Dissertations / Theses on the topic 'Blood endothelial cells'
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Al-Malki, Aysha Ibrahim. "Detection of endothelial cells in whole blood donations." Thesis, University of Sheffield, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.531130.
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Yuan, Yifan. "Enhancing Blood Outgrowth Endothelial Cells for Optimal Coating of Blood Contacting Surfaces." Thesis, Université d'Ottawa / University of Ottawa, 2017. http://hdl.handle.net/10393/36837.
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Implantable cardiovascular biomaterials have been widely applied in multiple cardiovascular disorders such as coronary artery disease, heart failure, and abdominal aortic aneurysms. However the failure modes of cardiovascular biomaterials are not uncommon, which is mainly due to the complications on blood-contacting surfaces such as thrombosis, calcification, and inflammation. Endothelium locates the inner surface of vessel lumen and is a critical regulator of vascular homeostasis. However, a readily available functional autologous source of endothelium has been hard to achieve. Human blood outgrowth endothelial cells (BOECs), cultured from peripheral blood mononuclear cells are proliferative and express endothelial protein profiles and as such are a very promising novel cell source for cardiovascular biomaterials coating. Endothelial nitric oxide synthase (eNOS) is an important regulator of vascular homeostasis and loss of eNOS activity is a hallmark of endothelial dysfunction. My data demonstrated that BOECs express markedly lower eNOS protein, mRNA as well as activity levels when compared to mature endothelial cells (ECs). My first project was to use transient transfection methods along with minicircle DNA to enhance eNOS expression levels in BOECs. Two promoters were tested in BOECs, the CMV promoter (pMini-CMV-eNOS) and the EF1α promoter (pMini-EF1α-eNOS). Transfection with pMini-CMV-eNOS achieved 24.8 ± 5.1 times more eNOS expression when compared to null transfected cells at 24 hours, a marked improvement over that achieved with conventional PVAX plasmid (10.2 ± 4.7 fold increase) or pMini-EF1α-eNOS (8.2 ± 1.2 fold increase both compared to null transfected control). pMini-CMV-eNOS mediated overexpression improved cell migration and network formation. When cultured on Osteopontin (OPN) coated surfaces, transient transfection with plasmid eNOS in BOECs can markedly enhance cell spreading and adhesion to ECM modified surfaces. These results suggest that eNOS expression in BOECs is suboptimal and BOECs may be functionally improved by techniques to enhance expression of this critical homeostatic regulator.
Extracellular matrix (ECM) proteins have been shown to negatively regulate eNOS expression and NO production in mature ECs. In addition, the deposition of Col IV and Col I in BOECs is higher compared to that in mature ECs. Thus, I have proposed that the lower eNOS expression/activity in BOECs compared to mature ECs is due to higher ECM deposition. When grown on fibronectin, type I collagen, type IV collagen and laminin, significantly decreased eNOS protein in HUVECs were found compared to cells on polystyrene. Interestingly, when cultured on polystyrene, BOECs express significantly more extracellular matrix (ECM) proteins especially type I collagen compared to mature ECs. Blocking collagen synthesis significantly enhanced eNOS expression in BOECs (1.77 ± 0.41 fold increase). My results suggest that the regulation of eNOS in BOECs and mature ECs is similar and the reduced eNOS level in BOECs may be due to their increased collagen production.
ECM proteins regulate intracellular signaling transduction primarily through integrin signaling associated with focal adhesion complexes. I have proposed that ECM proteins regulation on eNOS signaling in BOECs and mature ECs is through integrin and integrin-associated proteins. Matrix mediated eNOS downregulation was blocked by β1 integrin siRNA and focal adhesion kinase siRNA transfection in both BOECs and HUVECs. In addition, inhibitors of actin polymerization (e.g. ROCK inhibitors and cytochalasin D) block the effect of ECM on eNOS signaling. Taken together, my results suggest that ECM proteins regulate eNOS expression via a β1 integrin/FAK/actin polymerization dependent mechanism.
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Lester, Elizabeth Ann. "Consequences of biomaterial activation of blood cells on endothelial cell proinflammatory phenotype." Thesis, Georgia Institute of Technology, 2001. http://hdl.handle.net/1853/11869.
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Prasad, Raju. "Endothelial progenitor cells, vascular function, and exercise." Access to citation, abstract and download form provided by ProQuest Information and Learning Company; downloadable PDF file, 59 p, 2009. http://proquest.umi.com/pqdweb?did=1654501181&sid=4&Fmt=2&clientId=8331&RQT=309&VName=PQD.
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Zolle, Lapuente Olga C. "Cyclic GMP and calcium homeostasis in endothelial cells." Thesis, University of Liverpool, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.367654.
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Doty, Sherry D. "Fluid shear stress effects on fibronectin in endothelial cells." Thesis, Georgia Institute of Technology, 1989. http://hdl.handle.net/1853/19073.
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Tretiach, Marina Louise. "Bovine Models of Human Retinal Disease: Effect of Perivascular Cells on Retinal Endothelial Cell Permeability." University of Sydney, 2005. http://hdl.handle.net/2123/1153.
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Doctor of Philosophy (Medicine)Background: Diabetic vascular complications affect both the macro- and microvasculature. Microvascular pathology in diabetes may be mediated by biochemical factors that precipitate cellular changes at both the gene and protein levels. In the diabetic retina, vascular pathology is found mainly in microvessels, including the retinal precapillary arterioles, capillaries and venules. Macular oedema secondary to breakdown of the inner blood-retinal barrier is the most common cause of vision impairment in diabetic retinopathy. Müller cells play a critical role in the trophic support of retinal neurons and blood vessels. In chronic diabetes, Müller cells are increasingly unable to maintain their supportive functions and may themselves undergo changes that exacerbate the retinal pathology. The consequences of early diabetic changes in retinal cells are primarily considered in this thesis. Aims: This thesis aims to investigate the effect of perivascular cells (Müller cells, RPE, pericytes) on retinal endothelial cell permeability using an established in vitro model. Methods: Immunohistochemistry, cell morphology and cell growth patterns were used to characterise primary bovine retinal cells (Müller cells, RPE, pericytes and endothelial cells). An in vitro model of the blood-retinal barrier was refined by coculturing retinal endothelial cells with perivascular cells (Müller cells or pericytes) on opposite sides of a permeable Transwell filter. The integrity of the barrier formed by endothelial cells was assessed by transendothelial electrical resistance (TEER) measurements. Functional characteristics of endothelial cells were compared with ultrastructural morphology to determine if different cell types have barrier-enhancing effects on endothelial cell cultures. Once the co-culture model was established, retinal endothelial cells and Müller cells were exposed to different environmental conditions (20% oxygen, normoxia; 1% oxygen, hypoxia) to examine the effect of perivascular cells on endothelial cell permeability under reduced oxygen conditions. Barrier integrity was assessed by TEER measurements and permeability was measured by passive diffusion of radiolabelled tracers from the luminal to the abluminal side of the endothelial cell barrier. A further study investigated the mechanism of laser therapy on re-establishment of retinal endothelial cell barrier integrity. Müller cells and RPE, that comprise the scar formed after laser photocoagulation, and control cells (Müller cells and pericytes, RPE cells and ECV304, an epithelial cell line) were grown in long-term culture and treated with blue-green argon laser. Lasered cells were placed underneath confluent retinal endothelial cells growing on a permeable filter, providing conditioned medium to the basal surface of endothelial cells. The effect of conditioned medium on endothelial cell permeability was determined, as above. Results: Co-cultures of retinal endothelial cells and Müller cells on opposite sides of a permeable filter showed that Müller cells can enhance the integrity of the endothelial cell barrier, most likely through soluble factors. Low basal resistances generated by endothelial cells from different retinal isolations may be the result of erratic growth characteristics (determined by ultrastructural studies) or the selection of vessel fragments without true âbarrier characteristicsâ in the isolation step. When Müller cells were co-cultured in close apposition to endothelial cells under normoxic conditions, the barrier integrity was enhanced and permeability was reduced. Under hypoxic conditions, Müller cells had a detrimental effect on the integrity of the endothelial cell barrier and permeability was increased in closely apposed cells. Conditioned medium from long-term cultured Müller cells and RPE that typically comprise the scar formed after lasering, enhanced TEER and reduced permeability of cultured endothelial cells. Conclusions: These studies confirm that bovine tissues can be used as a suitable model to investigate the role of perivascular cells on the permeability of retinal endothelial cells. The dual effect of Müller cells on the retinal endothelial cell barrier under different environmental conditions, underscores the critical role of Müller cells in regulating the blood-retinal barrier in health and disease. These studies also raise the possibility that soluble factor(s) secreted by Müller cells and RPE subsequent to laser treatment reduce the permeability of retinal vascular endothelium. Future studies to identify these factor(s) may have implications for the clinical treatment of macular oedema secondary to diseases including diabetic retinopathy.
