Dissertations / Theses: 'Cell transdifferentiation'

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Cambuli, Francesco. "Comparative analysis of ES cell transdifferentiation to TS-like cells." Thesis, University of Cambridge, 2014. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.648815.

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Yuan, Songyang. "Differentiation and transdifferentiation of adult pancreatic cells." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp02/NQ30425.pdf.

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Madhavan, Mayur C. "Mechanisms of Transdifferentiation and Regeneration." Miami University / OhioLINK, 2005. http://rave.ohiolink.edu/etdc/view?acc_num=miami1133554812.

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Fakhry, Maya. "Molecular mechanisms of vascular smooth muscle cell transdifferentiation into osteochondrocyte-like cells." Thesis, Lyon 1, 2015. http://www.theses.fr/2015LYO10246.

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Chez les patients souffrant d'insuffisance rénale chronique, les calcifications vasculaires représentent la première cause de mortalité. Elles résultent de la trans-différenciation des cellules musculaires lisses (CMLs) en cellules de type ostéoblastique et/ou chondrocytaire, en réponse à des cytokines inflammatoires ou à une hyperphosphatémie. Les CMLs forment alors des cristaux par l'activité de la phosphatase alcaline non-spécifique du tissu (TNAP). A la lumière de résultats récents, nous avons émis l'hypothèse que la TNAP module la trans différenciation des CMLs. Nos objectifs étaient donc de déterminer l'effet de la TNAP dans la trans-différenciation des CMLs, et d'étudier les mécanismes impliqués dans son induction, avec un intérêt particulier pour les microRNAs. Nous avons observé que l'ajout de phosphatase alcaline purifiée ou la surexpression de TNAP stimule l'expression de marqueurs chondrocytaires en culture de CMLs et de cellules souches mésenchymateuses. De plus, l'inhibition de la TNAP bloque la maturation de chondrocytes primaires. Nous excluons un rôle des cristaux formés par la TNAP, puisque l'ajout de cristaux seuls ou associés à une matrice collagénique n'a pas reproduit les effets de la TNAP. Nous suspectons que la TNAP agit en hydrolysant le pyrophosphate inorganique (PPi). En effet, c'est la TNAP qui hydrolyse le PPi en culture de CMLs et de chondrocytes, et le PPi mime les effets de l'inhibition de TNAP en culture de chondrocytes. Enfin, nous rapportons le profil de microRNA des artères cultivées en conditions hyperphosphatémiques. Ces résultats pourraient être particulièrement importants dans le développement de nouvelles approches thérapeutiques
In patients with chronic kidney disease (CKD), vascular calcification represents the main cause of mortality. Vascular calcification results from the trans-differentiation of vascular smooth muscle cells (VSMCs) into cells similar to osteoblasts and/or chondrocytes, in response to inflammatory cytokines or hyperphosphatemia. Calcifying VSMCs form calcium phosphate crystals through the activity of tissue nonspecific alkaline phosphatase (TNAP). In light of recent findings, we hypothesized that TNAP also modulates VSMC trans-differentiation. Our objectives were therefore to determine the effect of TNAP activity on VSMC trans-differentiation, and secondly to investigate the molecular mechanisms involved in TNAP expression in aortas, with a particular interest in microRNAs. We first observed that addition of purified alkaline phosphatase or TNAP over-expression stimulates the expression of chondrocyte markers in culture of the mouse and rat VSMC lines, and of mesenchymal stem cells. Moreover, TNAP inhibition blocks the maturation of mouse primary chondrocytes and reduces mineralization. We exclude a role for crystals in TNAP effects, since addition of crystals alone or associated to a collagenous matrix fails to mimic TNAP effects. We rather suspect that TNAP acts through the hydrolysis of inorganic pyrophosphate (PPi). Indeed, PPi is hydrolyzed by TNAP in VSMCs and chondrocytes and addition of PPi mimics the effects of TNAP inhibition on chondrocyte maturation. Finally, we report microRNA signature of aortic explants treated under hyperphosphatemic conditions that induce vascular calcification. These results could be of particular importance in patients with CKD

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Mojica, Jorge Ricardo. "Commitment and transdifferentiation of adult progenitor cells: The contributing pathways." Thesis, University of Ottawa (Canada), 2008. http://hdl.handle.net/10393/27717.

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C2C12 murine progenitors are a model for transdifferentiation. This project investigated signaling events that regulate this process in culture by monitoring specific markers for transdifferentiation (alkaline phosphatase [ALP] and myogenin for the osteoblastic and myoblaststic fates respectively) in the presence of inducers and inhibitors. Antibodies specific for proteins implicated in signaling were used to monitor the differentiation process. BMP-2 addition caused transdifferentiation to the osteoblastic fate except for a small fraction of cells that may represent the so-called MyoD negative cells or the side population (SP). The timing of short term commitment of C2C12 to osteoblastic transdifferentiation was between 6 to 24h under our culture conditions. BMP-2-induced ALP activity increase and the repression of myogenin expression were not simultaneous. Differentiation as measured by ALP activity may be dependent on the interaction of BMP-2/Smad, p38 MAPK and TGF-beta1 pathways. Smads 1, 5 and/or 8 were almost always found under their phosphorylated form (pSmads) and thus almost invariably located in nuclei suggesting fast nucleoplasmic shuttling from the cytoplasm and nuclear retention of the phosphorylated forms. Phosphorylated forms were also found in the nucleus under proliferative conditions without exogenous BMP-2. In this case, it is suggested that this phosphorylation of Smads may be triggered by the reported autocrine secretion of TGF-beta by the C2C12 cells or by the TGF-beta present in the culture serum.

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Yuan, Yuan. "Small-Molecule Modulators of Pancreatic Ductal Cells: Histone Methyltransferases and \(\beta\)-Cell Transdifferentiation." Thesis, Harvard University, 2012. http://dissertations.umi.com/gsas.harvard:10637.

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Small molecules are important not only for treating human diseases but also for studying disease-related biological processes. This dissertation focuses on the effects of small molecules on pancreatic ductal adenocarcinoma cells. Here, I describe the discovery of two small-molecule tool compounds and their applications for interrogating the biological processes related to two distinct diseases in the human pancreas. First, BRD4770 was identified as a histone methyltransferase inhibitor through a target-based biochemical approach, and was used as a probe to study the function of methyltransferases in cancer cells. Second, BRD7552 was discovered as an inducer of Pdx1 using a cell-based phenotypic screening approach, and was used to induce the expression of Pdx1, a master regulatory transcription factor required for \(\beta\)-cell transdifferentiation. This compound is particularly interesting for the study of type-1 diabetes (T1D). The histone methyltransferase G9a catalyzes methylation of lysine 9 on histone H3, a modification linked to aberrant silencing of tumor-suppressor genes. The second chapter describes the collaborative effort leading to the identification of BRD4770 as a probe to study the function of G9a in human pancreatic cancer cells. BRD4770 induces cellular senescence and inhibits both anchorage-dependent and -independent proliferation in PANC-1 cell line, presumably mediated through ATM-pathway activation. Chapter three describes the study of a natural product gossypol, which significantly enhances the BRD4770 cytotoxicity in p53-mutant cells through autophagic cell death. The up-regulation of BNIP3 might be responsible for the synergistic cell death, suggesting that G9a inhibition may help overcome drug resistance in certain cancer cells. Ectopic overexpression of Pdx1, Ngn3, and MafA can reprogram pancreatic exocrine cells to insulin-producing cells in mice, which sheds light on a new avenue for treating T1D. The fourth chapter focuses on a gene expression-based assay using quantitative real-time PCR technique to screen >60,000 compounds for induction of one or more of these three transcription factors. A novel compound BRD7552 which up-regulated Pdx1 mRNA and protein levels in PANC-1 cells was identified. BRD7552 induces changes of the epigenetic markers within the Pdx1 promoter region consistent with transcriptional activation. Furthermore, BRD7552 partially complements Pdx1 in cell culture, enhancing the expression of insulin induced by the introduction of the three genes in PANC-1 cells. In summary, the central theme of my dissertation is to identify novel bioactive small molecules using different screening approaches, as well as to explore their effects in pancreatic ductal cells.
Chemistry and Chemical Biology

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Barbosa-Sabanero, Karla Y. "Dedifferentiation and transdifferentiation: a study of the RPE cell identity." Miami University / OhioLINK, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=miami1468660645.

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Gsour, Amna. "Differentiation of human cell line towards a pancreatic endocrine lineage." Thesis, University of Manchester, 2016. https://www.research.manchester.ac.uk/portal/en/theses/differentiation-of-human-cell-line-towards-a-pancreatic-endocrine-lineage(0c2c21fe-724d-449f-804c-02741c89828c).html.

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Islet transplantations have been successful in restoring glucose homeostasis in patients with diabetes; however, the limited number of donor organs limits the success of this treatment. The lineage reprograming of different cell sources to beta cells potentially provides an unlimited supply of insulin-producing cells for regenerative therapy for patients with diabetes. The aim of this study was to investigate the ability to transdifferentiate two cell lines into an endocrine lineage. Insulin production in pancreatic beta cells can be increased using a small molecule, 3,5-disubstituted isoxazole, N-cyclopropyl-t-(thiophen-2-yl)isoxazole-3-carboxamide (isoxazole) but its effect on other cell types has not been reported. Here, we investigated the lineage reprogramming of PANC-1 pancreatic ductal cells to insulin producing cells by isoxazole treatment. Gene expression was performed using RT-PCR and qPCR for approximately 30 genes critical to beta cell development and function. In addition, quantitative proteomic profiling was performed using LC-MS by monitoring protein abundance in isoxazole-treated PANC-1 cells compared to time-matched controls. Isoxazole treatment stimulated PANC-1 cells to aggregate into islet-like clusters and gene expression analysis revealed induction of important developmental beta cell markers including NGN3, NEUROD1 and INSULIN. In addition, beta cell surface markers were also upregulated such as CD200, GPR50, TROP-2, GLUT2 and SLC30A8. Using LC-MS a catalogue of approximately 2400 identified proteins was generated; 257 proteins were differentially expressed in isoxazole-treated cells compared to DMSO-vehicle controls at p < 0.05. Amongst the proteins upregulated were molecules that regulate metabolic processes and cytoskeletal reorganisation. The expression of the majority of these proteins has not been previously reported or studied in the context of beta cell differentiation. Functional analysis of the relative protein changes was determined using Ingenuity Pathway Analysis, IPA, and gene ontology, GO, software, which revealed the regulation of several cellular canonical pathways including metabolic pathways, cell adhesion, remodelling of epithelial adherens junctions and actin cytoskeleton signalling. The effects of isoxazole were further studied in the A549 lung cancer cell line. Similar effects were observed, such as the induction of pro-endocrine markers NGN3 and NEUROD1 and endocrine-specific hormones INS and GCG. These results indicate that isoxazole has the capacity to transdifferentiate pancreatic and non-pancreatic cell origins into an endocrine lineage. This study reveals the powerful induction capacity of isoxazole in inducing cellular reprogramming events.

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Rapino, Francesca 1982. "Induced transdifferentiation of human B-leukemia/lymphoma cell lines and inhibition of leukemogenicity." Doctoral thesis, Universitat Pompeu Fabra, 2013. http://hdl.handle.net/10803/128575.

