Dissertations / Theses: 'Cel Hypoxia'

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Ljungkvist, Anna. "Imaging the tumor microenvironment : the dynamics and modification of hypoxia." Doctoral thesis, Umeå : Univ, 2003. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-106.

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Lawrentschuk, Nathan Leo. "Hypoxia and angiogenesis in renal cell carcinoma." Connect to thesis, 2009. http://repository.unimelb.edu.au/10187/6790.

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Hypoxia is one of the hallmarks of cancer. It was first postulated to occur in solid tumours by Thomlinson and Gray in 1955.1 The presence of hypoxia has been demonstrated in different types of solid tumours.2 Intratumoral hypoxia is caused by the lack of functional blood vessels in proliferating tumour tissue, resulting in low intratumoral oxygen concentrations. If hypoxia is severe or prolonged, cell death occurs.3 Malignant cells can undergo genetic and adaptive changes that allow them to escape from dying of oxygen deprivation. These changes are associated with a more aggressive malignant phenotype 4,5 conferring resistance to radiation 6,7 and chemotherapeutic agents.3,8,9 Hence hypoxia is known to be a key factor responsible for tumour resistance in humans.
Invasive polarographic oxygen sensor measurements have demonstrated hypoxia in solid tumours and it is generally defined to occur at an oxygen tension less than ten mmHg.10 Perhaps of more importance is that hypoxia has been demonstrated to be a prognostic indicator for local control after treatment with radiotherapy in glioma, head and neck and cervical cancers.11-13 It has also been able to predict for survival and the presence of distant metastases in soft tissue sarcomas.14 Finally, the significance of hypoxia in the activation and induction of functional molecules such as hypoxia inducible factors (HIFs) and VEGF, the modulation of gene expression (e.g. carbonic anhydrase IX), increased proto-oncogene levels, activation of nuclear factors and accumulation of other proteins (e.g. TP53) although progressing, is yet to be defined.15,16
Thus, it is of clinical interest to understand the levels of hypoxia and numbers of hypoxic cell populations in tumours, particularly those resistant to radiation and chemotherapy. In doing so clinicians and researchers may formulate more accurate prognostic information and develop treatments targeting hypoxic cells. Renal cell carcinoma (RCC) is a tumour resistant to radiation and chemotherapy that is yet to have its oxygen status investigated.
Although the “gold standard” of oxygen tension measurement is the Polarographic Oxygen Sensor (POS or Eppendorf pO2 histograph), non-invasive means of measuring oxygen status via imaging, immunohistochemistry or serum tumour markers are more practical. As highlighted by Menon and Fraker, it is imperative that reliable, globally usable, and technically simplistic methods be developed to yield a consistent, comprehensive, and reliable profile of tumour oxygenation. Until newer more reliable techniques are developed, existing independent techniques or appropriate combinations of techniques should be optimized and validated using known endpoints in tumour oxygenation status and/or treatment outcomes.17
Hanahan and Weinberg 18 surmised that the field of cancer research has largely been guided by a reductionist focus on cancer cells and the genes within them- a focus that has produced an extraordinary body of knowledge. Looking forward in time, they believe that progress in cancer research would come from regarding tumours as complex tissues in which mutant cancer cells have conscripted and subverted normal cell types (endothelial cells, immune cells, fibroblasts) to serve as active collaborators in their neoplastic agenda. The interactions between the genetically altered malignant cells and these supporting coconspirators will prove critical to understanding cancer pathogenesis and to the development of novel, effective therapies.18
Essentially, the background outlined here not only highlights the core aim of this thesis: to better understand the oxygen status of renal cell carcinoma and the relationship of this to angiogenesis so that better targeted therapies may be pursued in the future; but it also places this research in the context of the future proposed by Hanahan and Weinberg,18 by clearly focusing on collaborators in the neoplastic agenda, rather than just tumour cells themselves, to better understand RCC.

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Lester, Robin D. "Hypoxia activated cell signaling receptors in cancer." Diss., Connect to a 24 p. preview or request complete full text in PDF format. Access restricted to UC campuses, 2008. http://wwwlib.umi.com/cr/ucsd/fullcit?p3297526.

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Thesis (Ph. D.)--University of California, San Diego, 2008.
Title from first page of PDF file (viewed April 28, 2008). Available via ProQuest Digital Dissertations. Vita. Includes bibliographical references (p. 114-134).

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Schioppa, Tiziana. "Effects of tumour hypoxia on cell migration." Thesis, Open University, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.434200.

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Cell adaptation to hypoxia requires activation of transcriptional programs that coordinate expression of genes involved in oxygen delivery (via- angiogenesis) and metabolic adaptation (via glycolisis). During migration and invasion of normal and pathological tissues, cells may encounter different oxygen levels, due to poor or altered vascularization, and recent evidence has suggested that chemotaxis is a cell function which may be affected by oxygen availability. This thesis describes how oxygen avaibility is a determinant parameter in the setting of chemotactic responsiveness to Stromal-Derived Factor 1 (SDF-1, CXCL12). Low oxygen concentration induces high expression of the CXCL12 receptor CXCR4, in different cell types (monocytes, monocyte-derived macrophages, tumor associated macrophages, endothelial cells, cancer cells and dendritic cells) as both mRNA and protein expression, which is paralleled by increased chemotactic responsiveness to its specific ligand. Furthermore, preliminary results on dendritic cells (DC) show that hypoxia may affect their maturation (CCR7'°'/CCR5h'gh) and functions. In particular, hypoxia-derived DC do not migrate in response to the CCR7 ligand CCL 19, while they do express higher levels of pro-inflammatory cytokines (IL-12, TNF-a), as compared to normoxia-derived DC. CXCR4 induction by hypoxia is dependent on both activation of hypoxia-inducible factor 1 (HIF-la) and transcript stabilization. Our data identify the hypoxia/HIF-1/CXCR4 pathway as a relevant molecular circuit in the functional tuning of the chemokine system and provide novel insights into the mechanisms controlling cell migration in hypoxic regions, with potential relevance in the pathogenesis of human diseases, including chronic inflammatory diseases and cancer

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Ledaki, Ioanna I. "Heterogeneity of tumour response to hypoxia : carbonic anhydrase IX induction defines a subpopulation of hypoxic cells with stem cell properties and drug resistance." Thesis, University of Oxford, 2013. http://ora.ox.ac.uk/objects/uuid:757a8e79-b20d-496c-b69b-4d6a3b7b56e3.

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Carbonic anhydrase IX (CA9) is strongly induced by hypoxia and its overexpression is associated with poor therapeutic outcome in cancer. The function of CAIX is to catalyze the reversible hydration of CO2 to bicarbonate and a proton. This helps hypoxic tumours to maintain a more neutral intracellular pH (pH_i) promoting survival, but produces a more acidic extracellular (pH_e) which promotes invasion and metastasis. Recent evidence has expanded on the role of hypoxia and CAIX by relating them to stem cell niches. In this study, taking advantage of the transmembrane location of CAIX, we show for the first time, a direct marked heterogeneity in response to hypoxia within each tumour cell population studied, associated with major biological differences. Based on CAIX expression pattern under hypoxic conditions, we identify, isolate and characterize two distinct populations of tumour cells, one that express CAIX and the other that does not. Interestingly, we discover that the CAIX positive population is enriched with cells expressing cancer stem cell markers. These include ALDHA1, IGF1, LIN28 and genes involved in epithelial-mesenchymal transition (EMT) and multi-drug resistance (i.e. WNT2, TWIST1, and ABCC2). Accordingly, CAIX+ve cells show higher self-renewal capacity and form tumours significantly faster compared to the CAIX-ve population. Importantly, functional suppression of CAIX in vitro and in vivo, in two breast cancer cell lines resulted in the downregulation of breast cancer stem cell signatures, suggesting that CAIX is not just a marker of stemness but also a regulator of stemness. The molecular mechanism underlying the differential expression of CAIX in the two populations is not HIF-1α-dependent, but instead driven by hypoxia-induced reorganization of chromatin structure. In line with this, we provide experimental evidence showing that the genomic locus encoding CA9 has a more “open” and transcriptionally active chromatin structure in CAIX+ve cells, and a condense and transcriptionally silent chromatin structure in the CAIX-ve cells. Given that HIF induces the transcription of CA9 by binding to hypoxia response elements (HREs) in its promoter we show a significant reduction in binding of HIF to the CA9 promoter of the negative population. We suggest that the reduce HIF binding is a result of the compact chromatin structure of CA9 promoter of the negative cells. Analysis of the transcriptome of the positive and negative populations suggests a symbiotic relationship between these two subpopulations and their environment, likely required to promote tumour growth. This is based on the following observations: Firstly, we identified that CAIX-ve cells express high levels of cytokines and based on this, we suggest that the cytokines secreted by CAIX-ve cells may transmit paracrine signals that regulate the CAIX+ve cells, thus providing a wider hypoxia tolerant microenvironment to protect the stem cell population. Secondly, we identified a metabolic heterogeneity between the CAIX+ve and CAIX-ve cells. The CAIX+ve cells show an upregulation of genes implicated in oxidative phosphorylation, TCA cycle and fatty acid synthesis. Whereas in CAIX-ve cells there is an upregulation of genes implicated in autophagy and mitophagy. Given the above together with the upregulation of oxidative phosphorylation and TCA cycle in the CAIX+ve cells, we proposed the existence of a metabolic symbiosis between the CAIX+ve and CAIX-ve cells. We postulate that the catabolic process such as autophagy and mitophagy in the CAIX-ve cells may results in the overproduction of high-energy metabolites such as lactate, glutamine and ketone bodies which in turns they are been utilized by CAIX+ve cells to fuel mitochondria respiration. Finally, we also demonstrated that in the CAIX+ve cells mTORC1 signaling is upregulated, and contributes to the regulation of CAIX expression. Given the role of mTORC1 in stem cell maintenance and EMT as well as the interdependence of mTORC1 and CAIX expression in the CAIX+ve cells we suggest that mTORC1 signaling may be the critical factor by which CAIX regulates stemness. Interestingly, the subpopulations show a differential sensitivity to HDAC inhibitors, NaBu and SAHA as treatment of MCF7 breast cancer cell line and HCT116 colon cancer cell line leads to elimination of the CAIX+ve population. This is not driven by the downregulation of HIF-1α, the major transcriptional regulator of CAIX. In contrast, we demonstrate that SAHA causes downregulation mTORC1. This suggests that SAHA-induced downregulation of CAIX expression could be due to its effect on mTORC1 pathway. Of wider significance, our findings show that tumours are not homogenous in their response to hypoxia, and distinct signal transduction networks regulate different populations of cells within the tumour. This highlights the need for the utilization of biomarkers, which reveal distinct functional hypoxia profiles of human cancers, and permit the stratification of tumours. Furthermore, the identification of epigenetic regulation of the histones in response to hypoxia for highly selective gene regulation, provides a connection between the epigenetic mechanisms under environmental stress and cancer progression, and is model for development of novel epigenetic cancer therapeutic drugs.

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Nilsson, Ingrid. "Hypoxia, PDGF and VEGF in Vascular Development." Doctoral thesis, Uppsala : Acta Universitatis Upsaliensis, 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-6894.

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Bedessem, Baptiste. "Contributions à l'étude de la réponse moléculaire à l'hypoxie : Modélisation mathématique et expérimentations sur cellules FUCCI." Thesis, Université Grenoble Alpes (ComUE), 2015. http://www.theses.fr/2015GREAS024/document.

