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1
Robinson, Simon N. "Proliferation regulation of haematopoietic stem cells in normal and leukaemic haematopoiesis." Thesis, University of St Andrews, 1992. http://hdl.handle.net/10023/14965.
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The cellular integrity of the blood is maintained by the cellular output of the haematopoietic stem cell population which produces the specialized precursors and differentiated cells which constitute the blood. The investigation of haematopoietic stem cell behaviour and regulation has been hampered by both the difficulty in their identification and the development of relevant assay systems. The purpose of this investigation was to study the behaviour and regulation of the haematopoietic stem cell population in normal and leukaemic haematopoiesis using an in vitro assay of a primitive haematopoietic precursor. The use of a combination of haematopoietic colony-stimulating factors [interleukin 3 (IL3)/multi-CSF and macrophage colony-stimulating factor (M-CSF/CSF-1)] in semi-solid agar culture of murine haematopoietic tissue, stimulated the proliferation of a haematopoietic colony-forming cell, defined as the "HPP-CFCIL3+CSF-1" population, which was characterized by a high proliferative potential, a multipotency and behavioural and regulatory properties consistent with its being a primitive haematopoietic precursor and possibly a component of the haematopoietic stem cell population. The proportion of the in vitro HPP-CFCIL3+csf-1 population in S-phase in normal murine marrow, was determined to be relatively low at approximately 10%, increasing to approximately 40% in sublethally X-irradiated, regenerating murine marrow and the respective presence of the haematopoietic stem cell proliferation inhibitor and stimulator was demonstrable by the induction of appropriate kinetic changes in the in vitro HPP-CFCIL3+CSF-1 population. In leukaemic haematopoiesis, leukaemic proliferation often occurs at the expense of apparently suppressed normal haematopoiesis. In vitro HPP-CFCIL3+CSF-1 assay of the haematopoietic stem cell proliferation regulators in a number of murine, myeloid leukaemic cell lines, failed to demonstrate either increased levels of the haematopoietic stem cell proliferation inhibitor, or evidence of a direct-acting, leukaemia- associated proliferation inhibitor, however, evidence of a leukaemia- associated impairment of inhibitor and stimulator production was observed and this may be a possible mechanism by which the leukaemic population develops a proliferative advantage over normal haematopoietic tissue. The identification of a possible mechanism of leukaemic progression and suppression of normal haematopoiesis may subsequently allow the development of potentially more effective disease treatment and management regimes. The endogenous haemoregulatory tetrapeptide: Acetyl-N-Ser- Asp-Lys-Pro [AcSDKP, Mr=487 amu] is reported to prevent the G0-G1 transition of haematopoietic stem cells into S-phase. The mechanism of action of AcSDKP and a number of related peptides, was investigated in relation to the stem cell proliferation stimulator and inhibitor. AcSDKP demonstrated no direct haemoregulatory role against the in vitro HPP-CFCIL3+CSF-1 population, which is consistent with reports that AcSDKP is not active against cells already in late G1, or S-phase, rather it appeared to act indirectly by impairing the capacity of the haematopoietic stem cell proliferation stimulator to increase the proportion of the in vitro HPP-CFCIL3+CSF-1 population in S-phase. An apparent impairment of stimulator action may explain the reported AcSDKP-associated 'block' of haematopoietic stem cell recruitment. A putative endogenous AcSDKP precursor and synthetic and degradative enzyme systems have been reported and the possible physiopathological role of AcSDKP in a number of myeloproliferative disorders has been implicated. The potential application of AcSDKP as a 'haemoprotective' agent administered prior to the use of S-phase- specific chemotherapy may be of clinical significance. The in vitro HPP-CFCIL3+CSF-1 assay of a primitive haematopoietic precursor cell population, which may be a component of the haematopoietic stem cell population, should play a significant role in the investigation of haematopoietic stem cell behaviour and regulation in both normal and aberrant haematopoiesis. With the characterization of the mechanism(s) of action of the haematopoietic stem cell proliferation inhibitor and stimulator and the haemoregulatory tetrapeptide AcSDKP, the manipulation of the haematopoietic system to clinical advantage can be envisaged, while the identification of the aberrant regulatory mechanism(s) in haematopoietic dysfunction may allow, the development of more effective disease treatment and management regimes.
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Samitsch, Marina. "Dissecting human haematopoietic progenitors." Thesis, University of Oxford, 2013. http://ora.ox.ac.uk/objects/uuid:58511f8d-cb36-4acf-b706-c465c50f5404.
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Human haematopoiesis resembles a complex hierarchy, however most intermediate stages are only poorly defined. Efforts to characterise human progenitors have been inconsistent and failed to integrate previous knowledge. Furthermore, characterisation of normal progenitors has important implications in acute myeloid leukaemia (AML) biology. We previously established that leukaemic stem cells (LSCs) resemble the immunophenotypic progenitor compartments more closely than the stem cell fraction. Therefore, I set out to characterise human stem and progenitor cells (HSCPs) on phenotypic, molecular and functional level to complete the picture of human haematopoiesis. I purified HSPCs based on their immunophenotype from adult bone marrow (BM) and umbilical cord blood (CB) to investigate steady state and neonatal haematopoiesis. To define differentiation potentials, HSPCs were subjected to functional in vitro assays on bulk and clonal level. Limit dilution assays were used to determine the frequency of cells with multiple differentiation potentials. RNA sequencing revealed underlying lineage priming and specific gene expression signatures. I successfully characterized the incompletely defined Lin-CD34+CD38-CD45RA+ fraction in BM and CB, containing a CD10lo lymphoid-primed multipotent progenitor (LMPP) with T cell, B cell, NK cell, granulocytic and monocytic differentiation potential, and succeeded in placing it in the haematopoietic hierarchy with relation to similar lympho-myeloid progenitors defined by other groups. This research lays the foundation to characterise early human progenitors with a comprehensive toolkit on a phenotypic, molecular and functional level. Findings from this thesis might provide knowledge about potential targets in LSCs.
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Bureau, Emilie Aurelie. "Investigation of the role of haematopoietic cell kinase in normal and leukaemic haematopoiesis." Thesis, University College London (University of London), 2008. http://discovery.ucl.ac.uk/1444125/.
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Acute myeloid leukaemia (AML) is characterised by an accumulation of immature blasts that fail to fully differentiate. Leukaemia is organised as a hierarchy, which is maintained by leukaemic stem cells (LSC). To identify molecular differences between normal haematopoietic stem cells (HSC) and LSC, we performed microarray analysis and found that haematopoietic cell kinase (HCK) is overexpressed in LSC. Thus, by knocking-down HCK in leukaemic cells or overexpressing it in stem cell enriched fractions, we should be able to evaluate its role in leukaemogenesis. Since LSC are difficult to culture in vitro, we started to validate HCK silencing in leukaemic cell lines, Mono-mac-6 (MM6), U937 and Fujioka/P31, which highly express HCK. In all cell lines studied, no more than 50% silencing could be achieved, even when a short-hairpin anti-HCK was cloned into a lentiviral backbone to follow the long-term effect of HCK silencing. Decrease in kinase activity was confirmed using kinase assay and phospho-specific antibody recognising the activated HCK kinase. We show that HCK silencing does not affect the cell cycle, proliferation, differentiation or apoptosis of the cell lines. However, using methylcellulose assay, we observe a significant change in MM6 colony morphology caused by a decrease in their migration properties. Moreover, using phospho-flow cytometry, G-CSF and GM-CSF signal transduction towards STAT5 could be proven to occur via HCK in MM6, but not in Fujioka/P31 or U937 cells. HSC enriched umbilical cord blood cells were also transduced with a lentivirus encoding p59HCK. Overexpression of p59HCK in these cells led to their enhanced erythroid differentiation at the expense of myeloid differentiation underlined by the activation of c-Raf, ERK and STATS. Overall, this study can be used as a preliminary set up for further investigation of the role of HCK in normal human stem cells and in primary AML samples in vivo.
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Ivanovs, Andrejs. "Development of haematopoietic stem cells in the human embryo." Thesis, University of Edinburgh, 2012. http://hdl.handle.net/1842/8839.
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Haematopoietic stem cells (HSCs) emerge during embryogenesis and maintain hematopoiesis in the adult organism. Qualitative and quantitative assessment of HSCs can only be performed functionally using the in vivo long-term repopulation assay. Due to the lack of such data, little is known about the development of HSCs in the human embryo, which is a prerequisite for the development of new therapeutic strategies. Employing the xenotransplantation assay, I have performed here the spatio-temporal mapping of HSC activity within the human embryo and have shown that human HSCs emerge first in the aorta-gonad-mesonephros (AGM) region, specifically in the ventral wall of the dorsal aorta, and only later appear in the yolk sac, liver and placenta. Human AGM region HSCs transplanted into immunodeficient mice provide long-term high-level multilineage haematopoietic repopulation. These cells, although present in the AGM region in low numbers, exhibit a very high self-renewal potential. A single HSC derived from the AGM region generates around 600 daughter HSCs in primary recipient mice, which disseminate throughout the entire recipient bone marrow and are retransplantable. These findings highlight the vast regenerative potential of the earliest human HSCs and set a new standard for in vitro generation of HSCs from pluripotent stem cells for the purpose of regenerative medicine. I have also established a preliminary immunophenotype of the earliest human HSC. These data will be useful for my future studies on the mechanisms underlying the high potency of human embryonic HSCs and on the characterisation of embryonic HSC niche.