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B-cell malignancies encompass a wide variety of distinct diseases including Non Hodgkin lymphoma (NHL) and leukemia. Currently, chemotherapy, radiation and anti-CD20 antibody treatment are the mainstays of B-cell lymphoma and leukemia therapy. However, the fact that a large number of patients are eventually not cured justifies the search for novel and more effective therapeutic approaches. Although induction of differentiation has been shown to be effective in several tumors such as acute promyelocytic leukemia, it has not been tested yet in NHL and leukemia. We therefore hypothesized that transdifferentiation of malignant B cells could be proposed as a novel therapeutic approach. Earlier work of our laboratory demonstrated that the transcription factor C/EBPα could convert immature and mature murine B lineage cells into functional macrophages at high efficiencies. Here we show that the ectopic expression of C/EBPα can likewise induce the conversion of selected human lymphoma and leukemia B-cell lines into macrophages. The reprogrammed cells are functional and quiescent. Importantly, the tumorigenicity of transdifferentiated lymphoma and leukemia cell lines was impaired after transplantation into immunodeficient mice, even when C/EBPα was activated in vivo. In summary, our experiments show for the first time that human cancer cells can be induced to transdifferentiate by C/EBPα into seemingly normal cells at high frequencies, thus proposing transdifferentiation as novel therapeutic approach. In line with this, we believe that the finding of a small molecule that mimics C/EBPα overexpression will open new horizons for the cure of patients affected by B cell malignancies.
Las neoplasias malignas de células B abarcan una amplia variedad de enfermedades diferentes, incluyendo el linfoma no Hodgkin (LNH) y leucemia. Actualmente, la quimioterapia, la radiación y el tratamiento con anticuerpos anti-CD20 son los pilares de la terapia contra el linfoma y la leucemia de células B. Sin embargo, el hecho de que un gran porcentaje de pacientes no se cura con estos tratamientos, justifica la búsqueda de nuevas terapias más eficaces. Aunque la inducción de la diferenciación ha demostrado ser eficaz en el tratamiento de varios tumores tales como la leucemia promielocítica aguda, esta técnica no se ha probado aún en el tratamiento del LNH o de la leucemia. Por lo tanto, la transdiferenciación de las células B malignas podría ser propuesta como un nuevo enfoque terapéutico. Trabajos anteriores de nuestro laboratorio han demostrado que el factor de transcripción C/EBPα puede convertir células de linaje B murinas inmaduras y maduras en macrófagos funcionales con una alta eficiencia. En este trabajo mostramos que la expresión ectópica de C/EBPα puede inducir la conversión de ciertas líneas de linfoma y leucemia humana en macrófagos. Las células reprogramadas son funcionales y quiescientes. Es importante destacar que la tumorigenicidad de linfoma transdiferenciados y líneas celulares de leucemia se vio afectada después del trasplante en ratones inmunodeficientes, incluso cuando C/EBPα se activó in vivo. En resumen, nuestros experimentos muestran por primera vez que las células de cáncer humano pueden ser inducidas por C/EBPα a transdifferenciarse en células aparentemente normales con una alta frecuencia, proponiendo así la transdiferenciación como nuevo enfoque terapéutico. En línea con esto, creemos que el hallazgo de una pequeña molécula que sea capaz de imitar la sobreexpresión de C/EBPα abrirá nuevos horizontes para la cura de los pacientes afectados por tumores malignos de células B.

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Gharibi, Borzo. "Adenosine receptor expression and function during mesenchymal stem cell differentiation and osteoblast transdifferentiation." Thesis, Cardiff University, 2010. http://orca.cf.ac.uk/54352/.

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The mechanisms involved in osteoblast and adipocyte differentiation from the common progenitor, mesenchymal stem cell (MSC) are not fully understood. The nucleoside, adenosine, exists in all cells and is known to be involved in cell growth, proliferation and apoptosis by interacting with four distinct receptors (Ai, A2A, A2b and A3). The aims of this study was to investigate the expression and function of adenosine receptors in 1) MSCs and as they differentiated into osteoblasts and adipocytes and in 2) mouse 7F2 and human osteoblasts and during their transdifferentiation to adipocytes. Rat MSCs and 7F2 osteoblasts expressed all four adenosine receptors. Osteoblast differentiation was associated with increases in A2A and A2B receptor expression and their activation stimulated the expression of alkaline phosphatase, core binding factor a1 and mineralisation. Adenosine also stimulated adipogenesis (lipid accumulation, peroxisome proliferator-activated receptor, CCAAT/enhancer binding protein a and lipoprotein lipase expression) of MSCs which was accompanied by increased A1 and A2A receptor expression. Transdifferentiation of 7F2 cells to adipocytes was associated with increased Ai, but decreased A2A and A2b receptor expression. Loss of A2 receptors in adipocytes was supported by reduced cAMP and extracellular signal regulated kinase responses to adenosine. Adenosine also stimulated transdifferentiation of human osteoblasts to adipocytes by inducing lipoprotein lipase and inhibiting alkaline phosphatase and osteocalcin expression. Overexpression of Ai receptors in 7F2 cells stimulated adipogenesis (lipid accumulation and lipoprotein lipase expression) whereas overexpression of A2b receptors stimulated alkaline phophatase expression and inhibited adipogenesis. These results show that adenosine receptor expression and function is involved in lineage specific differentiation or transdifferentiation A2B receptors are associated with MSCs and osteoblasts and Ai receptors with adipocytes. Targeting adenosine signal pathways may thus be useful as an adjunct therapy for the prevention or treatment of conditions in which there is insufficient bone or excessive adipocyte formation.

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Werner, Mimmi. "Hair cell regeneration in vestibular epithelia : a study in an in vitro model." Doctoral thesis, Umeå universitet, Öron- näs- och halssjukdomar, 2016. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-124077.

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Background Hair cells (HCs) are the sensory receptors in both the auditory and the vestibular organs of the inner ear. Supporting cells (SCs) are non-sensory cells embracing the HCs. Injuries of the HCs by aging, acoustic trauma or ototoxic drugs (mainly aminoglycosides, e.g. gentamicin) and cisplatin, often cause permanent impairment of hearing and balance. Birds and amphibians can regenerate their auditory and vestibular HCs after injury through proliferation of SCs or direct transdifferentiation of a SC into a HC. For mammals this ability is limited and spontaneous HC regeneration occurs only in the vestibular sensory epithelia. The utricle is one of the five vestibular organs and contributes to our balance by registering linear acceleration and head tilts. The aim of this PhD thesis was to investigate morphological and morphometric events during spontaneous HC regeneration following gentamicin exposure in neonatal rat utricular explants. Methods Long-term organ culture of macula utriculi, which is stable and reproducible for up to 28 days in vitro (DIV), was used in all papers in the thesis. HC damage was induced by gentamicin. On 2 DIV the explanted utricular maculae were divided into two groups, a control group and a gentamicin-exposed group. In the latter group macular explants were exposed to gentamicin for 48 hours during 2-3 DIV and then allowed to recover. Morphologic and morphometric evaluations were done from utricles harvested at various time points during 28 DIV. Imaging techniques used were light microscopy, including immunohistochemistry, and transmission electron microscopy. Results In the control group the epithelia were well preserved with a slight decline in HC density after 14 DIV. In the gentamicin-exposed group there was an initial substantial decline in HC density and thereafter the proportion of HCs in relation to SCs increased significantly. Using BrdU as a proliferation marker and myosin 7a as a HC marker, we found no cells that were double marked. At the ultrastructural level, the apical occlusion of the explanted epithelia was intact in both the control and the gentamicin exposed group during the entire in vitro period. Cells that seemed to be in a transitional state, transforming from SCs into HCs were observed in the gentamicin-exposed group. These cells had cytoplasmic extensions basally i.e. foot processes, an assembly of mitochondria basally in the cell or in these foot processes, and often apical SC extensions covering the HC. HCs classified as transitional cells had an increased number of SC connections to their basal parts compared to mature HCs. Conclusions In these neonatal rat utricular explants: - The morphological structure of the sensory epithelia was well preserved during long-term culture. - The renewal of hair cells after gentamicin exposure occurred through direct transdifferentiation of supporting cells into hair cells. - There was also a proliferative response by the supporting cells, but this supporting cell proliferation did not contribute to the generation of new hair cells. - Cells in a transitional state, showing a characteristic morphology, were observed during the process of transdifferentiation from supporting cells into hair cells. - The tight junctional seal of the epithelia stayed morphologically intact also after gentamicin exposure. - Gap junctions were observed in between supporting cells but not found in between hair cells and supporting cells or between transitional cells and supporting cells.

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Hazrati, Ali. "The role of transforming growth factor-beta (TGF-) in the transdifferentiation of islets of Langerhans to duct-like epithelial structures /." Thesis, McGill University, 2002. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=33773.

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The process of islet isolation destroys extracellular matrix and eliminates potentially important inter-cellular relationships. We have previously shown that isolated islets embedded in a type I collagen gel, in the presence of a defined medium, undergo a phenotypic switch to duct-like epithelial structures through a process known as transdifferentiation. The aim of this study was to characterize the specific effectors implicated in islet cell transdifferentiation in order to better understand the factors that confer morphogenetic stability on cells in the isolated islets.
We demonstrated cytoplasmic immunoreactivity for TGF-beta isoforms over 8 days post isolation using canine islets. Islet-to-duct epithelial transdifferentiation was correlated with the total amount of TGF-beta and was maximal at 48 h of culture. Up regulation of TGF-betaRI and TGF-betaRII expression on day 2 post-isolation was demonstrated by immunohistochemistry and Western blot analysis, and correlated temporally with the induction of cell proliferation. The presence of TGF-beta1 in culture supernatants was detected using the PAI/L assay. The peak TGF-beta1 level was 10.94 +/- 2.27 pM (active form) and 52.23 +/- 1.57 pM (total TGF-beta) at 48h. Addition of exogenous TGF-beta1 at different concentrations caused an accelerated and more pronounced epithelial transformation at 5--10 ng/mL compared to lower concentrations (0.5--1 ng/mL).
These studies confirm the biological potential of islets of Langerhans to transdifferentiate to duct epithelial structures. TGF-beta signal transduction appears to play an important role in this process.

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Imamura, Hirotoshi. "Transdifferentiation of bone marrow-derived endothelial progenitor cells into the smooth muscle cell lineage mediated by tansforming growth factor-β1." Kyoto University, 2010. http://hdl.handle.net/2433/120926.

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Matsunaga, Mami. "Initiation of Supporting Cell Activation for Hair Cell Regeneration in the Avian Auditory Epithelium: An Explant Culture Model." Doctoral thesis, Kyoto University, 2021. http://hdl.handle.net/2433/263555.

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Cruz, Raphael Marques de Almeida Rosa da. "Diferenciação neuronal de células-tronco de dente decíduo esfoliado." Universidade de São Paulo, 2016. http://www.teses.usp.br/teses/disponiveis/10/10132/tde-11012017-160059/.

