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Franklin JL, Amsler MO, Messina JL. Regulation of glucose responsive protein (GRP) gene expression by insulin. Cell Stress Chaperones 2022; 27:27-35. [PMID: 34755306 PMCID: PMC8821767 DOI: 10.1007/s12192-021-01243-z] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/13/2021] [Revised: 09/08/2021] [Accepted: 10/15/2021] [Indexed: 11/28/2022] Open
Abstract
While screening for insulin-induced genes, we identified two members of a family of stress-induced genes referred to as glucose-regulated proteins (GRPs). GRPs are members of the stress-responsive superfamily of genes which also includes heat shock proteins (HSPs). The GRP proteins are not normally heat-inducible, but are overproduced when cells are starved of glucose. The two major GRP proteins, GRP78 and GRP94, are highly conserved among vertebrates. We have found that physiological concentrations of insulin stimulate the transcription of GRP78 and GRP94 in rat H4IIE hepatoma cells. The regulation of GRP78 transcription was rapid, with the first induction within minutes, and a further induction after several hours, and both occurred in the presence of glucose. GRP78 transcription was more greatly induced by insulin in the presence of SB202190, a specific p38-MAPK inhibitor. Transcription of GRP94 was also induced, but only after several hours. Calcimycin (A23187) and anisomycin were used to induce endoplasmic reticulum (ER)/cellular stress, and both induced GRP78 and GRP94 transcription.
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Affiliation(s)
- J Lee Franklin
- Department of Pathology, Division of Pathobiology and Molecular Medicine, University of Alabama at Birmingham, 1670 University Blvd., Volker Hall G019, Birmingham, AL, 35294-0019, USA
- Veterans Administration Medical Center, Birmingham, AL, 35294, USA
| | - Margaret O Amsler
- Department of Biology, University of Alabama at Birmingham, Birmingham, AL, 35294, USA
| | - Joseph L Messina
- Department of Pathology, Division of Pathobiology and Molecular Medicine, University of Alabama at Birmingham, 1670 University Blvd., Volker Hall G019, Birmingham, AL, 35294-0019, USA.
- Veterans Administration Medical Center, Birmingham, AL, 35294, USA.
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Zhang Q, Yang Y, Lu Y, Cao Z. iTRAQ-based quantitative proteomic analyses the cycle chronic heat stress affecting liver proteome in yellow-feather chickens. Poult Sci 2021; 100:101111. [PMID: 33965807 PMCID: PMC8120948 DOI: 10.1016/j.psj.2021.101111] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2020] [Revised: 12/16/2020] [Accepted: 03/02/2021] [Indexed: 02/07/2023] Open
Abstract
Heat stress (HS) is one of the main environmental factors affecting the efficiency of poultry production. The yellow-feather chickens (YFC) as an indigenous strain of chicken is a popular poultry breed in China. Our previous study used the RNA-seq to analyze the gene expression profiles of male YFC under HS and showed that the lipid and energy metabolism pathways are activated in livers of YFC exposed to acute HS (38°C, 4 h and 25°C recovery 2 h). In this study, we used quantitative proteome analysis based on iTRAQ to study the liver response of YFC to cycle chronic HS (38 ± 1°C, 8 h/d, 7 d, CyCHS). The male YFCs treatment used the CyCHS from 22 to 28 days of age. The liver tissue samples were collected at 28 d old. A total of 39,327 unique peptides matches were detected using iTRAQ analysis and 4,571 proteins exhibited a false discovery rate of 1% or less. Forty-six significant differentially expressed proteins (DEPs) were detected in the CyCHS group compared with the control group for the liver samples, including up- and down-regulated DEPs were 18 and 28, respectively. We found that the enriched biological process terms of the DEPs expressed in the liver were related to DNA metabolic process, oxidation-reduction process, oxidative stress and gluconeogenesis. In KEGG pathway analysis. Most of the hepatic DEPs were annotated to glutathione metabolism and TCA cycle in response to CyCHS. The up-regulation of 5 DEPs (GPX1, GSTT1, GSTT1L, RRM2, and LOC100859645) in the glutathione metabolism pathway likely reflects an attempt to deal with oxidative damage by CyCHS. The down-regulation of 3 DEPs (Isocitrate dehydrogenase [IDH3A], IDH3B, and phosphoenolpyruvate carboxykinase 1) in the TCA cycle pathway contributes to the regulation mechanism of energy metabolism and probably to cope with the balance of heat production and dissipation during CyCHS in order to adapt to high temperature environments. Our results provide insights into the potential molecular mechanism in heat-induced oxidative stress and energy in YFCs and future studies will investigate the functional genes associated with the response to HS.
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Affiliation(s)
- Quan Zhang
- College of Coastal Agricultural Sciences, Guangdong Ocean University, Zhanjiang, China.
| | - YuZe Yang
- Beijing General Station of Animal Husbandry, Beijing, China
| | - YongQiang Lu
- Beijing General Station of Animal Husbandry, Beijing, China
| | - ZiWen Cao
- College of Coastal Agricultural Sciences, Guangdong Ocean University, Zhanjiang, China
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Zhang K, Zhao P, Guo G, Guo Y, Li S, He Y, Sun X, Chai H, Zhang W, Xing M. Arsenic Trioxide Exposure Induces Heat Shock Protein Responses in Cock Livers. Biol Trace Elem Res 2016; 170:459-65. [PMID: 26329997 DOI: 10.1007/s12011-015-0487-0] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/15/2015] [Accepted: 08/18/2015] [Indexed: 12/15/2022]
Abstract
Arsenic is a trace element widely found in nature, and there are several forms of arsenic, including the most toxic form of trivalent arsenic. Arsenic trioxide (As2O3) is widespread in nature and tends to accumulate in animals and humans, thus causing great harm. Although the important role of heat shock proteins (HSPs) has been demonstrated in many types of mammals exposed to As2O3, the function of these proteins in poultry, especially in cocks, remains unclear. In this study, we used experimental animals (male chickens), which were fed a diet including 0, 7.5, 15, and 30 mg kg(-1) As2O3, respectively, in the control, low, middle, and high groups. The livers were collected after the cocks were treated with arsenic for 30, 60, and 90 days. We detected HSP27, HSP60, HSP70, and HSP90 levels in the livers of the cocks by real-time PCR and HSP60 and HSP70 levels by Western blot. The results showed that the messenger RNA and protein expression of HSPs exposed to As2O3 had obviously increased. These results demonstrated that arsenic toxicity affected the expression of HSP levels in cock livers.
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Affiliation(s)
- Kexin Zhang
- College of Wildlife Resources, Northeast Forestry University, 26 Hexing Road, Harbin, Heilongjiang Province, 150040, China
| | - Panpan Zhao
- College of Wildlife Resources, Northeast Forestry University, 26 Hexing Road, Harbin, Heilongjiang Province, 150040, China
| | - Guangyang Guo
- College of Wildlife Resources, Northeast Forestry University, 26 Hexing Road, Harbin, Heilongjiang Province, 150040, China
| | - Ying Guo
- College of Wildlife Resources, Northeast Forestry University, 26 Hexing Road, Harbin, Heilongjiang Province, 150040, China
| | - Siwen Li
- College of Wildlife Resources, Northeast Forestry University, 26 Hexing Road, Harbin, Heilongjiang Province, 150040, China
| | - Ying He
- College of Wildlife Resources, Northeast Forestry University, 26 Hexing Road, Harbin, Heilongjiang Province, 150040, China
| | - Xiao Sun
- College of Wildlife Resources, Northeast Forestry University, 26 Hexing Road, Harbin, Heilongjiang Province, 150040, China
| | - Hongliang Chai
- College of Wildlife Resources, Northeast Forestry University, 26 Hexing Road, Harbin, Heilongjiang Province, 150040, China.
| | - Wen Zhang
- College of Wildlife Resources, Northeast Forestry University, 26 Hexing Road, Harbin, Heilongjiang Province, 150040, China.
| | - Mingwei Xing
- College of Wildlife Resources, Northeast Forestry University, 26 Hexing Road, Harbin, Heilongjiang Province, 150040, China.
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Rhoads RP, Baumgard LH, Suagee JK, Sanders SR. Nutritional interventions to alleviate the negative consequences of heat stress. Adv Nutr 2013; 4:267-76. [PMID: 23674792 PMCID: PMC3650495 DOI: 10.3945/an.112.003376] [Citation(s) in RCA: 135] [Impact Index Per Article: 11.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
Energy metabolism is a highly coordinated process, and preferred fuel(s) differ among tissues. The hierarchy of substrate use can be affected by physiological status and environmental factors including high ambient temperature. Unabated heat eventually overwhelms homeothermic mechanisms resulting in heat stress, which compromises animal health, farm animal production, and human performance. Various aspects of heat stress physiology have been extensively studied, yet a clear understanding of the metabolic changes occurring at the cellular, tissue, and whole-body levels in response to an environmental heat load remains ill-defined. For reasons not yet clarified, circulating nonesterified fatty acid levels are reduced during heat stress, even in the presence of elevated stress hormones (epinephrine, glucagon, and cortisol), and heat-stressed animals often have a blunted lipolytic response to catabolic signals. Either directly because of or in coordination with this, animals experiencing environmental hyperthermia exhibit a shift toward carbohydrate use. These metabolic alterations occur coincident with increased circulating basal and stimulated plasma insulin concentrations. Limited data indicate that proper insulin action is necessary to effectively mount a response to heat stress and minimize heat-induced damage. Consistent with this idea, nutritional interventions targeting increased insulin action may improve tolerance and productivity during heat stress. Further research is warranted to uncover the effects of heat on parameters associated with energy metabolism so that more appropriate and effective treatment methodologies can be designed.
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Affiliation(s)
- Robert P Rhoads
- Department of Animal and Poultry Sciences, Virginia Tech, Blacksburg, VA, USA.
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Shen M, Lin F, Zhang J, Tang Y, Chen WK, Liu H. Involvement of the up-regulated FoxO1 expression in follicular granulosa cell apoptosis induced by oxidative stress. J Biol Chem 2012; 287:25727-40. [PMID: 22669940 DOI: 10.1074/jbc.m112.349902] [Citation(s) in RCA: 147] [Impact Index Per Article: 11.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
Follicular atresia is common in female mammalian ovaries, where most follicles undergo degeneration at any stage of growth and development. Oxidative stress gives rise to triggering granulosa cell apoptosis, which has been suggested as a major cause of follicular atresia. However, the underlying mechanism by which the oxidative stress induces follicular atresia remains unclear. FoxO transcription factors are known as critical mediators in the regulation of oxidative stress and apoptosis. In this study, the involvement of FoxO1 in oxidative stress-induced apoptosis of mouse follicular granulosa cells (MGCs) was investigated in vivo and in vitro. It was observed that increased apoptotic signals correlated with elevated expression of FoxO1 in MGCs when mice were treated with the oxidant. Correspondingly, the expressions of FoxO1 target genes, such as proapoptotic genes and antioxidative genes, were also up-regulated. In primary cultured MGCs, treatment with H(2)O(2) led to FoxO1 nuclear translocation. Further studies with overexpression and knockdown of FoxO1 demonstrated the critical role of FoxO1 in the induction of MGC apoptosis by oxidative stress. Finally, inactivation of FoxO1 by insulin treatment confirmed that FoxO1 induced by oxidative stress played a pivotal role in up-regulating the expression of downstream apoptosis-related genes in MGCs. Our results suggest that up-regulation of FoxO1 by oxidative stress leads to apoptosis of granulosa cells, which eventually results in follicular atresia in mice.