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Les effets biologiques de l'hypoxie sont très étudiés aujourd'hui, principalement en raison du rôle crucial que jouent les conditions d'oxygénation dans le développement des cancers.Depuis plusieurs années, une littérature foisonnante tente ainsi de décrire les multiples aspects de la réponse moléculaire, cellulaire et physiologique à l'hypoxie. La complexité des voies de signalisation impliquées et la diversité de leurs effets cellulaires rendent la tâche délicate. Cet état de fait se reflète dans la pluralité des méthodes utilisées, depuis les simulations numériques jusqu'aux approches expérimentales.Dans cette thèse, j'ai abordé ce sujet sur la base de deux outils: la modélisation mathématique et une démarche expérimentale utilisant les cellules HeLa-FUCCI. Cette lignée cellulaire récemment développée est en effet un instrument de choix encore peu exploité. Par une construction génétique liant des protéines du cycle cellulaire à un fluorophore, elle rend possible l'étude, en continue, de la dynamique du cycle en microscopie de fluorescence. Nous avons ainsi pu analyser plusieurs aspects de la réponse cellulaire à l'hypoxie, dans un contexte tumoral.Dans un premier temps, nous avons cherché à caractériser mathématiquement les liens tissés entre le cycle cellulaire et les voies de signalisation de l'hypoxie, centrées sur le facteur de transcription HiF-1. Ce modèle propose un explication simple à l'arrêt du cycle observé notamment dans les cellules tumorales en conditions hypoxiques. Nous avons ainsi montré que l'induction de chimiorésistance pouvait se concevoir comme une entrée facilitée en quiescence des cellules cancéreuses. Dans le but de valider ces observations, nous avons ensuite cherché à quantifier expérimentalement la dynamique de la prolifération cellulaire en utilisant les cellules HeLa-FUCCI. Comme il est apparu que les fluorophores qu'elles portent sont sensibles au manque d'oxygène, nous avons testé différentes molécules couramment utilisées pour induire HiF-1 et mimer l'hypoxie (DFO et CoCl2). De cette étude ont émergé des résultats originaux quant à la dynamique de blocage du cycle des cellules HeLa en présence de chélateurs du fer.Si les conditions hypoxiques ne sont pas favorables à l'utilisation des cellules FUCCI, nous avons pu en revanche montrer qu'elles étaient tout à fait adaptées à l'étude de la dynamique du cycle cellulaire en condition de réoxygénation. De manière intéressante, nous avons alors pu observer un ralentissement significatif de la phase S après retour à la normoxie. Afin d'apporter un éclairage théorique à cette observation, nous avons proposé un modèle mathématique de la dynamique de régulation de HiF-1 en conditions d'oxygène fluctuantes, basé sur le couple HiF-1/pVHL, dont les relations sont pensées dans un cadre compartimenté (noyau/cytoplasme). Ce modèle simple reproduit fidèlement les caractéristiques principales de la réponse cellulaire à l'hypoxie. En outre, en simulant les conséquences d'une réoxygénation brutale, nous avons observé la genèse de fortes instabilités du niveau intracellulaire de HiF-1. Enfin, nous avons mené une étude expérimentale de la compartimentation de HiF-1. L'outil FUCCI permet en effet d'observer simultanément l'avancement du cycle (en microscopie de fluorescence), et la localisation intra-cellulaire de HiF-1(par immunomarquage). Nous avons pu montrer que la variabilité de la localisation de HiF-1α n'était pas due à la progression dans le cycle. Elle est donc certainement liée soit à des différences génétiques inter-cellulaire, soit à une stochasticité de la régulation de HiF-1
The biological effects of hypoxia are intensively studied today, mainly because of the crucial role played by oxygenation conditions during the development of cancers.For several years, a huge literature aims at describing the multiple aspects of the molecular, cellular and physiological responses to hypoxia. The complexity of the pathways which are involved and the diversity of their cellular effects make this task difficult.This situation is reflected in the plurality of the methods used, from the numerical simulations to the experimental approaches.In this thesis, I studied this subject using two tools: mathematical modeling and experimental approaches using HeLa-FUCCI cells.This recently developed cell line is an interesting tool not yetmuch exploited. By a genetic construction linking cell cycle proteins to a fluorophore, it makes possible the study of cell cycle dynamics using fluorescent microscopy.We could analyze various aspects of the cellular response to hypoxia, in a tumoral context. In a first time,we tried to mathematically characterize the links existing between cell cycle and the hypoxia pathways,driven by HiF-1.This model proposed a simple explanation to the cell cycle arrest notably observed in the tumor cells in hypoxicconditions.We then showed that the induction of chemoresistances could be considered as an entry into quiescence of tumor cells.In order to validate these observations we then tried to experimentally quantify the dynamics of cell proliferation using HeLa-FUCCI cells. As it appeared that the fluorophores were sensitive tothe lack of oxygen, we tested different molecules currently used to induceHiF-1 and mimic hypoxia (DFO and COCl2).From this study have emerged original results about the dynamics of cell cyclearrest of HeLa cells in presence of iron-chelators.If hypoxic conditions are not favorable to the use of HeLa-FUCCI cells, we could show that they were totally adapted to the study of cell cycle dynamics during reoxygenation.Interestingly, we then could observe a significant slowing down of the S-phase after the return to normoxia. In order to bring theoretical elements to this observation, we proposed a mathematical model of the dynamics of HiF-1 regulation in fluctuating oxygen conditions, based on thepVHL/HiF-1 couple, in the frame of a nucleo-cytoplasmic compartmentalization of HiF-1.This simple model well reproduce the main characteristics of the cell response to hypoxia.Besides, by simulating the consequences of a sudden reoxygenation, we observed the genesis of strong instabilities of HiF-1 intracellular level.Finally, we propose an experimental study of HiF-1 compartmentalization.Indeed, the FUCCI cells allow to simultaneously observe cell cycle progression (using fluorescent microscopy),and HiF-1 intra-cellular localization (with immunomarkage). We then could show that the variability of HiF-1 localization was not due to the progression into the cell cycle. Then, it is certainly linked to inter-cellular genetic differences, or to a stochasticity of HiF-1 regulation

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Lidgren, Anders. "Hypoxia inducible factor-1α in renal cell carcinoma." Doctoral thesis, Umeå universitet, Kirurgisk och perioperativ vetenskap, 2007. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-1462.

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Hypoxia Inducible Factor-1α in Renal Cell Carcinoma Departments of Surgical and Perioperative Sciences, Urology and Andrology; Radiation Sciences, Oncology; Medical Biosciences, Pathology; and Medical Biosciences, Clinical Chemistry, Umeå University, Umeå, Sweden Background: Renal cell carcinoma (RCC) accounts for approximately 2-3% of all human cancers. A distinguished feature of RCC is vascularisation and among the three dominating RCC types conventional RCC (cRCC) generally is more vascularised than papillary RCC (pRCC) and chromophobe RCC (chRCC). Angiogenesis is a critical step in tumour progression controlled by a balance involving molecules that have positive and negative regulatory activity. A balance distorted by metabolic stress such as hypoxia, acidosis, and inflammation. Hypoxia-Inducible Factor 1α (HIF-1α) is a key transcription factor in angiogenesis and tumour progression, targeting more than a 100 genes involved in vascular growth and regulation, iron metabolism and erythropoesis, collagen matrix formation, regulation of extracellular pH, glucose uptake and metabolism, proliferation, apoptosis, differentiation, and cell viability. Methods: Tumour tissue and corresponding kidney cortex from nephrectomised RCC patients was used in order to characterize HIF-1α expression and one of its target genes, Glucose Transporter 1 (GLUT-1). All tumour samples were thoroughly described regarding tumour type, TNM stage, nuclear grade, tumour size, vein invasion, and patient survival. Utilizing RT-PCR, Westen Blot and Tissue micro array (TMA) we studied HIF-1α mRNA and protein expression as well as GLUT-1 protein expression, correlating them to each other and clinicopathological parameters. Results: Using Western Blot, HIF-1α protein expression differed significantly between the different RCC types and kidney cortex. In cRCC, high expression of HIF-1α was an independent prognostic factor for favourable prognosis. TMA is a useful method to analyze HIF-1α protein expression in RCC. HIF-1α levels were significantly lower in locally aggressive cRCC and patients with high levels of HIF-1 tended to have a better prognosis. GLUT-1 levels were higher in cRCC than in other RCC types and for cRCC a correlation to HIF-1α was seen. HIF-1α mRNA levels were significantly lower in cRCC compared to other RCC types and kidney cortex. An inverse correlation between HIF-1α protein expression and mRNA levels was observed. Summary: These results demonstrate a discrepancy between RCC types, highlighting the need to separately evaluate biological events in different RCC types. Overexpression of HIF-1α protein is not necessarily all bad and translational regulation appears more critical than anticipated. Further studies are encouraged to clarify angiogenic pathways in RCC.

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Del, Sole Marianna <1981&gt. "Effect of hypoxia and hyperglycemia on cell bioenergetics." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2012. http://amsdottorato.unibo.it/4732/.

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Mitochondria have a central role in energy supply in cells, ROS production and apoptosis and have been implicated in several human disease and mitochondrial dysfunctions in hypoxia have been related with disorders like Type II Diabetes, Alzheimer Disease, inflammation, cancer and ischemia/reperfusion in heart. When oxygen availability becomes limiting in cells, mitochondrial functions are modulated to allow biologic adaptation. Cells exposed to a reduced oxygen concentration readily respond by adaptive mechanisms to maintain the physiological ATP/ADP ratio, essential for their functions and survival. In the beginning, the AMP-activated protein kinase (AMPK) pathway is activated, but the responsiveness to prolonged hypoxia requires the stimulation of hypoxia-inducible factors (HIFs). In this work we report a study of the mitochondrial bioenergetics of primary cells exposed to a prolonged hypoxic period . To shine light on this issue we examined the bioenergetics of fibroblast mitochondria cultured in hypoxic atmospheres (1% O2) for 72 hours. Here we report on the mitochondrial organization in cells and on their contribution to the cellular energy state. Our results indicate that prolonged hypoxia cause a significant reduction of mitochondrial mass and of the quantity of the oxidative phosphorylation complexes. Hypoxia is also responsible to damage mitochondrial complexes as shown after normalization versus citrate synthase activity. HIF-1α plays a pivotal role in wound healing, and its expression in the multistage process of normal wound healing has been well characterized, it is necessary for cell motility, expression of angiogenic growth factor and recruitment of endothelial progenitor cells. We studied hypoxia in the pathological status of diabetes and complications of diabetes and we evaluated the combined effect of hyperglycemia and hypoxia on human dermal ﬁbroblasts (HDFs) and human dermal micro-vascular endothelial cells (HDMECs) that were grown in high glucose, low glucose concentrations and mannitol as control for the osmotic challenge.
I mitocondri hanno un ruolo fondamentale nella produzione di energia nella cellula, ma sono coinvolti anche in altri processi tra cui la produzione di ROS e l’apoptosi. Disfunzioni del metabolismo mitocondriale sono state associate a diversi disordini, tra cui: diabete di tipo II, malattia si Alzheimer, infiammazione, cancro ed ischemia cardiaca. Quando i livelli di ossigeno nella cellula diventano limitanti, la funzione mitocondriale viene modulata per consentire l’adattamento biologico. La via dell’AMP- activated protein kinase (AMPK) ha il compito di monitorare lo stato energetico della cellula mantenendo i livelli fisioligici di ATP/ADP. In seguito all’esposizione prolungata in ambiente ipossico, l’attivazione di HIF-1 e’ in grado di upregolare diversi geni coinvolti nella sopravvivenza cellulare a basse concentrazioni di ossigeno. In questo lavoro, e’ stata valutata la bioenergetica mitocondriale in fibroblasti primari coltivati a basse concentrazioni di ossigeno (1 % O2) per 72 ore; in particolare, abbiamo preso in considerazione l’organizzazione mitocondriale nella cellula e il loro contributo nel mantenere lo stato energetico cellulare. I nostri risultati indicano che l’esposizione prolungata all’ipossia causa una significativa riduzione della massa mitocondriale e della quantita’ dei complessi della fosforilazione ossidativa, nonostante le cellule siano in grado di mantenere i livelli intracellulari di ATP. Inoltre abbiamo studiato l’ipossia nel contesto patologico del diabete ed in particolare delle complicanze del diabete. E’ noto che l’iperglicemia e l’ipossia, dovuta ad ischemia a danni vascolari, hanno un ruolo importante nell’insorgenza delle complicanze del diabete. HIF-1α rappresenta uno stimolo nella rigenerazione delle ferite, in quanto stimola la vascolarizzazione e la migrazione dei cheranociti ed e’ stato ipotizzato che le cellule perdano la capacita’ di adattarsi e rispondere all’ipossia quando sono coltivate in presenza di elevate concentrazioni di glucosio (>25 mM). Abbiamo valutato il ruolo della destabilizzazione di HIF-1α nella produzione di ROS, considerati i principali responsabili della progressione del diabete.

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Milani, Manuela. "Cell stress response and hypoxia in breast cancer." Thesis, University of Oxford, 2011. http://ora.ox.ac.uk/objects/uuid:74d3bf91-9888-4e9e-b5e1-7d5d2d476174.

