spinal dermatome remain unknown. This study aimed to investigate whether burns induce apoptosis of spinal cord ventral horn motor neurons (VHMNs and consequently cause skeletal muscle wasting. Methods. Third-degree hindpaw burn injury with 1% total body surface area (TBSA rats were euthanized 4 and 8 weeks after burn injury. The apoptosis profiles in the ventral horns of the lumbar spinal cords, sciatic nerves, and gastrocnemius muscles were examined. The Schwann cells in the sciatic nerve were marked with S100. The gastrocnemius muscles were harvested to measure the denervation atrophy. Result. The VHMNs apoptosis in the spinal cord was observed after inducing third-degree burns in the hindpaw. The S100 and TUNEL double-positive cells in the sciatic nerve increased significantly after the burn injury. Gastrocnemius muscle apoptosis and denervation atrophy area increased significantly after the burn injury. Conclusion. Local hindpaw burn induces apoptosis in VHMNs and Schwann cells in sciatic nerve, which causes corresponding gastrocnemius muscle denervation atrophy. Our results provided an animal model to evaluate burn-induced muscle wasting, and elucidate the underlying mechanisms.

  12. Modeling of kiwifruit metallothionein kiwi503

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Kiwifruit metallothionein, kiwi503, is a typical plant metallothionein protein. It has 63 amino acid residues in two cysteine-rich regions and one spacer region of about 32 residues. In this note, the two cysteine-rich regions and the spacer region have been modeled separately by the distance geometry and the homology method. The three parts are then connected to generate a three-dimensional structural model of kiwifruit metallothionein kiwi503. The result shows that there is no structural or energy barrier preventing either cysteine rich domain from independently forming a metal-sulfur chelating cluster. The method can also be applied to predicting the structures of the same type of other proteins.

  13. Tetramethylpyrazine suppresses transient oxygen-glucose deprivation-induced connexin32 expression and cell apoptosis via the ERK1/2 and p38 MAPK pathway in cultured hippocampal neurons.

    Directory of Open Access Journals (Sweden)

    Gu Gong

    Full Text Available Tetramethylpyrazine (TMP has been widely used in China as a drug for the treatment of various diseases. Recent studies have suggested that TMP has a protective effect on ischemic neuronal damage. However, the exact mechanism is still unclear. This study aims to investigate the mechanism of TMP mediated ischemic hippocampal neurons injury induced by oxygen-glucose deprivation (OGD. The effect of TMP on hippocampal neurons viability was detected by MTT assay, LDH release assay and apoptosis rate was measured by flow cytometry. TMP significantly suppressed neuron apoptosis in a concentration-dependent manner. TMP could significantly reduce the elevated levels of connexin32 (Cx32 induced by OGD. Knockdown of Cx32 by siRNA attenuated OGD injury. Moreover, our study showed that viability was increased in siRNA-Cx32-treated-neurons, and neuron apoptosis was suppressed by activating Bcl-2 expression and inhibiting Bax expression. Over expression of Cx32 could decrease neurons viability and increase LDH release. Furthermore, OGD increased phosphorylation of ERK1/2 and p38, whose inhibitors relieved the neuron injury and Cx32 up-regulation. Taken together, TMP can reverse the OGD-induced Cx32 expression and cell apoptosis via the ERK1/2 and p38 MAPK pathways.

  14. L-Satropane Prevents Retinal Neuron Damage by Attenuating Cell Apoptosis and Aβ Production via Activation of M1 Muscarinic Acetylcholine Receptor.

    Science.gov (United States)

    Yu, Ping; Zhou, Wei; Liu, Lu; Tang, Ya-Bin; Song, Yun; Lu, Juan-Juan; Hou, Li-Na; Chen, Hong-Zhuan; Cui, Yong-Yao

    2017-09-01

    Muscarinic acetylcholine receptor (mAChR) agonists have been used to treat glaucoma due to their intraocular pressure-lowering effects. Recently, it has been reported that retinal mAChRs activation can also stimulate neuroprotective pathways. In our study, we evaluated the potential neuroprotective effect of L-satropane, a novel mAChR agonist, on retinal neuronal injury induced by cobalt chloride (CoCl2) and ischemia/reperfusion (I/R). CoCl2-induced hypoxia injury in cultured cell models and I/R-induced retinal neuronal damage in rats in vivo were used to evaluate the abilities of L-satropane. In detail, we measured the occurrence of retinal pathological changes including molecular markers of neuronal apoptosis and Aβ expression. Pretreatment with L-satropane protects against CoCl2-induced neurotoxicity in PC12 and primary retinal neuron (PRN) cells in a dose-dependent manner by increasing retinal neuron survival. CoCl2 or I/R-induced cell apoptosis by upregulating Bax expression and downregulating Bcl-2 expression, which resulted in an increased Bax/Bcl-2 ratio, and upregulating caspase-3 expression/activity was significantly reversed by L-satropane treatment. In addition, L-satropane significantly inhibited the upregulation of Aβ production in both retinal neurons and tissue. We also found that I/R-induced histopathological retinal changes including cell loss in the retinal ganglion cell layer (GCL) and increased TUNEL positive retinal ganglion cells in GCL and thinning of the inner plexiform layer (IPL) and inner nuclear layer (INL) were markedly improved by L-satropane. The effects of L-satropane were largely abolished by the nonselective mAChRs antagonist atropine and M1-selective mAChR antagonist pirenzepine. These results demonstrated that L-satropane might be effective in preventing retinal neuron damage caused by CoCl2 or I/R. The neuroprotective effects of L-satropane may be attributed to decreasing cell apoptosis and Aβ production through activation

  15. Protection of ischemic postconditioning against neuronal apoptosis induced by transient focal ischemia is associated with attenuation of NF-κB/p65 activation.

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    Jianmin Liang

    Full Text Available BACKGROUND AND PURPOSE: Accumulating evidences have demonstrated that nuclear factor κB/p65 plays a protective role in the protection of ischemic preconditioning and detrimental role in lethal ischemia-induced programmed cell death including apoptosis and autophagic death. However, its role in the protection of ischemic postconditioning is still unclear. METHODS: Rat MCAO model was used to produce transient focal ischemia. The procedure of ischemic postconditioning consisted of three cycles of 30 seconds reperfusion/reocclusion of MCA. The volume of cerebral infarction was measured by TTC staining and neuronal apoptosis was detected by TUNEL staining. Western blotting was used to analyze the changes in protein levels of Caspase-3, NF-κB/p65, phosphor- NF-κB/p65, IκBα, phosphor- IκBα, Noxa, Bim and Bax between rats treated with and without ischemic postconditioning. Laser scanning confocal microscopy was used to examine the distribution of NF-κB/p65 and Noxa. RESULTS: Ischemic postconditioning made transient focal ischemia-induced infarct volume decrease obviously from 38.6% ± 5.8% to 23.5% ± 4.3%, and apoptosis rate reduce significantly from 46.5% ± 6.2 to 29.6% ± 5.3% at reperfusion 24 h following 2 h focal cerebral ischemia. Western blotting analysis showed that ischemic postconditioning suppressed markedly the reduction of NF-κB/p65 in cytoplasm, but elevated its content in nucleus either at reperfusion 6 h or 24 h. Moreover, the decrease of IκBα and the increase of phosphorylated IκBα and phosphorylated NF-κB/p65 at indicated reperfusion time were reversed by ischemic postconditioning. Correspondingly, proapoptotic proteins Caspase-3, cleaved Caspase-3, Noxa, Bim and Bax were all mitigated significantly by ischemic postconditioning. Confocal microscopy revealed that ischemic postconditioning not only attenuated ischemia-induced translocation of NF-κB/p65 from neuronal cytoplasm to nucleus, but also inhibited the abnormal

  16. Microglia-derived proinflammatory cytokines tumor necrosis factor-alpha and interleukin-1beta induce Purkinje neuronal apoptosis via their receptors in hypoxic neonatal rat brain.

    Science.gov (United States)

    Kaur, Charanjit; Sivakumar, Viswanathan; Zou, Zhirong; Ling, Eng-Ang

    2014-01-01

    The developing cerebellum is extremely vulnerable to hypoxia which can damage the Purkinje neurons. We hypothesized that this might be mediated by tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) derived from activated microglia as in other brain areas. One-day-old rats were subjected to hypoxia following, which the expression changes of various proteins in the cerebellum including hypoxia inducible factor-1α, TNF-α, IL-1β, TNF-R1 and IL-1R1 were analyzed. Following hypoxic exposure, TNF-α and IL-1β immunoexpression in microglia was enhanced coupled by that of TNF-R1 and IL-1R1 in the Purkinje neurons. Along with this, hypoxic microglia in vitro showed enhanced release of TNF-α and IL-1β whose receptor expression was concomitantly increased in the Purkinje neurons. In addition, nitric oxide (NO) level was significantly increased in the cerebellum and cultured microglia subjected to hypoxic exposure. Moreover, cultured Purkinje neurons treated with conditioned medium derived from hypoxic microglia underwent apoptosis but the incidence was significantly reduced when the cells were treated with the same medium that was neutralized with TNF-α/IL-1β antibody. We conclude that hypoxic microglia in the neonatal cerebellum produce increased amounts of NO, TNF-α and IL-1β which when acting via their respective receptors could induce Purkinje neuron death.

  17. Linoleic acid derivative DCP-LA protects neurons from oxidative stress-induced apoptosis by inhibiting caspase-3/-9 activation.

    Science.gov (United States)

    Yaguchi, Takahiro; Fujikawa, Hirokazu; Nishizaki, Tomoyuki

    2010-05-01

    The present study aimed at understanding the effect of the linoleic acid derivative 8-[2-(2-pentyl-cyclopropylmethyl)-cyclopropyl]-octanoic acid (DCP-LA) on oxidative stress-induced neuronal death. Sodium nitroprusside (SNP; 1 mM) reduced viability of cultured rat cerebral cortical neurons to 50% of basal levels, but DCP-LA significantly prevented the SNP effect in a concentration (1-100 nM)-dependent manner. In addition, DCP-LA (100 nM) rescued neurons from SNP-induced degradation. SNP (1 mM) activated caspase-3 and -9 in cultured rat cerebral cortical neurons, but DCP-LA (100 nM) abolished the caspase activation. For a mouse model of middle cerebral artery occlusion, oral administration with DCP-LA (1 mg/kg) significantly diminished degraded area due to cerebral infarction. The results of the present study, thus, demonstrate that DCP-LA protects neurons at least in part from oxidative stress-induced apoptosis by inhibiting activation of caspase-3/-9.

  18. Telmisartan protects central neurons against nutrient deprivation-induced apoptosis in vitro through activation of PPARγ and the Akt/GSK-3β pathway

    Science.gov (United States)

    Pang, Tao; Sun, Li-xin; Wang, Tao; Jiang, Zhen-zhou; Liao, Hong; Zhang, Lu-yong

    2014-01-01

    Aim: To determine whether angiotensin II receptor blockers (ARBs) could protect central neurons against nutrient deprivation-induced apoptosis in vitro and to elucidate the underlying mechanisms. Methods: Primary rat cerebellar granule cells (CGCs) underwent B27 (a serum substitute) deprivation for 24 h to induce neurotoxicity, and cell viability was analyzed using LDH assay and WST-1 assay. DNA laddering assay and TUNEL assay were used to detect cell apoptosis. The expression of caspase-3 and Bcl-2, and the phosphorylation of Akt and GSK-3β were detected using Western blot analysis. AT1a mRNA expression was determined using RT-PCR analysis. Results: B27 deprivation significantly increased the apoptosis of CGCs, as demonstrated by LDH release, DNA laddering, caspase-3 activation and positive TUNEL staining. Pretreatment with 10 μmol/L ARBs (telmisartan, candesartan or losartan) partially blocked B27 deprivation-induced apoptosis of CGCs with telmisartan being the most effective one. B27 deprivation markedly increased the expression of AT1a receptor in CGCs, inhibited Akt and GSK-3β activation, decreased Bcl-2 level, and activated caspase-3, which were reversed by pretreatment with 1 μmol/L telmisartan. In addition, pretreatment with 10 μmol/L PPARγ agonist pioglitazone was more effective in protecting CGCs against B27 deprivation-induced apoptosis, whereas pretreatment with 20 μmol/L PPARγ antagonist GW9662 abolished all the effects of telmisartan in CGCs deprived of B27. Conclusion: ARBs, in particular telmisartan, can protect the nutrient deprivation-induced apoptosis of CGCs in vitro through activation of PPARγ and the Akt/GSK-3β pathway. PMID:24793312

  19. Umbelliferone reverses depression-like behavior in chronic unpredictable mild stress-induced rats by attenuating neuronal apoptosis via regulating ROCK/Akt pathway.

    Science.gov (United States)

    Qin, Tingting; Fang, Fang; Song, Meiting; Li, Ruipeng; Ma, Zhanqiang; Ma, Shiping

    2017-01-15

    There is increasing evidence that major depressive disorder (MDD) is also a progressive neurodegeneration disorder and neuronal damage is the major pathology of MDD. Umbelliferone, a coumarin derivative, was found in a range of plants with proved anti-oxidative, anti-inflammatory and neuroprotective effects. The primary purpose of this investigation was to evaluate whether umbelliferone could confer an antidepressant-like effect on the depressive model in rats developed by chronic unpredictable mild stress (CUMS) and explore the possible mechanism involved in its neuroprotective effects. We found that treatments with umbelliferone (15mg/kg, 30mg/kg) significantly ameliorated CUMS-induced depressive-like behaviors, such as decreased sucrose consumption, reduced locomotor activity and prolonged immobility time. Rats under CUMS stimulation treated with umbelliferone (15mg/kg, 30mg/kg) showed reduced neuronal apoptosis, as well as inhibited inflammatory cytokines levels by down-regulating Rho-associated protein kinase (ROCK) signaling and up-regulating protein kinase B (Akt) signaling. In conclusion, umbelliferone showed neuroprotective effects on CUMS-induced model of depression, which was associated with the inhibition of neuronal apoptosis modulated by ROCK/Akt pathway.

  20. Structure and Function of Vertebrate Metallothioneins

    DEFF Research Database (Denmark)

    Penkowa, Milena; Vasak, Milan; Hidalgo, Juan

    2009-01-01

    In 1957, Margoshes and Vallee reported on the isolation of a protein from horse kidney, which showed a high affinity for cadmium, and soon thereafter the protein was named metallothionein (MT) by the leading scientists Ka¨ gi and Vallee. Fifty years of intense research has dissected out many of t...

  1. Metallothioneins are multipurpose neuroprotectants during brain pathology

    DEFF Research Database (Denmark)

    Penkowa, Milena

    2006-01-01

    Metallothioneins (MTs) constitute a family of cysteine-rich metalloproteins involved in cytoprotection during pathology. In mammals there are four isoforms (MT-I - IV), of which MT-I and -II (MT-I + II) are the best characterized MT proteins in the brain. Accumulating studies have demonstrated MT...

  2. Metallothionein treatment reduces proinflammatory cytokines IL-6 and TNF-alpha and apoptotic cell death during experimental autoimmune encephalomyelitis (EAE).

    Science.gov (United States)

    Penkowa, M; Hidalgo, J

    2001-07-01

    Experimental autoimmune encephalomyelitis (EAE) is an animal model for the human autoimmune disease multiple sclerosis (MS). Proinflammatory cytokines such as interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha) are considered important for induction and pathogenesis of EAE/MS disease, which is characterized by significant inflammation and neuroglial damage. We have recently shown that the exogenous administration of the antioxidant protein zinc-metallothionein-II (Zn-MT-II) significantly decreased the clinical symptoms, mortality, and leukocyte infiltration of the CNS during EAE. However, it is not known how EAE progression is regulated nor how cytokine production and cell death can be reduced. We herewith demonstrate that treatment with Zn-MT-II significantly decreased the CNS expression of IL-6 and TNF-alpha during EAE. Zn-MT-II treatment could also significantly reduce apoptotic cell death of neurons and oligodendrocytes during EAE, as judged by using TUNEL and immunoreactivity for cytochrome c and caspases 1 and 3. In contrast, the number of apoptotic lymphocytes and macrophages was less affected by Zn-MT-II treatment. The Zn-MT-II-induced decrease in proinflammatory cytokines and apoptosis during EAE could contribute to the reported diminution of clinical symptoms and mortality in EAE-immunized rats receiving Zn-MT-II treatment. Our results demonstrate that MT-II reduces the CNS expression of proinflammatory cytokines and the number of apoptotic neurons during EAE in vivo and that MT-II might be a potentially useful factor for treatment of EAE/MS.

  3. The role of metallothionein in oncogenesis and cancer treatment

    Directory of Open Access Journals (Sweden)

    Anna Bizoń

    2017-02-01

    Full Text Available Metallothionein is cysteine-rich low molecular mass protein. The involvement of MT in many physiological and pathophysiological processes such as apoptosis, proliferation, angiogenesis, and the detoxification of heavy metals suggested participation of this protein in carcinogenesis and tumor therapy.Depending on the type of tissue and classification of carcinoma various it was observed relation between MT expression and tumor type, stage, grade, poor prognosis and body resistance to radiotherapy and chemotherapy. MT in tumor cell plays important role in defense mechanism against the effect of radiation by inhibiting the processes that lead to the apoptosis. A number of studies have shown an increased expression of MT in various human tumors of larynx, pancreas, kidney, uterus and breast, whereas lower MT expression was detected in liver tumors. Variable MT expression was detected in case of thyroid, prostate, lung, stomach and central nervous system tumors.Also MT plays crucial role in the cytostatics treatment. MT can bind cis-platinum compounds and removes them from the cells, which may lead to multidrug resistance. However, the same functions of MT protect against the negative effects of chemotherapeutic treatment. It is especially important in case of heart cells.Analysis of MT expression in tumor cells may be useful in choosing method of treatment. It is difficult to determine whether increased expression of MT is only a inducing factor of the development of the carcinogenesis, its malignances and multidrug resistance, or it is a factor inhibiting the induction and development of cancer.

  4. The regulation of p53 up-regulated modulator of apoptosis by JNK/c-Jun pathway in β-amyloid-induced neuron death.

    Science.gov (United States)

    Akhter, Rumana; Sanphui, Priyankar; Das, Hrishita; Saha, Pampa; Biswas, Subhas Chandra

    2015-09-01

    Neuronal loss in selective areas of brain underlies the pathology of Alzheimer's disease (AD). Recent evidences place oligomeric β-amyloid (Aβ) central to the disease. However, mechanism of neuron death in response to Aβ remains elusive. Activation of the c-Jun N-terminal kinase (JNK) pathway and induction of the AP-1 transcription factor c-Jun are reported in AD. However, targets of JNK/c-Jun in Aβ-induced neuron death are mostly unknown. Our study shows that pro-apoptotic proteins, Bim (Bcl-2 interacting mediator of cell death) and Puma (p53 up-regulated modulator of apoptosis) are targets of c-Jun in Aβ-treated neurons. We demonstrate that the JNK/c-Jun pathway is activated, in cultures of cortical neurons following treatment with oligomeric Aβ and in AD transgenic mice, and that inhibition of this pathway by selective inhibitor blocks induction of Puma by Aβ. We also find that both JNK and p53 pathways co-operatively regulate Puma expression in Aβ-treated neurons. Moreover, we identified a novel AP1-binding site on rat puma gene which is necessary for direct binding of c-Jun with Puma promoter. Finally, we find that knocking down of c-Jun by siRNA provides significant protection from Aβ toxicity and that induction of Bim and Puma by Aβ in neurons requires c-Jun. Taken together, our results suggest that both Bim and Puma are target of c-Jun and elucidate the intricate regulation of Puma expression by JNK/c-Jun and p53 pathways in neurons upon Aβ toxicity. JNK/c-Jun pathway is shown to be activated in neurons of the Alzheimer's disease (AD) brain and plays a vital role in neuron death in AD models. However, downstream targets of c-Jun in this disease have not been thoroughly elucidated. Our study shows that two important pro-apoptotic proteins, Bim (Bcl-2 interacting mediator of cell death) and Puma (p53 up-regulated modulator of apoptosis) are targets of c-Jun in Aβ-treated neurons. We demonstrate that the JNK/c-jun pathway is activated, in cultures

  5. Intraneuronal signaling pathways of metallothionein

    DEFF Research Database (Denmark)

    Asmussen, Johanne Wirenfeldt; Von Sperling, Marie Louise; Penkowa, Milena

    2009-01-01

    in animals receiving MT treatment and in mice with endogenous MT overexpression or null mutation during various experimental models of neuropathology, and also in patients with Alzheimer's disease and amyotrophic lateral sclerosis. Exogenously applied MT increases neurite outgrowth and neuronal survival...

  6. Glutamate-induced apoptosis in primary cortical neurons is inhibited by equine estrogens via down-regulation of caspase-3 and prevention of mitochondrial cytochrome c release

    Directory of Open Access Journals (Sweden)

    Zhang YueMei

    2005-02-01

    Full Text Available Abstract Background Apoptosis plays a key role in cell death observed in neurodegenerative diseases marked by a progressive loss of neurons as seen in Alzheimer's disease. Although the exact cause of apoptosis is not known, a number of factors such as free radicals, insufficient levels of nerve growth factors and excessive levels of glutamate have been implicated. We and others, have previously reported that in a stable HT22 neuronal cell line, glutamate induces apoptosis as indicated by DNA fragmentation and up- and down-regulation of Bax (pro-apoptotic, and Bcl-2 (anti-apoptotic genes respectively. Furthermore, these changes were reversed/inhibited by estrogens. Several lines of evidence also indicate that a family of cysteine proteases (caspases appear to play a critical role in neuronal apoptosis. The purpose of the present study is to determine in primary cultures of cortical cells, if glutamate-induced neuronal apoptosis and its inhibition by estrogens involve changes in caspase-3 protease and whether this process is mediated by Fas receptor and/or mitochondrial signal transduction pathways involving release of cytochrome c. Results In primary cultures of rat cortical cells, glutamate induced apoptosis that was associated with enhanced DNA fragmentation, morphological changes, and up-regulation of pro-caspase-3. Exposure of cortical cells to glutamate resulted in a time-dependent cell death and an increase in caspase-3 protein levels. Although the increase in caspase-3 levels was evident after 3 h, cell death was only significantly increased after 6 h. Treatment of cells for 6 h with 1 to 20 mM glutamate resulted in a 35 to 45% cell death that was associated with a 45 to 65% increase in the expression of caspase-3 protein. Pretreatment with caspase-3-protease inhibitor z-DEVD or pan-caspase inhibitor z-VAD significantly decreased glutamate-induced cell death of cortical cells. Exposure of cells to glutamate for 6 h in the presence or

  7. Ecdysterone protects gerbil brain from temporal global cerebral ischemia/reperfusion injury via preventing neuron apoptosis and deactivating astrocytes and microglia cells.

    Science.gov (United States)

    Wang, Wei; Wang, Tao; Feng, Wan-Yu; Wang, Zhan-You; Cheng, Mao-Sheng; Wang, Yun-Jie

    2014-01-01

    Ecdysterone (EDS), a common derivative of ecdysteroid, has shown its effects on alleviating cognitive impairment and improving the cognition and memory. However, the mechanisms remain unknown. Using temporal global forebrain ischemia and reperfusion-induced brain injury as a model system, we investigated the roles of EDS in improving cognitive impairment in gerbil. Our results demonstrated that intraperitoneal injection of EDS obviously increased the number of surviving neuron cells by Nissl and neuronal nuclei (NeuN) staining. Indeed, the protecting effects of EDS are because of its ability to prevent the apoptosis of neuron cells as evidenced by TUNEL staining and caspase-3 deactivation in the brain of temporal global forebrain ischemia/reperfusion-treated gerbil. Moreover, EDS administration suppressed the ischemia stimulated activity of astrocytes and microglia cells by inhibiting the production of tumor necrosis alpha (TNF-α) in the brain of gerbil. More importantly, these actions of neurons and astrocytes/microglia cells in response to EDS treatment played pivotal roles in ameliorating the cognitive impairment in the ischemia/reperfusion-injured gerbil. In view of these observations, we not only decipher the mechanisms of EDS in reducing the syndrome of ischemia, but also provide novel perspectives to combat ischemic stroke.

  8. Neuronal apoptosis and synaptic density in the dentate gyrus of ischemic rats' response to chronic mild stress and the effects of Notch signaling.

    Directory of Open Access Journals (Sweden)

    Shaohua Wang

    Full Text Available Our previous research highlighted an inconsistency with Notch1 signaling-related compensatory neurogenesis after chronic mild stress (CMS in rodents suffering from cerebral ischemia, which continue to display post-stroke depressive symptoms. Here, we hypothesize that CMS aggrandized ischemia-related apoptosis injury and worsened synaptic integrity via gamma secretase-meditated Notch1 signaling. Adult rats were exposed to a CMS paradigm after left middle cerebral artery occlusion (MCAO. Open-field and sucrose consumption testing were employed to assess depression-like behavior. Gene expression of pro-apoptotic Bax, anti-apoptotic Bcl-2, and synaptic density-related synaptophysin were measured by western blotting and real-time PCR on Day 28 after MCAO surgery. CMS induced depressive behaviors in ischemic rats, which was accompanied by an elevation in Bax/bcl-2 ratio, TUNEL staining in neurons and reduced synaptophysin expression in the dentate gyrus. These collective effects were reversed by the gamma-secretase inhibitor DAPT (N-[N-(3,5-difluorophenacetyl-L-alanyl]-S-phenyl-glycine t-butyl ester. We found that post-stroke stressors made neurons in the dentate gyrus vulnerable to apoptosis, which supports a putative role for Notch signaling in neural integrity, potentially in newborn cells' synaptic deficit with regard to preexisting cells. These findings suggest that post-stroke depression therapeutically benefits from blocking gamma secretase mediated Notch signaling, and whether this signaling pathway could be a therapeutic target needs to be further investigated.

  9. Caspase-3 dependent nitrergic neuronal apoptosis following cavernous nerve injury is mediated via RhoA and ROCK activation in major pelvic ganglion.

    Science.gov (United States)

    Hannan, Johanna L; Matsui, Hotaka; Sopko, Nikolai A; Liu, Xiaopu; Weyne, Emmanuel; Albersen, Maarten; Watson, Joseph W; Hoke, Ahmet; Burnett, Arthur L; Bivalacqua, Trinity J

    2016-07-08

    Axonal injury due to prostatectomy leads to Wallerian degeneration of the cavernous nerve (CN) and erectile dysfunction (ED). Return of potency is dependent on axonal regeneration and reinnervation of the penis. Following CN injury (CNI), RhoA and Rho-associated protein kinase (ROCK) increase in penile endothelial and smooth muscle cells. Previous studies indicate that nerve regeneration is hampered by activation of RhoA/ROCK pathway. We evaluated the role of RhoA/ROCK pathway in CN regulation following CNI using a validated rat model. CNI upregulated gene and protein expression of RhoA/ROCK and caspase-3 mediated apoptosis in the major pelvic ganglion (MPG). ROCK inhibitor (ROCK-I) prevented upregulation of RhoA/ROCK pathway as well as activation of caspase-3 in the MPG. Following CNI, there was decrease in the dimer to monomer ratio of neuronal nitric oxide synthase (nNOS) protein and lowered NOS activity in the MPG, which were prevented by ROCK-I. CNI lowered intracavernous pressure and impaired non-adrenergic non-cholinergic-mediated relaxation in the penis, consistent with ED. ROCK-I maintained the intracavernous pressure and non-adrenergic non-cholinergic-mediated relaxation in the penis following CNI. These results suggest that activation of RhoA/ROCK pathway mediates caspase-3 dependent apoptosis of nitrergic neurons in the MPG following CNI and that ROCK-I can prevent post-prostatectomy ED.

  10. Inhibition of BCL-2 leads to increased apoptosis and delayed neuronal differentiation in human ReNcell VM cells in vitro.

    Science.gov (United States)

    Fröhlich, Michael; Jaeger, Alexandra; Weiss, Dieter G; Kriehuber, Ralf

    2016-02-01

    BCL-2 is a multifunctional protein involved in the regulation of apoptosis, cell cycle progression and neural developmental processes. Its function in the latter process is not well understood and needs further elucidation. Therefore, we characterized the protein expression kinetics of BCL-2 and associated regulatory proteins of the intrinsic apoptosis pathway during the process of neuronal differentiation in ReNcell VM cells with and without functional inhibition of BCL-2 by its competitive ligand HA14-1. Inhibition of BCL-2 caused a diminished BCL-2 expression and higher levels of cleaved BAX, activated Caspase-3 and cleaved PARP, all pro-apoptotic markers, when compared with untreated differentiating cells. In parallel, flow cytometric analysis of HA14-1-treated cells revealed a delayed differentiation into HuC/D+ neuronal cells when compared to untreated differentiating cells. In conclusion, BCL-2 possess a protective function in fully differentiated ReNcell VM cells. We propose that the pro-survival signaling of BCL-2 is closely connected with its stimulatory effects on neurogenesis of human neural progenitor cells.

  11. miR-200a-3p promotes β-Amyloid-induced neuronal apoptosis through down-regulation of SIRT1 in Alzheimer’s disease

    Indian Academy of Sciences (India)

    QI-SHUN ZHANG; WEI LIU; GUANG-XIU LU

    2017-09-01

    The aberrantly expressed microRNAs (miRNAs) including miR-200a-3p have been reported in the brains of Alzheimer’sdisease (AD) patients in recent researches. Nevertheless, the role of miR-200a-3p in AD has not been characterized. Thepurpose of this study was to examine whether miR-200a-3p regulated b-Ameyloid (Ab)-induced neuronal apoptosis bytargeting SIRT1, a known anti-apoptotic protein. An increased level of miR-200a-3p and a decreased level of SIRT1 in thehippocampus of APPswe/PSDE9 mice (a model for AD) were observed. To construct an in vitro cell model of AD, PC12cells were cultured in presence of Ab25-35. The results of flow cytometry analysis showed that the apoptosis rate andcleaved-caspase-3 expression in PC12 cells exposed to Ab25-35 were remarkably increased, but the apoptosis rate andcleaved-caspase-3 activity were decreased when cells were transfected with anti-miR-200a-3p. On the other hand, MTTassay showed that the cell survival rate was increased in the Ab25-35 ? anti-miR-200a-3p group compared with the Ab25-35 ? anti-miR-NC group. Dual-luciferase reporter gene assay validated the predicted miR-200a-3p binding sites in the 30-UTR of SIRT1 mRNA. In addition, downregulation of SIRT1 promoted Ab25-35-induced neuronal apoptosis and cleavedcaspase-3 level in PC12 cells, whereas anti-miR-200a-3p reversed these effects. Knockdown of SIRT1 decreased theinhibitory effect of Ab25-35 on cell viability, while anti-miR-200a-3p attenuated this effect. Overall, the results suggest thatsuppression of miR-200a-3p attenuates Ab25-35-induced apoptosis in PC12 cells by targeting SIRT1. Thus, miR-200a-3pmay be a potential therapeutic target for treatment of AD.

  12. Tetrahymena metallothioneins fall into two discrete subfamilies.

    Directory of Open Access Journals (Sweden)

    Silvia Díaz

    Full Text Available BACKGROUND: Metallothioneins are ubiquitous small, cysteine-rich, multifunctional proteins which can bind heavy metals. METHODOLOGY/PRINCIPAL FINDINGS: We report the results of phylogenetic and gene expression analyses that include two new Tetrahymena thermophila metallothionein genes (MTT3 and MTT5. Sequence alignments of all known Tetrahymena metallothioneins have allowed us to rationalize the structure of these proteins. We now formally subdivide the known metallothioneins from the ciliate genus Tetrahymena into two well defined subfamilies, 7a and 7b, based on phylogenetic analysis, on the pattern of clustering of Cys residues, and on the pattern of inducibility by the heavy metals Cd and Cu. Sequence alignment also reveals a remarkably regular, conserved and hierarchical modular structure of all five subfamily 7a MTs, which include MTT3 and MTT5. The former has three modules, while the latter has only two. Induction levels of the three T. thermophila genes were determined using quantitative real time RT-PCR. Various stressors (including heavy metals brought about dramatically different fold-inductions for each gene; MTT5 showed the highest fold-induction. Conserved DNA motifs with potential regulatory significance were identified, in an unbiased way, upstream of the start codons of subfamily 7a MTs. EST evidence for alternative splicing in the 3' UTR of the MTT5 mRNA with potential regulatory activity is reported. CONCLUSION/SIGNIFICANCE: The small number and remarkably regular structure of Tetrahymena MTs, coupled with the experimental tractability of this model organism for studies of in vivo function, make it an attractive system for the experimental dissection of the roles, structure/function relationships, regulation of gene expression, and adaptive evolution of these proteins, as well as for the development of biotechnological applications for the environmental monitoring of toxic substances.

  13. Epilepsy-induced neuronal injury: apoptosis and necrosis%癫痫发作后神经元的损伤:凋亡与坏死

    Institute of Scientific and Technical Information of China (English)

    孙建英; 刘学伍; 迟兆富

    2005-01-01

    OBJECTIVE: Epileptic attack can cause neuronal damage and increase the risk of potential seizure. Analysis of the possible mechanism of neuronal damage following epileptic seizure may provide evidences for implementing preventive measures against brain damage due to epileptic seizures.DATA SOURCES: A computer-based search of the related publications in PubMed database between June 1995 and June 2004 with different combinations of the key words of "epilepsy", "neuron damage", "necrosis"and "apoptosis", limiting the results to the language of English.STUDY SELECTION: The retrieved articles were examined at first to select reports of experimental study on human and animals related to epilepsy and the subsequent neuronal damages, and their full-text publications were obtained with the other unrelated articles excluded.DATA EXTRACTION: Eighteen articles documenting randomized controlled experiment immediately related to neuronal damage after epilepsy seizure, 4 reporting non-randomized controlled experiments related to central neuronal excitatory toxic damage, and 3 concerning neuronal damage were collected for this review.DATA SYNTHESIS: In the 14 randomized controlled experiments, chemical or electric methods were used to induce epilepsy in the animal models in which the ultrastructural changes of the neurons and cell organelles were observed and the expression of apoptosis-related factors determined.In the 4 non-randomized controlled experiments, central neuronal ischemic and hypoxic models were adopted for observing the expression of various apoptotic factors in the neurons due to different damages with the assistance of electron microscope, to provide direct evidences for the mechanism of central neuronal excitatory toxic damage. The other three related literatures introduced the pathways of neuronal damages and the expression of the related factors.CONCLUSION: Neuronal death after epileptic seizure is correlated with the severity of the damage and mitochondrial

  14. TPEN, a Specific Zn(2+) Chelator, Inhibits Sodium Dithionite and Glucose Deprivation (SDGD)-Induced Neuronal Death by Modulating Apoptosis, Glutamate Signaling, and Voltage-Gated K(+) and Na(+) Channels.

    Science.gov (United States)

    Zhang, Feng; Ma, Xue-Ling; Wang, Yu-Xiang; He, Cong-Cong; Tian, Kun; Wang, Hong-Gang; An, Di; Heng, Bin; Xie, Lai-Hua; Liu, Yan-Qiang

    2017-03-01

    Hypoxia-ischemia-induced neuronal death is an important pathophysiological process that accompanies ischemic stroke and represents a major challenge in preventing ischemic stroke. To elucidate factors related to and a potential preventative mechanism of hypoxia-ischemia-induced neuronal death, primary neurons were exposed to sodium dithionite and glucose deprivation (SDGD) to mimic hypoxic-ischemic conditions. The effects of N,N,N',N'-tetrakis (2-pyridylmethyl) ethylenediamine (TPEN), a specific Zn(2+)-chelating agent, on SDGD-induced neuronal death, glutamate signaling (including the free glutamate concentration and expression of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptor (GluR2) and N-methyl-D-aspartate (NMDA) receptor subunits (NR2B), and voltage-dependent K(+) and Na(+) channel currents were also investigated. Our results demonstrated that TPEN significantly suppressed increases in cell death, apoptosis, neuronal glutamate release into the culture medium, NR2B protein expression, and I K as well as decreased GluR2 protein expression and Na(+) channel activity in primary cultured neurons exposed to SDGD. These results suggest that TPEN could inhibit SDGD-induced neuronal death by modulating apoptosis, glutamate signaling (via ligand-gated channels such as AMPA and NMDA receptors), and voltage-gated K(+) and Na(+) channels in neurons. Hence, Zn(2+) chelation might be a promising approach for counteracting the neuronal loss caused by transient global ischemia. Moreover, TPEN could represent a potential cell-targeted therapy.

  15. The Neuron-Specific Rai (ShcC) Adaptor Protein Inhibits Apoptosis by Coupling Ret to the Phosphatidylinositol 3-Kinase/Akt Signaling Pathway

    Science.gov (United States)

    Pelicci, Giuliana; Troglio, Flavia; Bodini, Alessandra; Melillo, Rosa Marina; Pettirossi, Valentina; Coda, Laura; De Giuseppe, Antonio; Santoro, Massimo; Pelicci, Pier Giuseppe

    2002-01-01

    Rai is a recently identified member of the family of Shc-like proteins, which are cytoplasmic signal transducers characterized by the unique PTB-CH1-SH2 modular organization. Rai expression is restricted to neuronal cells and regulates in vivo the number of postmitotic sympathetic neurons. We report here that Rai is not a common substrate of receptor tyrosine kinases under physiological conditions and that among the analyzed receptors (Ret, epidermal growth factor receptor, and TrkA) it is activated specifically by Ret. Overexpression of Rai in neuronal cell lines promoted survival by reducing apoptosis both under conditions of limited availability of the Ret ligand glial cell line-derived neurotrophic factor (GDNF) and in the absence of Ret activation. Overexpressed Rai resulted in the potentiation of the Ret-dependent activation of phosphatidylinositol 3-kinase (PI3K) and Akt. Notably, increased Akt phosphorylation and PI3K activity were also found under basal conditions, e.g., in serum-starved neuronal cells. Phosphorylated and hypophosphorylated Rai proteins form a constitutive complex with the p85 subunit of PI3K: upon Ret triggering, the Rai-PI3K complex is recruited to the tyrosine-phosphorylated Ret receptor through the binding of the Rai PTB domain to tyrosine 1062 of Ret. In neurons treated with low concentrations of GDNF, the prosurvival effect of Rai depends on Rai phosphorylation and Ret activation. In the absence of Ret activation, the prosurvival effect of Rai is, instead, phosphorylation independent. Finally, we showed that overexpression of Rai, at variance with Shc, had no effects on the early peak of mitogen-activated protein kinase (MAPK) activation, whereas it increased its activation at later time points. Phosphorylated Rai, however, was not found in complexes with Grb2. We propose that Rai potentiates the MAPK and PI3K signaling pathways and regulates Ret-dependent and -independent survival signals. PMID:12242309

  16. iNOS participates in apoptosis of spinal cord neurons via p-BAD dephosphorylation following ischemia/reperfusion (I/R) injury in rat spinal cord.

