Дисертації з теми "Secretion stress"
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1
Smart, Darren. "Stressors & LH secretion in the ewe." Thesis, University of Liverpool, 1992. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.333607.
2
Parker, Victoria Joanne. "Hypothalamic mechanisms mediating inhibition of prolactin secretion following stress in early pregnant mice." Thesis, University of Edinburgh, 2012. http://hdl.handle.net/1842/6485.
Анотація:
In early pregnancy prolonged exposure to stress is known to have profound adverse effects on reproduction and is associated with suppressed progesterone secretion and consequent disturbance of the pregnancy-protective cytokine milieu, thus threatening early pregnancy maintenance. Maternal neuroendocrine responses to stress in early pregnancy are poorly understood. Therefore, we designed experiments to (1) study the hypothalamo-pituitary-adrenal (HPA) axis responses to stress in early pregnant mice, to discover whether and how responses change and (2) to determine the effect of stress in early gestation on pregnancy hormones, with a particular focus on the secretion and regulation of prolactin. To establish the effects of stress in early pregnancy (day 5.5) two different ethologically relevant stressors were used: lipopolysaccharide (LPS) or 24h fast stress, to mimic situations that may potentially arise during pregnancy in women: infection or hunger. HPA axis secretory responses to immune stress in early-mid pregnancy were robust and comparable to that in virgins. Vasopressin rather than the usual CRH neurone responses play a key role in maintaining this. However, the mode of action of glucocorticoids in mediating pregnancy complications is not yet established. Prolactin, and its hypothalamic control mechanisms, is a key candidate to mediate brain-to-body responses to stress. Prolactin has important roles in progesterone secretion, pregnancy establishment and immune regulation. We hypothesised that stress would negatively affect prolactin and its neuroendocrine control systems. Prolactin is mainly under the inhibitory control of dopamine, released predominantly from the tuberoinfundibular dopamine (TIDA) neurones. Prolactin also negatively feeds back on itself via prolactin receptors on the TIDA neurones and janus kinase (JAK)2/signal transducer and activator of transcription (STAT)5 signalling. Both immune and fasting stressors strongly inhibited basal prolactin secretion in early pregnancy, accompanied by a mild increase in activation of TIDA as shown by elevated Fos expression, compared to virgins. In addition, pregnancy attenuated LPS-induced recruitment of parvocellular paraventricular nucleus neurones and increased activation of brainstem noradrenergic nuclei which could potentially contribute to altered control of the dopamine-prolactin system. Following either immune or fast stress in early pregnancy ovine prolactin was able to drive enhanced expression of phosphorylated (p)STAT5. However, stress alone did not alter pSTAT5 implying it is not exclusively responsible for the stress-reduced prolactin observed in early pregnancy and another stress-induced stimulus must be activating TIDA neurones in these mice. LPS did not alter dopamine activity the median eminence (DOPAC: dopamine ratio) suggesting dopamine does not underlie stress-reduced prolactin secretion and other mechanisms must be considered. Direct effects of LPS, or its associated cytokines, on pituitary lactotrophs to inhibit prolactin secretion is a possible candidate. To investigate the effect of proinflammatory cytokines on the prolactin system in early pregnancy, d5.5 mice were administered TNF-alpha (a) or interleukin (IL)-6. Both cytokines increased TIDA activation, however, only TNF-a decreased plasma prolactin and progesterone, suggesting additional TNF-alpha action at the pituitary. As prolactin is anxiolytic we further proposed that stress would have a more profound effect on elevated plus maze performance in pregnant mice. However, early pregnant mice were generally more anxious vs. virgins regardless of LPS treatment. Taken together data show that stress in early pregnancy reduces prolactin and progesterone secretion, contributing to pregnancy complications/failure, but the neuroendocrine stress-related mechanism behind this suppression is yet to be determined.
3
Kim, Beob Gyun. "INFLUENCES OF CHROMIUM (III) PICOLINATE ON PIGS UNDER THERMAL, IMMUNE OR DIETARY STRESS, AND ON ADRENAL STEROID SECRETION." UKnowledge, 2007. http://uknowledge.uky.edu/gradschool_diss/560.
Анотація:
The objectives were to investigate the effects of chromium (III) picolinate (CrPic; up to 2,000 ppb of Cr) on growing pigs subjected to a variety of stressors including thermal, immune, or dietary stress and to examine the effects of CrPic on steroidogenesis from adrenocortical cells. In the thermal stress study, high ambient temperature caused reduced weight gain and feed consumption (P andlt; 0.01), and low ambient temperature caused increased feed intake and feed:gain (P andlt; 0.01). However, these effects were not moderated by CrPic, and respiratory rate, plasma cortisol, or plasma glucose were unaffected by CrPic. In the immune stress study, pigs challenged with lipopolysaccharide (LPS) lost 951 g during 12 hours post injection, while the phosphate buffer saline (PBS) injected group gained 170 g (P andlt; 0.001). The LPS group showed higher rectal temperature (P andlt; 0.05), higher respiratory rate (P andlt; 0.05), greater plasma cortisol (P andlt; 0.001), and lower plasma glucose (P andlt; 0.05) than the PBS group. These effects were not ameliorated by CrPic. In the dietary stress study, pigs fed the high-fat diet (HFD) gained weight faster (P andlt; 0.05), consumed less feed (P andlt; 0.001), and had lower feed:gain (P andlt; 0.001). Plasma insulin concentration on d 14 decreased with CrPic (P andlt; 0.05) in a linear manner (P = 0.05). Consumption of the HFD resulted in increases of slaughter weight, perirenal fat, and back fat measurements (P andlt; 0.01). The CrPic resulted in linear reductions of carcass weight, last rib fat, last lumbar fat and average backfat (P andlt; 0.10). The effects of CrPic on carcass fat measurements were more significant in barrows than gilts. In the adrenocortical cell study, forskolin stimulated cortisol and DHEAs secretion from H295R cells. CrPic inhibited aspects of steroidogenesis in agonist-stimulated adrenocortical cells. Overall, dietary CrPic was unable to moderate the stress related effects due to high ambient temperature, low ambient temperature, or an endotoxin challenge. However, CrPic attenuated effects of HFD, mainly on body fat accretion of pigs, especially in barrows, and CrPic inhibited steroidogenesis in stimulated adrenocorticoid cells.
4
Kosti, Ourania. "Intra-adrenal mechanisms in the control of steroid secretion and the response to stress." Thesis, Queen Mary, University of London, 2004. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.411344.
5
Weber, Barbara. "Stress response and virulence in Vibrio anguillarum." Doctoral thesis, Umeå : Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet), Umeå universitet, 2010. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-33269.
6
Marin, Marie-France. "Immediate and delayed effects of stress on a reactivitated declarative long-term memory trace." Thesis, McGill University, 2009. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=116034.
Анотація:
In 1968, a study demonstrated that consolidated memories can be affected again if they are reactivated. Given the importance of the stress hormones glucocorticoids (GCs) on memory consolidation, the goal of the current study was to assess whether GCs had the capacity to affect a reactivated long-term memory and whether neutral and emotional memories were affected differently. At the first session, participants encoded a movie containing neutral and emotional scenes. Two days later, they recalled the story. Half of them were then exposed to a psychosocial stressor. Memory performance was assessed again right after the stressor and five days later. The stressed group recalled less neutral material five days after the stressor compared to controls. Immediately after the stressor, the stressed group recalled more emotional material than controls. Moreover, this enhanced memory trace was maintained across time. This highlights the importance of minimizing exposure to stressful contexts when reactivating emotional memories.
7
Pilgrim, Kamala. "Mechanisms underlying cortisol reactivity to stress in low and high socioeconomic status individuals : role of naturally-occurring attentional biases." Thesis, McGill University, 2008. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=116040.
Анотація:
This Master's dissertation explored whether a rapid orienting of attention toward or away from social stress information during a restful state, relates to the magnitude of glucocorticoids (GC) released in response to a stressor, the Trier Social Stress Test (TSST). It also assessed whether childhood rearing in a low socioeconomic status (SES) context mediates this relationship. Subjects rested for 45 minutes during which time they completed a modified version of Posner's attention paradigm, comprising social stress words. Immediately following, participants were exposed to the stressor. Results indicated that a rapid attentional engagement toward social stress words associated with pronounced GC responses to the TSST. Fast engagers displayed lower self-esteem and did not differ in terms of their past SES. These findings demonstrate that attentional biases for social stress information at rest combine with diminished self-esteem to predict the magnitude of GC released during psychological stress irrespective of early SES conditions.
8
Valiquette, Luc François. "Association between self-reported childhood maltreatment and cortisol profiles in psychotic patients." Thesis, McGill University, 2008. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=112314.
Анотація:
Childhood maltreatment is extremely common in patients diagnosed with psychotic disorders. Moreover, it has been linked with impaired functioning of the Hypothalamic-Pituitary-Adrenal axis. Furthermore, abnormality of the HPA has been found in psychotic patients. Presence of childhood maltreatment could then explain why the HPA axis is dysfunctional in these subjects. Our objective was to clarify the role of childhood trauma in the cortisol profiles of psychotic patients. Thirty-one patients underwent assessments of childhood maltreatment. Diurnal cortisol and cortisol after a controlled psychosocial stress were also collected. Our results show that childhood trauma is associated with lower cortisol levels during the morning and during 24 hours. In men diagnosed with psychosis, childhood trauma is also associated with a higher cortisol response during psychosocial stress. This suggests an alteration of the HPA axis in psychotic patients, resulting from early trauma. Moreover, our results suggest that looking at specific types of childhood abuse may also be important.
9
Alharake, Jawad. "Study of genetic factors involved in enzyme secretion in hyperproductive strains of Trichoderma reesei." Electronic Thesis or Diss., université Paris-Saclay, 2023. http://www.theses.fr/2023UPASB061.
Анотація:
Les combustibles fossiles sont un contributeur majeur au réchauffement climatique, et leur nature non renouvelable est un obstacle à la construction de sociétés durables. Dans ce contexte, les biocarburants de seconde génération se présentent comme une alternative attractive et plus respectueuse de l'environnement. Le processus de production de biocarburants de deuxième génération comprend plusieurs étapes incluant le prétraitement physico-chimique de la biomasse lignocellulosique (non comestible), l'hydrolyse enzymatique de la cellulose en glucose et par la fermentation de sucres simples en biocarburants, comme le bioéthanol. Un des problèmes principaux pour une large application est cependant le coût relativement élevé des enzymes hydrolytiques, les cellulases, utilisées pour déconstruire la biomasse lignocellulosique prétraitée en sucres fermentescibles.Le champignon filamenteux Trichoderma reesei est le choix privilégié pour la production industrielle de cellulases car il possède des capacités d'hyperproduction et d'hypersécrétion. Les souches industrielles de T. reesei peuvent sécréter jusqu'à 100 g/L de cellulases dans des fermenteurs industriels contrôlés. En particulier, la souche mutante Rut-C30 est une souche hyperproductrice de référence, mais notre compréhension incomplète de son système de sécrétion performant complique l'amélioration de sa capacité d'hypersécrétion par génie génétique. C'est pourquoi, ce travail visait à éclaircir les voies de régulation contrôlant la sécrétion et les réponses au stress de sécrétion pour identifier des goulots d'étranglement et pour développer de nouvelles souches dotées d'une capacité de sécrétion supérieure dans le futur.Dans ce but, des données transcriptomiques étaient générées à partir de cultures de T. reesei Rut-C30 dans différentes conditions de stress de sécrétion et ont permis d'identifier de composants potentiels de régulation de la sécrétion qui pouvaient être ciblés pour invalidation. Pour compléter cette approche, un datamining de données transcriptomiques obtenues avec d'autres champignons filamenteux cultivés dans des conditions de stress de sécrétion a révélé d'autres gènes cibles potentiellement impliqués dans la régulation de la voie de sécrétion. Finalement, neuf gènes ont été invalidés dans la souche Rut-C30 et les mutants obtenus ont été caractérisés phénotypiquement. Tous ont montré une croissance ralentie et un comportement de sécrétion altéré. Un séquençage de l'ARN a été réalisé sur les mutants ∆res2, ∆rpn4 and ∆snd1 et comparé à celui de Rut-C30 dans les mêmes conditions de culture. Aucun des trois facteurs de transcription n'impacte la transcription des gènes impliqués dans la sécrétion ou dans la réponse au stress de sécrétion dans nos conditions. En revanche, des gènes codant pour des enzymes du métabolisme des lipides sont différentiellement exprimés dans les trois mutants ce qui pourrait affecter la sécrétion indirectement. Les résultats délivrent des premiers indices pour atténuer les goulots d'étranglement de la sécrétion dans T. reesei Rut-C30 et ouvrent la voie vers le développement de souches possédant une capacité de sécrétion amélioréeFossil fuels are a major contributor to global warming, and their non-renewable nature is an impediment for building sustainable societies. In this context, second generation biofuels represent an attractive and more environmentally friendly alternative. The process of second-generation biofuel production consists of several steps including a physicochemical pretreatment of lignocellulosic (non-edible) biomass, the enzymatic hydrolysis of cellulose into glucose and the fermentation of simple sugars into biofuels, such as bioethanol. One of the main bottlenecks for a large implementation of this process is, however, the relatively high cost of hydrolytic enzymes, namely cellulases, used to deconstruct the pretreated lignocellulosic biomass into fermentable sugars.The filamentous fungus Trichoderma reesei is the preferred choice for industrial production of cellulases since it has hyperproduction and hypersecretion capacities. Industrial strains of T. reesei can secrete up to 100 g/L of cellulases in controlled industrial fermenters. In particular, the mutant strain Rut- C30 is a reference hyperproducer strain, but our incomplete understanding of its enhanced secretion system complicates further improvement of its hypersecretion capacity by genetic engineering. Therefore, this work aimed at unravelling the regulatory pathways controlling secretion and the secretion stress response in order to identify bottlenecks and to develop new strains with enhanced secretion capacity in the future.To this end, transcriptomic data were generated from cultivations of T. reesei Rut-C30 in different secretion stress conditions which allowed to identify potential components of secretion regulation that were targeted for deletion. As a complementary approach, mining of transcriptomic data obtained with other filamentous fungi in secretion stress conditions revealed further target genes potentially involved in the regulation of the secretion pathway. Finally, nine genes were deleted in the Rut-C30 strain, and the resulting strains phenotypically characterized. All of them displayed reduced growth and showed altered protein secretion behavior. RNA sequencing was performed on ∆res2, ∆rpn4 and ∆snd1 mutant strains and compared to that of Rut-C30 in the same culture conditions. Neither of the three transcription factors impacts transcription of genes involved in secretion or the secretion stress response in our conditions. However, in all three mutants, genes encoding enzymes of lipid metabolism are differentially expressed which could affect secretion in an indirect way. The results represent first clues to alleviate bottlenecks in secretion in T. reesei Rut-C30 and pave the way to develop strains with still improved secretion capacity
10
Solà, Tapias Núria. "The involvement of the three main inflammatory bowel disease pathways and the secretion of trypsin proteolytic activity on intestinal epithelial cells." Thesis, Toulouse 3, 2018. http://www.theses.fr/2018TOU30049/document.