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As células-tronco (CT) são células que apresentam propriedades de auto-renovação, capazes de gerar diferentes tipos celulares e reconstituir diversos tipos de tecidos, por isso têm sido vistas como uma alternativa terapêutica para o tratamento de muitas doenças cujo tratamento convencional não é eficiente. Além disso, por sua plasticidade, as CT podem ser usadas para produzir modelos in vitro para o estudo de doenças, o que tem sido particularmente interessante para doenças que acometem o sistema nervoso. As células-tronco humanas de dente decíduo esfoliado (Stem cell from Human Esfoliated Deciduous teeth, SHED) são descritas em alguns trabalhos como capazes de se diferenciarem em células neuronais através da transdiferenciação direta, um processo simples que possibilita a diferenciação de um tipo celular em outro por meio da utilização de agentes químicos. No entanto, a diferenciação de SHED em neurônios é contestada como sendo um artefato de cultura devido ação citotóxica destes agentes. Com base nesta premissa, o objetivo deste trabalho foi realizar protocolos para a produção de neurônios a partir das SHED utilizando modificações de métodos descritos na literatura e utilizados na diferenciação neuronal. Durante um dos protocolos utilizados, foi possível verificar a morfologia de células semelhantes à neurônio, porém após alguns dias de manutenção, as células voltavam ao formato fusiforme original. Com este trabalho podemos afirmar que, de acordo com os resultados obtidos, as SHED não geram neurônios funcionais
Stem cells (SC) are cells that present self-renewal properties, capable of generating cells types and reconstitute several tissue types. For this reason, they have been seen as a therapeutic alternative to the treatment of many diseases which conventional treatment is not efficient. Besides, because of its plasticity, SC may be used to produce in vitro models in order to study diseases, what is interesting to diseases that affect the nervous system. Human SC from SHED (Stem cell from Human Exfoliated Deciduous teeth, SHED) are described in some papers as being capable of self-differentiation in neuronal cells through the direct transdifferentiation, a simple process that makes possible the differentiation of a cell type in another through the use of chemical agents. Nevertheless, SHED differentiation is refuted as being a artifact of culture due to cytotoxic action of these agents. Based on this premise, the aim of this study was to carry out protocols in order to produce neurons from SHED, using modifications and methods of neuronal differentiation described in the literature and used in the neuronal differentiation. During one of the protocols used, it was possible to verify the morphology of the cells as neuron-like, however, after some days of maintenance, they turn back to their original fusiform format. In this study, we can state, according to the obtained results, that SHED do not generate functional neurons

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Son, Yesde. "Exploring the Plasticity of Cellular Fate Using Defined-Factor Reprogramming." Thesis, Harvard University, 2012. http://dissertations.umi.com/gsas.harvard:10309.

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Cellular fate, once established, is usually stable for the lifetime of the cell. However, the mechanisms that restrict the developmental potential of differentiated cells are in principle reversible, as demonstrated by the success of animal cloning from a somatic genome through somatic cell nuclear transfer (SCNT). An increased understanding of the molecular determinants of cell fate has also enabled the reprogramming of cell fate using defined transcription factors; recently, these efforts have culminated in the discovery of four genes that convert somatic cells into induced pluripotent stem cells (iPSCs), which resemble embryonic stem cells (ESCs) and can give rise to all the cell types in the body. As a first step toward generating clinically useful iPSCs, we identified a small molecule, RepSox, that potently and simultaneously replaces two of the four exogenous reprogramming factors, Sox2 and cMyc. This activity was mediated by the inhibition of the Transforming Growth Factor-\(\beta\) \((Tgf-\beta)\) signaling pathway in incompletely reprogrammed intermediate cells. By isolating these stable intermediates, we showed that RepSox acts on them to rapidly upregulate the endogenous pluripotency factor, Nanog, allowing full reprogramming to pluripotency in the absence of Sox2. We also explored lineage conversion as an alternative approach for producing a target cell type in a patient-specific manner, without first generating iPSCs. A combination of pro-neural as well as motor neuron-selective factors could convert fibroblasts directly into spinal motor neurons, the cells that control all voluntary movement. The induced motor neurons (iMNs) displayed molecular and functional characteristics of bona fide motor neurons, actuating muscle contraction in vitro and even engrafting in the developing chick spinal cord when transplanted. Importantly, functional iMNs could be produced from fibroblasts of adult patients with the fatal motor neuron disease, amyotrophic lateral sclerosis (ALS). Given the therapeutic value of generating patient-specific cell types on demand, defined-factor reprogramming is likely to serve as an important tool in regenerative medicine. It is hoped that the different approaches presented here can complement existing technologies to facilitate the study and treatment of intractable human disorders.

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Keuylian, Zela Talar. "The implication of adenylyl cyclase isoform 8 and its regulation by the Notch pathway in vascular smooth muscle cell transdifferentiation and pathological vesel remodeling." Paris 6, 2013. http://www.theses.fr/2013PA066110.

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L’athérosclérose se caractérise par le rétrécissement de la taille de la lumière des vaisseaux artériels, appelé “sténose”, par la formation de plaques d’athérome dans la paroi des artères. L’un des facteurs majeurs qui contribue à la progression de la formation des lésions est le changement phénotypique des cellules musculaires lisses médiales. Ce processus permet leur transition d’un phénotype quiescent/contractile à un phénotype sécrétoire, prolifératif et migratoire. Mes travaux consistaient à élucider une partie des mécanismes qui régule ce changement. Nous avons montré que l’expression d’une isoforme de l’enzyme adenylyl cyclase (AC), l’AC8, est impliquée dans l’inflammation et la migration des cellules musculaires lisses vasculaires. Chez l’homme, nous avons montré que les plaques d’athérosclérose expriment fortement l’AC8, principalement dans les cellules musculaires lisses de l’intima. L’AC8 était peu présent dans la média, que ce soit dans les artères pathologiques ou saines. Chez le rat, dans le modèle de resténose, nous avons montré une expression transitoire de l’AC8 régulée par la voie Notch ; in vitro, nous avons démontré que l’inhibition de la voie Notch a potentialisé l’effet de l’IL1β sur l’expression de l’AC8 et a inhibé l’expression des gènes cibles de Notch, Hrt1 et Hrt3. Dans le même modèle de resténose, l’expression transitoire de l’AC8 a coïncidé avec l’inhibition de Notch3. Ces expériences ont démontré que la voie Notch inhibe l’expression de l’AC8 induit par IL1β, et suggèrent que l’expression de l’AC8, en plus d’avoir été induite par la cytokine IL1β, dépend de l’inhibition de la voie Notch dans un contexte inflammatoire
Atherosclerosis is characterized by the narrowing of the arterial lumen termed “stenosis”, due to the expansion of arterial plaques. One of the major contributing factors to the formation of lesions and the neo-intima during post-angioplasty restenosis is the phenotypic change of medial vascular smooth muscle cells. This process switches them from a quiescent/contractile phenotype to a secretory, proliferative, migratory one. My work consisted of elucidating some of the molecular mechanisms implicated in this switch. We showed that the expression of an adenylyl cyclase (AC) isoform, AC8, is implicated in both the inflammatory and migratory properties acquired by trans-differentiated VSMCs. In human atherosclerotic arteries we showed that only intimal VSMCs strongly express AC8; very few AC8 positive VSMCs were detected in the medial layer, either in atherosclerotic or healthy arteries. In the rat balloon injury model of restenosis, we showed a transitory increase of AC8 expression. In vitro, we demonstrated that AC8 expression is regulated by the Notch pathway; inhibiting Notch amplified AC8 expression and decreased Notch target genes Hrt1 and Hrt3. In the same model of restenosis, the transitory up-regulation of AC8 expression coincided with Notch3 down-regulation. These set of experiments demonstrated that the Notch pathway decreases IL1β-mediated AC8 up-regulation in trans-differentiated VSMCs and suggests that AC8 expression, besides being induced by the proinflammatory cytokine IL1β,also depends on the down-regulation of the Notch pathway occurring in an inflammatory context. As a whole, my studies attribute a new role for AC8 in pathological vascular remodeling

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18

Taylor, Sean R. "Co-expression of HB-EGF and ADAM 12S displays a brown adipose phenotype in mouse and human cell lines." Miami University / OhioLINK, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=miami1524347780576749.

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19

Mongeon, Kevin. "The Study of Hereditary Spastic Paraplegia-Causing Gene DDHD2 Using Cell Models." Thesis, Université d'Ottawa / University of Ottawa, 2018. http://hdl.handle.net/10393/37474.

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Hereditary spastic paraplegia type 54 is a rare autosomal recessive neurological gait disorder characterized by paraplegia, muscle spasticity, and intellectual disability. This length-dependent distal axonopathy is caused by mutations in the DDHD2 gene, which encodes the intracellular phospholipase A1 DDHD2. Little is known about the molecular function of the DDHD2 protein, especially in the context of HSP54. Thus, there is a need to further investigate its molecular functions and investigate the impact of DDHD2 deficiency in disease-relevant cells. Here, lipidomic profiling of dermal fibroblasts derived from three unrelated patients has revealed 19 glycerophosphoethanolamine species at differential levels in patients relative to unaffected controls. However, patient cells appear to have an unaffected Golgi apparatus morphology and lipid droplet formation, despite DDHD2’s proposed roles in these processes. To study the gene function in neuronal cells, I transdifferentiated the fibroblasts into induced neuronal precursor cells and found all the patient cells arrested in the G0/G1 phase of upon conversion. Given that these cell lines are unsustainable, I generated a stable knockdown cell line in the highly proliferative HEK293A to study the molecular biology of DDHD2. The knockdown cells had a reduced growth, were delayed in the G2/M phase of the cell cycle, and became multinucleated. I then treated the cells with antineoplastic compounds paclitaxel and nocodazole and found more knockdown cells in G0/G1 than controls, suggesting the possible occurrence of mitotic slippage. Lastly, I report a novel subcellular localization for DDHD2 at the microtubule organization center.

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20

Deshors, Pauline. "Transdifférenciation radio-induite des cellules souches de glioblastome en cellules endothéliales." Thesis, Toulouse 3, 2019. http://www.theses.fr/2019TOU30127.

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Les Glioblastomes (GBM) sont les tumeurs cérébrales les plus fréquentes chez l'adulte. Ces tumeurs sont de mauvais pronostic malgré un traitement lourd associant chirurgie, chimio- et radio-thérapie. En effet, ces tumeurs récidivent inévitablement. La présence d'une sous-population de cellules ayant des caractères souches appelées cellules souches de glioblastome (CSG) peut en partie expliquer l'agressivité de ces tumeurs et l'échec thérapeutique. D'autre part, les GBM sont caractérisés par une vascularisation abondante et anormale. Il a été montré que les CSG se transdifférenciaient en cellules ayant des caractéristiques endothéliales (appelées TDEC) qui étaient des facteurs clés de la croissance tumorale. Les CSG étant résistantes au traitement (et notamment à la radiothérapie), nous avons émis l'hypothèse que les radiations ionisantes pouvaient favoriser la transdifférenciation des CSG en TDEC. Ces cellules pourraient participer, après traitement, à l'apparition de nouveaux vaisseaux essentiels aux cellules tumorales résiduelles et à l'apparition des récidives. Au cours de ma thèse, à partir de primocultures de CSG, j'ai montré in vitro et in vivo que des radiations ionisantes potentialisaient les fonctions proangiogéniques des TDEC. De plus j'ai identifié la voie de signalisation du récepteur angiogénique Tie2 comme un facteur essentiel à cette potentialisation radio-induite. Par une stratégie d'inhibition à l'aide d'un inhibiteur pharmacologique spécifique de Tie2, j'ai confirmé in vitro et in vivo l'implication de cette voie de signalisation dans ce mécanisme. En conclusion, mes résultats de thèse ont permis d'identifier un nouveau mécanisme potentiellement impliqué dans la radiorésistance des GBM via la transdifférenciation des CSG survivantes, permettant la mise en place de nouveaux vaisseaux tumoraux. La voie de signalisation de Tie2 est impliquée dans cette potentialisation radio-induite des caractéristiques proangiogéniques des TDEC. De nouvelles stratégies thérapeutiques associant radiothérapie/témozolomide et inhibiteur de la voie de signalisation de Tie2 devraient être envisagées dans de prochains essais cliniques
Glioblastomas (GBM) are brain tumors which display a bad prognosis despite conventional treatment associating surgical resection and subsequent radio-chemotherapy. Indeed, these invasive tumors recur almost inevitably. The presence of a radioresistant and tumorigenic GBM Stem Cells (GSC) subpopulation could contribute to explain this clinical impasse and the recurrence of these tumors. Furthermore, GBM are characterized by an important and abnormal vascularization. It has been shown that GSC could transdifferentiate into cells with endothelial features named Tumor Derived Endothelial Cells (TDEC) which are key compounds of tumor growth. Since GSC are radioresistant, we hypothesized that ionizing radiations are able to facilitate transdifferentiation of GSC into TDEC. TDEC appearing within the irradiation field could thus participate to the formation of new vessels and could help remaining tumor cells to develop a new aggressive tumor. During my PhD, I first showed in vitro and in vivo that irradiation potentiates proangiogenic functions of TDEC. At the molecular level, I highlighted Tie2 signaling pathway as an important actor of this potentialization of proangiogenic features of TDEC. By using a specific inhibitor of Tie2 kinase activity, I confirmed Tie2 signaling pathway involvment in this mechanism in vitro and in vivo. In conclusion, my doctoral work highlights the existence of a new mechanism of radioresistance in GBM through the transdifferentiation of the surviving GSC after treatment, leading to the formation of a new tumor vasculature. The radio-induced potentialization of proangiogenic features of TDEC appeared to be supported by Tie2 signaling pathway. New therapeutic strategies associating radiotherapy/temozolomide and an inhibitor of Tie2 signaling pathway should be considered for forthcoming trials

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Tolentino, Felipe Thadeu 1983. "Produção e avaliação de vetores retrovirais visando à diferenciação de neurônios olfativos in vitro pela superexpressão de fatores de transcrição definidos." [s.n.], 2014. http://repositorio.unicamp.br/jspui/handle/REPOSIP/316718.