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Affiliation(s)
- Ming Shen
- College of Animal Science and Technology, Nanjing Agricultural University, Nanjing 210095, China
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Voolstra CR, Schnetzer J, Peshkin L, Randall CJ, Szmant AM, Medina M. Effects of temperature on gene expression in embryos of the coral Montastraea faveolata. BMC Genomics 2009; 10:627. [PMID: 20030803 PMCID: PMC2807443 DOI: 10.1186/1471-2164-10-627] [Citation(s) in RCA: 95] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/02/2009] [Accepted: 12/23/2009] [Indexed: 11/10/2022] Open
Abstract
BACKGROUND Coral reefs are expected to be severely impacted by rising seawater temperatures associated with climate change. This study used cDNA microarrays to investigate transcriptional effects of thermal stress in embryos of the coral Montastraea faveolata. Embryos were exposed to 27.5 degrees C, 29.0 degrees C, and 31.5 degrees C directly after fertilization. Differences in gene expression were measured after 12 and 48 hours. RESULTS Analysis of differentially expressed genes indicated that increased temperatures may lead to oxidative stress, apoptosis, and a structural reconfiguration of the cytoskeletal network. Metabolic processes were downregulated, and the action of histones and zinc finger-containing proteins may have played a role in the long-term regulation upon heat stress. CONCLUSIONS Embryos responded differently depending on exposure time and temperature level. Embryos showed expression of stress-related genes already at a temperature of 29.0 degrees C, but seemed to be able to counteract the initial response over time. By contrast, embryos at 31.5 degrees C displayed continuous expression of stress genes. The genes that played a role in the response to elevated temperatures consisted of both highly conserved and coral-specific genes. These genes might serve as a basis for research into coral-specific adaptations to stress responses and global climate change.
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Affiliation(s)
- Christian R Voolstra
- 1Red Sea Research Center, King Abdullah University of Science and Technology (KAUST), Thuwal, Saudi Arabia.
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Papathanassoglou EDE, Bozas E, Giannakopoulou MD. Multiple organ dysfunction syndrome pathogenesis and care: a complex systems' theory perspective. Nurs Crit Care 2009; 13:249-59. [PMID: 18816311 DOI: 10.1111/j.1478-5153.2008.00289.x] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
AIMS AND OBJECTIVES To discuss multiple organ dysfunction syndrome (MODS) from a complex systems' theory perspective and to delineate a conceptual framework for the development and care of MODS. BACKGROUND MODS is an intricate and devastating manifestation of critical illness characterized by widespread aberrant molecular, cellular and systemic responses. DESIGN AND METHODS Narrative literature review (MEDLINE, CINAHL databases) and knowledge synthesis with the theoretical assertions of chaos and complex systems' theory. Cellular and systemic response paradoxes in MODS (including cellular hypoxia, cell death and signalling) are reviewed. RESULTS The diseased person is depicted as a complex adaptive system. The relevancy of some of the principles of complex chaotic systems' theory to the proposed model is illustrated, including sensitive dependence on initial conditions, emergence, attractors, self-organization, self-organized criticality and emerging order. The transition from life-supporting to death-related organismic responses is illustrated as a critical event in MODS and care implications are drawn. CONCLUSIONS Patient responses in MODS appear to conform to the principles of chaotic systems. Death is illustrated not as a consequence of homeostatic failure but as a 'deliberate' self-organized phenomenon entailing multiple dynamically evolving mechanisms. RELEVANCE TO CLINICAL PRACTICE Some of the principles of chaotic complex systems may need to be taken into account to advance care in MODS. An alternative theoretical perspective may support nurses to conceptualize both MODS and their role in a way that will help them to cope better with this devastating syndrome and develop practice.
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8
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Mujahid A, Akiba Y, Warden CH, Toyomizu M. Sequential changes in superoxide production, anion carriers and substrate oxidation in skeletal muscle mitochondria of heat-stressed chickens. FEBS Lett 2007; 581:3461-7. [PMID: 17612532 DOI: 10.1016/j.febslet.2007.06.051] [Citation(s) in RCA: 86] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/24/2007] [Revised: 06/20/2007] [Accepted: 06/21/2007] [Indexed: 10/23/2022]
Abstract
We have shown that heat-stressed birds exhibit increased superoxide production in skeletal muscle mitochondria. To determine the precise mechanism for this effect, here we studied not only progressive, but also sequential changes in superoxide production, anion carriers and substrate oxidation in mitochondria of heat-stressed chickens. Exposure to acute heat stress (34 degrees C for 6, 12 and 18h) stimulated pectoralis muscle mitochondrial superoxide production. Heat stress-induced downregulations of avUCP gene transcripts and mitochondrial avUCP protein content were time-dependent: avUCP gene transcript was decreased after 6h, while avUCP protein content was only downregulated after 12h of heat stress. Avian adenine nucleotide translocator (avANT) gene transcripts were not changed on exposure to heat stress, suggesting that avANT may not be involved in the regulation of superoxide production in the muscle mitochondria of heat-stressed chickens. During the initial stage of acute heat stress beta-oxidation enzymes gene transcripts and activity were upregulated, with elevated plasma non-esterified fatty acid levels and increased expression of mitochondrial fatty acid transport genes. This sudden surge in mitochondrial substrate oxidation resulted in higher superoxide production: the avUCP expression at 6h after heat stress might have not been large enough to alleviate the overproduction of reactive oxygen species (ROS) even though a small amount of endogenous FFA, a potential uncoupler, might have been present in the mitochondria. Thereafter, avUCP content was downregulated while substrate oxidation returned to control levels. This downregulation of avUCP may have caused increased mitochondrial superoxide production, keeping the superoxide production high in the later stages of heat stress. These results suggest that overproduction of mitochondrial ROS in chicken skeletal muscle under the heat stress might result from enhanced substrate oxidation and downregulation of avUCP in a time-dependent manner.
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Affiliation(s)
- Ahmad Mujahid
- Science of Biological Function, Life Science, Graduate School of Agricultural Science, Tohoku University, 1-1 Tsutsumidori-Amamiyamachi, Aoba-ku, Sendai 981-8555, Japan
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9
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Gaitanaki C, Pliatska M, Stathopoulou K, Beis I. Cu2+ and acute thermal stress induce protective eventsviathe p38-MAPK signalling pathway in the perfusedRana ridibundaheart. J Exp Biol 2007; 210:438-46. [PMID: 17234613 DOI: 10.1242/jeb.02680] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
SUMMARYIn the present study, we investigated the induction of the p38-MAPK signalling pathway by copper, as exemplified by CuCl2, in the isolated perfused heart of the amphibian Rana ridibunda. We found that p38-MAPK phosphorylation by CuCl2 occurs in a dose-dependent manner, with maximum activation (8.73±1.43-fold relative to control values) attained by perfusion with 500 μmol l–1CuCl2 for 15 min, while this activation sustained even after 60 min of reperfusion with normal bicarbonate buffer. CuCl2 also induced the phosphorylation of the small heat shock protein 27 (Hsp27) in a p38-MAPK dependent manner, as revealed by experiments using the p38-MAPK inhibitor SB203580. p38-MAPK and Hsp27 phosphorylations were also strongly induced by hyperthermia (42°C), while the simultaneous use of hyperthermia and CuCl2 had a synergistic effect on p38-MAPK activation. Furthermore,perfusions with the potent antioxidant L-ascorbic acid (100 μmol l–1), the antioxidant enzymes catalase (CAT) (150 U ml–1) or superoxide dismutase (SOD) (30 U ml–1) in the presence of 500 μmol l–1CuCl2 did not attenuate the CuCl2-induced p38-MAPK activation, implying that at least the reactive oxygen species (ROS) scavenged by these agents are not implicated in this kinase activation. The p38-MAPK phosphorylation induced by the combined action of CuCl2 and hyperthermia was partially inhibited by catalase, indicating that hyperthermia possibly activates the kinase through the production of H2O2. Caspase-3, an effector protease of apoptosis,remained inactive in hearts perfused at normal or hyperthermic conditions, in the absence or presence of 500 μmol l–1 CuCl2. All the above results suggest that, in the amphibian Rana ridibundaheart, p38-MAPK activation by copper has a possible protective role through the small Hsp27.
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Affiliation(s)
- Catherine Gaitanaki
- Department of Animal and Human Physiology, School of Biology, Faculty of Sciences, University of Athens, Panepistimioupolis, Athens 157 84, Greece
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Mujahid A, Akiba Y, Toyomizu M. Acute Heat Stress Induces Oxidative Stress and Decreases Adaptation in Young White Leghorn Cockerels by Downregulation of Avian Uncoupling Protein. Poult Sci 2007; 86:364-71. [PMID: 17234852 DOI: 10.1093/ps/86.2.364] [Citation(s) in RCA: 87] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022] Open
Abstract
Reactive oxygen species-induced damage of cells and molecules is one of the mechanisms responsible for the decline in an animal's performance due to heat stress. Mitochondria are the main producers of cellular superoxide, a process that is sensitive to proton motive force, and this superoxide production can be decreased by mild uncoupling. We studied the effects of heat stress on the production of mitochondrial superoxide as well as heat stress effects on the expression of avian uncoupling protein (avUCP) and avian A nucleotide translocator (avANT) in skeletal muscles of chicks and young cockerels. Male White Leghorn (Julia) chicks at 16 d and cockerels at 87 d of age were exposed to acute heat stress, 34 degrees C for 18 h, or kept at moderate ambient temperature (25 and 21 degrees C, respectively). There was no difference in mitochondrial superoxide production between heat-exposed and control chicks, whereas significant differences were observed in the case of young cockerels. Greater substrate-independent superoxide production was found in muscle mitochondria from heat-stressed young cockerels. In chicks, neither avUCP nor avANT transcript expression was changed by heat exposure, whereas in young cockerels avUCP transcript was decreased, but avANT transcript level was not changed. Thus, in heat-stressed young cockerels, increased mitochondrial superoxide production was accompanied by downregulation of avUCP. Taken together, these results suggest that exposure of young cockerels to heat stress stimulates mitochondrial superoxide production, possibly via downregulation of avUCP. Chicks with persistent avUCP expression, on the other hand, are relatively better adapted to high temperature. It can be assumed that appropriate expression of avUCP may alleviate overproduction of mitochondrial superoxide and could help birds adapt to oxidative stress resulting from acute heat stress.