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During severe hypoxia (<0.01% oxygen) the protein folding machinery becomes dysfunctional, resulting in the accumulation of unfolded proteins with consequent endoplasmic reticulum (ER) stress and activation of the unfolded protein response (UPR) and autophagy, a process involved in the physiological turnover of cytoplasmic components. The link between the UPR and autophagy is not clearly defined. The aim of this thesis is to investigate the role of the induction of UPR under severe hypoxia in tumour survival and resistance to therapy. The results of this research suggest that the activating transcription factor 4 (ATF4), a component of the PKR-like ER kinase (PERK) pathway, fundamental in the UPR, is required for the ER-stress induced upregulation of autophagy. Mechanisms other than hypoxia for UPR induction were investigated, using the proteasome inhibitor bortezomib (BZ). BZ treatment increased ATF4 protein levels in MCF7 cells, even transfected with short-interference RNA (siRNA) against the classical UPR activator PERK, suggesting that the proteasomal stabilization is likely the main mechanism for ATF4 protein accumulation. The induction of autophagy by BZ is dependent upon the upregulation of the microtubule-associated protein 1 light chain 3B (LC3B), an autophagy marker, by ATF4 and acts as a survival mechanism. Hypoxia, UPR and autophagy markers (such as Pimonidazole, carbonic anhydrases IX (CAIX), C/EBP homologous protein (CHOP) and LC3B) were evaluated by immunohistochemical approach in spheroids, xenografts models and breast cancer samples. CHOP immunohistochemical staining was performed in breast cancer sections from a series of patients. CHOP was expressed in cells surrounding necrotic areas. No correlation were found with clinical outcome and further studies are needed.

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Ibegbu, Augustine. "The effects of hypoxia on neuronal cell signalling." Thesis, Queen Margaret University, 2009. https://eresearch.qmu.ac.uk/handle/20.500.12289/7367.

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Hypoxia adversely affects cells and tissues, and neuronal cells in particular have been shown to be more susceptible to the injurious effects of hypoxia i.e. they may begin to die when oxygen supply is reduced or completely eliminated. Cannabinoid (CB1) receptor and opioid (μ, δ and κ) receptor agonists have been shown to elicit several central nervous system (CNS) effects, mediated via G protein-coupled receptors. The aim of the research presented in this thesis was to study the effect of hypoxia on neuronal cell signalling and the consequent neuroprotectant effects of cannabinoid and opioid receptor agonists against hypoxia in the rat cortical neuronal cell line (B50) in culture. The B50 cells cultured in hypoxic conditions were treated and concurrently cultured with cannabinoid and opioid receptor agonists to determine the effects of these drugs on hypoxia-induced changes using downstream signalling activities such as cellular morphogenesis, growth, proliferation, differentiation, lactate dehydrogenase (LDH) leakage, second messenger (cAMP) and extracellular signalregulated kinases (ERK1/2) quantification, to assess the level of cellular damage and injury, repair and protection. Cortical B50 cells were cultured in either a normal incubator (21%O2; 5% CO2) as the normoxic control group, or a hypoxic incubator (5%O2; 5% CO2) as the experimental group. Three cannabinoid agonists [Win55,212- 2 mesylate (Win), anandamide or arachidonoylethanolamide (AEA), and 2- arachidonylglycerol (2-AG)] and three opioid agonists [DAMGO (μ), DSLET (δ) and ICI-199,441 hydrochloride (κ)], were selected and administered to the cells as treatment group for 48 hours after 48 hours of initial culture for a total of 96 hours of culture and pre-treatment group treated at 0 hour for a total of 96 hours in hypoxic conditions at concentrations of 10nM, 50nM and 100nM for cannabinoid agonists, and 10μM, 50μM and 100μM for opioid agonists. Neuronal viability, proliferation, differentiation and second messenger activity were assessed using morphological same-field assessment, LDH leakage, cellular proliferation assay, second messenger (cAMP) assay, and phospho-ERK1 & 2 assay and dibutyryl cyclic adenosine monophosphate (DbcAMP) induced differentiation method. Levels of G-protein coupled receptor (cannabinoid, CB1 and mu opioid, MOR) mRNAs were assessed using the RT-PCR method. The results showed that hypoxia induced a 4-fold increase in LDH leakage from B50 cells cultured in hypoxia when compared to the cells xxviii cultured in normoxic conditions (440% versus 100%, respectively; p<0.05). Cannabinoid receptor agonist treatment was able to reduce the LDH release in hypoxic cells to between 2-to 4-folds: 100nM AEA (69%), 100nM 2-AG (103%) and 10nM Win (217%), when compared to untreated hypoxic B50 cells (440% versus cannabinoid treated; p<0.05). The results of opioid administration showed a 3-fold decrease in the level of LDH leakage in B50 cells cultured in hypoxia when compared to untreated hypoxic cells (587%). The results of hypoxic treated B50 cells with opioid agonists are 100μM ICI-199,441 (318%); 50μM DSLET (339%) and 50μM DAMGO (352%) (p<0.05; untreated hypoxia versus opioid treated). The result of cAMP quantification in B50 cells in culture showed a reduction in cAMP concentration in untreated hypoxic B50 cells when compared to normoxic cells (0.7 pmol/ml versus 3.0 pmol/ml; p<0.05). Cannabinoid treated hypoxic cells showed increases in cAMP concentration: 2-AG 10nM (3.5 pmol/ml), 50nM (3.1 pmol/ml) and 100nM (0.9 pmol/ml), (p<0.05; Cannabinoid treated versus hypoxia untreated). The cAMP concentration in B50 cells treated in hypoxia with opioid agonist, ICI 199,441 hydrochloride, was significantly increased when compared to untreated hypoxic B50 cells (0.7 pmol/ml). The treatment with ICI 199,441 hydrochloride are 10μM (10.0 pmol/ml), 50μM (3.15 pmol/ml) and 100μM (1.15 pmol/ml), (p<0.05; opioid treated versus hypoxia untreated). The result of phospho-ERK1&2 assay in B50 cells showed decrease in phospho-ERK1&2 in untreated hypoxic cells when compared to normoxic untreated cells (6.0 units/ml versus 87.0 units/ml; p<0.05). The result of cannabinoid treated hypoxic cells showed increases in phospho-ERK1&2 when compared with the hypoxic untreated B50 cells: Win 10nM (98 units/ml), Win 100nM (27 units/ml), AEA 10nM (62 units/ml), AEA 100nM (60.5 units/ml), 2-AG 10nM (45 units/ml) and 2-AG 100nM (68 units/ml) (cannabinoid treated versus untreated hypoxia; p<0.05). The phospho-ERK1&2 in hypoxic B50 cells treated with opioid showed increase with DAMGO 10μM (22 units/ml), DSLET 10μM (16 units/ml) and ICI 199,441 hydrochloride 10μM (23.5 units/ml) (P<0.05; opioid treated versus hypoxia untreated). The result showed a decrease in cellular proliferation in untreated hypoxic cells when compared to the normoxic cells (7x106 cells/ml versus 20x106 cells/ml; p<0.05), while cannabinoid and opioid treatments was able to increase cell proliferation in hypoxic treated cells with: Win 10nM (11x106 cells/ml), AEA 100nM (12x106 cells/ml) and 2-AG 100nM (13.8x106 cells/ml), DAMGO 10μM (16x106 cells/ml), DSLET 10μM (20x106 cells/ml) and ICI xxix 199,441 100μM (21.5x106 cells/ml) when compared to hypoxic untreated cells (7x106 cells/ml) (hypoxia untreated versus hypoxia treated; p<0.05). Some of these changes were shown to be concentration-dependent between the normal and hypoxic B50 neurons, and between treated and untreated hypoxic B50 cells in culture, while the CB1 and MOR mRNA levels showed no appreciable change. The results show that B50 neuronal cells are susceptible to damage and injurious effects of hypoxia, as are most brain cells, while the results of the administration of cannabinoid and opioid agonists suggest that these agents have some potential therapeutic and protective benefits in the treatment and prevention of hypoxia-induced toxicity in neuronal B50 cells in culture. This could be of potential benefit in the treatment and protection against hypoxia-related neurodegenerative diseases and disorders such as stroke, dementias, ageing, Alzheimer’s and Parkinson’s diseases.

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Webster, Lynne. "Hypoxia and proliferation in murine tumour models." Thesis, University College London (University of London), 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.336415.

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13

Englund, Marita. "Effects of hypoxia and antiepileptic drugs on electrophysiological properties of CA1 neurons in hippocampus /." Stockholm, 2007. http://diss.kib.ki.se/2007/978-91-7357-237-8/.

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Wilcock, Paul. "A systems biology approach for investigating oral squamous cell carcinoma (OSCC)." Thesis, University of Manchester, 2013. https://www.research.manchester.ac.uk/portal/en/theses/a-systems-biology-approach-for-investigating-oral-squamous-cell-carcinoma-oscc(8ec3728b-1928-450f-b467-76996fa970fb).html.

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A systems biology approach was adopted in order to assess various aspects of the disease oral squamous cell carcinoma. Three main aims were addressed; assess the ability of CoCl2 to mimic the hypoxic response in a eukaryotic cell line, assess the role of PDE4D in oral squamous cell carcinoma (OSCC) and the construction of a normoxic/hypoxic mathematical model to identify therapeutic targets.Cancer cells often acquire a revised metabolism which aids in initiation, survival and progression of the tumour. This is predominantly due to the transcription factor HIF-1 which is activated under hypoxic conditions. Certain compounds such as cobalt chloride (CoCl2) have been used extensively to inhibit the degradation of HIF-1α and simulate hypoxia. CoCl2 is likely to have off-target effects on metabolism; these effects were examined when exposing human telomerase reverse transcriptase (hTERT) cells to 100μM CoCl2. Gas chromatography-mass spectrometry (GC-MS), liquid chromatography-mass spectrometry (LC-MS) based metabolomics were utilised in combination with ELISA assays for HIF-1α and ATP. Central metabolism was accurately mimicked when hTERT cells were subjected to 100μM CoCl2, however; it was apparent that this concentration of CoCl2 does not induce an equal extent of hypoxia as 1% oxygen. A number of off-target effects of CoCl2 were observed in secondary metabolism, specifically in lipids and fatty acids. In conclusion, CoCl2 should be used with caution as a hypoxic mimicker with the caveat that interpretation of results should be restricted to its effects on central metabolism.The transcription factor CREB has the ability to regulate approximately 4000 genes, a number of which are associated with cancer initiation and progression. Cyclic adenosine monophosphate (cAMP) is required to activate CREB and is partially regulated through its degradation via the enzyme phosphodiesterase type 4D (PDE4D). A homozygous deletion of PDE4D has been associated with OSCC; however; the exact consequence of this deletion has not been fully elucidated. PDE4D was knocked down in the OSCC cell line BicR16 and cellular proliferation, migration, resistance to ionising radiation and central metabolism was investigated using MTT, scratch, clonogenic and GC-MS, respectively. The knockdown resulted in an increase in proliferation, migration and radiation resistance suggesting the role of PDE4D as a TSG. Amino acids, cholesterol, fatty acids, carbohydrates and TCA intermediates were found to be altered in concentration.A mathematical model of glycolysis, TCA and glutaminolysis under normoxia and hypoxia was constructed through the amalgamation of two established models from the literature. New reactions, parameters and metabolite concentrations were added and unnecessary entities were deleted. COmplex PAthway SImulator (COPASI) was utilised to construct the model before validating the model using experimental data from the literature and steady state and flux analyses. Sensitivity analysis and a reduction in external glucose and glutamine were mimicked and the alterations in hypoxic and normoxic metabolism analysed. The reactions vCSII, vGS, vPGK and vGII were identified as potential therapeutic targets which may affect metabolism in hypoxia only. However, certain validation methods proved unsuccessful and hence the model requires further work before attempting the analyses again.

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Chen, Yixuan. "Effect of hypoxia on dendritic cell function and differentiation." Thesis, University of Oxford, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.426446.

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Salvadore, Christopher P. "Brain cell injury: metabolic dysfunction in ischemia and hypoxia." Thesis, Boston University, 1988. https://hdl.handle.net/2144/38099.

