Suzanne M Watt

NHS Blood and Transplant, Watford, England, United Kingdom

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Publications (136)590.39 Total impact

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    ABSTRACT: Transcriptional Profiling of Human Cord Blood CD133� and Cultured Bone Marrow Mesenchymal Stem Cells in Response to Hypoxia ABSTRACT Umbilical cord blood (UCB) and bone marrow (BM)-derived stem and progenitor cells possess two characteristics required for successful tissue regeneration: extensive proliferative capacity and the ability to differentiate into multiple cell lineages. Within the normal BM and in pathological conditions, areas of hypoxia may have a role in maintaining stem cell fate or determining the fine equilibrium between their proliferation and differentiation. In this study, the transcriptional profiles and proliferation and differentiation potential of UCB CD133� cells and BM mesenchymal cells (BMMC) exposed to normoxia and hypoxia were analyzed and compared. Both progenitor cell populations responded to hypoxic stimuli by stabilizing the hypoxia inducible factor (HIF)-1� protein. Short exposures to hypoxia increased the clonogenic myeloid capacity of UCB CD133� cells and promoted a significant increase in BMMC number. The differentiation potential of UCB CD133� clonogenic myeloid cells was unaltered by short exposures to hypoxia. In contrast, the chondrogenic differentiation potential of BMMCs was enhanced by hypoxia, whereas adipogenesis and osteogenesis were unaltered. When their transcriptional profiles were compared, 183 genes in UCB CD133� cells and 45 genes in BMMC were differentially regulated by hypoxia. These genes included known hypoxia-responsive targets such as BNIP3, PGK1, ENO2, and VEGFA, and other genes not previously described to be regulated by hypoxia. Several of these genes, namely CDTSPL, CCL20, LSP1, NEDD9, TMEM45A, EDG-1, and EPHA3 were confirmed to be regulated by hypoxia using quantitative reverse transcriptase polymerase chain reaction. These results, therefore, provide a global view of the signaling and regulatory network that controls oxygen sensing in human adult stem/progenitor cells derived from hematopoietic tissues. STEM CELLS 2007; 25:1003–1012
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    ABSTRACT: Motivation: Many important problems in cell biology require dense nonlinear interactions between functional modules to be considered. The importance of computer simulation in understanding cellular processes is now widely accepted, and a variety of simulation algorithms useful for studying certain subsystems have been designed. Expansion of hematopoietic stem and progenitor cells (HSC/HPC) in ex vivo culture with cytokines and small molecules is a method to increase the restricted numbers of stem cells found in umbilical cord blood while also enhancing the content of early engrafting neutrophil and platelet precursors. The efficacy of the expanded product depends on the composition of the cocktail of cytokines and small molecules used for culture. Testing the influence of a cytokine or small molecule on the expansion of HSC/HPC is a laborious and expensive process. We therefore developed a computational model based on cellular signaling interactions that predict the influence of a cytokine on the survival, duplication and differentiation of the CD133+ HSC/HPC subset from human umbilical cord blood (CB). Results: We have used results from in vitro expansion cultures with different combinations of one or more cytokines to develop an ordinary differential equation model that includes the effect of cytokines on survival, duplication and differentiation of the CD133+ HSC/HPC. Comparing the results of in vitro and in silico experiments, we show that the model can predict the effect of a cytokine on the fold expansion and differentiation of CB CD133+ HSC/HPC after 8 day culture on a 3D scaffold. Availability and implementation: The model is available visiting the following url: http://www.francescopappalardo.net/Bioinformatics_CD133_Model Contact: francesco.pappalardo{at}unict.it; suzanne.watt{at}nhsbt.nhs.uk
    Bioinformatics 04/2015; DOI:10.1093/bioinformatics/btv172 · 4.62 Impact Factor
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    ABSTRACT: Murine models of bone marrow transplantation show that pre-conditioning regimens affect the integrity of the bone marrow endothelium and that the repair of this vascular niche is an essential pre-requisite for successful haematopoietic stem and progenitor cell engraftment. Little is known about the angiogenic pathways that play a role in the repair of the human bone marrow vascular niche. We therefore established an in vitro humanized model, composed of bone marrow stromal and endothelial cells and have identified several pro-angiogenic factors, VEGFA, ANGPT1, CXCL8 and CXCL16, produced by the stromal component of this niche. We demonstrate for the first time that addition of CXCL8 or inhibition of its receptor, CXCR2, modulates blood vessel formation in our bone marrow endothelial niche model. Compared to wild type, Cxcr2(-/-) mice displayed a reduction in bone marrow cellularity and delayed platelet and leucocyte recovery following myeloablation and bone marrow transplantation. The delay in bone marrow recovery correlated with impaired bone marrow vascular repair. Taken together, our data demonstrate that CXCR2 regulates bone marrow blood vessel repair/regeneration and haematopoietic recovery, and clinically may be a therapeutic target for improving bone marrow transplantation. © 2015 The Authors. British Journal of Haematology published by John Wiley & Sons Ltd.