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Affiliation(s)
- A Mujahid
- Science of Biological Function, Life Science, Graduate School of Agricultural Science, Tohoku University, Aoba-ku, Sendai 981-8555, Japan.
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Mujahid A, Sato K, Akiba Y, Toyomizu M. Acute heat stress stimulates mitochondrial superoxide production in broiler skeletal muscle, possibly via downregulation of uncoupling protein content. Poult Sci 2006; 85:1259-65. [PMID: 16830867 DOI: 10.1093/ps/85.7.1259] [Citation(s) in RCA: 130] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/14/2022] Open
Abstract
Acute heat stress (34 degrees C for 18 h) resulted in increased levels of reactive oxygen species (ROS) in mitochondria isolated from the skeletal muscle of broilers. This occurred when glutamate-requiring complexes I, III, and IV of the electron transport chain or succinate-requiring complexes II, III and IV were used as the substrate. This result confirms our previous observation that exposure of broilers to 34 degrees C for 18 h results in increased superoxide production in skeletal (pectoralis) muscle, and extends this finding by showing that substrate-independent ROS generation occurs during the heat stress period. When broilers were exposed to heat stress, the levels of avian uncoupling protein (avUCP) mRNA in skeletal muscle were significantly decreased, to 28% of the levels found in untreated controls. This was accompanied by a significant decrease in the levels of the avUCP protein, to 37% of control levels. In contrast, avian adenine nucleotide translocator mRNA levels were not affected by exposure to heat stress. This finding is consistent with previous studies which showed that the increases in superoxide production that are observed in the presence of carboxyatractylate, a specific inhibitor of adenine nucleotide translocator, were the same for skeletal muscle mitochondria from both control and heat-stressed chickens. Taken together, these results suggest that acute heat stress stimulates mitochondrial superoxide production in broiler skeletal muscle, possibly via downregulation of avUCP. The present study provides the first evidence that synthesis of avUCP protein is downregulated in heat-stressed broilers.
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Affiliation(s)
- A Mujahid
- Science of Biological Function, Life Science, Graduate School of Agricultural Science, Tohoku University, Sendai, Japan.
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12
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Tiberio L, Tiberio GAM, Bardella L, Cervi E, Cerea K, Dreano M, Garotta G, Fra A, Montani N, Ferrari-Bravo A, Callea F, Grigolato P, Giulini SM, Schiaffonati L. Mechanisms of interleukin-6 protection against ischemia-reperfusion injury in rat liver. Cytokine 2006; 34:131-42. [PMID: 16814559 DOI: 10.1016/j.cyto.2006.04.009] [Citation(s) in RCA: 35] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/23/2006] [Revised: 04/03/2006] [Accepted: 04/11/2006] [Indexed: 01/20/2023]
Abstract
Numerous animal studies simulating liver injury have demonstrated that interleukin-6 (IL-6) exerts a protective effect. This study was designed to further analyze the molecular mechanisms underlying the protective role of IL-6 in a rat model of liver ischemia/reperfusion injury. We show that IL-6: (i) at high doses reduces cell damage which occurs in ischemic-reperfused liver, while at low doses displays only a limited protective capacity, (ii) anticipates and enhances hepatocyte compensatory proliferation seen in ischemic-reperfused liver also at a low, more pharmacologically acceptable dose, (iii) sustains the acute phase response which is dampened in ischemic-reperfused liver, (iv) strengthens the heat shock-stress response shown by ischemic-reperfused liver and (v) overcomes the dysfunctions of the unfolding protein response found in ischemic-reperfused liver. We also show that IL-6-enhanced STAT3 activation probably plays a crucial role in the potentiation of the different protective pathways activated in ischemic-reperfused liver. Our data confirm that IL-6 is a potential therapeutic in liver injury of different etiologies and reveal novel mechanisms by which IL-6 sustains liver function after ischemia/reperfusion injury.
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Affiliation(s)
- Laura Tiberio
- Department of Biomedical Sciences and Biotechnology, Division of General Pathology and Immunology, University of Brescia, Italy
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Sundar SV, Li YY, Rollwagen FM, Maheshwari RK. Hemorrhagic shock induces differential gene expression and apoptosis in mouse liver. Biochem Biophys Res Commun 2005; 332:688-96. [PMID: 15907801 DOI: 10.1016/j.bbrc.2005.05.008] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/04/2005] [Accepted: 05/05/2005] [Indexed: 01/07/2023]
Abstract
Comprehensive knowledge of the gene expression changes induced by hemorrhage in vital organs will greatly improve prognosis and therapy. Therefore, we used a mouse model of non-resuscitated hemorrhagic shock to study the pattern of stress-induced genes in liver at 1, 4, and 24 h following surgery. Hepatic injury was confirmed by assessment of liver injury markers and apoptotic cell death. We found that a variety of stress-regulated genes were differentially expressed, including seven genes that have not been reported previously as being regulated by hemorrhagic shock: ATF-2, alphaB-crystallin, GADD45, GADD45beta, Mdm2, p21Waf1, and TRPM-2. The changes in mRNA levels of the transcription factors AP-1, Egr-1, HSF-1, and NF-kappaB were transient but protein expression was noticeable at later time points. Our findings show that oxidative stress causes immediate upregulation of genes involved in a variety of cellular defense pathways. Complex interactions among them might determine the ultimate fate of the cell.
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Affiliation(s)
- Shirin V Sundar
- Department of Pathology, Uniformed Services University of the Health Sciences, Bethesda, MD 20814, USA
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Fallsehr C, Zapletal C, Kremer M, Demir R, von Knebel Doeberitz M, Klar E. Identification of differentially expressed genes after partial rat liver ischemia/reperfusion by suppression subtractive hybridization. World J Gastroenterol 2005; 11:1303-16. [PMID: 15761968 PMCID: PMC4250677 DOI: 10.3748/wjg.v11.i9.1303] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To identify potential diagnostic target genes in early reperfusion periods following warm liver ischemia before irreversible liver damage occurs.
METHODS: We used two strategies (SSH suppression subtractive hybridization and hybridization of cDNA arrays) to determine early changes in gene expression profiles in a rat model of partial WI/R, comparing postischemic and adjacent nonischemic liver lobes. Differential gene expression was verified (WI/R; 1 h/2 h) and analyzed in more detail after warm ischemia (1 h) in a reperfusion time kinetics (0, 1, 2 and 6 h) and compared to untreated livers by Northern blot hybridizations. Protein expression was examined on Western blots and by immunohistochemistry for four differentially expressed target genes (Hsp70, Hsp27, Gadd45a and IL-1rI).
RESULTS: Thirty-two individual WI/R target genes showing altered RNA levels after confirmation by Northern blot analyzes were identified. Among them, six functionally uncharacteristic expressed sequences and 26 known genes (12 induced in postischemic liver lobes, 14 with higher transcriptional expression in adjacent nonischemic liver lobes). Functional categories of the verified marker genes indicate on the one hand cellular stress and tissue damage but otherwise activation of protective cellular reactions (AP-1 transcription factors, apoptosis related genes, heat shock genes). In order to assign the transcriptional status to the biological relevant protein level we demonstrated that Hsp70, Hsp27, Gadd45a and IL-1rI were clearly up-regulated comparing postischemic and untreated rat livers, suggesting their involvement in the WI/R context.
CONCLUSION: This study unveils a WI/R response gene set that will help to explore molecular pathways involved in the tissue damage after WI/R. In addition, these genes especially Hsp70 and Gadd45a might represent promising new candidates indicating WI/R liver damage.
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Affiliation(s)
- Christine Fallsehr
- Institute of Molecular Pathology, University of Heidelberg, Heidelberg, Germany
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15
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Mujahid A, Yoshiki Y, Akiba Y, Toyomizu M. Superoxide radical production in chicken skeletal muscle induced by acute heat stress. Poult Sci 2005; 84:307-14. [PMID: 15742968 DOI: 10.1093/ps/84.2.307] [Citation(s) in RCA: 202] [Impact Index Per Article: 10.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022] Open
Abstract
Heat stress is of major concern for poultry, especially in the hot regions of the world because of the resulting poor growth performance, immunosuppression, and high mortality. To assess superoxide (O2*-) production in mitochondria isolated from skeletal muscle of chickens (n = 4 to 8) exposed to acute heat stress, electron spin resonance (ESR) spectroscopy using 5,5-dimethyl-1-pyrroline N-oxide (DMPO) as a spin trap agent and lucigenin-derived chemiluminescence (LDCL) method were applied. ESR spectra of suspensions containing mitochondria from control and acute heat-treated meat-type chickens showed similar hyperfine coupling constants (aN = 1.44 mT, aHbeta = 0.12 mT, and aHbeta = 0.11 mT) to those of DMPO-O2*- adducts observed in a hypoxanthine-xanthine oxidase system. Heat exposure resulted in enhancement of the DMPO-O2*- signal. The results using LDCL showed significantly enhanced superoxide production in heat stress-treated skeletal muscle mitochondria of meat-type chickens, whereas no such increase was observed in laying chickens. The enhancement of superoxide production in the former case was associated with heat-induced increments in rectal and muscle temperatures, leading to significant body weight loss. In contrast, the latter case showed no increase in temperatures, although there was a slight decrease in body weight gain. Percentage increases of superoxide production in the presence of carboxyatractylate, a specific inhibitor of adenine nucleotide translocator (ANT), were the same for skeletal muscle mitochondria from meat- and laying-type chickens from the control or heat stress-treated group. This finding suggests the irrelevance of ANT in the regulation of reactive oxygen species flux under heat stress conditions. The study provides the first evidence of superoxide anion production in the skeletal muscle mitochondria of meat-type chickens in response to acute heat stress.
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Affiliation(s)
- A Mujahid
- Life Science, Graduate School of Agriculture, Tohoku University, 1-1 Tsutsumidori-Amamiyamachi, Aoba-ku, Sendai 981-8555, Japan
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16
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Abstract
This paper presents a review of recent research on the hypothermic storage of hepatocytes. The first focus is on the diversity of methodologies currently employed in this area. The cell damage caused by hypothermic preservation and its possible mechanism are then investigated on both morphological and molecular biology. Later, the gene expressions on a mRNA level or enzyme level after hypothermic preservation are further discussed. Finally, the improvement of hypothermic storage by preconditioning, such as by increasing temperature, is explored.
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Affiliation(s)
- Qin Meng
- Department of Chemical Engineering and Biochemical Engineering, Zhejiang University, 38 Zheda Road, Hangzhou, Zhejiang, P. R. China.
