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Lazarus, Alan H. "Involvement of transferrin receptors in human natural killer cell specificity." Thesis, McGill University, 1988. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=75860.
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Natural Killer (NK) cells are able to recognize and lyse tumor cells without prior immunization or sensitization. The initial events leading to target cell lysis by NK cells involves a poorly defined recognition and binding phase. It has been hypothesized however that human NK cells may recognize transferrin receptors as target structures on tumor cells.To determine the possible involvement of transferrin receptors in human NK cell specificity, a correlation study between transferrin receptor expression and competitive activity for NK cell mediated lysis was undertaken. We have determined that the level of transferrin receptors expressed by different populations of K562 cells correlated well with their level of competitive activity for NK cell mediated lysis.
To investigate if these transferrin receptors could be recognized and bound by NK cells, a solid phase receptor binding assay was developed. As a model system, it was demonstrated that nitrocellulose immobilized transferrin retained its specific functional receptor binding capacity. This technique was quantitative and proved to be sufficiently sensitive to specifically detect nanogram quantities of transferrin receptor protein. Binding was assessed using an ELISA based system.
Human PBL were fractionated by discontinuous Percoll density centrifugation, bound to nitrocellulose, and evaluated for transferrin receptor binding capacity. A sample aliquot of cells from each Percoll fraction was retained to assess NK cell activity. It was observed that there was no positive relationship between NK cell activity and transferrin receptor binding capacity in these Percoll fractionated cells.
These findings indicate that while transferrin receptors may be involved in human NK cell specificity, they do not support a role for transferrin receptors in a high affinity mechanism between NK cells and tumor target cells.
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Cardoso, Aline Monticelli 1988. "Estudos sobre a internalização celular da STC1 humana." [s.n.], 2015. http://repositorio.unicamp.br/jspui/handle/REPOSIP/314360.
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Orientador: Jörg KobargDissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: A Stanniocalcina-1 (STC1) humana é uma glicoproteína homóloga a Stanniocalcina (STC) originalmente identificada como um hormônio regulador da homeostase de cálcio em peixes. A STC1 humana secretada atua em diferentes processos fisiológicos incluindo a angiogênese, a hipóxia e, principalmente, a carcinogênese, demonstrando assim uma atividade abrangente. Atualmente não se conhece o receptor da STC1 e pouco se sabe sobre o mecanismo de ação e de entrada nas células dessa proteína. Assim, o objetivo desse trabalho foi investigar um candidato a receptor de membrana dessa proteína, o receptor de transferrina (TfR1), uma proteína transmembrana responsável pela absorção de ferro nas células. Esse receptor é provavelmente expresso por todas as células em diferentes níveis, em destaque em células do sistema hematopoiético, em células em divisão celular e células neoplásicas. Assim, avaliou-se por citometria de fluxo o efeito do tratamento com STC1 em células não transfectadas e células transfectadas superexpressando o receptor de transferrina. Células tratadas com STC1 demonstraram um efeito semelhante ao tratamento com transferrina, um conhecido ligante desse receptor, no qual ambos diminuíram o número de células positivas para a marcação da superfície com transferrina conjugada com fluorocromo (transferrina-Alexa Fluor® 488 - Life Technologies). Em outro conjunto de experimentos de Western Blot foi demonstrado que a STC1 adicionada no sobrenadante das culturas de células é internalizada nas células e detectável no lisado celular, principalmente as células transfectadas para a superexpressão do receptor de transferrina. Complementarmente, em experimentos de localização subcelular por imunofluorescência a STC1 foi detectada em uma forma pontual e espalhada no citoplasma. Em conjunto, todos esses experimentos sugerem que STC1 e transferrina interferem na localização do receptor de transferrina na superfície celular e que possivelmente esse receptor está envolvido em mecanismos de internalização da própria STC1
Abstract: Human Stanniocalcin 1 (STC1) is the mammalian homologue of STC, which was originally identified as a calcium-regulating hormone in bony fishes. The human secreted Stanniocalcin acts on different physiological processes, including angiogenesis, hypoxia and especially carcinogenesis, facts that demonstrate their activity is wide. Currently there are few data on the mechanism of action of this protein or how it enters the cell. Thus, the aim of this study was to investigate transferrin receptor (TfR1) as a candidate to membrane receptor protein of STC1. This receptor is a membrane protein responsible for the iron uptake in cells. This receptor is probably expressed by all cells especially by cells in division and cancer cells, but its expression level may vary. We evaluated by flow cytometry the effect of STC1 treatment in non-transfected cells and cell with TfR1 overexpression. The treatment with STC demonstrated a similar effect to treatment with transferrin, a known ligand for receptor, which decreased the number of positive cells for staining with fluorochrome (transferrin conjugated to Alexa Fluor® 488 - Life Technologies). We also demonstrated by Western Blot that STC1 added to the supernatant of cultures of cells, especially cells that overexpress transferrin receptor, is internalized into the cells and detectable in the cell lysate. Additionally, in subcellular localization experiments by immunofluorescence STC1 was detected in a timely manner and scattered in the cytoplasm. Together all this information suggests that STC1 and transferrin interferes with the localization of the transferrin receptor in the cellular surface and perhaps this receptor is involved in the mechanism of internalization of STC1
Mestrado
Bioquimica
Mestra em Biologia Funcional e Molecular
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Kim, Jonghan. "Pharmacokinetics and pharmacodynamics of protein turnover and production in vivo." Connect to this title online, 2004. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1100554543.