    Science.gov (United States)

    Li, Yiming; Gu, Jun; Liu, Yuwen; Long, Hao; Wang, Guannan; Yin, Guoyong; Fan, Jin

    2013-06-17

    The pro-apoptotic effect of nitric oxide (NO) has been reported both in vivo and in vitro. Previous studies have revealed that NO, especially which produced by inducible nitric oxide synthase (iNOS), has an important effect on apoptosis of neurons in spinal cord ischemia/reperfusion (I/R) injury. To investigate the role of iNOS in this process, a randomized, controlled study was designed using a classical rat model of ischemic spinal cord injury. Fifty-four male Sprague-Dawley rats were randomly divided into three different groups: a sham-operated group (n=6), a vehicle group (I/R, n=24), and an iNOS inhibitor (aminoguanidine: AG) group (I/R+AG, n=24). Rats were sacrificed 6, 12, 24 and 72 h after reperfusion. We examined neurological motor function evaluated by 'Tarlov's score', assessed alterations in the morphology of spinal cord neurons by transmission electron microscopy (TEM), analyzed expression of iNOS at the levels of mRNA and protein, evaluated local concentrations and cellular locations of other key regulatory proteins, and investigated protein-protein interactions. In the vehicle group, iNOS expression, dephosphorylation of p-BAD (Ser 136), disassociation of BAD from p-BAD/14-3-3 dimers, and release of cytochrome c were all increased compared with the sham group. But in the AG group, all the performances above were decreased compared with the vehicle group. Similarly, rats in the sham group got a maximum score of 5 by Tarlov's motor scores evaluation. While the scores were higher in the AG group compared to the vehicle group because iNOS was inhibited. These results indicate that the activity of iNOS plays a critical role in the apoptosis of spinal cord neurons by influencing the dephosphorylation of p-BAD (Ser 136) and the interaction between BAD and 14-3-3.

  17. Activation of GSK-3β and Caspase-3 Occurs in Nigral Dopamine Neurons during the Development of Apoptosis Activated by a Striatal Injection of 6-Hydroxydopamine

    Science.gov (United States)

    Hernandez-Baltazar, Daniel; Mendoza-Garrido, Maria E.; Martinez-Fong, Daniel

    2013-01-01

    The 6-Hydroxydopamine (6-OHDA) rat model of Parkinson's disease is essential for a better understanding of the pathological processes underlying the human disease and for the evaluation of promising therapeutic interventions. This work evaluated whether a single striatal injection of 6-OHDA causes progressive apoptosis of dopamine (DA) neurons and activation of glycogen synthase kinase 3β (GSK-3β) and caspase-3 in the substantia nigra compacta (SNc). The loss of DA neurons was shown by three neuron markers; tyrosine hydroxylase (TH), NeuN, and β-III tubulin. Apoptosis activation was determined using Apostain and immunostaining against cleaved caspase-3 and GSK-3β pY216. We also explored the possibility that cleaved caspase-3 is produced by microglia and astrocytes. Our results showed that the 6-OHDA caused loss of nigral TH(+) cells, progressing mainly in rostrocaudal and lateromedial directions. In the neostriatum, a severe loss of TH(+) terminals occurred from day 3 after lesion. The disappearance of TH(+) cells was associated with a decrease in NeuN and β-III tubulin immunoreactivity and an increase in Apostain, cleaved caspase-3, and GSK-3β pY216 in the SNc. Apostain immunoreactivity was observed from days 3 to 21 postlesion. Increased levels of caspase-3 immunoreactivity in TH(+) cells were detected from days 1 to 15, and the levels then decreased to day 30 postlesion. The cleaved caspase-3 also collocated with microglia and astrocytes indicating its participation in glial activation. Our results suggest that caspase-3 and GSK-3β pY216 activation might participate in the DA cell death and that the active caspase-3 might also participate in the neuroinflammation caused by the striatal 6-OHDA injection. PMID:23940672

  18. Environmental Neurotoxic Pesticide Increases Histone Acetylation to Promote Apoptosis in Dopaminergic Neuronal Cells: Relevance to Epigenetic Mechanisms of Neurodegeneration

    OpenAIRE

    Song, C.; Kanthasamy, A.; Anantharam, V.; Sun, F.; Kanthasamy, A.G.

    2010-01-01

    Pesticide exposure has been implicated in the etiopathogenesis of Parkinson's disease (PD); in particular, the organochlorine insecticide dieldrin is believed to be associated with PD. Emerging evidence indicates that histone modifications play a critical role in cell death. In this study, we examined the effects of dieldrin treatment on histone acetylation and its role in dieldrin-induced apoptotic cell death in dopaminergic neuronal cells. In mesencephalic dopaminergic neuronal cells, dield...

  19. Efficient expression of the yeast metallothionein gene in Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Berka, T.; Shatzman, A.; Zimmerman, J.; Strickler, J.; Rosenberg, M.

    1988-01-01

    The yeast metallothionein gene CUP1 was cloned into a bacterial expression system to achieve efficient, controlled expression of the stable, unprocessed protein product. The Escherichia coli-synthesized yeast metallothionein bound copper, cadmium, zinc, indicating that the protein was functional. Furthermore, E. coli cells expressing CUP1 acquired a new, inducible ability to selectively sequester heavy metal ions from the growth medium.

  20. A Review of Metallothionein Isoforms and their Role in Pathophysiology

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    Senthil kumar M

    2011-05-01

    Full Text Available Abstract The Metallothionein (MT is a protein which has several interesting biological effects and has been demonstrated increase focus on the role of MT in various biological systems in the past three decades. The studies on the role of MT were limited with few areas like apoptosis and antioxidants in selected organs even fifty years after its discovery. Now acknowledge the exploration of various isoforms of MT such as MT-I, MT-II, MT-III and MT-IV and other isoforms in various biological systems. Strong evidence exists that MT modulates complex diseases and the immune system in the body but the primary function of MT still remains unknown. This review's main objective is to explore the capability to specifically manipulate MT levels in cells and in animals to provide answers regarding how MT could impact those complex disease scenarios. The experimental result mentioned in this review related among MT, zinc, cadmium, diabetic, heart disease, bone retardation, neuro toxicity, kidney dysfunction, cancer, and brain suggest novel method for exploration and contribute significantly to the growing scientist to research further in this field.

  1. Effect of Ganoderma lucidum spore on expression of insulin-like growth factor-1, nuclear factor-kappa B, and neuronal apoptosis in the epileptic rat brain

    Institute of Scientific and Technical Information of China (English)

    Shuang Zhao; Shuqiu Wang; Shengchang Zhang; Fafang Li

    2008-01-01

    BACKGROUND: It has been reported that Ganoderma lucidum spore powder, a very well known Chinese traditional medicine, can affect immunoregulation, free radical scavenging, and anti-hypoxia responses. OBJECTIVE: To investigate the effect of Ganoderma lucidum spore powder on expression of insulin-like growth factor-1 (IGF-1), nuclear factor-κB (NF-κB) and neuronal apoptosis in rats with pentylenetetrazol (PTZ)-induced epilepsy.DESIGN, TIME AND SETTING: A cellular and molecular biology experiment with randomized controlled study design was performed at the Central Laboratory of Basic Medical College of Jiamusi University from June to August 2005.MATERIALS: Thirty healthy, adult, male, Wistar rats were selected and randomly divided into 3 groups (10rats per group): control, epilepsy model, and Ganoderma lucidum spore powder. A sub-eclampsia PTZ dose (35 mg/kg) was intraperitoneally injected to induce epilepsy in the latter two groups. Wild Ganoderma lucidum spore powder (30 g/L) was provided by the wild Ganoderma lucidum plant nursery at Jiamusi,China. lmmunohistochemical detection and terminal deoxynucleotidyl transferase-mediate dUTP nick end-labeling (TUNEL) kits were purchased from Wuhan Boster Biological Technology Co., Ltd., China.METHODS: Ganoderma lucidum spore powder was intragastrically administered at a dose of 10.0 mL/kg,once a day for 28 days. In the epilepsy and control groups, an equivalent volume of normal saline was intragastrically administered.MAIN OUTCOME MEASURES: lmmunoreactivity for IGF-1 and NF-κB/P65 were detected by immunohistochemical staining. Neuronal apoptosis was detected using TUNEL methods.RESULTS: The hippocampus and cerebral cortex of rats with PTZ-induced epilepsy exhibited a higher number of apoptotic cells at high magnification (x400), compared with the control group. Expression of IGF-1and NF-κB were higher in the epilepsy group, compared with the control group (P < 0.01). In Ganoderma lucidum spore-treated rats, fewer

  2. Effect of melamine toxicity on Tetrahymena thermophila proliferation and metallothionein expression.

    Science.gov (United States)

    Li, Wei; Li, Hua; Zhang, Jie; Tian, Xuewen

    2015-06-01

    Melamine is a raw material in the chemical industry. Because of its high nitrogen content, melamine has been utilized by unscrupulous businessmen as a food additive to enhance the indices of protein content in food and feed testing. Tetrahymena has long been used as an excellent model organism in toxicological studies. The purpose of the present study was to determine the effect of melamine on Tetrahymena. In the present study, the effects of melamine on the proliferation and mating rate of Tetrahymena were examined by microscopic counting of the cell numbers. The comet assay and DAPI nuclear staining were performed to analyze the changes in the Tetrahymena genome. Flow cytometric analysis was conducted to detect apoptosis. Furthermore, RT-PCR was performed to determine the changes in the expression of the metallothionein gene in Tetrahymena that underwent stress treatment with varying concentrations of melamine. The results indicated that melamine affected the proliferation and sexual reproduction of Tetrahymena. High melamine concentrations damaged the Tetrahymena genome to a certain extent and induced apoptosis in the organism. Expression of the metallothionein gene was upregulated in Tetrahymena exposed to melamine stress to ameliorate melamine-induced damage. These results indicated that melamine displayed significant toxicity to Tetrahymena cells.

  3. Oxygen glucose deprivation post-conditioning protects cortical neurons against oxygen-glucose deprivation injury: role of HSP70 and inhibition of apoptosis.

    Science.gov (United States)

    Zhao, Jian-hua; Meng, Xian-li; Zhang, Jian; Li, Yong-li; Li, Yue-juan; Fan, Zhe-ming

    2014-02-01

    In the present study, we examined the effect of oxygen glucose deprivation (OGD) post-conditioning (PostC) on neural cell apoptosis in OGD-PostC model and the protective effect on primary cortical neurons against OGD injury in vitro. Four-h OGD was induced by OGD by using a specialized and humidified chamber. To initiate OGD, culture medium was replaced with de-oxygenated and glucose-free extracellular solution-Locke's medium. After OGD treatment for 4 h, cells were then allowed to recover for 6 h or 20 h. Then lactate dehydrogenase (LDH) release assay, Western blotting and flow cytometry were used to detect cell death, protein levels and apoptotic cells, respectively. For the PostC treatment, three cycles of 15-min OGD, followed by 15 min normal cultivation, were applied immediately after injurious 4-h OGD. Cells were then allowed to recover for 6 h or 20 h, and cell death was assessed by LDH release assay. Apoptotic cells were flow cytometrically evaluated after 4-h OGD, followed by re-oxygenation for 20 h (O4/R20). In addition, Western blotting was used to examine the expression of heat-shock protein 70 (HSP70), Bcl-2 and Bax. The ratio of Bcl-2 expression was (0.44±0.08)% and (0.76±0.10)%, and that of Bax expression was (0.51±0.05)% and (0.39±0.04)%, and that of HSP70 was (0.42±0.031)% and (0.72±0.045)% respectively in OGD group and PostC group. After O4/R6, the rate of neuron death in PostC group and OGD groups was (28.96±3.03)% and (37.02±4.47)%, respectively. Therefore, the PostC treatment could up-regulate the expression of HSP70 and Bcl-2, but down-regulate Bax expression. As compared with OGD group, OGD-induced neuron death and apoptosis were significantly decreased in PostC group (P<0.05). These findings suggest that PostC inhibited OGD-induced neuron death. This neuro-protective effect is likely achieved by anti-apoptotic mechanisms and is associated with over-expression of HSP70.

  4. Metallothionein deficiency aggravates depleted uranium-induced nephrotoxicity

    Energy Technology Data Exchange (ETDEWEB)

    Hao, Yuhui; Huang, Jiawei; Gu, Ying; Liu, Cong; Li, Hong; Liu, Jing; Ren, Jiong; Yang, Zhangyou [State Key Laboratory of Trauma, Burns and Combined Injury, Institute of Combined Injury, Chongqing Engineering Research Center for Nanomedicine, College of Preventive Medicine, Third Military Medical University, No. 30 Gaotanyan Street, Shapingba District, Chongqing 400038 (China); Peng, Shuangqing [Evaluation and Research Center for Toxicology, Institute of Disease Control and Prevention, Academy of Military Medical Science, 20 Dongdajie Street, Fengtai District, Beijing 100071 (China); Wang, Weidong, E-mail: wwdwyl@sina.com [Department of Radiation Oncology, Shanghai Jiao Tong University Affiliated Sixth People' s Hospital, Shanghai 200233 (China); Li, Rong, E-mail: yuhui_hao@126.com [State Key Laboratory of Trauma, Burns and Combined Injury, Institute of Combined Injury, Chongqing Engineering Research Center for Nanomedicine, College of Preventive Medicine, Third Military Medical University, No. 30 Gaotanyan Street, Shapingba District, Chongqing 400038 (China)

    2015-09-15

    Depleted uranium (DU) has been widely used in both civilian and military activities, and the kidney is the main target organ of DU during acute high-dose exposures. In this study, the nephrotoxicity caused by DU in metallothionein-1/2-null mice (MT −/−) and corresponding wild-type (MT +/+) mice was investigated to determine any associations with MT. Each MT −/− or MT +/+ mouse was pretreated with a single dose of DU (10 mg/kg, intraperitoneal injection) or an equivalent volume of saline. After 4 days of DU administration, kidney changes were assessed. After DU exposure, serum creatinine and serum urea nitrogen in MT −/− mice significantly increased than in MT +/+ mice, with more severe kidney pathological damage. Moreover, catalase and superoxide dismutase (SOD) decreased, and generation of reactive oxygen species and malondialdehyde increased in MT −/− mice. The apoptosis rate in MT −/− mice significantly increased, with a significant increase in both Bax and caspase 3 and a decrease in Bcl-2. Furthermore, sodium-glucose cotransporter (SGLT) and sodium-phosphate cotransporter (NaPi-II) were significantly reduced after DU exposure, and the change of SGLT was more evident in MT −/− mice. Finally, exogenous MT was used to evaluate the correlation between kidney changes induced by DU and MT doses in MT −/− mice. The results showed that, the pathological damage and cell apoptosis decreased, and SOD and SGLT levels increased with increasing dose of MT. In conclusion, MT deficiency aggravated DU-induced nephrotoxicity, and the molecular mechanisms appeared to be related to the increased oxidative stress and apoptosis, and decreased SGLT expression. - Highlights: • MT −/− and MT +/+ mice were used to evaluate nephrotoxicity of DU. • Renal damage was more evident in the MT −/− mice after exposure to DU. • Exogenous MT also protects against DU-induced nephrotoxicity. • MT deficiency induced more ROS and apoptosis after exposure to

  5. Insect peptide CopA3-induced protein degradation of p27Kip1 stimulates proliferation and protects neuronal cells from apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Nam, Seung Taek; Kim, Dae Hong; Lee, Min Bum; Nam, Hyo Jung; Kang, Jin Ku; Park, Mi Jung; Lee, Ik Hwan [Department of Life Science, College of Natural Science, Daejin University, Pocheon, Gyeonggido 487-711 (Korea, Republic of); Seok, Heon [Department of Biomedical Science, Jungwon University, Goesan, Chungcheongbukdo 367-700 (Korea, Republic of); Lee, Dong Gun [School of Life Sciences and Biotechnology, College of Natural Sciences, Kyungpook National University, Daegu 702-701 (Korea, Republic of); Hwang, Jae Sam [Department of Agricultural Biology, National Academy of Agricultural Science, RDA, Suwon 441-707 (Korea, Republic of); Kim, Ho, E-mail: hokim@daejin.ac.kr [Department of Life Science, College of Natural Science, Daejin University, Pocheon, Gyeonggido 487-711 (Korea, Republic of)

    2013-07-19

    Highlights: •CopA3 peptide isolated from the Korean dung beetle has antimicrobial activity. •Our study reported that CopA3 has anticancer and immunosuppressive effects. •We here demonstrated that CopA3 has neurotropic and neuroprotective effects. •CopA3 degrades p27Kip1 protein and this mediates effects of CopA3 on neuronal cells. -- Abstract: We recently demonstrated that the antibacterial peptide, CopA3 (a D-type disulfide dimer peptide, LLCIALRKK), inhibits LPS-induced macrophage activation and also has anticancer activity in leukemia cells. Here, we examined whether CopA3 could affect neuronal cell proliferation. We found that CopA3 time-dependently increased cell proliferation by up to 31 ± 2% in human neuroblastoma SH-SY5Y cells, and up to 29 ± 2% in neural stem cells isolated from neonatal mouse brains. In both cell types, CopA3 also significantly inhibited the apoptosis and viability losses caused by 6-hydroxy dopamine (a Parkinson disease-mimicking agent) and okadaic acid (an Alzheimer’s disease-mimicking agent). Immunoblotting revealed that the p27Kip1 protein (a negative regulator of cell cycle progression) was markedly degraded in CopA3-treated SH-SY5Y cells. Conversely, an adenovirus expressing p27Kip1 significantly inhibited the antiapoptotic effects of CopA3 against 6-hydroxy dopamine- and okadaic acid-induced apoptosis, and decreased the neurotropic effects of CopA3. These results collectively suggest that CopA3-mediated protein degradation of p27Kip1 may be the main mechanism through which CopA3 exerts neuroprotective and neurotropic effects.

  6. Neuronal Hyperactivity Disturbs ATP Microgradients, Impairs Microglial Motility, and Reduces Phagocytic Receptor Expression Triggering Apoptosis/Microglial Phagocytosis Uncoupling.

    Directory of Open Access Journals (Sweden)

    Oihane Abiega

    2016-05-01

    Full Text Available Phagocytosis is essential to maintain tissue homeostasis in a large number of inflammatory and autoimmune diseases, but its role in the diseased brain is poorly explored. Recent findings suggest that in the adult hippocampal neurogenic niche, where the excess of newborn cells undergo apoptosis in physiological conditions, phagocytosis is efficiently executed by surveillant, ramified microglia. To test whether microglia are efficient phagocytes in the diseased brain as well, we confronted them with a series of apoptotic challenges and discovered a generalized response. When challenged with excitotoxicity in vitro (via the glutamate agonist NMDA or inflammation in vivo (via systemic administration of bacterial lipopolysaccharides or by omega 3 fatty acid deficient diets, microglia resorted to different strategies to boost their phagocytic efficiency and compensate for the increased number of apoptotic cells, thus maintaining phagocytosis and apoptosis tightly coupled. Unexpectedly, this coupling was chronically lost in a mouse model of mesial temporal lobe epilepsy (MTLE as well as in hippocampal tissue resected from individuals with MTLE, a major neurological disorder characterized by seizures, excitotoxicity, and inflammation. Importantly, the loss of phagocytosis/apoptosis coupling correlated with the expression of microglial proinflammatory, epileptogenic cytokines, suggesting its contribution to the pathophysiology of epilepsy. The phagocytic blockade resulted from reduced microglial surveillance and apoptotic cell recognition receptor expression and was not directly mediated by signaling through microglial glutamate receptors. Instead, it was related to the disruption of local ATP microgradients caused by the hyperactivity of the hippocampal network, at least in the acute phase of epilepsy. Finally, the uncoupling led to an accumulation of apoptotic newborn cells in the neurogenic niche that was due not to decreased survival but to delayed

  7. Satratoxin-G from the black mold Stachybotrys chartarum induces rhinitis and apoptosis of olfactory sensory neurons in the nasal airways of rhesus monkeys.

    Science.gov (United States)

    Carey, Stephan A; Plopper, Charles G; Hyde, Dallas M; Islam, Zahidul; Pestka, James J; Harkema, Jack R

    2012-08-01

    Satratoxin-G (SG) is a trichothecene mycotoxin of Stachybotrys chartarum, the black mold suggested to contribute etiologically to illnesses associated with water-damaged buildings. We have reported that intranasal exposure to SG evokes apoptosis of olfactory sensory neurons (OSNs) and acute inflammation in the nose and brain of laboratory mice. To further assess the potential human risk of nasal airway injury and neurotoxicity, we developed a model of SG exposure in monkeys, whose nasal airways more closely resemble those of humans. Adult, male rhesus macaques received a single intranasal instillation of 20 µg SG (high dose, n = 3), or 5 µg SG daily for four days (repeated low dose, n = 3) in one nasal passage, and saline vehicle in the contralateral nasal passage. Nasal tissues were examined using light and electron microscopy and morphometric analysis. SG induced acute rhinitis, atrophy of the olfactory epithelium (OE), and apoptosis of OSNs in both groups. High-dose and repeated low-dose SG elicited a 13% and 66% reduction in OSN volume density, and a 14-fold and 24-fold increase in apoptotic cells of the OE, respectively. This model provides new insight into the potential risk of nasal airway injury and neurotoxicity caused by exposure to water-damaged buildings.

  8. Injection of Aβ1-40 into hippocampus induced cognitive lesion associated with neuronal apoptosis and multiple gene expressions in the tree shrew.

    Science.gov (United States)

    Lin, Na; Xiong, Liu-Lin; Zhang, Rong-Ping; Zheng, Hong; Wang, Lei; Qian, Zhong-Yi; Zhang, Piao; Chen, Zhi-Wei; Gao, Fa-Bao; Wang, Ting-Hua

    2016-05-01

    Alzheimer's disease (AD) can incur significant health care costs to the patient, their families, and society; furthermore, effective treatments are limited, as the mechanisms of AD are not fully understood. This study utilized twelve adult male tree shrews (TS), which were randomly divided into PBS and amyloidbetapeptide1-40 (Aβ1-40) groups. AD model was established via an intracerebroventricular (icv) injection of Aβ1-40 after being incubated for 4 days at 37 °C. Behavioral, pathophysiological and molecular changes were evaluated by hippocampal-dependent tasks, magnetic resonance imaging (MRI), silver staining, hematoxylin-eosin (HE) staining, TUNEL assay and gene sequencing, respectively. At 4 weeks post-injection, as compared with the PBS group, in Aβ1-40 injected animals: cognitive impairments happened, and the hippocampus had atrophied indicated by MRI findings; meanwhile, HE staining showed the cells of the CA3 and DG were significantly thinner and smaller. The average number of cells in the DG, but not the CA3, was also significantly reduced; furthermore, silver staining revealed neurotic plaques and neurofibrillary tangles (NFTs) in the hippocampi; TUNEL assay showed many cells exhibited apoptosis, which was associated with downregulated BCL-2/BCL-XL-associated death promoter (Bad), inhibitor of apoptosis protein (IAP), Cytochrome c (CytC) and upregulated tumor necrosis factor receptor 1 (TNF-R1); lastly, gene sequencing reported a total of 924 mobilized genes, among which 13 of the downregulated and 19 of the upregulated genes were common to the AD pathway. The present study not only established AD models in TS, but also reported on the underlying mechanism involved in neuronal apoptosis associated with multiple gene expression.

  9. The critical role of lipid rafts nanodomains in the cross-talk between calcium and reactive oxygen and nitrogen species in cerebellar granule neurons apoptosis by extracellular potassium deprivation

    Directory of Open Access Journals (Sweden)

    Carlos Gutierrez-Merino

    2016-01-01

    Full Text Available The apoptosis of cerebellar granule neurons (CGN induced by low-potassium in serum free medium in vitro has become a widely used model for neuronal apoptosis during in vivo brain development. In this review we shall summarize first the basic features of this model for neuronal apoptosis. Next, we shall focus on the L-type calcium channels (LTCC inactivation as the primary pro-apoptotic signal in low K+-induced CGN death. This apoptotic process can be split into two major and sequential cellular signaling phases: one reversible phase that offers a temporal window for therapeutic interventions to prevent neuronal death, and an irreversible later phase. Therefore, we shall comment next the critical role of reactive oxygen species (ROS production and major ROS sources triggering the entry of CGN in the irreversible stages of low K+-induced apoptosis. Then, we shall present the experimental evidences showing clustering of LTCC and ROS producing enzymes in plasma membrane lipid rafts of CGN matured in vitro, which have opened new perspectives for cell signaling in the early and reversible phase of this apoptosis. The role of lipid rafts nanodomains as fast response calcium/nitric oxide transducers of the switch of CGN to low K+ medium will be discussed next. The two major conclusions drawn from this review are: (1 deregulation of the pool of cytochrome b5 reductase associated to plasma membrane-lipid rafts, at least in part due to overexpression of cytochrome b5, can account for the critical superoxide anion overshot which triggers the entry in the irreversible phase of low K+ apoptosis of CGN, and (2 LTCC inactivation is rapidly transduced by lipid rafts nanodomains into a large drop of cytosolic calcium, a switch-off of nitric oxide production and subsequent inactivation of survival signaling pathways dependent on the activity of CaMKII, PKA and Akt/PKB kinases.

  10. Neuroprotective effects of Cyperus rotundus on SIN-1 induced nitric oxide generation and protein nitration: ameliorative effect against apoptosis mediated neuronal cell damage.

    Science.gov (United States)

    Hemanth Kumar, Kandikattu; Tamatam, Anand; Pal, Ajay; Khanum, Farhath

    2013-01-01

    Nitrosylation of tyrosine (3-nitro tyrosine, 3-NT) has been implicated in the pathophysiology of various disorders particularly neurodegenerative conditions and aging. Cyperus rotundus rhizome is being used as a traditional folk medicine to alleviate a variety of disorders including neuronal stress. The herb has recently found applications in food and confectionary industries also. In current study, we have explored the protective effects of C. rotundus rhizome extract (CRE) through its oxido-nitrosative and anti apoptotic mechanism to attenuate peroxynitrite (ONOO(-)) induced neurotoxicity using human neuroblastoma SH-SY5Y cells. Our results elucidate that pre-treatment of neurons with CRE ameliorates the mitochondrial and plasma membrane damage induced by 500 μM SIN-1 to 80% and 24% as evidenced by MTT and LDH assays. CRE inhibited NO generation by downregulating i-NOS expression. SIN-1 induced depletion of antioxidant enzyme status was also replenished by CRE which was confirmed by immunoblot analysis of SOD and CAT. The CRE pre-treatment efficiently potentiated the SIN-1 induced apoptotic biomarkers such as bcl-2 and caspase-3 which orchestrate the proteolytic damage of the cell. The ONOO(-) induced damage to cellular, nuclear and mitochondrial integrity was also restored by CRE. Furthermore, CRE pre-treatment also regulated the 3-NT formation which shows the potential of plant extract against tyrosine nitration. Taken together, our findings suggest that CRE might be developed as a preventive agent against ONOO(-) induced apoptosis.

  11. Ginsenoside Rg3 Improves Recovery from Spinal Cord Injury in Rats via Suppression of Neuronal Apoptosis, Pro-Inflammatory Mediators, and Microglial Activation

    Directory of Open Access Journals (Sweden)

    Dong-Kyu Kim

    2017-01-01

    Full Text Available Spinal cord injury (SCI is one of the most devastating medical conditions; however, currently, there are no effective pharmacological interventions for SCI. Ginsenoside Rg3 (GRg3 is one of the protopanaxadiols that show anti-inflammatory, anti-oxidant, and neuroprotective effects. The present study investigated the neuroprotective effect of GRg3 following SCI in rats. SCI was induced using a static compression model at vertebral thoracic level 10 for 5 min. GRg3 was administrated orally at a dose of 10 or 30 mg/kg/day for 14 days after the SCI. GRg3 (30 mg/kg treatment markedly improved behavioral motor functions, restored lesion size, preserved motor neurons in the spinal tissue, reduced Bax expression and number of TUNEL-positive cells, and suppressed mRNA expression of pro-inflammatory cytokines including tumor necrosis factor-α, interleukin (IL-1β, and IL-6. GRg3 also attenuated the over-production of cyclooxygenase-2 and inducible nitric oxide synthase after SCI. Moreover, GRg3 markedly suppressed microglial activation in the spinal tissue. In conclusion, GRg3 treatment led to a remarkable recovery of motor function and a reduction in spinal tissue damage by suppressing neuronal apoptosis and inflammatory responses after SCI. These results suggest that GRg3 may be a potential therapeutic agent for the treatment of SCI.

  12. Some commonly used brominated flame retardants cause Ca2+-ATPase inhibition, beta-amyloid peptide release and apoptosis in SH-SY5Y neuronal cells.

    Directory of Open Access Journals (Sweden)

    Fawaz Al-Mousa

    Full Text Available Brominated flame retardants (BFRs are chemicals commonly used to reduce the flammability of consumer products and are considered pollutants since they have become widely dispersed throughout the environment and have also been shown to bio-accumulate within animals and man. This study investigated the cytotoxicity of some of the most commonly used groups of BFRs on SH-SY5Y human neuroblastoma cells. The results showed that of the BFRs tested, hexabromocyclododecane (HBCD, tetrabromobisphenol-A (TBBPA and decabromodiphenyl ether (DBPE, all are cytotoxic at low micromolar concentrations (LC(50 being 2.7 ± 0.7 µM, 15 ± 4 µM and 28 ± 7 µM, respectively. They induced cell death, at least in part, by apoptosis through activation of caspases. They also increased intracellular [Ca(2+] levels and reactive-oxygen-species within these neuronal cells. Furthermore, these BFRs also caused rapid depolarization of the mitochondria and cytochrome c release in these neuronal cells. Elevated intracellular [Ca(2+] levels appear to occur through a mechanism involving microsomal Ca(2+-ATPase inhibition and this maybe responsible for Ca(2+-induced mitochondrial dysfunction. In addition, µM levels of these BFRs caused β-amyloid peptide (Aβ-42 processing and release from these cells with a few hours of exposure. These results therefore shows that these pollutants are both neurotoxic and amyloidogenic in-vitro.

  13. Dopamine-induced apoptosis in human neuronal cells: inhibition by nucleic acides antisense to the dopamine transporter

    Energy Technology Data Exchange (ETDEWEB)

    Porat, S.; Gabbay, M.; Tauber, M.; Ratovitski, T.; Blinder, E.; Simantov, R. [Department of Molecular Genetics, Weizmann Institute of Science Rehovot 76100 (Israel)

    1996-09-01

    Human neuroblastoma NMB cells take up [{sup 3}H]dopamine in a selective manner indicating that dopamine transporters are responsible for this uptake. These cells were therefore used as a model to study dopamine neurotoxicity, and to elucidate the role of dopamine transporters in controlling cell death. Treatment with 0.05-0.4 mM dopamine changed cells' morphology within 4 h, accompanied by retraction of processes, shrinkage, apoptosis-like atrophy, accumulation of apoptotic particles, DNA fragmentation and cell death. Cycloheximide inhibited dopamine's effect, suggesting that induction of apoptosis by dopamine was dependent upon protein synthesis. Dopamine cytotoxicity, monitored morphologically by flow cytometric analysis, and by lactate dehydrogenase released, was blocked by cocaine but not by the noradrenaline and serotonin uptake blockers desimipramine and imipramine, respectively. Attempting to inhibit dopamine transport and toxicity in a drug-free and highly selective way, three 18-mer dopamine transporter antisense phosphorothioate oligonucleotides (numbers 1, 2 and 3) and a new plasmid vector expressing the entire rat dopamine transporter complementary DNA in the antisense orientation were prepared and tested. Antisense phosphorothioate oligonucleotide 3 inhibited [{sup 3}H]dopamine uptake in a time- and dose-dependent manner. Likewise, transient transfection of NMB cells with the plasmid expressing dopamine transporter complementary DNA in the antisense orientation partially blocked [{sup 3}H]dopamine uptake. Antisense phosphorothioate oligonucleotide 3 also decreased, dose-dependently, the toxic effect of dopamine and 6-hydroxydopamine. Western blot analysis with newly prepared anti-human dopamine transporter antibodies showed that antisense phosphorothioate oligonucleotide 3 decreased the transporter protein level. These studies contribute to better understand the mechanism of dopamine-induced apoptosis and neurotoxicity. (Copyright (c) 1996

  14. Blockade of NMDA receptor subtype NR2B prevents seizures but not apoptosis of dentate gyrus neurons in bacterial meningitis in infant rats

    Directory of Open Access Journals (Sweden)

    Täuber Martin G

    2003-09-01

    Full Text Available Abstract Background Excitotoxic neuronal injury by action of the glutamate receptors of the N-methyl-d-aspartate (NMDA subtype have been implicated in the pathogenesis of brain damage as a consequence of bacterial meningitis. The most potent and selective blocker of NMDA receptors containing the NR2B subunit is (R,S-alpha-(4-hydroxyphenyl-beta-methyl-4-(phenylmethyl-1-piperid inepropanol (RO 25-6981. Here we evaluated the effect of RO 25-6981 on hippocampal neuronal apoptosis in an infant rat model of meningitis due to Streptococcus pneumoniae. Animals were randomized for treatment with RO 25-6981 at a dosage of either 0.375 mg (15 mg/kg; n = 28 or 3.75 mg (150 mg/kg; n = 15 every 3 h or an equal volume of sterile saline (250 μl; n = 40 starting at 12 h after infection. Eighteen hours after infection, animals were assessed clinically and seizures were observed for a period of 2 h. At 24 h after infection animals were sacrificed and brains were examined for apoptotic injury to the dentate granule cell layer of the hippocampus. Results Treatment with RO 25-6981 had no effect on clinical scores, but the incidence of seizures was reduced (P Conclusions Treatment with a highly selective blocker of NMDA receptors containing the NR2B subunit failed to protect hippocampal neurons from injury in this model of pneumococcal meningitis, while it had some beneficial effect on the incidence of seizures.

  15. Study of metallothionein-quantum dots interactions.

    Science.gov (United States)

    Tmejova, Katerina; Hynek, David; Kopel, Pavel; Krizkova, Sona; Blazkova, Iva; Trnkova, Libuse; Adam, Vojtech; Kizek, Rene

    2014-05-01

    Nanoparticles have gained increasing interest in medical and in vivo applications. Metallothionein (MT) is well known as a maintainer of metal ions balance in intracellular space. This is due to high affinity of this protein to any reactive species including metals and reactive oxygen species. The purpose of this study was to determine the metallothionein-quantum dots interactions that were investigated by spectral and electrochemical techniques. CuS, CdS, PbS, and CdTe quantum dots (QDs) were analysed. The highest intensity was shown for CdTe, than for CdS measured by fluorescence. These results were supported by statistical analysis and considered as significant. Further, these interactions were analysed using gel electrophoresis, where MT aggregates forming after interactions with QDs were detected. Using differential pulse voltammetry Brdicka reaction, QDs and MT were studied. This method allowed us to confirm spectral results and, moreover, to observe the changes in MT structure causing new voltammetric peaks called X and Y, which enhanced with the prolonged time of interaction up to 6 h.

  16. [Metallothionein-I/II in brain injury repair mechanism and its application in forensic medicine].

    Science.gov (United States)

    Li, Dong; Li, Ru-bo; Lin, Ju-li

    2013-10-01

    Metallothionein (MT) is a kind of metal binding protein. As an important member in metallothionein family, MT-I/II regulates metabolism and detoxication of brain metal ion and scavenges free radicals. It is capable of anti-inflammatory response and anti-oxidative stress so as to protect the brain tissue. During the repair process of brain injury, the latest study showed that MT-I/II could stimulate brain anti-inflammatory factors, growth factors, neurotrophic factors and the expression of the receptor, and promote the extension of axon of neuron, which makes contribution to the regeneration of neuron and has important effect on the recovery of brain injury. Based on the findings, this article reviews the structure, expression, distribution, adjustion, function, mechanism in the repair of brain injury of MT-I/II and its application prospect in forensic medicine. It could provide a new approach for the design and manufacture of brain injury drugs as well as for age estimation of the brain injury.

  17. Fipronil is a powerful uncoupler of oxidative phosphorylation that triggers apoptosis in human neuronal cell line SHSY5Y.

    Science.gov (United States)

    Vidau, Cyril; González-Polo, Rosa A; Niso-Santano, Mireia; Gómez-Sánchez, Rubén; Bravo-San Pedro, José M; Pizarro-Estrella, Elisa; Blasco, Rafael; Brunet, Jean-Luc; Belzunces, Luc P; Fuentes, José M

    2011-12-01

    Fipronil is a phenylpyrazole insecticide known to elicit neurotoxicity via an interaction with ionotropic receptors, namely GABA and glutamate receptors. Recently, we showed that fipronil and other phenylpyrazole compounds trigger cell death in Caco-2 cells. In this study, we investigated the mode of action and the type of cell death induced by fipronil in SH-SY5Y human neuroblastoma cells. Flow cytometric and western blot analyses demonstrated that fipronil induces cellular events belonging to the apoptosis process, such as mitochondrial potential collapse, cytochrome c release, caspase-3 activation, nuclear condensation and phosphatidylserine externalization. In addition, fipronil induces a rapid ATP depletion with concomitant activation of anaerobic glycolysis. This cellular response is characteristic of mitochondrial injury associated with a defect of the respiration process. Therefore, we also investigated the effect of fipronil on the oxygen consumption in isolated mitochondria. Interestingly, we show for the first time that fipronil is a strong uncoupler of oxidative phosphorylation at relative low concentrations. Thus in this study, we report a new mode of action by which the insecticide fipronil could triggers apoptosis.

  18. Arsenic metalation of seaweed Fucus vesiculosus metallothionein: the importance of the interdomain linker in metallothionein.

    Science.gov (United States)

    Ngu, Thanh T; Lee, Janice A; Rushton, Moira K; Stillman, Martin J

    2009-09-22

    The presence of metallothionein in seaweed Fucus vesiculosus has been suggested as the protecting agent against toxic metals in the contaminated waters it can grow in. We report the first kinetic pathway data for A3+ binding to an algal metallothionein, F. vesiculosus metallothionein (rfMT). The time and temperature dependence of the relative concentrations of apo-rfMT and the five As-containing species have been determined following mixing of As3+ and apo-rfMT using electrospray ionization mass spectrometry (ESI MS). Kinetic analysis of the detailed time-resolved mass spectral data for As3+ metalation allows the simulation of the metalation reactions showing the consumption of apo-rfMT, the formation and consumption of As1- to As4-rfMT, and subsequent, final formation of As5-rfMT. The kinetic model proposed here provides a stepwise analysis of the metalation reaction showing time-resolved occupancy of the Cys7 and the Cys9 domain. The rate constants (M(-1) s(-1)) calculated from the fits for the 7-cysteine gamma domain are k1gamma, 19.8, and k2gamma, 1.4, and for the 9-cysteine beta domain are k1beta, 16.3, k2beta, 9.1, and k3beta, 2.2. The activation energies and Arrhenius factors for each of the reaction steps are also reported. rfMT has a long 14 residue linker, which as we show from analysis of the ESI MS data, allows each of its two domains to bind As3+ independently of each other. The analysis provides for the first time an explanation of the differing metal-binding properties of two-domain MTs with linkers of varying lengths, suggesting further comparison between plant (with long linkers) and mammalian (with short linkers) metallothioneins will shed light on the role of the interdomain linker.

  19. Exposure to Cell Phone Radiation Up-Regulates Apoptosis Genes in Primary Cultures of Neurons and Astrocytes

    OpenAIRE

    Zhao, Tian-Yong; Zou, Shi-Ping; Pamela E Knapp

    2006-01-01

    The health effects of cell phone radiation exposure are a growing public concern. This study investigated whether expression of genes related to cell death pathways are dysregulated in primary cultured neurons and astrocytes by exposure to a working GSM (Global System for Mobile Communication) cell phone rated at a frequency of 1900 MHz. Primary cultures were exposed to cell phone emissions for 2 hrs. We used array analysis and real-time RT-PCR to show up-regulation of caspase-2, caspase-6 an...

  20. Neuronal apoptosis and inflammatory reaction in rat models of focal cerebral ischemia following 40-minute suspended moxibustion

    Institute of Scientific and Technical Information of China (English)

    Rixin Chen; Zhimai Lv; Mingren Chen; Xin An; Dingyi Xie; Jing Yi

    2011-01-01

    The treatment duration of heat-sensitive moxibustion (approximately 40 minutes on average) is longer than that of traditional suspended moxibustion.The present study investigated expression changes of three inflammatory and apoptosis-associated proteins (inducible nitric oxide synthase,cyclooxygenase-2 and caspase-3) in transient middle cerebral artery occlusion model rats following suspended moxibustion for 40 minutes,to explore the mechanisms underlying neuroprotective action of suspended moxibustion.The results indicated that suspended moxibustion at acupoint Dazhui (DU 14) for 40 minutes reduced the cortical expression of caspase-3,cyclooxygenase-2 and inducible nitric oxide synthase proteins of transient middle cerebral artery occlusion model rats,as well as decreasing infarct volume and ameliorating the neurological deficit score.Outcomes with 40 minutes of moxibustion were superior to the outcomes after suspended moxibustion for 15 minutes.