    British Journal of Haematology 03/2015; 169(4). DOI:10.1111/bjh.13335 · 4.96 Impact Factor
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    ABSTRACT: BACKGROUND: Expansion of human cord blood (CB) CD341 cells with thrombopoietin (TPO) can accelerate delayed platelet (PLT) recovery after transplantation into immunodeficient mice. Clinical implementation, however, will depend on practical and effective protocols. The best timing of TPO expansion in relation to cryopreservation in this respect is unknown. STUDY DESIGN AND METHODS: In this study, we evaluated whether the order of cryopreservation and TPO expansion affected the expansion rate and numbers of clonogenic hematopoietic progenitor cells in vitro or PLT and longer-term hematopoietic repopulation in NOD SCID mice in vivo. RESULTS: Our results demonstrate higher expansion rates and the generation of higher numbers of multilineage and megakaryocytic progenitors (granulocyte, erythrocyte, monocyte, megakaryocyte colony-forming units and megakaryocyte colony-forming units) in vitro when freshly isolated CB CD341 cells are first cultured with TPO and then cryopreserved and thawed as compared to TPO expansion after CD341 cell cryopreservation. In contrast, the cells produced with the latter strategy showed higher expression of CD62L and a superior stromal cell–derived factor-1a–mediated migration. This might play a role in an also observed superior early PLT recovery after transplantation of these cells into NOD SCID mice. The hematopoietic engraftment in the marrow 6 weeks after transplantation was not different between the two strategies. CONCLUSION: Although TPO expansion before cryopreservation would yield higher nucleated cell and clonogenic myeloid and megakaryocyte cell numbers and enable earlier availability, CB TPO expansion after cryopreservation is likely to be clinically more effective, despite the lower number of cells obtained after expansion. Moreover, the latter strategy is logistically more feasible.
    Transfusion 02/2015; DOI:10.1111/trf.13045 · 3.57 Impact Factor
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    ABSTRACT: The differentiation of human pluripotent stem cells to the B-cell lymphoid lineage has important clinical applications that include in-vitro modelling of developmental lymphogenesis in health and disease. Here, we first demonstrate the capacity of human induced pluripotent stem cells (hiPSCs) to differentiate into CD144+CD73-CD43/CD235a-, cells, characterised as hemogenic endothelium, and show this population is capable of differentiating to CD10+CD19+ B lymphocytes. We also demonstrate that B lymphocytes generated from hiPSCs are able to undergo full VDJ rearrangement and express surface IgM (sIgM+), thus representating an immature B-cell subset. Efficiency of sIgM expression on the iPS-derived B lymphocytes (c. 5% of CD19+ cells) was comparable with B lymphocytes generated from human umbilical cord blood hematopoietic progenitor cells. Importantly, when assessed by global transcriptional profiling, hiPSC-derived B-cells show a very high level of similarity when compared to their umbilical cord blood derived counterparts, such that from over 47,000 different transcripts, only 45 were significantly different (with a criteria adjusted P value P˂0.05, log FC ˃1.0 or 2.8 fold). This represents a unique in-vitro model to delineate critical events during lymphogeneisis in development and lymphoid diseases such as acute lymphocytic leukemia.
    Stem Cells and Development 12/2014; DOI:10.1089/scd.2014.0318 · 4.20 Impact Factor
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    ABSTRACT: Human cord blood (CB) hematopoietic stem (HSC) cell transplants demonstrate delayed early neutrophil and platelet recovery and delayed longer term immune reconstitution compared to bone marrow and mobilized peripheral blood transplants. Despite advances in enhancing early neutrophil engraftment, platelet recovery after CB transplantation is not significantly altered when compared to contemporaneous controls. Recent studies have identified a platelet biased murine HSC subset, maintained by thrombopoietin (TPO) which has enhanced capacity for short and long term platelet reconstitution, can self-renew, and can give rise to myeloid and lymphoid biased HSCs. In previous studies, we have shown that transplantation of human CB CD34+ cells pre-cultured in TPO as a single graft accelerates early platelet recovery as well as yielding long term repopulation in immune deficient mice. Here, using a double CB murine transplant model, we investigated whether TPO cultured human CB CD34+ cells have a competitive advantage or disadvantage over untreated human CB CD34+ cells in terms of i) short and longer term platelet recovery and ii) longer term hematological recovery. Our studies demonstrate that the TPO treated graft shows accelerated early platelet recovery without impairing the platelet engraftment of untreated CD34+ cells. Notably, this was followed by a dominant contribution to platelet production via the untreated CD34+ cell graft over the intermediate to longer term. Furthermore, although the contribution of the TPO treated graft to long term hematological engraftment was reduced, the TPO treated and untreated grafts both contributed significantly to long term chimerism in vivo.