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Rossouw, Sophia Catherine. "Apoptosis in Haematopoietic progenitors." Master's thesis, University of Cape Town, 2009. http://hdl.handle.net/11427/3179.
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Introduction: Intracranial pressure (ICP) monitoring is a cornerstone of care for patients with severe traumatic brain injury (TBI). The primary goal of ICP treatment is to preserve brain oxygenation, and since brain oxygenation is usually not measured, the control of ICP is used as a surrogate marker. However studies indicating that cerebral hypoxia/ischemia may occur in the face of adequate ICP and cerebral perfusion pressure (CPP) suggest that the interaction between ICP and brain oxygenation is poorly understood and warrants further investigation. This is of particular importance in the context of children in whom the interpretation of relationships between intracranial factors is even more complex due to changing physiological norms with age. To date little scientific data exists in children and treatment threshold values are often extrapolated from adult guidelines. This study aims to better understand the relationship between ICP and brain oxygenation measured as brain tissue oxygen tension (PbtO2) in a large paediatric cohort suffering from severe TBI. Specifically analysis 1) investigated ICP and PbtO2 profiles over time following TBI, 2) examined the relationship between ICP and PbtO2 from time-linked paired observations, 3) explored various critical thresholds for ICP and PbtO2, and 4) interrogated digital data trends depicting the relationship between ICP and PbtO2. The level of agreement between hourly recorded and high frequency electronic data for ICP and PbtO2 was also evaluated. Method: Paired ICP and PbtO2 data from 75 children with severe TBI were tested with correlation and regression. Additional analyses controlled for mean arterial pressure (MAP), arterial partial pressure of oxygen (PaO2), CPP, arterial partial pressure of carbon dioxide (PaCO2) and haemoglobin (Hb) using multivariate logistic regression analysis and general estimating equations. Various thresholds for ICP were examined; these included age-related thresholds to account for the potential influence of age. Receiver-operating curves (ROCs) were used to graphically demonstrate the relationships between various thresholds of ICP and various definitions of low PbtO2. These were constructed for pooled and individual patient data. Interrogation of electronically recorded data allowed for case illustrations examining the relationship between ICP and PbtO2 at selected time points. Hourly and electronic data were compared using Bland and Altman plots and by contrasting the frequency of ICP and PbtO2 perturbations recorded with each system. 5 Result: Analyses using over 8300 hours of paired observations revealed a weak relationship between ICP and PbtO2, with an initially positive but weak slope (r = 0.05) that trended downwards only at higher values of ICP. Controlling for inter-individual differences, as well as MAP, CPP, PaO2, PaCO2 and Hb did not strengthen this association. This poor relationship was further reflected in the examination of threshold ICP values with ROCs, no singular critical ICP threshold for compromised brain oxygenation was discernible. Using age-based thresholds did not improve this relationship and individual patient ROCs demonstrated inter-individual heterogeneity in the relationship between ICP and PbtO2. However, it was clear that in individual patients ICP did exhibit a strong negative relationship with PbtO2 at particular time points, but various different relationships between the 2 variables were also demonstrated. A high level of agreement was found between hourly and electronic data. Conclusion: These results suggest that the relationship between ICP and PbtO2 is highly complex. Although the relationship in individual children at specific time points may be strong, pooled data for the entire cohort of patients, and even for individual patients, suggest only a weak relationship. This is likely because several other factors affect PbtO2 outside of ICP, and some factors affect both independently of each other. These results suggest that more study should be directed at optimising ICP thresholds for treatment in children. The use of complimentary monitoring modalities may assist in this task. Depending on the adequacy of measures of brain perfusion, metabolism or oxygenation, it is possible that targeting a range of ICP values in individual patients may be appropriate; however this would require detailed investigation.
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Stocking, Carol E. "Autonomous growth of haematopoietic cells." Thesis, Brunel University, 1989. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.290956.
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Butler, Lisa H. "Chromosome translocations in haematopoietic neoplasms." Thesis, University of Oxford, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.360209.
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Trento, Cristina. "Interaction between haematopoietic and mesenchymal stroma." Thesis, Imperial College London, 2013. http://hdl.handle.net/10044/1/37557.
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Parenchyma and stroma represent the functional and structural units in every organ of the body respectively. Stromal cells of mesenchymal origin (MSC) have traditionally been associated with a structural support activity within the tissue, but it is only recently that more complex functions have been unveiled. Subsets of MSC have been shown to play a fundamental role in self-renewal and differentiation of haematopoietic stem cells (HSC). Recent findings show that MSC and bone marrow (BM) macrophages represent fundamental components in the niche, modulating egress and mobilization of HSC during normal or emergency myelopoiesis. Therefore, I have decided to investigate whether and how MSC contribute to the formation and function of myeloid cells. In an in vitro co-culture model I have observed that MSC have the ability to induce the expansion and differentiation of different subsets of mature myeloid cells from haematopoietic BM cells. Based on the differential expression of CD11b and Gr-1, three cell subsets recapitulating myeloid differentiation could be identified. MSC induced differentiation targets common myeloid progenitors (CMP) or granulocyte/macrophage progenitors (GMP) but not the primitive HSC. CD11b+ sorted cells obtained at the end of the co-culture exhibited a functional profile characterised by high levels of both anti- and pro-inflammatory markers, such as nitric oxide synthase 2 (NOS2) and arginase-1 (ARG-1). In order to identify the mechanisms involved in this phenomenon I have chosen to investigate a number of molecules involved in the regulation of haematopoietic differentiation by the microenvironment. I have demonstrated that whilst NOS2 and agrin, an ECM protein, play a key role in the differentiation of CD11b+ Gr-1- F4/80+ cells, complement appears to be primarily involved in the generation of CD11b+ Gr-1int-low F4/80- cells. My studies have shown that MSC differentiating activity is not confined to the BM but can also be detected in MSC from other tissues like skin and kidney. Overall these results suggest a key role for stromal cells as regulators of myeloid differentiation. Further investigation is under way to assess the importance of such a function in vivo.
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Goodman, Reyna Suzanne. "Immunogenetics of haematopoietic stem cell transplantation." Thesis, Anglia Ruskin University, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.478885.
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Wong, Gabriel K. "Haematopoietic clonality in common variable immunodeficiency." Thesis, University of Birmingham, 2016. http://etheses.bham.ac.uk//id/eprint/6935/.
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The aetiology of Common Variable Immunodeficiency (CVID) has fascinated immunologists since Dr. Janeway reported the first case in 1953. While the advances in molecular biology have enlightened us on the aetiology in some patients, the majority is not caused by inherited genetic disorders. A convincing mechanism accounting for the intrinsic variable and partial nature of the condition has yet been proposed. CVID separates itself from other primary antibody deficiencies by the procurement of an abnormal T-cell compartment. Data from this study support that both T-cells and B-cells are subjected to similar deficiency. Investigation of the T-cell receptor repertoire by next-generation sequencing and multi-parametric flow cytometry suggests a severe reduction in naïve T-cell output from the thymus. Similarly, the study of long-lived plasma cell generation and survival highlighted the greatest functional deficits in the naïve B-cell pool, altogether supporting an acquired arrest in lymphogenesis. Using DNA methylation as a surrogate marker for pre-VDJ clonality, this study further shows that some CVID patients exhibited clonal haematopoiesis, adjoining CVID to other clonal haematopoiesis related acquired haematological disorders. Further work is being focused on using high resolution techniques to confirm this association and mechanistically define the development of antibody deficiency in adulthood.
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Menzel, Ursula Rosa. "Gene expression in early haematopoietic development." Thesis, University of Edinburgh, 1998. http://hdl.handle.net/1842/11159.
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Under appropriate culture conditions, ES cells will spontaneously differentiate in vitro into a range of embryonic lineages, including the haematopoietic lineage. All haematopoietic lineages can result from ES cell differentiation including transplantable HSCs. The relative ease of transgenesis in ES cells has been exploited for the analysis of gene function and the identification of novel genes, by the use of gene targeting and gene trapping methodology, respectively. Conventional analysis of mutated ES cells is based on the production of chimeric embryos for in vivo studies. However, in vitro differentiation of mutated ES cells can provide an alternative and complementary approach to in vivo analysis. In particular an in vitro strategy for the screening of ES cells gene trap libraries could restrict the number of gene trap clones to be analyzed subsequently in chimeric embryos in vivo. Based on an established ES cell system for in vitro haematopoiesis, part of the present project has been the assessment of an in vitro prescreening strategy of a gene trap library for the identification of genes that may be involved in early haematopoietic differentiation. This was achieved by monitoring the temporal expression of a β-gal reporter gene in established gene trap lines after induction of haematopoietic differentiation by a morphogenic factor. The gene trap cell lines for use in this study were selected on the basis of their spatial expression patterns in chimeric embryos. The potential application of this strategy on a large scale has been tested by simplification of the culture procedures that support haematopoietic differentiation. The interaction of gene trap constructs into the ES cell genome facilitates the identification of the trapped endogenous gene but also allows the use of in situ hybridization for the analysis of reporter gene expression.
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Doostdar, Lobat. "Haematopoietic differentiation of embryonal stem cells." Thesis, University of Edinburgh, 1997. http://hdl.handle.net/1842/13690.