  1. Apoptosis and impairment of neurite network by short exposure of immature rat cortical neurons to unconjugated bilirubin increase with cell differentiation and are additionally enhanced by an inflammatory stimulus.

    Science.gov (United States)

    Falcão, Ana S; Silva, Rui F M; Pancadas, Sérgio; Fernandes, Adelaide; Brito, Maria A; Brites, Dora

    2007-05-01

    Nerve cell injury induced by unconjugated bilirubin (UCB) has been implicated in brain damage during severe neonatal hyperbilirubinemia, although the molecular mechanisms underlying UCB neurotoxicity are still not clarified. It has been suggested recently that there is an association between hyperbilirubinemia and long-term neurologic dysfunctions. We incubated immature neurons with UCB to evaluate the short- and long-term effects of UCB on apoptotic death and on neuritic outgrowth and ramification. We also evaluated whether mature neurons, exposed previously to UCB in an early stage of differentiation, are more sensitive to apoptosis or to neuritic breakdown when treated with inflammatory agents, such as lipopolysaccharide and tumor necrosis factor-alpha. Results show that exposure of immature neurons to UCB increased apoptosis and provoked a reduction of both neurite extension and number of nodes. These injurious effects observed in immature cells treated with UCB were increasingly perpetuated along cell differentiation, as compared to neurons incubated in the absence of UCB. In addition, neurons that were exposed to UCB when immature showed an increased susceptibility to death by apoptosis, as well as an additional decrease in neurite outgrowth when incubated with an inflammatory agent afterward. This work shows, for the first time, that UCB induces neurite changes consistent with neurodevelopment abnormalities. Furthermore, pre-exposure to UCB followed by an inflammatory stimulus leads to an enhanced susceptibility to long-term apoptosis, as well as a greater neuritic breakdown. These data support the association between neonatal hyperbilirubinemia and the later development of mental illness, such as schizophrenia.

  2. Isolation and characterization of a hepatic metallothionein from mice.

    Science.gov (United States)

    Nordberg, M; Nordberg, G F; Piscator, M

    1975-01-01

    Mice were given cadmium chloride orally for 18 weeks, after which they were given two subcutaneous injections of cadmium chloride. Cadmium-and zinc-containing proteins were obtained from livers by ultracentrifugation, Sephadex chromatography and isoelectric focusing. Metallothionein with apI of 4.2 at 8 degrees C was separated from other proteins (possibly including other forms of metallothionein) by the isoelectric procedure. The metallothionein with pI = 4.2 had a high E250, reflecting abundant Cd-SH bonds, in accord with amino acid analysis, which gave a cysteine content of 34.6 residues % and showed an absence of aromatic amino acids. These results indicated a very high purity of the form of metallothionein isolated after isoelectric focusing.

  3. Apoptosis in Critical Conditions

    Directory of Open Access Journals (Sweden)

    A. M. Golubev

    2006-01-01

    Full Text Available Apoptosis is a variant of programmed cell death. This term was introduced by Kerr et al. in 1972, but information on the important role of apoptosis of some cells in critical conditions has recently appeared. The review of literature considers the basic mechanisms of induction, development, and regulation of apoptosis. Based on a literature update, the authors analyze the role of apoptosis in the pathogenesis of various critical conditions: acute lung lesion (neutrophilic and epithelial hypotheses, sepsis, myocardial infarction, and ischemic stroke (apoptosis of tubular epithelial cells, hepatic dysfunction in sepsis, myopathies in critical conditions. The data of studies dealing with the effects of inhaled and non-inhaled anesthetics on the apoptosis of neurons of the brain and lymphocytes are given. The review of literature presents the options of therapeutic apoptosis modulation by pharmacological methods.  

  4. Treatment of Wilson's disease with zinc: X. Intestinal metallothionein induction.

    Science.gov (United States)

    Yuzbasiyan-Gurkan, V; Grider, A; Nostrant, T; Cousins, R J; Brewer, G J

    1992-09-01

    Oral zinc therapy is effective in controlling copper balance in patients with Wilson's disease and blocks the intestinal absorption of copper, as demonstrated by uptake of copper 64 and copper balance measurements. In this study, 64Cu uptake measurements were concomitantly carried out with intestinal biopsies to investigate the relationship of reduced copper absorption to the levels of intestinal metallothionein in patients with Wilson's disease at different stages of zinc therapy. A pronounced increase in intestinal metallothionein levels and a sharp drop in 64Cu absorption were found 4 to 5 days after the initiation of zinc treatment. Conversely, metallothionein levels decreased and 64Cu uptake increased on the discontinuation of zinc therapy. The data indicate that 64Cu absorption varies as a function of intestinal metallothionein level. Intestinal metallothionein levels were found to correlate linearly with urinary zinc levels, which reflect body zinc status. These findings support our hypothesis that intestinal metallothionein induction mediates decreased copper absorption observed during zinc therapy. The suppressive effect of zinc on copper absorption appears to have a half-life of about 11 days.

  5. Long Coding RNA XIST Contributes to Neuronal Apoptosis through the Downregulation of AKT Phosphorylation and Is Negatively Regulated by miR-494 in Rat Spinal Cord Injury.

    Science.gov (United States)

    Gu, Shixin; Xie, Rong; Liu, Xiaodong; Shou, Jiajun; Gu, Wentao; Che, Xiaoming

    2017-04-01

    Recent evidence has suggested that long non-coding RNAs (lncRNAs) may play a significant role in the pathogenesis of several neurological diseases, including spinal cord injury (SCI). However, little is known about the role of lncRNAs in SCI. The aim of the present study was to evaluate the potential functions of lncRNAs in SCI and to identify the underlying mechanisms of action. We firstly analyzed Gene Expression Omnibus (GEO) datasets to investigate aberrantly-expressed lncRNAs which might be involved in the pathogenesis of SCI. The long non-coding RNA X-inactive specific transcript (XIST) was found to be one of the most significantly upregulated lncRNAs in the GEO dataset analysis, and is associated with apoptosis. We, therefore, selected this as a candidate lncRNA and investigated its function. We found that knockdown of lncRNA-XIST by Lv-shRNA had a prominent protective effect on SCI recovery by suppressing apoptosis through reactivation of the PI3K/AKT signaling pathway in rat spinal cord tissue. In particular, our results suggested that lncRNA-XIST may act as a competitive endogenous RNA, effectively becoming a sink for miR-494, leading to derepression of its target gene, phosphatase and tensin homolog deleted on chromosome ten (PTEN). In addition, an inverse relationship between lncRNA-XIST and miR-494 was observed in spinal cord tissues of SCI rats. Further study demonstrated that antagomiR-494 could reverse the protective effects of lncRNA-XIST knockdown on SCI rats through blocking the PTEN/PI3K/AKT signaling pathway. These results suggested that lncRNA-XIST knockdown may play an important role in limiting neuronal apoptosis in rats following SCI, and that the observed protective effects of lncRNA-XIST knockdown might have been mediated by its regulation on the phosphorylation of AKT by competitively binding miR-494. These findings have revealed, for the first time, the importance of the XIST/miR-494/PTEN/AKT signaling axis in the pathogenesis of SCI

  6. Electroacupuncture Suppressed Neuronal Apoptosis and Improved Cognitive Impairment in the AD Model Rats Possibly via Downregulation of Notch Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Hai-dong Guo

    2015-01-01

    Full Text Available Acupuncture is a potential strategy for the treatment of Alzheimer’s disease (AD and the possible mechanisms worth to be explored. In this study, we proposed and tested the hypothesis that whether Notch signaling pathway is involved in the effect of electroacupuncture (EA treatment. Rats that received EA treatment on the acupoints of Baihui (Du 20 and Shenshu (BL 23 had shorter latency and remained in the original platform quadrant longer and crossed the former platform contained quadrant more frequently compared to the Aβ injection rats without EA treatment. EA obviously alleviated the cell apoptosis resulted by Aβ infusion in hippocampus CA1 regions through upregulating the expression of Bcl-2 and downregulating the expression of Bax. EA could further obviously promote the expression of synapsin-1 and synaptophysin in hippocampus. Aβ injection significantly increased the expression of Notch1, Jag1, and Hes1 mRNA, while EA treatment downregulated the level of Notch1 and Hes1 mRNA in hippocampus, but not Jag1 mRNA. Our data suggested that EA treatment improved learning and memory function in the AD rat model partially through downregulating Notch signaling pathway.

  7. Deletion of Metallothionein Exacerbates Intermittent Hypoxia-Induced Oxidative and Inflammatory Injury in Aorta

    Directory of Open Access Journals (Sweden)

    Shanshan Zhou

    2014-01-01

    Full Text Available The present study was to explore the effect of metallothionein (MT on intermittent hypoxia (IH induced aortic pathogenic changes. Markers of oxidative damages, inflammation, and vascular remodeling were observed by immunohistochemical staining after 3 days and 1, 3, and 8 weeks after IH exposures. Endogenous MT was induced after 3 days of IH but was significantly decreased after 8 weeks of IH. Compared with the wild-type mice, MT knock-out mice exhibited earlier and more severe pathogenic changes of oxidative damages, inflammatory responses, and cellular apoptosis, as indicated by the significant accumulation of collagen, increased levels of connective tissue growth factor, transforming growth factor β1, tumor necrosis factor-alpha, vascular cell adhesion molecule 1,3-nitrotyrosine, and 4-hydroxy-2-nonenal in the aorta. These findings suggested that chronic IH may lead to aortic damages characterized by oxidative stress and inflammation, and MT may play a pivotal role in the above pathogenesis process.

  8. Neuron Apoptosis Induced by 3,4-methylenedioxy Methamphetamine and Expression of Apoptosis-related Factors in Rat Brain%3,4-亚甲基二氧基甲基苯丙胺诱导大鼠神经元凋亡及凋亡相关因子的表达

    Institute of Scientific and Technical Information of China (English)

    王雪; 祝三平; 况伟宏; 李静; 孙啸; 黄明生; 孙学礼

    2009-01-01

    Objective To study the neuron apoptosis induced by I. P 3.4-methylenedioxy methamphetamine (MDMA) and the expression of apoptosis-related factors in rat brain. Methods Twenty rats were divided into 4 groups. In group A, the rats were injected intraperitoneally with single dosage of saline, while the rats of group B, C, D were injected I. P with MDMA in different regimen, which were 20 mg/kg, single injection in group B, 20 mg/ kg twice a day (8 am and 8 pm) , for 2 day in group C, as well as 20 mg/kg, twice a day (8 am and 8 pm) for 4 days. Neuron apoptosis were measured by TUNEL, and the expression of Caspase-3 and CytC were detected by immunohistochemistry. Results Compared with saline group, apoptosis neurons were detected at the related brain regions (such as frontal cortex, hippocampus and striatum) of the rats in MDMA treated groups;Expression of Caspase-3 and CytC was observed at different level. Compared with group B, the number of apoptosis neurons of group C and D increased, and also the apoptosis-related factors in the brain tissue increased (P<0. 05). Conclusion MDMA could induce neurons apoptosis and the expression of apoptosis-related factors such as Caspase-3 and CytC in rat brain.Objective To study the neuron apoptosis induced by I. P 3.4-methylenedioxy methamphetamine (MDMA) and the expression of apoptosis-related factors in rat brain. Methods Twenty rats were divided into 4 groups. In group A, the rats were injected intraperitoneally with single dosage of saline, while the rats of group B, C, D were injected I. P with MDMA in different regimen, which were 20 mg/kg, single injection in group B, 20 mg/ kg twice a day (8 am and 8 pm) , for 2 day in group C, as well as 20 mg/kg, twice a day (8 am and 8 pm) for 4 days. Neuron apoptosis were measured by TUNEL, and the expression of Caspase-3 and CytC were detected by immunohistochemistry. Results Compared with saline group, apoptosis neurons were detected at the related brain regions (such as frontal

  9. Phenylbutyric acid rescues endoplasmic reticulum stress-induced suppression of APP proteolysis and prevents apoptosis in neuronal cells.

    Science.gov (United States)

    Wiley, Jesse C; Meabon, James S; Frankowski, Harald; Smith, Elise A; Schecterson, Leslayann C; Bothwell, Mark; Ladiges, Warren C

    2010-02-09

    The familial and sporadic forms of Alzheimer's disease (AD) have an identical pathology with a severe disparity in the time of onset [1]. The pathological similarity suggests that epigenetic processes may phenocopy the Familial Alzheimer's disease (FAD) mutations within sporadic AD. Numerous groups have demonstrated that FAD mutations in presenilin result in 'loss of function' of gamma-secretase mediated APP cleavage [2], [3], [4], [5]. Accordingly, ER stress is prominent within the pathologically impacted brain regions in AD patients [6] and is reported to inhibit APP trafficking through the secretory pathway [7], [8]. As the maturation of APP and the cleaving secretases requires trafficking through the secretory pathway [9], [10], [11], we hypothesized that ER stress may block trafficking requisite for normal levels of APP cleavage and that the small molecular chaperone 4-phenylbutyrate (PBA) may rescue the proteolytic deficit. The APP-Gal4VP16/Gal4-reporter screen was stably incorporated into neuroblastoma cells in order to assay gamma-secretase mediated APP proteolysis under normal and pharmacologically induced ER stress conditions. Three unrelated pharmacological agents (tunicamycin, thapsigargin and brefeldin A) all repressed APP proteolysis in parallel with activation of unfolded protein response (UPR) signaling-a biochemical marker of ER stress. Co-treatment of the gamma-secretase reporter cells with PBA blocked the repressive effects of tunicamycin and thapsigargin upon APP proteolysis, UPR activation, and apoptosis. In unstressed cells, PBA stimulated gamma-secretase mediated cleavage of APP by 8-10 fold, in the absence of any significant effects upon amyloid production, by promoting APP trafficking through the secretory pathway and the stimulation of the non-pathogenic alpha/gamma-cleavage. ER stress represses gamma-secretase mediated APP proteolysis, which replicates some of the proteolytic deficits associated with the FAD mutations. The small molecular

  10. Phenylbutyric acid rescues endoplasmic reticulum stress-induced suppression of APP proteolysis and prevents apoptosis in neuronal cells.

    Directory of Open Access Journals (Sweden)

    Jesse C Wiley

    Full Text Available BACKGROUND: The familial and sporadic forms of Alzheimer's disease (AD have an identical pathology with a severe disparity in the time of onset [1]. The pathological similarity suggests that epigenetic processes may phenocopy the Familial Alzheimer's disease (FAD mutations within sporadic AD. Numerous groups have demonstrated that FAD mutations in presenilin result in 'loss of function' of gamma-secretase mediated APP cleavage [2], [3], [4], [5]. Accordingly, ER stress is prominent within the pathologically impacted brain regions in AD patients [6] and is reported to inhibit APP trafficking through the secretory pathway [7], [8]. As the maturation of APP and the cleaving secretases requires trafficking through the secretory pathway [9], [10], [11], we hypothesized that ER stress may block trafficking requisite for normal levels of APP cleavage and that the small molecular chaperone 4-phenylbutyrate (PBA may rescue the proteolytic deficit. METHODOLOGY/PRINCIPAL FINDINGS: The APP-Gal4VP16/Gal4-reporter screen was stably incorporated into neuroblastoma cells in order to assay gamma-secretase mediated APP proteolysis under normal and pharmacologically induced ER stress conditions. Three unrelated pharmacological agents (tunicamycin, thapsigargin and brefeldin A all repressed APP proteolysis in parallel with activation of unfolded protein response (UPR signaling-a biochemical marker of ER stress. Co-treatment of the gamma-secretase reporter cells with PBA blocked the repressive effects of tunicamycin and thapsigargin upon APP proteolysis, UPR activation, and apoptosis. In unstressed cells, PBA stimulated gamma-secretase mediated cleavage of APP by 8-10 fold, in the absence of any significant effects upon amyloid production, by promoting APP trafficking through the secretory pathway and the stimulation of the non-pathogenic alpha/gamma-cleavage. CONCLUSIONS/SIGNIFICANCE: ER stress represses gamma-secretase mediated APP proteolysis, which replicates

  11. Regulation of a metallothionein-growth hormone hybrid gene in bovine papilloma virus.

    OpenAIRE

    Pavlakis, G N; Hamer, D H

    1983-01-01

    We have constructed bovine papilloma virus recombinants carrying a hybrid gene in which human growth hormone structural sequences are fused to the promoter and presumptive control region of the mouse metallothionein-I gene. Mouse cells transformed with the recombinants synthesize metallothionein-growth hormone hybrid mRNA with the same 5' end as metallothionein mRNA. Hybrid mRNA is inducible by cadmium but not by dexamethasone, whereas the chromosomal metallothionein genes in the same cells a...

  12. Metallothionein (MT)-III: generation of polyclonal antibodies, comparison with MT-I+II in the freeze lesioned rat brain and in a bioassay with astrocytes, and analysis of Alzheimer's disease brains

    DEFF Research Database (Denmark)

    Carrasco, J; Giralt, M; Molinero, A;

    1999-01-01

    Metallothionein-III is a low molecular weight, heavy-metal binding protein expressed mainly in the central nervous system. First identified as a growth inhibitory factor (GIF) of rat cortical neurons in vitro, it has subsequently been shown to be a member of the metallothionein (MT) gene family...... that MT-III immunoreactivity was also present in microglia, monocytes and/or macrophages in the leptomeninges and lying adjacent to major vessels. In freeze lesioned rats, both MT-I+II and MT-III immunoreactivities increased in the ipsilateral cortex. The pattern of MT-III immunoreactivity significantly...

  13. Metallothionein treatment reduces proinflammatory cytokines IL-6 and TNF-alpha and apoptotic cell death during experimental autoimmune encephalomyelitis (EAE)

    DEFF Research Database (Denmark)

    Penkowa, M; Hidalgo, J

    2001-01-01

    , which is characterized by significant inflammation and neuroglial damage. We have recently shown that the exogenous administration of the antioxidant protein zinc-metallothionein-II (Zn-MT-II) significantly decreased the clinical symptoms, mortality, and leukocyte infiltration of the CNS during EAE...... apoptotic cell death of neurons and oligodendrocytes during EAE, as judged by using TUNEL and immunoreactivity for cytochrome c and caspases 1 and 3. In contrast, the number of apoptotic lymphocytes and macrophages was less affected by Zn-MT-II treatment. The Zn-MT-II-induced decrease in proinflammatory...

  14. Effects of Metallothionein on Isolated Rat Heart

    Institute of Scientific and Technical Information of China (English)

    SUN Zhongdong; XIA Jiahong; DONG Nianguo; DU Xinling; CHI Yifan; YANG Tienan; YANG Chenyuan

    2007-01-01

    To investigate the effects of metallothionein (MT) on isolated rat heart, 16 Wistar rats were randomly divided into 2 groups. In control group (group C), distilled water was injected intraperitoneally and 24 h later isolated hearts were perfused with Langendorff and stored at 4℃ for 3 h with histidine-tryptophan-ketoglutarate (HTK) solutions, and then isolated hearts were perfused for 2 h by Langendorff. In experimental group (group E), 3.6% ZnSO4 was injected intraperitoneally, 24 h later isolated hearts were perfused by Langendorff and stored at 4℃ for 3 h with HTK solutions, and then the isolated herts were perfused for 2 h with Langendorff. MT content, the recovery of hemodynamics, myocardial water content (MWC), lactate dehydrogenase (LDH) and creatine kinase (CK) leakage, adenosine triphosphate (ATP) and malondialdehyde (MDA) content, superoxide dismutase (SOD) activity, myocardial cell Ca2+ content, Ca2+-ATPase activity of mitochondria ([Ca2+-ATPase]m) and its Ca2+ content ([Ca2+]m), synthesizing ATP activity of mitochondria ([ATP]m), and the ultrastructure of cells were examined. There were a significant increase in group E in hemodynamic recovery, ATP content, SOD activity, [Ca2+-ATPase]m activity, [ATP]m activity, and substantial reduction in MWC, LDH and CK leakage, MDA content, myocardial cell Ca2+ content, [Ca2+]m content,and the ultrastructural injury were obviously milder than that of group C. This study demonstrated that MT has protective effects on isolated rat heart.

  15. Chromium increases pancreatic metallothionein in the rat.

    Science.gov (United States)

    Solis-Heredia, M J; Quintanilla-Vega, B; Sierra-Santoyo, A; Hernández, J M; Brambila, E; Cebrián, M E; Albores, A

    2000-01-03

    The ability of chromium (Cr) salts to increase metallothionein (MT) levels in rat liver, kidney and pancreas, and its relationship with the presence of toxic effects are reported here. Rats were injected subcutaneously with 0, 10, 20, 30, 40, or 50 mg K2Cr2O7/kg and sacrificed 24 h later. Total Cr accumulation followed a dose-dependent pattern, levels in kidney being higher than those in liver or pancreas, suggesting different tissue bioavailabilities and accumulation patterns. Cr(IV) administration resulted in a tissue-specific MT induction: pancreas and liver showed five- and 3.5-fold MT increases, respectively; no increase was observed in the kidney. A positive correlation was observed between zinc and MT concentrations in liver, and between total Cr and MT concentrations in pancreas. Serum alpha-amylase activity showed a dose-dependent increase starting from 20 mg/kg, whereas serum glucose levels increased at doses higher than 30 mg/kg. Serum aspartate aminotransferase and alanine aminotransferase activities were increased in a dose-dependent manner, from 20 and 30 mg/kg, respectively. Our results showed that treatment with Cr(VI) can induce MT synthesis in pancreas and suggests a subsequent binding of Cr to MT. Also, pancreas is a target organ for Cr toxicity, and the usefulness of alpha-amylase activity as a sensitive biomarker of Cr toxicity in human exposed populations merits further study.

  16. Metallothionein in rabbit kidneys preserved for transplantation

    Energy Technology Data Exchange (ETDEWEB)

    Elinder, C.G.; Lundgren, G.; Nordberg, M.; Palm, B.; Piscator, M.

    1984-03-01

    Thirteen rabbits were given cadmium injections to achieve cadmium concentrations in kidney cortex ranging from 0.05 to 1 mmole Cd/kg wet weight. Another four animals served as controls. One kidney from each animal was frozen directly to -70/sup 0/C whereas the other kidney was kept for 24 hr at +4/sup 0/C in a preservative (Sachs' solution) to simulate conditions for preservation of human donor kidneys before transplantation. Protein binding of cadmium, zinc and copper in kidney homogenates and the concentration of metallothionein (MT) were measured in the kidney that was frozen directly and in the kidney that had been preserved. No gross differences in either the protein binding of cadmium, zinc and copper or in the MT content were seen between the directly frozen and preserved kidney from the same animal. This indicates that MT is not rapidly broken down in rabbit kidneys which have been preserved similarly to human donor kidneys for 24 hr in a standard preservative solution prior to a transplantation. 27 references, 3 figures, 1 table.

  17. Metallothionein in rabbit kidneys preserved for transplantation.

    Science.gov (United States)

    Elinder, C G; Lundgren, G; Nordberg, M; Palm, B; Piscator, M

    1984-03-01

    Thirteen rabbits were given repeated cadmium injections to achieve cadmium concentrations in kidney cortex ranging from 0.05 to 1 mmole Cd/kg wet weight. Another four animals served as controls. One kidney from each animal was frozen directly to -70 degrees C whereas the other kidney was kept for 24 hr at +4 degrees C in a preservative (Sachs' solution) to simulate conditions for preservation of human donor kidneys before transplantation. Protein binding of cadmium, zinc and copper in kidney homogenates and the concentration of metallothionein (MT) were measured in the kidney that was frozen directly and in the kidney that had been preserved. No gross differences in either the protein binding of cadmium, zinc and copper or in the MT content were seen between the directly frozen and preserved kidneys from the same animal. This indicates that MT is not rapidly broken down in rabbit kidneys which have been preserved similarly to human donor kidneys for 24 hr in a standard preservative solution prior to a transplantation.

  18. Electro-acupuncture-modulated miR-214 prevents neuronal apoptosis by targeting Bax and inhibits sodium channel Nav1.3 expression in rats after spinal cord injury.

    Science.gov (United States)

    Liu, Jing; Wu, Yaochi

    2017-03-11

    Electro-acupuncture (EA) has been proven to contribute towards neurologic and functional recoveries in spinal cord injury (SCI), but the underlying mechanism remains largely unknown especially regarding the effects of preventing neuronal apoptosis and alleviating neuropathic pain involved in the development of EA. In this study, we evaluated the effect of EA treatment in an animal model of SCI using the Basso, Beattie, and Bresnahan (BBB) score method, lesion volume by cresyl violet staining and neuronal apoptosis by TUNEL staining. Our results showed that EA therapy improved functional recovery, and reduced tissue loss and neuronal apoptosis after SCI. Meanwhile, we found that proapoptotic proteins (cleaved-caspase-3, 9 and cleaved-PARP) were downregulated and antiapoptotic protein Bcl-2 was upregulated following EA. To further explore the antiapoptotic effect of EA treatment, we verified that a large set of microRNAs (miRNAs) expression were altered following EA treatment and the miR-214 was one of the miRNAs being most significantly upregulated. Importantly, we validated both apoptosis related protein Bax and pain related protein Nav1.3 as two functional targets of miR-214 in vitro and vivo. Furthermore, our data showed that EA attenuates SCI-induced Nav1.3 and Bax upregulation in injured spinal cord via upregulating miR-214. These results suggest that miR-214 played an important role after SCI in the process of EA therapy, and the miR-214 could become an attractive novel therapeutic target for the treatment of SCI.

  19. Apoptosis-inducing factor downregulation increased neuronal progenitor, but not stem cell, survival in the neonatal hippocampus after cerebral hypoxia-ischemia

    Directory of Open Access Journals (Sweden)

    Sun Yanyan

    2012-04-01

    Full Text Available Abstract Background A considerable proportion of all newly generated cells in the hippocampus will die before becoming fully differentiated, both under normal and pathological circumstances. The caspase-independent apoptosis-inducing factor (AIF has not been investigated previously in this context. Results Postnatal day 8 (P8 harlequin (Hq mutant mice, expressing lower levels of AIF, and wild type littermates were injected with BrdU once daily for two days to label newborn cells. On P10 mice were subjected to hypoxia-ischemia (HI and their brains were analyzed 4 h, 24 h or 4 weeks later. Overall tissue loss was 63.5% lower in Hq mice 4 weeks after HI. Short-term survival (4 h and 24 h of labeled cells in the subgranular zone was neither affected by AIF downregulation, nor by HI. Long-term (4 weeks survival of undifferentiated, BLBP-positive stem cells was reduced by half after HI, but this was not changed by AIF downregulation. Neurogenesis, however, as judged by BrdU/NeuN double labeling, was reduced by half after HI in wild type mice but preserved in Hq mice, indicating that primarily neural progenitors and neurons were protected. A wave of cell death started early after HI in the innermost layers of the granule cell layer (GCL and moved outward, such that 24 h after HI dying cells could be detected in the entire GCL. Conclusions These findings demonstrate that AIF downregulation provides not only long-term overall neuroprotection after HI, but also protects neural progenitor cells, thereby rescuing hippocampal neurogenesis.

  20. Environmental neurotoxic pesticide dieldrin activates a non receptor tyrosine kinase to promote PKCδ-mediated dopaminergic apoptosis in a dopaminergic neuronal cell model.

    Science.gov (United States)

    Saminathan, Hariharan; Asaithambi, Arunkumar; Anantharam, Vellareddy; Kanthasamy, Anumantha G; Kanthasamy, Arthi

    2011-10-01

    Oxidative stress and apoptosis are two key pathophysiological mechanisms underlying dopaminergic degeneration in Parkinson's disease (PD). Recently, we identified that proteolytic activation of protein kinase C-delta (PKCδ), a member of the novel PKC family, contributes to oxidative stress-induced dopaminergic degeneration and that phosphorylation of tyrosine residue 311 (tyr311) on PKCδ is a key event preceding the PKCδ proteolytic activation during oxidative damage. Herein, we report that a non-receptor tyrosine kinase Fyn is significantly expressed in a dopaminergic neuronal N27 cell model. Exposure of N27 cells to the dopaminergic toxicant dieldrin (60 μM) rapidly activated Fyn kinase, PKCδ-tyr311 phosphorylation and proteolytic cleavage. Fyn kinase activation precedes the caspase-3-mediated proteolytic activation of PKCδ. Pre-treatment with p60-tyrosine-specific kinase inhibitor (TSKI) almost completely attenuated dieldrin-induced phosphorylation of PKCδ-tyr311 and its proteolytic activation. Additionally, TSKI almost completely blocked dieldrin-induced apoptotic cell death. To further confirm Fyn's role in the pro-apoptotic function of PKCδ, we adopted the RNAi approach. siRNA-mediated knockdown of Fyn kinase also effectively attenuated dieldrin-induced phosphorylation of PKCδ-tyr311, caspase-3-mediated PKCδ proteolytic cleavage, and DNA fragmentation, suggesting that Fyn kinase regulates the pro-apoptotic function of PKCδ. Collectively, these results demonstrate for the first time that Fyn kinase is a pro-apoptotic kinase that regulates upstream signaling of the PKCδ-mediated apoptotic cell death pathway in neurotoxicity models of pesticide exposure. Copyright © 2011 Elsevier Inc. All rights reserved.

  1. Metallothionein dimers studied by nano-spray mass spectrometry.

    Science.gov (United States)

    Hathout, Yetrib; Reynolds, Kristy J; Szilagyi, Zoltan; Fenselau, Catherine

    2002-01-15

    Both transient and stable dimers of metallothionein have been characterized, based on earlier studies using NMR, circular dichroism and size-exclusion chromatography. Here additional characterization is provided by nanospray mass spectrometry. Rapid redistribution of metal ions between monomeric Cd7- and Zn7-metallothionein 2a is monitored by nanospray. An experiment in which theses two forms of the monomeric protein are separated by a dialysis membrane, which will pass metal ions but not proteins, confirms that a transient dimer must form for metal ions to be redistributed. On the other hand, size-exclusion chromatography of reconstituted Zn7- or Cd7-metallothionein revealed the presence of monomeric and dimeric species. These dimers do not equilibrate readily to form monomers and they are shown to be covalent.

  2. Apoptosis of midbrain neurons induced by sodium-selenite%亚硒酸钠诱发大鼠胚胎中脑神经细胞凋亡的观察

    Institute of Scientific and Technical Information of China (English)

    吴健民; 陈婉蓉; 傅中滇; 阮素云

    2001-01-01

    目的 研究亚硒酸钠对大鼠胚胎中脑神经细胞凋亡的影响,并探讨其中可能的机制。方法 采用原代培养的大鼠胚胎中脑神经细胞,加入不同浓度的亚硒酸钠(1、5、10、15、20μmol/L),观察亚硒酸钠对神经细胞凋亡的影响,用透射电镜观察神经细胞超微结构的变化,流式细胞仪分析法检测DNA含量及凋亡细胞百分率。结果高浓度的亚硒酸钠(≥10μmol/L)会导致神经细胞凋亡,并呈剂量依赖性。10 μmol/L的亚硒酸钠作用过的神经细胞呈现出典型的凋亡形态学改变,且凋亡细胞百分率达13.4%。结论高浓度的硒可通过激发产生氧自由基和调节有关凋亡基因(P53)的活性等多种途径诱发神经细胞凋亡。%Objective To investigate the effects of sodium-selenite on midbrain neurons apoptosis. Methods Sodium-selenite was exposed to primary cultured midbrain neurons from the rat embryo midbrain;Morphological changes were observed with transmision electron microscope; The DNA content and percentage of apoptosis were measured by flow cytometer. Results Sodiumselenite selectively mduced the apoptosis of neurons dependent on its concentration, typical apoptosis was observed in 10μmol/L group and the percentage of apoptosic cells was up to 13.4%. Conclusion The mechanisms of neurons apoptosis induced by sodium-selenite in high concentration may be through producing oxygen free radicals or regulating its activity of related apoptosic gene (P53) or other ways.

  3. Metallothioneins in human tumors and potential roles in carcinogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Cherian, M. George; Jayasurya, A.; Bay, Boon-Huat

    2003-12-10

    Metallothioneins (MT) are a group of low-molecular weight, cysteine rich intracellular proteins, which are encoded by a family of genes containing at least 10 functional isoforms in human. The expression and induction of these proteins have been associated with protection against DNA damage, oxidative stress and apoptosis. Moreover, MT may potentially activate certain transcriptional factors by donating zinc. Although MT is a cytosolic protein in resting cells, it can be translocated transiently to the cell nucleus during cell proliferation and differentiation. A number of studies have shown an increased expression of MT in various human tumors of the breast, colon, kidney, liver, lung, nasopharynx, ovary, prostate, salivary gland, testes, thyroid and urinary bladder. However, MT is down-regulated in certain tumors such as hepatocellular carcinoma and liver adenocarcinoma. Hence, the expression of MT is not universal to all human tumors, but may depend on the differentiation status and proliferative index of tumors, along with other tissue factors and gene mutations. In certain tumors such as germ cell carcinoma, the expression of MT is closely related to the tumor grade and proliferative activity. Increased expression of MT has also been observed in less differentiated tumors. Thus, expression of MT may be a potential prognostic marker for certain tumors. There are few reports on the expression of the different isoforms of MT which have been analyzed by specific gene probes. They reveal that certain isoforms are expressed in specific cell types. The factors which can influence MT induction in human tumors are not yet understood. Down-regulation of MT synthesis in hepatic tumors may be related to hypermethylation of the MT-promoter or mutation of other genes such as the p53 tumor suppressor gene. In vitro studies using human cancer cells suggest a possible role for p53 and the estrogen-receptor on the expression and induction of MT in epithelial neoplastic cells

  4. Dynamic changes of apoptosis in rat cerebral cortex neurons after hypoxia%大鼠大脑皮层神经元缺氧后细胞凋亡情况的动态观察

    Institute of Scientific and Technical Information of China (English)

    邹哲华; 陶陶; 徐坚; 刘智; 罗开俭

    2012-01-01

    Objective To observe the dynamic changes of apoptosis in rat cerebral cortex neurons after hypoxia. Methods Rat cerebral cortex neurons were primarily cultured from SD rats born within 24 h and then identified by immunocytochemical assay. Then the identified cells were cultured in the medium containing 100 μmol/L CoCl2 to simulate hypoxic condition. The cells cultivated in normal condition served as normal control ( normoxia group). Ultrastructural changes of the neurons were observed by transmission electron microscopy (TEM) . Neuronal apoptosis were observed by TUNEL assay. Results TEM displayed that the morphology of neurons was normal, so was the structure of chromatin, endoplasmic reticulum and mitochondria in normoxia group, while, cellular edema, organelle damage or disappearance were seen in the hypoxia group. TUNEL showed that obvious apoptosis were found in hypoxic cells, with significant difference with normoxia group ( P < 0. 01). The apoptosis reached its peak in 48 h after hypoxia (0. 187 ±0. 007) , significantly higher than those in 12, 24 and 72 h (P <0. 01). Conclusion Apoptosis is a dynamic process in hypoxic-ischemic brain injury, and an important pattern of neuronal death. Intervention for neuronal apoptosis should be performed in an appropriate time window to effectively treat hypoxic-ischemic encephalopathy.%目的 观察大鼠大脑皮层神经元缺氧后细胞凋亡动态变化.方法 制备大鼠大脑皮层神经元体外原代培养模型,免疫细胞化学鉴定大鼠大脑皮层神经元,透射电镜下观察不同时间点各实验组神经元超微结构的变化,TUNEL法观察不同时间点各实验组神经元凋亡情况.结果 正常对照组神经元透射电镜下形态及染色质正常、内质网、线粒体等结构正常,缺氧组神经元水肿,细胞器破坏或消失;TUNEL法检测神经元凋亡:缺氧后各组神经元凋亡明显增加,与相应正常对照组相比有显著差异(P<0.01),缺氧48 h

  5. Metallothionein in human gingival amalgam tattoos.

    Science.gov (United States)

    Lau, J C; Jackson-Boeters, L; Daley, T D; Wysocki, G P; Cherian, M G

    2001-11-01

    Amalgam tattoos occur when small particles of dental amalgam, composed largely of silver (Ag) and mercury (Hg), are inadvertently implanted into oral soft tissues during dental procedures. Metallothioneins (MTs) are ubiquitous, low molecular weight, cysteine-rich, metal-binding proteins that are inducible by many agents including metals and may be involved in the detoxification of toxic metals such as Hg. In this study, the correlation between MT expression and amalgam tattoos in human gingiva was investigated using energy-dispersive X-ray microanalysis (EDX) and immunohistochemical techniques. Light microscopically, amalgam tattoos presented as either fine granular particles or larger discrete opaque globular particles in connective tissues. EDX revealed the smaller particles to be silver sulphide (Ag(2)S), while the larger particles exhibited a shell of Ag(2)S that contained irregularly distributed masses of Ag and Hg. Particles of tin (Sn) were also found. No MT staining was observed in collagen, fibroblasts or blood vessels in areas exhibiting abundant amounts of embedded fine granular Ag(2)S particles. Blood vessels exhibiting relatively few amalgam particles stained positively for MT. Cells with the morphological features of histiocytes located directly adjacent to larger pieces of amalgam showed intense MT staining. These results indicate that amalgam tattoos contain no Hg or free Ag except in large globular pieces of amalgam, which still contain Hg and which induce MT expression in adjacent histiocytes. This suggests that Hg leaching from impacted dental amalgam particles induces MT, while residual Ag(2)S and Sn particles do not. MT may therefore act to reduce Hg exposure in patients with amalgam tattoos.

  6. Metallothioneins for correlative light and electron microscopy.

    Science.gov (United States)

    Fernández de Castro, Isabel; Sanz-Sánchez, Laura; Risco, Cristina

    2014-01-01

    Structural biologists have been working for decades on new strategies to identify proteins in cells unambiguously. We recently explored the possibilities of using the small metal-binding protein, metallothionein (MT), as a tag to detect proteins in transmission electron microscopy. It had been reported that, when fused with a protein of interest and treated in vitro with gold salts, a single MT tag will build an electron-dense gold cluster ~1 nm in diameter; we provided proof of this principle by demonstrating that MT can be used to detect intracellular proteins in bacteria and eukaryotic cells. The method, which is compatible with a variety of sample processing techniques, allows specific detection of proteins in cells with exceptional sensitivity. We illustrated the applicability of the technique in a series of studies to visualize the intracellular distribution of bacterial and viral proteins. Immunogold labeling was fundamental to confirm the specificity of the MT-gold method. When proteins were double-tagged with green fluorescent protein and MT, direct correlative light and electron microscopy allowed visualization of the same macromolecular complexes with different spatial resolutions. MT-gold tagging might also become a useful tool for mapping proteins into the 3D-density maps produced by (cryo)-electron tomography. New protocols will be needed for double or multiple labeling of proteins, using different versions of MT with fluorophores of different colors. Further research is also necessary to render the MT-gold labeling procedure compatible with immunogold labeling on Tokuyasu cryosections and with cryo-electron microscopy of vitreous sections.

  7. Neuronal and glial cell type-specific promoters within adenovirus recombinants restrict the expression of the apoptosis-inducing molecule Fas ligand to predetermined brain cell types, and abolish peripheral liver toxicity.