Анотація:
Les maladies inflammatoires chroniques de l'intestin (MICI) se caractérisent par une inflammation sévère de l'intestin grêle et du côlon et comprennent la maladie de Crohn (MC) et la rectocolite hémorragique (RCH). Les MICI sont des maladies complexes faisant intervenir des facteurs génétiques : certains senseurs bactériens, l'autophagie et le stress du réticulum endoplasmique. Un défaut de barrière de l'épithélium digestif est également fortement impliqué dans la physiopathologie du processus inflammatoire. La fonction barrière de l'épithélium digestif est assurée par plusieurs types cellulaires, synthétisant entre autres, des peptides antimicrobiens (PAM) et des mucines. Dans les MICI, une augmentation de la perméabilité intestinale et une perte de muco-sécrétion ont été décrites. Les protéases jouent un rôle fondamental dans la digestion du bol alimentaire mais également dans le maintien de l'homéostasie intestinale en activant ou dégradant divers motifs moléculaires, ou in induisant des signaux spécifiques aux cellules par l'activation de quatre récepteurs : les PARs (Protease-Activated Receptor). Dans les MICI, un excès d'activité protéolytique de type trypsine est observé. L'origine de cette activité est théoriquement attribuée aux cellules immunitaires, à une surproduction pancréatique ou au microbiote, mais les cellules épithéliales intestinales semblent également être une source majeure de protéases. L'objectif de mon projet de thèse visait à étudier l'impact des principales voies impliquées dans les MICI sur l'homéostasie des protéases épithéliales et le rôle de celles-ci dans la déstabilisation de la fonction de barrière. Nos résultats ont confirmé un excès de protéases à sérine dans les cellules épithéliales de patients atteint de MC ou de RCH. In vitro, sur des monocouches de cellules Caco-2, l'induction de l'autophagie diminuait la libération apicale de protéase de type trypsine, alors que le senseur bactériens NOD2 n'avait aucun effet. A l'inverse, une stimulation du Stress du réticulum endoplasmique (SRE) par la Thapsigargin, induisait une libération accrue de protéases actives de type trypsine au pôle apical des cellules. [...]Crohn's disease (CD) and Ulcerative colitis (UC) are two forms of Inflammatory Bowel Disease (IBD), a chronic inflammatory pathology affecting the digestive tract. Patients suffer from relapsing flares, diarrhea, abdominal pain and bleeding. Although the molecular mechanisms of IBD are poorly understood, recent data suggest that IBD occurs in genetically predisposed individuals developing an abnormal immune response to intestinal microbes after, being exposed to specific environmental triggers. Genetic studies have reported more than 170 polymorphisms susceptible to be involved in IBD pathogenesis. The strongest associations have highlighted three main pathways altered in IBD including bacterial sensing (NOD2, CD), autophagy (ATG16L1 and IRGM, CD) and endoplasmic reticulum stress (ER-Stress) (XBP1, UC). The role of intestinal barrier function is also strongly implicated in IBD pathogenesis, and is modulated by factors present in the lumen derived from microbiota, food or at a molecular level, by factors such as proteases. In IBD pathophysiology, the inflammatory process is characterized by impaired intestinal biology including disruption of tight junctions and leaky gut, decreased amount of Paneth and Goblet cells, and translocation of luminal antigens triggering inflammation. Previous studies have demonstrated an increased level of active serine proteases in the stools and tissues of IBD patients, supposing that proteases originate from infiltrated immune cells, pancreatic secretion or microbiota. However, our team has reported that intestinal epithelial cells are a major source of serine proteases, in particular trypsin-like enzymes, are released by a stressed epithelium in pathogenic context such as irritable bowel syndrome. In this project, we aimed at better understanding whether the three main pathways involved in IBD (Nod2, autophagy, ER-stress) could be linked to an epithelial release of trypsin and reciprocally, if epithelial trypsin is able to induce or modulate these three IBD pathways. We confirmed that trypsin-like activity was significantly higher in biopsies from UC and CD patients compared to healthy controls. In Caco-2 monolayers cultured in transwells, secreted trypsin-like proteolytic activity remained stable upon NOD2 stimulation but decreased under autophagy induction. Thapsigargin (Tg) stimulation a well-known ER-stress inducer, enhanced the apical release of trypsin-like activity in Caco2 cells. [...]
11
Hickman, Cristina Fontes Lindemann. "Environmental factors affecting interferon-τ expression and secretion by in vitro produced bovine blastocysts". Thesis, University of Edinburgh, 2010. http://hdl.handle.net/1842/4399.
Анотація:
Interferon (IFN)τ is the luteotrophic signal in ruminants and is secreted by bovine blastocysts both in vivo and in vitro. IFNτ secretion is highly variable and its control is only partly understood. Most studies on the effects of environmental factors on IFNτ production have evaluated IFNτ production during the time of embryo elongation and attachment. There is less knowledge of how IFNτ production at the blastocyst stage is modulated. Therefore, the hypothesis of this thesis was that the amounts of IFNτ expressed and/or secreted by bovine blastocysts produced in vitro were modulated by environmental factors. In the first set of experiments, bovine embryos were incubated with a cytokine (granulocyte macrophage colony stimulating factor, GM-CSF). GM-CSF had been shown previously to promote embryo viability in a range of species and to modulate IFNτ secretion by ovine blastocysts and thus was classified as a beneficial environmental factor. Three experiments were conducted to test whether GM-CSF stimulated bovine blastocyst development and IFNτ secretion. Embryos were incubated with a range of different concentrations of GM-CSF (2, 5, 10 and 50 ng mL-1) and at different stages of development (1 to 3 and 1 to 9 days post-insemination). Bovine embryos were unresponsive to GM-CSF in terms of IFNτ secretion, pyruvate oxidation, rate of development, blastocyst yield, morphological quality and apoptotic index, irrespective of timing of exposure and/or concentration of GM-CSF. In the second part of the thesis, bovine blastocysts were exposed to a mild heat treatment (42°C for four h) to determine whether heat stress affected IFNτ expression by bovine blastocysts. A novel multiplex reverse-transcription polymerase chain reaction methodology was validated to detect IFNτ and heat shock protein (HSP)70 mRNA in individual bovine embryos relative to an endogenous gene (YWHAZ) and an exogenous mRNA (α-globin) and results were expressed both in absolute terms and in relation to the endogenous control. Heat treatment upregulated IFNτ mRNA expression, suggesting that detrimental environmental factors may influence IFNτ expression. Heat treatment also caused an increase in HSP70 mRNA expression but did not affect blastocyst morphology, suggesting that the level of stress caused by the heat treatment was great enough to activate the cellular stress response, but mild enough not to cause a change in morphology. In addition, the positive correlation between HSP70 and IFNτ transcript levels and the higher IFNτ expression by embryos which showed signs of degeneration and collapse compared to those which progressed in development suggested that IFNτ expression may be indicative of stress. The relationship between IFNτ expression and secretion in vitro with morphology, pyruvate metabolism, apoptotic index and cell number was inconsistent, suggesting that IFNτ production did not correlate with ‘quality’ (defined as an index of viability). Blastocyst yield, day of blastulation and change in morphology index did account for at least part of the variation in IFNτ production, suggesting that some intrinsic factors may regulate IFNτ secretion. These intrinsic factors, however, did not explain all the variation in IFNτ secretion between blastocysts. Therefore, the amount of IFNτ secreted by bovine blastocysts is modulated by both intrinsic and environmental factors. A model was proposed where different levels of stress affect survivability to different extents, and the ability to respond to mild levels of stress may be indicative of improved survivability.
12
Zimmermann, Ulrich, Konstanze Spring, Hans-Ulrich Wittchen, Hubertus Himmerich, R. Landgraf, Manfred Uhr, and Florian Holsboer. "Arginine vasopressin and adrenocorticotropin secretion in response to psychosocial stress is attenuated by ethanol in sons of alcohol-dependent fathers." Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2013. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-110031.
Анотація:
Familial risk and environmental stress promote the development of alcohol dependence. We investigated whether a positive family history of alcoholism affects the neuroendocrine response to a standardized laboratory stress test in healthy subjects without alcohol use disorders. Twenty-four high-risk subjects with a paternal history of alcoholism (PHA) and 16 family history negative (FHN) controls were evaluated. Psychosocial stress was induced by having subjects deliver a 5-min speech and mental arithmetics in front of an audience on separate days, after drinking either placebo or ethanol (0.6 g/kg) in a randomized sequence. Adrenocorticotropin (ACTH) was measured in 10 plasma samples covering up to 75 min after the stress test. Plasma arginine vasopressin (AVP) was determined before the stressor, at the time of maximum ACTH secretion, and at 75 min after stress onset. The stress test induced a phasic increase in ACTH secretion. At the time of maximum ACTH, AVP was significantly increased in relation to baseline. Compared to placebo, alcohol administration significantly attenuated maximum ACTH concentration in PHA but not FHN subjects, and decreased AVP measured in the same samples in PHA but not FHN subjects. We conclude that activation of the hypothalamic–pituitary–adrenal system by psychosocial stress is accompanied by an increase in peripheral plasma AVP levels. Secretion of both ACTH and AVP suggest that alcohol attenuates the stress response selectively in PHA but not FHN subjects. This might imply some short-term positive alcohol effect in sons of alcoholics, but also constitute a mechanism by which their risk to develop alcohol use disorders is increased.
13
Gallo, Marco. "misc-1/OGC is a new stress response gene and regulator of apoptosis, germline stem cell proliferation and insulin secretion." Thesis, University of British Columbia, 2010. http://hdl.handle.net/2429/27092.
Анотація:
The present work produced new insights on the function of an ascaroside molecule and a gene that affect formation of dauer larvae, a diapause stage in Caenorhabditis elegans.
Our results indicate that the ascaroside daumone, although a component of the dauer pheromone, does not act through signalling pathways active in the cilia. Furthermore, daumone has toxic effects if the animals are exposed to dauer-inducing concentrations of the compound. This ascaroside is not able, by itself, to recapitulate the full spectrum of events that occur during dauer formation.
We identified misc-1 (MItochondrial Solute Carrier) as a novel suppressor of the dauer phenotype of daf-2/IGF1R mutants. We provide evidence that MISC-1 is the putative orthologue of mammalian OGC (2-OxoGlutarate Carrier), a metabolic carrier of the inner mitochondrial membrane. We show that a misc-1 null allele suppresses the dauer phenotype of daf-2 mutants by increasing insulin secretion. Consistent with this result, misc-1 mutants have increased proliferation of germline stem cells. Furthermore, we show that MISC-1 and OGC are involved in a phylogenetically conserved apoptosis pathway. Reduced levels of MISC-1 in C. elegans and OGC in human cells result in mitochondrial fragmentation. MISC-1 and human OGC physically interact with the anti-apoptotic Bcl-2-family members CED-9 and Bcl-xL, respectively, and are novel components of the mitochondrial permeability transition pore. Decreased levels of MISC-1 and OGC induce apoptosis in C. elegans and mouse cells. Finally, our experiments confirm that MISC-1/OGC has Reactive Oxygen Species (ROS)-detoxifying functions in vivo. We use misc-1 mutants to show that increased levels of ROS do not have negative effects on life span, contrary to current theories of aging. Using misc-1 mutants as a model, we show that extended life span of some C. elegans mitochondrial mutants is dependent on upregulation of extra-mitochondrial pathways of energy production. In conclusion, we show that the metabolic protein MISC-1/OGC affects insulin secretion, mitochondrial dynamics, apoptosis and ROS detoxification. We propose that MISC-1/OGC integrates metabolic and cell survival decisions by its physical interaction with the apoptotic machinery.
14
Zimmermann, Ulrich, Konstanze Spring, Hans-Ulrich Wittchen, Hubertus Himmerich, R. Landgraf, Manfred Uhr, and Florian Holsboer. "Arginine vasopressin and adrenocorticotropin secretion in response to psychosocial stress is attenuated by ethanol in sons of alcohol-dependent fathers." Technische Universität Dresden, 2004. https://tud.qucosa.de/id/qucosa%3A26808.