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Exploiting the ability of ES cells to differentiate toward haematopoietic lineages, the project sought to investigate gene expression with the view of identifying novel genes/proteins involved in the early stages of ES cell commitment to haematopoietic lineages. In the culture system used, it has been shown that under standard differentiation conditions 30-50% of embryoid bodies are able to give rise to haematopoietic colonies. This level of haematopoietic commitment was modulated with the addition of chemical inducers of the differentiation environment of embryoid bodies. Using the CFU-A assay to assess haematopoietic commitment and differentiation, we have found that the number of CFU-A colonies can be increased 2-fold after treatment of embryoid bodies with 1% DMSO during the first 48 hours of differentiation, and conversely reduced to below 5% after treatment with 10-8 all trans-retinoic acid (atRA) for the same period. Lineage commitment in untreated, DMSO and atRA embryoid bodies were further investigated using reverse transcription-polymerase chain reaction techniques. The mesoderm marker brachyury was found to be expressed in both DMSO and atRA treated samples to similar extents, suggesting neither compound to be exclusively exerting its effect by enhancing or inhibiting mesoderm commitment. However, the level of transcripts of various haematopoietic markers such as β-globin, PU-1 and CD45 were found to be raised in DMSO treated embryoid bodies compared to either untreated or atRA treated samples. Studies investigating the ability of cells from treated and untreated embryoid bodies, to reconstitute the haematopoietic system of lethally irradiated mice were also carried out. Overall these showed that mice injected with cells from DMSO treated embryoid bodies survived for longer periods of time than those injected with cells from either untreated or atRA treated embryoid bodies. It has been clearly demonstrated that the haematopoietic commitment of ES cells and their differentiated aggregates can be successfully modulated by the addition of chemical inducers. Consequently these effects will prove a useful means of screening for genes and proteins involved in haematopoietic commitment and differentiation.
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Patel, Sanjay. "Haematopoietic progenitor cells in carotid disease." Thesis, King's College London (University of London), 2012. https://kclpure.kcl.ac.uk/portal/en/theses/haematopoietic-progenitor-cells-in-carotid-disease(27973516-c623-44a0-86be-75dc20b4e6fe).html.
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Introduction Haematopoietic progenitor cells (HPCs) may attenuate the response to acute vascular injury by maintaining endothelial integrity and function. The aim of this study was to determine whether circulating HPC number and function, and mobilising cytokines, reflect significant carotid disease or correlate with restenosis following carotid endarterectomy (CEA). Methods Initially the assay conditions to measure HPC number and function were optimized. HPC numbers were measured by flow cytometry (CD133+ve/CD34+ve) and early colony forming unit assay (eCFU). HPC function was measured by migration assay and by staining for senescence associated B-galactosidase (SA-Bgal). HPC number and function was then measured in 62 patients undergoing CEA pre-operatively, 1 day post operatively and 6 weeks post-operatively. Restenosis was assessed by duplex scanning at 3, 6 and 12 months. The circulating profile of GM-CSF, PIGF, SDF1 and VEGF was measured by multiplex ELISA. Results HPC numbers (P<0.001) and eCFU counts (P=0.001) fell rapidly 24hrs after CEA. The percentage post-operative fall in CD133+ve/CD34+ve cell numbers negatively correlated with degree of restenosis at the 6 month scan (r= -0.38, p=0.013). The percentage fall in eCFU number negatively correlated with degree of restenosis at the 6 (R=-0.42, P=0.008) and 12 month scans(R=-0.49, P=0.026). The migration rate of HPCs isolated from pre-operative blood also negatively correlated with restenosis at the 6 (R=-0.5, P=0.009) and 12 month scans(R=-0.53, P=0.05). Pre-operative SDF1 levels correlated with falls in CD133+ve/CD34+ve number (R=0.42, P=0.044) and eCFU counts (R=0.56, P=0.004), though not with restenosis. Conclusion HPC function appears to be linked to the development of carotid artery restenosis following endarterectomy. These data support the concept that HPCs have a role in regulating remodelling of the unstable and injured arterial wall.
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Fanning, Niamh Catherine. "Novel ES cell differentiation system enables the generation of low-level repopulating haematopoietic stem cells with lymphoid and myeloid potential." Thesis, University of Edinburgh, 2014. http://hdl.handle.net/1842/17940.
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The potential of embryonic stem (ES) cells to generate any developmental or adult cell type holds much promise for regenerative medicine and in vitro modelling of development and disease. Haematopoietic stem cells (HSCs) regenerate all lineages of the blood throughout adult life and are essential for the treatment of a vast number of haematalogic disorders. Current sources of HSCs for clinical use and research, including adult bone marrow, peripheral blood stem cells and umbilical cord blood, are limited by the number of HSCs they contain and by the availability of a suitable donor. A system that generates a reliable source of HSCs from ES cells would therefore be an ideal alternative. While much progress has been made in the generation of downstream lineages of the haematopoietic system, progress in the derivation of HSCs capable of long-term self-renewal and multilineage reconstitution from ES cells has been limited. Understanding of the developmental steps leading to HSC emergence in the embryo has been advancing in recent years. In particular, precursors of HSCs (preHSCs) have been isolated from the mouse embryo, characterised and matured into HSCs ex vivo using the specialised conditions of aggregate culture systems (Taoudi et al 2008, Rybtsov et al 2011). We hypothesised that application of the aggregate culture system in the differentiation of ES cells could provide a missing link in the in vitro generation of HSCs. Here I have developed a novel ES cell differentiation system that employs the specialised conditions of the aggregate culture system, after an initial stage of mesoderm differentiation. I show that this system creates an environment for efficient haematopoietic and endothelial progenitor formation and generates cells of a preHSC type I (VE-Cadherin+CD45-CD41lo) and preHSC type II (VE-Cadhein+CD45+) surface phenotype. Notably, the system gives rise to cells that achieve low-levels of haematopoietic repopulation in sublethally irradiated NSG mice. The low-level repopulating cells persist for over 4 months in animals and show both myeloid and lymphoid potential. I identify genes that are expressed in cells of a preHSC II surface marker-phenotype from the E11.5 dorsal aorta, but not in cells of this phenotype from the E11.5 Yolk sac or differentiated ES cells. I also show that enforced expression of Notch downstream target Hes1 in Flk1+ mesoderm during ES cell differentiation does not improve levels of ES-derived repopulation.
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Gordon-Keylock, Sabrina Anne Megan. "Haematopoietic differentiation of murine embryonic stem cells." Thesis, University of Edinburgh, 2009. http://hdl.handle.net/1842/29123.
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This thesis aimed to determine which subregions of the E10.5 aorta-gonad-mesonephros (AGM) were responsible for the haematopoietic enhancing effects that primary AGM regions had on differentiating ES cells. To this end, a novel co-culture system has been established to test the enhancing effects of a panel of clonal stromal cell lines derived from different subregions of the midgestational AGM. Three stromal cell lines derived from the dorsal aorta and surrounding mesenchyme (AM) subregion of the AGM were able to significantly enhance the frequency of ES cell derived multipotent haematopoietic progenitors. Two stromal cell lines derived from the urogenital ridges (UG) of the AGM did not enhance haematopoietic differentiation of ES cells. The haematopoietic enhancing effects were not retained by extracellular matrices isolated from the AM stromal cell layers and the effects were dependent on direct ES cell-stromal cell contact. Co-culture of an ES cell line carrying a Brachyury-eGFP reporter gene demonstrated that the stromal lines mediated their effects post-Brachyury (mesoderm) induction in the ES cells. In addition, co-culture of sorted ES cell populations confirmed that Brachyury+, but not Brachyury-, cells gave rise to haematopoietic progenitors in AM co-culture, supporting the notion that ES cell differentiation recapitulated the in vivo pattern of lineage specification. Transplantation of co-cultured ES cells into irradiated adult NOD/SCID mouse recipients led to low levels of engraftment in the spleen and bone marrow. Adult bone marrow cells achieved repopulation more readily in the NOD/SCID animal model when transplanted intra-splenically, compared to intravenous injection. This suggests that transplantation of ES-derived haematopoietic cells directly into the haematopoietic niche, by intra-splenic or intra-femoral injection, could facilitate repopulation.
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Thorpe, Michael. "Haematopoietic Serine Proteases : A Cleavage Specificity Analysis." Doctoral thesis, Uppsala universitet, Institutionen för cell- och molekylärbiologi, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-221891.
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Mast cells are innate immune cells, historically involved in allergy responses involving IgE. Through this, they have earned a reputation as a fairly detrimental cell type. Their beneficial roles remain somewhat enigmatic although they clearly have the ability to modulate the immune system. This is due to their ability to synthesise many cytokines and chemokines as well as immediately release potent granule-stored mediators. One such mediator is a serine protease, chymase, which has been targeted by pharmaceutical companies developing inhibitors for use in inflammatory conditions. In order to address roles of the proteases, information regarding their cleavage specificity using substrate phage display can help find potential in vivo substrates. The human chymase cleaves substrates with aromatic amino acids in the P1 position and has a preference for negatively charged amino acids in the P2’ position. The molecular interactions mediating this P2’ preference was investigated by site-directed mutagenesis, where Arg143 and Lys192 had a clear effect in this selectivity. As humans express one chymase and rodents express multiple chymases, extrapolating data between species is difficult. Here, the crab-eating macaque was characterised, which showed many similarities to the human chymase including a near identical extended cleavage specificity and effects of human chymase inhibitors. Appropriate models are needed when developing human inhibitors for therapeutic use in inflammatory conditions. The effects of five specific chymase inhibitors in development were also tested. The selectivity of inhibitors was dependent on both Arg143 and Lys192, with a greater effect of Lys192. Identification of residues involved in specific inhibitor interactions is important for selective inhibitor development. Another innate cell type, the NK cell, is important in virus and tumour defence. In the channel catfish, a serine protease from an NK-like cell, granzyme-like I, was characterised. A strict preference for Met in the P1 position was seen, and caspase 6 was identified as a potential in vivo target. This may highlight a novel apoptosis-inducing mechanism from a similar cell type has been conserved for approximately 400 myr. Here, important residues mediating chymases’ specificity and interactions with inhibitors has been addressed, as well as finding a new animal model for providing ways to combat their roles in pathological settings.