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17
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Njemini R, Lambert M, Demanet C, Vanden Abeele M, Vandebosch S, Mets T. The induction of heat shock protein 70 in peripheral mononuclear blood cells in elderly patients: a role for inflammatory markers. Hum Immunol 2003; 64:575-85. [PMID: 12770787 DOI: 10.1016/s0198-8859(03)00068-5] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
The induction of heat shock proteins (Hsp) is the response to a plethora of stress signals including hyperthermia, physical stress, and various disease states. Although changes in Hsp expression are associated with certain diseases, the question as to whether this is an adaptation to a particular pathophysiologic state or a reflection of the suboptimal cellular environment associated with the disease remains open. In this study we have investigated the effects of inflammatory mediators on the induction of Hsp 70 in human peripheral mononuclear blood cells using flow cytometry. We demonstrate that without heat shock, the levels of the inflammatory mediators are positively related to Hsp 70 production in monocytes. On the contrary, negative correlations were found between heat induced Hsp 70 production and interleukin-6 (IL-6), as well as various markers of inflammation. These observations are in agreement with the antagonistic effects between heat stress and the inflammatory mediators on the activation of Hsp promoter.
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Affiliation(s)
- Rose Njemini
- Geriatric Unit, Academic Hospital, Free University of Brussels, Brussels, Belgium
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18
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Njemini R, Demanet C, Mets T. Determination of intracellular heat shock protein 70 using a newly developed cell lysate immunometric assay. J Immunol Methods 2003; 274:271-9. [PMID: 12609553 DOI: 10.1016/s0022-1759(03)00004-8] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022]
Abstract
Heat shock proteins (Hsp) have been associated to several clinical relevant conditions. Currently used methods to determine Hsp 70 possess certain drawbacks. Therefore, we developed a cell lysate immunometric assay (CLIA) for the quantification of intracellular Hsp 70. This CLIA uses a combination of two distinct monoclonal antibodies that recognize different epitopes on the Hsp 70 molecule. A recombinant human Hsp 70 was used as the standard material. The detection range of the CLIA was 4-4000 ng/ml. The intra- and interassay coefficients of variation were, on average, 5% and 12%, respectively. The recovery varied between 81% and 116%. The Hsp 70 levels assayed after serial dilution of cell lysates varied linearly with dilution (between 97% and 120%). The reliability of the CLIA was assessed by comparison with the values determined by flow cytometric procedure; these two sets of values showed a highly significant correlation (r=0.896, p<0.0001), indicating that the two methods are comparable. We conclude that this assay represents a low-cost alternative of the flow cytometric technique.
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Affiliation(s)
- Rose Njemini
- Geriatric Unit, Academic Hospital, Free University of Brussels (VUB), Laarbeeklaan 101, B-1090, Brussels, Belgium
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19
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Tacchini L, Fusar-Poli D, Sironi M, Mantovani A, Bernelli-Zazzera A. Activation of signal transducer and activator of transcription 3 in rat liver after heat shock and reperfusion stress. Int J Biochem Cell Biol 2003; 35:316-23. [PMID: 12531244 DOI: 10.1016/s1357-2725(02)00164-4] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Changes in transcription factors (TFs) accompany many types of cell stresses. By using electrophoretic mobility assays we show that the DNA binding of signal transducer and activator of transcription 3 (STAT3) is activated in rat liver by heat shock and ischemia-reperfusion. Northern blot and Western blot analysis reveal an increase of the mRNA and protein level of this transcription factor. Under both conditions the phosphorylation of pre-existing STAT3 is prompt and precedes the increase in the STAT3 protein. The activation: (1) is functional, i.e. is followed by the transcription of the target gene alpha(1)-acid glycoprotein (2) is strongly inhibited by pretreatment with the interleukin-1 receptor antagonist before heat shock but only slightly by pretreatment before ischemia-reperfusion (3) might, at least in part, be mediated by a cytokine cascade involving also interleukin-6. These results are consistent with the hypothesis that different kinds of stress can activate a number of common TFs.
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Affiliation(s)
- Lorenza Tacchini
- Istituto di Patologia Generale dell'Università degli Studi di Milano, Centro di Studio sulla Patologia Cellulare del CNR, via Mangiagalli 31, 20133 Milan, Italy
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20
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Land W. Allograft injury mediated by reactive oxygen species: from conserved proteins of Drosophila to acute and chronic rejection of human transplants. Part II: Role of reactive oxygen species in the induction of the heat shock response as a regulator of innate. Transplant Rev (Orlando) 2003. [DOI: 10.1053/trre.2003.2] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/09/2023]
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Marciano PG, Eberwine JH, Ragupathi R, Saatman KE, Meaney DF, McIntosh TK. Expression profiling following traumatic brain injury: a review. Neurochem Res 2002; 27:1147-55. [PMID: 12462413 DOI: 10.1023/a:1020973308941] [Citation(s) in RCA: 38] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
Abstract
Traumatic brain injury (TBI) elicits a complex sequence of putative autodestructive and neuroprotective cellular cascades. It is hypothesized that the genomic responses of cells in the injured brain serve as the basis for these cascades. Traditional methods for analyzing differential gene expression following brain trauma demonstrate that immediate early genes, cytokines, transcription factors, and neurotrophic factors can all participate in the brain's active and directed response to injury, and may do so concurrently. It is this complexity and multiplicity of interrelated molecular mechanisms that has demanded new methods for comprehensive and parallel evaluation of putative as well as novel gene targets. Recent advances in DNA microarray technology have enabled the simultaneous evaluation of thousands of genes and the subsequent generation of massive amounts of biological data relevant to CNS injury. This emerging technology can serve to further current knowledge regarding recognized molecular cascades as well as to identify novel molecular mechanisms that occur throughout the post-traumatic period. The elucidation of the complex alterations in gene expression underlying the pathological sequelae following TBI is of central importance in the design of future therapeutic agents.
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Affiliation(s)
- Paolo G Marciano
- Department of Neuroscience, University of Pennsylvania, Philadelphia, USA
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22
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Dai CL, Xia ZL, Kume M, Yamamoto Y, Yamagami K, Ozaki N, Yamaoka Y. Heat shock protein 72 normothermic ischemia, and the impact of congested portal blood reperfusion on rat liver. World J Gastroenterol 2001; 7:415-8. [PMID: 11819802 PMCID: PMC4688734 DOI: 10.3748/wjg.v7.i3.415] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Affiliation(s)
- C L Dai
- Department of Surgery, The Second Clinical College of China Medical University, No.36 San Hao Street, He-Ping District, Shenyang 110003, Liaoning Province,China
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23
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Saito T, Ishii S, Abe T, Tsuchiya T, Kanno H, Miyazawa M, Suzuki M, Gotoh M. Effect of preconditioning in the liver against ischemia/reperfusion injury, protection of sinusoidal cells and alterations of gene transcription. Transplant Proc 2001; 33:849. [PMID: 11267096 DOI: 10.1016/s0041-1345(00)02345-9] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Affiliation(s)
- T Saito
- Department of Surgery I, School of Medicine, Fukushima Medical University, Fukushima, Japan
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24
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White BC, Sullivan JM, DeGracia DJ, O'Neil BJ, Neumar RW, Grossman LI, Rafols JA, Krause GS. Brain ischemia and reperfusion: molecular mechanisms of neuronal injury. J Neurol Sci 2000; 179:1-33. [PMID: 11054482 DOI: 10.1016/s0022-510x(00)00386-5] [Citation(s) in RCA: 600] [Impact Index Per Article: 24.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022]
Abstract
Brain ischemia and reperfusion engage multiple independently-fatal terminal pathways involving loss of membrane integrity in partitioning ions, progressive proteolysis, and inability to check these processes because of loss of general translation competence and reduced survival signal-transduction. Ischemia results in rapid loss of high-energy phosphate compounds and generalized depolarization, which induces release of glutamate and, in selectively vulnerable neurons (SVNs), opening of both voltage-dependent and glutamate-regulated calcium channels. This allows a large increase in cytosolic Ca(2+) associated with activation of mu-calpain, calcineurin, and phospholipases with consequent proteolysis of calpain substrates (including spectrin and eIF4G), activation of NOS and potentially of Bad, and accumulation of free arachidonic acid, which can induce depletion of Ca(2+) from the ER lumen. A kinase that shuts off translation initiation by phosphorylating the alpha-subunit of eukaryotic initiation factor-2 (eIF2alpha) is activated either by adenosine degradation products or depletion of ER lumenal Ca(2+). Early during reperfusion, oxidative metabolism of arachidonate causes a burst of excess oxygen radicals, iron is released from storage proteins by superoxide-mediated reduction, and NO is generated. These events result in peroxynitrite generation, inappropriate protein nitrosylation, and lipid peroxidation, which ultrastructurally appears to principally damage the plasmalemma of SVNs. The initial recovery of ATP supports very rapid eIF2alpha phosphorylation that in SVNs is prolonged and associated with a major reduction in protein synthesis. High catecholamine levels induced by the ischemic episode itself and/or drug administration down-regulate insulin secretion and induce inhibition of growth-factor receptor tyrosine kinase activity, effects associated with down-regulation of survival signal-transduction through the Ras pathway. Caspase activation occurs during the early hours of reperfusion following mitochondrial release of caspase 9 and cytochrome c. The SVNs find themselves with substantial membrane damage, calpain-mediated proteolytic degradation of eIF4G and cytoskeletal proteins, altered translation initiation mechanisms that substantially reduce total protein synthesis and impose major alterations in message selection, down-regulated survival signal-transduction, and caspase activation. This picture argues powerfully that, for therapy of brain ischemia and reperfusion, the concept of single drug intervention (which has characterized the approaches of basic research, the pharmaceutical industry, and clinical trials) cannot be effective. Although rigorous study of multi-drug protocols is very demanding, effective therapy is likely to require (1) peptide growth factors for early activation of survival-signaling pathways and recovery of translation competence, (2) inhibition of lipid peroxidation, (3) inhibition of calpain, and (4) caspase inhibition. Examination of such protocols will require not only characterization of functional and histopathologic outcome, but also study of biochemical markers of the injury processes to establish the role of each drug.
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Affiliation(s)
- B C White
- Department of Emergency Medicine, Wayne State University School of Medicine, Detroit, MI, USA.