Анотація:
Familial risk and environmental stress promote the development of alcohol dependence. We investigated whether a positive family history of alcoholism affects the neuroendocrine response to a standardized laboratory stress test in healthy subjects without alcohol use disorders. Twenty-four high-risk subjects with a paternal history of alcoholism (PHA) and 16 family history negative (FHN) controls were evaluated. Psychosocial stress was induced by having subjects deliver a 5-min speech and mental arithmetics in front of an audience on separate days, after drinking either placebo or ethanol (0.6 g/kg) in a randomized sequence. Adrenocorticotropin (ACTH) was measured in 10 plasma samples covering up to 75 min after the stress test. Plasma arginine vasopressin (AVP) was determined before the stressor, at the time of maximum ACTH secretion, and at 75 min after stress onset. The stress test induced a phasic increase in ACTH secretion. At the time of maximum ACTH, AVP was significantly increased in relation to baseline. Compared to placebo, alcohol administration significantly attenuated maximum ACTH concentration in PHA but not FHN subjects, and decreased AVP measured in the same samples in PHA but not FHN subjects. We conclude that activation of the hypothalamic–pituitary–adrenal system by psychosocial stress is accompanied by an increase in peripheral plasma AVP levels. Secretion of both ACTH and AVP suggest that alcohol attenuates the stress response selectively in PHA but not FHN subjects. This might imply some short-term positive alcohol effect in sons of alcoholics, but also constitute a mechanism by which their risk to develop alcohol use disorders is increased.
15
Hammer, Jared Louis. "Changes In Threonyl-Trna Synthetase Expression And Secretion In Response To Endoplasmic Reticulum Stress By Monensin In Ovarian Cancer Cells." ScholarWorks @ UVM, 2017. http://scholarworks.uvm.edu/graddis/764.
Анотація:
Aminoacyl-tRNA synthetases (ARS) are a family of enzymes that catalyze the charging of amino acids to their cognate tRNA in an aminoacylation reaction. Many members of this family have been found to have secondary functions independent of their primary aminoacylation function. Threonyl-tRNA synthetase (TARS), the ARS responsible for charging tRNA with threonine, is secreted from endothelial cells in response to both vascular endothelial growth factor (VEGF) and tumor necrosis factor-α (TNF-α), and stimulates angiogenesis and cell migration. Here we show a novel experimental approach for studying TARS secretion, and for observing the role of intracellular TARS in the endoplasmic reticulum (ER) stress response and in angiogenesis.
Using Western blotting, immunofluorescence microscopy and RT-qPCR we were able to investigate changes in TARS protein and transcript levels. We initially hypothesized that TARS was secreted by exosomal release, and so we treated a human ovarian cancer cell line (CaOV-3) with monensin, an ionophore that increases exosome production, and VEGF to observe changes in intracellular and extracellular TARS protein. Monensin treatment consistently increased extracellular and intracellular TARS protein, however CD63, an exosome marker protein, levels were unaffected by monensin treatment. VEGF had no effect on intracellular TARS. We therefore hypothesized that the TARS response was a result of ER stress.
The unfolded protein response (UPR) is a series of signaling pathways that are activated upon ER stress. When CaOV-3 cells were treated with increasing concentrations of monensin, intracellular levels of TARS and p-eIF2α, a downstream UPR target, increased accordingly. Monensin increased intracellular TARS protein and transcript levels in CaOV-3 cells. Monensin also increased DNAJB9, an ER chaperone protein, transcript levels, further confirming ER stress. Interestingly, monensin increased VEGF transcript levels about 6-fold. Borrelidin, a natural TARS inhibitor, also increased VEGF transcript levels, and caused an increase in p-eIF2α protein.
Although the mechanism of TARS secretion remains unresolved, these data indicate that intracellular TARS expression increases in response to ER stress by monensin. Given TARS and VEGF transcript expression increased accordingly, it is possible that intracellular TARS may have pro-angiogenic function. Future directions may include investigating TARS interactions with translational control machinery.
16
Madec, Iltud. "Effets du sémiochimique MHUSA (Mother Hens' Uropygial Secretion Analogue) sur le stress des poulets de chair : approches zootechnique, physiologique et comportementale." Phd thesis, Toulouse, INPT, 2008. http://oatao.univ-toulouse.fr/7819/1/madec.pdf.
Анотація:
Constats et hypothèses L'Homme a détourné, et donc faussé, l'importance des relations mère/jeune. Les techniques de production actuelles sont telles que les poussins sont introduits dans des bâtiments d’élevage âgés d'un jour. Ils y sont élevés par lots homogènes jusqu'à leur abattage. A partir de la ponte, le contact avec la mère n'existe plus, ce qui peut modifier l'empreinte et le lien d'attachement. Certains comportements originels, issus de l’ancêtre du poulet actuel, sont encore fonctionnels, alors que d’autres sont déviants. Le stress, qui peut être caractérisé par différents indicateurs, a des conséquences mesurables sur les performances, la physiologie et le comportement du poulet. Ce dernier est capable de détecter et reconnaître des odeurs, notamment celle de son nid. Quant à la mère suitée, elle sécrète, par la glande uropygiale, un bouquet odorant caractéristique de son état. Partant de ces constats nous avons voulu tester les effets de la diffusion d’un analogue de cette sécrétion (MHUSA ou Mother Hens' Semiochemical Analogue) sur la réponse au stress chez le poulet domestique (Gallus gallus). Résultats zootechniques Les poids vifs finaux et intermédiaires sont plus élevés pour les poulets élevés sous MHUSA. La qualité du produit final est aussi améliorée : poids de carcasse supérieur avec une masse de filet plus importante sans être plus grasse et une couleur de viande plus uniforme sous MHUSA. L'indice de consommation n’est pas significativement influencé par MHUSA. Résultats physiologiques Les indicateurs de référence (ratio Hétérophiles/Lymphocytes ou corticostérone) montrent des niveaux de stress inférieurs sous MHUSA. Comme conséquence, on montre notamment que la production de certaines cytokines (et donc la réaction immunitaire) est influencée par une exposition à MHUSA (IFNγ et IL6). Résultats comportementaux Après une période d'adaptation, un poussin isolé se dirige dans une zone plus concentrée en MHUSA. Des poulets en croissance sous MHUSA s'adaptent mieux à leur environnement et montrent des réactions de peur moins prononcées. Conclusion L'ensemble de nos travaux montre que des poulets domestiques évoluant dans une atmosphère chargée en MHUSA, comparativement à des animaux non traités, ont des performances supérieures et une réponse au stress diminuée, à la fois au niveau physiologique et comportemental. Il semble que ces réactions ne nécessitent pas d'apprentissage de la part du poulet.
17
Gumpper, Kristyn Nicole. "Maintaining Cardiac and Gastric Physiology: TRIM Proteins as Central Factors in Regulation of Organ Homeostasis at the Cellular Level." The Ohio State University, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=osu1563287863754715.
18
Sá, Maria Joana Giraldes Pereira Côrte-Real Corrêa de. "Natural IgM secretion in health and disease : genetic control and role in type 1 diabetes." Doctoral thesis, Universidade Técnica de Lisboa. Faculdade de Medicina Veterinária, 2012. http://hdl.handle.net/10400.5/4232.
Анотація:
Tese de Doutoramento em Ciências Veterinárias, especialidade de Ciências Biológicas e BiomédicasGermline-encoded autoreactive natural antibodies (NAbs) of the IgM isotype are secreted and circulate as a result of basal immune system stimulation, and constitute an important first line of defense against microorganism invasion, bridging the innate and the adaptive immune responses. NAbs are mostly secreted by positively selected B1a cells, and have been claimed to have a protective role against autoimmunity. Nevertheless, NAbs binding to cell surface self-antigens could have implications in the initiation of autoimmunity. Article I focused on the genetic control of NAbs secretion in healthy mice. Importantly, interferon regulatory factor 4 (Irf4), a transcription factor required for plasma cell differentiation and antibody secretion, was identified as the most probable candidate for the control of homeostatic serum IgM levels in the mouse. Type 1 diabetes (T1D) is a complex autoimmune disease that develops spontaneously in humans and is pathogenically similar in the non-obese-diabetic (NOD) mouse model. B cells are necessary in the NOD diabetogenic process, and the presence of anti-pancreatic beta cell antibodies is the earliest manifestation of T1D. Article II revealed that NOD peritoneal cavity B1a cells are more prone to spontaneously secrete NAbs that recognize pancreatic beta cell autoantigens, which could promote T1D either by enhancing professional antigen presentation of islet antigens, by activating the complement cascade or by directly promoting beta cell damage and self-antigens release. The studies reported in article III have explored these possibilities and have proven that NAbs of NOD B1a cells origin could bind and directly induce oxidative stress on pancreatic beta cells. Moreover, these studies have shown that NOD B1a cells have a lower threshold for innate-like stimulation and have established a link between NOD B1a cells properties, NAbs specificities and impact of IgM binding on beta cells physiology. Finally, article IV provides evidence that early treatment with antibodies that evoke NOD B1a cells proliferation and differentiation into IgM secreting cells correlates with T1D precipitation. In conclusion, this thesis has shown that Irf4 is a critical player in the genetic network that controls IgM secretion in healthy individuals, and that in the NOD mouse model of T1D, a lower threshold for innate like stimulation of peritoneal cavity B1a cells contributes to a naturally increased state of B1a cells activation and autoreactive IgM secretion, determining the initiation and/or contributing to the fueling of beta cells autoimmunity.
RESUMO Secreção de IgM natural na saúde e na doença: controlo genético e papel na diabetes tipo 1 - Os autoanticorpos naturais (NAbs) da classe IgM existem no organismo na ausência de imunização e constituem uma primeira linha de defesa fundamental contra infecções. Os NAbs são secretados maioritariamente por células B1a e a sua ligação a autoantigénios na superfície celular pode ter implicações para a iniciação de autoimunidade. O trabalho descrito no artigo I focou-se na compreensão do controlo genético da secreção de NAbs em murganhos saudáveis. Este estudo identificou o interferon regulatory factor 4 (Irf4), um factor de transcrição necessário para a diferenciação de plasmócitos e secreção de anticorpos, como o candidato mais provável para o controlo da homeostasia dos níveis de IgM circulante no murganho. A diabetes tipo 1 (T1D) é uma doença autoimune complexa que se desenvolve espontaneamente nos humanos e que tem uma patogenia semelhante no murganho NOD (non-obese-diabetic). Os linfócitos B são necessários para o processo diabético do NOD, em que a presença de anticorpos anti-células beta pancreáticas é uma das manifestações mais precoces. O artigo II revelou que as células B1a da cavidade peritoneal do NOD têm uma elevada predisposição para secretarem NAbs que reconhecem autoantigénios de células beta pancreáticas e que podem promover o desenvolvimento de T1D quer pelo aumento da apresentação de autoantigénios, quer pela activação da cascata do sistema de complemento, quer pela indução directa de danos nas células beta pancreáticas. A investigação descrita no artigo III provou que os NAbs secretados por células B1a do NOD têm a capacidade de se ligarem e induzirem stress oxidativo nas células beta do pâncreas. Estes estudos revelaram ainda que as células B1a do NOD têm um limiar reduzido para activação inata e estabeleceram uma relação entre as propriedades das células B1a do NOD, as especificidades dos NAbs e o impacto da ligação de IgM na fisiologia das células beta. Finalmente, o artigo IV evidenciou que a indução de proliferação e diferenciação das células B1a em células secretoras de IgM contribui para o início da T1D. Esta tese demonstrou que o Irf4 é um factor de transcrição com um papel fundamental no controlo da secreção de IgM em animais saudáveis e que, no murganho NOD, as células B1a da cavidade peritoneal têm um menor limiar para estimulação inata, que contribui para o seu estado de activação e para a secreção de IgM autoreactiva, determinando a iniciação e/ou contribuindo para a progressão da diabetes tipo 1.
Fundação para a Ciência e Tecnologia; Instituto Gulbenkian da Ciência
19
Patrick-Melin, Amy J. "Effect of 7 Days Aerobic Exercise on Insulin Sensitivity, Oxidative Stress, TLR2/TLR4 Cell Surface Expression and Cytokine Secretion in Sedentary Obese Adults." Kent State University / OhioLINK, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=kent1310918543.
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Roma, Leticia Prates. "Mecanismos moleculares do efeito citotoxico da dexametasona em linhagens de celulas beta e ilhotas pancreaticas." [s.n.], 2009. http://repositorio.unicamp.br/jspui/handle/REPOSIP/314413.