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Thesis (M.A.)--Boston University
PLEASE NOTE: Boston University Libraries did not receive an Authorization To Manage form for this thesis or dissertation. It is therefore not openly accessible, though it may be available by request. If you are the author or principal advisor of this work and would like to request open access for it, please contact us at open-help@bu.edu. Thank you.
A sequence of biochemical events responsible for the destruction of brain cells during oxygen deprivation has been proposed based on available experimental evidence reviewed in the text. Oxygen deficiency results in a diminished p02 within the cell, resulting in inhibition of the electron transport chain and depletion of ATP pools. The decrease in cellular energy charge mediates a reduction in phospholipid synthesis, as well as the collapse of ionic gradients across the plasma and organelle membranes. Highly elevated levels of cytosolic Ca++ follow, activating membrane bound phospholipases which progressively deplete the membrane of its phospholipids. Membrane structure becomes severely compromised, resulting in loss of function. Cell death ensues.
2031-01-01

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McAleer, James Joseph Anthony. "The hypoxic tumour cell : a therapeutic challenge." Thesis, Queen's University Belfast, 1991. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.317447.

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Sandlund, Johanna. "Angiogenesis in human renal cell carcinoma : hypoxia, vascularity and prognosis." Doctoral thesis, Umeå : Univ, 2007. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-1331.

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Heddleston, John Michael. "The Role of Hypoxia in Modulating Glioma Cell Tumorigenic Potential." Case Western Reserve University School of Graduate Studies / OhioLINK, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=case1310043767.

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Wilson, William J. "Hypoxia inducible factor 1a : molecular mechanisms of regulation /." Stockholm, 2002. http://diss.kib.ki.se/2002/91-7349-223-X.

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Davis, Brandon James. "VEGF signaling mechanisms in increased blood brain barrier permeability following hypoxia." Diss., Search in ProQuest Dissertations & Theses. UC Only, 2007. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3261273.

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Givelet, Maëlle. "Étude des cellules souches germinales : caractérisation des cellules souches germinales humaines et effets de l'hypoxie Transcriptional landscape of spermatogonial stem cells and progenitors in human spermatogenesis Impact of hypoxia on the proliferation and colony-formation capacity of SSCs in culture Spermatogonial stem cells and progenitors are refractory to reprogramming to pluripotency by the transcription factors Oct3/4, c-Myc, Sox2 and Klf4." Thesis, Sorbonne Paris Cité, 2018. http://www.theses.fr/2018USPCB038.

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Tout au long de la vie, les spermatozoïdes sont produits dans le testicule à partir des cellules souches germinales (CSGs), au cours du processus de spermatogenèse. Les CSGs ont la capacité de s'auto-renouveler pour maintenir le stock de cellules souches et d'entrer en différenciation. L'infertilité masculine est responsable dans un cas sur deux des difficultés à enfanter au sein du couple. Chez les patients traités par radiothérapie et/ou chimiothérapie (plus particulièrement chez les enfants atteints de cancer), un des effets secondaires majeurs qui altèrent la qualité de vie après guérison est l'atteinte du stock de CSGs et les problèmes d'infertilité qui en découlent. Des solutions thérapeutiques telles que la transplantation de CSGs provenant de biopsies testiculaires réalisées avant les traitements anticancéreux sont actuellement étudiées. Des travaux récents de transplantation testiculaire de CSGs chez le primate non humain ont permis de restaurer, chez l'animal stérilisé, une spermatogénèse permettant la fécondation in vitro d'ovocytes à partir de spermatozoïdes issus des CSGs transplantées. Ces résultats très encourageants font entrer pour la première fois la transplantation des CSGs dans le champ de l'application préclinique. Cependant, l'identité du pool de CSGs humaines et les mécanismes moléculaires gouvernant leur auto-renouvellement restent peu connus. Mon travail de thèse a porté sur la caractérisation des CSGs chez l'Homme. Au cours de ce projet j'ai également utilisé le modèle murin afin de mieux appréhender certains mécanismes de régulation physiologique des CSGs. Dans un premier temps, ce travail de thèse a contribué à définir, à l'aide d'une combinaison de marqueurs cellulaires et d'un test fonctionnel, la transplantation testiculaire, une population de spermatogonies immatures enrichie en CSGs. Le profil d'expression de cette population a été réalisé par une analyse transcriptomique, et nous a permis de définir un réseau de régulateurs de la transcription préférentiellement exprimé dans cette population. Parmi ces régulateurs, nous nous sommes focalisés sur l'étude du répresseur de transcription bHLH Hes1 dans le modèle murin. Dans des conditions de culture de privation de sérum et de facteurs de croissance, induisant un arrêt prolifératif et la quiescence, notre étude tend à montrer un rôle protecteur de Hes1 contre la mort cellulaire. Nous avons également observé une diminution du nombre de cellules ayant le potentiel à régénérer une spermatogenèse dans les cultures de CSGs lorsque l'expression de Hes1 est diminuée à l'aide de siRNA. Dans un second temps, nous avons étudié l'effet de l'hypoxie sur l'auto-renouvellement et la différenciation des CSGs murines en culture. En effet, l'hypoxie est une composante importante de la niche des cellules souches qui régule leur destin cellulaire. Nous avons observé qu'une forte hypoxie (1% et 0,1% d'oxygène) a un effet délétère sur la capacité des CSGs murines à former des colonies, et qu'elle induit modérément la quiescence et un début de différenciation. Un effet positif sur la fonctionnalité des CSGs adultes murines a été observé à 3,5% d'oxygène, mais ces conditions n'ont pas d'effets bénéfiques sur la prolifération et le maintien des CSGs humaines à long terme. Enfin, mon travail de thèse a contribué à une meilleure compréhension des mécanismes responsables de la reprogrammation spontanée vers la pluripotence des CSGs in vitro. Si les facteurs de Yamanaka sont efficaces pour la reprogrammation des cellules somatiques testiculaires, ils ne permettent pas celle des CSGs adultes. Ces résultats suggèrent donc l'existence d'un mécanisme spécifique empêchant la reprogrammation par les facteurs de Yamanaka des cellules germinales adultes en un état pluripotent
Throughout life, sperm cells are produced from germinal stem cells (GSCs) during the process of spermatogenesis in the testis. GSCs have the ability to self-renew to maintain stem cell stock and to differentiate. Male infertility is responsible in one out of two cases of issues of procreation. In patients treated with radiotherapy and/or chemotherapy (particularly in children with cancer), one of the major side effects that affects the quality of life after healing is the attrition of the GSCs stock and the subsequent problems of infertility. The most severe infertility issue, the Sertoli-Cell-Only syndrome with a complete germ cell aplasia, results from the exhaustion of the GSCs population. Therapeutic solutions such as transplantation of GSCs obtained from testicular biopsies harvested prior to cancer treatments are being studied. Recent studies on testicular transplantation of GSCs in non-human primates have shown the restoration, in the sterilized animal, of spermatogenesis allowing in vitro fertilization of oocytes by intracytoplasmic microinjection of spermatozoa derived from transplanted GSCs. These very encouraging results bring for the first time the transplantation of GSCs into the field of preclinical application. However, the identity of the pool of human GSCs and the molecular mechanisms governing their self-renewal remain poorly known. My thesis work focused on the characterization of GSCs in humans. During this project I also used the mouse model of GSCs to better understand some mechanisms of the physiological regulation of GSCs. As a first step, this thesis work has contributed to define, using a combination of cell markers and the testicular transplantation, as functional test, a population of immature spermatogonia enriched in GSCs. The specific expression profile of this population was performed by transcriptomic analysis, and allowed us to define a transcription regulator network preferentially expressed in this population. Among the enriched transcriptional regulators in the human immature spermatogonia population, we focused on the study of the bHLH transcriptional repressor Hes1 in the murine model. Under serum and growth factors-deprived conditions of culture inducing proliferative arrest and quiescence, our study tends to show a protective role of HES1 against cell death. In addition, we also observed a decrease in the number of cells with the potential to regenerate spermatogenesis in GSCs cultures when Hes1 expression is decreased. Secondly, we studied the effect of hypoxia on the self-renewal and differentiation of murine GSCs in culture. Indeed, hypoxia is an important component of the stem cell niche that regulates their cell fate. We observed that high hypoxia (1% and 0.1% oxygen) has a deleterious effect on the ability of murine GSCs to form colonies, and moderately induce quiescence and the onset of differentiation of GSCs. A positive effect on the functionality of murine adult GSCs has been observed at 3.5% oxygen, but these conditions do not support the proliferation and maintenance at long-term of human GSCs. Finally, this work has contributed to a better understanding of the mechanisms responsible for the spontaneous reprogramming to pluripotency of GSCs in vitro. While Yamanaka factors are effective for reprogramming testicular somatic cells, they do not allow reprogrammation of adult GSCs and spermatogonial progenitors. These results suggest the existence of a specific mechanism preventing reprogramming by Yamanaka factors of adult germ cells into a pluripotent state

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Hernandez, Ivan. "PRIMING CARDIOVASCULAR STEM CELLS FOR TRANSPLANTATION USING SHORT-TERM HYPOXIA." CSUSB ScholarWorks, 2016. https://scholarworks.lib.csusb.edu/etd/348.

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Conventional medical treatments fail to address the underlying problems associated with the damage inflicted by a coronary event. Thus, the long-term prognosis of patients admitted for heart failure is disheartening, with reported survival rates of 25 percent. Recent advances in stem cell research highlight the potential benefits of autologous stem cell transplantation for stimulating repair in heart tissue. However, a majority of those suffering from cardiovascular diseases are older adults whose autologous cells no longer possess optimum functional capacity. Additional work is needed to identify the optimal cell types or conditions that will promote cardiovascular regeneration across all age groups. A pretreatment, such as short-term hypoxia, and concurrent implementation of a novel progenitor, such as those that co-express Isl-1 and c-Kit, may enhance the results reported in clinical trials completed to date. However, the effects of short-term hypoxia in this novel cell type are unknown and warrant investigation in vitro. Cloned adult and neonatal Isl-1+ c-Kit+ human cardiovascular progenitor cells were characterized and expanded for study. Populations from both age groups were preconditioned using short-term hypoxia (1% O2 for six hours) and, to identify shifts in gene expression, compared to their respective control (21% O2 at 37 °C) via qRT-PCR. Flow cytometry and western blot analysis was utilized to measure phosphorylation of Akt. Progression through the cell cycle was also analyzed by flow cytometry. Cellular function was evaluated by the use of a TUNEL assay and Transwell® invasion assay. Hypoxia-mediated alterations of a genetic or functional nature in Isl-1+ c-Kit+ human cardiac progenitors are clearly age-dependent. Although both age groups accrued benefit, the neonatal progenitors procured significantly greater improvements. Short-term hypoxia significantly elevated Akt phosphorylation in neonatal Isl-1+ c-Kit+ human cardiac progenitors. Benefits afforded to both age groups by hypoxic pretreatment included significant upregulation of pro-survival transcripts, and enhanced invasion capabilities in vitro. Therefore, prior to transplantation, hypoxic preconditioning may improve the ability of transplanted stem cells to home towards damaged areas of the heart and support cardiac regeneration in vivo.

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Noman, Muhammad zaeem. "Influence of hypoxia on tumour cell susceptibility to cytotoxic T lymphocyte mediated lysis." Thesis, Paris 11, 2012. http://www.theses.fr/2012PA11T051/document.