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25
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Wieland E, Oellerich M, Braun F, Schtüz E. c-fos and c-jun mRNA expression in a pig liver model of ischemia/reperfusion: effect of extended cold storage and the antioxidant idebenone. Clin Biochem 2000; 33:285-90. [PMID: 10936587 DOI: 10.1016/s0009-9120(00)00070-9] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Abstract
OBJECTIVES Expression of immediate early genes has been reported during reperfusion after ischemia in rat livers due to oxygen radical formation. This study investigates in perfused pig livers the effect of the antioxidant idebenone and of cold ischemia time on the gene expression of c-fos and c-jun. DESIGN AND METHODS Livers were perfused for 210 min after 0.5 h or 20 h ischemic storage (4 degrees C). One group of pigs was fed idebenone (280 mg/day/7days) prior to organ harvesting. C-fos and c-jun mRNA were determined by RT-PCR at 3, 30, 60, 120 180, 210 min during reperfusion. RESULTS Lipid peroxidation increased in liver tissue form 0.54 +/- 0.21 to 1. 09 +/- 0.54 nmol MDA/mg protein during reperfusion after 20 h compared to 0.5 h cold storage. This was antagonized by idebenone (0. 68 +/- 0.20 nmol/MDA/mg protein). C-fos and c-jun were strongly induced in livers stored for 20 h, which was attenuated by idebenone (p < 0.05). CONCLUSIONS These findings suggest that cold ischemia time and oxygen radicals are critical for immediate early gene expression and that application of an effective antioxidant can attenuate this early stress reaction of the pig liver.
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Affiliation(s)
- E Wieland
- Abteilung Klinische Chemie, Georg-August-Universität, Göttingen, Germany.
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26
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Itoh H, Yagi M, Fushida S, Tani T, Hashimoto T, Shimizu K, Miwa K. Activation of immediate early gene, c-fos, and c-jun in the rat small intestine after ischemia/reperfusion. Transplantation 2000; 69:598-604. [PMID: 10708117 DOI: 10.1097/00007890-200002270-00022] [Citation(s) in RCA: 40] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
BACKGROUND Activated immediate early genes (IEGs) play key roles in mediating cellular response after ischemia/reperfusion (I/R) injuries in some organs such as liver, heart and kidney. However, there is no report investigating an association between the activation of IEGs and cellular regeneration or programmed cell death after I/R in small intestine. METHODS We examined a sequential expression of c-fos and c-jun after I/R in rat small intestine using reverse transcription-polymerase chain reaction and Northern blot analysis, and compared the patterns with coexistent two parameters: (1) regeneration determined by immunohistochemical detection of proliferating cell nuclear antigen, (2) programmed cell death determined with the terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end-labeling (TUNEL) method and DNA fragmentation. RESULTS The expression of c-fos and c-jun mRNA increased markedly 15 min after reperfusion and was, respectively, 6.3 and 4.4 times higher than in controls. Proliferating cell nuclear antigen expression was significantly elevated between 5 min and 4 hr, peaking at 30 min after reperfusion. Apoptosis showed a peak 60 min after reperfusion. Apoptosis after I/R was detected in the nuclei of absorptive epithelial cells by the TUNEL method, and these apoptotic signals were consistent with the expression of c-Fos and c-Jun proteins using an immunohistochemical method. CONCLUSIONS These results suggest that overexpression of c-fos and c-jun after I/R in the small intestine correlates with programmed cell death and subsequent cellular regeneration.
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Affiliation(s)
- H Itoh
- Department of Surgery (II), School of Medicine, Kanazawa University, Ishikawa, Japan
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Papathanassoglou ED, Moynihan JA, Ackerman MH. Does programmed cell death (apoptosis) play a role in the development of multiple organ dysfunction in critically ill patients? a review and a theoretical framework. Crit Care Med 2000; 28:537-49. [PMID: 10708197 DOI: 10.1097/00003246-200002000-00042] [Citation(s) in RCA: 128] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
OBJECTIVES To critically review the current understanding of the pathophysiologic events leading to the development of secondary multiple organ dysfunction (MODS) in critical illness and to examine the role of apoptosis (programmed cell death) as a mechanism involved in the progression of MODS. DATA SOURCES Research and review articles published since 1982 on the pathophysiology of MODS, particularly the role of cytokines, reactive oxygen species, heat shock proteins, and apoptosis. Research and review articles on the physiology of apoptosis. Articles include human/animal and in vitro/in vivo studies. DATA EXTRACTION The most prevalent mediating factors of MODS were examined for their potential to induce apoptosis, as reported in the literature. The combination of several of the above factors was also examined in terms of apoptosis-triggering potential. DATA SYNTHESIS Specific pathophysiologic conditions related to the onset of MODS have been shown to affect apoptotic rates in organ tissue cells and their respective endothelial cells in animal and in vitro models. These conditions include the following: a) increased release of inflammation-related cytokines; b) increased production of oxygen free radicals associated with ischemia/reperfusion injury and states of low tissue perfusion; c) expression and release of heat shock proteins from tissue cells and the liver; d) elevated glucocorticoid concentrations after adrenal cortex activation; and e) release of bacterial products into the systemic circulation. CONCLUSION The most important MODS-related pathophysiologic conditions known to date have been shown to affect programmed cell death rates in almost all cell types. Organ-specific cell death involving both parenchymal and microvasculature endothelial cells is conceivably underlying organ dysfunction. The hypothesis that increased apoptotic rates are involved in organ dysfunction may provide a unifying theory for the pathophysiology of MODS.
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Affiliation(s)
- E D Papathanassoglou
- Division of Endocrinology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215, USA.
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28
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Tunici P, Schiaffonati L, Rabellotti E, Tiberio L, Perin A, Sessa A. In Vivo Modulation of 73 kDa Heat Shock Cognate and 78 kDa Glucose-Regulating Protein Gene Expression in Rat Liver and Brain by Ethanol. Alcohol Clin Exp Res 1999. [DOI: 10.1111/j.1530-0277.1999.tb04084.x] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
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Tacchini L, Radice L, Bernelli-Zazzera A. Differential activation of some transcription factors during rat liver ischemia, reperfusion, and heat shock. J Cell Physiol 1999; 180:255-62. [PMID: 10395295 DOI: 10.1002/(sici)1097-4652(199908)180:2<255::aid-jcp13>3.0.co;2-l] [Citation(s) in RCA: 38] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
Cells respond to external stimuli by changes in gene expression that are largely dependent on transcription factors (TFs). We studied the behavior of some TFs in rat liver during ischemia, postischemic reperfusion, and heat shock. Knowledge of the conditions at the end of ischemia is essential to understand changes occurring at reperfusion. The TFs investigated are known to be typically responsive to heat shock (HSF), hypoxia (HIF-1), pro- and antioxidant conditions (AP-1), or to various environmental changes (HNF-1 and ATF/CREB family). The most relevant new information includes the following: 1) Liver ischemia activates extremely rapidly the DNA binding capacity of HSF, soon followed by analogous activation of HIF-1 and AP-1. 2) After a certain lag time from the activation of HIF-1, mRNAs accumulate for two glycolytic enzymes, in particular Aldolase A and Heme Oxygenase 1, which contain HIF-1 sequences in their promoters. 3) Reperfusion, which is known to further increase the binding of HSF and to induce NFkappaB binding, abrogates or decreases the binding of HIF-1 and AP-1, stimulated by ischemia, and activates the binding of ATF/CREB. Later on, a second peak of AP-1 binding is induced. 4) Heat shock activates both ischemia-responsive and reperfusion-responsive TFs. 5) Preliminary experiments of supergelshift reveal that the activation of AP-1 at reperfusion or upon heat shock may result from the different involvement of the component subunits.
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Affiliation(s)
- L Tacchini
- Istituto di Patologia Generale dell'Università degli Studi di Milano, Centro di Studio sulla Patologia Cellulare del CNR, Milan, Italy
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30
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Dai CL, Kume M, Yamamoto Y, Yamagami K, Yamamoto H, Nakayama H, Ozaki N, Shapiro AM, Yamamoto M, Yamaoka Y. Heat shock protein 72 production in liver tissue after experimental total hepatic inflow occlusion. Br J Surg 1998; 85:1061-5. [PMID: 9717996 DOI: 10.1046/j.1365-2168.1998.00771.x] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
BACKGROUND The pathogenesis of hepatic ischaemia-reperfusion injury is incompletely understood. This study examined the effects of reperfusion with congested portal blood on ischaemia-reperfusion injury of the liver following Pringle's manoeuvre, as monitored by heat shock protein (HSP) 72 production in rat liver tissue. METHODS Rats were randomized to three groups. In group 1 hepatic ischaemia with portal congestion was induced by Pringle's manoeuvre for 15 min; in group 2 Pringle's manoeuvre was applied for 15 min with an extracorporeal portasystemic shunt; and in group 3 the superior mesenteric vein was occluded for 15 min. The production of HSP72 in liver tissue was measured by Western blotting at 48 h after each intervention. Conventional parameters for hepatic function were examined at 1, 3 and 48 h after reperfusion. RESULTS There was marked HSP72 expression in group 1, but not in group 2 or 3, showing that a combination of liver ischaemia and reperfusion of congested portal blood is required to induce strong expression of HSP72 in the tissue. On the other hand, biochemical parameters were raised equally in both groups 1 and 2, reflecting a similar degree of ischaemic hepatocyte injury. CONCLUSION The additional stress impact of temporary portal occlusion upon ischaemia-reperfusion injury of the liver was clearly detected by in situ hepatic HSP72 production in this study.
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Affiliation(s)
- C L Dai
- Department of Gastroenterological Surgery, Kyoto University Graduate School of Medicine, Sakyo, Japan
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Saito T, Matsumoto I, Goto S, Kamada N, Motoki R, Wilce PA. The differential induction of two immediate early genes, c-fos and c-jun, after systemic hypovolemic shock/resuscitation in the rat liver and kidney. Surg Today 1998; 28:608-17. [PMID: 9681610 DOI: 10.1007/s005950050193] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
The aim of this study was to investigate the expression of the immediate early genes (IEGs), c-fos and c-jun, in the rat kidney and liver in two types of hemorrhage shock/resuscitation models. In the first group, hemorrhagic shock was induced by the withdrawal of blood through the carotid artery. A mean arterial blood pressure (MAP) of 40mmHg was maintained for 1h before blood was reperfused. In the second group, the MAP was maintained at the same level for 2h. Animals were resuscitated with Ringer's lactate solution. In the first group, a rapid and transient induction of c-fos and c-jun mRNAs in both the liver and kidney was observed, peaking 0 to 2 h after reperfusion. In the second group, a more protracted pattern of induction was evident in both organs. In both models, the induction of c-fos mRNA was distinctly different in the liver and kidney. These results indicated, first, that with respect to IEG expression, organs respond differently to a systemic shock/resuscitation stimuli, and second, that alterations in the pattern of IEG expression might represent an indication of the degree of organ damage or the repair processes subsequent to hypotension/reperfusion.