    Stem Cells and Development 08/2014; DOI:10.1089/scd.2014.0294 · 4.20 Impact Factor
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    ABSTRACT: Proangiogenic factors, VEGF and FGF-2, prime endothelial cells to respond to 'hematopoietic' chemokines and cytokines by inducing/upregulating expression of the respective chemokine/cytokine receptors. Co-culture of human endothelial colony forming cell (ECFC)-derived cells with human stromal cells in the presence of VEGF and FGF-2 for 14 days resulted in upregulation of the 'hematopoietic' chemokine CXCL12 and its CXCR4 receptor by day 3 of co-culture. Chronic exposure to the CXCR4 antagonist, AMD-3100, in this vasculo/angio-genesis assay, significantly reduced vascular tubule formation, an observation recapitulated by delayed AMD3100 addition. While AMD3100 did not affect ECFC-derived cell proliferation, it did demonstrate a dual action. First, over the later stages of the 14 day co-cultures, AMD3100 delayed tubule organization into maturing vessel networks, resulting in enhanced endothelial cell retraction and loss of complexity as defined by live cell imaging. Secondly, at earlier stages of co-cultures, we observed that AMD3100 significantly inhibited the integration of exogenous ECFC-derived cells into established, but immature, vascular networks. Comparative proteome profiler array analyses of ECFC-derived cells treated with AMD3100 identified changes in expression of potential candidate molecules involved in adhesion and/or migration. Blocking antibodies to CD31, but not CD146 or CD166, reduced the ECFC-derived cell integration into these extant vascular networks. Thus, CXCL12 plays a key role not only in endothelial cell sensing and guidance, but also in promoting the integration of ECFC-derived cells into developing vascular networks.
    Stem Cells and Development 06/2014; 23(22). DOI:10.1089/scd.2014.0005 · 4.20 Impact Factor
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    ABSTRACT: Human induced pluripotent stem cells (hiPSCs), like embryonic stem cells, are under intense investigation for novel approaches to model disease and for regenerative therapies. Here, we describe the derivation and characterization of hiPSCs from a variety of sources and show that, irrespective of origin or method of reprogramming, hiPSCs can be differentiated on OP9 stroma towards a multi-lineage haemo-endothelial progenitor that can contribute to CD144+ endothelium, CD235a+ erythrocytes (myeloid lineage) and CD19+ B lymphocytes (lymphoid lineage). Within the erythroblast lineage, we were able to demonstrate by single cell analysis (flow cytometry), that hiPSC-derived erythroblasts express alpha globin as previously described, and that a sub-population of these erythroblasts also express haemoglobin F (HbF), indicative of fetal definitive erythropoiesis. More notably however, we were able to demonstrate that a small sub-fraction of HbF positive erythroblasts co-expressed HbA in a highly heterogeneous manner, but analogous to cord blood-derived erythroblasts when cultured using similar methods. Moreover, the HbA expressing erythroblast population could be greatly enhanced (44·0 ± 6·04%) when a defined serum-free approach was employed to isolate a CD31+ CD45+ erythro-myeloid progenitor. These findings demonstrate that hiPSCs may represent a useful alternative to standard sources of erythrocytes (RBCs) for future applications in transfusion medicine.
    British Journal of Haematology 05/2014; 166(3). DOI:10.1111/bjh.12910 · 4.96 Impact Factor
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    ABSTRACT: Cryopreserved umbilical cord blood (CB) is increasingly used as a cell source to reconstitute marrow in hematopoietic stem cell transplant patients. Delays in cryopreservation may adversely affect cell viability, thereby reducing their potential for engraftment after transplantation. The impact of delayed cryopreservation for up to 3 days on the viability of both CD45+ and CD34+ cell populations in 28 CB donations with volumes of 58.40 ± 15.4 mL (range, 39.4-107.4 mL) was investigated to establish whether precryopreservation storage time could be extended from our current time of 24 to 48 hours in line with other CB banks. Viability was assessed on 3 consecutive days, both before and after cryopreservation, by flow cytometry using 7-aminoactinomycin D (7-AAD) and annexin V methods. The results using 7-AAD and annexin V indicated the viability of CD34+ cells before cryopreservation remained high (>92.33 ± 4.11%) over 3 days, whereas the viability of CD45+ cells decreased from 86.36 ± 4.97% to 66.24 ± 7.78% (p < 0.0001) by Day 3. Storage time significantly affected the viability of CD34+ cells after cryopreservation. Using 7-AAD, the mean CD34+ cell viability decreased by approximately 5% per extra day in storage from 84.30 ± 6.27% on Day 1 to 79.01 ± 7.44% (p < 0.0057) on Day 2 and to 73.95 ± 7.54% (p < 0.0001) on Day 3. With annexin V staining CD34+ cell viability fell by approximately 7% per extra day in storage from 77.17 ± 8.47% on Day 1 to 69.56 ± 13.30% (p < 0.0194) on Day 2 and to 62.89 ± 15.22% (p < 0.0002) on Day 3. This study demonstrates that extended precryopreservation storage adversely affects viability and should be avoided.