    Science.gov (United States)

    Morelli, A E; Larregina, A T; Smith-Arica, J; Dewey, R A; Southgate, T D; Ambar, B; Fontana, A; Castro, M G; Lowenstein, P R

    1999-03-01

    Gene therapy using Fas ligand (FasL) for treatment of tumours and protection of transplant rejection is hampered because of the systemic toxicity of FasL. In the present study, recombinant replication-defective adenovirus vectors (RAds) encoding FasL under the control of either the neuronal-specific neuronal-specific enolase (NSE) promoter or the astrocyte-specific glial fibrillary acidic protein (GFAP) promoter have been constructed. The cell type-specific expression of FasL in both neurons and glial cells in primary cultures, and in neuronal and glial cell lines is demonstrated. Furthermore, transgene expression driven by the neuronal and glial promoter was not detected in fibroblastic or epithelial cell lines. Expression of FasL driven by a major immediate early human cytomegalovirus promoter (MIEhCMV) was, however, achieved in all cells tested. As a final test of the stringency of transgene-specific expression, the RAds were injected directly into the bloodstream of mice. The RAds encoding FasL under the control of the non-cell type-specific MIEhCMV promoter induced acute generalized liver haemorrhage with hepatocyte apoptosis, while the RAds containing the NSE or GFAP promoter sequences were completely non-toxic. This demonstrates the specificity of transgene expression, enhanced safety during systemic administration, and tightly regulated control of transgene expression of highly cytotoxic gene products, encoded within transcriptionally targeted RAds.

  8. Chronic intermittent hypoxia-induced neuronal apoptosis in the hippocampus is attenuated by telmisartan through suppression of iNOS/NO and inhibition of lipid peroxidation and inflammatory responses.

    Science.gov (United States)

    Yuan, Xiao; Guo, Xueling; Deng, Yan; Zhu, Die; Shang, Jin; Liu, Huiguo

    2015-01-30

    Obstructive sleep apnea syndrome (OSAS) plays a critical role in the initiation and progression of Alzheimer׳s disease (AD), but little is known about the precise mechanism of OSAS-induced AD. Nitric oxide synthase (NOS) and nitric oxide (NO) are known to play key roles in the development of AD. Several studies have confirmed that an angiotensin II type 1 receptor blocker, telmisartan, beneficially regulates NOS and NO. Here, we examined the neuroprotective effects of telmisartan against hippocampal apoptosis induced by chronic intermittent hypoxia (CIH), the most characteristic pathophysiological change of OSAS. Adult male Sprague Dawley rats were subjected to 8h of intermittent hypoxia per day with or without telmisartan for eight weeks. Neuronal apoptosis in the hippocampal CA1 region, NOS activity, NO content, and the presence of inflammatory agents and radical oxygen species in the hippocampus were determined. The results showed that CIH activated inducible nitric oxide synthase (iNOS), increased NO content, and enhanced lipid peroxidation and inflammatory responses in the hippocampus. Treatment with telmisartan inhibited excessive iNOS and NO generation and reduced lipid peroxidation and inflammatory responses. In addition, telmisartan significantly ameliorated the hippocampal apoptosis induced by CIH. In conclusion, Pre-CIH telmisartan administration attenuated CIH-induced hippocampal apoptosis partly by regulating NOS activity, inhibiting excessive NO generation, and reducing lipid peroxidation and inflammatory responses.

  9. Barley metallothioneins differ in ontogenetic pattern and response to metals

    DEFF Research Database (Denmark)

    Schiller, Michaela; Hegelund, Josefine Nymark; Pedas, Pai

    2014-01-01

    The barley genome encodes a family of 10 metallothioneins (MTs) that have not previously been subject to extensive gene expression profiling. We show here that expression of MT1a, MT2b1, MT2b2 and MT3 in barley leaves increased more than 50-fold during the first 10 d after germination. Concurrent...

  10. Label-free bead-based metallothionein electrochemical immunosensor.

    Science.gov (United States)

    Nejdl, Lukas; Nguyen, Hoai Viet; Richtera, Lukas; Krizkova, Sona; Guran, Roman; Masarik, Michal; Hynek, David; Heger, Zbynek; Lundberg, Karin; Erikson, Kristofer; Adam, Vojtech; Kizek, Rene

    2015-08-01

    A novel microfluidic label-free bead-based metallothionein immunosensors was designed. To the surface of superparamagnetic agarose beads coated with protein A, polyclonal chicken IgY specifically recognizing metallothionein (MT) were immobilized via rabbit IgG. The Brdicka reaction was used for metallothionein detection in a microfluidic printed 3D chip. The assembled chip consisted of a single copper wire coated with a thin layer of amalgam as working electrode. Optimization of MT detection using designed microfluidic chip was performed in stationary system as well as in the flow arrangement at various flow rates (0-1800 μL/min). In stationary arrangement it is possible to detect MT concentrations up to 30 ng/mL level, flow arrangement allows reliable detection of even lower concentration (12.5 ng/mL). The assembled miniature flow chip was subsequently tested for the detection of MT elevated levels (at approx. level 100 μg/mL) in samples of patients with cancer. The stability of constructed device for metallothionein detection in flow arrangement was found to be several days without any maintenance needed.

  11. Role of metallothioneins in peripheral nerve function and regeneration

    DEFF Research Database (Denmark)

    Ceballos, D; Lago, N; Verdú, E;

    2003-01-01

    The physiological role of the metallothionein (MT) family of proteins during peripheral nerve injury and regeneration was examined in Mt1+ 2 and Mt3 knockout (KO) mice. To this end, the right sciatic nerve was crushed, and the regeneration distance was evaluated by the pinch test 2-7 days...

  12. CNS wound healing is severely depressed in metallothionein I- and II-deficient mice

    DEFF Research Database (Denmark)

    Penkowa, M; Carrasco, J; Giralt, M

    1999-01-01

    . In contrast to normal mice, at 20 dpl no wound healing had occurred. The rate of apoptosis, as determined by using terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling, was drastically increased in neurons of ipsilateral cortex of the MT-I+II null mice. Our results demonstrate that MT......-I+II are essential for a normal wound repair in the CNS, and that their deficiency impairs neuronal survival....

  13. Facial paralysis induced by herpes simplex virus type 1 and the study of facial neurons apoptosis in mice%小鼠病毒性面神经炎模型中面神经元凋亡的研究

    Institute of Scientific and Technical Information of China (English)

    刘稳; 高志强; 神平; 冯国栋; 查洋; 张彦东; 刘佩毛

    2011-01-01

    目的 探讨1型单纯疱疹病毒(herpes simplex virus type 1,HSV-1)性面神经炎动物模型中面神经运动神经元(facial motor neurons,FMN)的凋亡情况以及凋亡相关基因bcl-2和bax的表达。方法 Balb/c小鼠84只,分别进行HSV-1接种和面神经切断处理。在操作后第1、3、7、10、15、20及30天分批处死动物,对面神经核进行HE染色、尼氏染色;采用原位末端转移酶标记(terminaldeoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling,TUNEL)技术检测FMN的凋亡;免疫组化染色检测凋亡相关基因bcl-2和bax在FMN中的表达。结果 面神经切断后,同侧FMN出现凋亡,在第10、15天达高峰,Bcl-2表达降低,Bax表达升高,Bcl-2/Bax比值下降;接种病毒后,同侧FMN没有明显的细胞凋亡,Bcl-2表达增高,在第15天达高峰,Bcl-2/Bax比值上升。结论 小鼠面神经接种HSV-1后FMN没有明显凋亡,可能与神经损伤程度轻以及HSV-1抑制宿主细胞凋亡有关,Bcl-2、Bax可能参与调控FMN的凋亡。%Objective To study the apoptosis of facial motor neurons and the expression of apoptosis-related genes, Bcl-2 and Bax, in the animal model of viral facial paralysis. Methods Total of 84Balb/c mice were divided into viral inoculation group and nerve transaction group. The animals were executed 1, 3, 7, 10, 15, 20 and 30 days after being operated respectively. The histopathological features of facial neurons in brain stem were observed by HE and Nissl stain. The changes of facial neuronal apoptosis were observed by TUNEL The changes of expression of Bcl-2 and Bax genes in facial neurons were observed by immunohistochemistry staining. Results After nerve transection, increased apoptotic cells were found in homolateral facial motor neucleus and the peak appeared at 10 and 15 days. The level of Bcl-2 expression inneurons declined while the expression of Bax increased gradually. Correspondingly, the ratio of

  14. The essential role of p53-up-regulated modulator of apoptosis (Puma) and its regulation by FoxO3a transcription factor in β-amyloid-induced neuron death.

    Science.gov (United States)

    Akhter, Rumana; Sanphui, Priyankar; Biswas, Subhas Chandra

    2014-04-11

    Neurodegeneration underlies the pathology of Alzheimer disease (AD). The molecules responsible for such neurodegeneration in AD brain are mostly unknown. Recent findings indicate that the BH3-only proteins of the Bcl-2 family play an essential role in various cell death paradigms, including neurodegeneration. Here we report that Puma (p53-up-regulated modulator of apoptosis), an important member of the BH3-only protein family, is up-regulated in neurons upon toxic β-amyloid 1-42 (Aβ(1-42)) exposure both in vitro and in vivo. Down-regulation of Puma by specific siRNA provides significant protection against neuron death induced by Aβ(1-42). We further demonstrate that the activation of p53 and inhibition of PI3K/Akt pathways induce Puma. The transcription factor FoxO3a, which is activated when PI3K/Akt signaling is inhibited, directly binds with the Puma gene and induces its expression upon exposure of neurons to oligomeric Aβ(1-42). Moreover, Puma cooperates with another BH3-only protein, Bim, which is already implicated in AD. Our results thus suggest that Puma is activated by both p53 and PI3K/Akt/FoxO3a pathways and cooperates with Bim to induce neuron death in response to Aβ(1-42).

  15. 吗啡成瘾时脑细胞凋亡数目增加%Neuron apoptosis increases in morphine-dependent rats

    Institute of Scientific and Technical Information of China (English)

    刘立伟; 王新华; 傅强; 吴青华; 傅舒昆

    2012-01-01

    deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNED method. Immunohistochemistry was used to detect the expression of Caspase-3 protein. Results The score of abstinent symptom in morphine-dependent group is not statistically different from control group (P>0. 05), while it was significantly lower than that in morphine-abstinent group (P0.05). Conclusion Chronic morphine can promote abnormal neuronal apoptosis in rat brain, which might be related to opiate-induced nerve damage.

  16. Metallothionein-I/II Promotes Axonal Regeneration in the Central Nervous System*

    Science.gov (United States)

    Siddiq, Mustafa M.; Hannila, Sari S.; Carmel, Jason B.; Bryson, John B.; Hou, Jianwei; Nikulina, Elena; Willis, Matthew R.; Mellado, Wilfredo; Richman, Erica L.; Hilaire, Melissa; Hart, Ronald P.; Filbin, Marie T.

    2015-01-01

    The adult CNS does not spontaneously regenerate after injury, due in large part to myelin-associated inhibitors such as myelin-associated glycoprotein (MAG), Nogo-A, and oligodendrocyte-myelin glycoprotein. All three inhibitors can interact with either the Nogo receptor complex or paired immunoglobulin-like receptor B. A conditioning lesion of the sciatic nerve allows the central processes of dorsal root ganglion (DRG) neurons to spontaneously regenerate in vivo after a dorsal column lesion. After a conditioning lesion, DRG neurons are no longer inhibited by myelin, and this effect is cyclic AMP (cAMP)- and transcription-dependent. Using a microarray analysis, we identified several genes that are up-regulated both in adult DRGs after a conditioning lesion and in DRG neurons treated with cAMP analogues. One gene that was up-regulated under both conditions is metallothionein (MT)-I. We show here that treatment with two closely related isoforms of MT (MT-I/II) can overcome the inhibitory effects of both myelin and MAG for cortical, hippocampal, and DRG neurons. Intrathecal delivery of MT-I/II to adult DRGs also promotes neurite outgrowth in the presence of MAG. Adult DRGs from MT-I/II-deficient mice extend significantly shorter processes on MAG compared with wild-type DRG neurons, and regeneration of dorsal column axons does not occur after a conditioning lesion in MT-I/II-deficient mice. Furthermore, a single intravitreal injection of MT-I/II after optic nerve crush promotes axonal regeneration. Mechanistically, MT-I/II ability to overcome MAG-mediated inhibition is transcription-dependent, and MT-I/II can block the proteolytic activity of α-secretase and the activation of PKC and Rho in response to soluble MAG. PMID:25947372

  17. Comparison of gene expression of metallothioneins, ubiquitin and p53 in fibroblasts from lung and skin of rats of different age

    Directory of Open Access Journals (Sweden)

    Y. G. Kot

    2015-09-01

    Full Text Available We studied gene expression of five metallothioneins (MT 1-5, ubiquitin and protein p53 and their products in fibroblasts culture of the skin and lungs of white rats of different ages (2 weeks, 1, 3, and 24 months and determined its (metallothionein 1-5 types, ubiquitin, p53 product quantity. All these proteins are protective ones, but perform their functions by using different mechanisms. Metallothionein bind, transport and excrete ions of bivalent metals, ubiquitin controls the cleavage of the defective and short-lived proteins in the proteasome, protein p53 controls apoptosis, thus ensuring the genome stability. The similarity of age dynamics of gene expression of ubiquitin and MT of cells of both sources has been shown – maximum at 3 months. Expression of p53 gene has a difference: both in the skin and lungs expression increases up to 24 months. Product quantity of p53 has a minimum in the skin at 3 months and remains constant; in the lungs, this value has a maximum at 1 month.

  18. The relationship between the expression of CIDE-B and the neuronal apoptosis following cerebral ischemia reperfusion in rats%脑缺血再灌注后CIDE-B表达与神经元凋亡的关系

    Institute of Scientific and Technical Information of China (English)

    于竹芹; 杨丽艳; 刘广义

    2013-01-01

    目的探讨大鼠脑缺血/再灌注损伤后细胞死亡DNA片段裂解因子45样因子B(CIDE-B)表达与海马神经元凋亡的关系。方法成年健康雄性Wistar大鼠,应用线栓法建立大鼠大脑中动脉缺血再灌注模型,TUNEL法染色观察海马神经元凋亡,免疫组化法和Western blot法检测海马神经元CIDE-B蛋白表达,RT-PCR检测CIDE-B mRNA表达。结果与假手术组比较,脑缺血2h再灌注6h,1d,3d,7d,14d,凋亡神经元显著增加(P<0.01), CIDE-B mRNA和CIDE-B蛋白表达明显增强加(P<0.01);至缺血2h再灌注28d神经元凋亡数量显著较少, CIDE-B mRNA和蛋白表达明显减弱(P<0.01),细胞凋亡与CIDE-B基因表达的时相一致。结论脑缺血再灌注损伤后CIDE-B表达与神经元凋亡呈时相依赖性。%Objective To explore the relationship between the cell-death-inducing DNA-fragmentation factor (DFF45)-1ike effecter-B (CIDE-B) and the neuronal apoptosis after cerebral ischemia reperfusion in rats. Methods The middle cerebral artery occlusion and reperfusion (MCAO/R) models were established by inserting monofilament suture from left external-internal carotid artery in adult healthy male Wistar rats. The neuronal apoptosis in hippocampus was detected by TUNEL assay, the expression of CIDE-B protein were determined by immunohistochemical staining and Western blot and CIDE-B mRNA by RT-PCR respectively. Results After ischemia 2h and reperfusion 6h, 1d, 3d, 7d, 14d, the number of neuronal apoptosis was significantly increased (P<0.01) and the expressions of CIDE-B mRNA and protein were evaluated (P < 0.01), while all reduced significantly than those in the control group at reperfusion 28d (P < 0.01). The time-phase of neuronal apoptosis was coincident with the expressions of CIDE-B gene. Conclusion The neuronal apoptosis was timely dependent on the expression of CIDE-B gene after cerebral ischemia reperfusion in rats.

  19. Zinc or copper deficiency-induced impaired inflammatory response to brain trauma may be caused by the concomitant metallothionein changes

    DEFF Research Database (Denmark)

    Penkowa, Milena; Giralt, M.; Thomsen, Pernille Sjølin;

    2001-01-01

    damage to neurons as indicated by neuron specific enolase (NSE) ir, and the number of cells undergoing apoptosis (measured by TUNEL) was dramatically increased. Zinc deficiency significantly altered brain response to TBI, potentiating the microgliosis and reducing the astrogliosis, while increasing...

  20. Effects of Osthole on Neuronal Apoptosis and Cell Cycle in AD Rats%蛇床子素对AD大鼠神经元凋亡及细胞周期的影响

    Institute of Scientific and Technical Information of China (English)

    董晓华; 孟宪勇; 张力; 杨建明

    2012-01-01

    Objective:To study the neuroprotective effects and its mechanism of osthole by observing neuronal apoptosis and cell cycle in Alzheimer’s disease(AD)rats. Methods:An intracerebroventricular(icv)injection ofβ-amyloid peptide(Aβ25-35)was administrated to establish AD rat model. Osthole(12.5,25.0 mg·kg-1)was injected intraperitoneally to rats. Cognitive functions,neuronal apoptosis and cell cycle of AD rats were observed. Results:Osthole could improve spatial learning and memory abilities of AD rats,reduce neuronal apoptosis,increase the percentage of S phase cells,promote the G2/M phase cells to divide further,strengthen the cell proliferation activity,regulate cell cycle and benefit to the maintenance of normal physiological function of neurons. Conclusion:Osthole has neuronal protection through reducing neuronal apoptosis and regulating cell cycle,which may be one of mechanism of improving learning and memory disorder in AD rats.%目的:观察蛇床子素对阿尔茨海默病(Alzheimer’s disease,AD)模型大鼠神经元凋亡及细胞周期的影响,探讨蛇床子素的神经保护作用及其作用机制。方法:采用一次性侧脑室注射(icv)聚集态β淀粉样肽(β-amyloid peptide,Aβ25-35)建立AD大鼠模型,腹腔注射(ip)蛇床子素12.5,25.0 mg·kg-1进行干预,观察大鼠认知功能、神经元凋亡及细胞周期变化。结果:蛇床子素能明显改善AD模型大鼠空间学习记忆能力,减少神经元凋亡,增加S期细胞百分率,促进G2/M期细胞进一步分裂,增强细胞增殖活性,调节细胞周期,有利于维持神经元正常的生理功能。结论:蛇床子素可通过减少神经元凋亡、调节细胞周期发挥神经保护作用,这可能是其改善AD大鼠学习记忆障碍的作用机制之一。

  1. 硝普钠控制性降压对兔脑神经元c-fos及细胞凋亡的影响%Effect of induced hypotension on brain neuronal c-fos and apoptosis in rabbits

    Institute of Scientific and Technical Information of China (English)

    黄宏艳; 肖晓山; 周代伟

    2011-01-01

    Objective To explore the brain's tolerance and the safe low limit to the hypotension through observing the effect of four different levels of sodium nitroprusside induced hypotension on neuronal ultrastructure and apoptosis and expression of c-fos in rabbits' hippocampal CA1 field. Methods Thirty New Zealand rabbits were equally randomized into five groups: mean arterial pressure fell to 70% (group I), 60% (group Ⅱ), 50% (group Ⅱ ) and 45% (group Ⅳ) of the baseline and normal control group (group V ). After maintaining target blood pressure for 1 h, the rabbits were sacrificed 2 h after boosting pressure. The ultrastructure of CA1 field was observed with transmission electron microscopy. The expression of c-fos was assayed by immunohistochemical SP and apoptosis was detected by Tunel immunofluorescence in hippocampal CA1 neuron. Results In the transmission electron microscopy of hippocampal CA1 neuron, group [I appeared cellular swelling and groups Ⅲ and Ⅳ appeared karyopyknosis and apoptosis. The c-fos expression in CA1 neuron significantly increased in hypotension groups (P<0. 01). The TUNEL mean fluorescence intensity was much strengthen in groups H , III and IV than that in groups I and V ( P <0.01). Conclusion Controlled hypotension with sodium nitroprusside can cause apoptosis in partial hippocampal CA1 neurons. C-fos can make rapid respond according to changes of brain blood and oxygen supply, and its expression may produce protective effects on neurons.%通过观察硝普钠诱导的四种低血压水平对兔脑海马CA1区神经元超微结构、细胞凋亡及c-fos表达的影响,了解脑组织对低血压的耐受性以及探讨安全的降压低限.方法30只新西兰兔随机均分为五组,MAP分别降至基础值的70%(Ⅰ组)、60%(Ⅱ组)、50%(Ⅲ组)、45%(Ⅳ组)和不降压(Ⅴ组).目标血压维持1h,复压2h处死兔.透射电镜观察海马CA1区神经元超微结构,SP法测定该

  2. Preparation and functional analysis of recombinant protein transduction domain-metallothionein fusion proteins.

    Science.gov (United States)

    Lim, Kwang Suk; Won, Young-Wook; Park, Yong Soo; Kim, Yong-Hee

    2010-08-01

    In order for proteins to be used as pharmaceuticals, delivery technologies need to be developed to overcome biochemical and anatomical barriers to protein drug transport, to protect proteins from systemic degradation, and to target the drug action to specific sites. Protein transduction domains (PTDs) are used for the non-specific transduction of bio-active cargo, such as proteins, genes, and particles, through cellular membranes to overcome biological barriers. Metallothionein (MT) is a low molecular weight intra-cellular protein that consists of 61 amino acids, including 20 cysteine residues, and is over-expressed under stressful conditions. Although MT has the potential to improve the viability of islet cells and cardiomyocytes by inhibiting diabetic-induced apoptosis and by removing reactive oxygen species (ROS), and thereby prevent or reduce diabetes and diabetic complications, all MT applications have been made for gene therapy or under induced over-expression of endogenous MT. To overcome the drawbacks of ineffective intra-cellular MT protein uptake, a human MT gene was cloned and fused with protein transduction domains (PTDs), such as HIV-1 Tat and undeca-arginine, in a bacterial expression vector to produce PTD-MT fusion proteins. The expression and purification of three types of proteins were optimized by adding Zn ions to maintain their stability and functionality mimicking intra-cellular stable conformation of MT as a Zn-MT cluster. The Zn-MT cluster showed better stability than MT in vitro. PTD-MT fusion proteins strongly protected Ins-1 beta cells against oxidative stress and apoptosis induced by glucolipotoxicity with or without hypoxia, and also protected H9c2 cardiomyocytes against hyperglycemia-induced apoptosis with or without hypoxia. PTD-MT recombinant fusion proteins may be useful protein therapeutics for the treatment or prevention of diabetes and diabetes-related complications.

  3. Immunochemical Detection of Metallothionein in Specific Epithelial Cells of Rat Organs

    Science.gov (United States)

    Danielson, Keith G.; Ohi, Seigo; Huang, P. C.

    1982-04-01

    The distribution of a heavy metal binding protein, metallothionein, was studied immunocytochemically by using antimetallothionein antibody and the immunoperoxidase staining technique on histological sections of liver, kidney, intestine, lung, and testis from cadmium-treated rats. These tissues either accumulate heavy metals (e.g., liver, kidney, and testis) or are exposed to metal by ingestion or inhalation (intestine and lung). Staining for metallothionein was observed intracellularly in epithelial parenchymal cells of the liver and kidney; all hepatocytes and most renal tubular cells stained for the protein. Accumulation of metallothionein was not seen in connective tissue cells surrounding either blood vessels or renal tubules. Extracellular localization of metallothionein was also observed in the liver sinusoids and within the lumina of the renal tubules, suggesting a metal transport or excretory function for this protein. Surface columnar epithelial cells of the intestinal villi indicated the presence of metallothionein but connective tissue cells of the lamina propria were negative for the protein. The granular secretory Paneth cells of the small intestine also stained strongly for metallothionein as did respiratory epithelial cells of the lung. In the testis, metallothionein was detected in the Sertoli cells and interstitial cells but not within the spermatogonia. Sertoli cells are closely associated with the developing spermatogonia and appear to serve a nutritive role in spermatogenesis. Because of the secretory, absorptive, or nutritive function of the metallothionein-localizing cells in the organs studied, we suggest that metallothionein may be involved in metal storage or transport in addition to its commonly proposed detoxification role.

  4. JNK抑制剂及NAC对氧糖剥离后复氧复糖所致脊髓神经元损伤的保护作用%The study of JNK inhibitor and NAC on oxygen-glucose deprivation/regain induced neuronal apoptosis in cultured spinal neuronal cells

    Institute of Scientific and Technical Information of China (English)

    华政哲; 张子韬; 张宁; 胡志毅; 蔡卫华; 殷国勇

    2011-01-01

    Objective:To study the protective effects of JNK (the c-Jun NH-tenninal kinase)inhibitor and NAC (N-acetylcysteine) on neuronal apoptosis induced by oxyen-glucose deprivation/regain in vitro. Methods:The oxygen-glucose deprivation/regain (OGD/ R) models were established in vitro. The neurons were exposed to OGD 4h before being placed into normal culture medium. Group A was taken as a blank control. Spinal cord neurons received the JNK inhibitor and NAC treatment before regaining the oxygen-glucose in group B and C, respectively. The neuronal apoptosis were detected by MTT assay in different time points (4 h, 12 h,24 h after regaining oxygen-glucose). Results:The model of OGD/R in spinal cord neurons was successfully established. Cell viability in MTT assay were higher in group B and group C than that in group A, and there was no significant difference between group B and group C. Conclusion:The JNK inhibitor and NAC can effectively protect neurons from apoptosis induced by OGD/R.%目的:初步探讨JNK(The c-Jun NH-terminal kinase)抑制剂及N-乙酰半胱氨酸(N-acetylcysteine,NAC)对脊髓神经细胞氧糖剥夺复糖复氧的保护作用。方法:离体脊髓神经元氧糖剥夺复糖复氧损伤(oxggen-glucose deprivation/regain,OGD/R)模型建立。随机分组:A组对照组(n=6);B组JNK抑制剂组(n=6);C组NAC处理组(n=6)。分别于脊髓神经元氧糖剥夺(oxygen-glucose deprivation,OGD)4 h后,再复氧复糖4、12、24 h,应用MTT法检测细胞凋亡。结果:成功建立OGD/R模型;脊髓神经元OGD/R后,B组和C组MTT各时间点吸光值均高于A组,但B组和C组间差异不显著。结论:JNK抑制剂及NAC能有效抑制体外培养的脊髓神经元氧糖剥夺复糖复氧所引起的凋亡。

  5. Exercise-induced metallothionein expression in human skeletal muscle fibres

    DEFF Research Database (Denmark)

    Penkowa, Milena; Keller, Pernille; Keller, Charlotte;

    2005-01-01

    in both type I and II muscle fibres. This is the first report demonstrating that MT-I + II are significantly induced in human skeletal muscle fibres following exercise. As MT-I + II are antioxidant factors that protect various tissues during pathological conditions, the MT-I + II increases post exercise......Exercise induces free oxygen radicals that cause oxidative stress, and metallothioneins (MTs) are increased in states of oxidative stress and possess anti-apoptotic effects. We therefore studied expression of the antioxidant factors metallothionein I and II (MT-I + II) in muscle biopsies obtained...... in response to 3 h of bicycle exercise performed by healthy men and in resting controls. Both MT-I + II proteins and MT-II mRNA expression increased significantly in both type I and II muscle fibres after exercise. Moreover, 24 h after exercise the levels of MT-II mRNA and MT-I + II proteins were still highly...

  6. Urinary metallothionein as a biological indicator of occupational cadmium exposure

    Energy Technology Data Exchange (ETDEWEB)

    Tohyama, C.; Shaikh, Z.A.; Ellis, K.J.; Cohn, S.H.

    1981-01-01

    Radioimmunoassay and neutron activation data indicate that the urinary metallothionein concentration is related to the liver Cd concentration in occupational Cd exposure. It is also related to the kidney Cd content - but only before the onset of renal dysfunction. Further epidemiological studies are needed to establish a dose-response relationship, which may be useful in minimizing the hazard of Cd-induced renal dysfunction.

  7. Trimethyltin-evoked apoptosis of murine hippocampal granule neurons is accompanied by the expression of interleukin-1beta and interleukin-1 receptor antagonist in cells of ameboid phenotype, the majority of which are NG2-positive.

    Science.gov (United States)

    Fiedorowicz, Anna; Figiel, Izabela; Zaremba, Małgorzata; Dzwonek, Karolina; Schliebs, Reinhard; Oderfeld-Nowak, Barbara

    2008-09-05

    Interleukin-1beta (IL-1beta) has been implicated in various neuropathologies, while IL-1 receptor antagonist (IL-1ra) has been shown to reduce neuronal injury. We investigated the pattern of expression of both cytokines in murine hippocampus after trimethyltin (TMT) intoxication. Using a ribonuclease protection assay, we demonstrated induction of transcription of IL-1beta and IL-1ra 3 days following TMT treatment which correlated with the peak of neuronal apoptosis. At this time, immunocytochemical staining revealed enhanced expression of both cytokines in NG2 proteoglycan expressing ameboid cells located at the site of neurotoxic insult, some of which bound also the microglial marker, lectin. There was some overlap between NG2 and lectin staining. Our results suggest that the two cytokines are involved in apoptotic processes in dentate granule cells and indicate that the pro-apoptotic effect of IL-1beta prevails over the presumed protective action of IL-1ra. The novel finding of expression of both cytokines in NG2(+) cells of ameboid phenotype indicates that these cells, through the regulatory roles of pro- and anti-inflammatory cytokines, may be involved in control of neuronal death or survival after injury.

  8. Assessment of metallothionein and antibodies to metallothionein in normal and autistic children having exposure to vaccine-derived thimerosal.

    Science.gov (United States)

    Singh, Vijendra K; Hanson, Jeff

    2006-06-01

    Allergic autoimmune reaction after exposure to heavy metals such as mercury may play a causal role in autism, a developmental disorder of the central nervous system. As metallothionein (MT) is the primary metal-detoxifying protein in the body, we conducted a study of the MT protein and antibodies to metallothionein (anti-MT) in normal and autistic children whose exposure to mercury was only from thimerosal-containing vaccines. Laboratory analysis by immunoassays revealed that the serum level of MT did not significantly differ between normal and autistic children. Furthermore, autistic children harboured normal levels of anti-MT, including antibodies to isoform MT-I (anti-MT-I) and MT-II (anti-MT-II), without any significant difference between normal and autistic children. Our findings indicate that because autistic children have a normal profile of MT and anti-MT, the mercury-induced autoimmunity to MT may not be implicated in the pathogenesis of autism.

  9. The mechanism of long non-coding RNA MEG3 for neurons apoptosis caused by hypoxia: mediated by miR-181b-12/15-LOX signaling pathway

    Directory of Open Access Journals (Sweden)

    Xiaomin Liu

    2016-09-01

    Full Text Available Objective: lncRNAs are recently thought to play a significant role in cellular homeostasis during pathological process of diseases by competing inhibiting miRNA function. The aim of present study was to assess the function of long non-coding RNA (lncRNA MEG3 and its functional interaction with microRNA-181b in cerebral ischemic infarct of mice and hypoxia-induced neurons apoptosis. Methods: To address this question, we performed the experiments with in vivo middle cerebral artery occlusion (MCAO mice model and in vitro oxygen-glucose deprivation (OGD-cultured neuronal HT22 cell line. Relative expression of MEG3, miR-181b and 12/15-LOX (lipoxygenase mRNA was determined using quantitative RT-PCR. Western blot was used to evaluate 12/15-LOX protein expression. TUNEL assay was performed to assess cell apoptosis.Results: In both MCAO mice and OGD-cultured HT22 cell, ischemia or hypoxia treatment results in a time-dependent increase in MEG3 and 12/15-LOX expression and decrease in miR-181b expression. Knockdown of MEG3 contributes to attenuation of hypoxia-induced apoptosis of HT22 cell. Also, expression level of MEG3 negatively correlated with miR-181b expression and positively correlated with 12/15-LOX expression. In contrary to MEG3, miR-181b overexpression attenuated hypoxia-induced HT22 cell apoptosis, as well as suppressed hypoxia-induced increase in 12/15-LOX expression. By luciferase reporter assay, we concluded that miR-181b directly binds to 12/15-LOX 3’-UTR, thereby negatively regulates 12/15-LOX expression. Conclusion: Our data suggested that long non-coding RNA MEG3 functions as a competing endogenous RNA for miR-181b to regulate 12/15-LOX expression in middle cerebral artery occlusion-induced ischemic infarct of brain nerve cells.

  10. Age-related changes of metallothionein 1/2 and metallothionein 3 expression in rat brain.

    Science.gov (United States)

    Scudiero, Rosaria; Cigliano, Luisa; Verderame, Mariailaria

    2017-01-01

    Neurodegeneration is one of the main physiological consequences of aging on brain. Metallothioneins (MTs), low molecular weight, cysteine-rich proteins that bind heavy-metal ions and oxygen-free radicals, are commonly expressed in various tissues of mammals. MTs are involved in the regulation of cell proliferation and protection, and may be engaged in aging. Expression of the ubiquitous MTs (1 and 2) and the brain specific MT3 have been studied in many neurodegenerative disorders. The research results indicate that MTs may play important, although not yet fully known, roles in brain diseases; in addition, data lack the ability to identify the MT isoforms functionally involved. The aim of this study was to analyse the level of gene expression of selected MT isoforms during brain aging. By using real-time PCR analysis, we determined the MT1/2 and MT3 expression profiles in cerebral cortex and hippocampus of adolescent (2months), adult (4 and 8months), and middle-aged (16months) rats. We show that the relative abundance of all types of MT transcripts changes during aging in both hippocampus and cortex; the first effect is a generalized decrease in the content of MTs transcripts from 2- to 8-months-old rats. After passing middle age, at 16months, we observe a huge increase in MT3 transcripts in both cortical and hippocampal areas, while the MT1/2 mRNA content increases slightly, returning to the levels measured in adolescent rats. These findings demonstrate an age-related expression of the MT3 gene. A possible link between the increasing amount of MT3 in brain aging and its different metal-binding behaviour is discussed.

  11. The retrograde delivery of adenovirus vector carrying the gene for brain-derived neurotrophic factor protects neurons and oligodendrocytes from apoptosis in the chronically compressed spinal cord of twy/twy mice.

    Science.gov (United States)

    Uchida, Kenzo; Nakajima, Hideaki; Hirai, Takayuki; Yayama, Takafumi; Chen, Kebing; Guerrero, Alexander Rodriguez; Johnson, William Eustace; Baba, Hisatoshi

    2012-12-15

    The twy/twy mouse undergoes spontaneous chronic mechanical compression of the spinal cord; this in vivo model system was used to examine the effects of retrograde adenovirus (adenoviral vector [AdV])-mediated brain-derived neurotrophic factor (BDNF) gene delivery to spinal neural cells. To investigate the targeting and potential neuroprotective effect of retrograde AdV-mediated BDNF gene transfection in the chronically compressed spinal cord in terms of prevention of apoptosis of neurons and oligodendrocytes. Several studies have investigated the neuroprotective effects of neurotrophins, including BDNF, in spinal cord injury. However, no report has described the effects of retrograde neurotrophic factor gene delivery in compressed spinal cords, including gene targeting and the potential to prevent neural cell apoptosis. AdV-BDNF or AdV-LacZ (as a control gene) was injected into the bilateral sternomastoid muscles of 18-week old twy/twy mice for retrograde gene delivery via the spinal accessory motor neurons. Heterozygous Institute of Cancer Research mice (+/twy), which do not undergo spontaneous spinal compression, were used as a control for the effects of such compression on gene delivery. The localization and cell specificity of β-galactosidase expression (produced by LacZ gene transfection) and BDNF expression in the spinal cord were examined by coimmunofluorescence staining for neural cell markers (NeuN, neurons; reactive immunology protein, oligodendrocytes; glial fibrillary acidic protein, astrocytes; OX-42, microglia) 4 weeks after gene injection. The possible neuroprotection afforded by retrograde AdV-BDNF gene delivery versus AdV-LacZ-transfected control mice was assessed by scoring the prevalence of apoptotic cells (terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling-positive cells) and immunoreactivity to active caspases -3, -8, and -9, p75, neurofilament 200 kD (NF), and for the oligodendroglial progenitor marker, NG2. RESULTS

  12. Role of Calpain in Apoptosis

    Directory of Open Access Journals (Sweden)

    Hamid Reza Momeni

    2011-01-01

    Full Text Available Apoptosis, a form of programmed cell death that occurs under physiologicalas well as pathological conditions, is characterized by morphological and biochemicalfeatures. While the importance of caspases in apoptosis is established,several noncaspase proteases (Ca2+-dependent proteases such as calpain mayplay a role in the execution of apoptosis. The calpain family consists of twomajor isoforms, calpain I and calpain II which require μM and mM Ca2+ concentrationsto initiate their activity. An increase in intracellular Ca2+ level isthought to trigger a cascade of biochemical processes including calpain activation.Once activated, calpains degrade membrane, cytoplasmic and nuclear substrates,leading to the breakdown of cellular architecture and finally apoptosis.The activation of calpain has been implicated in neuronal apoptosis followingspinal cord injuries and neurodegenerative diseases. This review focuses oncalpain with an emphasis on its key role in the proteolysis of cellular proteinsubstrates following apoptosis.

  13. Sevoflurane post-conditioning protects primary rat cortical neurons against oxygen-glucose deprivation/resuscitation via down-regulation in mitochondrial apoptosis axis of Bid, Bim, Puma-Bax and Bak mediated by Erk1/2.

    Science.gov (United States)

    Zhang, Li-Min; Zhao, Xiao-Chun; Sun, Wen-Bo; Li, Rui; Jiang, Xiao-Jing

    2015-10-15

    Temporal post-conditioning helps provide neuroprotection against brain injury secondary to ischemia-reperfusion and is considered an effective intervention, but the exact mechanism of sevoflurane post-conditioning is unclear. The essential axis involves activator Bid, Bim, Puma (BH3s), Bax, and Bak; activates the mitochondrial death program; and might be involved in a cell death signal. Extracellular signal-related kinases 1/2 (Erk1/2) play a pivotal role in cell growth and proliferation. We hypothesized that sevoflurane post-conditioning might inhibit Bid, Bim, Puma, Bax, and Bak expression and is activated by phosphor-Erk1/2 to decrease neuronal death. To test this hypothesis, we exposed primary cortical neuron cultures to oxygen-glucose deprivation for 1h, along with resuscitation for 24h (OGD/R). MTT assays, propidium iodide uptake (PI), JC-1 fluorescence, and Western blot indicated the following: decreased cell viability (PPuma, Bax, and Bak expression with OGD/R exposure. Inhibition of Erk1/2 phosphorylation could attenuate sevoflurane post-conditioning that mediated an increase in neuronal viability and mitochondrial membrane potential, as well as a decrease in cell death and Bid, Bim, Puma, Bax, and Bak expression after OGD/R treatment. The results demonstrated that sevoflurane post-conditioning caused a marked decrease in cortical neuronal death secondary to OGD/R exposure through the downregulation of the mitochondrial apoptosis axis involving Bid, Bim, Puma, Bax, and Bak that was mediated by the phosphorylation/activation of Erk1/2.