Анотація:
Orientador: Kleber Luiz de Araujo e SouzaTese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia
Made available in DSpace on 2018-08-13T09:13:30Z (GMT). No. of bitstreams: 1 Roma_LeticiaPrates_D.pdf: 1071638 bytes, checksum: 52777a4c39261ec7137298200cb2b319 (MD5) Previous issue date: 2009
Resumo:Introdução/Objetivos. A produção de espécies reativas de oxigênio (EROs) faz parte de diversos processos fisiológicos. Nos últimos anos, o aumento de EROs têm sido associado ao desenvolvimento de diversas doenças, dentre elas o Diabetes Mellitus Tipo 2. As células beta pancreáticas são notadamente mais suscetíveis ao estresse oxidativo devido a sua baixa capacidade antioxidativa, resultado da menor expressão e atividade de enzimas antioxidantes como superóxido dismutase e peroxidases. A dexametasona, um glicocorticóide sintético, tem efeitos diabetogênicos e citotóxicos em células produtoras de insulina e ilhotas pancreáticas. Entretanto, os mecanismos pelos quais a dexametasona atua sobre as células-alvo não estão bem esclarecidos. Dessa forma, nosso objetivo foi analisar se a dexametasona induz estresse oxidativo em células produtoras de insulina RINm5F e ilhotas pancreáticas. Utilizamos três modelos: 1) células RINm5F controle, que são extremamente sensíveis ao estresse oxidativo; 2) células RINm5F superexpressando a enzima catalase (RINm5F.Cat), que são resistente ao estresse oxidativo e 3) ilhotas de ratos adultos cultivadas por 72 h com dexametasona (Dexa) e ilhotas tratadas concomitantemente com dexametasona e o antioxidante N-acetilcisteína (Dexa+NAC). Resultados: Aumento na produção de EROs foi observado em células RINm5F tratadas com dexametasona. O tratamento com dexametasona aumentou a atividade/clivagem da caspase-3 e apoptose em células RINm5F após 3 dias de cultura. Expressão protéica e atividade de Cu/ZnSOD estava aumentada após o tratamento com dexametasona, enquanto que a expressão/atividade de MnSOD não foi modulada pelo corticóide. A superexpressão da catalase em linhagens de célula beta previniu todos os efeitos citotóxicos da dexametasona, inclusive a morte celular. Elevados níveis de Cu/ZnSOD podem favorecer o aumento na geração de EROs e conseqüentemente, apoptose. Da mesma forma, ilhotas tratadas com dexametasona apresentaramaumento na produção de EROs, efeito que foi revertido quando as ilhotas foram tratadas concomitantemente com dexametasona e NAC. Redução na secreção de insulina estimulada por glicose foi observada em ilhotas cultivadas com dexametasona. O tratamento com dexametasona e NAC restaurou a secreção de insulina a níveis próximos aos controles. Uma menor produção deNAD(P)H no grupo Dexa foi observado, sendo que o grupo Dexa+NAC mostrou níveis semelhantes ao grupo controle. Não ocorreram diferenças nas concentrações intracelulares de cálcio estimulado por glicose em nenhum dos grupos. A dexametasona reduziu a expressão gênica da sinaptotagmina VII, enquanto no grupo Dexa+NAC houve um aumento da expressão desse gene em ilhotas pancreáticas. Interessantemente, o tratamento com NAC diminuiu a expressão gênica da Cu/ZnSOD. Conclusões: Nossos resultados indicam que as ações da dexametasona em células produtoras de insulina e ilhotas pancreáticas são mediadas através do aumento do estresse oxidativo, sendo a Cu/ZnSOD importante nesse processo. A superexpressão da catalase e o uso do antioxidante n-acetilcisteína previnem contra os efeitos citotóxicos do glicocorticóide.
Abstract: Introduction/Aims: Reactive oxygen species (ROS) play a dual role on living organisms, being involved in many physiological processes and also being linked to the development of several pathologies, including the type 2 diabetes mellitus. Pancreatic beta cells are very sensitive to oxidative stress because of their low antioxidant capacity, wich results from their low expression and activity of antioxidant enzymes, especially peroxidases. Dexamethasone is a synthetic diabetogenic glucocorticoid that induces cytotoxic effects on pancreatic beta cells. However, the precise mechanisms of dexamethasone toxicity on target cells are not fully understood. The aim of the present study was to analyzed whether dexamethasone induces oxidative stress in insulinproducing cells and pancreatic islets. Experimental design: The experiments were performed using 3 models: 1) RINm5F control cells, extremely sensitive to oxidative stress; 2) RINm5F cells overexpressing the enzyme catalase (RINm5F.Cat), very resistant to oxidative stress and 3) rat pancreatic islets cultured for 72 h with dexamethasone (Dexa) or cultured concomitantly with dexamethasone and the antioxidant N-acetylcysteine (Dexa+NAC). Results: An increased generation of reative oxygen species (ROS) was observed in dexamethasone-treated insulinproducing cells together with an increase in caspase-3 activity and apoptosis rate. Interestingly, exposure to dexamethasone increased the cytosolic superoxide dismutase Cu/ZnSOD protein expression and activity, while the mitochondrial MnSOD isoform was not affected by the glucocorticoid. Overexpression of catalase in insulin-producing cells prevented all the cytotoxic effects of dexamethasone. Pancreatic islets cultured in the presence of dexamethasone (Dexa) for 72 h showed increased ROS production. Glucose-stimulated insulin secretion was decreased after Dexa treatment. Intracellular ROS levels were decreased and the insulin secretion capacitywas recovered by concomitant treatment with Dexa+NAC. The total insulin content and intracellular Ca+2 levels were not modulated in either Dexa or Dexa+NAC groups. There was a decrease in the NAD(P)H production rate, used as an indicator of viability, after dexamethasone treatment. Concomitant incubation with NAC returned viability to control levels. Dexamethasone also decreased SYT VII gene expression; in contrast, the Dexa+NAC group showed increased expression of SYT VII compared to controls. Surprisingly, treatment with NAC decreased the gene expression of the antioxidant enzyme, Cu/ZnSOD. Conclusions: The cytotoxic effects of dexamethasone in RINm5F insulin-producing cells and pancreatic islets are primarily ROS-mediated. High levels of expression and activity of the Cu/ZnSOD might favour the generation of ROS. The overexpression of catalase and the use of the antioxidant Nacetylcysteine counteract the cytotoxic effects of dexamethasone.
Doutorado
Fisiologia
Doutor em Biologia Funcional e Molecular
21
Alvarenga, Santos Buiate Ester. "ESTABLISHMENT OF BIOTROPHY BY THE MAIZE ANTHRACNOSE PATHOGEN COLLETOTRICHUM GRAMINICOLA: USE OF BIOINFORMATICS AND TRANSCRIPTOMICS TO ADDRESS THE POTENTIAL ROLES OF SECRETION, STRESS RESPONSE, AND SECRETED PROTEINS." UKnowledge, 2015. http://uknowledge.uky.edu/plantpath_etds/17.
Анотація:
Colletotrichum graminicola is a hemibiotrophic pathogen of maize that causes anthracnose leaf and stalk rot diseases. The pathogen penetrates the host and initially establishes an intracellular biotrophic infection, in which the hyphae are separated from the living host cell by a membrane that is elaborated by the host, apparently in response to pathogen signals. A nonpathogenic mutant (MT) of C. graminicola was generated that germinates and penetrates the host normally, but is incapable of establishing a normal biotrophic infection. The mutated gene is Cpr1, conserved in eukaryotes and predicted to encode a component of the signal peptidase complex. How can we explain why the MT is normal in culture and during early stages of pathogenicity, but is deficient specifically in the ability to establish biotrophy? To address this, first I characterized the insertion in the 3’ UTR of the MT strain in detail, something that had not been done before. The wild-type (WT) transcript did not differ from predictions, but the MT produced several aberrant transcript species, including truncated and non-spliced transcripts, and the normal one. Aberrant splicing of MT cpr1 was observed both in RNAseq transcriptome data and reverse-transcription polymerase chain reaction (RT-PCR), under different growth conditions and in planta. I also conducted a bioinformatic analysis of other conserved components of the secretory pathway in the MT and WT in planta. One explanation for nonpathogenicity of the MT is that it cannot cope with an increase in secretory activity during infection, and fails to produce necessary pathogenicity factors. With the transcriptome data, I was able to identify effector proteins that were expressed in the WT but not in the MT. Another possible explanation for the MT phenotype is that the MT can’t adapt to stress imposed by the plant. I developed a growth assay to characterize the effect of chemical stressors in vitro. The MT was more sensitive to most stressors, when compared to the WT. The transcriptome data indicates that the genes involved in different stress pathways are expressed in planta in both WT and MT, although very few genes are differentially expressed across the different growth stages.
22
Stewart, Benjamin J. "Characterization of the effects of the lipid peroxidation products 4-hydroxynonenal and 4-oxononenal on hepatic lipid accumulation, VLDL assembly, secretion, and microtubules : relevance to alcoholic liver disease /." Connect to full text via ProQuest. Limited to UCD Anschutz Medical Campus, 2008.
Анотація:
Thesis (Ph.D. in Toxicology) -- University of Colorado Denver, 2008.Typescript. Includes bibliographical references (leaves 111-122). Free to UCD affiliates. Online version available via ProQuest Digital Dissertations;
23
Botermans, Jos A. M. "Feeding environment for growing-finishing pigs : effects of competition for feed and feeding frequency on performance, behaviour, injuries, plasma cortisol and exocrine pancreatic secretion /." Alnarp : Swedish Univ. of Agricultural Sciences (Sveriges lantbruksuniv.), 1999. http://epsilon.slu.se/avh/1999/91-576-5744-0.pdf.
24
Khalili-Mahani, Najmeh 1971. "Observing the stressed brain : magnetic resonance imaging of the neural correlates of hypothalamic pituitary adrenal axis function." Thesis, McGill University, 2009. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=115857.
Анотація:
The Hypothalamic Pituitary Adrenal (HPA) axis is the coordinator of adaptive responses to physical and psychological stress. The central nervous system plays a key role in modulation of both basal and adaptive HPA axis functions. In fact, since long ago, animal studies have shown that acute and chronic exposure to glucocorticoids (a stress hormone released due to HPA axis activation, cortisol in humans) affects the function and the morphology of brain areas such as the hippocampus and the cingulate cortex. This thesis is based on novel neuroimaging methodologies used to investigate the interactions of psychological stress, cortisol and the brain. It consists of three functional studies and a morphometric one. In the first functional study we show that the hippocampus (where glucocorticoid receptors are most abundant) plays a role in initiation of an HPA axis stress response. In the second study, we provide evidence that besides hippocampus, the neural activity in the so-called "default mode network" (DMN), especially the anterior cingulate cortex (ACC), relates to interindividual variations in HPA axis response to psychological stress. In the third study we have investigated the cortisol-modulation of the DMN. Again, we provide evidence for a role of the ACC and the orbitofrontal cortex in negative feedback inhibition of the HPA axis activity. Finally, we show a morphological link between the ACC and the cortisol response to awakening which is an index of basal HPA axis activity. Overall, our findings confirm the critical role of the ACC and mesolimbic system in HPA axis regulation. These findings also draw attention to the interactions between functional subregions of the medial prefrontal cortex and states of HPA axis function prior to stress onset---suggesting an interplay of the monitoring and the executive planning roles of the medial prefrontal cortex in behavioral adaptation to stress. Beyond stress research, our findings offer a framework for combining neuroimaging and neuroendocrinology to better understand the interindividual variances in behavior, and perhaps to better identify subgroups at risk of psychological disorders.
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Schulz, Simone [Verfasser], and Reinhold [Akademischer Betreuer] Läßle. "Validity and Maintenance of Binge Eating Disorder. Laboratory and naturalistic studies on the role of negative affect, stress-induced eating and cortisol secretion in obese women with BED / Simone Schulz ; Betreuer: Reinhold Läßle." Trier : Universität Trier, 2014. http://d-nb.info/119780692X/34.
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Kuehn, Carina Brigitte. "Développement et études comparatives de méthodes pour améliorer la survie et les fonctions de cellules productrices d'insuline et d'îlots pancréatiques endocriniens porcins en conditions de culture in vitro et de stress apoptotiques." Thèse, Université de Sherbrooke, 2014. http://hdl.handle.net/11143/5411.
Анотація:
Résumé : Durant les dernières années, l’encapsulation d’îlots pancréatiques endocriniens a reçu une grande attention parce qu’elle pourrait constituer une solution pour diminuer les taux d'échecs des transplantations. Dans le contexte de la perte de la matrice extracellulaire (MEC) native des îlots lors de leur isolation et le rejet de greffes par le système immunitaire du receveur, cette thèse vise à améliorer la compréhension des interactions entre la MEC et les cellules des îlots pancréatiques endocriniens ainsi qu’à étudier les effets de stress apoptotiques associés à des éléments du système immunitaire sur la survie et les fonctions des îlots. Ces études pourraient permettre de raffiner notre compréhension des mécanismes associés au rejet des greffes d'îlots de Langerhans.
Dans cette thèse, le premier chapitre constitue une revue de la littérature permettant de mettre en lumière les rôles réciproques de la MEC dans l'action des cellules immunitaires et l'influence de ces rôles sur le diabète de type 1 (DT1) et sur la transplantation d'îlots. Ce premier chapitre a été publié dans la revue Pathologie Biologie.
Le premier travail expérimental comprend la culture de cellules d'insulinomes de rat (INS-1) sur des surfaces composées de carboxyméthyl dextrane (CMD) recouvertes de fibronectine, RGD ou YIGSR, un peptide synthétique de la laminine. Dans cette étude, l'effet bénéfique d’éléments de la MEC sur ces cellules productrices d'insuline a été démontré. Les cellules INS-1 ont davantage proliféré sur ces surfaces et sécrétaient plus d’insuline que les cellules INS-1 cultivées sur les surfaces contrôle de CMD, CMD+RGE et dans les plaques à multi-puits de polystyrène vendues pour la culture tissulaire (TCPS). Cette première étude a été publiée dans Acta Biomaterialia.
La deuxième étude expérimentale avait pour objectif d’étudier l’effet protecteur de gels de fibrine pour enrober des îlots pancréatiques endocriniens isolés de jeunes porcs et exposés à deux concentrations de peroxyde d'hydrogène (H[indice inférieur 2]O[indice inférieur 2]). L’enrobage dans la fibrine a permis de réduire l'apoptose chez les cellules des îlots et d’améliorer la sécrétion d'insuline par ceux-ci lorsque les résultats étaient comparés à ceux des îlots non-enrobés. Ce travail a été publié dans la revue Islets.
