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De, Mattos Coelho Aguiar Juliana. "Mesenchymal potentials of the trunk neural crest cells." Phd thesis, Université Paris Sud - Paris XI, 2012. http://tel.archives-ouvertes.fr/tel-00982495.
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The neural crest (NC) derives from the dorsal borders of the vertebrate neural tube. During development, the NC cells migrate and contribute to the formation of different tissues and organs. Along the anteroposterior axis, the NC gives rise to neurons and glia of the peripheral nervous system and to melanocytes. Furthermore, the cephalic NC yields mesenchymal tissues, which form all facial cartilages and bones, the large part of skull, facial dermis, fat cells and smooth muscle cells in the head. In the trunk of amniotes Vertebrates, these tissues are derived from the mesoderm, not from the NC. In lower Vertebrates, however, the trunk NC generates some mesenchymal tissues, such as in the dorsal fins of zebrafish. The question therefore is raised whether the ability of the NC to produce mesenchymal cells was totally lost in the trunk of amniote Vertebrates during evolution, or if it can still be achieved under specific conditions. This work is interested in uncovering the mesenchymal potential of the avian trunk NC, with special interest in the differentiation into osteoblasts and adipocytes.Our experimental approach was to examine the skeletogenic and adipogenic differentiation potentials of quail trunk NC cells after in vitro culture. Cell differentiation was evidenced by the analysis of lineage-specific genes and markers using in situ hybridization (ISH), immunocytochemistry and RT-PCR. The established culture conditions allowed observation of both skeletogenesis and adipogenesis. Osteogenesis was initially characterized by expression of Runx2, the first transcription factor specific of the osteoprogenitors, which was detected by ISH from 5 days of culture. Later, we observed osteoblast maturation, with the expression of collagen1 protein, osteopontin mRNA and alkaline phosphatase mRNA, until the bone matrix mineralization stage. The trunk NC cells also underwent chondrogenesis, as demonstrated by Sox9, aggrecan and collagen10 mRNA expression, and Alcian blue staining. The observation of the mineralized areas and chondrogenesis suggested that the trunk NC cells in vitro are able to perform endochondral and membranous ossifications. In same culture conditions, the cells differentiated also into adipocytes, identified from 10 days of culture by Oil Red O staining. The mRNAs of the CEBP, PPAR and FABP4 adipogenic markers were detected by RT-PCR from 3 days of culture. For the characterization of bone and adipocyte progenitors, we evaluated the differentiation potential of individual trunk NC cells. The phenotypic analysis of these clonal cultures showed that 76% of the cells generated Runx2-positive osteoblasts. Moreover, most of the clone-forming trunk NC cells were multipotent progenitors endowed with both neural and osteogenic potentials. Furthermore, in another clonal culture condition, adipocytes were found in 35.3% of the clones, and approximately half of them also contained glial and/or melanogenic cells.These results show that the trunk NC cells in vitro are able to differentiate not only in their classical derivatives found in vivo (melanocytes, neurons and glial cells), but also in mesenchymal phenotypes, including adipocytes and osteoblasts. Importantly, as in cephalic NC cells, mesenchymal phenotypes differentiated from multipotent progenitor cells, suggesting that, during evolution, the NC stem cells intended for both mesenchymal and neural fates, had the expression of their mesenchymal potential inhibited in the trunk. Thus, although at the dormant state and not expressed in vivo, a significant mesenchymal potential is present in the trunk NC cells of amniotes Vertebrates and can be disclosed in vitro
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Allardyce, Joanna Marie. "Analysis of Wt1 expression in neural crest cells." Thesis, University of Liverpool, 2012. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.569213.
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The neural crest is a transient collection of cell, termed neural crest cells (NCCs), which develop during neurulation at the outer extremities of the neural folds between surface ectoderm and the developing neural tube. NCCs del aminate from the crest and migrate throughout the developing embryo and differentiate into many cell types such as melanocytes, peripheral neurons, osteocytes, muscle cells and enteric neurons and glia. With the use of a lineage tracing system (Wtl-Cre X Rosa26R mouse line) it was previously found that cells derived from Wtl-expressing cells have contributed to the post-natal enteric nervous system (ENS), indicating that Wtl must have been expressed in progenitor cells of the ENS during embryonic development. The goal of this project was therefore to identify when and where Wtl is expressed during this process. Data from immunofluorescence studies revealed that Wtl is transiently expressed in Sox10-expressing NCCs when they first begin their migration from the neural crest, at E8.5 in the mouse. Wt1 is then down-regulated in NCCs before they enter the foregut at E9.5. This data has been supported by in situ hybridisation studies, where Wtl has been found in cells of the neural crest at the same time point (E8.5) but Wtl mRNA was not shown to be present at any embryonic stage later than this. In vitro investigations were carried out in order to characterise Wtl in vagal level NCCs, as it is NCCs from this region (opposite somites 1-7) and the sacral neural crest (caudal to somite 28) which have been established as the origin of the ENS. NCCs were characterised by morphology and marker expression over a time-period of 7 days. The results from immunofluorescence experiments revealed co- expression of Wtl and NCC markers, SoxlO, up to 96 hours in culture. After this time-point it was no longer possible to detect these proteins in cultured explants. The neuronal marker ~III Tubulin was detected from 48 hours and was still found to be expressed at high levels after 96 hours when Wtl and NCC markers had ceased to be expressed, suggesting differentiation ofNCCs. Migration assays whereby the rate of migration was determined in NCCs in culture over a period of 48 hours revealed a mean migration rate of oum/hour. These data are relevant for future siRNA Wtl knock-down experiments in NCCs in vitro to investigate the effect of loss of Wtl function on migration rates, cell morphology, and expression patterns following preliminary experiments carried out on kidney stem cells.
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Ngamjariyawat, Anongnad. "The beneficial Effects of Neural Crest Stem Cells on Pancreatic β–cells." Doctoral thesis, Uppsala universitet, Institutionen för neurovetenskap, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-233157.
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Patients with type-1 diabetes lose their β-cells after autoimmune attack. Islet transplantation is a co-option for curing this disease, but survival of transplanted islets is poor. Thus, methods to enhance β-cell viability and function as well as methods to expand β-cell mass are required. The work presented in this thesis aimed to study the roles of neural crest stem cells or their derivatives in supporting β-cell proliferation, function, and survival. In co-culture when mouse boundary cap neural crest stem cells (bNCSCs) and pancreatic islets were in direct contact, differentiating bNCSCs strongly induced β-cell proliferation, and these proliferating β-cells were glucose responsive in terms of insulin secretion. Moreover, co-culture of murine bNCSCs with β-cell lines RIN5AH and β-TC6 showed partial protection of β-cells against cytokine-induced β-cell death. Direct contacts between bNCSCs and β-cells increased β-cell viability, and led to cadherin and β-catenin accumulations at the bNCSC/β-cell junctions. We proposed that cadherin junctions supported signals which promoted β-cell survival. We further revealed that murine neural crest stem cells harvested from hair follicles were unable to induce β-cell proliferation, and did not form cadherin junctions when cultured with pancreatic islets. Finally, we discovered that the presence of bNCSCs in co-culture counteracted cytokine-mediated insulin-producing human EndoC-βH1 cell death. Furthermore, these two cell types formed N-cadherin, but not E-cadherin, junctions when they were in direct contact. In conclusion, the results of these studies illustrate how neural crest stem cells influence β-cell proliferation, function, and survival which may improve islet transplantation outcome.
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Ballard, Victoria. "The contribution of extracardiac cells to the developing heart." Thesis, University of Surrey, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.250728.
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Okeke, Chukwuebuka. "Role of Nr2f Nuclear Receptors in Controlling Early Neural Crest and Ectomesenchyme Gene Regulation." University of Cincinnati / OhioLINK, 2021. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1627660719070357.
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Johnston, D. A. "The avian neural crest : behaviour and long-term survival in culture." Thesis, University of Southampton, 1986. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.376464.
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Nekooie, Marnany Nioosha. "The Intersection of Metabolism and Neural Crest Cell Development." Electronic Thesis or Diss., Paris 12, 2022. http://www.theses.fr/2022PA120066.
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Le métabolisme en tant que clé de voûte du destin des cellules souches fournit non seulement des demandes d'énergie et de molécules précurseurs, mais joue également un rôle dans le remodelage de la chromatine. Dans les embryons de vertébrés, les cellules de la crête neurale (NC) constituent une population remarquable de progéniteurs embryonnaires qui, lors de la délamination du tube neural dorsal, d'une migration et d'une différenciation étendues, donnent lieu à des dérivés neuraux/neuronaux et mésenchymateux. Le potentiel de différenciation des cellules NC nécessite un remodelage épigénétique et des signaux environnementaux. En conséquence, l'intersection du métabolisme et de la lasticité NC fournira des informations essentielles sur la régulation de l'identité et du développement des cellules NC. Ainsi, j'avais l'intention de comprendre le rôle du métabolisme dans l'aspect développemental d'une sous-population de cellules NC, le tronc NC. La première partie de mon étude a abouti à une vision générale des impacts métaboliques sur toutes les étapes de développement de la CN. J'ai mis en évidence que l'oxydation du glucose est un profil métabolique essentiel régissant la délamination, l'adhésion, la migration, la prolifération, le maintien de la tige et la différenciation généralisée des NC. Compte tenu de l'incidence de la transition G1 / S sur l'EMT dans les cellules NC du tronc, l'inhibition de la voie des pentoses phosphates (PPP) n'a pas pu influencer la délamination NC, suggérant une adaptation métabolique pour maintenir les étapes de développement et la survie. Par conséquent, dans l'étape suivante, j'ai cherché à apprécier comment les voies métaboliques s'intègrent dans la délamination NC. Le recâblage de la voie dela glycolyse sous inhibition du PPP au stade de délaminage a fourni un support pour les voies métaboliques multiples recrutées par les progéniteurs NC en réponse au stress métabolique. Mon étude a également élucidé la reprogrammation métabolique du PPP à l'oxydation du glucose dans les cellules NC du tronc, alignée sur la délamination NC à la transition migratoire. De plus, outre le glucose, la glutamine joue un rôle de premier plan dans l'acquisition pluripotente et la délamination des progéniteurs NC qui déclenchent la localisation nucléaire de la glutaminase (GLS) lors de l'étape de délaminage. Par conséquent, la localisation nucléaire du GLS lors de la délamination des cellules NC pré-igratoiressuggère la fonction de régulation du gène pour le GLS. Dans l'ensemble, mes résultats ont indiqué l'intersection du métabolisme et de la reprogrammation NC de l'étape pluripotente à l'engagement NC, définis respectivement par le PPP promu et la localisation nucléaire de GLS au phénotype OXPHOS à base de glucose avec localisation GLS cytoplasmique. De plus, l'interaction possible entre le GLS et la B-caténine a favorisé le nouveau concept sur la contribution du GLS à la signalisation Wnt, prometteuse pour comprendre l'étiologie de nombreuses neurocristopathiesMetabolism as a keystone of stem cells' fate not only supplies demands for energy and precursor molecules but also has roles in chromatin remodeling. In vertebrate embryos, neural crest (NC) cells constitute a remarkable population of embryonic progenitors, which upon delamination from dorsal neural tube, extensive migration and differentiation give rise to both neural/neuronal and mesenchymal derivatives. The developmental potential of NC cells necessitates epigenetic remodeling and environmental cues. Accordingly, the intersection of metabolism and NC plasticity will provide critical insights into the regulation of NC cell identity and development. Thus, I intended to figure out the metabolism role in the developmental aspect of one sub-population of NC cells, trunk type. The first part of my study resulted in a general view of the metabolic impacts on all developmental NC steps. I evidenced that glucose oxidation is a pivotalmetabolic profile governing NC delamination, adhesion, migration, proliferation, maintenance of stemness, and widespread differentiation. Given the incidence of G1/S transition upon EMT in trunk NC cells, the inhibition of pentose phosphate pathway (PPP) was unable to influence the NC delamination, suggesting a metabolic adaptation to maintain developmental steps and survival. Hence, In the next step, I sought to appreciate how metabolic pathways integrate into the NC delamination. The rewiring of glycolysis pathway under PPP suppression in delaminating stage provided support for multi metabolic pathways recruited by NC progenitors in response to the metabolic stress. My study also elucidated the metabolic reprograming from PPP to glucose oxidation in trunk NC cells, aligned with delaminating to migratory transition of these cells. Additionally, besides glucose, glutamine had a prominent role in pluripotent acquisition anddelamination of NC progenitors that triggers the nuclear localization of glutaminase (GLS) upon EMT step. Therefore, the nuclear GLS localization of pre-migratory NC cells in delaminating stage suggests the gene regulatory function for GLS. Altogether, my results indicated the intersection of metabolism and NC reprograming from pluripotent step to the NC commitment, defined respectively by promoted PPP and nuclear localization of GLS to glucose-based OXPHOSphenotype with cytoplasmic GLS localization. Moreover, the possible interaction between GLS and B-catenin fostered the new concept about the contribution of GLS to Wnt signaling, holding promise for understanding the etiology of many neurocristopathies
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Schock, Elizabeth N. B. S. "The Role of Primary Cilia in Neural Crest Cell Development." University of Cincinnati / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1504800027927076.