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Orientador: Fabio Papes
Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia
Made available in DSpace on 2018-08-24T14:16:00Z (GMT). No. of bitstreams: 1 Tolentino_FelipeThadeu_M.pdf: 9244448 bytes, checksum: deea9f7963e05d8a997d9b5a554f9708 (MD5) Previous issue date: 2014
Resumo: O Sistema Sensorial Olfativo de mamíferos é composto por vários subsistemas na cavidade nasal. Dentre estes, destacam-se o sistema olfativo principal e o sistema olfativo acessório ou vomeronasal. O primeiro realiza a detecção geral de odores e parece participar também da detecção de algumas substâncias que levam a respostas comportamentais instintivas (feromônios), enquanto o último é especializado na detecção desta classe de semioquímicos. A detecção dos estímulos sensoriais olfativos resulta em informações importantes que dependem de vias complexas para sua interpretação e para a geração de respostas apropriadas por parte do sistema nervoso central. Existem vários pontos ainda desconhecidos sobre o funcionamento do sistema olfativo, tanto no que diz respeito aos mecanismos moleculares subjacentes à escolha dos receptores a serem expressos por um dado neurônio sensorial ¿ sendo que cada neurônio olfativo expressa apenas um receptor dentro de uma grande família multi-gênica ¿ quanto em relação ao processamento da informação sensorial em centros cerebrais superiores. Neurônios sensoriais olfativos cultivados eficientemente in vitro seriam extremamente úteis, pois poderiam ser utilizados como ferramenta para o estudo destes problemas, como a investigação da atividade das células sensoriais olfativas, possibilitando, por exemplo, uma melhor compreensão dos mecanismos genéticos e moleculares por trás da expressão dos receptores olfativos e de suas propriedades de detecção. Neste trabalho foram desenvolvidas ferramentas baseadas em vetores retrovirais com o objetivo de induzir a diferenciação celular de neurônios olfativos in vitro, utilizando uma combinação de fatores de transcrição, por meio de transdução viral em células-alvo (fibroblastos murinos). Os retrovírus produzidos foram testados e algumas combinações de fatores de transcrição foram preliminarmente testadas, sendo capazes de induzir mudanças moleculares em fibroblastos acompanhadas da expressão de marcadores de neurônios sensoriais olfativos
Abstract: The mammalian Olfactory System enables the vast majority of animal species to identify the presence and quality of food, predators, competitors, conspecifics and potential mates in the environment. Olfactory stimuli detected by sensory neurons are interpreted by brain processing pathways to generate appropriate behavioral and endocrine responses. Despite its central importance in mammalian physiology, several aspects about the biology of this sensory system remain uncharacterized. For example, it is known that each olfactory sensory neuron (OSN) in the nasal cavity expresses only one gene out of a large multi-gene family coding for receptors involved in odorant and pheromone detection. However, the molecular mechanisms behind this process of olfactory receptor gene choice are not fully understood. The study of this and many other aspects of olfaction has been made difficult by the lack of appropriate in vitro cellular models. An efficient way to obtain cultured OSNs would thus be extremely useful, enabling researchers to investigate the sensory neuron¿s activity in a controllable environment, avoiding obstacles imposed by the cellular heterogeneity found in sensory organs in vivo. In this study, we aimed at obtaining OSNs directly differentiated from mouse embryonic fibroblasts (MEF) using the forced expression of specific transcription factors via retroviral vectors. We therefore developed tools based on retroviral vectors with the objective of differentiating olfactory sensory neurons in vitro, using viral transduction in target cells (murine fibroblasts) with combinations of select transcription factors. Retroviruses were tested and some combinations of transcription factors were tested on a preliminary basis, which were capable of inducing molecular alterations on fibroblasts followed by the expression of olfactory sensory neuron markers
Mestrado
Genetica Animal e Evolução
Mestre em Genética e Biologia Molecular

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22

Wung, Nelly. "Tissue engineering of the liver." Thesis, University of Bath, 2017. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.715264.

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Currently, the only cure for liver failure is orthotopic liver transplantation. However, there are insufficient donor organs available to treat every patient on the transplant list and many die before they are able to receive a liver transplant. The bioartificial liver (BAL) device is a potential extracorporeal treatment strategy utilising hepatocytes or hepatocyte-like cells (HLCs) within a bioreactor to recapitulate normal liver function and therefore ‘bridge’ a patient with liver failure until they receive a transplant. The work in this thesis utilised tissue engineering methods to develop novel approaches to BAL device design through development and characterisation of a polymer membrane scaffold (“PX”) for hollow fibre bioreactor (HFB) culture and a HLC source generated from the transdifferentiation of pancreatic AR42J-B13 (B13) cells. A flat sheet membrane model was used for the development of asymmetrical, hydrophobic polystyrene (PS) phase inversion membranes. Oxygen plasma significantly increased PS membrane surface wettability through addition of oxygen functional groups to create an environment conducive for cell culture. The treated membrane was henceforth referred to as “PX”. The culture medium HepatoZYME+ was investigated for its ability to induce transdifferentiation of B13 cells to HLCs and maintain the hepatic phenotype. Overall, HepatoZYME+-cultured cells experienced viability loss. A diluted version, “50:50”, showed induction of the hepatic markers carbamoylphosphate synthetase-1 (CPS-1) and HNF4α, as well as a change towards a HLC morphology. When using 50:50 as a maintenance medium, transdifferentiated HLCs retained loss of pancreatic amylase and also induction of hepatic markers, with comparable serum albumin secretion to the established Dex + OSM treatment. However, culture viability in 50:50 was still compromised. Therefore, HepatoZYME+ based media were deemed unsuitable for induction and maintenance compared to Dex-based protocols. PX flat sheet membranes were able to support culture of B13 cells and also the human osteosarcoma cell line, MG63, demonstrating improved cell attachment over non-surface treated PS membranes. PX membranes supported transdifferentiation of B13 cells to HLCs, presenting with loss of pancreatic amylase, induction of the hepatic markers transferrin, GS and CPS-1 and serum albumin secretion. Furthermore, PX showed no change in mass or loss of culture surface area over 15 days in culture conditions. Together, the novel membrane material and the media formulation and feeding regime developed have strong potential to be translated to a HFB setting and guide future BAL device design.

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23

Cassady, John P. "Transdifferentiation of fibroblasts to neural stem cells." Thesis, Massachusetts Institute of Technology, 2013. http://hdl.handle.net/1721.1/83634.

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Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, 2013.
This electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections.
Cataloged from student-submitted PDF version of thesis.
Includes bibliographical references.
The developmental process is carefully controlled by transcriptional and epigenetic changes that occur as a zygote transforms into an adult organism. This process can be reversed by the overexpression of transcription factors Oct4, Sox2, Klf4, and c-Myc, which reprogram a differentiated cell!s nucleus to one that is transcriptionally and epigenetically indistinguishable from an embryonic stem (ES) cell. However, it is still unclear if transcription factors can completely convert the nucleus of a differentiated cell into that of a distantly related somatic cell type with complete transcriptional and epigenetic reprogramming maintained in the absence of exogenous factor expression. To test this idea, we generated doxycyline (dox)-inducible vectors encoding neural stem cell-expressed factors. We found that stable, self-maintaining NSC-like cells could be induced under defined growth conditions. These cells were characterized in the absence of exogenous factor induction and were shown to be transcriptionally, epigenetically, and functionally similar to endogenous embryonic cortical NSCs. Additionally, a cellular system was created for reproducible generation of doxindependent iNSCs without additional factor transduction. Our results show that a transcriptionally and epigenetically reprogrammed somatic nucleus can be stabilized in vitro and provides a tool to study the mechanism of somatic cell conversion.
by John P. Cassady.
Ph.D.

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24

Di, Stefano Bruno 1984. "C/EBPα poises B cells for rapid reprogramming into induced pluripotent stem cells." Doctoral thesis, Universitat Pompeu Fabra, 2014. http://hdl.handle.net/10803/283484.