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L’hypoxie est une caractéristique commune des tumeurs solides et l’une des spécificités du micro environnement tumoral. L’hypoxie tumorale joue un rôle important dans l’angio génèse, la progression maligne, le développement de métastases, la chimio/radio-résistance et favorise l’échappement au système immunitaire du fait de l’émergence de variant tumoraux avec un potentiel de survie et de résistance à l’apoptose augmenté. Cependant, très peu de travaux ont étudié l’impact de l’hypoxie tumorale sur la régulation de la susceptibilité des tumeurs à la lyse induite par la réponse immune cytotoxique. Nous nous sommes donc demandé si l’hypoxie pouvait conférer aux tumeurs une résistance à la lyse induite par les lymphocytes T cytotoxiques (CTL). Nous avons démontré que l’exposition de cellules cibles tumorales à l’hypoxie possédait un effet inhibiteur sur la lyse de ces cellules tumorales par des CTL autologues. Cette inhibition n’est pas associée à des altérations de la réactivité de CTL ou de la reconnaissance des cellules cibles. Cependant, nous avons montré que l’induction hypoxique concomitante de la phosphorylation de STAT3 (pSTAT3) au niveau de la tyrosine 705 et du facteur HIF-1α (Hypoxia Inducible Factor-1 alpha) est liée fonctionnellement à l’altération de la susceptibilité de cellules tumorales bronchiques non à petites cellules (NSCLC) à la mort induite par les CTL. Nous avons aussi montré que la résistance de cellules tumorales bronchiques à la lyse CTL induite par l’hypoxie était associée à une induction d’autophagie dans les cellules cibles. En effet, l’inhibition de l’autophagie empêche la phosphorylation de STAT3 (via l’inhibition de la kinase Src) et restaure la susceptibilité des cellules tumorales hypoxiques à la lyse induite par les CTL. De plus, l’inhibition in vivo de l’autophagie par l’hydroxychloroquine (HCQ) dans le modèle murin portant la tumeur B16F10 and chez les souris vaccinée avec le peptide TRP2 augmente de façon drastique l’inhibition de la croissance tumorale. Collectivement, cette étude établit un nouveau lien fonctionnel entre l’autophagie induite par l’hypoxie et la régulation de la lyse induite par les cellules T spécifique d’antigènes et souligne le rôle majeur de l’autophagie dans le contrôle de la croissance tumorale in vivo.Finalement, étant donné que le la résistance tumorale à la lyse induite par les cellules tueuses est très probablement régulée par de multiples facteurs, nous avons aussi eu pour but d’identifier les micro-ARNs (miRs) régulés par l’hypoxie dans des modèles de NSCLC et de mélanome et leur implication putative dans la régulation de la susceptibilité tumorale à la lyse induite par les cellules T spécifique d’antigènes. Le micro-ARN 210 (miR-210) est ainsi significativement induit de manière dépendante de HIF-1α dans des cellules de NSCLC et de mélanome, et miR-210 est exprimé dans les zones hypoxiques de tissus issus de NSCLC. De plus, nous avons démontré que l’induction de miR-210 par l’hypoxie régule la susceptibilité tumorale à la lyse induite par les CTL en partie grâce à l’inhibition de l’expression de PTPN, HOXA1 et TP53I11, indiquant que miR-210 joue un rôle potentiel dans la régulation de la réponse immune antitumorale
Hypoxia is a common feature of solid tumors and one of the hallmarks of tumor microenvironment. Tumor hypoxia plays an important role in angiogenesis, malignant progression, metastatic development, chemo-radio resistance and favours immune evasion by the emergence of tumor variants with increased survival and anti-apoptotic potential. There is very little work done on the impact of tumor hypoxia on the regulation of tumor susceptibility to the lysis induced by cytotoxic antitumor response. Therefore, we asked whether hypoxia confers tumor resistance to cytotoxic T lymphocyte (CTL)-mediated killing. We demonstrated that exposure of target cells to hypoxia has an inhibitory effect on the CTL-mediated autologous target cell lysis. Such inhibition was not associated with an alteration of CTL reactivity and tumor target recognition. We also showed that the concomitant hypoxic induction of Signal transducer and activator of transcription 3 (STAT3) phosphorylation on tyrosine 705 residue (pSTAT3) and hypoxia inducible factor 1 alpha (HIF-1α) is functionally linked to the alteration of Non small cell lung carcinoma (NSCLC) target susceptibility to CTL-mediated killing. We also showed that hypoxia-induced resistance of lung tumor to CTL-mediated lysis was associated with autophagy induction in target cells. Inhibition of autophagy resulted in impairment of pSTAT3 (via inhibition Src kinase) and restoration of hypoxic tumor cell susceptibility to CTL-mediated lysis. Moreover, in vivo inhibition of autophagy by hydroxychloroquine (HCQ) in B16F10 tumor bearing mice and mice vaccinated with TRP2 peptide dramatically increased tumor growth inhibition. Collectively, the current study establishes a novel functional link between hypoxia-induced autophagy and the regulation of antigen specific T cell lysis and points to a major role of autophagy in the control of in vivo tumor growth.Finally, as resistance of tumor targets to killer cells is likely to be regulated by multiple factors, we further aimed to identify the microRNA’s regulated by hypoxia in NSCLC and melanoma and their putative involvement in the regulation of tumor susceptibility to antigen-specific CTL-mediated killing. MicroRNA-210 (miR-210) was significantly induced in a HIF-1α dependent manner in NSCLC and melanoma cells and miR-210 was expressed in hypoxic zones of human NSCLC tissues. Moreover, we demonstrated that hypoxia-induced miR-210 regulates tumor cell susceptibility to CTL-mediated lysis in part by suppressing PTPN, HOXA1 and TP53I11 expression indicating that miR-210 plays a potential role in the regulation of anti-tumor immune response

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Kukucka, Mark A. "Mechanisms by which hypoxia augments Leydig cell viability and differentiated cell function in vitro." Diss., Virginia Tech, 1993. http://hdl.handle.net/10919/38407.

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The 1980's heralded the discovery and identification of extra-pituitary sources of the neurohypophysial hormone oxytocin in non-neural tissues of several animal species. The presence, location and biosynthesis of significant amounts of oxytocin in the ovarian corpus luteum was followed by the immunocytochemical demonstration of an oxytocin-like peptide in the testicular interstitial cells. Leydig cells, which comprise up to 80% of the testicular intertubular cell population, are known to synthesize testosterone in situ. Indirect evidence indicated that an oxytocin-like peptide was also present in Leydig cells. The question arose whether this peptide was synthesized de novo by Leydig cells or was taken up and stored by the cells following biosynthesis at some other intra- and/or extra-gonadal source(s). Since luteinizing hormone (LH) and ascorbate are known to augment the production of oxytocin in ovarian granulosa cells, varying concentrations of these two stimulants were used to monitor the biosynthesis of oxytocin from isolated Leydig cells in culture.
Ph. D.

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Kukucka, Mark Anthony. "Mechanisms by which hypoxia augments Leydig cell viability and differentiated cell function in vitro /." This resource online, 1993. http://scholar.lib.vt.edu/theses/available/etd-06062008-170416/.

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27

Guimbellot, Jennifer S. "Role of hypoxia in epithelial gene regulation." Thesis, Birmingham, Ala. : University of Alabama at Birmingham, 2007. https://www.mhsl.uab.edu/dt/2009r/guimbellot.pdf.

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Schmidt, Dirk. "Role of JunB in hypoxia-mediated cell response and tumour angiogenesis." [S.l.] : [s.n.], 2001. http://www.freidok.uni-freiburg.de/volltexte/327.

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Peurala, E. (Emmi). "Regulators of hypoxia response and the cell cycle in breast cancer." Doctoral thesis, Oulun yliopisto, 2013. http://urn.fi/urn:isbn:9789526202709.

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Abstract Breast cancer is the most common cancer affecting the female population of the Western world. It is a heterogeneous disease entity that encompasses tumors with remarkably different forms of behaviour, and it is therefore vital to distinguish patients with good and poor prognoses. The classical prognostic and predictive factors for breast cancer serve as tools for clinical oncologists when planning treatment, but the growing awareness of breast cancer biology is bringing about a need for novel prognostic and predictive biomarkers. This thesis examines the prognostic significance of hypoxia response and cell cycle regulators in ductal breast cancer and in triple-negative breast cancer (negative for hormone receptors and human epidermal growth factor receptor 2), concluding that PHD2 and PHD3 are associated with a good prognosis, while the role of PHD1 is controversial, as it is associated with proliferation in ductal breast cancer but with node-negative status in triple-negative breast cancer. In our experiments HIF-1α redeemed its role as a marker of an adverse prognosis, whereas the role of HIF-2α appeared to be the opposite. Our data suggest that PHDs can have other targets than the HIF-αs, and that triple-negative breast tumors express more HIF-1α and less HIF-2α and PHD3 than those with a good prognosis. Furthermore, we identified cyclin D1 as a biomarker with independent prognostic significance in ductal breast cancer, being associated with good prognostic factors and a better outcome, whereas the opposite was seen in triple-negative breast cancer. CDK4 was associated with high proliferation in triple-negative breast cancer. In addition, high levels of p16 correlated with increased survival in breast cancer patients independently of receptor status
Tiivistelmä Rintasyöpä on naisten yleisin syöpä läntisessä maailmassa. Rintasyöpä on heterogeeninen tautiryhmä, jossa kasvaimet vaihtelevat biologiselta käyttäytymiseltään huomattavasti. Tästä syystä on tärkeää erottaa hyvä- ja huonoennusteiset potilaat. Syöpälääkärit käyttävät klassisia ennustetekijöitä hoitopäätöksiä tehdessään, mutta lisääntynyt tieto rintasyövän biologiasta on saanut aikaan tarpeen löytää uusia ennustetekijöitä. Tässä väitöskirjatyössä tutkimme hypoksiavasteen ja solusyklin säätelijöiden ennusteellisuutta duktaalisessa rintasyövässä sekä kolmoisnegatiivisessa (ei ilmennä hormonireseptoreita eikä epidermaalikasvutekijäreseptoria) rintasyövässä. PHD2 ja PHD3:n vahva ilmentyminen liittyi parempaan ennusteeseen, mutta PHD1:n esiintymisen vaikutus oli ristiriitainen. PHD1:n ilmentyminen liittyi lisääntyneeseen solujakautumiseen duktaalisessa rintasyövässä, mutta kolmoisnegatiivisessa rintasyövässä sen esiintyminen liittyi vähentyneeseen imusolmukemetastasointiin. Tutkimuksessamme HIF-1α osoittautui huonon ennusteen merkiksi. Sitä vastoin HIF-2α:n ilmentymisen vaikutus näytti liittyvän parempaan ennusteeseen. Tuloksemme osoittavat, että PHD-entsyymeillä on mahdollisesti muitakin kohteita kuin HIF-α:t. Osoitimme myös, että HIF-1α:n ilmentyminen on yleisempää ja HIF-2α:n sekä PHD3:n ilmentyminen vähäisempää kolmoisnegatiivisessa kuin duktaalisessa rintasyövässä. Lisäksi totesimme, että sykliini D1 on itsenäinen ennustetekijä liittyen parempaan ennusteeseen duktaalisessa rintasyövässä. Huomioitavaa on kuitenkin, että kolmoisnegatiivisessa rintasyövän alaryhmässä sykliini D1:n esiintyminen oli huonon ennusteen merkki. CDK4 osoittautui voimakkaan proliferaation merkiksi kolmoisnegatiivisessa rintasyövässä. Lisäksi osoitimme, että p16:n ilmentyminen liittyy parempaan ennusteeseen sekä duktaalisessa rintasyövässä että kolmoisnegatiivisessa rintasyövässä

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Chiarotto, James Anthony. "Hypoxia-induced upregulation of VEGF mRNA in cervical cancer cell lines." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape10/PQDD_0010/MQ40781.pdf.

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31

Ahmed, A. "Regulation of p53-dependent cell death responses in normoxia and hypoxia." Thesis, University College London (University of London), 2011. http://discovery.ucl.ac.uk/1298195/.

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Hypoxia, defined as low oxygen tension, is a common characteristic of growing tumours. Tumour cells adapt to their hypoxic microenvironment by inducing angiogenesis and escaping cell death. In doing so, tumour cells become resistant to radiotherapy and many forms of chemotherapy. Hypoxia signalling and angiogenesis is mediated by the hypoxia-inducible factor (HIF) transcriptional complex. HIF can crosstalk to the p53 tumour suppressor protein, a critical regulator of cell cycle and cell death responses to stress. This study aims to understand how cell death responses are regulated in tumour cells by HIF and p53, in normoxia and in hypoxia. Recently, activation of p53 by the small molecule RITA has been investigated. Flow cytometry, comet assays and western blot analysis have been used to reveal a novel p53-dependent DNA damage response that activates cell cycle checkpoints, and induces significant cell death of hypoxic tumour cells. Activation of p53 also achieves anti-angiogenic effects, both in vitro and in vivo by inhibition of HIF-1α protein synthesis and HIF target genes, including VEGF. The MEK-ERK MAPK pathway has also been investigated as a critical modulator of p53-dependent cell death in normoxia and hypoxia. Inhibition of MEK1/2 by the MAPK signalling inhibitor PD98059 significantly inhibits p53 induction and cell death responses by RITA. The anti-tumour effect of p53 activation in response to RITA is therefore dependent on MEK-ERK signalling. By understanding p53 interactions with HIF signalling, and the role of p53 in responding to DNA damage and apoptotic signals, this study has potential to improve the therapeutic targeting of resistant tumours with deregulated angiogenic and cell death pathways.