  14. Metallothionein turnover, cytosolic distribution and the uptake of Cd by the green mussel Perna viridis.

    Science.gov (United States)

    Ng, T Y-T; Rainbow, P S; Amiard-Triquet, C; Amiard, J C; Wang, W-X

    2007-08-30

    We examined the relationship between Cd kinetics (uptake from solution and diet, and efflux), metallothionein turnover, and changes in the cytosolic distribution of accumulated Cd between protein fractions in the green mussel Perna viridis. We pre-exposed the mussels to 5, 20, 50 and 200 microg l(-1) of Cd for 1 week and determined the biokinetics of Cd uptake and efflux in the mussels. The dietary assimilation efficiency of Cd increased by 2 times following exposure to 20-200 microg l(-1) Cd, but the dissolved uptake rate was unchanged by pre-exposure to any Cd concentrations. The efflux rate of Cd was also similar among control and Cd pre-exposed mussels. The cytosolic distribution of Cd in the mussels that had been exposed to dissolved Cd, showed that besides metallothionein (7000 - 20,000 Da), high molecular weight proteins (>20,000 Da) were important for Cd binding and depuration. In general, the Cd pre-exposed mussels had higher metallothionein turnover with a higher metallothionein synthesis rate, but similar metallothionein breakdown rates as the control mussels. Metallothionein synthesis rate was correlated to the dietary assimilation of Cd, whereas metallothionein breakdown and Cd efflux rate were independent of each other. This study provides important new information for the role of metallothionein turnover on Cd kinetics in an aquatic invertebrate.

  15. Differential expression of metallothioneins in the CNS of mice with experimental autoimmune encephalomyelitis

    DEFF Research Database (Denmark)

    Espejo, C; Carrasco, J; Hidalgo, J

    2001-01-01

    , and to a lower extent in the brain. Interferon-gamma receptor knockout mice suffered from a more severe experimental autoimmune encephalomyelitis, and interestingly showed a higher metallothioneins-I+II induction in both white and grey matter of the spinal cord and in the brain. In contrast...... during experimental autoimmune encephalomyelitis in interferon-gamma receptor knockout mice with two different genetic backgrounds: 129/Sv and C57BL/6x129/Sv.Mice with experimental autoimmune encephalomyelitis showed a significant induction of metallothioneins-I+II in the spinal cord white matter...... to the metallothioneins-I+II isoforms, metallothionein-III expression remained essentially unaltered during experimental autoimmune encephalomyelitis; interferon-gamma receptor knockout mice showed an altered metallothionein-III expression (a slight increase in the spinal cord white matter) only in the C57BL/6x129/Sv...

  16. One-step purification of metallothionein extracted from two different sources.

    Science.gov (United States)

    Honda, Rubens T; Araújo, Roziete Mendes; Horta, Bruno Brasil; Val, Adalberto L; Demasi, Marilene

    2005-06-25

    We describe a one-step purification of hepatic metallothionein from the Amazon fish Colossoma macropomum injected with cadmium and from the copper-loaded metallothionein from the yeast Saccharomyces cerevisiae, performed by affinity chromatography through metal-chelating columns. Yeast metallothionein was purified from Cu2+-loaded resin and eluted by a continuous EDTA gradient whereas hepatic metallothionein extracted from fishes was purified by Ni2+-loaded resin and eluted by a continuous imidazol gradient. Purified metallothioneins were evaluated by SDS-PAGE and characterized by UV spectra of the apo- and Cd2+-loaded protein. This method allowed high purity and yield as well as rapid one-step extraction of both metal-loaded and apoprotein.

  17. Neuroprotective action of N-acetyl serotonin in oxidative stress-induced apoptosis through the activation of both TrkB/CREB/BDNF pathway and Akt/Nrf2/Antioxidant enzyme in neuronal cells.

    Science.gov (United States)

    Yoo, Jae-Myung; Lee, Bo Dam; Sok, Dai-Eun; Ma, Jin Yuel; Kim, Mee Ree

    2017-04-01

    N-acetyl serotonin (NAS) as a melatonin precursor has neuroprotective actions. Nonetheless, it is not clarified how NAS protects neuronal cells against oxidative stress. Recently, we have reported that N-palmitoyl serotonins possessed properties of antioxidants and neuroprotection. Based on those, we hypothesized that NAS, a N-acyl serotonin, may have similar actions in oxidative stress-induced neuronal cells, and examined the effects of NAS based on in vitro and in vivo tests. NAS dose-dependently inhibited oxidative stress-induced cell death in HT-22 cells. Moreover, NAS suppressed glutamate-induced apoptosis by suppressing expression of AIF, Bax, calpain, cytochrome c and cleaved caspase-3, whereas it enhanced expression of Bcl-2. Additionally, NAS improved phosphorylation of tropomyosin-related kinase receptor B (TrkB) and cAMP response element-binding protein (CREB) as well as expression of brain-derived neurotrophic factor (BDNF), whereas the inclusion of each inhibitor of JNK, p38 or Akt neutralized the neuroprotective effect of NAS, but not that of ERK. Meanwhile, NAS dose-dependently reduced the level of reactive oxygen species, and enhanced the level of glutathione in glutamate-treated HT-22 cells. Moreover, NAS significantly increased expression of heme oxygenase-1, NAD(P)H quinine oxidoreductase-1 and glutamate-cysteine ligase catalytic subunit as well as nuclear translocation of NF-E2-related factor-2. Separately, NAS at 30mg/kg suppressed scopolamine-induced memory impairment and cell death in CA1 and CA3 regions in mice. In conclusion, NAS shows actions of antioxidant and anti-apoptosis by activating TrkB/CREB/BDNF pathway and expression of antioxidant enzymes in oxidative stress-induced neurotoxicity. Therefore, such effects of NAS may provide the information for the application of NAS against neurodegenerative diseases.

  18. Neuroprotective action of N-acetyl serotonin in oxidative stress-induced apoptosis through the activation of both TrkB/CREB/BDNF pathway and Akt/Nrf2/Antioxidant enzyme in neuronal cells

    Directory of Open Access Journals (Sweden)

    Jae-Myung Yoo

    2017-04-01

    Full Text Available N-acetyl serotonin (NAS as a melatonin precursor has neuroprotective actions. Nonetheless, it is not clarified how NAS protects neuronal cells against oxidative stress. Recently, we have reported that N-palmitoyl serotonins possessed properties of antioxidants and neuroprotection. Based on those, we hypothesized that NAS, a N-acyl serotonin, may have similar actions in oxidative stress-induced neuronal cells, and examined the effects of NAS based on in vitro and in vivo tests. NAS dose-dependently inhibited oxidative stress-induced cell death in HT-22 cells. Moreover, NAS suppressed glutamate-induced apoptosis by suppressing expression of AIF, Bax, calpain, cytochrome c and cleaved caspase-3, whereas it enhanced expression of Bcl-2. Additionally, NAS improved phosphorylation of tropomyosin-related kinase receptor B (TrkB and cAMP response element-binding protein (CREB as well as expression of brain-derived neurotrophic factor (BDNF, whereas the inclusion of each inhibitor of JNK, p38 or Akt neutralized the neuroprotective effect of NAS, but not that of ERK. Meanwhile, NAS dose-dependently reduced the level of reactive oxygen species, and enhanced the level of glutathione in glutamate-treated HT-22 cells. Moreover, NAS significantly increased expression of heme oxygenase-1, NAD(PH quinine oxidoreductase-1 and glutamate-cysteine ligase catalytic subunit as well as nuclear translocation of NF-E2-related factor-2. Separately, NAS at 30 mg/kg suppressed scopolamine-induced memory impairment and cell death in CA1 and CA3 regions in mice. In conclusion, NAS shows actions of antioxidant and anti-apoptosis by activating TrkB/CREB/BDNF pathway and expression of antioxidant enzymes in oxidative stress-induced neurotoxicity. Therefore, such effects of NAS may provide the information for the application of NAS against neurodegenerative diseases.

  19. Orexin-A promotes Glu uptake by OX1R/PKCα/ERK1/2/GLT-1 pathway in astrocytes and protects co-cultured astrocytes and neurons against apoptosis in anoxia/hypoglycemic injury in vitro.

    Science.gov (United States)

    Shu, Qing; Zhang, Jianhuai; Ma, Wei; Lei, Youying; Zhou, Dan

    2017-01-01

    Orexin-A, which is an endogenous neuropeptide, is reported to have a protective role in ischemic stroke. High-concentration glutamic acid (Glu) induced by hypoxia injury in ischemic stroke can be inhibited by glial glutamate transporter GLT-1 which is only expressed in astroglia cells. A previous study reported that Orexin-A may regulate GLT-1 expression. However, the role of orexin-A in the regulation of GLT-1 in ischemic stroke still remains unclear. In this study, we aimed to investigate the effect and the underlying mechanism of orexin-A on Glu uptake in astrocytes in vitro and this effect on protecting the neurons from anoxia/hypoglycemic injury. The expression of GLT-1 significantly increased in the astrocytes with orexin-A treatment under anoxia/hypoglycemic conditions, promoting the uptake of Glu and inhibiting the apoptosis of co-cultured cells of astrocytes and neurons. However, these effects were significantly weakened by treatment with orexin-A receptor 1 (OX1R) antagonist. Orexin-A significantly up-regulated the expressions of PKCα and ERK1/2 under anoxia/hypoglycemic conditions in astrocytes, whereas the OX1R antagonist markedly reversed the effect. Furthermore, PKCα or ERK1/2 inhibitor significantly constrained the GLT-1 expression in astrocytes and facilitated the apoptosis of co-cultured cells, and GLT-1 overexpression could reverse those effects of PKCα or ERK1/2 inhibitor. Taken together, orexin-A promoted the GLT-1 expression via OX1R/PKCα/ERK1/2 pathway in astrocytes and protected co-cultured cells against anoxia/hypoglycemic injury.

  20. Phytic Acid Protects against 6-Hydroxydopamine-Induced Dopaminergic Neuron Apoptosis in Normal and Iron Excess Conditions in a Cell Culture Model.

    Science.gov (United States)

    Xu, Qi; Kanthasamy, Anumantha G; Reddy, Manju B

    2011-02-07

    Iron may play an important role in Parkinson's disease (PD) since it can induce oxidative stress-dependent neurodegeneration. The objective of this study was to determine whether the iron chelator, phytic acid (IP6) can protect against 6-hydroxydopamine- (6-OHDA-) induced apoptosis in immortalized rat mesencephalic dopaminergic cells under normal and iron-excess conditions. Caspase-3 activity was increased about 6-fold after 6-OHDA treatment (compared to control; P IP6 pretreatment decreased it by 38% (P IP6 pretreatment. Under iron-excess condition, a 6-fold increase in caspase-3 activity (P IP6. Together, our data suggest that IP6 protects against 6-OHDA-induced cell apoptosis in both normal and iron-excess conditions, and IP6 may offer neuroprotection in PD.

  1. Shaping mechanisms of metal specificity in a family of metazoan metallothioneins: evolutionary differentiation of mollusc metallothioneins

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    Atrian Sílvia

    2011-01-01

    Full Text Available Abstract Background The degree of metal binding specificity in metalloproteins such as metallothioneins (MTs can be crucial for their functional accuracy. Unlike most other animal species, pulmonate molluscs possess homometallic MT isoforms loaded with Cu+ or Cd2+. They have, so far, been obtained as native metal-MT complexes from snail tissues, where they are involved in the metabolism of the metal ion species bound to the respective isoform. However, it has not as yet been discerned if their specific metal occupation is the result of a rigid control of metal availability, or isoform expression programming in the hosting tissues or of structural differences of the respective peptides determining the coordinative options for the different metal ions. In this study, the Roman snail (Helix pomatia Cu-loaded and Cd-loaded isoforms (HpCuMT and HpCdMT were used as model molecules in order to elucidate the biochemical and evolutionary mechanisms permitting pulmonate MTs to achieve specificity for their cognate metal ion. Results HpCuMT and HpCdMT were recombinantly synthesized in the presence of Cd2+, Zn2+ or Cu2+ and corresponding metal complexes analysed by electrospray mass spectrometry and circular dichroism (CD and ultra violet-visible (UV-Vis spectrophotometry. Both MT isoforms were only able to form unique, homometallic and stable complexes (Cd6-HpCdMT and Cu12-HpCuMT with their cognate metal ions. Yeast complementation assays demonstrated that the two isoforms assumed metal-specific functions, in agreement with their binding preferences, in heterologous eukaryotic environments. In the snail organism, the functional metal specificity of HpCdMT and HpCuMT was contributed by metal-specific transcription programming and cell-specific expression. Sequence elucidation and phylogenetic analysis of MT isoforms from a number of snail species revealed that they possess an unspecific and two metal-specific MT isoforms, whose metal specificity was

  2. Role of Metallothionein1H in Cisplatin Resistance of Non-Small Cell Lung Cancer Cells

    Institute of Scientific and Technical Information of China (English)

    Xin-fang Hou; Qing-xia Fan; Liu-xing Wang; Shi-xin Lu

    2009-01-01

    Objective: Despite platinum-based adjuvant chemotherapy has improved greatly patients' outcomes, drug resistance poses a major impediment to the successful use of such an effective agent. Metallothioneins(MTs) are known to play putative roles in cancer cell proliferation, apoptosis, differentiation, drug resistance and prognosis. The present studiy was to investigte the role of metallethioein1H(MT1H) in cisplatin resistance of human non-small cell lung cancer(NSCLC) cell lines in vitro or its possible molecular mechanisms. Methods: MT1H mRNA expression in A549 and A549/DDP cells was detected by RT-PCR. A recombinant eukaryotic expression plasmid pcDNA3.1(-)-MT1H was constructed and transfected into A549 cells which express no MT1H. MT1H siRNA was transfected into A549/DDP cells which express MT1H highly. MT1H expression was detected by RT-PCR and Immunoblot. The chemosensitivity to cisplatin was assessed by MTT assay. Apoptosis rate was determined by Tunel and FCM. Bcl-2 and Bax were determined by immunohistochemistry. Results: MT1H mRNA was expressed in A549/DDP but not in A549. After transfection of MT1H, MT1H expression was enhanced and the chemosensitivity to cisplatin was decreased in A549 cells. Inversely, after transfection of MT1H siRNA, MT1H expression was decreased and the chemosensitivity to cisplatin was increased in A549/DDP. The apoptosis rate induced by cisplatin was increased and Bcl-2 was down-regulated but Bax showed little change in A549/DDP cells interferred with MT1H siRNA. Conclusion: MT1H overexpression can promote drug resistance in A549 cells . Down-regulation of MT1H interfered with siRNA can effectively reverses the drug resistance in A549/DDP cells by down-regulating the expression of Bcl-2 and increasing cisplatin induced apoptosis. SiRNA targeting MT1H combined with chemotherapy may be a very promising strategy for treatment of lung cancer.

  3. Tetramethylpyrazine Analogue CXC195 Protects Against Dopaminergic Neuronal Apoptosis via Activation of PI3K/Akt/GSK3β Signaling Pathway in 6-OHDA-Induced Parkinson's Disease Mice.

    Science.gov (United States)

    Chen, Lin; Cheng, Li; Wei, Xinbing; Yuan, Zheng; Wu, Yanmei; Wang, Shuaishuai; Ren, Zhiping; Liu, Xinyong; Liu, Huiqing

    2016-12-22

    Parkinson's disease (PD) is a progressive neurodegenerative disorder and characterized by motor system disorders resulting in loss of dopaminergic (DA) neurons. CXC195, a novel tetramethylpyrazine derivative, has been shown strongest neuroprotective effects due to its anti-apoptotic activity. However, whether CXC195 protects against DA neuronal damage in PD and the mechanisms underlying its beneficial effects are unknown. The purpose of our study was to investigate the potential neuroprotective role of CXC195 and to elucidate its mechanism of action against 6-hydroxydopamine (6-OHDA)-induced mouse model of PD. CXC195 administration improved DA neurodegeneration in PD mice induced by 6-OHDA. Our further findings confirmed treatment of CXC195 at the dose of 10 mg/kg significantly inhibited the apoptosis by decreasing the level of cleaved caspase-3 and Bax, and increasing the level of Bcl-2 in 6-OHDA-lesioned mice. Meanwhile, 6-OHDA also decreased the amount of phosphorylated Akt while increased GSK-3β activity (the amount of phosphorylated GSK-3β at Ser9 was decreased) which was prevented by CXC195. Wortmannin, a specific PI3K inhibitor, dramatically abolished the changes induced by CXC195. Our study firstly demonstrated that CXC195 protected against DA neurodegeneration in 6-OHDA-induced PD model by its anti-apoptotic properties and PI3K/Akt/GSK3β signaling pathway was involved in it.

  4. Metallothionein expression in placental tissue in Menkes' disease

    DEFF Research Database (Denmark)

    Hærslev, T.; Krag Jacobsen, G.; Horn, N.

    1995-01-01

    Menkes' disease is a recessive X-linked disturbance of copper metabolism, resulting in accumulation of copper in several extra-hepatic tissues including the placenta. Metallothionein (MT) is a low-molecular weight protein with a high affinity for group II metal ions, such as copper. Its synthesis....... The avidin-biotin-complex (ABC)-technique was used. The copper content was measured by neutron activation analysis (NAA). In all placental tissue sections positive MT immunostaining appeared only in the trophoblast and only in proliferating cells. In placental tissue sections obtained from foetuses...

  5. Promiscuity and preferences of metallothioneins: the cell rules

    Directory of Open Access Journals (Sweden)

    Robinson Nigel J

    2011-04-01

    Full Text Available Abstract Metalloproteins are essential for many cellular functions, but it has not been clear how they distinguish between the different metals to bind the correct ones. A report in BMC Biology finds that preferences of two metallothionein isoforms for two different cations are due to inherent properties of these usually less discriminating proteins. Here these observations are discussed in the context of the cellular mechanisms that regulate metal binding to proteins. See research article: http://www.biomedcentral.com/1741-7007/9/4

  6. Phytic Acid Protects against 6-Hydroxydopamine-Induced Dopaminergic Neuron Apoptosis in Normal and Iron Excess Conditions in a Cell Culture Model

    Directory of Open Access Journals (Sweden)

    Qi Xu

    2011-01-01

    Full Text Available Iron may play an important role in Parkinson's disease (PD since it can induce oxidative stress-dependent neurodegeneration. The objective of this study was to determine whether the iron chelator, phytic acid (IP6 can protect against 6-hydroxydopamine- (6-OHDA- induced apoptosis in immortalized rat mesencephalic dopaminergic cells under normal and iron-excess conditions. Caspase-3 activity was increased about 6-fold after 6-OHDA treatment (compared to control; <.001 and 30 μmol/L IP6 pretreatment decreased it by 38% (<.05. Similarly, a 63% protection (<.001 against 6-OHDA induced DNA fragmentation was observed with IP6 pretreatment. Under iron-excess condition, a 6-fold increase in caspase-3 activity (<.001 and a 42% increase in DNA fragmentation (<.05 with 6-OHDA treatment were decreased by 41% (<.01 and 27% (<.05, respectively, with 30 μmol/L IP6. Together, our data suggest that IP6 protects against 6-OHDA-induced cell apoptosis in both normal and iron-excess conditions, and IP6 may offer neuroprotection in PD.

  7. Metallothionein metabolism in the streptozotocin-diabetic rat

    Energy Technology Data Exchange (ETDEWEB)

    Chen, M.L.; Failla, M.L.

    1986-03-05

    Earlier reports from their laboratory showed the induction of the insulin-deficient diabetic state in adult rats was associated with an accumulation of zinc, copper, and a metallothionein-like zinc and copper binding protein in the soluble fraction of liver and kidney. Based upon chromatographic and electrophoretic properties, -SH to metal ratio and amino acid composition, they now report that elevated concentrations of metallothioneins (MT)-I and -II are indeed present in diabetic rat liver and kidney cytosol. The relative rates of MT synthesis in tissues from diabetic and control rats were measured by comparing incorporation of /sup 35/S-cysteine into MT vs. total cytoplasmic proteins at 5 h after injection of the precursor. The relative rates of MT synthesis in livers from rats diabetic for 10 d and fed either chow or purified diet containing 13 or 35 ppm copper were 1.4, 2.3 and 2.8 times greater, respectively, than control rats fed the same diets. Higher relative rates of MT synthesis were also observed in kidneys from diabetic rats fed purified diets compared to controls. Maximal relative rates of MT synthesis in diabetic liver and kidney were observed at 4 and 10 d, respectively, after onset of diabetes. The half-lives of cytoplasmic MT in liver and kidney from diabetic (10 d) rats were 1.3 and 2.6 days, respectively; half-lives of MT in control liver and kidney were 5.0 and 2.1 days, respectively.

  8. Effects of nitric oxide on spontaneous pain reaction and neuronal apoptosis in the spinal cord of rats induced by formalin inflammatory pain%NOS抑制剂对甲醛炎性痛诱导的大鼠痛反应及脊髓神经元凋亡的影响

    Institute of Scientific and Technical Information of China (English)

    褚赛纯; 胡玉燕; 李清君; 李慧娜; 李文斌

    2011-01-01

    Objective: To observe whether formalin inflammatory pain can induce neuron apoptosis in rats spinal cord or not and the effects of nitric oxide on the spontaneous pain reaction and neuron apoptosis in the spinal cord of rats with formalin inflammatory pain. Methods: Formalin-induced paw licking time was used to reflect the degree of spontaneous pain of rats, and the flow cytometry was used to detecte neuron apoptosis rate of spinal cord. Results: Compared with control group, the apoptosis ratio of spinal neuron was increased in the rats with formalin inflammatory pain, and peaked at 3d after formalin injection. Pre-intrathecal injection of NOS inhibitor L-NAME inhibited the nocicepu've behavioural response in double phases induced by formalin injection and cut down the neuron apoptosis ratio of spinal cord of rats with formalin inflammatory pain. Nocicepu've behavioural response and incraesed neuron apoptosis in the spinal cord were induced by intrathecal injection of L-Arg in normal rats. Conclusion: The results indicated that formalin inflammatory pain could induce the apoptosis of spinal neurons. The neurons apoptosis was the most significant on the third day after formalin injection. The increased pruduction of NO in spinal cord could promote the transmit of nociceptive information and participate the induction of neuronal apoptosis during the formalin inflammatory pain.%目的:观察甲醛炎性痛是否可诱导脊髓神经元凋亡以及一氧化氮(NO)对甲醛炎性痛诱导的大鼠痛反应以及脊髓神经元凋亡的影响.方法:采用行为学方法观察大鼠自发痛反应,流式细胞术检测脊髓神经元凋亡率.结果:与正常大鼠比较,甲醛炎性痛可诱导大鼠脊髓神经元凋亡率明显增加,于注射甲醛后3d时最为明显.预先鞘内注射NOS抑制剂L-NAME可剂量依赖性抑制足底注射甲醛诱导的大鼠第一相和第二相痛反应,并可剂量依赖性抑制足底注射甲醛诱导的脊髓神经元凋亡

  9. Effect of L-arginine on neuronal apoptosis induced by focal cerebral ischemia in rats%L-精氨酸对局灶性脑缺血大鼠神经元凋亡的影响

    Institute of Scientific and Technical Information of China (English)

    张建新; 李兰芳; 李永辉; 张会欣

    2008-01-01

    Objective To investigate the effect of L-arginine(L-arg)on neuronal apoptosis induced by focal cerebral ischemia in rats.Methods Fifly-six male SD rats,weighing 250-300 g,were randomly divided into 7 groups(n=8 each):sham operation group(SH),2 h cerebral ischemia group(IS1),2 h cerebral ischemia and L-arg treatment group(L-arg1),6 h cerebral ischemia group(IS2),6 h cerebral ischemia and L-arg treatment group(L-arg2),12 h cerebral ischemia group(IS3),and 12 h cerebral ischemia and L-arg treatment group (L-arg3).Focal cerebral ischemia was produced by middle cerebral artery occlusion for 12 h.A nylon thread with rounded tip was inserted into left internal carotid artery cranial until resistance was met.The distance from bifurcation of common carotid artery to the tip of the thread was about 18-19 mm.Focal cerebral ischemia was confirmed by left Horner's syndrome and right side hemiplegia.In each L-arg group,L-arg 500 mg/kg was given intraperitoneally after cerebral ischemia twice a day for 3 days.In IS group,normal saline was given instead of L-arg.The animals were decapitated 3 days after treatment.The brains were immediately removed for the determination of the neuronal apoptosis rate in the ischemic region and the expression of Caspase-3,Bcl-2 and Bax protein.Results The neuronal apoptosis rate and the expression of Caspase-3 and Bax protein were significantly higher,while the ratio of Bcl-2/Bax protein was significantly lower in IS1,IS2 and IS3 group than in SH group(P <0.01).The neuronal apoptosis rate and the expression of Caspase-3 and Bax protein were significantly lower,while the expression of Bcl-2 protein and the ratio of Bcl-2/Bax protein were significantly higher in L-arg1 and L-arg2 group than in IS1 and IS2 group(P<0.01).There was no significant difference in the indexes mentioned above between IS3 and L-arg3 group(P>0.05).Conclusion L-arg can reduce the neuronal apoptosis and has certain therapeutic effect during the early cerebral ischemia

  10. Protective/detoxicative function of metallothionein in the rat brain and blood induced by controlled cadmium doses

    Directory of Open Access Journals (Sweden)

    H. N. Shiyntum

    2015-09-01

    Full Text Available Cadmiumclassified as a major carcinogen is considered a poisonous and unwanted heavy metal to a lot of tissues in many organisms. Of many publications already available, the general consensus is that the cadmium attenuating element is metallothionein (MT through its interchangeable mechanism with Zn triggered by the presence of Cd, providing binding sites for Cd ions. MT was first discovered in the kidney cortex of the horse; it represents a low molecular weight protein, rich in cysteine residues which effectively bind with metals. Its functions consist in detoxification of heavy metals like mercury, arsenic, cadmium, homeostasis of essential metals including copper and zinc, anti-oxidation against reactive oxygen species, protection against DNA damage, oxidative stress, cell survival, angiogenesis, apoptosis, and increase of proliferation. In this work, we sought to highlight the protective function of MT in the brain and serum of rats by means of detoxification under induced effects of controlled Cd doses. We have done this by exposing Wistar rats to Cd at different doses in drinking water at different time intervals. In two independent experiments, 58 rats were subjected to 0.1 or 1.0 µg Cd2+/kg of body weight for 15 or 36 days under different conditions. The obtained data indicates the different functioning systems for the brain and the blood for MT metabolism under Cd effect. Our results indicate significant loss of metallothionein level in the brain and important increases in the amount of MT in serum proving that even minimal ingestion of toxic Cd is enough to trigger the release of MT protein in blood.

  11. Pinocembrin protects against β-amyloid-induced toxicity in neurons through inhibiting receptor for advanced glycation end products (RAGE-independent signaling pathways and regulating mitochondrion-mediated apoptosis

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    Liu Rui

    2012-09-01

    depressed the activation of p38 mitogen-activated protein kinase (MAPK-MAPKAP kinase-2 (MK2-heat shock protein 27 (HSP27 and stress-activated protein kinase (SAPK/c-Jun N-terminal kinase (JNK-c-Jun pathways and the downstream nuclear factor κB (NFκB inflammatory response subsequent to Aβ-RAGE interaction. In addition, pinocembrin significantly alleviated mitochondrial dysfunction through improving mitochondrial membrane potential and inhibiting mitochondrial oxidative stress, and regulated mitochondrion-mediated apoptosis by restoration of B cell lymphoma 2 (Bcl-2 and cytochrome c and inactivation of caspase 3 and caspase 9. Conclusions Pinocembrin was shown to infer cognitive improvement and neuronal protection in AD models. The mechanisms of action of the compound were illustrated on RAGE-dependent transduction inhibition and mitochondrion protection. It appears to be a promising candidate for the prevention and therapy of AD.

  12. Pinocembrin protects against β-amyloid-induced toxicity in neurons through inhibiting receptor for advanced glycation end products (RAGE)-independent signaling pathways and regulating mitochondrion-mediated apoptosis.

    Science.gov (United States)

    Liu, Rui; Wu, Cai-Xia; Zhou, Dan; Yang, Fan; Tian, Shuo; Zhang, Li; Zhang, Tian-Tai; Du, Guan-Hua

    2012-09-18

    -activated protein kinase (MAPK)-MAPKAP kinase-2 (MK2)-heat shock protein 27 (HSP27) and stress-activated protein kinase (SAPK)/c-Jun N-terminal kinase (JNK)-c-Jun pathways and the downstream nuclear factor κB (NFκB) inflammatory response subsequent to Aβ-RAGE interaction. In addition, pinocembrin significantly alleviated mitochondrial dysfunction through improving mitochondrial membrane potential and inhibiting mitochondrial oxidative stress, and regulated mitochondrion-mediated apoptosis by restoration of B cell lymphoma 2 (Bcl-2) and cytochrome c and inactivation of caspase 3 and caspase 9. Pinocembrin was shown to infer cognitive improvement and neuronal protection in AD models. The mechanisms of action of the compound were illustrated on RAGE-dependent transduction inhibition and mitochondrion protection. It appears to be a promising candidate for the prevention and therapy of AD.

  13. 灵芝孢子粉对杏仁核点燃模型大鼠神经元凋亡的影响%The effect of Reishi Mushroom Powder on neuronal apoptosis of the amygdala kindling model

    Institute of Scientific and Technical Information of China (English)

    张继国; 王艺; 张静; 邱彦龙

    2012-01-01

    Purpose To observe the effect of Reishi Mushroom Powder on neuronal apoptosis of the a-mygdala kindling model. Methods 24 male amygdala kindling rats were divided into Reishi Mushroom Powder (treatment group) , saline ( negative control) , and phenobarbital gavage ( positive control). After 2 weeks treatment, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay (TUNEL) was used to collect apoptotic cells of the hippocampus. Results The number of apoptotic cells in the treatment group, negative control group and the positive control group were (21. 25 ± 4. 03 ) , (42.08 ±4.29) and (23. 17 ±3. 73)/mm2 .respectively. Compared with the negative control group,apoptotic cells in the treatment group and the positive control group were significantly lower(P<0.05). No significant difference was seen between treatment group and the positive control group (P>0.05). Conclusion The Reishi Mushroom Powder can significantly reduce epilepsy-induced neuronal apoptosis and has protection function on the nervous system.%目的 观察灵芝孢子粉治疗后杏仁核点燃模型大鼠神经元凋亡状况.方法 雄性大鼠24只,制备杏仁核点燃模型后,分为灵芝孢子粉组、生理盐水(阴性对照)组、苯巴比妥(阳性对照)组,灌胃给药2周后,采用原位杂交末端标记(TUNEL)法检测大鼠脑海马组织细胞凋亡状况.结果 灵芝孢子粉组、阴性对照组及阳性对照组的凋亡细胞数分别为(21.25±4.03),(42.08±4.29)和(23.17±3.73)个/mm2,灵芝孢子粉组和阳性对照组凋亡指教均明显低于阴性对照组(P<0.05),给药组与阳性对照组之问无明显差异(P>0.05).结论 灵芝孢子粉能够明显减少癫痫引起的神经元凋亡,对神经系统具有保护功能.

  14. The effect of caffeine on the primarily cultured cortical neuron apoptosis in neonate mice%咖啡因对乳鼠脑皮质神经元凋亡的作用

    Institute of Scientific and Technical Information of China (English)

    汪岩; 卢延旭; 耿广军; 喻洪江

    2009-01-01

    Objective To examine the effect of Caffeine on the cultured cortical neuron apoptosis in neonatal rats.Methods The primary cerebral cortex neurons for cultures were obtained from neonatal mice 2-3 days after birth,Caffeine reconstituted at final concentrations 300μmol/L and 1 000μmol/L was added to the cell cultures and continuously co-incubated for 6-36 h,respectively after the cortical neurons were continuously cultivated 7 days after incubation under temperature of 37℃ incubator with 5% CO_2 and 100% relative humidity,the intracellular calcium concentration,mitochondrial membrane potential and apoptosis rate were determined by the flow cytometry.The activity of Caspase-9 was assayed by enzyme-labeled instrument,and Caspase-9 activity by the enzyme-1inked analyzer.Cell morphological changes were observed under electron microscope and fluorescent microscope after being stained with Hoechst 33258 fluorescent dye.Results Compared with the control group,the average increase in intracellular calcium fluorescence intensity was most significant(P<0.05),which elevated from the normal value 43.13±2.02 to 45.28±1.16 and 46.92±1.99,respectively at 6 h;mitochondrial membrane potentials were reduced most significandy(P<0.05).from the base value 443.58 ±11.77 down to 289.53±16.47 and 165.14±14.72,respectively at 8h.Caspase-9 activity was peaked(P<0.05),from the normal value 1.00±0.000 to 5.33±1.02 and 8.33±0.92,respectively at 10 h.The neuronal apoptosis ratio was increased significantly (P<0.05),from the normal value 4.94±1.74 to 15.98±2.03 and 18.70±2.09,at 36h.The apoptotic bodies were observed at 24 h after administration of 300 μmol/L and 1000 μmol/L Caffeine.Conclusion Caffeine may promote neuronal apoptosis in neonate mice.%目的 考察咖啡因对乳鼠脑皮质神经元凋亡的作用.方法 取出生后2~3d的乳鼠脑皮质神经元,在37℃、5%CO_2、100%相对湿度的培养箱中培养7d后,分别加入终浓度为300μmol/L和1 000μmol/L

  15. 逍遥散抗慢性心理应激海马神经元凋亡的机理研究%Mechanism of Xiaoyao Powder in Counteracting Apoptosis of Hippocampus Neurons Caused by Chronic Psychological Stress

    Institute of Scientific and Technical Information of China (English)

    王文竹; 杨忠华; 徐志伟; 苏俊芳

    2011-01-01

    Objective To investigate the effect of Xiaoyao Powder(XP) on apoptosis of hippocampus neurons and glucocorticoid receptor(GR) mRNA expression caused by chronic psychological stress. Methods Wistar rats were randomized to 3 groups: normal group(without any stimulation,gastric gavage of 2 mL normal saline per day), model group( chronic mild unpredictable stress, gastric gavage of 2 mL normal saline per day) and XP group(gastric gavage of 2 mL XP one hour before stress). The changes of body weight and sucrose preference were observed. The apoptotic rate of hippocampal neurons were detected by PI staining method and GR mRNA expression by in-situ hybridization.Results After treatment with chronic mild unpredictable stress for 21 days, the body weight as well as the weight gain was lower in the model group and XP group than that in the normal group(P < 0.01 ), and the sucrose preference was decreased in the model group (P < 0.01 ). In the model group, the apoptotic rate of hippocampal neurons was increased(P < 0.05), and GR mRNA expression was decreased(P < 0.01 ). XP decreased the apoptotic rate (P < 0.05) and increased GR mRNA expression(P < 0.01 ). Conclusion Chronic mild unpredictable stress can significantly decrease sucrose preference. XP can counteract the decrease of sucrose preference and significantly improve the apoptosis of hippocampus neurons caused by chronic psychological stress, which mechanism maybe related to the regulation of GR mRNA expression.%目的 观察逍遥散抗慢性心理应激海马神经元凋亡及糖皮质激素受体(GR)表达的影响.方法 Wistar 大鼠按体重分为正常组、模型组、逍遥散组.正常组及模型组灌胃等量生理盐水,逍遥散组给予逍遥散7.02g/kg.观察实验前后大鼠体重及糖水偏爱度的变化,采用PI法检测大鼠海马神经元凋亡,原位杂交法检测GR mrRNA表达.结果 经过21 d慢性轻度不可预计应激(CMUS)造模后,模型组、逍遥散组体重及

  16. Metallothioneins 1 and 2 Modulate Inflammation and Support Remodeling in Ischemic Cardiomyopathy in Mice

    Science.gov (United States)

    Dewald, Daniela; Schmitz, Eva J.; Verfuerth, Luise; Keppel, Katharina; Peigney, Christine; Ghanem, Alexander; Welz, Armin; Dewald, Oliver

    2016-01-01

    Aims. Repetitive brief ischemia and reperfusion (I/R) is associated with left ventricular dysfunction during development of ischemic cardiomyopathy. We investigated the role of zinc-donor proteins metallothionein MT1 and MT2 in a closed-chest murine model of I/R. Methods. Daily 15-minute LAD-occlusion was performed for 1, 3, and 7 days in SV129 (WT)- and MT1/2 knockout (MT−/−)-mice (n = 8–10/group). Hearts were examined with M-mode echocardiography and processed for histological and mRNA studies. Results. Expression of MT1/2 mRNA was transiently induced during repetitive I/R in WT-mice, accompanied by a transient inflammation, leading to interstitial fibrosis with left ventricular dysfunction without infarction. In contrast, MT−/−-hearts presented with enhanced apoptosis and small infarctions leading to impaired global and regional pump function. Molecular analysis revealed maladaptation of myosin heavy chain isoforms and antioxidative enzymes in MT1/2−/−-hearts. Despite their postponed chemokine induction we found a higher total neutrophil density and macrophage infiltration in small infarctions in MT−/−-hearts. Subsequently, higher expression of osteopontin 1 and tenascin C was associated with increased myofibroblast density resulting in predominately nonreversible fibrosis and adverse remodeling in MT1/2−/−-hearts. Conclusion. Cardioprotective effects of MT1/2 seem to be exerted via modulation of contractile elements, antioxidative enzymes, inflammatory response, and myocardial remodeling. PMID:27403038

  17. Autophagy of metallothioneins prevents TNF-induced oxidative stress and toxicity in hepatoma cells.

    Science.gov (United States)

    Ullio, Chiara; Brunk, Ulf T; Urani, Chiara; Melchioretto, Pasquale; Bonelli, Gabriella; Baccino, Francesco M; Autelli, Riccardo

    2015-01-01

    Lysosomal membrane permeabilization (LMP) induced by oxidative stress has recently emerged as a prominent mechanism behind TNF cytotoxicity. This pathway relies on diffusion of hydrogen peroxide into lysosomes containing redox-active iron, accumulated by breakdown of iron-containing proteins and subcellular organelles. Upon oxidative lysosomal damage, LMP allows relocation to the cytoplasm of low mass iron and acidic hydrolases that contribute to DNA and mitochondrial damage, resulting in death by apoptosis or necrosis. Here we investigate the role of lysosomes and free iron in death of HTC cells, a rat hepatoma line, exposed to TNF following metallothionein (MT) upregulation. Iron-binding MT does not normally occur in HTC cells in significant amounts. Intracellular iron chelation attenuates TNF and cycloheximide (CHX)-induced LMP and cell death, demonstrating the critical role of this transition metal in mediating cytokine lethality. MT upregulation, combined with starvation-activated MT autophagy almost completely suppresses TNF and CHX toxicity, while impairment of both autophagy and MT upregulation by silencing of Atg7, and Mt1a and/or Mt2a, respectively, abrogates protection. Interestingly, MT upregulation by itself has little effect, while stimulated autophagy alone depresses cytokine toxicity to some degree. These results provide evidence that intralysosomal iron-catalyzed redox reactions play a key role in TNF and CHX-induced LMP and toxicity. The finding that chelation of intralysosomal iron achieved by autophagic delivery of MT, and to some degree probably of other iron-binding proteins as well, into the lysosomal compartment is highly protective provides a putative mechanism to explain autophagy-related suppression of death by TNF and CHX.

  18. Pengaruh stresor fisik terhadap distribusi SERT dan indeks apoptosis neuron hipokampus, serta distribusi TNF-± gaster tikus dengan mediasi kortisol dan IL-6

    Directory of Open Access Journals (Sweden)

    Suparno Suparno

    2012-02-01

    Full Text Available Generally, distres which followed by psychiatric disorders still more medicinal practice controversial especially physically stressor roles on holistic health condition. Therefore, researches still more needed to explain and clarify the infl uences and the role of physically stressors on psychiatric disorders. This study used an psychoneuroimmunologic paradigm, which used posttest-only true experiment design for the research. About 24 female Rattus norvegicus strain Wistar had been used, they were devided into six groups. To investigate functional changes, we had been used hippocampus SERT distribution changes as mirror of central serotonergic malfunction changes. While, to investigate structural changes we had been inspected apoptosis index of hippocampus CA3 regions (hippocampus slicing. We also took the blood from the rats heart to examinate IL – 6 and cortisol level, and gastric tissues for TNF-α level examination. Physical stressors lead to arise functional and structural changes include hippocampus SERT distribution, increases hippocampus apoptotic index, and increases TNF-α level, that fi nally arise the distress. These results of statistical analyses demonstrate that physical stressors tend to be mediated by serotonergic neurotransmission pathway. This study also support assumptions that psychological stressors caused and or related with gastrical changes.