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Ahmed, Syed Rumel. "Characterising lymphocyte trafficking across blood vascular and lymphatic endothelial cells." Thesis, University of Birmingham, 2012. http://etheses.bham.ac.uk//id/eprint/3846/.
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The recruitment of peripheral blood lymphocytes (PBL) to sites of inflammation and their subsequent traffic into the lymphatic circulation is important in host defense. However, surprisingly little is known about their recruitment from the blood vasculature into inflamed tissue, and almost nothing about their egress from inflamed tissue via the lymphatic circulation. We showed that both human macrovascular and microvascular endothelial cells stimulated by TNF\(\alpha\) and IFN\(\gamma\), preferentially recruited memory T-lymphocytes (CD45RO positive cells) from a mixed pool of PBL. T-cells that had migrated across vascular endothelial cells subsequently utilised a combination of \(\beta\)1 and \(\beta\)2 integrins to traverse cytokine activated lymphatic endothelium. In addition we provide evidence that PGD2 was critical for the transmigration of lymphocytes through vascular endothelium. The process of trans-lymphatic migration was also significantly retarded in the presence of a function neutralising antibody against CCR7. Most importantly, we observed that memory T-cells showed a markedly enhanced capacity to migrate across lymphatic endothelium if they had first traversed a vascular endothelial cell barrier. We have shown that addition of exogenous PGD2 to isolated lymphocytes is able to restore the enhanced migration capacity of lymphocytes that have previously migrated through a vascular monolayer. The nature of the priming signal delivered by the process of migration across blood vessel endothelium remains to be fully identified, but is likely to be important in regulating the dynamics of an inflammatory response.
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Helmlinger, Gabriel. "Effects of pulsatile laminar shear stress on cultured vascular endothelial cells." Thesis, Georgia Institute of Technology, 1990. http://hdl.handle.net/1853/16738.
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Seetharaman, Seeta Lakshmy. "Multidrug transporter expression in endothelial cells of the blood-brain barrier." Thesis, University of Cambridge, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.621690.
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Joseph, Laurie B. "Comparison of stressed human endothelial cells derived from different vascular beds /." The Ohio State University, 1986. http://rave.ohiolink.edu/etdc/view?acc_num=osu1487266691093865.
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Jiang, Liying. "Exposure of endothelial cells to shear stress stimulates protein tryosine phosphorylation." Thesis, Georgia Institute of Technology, 1996. http://hdl.handle.net/1853/25421.
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Afzal-Ahmed, Iram. "Redox regulation in circulating blood cells and foetal endothelial cells in normotensive and pre-eclamptic pregnancies." Thesis, King's College London (University of London), 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.497287.
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Magid, Richard. "Engineering molecular reporters to investigate the effects of shear stress upon endothelial cells." Thesis, Georgia Institute of Technology, 2000. http://hdl.handle.net/1853/13754.
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Ensley, Ann Elizabeth. "Functional evaluation of circulating endothelial progenitor cells for vascular tissue engineering." Diss., Available online, Georgia Institute of Technology, 2006, 2006. http://etd.gatech.edu/theses/available/etd-04042006-145611/.
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Thesis (Ph. D.)--Biomedical Engineering, Georgia Institute of Technology, 2006.Vito, Raymond, Committee Member ; Nerem, Robert, Committee Chair ; Eskin, Suzanne, Committee Member ; Hanson, Stephen, Committee Member ; Gibbons, Gary, Committee Member.
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Tan, Kevin S. "Role of Circulating Peripheral Blood-Derived Endothelial Colony-Forming Cells in Patients with Proliferative Diabetic Retinopathy." Case Western Reserve University School of Graduate Studies / OhioLINK, 2009. http://rave.ohiolink.edu/etdc/view?acc_num=case1238452739.
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Mitchell, Andrew Joseph. "Understanding the role of endothelial progenitor cells in vascular injury and repair." Thesis, University of Edinburgh, 2018. http://hdl.handle.net/1842/33310.
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Introduction: Vascular injury is the crucial initiating event in atherosclerosis and is universal following percutaneous coronary intervention. The cellular response to this injury largely determines vessel outcome. Endothelial progenitor cells (EPCs) and their progeny, late outgrowth endothelial cells (EOCs) are thought to play an important role in this process and characterising this role would be valuable in better understanding vascular injury and repair. Methods: The radial artery in the context of transradial cardiac catheterisation was examined as a model of vascular injury with characterisation of structural injury, longitudinal function and EPC populations. To examine the role of late outgrowth endothelial cells a method for GMP-compliant cell culture and labelling with F18Fluorodeoxyglucose was developed with a view to conducting a cell-tracking study of human administration. Results: Radial artery function was reduced following transradial cardiac catheterisation with recovery over a period of three months. There was no correlation between recovery of arterial function and EPC populations as defined by conventional surface markers. A research grade protocol for EOC culture was successfully translated to a GMP-compliant process producing a viable, phenotypically homogeneous EOC product. Cells were successfully labelled with F18Fluorodeoxyglucose and whilst proliferation was reduced, acute viability and function were not compromised. Conclusion: The radial artery in the context of transradial cardiac catheterisation is a useful model of vascular injury and repair although recovery of vascular function does not appear to be influenced by EPC populations. GMP-compliant culture and labelling of EOCs is feasible and will allow examination of the physiology of these cells in vivo in man.
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Robinson, Scott Thomas. "Determining the role of endothelial progenitor cells in post-natal neovascularization." Diss., Georgia Institute of Technology, 2010. http://hdl.handle.net/1853/37178.
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Endothelial Progenitor Cells (EPCs) were first identified from human blood samples as a population of circulating mononuclear cells capable of displaying a mature endothelial cell phenotype in culture. Subsequent studies have established that EPCs arise from the bone marrow (BM) and incorporate into the endothelium at sites of blood vessel growth, suggesting a potential role for these cells in neovascularization. Furthermore, a decline in EPC count has been correlated to multiple vascular pathologies, indicating that EPC number could serve as a biomarker of cardiovascular disease. Unfortunately, due to the variability in techniques used for EPC isolation and identification, considerable heterogeneity exists within the population of cells commonly defined as EPCs. In order for the clinical potential of EPCs to be fully realized, thorough characterization of the BM-derived cell populations involved in neovascularization is required.
The objective of our study was to determine the functional significance of circulating EPCs in postnatal vascular growth and repair. Two separate strategies were employed to achieve this objective. In the first, we attempted to generate a novel mouse model where the pool of bone marrow-derived endothelial precursors was drastically reduced or eliminated. Our overall approach was to deliver a "suicide" gene, under control of an endothelial cell-specific promoter, to bone marrow cells for use in bone marrow transplantation (BMT) experiments. Mice receiving BMTs would therefore lack the ability to deliver viable BM-derived EPCs to sites of neovascularization. Our central hypothesis for this study was that a reduction in EPC viability would hinder endogenous vascular repair mechanisms, thereby exacerbating cardiovascular disease. In the second strategy, we attempted to identify novel progenitor cell populations based on the transcriptional regulation of pro-angiogenic genes. Our overall approach was to transduce BM with a retrovirus containing a fluorescent reporter gene under control of pro-angiogenic promoters for use in transplantation experiments. Our central hypothesis for this study was that unique populations of BM-derived cells could be identified by expression of the fluorescent reporter gene directed by the Vascular Endothelial Growth Factor (VEGF), endothelial Nitric Oxide Synthase (eNOS) and Vascular Endothelial (VE) Cadherin promoters.