    Transfusion 11/2013; 54(5). DOI:10.1111/trf.12481 · 3.57 Impact Factor
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    ABSTRACT: Adoptive cell therapy employing gene-modified T-cells expressing chimeric antigen receptors (CARs) has shown promising preclinical activity in a range of model systems and is now being tested in the clinical setting. The manufacture of CAR T-cells requires compliance with national and European regulations for the production of medicinal products. We established such a compliant process to produce T-cells armed with a first-generation CAR specific for carcinoembryonic antigen (CEA). CAR T-cells were successfully generated for 14 patients with advanced CEA(+) malignancy. Of note, in the majority of patients, the defined procedure generated predominantly CD4(+) CAR T-cells with the general T-cell population bearing an effector-memory phenotype and high in vitro effector function. Thus, improving the process to generate less-differentiated T-cells would be more desirable in the future for effective adoptive gene-modified T-cell therapy. However, these results confirm that CAR T-cells can be generated in a manner compliant with regulations governing medicinal products in the European Union.
    Cancer Immunology and Immunotherapy 11/2013; 63(2). DOI:10.1007/s00262-013-1492-9 · 3.94 Impact Factor
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    ABSTRACT: Background Blood vessel formation is fundamental to development, while its dysregulation can contribute to serious disease. Expectations are that hundreds of millions of individuals will benefit from therapeutic developments in vascular biology. MSCs are central to the three main vascular repair mechanisms.Sources of dataKey recent published literature and ClinicalTrials.gov.Areas of agreementMSCs are heterogeneous, containing multi-lineage stem and partly differentiated progenitor cells, and are easily expandable ex vivo. There is no single marker defining native MSCs in vivo. Their phenotype is strongly determined by their specific microenvironment. Bone marrow MSCs have skeletal stem cell properties. Having a perivascular/vascular location, they contribute to vascular formation and function and might be harnessed to regenerate a blood supply to injured tissues.Areas of controversyThese include MSC origin, phenotype and location in vivo and their ability to differentiate into functional cardiomyocytes and endothelial cells or act as vascular stem cells. In addition their efficacy, safety and potency in clinical trials in relation to cell source, dose, delivery route, passage and timing of administration, but probably even more on the local preconditioning and the mechanisms by which they exert their effects.Growing pointsUnderstanding the origin and the regenerative environment of MSCs, and manipulating their homing properties, proliferative ability and functionality through drug discovery and reprogramming strategies are important for their efficacy in vascular repair for regenerative medicine therapies and tissue engineering approaches.Areas timely for developing researchCharacterization of MSCs' in vivo origins and biological properties in relation to their localization within tissue niches, reprogramming strategies and newer imaging/bioengineering approaches.
    British Medical Bulletin 10/2013; DOI:10.1093/bmb/ldt031 · 3.95 Impact Factor

Publication Stats

4k Citations
590.39 Total Impact Points

Institutions

  • 2006–2015
    • NHS Blood and Transplant
      Watford, England, United Kingdom
  • 1999–2014
    • University of Oxford
      • • Chemical Research Laboratory
      • • Nuffield Division of Clinical Laboratory Sciences
      • • Weatherall Institute of Molecular Medicine
      • • Molecular Haematology Unit
      Oxford, England, United Kingdom
  • 1995–2014
    • Oxford University Hospitals NHS Trust
      • • Nuffield Department of Clinical Laboratory Sciences
      • • Department of Paediatrics
      Oxford, England, United Kingdom
  • 2008
    • The Bracton Centre, Oxleas NHS Trust
      Дартфорде, England, United Kingdom
  • 1983–2001
    • University of Cambridge
      • • Department of Biochemistry
      • • Department of Pathology
      Cambridge, England, United Kingdom
  • 1991
    • Cancer Research UK
      Londinium, England, United Kingdom
  • 1978–1981
    • Royal Melbourne Hospital
      Melbourne, Victoria, Australia
  • 1979
    • The Walter and Eliza Hall Institute of Medical Research
      Melbourne, Victoria, Australia