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Dickens, Claire Julia. "A study of ion regulatory mechanisms in neural crest cells and fibroblasts." Thesis, University of Newcastle Upon Tyne, 1990. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.287255.
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Rossi, Christy Cortez. "Early development of two cell populations at the neural plate border : rohon-beard sensory neurons and neural crest cells /." Connect to full text via ProQuest. Limited to UCD Anschutz Medical Campus, 2008.
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Thesis (Ph.D. in Neuroscience) -- University of Colorado Denver, 2008.Includes bibliographical references (leaves 112-120). Free to UCD affiliates. Online version available via ProQuest Digital Dissertations;
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McLennan, Rebecca. "Expression and function of EphA4 and ephrin-As in avian trunk neural crest migration /." free to MU campus, to others for purchase, 2004. http://wwwlib.umi.com/cr/mo/fullcit?p3144440.
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Darrigrand, Jean-François. "Influence of BMP signaling on neural crest cells during heart outflow tract septation." Electronic Thesis or Diss., Sorbonne université, 2019. https://accesdistant.sorbonne-universite.fr/login?url=https://theses-intra.sorbonne-universite.fr/2019SORUS085.pdf.
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La voie d’éjection cardiaque (VEC) est initialement un tube unique, qui se divise ensuite en artère aorte (Ao) et pulmonaire (Pa) au cours du développement embryonnaire. Cette morphogenèse est régulée par les cellules de crête neurale (CCN) qui colonisent la VEC et se condensent le long de l’endocarde, entraînant sa rupture et la formation des Ao et Pa. Des recherches ont montré que les Bone Morphogenetic Proteins (BMP) contrôlent le comportement des CCN durant ce processus. Cependant, les cascades moléculaires impliquées sont méconnues. Afin de mieux comprendre ces cascades moléculaires nous avons étudié le rôle de Dullard, une phosphatase identifiée comme inhibiteur intracellulaire de la voie BMP, au cours de la morphogenèse de la VEC. Nos résultats montrent que la délétion de Dullard dans les CCN augmente la signalisation BMP dans ces cellules, induit une septation asymétrique et prématurée de la VEC, l’obstruction de Pa et la mort embryonnaire. Cette suractivation de la voie BMP dans les CCN entraîne la diminution de leurs marqueurs mésenchymateux et augmente l’expression d’une cytokine appelée Sema3c, qui elle-même induit une compaction prématurée des CCN à l’endocarde. En parallèle, la différentiation asymétrique du myocarde sous-pulmonaire entraîne la rupture asymétrique de l’endocarde et l’obstruction de Pa. Enfin, nos résultats montrent qu’un gradient d’activation de la voie BMP et d’expression de Sema3c dans les CCN le long de l’axe de la VEC régule sa septation disto-proximale. Ainsi, nous mettons en évidence que la régulation des BMP dans les CCN orchestre spatiotemporellement la septation de la VECThe heart outflow tract (OFT) is originally a solitary tube, which is septated into the aortic and pulmonary artery (Pa) during embryonic development. This morphogenesis is regulated by the cardiac neural crest cells (cNCC), which colonize the OFT and condense towards the endocardium, triggering its rupture and the fromation of the two arteries. Investigations to identify the molecular cues controlling cNCC behaviour in the OFT mesenchyme have established the importance of the Bone Morphogenic Proteins (BMP). However, little is known on the molecular cascades triggered by BMP signaling responsible for the cNCC mediated OFT septation. To get insights into these molecular cascades, we decided to dissect the role of Dullard, a perinuclear phosphatase uncovered as a BMP intracellular signaling inhibitor, during OFT morphogenesis. Our results show that deletion of Dullard in the cNCC increases BMP intracellular signaling, leading to premature and asymmetric septation of the OFT, Pa obstruction and embryonic death. This BMP overactivation in the cNCC triggers the downregulation of mesenchymal markers and the upregulation of a cytokine called Sema3c, which in turn results in premature cNCC compaction at the endocardium. In addition, asymmetric differentiation of the distal subpulmonary myocardium contributes to asymmetrical rupture of the endocardium and Pa obstruction. Finally, our data converge to a model whereby graded BMP activity and Sema3c expression in the cNCC along the OFT axis set the tempo of OFT septation from its distal to its proximal regions. Hence, our findings reveal that fine tuning of BMP signaling levels in cNCC orchestrate OFT septation in time and space
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Frith, Thomas J. R. "Delineating the signals in anterior-posterior patterning of hPSC derived neural crest cells." Thesis, University of Sheffield, 2017. http://etheses.whiterose.ac.uk/19471/.
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Yang, Xiu. "ALTERED NEURONAL LINEAGES IN THE FACIAL GANGLIA OF Hoxa2 MUTANT MICE." Case Western Reserve University School of Graduate Studies / OhioLINK, 2008. http://rave.ohiolink.edu/etdc/view?acc_num=case1207189742.
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Mills, Alexandra Noelle. "Wolf-Hirschhorn Syndrome related genes are implicated in neural crest cell migration during development." Thesis, Boston College, 2018. http://hdl.handle.net/2345/bc-ir:108021.
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Thesis advisor: Laura Anne LoweryWolf Hirschhorn Syndrome (WHS) is a neurodevelopmental disorder characterized by craniofacial malformations, heart and skeletal defects, intellectual disability as well as seizure disorders. While this disorder is thought to arise from a deletion of a region on the short arm of chromosome 4, which includes the four genes WHSC1, WHSC2, LETM1 and TACC3, the mechanism by which loss of these genes results in WHS is not understood. Given that these genes have been linked to cell migration and that affected tissues include those derived from the neural crest, we propose that WHS results from a defect in neural crest cell migration. Here, we show that WHSC1, WHSC2, TACC3 and LETM1 are all expressed along the neural tube and developing neural folds during Xenopus embryonic development. These genes are additionally enriched in the pharyngeal arches, which are migrating neural crest cells. The knockdown of these WHS-related genes leads to variable defects in craniofacial and cartilage morphology. Moreover, the loss of WHS gene expression causes defects in forebrain and midbrain development. This implicates these four genes in the WHS phenotype. Further analysis of both WHSC1 and TACC3 function show that their individual knockdown causes defective neural crest cell migration both in vivo and in vitro. This supports the notion that the WHS phenotype is a result of erroneous neural crest cell motility. Our analysis shows that the WHS related genes; WHSC1, WHSC2, LETM1 and TACC3, play a role in the WHS phenotype of craniofacial malformation, skeletal abnormality, and microcephaly. Further analysis of these genes will determine the combinatorial effects of their knockdown on neural crest cell migration during embryonic development to further elucidate the mechanism through which WHS develops
Thesis (BS) — Boston College, 2018
Submitted to: Boston College. College of Arts and Sciences
Discipline: Departmental Honors
Discipline: Biology
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Fu, Ming, and 付明. "Neural crest cell development in the nervous system of normal gut and in Hirschsprung's disease." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2003. http://hub.hku.hk/bib/B29281970.
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McKenzie, Ian 1977. "The neural crest origins of skin-derived precursors : an accessible source of myelinating Schwann cells." Thesis, McGill University, 2005. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=100654.
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Skin-derived precursors are multipotent stem cells capable of differentiation into neural and mesodermal progeny. Described here is evidence that SKPs are of a neural crest origin; SKPs neural progeny are peripheral in subtype including peripheral catecholaminergic neurons and Schwann cells, SKPs express embryonic neural crest transcription factors, SKPs migrate via neural crest migratory pathways when transplanted into chick embryos and SKPs respond to factors know to influence neural crest stem cell differentiation in a similar manner. The expression of the neural crest transcription factors in situ identified the dermal papillae of hair follicles as a potential niche for SKPs in the skin. Cells isolated from microdissected vibrissae papillae generated spheres with SKPs properties when cultured under SKPs conditions and like SKPs, could be differentiated into neurons and smooth muscle. In agreement with the hypothesis that SKPs are of a neural crest origin, transgenic mice expressing beta-galactosidase in all neural crest progeny are characterized by beta-galactosidase expression in the dermal papillae, a potential SKPs niche and SKPs cultured from the whisker pads of these mice were also beta-galactosidase-positive. In order to demonstrate the functionality of SKP progeny as a proof of principle of their utility for potential therapeutic cell transplantation therapies, SKPs and SKP-derived Schwann cells were transplanted into various injury and disease models to assess their ability to myelinate. Cell culture protocols based on glial differentiation from the neural crest stem cell literature were adapted to selectively promote and expand Schwann cell differentiation from SKPs. SKP-derived Schwann cells transplanted into injured nerve were able to myelinate regenerating axons. SKPs and SKP-derived Schwann cells transplanted into congenitally dysmyelinated neonatal brains were able to form normal compact myelin. These data demonstrate the SKPs are a neural crest precursor that persists in adult skin that represents an accessible, autologous source of precursors capable of generating functional myelinating cells that could be used to treat disease and injury of the nervous system.