The BMT strategy utilized to address our first hypothesis was unsuccessful due to the use of a truncated form of the pro-apoptotic Bax as our suicide gene target. A plasmid encoding GFP fused to the truncated Bax fragment (ΔN-Bax, consisting of amino acids 112-192 of the full length protein) was used in transfection experiments to assess ΔN-Bax function. The GFP:ΔN-Bax fusion protein formed distinct extranuclear aggregates (presumably due to mitochondrial translocation) but did not induce apoptosis in transfected cells. The ΔN-Bax fragment also did not induce cell death when targeted to endothelial cells with retoviral-mediated gene delivery or in a transgenic mouse setting.
To address our second hypothesis, we generated retroviral vectors containing the fluorescent tdTomato reporter under control of the VEGF, eNOS and VE Cadherin promoters. Significant fluorescence was detected in cultured endothelial cells and ex vivo-expanded BM cells. Following transplantation of transduced BM cells into lethally irradiated recipient mice, we were able to identify circulating populations of tdTomato-positive cells using flow cytometry. With these results we have identified novel subpopulations of circulating BM-derived cells which may play a significant role in post-natal neovascularization in mice. Therefore, results acquired from these studies could lead to improved cell therapy techniques for treatment of vascular disease.
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Lafleur, Marc Andre. "The involvement of matrix metalloproteinases in angiogenesis." Thesis, University of East Anglia, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.327541.
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Krenning, Guido. "Endothelial progenitor cells in vascular regenerative medicine towards 'designer blood vessels' and 'therapeutic neovascularization' /." [S.l. : [Groningen : s.n.] ; University of Groningen] [Host], 2009. http://irs.ub.rug.nl/ppn/.
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Helmlinger, Gabriel. "Effect of pulsatile flow on the intracellular free calcium concentration of cultured vascular endothelial cells." Diss., Georgia Institute of Technology, 1994. http://hdl.handle.net/1853/16707.
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Rajani, Rikesh Mukesh. "Is small vessel disease a disease of the blood brain barrier?" Thesis, University of Edinburgh, 2016. http://hdl.handle.net/1842/25866.
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Cerebral small vessel disease (SVD) is a vascular neurodegenerative disease which is the leading cause of vascular dementia and causes 20% of strokes. 20-30% of those over 80 show signs of the disease as white matter hyperintensities on MRI scans, doubling their risk of stroke and trebling their risk of dementia. Sporadic SVD is thought to be caused by hypertension but 30% of sufferers are normotensive and an alternative hypothesis implicates loss of integrity of the blood brain barrier (BBB). To investigate this, I studied brains from normotensive people with early stage SVD and found reduced capillary endothelial claudin-5 (a BBB tight junction protein), more oligodendrocyte precursor cells (OPCs; the precursors to myelinating oligodendrocytes), and more microglia/macrophages compared to controls. Furthermore, in a relevant rat model of spontaneous SVD, the Stroke Prone Spontaneously Hypertensive Rat (SHRSP; disease model; DM) I found that reduced endothelial claudin-5 was the earliest change, appearing at 3 weeks of age, followed by OPC proliferation, appearing at 4 weeks, and then increased number of microglia/macrophages, appearing at 5 weeks. Importantly, all these changes occurred at a young age (< 5 weeks), before any measurable hypertension. These changes were confirmed in an ex vivo slice culture model (i.e. removing blood flow), ruling out direct damage by leakage of blood components through an impaired BBB and suggesting an inherent endothelial cell dysfunction as the primary cause, with secondary BBB defects. This hypothesis of endothelial dysfunction is supported by increased endothelial cell proliferation in both human SVD tissue and the DM rats, and lower levels of endothelial nitric oxide synthase (eNOS) in brains of DM rats. To study this further I isolated primary brain microvascular endothelial cells (BMECs) from DM and control rats and found that those from DM rats formed less mature tight junctions (less membranous claudin-5) than control BMECs. I also found that conditioned media (CM) from DM BMECs causes OPCs in culture to proliferate more and mature less. This indicates that the endothelial dysfunction is inherent to the endothelial cells, rather than induced by other cell types, and through secreted factors causes OPC changes mirroring what is seen in vivo. Using an antibody array, I identified HSP90α as a candidate secreted factor and showed that it is necessary (by blocking the protein in CM) and sufficient (by adding recombinant HSP90α) to induce the maturation phenotype in OPCs, but not the proliferation phenotype. The idea that endothelial dysfunction causes SVD begs the question of what causes endothelial dysfunction, especially in our inbred DM rat strain. To establish this, I reanalysed sequencing data of the DM and control rats from a previously published study, searching for mutations which lead to truncated proteins in genes expressed in brain endothelial cells. We confirmed the candidate gene Atp11b, a phospholipid flippase, was mutated as predicted. I found that knocking down Atp11b using siRNA in a control endothelial cell line caused endothelial dysfunction and a loss of tight junction maturity, and that CM from these cells causes OPCs to proliferate more and mature less, mirroring what we see in primary DM BMECs and suggesting that Atp11b has a key function in promoting normal endothelial function. Furthermore, I showed that knocking down Atp11b causes cells to secrete increased levels of HSP90α. I propose a mechanism whereby ATP11B regulates the retention of HSP90α within endothelial cells, which in turns regulates eNOS levels and activity, as has been shown previously. In summary, this work shows that there are many pre-symptomatic changes which occur in the brain in the development of SVD in DM rats, and that these are ultimately caused by endothelial dysfunction. As these changes are similar to those found in spontaneous human SVD, I propose that endothelial dysfunction is a key mechanism of human SVD, which may in the future lead to new therapies.
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Jenkins, Meredith E. "An examination of the human fibrinogen-like protein2 sequence variations and genetic expression by human endothelial cells /." unrestricted, 2005. http://etd.gsu.edu/theses/available/etd-07222005-093438/.
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Thesis (M.S.)--Georgia State University, 2005.Title from title screen. Roberta Attanasio, committee chair; P.C. Tai, W.C. Hooper, committee members. Electronic text (57 p. : col. ill.) : digital, PDF file. Description based on contents viewed Aug. 15, 2007. Includes bibliographical references (p. 55-57).
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Thomas, Kelly Angelique. "The effects of Ethanol and Aspalathus linearis on immortalized mouse brain endothelial cells (bEnd5)." Thesis, University of the Western Cape, 2015. http://hdl.handle.net/11394/4959.