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One of the major goals of current stem cell research is understanding the mechanism of somatic cell reprogramming by Oct4, Sox2, Klf4 and Myc (OSKM) into induced pluripotent stem cells (iPSCs). However, the finding that only a small proportion of the cells become reprogrammed, typically requiring >12 days, has hampered progress towards this goal. C/EBPα is a transcription factor specifically expressed in myelomonocytic cells within the hematopoietic system whose forced expression in B cells efficiently induces transdifferentiation into macrophages. We have now found that an 18-hour pulse of C/EBPα expression followed by OSKM activation induces an approximately 100-fold increase in the iPSC reprogramming efficiency, involving up to 95% of the cells within a week. Concomitantly, the cells undergo an epithelial-mesenchymal transition and pluripotency genes become upregulated to levels comparable to embryonic stem and iPS cells. In serum-free conditions the process is further accelerated, with 60% of the poised and OSKM induced B cells becoming Oct4-GFP positive within 2 days. These results are consistent with the idea that the C/EBPα pulse helps to overcome the stochastic phase of iPSC reprogramming. In addition, our work shed new light on the role of C/EBPα in induced pluripotency. Our data indicate that C/EBPα acts as a pathbreaker, at least in part mediated by the dioxygenase Tet2. C/EBPα binds to the Tet2 gene, induces its expression and translocates the protein to the nucleus. Here Tet2 binds to regulatory regions of pluripotency genes and converts methylated cytosine residues into hydroxymethylated cytosines. The pulse also renders the chromatin at regulatory sites of pluripotency genes accessible to DNase I digestion and, following OSKM induction, leads to local demethylation and to the binding of Oct4, correlating with the observed rapid upregulation of pluripotency genes. In line with an important role of Tet2 as a mediator of reprogramming, coexpression of the gene with OSKM enhanced B cell reprogramming substantially. The rapid and highly efficient iPSC reprogramming approach described herein should help to fully elucidate the early events of reprogramming to pluripotency and, if applicable to human cells, could have potential clinical applications.
Actualmente uno de los principales objetivos de la investigación con células madre es la comprensión de los mecanismos por los cuales las células somáticas se pueden reprogramar a células madre pluripotentes inducidas (iPSCs) por la acción de los factores de transcripción Oct4, Sox2, Klf4 y Myc (OSKM). Sin embargo, la baja eficiencia de este proceso, que tiene lugar sólo en un pequeño porcentaje de células y que típicamente requiere más de 12 días para llevarse a cabo, ha impedido la consecución de grandes avances en este campo en los últimos años. C/EBPα es un factor de transcripción específico de células del linaje mielo-monocítico del sistema hematopoyético. La expresión ectópica de esta proteína en células B puede inducir su transdiferenciación a macrófagos. En nuestro estudio de investigación hemos descubierto que la exposición de C/EBPα durante 18 horas seguida de la activación de OSKM, aumenta en 100 veces la eficiencia de reprogramación de las iPSC, resultando en la reprogramación del 95% de las células después de una semana. En detalle, durante este proceso de reprogramación las células experimentan una transición epitelio-mesénquima y los genes de pluripotencia se expresan en niveles comparables a los expresados en células madre embrionarias y iPSC. Cuando la reprogramación se lleva a cabo en medio de cultivo sin suero el proceso es aún más rápido, de tal modo que el 60% de las células B inducidas por C/EBPα y OSKM son positivas para Oct4-GFP en tan sólo dos días. Estos resultados apoyan la idea de que una exposición transitoria de C/EBPα ayuda a superar la fase estocástica de la reprogramación de las iPSC. Además, nuestros descubrimientos aclaran el papel de C/EBPα en el proceso de pluripotencia inducida, indicando que actúa como un catalizador, mediado en parte por la actividad de la dioxigenasa Tet2. De tal modo, que C/EBPα se une a regiones reguladoras del locus de Tet2, induciendo de esta manera su expresión y translocando la proteína al núcleo. Una vez en el núcleo, Tet2 se une a las regiones regulatorias de los genes de pluripotencia y convierte los residuos de citosinas metilados existentes en estas regiones en citosinas hidroximetiladas. Además, la exposición transitoria de C/EBPα deja la cromatina más accesible a la digestión con DNasa I alrededor de las regiones regulatorias de los genes de pluripotencia y, tras la inducción con OSKM, desencadena una demetilación local favoreciendo la posterior unión de Oct4 a estas regiones. Todo ello finalmente promueve la expresión concomitante de los genes de pluripotencia. Adicionalmente, en nuestro estudio se demuestra que la coexpresión de Tet2 y OSKM aumenta significativamente la reprogramación de las células B, lo cual se encuentra en línea con un papel importante de Tet2 en la reprogramación. En resumen, en este estudio se presenta el sistema de reprogramación de iPSC más rápido y eficiente descrito a día de hoy. El cual, facilitará la comprensión de los eventos precoces en el proceso de reprogramación a pluripotencia y, en el caso de que se pueda extrapolar a células humanas, podrá tener aplicaciones clínicas relevantes en el campo de la medicina regenerativa.

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Silva, Thaís Girão da. "Mecanismos moleculares envolvidos no fenótipo endotelial em resposta a estímulos físicos e químicos." Universidade de São Paulo, 2018. http://www.teses.usp.br/teses/disponiveis/5/5166/tde-22102018-104726/.

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O endotélio reveste a parede vascular e possui função essencial na manutenção da homeostase. A célula endotelial é capaz de perceber estímulos extracelulares, como fatores químicos e mecânicos, transmitir a informação para dentro da célula e regular sua função e fenótipo. Neste sentido, investigamos os mecanismos moleculares associados as células endoteliais em dois contextos importantes de intervenções vasculares 1) nos stents farmacológicos, onde a rapamicina exerce funções antiproliferativas e pró-trombogênicas, e 2) na revascularização cardíaca por ponte de safena, onde o alto estiramento mecânico exerce grande impacto no remodelamento vascular e no fenótipo da célula endotelial. A rapamicina pertence à classe de drogas limus, bastante utilizadas nos stents farmacológicos usados no procedimento de desobstrução vascular. Além de sua função antiproliferativa, exploramos os efeitos deletérios associados a pró-trombogênese. Os dados demonstraram que a rapamicina ativa o receptor de TGF independentemente de seu ligante TGFbeta, promovendo aumento na expressão da PAI-1 (pró-trombogênica), alteração no fenótipo endotelial (Transição endotélio-mesenquimal - EndMT) e na formação de fibras de estresse. Os efeitos observados são dependentes da ativação de Smad2 e independentes da via clássica antiproliferativa por mTOR. Experimentos in vivo mostraram que o tratamento com inibidor do receptor de TGF diminui os efeitos pró-trombogênicos e a expressão de PAI-1 induzidos pela rapamicina em artérias carótidas de camundongos. A ponte de safena é um procedimento bastante utilizado na cirurgia de revascularização cardíaca e a arterialização do segmento venoso submetido ao estresse hemodinâmico arterial resulta em remodelamento vascular, que influencia o sucesso do procedimento. Nossos dados demonstram que a célula endotelial humana de veia safena humana (hSVEC), susceptível as modificações do tipo EndMT induzido quimicamente (estímulo pró-fibrótico e pró-inflamatório), não expressou o mesmo comportamento em resposta ao aumento de estiramento mecânico que ocorre durante a arterialização venosa. Entretanto, detectamos uma pronunciada redução dos filamentos de actina, modulação no padrão de ativação da cofilina e na proporção de actina glomerular (G-actina) entre citoplasma e núcleo, com redução da biodisponibilidade de NO. De modo interessante, demonstramos que a redução no filamento de actina é específica para a célula endotelial venosa, não sendo observado em células endoteliais de origem arterial de aorta e coronária. Em conjunto, os dados mostram que 1) efeitos pró-trombogênicos associados a rapamicina são mediados por ativação do receptor de TGF independente do seu ligante e da atividade antiproliferativa da droga e 2) a adaptação da célula endotelial venosa ao estiramento mecânico envolve modulação da síntese/degradação de filamentos de actina e redução na biodisponibilidade de NO. Estes novos elementos sobre o mecanismo de transdução de estímulos químicos e físicos pelo endotélio poderão ser explorados terapeuticamente para modular a plasticidade endotelial em disfunções cardiovasculares
Endothelium is the inner layer in vascular wall and displays an essential role in the maintenance of cardiovascular homeostasis. Endothelial cell senses the extracellular stimuli, such as chemical and mechanical factors, transduce and process these signals to regulate cell function and phenotype. Here, we investigated molecular underpinning of the endothelial cells under two important scenarios: 1) in drug-eluting stents, where rapamycin exerts antiproliferative and undesirable prothrombogenic functions, and 2) in vein graft bypass surgery, where increased stretch modulates vascular remodeling and endothelial cell phenotype. Rapamycin belongs to the class of limus drugs and is widely used in drug eluting stents (DES) to vascular restenosis. In addition to its antiproliferative function, we explore the deleterious effects associated with prothrombogenesis. Our data demonstrated that rapamycin activates TGF receptor independent of its ligand TGFbeta, in concert with promotion of PAI-1 expression (prothrombogenic), changes in endothelial phenotype (Endothelial to Mesenchymal Transition - EndMT) and stress fibers induction. These effects are Smad2 dependent and independent of the classical antiproliferative mTOR pathway of rapamycin. Our in vivo experiments showed that TGF receptor inhibitor treatment decreases prothrombogenic effects and PAI-1 expression induced by rapamycin in mice carotid arteries. Saphenous vein is widely used in coronary artery bypass surgery (CABG) and the vein arterialization remodeling in response to the increased stress influences graft patency. Our data demonstrated that human saphenous vein endothelial cell (hSVEC) is susceptible to chemically induced endothelial-to-mesenchymal transition (EndMT) by pro-fibrotic and pro-inflammatory stimuli. On the other hand, physical stimulus associated with high stretch failed to induce EndMT. However, we detected a pronounced decrease of actin filaments, modulation of the cofilin activation, changes in the proportion of glomerular actin (G-actin) between cytoplasm and nucleus, and reduction of NO bioavailability. Interestingly, the reduction of actin fibers by high stretch is specific to venous endothelial cell since arterial endothelial cells from aorta, and coronary artery failed to display the response. Altogether, our data show that 1) the thrombogenic effects of rapamycin are mediated by TGF receptor activation independent of its ligand and independent of the antiproliferative pathway of the drug, and 2) the adaptation of venous endothelial cell to mechanical stretch involves synthesis/degradation of actin filaments and reduced NO bioavailability. These new elements on signal transduction of endothelial cells in response to chemical and physical stimuli may be therapeutically explored to modulate endothelial plasticity in cardiovascular disorders

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26

O'Neill, Kathy. "Reprogramming hepatocytes into duct-like cells." Thesis, University of Bath, 2010. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.528369.

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Primary hepatocytes maintained in culture progressively down regulate liver-specific genes and lose their characteristic function and morphology. This process, termed dedifferentiation, is a hindrance to in vitro modelling of systems such as xenobiotic metabolism, liver disease and regeneration. However the results presented here demonstrate that dedifferentiated hapatocytes spontaneously induce expression of ductal genes, and therefore represent a useful model of cell reprogramming.

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de, Back Walter, Roland Zimm, and Lutz Brusch. "Transdifferentiation of pancreatic cells by loss of contact-mediated signaling." Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2014. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-127265.

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Background: Replacement of dysfunctional β-cells in the islets of Langerhans by transdifferentiation of pancreatic acinar cells has been proposed as a regenerative therapy for diabetes. Adult acinar cells spontaneously revert to a multipotent state upon tissue dissociation in vitro and can be stimulated to redifferentiate into β-cells. Despite accumulating evidence that contact-mediated signals are involved, the mechanisms regulating acinar-to-islet cell transdifferentiation remain poorly understood. Results: In this study, we propose that the crosstalk between two contact-mediated signaling mechanisms, lateral inhibition and lateral stabilization, controls cell fate stability and transdifferentiation of pancreatic cells. Analysis of a mathematical model combining gene regulation with contact-mediated signaling reveals the multistability of acinar and islet cell fates. Inhibition of one or both modes of signaling results in transdifferentiation from the acinar to the islet cell fate, either by dedifferentiation to a multipotent state or by direct lineage switching. Conclusions: This study provides a theoretical framework to understand the role of contact-mediated signaling in pancreatic cell fate control that may help to improve acinar-to-islet cell transdifferentiation strategies for β-cell neogenesis.

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Li, Wan-Chun. "In vitro transdifferentiation of liver into functional pancreatic-like cells." Thesis, University of Bath, 2006. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.425508.

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de, Back Walter, Roland Zimm, and Lutz Brusch. "Transdifferentiation of pancreatic cells by loss of contact-mediated signaling." BioMed Central, 2013. https://tud.qucosa.de/id/qucosa%3A27291.

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Background: Replacement of dysfunctional β-cells in the islets of Langerhans by transdifferentiation of pancreatic acinar cells has been proposed as a regenerative therapy for diabetes. Adult acinar cells spontaneously revert to a multipotent state upon tissue dissociation in vitro and can be stimulated to redifferentiate into β-cells. Despite accumulating evidence that contact-mediated signals are involved, the mechanisms regulating acinar-to-islet cell transdifferentiation remain poorly understood. Results: In this study, we propose that the crosstalk between two contact-mediated signaling mechanisms, lateral inhibition and lateral stabilization, controls cell fate stability and transdifferentiation of pancreatic cells. Analysis of a mathematical model combining gene regulation with contact-mediated signaling reveals the multistability of acinar and islet cell fates. Inhibition of one or both modes of signaling results in transdifferentiation from the acinar to the islet cell fate, either by dedifferentiation to a multipotent state or by direct lineage switching. Conclusions: This study provides a theoretical framework to understand the role of contact-mediated signaling in pancreatic cell fate control that may help to improve acinar-to-islet cell transdifferentiation strategies for β-cell neogenesis.