Dans la troisième étude expérimentale, des îlots porcins étaient enrobés dans des gels de fibrine et d'alginate et exposés à des monocytes humains pour comparer l’effet de l’enrobage par ces deux matériaux sur la survie et les fonctions des îlots. Les monocytes sécrétaient des
concentrations importantes de cytokines TNFα, IL-6, IL-1β en réponse à la fibrine seule et aux îlots. Les cellules des îlots enrobés dans les gels de fibrine et d'alginate étaient moins apoptotiques et sécrétaient plus d'insuline que leurs contrôles respectifs non-enrobés. Cette étude a été acceptée dans la revue Pathologie Biologie. // Abstract : In recent years, the encapsulation of endocrine pancreatic islets has received enhanced attention as it might constitute a solution for islet transplantation failure. In the context of the loss of the native islet extracellular matrix (ECM) and graft rejection by the recipient’s immune system, this thesis aims to improve the understanding of ECM-islet cell interactions and immune system-related implications in islet survival and function in the context of type 1 diabetes mellitus (T1DM) and islet graft rejection. In the first chapter, a literature review introduces the reciprocal roles of the ECM in immune cell action and the influence of these interactions on T1DM and islet transplantation. The most important ECM components are discussed followed by an overview of immune cells and their possible implication in diabetes. Immune cell integrins and cytokines and their communication with and influence on ECM are highlighted, concluding in a brief discussion of the significance of these interactions for islet transplantation and encapsulation. This review has been accepted for publication by Pathologie Biologie. The first experimental work comprises the culture of rat insulinoma cells (INS-1) on welldefined low-fouling carboxymethyl-dextran (CMD) surfaces covalently grafted with fibronectin, RGD and YIGSR, a synthetic laminin peptide, resulting in higher cell proliferation and insulin secretion of INS-1 cells when compared to the controls CMD, CMD+RGE and tissue culture polystyrene (TCPS) plates. With this work, the beneficial effect of ECM cues on insulin-producing cells was proven. This study has been published in Acta Biomaterialia. The second experimental work aimed to study the effect of fibrin gels when used to embed endocrine pancreatic islets isolated from young pigs and exposed to hydrogen peroxide (H[subscript 2]O[subscript 2]). Fibrin-embedded islets showed less apoptosis and higher relative insulin secretion than islets on TCPS, verifying the protective effect of fibrin towards islets. This study has been published in Islets. In the third experimental study, porcine islets were encapsulated in fibrin and alginate gels and exposed to human monocytes to compare the two materials and to further investigate the immune protective properties of fibrin and alginate. Monocytes secreted high concentrations of TNFα, IL-6, and IL-1β in response to fibrin, but at the same time islets in both fibrin and alginate gels were less apoptotic and secreted more insulin then their TCPS controls. This study has been submitted to Pathologie Biologie.
27
Cerrato, Giulia. "Oleate : An Atypical Cellular Stress Inducer That Stalls Protein Secretion Oleate-Induced Aggregation of LC3 at the Trans-Golgi Network Is Linked to a Protein Trafficking Blockade A Genome-Wide RNA Interference Screen Disentangles the Golgi Tropism of LC3 Live Cell Imaging of LC3 Dynamics." Thesis, université Paris-Saclay, 2021. http://www.theses.fr/2021UPASL023.
Анотація:
Les diverses classes d’acides gras (chaines carbonées saturées ou cis-/trans- insaturées) influencent la physiologie au niveau de la cellule et de l’organisme de façon différente. Curieusement, ces catégories distinctes ont un effet important (mais différent) sur l’autophagie, le mécanisme intracellulaire de dégradation qui maintient l’homéostasie énergétique et protège les cellules contre le stress. L’oléate, l’acide gras cis-insaturé endogène et alimentaire le plus abondant, possède la propriété atypique d’induire une redistribution de la protéine LC3 (signe particulier d’autophagie) de manière non-canonique et préférentiellement dans l’appareil de Golgi. Puisqu’il a été montré que, d’une part, les acides gras cis-insaturés présentent des effets bénéfiques pour la santé et que, d’autre part, les acides gras trans-insaturés et saturés induisent des effets toxiques, nous avons décidé d’explorer les mécanismes à la base de la redistribution de LC3 au niveau de l’appareil de Golgi induite par l’oléate. Cette analyse pourrait nous éclairer sur l’origine des différents effets des acides gras sur la santé. Pour cela, un criblage robotisé du génome entier par ARNs interférents a permis d’identifier plusieurs gènes impliqués dans le transport des protéines lié à l’appareil de Golgi, et également dans la réponse intégrée au stress.Des expériences supplémentaires ont montré que l’oléate impacte la morphologie subcellulaire de l’appareil de Golgi, en corrélation avec le blocage de la sécrétion protéique conventionnelle (dépendante du Golgi) lorsque que la cargaison est bloquée au niveau du réseau trans-golgien. L’inhibition de la sécrétion protéique a été observée dans plusieurs systèmes expérimentaux, tant in vitro qu’in vivo. De plus, un criblage visant à rechercher des agents chimiques capables d’induire les mêmes effets cellulaires que l’oléate, a permis d’identifier plusieurs composés appartenant à diverses classes pharmacologiques. De la même manière que l’oléate ces composés induisent un blocage de la sécrétion protéique conventionnelle, renforçant l’idée que cette voie de perturbation du Golgi joue un rôle pharmacologique important. En conclusion, ces résultats montrent que l’oléate représente une classe de molécules agissant sur l’appareil de Golgi pour y induire l’agrégation de LC3, tout en bloquant en même temps la sécrétion protéiqueDistinct classes of fatty acids (FAs) (saturated or cis-/trans-unsaturated carbon chains) impact on cellular and organismal physiology in a different manner. Interestingly, these diverse categories have a profound (but different) effect on autophagy, the conserved intracellular degradation mechanism that maintains energy homeostasis and protects cells against stress. Oleate, the most abundant endogenous and dietary cis-unsaturated FA, has the atypical property to induce the redistribution of the LC3 protein (peculiar sign of autophagy) in a non-canonical fashion and preferentially to the Golgi apparatus. Intrigued by these observations, which might be related to the health-improving effects of cis-unsaturated FAs (and the notorious toxicity of trans-unsaturated and saturated FAs), we decided to explore the mechanisms causing the oleate-induced relocation of LC3 to the Golgi apparatus. To achieve this goal, a robotized RNA interference genome-wide screen led to the identification of multiple genes involved in the Golgi-related protein transport, as well as in the integrated stress response. Follow-up experiments revealed that oleate affected the subcellular morphology of the Golgi apparatus, correlating with a blockade of conventional (Golgi-dependent) protein secretion that caused secretory cargo to be stalled at the level of the trans-Golgi network. The inhibition of protein secretion was observed using several experimental systems, both in vitro and in vivo. Moreover, a systematic screen searching for other chemical entities that mimic the oleate-induced cellular effects led to the identification of several compounds belonging to rather different pharmacological classes. These “oleate mimetics” also shared with oleate the capacity to block conventional protein secretion, supporting the notion that this pathway of Golgi perturbation is indeed of pharmacological relevance. In conclusion, this research work shows that oleate represents a class of molecules that act on the Golgi apparatus to cause the recruitment of LC3 and to stall protein secretion
28
Nyblom, Hanna K. "Glucotoxicity in Insulin-Producing β-Cells". Doctoral thesis, Uppsala University, Department of Medical Cell Biology, 2007. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-8309.
Анотація:
Background and aims: Type 2 diabetes mellitus is connected with elevated glucose levels, which cause impaired glucose-stimulated insulin secretion (GSIS) and degeneration of β-cells. Mechanisms for such glucotoxic effects were explored in the present study.
Materials and methods: INS-1E cells were cultured for 5 days in 5.5, 11, 20 or 27 mM glucose in the presence or absence of AMPK-agonist AICAR. GSIS was determined from INS-1E cells and islets obtained from type 2 diabetes and control donors. Human islets and INS-1E cells were functionally characterized (GSIS) and protein profiled (SELDI-TOF MS). Glucose-induced de novo synthesis of fatty acyls (HR-MAS NMR spectroscopy), fatty acid composition (GC-MS), triglyceride content and specific proteins (Western blotting) were determined in INS-1E cells.
Results: Impaired GSIS was observed from INS-1E cells exposed to chronic hyperglycaemia and islets isolated from type 2 diabetics compared to INS-1E cells cultured at normal glucose levels and control islets, respectively. Several glucose-regulated proteins were found when type 2 diabetes and control islets or mitochondria from INS-1E cells cultured at different glucose concentrations were protein profiled. Glucose induced lipid de novo synthesis of both saturated and unsaturated fatty acids in specific proportions. Glucose-induced impairment of function and mass was reverted by inclusion of AICAR, which lowered levels of pro-apoptotic protein CHOP but left triglyceride content unaffected.
Conclusions: Impaired GSIS and increased apoptosis observed in β-cells after prolonged exposure to elevated glucose concentrations involved accumulation of lipid species in specific proportions, AMPK-inactivation, ER-stress activation and complex, coordinated changes in expression patterns of mitochondrial and human islet proteins.
29
Mühlhan, Markus, Ulrike Lüken, Jens Siegert, Hans-Ulrich Wittchen, Michael N. Smolka, and Clemens Kirschbaum. "Enhanced Sympathetic Arousal in Response to fMRI Scanning Correlates with Task Induced Activations and Deactivations." Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2014. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-127271.
Анотація:
It has been repeatedly shown that functional magnetic resonance imaging (fMRI) triggers distress and neuroendocrine response systems. Prior studies have revealed that sympathetic arousal increases, particularly at the beginning of the examination. Against this background it appears likely that those stress reactions during the scanning procedure may influence task performance and neural correlates. However, the question how sympathetic arousal elicited by the scanning procedure itself may act as a potential confounder of fMRI data remains unresolved today. Thirty-seven scanner naive healthy subjects performed a simple cued target detection task. Levels of salivary alpha amylase (sAA), as a biomarker for sympathetic activity, were assessed in samples obtained at several time points during the lab visit. SAA increased two times, immediately prior to scanning and at the end of the scanning procedure. Neural activation related to motor preparation and timing as well as task performance was positively correlated with the first increase. Furthermore, the first sAA increase was associated with task induced deactivation (TID) in frontal and parietal regions. However, these effects were restricted to the first part of the experiment. Consequently, this bias of scanner related sympathetic activation should be considered in future fMRI investigations. It is of particular importance for pharmacological investigations studying adrenergic agents and the comparison of groups with different stress vulnerabilities like patients and controls or adolescents and adults.
30
Mühlhan, Markus, Ulrike Lüken, Jens Siegert, Hans-Ulrich Wittchen, Michael N. Smolka, and Clemens Kirschbaum. "Enhanced Sympathetic Arousal in Response to fMRI Scanning Correlates with Task Induced Activations and Deactivations." Public Library of Science, 2013. https://tud.qucosa.de/id/qucosa%3A27292.
Анотація:
It has been repeatedly shown that functional magnetic resonance imaging (fMRI) triggers distress and neuroendocrine response systems. Prior studies have revealed that sympathetic arousal increases, particularly at the beginning of the examination. Against this background it appears likely that those stress reactions during the scanning procedure may influence task performance and neural correlates. However, the question how sympathetic arousal elicited by the scanning procedure itself may act as a potential confounder of fMRI data remains unresolved today. Thirty-seven scanner naive healthy subjects performed a simple cued target detection task. Levels of salivary alpha amylase (sAA), as a biomarker for sympathetic activity, were assessed in samples obtained at several time points during the lab visit. SAA increased two times, immediately prior to scanning and at the end of the scanning procedure. Neural activation related to motor preparation and timing as well as task performance was positively correlated with the first increase. Furthermore, the first sAA increase was associated with task induced deactivation (TID) in frontal and parietal regions. However, these effects were restricted to the first part of the experiment. Consequently, this bias of scanner related sympathetic activation should be considered in future fMRI investigations. It is of particular importance for pharmacological investigations studying adrenergic agents and the comparison of groups with different stress vulnerabilities like patients and controls or adolescents and adults.
31
Herson, Paco S. "Oxidative stress activates a novel non-selective cation channel in insulin-secreting cells." Thesis, University of Aberdeen, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.265376.
Анотація:
Single channel recordings from CRI-G1 insulin-secreting cells were used to characterize a novel ion channel. The presence of both Ca2+ and β-NAD+ at the cytoplasmic aspect of the membrane are required for channel activity. This is the first ion channel described which requires internal β-NAD+ for activity (thus termed NSNAD). The channel was found to be permeable to all monovalent (Na+, K+ and Cs+) and divalent cations tested (Ca2+, Mg2+, Ba2+, and Mn2+). The slope conductance is relatively large (70 - 90pS) compared to other non-selective cation channels and also has extremely slow kinetics (open and closed times in the range of seconds). Whole-cell voltage clamp experiments illustrate that internal β-NAD+ activates a cation current consistent with activation of the NSNAD channel. Similar to the single NSNAD channel, the β-NAD+-activated current was sensitive to the internal concentrations of both Ca2+ and β-NAD+. The non-selective nature of this cation current was confirmed by replacement of the internal K+ with Cs+ which did not diminish the β-NAD+-activated current. Additionally, replacement of external cations with the impermeant NMDG abolished the β-NAD+-activated current. The diabetogenic agent alloxan was found to irreversibly depolarize CRI-GI cells by opening a non-selective cation channel with characteristics similar to the NSNAD channel. The channel activated by alloxan is characterized by a slope conductance of approximately 70 pS and very slow (seconds) kinetics. Channel activity is lost upon excision of the patch, but can be re-activated by the application of internal β-NAD+. The mechanism of alloxan-induced depolarization and channel activation appears to be through the production of reactive oxygen species (ROS). This data indicates that oxidative stress generated by both alloxan and H2O2 causes the activation of the NSNAD channel which results in irreversible collapse of the membrane potential and massive Ca+ influx leading to eventual cell death. This may represent a component of the destruction of pancreatic β-cells during type I diabetes and possibly other pathologies in which oxidative stress is implicated.
32
DeAngelis, Cara Marie. "Characterization of the Vibrio cholerae Phage Shock Protein Response." University of Toledo Health Science Campus / OhioLINK, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=mco1556723496251854.
33
Lipson, Kathryn L. "The Role of Endoplasmic Reticulum Stress Signaling in Pancreatic Beta Cells: a Dissertation." eScholarship@UMMS, 2008. https://escholarship.umassmed.edu/gsbs_diss/363.