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Kostadima, Myrto Areti. "Analysis of the haematopoietic transcriptome in development." Thesis, University of Cambridge, 2015. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.708506.
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MacLean, Adam L. "Modelling haematopoietic stem cells in their niche." Thesis, Imperial College London, 2014. http://hdl.handle.net/10044/1/24927.
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Modelling haematopoietic stem cells (HSCs) mathematically allows us to probe their behaviour, test hypotheses and make predictions. HSCs are essential and elusive; despite many recent advances much remains unknown, including how the microenvironment (niche) influences HSC behaviour, and the nature of complex interactions between HSCs, the niche and disease. This thesis comprises three sections. In the first we present new methods for the analysis of ODE systems that allow us to characterise steady state properties such as the probability of a state being stable (over some parameter range). We apply these methods to models of stem cell dynamics that differ in the form of regulation (feedback) imposed on the system and study the steady states of these systems. Competition within the niche between HSCs and invading cancer cells disrupts the system. In the second section we model this using ODEs that describe the population dynamics of cellular species from an ecological perspective. From the analysis of two models differing in their treatment of the niche, using Bayesian inference, we find that maintaining a viable HSC population is necessary and almost sufficient in order to outcompete leukaemia and restore healthy haematopoiesis. In the third section we extend the analysis of interactions between HSCs, leukaemia and the niche: performing a comparison of three models that describe the dynamics of chronic myeloid leukaemia. We study heterogeneous data from a clinical trial of the disease. All models fit these data, but do so in different ways. One model is discarded due to its unrealistic predictions, suggesting that direct competition between species is important. Each of the remaining two models makes testable predictions, but validating these is at current experimental limits. In the future, the results presented will aid experimental design and provide a framework from which to identify new therapeutic targets for haematopoietic diseases.
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Maleszewska, Marta. "Epigenetic regulation of haematopoietic stem cell differentiation." Paris 7, 2009. http://www.theses.fr/2009PA077097.
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Le rôle des événements épigénétiques dans le maintien de la multipotentialité et la détermination cellulaire des cellules souches hématopoïétiques (CSH) reste mal connu. L'objet de ma thèse a été de définir les éléments épigénétiques qui sous-tendent la multipotentialité des CSH. Les modifications des histones, la méthylation de l'ADN et la localisation sub-nucléaire de loci spécifiques des lignées hématopoïétiques ont été étudiées dans les CSH CD34+CD381o et dans des précurseurs hématopoïétiques. Nous montrons que dans les CSH les gènes hématopoïétiques ont une structure chromatinienne particulière, avec de hauts niveaux d'acétylation de l'histone H4 et de H3K4me2, en l'absence des marqueurs actif H3K4me3 et répressifs H3K9me3 et H3K27me3. La différenciation des cellules souches est associée à une inhibition épigénétique des loci non spécifiques de la lignée (diminution des marqueurs d'activation et augmentation des marqueurs d'inhibition) et à un enrichissement des gènes spécifiques de la lignée en marqueurs d'activation, notamment acétylation de l'histone H3 et H3K4me3. Les modifications des histones aux loci D -globine et Ig précèdent leur changement de localisation sub-nucléaire aux cours de l'hématopoïèse. Nous montrons que la région JH du locus IgH est méthylée de façon hétérogène en tout-ou-rien dans les cellules CD34+CD381o, du fait soit de différences spécifiques d'allèle dans la méthylation de l'ADN soit d'une hétérogénéité au sein des cellules CD34+CD381o. Au total, les CSH présentent des caractéristiques épigénétiques qui pourraient contribuer à établir la multipotentialité de ces cellules souchesThe role of chromatin in haematopoietic stem cell (HSC) cell fate decisions is poorly understood. My PhD studies were aimed at defining epigenetic correlates that underlie the multilineage potential of HSC. To this end, the level of histone modifications, DNA methylation and subnuclear localisation of lineage-specific haematopoietic loci were analysed in CD34+CD381o HSC and lineage committed precursors. This study shows that HSC maintain haematopoietic genes in a distinct chromatin conformation, characterised by the presence of histone H4 acetylation and H3K4me2 in the absence of active H3K4me3 or repressive H3K27me3 or H3K9me3 histone marks. This chromatin structure appears to describe a transcriptionally competent "ground state" for these genes keeping them silent but poised for expression at later stages of HSC differentiation. Progressive lineage restriction and differentiation of HSC was accompanied by epigenetic silencing of lineage-inappropriate genes associated with loss of active and addition of repressive histone marks, while lineage-specific genes were further enriched for active histone H3 acetylation and H3K4me3 marks. Furthermore, we found that changes in histone modifications at the β-globin and Ig loci precede changes in subnuclear localisation during HSC differentiation. DNA methylation analysis indicted that the IgH JH region is heterogeneously methylated in CD34+CD381o progenitors, perhaps reflecting allele-specific differences in HSC or heterogeneity within the CD34+CD381o population. All together, the results of this study have identified epigenetic features that may serve to establish and maintain the multilineage potential of HSC
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Atarod, Sadaf. "MicroRNAs in haematopoietic stem cell transplantation outcome." Thesis, University of Newcastle upon Tyne, 2015. http://hdl.handle.net/10443/2936.
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Allogeneic haematopoietic stem cell transplantation (allo-HSCT) is a curative treatment for numerous haematological malignancies. Graft-versus-host disease (GVHD) is the major complication causing mortality and is classified into acute (aGVHD) and chronic. MicroRNAs play a significant role in inflammation and have reported potential as biomarkers of different diseases. This study has investigated the role of microRNAs in allo-HSCT outcomes and had two main aims; (1) identification of microRNAs specific to the aGVHD target organ, skin and (2) an investigation into the role of immune specific miRNAs (miR-146a and miR-155) in peripheral blood. Initially, pathway mining was performed on a list of 18 genes that were shown previously, to have deregulated expression levels with regards to GVHD. The pathway mining identified specific immunological pathways in relation to the genes and potential microRNA targets. Global microRNA profiling was performed on a discovery cohort that identified a signature microRNA list in skin biopsies obtained from patients at the time of cutaneous histopathological aGVHD onset (grades I-III) and healthy volunteers. Twelve microRNAs were selected for further validation and it was shown that miR-34a-5p, miR-34a-3p, miR-503-5p and let-7c-5p were elevated and significantly involved in allo-HSCT outcomes. There was an interaction between miR-34a-3p and miR-503-5p which was significantly diagnostic of aGVHD and let-7c-5p was significantly predictive of disease relapse. MiR-34a-5p protein targets; p53 and c-Myc were then evaluated in the same cohort. MiR-34a-5p expression levels and cells stained positively for p53 were significantly correlated in the epidermis. Preliminary optimization of miR-34a-5p knockdown study was successfully conducted which showed promising results in the reduction of T cell proliferation. The whole blood study showed that miR-146a-5p and its interaction with miR-155-5p was predictive of aGVHD incidence in pre-disease onset (Day+28) samples. Interestingly, the expression levels of miR-146a-5p and miR-155-5p negatively correlated with SPI1 (PU.1). In conclusion, these investigations showed that (1) the microRNAs studied in this investigation may regulate the expression levels of the selected 18 genes, (2) microRNA expression levels in clinical skin biopsies obtained at the time of aGVHD onset could potentially be used as diagnostic biomarkers for aGVHD and as predictive biomarkers for overall survival as well as relapse and (3) miR-146a-5p and miR-155-5p expression levels in whole blood could be used as predictive biomarkers for aGVHD incidence.
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Tura-Ceide, Olga. "Recognition and manipulation of adult stem cells through haematopoietic and non-haematopoietic differentiation pathways for intended therapeutic clinical applications." Thesis, University of Edinburgh, 2007. http://hdl.handle.net/1842/25261.
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Studies are presented which focus on autologous stem cells for autograft cellular therapies in (1) improvement of early haematopoietic reconstitution by ex vivo manipulation of graft cells, and (2) assessment of the angiogenic potential of available clinical sources for development of therapies for vascular regeneration in ischaemic tissue. There remains a period following HSC autograft when patients are neutropenic and thrombocytopenic. A number of studies have attempted simultaneous ex vivo expansion of progenitors of neutrophil and megakaryocyte lineages to reduce post transplant neutropenia and thrombocytopenia. Our results show that CD34+ HSCs could expand into mature functional neutrophils under the influence of SCF + Flt3-L + G-CSF, but that the addition of other cytokines did not improve CD34+ expansion, and the megakaryocytic growth factor TPO reduced neutrophil maturation. Endothelial progenitor cells (EPCs) are stem cells with the potential to proliferate and differentiate into mature endothelial cells and to form blood vessels in a process resembling embryonic vasculogenesis. Autologous EPC transplantation may be used to promote endothelial reconstitution in patients with ischaemic or infracted tissue. EPC appear to share many properties with HSC, but while haematopoietic potential is now assessed by numbers of HSC expressing CD34, EPC determination remains ambiguous. Bone marrow and mobilised peripheral blood are HSC-rich sources which have been used for vasculogenic therapy. The studies presented here show that there is no correlation between the diverse EPC phenotype definitions proposed in the literature. There is no obvious relationship between the numbers of haematopoiesis related CD34+ or CD133+ cells and outcome of the EPC-CFU assay. The cells responsible for these early outgrown EPC-CFU colonies were CD14+ colonies were CD14+ plastic-adherent monocyte-like cells. Recent reports indicate that a peripheral blood myelocyte monocytes population may also be considered as an EPC source with potential angiogenic clinical capability. Further work is required to better define the relative endothelial potential of these sources for development of clinical autograft vascular regenerative therapies.