  19. Metallothionein Induction by Cd(Ⅱ) and Zn(Ⅱ)

    Institute of Scientific and Technical Information of China (English)

    Xiao Yan ZHANG; Xin GAO; Yan LI; Xiao Dong FAN

    2006-01-01

    The induction of MT(metallothionein) by Cd(Ⅱ) and Zn(Ⅱ) in renal proximal tubule cells (LLC-PKI) and the effects of simultaneous administration of Cd(Ⅱ) and Zn(Ⅱ) on MT induction were discussed. The two metals administered together are able to induce MT to almost 40 μg MT/mg protein. The additive production of MT may be due to two different effects by Cd(Ⅱ) and Zn(Ⅱ) on the activation of MT transcription. The experiments of simultaneous administration of Cd(Ⅱ) and Zn(Ⅱ) to the renal proximal tubule cells demonstrated the interaction of these metals, MT induction and proximal tubule toxicity.

  20. The involvement of metallothionein in the development of aquatic invertebrate

    Energy Technology Data Exchange (ETDEWEB)

    Mao Huan; Wang Dahui [Sperm Laboratory, College of Life Sciences, Zhejiang University, Hangzhou 310058 (China); Yang Wanxi, E-mail: wxyang@spermlab.org [Sperm Laboratory, College of Life Sciences, Zhejiang University, Hangzhou 310058 (China)

    2012-04-15

    The many documents on metallothioneins (MTs) in aquatic organisms focus especially on their use as biomarkers in environmental monitoring programs, but there are a few papers that summarize the physiological role of MTs in aquatic organisms especially in their development. The multifaceted role of MTs include involvement in homeostasis, protection against heavy metals and oxidant damage, metabolic regulation, sequestration and/or redox control. MTs could be induced by heavy metals which are able to hinder gametogenesis, suppress embryogenesis, and hamper development. Here we pay more attention on the non-essential metal cadmium, which is the most studied heavy metal regarding MTs, and its effects on the development of aquatic invertebrates. In this paper, we have collected published information on MTs in aquatic organisms - mollusks, crustaceans, etc., and summarize its functions in aquatic invertebrates, especially those related to their development.

  1. Determination of the cadmium and copper content inherent to metallothionein

    Energy Technology Data Exchange (ETDEWEB)

    Raspor, B.; Kozar, S.; Pavicic, J.; Juric, D. [Ruder Boskovic Institute, Center for Marine Research Zagreb, P.O.B. 1016, HR-10 001 Zagreb (Croatia)

    1998-05-01

    The reliability of the voltammetric determination of the cadmium and copper content (at pH 1.0), inherent to metallothionein (MT) isolated from the digestive gland of Mytilus galloprovincialis, was investigated. An artifact signal enhancement of copper, caused by the cupric-thionein complex adsorption at the mercury electrode, was established. This artifact was removed by UV-digestion of the sample for 15-20 h prior to analysis. A similar artifact was not detected for cadmium, because at this pH the cadmium-thionein complex has dissociated, and cadmium exists in the ionic form. Therefore, the voltammetric analysis of the cadmium content can be performed directly at pH 1.0, without prior UV-digestion of the sample. (orig.) With 3 figs., 1 tab., 12 refs.

  2. Separation of two forms of rabbit metallothionein by isoelectric focusing.

    Science.gov (United States)

    Nordberg, G F; Nordberg, M; Piscator, M; Vesterberg, O

    1972-02-01

    Rabbits were given repeated injections of cadmium chloride. Cadmium- and zinc-containing protein fractions were obtained from the livers of these animals by precipitation procedures and Sephadex G-75 chromatography. The protein thus obtained showed several characteristics similar to those of the earlier described protein metallothionein. Further separation by isoelectric focusing showed two main protein peaks with isoelectric points at 3.9 and 4.5 respectively. Amino acid analysis of these two forms showed similar content of most amino acids [residues per cent.: cysteine (28%), aspartate (8%), threonine (5-6%), serine (12%), glycine (7%), alanine (13%), methionine (2%), isoleucine (2%)] but with a small difference in content of lysine (12 and 13% respectively), proline (9 and 5% respectively) and glutamate (2 and 4% respectively). The two forms of the protein both contained cadmium, but only the one with pI4.5 contained also significant amounts of zinc.

  3. Temporal variations in metallothionein concentration and subcellular distribution of metals in gills and digestive glands of the oyster Crassostrea angulata

    Directory of Open Access Journals (Sweden)

    Chiara Trombini

    2010-11-01

    Full Text Available The metallothionein levels and metal concentrations in whole body, digestive gland and gills of Crassostrea angulata were analyzed in field samples collected from the River Guadalquivir estuary over several years following a mining waste spill upstream. The subcellular distribution of metals was analyzed to determine the mechanisms involved in the detoxification process. The highest metallothionein levels were reported in the digestive gland shortly after the mining contamination event. In this organ, metals are stored preferentially in the non-cytosolic fraction when increased bioaccumulation takes place. In the cytosol of the gills, metals are associated with metallothionein, whereas in the digestive gland, the distribution of metals between metallothioneins and high molecular weight proteins is similar. Metallothionein variation cannot be explained by metals alone; other abiotic factors must be taken into account. In order to use metallothionein as a metal exposure biomarker in field studies, natural variability needs to be taken into account for the correct interpretation of results.

  4. Metallothionein expression in chloroplasts enhances mercury accumulation and phytoremediation capability.

    Science.gov (United States)

    Ruiz, Oscar N; Alvarez, Derry; Torres, Cesar; Roman, Laura; Daniell, Henry

    2011-06-01

    Genetic engineering to enhance mercury phytoremediation has been accomplished by expression of the merAB genes that protects the cell by converting Hg[II] into Hg[0] which volatilizes from the cell. A drawback of this approach is that toxic Hg is released back into the environment. A better phytoremediation strategy would be to accumulate mercury inside plants for subsequent retrieval. We report here the development of a transplastomic approach to express the mouse metallothionein gene (mt1) and accumulate mercury in high concentrations within plant cells. Real-time PCR analysis showed that up to 1284 copies of the mt1 gene were found per cell when compared with 1326 copies of the 16S rrn gene, thereby attaining homoplasmy. Past studies in chloroplast transformation used qualitative Southern blots to evaluate indirectly transgene copy number, whereas we used real-time PCR for the first time to establish homoplasmy and estimate transgene copy number and transcript levels. The mt1 transcript levels were very high with 183,000 copies per ng of RNA or 41% the abundance of the 16S rrn transcripts. The transplastomic lines were resistant up to 20 μm mercury and maintained high chlorophyll content and biomass. Although the transgenic plants accumulated high concentrations of mercury in all tissues, leaves accumulated up to 106 ng, indicating active phytoremediation and translocation of mercury. Such accumulation of mercury in plant tissues facilitates proper disposal or recycling. This study reports, for the first time, the use of metallothioneins in plants for mercury phytoremediation. Chloroplast genetic engineering approach is useful to express metal-scavenging proteins for phytoremediation.

  5. Triclosan activates aryl hydrocarbon receptor (AhR)-dependent apoptosis and affects Cyp1a1 and Cyp1b1 expression in mouse neocortical neurons.

    Science.gov (United States)

    Szychowski, Konrad A; Wnuk, Agnieszka; Kajta, Małgorzata; Wójtowicz, Anna K

    2016-11-01

    Triclosan (TCS) is an antimicrobial agent that is used extensively in personal care and in sanitizing products, such as soaps, toothpastes, and hair products. A number of studies have revealed the presence of TCS in human tissues, such as fat, liver and brain, in addition to blood and breast milk. The aim of the present study was to investigate the impact of TCS on AhR and Cyp1a1/Cyp1b1 signaling in mouse neocortical neurons in primary cultures. In addition to the use of selective ligands and siRNAs, expression levels of mRNA and proteins as well as caspase-3 activity, reactive oxygen species (ROS) formation, and lactate dehydrogenase (LDH) release have been measured. We also studied the involvement of the AhR in TCS-induced LDH release and caspase-3 activation as well as the effect of TCS on ROS generation. Cultures of neocortical neurons were prepared from Swiss mouse embryos on day 15/16 of gestation. The cells were cultured in phenol red-free Neurobasal medium with B27 and glutamine, and the neurons were exposed to 1 and 10µM TCS. Our experiments showed that the expression of AhR and Cyp1a1 mRNA decreased in cells exposed to 10µM TCS for 3 or 6h. In the case of Cyp1b1, mRNA expression remained unchanged compared with the control group following 3h of exposure to TCS, but after 6h, the mRNA expression of Cyp1b1 was decreased. Our results confirmed that the AhR is involved in the TCS mechanism of action, and our data demonstrated that after the cells were transfected with AhR siRNA, the cytotoxic and pro-apoptotic properties of TCS were decreased. The decrease in Cyp1a1 mRNA and protein expression levels accompanied by a decrease in its activity. The stimulation of Cyp1a1 activity produced by the application of an AhR agonist (βNF) was attenuated by TCS, whereas the addition of AhR antagonist (αNF) reversed the inhibitory effects of TCS. In our experiments, TCS diminished Cyp1b1 mRNA and enhanced its protein expression. In case of Cyp1a1 we observed

  6. Mild Hypothermia Combined with Hydrogen Sulfide Treatment During Resuscitation Reduces Hippocampal Neuron Apoptosis Via NR2A, NR2B, and PI3K-Akt Signaling in a Rat Model of Cerebral Ischemia-Reperfusion Injury.

    Science.gov (United States)

    Dai, Hai-Bin; Xu, Miao-Miao; Lv, Jia; Ji, Xiang-Jun; Zhu, Si-Hai; Ma, Ru-Meng; Miao, Xiao-Lei; Duan, Man-Lin

    2016-09-01

    We investigated whether mild hypothermia combined with sodium hydrosulfide treatment during resuscitation improves neuron survival following cerebral ischemia-reperfusion injury beyond that observed for the individual treatments. Male Sprague-Dawley rats were divided into seven groups (n = 20 for each group). All rats underwent Pulsinelli 4-vessel occlusion. Ischemia was induced for 15 min using ligatures around the common carotid arteries, except for the sham group. Immediately after initiating reperfusion, the mild hypothermia (MH), sodium hydrosulfide (NaHS), hydroxylamine (HA), MH + NaHS, MH + HA, and ischemia-reperfusion (I/R) control groups received an intraperitoneal injection of saline, sodium hydrosulfide, hydroxylamine, sodium hydrosulfide, hydroxylamine, and saline, respectively, and mild hypothermia (32 to 33 °C) was induced in the MH, MH + NaHS, and MH + HA groups for 6 h. The levels of NR2A, NR2B, p-Akt, and p-Gsk-3β in the hippocampus of the MH, NaHS, and MH + NaHS groups were higher than those in the I/R control group, with the highest levels observed in the MH + NaHS group (P sodium hydrosulfide treatment for resuscitation following ischemia-reperfusion injury was more beneficial for reducing hippocampal apoptosis and pathology than that of mild hypothermia or hydrogen sulfide treatment alone.

  7. P2X7 Cell Death Receptor Activation and Mitochondrial Impairment in Oxaliplatin-Induced Apoptosis and Neuronal Injury: Cellular Mechanisms and In Vivo Approach.

    Directory of Open Access Journals (Sweden)

    France Massicot

    Full Text Available Limited information is available regarding the cellular mechanisms of oxaliplatin-induced painful neuropathy during exposure of patients to this drug. We therefore determined oxidative stress in cultured cells and evaluated its occurrence in C57BL/6 mice. Using both cultured neuroblastoma (SH-SY5Y and macrophage (RAW 264.7 cell lines and also brain tissues of oxaliplatin-treated mice, we investigated whether oxaliplatin (OXA induces oxidative stress and apoptosis. Cultured cells were treated with 2-200 µM OXA for 24 h. The effects of pharmacological inhibitors of oxidative stress or inflammation (N-acetyl cysteine, ibuprofen, acetaminophen were also tested. Inhibitors were added 30 min before OXA treatment and then in combination with OXA for 24 h. In SH-SY5Y cells, OXA caused a significant dose-dependent decrease in viability, a large increase in ROS and NO production, lipid peroxidation and mitochondrial impairment as assessed by a drop in mitochondrial membrane potential, which are deleterious for the cell. An increase in levels of negatively charged phospholipids such as cardiolipin but also phosphatidylserine and phosphatidylinositol, was also observed. Additionally, OXA caused concentration-dependent P2X7 receptor activation, increased chromatin condensation and caspase-3 activation associated with TNF-α and IL-6 release. The majority of these toxic effects were equally observed in Raw 264.7 which also presented high levels of PGE2. Pretreatment of SH-SY5Y cells with pharmacological inhibitors significantly reduced or blocked all the neurotoxic OXA effects. In OXA-treated mice (28 mg/kg cumulated dose significant cold hyperalgesia and oxidative stress in the tested brain areas were shown. Our study suggests that targeting P2X7 receptor activation and mitochondrial impairment might be a potential therapeutic strategy against OXA-induced neuropathic pain.

  8. 罗布麻提取物对大鼠脑缺血再灌注后细胞凋亡的影响%Effects of apocynum venetum L extract on neuron apoptosis after cerebral ischemia and reperfusion in rats

    Institute of Scientific and Technical Information of China (English)

    陈红艳; 耿淼; 王建华

    2013-01-01

    tissue in the AVE treated groups obviously decreased compared with those of the untreated group( P <0. 05). Drug groups could markedly upregulate the expression of anti-apoptosis gene Bcl-2 and restrain the expression of Bax. Conclusion AVE could inhibit neuron apoptosis by improving the expression of Bel-2 protein and down-regulating the expression of Bax protein, which maybe one of the mechanisms of neuron protective effects of AVE on cerebral ischemia and reperfusion injury.

  9. Metallothionein-3 regulates lysosomal function in cultured astrocytes under both normal and oxidative conditions.

    Science.gov (United States)

    Lee, Sook-Jeong; Park, Mi-Ha; Kim, Hyun-Jae; Koh, Jae-Young

    2010-08-01

    Cellular zinc plays a key role in lysosomal change and cell death in neurons and astrocytes under oxidative stress. Here, using astrocytes lacking metallothionein-3 (MT3), a potential source of labile zinc in the brain, we studied the role of MT3 in oxidative stress responses. H(2)O(2) induced a large increase in labile zinc in wild-type (WT) astrocytes, but stimulated only a modest rise in MT3-null astrocytes. In addition, H(2)O(2)-induced lysosomal membrane permeabilization (LMP) and cell death were comparably attenuated in MT3-null astrocytes. Expression and glycosylation of Lamp1 (lysosome-associated membrane protein 1) and Lamp2 were increased in MT3-null astrocytes, and the activities of several lysosomal enzymes were significantly reduced, indicating an effect of MT3 on lysosomal components. Consistent with lysosomal dysfunction in MT3-null cells, the level of LC3-II (microtubule-associated protein 1 light chain 3), a marker of early autophagy, was increased by oxidative stress in WT astrocytes, but not in MT3-null cells. Similar changes in Lamp1, LC3, and cathepsin-D were induced by the lysosomal inhibitors bafilomycin A1, chloroquine, and monensin, indicating that lysosomal dysfunction may lie upstream of changes observed in MT3-null astrocytes. Consistent with this idea, lysosomal accumulation of cholesterol and lipofuscin were augmented in MT3-null astrocytes. Similar to the results seen in MT3-null cells, MT3 knockdown by siRNA inhibited oxidative stress-induced increases in zinc and LMP. These results indicate that MT3 may play a key role in normal lysosomal function in cultured astrocytes.

  10. Osthole promotes differentiation into neurons and reduces neuronal apoptosis via Wnt/β-catenin signaling pathway in APP transduced neural stem cells%蛇床子素通过Wnt/β-catenin信号通路促进转染APP基因的神经干细胞分化为更多神经元且减少神经元凋亡

    Institute of Scientific and Technical Information of China (English)

    姚璎珈; 孔亮; 教亚男; 李少恒; 陶震宇; 闫宇辉; 杨静娴

    2015-01-01

    Aim To investigate the effects of osthole ( Ost) on the ability of proliferation and differentiation in APP transduced neural stem cells( NSCs) , and neu-ronal apoptosis, in order to find related mechanism. Methods A model of Alzheimer′s disease( AD) cells was successfully established by transducing APP gene into NSCs in vitro. The ability of proliferation and dif-ferentiation was tested by staining. The viability of NSCs was determined by using CCK-8 assay. The cell apoptosis was tested by Hoechst 33258 staining. The expression of GSK-3β and β-catenin mRNA was deter-mined by RT-PCR. The expression of GSK-3β and β-catenin protein was determined by Western blot. Re-sults The ability of proliferation had increased by 10 . 24% with Ost treatment, compared with APP group. The ability of differentiation had increased by 6 . 74%with Ost treatment, compared with APP group. The vi-ability of NSCs had increased and cell apoptotic rate had decreased significantly. From the results of RT-PCR and Western blot, we could find the expression of GSK-3βmRNA and protein had decreased, and the ex-pression of β-catenin mRNA and protein had increased significantly, compared with APP group. Conclusion Ost could enhance the ability of proliferation and dif-ferentiation into more neurons of NSCs transducing APP gene, and reduce neuronal apoptosis. It might be relat-ed with activiting Wnt/β-catenin signaling pathway.%目的:通过转染APP基因于神经干细胞,研究蛇床子素对其增殖和分化能力的影响,及对神经元凋亡的影响,并研究其机制。方法体外建立阿尔茨海默病的神经干细胞模型,利用免疫组化染色法,研究蛇床子素对转染APP基因的神经干细胞增殖和分化能力的影响;通过CCK-8法检测神经干细胞存活率;Hoechst 33258法检测神经元凋亡情况;RT-PCR法检测 GSK-3β和β-catenin mRNA 的表达变化情况;Western blot法检测 GSK-3β和β-catenin 蛋白的表达变化情况

  11. 全麻/控制性降压手术对海马神经元凋亡的影响%Effect of surgical operation with general anesthesia/controlled hypotension on neuronal apoptosis in hippocampus

    Institute of Scientific and Technical Information of China (English)

    王改梅; 刘喆; 沈颖洁; 方剑乔

    2013-01-01

    maintained at 40% of the baseline level for 1 h. The animals in both groups underwent exploratory laparotomy in the corresponding period. MAP was controlled to return to baseline level after surgery, and then the brain tissues were taken out 72 h later. The levels of IL-1β, TNF-α, Bcl-2, Bax and cleaved caspase-3 were measured by the method of immunohistochemistry, and neuronal apoptosis was detected by TUNEL in CA1 and CA3 regions of hippocampus. RESULTS: The ratios of IL-1β and TNF-α positive cells in CA1 and CA3 regions of hippocampus were significantly increased in group C than those in group A. The ratio of Bcl-2/Bax in CA1 and CA3 regions of hippocampus was lower in group C than that in group A. In contrast, the ratio of caspase-3 positive cells in group C was significantly higher than that in group A. The apoptotic index of hippocampal neurons in group C was significantly higher than that in group A. CONCLUSION: The exaggerated expression of proinflammatory cytokines in hippocampus may be induced by surgical operation with general anesthesia/controlled hypotension, leade to reduce the Bcl-2/Bax ratio, and to promote the expression of caspase-3, so as to resulting in neuronal apoptosis in hippocampus. These results suggest that postoperative neuronal damage may be related to hippocampal neuro-inflammation caused by surgical operation with general anesthesia/controlled hypotension.

  12. Studies on the Apoptosis of Retinal Neurons Following Optic Nerve Injure in Zebra fish (Brachydanio rerio )%视神经损伤引起斑马鱼视网膜神经细胞凋亡的研究

    Institute of Scientific and Technical Information of China (English)

    陈玲玲; 刘明学; 王子仁

    2001-01-01

    The changes of the density of retinal ganglion cells, rod cellsand co ne cells of Zebra fish following optic nerve crush or section were quantitativel y studied with paraffin continuous sections stained by hematxylin.The results sh owed that the density of nucleus of the three-type retinal neurons was decreased during 7~21 days after optic nerve injure.The decreased ratio of the GCs was b igger than that of photoreceptors (including CCs and RCs), especially in CCs.In both the crush and section conditioning , the change of optic nerves was m ore o bvious in the latter (section condition).These changes indicated that optic ner ve injure could not only affect the retinal ganglion cells directly, but could transneuronally affect the changes of photoreceptors. It can be supposed from all the results above that the decrease of the de nsity of retinal neurons may be due to the loss of thei r target tissues. It is an expression of apoptosis of retinal neurons.%用石蜡连续切片苏木精染色法,通过定量分析研究夹伤和切断视神经后,斑马鱼视网膜神经节细胞、视杆和视锥细胞密度的变化.结果发现,在损伤视神经7~21d后,上述3种细胞的细胞核密度均呈减少趋势,节细胞减少的比率大于感光细胞,而感光细胞中视锥细胞所受影响比视杆细胞更为明显;在夹伤和切断视神经两种情况中,后者引起视网膜神经节细胞核密度的减少更为显著.上述结果表明,损伤视神经不但影响与其相连的神经节细胞,而且可逆向跨神经元地影响感光细胞的变化.由上述结果推测,由于损伤视神经使视网膜神经节细胞失去靶组织而引起的各种神经细胞密度减少是视网膜中神经细胞凋亡的表现.

  13. 内源性内皮衍生超极化因子对缺氧/再给氧致大鼠海马神经细胞凋亡的抑制作用%The inhibition of endogenous endothelium-derived hyperpolarizing factor on hypoxia/reoxygenation-induced apoptosis of hippocampal neurons in rats

    Institute of Scientific and Technical Information of China (English)

    余小蒙; 吴剑; 陈志武; 郭岩

    2014-01-01

    Objective To study the effect of acetylcholine ( ACh) induced endogenous endothelium-derived hyper-polarizing factor ( EDHF ) on apoptosis of rat hippocampal neurons. Methods Primarily cultured hippocampal neu-rons were insulted by hypoxia/reoxygenation ( H/R) , and EDHF from rat MCA segment was induced by acetylcho-line ( ACh) in the presence of NG-nitro-L-argininemethyl ester ( L-NAME) and indomethacin ( Indo) , the rate of hippocampal neuron apoptosis was detected by Hoechst staining and flow cytometry ( FC) methods. Results Ex-aminations of FC and Hoechst staining showed that the apoptosic rate of rat hippocampal neurons was very low in the sham group, only about 3. 1% and 2. 2% respectively,but H/R injury caused an obvious increase of apoptosis of rat hippocampal neurons. Co-adminstration of endothelium-intact MCA segment with 1 μmol/L ACh or co-admin-stration of endothelium-intact MCA segment with 1 μmol/L ACh + 30 μmol/L L-NAME + 10 μmol/L Indo markedly inhibits H/R-induced apoptosis of rat hippocamal neurons. However, administration of ACh or MCA seg-ment separately had not effect on H/R-induced apoptosis of rat hippocamal neurons, and neither did co-adminstra-tion of endothelium-denuded MCA segment with ACh. Conclusion Endogenous EDHF released from ACh-induced vascular endothelial cells significantly inhibits H/R-caused apoptosis of rat primarily cultured neurons.%目的研究乙酰胆碱( ACh)诱导内源性内皮衍生超极化因子( EDHF)对缺氧/再给氧大鼠海马神经细胞凋亡的影响。方法取原代培养的大鼠海马神经细胞制备缺氧/再给氧损伤模型;大鼠大脑中动脉( MCA)血管段用一氧化氮合酶抑制剂NG-nitro-L-argininemethyl ester ( L-NAME)和前列环素抑制剂indomethacin ( Indo)预处理,用ACh诱导血管内皮细胞合成释放内源性EDHF;分别用流式细胞仪法和Ho-echst荧光法检测海马神经元凋亡。结果流式细胞仪法和Hoechst荧光法检测均表明假缺氧组海

  14. METALLOTHIONEINS AS SENSORS AND CONTROLS EXCHANGE OF METALS IN THE CELLS

    Directory of Open Access Journals (Sweden)

    V. A. Kutyakov

    2014-01-01

    Full Text Available The basic information on the classification, structure, induction and degradation, functions of the protein family – metallothionein (MT, including CNS in health and disease are presented in this review. It was found that four major isoforms of metallothionein perform different biological roles, are localized in dif- ferent tissues. Induction of MT is a universal reaction to the impact of a variety of stress factors. In recent years, understanding of the role of metallothioneins in metal homeostasis in the tissues in normal and pathological conditions have changed significantly. Notes polyfunctionality metallothioneins (transport of metal ions, maintaining redox reactions, tread, signal, modulated and regulatory functions and their im- pact on basic cellular functions such as proliferation, differentiation, programmed cell death. Further- more, a special role is shown MT in the pathogenesis of cardiovascular, neurodegenerative and neoplastic disorders.Currently, these molecules are increasingly considered as potential targets for therapy of a wide range of diseases and the development of targeted approaches to the regulation of expression of MT – one of the promising areas of pharmacology and toxicology. Stressed the safety of metallothioneins as therapeutic agents.

  15. Influence of metallothioneins on zinc and copper distribution in brain tumours.

    Science.gov (United States)

    Floriańczyk, Bolesław; Osuchowski, Jacek; Kaczmarczyk, Robert; Trojanowski, Tomasz; Stryjecka-Zimmer, Marta

    2003-01-01

    Metallothioneins take part in the homeostasis of the ions of the metals which are necessary for the proper metabolism of the organism (zinc, copper), in biosynthesis regulation of the zinc-containing proteins and also in the detoxication of metals from the tissues. They also protect the tissue from the effects of free radicals, radiation, electrophilic pharmacological agents used in the cancer therapy and from mutagens. The experimental materials were brain astrocytomas, benign gliomas and malignant gliomas. The levels of the metallothionein were determined by cadmium-haemoglobin affinity assay using the cadmium isotope (109Cd). The values of zinc and copper were determined by means of atomic absorption spectrophotometry. In our studies, the level of metallothioneins in the group of malignant neoplasms was slightly higher than the level of these proteins in the group of benign neoplasms. The correlation coefficient of the studied parameters proved an interrelation between the levels of zinc and copper and the content of metallothioneins. In malignant neoplasms, the level of zinc showed a positive relationship with the metallothionein level, whereas the copper content showed an inverse relationship. There was a statistical difference, but no significant difference, in the levels of copper between malignant and benign groups.

  16. A highly sensitive fluorescence probe for metallothioneins based on tiron-copper complex.

    Science.gov (United States)

    Xiao, Xilin; Xue, Jinhua; Liao, Lifu; Huang, Mingyang; Zhou, Bin; He, Bo

    2015-06-15

    The fabrication of tiron-copper complex as a novel fluorescence probe for the sensitive directly detection of metallothioneins at nanomolar levels was demonstrated. In Britton-Robinson (B-R) buffer (pH 7.50), the interaction of bis(tiron)copper(II) complex cation [Cu(tiron)2](2+) and metallothioneins enhanced the fluorescence intensity of the system. The fluorescence enhancement at 347 nm was proportional to the concentration of metallothioneins. The mechanism was studied and discussed in terms of the fluorescence spectra. Under the optimal experimental conditions, at 347 nm, there was a linear relationship between the fluorescence intensity and the concentration of the metallothioneins in the range of 8.80 × 10(-9)-7.70 × 10(-7)mol L(-1), with a correlation coefficient of r=0.995 and detection limit 2.60 × 10(-9)mol L(-1). The relative standard deviation was 0.77% (n=11), and the average recovery 94.4%. The method proposed was successfully reliable, selective and sensitive in determining of trace metallothioneins in fish visceral organ samples with the results in good agreement with those obtained by HPLC.

  17. Shapes of Differential Pulse Voltammograms and Level of Metallothionein at Different Animal Species

    Science.gov (United States)

    Adam, Vojtech; Beklova, Miroslava; Pikula, Jiri; Hubalek, Jaromir; Trnkova, Libuse; Kizek, Rene

    2007-01-01

    Metallothioneins play a key role in maintaining homeostasis of essential metals and in protecting of cells against metal toxicity as well as oxidative damaging. Excepting humans, blood levels of metallothionein have not yet been reported from any animal species. Blood plasma samples of 9 animal species were analysed by the adsorptive transfer stripping technique to obtain species specific voltammograms. Quite distinct records were obtained from the Takin (Budorcas taxicolor), while other interesting records were observed in samples from the European Bison (Bison bonasus bonasus) and the Red-eared Slider (Trachemys scripta elegans). To quantify metallothionein the catalytic peak Cat2 was used, well developed in the Domestic Fowl (Gallus gallus f. domestica) and showing a very low signal in the Red Deer (Cervus elaphus). The highest levels of metallothionein reaching over 20 µM were found in the Domestic Fowl. High levels of MT were also found in the Bearded Dragon (Pogona vitticeps) and the Grey Wolf (Canis lupus lupus). The lowest values of about 1-3 µM were determined in the Red-eared Slider, Takin and Red Deer. Employing a simple electrochemical detection it was possible to examine variation in blood metallothionein in different species of vertebrates. PMID:28903235

  18. Shapes of Differential Pulse Voltammograms and Level of Metallothionein at Different Animal Species

    Directory of Open Access Journals (Sweden)

    Rene Kizek

    2007-10-01

    Full Text Available Metallothioneins play a key role in maintaining homeostasis of essential metalsand in protecting of cells against metal toxicity as well as oxidative damaging. Exceptinghumans, blood levels of metallothionein have not yet been reported from any animalspecies. Blood plasma samples of 9 animal species were analysed by the adsorptive transferstripping technique to obtain species specific voltammograms. Quite distinct records wereobtained from the Takin (Budorcas taxicolor, while other interesting records were observedin samples from the European Bison (Bison bonasus bonasus and the Red-eared Slider(Trachemys scripta elegans. To quantify metallothionein the catalytic peak Cat2 was used,well developed in the Domestic Fowl (Gallus gallus f. domestica and showing a very lowsignal in the Red Deer (Cervus elaphus. The highest levels of metallothionein reachingover 20 μM were found in the Domestic Fowl. High levels of MT were also found in theBearded Dragon (Pogona vitticeps and the Grey Wolf (Canis lupus lupus. The lowestvalues of about 1-3 μM were determined in the Red-eared Slider, Takin and Red Deer. Employing a simple electrochemical detection it was possible to examine variation in blood metallothionein in different species of vertebrates.

  19. The Functions of Metamorphic Metallothioneins in Zinc and Copper Metabolism

    Directory of Open Access Journals (Sweden)

    Artur Krężel

    2017-06-01

    Full Text Available Recent discoveries in zinc biology provide a new platform for discussing the primary physiological functions of mammalian metallothioneins (MTs and their exquisite zinc-dependent regulation. It is now understood that the control of cellular zinc homeostasis includes buffering of Zn2+ ions at picomolar concentrations, extensive subcellular re-distribution of Zn2+, the loading of exocytotic vesicles with zinc species, and the control of Zn2+ ion signalling. In parallel, characteristic features of human MTs became known: their graded affinities for Zn2+ and the redox activity of their thiolate coordination environments. Unlike the single species that structural models of mammalian MTs describe with a set of seven divalent or eight to twelve monovalent metal ions, MTs are metamorphic. In vivo, they exist as many species differing in redox state and load with different metal ions. The functions of mammalian MTs should no longer be considered elusive or enigmatic because it is now evident that the reactivity and coordination dynamics of MTs with Zn2+ and Cu+ match the biological requirements for controlling—binding and delivering—these cellular metal ions, thus completing a 60-year search for their functions. MT represents a unique biological principle for buffering the most competitive essential metal ions Zn2+ and Cu+. How this knowledge translates to the function of other families of MTs awaits further insights into the specifics of how their properties relate to zinc and copper metabolism in other organisms.

  20. Metallothionein-I induction by stress in specific brain areas.

    Science.gov (United States)

    Hidalgo, J; Campmany, L; Martí, O; Armario, A

    1991-10-01

    The distribution of metallothionein-I (MT) in several areas of the brain and its induction by immobilization stress has been studied in the rat. MT content was highest in hippocampus and midbrain and lowest in frontal cortex and pons plus medulla oblongata. Immobilization stress for 18 hours (which was accompanied by food and water deprivation) significantly increased MT levels in the frontal cortex, pons plus medulla oblongata and hypothalamus, but not in midbrain and hippocampus. The effect of stress on MT levels was specific as food and water deprivation along had no significant effect on MT levels in any of the brain areas studied. The effect of stress on MT levels was independent of changes in cytosolic Zn content; this was generally unaffected by stress or food and water deprivation but decreased in pons plus medulla oblongata from stressed rats. The results suggest that MT is induced more significantly in the brain areas that are usually involved in the response of animals to stress.

  1. Gold resistance in cultured human cells possible role of metallothionein.

    Science.gov (United States)

    Glennås, A

    1983-01-01

    Insufficient therapeutic effect of auranofin (AF), used in the treatment of rheumatoid arthritis, is found in about 8% of the patients included in clinical trials until now. The mechanisms of resistance to gold-containing drugs are not known, but one reason might be acquired drug resistance. We have studied the relationship between the effects of gold and concentration of the cytoplasmic metal-binding protein metallothionein (MT), in order to evaluate MT as a possible contributing factor to resistance against AF. Different strains of cultured human epithelial cells derived from normal skin, treated with AF, were used as models. The experiments indicate two possible mechanisms for resistance against AF in cells: 1) binding of gold to pre-existent cadmium-induced MT or to de novo AF-induced MT, and 2) the cells' ability to keep the intracellular gold concentration at a low level. AF apparently causes a rapid and pronounced increase of MT-content in these cells. Preliminary results also indicated that AF causes increase of MT-content in human rheumatoid synovial cells, grown as primary cultures. These findings may have two clinical implications: 1) AF-induced MT may decrease therapeutic response, and 2) decrease the toxicity of AF.

  2. Purification of human brain metallothionein by organic and reversed-phase high-performance liquid chromatography under acidic conditions.

    Science.gov (United States)

    Cartel, N J

    1996-02-09

    A simplified high-performance liquid chromatographic method for the detection of metallothioneins, notably metallothionein-III, has been developed. In order to purify metallothionein, differential acetone precipitation at 50% (v/v) and at 80% (v/v) was employed on a 20% normal human brain homogenate. The reconstituted pellet was injected into a C18 microbore reversed-phase HPLC column, equilibrated with 0.1% trifluoroacetic acid, and developed at a flow-rate of 800 microliter/min with a linear gradient from 0% to 60% acetonitrile in 0.094% trifluoroacetic acid for 60 min. Western blots indicated that metallothioneins-I and II eluted at 16% acetonitrile and metallothionein-III eluted at 37% acetonitrile.

  3. 金属硫蛋白在缺血性脑损伤过程中的调控作用%Regulatory role of metallothionein in process of ischemic brain injury

    Institute of Scientific and Technical Information of China (English)

    郭家彬; 冯敏; 张丽; 彭双清

    2014-01-01

    Metallothionein ( MT ) is a cysteine-rich and low-molecular metal binding protein. Three isoforms of MT have been found in the central nervous system, including MT-Ⅰ, Ⅱ, and Ⅲ. MT is widely involved in many critical activities in the central nervous system, such as neuronal growth, auto-defensive reaction, immune-regulation, and repair of cerebral injury. MT exerts many important biological functions like scavenging of free radicals, regulation of ion homeostasis in brain cells, detoxification of heavy metals, anti-inflammation, and anti-apoptosis. Recently, MT has been increasingly shown to have protective effects against cerebral ischemia. MT promises to be an important target for prevention and/or treatment of cerebral ischemic disease. ln this review, the expression and regulation characteristics, and the effect of cerebral ischemic stress on MT expression have been summarized, with focus on the neuro-protective effect of MT and its possible underlying mechanisms.%金属硫蛋白( MT)是一类富含半胱氨酸残基的低分子量金属连接蛋白。在中枢神经系统中主要有MT-Ⅰ,Ⅱ和Ⅲ亚型。 MT可广泛参与中枢神经系统的神经细胞生长、自主防御反应、免疫调节和脑损伤修复等活动。 MT具有清除自由基、调节脑细胞离子稳态、重金属解毒、抗炎症和抗细胞凋亡等多种重要生物学功能。脑缺血应激可显著诱导脑组织细胞MT的表达。近年来越来越多的研究表明, MT对脑缺血损伤具有重要保护作用,有可能成为预防和(或)治疗脑缺血疾病的重要靶点。本文简要综述中枢神经系统中MT的表达与调控特点以及脑缺血应激对MT表达的影响,重点讨论MT对脑缺血损伤的保护作用及其可能机制。

  4. Metal release in metallothioneins induced by nitric oxide: X-ray absorption spectroscopy study.

    Science.gov (United States)

    Casero, Elena; Martín-Gago, José A; Pariente, Félix; Lorenzo, Encarnación

    2004-12-01

    Metallothioneins (MTs) are low molecular weight proteins that include metal ions in thiolate clusters. The capability of metallothioneins to bind different metals has suggested their use as biosensors for different elements. We study here the interaction of nitric oxide with rat liver MTs by using in situ X-ray absorption spectroscopy techniques. We univocally show that the presence of NO induces the release of Zn atoms from the MT structure to the solution. Zn ions transform in the presence of NO from a tetrahedral four-fold coordinated environment in the MT into a regular octahedral six-fold coordinated state, with interatomic distances compatible with those of Zn solvated in water.

  5. Metallothionein I+II expression and their role in experimental autoimmune encephalomyelitis

    DEFF Research Database (Denmark)

    Penkowa, M; Hidalgo, J

    2000-01-01

    We examined the expression and roles of neuroprotective metallothionein-I+II (MT-I+II) in the rat CNS in experimental autoimmune encephalomyelitis (EAE), an animal model for the human autoimmune disease, multiple sclerosis (MS). EAE caused significant macrophage activation, T-lymphocyte infiltrat......We examined the expression and roles of neuroprotective metallothionein-I+II (MT-I+II) in the rat CNS in experimental autoimmune encephalomyelitis (EAE), an animal model for the human autoimmune disease, multiple sclerosis (MS). EAE caused significant macrophage activation, T...

  6. Single and double metallothionein knockout in the nematode C. elegans reveals cadmium dependent and independent toxic effects on life history traits

    Energy Technology Data Exchange (ETDEWEB)

    Hughes, Sam [School of Biosciences, Cardiff University, Main Building, Park Place, Cardiff CF10 3TL (United Kingdom); School of Biomedical and Health Sciences, Pharmaceutical Sciences Research Division, King' s College London, 150 Stamford Street, London SE1 9NH (United Kingdom); Stuerzenbaum, Stephen R. [School of Biosciences, Cardiff University, Main Building, Park Place, Cardiff CF10 3TL (United Kingdom) and School of Biomedical and Health Sciences, Pharmaceutical Sciences Research Division, King' s College London, 150 Stamford Street, London SE1 9NH (United Kingdom)]. E-mail: stephen.sturzenbaum@kcl.ac.uk

    2007-01-15

    The genome of the nematode Caenorhabditis elegans contains two metallothionein genes, both involved in metal homeostasis and/or detoxification. Single metallothionein knockout mutants have been created and now, for the first time, a double mutant has been isolated. Life history studies in the presence or absence of cadmium showed that all metallothionein mutants are viable. Although cadmium did not influence longevity, a dose dependent reduction in total brood size and volumetric growth was observed in wild type animals, which was magnified in single knockouts and further exacerbated in the double knockout. However, the metallothionein deletion caused two effects that are independent of cadmium exposure, namely all knockout strains displayed a reduced total brood size and the deletion of both metallothionein loci caused a significant reduction in volumetric growth. In summary, metallothionein is undoubtedly an important player in cadmium detoxification, but evidently also an important factor in cadmium independent pathways. - Metallothionein is a modifier of life-history parameters.