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Magister Scientiae (Medical Bioscience) - MSc(MBS)The blood brain barrier (BBB) is a signaling interface between the blood and the central nervous system (CNS), which prohibits the entry of harmful blood-borne substances into the brain micro-environment, thus maintaining brain homeostasis. The crucial role of the BBB is protecting the CNS, which may adversely be affected by alcohol. The central component of the BBB, endothelial cells (ECs), regulates BBB transport by regulating the permeability both transcellularly and through their paracellular junctions, by structures called tight junctions (TJs) that are composed of proteins. The aim of this study was to investigate the in vitro effects of ethanol (EtOH) and fermented rooibos (Rf) on a monolayer of bEnd5 mouse brain ECs, by determining the effects of EtOH and Rf on bEnd5 (i) cell viability (ii) cell proliferation (iii) rate of cell division (iv) cell toxicity (v) claudin-5 transcription (vi) permeability across a monolayer of bEnd5 ECs and (vii) morphology, for a selected experimental timeline of 24, 48, 72, and 96hrs. We then investigated if the simultaneous exposure of Rf and EtOH could reverse or alleviate the EtOHinduced effects on the bEnd5 ECs. EtOH metabolism induces oxidative stress and results in a range of adverse physiological effects. Aspathalus linearis (rooibos) contains many phenolic compounds, of which the main antioxidant activity is attributed to aspalathin. Our underlining hypothesis is that the antioxidants in an aqueous rooibos extract may therefore protect against the potential oxidant damaging effects of alcohol on the BBB. Cells were exposed for 24hrs to selected concentrations of EtOH (25mM and 100mM), a concentration of Rf containing equivalent of 1.9nM aspalathin, and the combinations of EtOH and Rf. Cell viability and cell toxicity was determined, while cell proliferation and rate of cell division was estimated using the trypan blue exclusion assay. Real time quantitative PCR was implemented to quantify claudin-5 transcription, normalized against housekeeping genes, GAPDH and HPRT. Transepithelial electrical resistance (TEER) was measured using the Ohm Millicell-electrical resistance system, while bEnd5 monolayer morphology was analysed using the Zeiss scanning electron microscope. Both concentrations of EtOH led to an overall decrease in cell viability, and a decreased number of live cells across 72hrs. Consistent with this, EtOH resulted in increased cell toxicity across the 96hr experimental timeframe and a diminished rate of cell division. The transcription of claudin-5 in bEnd5 ECs exposed to 25mM and 100mM EtOH varied dramatically across the 96hr timeframe. While 25mM EtOH resulted in an overall decrease in TEER, cells exposed to 100mM EtOH only decreased TEER between 48 and 96hrs. Morphologically, both concentrations of EtOH led to compromised paracellular spaces as endorsed by high definition SEM analysis. The administration of Rf on its own resulted in an initial decrease in viability, followed by recovery between 72 and 96hrs. Exposure to Rf diminished live cell numbers at 72 and 96hrs, accompanied by a compromised rate of cell division and an overall increase in cell toxicity. In addition, Rf down-regulated claudin-5 transcription across the course of the experiment, particularly between 24 and 48hrs. In alignment with this, Rf also led to an increase in BBB permeability from 24 to 96hrs. However, SEM studies were not able to discriminate any differences between control and Rf treated cells. Our study showed that the BBB could be protected against the adverse effects of EtOH, and this at the plasma concentration induced by 500ml’s of Rooibos tea. The simultaneous exposure of Rf and EtOH was able to negate the effects of EtOH on cell viability, cell proliferation, and cell toxicity but exacerbated the effects of EtOH on claudin-5 transcription and paracellular permeability. Morphologically, co-exposure with Rf only reversed the effects of 25mM EtOH while exacerbating the effects of 100mM EtOH at 96hrs. In conclusion, EtOH was shown to be detrimental to the integrity of bEnd5 ECs, and the addition of a minuscule quantity of t h e Rf extract was able to partially alleviate excess ROS-induced effects.
National Research Foundation (NRF)
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Plant, Stuart D. "The response of human umbilical vein endothelial cells and blood platelets to modified NiTi surfaces." Thesis, University of Nottingham, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.275630.
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Cardinal, Kristen O'Halloran. "Development and Utilization of a Tissue Engineered Blood Vessel Mimic to Assess the Neointimal Response to Intravascular Stents." Diss., The University of Arizona, 2007. http://hdl.handle.net/10150/195387.
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The use of intravascular stents to restore blood flow through restricted vessels in patients with coronary artery disease has become the preferred method for treating a variety of lesion locations and pathologies. As new stent configurations and coatings are developed, a great need exists for high-throughput preclinical evaluation techniques that can interface human tissue with three-dimensional devices. Thus, the goals of this dissertation research were 1) to develop an in vitro blood vessel mimic composed of human cells for preclinical evaluation of intravascular devices, and 2) to utilize the mimic to assess neointimal responses to implanted stents.Experiments in support of these goals were broken into four specific aims. The first aim was to develop an in vitro human blood vessel mimic based on techniques for creating tissue engineered vascular grafts. The second aim was to determine the feasibility of utilizing this vessel mimic for bare metal stent evaluation. The third aim was to use the in vitro vessel to evaluate the cellular response to protein-coated stents. The fourth aim was to take advantage of the ability to control the in vitro vessel environment in order to evaluate the effect of shear rate on the neointimal response to implanted stents.Human blood vessel mimics were created by sodding fat-derived microvascular endothelial cells onto expanded polytetrafluoroethylene grafts and cultivating the vessels in bioreactor systems. This resulted in the development of a luminal lining of endothelial cells with sub-endothelial smooth muscle and mesenchymal cells. Deployment and assessment of bare metal stents within blood vessel mimics supported the feasibility of using the model for stent evaluation, and demonstrated that cell coverage of the device surface could be observed and measured. Protein-modified stents were created by submerging devices in enriched medium, and following implantation in the blood vessel mimic exhibited increased cell coverage and increased tissue thickness as compared with bare metal stents. Finally, an increase in shear rate lead to decreased neointimal coverage of implanted bare metal and modified stents. Overall, this dissertation demonstrates that in vitro human blood vessel mimics can be created and utilized for preclinical device evaluation.
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Ziegler, Thierry. "Co-culture of endothelial cells with smooth muscle cells in a matrix of collagen : Effect of a steady, laminar stress on the cell behavior." Diss., Georgia Institute of Technology, 1994. http://hdl.handle.net/1853/17618.
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Arief, Melissa Suen. "Human Tissue Engineered Small Diameter Blood Vessels." Yale University, 2010. http://ymtdl.med.yale.edu/theses/available/etd-03152010-144428/.
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The engineering of human vascular grafts is an intense area of study since there is crucial need for alternatives to native vein or artery for vascular surgery. This current study sought to prove that a tissue engineered blood vessel (TEBV) 1mm in diameter could be developed from human smooth muscle cells and that endothelial progenitor cells (EPCs) could be cultured and used to endothelialize these grafts. This project had four specific aims: the isolation and characterization of EPCs, the seeding of a novel scaffold with EPCs and exposure to physiologic shear stress in vitro, the development of TEBV from human smooth muscle cells that are strong enough to implant in vivo, and the in vivo implantation of TEBV into the rat aortic model with a comparison of EPC seeded TEBVs pretreated with shear stress and unseeded TEBVs. The results yielded isolation of four EPC lines and a flow system design capable of seeding EPCs onto a novel scaffold with preliminary studies indicating that it is capable of exposing the EPCs to physiologic shear stress, although further studies require more optimization. The development of mechanically strong TEBV was highly successful, yielding TEBVs comparable to native vessels in collagen density and burst pressure, but with much lower compliance. Current implantation studies indicated that unseeded TEBV grafts implanted into the rat aorta without anticoagulation is highly thrombogenic. However, anticoagulation using Plavix may be capable of maintaining graft patency. These TEBVs did not rupture or form aneurysm in vivo and the future completion of the in vivo studies are likely to demonstrate the high potential of these grafts.
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Braddon, Linda Greer. "The effect of shear stress on a co-culture of endothelial cells and fibroblasts in a biodegradable polymer scaffold." Diss., Georgia Institute of Technology, 1997. http://hdl.handle.net/1853/17850.
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Suidan, Georgette Leila. "CD8 T cells mediate CNS vascular permeability under neuroinflammatory conditions." University of Cincinnati / OhioLINK, 2009. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1242912012.
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Johnson, Tiffany Lynn. "Endothelial Cell Function Using a Tissue Engineered Blood Vessel Model: A Case Study of Cell-Cell Communication." Diss., Available online, Georgia Institute of Technology, 2006, 2006. http://etd.gatech.edu/theses/available/etd-04032006-120602/.
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Thesis (Ph. D.)--Biomedical Engineering, Georgia Institute of Technology, 2006.Pollman, Matthew, Committee Member ; Galis, Zorina, Committee Member ; McIntire, Larry, Committee Member ; Taylor, W Robert, Committee Member ; Jo, Hanjoong, Committee Member ; Nerem, Robert, Committee Chair.
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Okyere, Benjamin. "Eph-mediated restriction of cerebrovascular arteriogenesis." Diss., Virginia Tech, 2019. http://hdl.handle.net/10919/89222.