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Jing, Duohui. "Mobilisation, Isolation and Coculture of Haematopoietic Stem Cells." Doctoral thesis, Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2011. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-39915.
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Since decades, hematopoietic stem cell transplantation (HSCT) has become a well established treatment modality for hematological malignancies and non-malignant disorders. Autologous and allogeneic hematopoietic stem cells (HSCs) mobilized into the peripheral blood (PB) have been used as a preferred source of transplantable stem cells1-3. And umbilical cord blood (UCB) has been introduced as a more attractive HSC source for HSCT, because fetal stem cells in UCB are speculated to be more primitive in comparison to adult stem cells. However the limited amount of HSCs is limiting their application for stem cell therapy in clinic. Therefore, people started to utilize extra-embryonic tissue to harvest more fetal stem cells, while people also tried to optimize the clinical protocol to mobilize more adult stem cells out of adult bone marrow. The innovative strategies and feasible procedures were discussed in this thesis.
The axis of the chemokine receptor CXCR4 and its ligand SDF-1 is important for trafficking and homing of HSCs. It has already been demonstrated that the bicyclam AMD3100, a CXCR4 antagonist, in combination with G-CSF is able to induce a significant mobilization of CD34+ cells4. And human placenta is a potent hematopoietic niche containing hematopoietic stem and progenitor cells throughout development5. The homing of HSCs to the placenta is probably also mediated by the expression of SDF-1 as demonstrated for the bone marrow niche. In this study (part 1 of the chapter “Results and discussions”), we utilized AMD3100 to mobilize HSCs from placenta. And we can demonstrate that the CXCR4 antagonist AMD3100 mobilise placenta derived CD34+ cells ex utero already after 30 min of incubation and may further enhance the efficacy of harvesting placenta-derived HSC.
The alpha4 integrin CD49d is involved in migration and homing of hematopoietic stem cells (HSC). Therapeutic application of natalizumab, an anti-CD49d antibody, in patients with multiple sclerosis (MS) has been associated with increased levels of circulating CD34+ progenitors. In our study (part 2 of the chapter “Results and discussions”), we compared circulating HSCs from MS patients after natalizumab treatment and HSCs mobilized by G-CSF in healthy volunteers, with regard to their migratory potential, clonogenicity and gene expression. CD34+ cells in the blood and marrow of natalizumab-treated patients expressed less of the stem cell marker CD133, were enriched for erythroid progenitors (CFU-E) and expressed lower levels of adhesion molecules. The level of surface CXCR-4 expression on CD34+ cells from patients treated with natalizumab was higher compared to that of CD34+ cells mobilized by granulocyte-colony stimulating factor (G-CSF) (median 43.9% vs. 15.1%). This was associated with a more than doubled migration capacity towards a chemokine stimulus. Furthermore, CD34+ cells mobilized by natalizumab contained more m-RNA for p21 and less MMP9 compared to G-CSF mobilised HSC. Our data indicate that G-CSF and CD49d blockade mobilize different HSC subsets and suggest that both strategies may be differentially applied in specific cell therapy approaches.
In order to further improve the clinical outcome of HSC transplantation, many groups are focusing on ex vivo maintain or expand HSC. Unfortunately, the maintenance of HSC in vitro is difficult to achieve because of their differentiation. This is presumably caused by a lack of appropriate cues that are provided in vivo by the microenvironment. Indeed, HSCs located in the bone marrow are interacting with a specific microenvironment referred to as the stem cell niche, which regulates their fate in terms of quiescence, self-renewal and differentiation. An orchestra of signals mediated by soluble factors and/or cell-to-cell contact keeps the balance and homeostasis of self-renewal, proliferation and differentiation in vivo. To investigate the communication between HSCs and the niche, coculture assays with mesenchymal stromal cells (MSCs) were performed in vitro. Here, we can demonstrate that cell-to-cell contact has a significant impact on hematopoietic stem cells expansion, migratory potential and stemness. In this study (part 3 of the chapter “Results and discussions”), we investigated in more detail the spatial relationship between hematopoietic stem cells and mesenchymal stromal cells during ex-vivo expansion. And we defined three distinct localizations of HSCs relative to MSC layer: (i) those in supernatant (non-adherent cells); (ii) cells adhering on the surface of mesenchymal stromal cells (phase-bright cells) and (iii) cells beneath the mesenchymal stromal cells (phase-dim cells). Our data suggest that the mesenchymal stromal cell surface is the dominant location where hematopoietic stem cells proliferate, whereas the compartment beneath the mesenchymal stromal cell layer seems to be mimicking the stem cell niche for more immature cells. Our data provide novel insight into the construction and function of three-dimensional HSC–MSC microenvironments.
In summary, we provided a new method to isolate fetal stem cells from extra-embryonic tissue (i.e. placenta) in the first part, then we discussed an innovative strategy with CD49d blockade to improve clinical modality for adult stem cell mobilization in the second part, and finally we investigated HSC maintenance and expansion in vitro and provided feasible way to mimic HSC niche in vitro in the last part.
This thesis contributes to HSC-based stem cell therapy in two aspects, i.e. 1) fetal and adult stem cell isolation holding great therapeutic potential for blood diseases; 2) ex vivo stem cell manipulation providing a valuable platform to model HSC niche regulation.
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Magill, E. L. "The role of the proteasome in haematopoietic malignancy." Thesis, Queen's University Belfast, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.273171.
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Escobedo, Cousin M. H. "Ancillary cell help in haematopoietic cord blood engraftment." Thesis, University College London (University of London), 2014. http://discovery.ucl.ac.uk/1424406/.
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Cord blood (CB) has served as a source of haematopoietic stem cells (HSC) to treat haematological diseases for the last two decades. Low incidence and severity of graft-versus-host disease, and a robust graft-versus-leukaemia effect are some of CB advantages as a source for HSC. However, its main disadvantages are a limited number of HSC per unit and delayed immune reconstitution and infections after CB transplantation (CBT). In order to improve engraftment and immune reconstitution after CBT different approaches have been explored like HSC expansion, double CB transplantation, CBT plus third party donor HSC or intra-bone infusion. It seems as if the interaction of cord blood stem cells (CB SC) with other cells is required for CB SC to engraft. On this basis, the effect of accessory cells, particularly natural killer (NK) cells, on CB SC engraftment was analysed. It was observed that NK cell co-infusion with CB SC led to higher levels of engraftment in NSG mice. This in parallel with a higher level of expression of CXCR4 receptor, a higher migration index, a higher number of colony forming units (CFU) after long-term culture initiating cells assay (LTC-IC) and a trend to a higher number of CFU when CB SC were co-cultured with activated NK cells (aNK). The effect observed on CBSC clonogenic capacity was maintained even after treatment with pertussis toxin (PTX), which blocks G-protein coupled receptors signalling, such as CXCR4. The results presented in this work offer the basis for an alternative approach that could help to improve CBT.
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Paruzina, Daria. "Factors affecting optimal culture of haematopoietic stem cells." Thesis, University of Edinburgh, 2016. http://hdl.handle.net/1842/15917.
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Haematopoietic stem cells (HSC) are invaluable, due to their potential to treat malignant and non-malignant diseases. Modern medicine requires a reliable source of human HSCs (hHSCs) for efficient transplantations, which in many cases cannot be obtained from a single donor. Therefore, the ability to amplify donor hHSCs ex vivo would be an ideal alternative. Past attempts to expand hHSCs in vitro, demonstrated that the protocols developed so far have limited success. My research studied the factors which can affect the optimal culture of transplantable HSCs using a 3D culture system that had previously been used to culture HSCs derived from the aorta-gonad-mesonephros (AGM) region of the mouse embryo. This system involved cell culturing at the gas-liquid interface which is particularly sensitive to mechanical disturbances. To overcome this problem, floating Polypropylene support (rings) were designed and tested and I demonstrated that this was able to prolong aggregate culturing for up to 21 days. Further optimisation tests included altering factors such as oxygen levels, and the presence of antioxidants and apoptosis inhibitors in mouse HSCs culture. I have shown that moderate hypoxia (6% O2) did not affect HSCs in culture, while 2% of O2 led to a significant decrease of HSCs activity. Normoxia resulted in higher reactive oxygen species generation, which would likely be detrimental to cells. However, unexpectedly no improvement in repopulation efficiency of cultured HSCs was achieved by the addition of antioxidant. I also found that when the AGM region was dissociated and co-aggregated in the presence of Rho kinase inhibitor a higher level of repopulation was achieved. In addition, troloxpifitrin-a and p38 inhibitor blocked HSC development without affecting progenitor frequency or the total number of live cells. Subclones of mouse stromal cell line (OP9) were used to create a defined haematopoietic niche for hHSC. Functional screening of these lines in co-aggregate culture re- vealed that 3 of the 34 subclones tested were able to maintain hHSC in culture and repopulate immunodeficient mice at a comparable level to uncultured CD34+ cells. The repopulation in engrafted recipients persisted for over 6 months and showed both myeloid and lymphoid potential. These 3 subclones therefore appeared to create a functional niche for hHSCs and were subsequently used to study the impact of a number of factors including SCF, rock inhibitor, TGFb inhibitor, StemRegenin1 (SR), and prolonged culture technique on hHSC expansion. A significant level of fluctuation between experiments was observed and no definitive conclusions could be drawn. I also attempted to establish stromal cell lines from the human AGM region, more specifically from the ventral (AoV) and dorsal (AoD) regions of the dorsal aorta. Despite attempts to immortalise primary stromal cells, all lines went through a growth crisis. Nevertheless, 30 lines were screened for their ability to support haematopoietic cells in co-aggregate culture with results suggesting that lines derived from AoV expanded haematopoietic precursors more efficiently than AoD lines and OP9 control. Many of the tested lines were able to maintain long-term repopulating human HSCs but the level of repopulation was not as high as that achieved from uncultured CD34+ cells. Unfortunately, these human stromal cell lines have an unstable karyotype which may have an impact on their functional characteristics and they may not represent the nature of the primary cells.