  7. 脊髓损伤后Bcl-2抗神经元凋亡的研究%Study on the role of Bcl-2 in anti- neuronal apoptosis after spinal cord injury

    Institute of Scientific and Technical Information of China (English)

    王瑛; 孙志扬; 张夔鸣; 许国强; 李光

    2010-01-01

    ,functional deficits were evaluated with BBB scales, and the apoptosis of neurons was investigated by using TUNEL method. Another three mice of control group were only treated with laminectomy without SCI for comparison. Results The mean functional scores in the control mice were lower than those in the Bcl-2 TG mice, although the unpaired T -test revealed no significant differences. On the other hand, the number of TUNEL positive neurons and IOD(Integrated Optical Density)score in the Bcl-2 TG mice were both significantly lower than those in the control mice. Conclusions This experiment suggests that overexpression of Bcl-2 may suppress neuronal apoptosis after SCI. The Bcl-2 may be an important factor in relieving the damage within CNS after trauma.

  8. Cadmium modulates adipocyte functions in metallothionein-null mice

    Energy Technology Data Exchange (ETDEWEB)

    Kawakami, Takashige; Nishiyama, Kaori; Kadota, Yoshito; Sato, Masao; Inoue, Masahisa; Suzuki, Shinya, E-mail: suzukis@ph.bunri-u.ac.jp

    2013-11-01

    Our previous study has demonstrated that exposure to cadmium (Cd), a toxic heavy metal, causes a reduction of adipocyte size and the modulation of adipokine expression. To further investigate the significance of the Cd action, we studied the effect of Cd on the white adipose tissue (WAT) of metallothionein null (MT{sup −/−}) mice, which cannot form atoxic Cd–MT complexes and are used for evaluating Cd as free ions, and wild type (MT{sup +/+}) mice. Cd administration more significantly reduced the adipocyte size of MT{sup −/−} mice than that of MT{sup +/+} mice. Cd exposure also induced macrophage recruitment to WAT with an increase in the expression level of Ccl2 (MCP-1) in the MT{sup −/−} mice. The in vitro exposure of Cd to adipocytes induce triglyceride release into culture medium, decrease in the expression levels of genes involved in fatty acid synthesis and lipid hydrolysis at 24 h, and at 48 h increase in phosphorylation of the lipid-droplet-associated protein perilipin, which facilitates the degradation of stored lipids in adipocytes. Therefore, the reduction in adipocyte size by Cd may arise from an imbalance between lipid synthesis and lipolysis. In addition, the expression levels of leptin, adiponectin and resistin decreased in adipocytes. Taken together, exposure to Cd may induce unusually small adipocytes and modulate the expression of adipokines differently from the case of physiologically small adipocytes, and may accelerate the risk of developing insulin resistance and type 2 diabetes. - Highlights: • Cd causes a marked reduction in adipocyte size in MT-null mice. • Cd enhances macrophage migration into adipose tissue and disrupt adipokine secretion. • MT gene alleviates Cd-induced adipocyte dysfunctions. • Cd enhances the degradation of stored lipids in adipocytes, mediated by perilipin. • Cd induces unusually small adipocytes and the abnormal expression of adipokines.

  9. Role of Metallothionein in Post-Burn Inflammation.

    Science.gov (United States)

    Zhang, Wei; Xie, Yongjun; Liu, Weihua; Xu, Xuefeng; Chen, Xuelian; Liu, Hairong; Liu, Yueming

    2016-04-01

    Metallothioneins (MTs) are a family of low molecular-weight and cysteine-rich metalloproteins that regulate metal metabolism and protect cells from oxygen free radicals. Recent studies suggested that MTs have some anti-inflammatory effects. However, the role of MTs in post-burn inflammation remains unclear. This study is designed to investigate the role of MTs in post-burn inflammation in a mouse burn model. MT-I/II null (-/-) and C57BL/6 wild-type (WT) mice were randomly divided into sham burn, burn, Zn treated, and Zn-MT-2 treated groups. The inflammatory cytokines levels were measured by enzyme-linked immunosorbent assay (ELISA). Myeloperoxidase (MPO) activity was determined by spectrophotometry. In in vitro study, exogenous MT-2 was added to macrophages that were stimulated with burn serum in the presence or absence of a p38 MAPK inhibitor SB203580. The IL-6 and TNF-α messenger RNA (mRNA) expression were detected by quantitative real-time polymerase chain reaction. The levels of p38 expression were determined by Western blot. Burn induced increased inflammatory cytokines such as interleukin (IL)-1β, IL-6, tumor necrosis factors-α, and macrophage chemoattractant protein-1 production in burn wound and serum. The MPO activities in the lung and heart were also increased after burn. These effects were significantly more prominent in MT (-/-) mice than in WT mice. Furthermore, these effects were inhibited by administration of exogenous MT-2 to both WT and MT (-/-) mice. Exogenous MT-2 inhibited the p38 expression and abrogated the increase of IL-6 and TNF-α mRNA expression from macrophages that were stimulated with burn serum. The effect of MT-2 was not further strengthened in the presence of SB203580. MTs may have a protective role against post-burn inflammation and inflammatory organ damage, at least partly through inhibiting the p38 MAPK signaling.

  10. Minimal role of metallothionein in decreased chelator efficacy for cadmium.

    Science.gov (United States)

    Waalkes, M P; Watkins, J B; Klaassen, C D

    1983-05-01

    Chelator efficacy in Cd poisoning drops precipitously if therapy is not commenced almost immediately after exposure. Metallothionein (MT), a low-molecular-weight metal-binding protein with high affinity for Cd, may be important for this phenomenon. To more fully assess this role of MT in the acute drop in chelator efficacy following Cd poisoning, rats were injected iv with radioisotopic Cd (1mg/kg as CdCl2; 50 muCi/kg) followed by diethylenetriaminepentaacetic acid (DTPA; 90 mg/kg ip) at various times (0, 15, 30, 60, and 120 min) after Cd. Ther percentage of the Cd dose remaining in major organs 24 hr following Cd was determined. Although DTPA reduced Cd content in the various organs when given immediately after Cd, the chelator was ineffective at all later times. Increases in hepatic and renal MT did not occur until 2 hr after Cd, and did not coincide with the earlier drop in chelator efficacy. Blockade of MT synthesis by actinomycin D treatment (1.25 mg/kg, 1 hr before Cd) failed to prolong the chelators effectiveness. Furthermore, newborn rats have high levels of hepatic MT which had no effect on the time course of chelator effectiveness since DTPA still decreased Cd organ contents if given immediately following Cd but had no effect if given 2 hr after Cd. Therefore, if appears that MT does not have an important role in the acute decrease in efficacy of chelation therapy for Cd poisoning. The quick onset of chelator ineffectiveness may be due to the rapid uptake of Cd into tissues which makes it relatively unavailable of chelation.

  11. Ankaferd Blood Stopper induces apoptosis and regulates PAR1 and ...

    African Journals Online (AJOL)

    Mine Mumcuoglu

    2014-12-16

    Dec 16, 2014 ... Besides its widely accepted role in plate- ... important role in cell cycle arrest and apoptosis [17]. ABS has ..... lial cells, fibroblasts, neuronal cells and tumor cells by either inducing or ... hemorrhages and wound healing.

  12. APOPTOSIS AFTER SPINAL CORD INJURY IN RATS

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Objective To confirm the role played by apoptosis in spinal cord injury. Methods 36 rats models of spinal cord injury were made by Allen method. Histological examinations using HE staining and in situ end-labeling were used to observe apoptosis in spinal cord tissues from 1h to 21d after injury. Results HE staining sections showed hemorrhage and necrosis, neuronal degeneration and gliai cell proliferation. In situ end-labeling sections showed the appearance of apoptosis in both gray and white matter as well as in both central and surrounding region. The number of apoptotic cells increased from 12h after injury, increased to the peak at 4d and declined to normal at 21d. Conclu sion The results suggest that apoptosis, especially glial apoptosis, plays a role in the pathogenesis of spinal cord in jury.

  13. Induction by mercury compounds of brain metallothionein in rats: Hg{sup 0} exposure induces long-lived brain metallothionein

    Energy Technology Data Exchange (ETDEWEB)

    Yasutake, Akira; Nakano, Atsuhiro [Biochemistry Section, National Institute for Minamata Disease, Kumamoto (Japan); Hirayama, Kimiko [Kumamoto University, College of Medical Science (Japan)

    1998-03-01

    Metallothionein (MT) is one of the stress proteins which can easily be induced by various kind of heavy metals. However, MT in the brain is difficult to induce because of blood-brain barrier impermeability to most heavy metals. In this paper, we have attempted to induce brain MT in rats by exposure to methylmercury (MeHg) or metallic mercury vapor, both of which are known to penetrate the blood-brain barrier and cause neurological damage. Rats treated with MeHg (40 {mu}mol/kg per day x 5 days, p.o.) showed brain Hg levels as high as 18 {mu}g/g with slight neurological signs 10 days after final administration, but brain MT levels remained unchanged. However, rats exposed to Hg vapor for 7 days showed 7-8 {mu}g Hg/g brain tissue 24 h after cessation of exposure. At that time brain MT levels were about twice the control levels. Although brain Hg levels fell gradually with a half-life of 26 days, MT levels induced by Hg exposure remained unchanged for >2 weeks. Gel fractionation revealed that most Hg was in the brain cytosol fraction and thus bound to MT. Hybridization analysis showed that, despite a significant increase in MT-I and -II mRNA in brain, MT-III mRNA was less affected. Although significant Hg accumulation and MT induction were observed also in kidney and liver of Hg vapor-exposed rats, these decreased more quickly than in brain. The long-lived MT in brain might at least partly be accounted for by longer half-life of Hg accumulated there. The present results showed that exposure to Hg vapor might be a suitable procedure to provide an in vivo model with enhanced brain MT. (orig.) With 4 figs., 1 tab., 27 refs.

  14. The Cd(II)-binding abilities of recombinant Quercus suber metallothionein: bridging the gap between phytochelatins and metallothioneins.

    Science.gov (United States)

    Domènech, Jordi; Orihuela, Rubén; Mir, Gisela; Molinas, Marisa; Atrian, Sílvia; Capdevila, Mercè

    2007-08-01

    In this work, we have analyzed both at stoichiometric and at conformational level the Cd(II)-binding features of a type 2 plant metallothionein (MT) (the cork oak, Quercus suber, QsMT). To this end four peptides, the wild-type QsMT and three constructs previously engineered to characterize its Zn(II)- and Cu(I)-binding behaviour, were heterologously produced in Escherichia coli cultures supplemented with Cd(II), and the corresponding complexes were purified up to homogeneity. The Cd(II)-binding ability of these recombinant peptides was determined through the chemical, spectroscopic and spectrometric characterization of the recovered clusters. Recombinant synthesis of the four QsMT peptides in cadmium-rich media rendered complexes with a higher metal content than those obtained from zinc-supplemented cultures and, consequently, the recovered Cd(II) species are nonisostructural to those of Zn(II). Also of interest is the fact that three out of the four peptides yielded recombinant preparations that included S(2-)-containing Cd(II) complexes as major species. Subsequently, the in vitro Zn(II)/Cd(II) replacement reactions were studied, as well as the in vitro acid denaturation and S(2-) renaturation reactions. Finally, the capacity of the four peptides for preventing cadmium deleterious effects in yeast cells was tested through complementation assays. Consideration of all the results enables us to suggest a hairpin folding model for this typical type 2 plant Cd(II)-MT complex, as well as a nonnegligible role of the spacer in the detoxification function of QsMT towards cadmium.

  15. Metallothionein induction in human CNS in vitro: neuroprotection from ionizing radiation.

    Science.gov (United States)

    Cai, L; Cherian, M G; Iskander, S; Leblanc, M; Hammond, R R

    2000-07-01

    There have been extensive studies on the regulation of metallothionein (MT) synthesis, and its biological role in liver and kidney. Although there are few reports on brain MT, there is a growing interest in the role of MT in brain. There have been no publications to date on MT synthesis in the human central nervous system (CNS) following exposure to ionizing radiation. In the present study, primary human CNS cultures were used to examine the effect of ionizing radiation on MT mRNA and protein synthesis. In the same cultures, the neuroprotective effects of zinc (Zn) and cadmium (Cd)-induced MT synthesis from high-dose radiation were also examined. Primary, serum-free, human CNS cultures were exposed to 30 or 60 Gy gamma-rays. The total MT protein was then measured by a Cd-heme assay, and mRNA for MT-II and MT-III was detected by reverse transcription polymerase chain reaction (RT-PCR). Cytotoxicity was measured by LDH release and apoptotic cell death by DNA fragmentation analysis. Sublethal neuroglial injury was assessed morphologically using specific astrocytic (glial fibrillary acidic protein--GFAP) and neuronal (microtubule-associated protein 2--MAP2) immunohistochemical markers. The total MT protein content was increased 12h after exposure to 30Gy. The increase in MT content in response to 60Gy was not statistically significant. MT-II mRNA levels increased at 3 and 6h after exposure to 30Gy gamma-rays, with a maximum expression at 12-24 h. MT-III mRNA was not significantly affected. Exposure to 60 Gy, but not 30 Gy, caused a marked increase in LDH release. Cells exposed to 30 Gy or less showed some apoptotic cell death by DNA fragmentation analysis, while exposure to 60 Gy resulted in a DNA smear confirmed by LDH assays. Preinduction of MT by 5 microM Cd or 100 microM Zn resulted in a significant reduction in radiation-induced LDH release. Morphological evaluations revealed that Cd or Zn preincubation led to relative preservation of MAP2 staining and GFAP. Both

  16. Amount and metal composition of midgut gland metallothionein in shore crabs (Carcinus maenas) after exposure to cadmium in the food

    Energy Technology Data Exchange (ETDEWEB)

    Pedersen, Knud Ladegaard; Bach, Louise Thornhøj; Bjerregaard, Poul, E-mail: poul@biology.sdu.dk

    2014-05-01

    Highlights: • Crabs were fed with Cd in concentrations of 1.1–5.1 μg g⁻¹ food. • Metallothionein concentrations only increased at 5.1 μg g⁻¹. • Cd contents of metallothionein increased linearly with exposure. • A marked influence by the variable Cu contents on metal composition was recorded. • Digestive gland metallothionein is a poor biomarker for Cd exposure. - Abstract: Accumulation of cadmium in aquatic invertebrates may compromise human food safety and anthropogenic additions of cadmium to coastal areas cause concern. Induction of crustacean metallothionein has been suggested as a useful biomarker for contamination of the aquatic environment with cadmium. We investigated how exposure to low concentrations of cadmium in the food affects the subcellular binding of cadmium with the shore crab Carcinus maenas as model organism. Approximately 80% of the assimilated cadmium was bound in the soluble fraction of the midgut gland and of this, 82% was found in the metallothionein fraction. Metallothionein synthesis was only induced at the highest exposure level. However, the number of cadmium atoms bound per molecule of metallothionein increased linearly with exposure, from approximately 0.18 in the control group to 1.4 in a group administered food containing 5.1 μg Cd g⁻¹. We noted a marked interaction between the presence of copper and zinc in the midgut gland and the binding of cadmium. The usefulness of crustacean midgut gland metallothionein as a biomarker for cadmium exposure at modest levels was questioned since exposures at levels producing significant increases in the tissue contents of the metal did not result in elevated concentrations of metallothionein in the midgut gland.

  17. CNS wound healing is severely depressed in metallothionein I- and II-deficient mice

    DEFF Research Database (Denmark)

    Penkowa, M; Carrasco, J; Giralt, M

    1999-01-01

    To characterize the physiological role of metallothioneins I and II (MT-I+II) in the brain, we have examined the chronological effects of a freeze injury to the cortex in normal and MT-I+II null mice. In normal mice, microglia/macrophage activation and astrocytosis were observed in the areas surr...

  18. Metallothionein-1+2 protect the CNS after a focal brain injury

    DEFF Research Database (Denmark)

    Giralt, Mercedes; Penkowa, Milena; Lago, Natalia;

    2002-01-01

    We have evaluated the physiological relevance of metallothionein-1+2 (MT-1+2) in the CNS following damage caused by a focal cryolesion onto the cortex. In comparison to normal mice, transgenic mice overexpressing the MT-1 isoform (TgMTI* mice) showed a significant decrease of the number of activa...

  19. Purification and characterization of a cadmium-induced metallothionein from the shore crab Carcinus maenas (L.)

    DEFF Research Database (Denmark)

    Pedersen, K L; Pedersen, S N; Højrup, P

    1994-01-01

    Two metallothionein variants were purified from the midgut gland of crabs (Carcinus maenas) exposed to a high cadmium concentration (2 p.p.m.). One of the variants was purified from crabs exposed to a low cadmium concentration (0.5 p.p.m.). The purification method involved acetone precipitation, ...

  20. Metallothionein 1+2 protect the CNS during neuroglial degeneration induced by 6-aminonicotinamide

    DEFF Research Database (Denmark)

    Penkowa, Milena; Giralt, Mercedes; Camats, Jordi

    2002-01-01

    6-Aminonicotinamide (6-AN) is a niacin antagonist, which leads to degeneration of gray matter astrocytes. Metallothionein 1+2 (MT-1+2) are neuroprotective factors in the central nervous system (CNS), and to determine the roles for MT after 6-AN, we have examined transgenic mice overexpressing MT-1...

  1. The aryl hydrocarbon receptor and glucocorticoid receptor interact to activate human metallothionein 2A

    Energy Technology Data Exchange (ETDEWEB)

    Sato, Shoko, E-mail: satosho@rs.tus.ac.jp [Laboratory of Nutrition, Graduate School of Agricultural Science, Tohoku University, Sendai 981-8555 (Japan); Shirakawa, Hitoshi, E-mail: shirakah@m.tohoku.ac.jp [Laboratory of Nutrition, Graduate School of Agricultural Science, Tohoku University, Sendai 981-8555 (Japan); Tomita, Shuhei, E-mail: tomita@med.tottori-u.ac.jp [Division of Molecular Pharmacology, Department of Pathophysiological and Therapeutic Science, Yonago 683-8503 (Japan); Tohkin, Masahiro, E-mail: tohkin@phar.nagoya-cu.ac.jp [Department of Medical Safety Science, Graduate School of Pharmaceutical Science, Nagoya City University, Nagoya 267-8603 (Japan); Gonzalez, Frank J., E-mail: gonzalef@mail.nih.gov [Laboratory of Metabolism, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892 (United States); Komai, Michio, E-mail: mkomai@m.tohoku.ac.jp [Laboratory of Nutrition, Graduate School of Agricultural Science, Tohoku University, Sendai 981-8555 (Japan)

    2013-11-15

    Although the aryl hydrocarbon receptor (AHR) and glucocorticoid receptor (GR) play essential roles in mammalian development, stress responses, and other physiological events, crosstalk between these receptors has been the subject of much debate. Metallothioneins are classic glucocorticoid-inducible genes that were reported to increase upon treatment with AHR agonists in rodent tissues and cultured human cells. In this study, the mechanism of human metallothionein 2A (MT2A) gene transcription activation by AHR was investigated. Cotreatment with 3-methylcholanthrene and dexamethasone, agonists of AHR and GR respectively, synergistically increased MT2A mRNA levels in HepG2 cells. MT2A induction was suppressed by RNA interference against AHR or GR. Coimmunoprecipitation experiments revealed a physical interaction between AHR and GR proteins. Moreover, chromatin immunoprecipitation assays indicated that AHR was recruited to the glucocorticoid response element in the MT2A promoter. Thus, we provide a novel mechanism whereby AHR modulates expression of human MT2A via the glucocorticoid response element and protein–protein interactions with GR. - Highlights: • Aryl hydrocarbon receptor forms a complex with glucocorticoid receptor in cells. • Human metallothionein gene is regulated by the AHR and GR interaction. • AHR–GR complex binds to glucocorticoid response element in metallothionein gene. • We demonstrated a novel transcriptional mechanism via AHR and GR interaction.

  2. Metallothionein Zn(2+)- and Cu(2+)-clusters from first-principles calculations

    DEFF Research Database (Denmark)

    Greisen, Per Junior; Jespersen, Jakob Berg; Kepp, Kasper Planeta

    2012-01-01

    Detailed electronic structures of Zn(ii) and Cu(ii) clusters from metallothioneins (MT) have been obtained using density functional theory (DFT), in order to investigate how oxidative stress-caused Cu(ii) intermediates affect Zn-binding to MT and cooperatively lead to Cu(i)MT. The inferred accura...

  3. Expression of metallothionein-I, -II, and -III in Alzheimer disease and animal models of neuroinflammation

    DEFF Research Database (Denmark)

    Hidalgo, Juan; Penkowa, Milena; Espejo, Carmen

    2006-01-01

    In recent years it has become increasingly clear that the metallothionein (MT) family of proteins is important in neurobiology. MT-I and MT-II are normally dramatically up-regulated by neuroinflammation. Results for MT-III are less clear. MTs could also be relevant in human neuropathology. In Alz...

  4. Unexpected Interactions of the Cyanobacterial Metallothionein SmtA with Uranium.

    Science.gov (United States)

    Acharya, Celin; Blindauer, Claudia A

    2016-02-15

    Molecules for remediating or recovering uranium from contaminated environmental resources are of high current interest, with protein-based ligands coming into focus recently. Metallothioneins either bind or redox-silence a range of heavy metals, conferring protection against metal stress in many organisms. Here, we report that the cyanobacterial metallothionein SmtA competes with carbonate for uranyl binding, leading to formation of heterometallic (UO2)(n)Zn4SmtA species, without thiol oxidation, zinc loss, or compromising secondary or tertiary structure of SmtA. In turn, only metalated and folded SmtA species were found to be capable of uranyl binding. (1)H NMR studies and molecular modeling identified Glu34/Asp38 and Glu12/C-terminus as likely adventitious, but surprisingly strong, bidentate binding sites. While it is unlikely that these interactions correspond to an evolved biological function of this metallothionein, their occurrence may offer new possibilities for designing novel multipurpose bacterial metallothioneins with dual ability to sequester both soft metal ions including Cu(+), Zn(2+), Cd(2+), Hg(2+), and Pb(2+) and hard, high-oxidation state heavy metals such as U(VI). The concomitant protection from the chemical toxicity of uranium may be valuable for the development of bacterial strains for bio-remediation.

  5. GENETIC BACKGROUND BUT NOT METALLOTHIONEIN PHENOTYPE DICTATES SENSITIVITY TO CADMIUM-INDUCED TESTICULAR INJURY IN MICE

    Science.gov (United States)

    Genetic Background but not Metallothionein Phenotype Dictates Sensitivity to Cadmium-Induced Testicular Injury in MiceJie Liu1,2, Chris Corton3, David J. Dix4, Yaping Liu1, Michael P. Waalkes2 and Curtis D. Klaassen1 ABSTRACTParenteral administrati...

  6. Teratogenicity of cadmium-metallothionein in the rat

    Energy Technology Data Exchange (ETDEWEB)

    Webb, M.; Holt, D.; Brown, N.; Hard, G.C.

    1988-06-01

    A single dose in the range 0.25-1.9 mg metallothionein-bound cadmium (MT-Cd)/kg body weight, when administered parenterally to the rat between day 8 and day 14 of gestation, is teratogenic. In vitro, the development of the isolated rat conceptus is unaffected by the addition of 1.5 ..mu..M MT-Cd to the culture medium whereas the same concentration of ionic Cd (as CdCl/sub 2/) is lethal. At short times after injection of 0.25 mg MT-Cd/kg body weight on gd 12, the maximal foetal and placental contents of Cd are low in comparison with those after a teratogenic dose of CdCl/sub 2/ and are of the same order as those in the embryo and placenta + yolk sac of the rat conceptus, cultured in the presence of the highest no-effect concentration of CdCl/sub 2/. From this evidence, it is concluded that the uptake by the conceptus in vivo of either CdMT, or of Cd liberated therefrom, is unlikely to contribute to the teratogenic response. In the pregnant, as in the non-pregnant rat, the kidney appears to be the only organ that is affected directly by the metalloprotein. All doses in the range 0.25-1.0 mg MT-Cd/kg body weight are nephrotoxic and result in prolonged anorexia in the pregnant animal. While some of the foetal deformities that occur in the CdMT-dosed animal seem to be direct consequences of the renal dysfunction, others apparently are secondary to the maternal anorexia. In rats that are injected i.p on gd 12 with 0.25 mg MT-Cd/kg renal uptake of Cd is slower, but the final concentration is higher than in animals that are given the same dose i.v. At this and the higher dose levels structural and/or functional damage to the kidneys also is greater in i.p.-, than in i.v.-dosed animals. The incidence of foetal malformations, however, is similar in the i.p. and i.v. groups and varies little over the dose range.

  7. Metallothionein - immunohistochemical cancer biomarker: a meta-analysis.

    Directory of Open Access Journals (Sweden)

    Jaromir Gumulec

    Full Text Available Metallothionein (MT has been extensively investigated as a molecular marker of various types of cancer. In spite of the fact that numerous reviews have been published in this field, no meta-analytical approach has been performed. Therefore, results of to-date immunohistochemistry-based studies were summarized using meta-analysis in this review. Web of science, PubMed, Embase and CENTRAL databases were searched (up to April 30, 2013 and the eligibility of individual studies and heterogeneity among the studies was assessed. Random and fixed effects model meta-analysis was employed depending on the heterogeneity, and publication bias was evaluated using funnel plots and Egger's tests. A total of 77 studies were included with 8,015 tissue samples (4,631 cases and 3,384 controls. A significantly positive association between MT staining and tumors (vs. healthy tissues was observed in head and neck (odds ratio, OR 9.95; 95% CI 5.82-17.03 and ovarian tumors (OR 7.83; 1.09-56.29, and a negative association was ascertained in liver tumors (OR 0.10; 0.03-0.30. No significant associations were identified in breast, colorectal, prostate, thyroid, stomach, bladder, kidney, gallbladder, and uterine cancers and in melanoma. While no associations were identified between MT and tumor staging, a positive association was identified with the tumor grade (OR 1.58; 1.08-2.30. In particular, strong associations were observed in breast, ovarian, uterine and prostate cancers. Borderline significant association of metastatic status and MT staining were determined (OR 1.59; 1.03-2.46, particularly in esophageal cancer. Additionally, a significant association between the patient prognosis and MT staining was also demonstrated (hazard ratio 2.04; 1.47-2.81. However, a high degree of inconsistence was observed in several tumor types, including colorectal, kidney and prostate cancer. Despite the ambiguity in some tumor types, conclusive results are provided in the tumors of

  8. Determination of the Cd/S cluster stoichiometry in Fucus vesiculosus metallothionein.

    Science.gov (United States)

    Merrifield, Maureen E; Chaseley, Jennifer; Kille, Peter; Stillman, Martin J

    2006-03-01

    The seaweed Fucus vesiculosus is unusual when compared with other algal species, in that it can survive in toxic-metal-contaminated aquatic environments. The metallothionein gene has been identified in F. vesiculosus by Kille and co-workers (Morris, C. A., Nicolaus, B., Sampson, V., Harwood, J. L., and Kille, P. (1999) Biochem. J. 338, 553), which suggests a possible protective mechanism against toxic metals for this species. We report the first detailed study of the metal binding properties of F. vesiculosus metallothionein using UV absorption, circular dichroism (CD), and electrospray mass spectral techniques. The overall metal-to-sulfur ratios of this novel algal protein when bound to divalent cadmium and zinc were determined to be Cd(6)S(16) and Zn(6)S(16), respectively. Mixed Cd/Zn species were also formed when Cd(2+) was added to the Zn-containing Fucus metallothionein. Only one conformation was identified at low pH for the native protein. Analysis of the UV absorption, CD, and ESI-MS spectral data recorded during stepwise, acid-induced demetalation supports a two-domain structure for the protein, with two 3-metal binding sites. The data suggest that one of the domains is significantly less stable than the other, and we tentatively propose from the arrangement of cysteines in the sequence that the two domains are M(3)S(7) and M(3)S(9) (where M = Cd(2+) or Zn(2+)). While the M(3)S(9) cluster is known in the beta clusters of crab, lobster, and mammalian metallothioneins, the M(3)S(7) is a hitherto unknown cluster structure. Metallothionein in F. vesiculosus is thought to act as a protective mechanism against incoming toxic metals. The metal binding studies reported are a putative model for metal binding in vivo.

  9. Expression of the zinc transporters genes and metallothionein in obese women.

    Science.gov (United States)

    dos Santos Rocha, Paula Beatriz Krebsky; de Castro Amorim, Amanda; de Sousa, Artemizia Francisca; do Monte, Semíramis Jamil Hadad; da Mata Sousa, Luiz Claudio Demes; do Nascimento Nogueira, Nadir; Neto, José Machado Moita; do Nascimento Marreiro, Dilina

    2011-11-01

    Research has investigated the participation of zinc transport proteins and metallothionein in the metabolism of this mineral. However, studies about the genetic expression of these proteins in obese patients are scarce. The study determined the expression of zinc transporter protein codifying genes (ZnT-1, Zip-1 and Zip-3) and of metallothionein in 55 obese women, aged between 20 and 56 years. The assessment of body composition was carried out using anthropometric measurements and bioelectrical impedance. Zinc intake was obtained by recording diet over a 3-day period, and the nutritional analysis was carried out using NutWin software version 1.5. The plasmatic and erythrocytary zinc were analyzed by atomic absorption spectrophotometry (λ=213. 9 nm). The determination of mRNA expression of the zinc transporter proteins and metallothionein was carried out using blood, using the RT-PCR method. The mean values of body mass index were 37.9±5.5 kg/m2. The average intake of zinc was 9.4±2.3 mg/day. The analysis of the zinc plasma concentrations showed values of 58.4±10.9 μg/dL. The mean values of zinc in the erythroytes were 38.7±9.1 μg/g Hb. The metallothionein gene had a higher expression in the blood, when compared to zinc transporters ZnT-1, Zip-1, and Zip-3 (p=0.01). The study shows that there are alterations in the biochemical parameters of zinc in obese patients assessed, as well as higher expression of the codifying gene metallothionein, when compared to the investigated zinc transporters.

  10. Characterization of mercury bioremediation by transgenic bacteria expressing metallothionein and polyphosphate kinase

    Directory of Open Access Journals (Sweden)

    Gonzalez-Ruiz Gloriene

    2011-08-01

    Full Text Available Abstract Background The use of transgenic bacteria has been proposed as a suitable alternative for mercury remediation. Ideally, mercury would be sequestered by metal-scavenging agents inside transgenic bacteria for subsequent retrieval. So far, this approach has produced limited protection and accumulation. We report here the development of a transgenic system that effectively expresses metallothionein (mt-1 and polyphosphate kinase (ppk genes in bacteria in order to provide high mercury resistance and accumulation. Results In this study, bacterial transformation with transcriptional and translational enhanced vectors designed for the expression of metallothionein and polyphosphate kinase provided high transgene transcript levels independent of the gene being expressed. Expression of polyphosphate kinase and metallothionein in transgenic bacteria provided high resistance to mercury, up to 80 μM and 120 μM, respectively. Here we show for the first time that metallothionein can be efficiently expressed in bacteria without being fused to a carrier protein to enhance mercury bioremediation. Cold vapor atomic absorption spectrometry analyzes revealed that the mt-1 transgenic bacteria accumulated up to 100.2 ± 17.6 μM of mercury from media containing 120 μM Hg. The extent of mercury remediation was such that the contaminated media remediated by the mt-1 transgenic bacteria supported the growth of untransformed bacteria. Cell aggregation, precipitation and color changes were visually observed in mt-1 and ppk transgenic bacteria when these cells were grown in high mercury concentrations. Conclusion The transgenic bacterial system described in this study presents a viable technology for mercury bioremediation from liquid matrices because it provides high mercury resistance and accumulation while inhibiting elemental mercury volatilization. This is the first report that shows that metallothionein expression provides mercury resistance and

  11. NF-κB在氯胺酮诱导大鼠原代培养大脑皮层神经元凋亡中的作用%Role of NF-κB in ketamine-induced apoptosis in rat cortical neurons in vitro

    Institute of Scientific and Technical Information of China (English)

    王娴; 周志强; 杨建军; 徐建国

    2010-01-01

    Objective To investigate the role of NF-κB in ketamine-induced apoptosis in rat cortical neurons in vitro. Methods Cortical neurons were isolated from newborn pathogen-free SD rats according to the method described by Kimney. After being cultured for 6 days the cortical neurons were randomly divided into 3 groups:group Ⅰ control (C); group Ⅱ ketamine (K) and group Ⅲ ketamine + SN50 (NF-κB translocation inhibitor) (KS). The cortical neurons were exposed to ketamine l mmol/L and ketamine 1 mmol/L + SN502.5 μmol/L for 24 h in group K and KS respectively. Twenty-four hours after the drugs were washed out, NF-κB activity, the expression of Bcl-XL and Bax and apoptosis in the neurons were measured. The ultrastructure of the neurons were examined by electron microscopy. Results NF-κB activity and the percentage of apoptotic neurons were significantly higher and Bcl-XL/Bax ratio was significantly lower in group K than in group C. There was no significant difference in NF-κB activity, Bcl-XL/Bax ratio and percentage of apoptotic neurons between group C and KS. Conclusion NF-κB is involved in ketamine-induced apoptosis in contical neurons in vitro.%目的 评价NF-κB在氯胺酮诱导大鼠原代培养大脑皮层神经元凋亡中的作用.方法 体外培养6 d的大鼠大脑皮层神经元15孔,随机分为3组(n=5):对照组(C组)正常培养;氯胺酮组(K组)和氯胺酮+NF-κB抑制剂SN50组(KS组)分别加入1 mmol/L氯胺酮、1 mmol/L氯胺酮+2.5 μmol/L SN50,作用24 h.洗脱后24 h时采用凝胶迁移实验检测神经元NF-κB活性,采用Westernblot法测定凋亡相关蛋白Bcl-XL和Bax的表达,计算Bcl-XL/Bax,计数凋亡神经元,电镜下观察神经元的超微结构.结果 与C组相比,K组NF-κB活性、Bcl-XL/Bax、神经元凋亡率升高(P<0.05),KS组上述指标差异无统计学意义(P>0.05);与K组相比,KS组NF-κB活性、Bcl-XL/Bax、神经元凋亡率降低(P<0.05).电镜结果显示:KS组神经元损伤较K

  12. 丹参注射液对缺氧神经干细胞凋亡和Caspase-3活性的影响%Effects of salvia miltiorrhizae injection on hypoxia-induced apoptosis or cultured rat neuronal stem cells and activity of Caspase-3

    Institute of Scientific and Technical Information of China (English)

    黄涛; 韩富; 张志强; 谭齐家; 谢才军; 谢绍盈; 朱灿辉

    2008-01-01

    Objective To explore the effects of salvia miltiorrhizae (SM) injection on the apoptosis of cultured rat neuronal stem cells induced by hypoxia and the activity of Caspase-3, in order to provide the further evidence for the molecular mechanism of neuroprotection of SM injection. Methods The neuronal stem cells from neonatal rat hippocampus were cultured and divided randomly into normal control group, hypoxia group and SM treatment group. After Hoechst staining, the apoptotic morphological change and apoptosis percentage were observed under fluorescence microscope. The activities of Caspase-3 in the 3 groups were evaluated by the colorimetric assay. Results Compared with normal control group [(2.75±0.28)%, 1.16±0.07], the percentage of apoptosis and the activity of Caspase-3 were increased significantly in neuronal stem cells cultured in hypoxia [(30.12%±2.09)%,3.85±0.41, P<0.05). Application of SM injection reduced markedly the percentage of apoptosis and the activity of Caspase-3 of the neuronal stem cells cultured in hypoxia [(9.16±1.34)%, 1.50±0.09, P<0.05].Conclusion SM injection can depress the apoptosis of the rat neuronal stem cells induced by hypoxia,so as to exert the neuroprotection.%目的 探讨丹参注射液对缺氧培养大鼠神经干细胞的凋亡及Caspase-3活性的影响,以进一步明确丹参注射液神经保护作用的分子机制.方法 体外培养新生大鼠海马神经干细胞,将其分为正常对照组,缺氧培养组及丹参注射液处理组.Hoechst染色后荧光显微镜下观察并计算细胞凋亡率:比色法检测各组细胞Caspase-3的相对活性.结果 缺氧培养大鼠神经干细胞的细胞凋亡率(30.12%±2.09%)及Caspase-3活性(3.85±0.41)均较正常对照组(2.75%±0.28%,1.16±0.07)明显升高,差异有统计学意义(P<0.05);施加丹参注射液后,大鼠神经干细胞的细胞凋亡率(9.16%±1.34%)和Caspase-3活性(1.50±0.09)均较缺氧培养组明显下降,差异有统计学意义(P<0

  13. Incorporation of sulfide ions into the cadmium(II) thiolate cluster of Cicer arietinum metallothionein2.

    Science.gov (United States)

    Wan, Xiaoqiong; Freisinger, Eva

    2013-01-18

    The plant metallothionein2 from Cicer arietinum (chickpea), cic-MT2, is known to coordinate five divalent metal ions such as Zn(II) or Cd(II), which are arranged in a single metal thiolate cluster. When the Zn(II) form of the protein is titrated with Cd(II) ions in the presence of sulfide ions, an increased Cd(II) binding capacity and concomitant incorporation of sulfide ions into the cluster are observed. The exact stoichiometry of this novel cluster, its spectroscopic properties, and the significantly increased pH stability are analyzed with different techniques, including UV and circular dichroism spectroscopy and colorimetric assays. Limited proteolytic digestion provides information about the spacial arrangement of the cluster within the protein. Increasing the Cd(II) scavenging properties of a metallothionein by additionally recruiting sulfide ions might be an economic and very efficient detoxification strategy for plants.

  14. Change in metallothionein phosphorylation state in Mya arenaria clams: implication in metal metabolism and oxidative stress

    OpenAIRE

    Gagné, F.; M Gélinas; Gagnon, C.; André, C.; C Blaise

    2010-01-01

    The contamination of the benthic environment poses a threat to long-lived sessile organisms such as clams. The purpose of this study was to investigate metal contamination in tissues and changes in metallothioneins (MT) in respect to its redox status in Mya arenaria clams collected at three polluted sites. The phosphorylation state of MT was also investigated to determine whether this state is changed in clams collected at heavy-metal contaminated site and its involvement in cytoprotective si...

  15. Metallothionein-1 and nitric oxide expression are inversely correlated in a murine model of Chagas disease

    OpenAIRE

    2014-01-01

    Chagas disease, caused by Trypanosoma cruzi, represents an endemic among Latin America countries. The participation of free radicals, especially nitric oxide (NO), has been demonstrated in the pathophysiology of seropositive individuals with T. cruzi. In Chagas disease, increased NO contributes to the development of cardiomyopathy and megacolon. Metallothioneins (MTs) are efficient free radicals scavengers of NO in vitro and in vivo. Here, we developed a murine model of the chronic phase of C...

  16. Metallothioneins, Unconventional Proteins from Unconventional Animals: A Long Journey from Nematodes to Mammals †

    OpenAIRE

    Gloria Isani; Emilio Carpenè

    2014-01-01

    Metallothioneins (MTs) are ubiquitous low molecular weight cysteine-rich proteins characterized by high affinity for d10 electron configuration metals, including essential (Zn and Cu) and non-essential (Cd and Hg) trace elements. The biological role of these ancient and well-conserved multifunctional proteins has been debated since MTs were first discovered in 1957. Their main hypothesized functions are: (1) homeostasis of Zn and Cu; (2) detoxification of Cd, and Hg; and (3) free radical scav...