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Meyer, Lionel. "Contribution des cellules souches de glioblastome à l'hétérogénéité tumorale : aspect thérapeutique et développement d'un système d'expression mosaïque fluorescent." Thesis, Strasbourg, 2016. http://www.theses.fr/2016STRAJ109/document.

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Le glioblastome (GBM) est la tumeur cérébrale primaire la plus agressive comportant une sous-population de cellules souches tumorales (CSG). Elles sont capables d’auto-renouvellement, de prolifération, de différenciation en cellules exprimant les marqueurs neuraux et de trans-différenciation en cellules de types vasculaires. Dans ce contexte, j’ai dérivé et caractérisé plusieurs lignées de CSG à partir de biopsies de patients. Puis j’ai évalué l’impact des peptides thérapeutiques transmembranaires développés au laboratoire, visant les plateformes de récepteurs de neuropiline-1 et de plexine-A1 surexprimées dans les CSG. Les deux peptides diminuent la croissance des CSG in vitro et in vivo. Finalement, j’ai développé un outil génétique fluorescent permettant de suivre le destin des CSG en direct. Basé sur l’expression de 4 rapporteurs fluorescents contrôlés par des promoteurs spécifiques des types cellulaires, il permet d’identifier l’hétérogénéité de ces cellules en différenciation
The glioblastoma multiforme (GBM) is the most aggressive primary brain tumor and includes a subpopulation of tumoral stem cells (CSG). Those cells can self-renew, proliferate and differentiate by expressing specific neural markers and/or transdifferentiate into vascular-like cells. In this context, my work consisted first to produce and characterize several CSG lines from patient biopsies to constitute a bank of cell lines with different properties. We also evaluated the impact of in house therapeutic transmembrane peptides targeting the neuropilin-1 / plexin-A1 receptor platforms overexpressed in GBM. We thus showed that both targeting peptides decrease the growth of GSC in in vitro and in vivo models. Finally, I developed an inducible mosaic expression system to track the live differentiation of CSG. This system is based on the expression of four different fluorescent reporters controlled by the activity of cell type specific promoters

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Prahalad, Priya. "9-cis-retinoic acided mediated endothelial transdifferentiation in breast cancer cells." Connect to Electronic Thesis (CONTENTdm), 2008. http://worldcat.org/oclc/645463979/viewonline.

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Al-Adsani, Amani Mustafa. "Transdifferentiation of pancreatic AR42J-B13 cells to hepatocyte and ductal phenotypes." Thesis, University of Bath, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.503389.

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Dawes, Lucy Jean. "TGFÎ² induced transdifferentiation and matrix contraction of human lens epithelial cells." Thesis, University of East Anglia, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.433913.

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34

Martínez, Romero Carles. "Polycomb group proteins Bmi1 and Ring1B are involved in cell plasticity and tumorigenesis of the pancreas." Doctoral thesis, Universitat Pompeu Fabra, 2009. http://hdl.handle.net/10803/7190.

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L'adenocarcinoma ductal pancreàtic (PDAC) és un dels càncers més letals. Per tal de millorar el diagnòstic precoç, s'estan investigant les etapes inicials de la formació del càncer, com és el cas de les lesions preneoplàstiques, i es vol desxifrar l'origen cel·lular de la malaltia. Les proteïnes Polycomb constitueixen una família de silenciadors epigenètics que es troben en una varietat de tumors sòlids. La hipòtesi principal és que Polycomb pot estar participant en els processos preneoplàstics del pàncreas i en l'aparició i progressió del tumor. La expressió de Bmi1 i Ring1B fou analitzada durant el desenvolupament del pàncreas, en teixit pancreàtic de diferents models murins de la malaltia i en mostres humans de teixit pancreàtic. Es va dur a terme l'anàlisi del mecanisme de Bmi1 mitjançant models in vitro i induint la depleció de Bmi1. Bmi1 i Ring1B s'expressaren en precursors pancreàtics durant etapes primerenques del desenvolupament i en cèl·lules ductals i dels illots,
però no en els acins, en el pàncrees adult. Bmi1 s'induí en cèl·lules acinars durant lesió aguda, en lesions metaplàstiques acinoductals, en neoplàsies intraepitelials pancreàtiques (PanIN) i en PDAC. Ring1B s'incrementà significativament en PanINs de grau alt i en PDAC. La disminució dels nivells de Bmi1 en la línia cel·lular acinar canvià l'expressió dels enzims digestius pancreàtics. Aquests resultats suggereixen que Bmi1 i Ring1B podrien estar contribuint de diferent manera en la progressió tumoral.
Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers. To improve early diagnosis, research efforts are focused in characterising early events of cancer formation like preneoplastic lesions and deciphering the cell origin of the malignancy. Polycomb proteins constitute a family of epigenetic silencers found in a variety of solid tumours. The main hypothesis is that Polycomb might play a role in preneoplastic states in the pancreas and in tumour development and progression. The expression of Bmi1 and RingB was analysed during pancreatic development, in pancreatic tissue from mouse models of disease and in human pancreatic tissue samples. Mechanistic insights of Bmi1 were performed using in vitro models and with induced Bmi1 depletion. Bmi1 and Ring1B were expressed in pancreatic exocrine precursors during early development and in ductal and islet cells, but not in acinar cells, in the adult pancreas. Bmi1 was induced in acinar cells during acute injury, in acinar-ductal metaplastic lesions, in pancreatic intraepithelial neoplasia (PanIN) and PDAC. In contrast, Ring1B was significantly increased in high-grade PanINs and in PDAC. Bmi1 knockdown in acinar cell line changed the expression of pancreatic digestive enzymes. These results suggest that Bmi1 and Ring1B could contribute differently to tumour development.

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Sangan, Caroline Beth. "Reprogramming of hepatic and pancreatic cells." Thesis, University of Bath, 2012. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.582549.

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Cell therapy involving treatment of diseases with the body’s own cells would benefit both liver diseases and Type 1 diabetes. Liver diseases are associated with a marked reduction in hepatocytes whilst Type 1 diabetes is characterized by the loss of functional insulin-producing β-cells. Treatment is currently achieved by whole organ liver (or hepatocyte) and islet transplantation methods respectively. However the major limitation to this approach is the shortage of organ donors, thus alternative sources of cells must be found. Potential sources with enormous therapeutic potential are existing cells in the liver and pancreas involved during the regeneration process. In vivo studies have shown progenitor oval cells differentiate into hepatocytes during liver regeneration and α-cells transdifferentiate into β-cells during pancreas regeneration. However neither can be fully exploited until the molecular mechanisms governing their proliferation and trans/differentiation are fully elucidated. Herein we characterise two in vitro cell models, a mouse adult oval cell line, known as BMOL-TAT1.1, and mouse adult pancreatic α-cell line, known as α-TC19 by RT-PCR and immunofluorescent staining. We found that under proliferating culture conditions BMOL-TAT1.1 were heterogenous consisting of two distinct cell types with different β-catenin signalling pathway activation. Inducible differentiation (dexamethasone) induced hepatic and non-hepatic markers in specific cell subtypes, indicating multi-potentiality. Ectopic expression of transcription factor HNF4α in homogenous small BMOL-TAT1.1 cells revealed no hepatic differentiation but potent expression of intestinal markers (Villin, ALPi, ApoAIV). HNF4α was identified as a candidate transcriptional regulator in α- to β-cell transdifferentiation, as ectopic expression in α-TC19 cells, suppressed glucagon and induced expression of several functionally important β-cell markers (GLUT2, GCK, insulin). The contribution of chromatin histone acetylation was also assessed, due to its importance in endocrine fate regulation. In toto these results have important implications for the development of potential therapies to treat liver diseases and Type 1 diabetes.

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Simonneau, Lionel. "Etude de l'expression des cristallines et de leurs proprietes aggregatives dans les cultures de cellules epitheliales de cristallin de boeuf et de la neurotine embryonnaire de caille normale ou transformee par des retrovirus aviaires." Paris 7, 1988. http://www.theses.fr/1988PA077154.

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37

Duran, Jason Mathew. "Bone-derived stem cells repair the heart after myocardial infarction through transdifferentiation and paracrine signaling mechanisms." Diss., Temple University Libraries, 2015. http://cdm16002.contentdm.oclc.org/cdm/ref/collection/p245801coll10/id/253042.

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Physiology
Ph.D.
Rationale: Autologous bone marrow- or cardiac-derived stem cell therapy for heart disease has demonstrated safety and efficacy in clinical trials but has only offered limited functional improvements. Finding the optimal stem cell type best suited for cardiac regeneration remains a key goal toward improving clinical outcomes. Objective: To determine the mechanism by which novel bone-derived stem cells support the injured heart. Methods and Results: Cortical bone stem cells (CBSCs) and cardiac-derived stem cells (CDCs) were isolated from EGFP+ transgenic mice and were shown to express c-kit and Sca-1 as well as 8 paracrine factors involved in cardioprotection, angiogenesis and stem cell function. Wild-type C57BL/6 mice underwent sham operation (n=21) or myocardial infarction (MI) with injection of CBSCs (n=57), CDCs (n=31) or saline (n=57). Cardiac function was monitored using echocardiography with strain analysis. EGFP+ CBSCs in vivo were shown to express only 2/8 factors tested (basic fibroblast growth factor and vascular endothelial growth factor) and this expression was associated with increased neovascularization of the infarct border zone. CBSC and CDC therapy improved survival, cardiac function, attenuated adverse remodeling, and decreased infarct size relative to saline-treated MI controls. CBSC treated animals showed the most pronounced improvements in all parameters. By 6 weeks post-MI, EGFP+ cardiomyocytes, vascular smooth muscle cells and endothelial cells could be identified on histology in CBSC-treated animals but not in CDC-treated animals. EGFP+ myocytes isolated from CBSC-treated animals were smaller, more frequently mononucleated, and demonstrated fractional shortening and calcium currents indistinguishable from EGFP- myocytes from the same hearts. Conclusions: CBSCs improve survival, cardiac function, and attenuate remodeling more so than CDCs and this occurs through two mechanisms: 1) secretion of the proangiogenic factors bFGF and VEGF (which stimulates endogenous neovascularization), and 2) differentiation into functional adult myocytes and vascular cells.
Temple University--Theses

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38

Bhavsar, Rital. "Newt Lens Regeneration: Role of Oct-4 in Newt Regenerating Tissue and Proteome Analysis of Regeneration Competent Vs. Regeneration Incompetent Cells." University of Dayton / OhioLINK, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=dayton1397131576.

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39

Rako, Zvonimir Andelko [Verfasser]. "miRNA-154 mediates the transdifferentiation of alveolar type II to alveolar type I cells in the mouse model of Bronchopulmonary Dysplasia / Zvonimir Andelko Rako." Gießen : Universitätsbibliothek, 2020. http://d-nb.info/1219983101/34.

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40

Rako, Zvonimir A. [Verfasser]. "miRNA-154 mediates the transdifferentiation of alveolar type II to alveolar type I cells in the mouse model of Bronchopulmonary Dysplasia / Zvonimir Andelko Rako." Gießen : Universitätsbibliothek, 2020. http://d-nb.info/1219983101/34.

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Baddour, Joelle. "An Approach to Lens Regeneration in Mice Following Lentectomy and the Implantation of a Biodegradable Hydrogel Encapsulating Iris Pigmented Tissue in Combination with Basic Fibroblast Growth Factor." University of Dayton / OhioLINK, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=dayton1335916825.

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42

Santos, Filipa Inês Rodrigues dos. "Towards direct transdifferentiation of adult human cells to the pancreatic B-cell fate." Master's thesis, 2013. http://hdl.handle.net/10400.1/7273.