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Foster, K. E. "The role of neural crest cells in the development, organisation and migration of the thymus." Thesis, University College London (University of London), 2010. http://discovery.ucl.ac.uk/19301/.
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Neural Crest (NC) derived mesenchyme has previously been shown to play an important role in the early development of the foetal thymus. Using Wnt1-Cre and Sox10-Cre mice crossed to Rosa26eYfp reporter mice, NC derived mesenchymal cells were revealed in the adult murine thymus. It is reported here that NC derived cells infiltrate the thymus before E13.5, and differentiate into cells with characteristics of smooth muscle cells associated with large vessels, and pericytes associated with capillaries. In the adult organ at three months of age, NC derived perivascular cells continue to be associated with the vasculature providing structural support to the blood vessels and possibly regulating endothelial cell function. Thymus organogenesis requires co-ordinated interactions of multiple cell types including NC cells that orchestrate the formation, separation and subsequent migration of the developing thymus from the third pharyngeal pouch to the thoracic cavity. The molecular mechanisms driving these processes are unclear, however NC derived mesenchyme has been shown to be important. Here, it is shown that the separation process of the thymus from the pouch is independent of ephrin-B2 expression on thymic NC derived mesenchyme, however in its absence the thymus remains in the cervical area instead of migrating into the thoracic cavity. Analyses of individual NC derived thymic mesenchymal cells shows that the absence of ephrin-B2 impairs their polarisation, and thus motility, as a result of defective EphB receptor signalling. This implies a NC derived cell specific role of EphB-ephrin-B2 interactions in the collective migration of the thymic rudiment during organogenesis.
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Millington, Grethel. "Primary Cilia-dependent Gli Processing in Neural Crest Cells is Required for Early Tongue Development." University of Cincinnati / OhioLINK, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1479815997983138.
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Marathe, Himangi. "SWI/SNF Chromatin Remodeling Enzymes as Regulators of Neural-crest Derived Cell Differentiation." University of Toledo Health Science Campus / OhioLINK, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=mco1374233816.
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Yoshimatsu, Masayoshi. "In vivo regeneration of rat laryngeal cartilage with mesenchymal stem cells derived from human induced pluripotent stem cells via neural crest cells." Doctoral thesis, Kyoto University, 2021. http://hdl.handle.net/2433/265189.
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京都大学新制・課程博士
博士(医学)
甲第23417号
医博第4762号
新制||医||1052(附属図書館)
京都大学大学院医学研究科医学専攻
(主査)教授 松田 秀一特定拠点, 教授 妻木 範行, 教授 安達 泰治
学位規則第4条第1項該当
Doctor of Medical Science
Kyoto University
DFAM
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Ainsworth, Sophie Jane. "Avian development and disease : the importance of cranial neural crest cells in understanding cartilage repair strategies." Thesis, University of Brighton, 2009. https://research.brighton.ac.uk/en/studentTheses/3ab2e0e6-4897-4350-b5f4-09353049d1e0.
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Improving cartilage repair is a problem faced by scientists and clinicians alike who study and treat a wide variety of diseases including osteoarthritis (OA). OA has recently been seen with greater frequency in the western world due to the increasingly aging population. At present the possibility of using multipotent cells in treatments for OA is an exciting possibility that has been met with difficulties.
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Brindley, Gemma. "The role of cardiac neural crest cells in the development of the outflow tract of the heart." Thesis, University of Newcastle Upon Tyne, 2004. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.413391.
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Komatsu, Koji. "Meltrin β expressed in cardiac neural crest cells is required for ventricular septum formation of the heart." Kyoto University, 2007. http://hdl.handle.net/2433/135667.
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Losa, Llabata Marta. "Gene regulation in embryonic development." Thesis, University of Manchester, 2016. https://www.research.manchester.ac.uk/portal/en/theses/gene-regulation-in-embryonic-development(8a9efb79-1ca9-409e-89b9-9d66213e593f).html.
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Branchial arches (BAs) are a series of transient structures that develop on the ventro-lateral surface of the head in vertebrate embryos. BAs initially appear as a series of similar segments; as development proceeds each BA will contribute to different structures. Here, it was investigated the transcriptional mechanisms that instruct the different fates of the BAs in development. Initially, each BA contains a blood vessel, known as aortic arch (AA) artery, that connects the dorsal aorta with the heart. Remodelling of the AAs is crucial to form the adult heart circulation. This process leads to regression of the anterior AAs, running though the first and second BAs (BA1 and BA2), and persistence of the AAs contained in more posterior BAs (PBA). To identify the mechanisms that control remodelling of the AAs, we compared the transcriptomes and epigenomic landscapes of different BAs. Using RNA-seq and H3K27Ac ChIP-seq, we uncovered the activation of a vascular smooth muscle cell (VSMC) differentiation transcriptional program exclusively in the PBAs (and not in BA1/BA2). In support of this finding, we show that VSMC differentiation occurs specifically in the PBAs, but not BA1-2 in mouse embryonic development. Despite the absence of VSMC differentiation in developing BA1-2, cells harvested from these tissues reveal a spontaneous tendency to differentiate towards VSMC fate when grown in vitro, and activate several VSMC-specific genes (Myocd, Acta2, Tagln, Jag1). Together, our results suggest that forming VSMCs is a key process for the persistence of AAs. We also showed that cells derived from all BAs have the potential to differentiate to VSMCs in vitro. However, only cells in the PBAs differentiate to VSMCs in vivo, resulting in the maintenance of posterior AAs. In this study, we also uncovered a novel transcriptional principle that specifies the fate of BA2. Using ChIP-seq, we found that binding of Meis transcription factors establish a ground pattern in the BAs. Hoxa2, which specifies BA2 identity, selects a subset of Meis-bound sites. Meis binding is strongly increased at these sites, which coincide with active enhancers, linked to genes highly expressed in the BA2 and regulated by Hoxa2. Thus, Hoxa2 modifies a ground state binding of Meis to instruct segment-specific transcriptional programs.
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Philip, Beatrice. "An Investigation Into the Molecular Mechanisms and Genes Involved in Hypoxic Signalling Pathways in Neural Crest Derived Tumours." Thesis, Griffith University, 2014. http://hdl.handle.net/10072/367341.
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Hypoxia-inducing factors (HIF’s) are master regulators of metastasis in most solid tumours because of their ability to cause rapid cell proliferation, glycolytic metabolism, angiogenesis, and tumour invasiveness. Considerable progress has been made in defining HIF function in many hallmarks of carcinogenesis, including primary and metastatic tumour aggressiveness, poor prognosis, radio and chemotherapeutic resistance, vascular remodelling, inflammation, and hypoxia/ischemia induced cell damage. Hypoxic activation of HIF’s is gaining prominence in many cancer types as an essential mediator of malignant progression to metastasis. However, in neuroendocrine tumours of neural crest origin, hypoxia is often associated with dysfunction in genes encoding key enzymes of the TCA cycle leading to abnormally high levels of particular metabolites such as succinate that cause HIF stabilization.Thesis (PhD Doctorate)
Doctor of Philosophy (PhD)
School of Medical Science
Griffith Health
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Matthews, Helen Katherine. "A role for Syndecan-4 and PCP signalling in controlling directional migration of neural crest cells in vivo." Thesis, University College London (University of London), 2009. http://discovery.ucl.ac.uk/15955/.
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The neural crest (NC) is an embryonic population of cells, which delaminate from the neural tube epithelium to become vigorous migratory cells that colonise the entire embryo and give rise to many different derivatives. Neural crest migration requires activation of the non-canonical Wnt/planar cell polarity (PCP) signalling pathway, but it is not known exactly how this pathway controls cell migration. Here I show that the PCP ligand, Wnt11R, and the downsteam PCP element, Dishevelled, are essential for neural crest migration in Xenopus laevis embryos. Additionally, the proteoglycan, Syndecan-4, interacts with Dishevelled to control NC migration. A detailed examination of neural crest cell behaviour in Xenopus and zebrafish embryos shows that, in the absence of Dishevelled or Syndecan-4, cells are motile but lack the persistent migration that allows them to reach their target tissue. Furthermore, Dishevelled and Syndecan-4 control directional migration by regulating the polarised formation of cell protrusions. They also regulate the formation of paxillin-containing focal contacts in vitro and in vivo. Rho GTPase activity was measured using FRET analysis in neural crest cells migrating in vitro and in vivo after interfering with Syndecan-4/PCP signalling. I demonstrate that Syndecan-4 acts as a potent inhibitor of Rac, while Dishevelled activates RhoA. In addition, I show that RhoA inhibits Rac in neural crest cells. So, modulation of Rac by Syndecan-4 and PCP signalling allows the polarised formation of cell protrusions required for persistent NC migration. Finally I show that cell-cell contact inhibition of locomotion, dependent on PCP signalling, contributes to the initial polarity of the cell by inhibiting cell protrusions. Thus I present a model whereby neural crest cells are able to establish and maintain a directed migration by the integration of signals from cell-cell interactions mediated by PCP signalling and from the extracellular matrix via Syndecan-4.
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Ngamjariyawat, Anongnad, Kyril Turpaev, Svitlana Vasylovska, Elena N. Kozlova, and Nils Welsh. "Co-Culture of Neural Crest Stem Cells (NCSC) and Insulin Producing Beta-TC6 Cells Results in Cadherin Junctions and Protection against Cytokine-Induced Beta-Cell Death." Uppsala universitet, Neuroanatomi, 2013. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-198839.
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PURPOSE: Transplantation of pancreatic islets to Type 1 diabetes patients is hampered by inflammatory reactions at the transplantation site leading to dysfunction and death of insulin producing beta-cells. Recently we have shown that co-transplantation of neural crest stem cells (NCSCs) together with the islet cells improves transplantation outcome. The aim of the present investigation was to describe in vitro interactions between NCSCs and insulin producing beta-TC6 cells that may mediate protection against cytokine-induced beta-cell death. PROCEDURES: Beta-TC6 and NCSC cells were cultured either alone or together, and either with or without cell culture inserts. The cultures were then exposed to the pro-inflammatory cytokines IL-1β and IFN-γ for 48 hours followed by analysis of cell death rates (flow cytometry), nitrite production (Griess reagent), protein localization (immunofluorescence) and protein phosphorylation (flow cytometry). RESULTS: We observed that beta-TC6 cells co-cultured with NCSCs were protected against cytokine-induced cell death, but not when separated by cell culture inserts. This occurred in parallel with (i) augmented production of nitrite from beta-TC6 cells, indicating that increased cell survival allows a sustained production of nitric oxide; (ii) NCSC-derived laminin production; (iii) decreased phospho-FAK staining in beta-TC6 cell focal adhesions, and (iv) decreased beta-TC6 cell phosphorylation of ERK(T202/Y204), FAK(Y397) and FAK(Y576). Furthermore, co-culture also resulted in cadherin and beta-catenin accumulations at the NCSC/beta-TC6 cell junctions. Finally, the gap junction inhibitor carbenoxolone did not affect cytokine-induced beta-cell death during co-culture with NCSCs. CONCLUSION: In summary, direct contacts, but not soluble factors, promote improved beta-TC6 viability when co-cultured with NCSCs. We hypothesize that cadherin junctions between NCSC and beta-TC6 cells promote powerful signals that maintain beta-cell survival even though ERK and FAK signaling are suppressed. It may be that future strategies to improve islet transplantation outcome may benefit from attempts to increase beta-cell cadherin junctions to neighboring cells.