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Mavin, Emily. "Regulatory T cells in haematopoietic stem cell transplantation." Thesis, University of Newcastle upon Tyne, 2014. http://hdl.handle.net/10443/2731.
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Graft-versus-host disease (GvHD) remains the main complication associated with haematopoietic stem cell transplantation (HSCT). GvHD is caused by allo-reactive donor T cells mounting an attack against specific target tissues. CD4+CD25HiFoxp3+ regulatory T cells have been shown to modulate GvHD in vitro and also in vivo animal models. More recently early stage clinical trials have described the successful use of Treg to reduce the incidence of GvHD following HSCT. The aim of this study was to investigate further the suppressive mechanisms by which Treg are able to modulate GvHD and assess the influence of Treg on the beneficial graft-versus-leukaemia (GvL) effect therefore providing further insight into the use of Treg in the therapeutic management of GVHD. Data presented in this thesis demonstrates the successful isolation and expansion of a highly pure Treg population which maintained suppressive capacity throughout culture. We also confirmed that Treg retain suppressive capacity following cryopreservation resulting in reduced workload and increased consistency when used for in vitro functional studies. We also provide the first human in vitro evidence that Treg are able to prevent cutaneous GvH reaction by blocking the migration of effector T cells into the target tissues. The presence of Treg during allo-stimulation caused reduced effector cell activation, proliferation, IFNγ secretion and decreased skin homing receptor expression. Further investigation into the Treg modulation of dendritic cells demonstrated, for the first time in experimental in vitro human GvHD, that this was due to ineffective effector T cell priming in the presence of Treg caused by impairment of dendritic cell functions. Comprehensive phenotypic and functional analysis of Treg treated moDC showed their decreased antigen processing ability and allostimulatory capacity, resulting in a less severe GvH reaction in the skin explant model. Furthermore, this work has revealed that despite Treg impairing in vitro GvL mechanisms at a cellular level there was no association observed between increased Treg levels and the incidence of relapse in a small clinical cohort of HSCT patients. In conclusion this study has provided further insight into the mechanisms by which Treg are able to modulate GvHD. This would inform future clinical trials using Treg as a therapeutic alternative to current GvHD treatment and prophylaxis.
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Pina, Cristina Maria Correia Antunes. "Transcriptional programming in cord blood-derived haematopoietic stem cells." Thesis, University of Oxford, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.492057.
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Molecular regulators of HSC fate decisions have been identified through involvement in leukaemia or by post-genomic gene discovery approaches. Most candidate regulators have been studied in mouse, but the clinical potential of HSC fate modulation emphasises the need to study regulatory mechanisms in the human context. In order to identify regulators of human haematopoiesis, the global expression profile of cord blood (CB) CD133+Go cells was characterised. Less immature CD133+G1 cells served as comparison. CD133+Go cells had been shown to be highly enriched for LTC-IC, a surrogate in vitro assay of HSC function. Further functional characterisation of CD133+Go cells revealed enhanced mixed-lineage and erythroid progenitor activity. In line with the functional data, CD133+Go-enriched transcripts contained an erythroid expression signature. Moreover, global expression of myelo-Iymphoidaffiliated genes was reduced in CD133+Go cells, suggesting that myelo-Iymphoid programmes may be activated downstream of the HSC compartment. A significant proportion of transcripts enriched in CD133+Go cells were associated with transcription regulation. These included MLLT3, a common fusion partner of MLL in acute myeloid leukaemia. Analysis of MLLT3 expression in prospectively isolated progenitors from human CB revealed higher expression in HSC and megakaryocytic (Meg)/E progenitors (MEP). This suggested a role for MLLT3 in E/Meg fate decisions; it may also underline an association between E/Meg and HSC compartments. Lentiviral vectors were used to modulate MLLT3 expression in primitive human CB cells. Forced MLLT3 expression promoted E/Meg progenitor output and analysis of MLLT3 mutants suggested that MLLT3 functional effects depend on its transcription regulatory activity. Gene expression and cis-regulatory element analyses revealed cross-regulatory interactions between MLLT3 and E/Megaffiliated transcription factor GATA1. MLLT3 expression knockdown indicated a requirement for MLLT3 in the elaboration of E/Meg lineages. Gene expression knockdown also impaired multilineage engraftment of human HSC in immunodeficient NOD/SCID mice, with more significant effects on B-cell than on myelo-monocytic output. The latter observation may have implications for the lineage affiliation of MLUMLLT3 leukaemia. In summary, MLLT3 was identified as a novel regulator of human E/Meg lineage decisions and positioned in a regulatory circuit with GATA1. The data supports the notion that E/Meg commitment occurs hierarchically upstream of myelolymphoid lineage decisions and suggests that recently revised hierarchical models of murine haematopoiesis may also apply to the human system.
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Remberger, Mats. "Cytokine production in allogeneic haematopoietic stem-cell transplantation patients /." Stockholm, 1999. http://diss.kib.ki.se/1999/91-628-3749-4/.
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Petropoulos, Antonios. "Assessment of pharmacogenetic polymorphisms in haematopoietic stem cell transplantation." Thesis, University of Newcastle Upon Tyne, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.515029.
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Bowles, K. M. "Generation of haematopoietic cells from human embryonic stem cells." Thesis, University of Cambridge, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.596829.
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Culture of hESCs on murine stromal layers or in stromal free conditions as embryoid bodies results in low levels of haematopoietic cells. Here it is demonstrated that overexpression of the transcription factor HOXB4 considerably augments haematopoetic development of hESCs. Stable HOXB4 expressing hESC clones were generated by lipofection and could be maintained in the undifferentiated state for prolonged passages. Moreover, differentiation of hESCs as embryoid bodies in serum containing medium without the use of additional of cytokines led to sequential expansion of first erythroid then myeloid and monocytic progenitors. These cells retained the capacity to develop into formed blood elements during in vitro culture. Consistent with the development of committed haematopoietic cells the expression of transcription factors known to be critical for haematopoietic development was observed. The successful use of enforced gene expression to promote the differentiation of hESCs into a terminally differentiated tissue is thus demonstrated, thereby revealing an important role for HOXB4 in supporting their in vitro development along the haematopoietic pathway. Once a method for producing significant numbers of haematopoietic cells from hESCs in vitro was established, hESCs were used to study the role of stem cell leukaemia gene (SCL) in human blood and endothelial development. hESC lines were generated in which the expression of SCL, under control of an enhancer previously shown in mice to target expression to blood and endothelial progenitors, was evident through green fluorescent protein (GFP) expression. The enhancer directed GFP expression inhuman K562 cells and some differentiated progeny of HOXB4 transfected hESCs. However a more thorough assessment of GFP positive cells was hindered by problems with transgene silencing.
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Anderson, Claire L. "Molecular control of apoptosis in cytokine-dependent haematopoietic cells." Thesis, Keele University, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.392178.
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Ramachandra, Durrgah Latchumi. "The haematopoietic potential of human amniotic fluid stem cells." Thesis, University College London (University of London), 2017. http://discovery.ucl.ac.uk/10040028/.