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Affiliation(s)
- T Saito
- First Department of Surgery, Fukushima Medical College, Hikarigaoka, Japan
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32
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Flanagan SW, Moseley PL, Buettner GR. Increased flux of free radicals in cells subjected to hyperthermia: detection by electron paramagnetic resonance spin trapping. FEBS Lett 1998; 431:285-6. [PMID: 9708920 DOI: 10.1016/s0014-5793(98)00779-0] [Citation(s) in RCA: 150] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023]
Abstract
It has been hypothesized that hyperthermia promotes oxygen-centered free radical formation in cells; however, to date there is no direct evidence of this heat-induced increase in oxygen free radical flux. Using electron paramagnetic resonance (EPR) spin trapping, we sought direct evidence for free radical generation during hyperthermia in intact, functioning cells. Rat intestinal epithelial cell monolayers were exposed to 45 degrees C for 20 min, after which the nitrone spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) was added. Compared to control cells at 37 degrees C, heat-exposed cells had increased free radical EPR signals, consistent with the formation of DMPO/.OH (aN = aH = 14.9 G). These findings indicate that heat increases the flux of cellular free radicals and support the hypothesis that increased generation of oxygen-centered free radicals and the resultant oxidative stress may mediate in part, heat-induced cellular damage.
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Affiliation(s)
- S W Flanagan
- Department of Exercise Science, The University of Iowa, Iowa City 52242, USA.
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33
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Hori T, Wanibuchi H, Yano Y, Otani S, Nishikawa A, Osugi H, Kinoshita H, Fukushima S. Epithelial cell proliferation in the digestive tract induced by space restriction and water-immersion stress. Cancer Lett 1998; 125:141-8. [PMID: 9566708 DOI: 10.1016/s0304-3835(97)00504-1] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
The effects of space restriction and water-immersion stress on epithelial cell proliferation in the digestive tract, with special attention to the esophagus, stomach and duodenum, in 8-week-old SD male rats were examined. Histological assessment revealed spotted hemorrhagic lesions in the fundus of the glandular stomach, accompanied by statistically increased 5-bromo-2'-deoxyuridine (BrdU) labeling index in the fundic and pyloric regions. Furthermore, biochemical analysis demonstrated an increased activity of ornithine decarboxylase (ODC) and spermidine/spermine N1-acetyltransferase (SAT), known as key late-limiting enzymes of the polyamine pathway, in the gastric fundus. The stress may induce a remarkable increase in expression of c-fos, c-jun and c-myc mRNAs in both fundic and pyloric regions of the glandular stomach. There were no remarkable changes in the esophagus. These results indicate that space restriction and water-immersion stress induced cell proliferation in the glandular stomach through overexpression of proto-oncogenes and increased ODC and SAT activities that might be related to the promotion of gastric carcinogenesis.
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Affiliation(s)
- T Hori
- First Department of Pathology, Osaka City University Medical School, Osaka, Japan
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34
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Schütz E, Wieland E, Heine L, Hensel A, Schmiedl A, Armstrong VW, Richter J, Schuff-Werner P, Günther E, Oellerich M. Acceleration of hepatocellular energy by idebenone during early reperfusion after cold preservation ameliorates heat shock protein 70 gene expression in a pig liver model. Transplantation 1997; 64:901-7. [PMID: 9326418 DOI: 10.1097/00007890-199709270-00019] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/05/2023]
Abstract
BACKGROUND Heat shock proteins (HSPs) are induced in the liver after warm ischemia/reperfusion and are thought to be markers of hepatocellular injury and oxidative stress. METHODS The influence of variable periods of cold storage followed by reperfusion on the expression of HSP70 was studied in the isolated perfused pig liver. Organs were harvested and stored in histidine-tryptophan-ketoglutarate solution at 4 degrees C and then perfused (210 min) in a closed water bath (38 degrees C), which subjects the liver to fluctuating outer pressure. The role of energy depletion, reactive oxygen intermediates, Kupffer cells, and circulating leukocytes in HSP70 expression was determined. RESULTS HSP70 expression was not detectable in liver tissue before explantation or before reperfusion by Northern blot analysis using a pig HSP70 gene probe. HSP70 expression was observed after reperfusion depending on cold storage time. Kinetics of HSP70 expression monitored by reverse transcriptase polymerase chain reaction showed a rapid increase of mRNA within 1 hr, which was closely associated with delayed recovery of hepatocellular energy charge, as assessed by the ketone body ratio. The inactivation of Kupffer cells, the presence or absence of leukocytes, and the suppression of oxidative stress with the antioxidant idebenone, given during reperfusion, had no influence. However, feeding the animals with idebenone over 7 days before explantation led to a faster recovery of ketone body ratio, paralleled by a substantial suppression of HSP70 expression. CONCLUSIONS Our data show that HSP70 expression during reperfusion is mainly dependent on the preceding cold storage time and the consecutive delayed recovery of the hepatocellular energy charge.
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Affiliation(s)
- E Schütz
- Abteilung Klinische Chemie, Georg-August-Universität, Göttingen, Germany
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Schiaffonati L, Tiberio L. Gene expression in liver after toxic injury: analysis of heat shock response and oxidative stress-inducible genes. LIVER 1997; 17:183-91. [PMID: 9298488 DOI: 10.1111/j.1600-0676.1997.tb00804.x] [Citation(s) in RCA: 56] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/05/2023]
Abstract
In the liver, CCl4 induces cell necrosis followed by regeneration. Cell injury is caused by free radical damage and may be due, at least in part, to oxidative stress and the subsequent formation of reactive oxygen intermediates (ROIs). In a rat model of acute CCl4-induced hepatic injury, we examined the expression of genes involved in cellular response to different kinds of stress, including oxidative stress (hsp 70 family, heme oxygenase), in free radical detoxification (Mn superoxide dismutase and Cu/ Zn superoxide dismutase), in iron homeostasis (H and L ferritin subunits) and in the cell cycle (c-fos, c-jun, histone H3). As an experimental approach, we first analysed the pattern of protein synthesised by liver slices in vitro. Then we studied the mechanisms regulating the expression of different genes, by analysing both mRNA steady state levels and transcription rates. Activation of the specific heat shock transcription factor (HSF) by CCl4 was also investigated. We observed that different members of the hsp70 family (hsp70, hsc73, grp78) are activated by different kinetics and are regulated mainly at the transcriptional level. Induction of the hsp70 gene occurs rapidly and transiently and is preceded by the activation of HSF DNA-binding activity. We demonstrated an increase in the steady-state levels of mRNAs for heme oxygenase, Mn and Cu/Zn superoxide dismutases and H and L ferritin subunits. However, different kinetics and regulatory mechanisms occurred with different genes. We showed that induction of c-fos and c-jun protooncogenes is the earliest event after CCl4 administration, whereas histone H3 expression peaked at 24-48 h. The results of this study are interpreted as evidence that activation of specific stress response genes is primarily related to the defence against the rapidly occurring cell damage, but may also be related to subsequent processes of tissue inflammation and cell proliferation.
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Affiliation(s)
- L Schiaffonati
- Dipartimento di Scienze Biomediche e Biotecnologie, Università degli Studi di Brescia, Italy
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36
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Laderoute KR, Webster KA. Hypoxia/reoxygenation stimulates Jun kinase activity through redox signaling in cardiac myocytes. Circ Res 1997; 80:336-44. [PMID: 9048653 DOI: 10.1161/01.res.80.3.336] [Citation(s) in RCA: 144] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/03/2023]
Abstract
Hypoxia and reoxygenation are principal components of myocardial ischemia and reperfusion and have distinctive effects on the tissue. Both conditions have been associated with inflammation, necrosis, apoptosis, and myocardial infarction. Using a cell culture model of ischemia and reperfusion in which cardiac myocytes were exposed to cycles of hypoxia and reoxygenation, we report here that reoxygenation, but not hypoxia alone, caused sustained approximately 10-fold increases in phosphorylation of the amino-terminal domain of the c-jun transcription factor. The activation was similar to treatments with anisomycin or okadaic acid and correlated with the hypoxia-mediated depression of intracellular glutathione. Reoxygenation-induced c-Jun kinase activity was reduced by preincubating myocytes during the hypoxia phase with the spin-trap agent alpha-phenyl N-tert-butylnitrone or with N-acetylcysteine. The kinase activation was also inhibited by the tyrosine kinase inhibitor genistein but not by other protein kinase inhibitors. These results implicate unquenched reactive oxygen intermediates as the stimulus that initiates a kinase pathway involving the stress-activated protein kinases (JNKs/SAPKs) in reoxygenated cardiac myocytes.
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Affiliation(s)
- K R Laderoute
- Department of Cell and Molecular Biology, SRI International, Menlo Park, Calif, USA
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37
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Aoe T, Inaba H, Kon S, Imai M, Aono M, Mizuguchi T, Saito T, Nishino T. Heat shock protein 70 messenger RNA reflects the severity of ischemia/hypoxia-reperfusion injury in the perfused rat liver. Crit Care Med 1997; 25:324-9. [PMID: 9034272 DOI: 10.1097/00003246-199702000-00022] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/03/2023]
Abstract
OBJECTIVES To determine whether ischemia-reperfusion and hypoxia-reoxygenation cause cellular damages and stress responses in an isolated perfused rat liver model. To determine whether the increased synthesis of stress protein messenger RNA reflects cellular injury. DESIGN Prospective, controlled study. SETTING Institutional laboratories. SUBJECTS Male Sprague-Dawley rats. INTERVENTIONS Isolated rat livers with cell free perfusion were exposed to various periods of ischemia-reperfusion or hypoxia-reoxygenation. MEASUREMENTS AND MAIN RESULTS We measured hepatic oxygen consumption and alanine aminotransferase leakage from liver during perfusion. We analyzed the gene expression of heat shock protein 70, a major stress protein, of the liver by Northern blotting after perfusion. The expression of heat shock protein 70 messenger RNA augmented as the reperfusion period increased. The expression level after graded ischemia or hypoxia significantly correlated with the calculated hepatic oxygen debt (r2 = .737; p < .001; n = 21), or with the accumulated alanine aminotransferase leakage from the liver (r2 = .509; p < .001; n = 21). CONCLUSIONS These results suggest that the accumulation of heat shock protein 70 messenger RNA reflects the severity of ischemia-reperfusion and hypoxia-reoxygenation injuries, and that a stress response in reperfusion can be triggered without formed elements of blood.
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Affiliation(s)
- T Aoe
- Department of Anesthesiology, Chiba University School of Medicine, Japan
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38
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Gasbarrini A, Colantoni A, Di Campli C, De Notariis S, Masetti M, Iovine E, Mazziotti A, Massari I, Gasbarrini G, Pola P, Bernardi M. Intermittent anoxia reduces oxygen free radicals formation during reoxygenation in rat hepatocytes. Free Radic Biol Med 1997; 23:1067-72. [PMID: 9358250 DOI: 10.1016/s0891-5849(97)00141-x] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/05/2023]
Abstract
The sensitivity of liver cells to anoxia is a major problem afflicting liver preservation and transplantation. Intermittent ischemia has been proposed to reduce reperfusion injury. The aim of the study was to assess oxygen free radical formation and cell injury during continuous or intermittent anoxia/reoxygenation in rat hepatocytes. Anion superoxide was measured by lucigenin-enhanced chemiluminescence and cell damage by LDH release and trypan blue uptake. During anoxia, superoxide generation dropped to background level in both groups; trypan blue uptake and LDH release, which increased progressively, were significantly greater in hepatocytes exposed to continuous compared to intermittent anoxia. During reoxygenation, a massive generation of superoxide anion formation, followed by a sharp increase in LDH release, was observed in both groups. However, both oxyradical generation and cell injury were significantly greater in cells exposed to continuous compared to intermittent anoxia. The data, showing that intermittent oxygen deprivation reduce liver cell injury and oxygen free radical formation determined by anoxia/reoxygenation, suggest a novel possible approach to the reduction of reperfusion injury.