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Dissertação de mestrado, Ciências Biomédicas, Departamento de Ciências Biomédicas e Medicina, Universidade do Algarve, 2013
Type 1 Diabetes mellitus (T1DM) is one of the most widespread metabolic disorders with epidemic dimension affecting almost 6% of the world’s population. Autoimmune reaction causes selective destruction of the insulin-producing β-cells within the pancreatic islets, leading to both acute and long-term complications. Daily insulin injections treat but no dot cure diabetes. Any progress in obtaining large number of transplantable insulin producing cells would be a major advance towards a cure for the disease. In order to create an alternative source of β-cells we are developing a method to transdifferentiate adult human cells to the beta cell phenotype by direct reprogramming mediated by forced expression of an optimized set of pancreas specific transcription factors. The underlying experimental rationale is that sequential or combinatorial ectopic expression of transcription factors can induce recipient cells to establish a β-cell regulatory state. We cloned a set of transcription factors known to be involved in pancreatic development into viral vectors and used them to transdifferentiate adult human cell types in conditions known to favor β-cell differentiation
A diabetes mellitus tipo 1 (DM1) é uma das alterações metabólicas mais comum a nível mundial, afectando quase 6% da população mundial. É uma doença auto-imune que tem como resultado a destruição das células β do pâncreas, produtoras de insulina. Caracteriza-se por hiperglicémia e insuficiência na produção de insulina, levando a complicações quer a curto quer a longo prazo. Não existe cura para a DM1. O único tratamento para os doentes com DM1 é a administração diária de insulina. No entanto, a administração de insulina está frequentemente associada a episódios graves de hipoglicémia, para além de que não previne o aparecimento das complicações crónicas associadas à doença, tais como a retinopatia diabética e a doença renal terminal. A obtenção em larga escala de células produtoras de insulina poderá ser um grande avanço para a cura da diabetes. Um dos métodos mais inovadores para a obtenção de células β é a transdiferenciação de células humanas adultas em células β produtoras de insulina, através da expressão ectópica de um conjunto de factores de transcrição que estão envolvidos no estabelecimento destas células. O procedimento experimental baseou-se na expressão ectópica destes factores de transcrição para induzir células humanas adultas a estabelecer o estadio regulatório que define as células β. Para tal, foi clonada uma coleção de factores de transcrição envolvidos no desenvolvimento pancreático em vectores virais. Estes vectores foram posteriormente utilizados na transdiferenciação de células humanas adultas em condições que favorecem a diferenciação das células β.

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Medina, Abelardo. "Circulating bone marrow-derived precursor cells modulate the wound healing outcome by cell transdifferentiation." 2009. http://hdl.handle.net/2429/12657.

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Normal wound healing process is a regulated biological response of multiple events whose common aim is to restore the integrity of injured tissues. Fibroblasts exert an important role in this process as target cells that allow either tissue remodeling through the predominant production of proteases (i.e., matrix metalloproteinases) or tissue fibrosis through the over-expression of extracellular matrix (ECM) components such as collagens. However, it remains unclear which factors activate such diversity of fibroblast responses and how this decision making process is made. Currently, there is no a well established model that integrally explains the diversity of responses from minimal to hypertropic scarring and keloids. Previous reports have demonstrated that some recruited cells can be locally transformed into fibrocytes, a pro-fibrogenic cells that stimulate resident fibroblasts to produce collagen accumulation and tissue fibrosis. However, recruited cells with anti-fibrogenic profile that can compete with and eventually reverse the local effects of fibrocytes have not been identified. How the skin maintains the cell transdifferentiation balance in normal wound healing and how this balance is modified in fibro-proliferative cutaneous disorders cannot be entirely explained by pro-fibrogenic fibrocytes without anti-fibrogenic counterparts. This doctoral thesis hypothesizes a novel mechanism in which bone marrow-derived cells recruited to the injured area modulate the expression of ECM components produced by resident fibroblasts. As a result of the tissue injury, a repertoire of systemic and local cytokines and growth factors induce epigenetic changes and cell transdifferentiation in circulating recruited cells. Thus, these locally transformed cells can acquire either pro- or anti-fibrogenic profile, and more importantly, they can induce the production of either collagens or MMPs by dermal fibroblasts. This study demonstrates that circulating stem cells and monocytes have the capacity to transdifferentiate into keratincoyte-like cells (KLCs), anti-fibrogenic cells that increase the expression MMPs by themselves and by the stimulation of dermal fibroblasts. The transdifferentiation of pro-fibrogenic fibrocytes into KLCs was also studied. In all cases the TGF-β deprivation was identified as crucial factor in the anti-fibrotic commitment of recruited cells. Finally, adipofascial groin flaps in rats were utilized as in vivo model to study the role of the tissue repair microenvironment in the cell transdifferentiation of recruited bone marrow-derived cells. The findings presented in this thesis are consistent with the existence of a "seesaw mechanism" in the regulation of MMPs/collagen production by dermal fibroblasts. Thus, during the wound-healing response, the local environment may induce epigenetic changes in recruited bone marrow-derived cells to follow either pro- or anti-fibrogenic pathways. Subsequently, these committed cells may trigger a fibro-proliferative switch on resident fibroblasts to predominantly develop either collagen accumulation with tissue fibrosis or collagen breakdown with tissues remodeling. Findings of this doctoral thesis provide new insights into the role of cell transdifferentiation and local environment not only in wound healing, but may also in other fibro-proliferative processes such as lung fibrosis, asthma, liver cirrhosis, chronic pancreatitis, and atherosclerosis, among others.

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Yin, Shu-Yi, and 尹書翊. "Use of Combinational Omics Approach for Studying Stimulatory Effects of Phyto-chemicals/-extracts on Dendritic cell immunity and Cell Transdifferentiation." Thesis, 2012. http://ndltd.ncl.edu.tw/handle/36421536270565608945.

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博士
國立中興大學
生物科技學研究所
100
Various omics approaches have been employed and are still considered as highly desirable and useful tools for herbal medicine research, presumably because conventional phytomedicines often utilized multi-factoral formulations, and such phytochemical mixtures are blieved to interact with multiple molecular targets, resulting in composite (e.g., immunomodulatory) effects. In order to globally and integratively evaluate the effects of specific herbal extracts or phytomedicines, in this study various omics approaches including genomics, proteomics, and the associated bio-informatics tools were systematically combined to develop my research systems. For the current study, a specific fraction extracted from a popular western medicinal plant, Echinacea purpurea, namely [BF/S+L/Ep], was investigated for its immunomodulatory effects on mouse bone marrow-derived dendritic cells (BMDCs). Various cellular and molecular effects were revealed using a network knowledge-based approach, and analyzed at the level of genome-wide transcriptome, specific proteome and functional phenotype activities. My experimental results suggest that [BF/S+L/Ep] can efficiently enhance DC mobility and the related cellular physiology in vivo. Moreover, the signaling networks and molecules highlighted in this study provided potentially useful targets for nutritional or clinical application of Echinacea or other medicinal plants for immune-modulation activities. The next part of my study was aimed at improving the in vitro differentiation technology for mouse BMDCs, an important pre-clinical model for future development of DC-based therapeutics. I demonstrate here that IL-4, a key driving factor for the in vitro differentiation of BMDCs, can specifically enhance the in vivo trafficking activity of BMDCs during a late stage in cultivation. With such temporal control of IL-4 stimulation, much less cytokine quantity is needed to generate a high yield of BMDCs with increased purity, increased secretion level of cytokines and a higher capacity to induce proliferation of allogeneic CD4+ T cells, as compared to the conventional method which utilizes a continuous supplement of IL-4 throughout the in vitro cultivation period. For the final part of my study, I investigate the in vivo biological effects of shikonin on mouse skin tissues, now appreciated as a highly important organ for metabolic regulations of the whole body system. Through a cross-examination between the transcriptome and microRNA data sets, I show that topical treatment of shikonin in skin with shikonin can drastically enhance epithelial–mesenchymal transition (EMT) activity and suppress the associated-microRNAs expression in vivo in test skin tissues. I consider these results and findings have provided us with good evidence in supporting of various previous observations on the wound-healing and immune-modulatory activities conferred by shikonins, in vivo and in vitro.

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45

"Role of NG2 expressing cells in murine terminal phalanx regeneration." Tulane University, 2013.

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Research using the adult mammalian model shows that regeneration in the limb is limited to the distal most portion of the terminal phalanx. Recent studies suggest that the cellular contributions made to the regenerating system are lineage restricted and that the niche bone marrow hematopoietic stem cell population’s contributions are minimal. These studies however, do not address other residing populations within the bone marrow, specifically the mesenchymal and endothelial stem cell populations. One of the residing populations, the reputed pericyte or perivascular cells, possesses the ability to differentiate into multiple other cell types. To assess the potential contribution of perivascular cells to the regeneration competency of the terminal phalanges, we began by identifying perivascular cells within the terminal phalanx by using two accepted pericyte markers: nerve-glial antigen 2 (NG2) and endosialin (TEM1). Using NG2 and TEM1 in conjunction with vascular marker Tie2 in the Tie2-EGFP murine reporter line, we confirm a large number of perivascular cells in the bone marrow’s unusually well-developed and organized vasculature and a lower density within the connective tissue microvasculature; implicating a great potential contribution from the bone marrow. Post-amputation, we observe a large population of NG2+ and TEM1+ cells within the regenerating blastema region. Co-immunohistochemical studies reveal the blastema have cells that co-express osteogenic and pericyte markers; strongly suggestive of a transdifferentiation event. We attempt to confirm our hypotheses made in our initial assessment by utilizing two independent cell tracing studies: a DiI labeling of the bone marrow of the terminal phalanx to identify a marrow derived cellular contribution to the regenerate and a genetic fate tracing study using transgenic NG2CreERTamR26REYFP mice to confirm a transdifferentiation event. Using a novel in vivo method , we DiI-label the bone marrow content before amputation and trace DiI labeled bone marrow derived cellular contributions to the regenerate. DiI labeled cells were observed within the blastema expressing either endothelial, perivascular, or osteogenic markers, confirming the bone marrow contributes multiple cell types during the regeneration process. Using a similar experimental design, we genetically label the terminal phalanx NG2 expressing cells using systemic tamoxifen induction of NG2CreERTamR26REYFP mice. We fate trace the initially labeled population during blastema formation and re-differentiation and observed transdifferentiation events of the perivascular cells into two distinctive lineages, endothelial and osteoprogenitor cells. Establishing a direct correlation between peri-vasculature and re-differentiation, we address NG2/perivascular necessity with a series of temporal loss of function studies using a blocking antibody (iNG2). We implant iNG2 soaked microcarrier beads into various regions of the terminal phalanx and during different stages of the regeneration process. The experiments confirm the necessity of NG2 expression for distal bone elongation, as well as ascertain the temporal nature of the NG2 expression in different microenvironments. These results establish the importance of NG2+ cells in the bone marrow during early stages of regeneration, with early iNG2 bone marrow implantation resulting in a complete failure of the regeneration process. In an attempt to rescue this iNG2 failed regeneration we employ an established position-specific fibroblast cell line that displays a surprising plasticity as a cell-based therapeutic. Through a series of RNAi lentiviral transfection of inhibitors of the TGFβ-BMP pathway we induce osteogenic plasticity in the line. These results reveal regeneration competency associated with the mammalian terminal phalanx is in part due to the ability to recruit local perivascular multipotent populations, which has great translational relevancy.
acase@tulane.edu

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46

Ye, Lihua. "The roles of pancreatic hormones in regulating pancreas development and beta cell regeneration." 2015. http://hdl.handle.net/1805/8030.