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Stroke is a leading cause of morbidity and long-term neurological disability in the U.S. Ischemic stroke, which accounts for approximately 90% of all strokes, is the result of an occlusion in the arteriole cerebrovascular network. No effective treatment options exist to provide neuroprotection from occlusion, and limited success has been seen clinically when attempting to restore blood flow to vulnerable neural tissue regions. Enhancement of pial collateral remodeling (Arteriogenesis) has recently been shown to improve blood flow and mitigate neural tissue damage following stroke (1-3). Arteriogenesis is the remodeling of pre-existing arteriole vessel which are able to re-route blood to blood-deprived regions of tissue. Arteriogenesis requires endothelial cell (EC) and smooth muscle cell proliferation, extracellular matrix degradation and recruitment of circulating bone marrow-derived cells (4-6). Unlike spouting angiogenesis, which requires weeks following occlusion to develop, arteriogenesis begins as early as 24-48hrs post-stroke (7, 8) and can expeditiously enhance blood flow to ischemic regions, making it an attractive target for therapeutic intervention. Our preliminary studies, in an EphA4 global knockout mouse model, indicated that EphA4 receptor tyrosine kinase severely limits pial arteriole collateral formation. The preliminary work also showed that activation of EC EphA4 receptor in vitro inhibited vascular formation. Additionally, ECs lining the collateral vessel have been shown to play a role in collateral remodeling (9). Taken together, the objective of this dissertation was to elucidate the cell autonomous role of the EphA4 receptor and given the central role of the EC in collateral remodeling, we postulated that EphA4 receptor on ECs the limits pial collateral formations. Using a cell-specific loss-of-function approach, we tested the hypothesis that EC-specific EphA4 plays an important role in pial collateral development and remodeling after induced stroke. The results from this dissertation show that (1) EphA4 expression on ECs suppress the formation of pial collaterals during development and limits EC growth via suppression of p-Akt in vitro (2) EC-specific EphA4 ablation leads to increased collateral remodeling, enhanced blood flow recovery, tissue protection and improved neurological behavioral outcomes after stroke and (3) Mechanistically, EphA4 limits pial collateral remodeling via attenuation of the Tie2/Angiopoietin-2 signaling pathway. The work presented in this dissertation demonstrate that EphA4 can be targeted therapeutically to increase pial collateral remodeling to alleviate neurological deficits after ischemic stroke.Doctor of Philosophy
Stroke is the fifth leading cause of death in the United States. Ischemic stroke is the most common type of stroke and occurs when blood flow to part of the brain is impeded. Lack of blood results in cell death and tissue damage in the brain. In an effort to restore blood flow, specialized blood vessels in the brain called collaterals remodel and become larger to allow re-routed blood to the blood-deprived region of the brain. The duration it takes to remodel these remarkable blood vessels and re-route blood varies in humans, and sometimes is not able to prevent adequate tissue damage. The current work explores novel therapeutic targets to accelerate collateral remodeling in an effort to reduce tissue loss after stroke. We present studies which show that a protein called EphA4, found on endothelial cells restricts remodeling, and when inhibited in the brain can increase collateral remodeling and reduced adverse effects after ischemic stroke.
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Lacroix-Desmazes, Sebastien. "Influence of flow environment on the production and secretion of metalloproteinases and urokinase-type plasminogen activator by cultured bovine aortic endothelial cells." Thesis, Georgia Institute of Technology, 1993. http://hdl.handle.net/1853/15826.
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Sun, Bing. "Characterization of CRGRP and 5-HT receptors in vascular tissues, and expression of CGRP in cultured human endothelial cells." Thesis, University of Cambridge, 1992. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.260599.
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Nguyen, Hieu Thi Minh. "The effect of cardiolipin synthase deficiency on the mitochondrial function and barrier properties of human cerebral capillary endothelial cells." Elsevier, 2014. http://hdl.handle.net/1993/30184.
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The blood brain barrier (BBB), formed by endothelial cells lining the lumen of the brain capillaries, is a restrictively permeable interface that only allows transport of specific compounds into the brain. Cardiolipin (CL) is a mitochondrial- specific phospholipid known to be required for the activity and integrity of the respiratory chain. The current study examined the role of cardiolipin in maintaining an optimal mitochondrial function that may be necessary to support the barrier properties of the brain microvessel endothelial cells (BMECs). Endothelial cells have been suggested to obtain most of their energy through an-aerobic glycolysis based on studies of cells that were obtained from the peripheral vasculatures. However, here, we showed that the adult human brain capillary endothelial cell line (hCMEC/D3) appeared to produce ~60% of their basal ATP requirement through mitochondrial oxidative phosphorylation. In addition, RNAi mediated knockdown of the CL biosynthetic enzyme cardiolipin synthase (CLS), although did not grossly affect the mitochondrial coupling efficiency of the hCMEC/D3 cells, did seem to reduce their ability to increase their mitochondrial function under conditions of increased demand. Furthermore, the knockdown appeared to have acted as a metabolic switch causing the hCMEC/D3 cells to become more dependent on glycolysis. These cells also showed increase in [3H]-2-deoxyglucose uptake under a low glucose availability condition, which might have served as a mechanism to compensate for their reduced energy production efficiency. Interestingly, the increase in glucose uptake appeared correlated to an increase in [3H]-2-deoxyglucose glucose transport across the knockdown confluent hCMEC/D3 monolayers grown on Transwell® plates, which was used in our study as an in vitro model for the human BBB. This suggests that changes in the brain endothelial energy status may play a role in regulating glucose transport across the BBB. These observations, perhaps, also explain why the brain capillary endothelial cells were previously observed to possess higher mitochondrial content than those coming from non-BBB regions (Oldendorf et al. 1977).
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Kojima, Hidenobu. "Establishment of practical recellularized liver graft for blood perfusion using primary rat hepatocytes and liver sinusoidal endothelial cells." Kyoto University, 2018. http://hdl.handle.net/2433/233836.
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Arzhangi, Zahra S. "Collagen biomaterials for the delivery of FVIII-producing blood outgrowth endothelial cells in the treatment of Haemophilia A." Thesis, University of Ottawa (Canada), 2011. http://hdl.handle.net/10393/28921.
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In this thesis, we describe the fundamental aspects to the development of molecularly-defined tailor-made scaffolds for the delivery of FVIII-expressing endothelial cells in the treatment of Haemophilia A. In particular, scaffolds prepared by chemical crosslinking of type I collagen, and growth factor incorporation is discussed. The general strategy was to prepare tailor-made biomaterials as a specific microenvironment to enable cells to produce FVIII and secrete this coagulation factor into the blood stream. As an initial step, pure materials of known concentrations were combined to develop two forms of collagen scaffolds: an injectable hydrogel that may be formed in situ in the presence of cells and a capsule into which the cell-housing hydrogel can be injected. Next, scaffolds were crosslinked using natural- genipin- and synthetic- carbodiimide (EDC)- chemicals. Crosslinking resulted in collagenase-resistant scaffolds. The tissue response to scaffolds was evaluated following subcutaneous implantations in mice. Crosslinked scaffolds maintained their integrity and supported the formation of new extracellular matrix and neovascularization during tissue remodeling. Collagen scaffolds loaded with fibroblast growth factor 2 significantly enhanced FVIII production during long-term encapsulation. Potential applications of these scaffolds for ischemic models are discussed in more detail.
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Biernacki, Katarzyna. "Blood-brain barrier endothelial cells as regulators of immune-mediated injury and repair in the central nervous system." Thesis, McGill University, 2004. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=85129.