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Bendinelli P, Piccoletti R, Maroni P, Bernelli-Zazzera A. The MAP kinase cascades are activated during post-ischemic liver reperfusion. FEBS Lett 1996; 398:193-7. [PMID: 8977105 DOI: 10.1016/s0014-5793(96)01228-8] [Citation(s) in RCA: 56] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/03/2023]
Abstract
We have investigated the involvement of MAP kinase cascades in the response of the liver to post-ischemic reperfusion. Both JNKs and ERKs are activated but the duration and magnitude of the increase in their activities appear to be different. JNK activation is more marked but shorter than that of ERKs. The increase observed in the phosphotyrosine content of the 52 kDa Shc protein, accompanied by an increased amount of co-immunoprecipitated Grb2, and the activation of Raf-1 kinase provide evidence of the involvement of a Ras-Raf-dependent pathway, with a time course that is similar to that of ERK activation. The treatment of rats with IL-1 receptor antagonist modified all of the described effects, suggesting that IL-1 plays a role in the response of the liver to reperfusion.
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Affiliation(s)
- P Bendinelli
- Istituto di Patologia Generale dell'Università degli Studi di Milano, Italy
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40
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Cairo G, Pietrangelo A. Nitric-oxide-mediated activation of iron-regulatory protein controls hepatic iron metabolism during acute inflammation. EUROPEAN JOURNAL OF BIOCHEMISTRY 1995; 232:358-63. [PMID: 7556182 DOI: 10.1111/j.1432-1033.1995.358zz.x] [Citation(s) in RCA: 46] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/25/2023]
Abstract
The molecular regulation of intracellular iron metabolism has been studied in the livers of rats undergoing an acute inflammatory reaction following turpentine injection. Treatment induced an increase in the steady-state level of the transferrin receptor (TfR) mRNA, peaking 18 h after treatment and returning to control levels 24 h after treatment, with no change in TfR gene transcription. RNA band-shift assays documented an activation of the cytoplasmic RNA-binding protein called the iron-regulatory protein (IRP), in parallel with a rise in the amount of TfR transcripts. A 2-3-fold increase in the amount of H and L ferritin subunit mRNAs was found 12-18 h after turpentine treatment. Surprisingly, higher accumulation of ferritin mRNAs did not result in appreciable differences in the liver ferritin content. This might be due to the concomitant rise of IRP activity, which is known to prevent ferritin mRNA translation. The absence of significant changes in the total iron and ferritin contents prompted us to investigate the role of nitric oxide (NO), an inflammatory mediator which is also known to modulate the activity of IRP. Northern-blot analysis showed a marked enhancement in the expression of the inducible form of nitric oxide synthase mRNA in turpentine-treated rats. Furthermore, the activation of IRP and the increase of the TfR mRNA content that occur in turpentine-treated rats were abolished by treatment with N5-nitro-L-arginine, a specific nitric oxide synthase inhibitor. The present data suggest that NO-mediated activation of IRP regulates alterations of hepatic iron homeostasis that occur in acute inflammation.
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Affiliation(s)
- G Cairo
- Centro di Studio sulla Patologia Cellulare, CNR, Milano, Italy
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41
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Tacchini L, Pogliaghi G, Radice L, Anzon E, Bernelli-Zazzera A. Differential activation of heat-shock and oxidation-specific stress genes in chemically induced oxidative stress. Biochem J 1995; 309 ( Pt 2):453-9. [PMID: 7626009 PMCID: PMC1135753 DOI: 10.1042/bj3090453] [Citation(s) in RCA: 39] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/26/2023]
Abstract
Post-ischaemic reperfusion increases the level of the major heat-shock (stress) protein hsp 70 and of its mRNA by transcriptional mechanisms, and activates the binding of the heat-shock factor HSF to the consensus sequence HSE. In common with CoCl2 treatment, post-ischaemic reperfusion increases the level of haem oxygenase mRNA, an indicator of oxidative stress, but CoCl2 does not seem to induce the expression of the hsp 70 gene [Tacchini, Schiaffonati, Pappalardo, Gatti and Bernelli-Zazzera (1993) Lab. Invest. 68, 465-471]. Starting from these observations, we have now studied the expression of two genes of the hsp 70 family and of other possibly related genes under conditions of oxidative stress. Three different chemicals, which cause oxidative stress by various mechanisms and induce haem oxygenase, enhance the expression of the cognate hsc 73 gene, but do not activate the inducible hsp 70 gene. Expression of the other genes that have been studied seems to vary in intensity and/or time course, in relation to the particular mechanism of action of any single agent. The pattern of induction of the early-immediate response genes c-fos and c-jun observed during oxidative stress differs from that found in post-ischaemic reperfused livers. Oxidative-stress-inducing agents do not promote the binding of HSF to its consensus sequence HSE, such as occurs in heat-shock and post-ischaemic reperfusion, and fail to activate AP-1 (activator protein 1). With the possible exception of Phorone, the oxidative stress chemically induced in rat liver activates NFkB (nuclear factor kB) and AP-2 (activator protein 2) transcription factors.
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Affiliation(s)
- L Tacchini
- Istituto di Patologia Generale dell'Università degli Studi di Milano, Centro di Studio sulla Patologia Cellulare del CNR, Italy
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42
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Joannidis M, Cantley LG, Spokes K, Medina R, Pullman J, Rosen S, Epstein FH. Induction of heat-shock proteins does not prevent renal tubular injury following ischemia. Kidney Int 1995; 47:1752-9. [PMID: 7643546 DOI: 10.1038/ki.1995.242] [Citation(s) in RCA: 32] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/26/2023]
Abstract
The possible protective effect of heat-shock proteins (HSPs) on ischemic injury to renal cells was assessed in two different experimental models: ischemia-reflow in intact rats and medullary hypoxic injury as seen in the isolated perfused rat kidney. Heat shock was induced by raising the core temperature of rats to 42 degrees C for 15 minutes. Following this, Northern blots showed enhanced gene expression of HSP70, HSP60 and ubiquitin at one hour and reaching a maximum by six hours after heat shock in all regions of the kidney, but most prominently in medulla and papilla. The HSP70 protein in the kidney, estimated by immunohistochemical means, was detectable 24 hours following heat shock and further increased at 48 hours following heat shock. In the first set of experiments, the animals underwent uninephrectomy followed by cross clamping of the remaining renal artery for 40 minutes prior to reflow. Serum creatinine and urea nitrogen rose to 3.15 +/- 0.98 and 126.4 +/- 62.5 mg/dl at 24 hours. No significant differences were observed at 24, 48 and 72 hours after reflow between these values in control rats and rats pretreated with heat shock 48 hours earlier. Severe morphological damage to proximal tubules of the renal cortex was observed to the same extent in both groups. In a second set of experiments, the right kidney was removed either 24 or 48 hours after heat shock and perfused in isolation for 90 minutes. Functional and morphological parameters were compared with those of isolated perfused kidneys obtained from animals that had not been subjected to heat shock.(ABSTRACT TRUNCATED AT 250 WORDS)
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Affiliation(s)
- M Joannidis
- Department of Medicine, Beth Israel Hospital, Boston, Massachusetts, USA
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Cairo G, Tacchini L, Pogliaghi G, Anzon E, Tomasi A, Bernelli-Zazzera A. Induction of ferritin synthesis by oxidative stress. Transcriptional and post-transcriptional regulation by expansion of the "free" iron pool. J Biol Chem 1995; 270:700-3. [PMID: 7822298 DOI: 10.1074/jbc.270.2.700] [Citation(s) in RCA: 261] [Impact Index Per Article: 8.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/27/2023] Open
Abstract
Ferritin, by regulating the "free" intracellular iron pool, controls iron-catalyzed generation of reactive oxygen species, but its role in oxidative damage is still unclear. We show that ferritin synthesis is significantly stimulated in the liver of rats subjected to oxidative stress by treatment with phorone, a glutathione-depleting drug. RNA-bandshift assays document reduced activity of iron regulatory factor, in particular of IRFB, the cytoplasmic protein that post-transcriptionally controls ferritin mRNA translation. Furthermore, Northern blot analysis shows increased accumulation of H and L subunit mRNAs, and nuclear run-on experiments provide evidence of transcriptional activation. Direct measurements of intracellular free iron levels by EPR indicate that the increased ferritin synthesis can be mediated by an expansion of the free iron pool. An early drop of ferritin content after phorone treatment indicates that part of the iron that fuels the free pool might derive from ferritin degradation. Present data seem to suggest that, under conditions of oxidative stress, liver ferritin can represent either a pro- or an anti-oxidant in a time-dependent manner. In fact, its early degradation contributes to expand the intracellular free iron pool that, later on, activates multiple molecular mechanisms to reconstitute ferritin content, thus limiting the pro-oxidant challenge of iron.
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Affiliation(s)
- G Cairo
- Centro di Studio sulla Patologia Cellulare, Università di Milano, Italy
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44
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Ikeda J, Nakajima T, Osborne OC, Mies G, Nowak TS. Coexpression of c-fos and hsp70 mRNAs in gerbil brain after ischemia: induction threshold, distribution and time course evaluated by in situ hybridization. BRAIN RESEARCH. MOLECULAR BRAIN RESEARCH 1994; 26:249-58. [PMID: 7854054 DOI: 10.1016/0169-328x(94)90097-3] [Citation(s) in RCA: 46] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/27/2023]
Abstract
Levels of mRNAs encoding the proto-oncogene, c-fos, and the 70 kDa stress protein, hsp70, were evaluated in gerbil brain following transient cerebral ischemia of varied duration by in situ and blot hybridization techniques. Blots of total hippocampal RNA obtained after 5 min ischemic insults confirmed a characteristic, transient time course of c-fos expression with a striking elevation within 1 h and a return to control levels by 3 h recirculation. Hsp70 hybridization was significant at 1 h and continued to increase until 3-6 h after the insult. Striking accumulation of c-fos mRNA was detected within 15 min recirculation in dentate granule cells, persisting through 1 h, and a weaker signal was evident in CA1 and CA3 pyramidal neurons of hippocampus, as well as in prepiriform/entorhinal cortex and neocortical regions, during the same interval. Hsp70 hybridization showed an identical distribution at 1 h recirculation. Ischemic insults of 1 min duration resulted in no detectable increase of either mRNA, while 2 min ischemia resulted in changes comparable to those seen after 5 min insults. This common threshold corresponds to the ischemic interval required for energy depletion and resultant failure of intracellular ion homeostasis. In contrast, expression of hsp70 mRNA was not observed under conditions of brief depolarization accompanying cortical or hippocampal spreading depression that were shown to induce c-fos. A delayed component of c-fos mRNA expression was not detected in this model, while persistent hsp70 hybridization, restricted to hippocampal CA1 neurons, was evident at 48 h after either 2 min or 5 min ischemic insults. The parallels in c-fos and hsp70 mRNA expression during early recirculation suggest that overlapping mechanisms triggered following postischemic depolarization contribute to their induction after transient ischemia.