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Indiana University-Purdue University Indianapolis (IUPUI)
Diabetes mellitus is a group of related metabolic diseases that share a common pathological mechanism: insufficient insulin signaling. Insulin is a hormone secreted from pancreatic β cells that promotes energy storage and consequently lowers blood glucose. In contrast, the hormone glucagon, released by pancreatic α cells, plays a critical complementary role in metabolic homeostasis by releasing energy stores and increasing blood glucose. Restoration of β cell mass in diabetic patients via β cell regeneration is a conceptually proven approach to finally curing diabetes. Moreover, in situ regeneration of β cells from endogenous sources would circumvent many of the obstacles encountered by surgical restoration of β cell mass via islet transplantation. Regeneration may occur both by β cell self-duplication and by neogenesis from non-β cell sources. Although the mechanisms regulating the β cell replication pathway have been highly investigated, the signals that regulate β cell neogenesis are relatively unknown. In this dissertation, I have used zebrafish as a genetic model system to investigate the process of β cell neogenesis following insulin signaling depletion by various modes. Specifically, I have found that after their ablation, β cells primarily regenerate from two discrete cellular sources: differentiation from uncommitted pancreatic progenitors and transdifferentiation from α cells. Importantly, I have found that insulin and glucagon play crucial roles in controlling β cell regeneration from both sources. As with metabolic regulation, insulin and glucagon play counter-balancing roles in directing endocrine cell fate specification. These studies have revealed that glucagon signaling promotes β cell formation by increasing differentiation of pancreas progenitors and by destabilizing α cell identity to promote α to β cell transdifferentiation. In contrast, insulin signaling maintains pancreatic progenitors in an undifferentiated state and stabilizes α cell identity. Finally, I have shown that insulin also regulates pancreatic exocrine cell development. Insufficient insulin signaling destabilized acinar cell fate and impairs exocrine pancreas development. By understanding the roles of pancreatic hormones during pancreas development and regeneration can provide new therapeutic targets for in vivo β cell regeneration to remediate the devastating consequences of diabetes.

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47

Coetzee, Magdalena. "Differential effects of arachidonic acid and docosahexaenoic acid on cell biology and osteoprotegerin synthesis in osteoblast-like cells." Thesis, 2005. http://upetd.up.ac.za/thesis/available/etd-03092006-114304.

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48

Schilling, Tatjana. "Transdifferentiation of Human Mesenchymal Stem Cells." Doctoral thesis, 2007. https://nbn-resolving.org/urn:nbn:de:bvb:20-opus-24299.

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With ageing, the loss of bone mass correlates with the expansion of adipose tissue in human bone marrow thus facilitating bone-related diseases like osteopenia and osteoporosis. The molecular mechanisms underlying these events are still largely unknown. Reduced osteogenesis and concurrently enhanced adipogenesis might not only occur due to the impairment of conventional osteogenic differentiation originating from mesenchymal stem cells (MSCs). Additionally, transdifferentiation of (pre-)osteoblasts into adipocytes could contribute to the fatty conversion. Therefore, the aim of the present study was to prove the existence of transdifferentiation between the adipogenic and osteogenic lineage and to elucidate molecular mechanisms underlying this phenomenon. At first, a cell culture system of primary human MSCs was established that allowed for differentiation into the adipogenic and osteogenic lineage and proved that the MSC-derived adipocytes and pre-osteoblasts were capable of transdifferentiation (reprogramming) from one into the other lineage. Thereby, lineage-specific markers were completely reversed after reprogramming of pre-osteoblasts into adipocytes. The osteogenic transdifferentiation of adipocytes was slightly less efficient since osteogenic markers were present but the adipogenic ones partly persisted. Hence, plasticity also reached into the differentiation pathways of both lineages and the better performance of adipogenic reprogramming further supported the assumption of its occurrence in vivo. The subsequent examination of gene expression changes by microarray analyses that compared transdifferentiated cells with conventionally differentiated ones revealed high numbers of reproducibly regulated genes shortly after initiation of adipogenic and osteogenic reprogramming. Thereof, many genes were correlated with metabolism, transcription, and signal transduction as FGF, IGF, and Wnt signalling, but only few of the established adipogenesis- and none of the osteogenesis-associated marker genes were detected within 24 h after initiation of transdifferentiation. To find possible key control factors of transdifferentiation amongst the huge amount of regulated genes, a novel bioinformatic scoring scheme was developed that ranked genes due to their potential relevance for reprogramming. Besides the reproducibility and level of their regulation, also the possible reciprocity between the adipogenic and osteogenic transdifferentiation pathway was taken into account. Fibroblast growth factor 1 (FGF1) that ranked as one of the leading candidates to govern reprogramming was proven to inhibit adipogenic differentiation as well as adipogenic transdifferentiation in our cell culture system. Further examination of the FGF signalling pathway and other highly ranked genes could help to better understand the age-related fatty degeneration at the molecular level and therefore provide target molecules for therapeutic modulation of the plasticity of both lineages in order to inhibit adipogenic degeneration and to enhance osteogenesis
Der Verlust an Knochenmasse im Alter ist mit der Ausbreitung von Fettgewebe im menschlichen Knochenmark assoziiert und fördert daher auch knochenspezifische Erkrankungen wie Osteopenie und Osteoporose. Die diesen Ereignissen zu Grunde liegenden Mechanismen sind immer noch weitgehend unbekannt. Die abnehmende Osteogenese und die gleichzeitig zunehmende Adipogenese treten wahrscheinlich nicht nur wegen der Beeinträchtigung der konventionellen osteogenen Differenzierung von mesenchymalen Stammzellen (MSZ) auf. Zusätzlich könnte auch die Transdifferenzierung (Reprogrammierung) von Osteoblasten(vorläufern) zu Adipozyten zur fettigen Umwandlung beitragen. Das Ziel der vorliegenden Studie war es daher, die Existenz der Transdifferenzierung zwischen dem adipogenen und osteogenen Differenzierungsweg nachzuweisen und die molekularen Mechanismen aufzuklären, die diesem Phänomen zu Grunde liegen. Zunächst wurde ein Zellkultursystem primärer mesenchymaler Stammzellen etabliert, in dem eine Differenzierung zu Adipozyten und Osteoblasten durchgeführt werden konnte, und nachgewiesen, dass aus MSZ erhaltene Adipozyten und Osteoblastenvorläufer von einer zur anderen Zelllinie transdifferenziert (reprogrammiert) werden können. Dabei wurden die zelllinienspezifischen Marker nach der Reprogrammierung von Osteoblastenvorläufern zu Adipozyten vollständig umgekehrt. Die osteogene Transdifferenzierung von Adipozyten war etwas weniger effizient, da die osteogenen Marker zwar vorhanden waren, aber auch die adipogenen Marker weiterhin auftraten. Die Plastizität erstreckte sich also auch auf die Differenzierungswege der beiden Zellpopulationen, wobei das bessere Ergebnis bezüglich der adipogenen Reprogrammierung die Annahme ihres Auftretens in vivo weiter unterstützte. Die nachfolgende Untersuchung von Genexpressionsänderungen mittels Mikroarray-Analysen, die transdifferenzierte mit konventionell differenzierten Zellen verglichen, führte kurz nach Initiation der adipogenen und osteogenen Transdifferenzierung zum Auffinden zahlreicher, reproduzierbar regulierter Gene. Viele dieser Gene standen mit Metabolismus, Transkription und Signaltransduktion wie dem FGF-, IGF- und Wnt-Signalweg in Zusammenhang, es wurden allerdings nur einige Adipogenese- und keinerlei Osteogenese-assoziierte Markergene innerhalb 24 h nach Initiation der Transdifferenzierung detektiert. Um unter der großen Zahl an regulierten Genen mögliche Schlüsselkontrollfaktoren der Transdifferenzierung zu finden, wurde ein neuartiges, bioinformatisches Punktesystem entwickelt, das Gene entsprechend ihrer potenziellen Relevanz für die Reprogrammierung auflistete. Dabei wurde neben der Reproduzierbarkeit und dem Ausmaß ihrer Regulation auch eine mögliche Reziprozität der Regulation zwischen dem adipogenen und osteogenen Transdifferenzierungsweg berücksichtigt. Es konnte nachgewiesen werden, dass der Fibroblastenwachstumsfaktor 1 (FGF1), der als einer der Hauptkandidaten für die Steuerung der Reprogrammierung eingeordnet worden war, in unserem Zellkultursystem sowohl die adipogene Differenzierung als auch die adipogene Transdifferenzierung hemmt. Die weitere Untersuchung des FGF-Signalwegs und anderer, hoch gelisteter Gene könnte zum besseren Verständnis der altersbezogenen fettigen Degeneration auf molekularer Ebene beitragen und daher Zielmoleküle liefern, die eine therapeutische Beeinflussung der Plastizität zwischen beiden Zelllinien zur Verhinderung der fettigen Degeneration und zur Förderung der Osteogenese erlauben

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49

Schilling, Tatjana [Verfasser]. "Transdifferentiation of human mesenchymal stem cells / vorgelegt von Tatjana Schilling." 2007. http://d-nb.info/985826525/34.

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50

Tsai, Hung-Li, and 蔡弘曆. "Extracellular Matrix-Cytokine Guides Transdifferentiation from Mesoderm Stem Cells to Neuroectoderm Stem Cells." Thesis, 2014. http://ndltd.ncl.edu.tw/handle/s3rpj5.

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博士
臺北醫學大學
醫學科學研究所
102
Mesenchymal stem cells (MSCs) can differentiate into neural stem cells (NSCs) under specific conditions. Wnt was reported to be crucial during embryonic development and adult tissue homeostasis. Wnts regulate neurogenesis, such as neurite and synapse formation, of the neural stem or progenitor cells. In addition to Wnts, tenascin (Tn) family members, TnC and TnR, have been demonstrated to regulate differentiation and migration, as well as neurite outgrowth and survival in numerous types of neurons and progenitor cells. Thus, we hypothesize that Wnt signaling and different forms of Tn regulate the terminal neuronal differentiation in neurotrophin-induced hMSCs. Wnt was also reported, to stimulate hMSC with neurotrophins containing medium (retinoic acid, nerve growth factor, and brain-derived neurotrophic factor; NBR) resulting in the expression of Wnt7a, which enhanced the hMSCs to express neuronal markers in our subsequent analysis, Synapsin-1 (SYN) was induced by Wnt7a and lithium, a glycogen synthase kinase (GSK)-3βinhibitor, in the NBR-induced hMSCs, but was further inhibited by the Wnt inhibitors. Furthermore, hrWnt7a triggered the formation of cholinergic, dopaminergic, GABAergic, and serotonergic neurons. hMSCs were then further cultured in media incorporated with soluble Tn, or on pre-coated Tn. In a qualitative PCR analysis, adding a soluble TnC and TnR mixture to the medium significantly enhanced the expression of neuronal and glial markers, whereas no synaptic markers were expressed. Conversely, in the groups of cells treated with coated TnC, hMSCs showed neurite outgrowth and synaptic marker expression. A combination of TnC and TnR significantly promoted hMSC differentiation in neurons or oligodendrocytes, induced neurite and synapse formation, and inhibited differentiation into astrocytes. In a functional blocking study, integrin α7 and α9β1 blocking antibodies inhibited, respectively, 80% and 20% of the mRNA expression by the hMSCs in the coated Tn mixture. In summary, our results demonstrated novel mechanisms and functions of Wnt7a and Tn for regulating neural differentiation. Data can be broadly employed in the use of MSC to treat neurodegenerative disease.

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Dissertations / Theses on the topic 'Cell transdifferentiation'

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