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The blood-brain barrier (BBB) is a specialized structure that restricts the passage of blood-borne cells and molecules into the central nervous system (CNS). BBB dysfunction and extensive perivascular leukocytic infiltration are critical events for lesion formation in multiple sclerosis (MS).To examine the mechanisms that regulate the migration of immune cells that participate in CNS injury and/or repair, I have used human adult brain-derived endothelial cells (HBECs) grown in Boyden chambers. I compared the migration of Th1 lymphocytes, disease-inducing cells, and Th2 lymphocytes, cells induced by several immuno-modulatory therapies aimed at reducing neuroinflammation, across an in vitro model of the BBB. I demonstrate that Th2 cells can efficiently cross the BBB, and do so at significantly higher rates compared to Th1 lymphocytes. This selectivity is dependent on HBEC chemokine CCL2, and the selective expression of the receptor CCR2 on Th2 cells. Adhesion molecules are not implicated in selective T cell recruitment, although migration of both subsets can be blocked with anti-ICAM-1 antibody. I also explored the consequences of T cell:HBEC interactions on HBEC properties. I show that supernatants derived from Th1 polarized cells significantly up-regulate a range of pro-inflammatory chemokines and adhesion molecules in HBECs. Conversely, Th2 supernatants had no such effects and did not suppress the effects induced by Thl lymphocytes. Together, this suggests that Th2 cell-mediated bystander suppression does not occur at the level of the BBB. I also show that Th1 and Th2 cell migration can enhance subsequent passage of cells and molecules across HBEC barriers. Furthermore, I demonstrate that lymphocyte contact induced HBEC production of the neurotrophin NGF but not BDNF, an effect potentiated by IFNbeta. The secretion of NGF was specific to HBECs; neither astrocytes nor microglia could be induced to secrete this neurotrophin. I also demonstrate an inverse correlation between HBEC NGF production induced by MS patient lymphocytes and measures of clinical disability and brain atrophy suggesting that HBECs may have a neuroprotective role in MS. Collectively, my data indicate that the BBB is a dynamic structure involved in the regula
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Alamu, Olufemi Akinyinka. "Differential toxicity of two murine endothelial cells to ROS duress: Understanding oxidative stress-induced blood-brain barrier dysfunction." University of the Western Cape, 2020. http://hdl.handle.net/11394/7876.
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Philosophiae Doctor - PhDThe blood-brain barrier (BBB) is a critical interface between the blood circulation and brain tissue which performs critical selection of circulating molecules that gain access to the brain tissue. Its unique ability to adjust to changes in the constituents of the blood circulation confer in the BBB a dynamic nature enabling changes in its properties to suit the homeostatic needs of the brain. Dysfunction of the BBB has been established to be pivotal to the initiation and/or maintenance of an array of neurological disorders, most of which involve the production of excess reactive oxygen species (ROS) and oxidative stress in their pathophysiology. Thus, clinical trials of exogenous antioxidant agents have been proposed and initiated, with most results being inconclusive. Extensive studies of the impact, capacity and plasticity of endogenous antioxidants in the cells that constitute the blood-brain barrier, especially the brain endothelial cells, therefore, became necessary for the rational choice, timing, and the mode of application of antioxidants in the management of oxidative stress-mediated neurological diseases.
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Al-Waili, Daniah I. "In Vitro Functional Study of YES-Associated Protein (YAP) in Murine Brain Endothelial Cells under Normal and Ischemic Conditions." University of Cincinnati / OhioLINK, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1448037357.
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Hedigan, Conor Charles. "Evaluation of Human Umbilical Vein Endothelial Cells in Blood Vessel Mimics Through Changes in Gene Expression and Caspase Activity." DigitalCommons@CalPoly, 2019. https://digitalcommons.calpoly.edu/theses/2081.
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Blood vessel mimics (BVMs) are simple tissue engineered blood vessel constructs intended for preclinical testing of vascular devices. This thesis developed and implemented methods to characterize two of these components. The first aim of this thesis investigated the effect of cell culture duration and flow conditions on endothelial cell gene expression, especially regarding endothelial-to-mesenchymal transition (EndMT). A trend of decreased endothelial marker gene expression and increased mesenchymal marker gene expression would indicate EndMT. qPCR analysis revealed that increased cell culture duration did not result in EndMT, and in fact increased endothelial marker expression as cell culture duration increased. Disturbed flow conditions decreased endothelial marker and increased mesenchymal marker expression relative to static culture.
The second aim of this thesis developed methods to determine cytotoxicity of, and endothelial cell adhesion to, novel BTEAC salt scaffolds. Immunostaining was used to visualize these scaffold effects. The cytotoxicity elution assay showed that BTEAC salt scaffolds were not more cytotoxic than the standard PLGA scaffold. Direct contact assays spanning several timepoints also found that BTEAC salt scaffolds were not more cytotoxic than standard scaffolds but had higher endothelial cell adhesion and coverage than standard scaffolds. Overall, this thesis developed and implemented methods to characterize the endothelial cells used in the BVM model.
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Chen, Lin Min. "Angiogenic activities of Drynaria fortunei-derived extract and isolated compounds on zebrafish in vivo and human umbilical vein endothelial cells in vitro." Thesis, University of Macau, 2017. http://umaclib3.umac.mo/record=b3690926.
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Mehra, Anupriya. "NMDA receptor of the blood brain barrier : mechanism of action and interaction with tPA." Thesis, Normandie, 2017. http://www.theses.fr/2017NORMC404/document.
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La neuroinflammation est un dénominateur commun de plusieurs troubles du système nerveux central. Les réactions inflammatoires sont souvent médiées par plusieurs voies de signalisation qui conduisent à l'ouverture de la barrière hémato-encéphalique. L'activateur tissulaire du plasminogène (tPA) est une serine protéase qui induit l'ouverture de la barrière hémato-encéphalique. Au cours des dernières années, il a également été montré que les récepteurs NMDA situés dans les cellules endothéliales peuvent jouer un rôle crucial dans la propagation de la réaction inflammatoire.Mon travail au cours de ma thèse a mis l'accent sur la découverte des mécanismes par lesquels le récepteur NMDA effectue une médiation de l'ouverture de la barrière hémato-encéphalique induite par le TPA. Dans notre première étude, nous montrons que les récepteurs NMDA endothéliaux sont des cibles thérapeutiques potentielles pour prévenir l'infiltration et l'inflammation des cellules immunitaires médiées par l'EAE. Nous montrons que l'anticorps monoclonal du récepteur NMDA spécifique à la souris, le Glunomab, pourrait protéger la barrière de la moelle épinière de dommages inflammatoires. Nous montrons également que les récepteurs NMDA sont exprimés en étroite association avec les protéines de jonction serrées dans les cellules endothéliales cérébrales. Dans notre deuxième étude, nous montrons pour la première fois que les récepteurs NMDA neuroendothéliaux peuvent présenter une action métabotropique lors de l'inflammation. Nous soulignons également que ces récepteurs sont en effet des récepteurs NMDA non conventionnels exprimant la sous unité GluN3A. En outre, nous rapportons que le tPA accélère l'ouverture de la barrière hémato-encéphalique en présence d'une agoniste rare de la glycine par un mécanisme dépendant de l'activation de RhoA. Les résultats de mon projet apportent une nouvelle vision du rôle des récepteurs NMDA métabotropiques dans les cellules endothéliales cérébrales. En outre, il fournit également des détails plus précis sur l'ouverture de la barrière hémato-encéphalique via l’activateur tissulaire du plasminogèneNeuroinflammation is a common denominator of several central nervous system disorders. Inflammatory reactions are often mediated by several signaling pathways which lead to the opening of the blood brain barrier. Tissue plasminogen activator (tPA) is a serine protease induces opening of the blood brain barrier. In recent years, it has also been shown that NMDA receptors located in endothelial cells can play a crucial role in propagation of inflammatory reaction. My doctoral study focused on the finding the underlying mechanisms of action(s) by which NMDA receptor mediates tPA induced opening of the blood brain barrier. In our first study we show that endothelial NMDA receptors are potential therapeutic targets to prevent EAE mediated immune cell infiltration and inflammation. We show that NMDA receptor specific mouse monoclonal antibody Glunomab could prevent the brain spinal cord barrier from inflammatory damage. We also show that NMDA receptors are expressed in close association of tight junction proteins in cerebral endothelial cells. In our second study, we show for the first time that, neuroendothelial NMDA receptors can exhibit metabotropic mode of action during inflammation. We also highlight that these receptors are indeed GluN3A expressing non-conventional NMDA receptors. In addition, we report that tPA accelerates the opening of blood brain barrier in presence of an uncommon agonist glycine by RhoA activation dependent mechanism.My project results provide a nouvelle insight for the role of metabotropic NMDA receptors in cerebral endothelial cells. In addition it also provides more precise details of blood brain barrier opening mediated by tissue plasminogen activator
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Punshon, G. "Development of an improved two-stage seeding process for a nanocomposite vascular graft using human peripheral-blood derived endothelial cells." Thesis, University College London (University of London), 2017. http://discovery.ucl.ac.uk/1560130/.