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Hjerling-Leffler, Jens. "Sensory neurons: stem cells and development /." Stockholm, 2006. http://diss.kib.ki.se/2006/91-7140-667-0/.
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Bradshaw, Lucy. "The-role of neural crest cells in the septation of the cardiac outflow and the effects of vitamin supplementation." Thesis, University of Newcastle upon Tyne, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.485859.
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Septation of the initially single cardiac outflow tract (OFT) into the distinct vessels of the aorta and pulmonary trunk is a crucial step in the development ofthe mature heart and is dependent upon two cell types; cardiac neural crest cells (NCe) and a contribution from the secondary heart field. Vitamin supplementation has been implicated as one of a number of environmental 'factors to influence the occurrence of congenital heart defects. Folic acid has been shown to rescue some neural tube and conotruncal heart defects whereas retinoic acid is critical for the formation of the outflow tract, but teratogenic in excess. The aim of the first part of this study was to investigate how the deficiency of NC~ in the Splotch2H mouse mutant causes OFT defects by. lineage tracing Wntl-expressing cells and how this may, in turn, affect the behaviour of SHF-derived cells. NCC's did not sufficiently colonise the caudal pharyngeal arches of Sp2H/Sp2H embryos, and as a consequence the 4th arch was completely absent. Reduced numbers of NCC were also observed in the dorsal wall of the aortic sac (aorto-pulmonary septum) and the proximal outflow cushion leading to a single, unseptated outflow vessel. This loss of NCC caused the ectopic localisation of Isletl-positive secondary heart field cells in the distal outflow tract, leading to abnormal myocardialisation and alignment defects. Thus, the outflow malformations seen in Sp2H/siH fetuses result from a combination of direct deficiency of NCC together with secondary effects on cells of SHF origin. Unravelling the relationship between these two cell types is likely to be of major importance in understanding the aetiology of malformations affecting the outflow region in humans. Secondly, the effects of supplementary exogenous vitamins on NCC were studied, to elucidate potential mechanisms for the rescue of neural-crest dependent OFT structures. Additional folic acid directly affected the NCC of Sp2H/Sp2H fetuses, exacerbating their intrinsic role to stabilise the developing pharyngeal arches, in particular the maintenance of right-sided arch structures. Retinoic acid did not alter the outflow tract defects ofsiH/sp2H fetuses, but did induce a variety of non-neural crest-dependent alignment/rotation defects in Sp2H/+ and wild-type fetuses at high doses, implicating the SHF as a potential target of Vitamin A teratogenicity. Although the exact mechanisms by which these vitamins exert their effects are still largely elusive, this study is an important step towards paving the way to primary prevention of a major group of congenital heart defects.
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Rabadán, Lozano M. Ángeles. "Genetic analysis of neural crest migration: Requirement of Dapper2-mediated inhibition of the Wnt canonical activity." Doctoral thesis, Universitat de Barcelona, 2012. http://hdl.handle.net/10803/83279.
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Numerous initiatives to improve our understanding of cancer biology have been lunched in different laboratories that aim to describe the interactome and gene-expression profile in different tumour cell line. It is now clear that the different strategies of cell migration observed in cancer are reminiscent of the different migratory strategies observed during embryo development. These similarities suggest that developmental program that has to be kept off after embryogenesis may be induced by spontaneous genetics modifications that produce tumour cells.
In this study we went inside the genetic network/profile that controls how neural crest cells eventually switch on the migration program and how they are able to arise into different lines with the propose of getting new ideas on how to prevent dissemination of tumour cells or how to treat advanced tumour that have already spread.
Neural progenitors of the dorsal neural tube that acquire the expression of specific neural crest determinants, delaminate from the neural tube and follow precise migratory pathways, to terminally differentiate into the various neural crest derivatives. Here we developed a novel resource for lineage trace and isolation of neural crest cells that allowed for a genome-wide expression screen in pre-migratory and migratory neural crest progenitors. We efficiently identified previously known neural crest specific genes. Expression profiling revealed new neural crest genes belonging to a wide range of cellular functions, with high representation of genes associated to cell motion. Additionally, we identified chick genes for which the human orthologues and/or paralogues are associated to Neuroblastoma formation.
In my thesis we identified new genes specifically expressed in the developing neural crest, and proposed a revised genomic signature for the normal neural crest cells. Furthermore, mutations on some of these genes are markers for Neuroblastoma tumour formation. Thus we propose this as a valid screen to identify candidates genes that contribute to the characterization of the Neuroblastoma cancer stem cells, and thus to the identification of specific targets to design new therapeutic strategies.
Getting in more detail into this genetic network, Wnt canonical signalling response has to been shown to be a key event in both cancer and neural crest cell development. Traditionally Wnt canonical pathway has been involved in neural crest induction process, but here we demonstrated that it is also critical for the onset migration of the neural crest cell. In fact, high levels of Wnt canonical activity prevents neural crest cell to delaminate and only through the inhibition of this activity mediate by dapper protein, neural crest cells can undergo into their normal migration pathways.
If this process has an implication in cancer is still unknown, but Dapper expression proteins have been already associated to different types of cancer.
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Poon, Hiu-ching. "A study of the regulatory roles of Hedgehog in the enteric nervous system development by the conditional knockout of Patched1 enteric gene in the enteric neural crest cells." Click to view the E-thesis via HKUTO, 2009. http://sunzi.lib.hku.hk/hkuto/record/B42841604.
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Zer, Heba al [Verfasser], and Ralf [Akademischer Betreuer] Smeets. "Enrichment and Characterization of Neural Crest - derived Dental Pulp Stem Cells from Human Dental Pulp / Heba Al-Zer. Betreuer: Ralf Smeets." Hamburg : Staats- und Universitätsbibliothek Hamburg, 2015. http://d-nb.info/106893154X/34.
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Poon, Hiu-ching, and 潘曉澄. "A study of the regulatory roles of Hedgehog in the enteric nervous system development by the conditional knockout of Patched1 entericgene in the enteric neural crest cells." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2009. http://hub.hku.hk/bib/B42841604.
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Acuña, Mendoza Soledad. "Sources alternatives de cellules souches pour la bio-ingénierie de la dent." Thesis, Sorbonne Paris Cité, 2015. http://www.theses.fr/2015USPCB101.
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Les cellules de la crête neurale (CN) sont une population de cellules multipotentes que pendant le développement embryonnaire vont migrer et se différencier vers divers lignages comme mélanocytes, muscle lisse, neurones périphériques et entériques, glie ainsi que tissus mésenchymateux cranio-faciaux y compris ceux de la dent. Dans le contexte de l’étude de modèles pour l’ingénierie tissulaire de la dent, nous avons établi une nouvelle lignée de cellules souches embryonnaires (ES) à partir de blastocystes issus de croisements entre un souris Wnt1-Cre et souris rapportrices fluorescentes, les Rosa26 mT/mT. Dans ce system, les cellules qui acquièrent l’identité CN et expriment le gène Wnt1 vont devenir fluorescentes grâce à l’activation de la protéine Tomato, ce qui permet de suivre 1) leur différenciation in vitro 2) isolement et 3) devenir lorsqu’elles sont utilisées dans de modèles in vivo. En parallèle, nous avons mis au point un nouveau protocole simplifié de différenciation (monocouche et milieu défini), vers un phénotype CN. Finalement nous avons tenté de développer un protocole d’induction d’une compétence odontogénique. Notre étude montre que la lignée Wnt1 Cre/Tomato 1) présentent toutes les caractéristiques d’une lignée ES classique i.e. expression de marqueurs de pluripotence, caryotype normal, capacité à se différencier in vitro et in vivo en tissus dérivés des 3 feuillets embryonnaires 2) acquièrent une identité CN, après induction in vitro avec notre protocole de différenciation. 3) Par l’intermédiaire de réassociations tissulaires in vitro, nous avons montré que ces cellules sont capables d’interagir avec un épithélium oral pour former des tissus squelettiques oro-faciaux. Ce nouvel outil cellulaire devrait aider à la compréhension des signaux impliqués dans le dialogue ectomésenchymateux qui sous-tend la formation des tissus durs de la face mais aussi plus généralement permettre suivir le devenir de cellules CN dans des modèles d’ingénierie tissulaireNeural crest cells are multipotent progenitor cells that, during embryogenic development, migrate and differentiate into diverse lineages such as melanocytes, smooth muscle, peripheral and enteric neurons, glial cells as well as craniofacial mesenchymatic components, including teeth. In the context of the development of an odontogenic model for tissue engineering, we have generated a new cell line of embryonic stem cells (ES) obtained from blastocysts from crossing Wnt1-CRE mice with fluorescent reporter Rosa26 mT/mT mice. In this Cre/Lox system the cells that have acquired a CN identity and thus expressing Wnt1, will become and remain fluorescent due to the activation of Tomato expression. We have generated a simplified protocol in a monolayer cell culture in defined serum-free medium in order to differentiate the cells into CN cells, named ES-CN cells. Second, we investigated the signals necessary for the odontogenic specification of these ES-CN cells. Our study provides evidence that the Wnt1-CRE/Tomato cell line 1) is a competent ES cell line with the expression of pluripotent markers, a stable karyotype and the ability to differentiate in vitro and in vivo into all the three embryonic germ layers, 2) acquires in vitro a CN identity after induction with our protocol, 3) expresses odontogenic markers in hypoxic culture conditions and 4) is able to interact with an oral epithelium in order to form orofacial skeletal tissues via the tissue reassociation in vitro. This novel cell model should facilitate the understanding of the mechanisms implicated in the ectomesenchymatic interaction, at the base for formation of orofacial skeletal tissues, and will provide the possibility to follow the fate of ES-CN cells tissue engineering models of wounded orofacial structures in general
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Sharma, Vipul. "Vangl2 as a key regulator of cell behaviour within the developing cardiac outflow tract : elaborating specific roles in second heart field and neural crest cells." Thesis, University of Newcastle upon Tyne, 2014. http://hdl.handle.net/10443/2611.