Анотація:
Protein folding in the endoplasmic reticulum (ER) is essential for proper cellular function. However, the sensitive environment in the ER can be perturbed by both pathological processes as well as by physiological processes such as a large biosynthetic load placed on the ER. ER stress is a specific type of intracellular stress caused by the accumulation of immature or abnormal misfolded or unfolded proteins in the ER. Simply defined, ER stress is a disequilibrium between ER load and folding capacity. Cells have an adaptive response that counteracts ER stress called the "Unfolded Protein Response” (UPR). The ability to adapt to physiological levels of ER stress is especially important for maintaining ER homeostasis in secretory cells. This also holds true for pancreatic β-cells, which must fold and process large amounts of the hormone insulin.
Pancreatic β-cells minimize abnormal levels of glycemia through adaptive changes in the production and regulated secretion of insulin. This process is highly sensitive, so that small degrees of hypo- or hyperglycemia result in altered insulin release. The frequent fluctuation of blood glucose levels in humans requires that β-cells control proinsulin folding in the ER with exquisite sensitivity. Any imbalance between the load of insulin translation into the ER and the actual capacity of the ER to properly fold and process the insulin negatively affects the homeostasis of β-cells and causes ER stress.
In this dissertation, we show that Inositol Requiring 1 (IRE1), an ER-resident kinase/endoribonuclease and a central regulator of ER stress signaling, is essential for maintaining ER homeostasis in pancreatic β-cells. Importantly, IRE1 has a crucial function in the body’s normal production of insulin in response to high glucose. Phosphorylation and subsequent activation of IRE1 by transient exposure to high glucose is coupled to insulin biosynthesis, while inactivation of IRE1 by siRNA or inhibition of IRE1 phosphorylation abolishes insulin biosynthesis. IRE1 signaling under these physiological ER stress conditions utilizes a unique subset of downstream components of IRE1 and has a beneficial effect on pancreatic β-cell homeostasis.
In contrast, we show that chronic exposure of β-cells to high glucose causes pathological levels of ER stress and hyperactivation of IRE1, leading to the degradation of insulin mRNA. The term “glucose toxicity” refers to impaired insulin secretion by β-cells in response to chronic stimulation by glucose and is characterized by a sharp decline in insulin gene expression. However, the molecular mechanisms of glucose toxicity are not well understood. We show that hyperactivation of IRE1 caused by chronic high glucose treatment or IRE1 overexpression leads to insulin mRNA degradation in pancreatic β-cells. Inhibition of IRE1 signaling using a dominant negative form of the protein prevents insulin mRNA degradation in β-cells. Additionally, islets from mice heterozygous for IRE1 retain expression of more insulin mRNA after chronic high glucose treatment than do their wild-type littermates.
This work suggests that the rapid degradation of insulin mRNA could provide immediate relief for the ER and free up the translocation machinery. Thus, this mechanism may represent an essential element in the adaptation of β-cells to chronic hyperglycemia. This adaptation is crucial for the maintenance of β-cell homeostasis and may explain in part why the β-cells of diabetic patients with chronic hyperglycemia stop producing insulin without simply undergoing apoptosis. This work implies that prolonged activation of IRE1 signaling is involved in the molecular mechanisms underlying glucose toxicity.
This work therefore reveals two distinct activities elicited by IRE1 in pancreatic β-cells. IRE1 signaling activated by transient exposure to high glucose enhances proinsulin biosynthesis, while chronic exposure of β-cells to high glucose causes hyperactivation of IRE1, leading to the degradation of insulin mRNA. Physiological IRE1 activation by transient high glucose levels in pancreatic β cells has a beneficial effect on insulin biosynthesis. However, pathological IRE1 activation by chronic high glucose or experimental drugs negatively affects insulin gene expression. In the future, a system to induce a physiological level of IRE1 activation, and/or reduce the pathological level of IRE1 activation could be used to enhance insulin biosynthesis and secretion in people with diabetes, and may lead to the development of new and more effective clinical approaches to the treatment of this disorder.
34
Fonseca, Sonya G. "Role of WFS1 in Regulating Endoplasmic Reticulum Stress Signaling: A Dissertation." eScholarship@UMMS, 2009. https://escholarship.umassmed.edu/gsbs_diss/414.
Анотація:
The endoplasmic reticulum (ER) is a multi-functional cellular compartment that functions in protein folding, lipid biosynthesis, and calcium homeostasis. Perturbations to ER function lead to the dysregulation of ER homeostasis, causing the accumulation of unfolded and misfolded proteins in the cell. This is a state of ER stress. ER stress elicits a cytoprotective, adaptive signaling cascade to mitigate stress, the Unfolded Protein Response (UPR). As long as the UPR can moderate stress, cells can produce the proper amount of proteins and maintain a state of homeostasis. If the UPR, however, is dysfunctional and fails to achieve this, cells will undergo apoptosis.
Diabetes mellitus is a group of metabolic disorders characterized by persistent high blood glucose levels. The pathogenesis of this disease involves pancreatic β-cell dysfunction: an abnormality in the primary function of the β-cell, insulin production and secretion. Activation of the UPR is critical to pancreatic β-cell survival, where a disruption in ER stress signaling can lead to cell death and consequently diabetes. There are several models of ER stress leading to diabetes. Wolcott-Rallison syndrome, for example, occurs when there is a mutation in the gene encoding one of the master regulators of the UPR, PKR-like ER kinase (PERK).
In this dissertation, we show that Wolfram Syndrome 1 (WFS1), an ER transmembrane protein, is a component of the UPR and is a downstream target of two of the master regulators of the UPR, Inositol Requiring 1 (IRE1) and PERK. WFS1 mutations lead to Wolfram syndrome, a non-autoimmune form of type 1 diabetes accompanied by optical atrophy and other neurological disorders. It has been shown that patients develop diabetes due to the selective loss of their pancreatic β-cells. Here we define the underlying molecular mechanism of β-cell loss in Wolfram syndrome, and link this cell loss to ER stress and a dysfunction in a component of the UPR, WFS1. We show that WFS1 expression is localized to the β-cell of the pancreas, it is upregulated during insulin secretion and ER stress, and its inactivation leads to chronic ER stress and apoptosis.
This dissertation also reveals the previously unknown function of WFS1 in the UPR. Positive regulation of the UPR has been extensively studied, however, the precise mechanisms of negative regulation of this signaling pathway have not. Here we report that WFS1 regulates a key transcription factor of the UPR, activating transcription factor 6 (ATF6), through the ubiquitin-proteasome pathway. WFS1 expression decreases expression levels of ATF6 target genes and represses ATF6-mediated activation of the ER stress response (ERSE) promoter. WFS1 recruits and stabilizes an E3 ubiquitin ligase, HMG-CoA reductase degradation protein 1 (HRD1), on the ER membrane. The WFS1-HRD1 complex recruits ATF6 to the proteasome and enhances its ubiquitination and proteasome-mediated degradation, leading to suppression of the UPR under non-stress conditions. In response to ER stress, ATF6 is released from WFS1 and activates the UPR to mitigate ER stress.
This body of work reveals a novel role for WFS1 in the UPR, and a novel mechanism for regulating ER stress signaling. These findings also indicate that hyperactivation of the UPR can lead to cellular dysfunction and death. This supports the notion that tight regulation of ER stress signaling is crucial to cell survival. This unanticipated role of WFS1 for a feedback loop of the UPR is relevant to diseases caused by chronic hyperactivation of ER stress signaling network such as pancreatic β-cell death in diabetes and neurodegeneration.
35
Oslowski, Christine M. "TXNIP is a Mediator of ER Stress-Induced β-Cell Inflammation and Apoptosis: A Dissertation". eScholarship@UMMS, 2012. https://escholarship.umassmed.edu/gsbs_diss/611.
Анотація:
Diabetes mellitus is a group of metabolic disorders characterized by hyperglycemia. The pathogenesis of these diseases involves β-cell dysfunction and death. The primary function of β-cells is to tightly regulate the secretion, production, and storage of insulin in response to blood glucose levels. In order to manage insulin biosynthesis, β-cells have an elaborate endoplasmic reticulum (ER).
The ER is an essential organelle for the proper processing and folding of proteins such as proinsulin. Proteins fold properly when the ER protein load balances with the ER folding capacity that handles this load. Disruption of this ER homeostasis by genetic and environmental stimuli leads to an accumulation of misfolded and unfolded proteins, a condition known as ER stress. Upon ER stress, the unfolded protein response (UPR) is activated. The UPR is a signaling network that aims to alleviate ER stress and restore ER homeostasis promoting cell survival. Hence, the UPR allows β-cells to handle the physiological fluctuations of insulin demand.
However upon severe unresolvable ER stress conditions such as during diabetes progression, the UPR switches to pathological outputs leading to β-cell dysfunction and apoptosis. Severe ER stress may also trigger inflammation and accumulating evidence suggests that inflammation also contributes to β-cell failure, but the mechanisms remain elusive.
In this dissertation, we demonstrate that thioredoxin interacting protein (TXNIP) mediates ER stress induced β-cell inflammation and apoptosis. During a DNA microarray analysis to identify novel survival and death components of the UPR, we identified TXNIP as an interesting proapoptotic candidate as it has been linked to glucotoxicity in β-cells. During our detailed investigation, we discovered that TXNIP is selectively expressed in β-cells of the pancreas and is strongly induced by ER stress through the IRE1α and PERK-eIF2α arms of the UPR and specifically its transcription is regulated by activating transcription factor 5 (ATF5) and carbohydrate response element binding protein (ChREBP) transcription factors.
As TXNIP has been shown to activate the Nod-like receptor protein 3 (NLRP3) inflammasome leading to the production of the inflammatory cytokine interleukin-1β (IL- 1β), we hypothesized that perhaps TXNIP has a role in IL-1β production under ER stress. We show that ER stress can induce IL-1β production and that IL-1β is capable of binding to IL-1 type 1 receptor (IL-1R1) on the surface of β-cells stimulating its own expression. More importantly, we demonstrate that TXNIP does indeed play a role in ER stress mediated IL-1β production through the NLRP3 inflammasome. Furthermore, we also confirmed that TXNIP is a mediator of β-cell apoptosis under ER stress partially through IL-1β signaling.
Collectively, we provide significant novel findings that TXNIP is a component of the UPR, mediates IL-1β production and autostimulation, and induces cell death under ER stress in β-cells. It is becoming clear that TXNIP has a role in the pathogenesis of diabetes and is a link between ER stress, oxidative stress and inflammation. Understanding the molecular mechanisms involved in TXNIP expression, activity, and function as we do here will shed light on potential therapeutic strategies to tackle diabetes.
36
Dugani, Aisha Mohammed. "Effects of dopamine agonists and antagonists on gastric acid secretion and stress ulcer formation." 1987. http://hdl.handle.net/1993/15446.
37
Chou, Sheng-Hsia, and 周聖夏. "Effects of docosahexaenoic acid deficiency on stress-induced glucocorticoid secretion in adult male rats." Thesis, 2013. http://ndltd.ncl.edu.tw/handle/44595451536292047639.
Анотація:
碩士國立臺灣大學
生理學研究所
101
Most previous studies have focused on rat docosahexaenoic acid (22:6n-3, DHA) deficiency created during brain development via maternal n-3 fatty acid-deficient diet from pregnancy and lactation on adult anxiety and depression behaviors. It has been suggested the anxiety and depression is associated with hypothalamic-pituitary-adrenal axis (HPA axis) responses. The aim of this study was to evaluate whether brain DHA deficiency during post-weaning period would enhance restrain-induced HPA axis responses and anxiety or depression-like behavior in adult male rats. After weaning at postnatal day 21, male rats fed an n-3 fatty acid-deficient or an n-3 fatty acid-adequate diet till sacrificed at postnatal day 70. The DHA levels were significant decreased in hypothalamus and hippocampus and other brain regions in rats fed with n-3 fatty acid-deficient diet compared to the rats fed with n-3 fatty acid-adequate diet during post-weaning period for 7 weeks. The anxiety-like behavior in the elevated plus-maze test and depressive-like behavior in the forced swim test were no difference between groups. The restraint-induced rectal temperature was increased during the restraint, but no difference between groups. The serum corticosterone was significant increased right after restraint in rats fed an n-3 fatty acid-deficient diet. The GR expression in hippocampus and hypothalamus were no difference between groups. The BDNF was significant decreased in hippocampus after restraint in rats fed n-3 fatty acid-deficient diet compared to the rats fed n-3 fatty acid-adequate diet. This study suggest that brain DHA level can be decreased in male rats fed n-3 fatty acid-deficient diet during post-weaning period, and the decreased brain DHA is associated with higher stress-induced serum corticosterone levels.
38
Huang, Lih-Rong, and 黃麗榕. "Effect of L-Glutamic Acid on Gastric Secretion and Stress-induced Gastric Lesions -- an in Vivo Study." Thesis, 1995. http://ndltd.ncl.edu.tw/handle/29291475582945156334.