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Pike, Luke R. G. "The role of ATF4 in hypoxia-induced cell death in cancer." Thesis, University of Oxford, 2011. http://ora.ox.ac.uk/objects/uuid:f32e03f9-0bd2-4dd1-8320-b082b9b2d363.

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Cancer cells survive the harsh oxygen and nutrient deprivation of the tumour microenvironment through the selection of apoptosis-resistant and glycolytic clones (Cairns et al., 2011; Graeber et al., 1996). In particular, the integrated stress response (ISR) has been shown to be pivotal in cancer cell survival in vivo and the resistance of cancer cells to therapy (Harding et al., 2003). In recent years, it has become apparent that increased autophagy is one mechanism by which the ISR can confer resistance to stress (Kroemer et al., 2010). ATF4 is a major transcriptional effector of the integrated stress response in severe hypoxia (<0.01% O₂). ATF4 is a well-established regulator of genes involved in oxidative stress, amino acid synthesis and uptake, lipid metabolism, protein folding, metastasis, and angiogenesis. Recent work has demonstrated an important role of ATF4 in promoting resistance to severe hypoxia through the transcriptional upregulation of MAP1LC3B and ATG5, essential components of the autophagy machinery (Rouschop et al., 2009b; Rzyski et al., 2010). In this work, the author describes several novel ATF4 target genes, and examines their role in the regulation of autophagy and the resistance of cancer cells to severe hypoxia. In the first part of this thesis, the author shows that three BH3-only members of the BCL-2 family of proteins--HRK, PUMA, and NOXA--are upregulated in response to severe hypoxia in an ATF4-dependent manner. In particular, the author shows that the poorly described BH3-only protein HRK is a direct target of transcriptional activation by ATF4, and that HRK induces autophagy in severe hypoxia, thereby providing the first evidence that the integrated stress response can transcriptionally trigger the autophagy process. In contrast to the previously described role of HRK in apoptosis, this thesis demonstrates that HRK can play a pro-survival role in the context of breast cancer cells. In the latter part of this thesis, the author identifies the essential autophagy gene ULK1 as an ISR target. The author shows that ULK1 expression in severe hypoxia is transcriptionally upregulated through direct activation by ATF4. The author identifies ULK1 as a crucial regulator of autophagy and mitophagy in both normoxia and severe hypoxia and shows that ULK1 plays a pivotal role in cancer cell survival. Furthermore, it is shown that human breast cancer patients with high levels of ULK1 relapse earlier than those with low levels of ULK1, thereby identifying ULK1 as a potential target for cancer therapy.

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Anderson, Scott James. "Beta-cell dedifferentiation following short term hypoxia and clinical islet transplantation." Thesis, University of Newcastle upon Tyne, 2017. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.750387.

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Individuals with insulin-requiring type 1 diabetes account for around 10% of all diabetes cases in the UK. Administration of exogenous insulin to maintain blood glucose homeostasis can lead to life-threatening hypoglycaemia in a subset of people with type 1 diabetes, who have lost hypoglycaemic awareness. Islet transplantation is a life-saving therapy for these individuals who suffer recurrent, hospital requiring, episodes of hypoglycaemia. Despite improving outcomes, mainly due to an improved immunosuppression regimen, the functional duration of islet transplants still fails to match the gold standard in endocrine replacement therapy, whole pancreas transplant. During pancreas retrieval, islet isolation and post-transplantation, islets are subjected to prolonged periods of low oxygen (hypoxia). Islets are highly vascularised within the pancreas, providing sufficient oxygen for appropriate beta-cell glucose sensing and appropriate insulin secretion. Islet hypoxia is associated with decreased beta-cell function and worse transplant outcomes. Despite this, the processes which cause betacell dysfunction during periods of hypoxia are not fully understood. The aim of this thesis was to elucidate the mechanisms of hypoxia-induced beta-cell dysfunction. Using the MIN6 functional beta-cell line, hypoxic stress was modelled in vitro. MIN6, subjected to short-term hypoxia, demonstrated decreased viability, increased expression of hypoxia-associated genes and reduction in beta-cell specific gene expression. Gene expression changes were associated with decreased in vitro function, decreased insulin content and nuclear-to-cytoplasmic translocation of betacell transcription factors Pdxl and Nkx6.1. Using optimised isolation protocols, freshly isolated human islets were established in normal and hypoxic culture conditions. Hypoxic cultured islets demonstrated decreased expression of beta-cell genes, decreased function and viability. Further analysis of beta-cell phenotype by immunofluorescence identified decreased coexpression of end-differentiated marker urocortin-3 and insulin, alongside coexpression of insulin and the mesenchymal marker vimentin in hypoxia treated islets. This model suggests short-term hypoxic exposure causes changes in end- differentiated islet beta-cell phenotype. To determine if these phenotypes are present following clinical islet transplantation we performed immunofluorescence analysis of islets engrafted within recipient liver. Transplanted islet beta-cells in two recipients with excellent graft function demonstrated loss of mature phenotype, with all insulin positive cells lacking coexpression of urocortin-3. Transplanted islet beta-cells also contained cells coexpressing insulin and vimentin, suggesting mature end-differentiated phenotype is lost following transplantation. To determine whether hypoxia-induced HIF1α stabilisation plays a role in loss of end- differentiated beta-cell phenotype, HIF1α stabilisation and knockdown models were established in the MIN6 cells. Using cobalt chloride as a hypoxia mimic leading to HIF1α stabilisation led to changes in beta-cell gene expression and function comparable to those induced by hypoxia. These changes persisted following HIF la knockdown demonstrating that changes in beta-cell phenotype during hypoxia are HIF1α independent. These studies offer new insight into loss of mature beta-cell phenotype as a novel cause of beta-cell dysfunction during islet hypoxia.

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Emery, Elizabeth Dorothy. "Regulation of stem cell marker LGR5 by hypoxia in colorectal cancer." Thesis, University of Bristol, 2015. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.682727.

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Leucine rich repeat containing G-protein coupled receptor 5 (LGR5) is a well-established stem cell marker in the normal intestine. Recent evidence suggests LGR5 is also a marker of cancer stem cells in colorectal tumours. Cancer stem cells propagate and maintain tumours and are hypothesized to be refractory to therapy. Solid tumours frequently experience hypoxia and an unresolved question remains as to how the cancer stem cell population survives this environmental stress. Several regulatory links are known to exist between hypoxic and stem cell signalling pathways. However, the role of hypoxia in regulating LGR5 is yet to be elucidated. Here it is reported that hypoxia down-regulates LGR5 expression. Both protein and mRNA expression is reduced in hypoxia. LGR5 down regulation was observed in cells derived from adenoma, primary carcinoma and metastases, suggesting this process occurs throughout tumourigenesis. LGR5 was found to be re-expressed following re-oxygenation, demonstrating tumour cells ability to switch between expressing LGR5 in normoxic conditions and reducing expression in hypoxia. LGR5 is an established target gene of the WNT signalling pathway and hypoxia has been reported previously to regulate the WNT pathway. Results presented here suggest hypoxic down-regulation of the WNT pathway mediates the hypoxic regulation of LGR5. Topflash WNT reporter activity and expression of a selection of additional WNT target genes decreased in hypoxia in the cell lines tested here. Preliminary ChiP experiments suggest WNT signalling effectors Beta-catenin and TCF4 are lost from the LGR5 promoter in hypoxia. It has been reported previously that HIF-1 interacts with the WNT pathway in hypoxia to down-regulate WNT target gene expression. However HIF-1 does not regulate LGR5 in the cell lines tested here. Highly related HIF-2, though, is critical in the hypoxic regulation of LGR5 in the LoVo cell line and not in other cell lines tested here. The reversible down regulation of stem cell markers during hypoxia may have important implications for targeting cancer stem cells in vivo where tumours are heterogeneous with fluctuating areas of hypoxia.

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Wangpaichitr, Medhi. "The Relevance of mTOR and Hypoxia Inducible Factor to 2-Deoxy-D-Glucose Toxicity in Lung Cancer Cell Lines Under Hypoxia." Scholarly Repository, 2008. http://scholarlyrepository.miami.edu/oa_dissertations/156.

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Hypoxic regions found in most solid tumors often contain cells which are resistant to various cancer therapies. However, hypoxia also forces cells to rely solely on the catabolism of glucose through glycolysis for ATP production and survival, thereby creating a therapeutic window that can be exploited by glycolytic inhibitors, such as 2-deoxy-D-glucose (2-DG). Previous studies in our lab demonstrated that activation of Hypoxia Inducible Factor (HIF-1) in hypoxic tumor cells confers resistance to glycolytic inhibition by 2-DG. In surveying a number of tumor types for differences in intrinsic levels of HIF-1 alpha under hypoxia, we found that pathways upstream of HIF -- i.e. AKT and mammalian target of rapamycin (mTOR) -- have significantly reduced activity in 2 human non-small lung cancer cell lines (NSCLC) as compared to 4 small cell lung cancer cell (SCLC) lines. This reduced activity of AKT and mTOR correlated with increased sensitivity to 2-DG under hypoxia. Since HIF-1 alpha translation is regulated by the mammalian target of rapamycin (mTOR), we examined the effects of blocking mTOR with an analog of rapamycin (CCI-779) in SCLC cells which express high levels of mTOR activity. Under hypoxia, treatment with CCI-779 resulted in HIF-1 alpha down-regulation. Furthermore, CCI-779 potentiated the cytotoxic effects of 2-DG in hypoxic SCLC cells. Conversely, CCI-779 did not increase 2-DG toxicity in NSCLC lines that do not express HIF, SCLC lines treated with siRNA against HIF-1 alpha, or HIF-deficient mutants. These latter results support the hypothesis that, although mTOR modulates numerous downstream pathways, mTOR inhibition by CCI-779 increases the toxicity of 2-DG in hypoxic cells through down-regulation of HIF-1 alpha. Overall, our findings show that CCI-779 hyper-sensitizes HIF-expressing hypoxic tumor cells to 2-DG. Additionally, our results suggest that the intrinsic expression of AKT, mTOR, and HIF in many tumor types may be important predictors of clinical responsiveness to 2-DG and could be used to guide future treatment decisions on whether to use 2-DG alone or in combination with an mTOR inhibitor.

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Bhogal, Ricky Harminder. "The role of oxidative stress and CD154-mediated reactive oxygen species in regulating hepatocyte cell death during hypoxia and hypoxia-reoxygenation." Thesis, University of Birmingham, 2013. http://etheses.bham.ac.uk//id/eprint/3857/.

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Hypoxia and hypoxia-reoxygenation (H-R) are pathogenic factors in many liver diseases and lead to hepatocyte death as a result of reactive oxygen species (ROS) accumulation. Activation of the Tumour Necrosis Factor-a (TNFa) super-family member CD40 by its cognate ligand CD 154 can induce hepatocyte apoptosis via induction of autocrine/paracrine F as Ligand/CD178 expression but the relationship between CD40 activation, ROS generation and hepatocyte cell death is poorly understood. Therefore, human hepatocytes were isolated from liver tissue and exposed to an in vitro model of hypoxia and H-R in the presence or absence of CD154 and/or various inhibitors. Hepatocyte ROS production, apoptosis, necrosis and autophagy were determined by a four-colour reporter flow cytometry assay. The in vivo regulation of liver injury by CD40 and CD 154 was determined using a murine model of partial liver ischaemia. Exposure of human hepatocytes to recombinant CD 154 or platelet-derived soluble CD 154 augmented ROS accumulation during H-R resulting in NADPH oxidase-dependent apoptosis and necrosis. The cyto-protectivc mechanism of autophagy limited apoptotic cell death during hypoxia and H-R. CD40 and CD 154 knockout mice but not wild type mice were protected from ischaemic liver injury. Hence, CD40:CD154 mediate hepatocytes cell death in vitro and in vivo during hypoxia and H-R.

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Swinson, Daniel. "Hypoxic markers in non-small cell lung cancer." Thesis, University of Leicester, 2004. http://hdl.handle.net/2381/29476.