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ii Abstract Vangl2 is a key member of the multi-protein planar cell polarity (PCP) pathway. Previous studies using the loop-tail (Lp) mouse, which carries a mutation in the Vangl2 gene, have shown that PCP is required for normal development of the cardiac outflow tract. The main cell types involved in development of the outflow tract are neural crest cells (NCC) and cells derived from the second heart field (SHF). The PCP pathway plays important roles in polarisation of cells within tissues and in directional cell movements. I hypothesised that PCP signalling is required for efficient movement of progenitor cells into the developing heart and that an abnormality in these processes is sufficient to cause common outflow tract defects. Whilst loss of Vangl2 in NCC has no affect on outflow tract development, deletion of Vangl2 from SHF cells (using Vangl2flox crossed with Isl1-Cre mice) recapitulates the shortened outflow tract and malalignment defects seen in Lp mice. The cellular distribution of Vangl2 changes as SHF cells pass from a progenitor state, still expressing Isl1 protein, to differentiated myocardium. When Vangl2 is lost from the cells derived from the SHF, the cells within the distal walls of the outflow tract show altered localisation of polarised molecules such as β-catenin, fibronectin and laminin, as well as PCP proteins including Dvl2 and Celsr1, suggesting disrupted cellular polarity. The expression of PKCζ and E-cadherin is also altered in the distal outflow tract walls of Vangl2flox/flox;Isl1-Cre embryos, supporting the idea that Vangl2 may regulate the polarity of this tissue. Together, these studies suggest that Vangl2 plays a role in imparting polarity on SHF cells as they contribute to the outflow and that this is important for its lengthening. Confirmation of the importance of the PCP pathway in regulating the polarity of the cells in the distal outflow tract, and its importance for outflow tract development, was obtained by examining upstream components (Wnt5a and Ror2) and downstream targets (Rac1 and ROCK) of the pathway, showing outflow defects and a similar expression pattern of polarised molecules.
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Atashpazgargari, S. "A CELL REPROGRAMMING-BASED APPROACH TO STUDY 7Q11.23 GENE DOSAGE IMBALANCES IN WILLIAMS BEUREN SYNDROME AND AUTISM SPECTRUM DISORDER." Doctoral thesis, Università degli Studi di Milano, 2015. http://hdl.handle.net/2434/264765.
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Symmetrical gene dosage imbalances at 7q11.23 chromosomal region cause two unique neurodevelopmental diseases, Williams Beuren Syndrome (WBS) and the 7q11.23 microduplication associated to autistic spectrum disorder (7dup-ASD). Although both these diseases share common features such as intellectual disability and craniofacial dysmorphism, they can be distinguished by distinct social and language abilities: WBS patients characterized by hypersociality and comparatively well-preserved language skills while 7dup-ASD is associated with impairment in social interaction and communicative skills. The involvement of same genetic interval in these disease, points out to small subset of dosage-sensitive genes affecting cognition, social behavior and communication skills.
Among the genes in the deleted region, some were shown to contribute to the abnormalities in these patients through transgenic mice models and individual case reports. However, the precise cellular and molecular phenotypes associated with these syndromes in disease-relevant cell-types are unknown due to the scarce availability of primary diseased tissues. Transcription factor induced somatic cell reprogramming has bypassed such fundamental limitation and has enabled us to model human diseases, elucidate their pathogenesis and discover new therapeutics by screening small chemicals/drugs on these models. During my PhD studies, I focused on the functional dissection of these complementary diseases at the level of transcriptional deregulation in patient-derived iPSC and its differentiated derivatives such as neural crest stem cells, mesenchymal stem cells, and neural progenitors. To this end, we have assembled a unique cohort of typical WBS, atypical WBS (patient with a partial deletion) and 7dup-ASD patients (along with unaffected relatives), and then I used mRNA reprogramming to establish and characterize at least 3 independent iPSC lines from a total of 12 individuals. High throughput mRNA sequencing on iPSC revealed critical transcriptional derangements in disease-relevant pathways already at the pluripotent state. These alterations found to be selectively amplified upon differentiation into disease-relevant lineages, thereby establishing the value of large iPSC cohorts in the elucidation of disease-relevant developmental pathways. Finally, we created an open-access web-based platform to make accessible our multi-layered datasets and integrate contributions by the entire community working on the molecular dissection of the 7q11.23 syndromes.
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Repele, Andrea. "Differentiation potential and metabolic analysis of satellite cells and amniotic fluid stem cells." Doctoral thesis, Università degli studi di Padova, 2012. http://hdl.handle.net/11577/3422458.
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We have recently characterized two distinct populations of Satellite Cells (SCs), defined as Low Proliferative Clones (LPC) and High Proliferative Clones (HPC), that differ for proliferation, egenerative potential and mitochondrial coupling efficiency. In here, we have deep investigated their cell biology and characterized features that remark their intrinsic differences retrievable also at the initial phases of their cloning. LPC and HPC can indeed be istinguished for characteristic mitochondrial membrane potential (ΔΨm) just after isolation from their parental fibre. This is merged by mitochondrial redox state measured via NAD+/NADH analysis- and alternative respiratory CO2 production in cloned cells, which are accountable for metabolic differences reflected by alternative expression of the glycolytic enzyme Pfkfb3. In addition also mitochondrial Ca2+ handling and the sensitivity to apoptosis triggered via the intrinsic pathway are modified as well as the size of the mitochondrial network. In conclusion, we were able to determine which clone represents the suitable stem cell within the SCs population. These further experimental observations report novel
physiological features in the cell biology of SCs populations before and after cloning, highlighting an intrinsic heterogeneity on which the stemness of the satellite cell is likely to depend.
In the second part of my work we have also investigated their potential to trans-differentiate into smooth muscle cells. Enteric Nervous System normally interacts with muscle cells to control the peristaltic and secretory activity of the gut wall. Incomplete gut colonization by neural crest cells causes Hirschsprung’s disease, characterized by aganglionosis of the distal bowel. Multipotent, self-renewing enteric precursor neurosphere-like bodies (NLBs) -capable of generating neurons and glia derived from the neural crest- can be isolated from the gut of mice, rats, and human and they are able to colonize the gut after transplantation. Our aim is to understand the relationship between satellite cells-derived muscle precursor cells (MPCs) and NLBs using an in vitro co-culture model: this will be useful in perspective of a tissue engineering approach for bowel regeneration and skeletal muscle. Our records highlighted that NLBs were able to form new myotubes in presence of MPCs. Co-cultures in myogenic medium showed a remarkable improvement of MPCs ifferentiation by NLBs, promoting the formation of sarcomeric striatures onto myotubes and increasing the desmin expression of MPCs. On the other side, using neurogenic medium MPCs-NLBs showed a neural-like phenotype. As future perspectives, we need to understand the relationship between MPCs and NLBs and if the synapses are involved in this process; to verify if the seeding on a biocompatible polymer influences the behaviour of neural cells; and we must confirm these data with an in vivo skeletal and smooth muscle differentiation.
We have finally explored the possibility of deriving smooth muscle cells from a different source, taking in consideration the difficulties related to the expansion of both skeletal and smooth muscle progenitors. Therefore, we aim to derive functional smooth muscle cells (SMCs) from non-muscle cells, such as human Amniotic Fluid Stem (hAFSC) cells. hAFSC were transduced using vector encoding ZsGreen under the αSMA promoter. SMhAFSC expressed significantly higher level of smooth muscle genes (such as αSMA, desmin, calponin and smoothelin expression) after selective culture condition. These features were confirmed by immunofluorescence, demonstrating a single lineage commitment; TEM established increased
intermediate filaments, dense bodies and glycogen deposits in SMhAFSC, similar pattern compared to SMCs; and sequential imaging analyses demonstrated that SMhAFSC have a higher contractile potential than hAFSC. Consecutive single cell sampling showed the presence of voltagedependent calcium activated potassium channels on differentiated SMhAFSC and showed a higher production of carbon dioxide. In conclusion, we were able to generate to functional SMCs starting from a non-muscle precursor; secondly the transduction process may represent a valuable tool to select SM committed population. This step may eventually overcome the well-known problem of expanding SM progenitors, making these cells amenable to tissue engineering.Il nostro gruppo ha recentemente caratterizzato due distinte popolazioni di cellule satelliti, classificate come cloni a bassa proliferazione (LPC) e ad alta proliferazione (HPC), che si differenziano in termini di proliferazione, potenziale rigenerativo e metabolismo mitocondriale. Nel mio lavoro di dottorato, abbiamo valutato e caratterizzato la loro biologia cellulare con particolare attenzione a quelle differenze intrinseche presenti anche prima della loro clonazione. Infatti, ambo le tipologie clonali possono essere distinte mediante il potenziale di membrana mitocondriale (ΔΨm) subito dopo l’isolamento dalla fibra. Questo dato è in accordo con lo stato ossido riduttivo mitocondriale misurato tramite NAD+/NADH e la quantificazione della produzione di CO2. Questi risultati sono responsabili delle differenze metaboliche e possono essere spiegati dalla diversa espressione dell’enzima glicolitico Pfkfb3. Inoltre la concentrazione mitocondriale del Ca2+ e la sensibilità all’apoptosi sono modificate così come la dimensione della rete mitocondriale. In conclusione, siamo stati in grado di determinare quale clone rappresenta la cellula staminale all’interno della popolazione di cellule satelliti. Queste nuove osservazioni sperimentali rivelano caratteristiche fisiologiche della biologia delle popolazioni delle cellule satelliti prima e dopo la clonazione, mettendo in luce un’eterogeneità intrinseca della cellula satellite. Nella seconda parte della mia tesi abbiamo esplorato la possibilità che le cellule satelliti possano, se opportunamente stimolate, trans-differenziarsi in cellule muscolari lisce. Il sistema nervoso enterico normalmente interagisce con le cellule muscolari per controllare l’attività peristaltica e secretoria della parete intestinale. L’incompleta colonizzazione dell’intestino da parte delle cellule della cresta neurale provoca la malattia di Hirschsprung, caratterizzata da aganglionosi del colon distale. Le neurosfere (NLBs), precursori enterici in grado di auto-rinnovarsi, possono generare neuroni e glia; essere isolate dall’intestino di topi, ratti e umani e sono in grado di colonizzare l'intestino dopo il trapianto. Il nostro obiettivo è di capire la relazione tra i precursori di cellule satelliti (MPCs) e NLBs utilizzando un modello in vitro di co-coltura: questo sarà utile in prospettiva di un approccio di ingegneria tissutale per la rigenerazione intestinale e muscolo scheletrico. I nostri dati hanno evidenziato che NLBs, in presenza di MPCs, sono in grado di formare nuovi miotubi. L’uso di terreni di coltura miogenici ha evidenziato un notevole aumento della differenziazione in senso muscolare, promuovendo la formazione di striature ed aumentando l’espressione di desmina. Dall’altra parte, l’utilizzo di terreni di coltura neurogenici ha mostrato un fenotipo simil neurale. Come prospettive future, dobbiamo comprendere ulteriormente la relazione tra MPCs e NLBs e se le sinapsi sono coinvolte in questo processo; si deve verificare se un loro utilizzo su polimeri biocompatibili ne possa influenzare il comportamento, ed infine è necessaria una conferma dei suddetti dati tramite un’analisi di differenziazione in vivo in muscolo scheletrico e liscio. Nella terza ed ultima fase del mio lavoro, ci siamo focalizzati ad esplorare la possibilità che cellule non-muscolari possano, se opportunamente stimolate, differenziare in senso muscolare liscio. Il nostro obiettivo è stato quello di ottenere cellule muscolari lisce (SMCs) partendo da cellule staminali del fluido amniotico umano (hAFSC). hAFSC sono state trasdotte utilizzando un virus codificante per ZsGreen sotto il promotore αSMA. SMhAFSC così ottenute hanno evidenziate un alto livello d’espressione dei geni del muscolo liscio (come αSMA, desmina, calponina e smoothelin). Queste caratteristiche sono state confermate da molteplici analisi: di immunofluorescenza, dimostrando la positività a marcatori specifici per il muscolo liscio; microscopia a trasmissione elettronica (TEM), dove si verificava l’aumento della presenza di filamenti intermedi, di corpi densi e depositi di glicogeno, modello simile rispetto alle SMCs. Analisi in timelapse di SMhAFSC hanno dimostrato che queste possiedono un potenziale contrattile superiore rispetto hAFSC e studi su singola cellula hanno evidenziato la presenza di canali calcio voltaggio-dipendenti attivati da potassio solamente su SMhAFSC. In conclusione, siamo stati in grado di generare di cellule muscolari lisce funzionali da un precursore nonmuscolare ed in secondo luogo il processo di trasduzione può rappresentare un valido strumento per distinguere e selezionare differenti popolazioni. Questa fase può eventualmente superare il ben noto problema dell’espansione di progenitori di cellule muscolari lisce, rendendo queste cellule suscettibili per approcci d’ingegneria tessutale.