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There is a constant demand for haematopoietic stem cells (HSC) for clinical applications. Amniotic fluid stem (AFS) cells serve as a potential autologous cell source for therapy. Previously murine and sheep AFS have shown to have significant haematopoietic activity after transplantation in immune deficient mice. The haematopoietic potential of Human AFS have never been established in vivo, and its use has been limited by the presence of debris and low cell number at sample collection. My thesis explored the (1) isolation of human amniotic fluid (AF), (2) haematopoietic potential of human AF (CD117/c-Kit+; AFSC) by reconstituting the haematopoietic system of NOD-SCID/IL2rγnull (NSG) mice in vivo and (3) expansion of haematopoietic human AFSC in vitro. Human AF samples (2nd and 3rd trimester, n=110) were collected for the study under an ethically approved project from women undergoing amniocentesis for prenatal diagnosis of congenital disease, or amniodrainage procedures for fetal abnormality. I have employed several strategies to eliminate the large amount of cellular debris from the collected human AF and provide a more homogeneous cell population. Percoll density centrifugation demonstrated a reduction in cell debris and enrichment of the CD117+ population. The haematopoietic potential of human AFSC was explored in vivo. Human AF (2nd and 3rd trimester) and cord blood (CB; control) were selected for CD117 and CD34 respectively. Sorted cells (104 in 200μl PBS) were injected intravenously into sub-lethally irradiated NSG mice (~n=6/group). Human AFSC engrafted the haematopoietic system of NSG mice at levels similar to those achieved with CB-HSC post-primary and secondary transplantation. Importantly, multi-lineage haematopoietic reconstitution was observed at 16 weeks post-primary and secondary transplantation. Moreover, the possibility of expanding haematopoietic progenitors from human AF in vitro was demonstrated with the use of a cytokine-based media and the generation of haematopoietic progenitors by AF derived-induced pluripotent stem cell (AF-iPS) lines. In conclusion, I showed that human AF could be isolated, have long-term multi-lineage haematopoietic potential that is similar to the current “gold-standard” stem cell source for haematopoietic transplantation as well as demonstrates haematopoietic expansion. These findings make human AFSC to be an alternative novel fetal cell source for pre- and post-natal cell or cell-based gene therapy for the treatment of haematological disorders in the future.
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Taoudi, Samir. "Emergence and expansion of embryonic definitive haematopoietic stem cells." Thesis, University of Edinburgh, 2006. http://hdl.handle.net/1842/14526.
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De novo generation and physiological expansion of definitive haematopoietic stem cells (dHSCs) occurs exclusively during embryogenesis and is initiated within the dorsal aorta/para-aortic mesenchyme (Ao) of the E11.5 aorta-gonad-mesonephros (AGM) region. The elucidation of the in vivo induction of dHSC emergence and regulation of expansion is likely to be a preparation to the success of generating dHSC in vitro and facilitating an effective and robust expansion of pre-existing dHSCs, thus providing a basis for potentially useful therapeutic translation. Utilising flow cytometry and the in vivo long-term haematopoietic reconstitution assays, we describe the endothelial affiliation of early dHSC as they emerge within the embryonic AGM region and yolk sac, fundamentally defined by a VE-cadherin +CD45+ immunophenotype, and the successive restriction to a haematopoietic identity upon hepatic colonisation. Well-defined haematopoietic colony forming unit-culture (CFU-C) and endothelial network forming assays were used to reveal that the early strong endothelio-haematopoietic promiscuity observed in the early embryo is lost in favour of a mutually exclusive lineage commitment. Experiments using a novel liquid suspension culture system reveal that successful in vitro myeloid differentiation of VE-cadherin +CD45+dHSCs/multipotent haematopoietic progenitors from the E11.5 AGM region is dependent on an interaction with the endothelial compartment. To directly test the predominating hypothesis of a dorso-ventral polarity in haematopoietic activity of the AGM region I have bisected the Ao from the E11.5 AGM region along the dorso-ventral axis and subjected the resultant ventral (AoV) and dorsal (AoD) aspects to in vivo and in vitro tests. Here I provide the first functional data to support the hypothesis of an anatomical polarity of dHSC distribution. Furthermore, I provide evidence to support an hypothesis that in vivo components of the E10.5-11.5 Ao only permit ventral emergence and expansion of dHSCs. Investigation into the transcriptional activity of the E11.5 AoV and AoD highlight the differential expression of GATA2, GATA3, HoxB4, Runxl, BMP4, Noggin and Chordin. These data demonstrate the conductive nature of the AGM region, which after structural disruption not only supports the ex vivo maintenance of dHSC activity but also exhibits a marked capacity for rapid stem cell expansion. Thus, in our hands we have a powerful system with the potential to allow useful investigation into the fundamental processes that occupy the field of stem cell biology; the mechanisms of stem cell emergence, induction and expansion.
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Stephenson, Rachel A. "Notch signalling in Xenopus laevis haematopoietic stem cell programming." Thesis, University of Oxford, 2013. http://ora.ox.ac.uk/objects/uuid:4a6db3d9-14ec-4ad2-ad7f-06705c49d32e.
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Multipotent haematopoietic stem cells (HSCs) originate in the dorsal aorta (DA) during vertebrate embryogenesis, and after migrating to a permanent niche, give rise to a continuous supply of mature blood cells of all lineages throughout adult life. Previous cell tracing experiments have shown that the cells of the DA migrate here from an early collection of haemangioblasts (bipotential precursors of blood and endothelial cells) which reside in the dorsolateral plate (DLP) mesoderm. Development of HSCs is tightly regulated by a number of key signalling pathways in both the DLP and the DA. In particular, notch signalling is considered an important factor in vascular, arterial and HSC development. Here, the relatively slow development and the spatial separation of definitive haematopoiesis from primitive haematopoiesis in Xenopus laevis has been exploited to reveal the first defect of reduced notch signalling in the Xenopus DA. Two notch inputs to HSC programming have been identified in Xenopus: notch4 and its target genes, esr7 and esr10, are expressed from stage 31, immediately after migrating haemangioblast cells reach the midline of the embryo to form the DA, whilst notch1 is expressed slightly later, from stage 34, and controls expression of two further notch target genes, esr1 and hesr1. Using both morpholino knockdown of these six genes, and chemical inhibition of notch signalling using a specific γ-secretase inhibitor, notch signalling has been demonstrated to be essential for HSC programming but dispensable for earlier haemangioblast and arterial programming. Furthermore, esr1, downstream of both notch1 and notch4, is shown to be responsible for repression of endothelial genes in the DA. Taken together, this demonstrates that a cascade of notch and notch effector genes are essential for the programming of Xenopus HSCs.
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Nottingham, Wade. "Transcriptional regulation of Runx1 in the developing haematopoietic system." Thesis, University of Oxford, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.670091.
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Dalakas, Evangelos. "Haematopoietic stem cell response in alcohol induced liver injury." Thesis, University of Edinburgh, 2009. http://hdl.handle.net/1842/29723.
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Aims: 1) To investigate the mobilisation and hepatic recruitment of HSCs in patients with alcohol induced liver injury and define their contribution to parenchymal and nonparenchymal liver cell lineages. 2) To establish that mobilised HSCs in alcohol induced liver injury are functional and demonstrate pluripotent stem cell properties. 3) To study the role of inflammatory cytokines and chemokine axes in regulating the mobilisation and hepatic recruitment of HSCs in alcohol induced liver injury. Methods: Liver biopsies from alcoholic hepatitis (AH) patients and male patients who had received a female liver transplant (and who subsequently developed AH) were analysed for HSC content using immunohistochemical and flurorescent in situ hybridisation techniques (FISH). FISH for Y-chromosome was performed on liver tissue, along with co-staining for hepatocyte, biliary, myofibroblasts (α-SMA) and hepatic parenchymal cells proliferation markers. Peripheral blood HSC (CS34+) levels were quantified in AH patients and normal controls (NC) using flow cytometry and CD34+/CD45+ HSCs were collected and cultured in colony forming unit (CFU) assays. CXCR3/CXCR4 receptor cell expression on mobilised CD34+ HSCs were quantified in AH patients using flow cytometry. Conclusions: Alcohol induced liver injury mobilises CD34+ stem cells into the peripheral circulation and recruits them into the liver. These bone marrow derived stem cells contribute to the hepatic myofibroblasts population but not to parenchymal lineages and their presence within the liver does not promote hepatocyte repair. Mobilised HSCs from AH patients were functional and displayed true stem cell potential at a level higher than control HSCs. Serum SDF-1, MMP-9 and G-CSF rather than chemokine receptor expression, plays a central role in regulating the mobilisation of CD34+ stem cells in alcohol induced liver injury.
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Ewels, Philip Andrew. "Spatial organisation of proto-oncogenes in human haematopoietic progenitor cells." Thesis, University of Cambridge, 2013. https://www.repository.cam.ac.uk/handle/1810/245861.
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The eukaryotic cell nucleus is a highly organised organelle, with distinct specialised sub- compartments responsible for specific nuclear functions. Within the context of this functional framework, the genome is organised, allowing contact between specific genomic regions and sub-compartments. Previous work has shown that genes in both cis and trans can make specific contacts with each other. I hypothesise that such a preferred juxtaposition may impact the propensity for specific cancerinitiating chromosomal translocations to occur. In this thesis, I describe how I have extended and developed a ligation based proximity assay known as enriched 4C. I have coupled this technique with high throughput sequencing to determine genomic regions that spatially co-associate with the proto-oncogenes MLL, ABL1 and BCR. In addition to further developing the laboratory protocol, I have created bioinformatics tools used in the analysis of the sequencing data. I find that the association profiles of the three genes show strong correlation to the binding profile of RNA polymerase II and other active marks, suggesting that transcribed genes have a propensity to associate with other transcribed regions of the genome. Each gene also exhibits a unique repertoire of preferred associations with specific regions of the genome. Significantly, I find that the most frequent trans association of BCR is telomeric chromosome 9, encompassing its recurrent translocation partner gene ABL1. Interestingly, ABL1 is not at the maximum point of interaction. I use DNA-fluorescence in-situ hybridisation to validate the e4C association. My data supports a hypothesis that gene transcription has a direct role on genome organisation. I suggest that preferred co-associations of genes at transcription factories may promote the occurrence of specific chromosomal translocations.
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Chandrashekran, Anil. "The development of retroviral vectors targeted to haematopoietic stem cells." Thesis, Imperial College London, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.404399.