  17. Characterization of mercury bioremediation by transgenic bacteria expressing metallothionein and polyphosphate kinase

    OpenAIRE

    Gonzalez-Ruiz Gloriene; Alvarez Derry; Ruiz Oscar N; Torres Cesar

    2011-01-01

    Abstract Background The use of transgenic bacteria has been proposed as a suitable alternative for mercury remediation. Ideally, mercury would be sequestered by metal-scavenging agents inside transgenic bacteria for subsequent retrieval. So far, this approach has produced limited protection and accumulation. We report here the development of a transgenic system that effectively expresses metallothionein (mt-1) and polyphosphate kinase (ppk) genes in bacteria in order to provide high mercury r...

  18. Metalloselenonein, the selenium analogue of metallothionein: synthesis and characterization of its complex with copper ions.

    OpenAIRE

    Oikawa, T; Esaki, N; Tanaka, H.; Soda, K

    1991-01-01

    We used an automated peptide synthesizer to produce a peptide, metalloselenonein, that contains selenocysteine residues substituted for all cysteine residues in Neurospora crassa copper metallothionein. Metalloselenonein binds 3 mol of Cu(I) per mol. This adduct shows a broad absorption band between 230 and 400 nm and a fluorescence band at 395 nm, which can be attributed to copper-selenolate coordination. The circular dichroism spectrum of the copper-metalloselenonein complex shows a positiv...

  19. Quantitative immunodetection of metallothioneins in relation to metals concentration in spiders from variously polluted areas.

    Science.gov (United States)

    Babczyńska, Agnieszka; Wilczek, Grażyna; Szulińska, Elżbieta; Franiel, Izabella

    2011-09-01

    Spiders inhabiting post industrial environments, such as waste heaps or ore-bearing areas, are exposed to high concentrations of metals, accumulated in the body of their prey and transferred along food chains. Therefore spiders are pressed to develop metal-neutralization strategies. Low-molecular, multifunction proteins: metallothioneins (MTs), often postulated as biomarkers of metal exposure, are known to bind metals and thus protect organisms against their toxic effects. Yet the proteins are still not well recognized in spiders. The aim of this study was to assess, by immunodetection method, ELISA, the concentration of metallothioneins in adult females of three web building spider species: Araneus diadematus (Araneidae), Agelena labyrinthica (Agelenidae) and Linyphia triangularis (Linyphiidae) from three variously polluted areas in southern Poland: Olkusz, ore-bearing post industrial site; Katowice-WeŁnowiec: post metallurgic waste heap, Pilica: the reference, rural, area. The concentration of metallothioneins has been analyzed in relation to the metal concentration in spiders body. The study gives the evidence that metallothioneins are reliably detectable by means of ELISA technique. The analysis of results obtained shows a strong species-dependence of the MTs level. Positive correlations between MTs concentration and metal body burden (mainly Zn and Pb) were found. This suggests that the proteins play an important role in the neutralization and regulation of metal ions in spiders. The same correlation indicate the possibility to consider MTs in spiders as biomarkers of metal exposure and effects. However, the species specificity as well as metal characteristics should be taken under account. Copyright © 2011 Elsevier Inc. All rights reserved.

  20. Alterations in prey capture and induction of metallothioneins in grass shrimp fed cadmium-contaminated prey

    Energy Technology Data Exchange (ETDEWEB)

    Wallace, W.G.; Hoexum Brouwer, T.M.; Brouwer, M.; Lopez, G.R.

    2000-04-01

    The aquatic oligochaete Limnodrilus hoffmeisteri from a Cd-contaminated cove on the Hudson River, Foundry Cove, New York, USA, has evolved Cd resistance. Past studies have focused on how the mode of detoxification of Cd by these Cd-resistant worms influences Cd trophic transfer to the grass shrimp Palaemonetes pugio. In the present study, the authors investigate reductions in prey capture in grass shrimp fed Cd-contaminated prey. They also investigate the induction of metal-binding proteins, metallothioneins, in these Cd-exposed shrimp. Grass shrimp were fed field-exposed Cd-contaminated Foundry Cove oligochaetes or laboratory-exposed Cd-contaminated Artemia salina. Following these exposures, the ability of Cd- dosed and control shrimp to capture live A. salina was compared. Results show that shrimp fed laboratory-exposed Cd-contaminated A. salina for 2 weeks exhibit significant reductions in their ability to successfully capture prey (live A. salina). Reductions in prey capture were also apparent, though not as dramatic in shrimp fed for 1 week on field-exposed Cd-contained Foundry Cove oligochaetes. Shrimp were further investigated for their subcellular distribution of Cd to examine if alterations in prey capture could be linked to saturation of Cd-metallothionein. Cd-dosed shrimp produced a low molecular weight CD-binding metallothionein protein in a dose- and time-dependent manner. Most importantly, successful prey capture decreased with increased Cd body burdens and increased Cd concentration bound to high molecular weight proteins.

  1. The liver in itai-itai disease (chronic cadmium poisoning): pathological features and metallothionein expression.

    Science.gov (United States)

    Baba, Hayato; Tsuneyama, Koichi; Yazaki, Megumi; Nagata, Kohei; Minamisaka, Takashi; Tsuda, Tatsuhiro; Nomoto, Kazuhiro; Hayashi, Shinichi; Miwa, Shigeharu; Nakajima, Takahiko; Nakanishi, Yuko; Aoshima, Keiko; Imura, Johji

    2013-09-01

    Cadmium (Cd) is a highly hepatotoxic heavy metal, which is widely dispersed in the environment. Acute Cd hepatotoxicity has been well studied in experimental animals; however, effects of prolonged exposure to Cd doses on the liver remain unclear. In the present study, to evaluate chronic Cd hepatotoxicity, we examined specimens from cases of itai-itai disease, the most severe form of chronic Cd poisoning. We compared 89 cases of itai-itai disease with 27 control cases to assess Cd concentration in organs. We also examined 80 cases of itai-itai disease and 70 control cases for histopathological evaluation. In addition, we performed immunohistochemistry for metallothionein, which binds and detoxifies Cd. Hepatic Cd concentration was higher than Cd concentration in all other organs measured in the itai-itai disease group, whereas it was second highest following renal concentration in the control group. In the liver in the itai-itai disease group, fibrosis was observed at a significantly higher rate than that in the control group. Metallothionein expression was significantly higher in the itai-itai disease group than that in the control group. Prolonged exposure to low doses of Cd leads to high hepatic accumulation, which can then cause fibrosis; however, it also causes high expression of metallothionein, which is thought to reduce Cd hepatotoxicity.

  2. Metallothionein response in earthworms Lampito mauritii (Kinberg) exposed to fly ash

    Energy Technology Data Exchange (ETDEWEB)

    Maity, S.; Hattacharya, S.; Chaudhury, S. [Visva Bharati, Santini Ketan (India)

    2009-10-15

    Among pollutants, the coal fly ash occupies a significant position in industrial wastes. The fly ash matrix is a complex mixture of various organic (polyhalogenated compounds) and inorganic (Si, Al, Fe, As, Cd, Bi, Hg, etc.) chemicals. The application of fly ash for agricultural purposes and as landfills may lead to the contamination of the land with some of the toxic chemical compounds present in fly ash. Thus prior to the application of fly ash for developmental activities, it requires bio-monitoring and risk characterization. In order to achieve this objective adult Lampito mauritii were exposed to different proportions of fly ash in soil for 30 d and the concentrations of metallothionein in earthworm were assessed. The results revealed that up to 50% of fly ash amendment does not apparently harm the earthworm in respect of their survival and growth. A significant increase in tissue metallothionein level was recorded in L mauritii exposed to fly ash amended soil without tissue metal accumulation indicating that metallothionein is involved in scavenging of free radicals and reactive oxygen species metabolites. It is concluded that this biochemical response observed in L mauritii exposed to fly ash amended soil could be used in ecotoxicological field monitoring.

  3. Identification of quantum dots labeled metallothionein by fast scanning laser-induced breakdown spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Konecna, Marie [Central European Institute of Technology, Brno University of Technology, Technicka 3058/10, CZ-616 00 Brno (Czech Republic); Department of Chemistry and Biochemistry, Faculty of Agronomy, Mendel University in Brno, Zemedelska 1, CZ-613 00 Brno (Czech Republic); Novotny, Karel [Central European Institute of Technology, Masaryk University, Kamenice 753/5, CZ-625 00 Brno (Czech Republic); Krizkova, Sona [Central European Institute of Technology, Brno University of Technology, Technicka 3058/10, CZ-616 00 Brno (Czech Republic); Department of Chemistry and Biochemistry, Faculty of Agronomy, Mendel University in Brno, Zemedelska 1, CZ-613 00 Brno (Czech Republic); Blazkova, Iva [Department of Chemistry and Biochemistry, Faculty of Agronomy, Mendel University in Brno, Zemedelska 1, CZ-613 00 Brno (Czech Republic); Kopel, Pavel [Central European Institute of Technology, Brno University of Technology, Technicka 3058/10, CZ-616 00 Brno (Czech Republic); Department of Chemistry and Biochemistry, Faculty of Agronomy, Mendel University in Brno, Zemedelska 1, CZ-613 00 Brno (Czech Republic); Kaiser, Jozef [Central European Institute of Technology, Brno University of Technology, Technicka 3058/10, CZ-616 00 Brno (Czech Republic); Institute of Physical Engineering, Brno University of Technology, Technicka 2, CZ-616 69 Brno (Czech Republic); Hodek, Petr [Department of Biochemistry, Faculty of Science, Charles University in Prague, Hlavova 2030/8, CZ-128 00 Prague,Czech Republic (Czech Republic); Kizek, Rene [Central European Institute of Technology, Brno University of Technology, Technicka 3058/10, CZ-616 00 Brno (Czech Republic); Department of Chemistry and Biochemistry, Faculty of Agronomy, Mendel University in Brno, Zemedelska 1, CZ-613 00 Brno (Czech Republic); and others

    2014-11-01

    The technique described in this paper allows detection of quantum dots (QDs) specifically deposited on the polystyrene surface by laser-induced breakdown spectroscopy (LIBS). Using LIBS, the distribution of QDs or their conjugates with biomolecules deposited on the surface can be observed, regardless of the fact if they exhibit fluorescence or not. QDs deposited on the specific surface of polystyrene microplate in the form of spots are detected by determination of the metal included in the QDs structure. Cd-containing QDs (CdS, CdTe) stabilized with mercaptopropionic (MPA) or mercaptosuccinic (MSA) acid, respectively, alone or in the form of conjugates with metallothionein (MT) biomolecule are determined by using the 508.58 nm Cd emission line. The observed absolute detection limit for Cd in CdTe QDs conjugates with MT in one spot was 3 ng Cd. Due to the high sensitivity of this technique, the immunoanalysis in combination with LIBS was also investigated. Cd spatial distribution in sandwich immunoassay was detected. - Highlights: • We describe determination of biomolecules labeled with quantum dots by LIBS. • LIBS and immunoassay are applied for the determination of metallothionein. • Metallothionein amount detected by LIBS is 10-times lower compared to ELISA.

  4. Apoptosis and oxidative stress in neurodegenerative diseases.

    Science.gov (United States)

    Radi, Elena; Formichi, Patrizia; Battisti, Carla; Federico, Antonio

    2014-01-01

    Neurodegenerative disorders affect almost 30 million individuals leading to disability and death. These disorders are characterized by pathological changes in disease-specific areas of the brain and degeneration of distinct neuron subsets. Despite the differences in clinical manifestations and neuronal vulnerability, the pathological processes appear similar, suggesting common neurodegenerative pathways. Apoptosis seems to play a key role in the progression of several neurologic disorders like Alzheimer's disease, Parkinson's disease, Huntington's disease, and amyotrophic lateral sclerosis as demonstrated by studies on animal models and cell lines. On the other hand, research on human brains reported contradictory results. However, many dying neurons have been detected in brains of patients with neurodegenerative diseases, and these conditions are often associated with significant cell loss accompanied by typical morphological features of apoptosis such as chromatin condensation, DNA fragmentation, and activation of cysteine-proteases, caspases. Cell death and neurodegenerative conditions have been linked to oxidative stress and imbalance between generation of free radicals and antioxidant defenses. Multiple sclerosis, stroke, and neurodegenerative diseases have been associated with reactive oxygen species and nitric oxide. Here we present an overview of the involvement of neuronal apoptosis and oxidative stress in the most important neurodegenerative diseases, mainly focusing the attention on several genetic disorders, discussing the interaction between primary genetic abnormalities and the apoptotic pathways.

  5. 依托咪酯乳剂对老年大鼠海马神经元凋亡及突触素表达的影响%Effects of etomidate emulsion on neuronal apoptosis and synaptophysin expression in aged rat hippocampus

    Institute of Scientific and Technical Information of China (English)

    冯舒韵; 杨承祥; 赵伟成; 陈立成; 郑雪琴; 李云

    2016-01-01

    Objective To investigate the effects of etomidate emulsion on neuronal apoptosis and synaptophysin expression in aged rat hippocampus.Methods Forty-five male and female SD rats aged 18 months were randomly divided into 3 groups (n=15):NS group(N group),Etomidate group(E group),Lipid group(L group).On the 1st day to 3rd day,animal received intraperitoneal (IP) injection of either etomidate 20 mg/kg,or lipid 20 mg/kg,or same volume NS depending group division once a day for 3 d.On the 4th day,all of the animals were killed and the brains were immediately removed for determination of neuronal apoptosis and synaptophysin expression in the hippocampus.Results The numbers of apoptosis cells and apoptosis index in hippocampus determined by TUNEL assay were not significant different among 3 groups (P>0.05).No significant difference was found in the expression of synaptophysin among 3 groups (P>0.05).The expression of activated caspase-3 was also not significantly different among 3 groups(P>0.05).Conclusions Etomidate(20 mg/kg) does not affect the neuronal apoptosis and synaptophysin expression in aged rat hippocampus.%目的 观察腹腔注射依托咪酯乳剂对老年大鼠海马神经元突触素表达的影响. 方法 选择18月龄的SD大鼠45只,用完全随机分组法分为3组(每组15只):生理盐水组(N组)、依托咪酯组(E组)、脂肪乳组(L组).实验第1~3天,E组腹腔注射依托咪酯(乳剂)20 mg/kg,L组腹腔注射20%脂肪乳20 mg/kg,N组腹腔注射同等体积的生理盐水,每天1次,连续3d.实验第4天,处死大鼠,取右侧脑海马组织,固定,石蜡切片,进行细胞凋亡TUNEL检验、天冬氨酸特异性半胱氨酸蛋白酶(cysteinyl aspartate specific proteinase-3,caspase-3)Western blot检测及突触素免疫组化检验. 结果 TUNEL结果:海马CA1区神经元凋亡,与N组、L组比较,E组海马神经元凋亡细胞及凋亡指数无明显增加,差异无统计学意义(P>0.05).caspase-3表达结果:海马CA1

  6. Trace metals and over-expression of metallothioneins in bladder tumoral lesions: a case-control study

    Directory of Open Access Journals (Sweden)

    Cymbron Teresa

    2009-07-01

    Full Text Available Abstract Background Previous studies have provided some evidence of a possible association between cancer and metallothioneins. Whether this relates to an exposure to carcinogenic metals remains unclear. Methods In order to examine the association between the expression of metallothioneins and bladder tumors, and to compare the levels of arsenic, cadmium, chromium, lead and nickel in animals with bladder tumors and animals without bladder tumors, 37 cases of bovine bladder tumors and 17 controls were collected. The detection and quantification of metallothioneins in bladder tissue of both cases and controls was performed by immunohistochemistry. And the quantification of metals in tissue and hair was assessed by inductively coupled plasma – mass spectrometry. Results Increased expression of metallothioneins was associated with bladder tumors when compared with non-tumoral bladder tissue (OR = 9.3, 95% CI: 1.0 – 480. The concentrations of cadmium, chromium, lead and nickel in hair of cases were significantly higher than those of controls. However, as for the concentration of metals in bladder tissue, the differences were not significant. Conclusion Though the sample size was small, the present study shows an association between bladder tumors and metallothioneins. Moreover, it shows that concentrations of metals such as cadmium, chromium, lead and nickel in hair may be used as a biomarker of exposure.

  7. Effects of Citicoline on the Rat's cognitive Function, Expression of Hippocampal Neuron BCL-2 and BAX and Apoptosis After Cerebral Dschemia Injury%胞二磷胆碱对大鼠脑缺血损伤后认知功能及海马神经元BCL-2,BAX表达和细胞凋亡的影响

    Institute of Scientific and Technical Information of China (English)

    孟勇; 杨建华; 苏子锐

    2013-01-01

    探讨胞二磷胆碱对脑缺血损伤后大鼠认知功能和海马神经元BCL-2,BAX表达的影响.取健康Sprague-Dawley成年雄性大鼠60只,随机分为假手术对照组,缺血组和胞二磷胆碱组3组,每组20只.对缺血组和胞二磷胆碱组采用Zea Longa等法改良复制大脑中动脉狭窄(middle cerebral artery occlusion,MCAO)动物模型,通过Morris水迷宫系统观察各组大鼠学习记忆功能;免疫组化检测BCL-2,BAX表达,Tunel法检测神经元凋亡.胞二磷胆碱使Bcl-2蛋白表达从缺血对照组的39.88±5.41增加至55.13±8.17,BAX蛋白表达从缺血对照组的62.38±8.47下降至36.13±5.94,凋亡神经元从缺血对照组的18.67±3.86下降至14.67±4.25.胞二磷胆碱对脑缺血损伤后大鼠的神经保护作用可能是通过上调海马神经元Bcl-2蛋白、下调Bax蛋白的表达,减少海马神经元凋亡进而改善脑缺血损伤后大鼠的学习记忆能力.%This paper investigates the effects of citicoline on the rat's cognitive function and expression of hippocampal neuron BCL-2 and BAX after cerebral ischemia injury.With the equal and random principle,sixty healthy male adult SpragueDawley rats were divided into three groups which include sham operation control group,and ischemia group and citicoline group,each group of 20.In the last two groups,middle cerebral artery occlusion model was established by the improved method of Zea Longa.Then we evaluate each group's learning and memory function through Morris water maze test,Immunohistochemistry was used to detect BCL-2 and BAX expression,and Tunel method to detect neuron apoptosis.Compared with the ischemia group,the protein expression of BCL-2 increased from (39.88±5.41) to (55.13±8.17),the protein expression of BAX decreased from (62.38±8.47) to (36.13±5.94),and the apoptosis of neurons dropped from (18.67±3.86) to (14.67±4.25) in the citicoline group.Citicoline can improve the learning and memory function of rats after cerebral

  8. Time window characteristics of cultured rat hippocampal neurons subjected to ischemia and reperfusion

    Institute of Scientific and Technical Information of China (English)

    XU Zhong; XU Ru-xiang; LIU Bao-song; JIANG Xiao-dan; HUANG Tao; DING Lian-shu; YUAN Jun

    2005-01-01

    Objective: To explore cell death and apoptosis in rat hippocampal neurons at different time points after ischemia, hypoxia and reperfusion injury and to elucidate time window characteristics in ischemia neuronal injury.Methods: Hippocampal neurons were obtained from rat embryo and were cultured in vitro. The ischemia and reperfusion of cultured rat hippocampal neurons were simulated by oxygen-glucose deprivation (OGD) and recovery. OGD at different time points (0.25 h to 3.0 h) and then the same recovery (24 h) were prepared. Annexin V-PI staining and flow cytometry examined neuron death and apoptosis at different time after injury. Results: After OGD and recovery, both necrosis and apoptosis were observed. At different times after OGD, there were statistically significant differences in neuron necrosis rate (P0.05). At recovery, survival rate of hippocampal neurons further decreased while apoptosis rate increased. Furthermore, apoptosis rates of different time differed greatly (P<0.05). Apoptosis rate gradually increased with significant difference among those of different time points (P<0.05). However, 2 h after ischemia, apoptosis rate decreased markedly.Conclusions: Apoptosis is an important pathway of delayed neuron death. The therapeutic time window should be within 2 h after cerebral ischemia and hypoxia.

  9. A novel perspective on neuron study: damaging and promoting effects in different neurons induced by mechanical stress.

    Science.gov (United States)

    Wang, Yazhou; Wang, Wei; Li, Zong; Hao, Shilei; Wang, Bochu

    2016-10-01

    A growing volume of experimental evidence demonstrates that mechanical stress plays a significant role in growth, proliferation, apoptosis, gene expression, electrophysiological properties and many other aspects of neurons. In this review, first, the mechanical microenvironment and properties of neurons under in vivo conditions are introduced and analyzed. Second, research works in recent decades on the effects of different mechanical forces, especially compression and tension, on various neurons, including dorsal root ganglion neurons, retinal ganglion cells, cerebral cortex neurons, hippocampus neurons, neural stem cells, and other neurons, are summarized. Previous research results demonstrate that mechanical stress can not only injure neurons by damaging their morphology, impacting their electrophysiological characteristics and gene expression, but also promote neuron self-repair. Finally, some future perspectives in neuron research are discussed.

  10. Apoptotic death of olfactory sensory neurons in the adult rat.

    Science.gov (United States)

    Deckner, M L; Risling, M; Frisén, J

    1997-01-01

    Olfactory sensory neurons only live for about 1 month in most mammals. It is not fully understood whether the short life span of these neurons is due to necrotic death, or if these cells die by apoptosis. One characteristic of cells undergoing apoptotic cell death is internucleosomal DNA-fragmentation. We have used TdT-mediated dUTP-digoxigenin nick end labeling (TUNEL) to detect cells undergoing DNA-fragmentation in situ. In the intact olfactory epithelium of adult rats a subpopulation of basal immature neuronal progenitor cells, as well as mature olfactory sensory neurons, showed DNA-fragmentation. The number of TUNEL-labeled neurons increased dramatically 1.5 days after transection of the fila olfactoria and declined to control levels by Day 4 after the injury. In order to relate DNA-fragmentation to ultrastructural characteristics of apoptosis we modified the TUNEL-labeling protocol to enable studies of TUNEL-labeled cells in the electron microscope. This confirmed that TUNEL-labeled neurons showed morphological characteristics of apoptosis. The data provide evidence for apoptotic death of neurons in the adult mammalian nervous system. The turnover of olfactory sensory neurons is, at least in part, regulated by apoptosis and disruption of the contact with the olfactory bulb results in massive apoptotic death of neurons in the olfactory epithelium.

  11. Induction, regulation, degradation, and biological significance of mammalian metallothioneins.

    Science.gov (United States)

    Miles, A T; Hawksworth, G M; Beattie, J H; Rodilla, V

    2000-01-01

    MTs are small cysteine-rich metal-binding proteins found in many species and, although there are differences between them, it is of note that they have a great deal of sequence and structural homology. Mammalian MTs are 61 or 62 amino acid polypeptides containing 20 conserved cysteine residues that underpin the binding of metals. The existence of MT across species is indicative of its biological demand, while the conservation of cysteines indicates that these are undoubtedly central to the function of this protein. Four MT isoforms have been found so far, MT-1, MT-2, MT-3, and MT-4, but these also have subtypes with 17 MT genes identified in man, of which 10 are known to be functional. Different cells express different MT isoforms with varying levels of expression perhaps as a result of the different function of each isoform. Even different metals induce and bind to MTs to different extents. Over 40 years of research into MT have yielded much information on this protein, but have failed to assign to it a definitive biological role. The fact that multiple MT isoforms exist, and the great variety of substances and agents that act as inducers, further complicates the search for the biological role of MTs. This article reviews the current knowledge on the biochemistry, induction, regulation, and degradation of this protein in mammals, with a particular emphasis on human MTs. It also considers the possible biological roles of this protein, which include participation in cell proliferation and apoptosis, homeostasis of essential metals, cellular free radical scavenging, and metal detoxification.

  12. Inlfammatory response and neuronal necrosis in rats with cerebral ischemia

    Institute of Scientific and Technical Information of China (English)

    Lingfeng Wu; Kunnan Zhang; Guozhu Hu; Haiyu Yang; Chen Xie; Xiaomu Wu

    2014-01-01

    In the middle cerebral artery occlusion model of ischemic injury, inlfammation primarily occurs in the infarct and peripheral zones. In the ischemic zone, neurons undergo necrosis and apop-tosis, and a large number of reactive microglia are present. In the present study, we investigated the pathological changes in a rat model of middle cerebral artery occlusion. Neuronal necrosis appeared 12 hours after middle cerebral artery occlusion, and the peak of neuronal apoptosis ap-peared 4 to 6 days after middle cerebral artery occlusion. Inlfammatory cytokines and microglia play a role in damage and repair after middle cerebral artery occlusion. Serum intercellular cell adhesion molecule-1 levels were positively correlated with the permeability of the blood-brain barrier. These ifndings indicate that intercellular cell adhesion molecule-1 may be involved in blood-brain barrier injury, microglial activation, and neuronal apoptosis. Inhibiting blood-brain barrier leakage may alleviate neuronal injury following ischemia.

  13. A Study on relationship between translocation of phospho-Rb and neuronal apoptosis following focal cerebral ischemia in rats%大鼠局灶性脑缺血后不同时间点磷酸化Rb蛋白与凋亡神经元的共定位研究

    Institute of Scientific and Technical Information of China (English)

    余樱; 张兆辉; 喻志源; 王伟

    2012-01-01

    目的 探讨大鼠局灶性脑缺血后磷酸化Rb蛋白(p-Rb,ser 795)的表达定位与神经元凋亡的时空关系.方法 制备大鼠大脑中动脉梗塞(MCAO)模型,分为假手术对照组、缺血1h再灌注12h,1d,3d,7d组.利用TUNEL法检测缺血周边区细胞凋亡情况;TUNEL与p-Rb荧光双标观察神经元凋亡与p-Rb表达、定位的关系.结果 缺血半暗带内大部分TUNEL阳性细胞为神经元;大鼠MCAO再灌注12h和1d,TUNEL与p-Rb分别以重叠和镶嵌的方式共定位;再灌注3d,7d发生p-Rb核浆转移的神经元与TUNEL染色细胞仍然分别维持在高水平,但是两者却没有明显的共定位关系.结论 p-Rb可能参与短暂局灶脑缺血后神经元早期凋亡过程,间接或者不参与神经元晚期凋亡过程.%Objective To investigate the temporal and spatial relationship between subcellular localization of phospho-Rb (ser 795) and neuronal apoptotic death in rats subjected to transient focal cerebral ischemia. Methods The model of middle cerebral artery occlusion (MCAO) was carried out. All animals were divided into five groups: sham-operated controls, 12 h, 1 d, 3 d, and 7 d after MCAO/ reperfusion. By analyzing DNA fragmentation with a TUNEL assay, apoptotic cells were examined in the peri-infarct area of the ischemic cerebral cortex. Double-labeling analysis of TUNEL and phospho-Rb staining was used to detect the temporal and spatial relationship between phospho-Rb and neuronal apoptotic death. Results TUNEL-positive cells were mainly neurons in the penumbra zone. Most phospho-Rb appeared to be localized within TUNEL staining cells at 12 h and 1 d after MCAO/reperfusion in an overlapping and mosaic pattern. There was no apparent colocalization of phospho-Rb and TUNEL staining at 3 d and 7 d, although the number of phospho-Rb translocated neurons and TUNEL stained cells were still at high levels at these time points. Conclusion Phospho-Rb may be involved in the early stages of neuronal apoptosis

  14. Basal and Oxidative Stress–Induced Expression of Metallothionein Is Decreased in Ascending Aortic Aneurysms of Bicuspid Aortic Valve Patients

    Science.gov (United States)

    Phillippi, Julie A.; Klyachko, Ekaterina A.; Kenny, John P.; Eskay, Michael A.; Gorman, Robert C.; Gleason, Thomas G.

    2017-01-01

    Background Bicuspid aortic valve (BAV) is a heritable condition that has been linked by an unknown mechanism to a predisposition for ascending aortic aneurysm. Matrix metalloproteinases have been implicated in this predisposition. Metallothionein is a poorly characterized, metal-binding protein that regulates matrix metalloproteinases and is an antioxidant known to be upregulated under oxidative stress. Methods and Results To determine putative factors involved in the pathogenesis of aortic aneurysm in BAV patients, our first goal was to identify genes that are dysregulated in ascending aortic aneurysms of BAV patients compared with tricuspid aortic valve patients and nondiseased (control) donors. By microarray analysis (22 000 probe sets), 110 dysregulated genes were identified in BAV compared with tricuspid aortic valve patients and control donors; 8 were genes of the metallothionein family. Metallothionein gene expression and protein expression were significantly lower in aortic tissue and cultured aortic smooth muscle cells from BAV patients compared with control subjects. Matrix metalloproteinase-9 expression was increased in BAV aortic samples relative to controls. BAV aorta was more susceptible to oxidative stress, and induction of metallothionein under oxidative stress was reduced in BAV patients compared with control subjects. Conclusions These results demonstrate dysregulated metallothionein expression in ascending aortic smooth muscle cells of BAV patients that may contribute to an inadequate response to oxidative stress and provoke aneurysm formation. We hypothesize that metallothionein plays a pivotal role in the response of ascending aortic smooth muscle cells to oxidative stress cues normally involved in the maintenance of the extracellular matrix, including the regulation of matrix metalloproteinase expression. PMID:19398671

  15. L-3-n-Butylphthalide Activates Akt/mTOR Signaling, Inhibits Neuronal Apoptosis and Autophagy and Improves Cognitive Impairment in Mice with Repeated Cerebral Ischemia-Reperfusion Injury.

    Science.gov (United States)

    Xu, Jing; Huai, Yaping; Meng, Nan; Dong, Yanhong; Liu, Zhijuan; Qi, Qianqian; Hu, Ming; Fan, Mingyue; Jin, Wei; Lv, Peiyuan

    2017-06-15

    L-3-n-Butylphthalide (L-NBP) exerts neuroprotective effects in animal models of cerebral ischemia, but its potential benefits in repeated cerebral ischemia-reperfusion (RCIR) injury remain unknown. We investigated the effect of L-NBP on cognitive impairment induced by RCIR in mice. Male C57Bl/6 mice received sham surgery or bilateral common carotid artery occlusion (3 times, 20 min each) and were orally administered preoperative L-NBP (30 mg/kg/day, 7 days), postoperative L-NBP (30 or 60 mg/kg/day, 28 days) or postoperative vehicle (28 days). Learning and memory were assessed by the Morris water maze task and step-down passive avoidance test. Nissl staining was used to identify pathologic changes in the hippocampal CA1 region. The expressions of proteins associated with signaling, apoptosis and autophagy were assessed by quantitative PCR and western blot. RCIR induced deficits in learning and memory that were alleviated by preoperative or postoperative L-NBP administration. Pathologic lesions in the hippocampal CA1 region induced by RCIR were less severe in mice treated with L-NBP. Preoperative or postoperative L-NBP administration in mice receiving RCIR promoted hippocampal expression of phospho-Akt and phospho-mTOR (suggesting activation of Akt/mTOR signaling), increased the Bcl-2/Bax ratio (indicating suppression of apoptosis) and reduced the LC3-II/LC3-I ratio (implying inhibition of autophagy). Preoperative or postoperative L-NBP administration also depressed hippocampal levels of beclin-1 mRNA (indicating suppression of autophagy). These findings suggest that the effect of L-NBP to alleviate learning and memory deficits in mice following RCIR may involve activation of Akt/mTOR signaling and regulation of the expressions of proteins related to apoptosis and autophagy.

  16. 益智仁挥发油抗帕金森模型小鼠黑质神经元凋亡的作用研究%Effects of Volatile Oil Extracted from Alpinia oxyphylla on the Apoptosis of Substantial Nigra Neuron in PD Mice Model

    Institute of Scientific and Technical Information of China (English)

    黄凌; 朱毅; 邝少轶

    2011-01-01

    OBJECTIVE: To study the effect of volatile oil extracted from Alpinia oxyphylla on the expression of Nissl's cyto-ryctes and TH and the apoptosis of substantial nigra neuron in Parkinson' s disease (PD) mice model. METHODS: Model mice were divided into 6 groups, I.e. normal control group (constant volume normal saline), model group (constant volume normal saline, model induced by intragastrical administration of normal saline once a day since 6th day for consecutive 7 days), selegiline group (10 mg·kg-1, model induced by intragastrical administration of selegiline once a day since 4th day for consecutive 7 days), A. oxyphylla volatile oil high-dose, medium-dose and low-dose groups (2.5, 0.833, 0.278 mL· kg-1,model induced by intragastrical administration of A. oxyphylla volatile oil once a day since 6th day for consecutive 7 days). MPTP was used to establish C57BL PD mice model. Nissl staining method was used to detect Nissl cytoryctes expression; SABC staining method was used to detect TH expression; TUNEL staining method was used to detect the apoptosis of substantial nigra neuron in mice. RESULTS: Compared with normal control group, Nissl's cytoryctes and TH expression of substantia nigra neurons in model group were significantly reduced, and the apoptosis of substantia nigra neurons in model mice were significantly increased; compared with model group, Nissl's cytoryctes of PD mice in A oxyphylla volatile oil high-dose and medium-dose groups increased significantly ;TH expression of PD mice in A. oxyphylla voletileoil high-dose,medium-dose, low-dose groups increased significantly, while the apoptosis of substantia nigra neurons were significantly reduced. CONCLUSION: It is suggested that A. oxyphylla volatile oil can relieve the apoptosis of substantia nigra neurons in PD mice.%目的:研究益智仁挥发油对帕金森(PD)模型小鼠黑质致密部尼氏小体、酪氨酸羟化酶(TH)表达的影响和神经元凋亡的对抗作用.方法:实验分为6

  17. Basal and copper-induced expression of metallothionein isoform 1,2 and 3 genes in epithelial cancer cells: The role of tumor suppressor p53.

    Science.gov (United States)

    Ostrakhovitch, E A; Song, Y P; Cherian, M G

    2016-05-01

    Metallothioneins (MTs) are a ubiquitous low-molecular weight, cysteine rich proteins with a high affinity for metal ions. The expression and induction of MTs have been associated with protection against DNA damage, oxidative stress, and apoptosis. Our past research had shown that p53 is an important factor in metal regulation of MTs. The present study was undertaken to explore further the interrelationship between p53 and MTs. We investigated whether silencing of p53 could affect expression pattern of basal and copper induced metallothioneins. The silencing of wild-type p53 (wt-p53) in epithelial breast cancer MCF7 cells affected the basal level of MT-2A RNA, whereas the levels of MT-1A and MT-1X RNA remained largely unchanged. The expression of MT-3 was undetectable in MCF7 with either functional or silenced p53. MCF7 cells with silenced wt-p53 failed to upregulate MT-2A in response to copper and showed a reduced sensitivity toward copper induced cell apoptotic death. Similarly in MCF7-E6 and MDA-MB-231 cells, the presence of inactive/mutated p53 halted MT-1A and MT-2A gene expression in response to copper. Constitutive expression of MT-3 RNA was detectable in the presence of mutated p53 (mtp53). Transient transfection of MDA-MB-231 cells with wt-p53 enabled copper induced upregulation of both MT-1A and MT-2A but not basal level of MT-2A, MT-1E, MT-1X and MT-3. Inactivation of p53 in HepG2 cells amplified the basal expression of studied MT isoforms, including MT-3, as well as copper-induced mRNA expression of MTs except MT-1H and MT-3. Presented data demonstrate a direct relation between p53 and MT-1A and MT-2A and they also indicate that wt-p53 might be a negative regulator of MT-3 in epithelial cancer cells.

  18. 脊髓神经元凋亡在鞘内注射血小板活化因子诱发大鼠痛敏中的作用%Effects of platelet-activating factor administered intrathecally on the expression of neuronal apoptosis in spi-nal cord of rats

    Institute of Scientific and Technical Information of China (English)

    杨京利; 段宏伟; 马国平

    2014-01-01

    Objective To investigate the roles of neuronal apoptosis in development of tactile allodynia and thermal hyperalgesia induced by platelet-activating factor administered intrathecally.Methods 60 Sprague-Dawley rats with intrathecal PE-10 catheters were randomly divided into two groups:control(artificial cerebral spinal fluid,10 μl)group,PAF (10 μg,dissolved in 10 μl ACSF)group.Paw withdrawal mechanical threshold and thermal latency were measured at-1 ,1 ,3,5,7 and 14 days after surgery .Changes of apoptosis of spinal neuron were observed by TUNEL.Results Intrathecally administered PAF induced development of tactile all-odynia and thermal hyperalgesia rapidly,The expression of spinal neuronal apoptotic index in control group were significantly lower than those in the PAF group(P <0.01).Conclusions Intrathecally administered PAF may induce development of tactile allodynia and thermal hyperalgesia in rats,and this process might be associ-ated with neuronal apoptosis in spinal cord.%目的:探讨脊髓神经元凋亡在鞘内注射血小板活化因子(PAF)诱发大鼠痛敏中的作用。方法鞘内置管成功的雄性 Sprague-Dawley 大鼠60只,随机分为2组:对照组,30只,鞘内注射人工脑脊液(arti-ficial cerebral spinal fluid,ACSF)10μl;PAF 组,30只,鞘内注射 PAF 10μg,溶解于10μl 人工脑脊液;分别于鞘内给药前1 d、给药后1、3、5、7、14 d 分别测定机械痛阈(PWMT)和热痛阈(PWTL)。取 L4-6脊髓,采用TUNEL 法观察脊髓神经元凋亡。结果鞘内注射血小板活化因子(PAF)可诱发出大鼠机械性触诱发痛和热痛觉过敏。PAF 组术后1 d 脊髓中开始有少量凋亡神经元出现,凋亡指数于术后3 d 开始迅速增加,术后5 d 达峰值,与对照组比较,有显著性差异(P <0.01)。结论鞘内注射 PAF 诱发大鼠触觉异常痛敏和热痛敏,脊髓神经元凋亡可能参与了鞘内注射 PAF 大鼠痛敏的形成。

  19. 舒肝解郁胶囊对抑郁模型大鼠海马神经元凋亡及脑组织caspase-3蛋白表达的影响%Effect of Shuganjieyu capsules on neuronal apoptosis in hippocampal CA3 area and the expression of caspase-3 in the brain of rat depression model

    Institute of Scientific and Technical Information of China (English)

    傅锦华; 刘勇; 王清勇; 赵靖平

    2012-01-01

    Objective: To evaluate the effect of "Shuganjieyu" (SGJY) capsules on neuronal apoptosis in hippocampal CA3 area and the expression of caspase-3 in the brain of rat depression model, and to investigate its pharmacological mechanisms in depression treatment. Methods: Adult male SD rats were randomly divided into 4 groups: a control, a model, a SGJY and a fluoxetine group. The rat depression model was established under chronic unpredictable mild stress (CUMS) and separate feeding. The behaviors were measured by open-field test, sucrose consumption and forced swimming test. We observed the neuronal morphology structure and neuronal apoptosis in the hippocampal CA3 area. We detected the rat caspase-3 expression level of medial prefrontal cortex ( mPFC) and hippocampal CA3 area by Western blot. Results: After 21-day stress, compared with the model group, spontaneous activity and sucrose consumption and preference percentage of the rats in the SGJY group significantly increased, while the immobility time in forced swimming test, the number of apoptotic cells and the protein levels of caspase-3 significantly reduced (P0.05). Conclusion: SGJY capsules can reduce the depression symptoms of CUMS and help to increase hippocampal neuron generation, survival and neogenesis, reduce the protein levels of caspase-3, and reverse neurocyte apoptosis in the rat depression model with the same efficacy as fluoxetine.%目的:研究舒肝解郁胶囊对抑郁模型大鼠海马神经元凋亡及脑组织caspase-3蛋白表达的影响,探讨其治疗抑郁症的作用机制.方法:将雄性SD大鼠随机分为正常对照组、模型组、舒肝解郁组和氟西汀组四组;采用慢性轻度不可预见性应激(CUMS)结合孤养建立抑郁大鼠模型,并用旷场、糖水消耗和强迫