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Schizas, Nikos. "Neuroprotection in the Injured Spinal Cord : Novel Strategies using Immunomodulation, Stem cell Transplantation and Hyaluronic acid Hydrogel carriers." Doctoral thesis, Uppsala universitet, Ortopedi, 2015. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-251477.
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The overall aim of this thesis was to establish strategies to minimize secondary damage to the injured spinal cord. Secondary damage that follows spinal cord injury (SCI) involves inflammatory and excitotoxic pathways. Regulation of these pathways using immunomodulatory and neuroprotective substances potentially protects the injured spinal cord from further damage. We also developed and studied resorbable biomaterials to be used as carriers for potential neuroprotectants to the injured spinal cord. We used transversal spinal cord slice cultures (SCSCs) derived from postnatal mice as a model. SCSCs were maintained on different biomaterials and were studied after treatment with immunomodulatory and/or neurotrophic factors. They were further excitotoxically injured and subsequently treated with interleukin-1 receptor antagonist (IL1RA) or by neural crest stem cell (NCSC)-transplantation. The results show that biocompatible and resorbable hydrogels based on hyaluronic acid (HA) preserved neurons in SCSCs to a much higher extent than a conventional collagen-based biomaterial or standard polyethylene terephthalate (PET) membrane inserts. Glial activation was limited in the cultures maintained on HA-based hydrogel. The anti-inflammatory factor IL1RA protected SCSCs from degenerative mechanisms that occur during in vitro incubation, and IL1RA also protected SCSCs from excitotoxic injury induced by N-Methyl-d-Aspartate (NMDA). IL1RA specifically protected neurons that resided in the ventral horn, while other neuronal populations such as dorsal horn neurons and Renshaw cells did not respond to treatment. Finally, transplantation of NCSCs onto excitotoxically injured SCSCs protected from neuronal loss, apoptosis and glial activation, while NCSCs remained undifferentiated. The results presented in this thesis indicate that carriers based on HA seem to be more suitable than conventional collagen-based biomaterials since they enhance neuronal survival per se. The observed neuroprotection is likely due to biomechanical properties of HA. IL1RA protects SCSCs from spontaneous degeneration and from NMDA-induced injury, suggesting that excitotoxic mechanisms can be modulated through anti-inflammatory pathways. Different neuronal populations are affected by IL1RA to various degrees, suggesting that a combination of different neuroprotectants should be used in treatment strategies after SCI. Finally, NCSCs seem to protect SCSCs from excitotoxic injury through paracrine actions, since they remain undifferentiated and do not migrate into the tissue during in vitro incubation. It seems that combinations of neuroprotectants and carrier substances should be considered rather than one single strategy when designing future treatments for SCI. Incorporation of neuroprotectants such as IL1RA combined with stem cells in injectable biocompatible carriers based on HA is the final goal of our group in the treatment of SCI.
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Ferronha, Tiago Guimaràes. "The contribution of LMO4 to neural development: generatin neuronal subtypes in the dorsal spinal cord and controlling EMT in neural crest cells and Neuroblastoma = La contribución de LMO4 al desarrollo neural: la generación de subtipos neuronales de la médula dorsal espinal y el control de la EMT en las células de la cresta neural y de Neuroblastoma." Doctoral thesis, Universitat de Barcelona, 2013. http://hdl.handle.net/10803/127186.
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The formation of the central nervous system (CNS) requires the generation of a remarkable diversity of neurons, which must be produced in adequate amounts in the location and timing during development. Studies on the development of the spinal cord, the caudal and simpler anatomically vertebrate CNS, have shown that the neuronal diversity is progressively acquired through the processes of cell type specification. In amniotes, patterning of the neural tube along the dorsal-ventral axis generate 11 distinct domains of neural progenitors. Thus, the developing spinal cord, may be subdivided into 6 dorsal domains of progenitors (dP1-6, from dorsal to ventral) and a ventral part consisting of 5 domains (p3, pMN, p2-0, of ventral to dorsal). Dorsal and ventral domains of progenitors generate classes spinal neurons involved in sensory and motor circuits, respectively. Thus, accurate identification of each of the molecules that regulate this process is crucial. First, in this study, we focus on the function of LMO4 during neurogenesis in the development of the dorsal spinal cord. Using electroporation in ovo, in chicken embryos, as a method for modulating the activity of LMO4 in vivo, we demonstrated that LMO4 is dispensable for maintaining patterns progenitor cells, but is necessary for proper generation of discrete classes of interneurons ie , dI1/3/5. Further we provided evidence that LMO4 activity is essential for the promotion of these identities by the canonical BMP activity and suggest that LMO4 and SMAD1 / 5 contribute to the multi-protein complexes that regulate the genetic program for the specification of interneurons dI1 / 3/5.
In the second part of this thesis, we study the peripheral nervous system, PNS, and one type of cancer that is derived from the PNS. Neuroblastoma (NB) is a neurological tumor that arises from neural crest cells (NC) and is the most common extracranial tumor in children. NB represents a very heterogeneous group of tumors in terms of biological, genetic and morphological characteristics. Clinically, these tumors can develop differently, since spontaneous remission differentiation or apoptosis (NC behavior resembling normal) to aggressive metastatic disease with low rates of overall survival. The neuroectodermal origin of NB suggests that these tumors can spread from its primary site using mechanisms similar to those involved in the delamination and dispersion of embryonic neural crest cells, and genes involved in cell migration of neural crest cells may also be involved in the acquisition of the phenotype of invasive NB. We developed marker for neural crest lineage and taking the hypothesis that the molecular mechanisms that mediate the neural crest delamination may also be involved in the spread of neuroblastoma, we were able to identify genes that are expressed in the neural crest development and formation of neuroblastoma. A subsequent search in the NBGS (neuroblastoma gene server) for the human orthologous of genes differentially expressed in neural crest chicken embryo, retrieved LMO4 which was expressed in both cell types tested. Functional experiments in these two systems, revealed that the activity LMO4 is required for delamination of the neural crest and for neuroblastoma cell invasion. Moreover, LMO4 identified as an essential cofactor in the cadherin repression mediated Snail2 and epithelial to mesenchymal transition from neural crest cells and neuroblastoma. Together, our results suggest that the association of high levels of LMO4 with aggressive neuroblastomas, depends on the regulation of cadherin expression by LMO4 and therefore tumor invasiveness.La formación del sistema nervioso central requiere la generación de una gran diversidad de neuronas, y la identificación de las señales moleculares que regulan este proceso es por lo tanto crucial. En esta tesis, me he centrado primero en estudiar la función de LMO4 durante la neurogénesis en la médula espinal dorsal. Mediante las electroporación in ovo, en embriones de pollo, como una metodología para modular la actividad LMO4 in vivo, se demostró que LMO4 era prescindible para el mantenimiento de los patrones de progenitores, pero que se requiere para la correcta generación de subtipos concretos de interneuronas dorsales (dI1/3/5). Por otra parte, se demostró que la actividad LMO4 es esencial para la promoción de estas identidades mediante la actividad BMP canónica, y proponemos que LMO4 y Smad1 / 5 forman parte de los complejos de proteínas que impulsan el programa genético de especificación de dI1/3/5. En la segunda parte de la tesis, he estudiado el sistema nervioso periférico, así como tumores que surgen de él. El neuroblastoma es un tumor embrionario derivado de células de la cresta neural. Aprovechando un nuevo marcador desarrollo para el linaje de cresta neural y en base a la hipótesis de que los mecanismos moleculares que median la delaminación de la cresta neural también están involucrados en la propagación del neuroblastoma, identificamos varios genes comunes en el desarrollo de la cresta neural y el neuroblastoma. Una búsqueda posterior de los ortólogos humanos del NBGS (genes expresados en neuroblastoma) observamos que LMO4 se expresa en ambos tipos de células analizadas. Experimentos funcionales en estos dos sistemas modelo revelaron que se requiere la actividad LMO4 durante la invasión de células del neuroblastoma y durante la delaminación de la cresta neural. Además, identificamos LMO4 como un cofactor esencial de Snail2, en la represión de la expresión de cadherina, y por tanto en la transición epitelio mesenchyma de la cresta neural y de las células de neuroblastoma. En conjunto, nuestros resultados sugieren que la asociación de altos niveles de LMO4 con neuroblastomas agresivos depende de la regulación de cadherina y, por tanto, de la invasividad tumoral.
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Loison-Robert, Ludwig. "Cellule souche gingivale : origine et multipotence." Thesis, Paris Est, 2016. http://www.theses.fr/2016PESC0083/document.