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O'Leary, H. A. ngharad E. S. G. "Heparan sulphate inhibits haematopoietic stem cell homing in mucopolysaccharidosis I." Thesis, University of Manchester, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.528510.
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Mortensen, Monika. "The role of autophagy in the haematopoietic system in vivo." Thesis, University of Oxford, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.514970.
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Green, Angela Lisa. "Role of the haematopoietic transcription factor SCL in mesoderm development." Thesis, University of Oxford, 2012. http://ora.ox.ac.uk/objects/uuid:dd7a6c6a-0a84-408b-96ad-8e4922f5ca80.
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During embryonic development, precursor cells commit to specific cell fates in response to environmental cues through the establishment of lineage-specific gene expression programmes. Transcription factors are important downstream effectors of signalling pathways that initiate and maintain cell fate decisions. The haematopoietic transcription factor SCL (TAL-1) is an essential regulator of embryonic blood development. However, the exact stage at which SCL is required, its mechanisms of action, and its genomic targets are poorly understood. Characterising, jiow SCL functions - , during haematopoietic development will provide insights into how stern cells are specified. Using the embryonic stem cell/embryoid body (ES/EB) system to model early mouse development, we describe a critical role for SCL in mesoderm patterning. SCL is first expressed in PDGFRa+ FLK1+ mesoderm populations which contain lateral, paraxial and cardiac precursors. Through loss- and gain-of-function studies, we show that SCL drives lateral mesoderm specification and activates the haematopoietic programme in a direct DNA-binding independent manner, while actively repressing alternative mesodermal fates, specifically cardiac development, in a DNA-binding dependent manner. At a molecular level, we have identified direct genomic targets of SCL in Flk-1 + mesoderm populations. These include haematopoietic and cardiac transcription factors, cardiac-specific structural proteins, signalling proteins and general transcriptional repressors; thereby strengthening the dual function of SCL in mesoderm patterning. Finally, we have shown that the cardiac transcription factor GATA4 acts in a reciprocal manner, specifying cardiac precursors while repressing a lateral mesoderm fate. Collectively, this implicates SCL as a critical transcriptional regulator of cell fate decisions in early mesodermal precursors, employing distinct molecular mechanisms to impose a blood programme. Moreover, and extending earlier reports, we document the existence of an antagonistic cross-talk between haematopoietic and cardiac lineages during mesoderm patterning. In conclusion, this work offers a cellular and molecular platform to begin to dissect the network of genetic interactions involved in these developmental processes.
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Batsivari, Antoniana. "Studying the cell cycle status during haematopoietic stem cell development." Thesis, University of Edinburgh, 2016. http://hdl.handle.net/1842/25802.
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In adults blood stem cells, called haematopoietic stem cells (HSC), give rise to all blood cells throughout life. The origin and biology of HSCs during embryo development has been an intensely studied topic. Definitive HSCs are generated intra-embryonically in the aorta-gonad-mesonephros (AGM) region of the mid-gestation embryo. Recent research revealed that HSCs emerge through multistep maturation of precursors: proHSC → preHSC I → preHSC II → definitive HSC (dHSC). A hallmark of the HSC emergence is the appearance of intra-aortic haematopoietic clusters that are considered to be sites of haematopoiesis. It was shown in vitro that the E11.5 HSCs are slowly cycling compared to progenitor cells. However, cell cycle status and its role during early HSC development remain unclear. Here I used Fucci transgenic mice that enable in vivo visualisation of the cell cycle. Functional and phenotypic analysis showed that in the early embryo the proHSC precursors cycle slowly, whereas committed progenitors are actively cycling. Meanwhile the preHSC I precursors arising in the E10.5 AGM region become more rapidly cycling. They are located closer to the luminal cavity of the dorsal aorta, while their ancestors, the proHSCs, are slowly cycling and are located at base of the clusters. Furthermore, in the mid-gestation embryo the preHSC I become slowly cycling and are closer to the endothelial lining of the aorta, while they give rise to the actively cycling preHSC II that are located to the luminal area of the artery. Finally, definitive HSCs are mainly slowly cycling at this stage like their foetal liver counterparts. As expected, HSCs in adult bone marrow are mainly dormant. The data suggest that transition from one precursor type to another is accompanied by distinct changes in cell cycle profile and that HSCs become progressively quiescent during development. To test the role of cell cycle in HSC maturation, we used inhibitors against signalling pathways known to play important roles in HSC development. Notch inhibitor affected the cell cycle status of haematopoietic precursors, by possibly promoting them to rapidly proliferate and potentially blocking the maturation from preHSC I to preHSC II precursors. Shh antagonist had the opposite effect and enhanced the HSC activity from the preHSC I precursors. Altogether these results suggest that the cell cycle status plays an important role in the HSC development. A better understanding of the molecules that control this process will allow us to optimize the culture condition for generation of functional HSCs in the laboratory.
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Turner, Marc Leighton. "Cell adhesion of human haematopoietic progenitors : development of assay techniques." Thesis, University of Edinburgh, 1996. http://hdl.handle.net/1842/21578.
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Elucidation of the mechanisms underlying the reciprocal phenomena of HPC homing and mobilisation is the objective of this thesis, and may allow the development of novel approaches to mobilisation regimens and/or ex vivo HPC manipulation. Accurate and reproducible assays for quantitative and qualitative studies of HPC were established. These included an immunocytometry based assay of CE34 antigen expression, dual colour immunocytometry studies of cell adhesion molecule, lineage and activation marker expression by HPC derived from different sources, and three colour immunocytometry of adhesion molecule expression within HPC subsets. A chromium51-labelling technique was developed as a functional assay with which to examine the adhesion of haematopoietic cell lines to extracellular matrix components, stromal and endothelial tissues in culture. A variety of techniques for adhesion blockade were explored. A protocol for high-purity enrichment of HPC was developed, and the feasibility of applying the chromium51 adhesion assay to these cells was examined. CD34 immunocytometry was confirmed as a valid method for defining HPC populations. Marrow HPC were found to express nine adhesion molecules, two of which were reduced by circulating cells. HPC derived from different sources showed variation in lineage and activation marker expression, and HPC subsets displayed differences in adhesion molecule expression. Haematopoietic cell lines adhered to fibronectin and thrombospondin, but not to other extracellular matrix components. Blockade of fibronectin adhesion was effected by divalent cation chelation, synthetic peptides and chondroitinase ABC. Cell line adhesion to stromal and endothelial tissue cultures was demonstrated, but highly-enriched HPC labelled poorly with chromium51.
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Garaycoechea, Amoroso Juan Ignacio. "Aldehyde genotoxicity underlies haematopoietic stem cell depletion in Fanconi anaemia." Thesis, University of Cambridge, 2014. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.708180.
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Hull, Katrusha. "Oral Long-Term Complications of Allogeneic Haematopoietic Stem Cell Transplantation." Thesis, Faculty of Dentistry, 2010. http://hdl.handle.net/2123/5930.
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Facchini, Raffaella Maria. "Investigating the specific roles of the growth factor kit ligand in the regulation of murine haematopoiesis." Thesis, University of Oxford, 2015. http://ora.ox.ac.uk/objects/uuid:35da4965-5da9-4007-bf97-8408f0f5ed06.
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McGarvey, Alison Clare. "Genome-wide transcriptional characterisation and investigation of the murine niche for developing haematopoietic stem cells." Thesis, University of Edinburgh, 2017. http://hdl.handle.net/1842/28741.
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Haematopoietic stem cells (HSCs) are capable of differentiation into all mature haematopoietic lineages, as well as long-term self-renewal and are consequently able to sustain the adult haematopoietic system throughout life. Currently, in the mouse, HSCs are understood to first appear in the aorta-gonad-mesonephros (AGM) region at embryonic day 11 via a process of maturation from precursors (pre-HSCs). This maturation within the AGM region involves the complex interplay of signalling between cells of the niche and maturing precursor cell populations, but is relatively little understood at a molecular level. Recently our understanding of the AGM region has been refined, identifying the progression from E9.5 to E10.5 and the polarity along the dorso-ventral axis as clear demarcations of the supportive environment for HSC maturation. In this thesis, I investigated the molecular characteristics of these spatio-temporal transitions in the AGM region through the application of RNA-sequencing. This enabled the identification of molecular signatures which may underlie the supportive functionality of the niche. I further compared these expression signatures to the transcriptional profile of an independent cell type, also capable of supporting HSC maturation, the OP9 stromal cell line. By combining this transcriptional information with an ex vivo culture system, I screened a number of molecules for their ability to support HSC maturation from early precursors, leading to the discovery of a novel regulator of HSC maturation: BMPER. Further characterisation of this molecule enabled the identification of its specific cellular source and the proposal that through its action as an inhibitor of BMP signalling it facilitates the maturation of precursors into HSCs. These results lend further detail and support to the role of BMP signalling in the regulation of HSC maturation as well as demonstrating the potential of these transcriptional profiles to yield novel mechanistic insight.
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Schmidt, Sarah Marianne. "The role of Kindlin-3 in cells of the haematopoietic system." Diss., lmu, 2011. http://nbn-resolving.de/urn:nbn:de:bvb:19-134375.
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Mahdipour, Elahe. "Haematopoietic stem/progenitor cell migration and differentiation in response to injury." Thesis, University of Manchester, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.518453.
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Hamid, Zariyantey Abdul. "Protein transduction for the in-vitro expansion of haematopoietic stem cells." Thesis, King's College London (University of London), 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.682779.
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