Анотація:
碩士台北醫學大學
醫學研究所
83
The effects of L-glutamic acid (L-Glu) on gastric acid secretion were incestigated in vivo. All SD rats were anesthetized with sodium pentobarbital (45 mg/kg, i.p.) after 24 hrs of fasting. The trachea、esophagus、femoral vein and duodenum were catheterized. The stomach was flushed throuth the esophagus cannula via a peristaltic pump with normal saline at room temperature. Acid output was determined by titration (autotitrator VIT90, Radiometer Corp., Corpenhagen, Denmark) of the perfusate with 0.01N NaOH to pH 7.0. after basal secretions were collected for 30 min, and various acid stimulator (histamine, oxotremorine or pentagastrin) were infused for 60 min. In addition, L-glutamic acid (L-Glu) and 6, 7-dinitroquinoxaline-2, 3-dione (DNQX) was infused for 30 min after the infusion of histamine for 30 min. The result was found that infusion with synthetic L-Glu alone had no effect on spontaneous acid secretion. The histamine-(2 mg/kg/hr) and oxotremorine-(1 ug/kg/hr) stimulated acid secretion and were markedly reduced by L-Glu (750 ug/kg/hr), whereas L-Glu had no effect on acid secretion stimulated by pentagestrin (5 ug/kg/hr). Furthermore, this inhibitory effect of L-Glu on hisstamine-stimulated acid secretion was blocked by DNQX (750 ug/kg/hr), a non-DMDA receptor antagonist. On the other hand, L-Glu and it's subtypes including N-meghy1-D-aspartate (NMDA), kainic acid (KA) and quisqualic acid (QA), which functions to protect mucosal damage and reduce cyclic AMP concentration were also investigated in stres-induced gastric lesions. L-Glu and it's eubtypes were administered before cold-restraint stress (4 + 1℃, 2 hrs). The cAMP concentration of cold restraint-induced mucosal lesion was significantly different from the normal stomach. The present study was conducted to evaluate the effect of these drugs on the development of cold and restraint-induced gastric ulcers. Two hours of restraint at 4 + 1℃ resulted in the production of gastric lesions in all mice. The cAMP concentration of cold restraint-induced was inhibited by L-Glu and it's subtypes. L-Glu at 2, 4 and 8 mg/kg i.p. injection 30 min before strss significantly and dose dependently prevented gastric lesions. NMDA at 0.2, 1 and 2 mg/kg , QA at 2 and 5 mg/kg or KA at 1, 2 and 5 mg/kg administered 30 min before stress prevented gastric ulcer in dose dependently. All these results suggest that L-Glu is involved in the inhibition of oxotremorine-and histamine-induced gastric acid secretion via ionotropic non-NMDA receptors. L-Glu and it's subtypes also may participate in a physiological modulation of the gastric mucosal barrier, by increasing resistance to cold restraint-induced gastric lesions in mice.
39
Diamond, Scott Lee. "Effect of laminar shear stress on gene regulation, protein synthesis, and protein secretion by cultured human endothelial cells." Thesis, 1990. http://hdl.handle.net/1911/16336.
Анотація:
To test the hypothesis that wall shear stress generated by blood flow may regulate endothelial cell expression of blood clot dissolving proteins or vasoactive proteins, an in vitro perfusion system was used to expose human umbilical vein endothelial cell monolayers to well defined, laminar fluid flow. Protein production studies utilized immunoassays, while semi-quantitative studies of messenger RNA levels in a small numbers of cells required a reverse transcription/polymerase chain reaction technique.
Secretion by endothelial cells of the two main regulators of the fibrinolytic (ie blood clot dissolving) system, tissue plasminogen activator (tPA) and plasminogen activator inhibitor, type 1 (PAI-1) were not affected by exposure to venous levels of shear stress (4 dynes/cm$\sp2$). However, at arterial shear stresses of 15 and 25 dynes/cm$\sp2$, the tPA secretion rate was 2.1 and 3.0 times greater, respectively, than the basal tPA secretion rate. PAI-1 secretion was unstimulated by shear stress over the entire physiological range. The tPA mRNA level was many fold higher ($>$10 fold) in endothelial cells sheared for 24 hours than in stationary controls. The mRNA level of the common house-keeping gene, glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was found to be the same in control and sheared cells. The fact that GAPDH was unregulated indicates selectively in cellular response to shear stress in addition to validating the PCR technique.
Endothelin (ET), a 21-amino acid peptide secreted by endothelial cells, has vasoconstrictor and mitogenic activity for vascular smooth muscle cells. Fluid shear stress of 25 dynes/cm$\sp2$ caused a rapid and sustained drop in endothelin production after only 2 hours exposure to shear stress. Endothelin secretion was not affected by venous shear stress of 4 dynes/cm$\sp2$. The mRNA level for endothelin in cells exposed to shear stress was almost undetectable, indicating that the drop in protein secretion is due to a drop in transcription of the message RNA for endothelin. The mRNA level of basic fibroblast growth factor (bFGF) was found to be the same in cells sheared for 24 hours as in controls.
Enhancement of the fibrinolytic potential of endothelial cells in response to hemodynamic forces involves transcriptional events and could explain the deposition of fibrin in low shear zones near arterial bifurcations. Flow-regulated ET expression may explain the inverse correlation of fluid shear stress with: (1) the localization of atherosclerotic lesions near vessel bifurcations and; (2) the severity of intimal hyperplasia in surgical vein bypass grafts and vessel anastomotic sites.
40
Subramaney, Ugasvaree. "Personality style, cortisol secretion and the inflammatory response to trauma exposure in a cohort of South African metro police cadets: a prospective, longitudinal study." Thesis, 2012. http://hdl.handle.net/10539/11051.
Анотація:
Literature investigating trauma exposure, Posttraumatic stress disorder (PTSD) and
cortisol secretion has produced conflicting results with regard to whether cortisol is
increased or decreased. With trauma there is also a pro- inflammatory response which
is intimately linked with the hypothalamic pituitary axis (HPA). The police population can
offer useful information in this regard as they represent a sample that will undergo
exposure to traumatic events as part of their normal duties. In South Africa few studies
have examined biological correlates of the traumatic stress response in the police
population.
This study sought to determine whether correlations exist between cortisol and the
inflammatory response in terms of the cytokines Interleukin 6 (IL6) and Tumour Necrosis
Factor (TNF) in response to trauma exposure in a cohort of newly enrolled metro police
officers, previously naïve to the duty related trauma exposure. Personality styles were
assessed, as coping skills and personality have been suggested as factors determining
responses to trauma.
The study participants were followed up for one year with repeated measures analysis of
urine, blood, and saliva cortisol as well as blood cytokine determination every 3 months.
Measures for PTSD [the Clinician Administered Scale for Posttraumatic stress disorder
(CAPS) and the revised version of the Impact of Event Scale (IES-R)] as well as for
depression [the Hamilton Depression Rating Scale (HAM-D)] were undertaken.
145 new recruits volunteered for the study, of which 120 completed all 5 visits. There
were slightly more females than males in the sample and almost 50% of the sample admitted to alcohol abuse. Trauma exposure on entry into the police force was
remarkably high with 99% having been exposed to at least one traumatic event in their
lives. The majority (61.1 %) had been exposed to more than one traumatic event. There
was evidence for the influence of prior trauma on responses to current traumatic events.
MVA’s were very common, both duty and non duty related. Certain traumas were
associated with greater changes in scores for PTSD and depression in relation to
baseline. Over the 5 visits, only a third submitted valid 24 hour urine samples. Of these,
the profile of the entire group indicated that 24 hour urine cortisol tended to initially
decrease, and then increase with time. Saliva and blood cortisol, which were more
reliably measured, tended to decrease with time.
Scores for depression and post traumatic stress disorder were generally low in response
to duty related traumatic events, and tended to decrease over time. However, the
prevalence of lifetime PTSD as measured by the CAPS was high.
There was a strong linear correlation between TNF and IL6. Results indicate a
proinflammatory response, particularly with regard to IL6. There were no significant
correlations between blood cortisol and HAM-D and between blood cortisol and CAPS
(lifetime). There was an inverse relationship between CAPS (current scores) and blood
cortisol. Cortisol and IES-R scores were significant at visit 3 (inverse relationship). For
saliva, there were no significant associations with any of the variables for PTSD and
depression.
For personality styles, aggressive and antisocial clinical patterns were associated with
lower current CAPS scores, while schizoid clinical pattern and the severe syndrome
scale of thought disorder showed an association with lower lifetime CAPS score. For the
IES-R, only narcissistic clinical pattern was associated with lower scores. A further analysis of those with low (less than 25% of the median) and high (greater than 25% of
the median) cortisol responses was undertaken.
The results indicate a similar trend to some studies showing lowered cortisol levels with
chronic trauma exposure, but this did not correlate with sufficiently high scores for PTSD
as measured by the CAPS. Similarly, proinflammatory cytokine increases are evident
with trauma exposure, but not with scores for PTSD and depression. There were more
variables significantly associated with the low cortisol responders than the high cortisol
responders; with a suggestion of cumulative trauma exposure correlating with low
cortisol response and a corresponding pro inflammatory response in terms of IL6.
The results are discussed with a view to assisting the metro police force with recruitment
and counseling strategies and important future research is recommended.
41
Chen, Yu-Ting, and 陳育廷. "Role of oxidative stress in the development of mild portal endotoxemia-induced chronic hepatic inflammation and impairment of pancreatic insulin secretion." Thesis, 2008. http://ndltd.ncl.edu.tw/handle/11640773136953732044.
Анотація:
碩士國防醫學院
生理學研究所
96
Portal endotoxemia has been considered as a key pathogenic factor in chronic hepatic inflammation, which have shown to highly associate with metabolic syndrome and type 2 diabetes. Our previous study demonstrated that low-dose intraportal endotoxin infusion could induce chronic hepatic inflammation and impair endocrine pancreatic function. This study was to further test the role of oxidative stress in the pathogenesis of the portal endotoxemia-induced damages in liver and pancreas. from normal rats and fructose-induced insulin resistant rats with fatty liver. Chronic portal infusion was performed under the status of normal and fatty liver condition with portal catheterization connected to lipopolysaccharide (LPS, 0.42 ng/kg/min) or saline-filled osmotic mini-pump for 4 weeks. The rats with intraportal LPS infusion were further divided into two subgroups combined with or without α-Lipoic acid treatment (60 mg/kg/day, P.O.). Our results showed that mild portal endotoxemia created by low-dose intraportal LPS infusion induced subacute hepatic and pancreatic inflammation indicated by the increases in tissue contents of TNF-alpha, IL-6 and superoxide both in normal and fructose-fed rats. Portal LPS infusion significantly attenuated glucose-stimulated insulin secretion shown in hyperglycermic clamp study, but increased plasma amylase, CRP, superoxide levels and WBC count in normal rats. α-Lipoic acid administration significantly alleviated the portal endotoxemia -induced pathological changes in liver and pancreas by histological examination. α-Lipoic acid also reduced plasma amylase, CRP and superoxide production in normal rats with portal LPS infusion. Fructose-induced attenuation in insulin-stimulated glucose uptake was further deteriorated in those with portal LPS infusion and was partially reversed in those LPS-treated rats with α-Lipoic acid administration. Fructose-induced increases in plasma amylase, CRP, superoxide, and WBC counts were further deteriorated in those with LPS treatment. α- Lipoic acid significantly attenuated the detrimental effects of portal endotoxemia on the above-mentioned parameters in fructose-fed rats. These results showed that α-Lipoic acid effectively attenuated portal endotoxemia-induced hepatic inflammation and pancreatic insulin secretion, implicating that augmentation of oxidative stress is crucial for the detrimental effects of chronic portal endotoxemia on liver and pancreas in normal rats and fructose-fed rats, an animal model of metabolic syndrome with fatty liver.
42
"Phycocyanin protects INS-1E pancreatic beta cells against human islet amyloid polypeptide-induced apoptosis through attenuating oxidative stress and mitochondrial dysfunction." Thesis, 2010. http://library.cuhk.edu.hk/record=b6075047.
Анотація:
Additionally, cyclosporin A, an inhibitor of the mitochondrial permeability transition (MPT) pore, failed to prevent hIAPP-induced DeltaPsim collapse, cytochrome c and AIF release and caspase-3 activation, indicating that the MPT pore was not involved in hIAPP-induced apoptosis. On the other hand, potential crosstalk between the extrinsic and intrinsic apoptotic pathways was demonstrated by cleavage of Bid by caspase-8 in the apoptotic process triggered by hIAPP.It is widely accepted that human islet amyloid polypeptide (hIAPP) aggregation plays an important role in the loss of insulin-producing pancreatic beta cells. Insulin secretion impairment and cell apoptosis can be due to mitochondrial dysfunction in pancreatic beta cells. hIAPP-induced cytotoxicity is mediated by the generation of reactive oxygen species (ROS). Phycocyanin (PC) is a natural compound from blue-green algae that is widely used as food supplement. Currently, little information is available about the effect of hIAPP on mitochondrial function of beta cells and protection of PC against hIAPP-induced cytotoxicity. In this thesis, I hypothesize that hIAPP may impair beta cell function with the involvement of mitochrondrial dysfunction, and this effects could be attenuated by PC. Therefore, the aim of this study was to investigate the role of mitochondria in hIAPP-induced apoptosis, the in vitro protective effects of PC and explore the underlying mechanisms.
It was found that hIAPP induced apoptosis in INS-1E cells with the disruption of mitochondrial function, as evidenced by ATP depletion, mitochondrial mass reduction, mitochondrial fragmentation and loss of mitochondrial membrane potential (DeltaPsim). Further molecular analysis showed that hIAPP induced changes in the expression of Bcl-2 family members, release of cytochrome c and apoptosis-inducing factor (AIF) from mitochondria into cytosol, activation of caspases and cleavage of poly (ADP-ribose) polymerase. Interestingly, the hIAPP-induced mitochondrial dysfunction in INS1-E cells was effectively restored by co-treatment with PC.
Our results showed that hIAPP inhibited the INS-1E cell growth in a dose-dependent manner. However, cytotoxicity of hIAPP was significantly attenuated by co-incubation of the cells with PC. hIAPP induced DNA fragmentation and chromatin condensation, which were key characteristics of cell apoptosis. These changes were inhibited by PC as examined by TUNEL assay and DAPI staining. Moreover, PC significantly prevented the hIAPP-induced overproduction of intracellular ROS and malonaldehyde (MDA), as well as changes of activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) enzymes. Furthermore, hIAPP triggered the activation of mitogen-activated protein kinases (MAPKs) such as c-Jun N-terminal kinase (JNK) and p38 kinase, and these effects were effectively suppressed by PC.
Taken together, I have demonstrated for the first time the involvement of mitochondrial dysfunction in hIAPP-induced INS-1E cell apoptosis, which was attenuated by PC through attenuating oxidative stress, modulating JNK and p38 pathways and reducing mitochondrial dysfunction.