Full textAbstract:
La gencive correspond à un modèle de régénération naturelle grâce notamment à sa capacité de cicatrisation « ad integrum ». Ce phénomène est permis par sa composition en fibroblastes gingivaux. Ces cellules, composante cellulaire principale du tissu conjonctif gingival, sont au cœur de la régulation des réponses inflammatoires et de la cicatrisation. Ce tissu contient, comme d’autres tissus mésenchymateux, des cellules souches ; qui expliquent en partie ces capacités de régénération. De plus, comme le tissu gingival est abondant et facilement accessible, l’utilisation de ces cellules souches pourraient être d’un intérêt prometteur en thérapie cellulaire ou pour de la modélisation in vitro. Au cours de cette thèse, nous avons pu montrer que les Cellules Souches dérivées de la Gencive Humaine (CSGH) possèdent des propriétés communes avec les cellules souches adultes dérivées des crêtes neurales. Ces cellules peuvent être qualifiées de « souche » par leur capacité d’auto-renouvèlement, d’adhésion au plastique et de multipotence. Premièrement, nous avons montré que la méthode ainsi que les produits de culture utilisés pour l’isolation des fibroblastes gingivaux in vitro à partir de biopsies de gencive avait une influence sur les cellules obtenues. Dans un second temps, une analyse clonale in vitro de populations de fibroblastes gingivaux a permis de montrer que les fibroblastes gingivaux sont composés de sous-populations qui expriment des marqueurs spécifiques des cellules souches et des crêtes neurales. Outre leur origine embryologique, l’étude de leur multipotence a aussi été caractérisée après expansion et en fonction des additifs utilisés. Pour finir, deux exemples d’utilisation de ces cellules comme modèle d’étude de la biocompatibilité de biomatériaux in vitro ont été développés; imitant la muqueuse buccale ainsi que les réactions dentaires (réparatrices et réactionnaire)Gingiva is a natural regeneration model thanks to its "ad integrum" healing capability. Gingival fibroblasts are the main actors of this property. These cells, the main cellular component of the gingival connective tissue, regulate the inflammatory responses and healing process. This tissue contains, like many others, mesenchymal stem cells; which also partly explain these regenerative abilities. Moreover, as the gingiva is abundant and easily accessible, the use of these stem cells may interest cell therapy or in vitro model tissues responses. In this work, we demonstrated that Stem Cells Derived from Human Gingiva (SCHG) have common properties with neural crest adult stem cells. These cells can be called "stem cells" for their ability to self-renew, adhere to plastic and to differentiate. First, we have shown that the method and the culture products used for isolation of gingival fibroblasts from gingival biopsy had an influence on the obtained cells. Secondly, an analysis of in vitro clonal populations of gingival fibroblasts has shown that gingival fibroblasts are composed of subpopulations that express specific markers of stem cells and neural crests. In addition to their embryological origin, the study of their multipotency was also characterized after expansion and depending on the used additives. Finally, two examples of using these cells and dental pulp stem cells as a model to study the in vitro biocompatibility of biomaterials have been developed, mimicking oral mucosa or dentin reactions (reparative or reactional)
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Workman, Michael J. "Generating 3D human intestinal organoids with an enteric nervous system." University of Cincinnati / OhioLINK, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1416570664.
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Grapensparr, Liza. "Auxiliary Cells for the Vascularization and Function of Endogenous and Transplanted Islets of Langerhans." Doctoral thesis, Uppsala universitet, Integrativ Fysiologi, 2017. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-327314.
Full textAbstract:
Type 1 diabetes develops through the progressive destruction of the insulin-producing beta-cells. Regeneration or replacement of beta-cells is therefore needed to restore normal glucose homeostasis. Presently, normoglycemia can be achieved by the transplantation of whole pancreas or isolated islets of Langerhans. Islet transplantation can be performed through a simple laparoscopic procedure, but the long-term graft survival is low due to poor revascularization and early cell death. This thesis examined the possibility of using different auxiliary cells (Schwann cells, endothelial progenitor cells, and neural crest stem cells) to improve the engraftment and function of endogenous and transplanted islets. Co-transplantation of Schwann cells with islets improved islet graft function early after transplantation, and caused an increased islet mass at one month posttransplantation. However, the vascular densities of these grafts were decreased, which also related to an impaired graft function. Islet grafts containing endothelial progenitor cells had a superior vascular density, with functional chimeric blood vessels and substantially higher blood perfusion and oxygen tension than control transplants. By culturing and transplanting islets together with neural crest stem cells it was found that islets exposed to these cells had a higher beta-cell proliferation compared with control islets. At one month posttransplantation, the grafts with neural crest stem cells also had a superior vascular- and neural density. The potential of intracardially injected neural crest stem cells to home to the pancreas and ameliorate hyperglycemia in diabetic mice was investigated. During a three-week period after such cell treatment blood glucose concentrations decreased, but were not fully normalized. Neural crest stem cells were present in more than 10% of the pancreatic islets at two days postinjection, at which time the beta-cell proliferation was markedly increased when compared with islets of saline-treated diabetic animals. Three weeks later, a doubled beta-cell mass was observed in animals receiving neural crest stem cells. In summary, islets can easily be transplanted together with different auxiliary cells. Some of these cells provide the possibility of improving vascular- and neural engraftment, as well as beta-cell growth and survival. Systemic administration of neural crest stem cells holds the potential of regenerating the endogenous beta-cells.
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Latta, Elizabeth Janet. "Studies of the tissues and molecules that regulate the migration of cranial neural crest cells in the chicken embryo : roles of Midline 1 and retinoic acid." Thesis, Open University, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.527449.
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Oldenburg, Denise [Verfasser], Beate [Gutachter] Brand-Saberi, and Carsten [Gutachter] Theiß. "The role of the cephalic neural crest cells in the development of the limbal stem cell niche of the chicken cornea / Denise Oldenburg ; Gutachter: Beate Brand-Saberi, Carsten Theiß ; Medizinische Fakultät." Bochum : Ruhr-Universität Bochum, 2018. http://d-nb.info/1171521901/34.
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Gazquez, Elodie. "Études des interactions fonctionnelles entre l'endothéline-3, les intégrines beta1 et les propriétés élastiques du tissu embryonnaire au cours du développement du système nerveux entérique." Thesis, Paris 6, 2016. http://www.theses.fr/2016PA066240/document.
Full textAbstract:
Le système nerveux entérique (SNE) provient des cellules de crête neurale entériques (CCNEs) qui migrent au sein de l'intestin embryonnaire, prolifèrent et se différencient en cellules gliales et neurones formant des ganglions interconnectés. Mon projet de thèse vise à comprendre comment les propriétés biochimiques et mécaniques de l'intestin embryonnaire influencent la colonisation et la différenciation des ccnes. L'absence d'endothéline-3 (EDN3), un facteur biochimique exprimé dans la paroi intestinale, est une des causes de la maladie de hirschsprung, caracterisée par une aganglionose du côlon distal. Nous montrons pour la première fois que l'EDN3 stimule l'adhésivité des CCNEs en augmentant leurs adhérences focales dépendantes des intégrines beta1 ainsi que la dynamique de leurs protrusions membranaires. De plus, nous avons mis en évidence l'existence d'une interaction génétique entre Edn3 et Itgb1 gouvernant le développement du SNE. Par ailleurs, les propriétés mécaniques du microenvironnement influençant la migration et la différenciation cellulaire , nous avons analysé par des approches biophysiques les propriétés élastiques de l'intestin embryonnaire et leurs impacts sur les comportements des ccnes. Nous avons montré que l'intestin embryonnaire se rigidifie au cours de son developpement et que la migration en 3D des CCNEs est inhibée lorsque la rigidité de l'environnement dépasse un certain seuil. Enfin, nous avons démarré l'analyse de l'effet de l'élasticité sur la différenciation des progéniteurs entériques. L'ensemble de nos résultats permettent de mieux comprendre les mécanismes contrôlant le développement du SNEThe enteric nervous system (ENS) is derived from enteric neural crest cells (ENCC) that migrate along the length of the intestine through the gut mesenchyme. During this process, ENCC proliferate and differentiate into glial cells and neurons, which aggregate into ganglia. The aim of my thesis is to study how biochemical and mechanical properties of the gut tissue influence ENCC colonization and fate during embryogenesis. The absence of endothelin-3 (EDN3), a small peptide trapped in the embryonic gut mesenchyme, is one of the causes leading to hirschsprung disease, characterized by an aganglionosis of the distal colon. We highlighted for the first time that EDN3 increases ENCC adhesion properties throught 1-integrins focal adhesions and modulates their protrusion dynamics. Moreover, we evidenced a genetic interaction between Edn3 and Itgb1 during ENS development. Also, it is now well established that mechanical properties of the microenvironment influence fundamental mechanisms such as cell migration and cell fate determination. Thus, we analysed whether the mechanical properties of the ENCC’s environment influence their behaviours. Using biophysical approaches, we evidenced a physiological stiffening of the embryonic gut during its development and showed that ENCC migration in 3D is inhibited above a certain rigidity threshold. Finally, we begun to analyse the influence of the elastic properties of the environment onto enteric progenitor cells differenciation, taking advantage of the neurosphere culture system. All together, our results contribute to the understanding of the molecular and cellular mechanisms driving physiological and pathological ENS ontogenesis
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Ferré, François. "Isolation et caractérisation des cellules souches gingivales : étude de leur potentiel multipotent." Phd thesis, Université René Descartes - Paris V, 2013. http://tel.archives-ouvertes.fr/tel-01017172.
Full textAbstract:
Les capacités de cicatrisation de la gencive en font un modèle de régénération tissulaire naturelle. Ces capacités sont liées en grande partie à l'activité des fibroblastes. Composante cellulaire principale du tissu conjonctif gingival, ils sont au cœur de la régulation des réponses inflammatoires et des processus de cicatrisation. Nous avons supposé que ce tissu pouvait contenir des cellules souches, pouvant expliquer en partie, ces capacités de réparation. Au cours de cette thèse, nous avons pu mettre en évidence la présence de cellules souches mésenchymateuses aux propriétés communes avec les cellules souches adultes dérivées des crêtes neurales. Ces cellules expriment des marqueurs spécifiques des cellules souches et des crêtes neurales. Par ailleurs, elles présentent des capacités d'auto-renouvellement et de multipotence. Elles sont, en effet, capables de se différencier en adipocytes, ostéocytes et chondrocytes. Nous nous sommes plus particulièrement intéressés à la différenciation chondro/endochondrale. La culture des cellules, sous forme de sphères en suspension, a permis de mettre en évidence leurs capacités de différenciation en tissus cartilagineux et articulaires. Elles s'organisent spontanément en plusieurs types cellulaires différents, générant notamment des chondrocytes hypertrophiques et des synoviocytes selon leur localisation au sein des sphères et du milieu de culture utilisé. Le comportement de ces cellules soumises à ces conditions a permis de montrer leurs facultés à reproduire, in vitro, des processus proches de ceux retrouvés au cours du développement. Ces résultats permettent une meilleure compréhension des phénomènes de différenciation des cellules souches adultes, ouvrant ainsi de nouvelles perspectives pour des applications en thérapie cellulaire articulaire et osseuse.
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Gazquez, Elodie. "Études des interactions fonctionnelles entre l'endothéline-3, les intégrines beta1 et les propriétés élastiques du tissu embryonnaire au cours du développement du système nerveux entérique." Electronic Thesis or Diss., Paris 6, 2016. https://accesdistant.sorbonne-universite.fr/login?url=https://theses-intra.sorbonne-universite.fr/2016PA066240.pdf.