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Background: Currently available prosthetic small calibre vascular grafts have poor medium and long term outcomes due to the development of neo-intimal hyperplasia caused by their non-compliant properties and lack of an endothelial cell lining. Here a nanocomposite based on polyhedral oligomeric silsesquioxane (POSS) attached by direct reaction onto a urethane segment was employed as a potential vascular graft material. It has been demonstrated to have similar viscoelastic properties to a native artery and be resistant to degradation. Initially Human Umbilical Vein Endothelial Cells (HUVEC) were employed as the cell source of choice after which investigations were carried out into the suitability of human peripheral blood derived circulating endothelial cells (CEC) and endothelial progenitor cells (EPC) for graft seeding. Aim: The aim of this study was to develop a system with the potential to deliver an EPC/CEC-seeded bypass graft in a realistic time-frame. Methods: The cytocompatibility of the nanocomposite was initially investigated using HUVEC as a cell source. Surface modification of the nanocomposite to improve cell adhesion and proliferation was then attempted using UV exposure. Seeded nanocomposite grafts were then exposed to flow and the effect of preconditioning investigated. Following this the use of human peripheral blood derived EPC and CEC as a potential cell seeding source was investigated. Studies were also carried out into the sterilization of the nanocomposite. Results: The nanocomposite was able to support the attachment and growth of both HUVEC and EPC/CEC. Nanocomposite grafts were successfully seeded and exposed to flow with cells being retained on the graft surface following exposure to flow. Conclusions: The results obtained suggest that the nanocomposite graft and the use of EPC/CEC derived from human peripheral blood process has potential both for a realistic and achievable two-stage seeding process for vascular bypass grafts.
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Mohd, Nasir Mohd Hamzah. "Activation of endothelial cells and its potential involvement in blood-brain barrier damage in cerebral malaria : an in vitro study." Thesis, Keele University, 2015. http://eprints.keele.ac.uk/3252/.
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One of the severe complications of a Plasmodium falciparum infection is cerebral malaria (CM). CM is characterised by the accumulation of mature infected red blood cells (RBC) in the brain microvasculature. One of the consistent detrimental effects of sequestration is the breakdown of the blood-brain barrier (BBB), often with a fatal outcome in children in endemic areas. This study investigates the mechanisms underlying BBB breakdown secondary to sequestration, using immortalised human brain microvascular endothelial cells (tHBEC) as an in-vitro model of BBB and ITG-strain Plasmodium falciparum. First, the tHBEC monolayer was co-cultured with Plasmodium falciparum infected red blood cell (PRBC) or uninfected red blood cells (uRBC) control for 20 hours and the supernatant was recovered for subsequent analysis. The co-culture supernatants showed upregulation of inflammatory mediators (MCP-1 and IL-8) and a member of metalloproteases (ADAMTS-1, ADAMTS-4, MMP-2 and MMP-9) in the PRBC-tHBEC co-culture supernatants. The PRBC-tHBEC co-culture supernatants induced loss of endothelial cell monolayer integrity, represented by real time reduction in the transendothelial electrical resistance, measured using Electrical Cell-Substrate Impedance Sensing (ECIS™). The same supernatants also increased the permeability of tHBEC monolayer to the fluorescently labelled 40 kDa dextran showing leakage across the tHBEC monolayer. Interestingly, the loss of barrier function of tHBEC monolayer is partially inhibited by the addition of protease inhibitors GM6001 and rhTIMP-3. Prolonged exposure to PRBC-tHBEC co-culture supernatants reduced the level of vinculin. This study demonstrates that the interactions between PRBC and tHBEC induces activation of tHBEC and the release of proteases that contribute to BBB breakdown in CM, and could be a potential drug target for adjunct therapy in CM.
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Rennel, Emma. "Molecular Mechanisms in Endothelial Cell Differentiation." Doctoral thesis, Uppsala : Acta Universitatis Upsaliensis : Univ.-bibl. [distributör], 2004. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-4059.
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Chen, Miao. "Endothelial Cell-Specific Knockout of Meis1 Protects Ischemic Hindlimb Through Vascular Remodeling." Diss., Virginia Tech, 2018. http://hdl.handle.net/10919/96188.
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Peripheral artery disease (PAD) affects more than 200 million people worldwide. PAD refers to illness due to a reduction or complete occlusion of blood flow in the artery, especially to the extremities in disease conditions, such as atherosclerosis or diabetes. Critical limb ischemia (CLI) is a severe form of PAD associated with high morbidity and mortality. Currently, no effective and permanent treatments are available for this disease. The current endovascular medications (e.g., angioplasty or stents) only relieve the clinical symptoms while the surgical therapies (e.g., bypass or endarterectomy) require grafting vessels from a healthy organ to the diseased limb of the patient. However, even with these therapeutic techniques, 30% of patients still undergo limb amputation within a year. Thus, understanding of disease mechanism and development of new therapeutic approaches are in urgent needs.
Meis1 (myeloid ecotropic viral integration site 1) gene belongs to the three-amino-acid loop extension subclass of homeobox gene families, and it is a highly conserved transcription factor in all eukaryotes. Up to date, little is known about the role of Meis1 in regulating vascular remodeling under ischemic condition. In this study, we aim to investigate the role and underlying mechanism of Meis1 in the regulation of arteriogenesis and angiogenesis using hindlimb ischemia model of transgenic neonatal mice. The long-term goal is to develop a new treatment for patients with PAD. Three separate but related studies were planned to complete the proposed research aims.
To better understand the role of Meis1, we reviewed, in the first chapter, all literature relevant to the recent advances of the Meis1 in normal hematopoiesis, vasculogenesis, and heart developments, which were mostly studied in zebrafish and mouse. Briefly, Meis1 is found to be highly expressed in the brain and retina in zebrafish and additional in the heart, nose, and limb in mouse during the very early developmental stage, and remains at a low level quickly after birth. Meis1 is necessary for both primitive and definitive hematopoiesis and required for posterior erythroid differentiation. The absence of Meis1 results in a severe reduction of the number of mature erythrocytes and weakens the heart beats in zebrafish. Meis1 deficiency mouse is dead as early as E11.5 due to the severe internal hemorrhage. In addition, Meis1 is essential in heart development. Knock-down of Meis1 can promote angiotensin II-induced cardiomyocytes (CMs) hypertrophy or CMs proliferation, which can be repressed by a transcription factor Tbx20. Meis1 appears to play a complicated role in the blood vessels. Although the major blood vessels are still normal when global deletion of Meis1, the intersegmental vessel cannot be formed in Meis1 morphants in the zebrafish, and the small vessels are either too narrow or form larger sinuses in Meis1 deficient mouse. The effects of Meis1 on the vascular network under normal and disease (ischemia) condition remain largely unknown, and the existing data in this field is limited.