Li, Xiaoling.
Adviser: Juliana Chung Ngor Chan.
Source: Dissertation Abstracts International, Volume: 73-01, Section: B, page: .
Thesis (Ph.D.)--Chinese University of Hong Kong, 2010.
Includes bibliographical references (leaves 150-159).
Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
Abstract also in Chinese.
43
Barbeau, Annie. "Rôle de l'estérification des acides gras dans la régulation de la sécrétion d'insuline et le stress métabolique induits par le glucose." Thèse, 2012. http://hdl.handle.net/1866/7063.
Анотація:
Le diabète est une maladie chronique de l’homéostasie du glucose caractérisée par une hyperglycémie non contrôlée qui est le résultat d’une défaillance de la sécrétion d’insuline en combinaison ou non avec une altération de l’action de l’insuline. La surnutrition et le manque d’activité physique chez des individus qui ont des prédispositions génétiques donnent lieu à la résistance à l’insuline. Pendant cette période dite de compensation où la concentration d’acides gras plasmatiques est élevée, l’hyperinsulinémie compense pleinement pour la résistance à l’insuline des tissus cibles et la glycémie est normale.
Le métabolisme du glucose par la cellule pancréatique bêta entraîne la sécrétion d’insuline. Selon le modèle classique de la sécrétion d’insuline induite par le glucose, l’augmentation du ratio ATP/ADP résultant de la glycolyse et de l’oxydation du glucose, induit la fermeture des canaux KATP-dépendant modifiant ainsi le potentiel membranaire suivi d’un influx de Ca2+. Cet influx de Ca2+ permet l’exocytose des granules de sécrétion contenant l’insuline. Plusieurs nutriments comme les acides gras sont capables de potentialiser la sécrétion d’insuline. Cependant, le modèle classique ne permet pas d’expliquer cette potentialisation de la sécrétion d’insuline par les acides gras.
Pour expliquer l’effet potentialisateur des acides gras, notre laboratoire a proposé un modèle complémentaire où le malonyl-CoA dérivé du métabolisme anaplérotique du glucose inhibe la carnitine palmitoyltransférase-1, l’enzyme qui constitue l’étape limitante de l’oxydation des acides gras favorisant ainsi leur estérification et donc la formation de dérivés lipidiques signalétiques. Le modèle anaplérotique/lipidique de la sécrétion d'insuline induite par le glucose prédit que le malonyl-CoA dérivé du métabolisme du glucose inhibe la bêta-oxydation des acides gras et augmente la disponibilité des acyl-CoA ou des acides gras non-estérifiés. Les molécules lipidiques agissant comme facteurs de couplage du métabolisme des acides gras à l'exocytose d'insuline sont encore inconnus.
Des travaux réalisés par notre laboratoire ont démontré qu’en augmentant la répartition des acides gras vers la bêta-oxydation, la sécrétion d’insuline induite par le glucose était réduite suggérant qu’un des dérivés de l’estérification des acides gras est important pour la potentialisation sur la sécrétion d’insuline. En effet, à des concentrations élevées de glucose, les acides gras peuvent être estérifiés d’abord en acide lysophosphatidique (LPA), en acide phosphatidique (PA) et en diacylglycérol (DAG) et subséquemment en triglycérides (TG).
La présente étude a établi l’importance relative du processus d’estérification des acides gras dans la production de facteurs potentialisant la sécrétion d’insuline. Nous avions émis l’hypothèse que des molécules dérivées des processus d’estérification des acides gras (ex : l’acide lysophosphatidique (LPA) et le diacylglycerol (DAG)) agissent comme signaux métaboliques et sont responsables de la modulation de la sécrétion d’insuline en présence d’acides gras. Afin de vérifier celle-ci, nous avons modifié le niveau d’expression des enzymes clés contrôlant le processus d’estérification par des approches de biologie moléculaire afin de changer la répartition des acides gras dans la cellule bêta. L’expression des différents isoformes de la glycérol-3-phosphate acyltransférase (GPAT), qui catalyse la première étape d’estérification des acides gras a été augmenté et inhibé. Les effets de la modulation de l’expression des isoenzymes de GPAT sur les processus d’estérifications, sur la bêta-oxydation et sur la sécrétion d’insuline induite par le glucose ont été étudiés.
Les différentes approches que nous avons utilisées ont changé les niveaux de DAG et de TG sans toutefois altérer la sécrétion d’insuline induite par le glucose. Ainsi, les résultats de cette étude n’ont pas associé de rôle pour l’estérification de novo des acides gras dans leur potentialisation de la sécrétion d’insuline. Cependant, l’estérification des acides gras fait partie intégrante d’un cycle de TG/acides gras avec sa contrepartie lipolytique. D’ailleurs, des études parallèles à la mienne menées par des collègues du laboratoire ont démontré un rôle pour la lipolyse et un cycle TG/acides gras dans la potentialisation de la sécrétion d’insuline par les acides gras.
Parallèlement à nos études des mécanismes de la sécrétion d’insuline impliquant les acides gras, notre laboratoire s’intéresse aussi aux effets négatifs des acides gras sur la cellule bêta. La glucolipotoxicité, résultant d’une exposition chronique aux acides gras saturés en présence d’une concentration élevée de glucose, est d’un intérêt particulier vu la prépondérance de l’obésité. L’isoforme microsomal de GPAT a aussi utilisé comme outil moléculaire dans le contexte de la glucolipotoxicité afin d’étudier le rôle de la synthèse de novo de lipides complexes dans le contexte de décompensation où la fonction des cellules bêta diminue.
La surexpression de l’isoforme microsomal de la GPAT, menant à l’augmentation de l’estérification des acides gras et à une diminution de la bêta-oxydation, nous permet de conclure que cette modification métabolique est instrumentale dans la glucolipotoxicité.Diabetes is a chronic disease of glucose homeostasis characterized by hyperglycemia and the result of a failure of insulin secretion in combination or not with impaired insulin action. Overnutrition and lack of physical activity in individuals who have acquired or inherited genetic predispositions lead to insulin resistance. During the period of compensation where the concentration of plasma fatty acids is high, hyperinsulinemia fully compensates for the insulin resistance of target tissues and blood sugar is normal. Glucose promotes insulin secretion through its metabolism by the pancreatic β cell. According to the classical model of glucose-induced insulin secretion, the increase in the ATP/ADP ratio resulting from glycolysis and glucose oxidation induces the closure of KATP channels thus changing membrane potential followed by an influx of Ca2+. This influx of Ca2+ allows the exocytosis of secretory granules containing insulin. Several nutrients like fatty acids are capable of potentiating insulin secretion. However, the classical model does not explain the potentiation of insulin secretion by fatty acids. To explain the potentiating effect of fatty acids, our laboratory has proposed a complementary model in which malonyl-CoA derived from glucose anaplerotic metabolism inhibits carnitine palmitoyltransferase 1, the enzyme catalyzing the limiting step of fatty acid oxidation, thereby promoting their esterification and thus the formation signaling derivatives. The anaplerotic model of insulin secretion predicts that malonyl-CoA derived from glucose metabolism inhibits β-oxidation of fatty acids and increases the availability of acyl-CoA or non esterified fatty acids. Thus, lipid molecules can act as coupling factors for insulin exocytosis. Fatty acid-derived signalling molecules that are active remain to be identified. Work performed by our laboratory has shown that increasing the partition of fatty acids toward β-oxidation reduced glucose-induced insulin secretion, suggesting that derivatives of fatty acid esterification are important for the potentiation of insulin secretion. Indeed, at high concentrations of glucose, fatty acids are esterified into lysophosphatidic acid (LPA), phosphatidic acid (PA) and diacylglycerol (DAG) and subsequently in triglycerides (TG). The present study established the relative importance fatty acid esterification in the production of factors potentiating insulin secretion. We hypothesized that molecules derived from the process of esterification of fatty acid (eg lysophosphatidic acid (LPA) and diacylglycerol (DAG)) act as metabolic signals and are responsible for the modulation of the secretion of insulin in the presence of fatty acids. Thus, the level of expression of key enzymes controlling the process of esterification has been altered by molecular biology approaches to increase distribution of fatty acids toward esterification in the β cell. The expression of various isoforms of glycerol-3-phosphate acyltransferase (GPAT), which catalyzes the first step of esterification of fatty acids was increased and inhibited. The effects of GPAT isoenzyme modulation on the esterification process, on β-oxidation and on glucose-induced insulin secretion were investigated. The various approaches we used have changed the levels of DAG and TG without altering insulin secretion induced by glucose in the presence or absence of fatty acids. Thus, the results of this study do not suggest a role for de novo synthesis of glycerolipid intermidiates via esterification of fatty acids in the potentiation of insulin secretion. However, the esterification of fatty acids is an integral part of a TG/fatty acid cycle with its counterpart lipolysis. Moreover, parallel studies conducted by colleagues of the laboratory have demonstrated a role for lipolysis and a cycle TG/fatty acid in the potentiation of insulin secretion by fatty acids. In parallel with our studies of the mechanisms of insulin secretion involving fatty acids, our laboratory is also interested in the negative effects of fatty acids on the β cell. The glucolipotoxicity resulting from chronic exposure to saturated fatty acids in the presence of high glucose concentrations is of particular interest in the context of obesity rates. The microsomal isoform of GPAT was also used as a molecular tool under glucolipotoxicity conditions to study the role of de novo synthesis of complex lipids in the context of decompensation when β-cell function decreases. Increased esterification of fatty acids by the overexpression of microsomal isoform of GPAT has increased the toxic effects of fatty acids in the context of glucolipotoxicity. Thus, our results allow us to conclude that the distribution of lipids toward esterification and a decrease in β-oxidation is instrumental in glucolipotoxicity.
44
Narayanan, Niju. "Molecular and Genetic Strategies to Enhance Functional Expression of Recombinant Protein in Escherichia coli." Thesis, 2009. http://hdl.handle.net/10012/4721.
Анотація:
The versatile Escherichia coli facilitates protein expression with relative simplicity, high cell density on inexpensive substrates, well known genetics, variety of expression vectors, mutant strains, co-overexpression technology, extracytoplasmic secretion systems, and recombinant protein fusion partners. Although, the protocol is rather simple for soluble proteins, heterologous protein expression is frequently encountered by major technical limitations including inefficient translation, formation of insoluble inclusion bodies, lack of posttranslational modification mechanisms, degradation by host proteases, and impaired cell physiology due to host/protein toxicity, in achieving functional expression of stable, soluble, and bioactive protein.. In this thesis, model protein expression systems are used to address the technical issues for enhancing recombinant protein expression in E. coli.
When yellow fluorescence protein (YFP) was displayed on E. coli cell surface, the integrity of the cell envelope was compromised and cell physiology was severely impaired, resulting in poor display performance, which was restored by the coexpression of Skp, a periplasmic chaperone. On the basis of monitoring the promoter activities of degP, rpoH, and cpxP under various culture conditions, it was demonstrated that the cell-surface display induced the σE extracytoplasmic stress response, and PdegP::lacZ was proposed to be a suitable “sensor” for monitoring extracytoplasmic stress.
Intracellular proteolysis has been recognized as one of the key factors limiting recombinant protein production, particularly for eukaryotic proteins heterologously expressed in the prokaryotic expression systems of E. coli. Two amino acids, Leu149 and Val223, were identified as proteolytically sensitive when Pseudozyma antarctica lipase (PalB) was heterologously expressed in Escherichia coli. The functional expression was enhanced using the double mutant for cultivation. However, the recombinant protein production was still limited by PalB misfolding, which was resolved by DsbA coexpression. The study offers an alternative genetic strategy in molecular manipulation to enhance recombinant protein production in E. coli.
To overcome the technical limitations of protein misfolding, ineffective disulfide bond formation, and protein instability associated with intracellular proteolysis in the functional expression of recombinant Pseudozyma antarctica lipase B (PalB) in Escherichia coli, an alternative approach was explored by extracellular secretion of PalB via two Sec-independent secretion systems, i.e. the α-hemolysin (Type I) and the modified flagellar (Type III) secretion systems, which can export proteins of interest from the cytoplasm directly to the exterior of the cell. Bioactive PalB was expressed and secreted extracellularly either as HlyA fusion (i.e. PalB-HlyA via Type I system) or an intact protein (via Type III system) with minimum impact on cell physiology. However, the secretion intermediates in the intracellular fraction of culture samples were non-bioactive even though they were soluble, suggesting that the extracellular secretion did mediate the development of PalB activity. PalB secretion via Type I system was fast with higher specific PalB activities but poor cell growth. On the other hand, the secretion via Type III system was slow with lower specific PalB activities but effective cell growth.
Functional expression of lipase from Burkholderia sp. C20 (Lip) in various cellular compartments of Escherichia coli was explored. The poor expression in the cytoplasm was improved by several strategies, including coexpression of the cytoplasmic chaperone GroEL/ES, using a mutant E. coli host strain with an oxidative cytoplasm, and protein fusion technology. Fusing Lip with the N-terminal peptide tags of T7PK, DsbA, and DsbC was effective in boosting the solubility and biological activity. Non-fused Lip or Lip fusions heterologously expressed in the periplasm formed insoluble aggregates with a minimum activity. Biologically active and intact Lip was obtained upon the secretion into the extracellular medium using the native signal peptide and the expression performance was further improved by coexpression of the periplasmic chaperon Skp. The extracellular expression was even more effective when Lip was secreted as a Lip-HlyA fusion via the α-hemolysin transporter. Finally, Lip could be functionally displayed on the E. coli cell surface when fused with the carrier EstA.
45
Décary, Simon. "Amélioration de la fonction pancréatique par l'activité physique chez le rat diabétique de type 2." Thèse, 2008. http://hdl.handle.net/1866/7740.