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Hypoxia is an important factor in the pathogenesis of solid tumours. Hypoxia inducible factors (HIF)-1alpha and HIF-2alpha are transcription factors that in part mediate the cellular response to hypoxia. These transcription factors are involved in the regulation of angiogenesis, anaerobic metabolism, pH homeostasis, erythropoiesis and cell death.;Immunohistochemical (IHC) assays were optimised for HIF-1alpha and one of its transcriptional targets, Carbonic Anhydrase (CA) IX. Attempts to optimise an IHC assay for HIF-2alpha failed to produce reproducible staining. A scoring system was also devised to assess the extent of tumour necrosis (TN) in tumour sections. The expression of these factors was assessed in a retrospective series of patients who had NSCLC tumours resected with curative intent. The expression of EGFR, p53, Bcl-2, MMP-2 and MMP-9 and angiogenesis had previously been assessed.;Extensive TN, perinuclear (p) CA IX and high HIF-1alpha expression were associated with a poor prognosis. PCA IX, stage, gender, MMP-9 and angiogenesis were independent prognostic factors.;The spatial relationship between membranous CA IX expression and TN and tumour microvessels support other studies proposing that CA IX is a marker of tumour hypoxia.;EGFR expression was associated with pCA IX, membranous (m)CA IX and HIF-1alpha expression. In vitro studies demonstrated that prolonged treatment with the EGFR tyrosine kinase inhibitor, ZD 1839 suppressed CA IX expression. These results suggest that activated EGFR may induce CA IX. As such co-expression of these factors may identify patients that are more likely to respond to EGFR targeted therapies.

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Charlier, Nico Nawid. "Hypoxie-induzierter Zelltod und Veränderungen der HIF-1-Aktivität in PC12-Zellen." Doctoral thesis, Humboldt-Universität zu Berlin, Medizinische Fakultät - Universitätsklinikum Charité, 2004. http://dx.doi.org/10.18452/15012.

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Der Transkriptionsfaktor hypoxia inducible factor-1 (HIF-1) trägt zur Expression von adaptiven Genen unter hypoxischen Bedingungen bei. Zusätzlich wurde vermutet, dass HIF-1 eine Rolle in der Regulation des späten neuronalen Zelltodes spielt. Suspensionszellen und adhärenten PC12-Zellen mit Nervenwachstumsfaktor (NGF) behandelt, wurden als ein experimentelles Modell für die Untersuchung der Beziehung zwischen Hypoxie induziertem Zelltod und Aktivität von HIF-1 herangezogen. Zelltod wurde durchflusszytometrisch mit einer Doppelfärbung (Annexin V und Propidium-Jodid) der Zellen und durch eine Analyse der allgemeinen Zelltodparameter wie LDH und die mitochondriale Dehydrogenase bestimmt. Parallel wurden Zellen mit einem Kontrollvektor und einem hypoxiesensitiven Vektor mit drei Hypoxie-bindenden-Elementen (HBE) transfiziert und die durch HIF-1 aktivierte Luciferase gemessen. Hypoxieexposition der NGF-behandelten PC12-Zellen resultierte in einer höheren Zelltodrate verglichen mit den unbehandelten Kontrollzellen. PC12 Zellen, zwei Tage mit NGF behandelt, zeigten eine bis zu 10-fach verminderte HIF-1-Aktivität. Diese Verminderung könnte zu dem erhöhten hypoxie-induzierten Zelltod durch verminderte Expression von HIF-1alpha-regulierten Genen, welche für die Anpassung an Hypoxie verantwortlich sind, beitragen. Die Verminderung der HIF-1 Aktivität und der Anstieg der Hypoxiesensitivität könnte darauf hinweisen, dass NGF als eine Art hierarchisch organisiertes Signalmolekül fungiert.
The transcription factor hypoxia-inducible factor-1 (HIF-1) strongly contributes to the expression of adaptive genes under hypoxic conditions. In addition, HIF-1 has been implicated in the regulation of delayed neuronal cell death. Suspension-grown and adherent PC12 cells treated with NGF were used as an experimental model for studying the relationship between hypoxia-induced cell death and activation of HIF-1. Cell damage was assessed by flow cytometry of double-stained (annexin V and propidiumiodide) cells, and by analysis of the overall death parameters LDH and mitochondrial dehydrogenase. In parallel, cells were transfected with a control and a three-hypoxia-responsive-elements (HRE)-containing vector and HIF-1-driven luciferase activity was determined. Exposure of NGF-treated PC12 cells to hypoxia resulted in a higher cell death rate when compared to untreated controls. PC12 cells exposed for 2 days to NGF exhibited a decrease of HIF-1 activity up to a factor of ten. This decrease may contribute to the enhanced hypoxia-induced cell death via reduced expression of HIF-1alpha-regulated genes resposible for adaptation to hypoxia, like those for glucose transport proteins and enzymes of the glycolytic chain. The decrease in HIF-1 activity and the increase in hypoxia sensitivity may suggest that NGF act as an hierachically organized signaling molecule.

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Pan, Minglin, Ying Han, Rui Si, Rui Guo, Ankit Desai, and Ayako Makino. "Hypoxia-induced pulmonary hypertension in type 2 diabetic mice." SAGE PUBLICATIONS INC, 2017. http://hdl.handle.net/10150/623894.

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Hypoxia-induced pulmonary hypertension (HPH) is a progressive disease that is mainly caused by chronic exposure to high altitude, chronic obstructive lung disease, and obstructive sleep apnea. The increased pulmonary vascular resistance and increased pulmonary arterial pressure result in increased right ventricular afterload, leading to right heart failure and increased morbidity. There are several clinical reports suggesting a link between PH and diabetes, insulin resistance, or obesity; however, it is unclear whether HPH is associated with diabetes as a progressive complication in diabetes. The major goal of this study is to examine the effect of diabetic ''preconditioning'' or priming effect on the progression of HPH and define the molecular mechanisms that explain the link between diabetes and HPH. Our data show that HPH is significantly enhanced in diabetic mice, while endothelium-dependent relaxation in pulmonary arteries is significantly attenuated in chronically hypoxic diabetic mice (DH). In addition, we demonstrate that mouse pulmonary endothelial cells (MPECs) isolated from DH mice exhibit a significant increase in mitochondrial reactive oxygen species (ROS) concentration and decreased SOD2 protein expression. Finally, scavenging mitochondrial ROS by mitoTempol restores endothelium-dependent relaxation in pulmonary arteries that is attenuated in DH mice. These data suggest that excessive mitochondrial ROS production in diabetic MPECs leads to the development of severe HPH in diabetic mice exposed to hypoxia.

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Gunawardena, A. H. L. A. N. "Investigation of cell death and aerenchyma formation in roots of maize (Zea mays l.)." Thesis, Oxford Brookes University, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.322185.

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Lequeux, Audrey. "Impact du ciblage du domaine de liaison de HIF-1α avec HIF-1β sur le paysage immunitaire du mélanome Targeting HIF-1 Alpha Transcriptional Activity Drives Cytotoxic Immune Effector Cells into Melanoma and Improves Combination Immunotherapy Hijacker of the Antitumor Immune Response: Autophagy is Showing its Worst Facet Impact of Hypoxic Tumor Microenvironment and Tumor Cell Plasticity on The Expression of Immune Checkpoints Improving Cancer Immunotherapy by Targeting the Hypoxic Tumor Microenvironment: New Opportunities and Challenges." Thesis, université Paris-Saclay, 2020. http://www.theses.fr/2020UPASL026.

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L’hypoxie est une caractéristique majeure des tumeurs solides et elle est capable d’induire un microenvironnement tumoral immunosuppressif. Nous avons étudié l’impact de l’inhibition du domaine de liaison de HIF-1α avec HIF-1β sur le paysage immunitaire du mélanome murin B16-F10. Le ciblage de ce domaine de liaison inhibe l’activité transcriptionnelle de HIF-1α dans les cellules B16-F10 in vitro. In vivo, l’inhibition de l’activité transcriptionnelle de HIF-1α dans le mélanome B16-F10 montre une diminution significative de la croissance tumorale et une amélioration consistante de la survie des souris. La croissance tumorale est restaurée dans les souris immunodéficientes, soulignant l’importance du système immunitaire dans le contrôle de la croissance du mélanome. L’étude du phénotype des cellules immunitaires intra-tumorales révèle une augmentation de l’infiltration des cellules Natural Killer (NK), des lymphocytes T CD4+, des T régulateurs, des macrophages de type M1 et M2 et des cellules dendritiques lorsque l’activité transcriptionnelle de HIF-1α est inhibée. La déplétion de cellules NK dans notre modèle expérimental restaure la croissance tumorale, soulignant le rôle des NK dans la surveillance du mélanome B16-F10. Le changement du paysage immunitaire observé dans notre modèle corrèle également avec une modification du réseau de cytokines caractérisé par une nette augmentation de la sécrétion de CCL5 et de CCL2. En conclusion, cette étude met en évidence le rôle de HIF-1α dans le contrôle de la croissance et le remodelage du paysage immunitaire du mélanome B16-F10. Elle indique la possibilité de combiner un inhibiteur de HIF-1α avec une immunothérapie basée sur le blocage des points de contrôle immunitaire pour étendre leur efficacité et leur bénéfice thérapeutiques à un plus grand nombre de patients cancéreux
Hypoxia is a major feature of solid tumors and is able to induce a tumor immunosuppressive microenvironment. Here, we investigated the impact of inhibiting of the binding domain of HIF-1α to HIF-1β on the immune landscape of B16-F10 melanoma. Targeting this binding domain inhibits the transcriptional activity of HIF-1α in B16-F10 cells in vitro. In vivo, inhibiting the transcriptional activity of HIF-1α in B16-F10 melanoma shows a significant decrease in tumor growth and a consistent improvement in mice survival. Tumor growth is restored in immunodeficient mice, highlighting the critical role of the immune system in controlling melanoma growth. The phenotyping of intra-melanoma immune cells reveals an increase in Natural Killer (NK), CD4+ T cells, regulatory T cells, M1 and M2 macrophages and dendritic cells. NK depletion restores tumor growth in our experimental model, highlighting the role of NK cells in melanoma surveillance. The alteration of the immune landscape that we observed also correlates with a clear increase of secreted CCL5 and CCL2. In conclusion, this study highlights the role of HIF-1α in controlling the growth and the immune landscape of B16-F10 melanoma. It indicates the opportunity of combining HIF-1α inhibitors with immune checkpoint blockade to extend immune checkpoint blockade efficiency and therapeutic benefit to a larger number of cancer patients

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Sipe, Conor W. "Cloning and Functional Characterization of Hypoxia-Inducible Factor 1alpha Upstream Regions in Xenopus laevis." W&M ScholarWorks, 2003. https://scholarworks.wm.edu/etd/1539626407.

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Faysal, Joanne M. "The Effects of Hypoxia with Concomitant Acidosis on Prostate Cancer Cell Survival." Scholarly Repository, 2010. http://scholarlyrepository.miami.edu/oa_theses/69.

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Prostate cancer is the second most common cancer among men in the United States. While treatments for prostate cancer exist, none are curative. As a solid tumor, prostate cancer can grow beyond the diffusion limits of oxygen, thereby resulting in a hypoxic environment. While hypoxia can cause death to a variety of cell types, tumor cells can develop resistance to hypoxia and survive under minimal oxygen conditions. Hypoxia in tumor cells has also been associated with poor prognosis, increased metastasis, and decreased efficacy of chemotherapy. BNIP3, a BH-3 only proapoptotic Bcl-2 family member, has been shown to play an important role in cell death under hypoxic conditions in a variety of cell types. In normoxia, BNIP3 shows little to no expression in both cardiomyocytes and many cancer cell types, but is then upregulated under hypoxic conditions. Previous work in our laboratory provides evidence that hypoxia alone, as well as the concomitant increase in BNIP3 expression, cannot cause death of rat neonatal cardiomyocytes. Instead, our studies found that hypoxia with concomitant intracellular acidosis is required. Further studies indicated that BNIP3 is also necessary for hypoxia-acidosis associated cell death in cardiomyocytes. Our results in rat neonatal cardiomyocytes led us to hypothesize that cell death could be induced in hypoxic prostate cancer cells if concomitant acidosis could be induced. Additionally, our intention was to determine if BNIP3 was required for any prostate cancer cell death that may occur under hypoxia-acidosis conditions.

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Sheares, Karen Kwie Kay. "The regulation of human pulmonary artery smooth muscle cell growth by hypoxia." Thesis, University of Cambridge, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.614833.