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Affiliation(s)
- J Ikeda
- Laboratory of Neuropathology and Neuroanatomical Sciences, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892
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45
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Schiaffonati L, Tacchini L, Pappalardo C. Heat shock response in the liver: expression and regulation of the hsp70 gene family and early response genes after in vivo hyperthermia. Hepatology 1994; 20:975-83. [PMID: 7927240 DOI: 10.1002/hep.1840200429] [Citation(s) in RCA: 19] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/27/2023]
Abstract
Heat shock response in cultured cells has been studied extensively; however few data are available on heat shock response in an intact organ of a living animal. In this study we analyzed the kinetics of expression of the heat shock protein 70 gene family (heat shock protein 70, heat shock cognate protein 73 and glucose-regulated protein 78) in the liver of the thermally stressed rat. New synthesis of heat shock protein 70 and heat shock cognate protein 73 was shown in liver slices pulse labeled in vitro with 35S-methionine. Accumulation of heat shock protein 70 and heat shock cognate protein 73 proteins was shown in total cellular extracts. 32P-labeled complementary DNA probes encoding heat shock protein 70, heat shock cognate protein 73 and glucose-regulated protein 78 were used to show that the levels of the corresponding messenger RNAs increase as a fraction of total RNA and in polysomes at different extents and with different kinetics. The induction of heat shock protein 70 and heat shock cognate protein 73 messenger RNAs reflected the increase in the synthesis of the corresponding proteins. Run-on transcription analysis indicated that the expression of heat shock protein 70 and heat shock cognate protein 73 genes was mainly regulated at the transcriptional level. On the contrary, both transcriptional and posttranscriptional regulatory mechanisms can explain the induction of the glucose-regulated protein 78 gene.(ABSTRACT TRUNCATED AT 250 WORDS)
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Affiliation(s)
- L Schiaffonati
- Istituto di Patologia Generale dell'Universitá di Milano, Italy
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46
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Webster KA, Discher DJ, Bishopric NH. Regulation of fos and jun immediate-early genes by redox or metabolic stress in cardiac myocytes. Circ Res 1994; 74:679-86. [PMID: 8137504 DOI: 10.1161/01.res.74.4.679] [Citation(s) in RCA: 61] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/29/2023]
Abstract
We have previously demonstrated coordinate inductions of c-fos, c-jun, jun B, and jun D in cardiac myocytes exposed to hypoxia for 2 to 4 hours. Induction of these transcripts occurred before any significant loss of intracellular ATP. In the present study, the origin of the signal(s) that regulates immediate-early gene induction was investigated by comparing the effects of hypoxia with those of the metabolic inhibitors cyanide, deoxyglucose and cyanide combined, and iodoacetic acid. Cyanide, an inhibitor of oxidative metabolism, closely mimicked the metabolic effects of hypoxia, with elimination of oxygen consumption, increased lactate production, and minimal decline in ATP levels under both conditions. Compared with hypoxia, cyanide mediated small transient inductions of fos and jun transcripts that followed a different time course. The combination of cyanide and deoxyglucose resulted in inhibition of lactate production as well as respiration, and ATP dropped rapidly to 20% of control levels. The loss of intracellular ATP was followed by fourfold inductions of c-fos and c-jun with minor changes in jun B and jun D transcript levels. Similarly, iodoacetic acid caused a major (90%) loss of ATP and irreversible cell damage as measured by leakage of creatine phosphokinase enzyme and loss of membrane arachidonic acid; ATP loss was followed by fivefold to sevenfold inductions of c-fos, c-jun and jun B transcripts.(ABSTRACT TRUNCATED AT 250 WORDS)
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Affiliation(s)
- K A Webster
- Department of Cell and Molecular Biology, SRI International, Menlo Park, CA 94025
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47
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Cairo G, Pietrangelo A. Transferrin receptor gene expression during rat liver regeneration. Evidence for post-transcriptional regulation by iron regulatory factorB, a second iron-responsive element-binding protein. J Biol Chem 1994. [DOI: 10.1016/s0021-9258(17)37386-6] [Citation(s) in RCA: 37] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/22/2022] Open
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48
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Ferrero M, Desiderio MA, Martinotti A, Melani C, Bernelli-Zazzera A, Colombo MP, Cairo G. Expression of a growth arrest specific gene (gas-6) during liver regeneration: molecular mechanisms and signalling pathways. J Cell Physiol 1994; 158:263-9. [PMID: 8106563 DOI: 10.1002/jcp.1041580208] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/28/2023]
Abstract
A set of growth arrest-specific (gas) genes negatively regulated by serum has been identified. To define the role of gas genes in a model of cell proliferation in vivo we analyzed the expression of one of these genes (gas-6) during liver regeneration after partial hepatectomy (PH). We found that gas-6 mRNA was down-regulated 4 hours after PH, within the G0 to G1 transition. Later on, gas-6 mRNA increased over the level found in normal liver with a peak at 16 hours, before the onset of DNA synthesis. This surge was probably triggered by an inflammatory response caused by the surgical trauma, because an increase of similar extent occurring with the same time course was present in livers of sham-operated and turpentine-treated rats. Comparison of mRNA steady state levels with nuclear transcription rates indicated that gas-6 expression is post-transcriptionally regulated. As we found that down-regulation of gas-6 expression was prevented by treatment with Actinomycin D, a labile protein might be involved in the determination of gas-6 mRNA stability. To investigate the mitogenic signals controlling gas-6 expression during liver regeneration we treated hepatectomized rats with a specific alpha-1-adrenoceptor blocker (prazosin) as well as with drugs which modify intracellular calcium levels. The decrease of gas-6 mRNA 4 hours after PH was prevented by prazosin and by neomycin, an inhibitor of calcium release from endogenous stores. These findings suggest that down-regulation of gas-6 expression during hepatic regeneration is triggered by catecholamines interaction with alpha-1-adrenergic receptors and by subsequent calcium release. In addition we found that the rise of gas-6 gene expression occurring at 16 hours after PH was not affected by prazosin but was inhibited by trifluoperazine. Therefore, we suggest that up-regulation of gas-6 gene expression is mediated by the interaction of calcium with calmodulin, independently of catecholamines.
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Affiliation(s)
- M Ferrero
- Centro di Studio sulla Patologia Cellulare C.N.R. Università di Milano, Italy
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Schoeniger LO, Andreoni KA, Ott GR, Risby TH, Bulkley GB, Udelsman R, Burdick JF, Buchman TG. Induction of heat-shock gene expression in postischemic pig liver depends on superoxide generation. Gastroenterology 1994; 106:177-84. [PMID: 8276180 DOI: 10.1016/s0016-5085(94)95209-4] [Citation(s) in RCA: 59] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/29/2023]
Abstract
BACKGROUND/AIMS Both hemorrhagic and cardiogenic shock are associated with hepatic shock gene expression at resuscitation. This study investigated the potential role of intravascular superoxide anion as a proximal trigger of heat shock protein gene expression. METHODS Preanesthetized pigs were subjected to 120 m of total warm hepatic ischemia. The survival model consisted of warm, total hepatic ischemia and reperfusion (with active portal-systemic bypass) followed by reperfusion and survival for 3 days. Serial hepatic biopsy samples were evaluated for the expression of heat shock protein 72 (HSP-72) messenger RNA (mRNA) by Northern and Western analysis and by in situ RNA hybridization. The possible role of intravascular O2- as a mediator of heat shock response was evaluated by its specific inhibition by the intravenous infusion of recombinant human superoxide dismutase (SOD). RESULTS Ischemia for 120 minutes followed by 60 minutes of reperfusion caused accumulation of HSP-72 mRNA. Transcripts were localized to hepatocytes. HSP-72 mRNA was detected neither following ischemia alone nor when SOD was infused for 15 minutes at reperfusion. Three days later, transcripts were not detectable, but HSP-72 protein accumulated irrespective of SOD administration. CONCLUSIONS Warm hepatic ischemia induces the hepatocyte expression of HSP-72 at reperfusion by a mechanism that is dependent upon the superoxide anion, probably generated intravascularly. However, the transient dismutation of superoxide is insufficient to suppress subsequent accumulation of HSP-72.
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Affiliation(s)
- L O Schoeniger
- Department of Surgery, Johns Hopkins Medical Institutions, Baltimore, Maryland
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Wang S, Longo FM, Chen J, Butman M, Graham SH, Haglid KG, Sharp FR. Induction of glucose regulated protein (grp78) and inducible heat shock protein (hsp70) mRNAs in rat brain after kainic acid seizures and focal ischemia. Neurochem Int 1993; 23:575-82. [PMID: 8281126 DOI: 10.1016/0197-0186(93)90106-f] [Citation(s) in RCA: 59] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/29/2023]
Abstract
Specific probes were obtained using PCR cloning from rat brain for the 78 kDa glucose regulated (grp78), inducible 72 kDa (hsp70) as well as constitutive 73 kDa (hsc73) heat shock mRNAs. Grp78 and hsc73 were expressed in normal rat brain whereas hsp70 was not. Subcutaneous injection kainic acid (10 mg/kg) produced seizures and induced all three mRNAs. The induction of grp78 and hsp70 mRNAs occurred within 2 h, peaked between 6-8 h, persisted for 48 h, and returned to control levels by 72 h. Expression of the grp78 and hsp70 mRNAs after focal ischemia progressively increased with occlusion durations from 15-120 min in the cerebral cortex. Though grp78 and hsp70 mRNAs were induced modestly in the striatum by 15 min of ischemia, longer durations of ischemia were characterized by little change in the grp78 mRNA levels and relatively lower levels of hsp70 expression. This result indicates that progressive increases in the duration of ischemia in brain, prior to infarction, may produce proportional increases in transcription of the heat shock genes. However, once the duration of ischemia is long enough to produce infarction, this severely limits the availability of ATP which blocks transcription of the heat shock genes. In conclusion, concurrent induction of the heat shock genes suggests that kainic acid seizures and focal ischemia induce several different stress responses in brain cells caused by denaturation of proteins, changes of protein synthesis, and changes of protein glycosylation.
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Affiliation(s)
- S Wang
- Department of Neurology (V127), University of California, San Francisco
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