Full textAbstract:
Le système nerveux entérique (SNE) provient des cellules de crête neurale entériques (CCNEs) qui migrent au sein de l'intestin embryonnaire, prolifèrent et se différencient en cellules gliales et neurones formant des ganglions interconnectés. Mon projet de thèse vise à comprendre comment les propriétés biochimiques et mécaniques de l'intestin embryonnaire influencent la colonisation et la différenciation des ccnes. L'absence d'endothéline-3 (EDN3), un facteur biochimique exprimé dans la paroi intestinale, est une des causes de la maladie de hirschsprung, caracterisée par une aganglionose du côlon distal. Nous montrons pour la première fois que l'EDN3 stimule l'adhésivité des CCNEs en augmentant leurs adhérences focales dépendantes des intégrines beta1 ainsi que la dynamique de leurs protrusions membranaires. De plus, nous avons mis en évidence l'existence d'une interaction génétique entre Edn3 et Itgb1 gouvernant le développement du SNE. Par ailleurs, les propriétés mécaniques du microenvironnement influençant la migration et la différenciation cellulaire , nous avons analysé par des approches biophysiques les propriétés élastiques de l'intestin embryonnaire et leurs impacts sur les comportements des ccnes. Nous avons montré que l'intestin embryonnaire se rigidifie au cours de son developpement et que la migration en 3D des CCNEs est inhibée lorsque la rigidité de l'environnement dépasse un certain seuil. Enfin, nous avons démarré l'analyse de l'effet de l'élasticité sur la différenciation des progéniteurs entériques. L'ensemble de nos résultats permettent de mieux comprendre les mécanismes contrôlant le développement du SNEThe enteric nervous system (ENS) is derived from enteric neural crest cells (ENCC) that migrate along the length of the intestine through the gut mesenchyme. During this process, ENCC proliferate and differentiate into glial cells and neurons, which aggregate into ganglia. The aim of my thesis is to study how biochemical and mechanical properties of the gut tissue influence ENCC colonization and fate during embryogenesis. The absence of endothelin-3 (EDN3), a small peptide trapped in the embryonic gut mesenchyme, is one of the causes leading to hirschsprung disease, characterized by an aganglionosis of the distal colon. We highlighted for the first time that EDN3 increases ENCC adhesion properties throught 1-integrins focal adhesions and modulates their protrusion dynamics. Moreover, we evidenced a genetic interaction between Edn3 and Itgb1 during ENS development. Also, it is now well established that mechanical properties of the microenvironment influence fundamental mechanisms such as cell migration and cell fate determination. Thus, we analysed whether the mechanical properties of the ENCC’s environment influence their behaviours. Using biophysical approaches, we evidenced a physiological stiffening of the embryonic gut during its development and showed that ENCC migration in 3D is inhibited above a certain rigidity threshold. Finally, we begun to analyse the influence of the elastic properties of the environment onto enteric progenitor cells differenciation, taking advantage of the neurosphere culture system. All together, our results contribute to the understanding of the molecular and cellular mechanisms driving physiological and pathological ENS ontogenesis
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Radu, Anca Gabriela. "Nouvelles régulations métaboliques exercées par la signalisation LKB1 dans les cellules polarisées : conséquences pour l’ontogénie tissulaire." Thesis, Université Grenoble Alpes (ComUE), 2018. http://www.theses.fr/2018GREAV011/document.
Full textAbstract:
Le suppresseur de tumeur et sérine/thréonine kinase LKB1 est un régulateur clé de la polarité cellulaire et du métabolisme énergétique en partie grâce à l'activation de sa kinase substrat AMPK. Cette protéine est un senseur métabolique pour adapter les apports énergétiques aux besoins nutritionnels des cellules confrontées à un stress. Pour cela, AMPK phosphoryle divers substrats qui activent les réactions cataboliques et inhibent les processus anaboliques dont la kinase mTOR.Au cours de ma thèse, via l’utilisation de modèles murins d’inactivation conditionnelle, j'ai découvert que Lkb1 est crucial pour la formation des cellules de crête neurale (CCN). Ces cellules multipotentes, originaires du tube neural, donnent naissance à divers dérivés, comme les cellules des os et cartilage de la face, les cellules pigmentées de la peau et les cellules gliales et neurales des nerfs périphériques et du système nerveux entérique. J'ai démontré que Lkb1 est essentiel pour la formation de la tête des vertébrés et pour la différenciation et le maintien des dérivés des CCN dans le système nerveux périphérique. J'ai également mis en évidence l’acétylation de LKB1 sur la lysine 48 par l'acétyltransférase GCN5 et son rôle dans l'ontogenèse des CCN céphaliques et la formation de la tête. De plus, j'ai découvert que Lkb1 contrôle la différenciation des cellules gliales en réprimant un programme de biosynthèse d’acides aminés couplé à la transamination du pyruvate en alanine, en amont de la voie de signalisation mTOR.Les phénotypes dus à la perte de Lkb1 dans les CCN récapitulent les caractéristiques cliniques de maladies humaines appelées neurocristopathies. L’activation anormale du suppresseur de tumeur p53 est également associée à certaines neurocristopathies et l’ablation de p53 sauve le phénotype pathologique. Ainsi, j'ai montré que Lkb1 dans les cellules gliales contrôle p53 en limitant les dommages à l’ADN. Lkb1 est aussi essentiel pour maintenir l’homéostasie lysosomale et le recyclage des protéines et ainsi empêcher la formation de granules nommés lipofuscine, chargés en protéines et lipides oxydés. De façon intéressante, les voies mTOR et LKB1/AMPK sont activées à la surface des lysosomes de façon dépendante des niveaux d’acides aminés. Des données récentes de la littérature suggèrent que les lysosomes constitueraient une plateforme de signalisation pour contrôler la protéolyse et le devenir cellulaire. Ainsi, nos données proposent que les signalisations Lkb1 et p53 pourraient réguler l'homéostasie lysosomale et en conséquence le vieillissement cellulaire.De façon intéressante, les cellules de Sertoli, des cellules somatiques épithéliales, localisées dans les tubes séminifères des testicules, et qui régissent la maturation des cellules germinales et l'homéostasie testiculaire, partagent des fonctions métaboliques similaires avec les cellules gliales. En effet, ces cellules sécrètent le lactate et l'alanine qui alimentent les mitochondries des cellules voisines (cellules germinales ou neurones respectivement) contrôlant ainsi leur survie et leur maturation. Au cours de ma thèse, nous avons observé que Lkb1 est requis pour l'homéostasie testiculaire et la spermatogenèse en régulant la polarité des cellules de Sertoli et leur métabolisme énergétique par le cycle pyruvate-alanine. Ces résultats suggèrent une conservation des régulations métaboliques par Lkb1 dans divers tissus.Dans leur ensemble, mes travaux de thèse ont apporté une meilleure connaissance des mécanismes sous-jacents des régulations métaboliques lors du devenir cellulaire. Ces résultats fournissent de nouvelles perspectives sur le développement des CCN et élargissent notre compréhension du contrôle métabolique exercé par LKB1. Enfin, mes projets de doctorat ont mis en évidence l'existence d'une communication entre les voie de signalisation Lkb1 et p53 et suggèrent l’importance de cette communication dans les pathologies humaines dues à des défauts des CCNThe tumor suppressor LKB1 codes for a serine/threonine kinase. It acts as a key regulator of cell polarity and energy metabolism partly through the activation of the AMP-activated protein kinase (AMPK), a sensor that adapts energy supply to the nutrient demands of cells facing situations of metabolic stress. To achieve metabolic adaptations, AMPK phosphorylates numerous substrates which inhibit anabolic processes while activating catabolic reactions. In particular, AMPK inhibits the mammalian target of rapamycin (mTOR).During my PhD, based on genetically engineered mouse models, I uncovered that Lkb1 signaling is essential for neural crest cells (NCC) formation. NCC are multipotent cells that originate from the neural tube and give rise to various derivatives including bones and cartilage of the face, pigmented cells in the skin and glial and neural cells in peripheral nerves and the enteric nervous system. I demonstrated that Lkb1 is essential for vertebrate head formation and for the differentiation and maintenance of NCC-derivatives in the peripheral nervous system. I also emphasized that LKB1 is acetylated on lysine 48 by the acetyltransferase GCN5 and that this acetylation could regulates cranial NCC ontogeny and head formation. Furthermore, I discovered that Lkb1 controls NCC-derived glial differentiation through metabolic regulations involving amino acid biosynthesis coupled to pyruvate-alanine cycling upstream of mTOR signaling.Phenotypes due to Lkb1 loss in NCC recapitulate clinical features of human disorders called neurocristopathies and therefore suggest that aberrant Lkb1 metabolic signaling underlies the etiology of these pathologies. Abnormal activation of the tumor suppressor p53 has been described in some NCC disorders and p53 inactivation in neurocristopathy mouse models rescues the pathological phenotype. By using a NCC line that can be cultivated as progenitors or differentiated in glial cells in vitro, I demonstrated that Lkb1 expression in NCC-derivatives controls p53 activation by limiting oxidative DNA damage and prevents the formation of lysosomes filled with oxidized proteins and lipids called lipofuscin granules. Interestingly, activation of mTOR and LKB1/AMPK pathways is governed by amino acid sensors and takes place at the lysosome surface. Lysosomes have been proposed as a signaling hub controlling proteolysis and aging. Thus Lkb1 and p53 signaling could converge especially through lysosome homeostasis thereby potentially impacting cellular aging.Strikingly, Sertoli cells, that are epithelial somatic cells, located in seminiferous tubules in testes, and which govern germ cells maturation and whole testis homeostasis, share similar metabolic functions with glial cells. For example, they secrete lactate and alanine to fuel mitochondria of neighboring cells (germ cells or neurons respectively) to control their survival and maturation. During my PhD, we highlighted that Lkb1 is essential for testis homeostasis and spermatogenesis by regulating Sertoli cell polarity and, as observed in glial cells, energy metabolism through pyruvate-alanine cycling. These data suggest that this particular Lkb1 metabolic regulation is conserved in tissues with similar function.Taken together, these studies reveal the underlying molecular mechanisms that coordinately regulate energy metabolism and cell fate. They provide new insights into NCC development and expand our understanding of the role of LKB1 as an energy metabolic regulator. Finally, my PhD projects uncover the existence of a crosstalk between Lkb1 and p53 and underline its importance in NCC disorders
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Bahm, Isabel. "PDGF signalling during Neural Crest Cell migration." Thesis, University College London (University of London), 2018. http://discovery.ucl.ac.uk/10041758/.
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Neural crest cells are a transient cell population, which migrates through the vertebrate embryonic body, and eventually gives rise to a many different cell types in the adult. Contact inhibition of locomotion (CIL) is a fundamental property of the collective migrating neural crest cells. CIL describes a process by which colliding cells change their direction upon collision and move away from each other, which has been linked to cell dispersion, boundary formation and metastasis. CIL is acquired in neural crest cells during Epithelial-to-Mesenchymal-Transition (EMT), by a switch in the expression of cadherins, from E to N-cadherin. To examine what governs this change I study PDGF signalling during Xenopus laevis cranial neural crest migration. Here I show that PDGFRα and its ligand PDGF-A are expressed in pre-migratory and migrating cranial neural crest cells. Inhibition of PDGF-A/PDGFRα impairs neural crest migration in vivo and cell dispersion in vitro. I find that PDGFRα inhibition leads to a decrease of N-cadherin levels in neural crest cells. Further, I demonstrate that PDGFRα signalling controls N-cadherin dependent CIL. This cellular response is controlled by the PI3K/AKT signalling pathway as a downstream effector of the PDGFRα cellular response in cranial neural crest cells. This data lead me to propose a novel mechanism by which PDGF signalling as a tissue-autonomous regulator of EMT is regulating N-cadherin dependent CIL during cranial neural crest cell migration in Xenopus laevis.