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Ahmed, Muhammad A. "Exploring the impact of hypoxia mimetic agents on multipotent stem cell biology." Thesis, Keele University, 2018. http://eprints.keele.ac.uk/4532/.

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Oxygen is an important molecule in life and is essential for a broad spectrum of physiological reactions that include, but are not restricted to, cell metabolism, respiration and growth. Under physiological conditions, oxygen levels vary from one tissue to another ranging from 0.002% to 10% and substantially lower than the atmospheric level of 21 % O2. Hypoxia is defined as a state of reduced O2 supplied to cells or tissues when compared to their normal physiological levels. Hypoxia mimetic agents (HMAs) are chemical used to induce pharmacological hypoxia without affecting environmental oxygen levels per se. The name HMA is routinely applied as these agents will activate the family of transcription factors which respond to reduced oxygen availability, Hypoxia Inducible Factors (HIF), which is taken as a surrogate indicator for hypoxia. These agents have been proposed as a cheaper alternative to engineered oxygen control measures including tri-gas incubators and workstation approaches. Multipotent stem cells (e.g. neuronal PC12 and human mesenchymal stem cells (hMSCs)) due their ability to differentiate into various cell types provide a means to develop better understanding of tissue development, repair, and protection. In addition, they provide better therapeutic perspectives and opportunities for the treatment of many diseases. Physiological oxygen plays a key role in the maintenance of cellular proliferation, behaviour and biology both in vivo and when mimiced in vitro. Physiological oxygen and hypoxia are routinely confused creating additional complexity in defining the role of oxygen in cell behaviours. This work has therefore evaluated the role of a panel of well-established HMAs (CoCl2, DFO, DMOG) and new agent (IOX2) vs. different reduced oxygen culture conditions. PC12 and hMSCs were both used to examinine the roles of HMAs on proliferation, metabolic activity, HIFs expression, nitroreductase activity, oxidative stress, apoptosis, death/necrosis and mitochondrial dynamic (burden, action potential and genome copy number) in comparison to physiological normoxia and intermittent hypoxia. HMAs induced HIF expression, apoptosis, trapped cell at G0/G1 phase and induced both ROS formation and nitroreductase activity in a manner that was not consistent with a reduced oxygen culture condition alone. Mitochondrial burden, mitochondrial action potential, and mitochondrial genome number also changed in response to HMA exposure in a manner that was indenependent of the oxygen culture conditions tested. In summary, HMAs do not provide an accurate replication of engineered oxygen control measures in PC12 and hMSC biology. This is reflected in biological alterations impacting on cell yield, behaviour and biology.

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Bowler, E. "The effect of hypoxia on alternative splicing in prostate cancer cell lines." Thesis, University of the West of England, Bristol, 2017. http://eprints.uwe.ac.uk/30029/.

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Hypoxia is defined as the state in which the availability or delivery of oxygen is insufficient to meet tissue demand. It occurs particularly in aggressive, fast-growing tumours in which the rate of new blood vessel formation (angiogenesis) cannot match the growth rate of tumour cells. Cellular stresses such as hypoxia can cause cells to undergo apoptosis; however some tumour cells adapt to hypoxic conditions and evade apoptosis. Tumour hypoxia has been linked to poor prognosis and to greater resistance to existing cancer therapies. This thesis provides evidence that alterations in alternative splicing patterns of key genes is one method tumour cells adapt to hypoxia. This study confirms a hypoxic-induced change in the alternative splicing of carbonic anhydrase IX (CA IX) following 1% oxygen treatment. CA IX is one of the best studied hypoxia markers, involved in maintaining an intracellular pH that favours tumour cell growth. Furthermore, evidence is provided here that in PC3 cells the regulation of CA IX splicing involves the SAFB1 and PRPF8 splice factors. Additionally, SAFB1 expression is shown to decrease in hypoxia. This study further demonstrates that alternative splicing patterns of previously documented cancer-associated genes are altered in hypoxia. PCR analysis showed that hypoxia significantly altered the alternative splicing of apoptotic-associated genes: caspase-9; Mcl-1; Bcl-x; survivin. The expression of the pro-apoptotic isoforms of the first two genes, and the anti-apoptotic isoforms of the latter two genes were favoured by hypoxia. Furthermore, high-throughput PCR analysis provided evidence of significant changes in the alternative splicing of several other cancer-associated genes in hypoxia: APAF1; BTN2A2; CDC42BPA; FGFR1OP; MBP; PTPN13; PUF60; RAP1GDS1; TTC23; UTRN. Most notably, the pro-oncogenic isoforms of APAF1, BTN2A2 and RAP1GDS1 were favoured in hypoxia. The majority of alternative splicing changes were found in the PC3 cell line. However changes in alternative splicing patterns that mirrored those in the PC3 cell line were also found in the VCaP (CDC42BPA, RAP1GDS1 and UTRN) and PNT2 (BTN2A2, CDC42BPA, FGFR1OP and TTC23) cell lines. The mRNA expression of splice factors (SRSF1, SRSF2, SRSF3, SAM68, HuR and hnRNP A1) and splice factor kinases (CLK1 and SRPK1) were shown to significantly increase in hypoxia. Subsequent experiments provided evidence that CLK1 and SRSF1 protein expression also increased in hypoxia. The phosphorylation of SRSF4 and SRSF5 were demonstrated to increase in hypoxia. However, the phosphorylation of SRSF6 was not. In addition, siRNAs and chemical inhibitors of CLK1 (TG003) and SRPK1 (SPHINX) were used to assess the effect of these splice factor kinases on the subsequent splicing of cancer-associated genes. There were no significant changes to splicing found with SRPK1 siRNA knockdown or SPHINX treatment. However CLK1 siRNA knockdown and TG003 treatment demonstrated a shift in FGFR1OP splicing that mirrored the effect of hypoxia on FGFR1OP splicing. This suggests that CLK1 activity is inhibited in hypoxia. Furthermore, in contrast to previous research CLK1 was found to be localised to the cytoplasm in both normoxia and hypoxia in the PC3 cell line. This work has uncovered factors and provided an insight into mechanisms that are involved in alternative splicing changes in hypoxia in mammalian cell lines. It is hoped that these novel research findings will aid in the understanding of how cells adapt to hypoxia especially in regards to alternative splicing, and may offer future therapeutic targets in hypoxic tumours.

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Gustafsson, Maria. "Signal integration between notch and hypoxia : insights into development and disease /." Stockholm, 2007. http://diss.kib.ki.se/2007/978-91-7357-090-9/.

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Barry, Michelle. "Beta cell viability and function in hypoxia : towards a clinically reflective model of beta cell transplantation." Thesis, University of Brighton, 2013. https://research.brighton.ac.uk/en/studentTheses/e738e627-bf59-4da9-8833-73982f74e209.

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Beta-cell survival is low following islet transplantation and this may be linked to a delay in revascularisation of donor cells. This decrease in oxygen supply is termed hypoxia, the result of which is detrimental to beta-cell survival. The current research sought to investigate post-transplant beta-cell viability and function by investigating the effects of low oxygen levels on MIN6 pancreatic beta-cells. MIN6 beta-cells were exposed to 1% oxygen (hypoxia) or 21% oxygen (normoxia) over a period of 72 hours. Viability was assessed by MTT assay and cell number was determined by haemocytometer count at 0 hours (normoxia), 24 hours, 48 hours and 72 hours. A Hoechst propidium iodide (HPI) stain was used to identify beta-cell apoptosis or necrosis during hypoxia. Western blot analyses were performed to determine the PI3K, pAkt, pAMPK, PDX-1, GLUT2 and pS6K protein levels. Real time PCR was used to estimate glut2, vegf, hif and insulin gene expression by MIN6 cells following exposure to hypoxia over various time points.

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Jensen, Gregory. "The Role of Serotonin (5-HT) in Regulating the Hypoxic Hyperventilatory Response of Larval Zebrafish." Thesis, Université d'Ottawa / University of Ottawa, 2016. http://hdl.handle.net/10393/35196.

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Serotonin (5-HT) containing neuroepithelial cells (NECs) are O2 sensitive chemoreceptors found throughout the skin of larval zebrafish (Danio rerio). Zebrafish larvae are sensitive to changes in ambient PO2 as early as 2 days post fertilisation (dpf) and hyperventilate in response to hypoxia beginning at 3 dpf. Tryptophan hydroxylase (tph) is the rate-limiting enzyme in 5-HT synthesis; three tph paralogs are present in zebrafish (tph1a, tph1b and tph2). Although 5-HT has been implicated as a key neurotransmitter mediating hypoxic hyperventilation, it has not been possible to discern the role of 5-HT specifically contained within the NECs in promoting hypoxic hyperventilation. The purpose of this study was to determine the role of NEC 5-HT in regulating the hypoxic ventilatory response in larval zebrafish. It was hypothesised that 5-HT is a key neurotransmitter released from NECs which contributes to hypoxic hyperventilation. Immunohistochemistry was used to determine the distribution of tph paralogs and their role in 5-HT production in NECs. Tph1a was present in NECs and nerves innervating NECs. Exposure to the non-selective tph inhibitor, para-chlorophenylalanine (pCPA), or translational gene knockdown of tph1a, diminished 5-HT expression within NECs. Exposure to acute hypoxia (PO2 = 30 mmHg) revealed a blunted hypoxic ventilatory response (reduced breathing frequency) in fish exhibiting depleted 5-HT in NECs. The hypoxic hyperventilatory response was rescued with application of 5-HT. The results of these experiments demonstrate that tph1a is responsible for 5-HT production in NECs of larval zebrafish, and that 5-HT released from NECs is involved in establishing their hypoxic hyperventilatory response.

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Bagnall, James Steven. "Single-cell imaging and mathematical modelling of the hypoxia-inducible factor signalling network." Thesis, University of Liverpool, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.569203.

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Hypoxia-inducible Factor (HIF) is an oxygen-sensitive transcription factor present in all multicellular life, including humans. It is responsible for mediating the cellular response to hypoxia, a situation in which oxygen supply falls below adequate levels. In the presence of oxygen HIF is constitutively degraded and inactivated. In the absence or reduction of oxygen, HIF escapes degradation and accumulates. Once accumulated, HIF elicits an adaptive response by inducing a diverse array of genes affecting the processes of cell fate, vascularisation and red blood cell production. However, there are many instances of the pathological activation of this pathway, which is known to cause serious and maladaptive issues, as found in cancer. HIF is functional as a heterodimer comprised of an Os-sensitive alpha subunit (HIF-α), of which there are two isoforms (-lα and -2α), and an Oe-insensitive beta subunit (HIF- β). The Os-dependent destabilisation of HIF-α is carried out by the prolyl hydroxylase domain proteins (PHDs). Importantly, several of the PHDs are themselves HIF target genes, thereby forming a delayed negative feedback loop. Delayed negative feedback loops are a common signalling motif able to confer dynamic properties to the signalling network. The resulting kinetics may have the capacity to encode additional information into the system and alter the course of downstream gene expression. The dissection and understanding of this requires a high-degree of characterisation of the temporal profile of response. Live-cell imaging offers an experimental system ideally suited to this approach. To date, the dynamics of HIF-l accumulation have not been investigated in this detail and so it was a main aim of this study to depict the temporal properties of the HIF signalling system. The dynamic response of HIF -a protein to hypoxia and re-oxygenation was imaged by live-cell time-lapse confocal microscopy. Interestingly, it was observed that the hypoxic accumulation of HIF-α was heterogeneous and transient. The duration of HIF- α accumulation was found to be a distinctive and robust temporal feature of the response. This assorted data was used in building a mathematical model depicting the HIF - PHD feedback motif. Subsequently, the model was utilised to accurately predict the consequence of loss of the PHD feedback. Additionally the model provided predictions on how HIF -α dynamics are differentially responsive to varied hypoxic and re- oxygenation conditions. Live-cell imaging of the HIF-α response also provided insight into the novel subcellular localisation pattern of the HIF-2α protein, which differs distinctly from HIF-la. HIF-2α was found to reside in several types of subcellular bodies. The dynamic interplay and flux of protein in and out of these bodies was investigated. Evidence gathered supports the involvement of HIF -2α in aggresomes and PML bodies and also the involvement of the subcellular landscape in the HIF regulatory network. Overall, research arising from this thesis offers new insights into the intracellular spatial and temporal dynamics of the HIF signalling system. 11

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