In the second chapter, we developed a method protocol to identify mice of all ages, especially neonates that we faced methodological difficulties to easily and permanently label prior to our major experiments. In this study, single- or 2-color tattooing (ear, tail, or toe or combinations) was performed to identify a defined or unlimited number of mice, respectively. Tail tattooing using both green and red pastes was suitable for identifying white-haired neonatal mice as early as postnatal day (PND) 1, whereas toe tattooing with green paste was an effective alternative approach for labeling black-haired mouse pups. In comparison, single-color (green) or 2-color (green and red) ear tattooing identified both white and black adult mice older than three weeks. Ear tattooing can be adapted to labeling an unlimited number of adult mice by adding the cage number. Thus, tattooing various combinations of the ears, tail, and toes provides an easy and permanent approach for identifying mice of all ages with minimal disturbance to the animals, which shows a new approach than any existing method to identify mouse at all ages, especially the neonatal pups used in the present study (Chapter 4).
Various formation of hindlimb ischemia with ligations of femoral artery or vein or both have been reported in the literature. The ischemic severity varies dependent on mouse strains and ligation methods. Due to the tiny body size of our experimental neonatal mice (PND2), it is technically challenging to separate the femoral artery from femoral vein without potential bleeding. In the third chapter, we aimed to explore a suitable surgical approach that can apply to neonatal mice. To this end, we compared the effects of femoral artery/vein (FAV) excision vs. femoral artery (FA) excision on hindlimb model using adult CD-1 mice. We showed during the 4-week period of blood reperfusion, no statistically significant differences were found between FAV and FA excision-induced ischemia regarding the reduction of limb blood flow, paw size, number of necrotic toes, or skeletal muscle cell size. We conclude that FAV and FA excision in CD-1 mice generate a comparable severity of hindlimb ischemia. In other words, FAV ligation is no more severe than FA ligation. These findings provide valuable information for researchers when selecting ligation methods for their neonate hindlimb models. Based on these findings, we selected FAV ligation of hindlimb ischemia approach to study the function of Meis1 in vascular remodeling of neonatal mice. In the fourth chapter (the main part of my dissertation), we investigated the roles of Meis1 in regulating arteriogenesis and angiogenesis of neonatal mouse under the ischemic condition. To this end, endothelial cell-specific deletion of Meis1 was generated by cross-breeding Meis1flox/flox mice with Tie2-Cre mice. Wild-type (WT, Meis1f/f) and endothelial cell-specific knock-out (KO, Meis1ec-/-Tie2-Cre+) C57BL/6 mice at the age of PND2 were used. Under the anesthesia, the pups were subject to hindlimb ischemia by excising FAV. Laser Doppler Imager was used to measure the blood flow pre- and post-surgery up to 28 days. Toe necrosis, skeletal regeneration, and vascular distributions were examined at the end of experiments (PND28 post-ischemia). Surprisingly, during 4-week periods after ischemia, the blood flow ratios (ischemic vs. control limb) in KO mice significantly increased compared to WT on PND14 and PND28, suggesting the inhibitory effects of Meis1 on blood flow recovery under ischemic condition. Meanwhile, WT mice showed more severe necrotic limb (lower ratio of limb length and area, and higher necrotic scores at PND7) than those in the KO mice. Furthermore, significant increases in diameters of Dil-stained arterioles of the skin vessel and the vessels on the ligation site were observed in KO mice, indicating the enhanced arteriogenesis in KO mice. To investigate the underlying mechanism, RNA from the ischemia and control limb was extracted and q-PCR was used to study the potential genes involved in the mechanism. Casp3 and Casp8 were found downregulated showing less apoptosis in the KO mice. On the other hand, endothelial cells (ECs) were isolated from the lungs of 3-5 WT and KO neonates using CD31 Microbeads. CD31+ cells were plated and treated with 0, 0.5, and 1μM doxorubicin for 24 hours and analyzed with various assays. Meis1-KO ECs demonstrated higher cell viability and formed a higher number of vascular tubes than those in WT ECs following 0.5μM Dox treatment, presenting the potential ability of angiogenesis in KO-ECs. Furthermore, the increased viability in KO ECs may be due to the decreased expression or activities of Casp8 and Casp3.
In conclusion, my present studies have developed a new methodology to easily and permanently identify all mice at any ages. The insignificant differences between FAV and FA ligations suggest that a relative-easy surgical approach could be used to generate hindlimb ischemic model, which potentially reduces the cost, decreases the surgical time and prevents damage of femoral nerve from surgical tools. More importantly, by using transgenic mice, we found that Meis1-KO dramatically increased blood flow and protected the ischemic hindlimb through vascular remodeling. Obviously, the molecular and cellular mechanisms underlying the above beneficial effects appear complicated and likely to involve multiple cellular remodeling processes and molecular signaling pathways to enhance arteriogenesis and angiogenesis and/or reduce cellular apoptosis through Meis1-mediated pathways. Our study demonstrated that under ischemic condition, knockout of Meis1 increases expression of Hif1a, which then activates Agt or VEGF, thus enhances arteriogenesis or angiogenesis; In addition, knockout of Meis1 activates Ccnd1, which subsequently promotes regeneration of skeletal muscle, and reduces expression of Casp8 and Casp3, thus preventing limb tissue from ischemia-induced apoptosis. Our innovative findings offer great potential to ultimately lead to new drug discovery or therapeutic approaches for prevention or treatment of PAD.PHD
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Djassemi, Navid. "Tissue Engineering A Blood Vessel Mimic While Monitoring Contamination Through Sterility Assurance Testing." DigitalCommons@CalPoly, 2012. https://digitalcommons.calpoly.edu/theses/831.
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Tissue Engineering A Blood Vessel Mimic While Monitoring Contamination Through Sterility Assurance Testing
Navid Djassemi
Tissue engineering blood vessel mimics has been proposed as a method to analyze the endothelial cell response to intravascular devices that are used in today’s clinical settings for the treatment of cardiovascular disease. Thus, the development of in vitro blood vessel mimics (BVMs) in Cal Poly’s Tissue Engineering Lab has introduced the possibility of assessing the characteristics of cellular response to past, present, and future intravascular devices that aim at treating coronary artery disease.
This thesis aimed at improving the methods and procedures utilized in the BVM model. Initial aspects of this project focused on using an expanded polytetrafluoroethylene (ePTFE) scaffold in conjunction with human endothelial cells to replicate the innermost intimal layer of a blood vessel. Human umbilical vein endothelial cells (HUVECs) were pressure sodded onto ePTFE scaffolds through cell sodding techniques in an attempt to effectively and consistently replicate and assess the intimal layer. Through each study ePTFE grafts were subjected to different culture times and steady flow rates to observe and compare the differences in the endothelial cell deposition. Results were inconsistent, although moderate cell adhesion was noted throughout each of the BVM setups. Each study exhibited a range of cell sodding density rates.
In the second phase of the thesis, contamination assessment protocols were implemented in the BVM lab. The intent of this part of the project was to assess the relative sterility in the cell culture lab, a critical component involved with the success or hindrance of cell and tissue cultures. Using microbiological validated methods, microbiological tests were conducted to examine the levels of microbial growth in and around the tissue engineering lab.
Results were tracked over a two month period in the lab with several observations of aerobic microorganism growth on various counter and lab surfaces. Higher growth trends were found on surfaces outside the cell culture lab, in the general TE lab area. These findings were used to provide overall suggestions on tracking microbes for long-term durations in ongoing BVM setups to directly improve the overall sterility assurance of the studies.
As the project reached its conclusion a look back at all the BVM setups and contamination assessments lead to a few suggestions for improving aseptic techniques within the TE lab, such as monitoring microbial growth in the culture processes, creating limit specifications, and creating a standardized way to regulate quality control within the lab environment. Furthermore, as the development BVM evolves, the findings from this report can be used with related research for improving the culture conditions of various BVM studies.
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Lu, Lingge. "Roles of the Shb and Cbl Proteins in Signal Transduction and Blood Vessel Formation." Doctoral thesis, Uppsala : Acta Universitatis Upsaliensis : Univ.-bibl. [distributör], 2003. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-3491.
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Lam, In Kei. "Anti-angiogenic activities of flavonoids from Pericarpium Citri Reticulatae on human umbilical vein endothelial cells (HUVECs) and zebrafish." Thesis, University of Macau, 2010. http://umaclib3.umac.mo/record=b2447360.
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