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Thesis (Ph. D.)--Ohio State University, 2004.Title from first page of PDF file. Document formatted into pages; contains xxi, 203 p.; also includes graphics. Includes bibliographical references (p. 191-203).
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Hälldin, Jonas. "Oxidative stress and alterations in the mammalian iron metabolism : a study on iron, inflammation, oxidative stress and neurodegeneration in cellular model systems /." Stockholm : Department of Neurochemistry, Stockholm University, 2007. http://urn.kb.se/resolve?urn=urn:nbn:se:su:diva-7037.
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Lazaron, Victor. "A Potential Role for the 70 kD Heat Shock Cognate Protein in Receptor Endocytosis." eScholarship@UMMS, 1996. http://escholarship.umassmed.edu/gsbs_diss/234.
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Nutrient and growth factor receptors internalize through dathrin coated pits. The signal sequences which mediate the association between receptors and the coated pit reside in receptor cytoplasmic tail domains. These signal sequences have been extensively investigated in nutrient receptors, and a minimal functional sequence has been identified consisting of a tyrosine residue in an exposed b turn. Protein-protein contacts between internalization signal sequences and components of the coated pit machinery have been proposed to mediate rapid internalization. In vitro evidence suggests the AP-2 adaptor may be that protein component. The signal sequences of growth factor receptors are less well understood. However, a growth factor- and temperature- dependent binding between the epideimal growth factor receptor and the AP-2 adaptor has been observed.
We identified Hsc70 as a cytosolic ligand for the cytoplasmic tail of the transferrin receptor. The binding was mapped to the internalization signal sequence of the receptor tail. Mutations within the signal sequence which inhibit internalization result in alteration of signal sequence secondary structure and reduction in stimulation of the Hsc70 ATPase. Co-immunoprecipitation analysis showed a population of transferrin receptors which are bound to Hsc70, suggesting an association in vivo.
We also showed binding of Hsc70 to the epidermal growth factor receptor by co-immunoprecipitation analysis. This binding was increased by treatment with EGF. The binding was transient, and occured prior to the binding of the receptor to AP-2 adaptors. Other agents which induce EGF receptor clustering and internalization also stimulate the transient increase in Hsc70 binding and the later AP-2 binding, suggesting a role in early endocytosis.
These data support the hypothesis that Hsc70 is associated with the receptors for transferrin and epidermal growth factor in vitro and in vivo. We propose a role for the 70 kD heat shock protein in the assembly/disassembly of protein complexes involved in receptor signalling and/or internalization.
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Carvalho, Beatriz Assis. "Estado nutricional de ferro de lactentes atendidos em unidades básicas de saúde." Universidade Federal de Goiás, 2015. http://repositorio.bc.ufg.br/tede/handle/tede/4498.
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Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq
To evaluate the nutritional status of iron and its related factors in children 12 to 15 months assisted in Health Units in Goiânia, Goiás. METHODS: This is a cross-sectional study nested in research "Effectiveness of home fortification with vitamins and minerals in the prevention of iron deficiency and anemia in children under one year of age: a multicenter study in Brazilian cities ". The study was conducted with 230 children, aged between 12 and 15 months, assisted in Health Units in Goiânia, from June 2012 to February 2013. The prevalence of iron deficiency, iron deficiency anemia and anemia were assessed by the plasma means concentration of ferritin and transferrin receptor, hemoglobin and C-reactive protein. Multiple linear regression was used to estimate the effect of independent variables on the log plasma concentrations of ferritin. These variables were socioeconomic, demographic, maternal, pregnancy, anthropometric, breastfeeding, use of supplement, and biochemical parameters. RESULTS: Regarding the iron status, iron deficiency and iron deficiency anemia prevalence was 14.1% and 1.5%, respectively. Also, anemia prevalence was 5.6% of the infants studied. The predictors of ferritin were folate, vitamin B12 and the use of iron supplement at the time of collection, which each unit raised the log plasma concentration of ferritin in 0.009 mg/L, 0.001 mg/L and 0.315 mg/L, respectively. CONCLUSION: The results of this study showed low prevalence of iron deficiency and anemia in children studied. The use of iron supplements and serum concentrations of vitamin B12 and folate correlated ferritin concentrations and consequently the iron status in this population. Keywords: Iron Deficiency; Ferritins; Receptors, transferrin; Folic Acid; Vitamin B 12; Infant.
Avaliar o estado nutricional de ferro e os seus fatores relacionados em crianças de 12 a 15 meses atendidas em Unidades Básicas de Saúde de Goiânia, Goiás. MÉTODOS: Trata-se de um estudo transversal aninhado a pesquisa “Efetividade da fortificação caseira com vitaminas e minerais na prevenção da deficiência de ferro e anemia em crianças menores de um ano: estudo multicêntrico em cidades brasileiras”. O trabalho foi realizado com 230 crianças, de 12 e 15 meses, atendidas em Unidades Básicas de Saúde de Goiânia, no período de junho de 2012 a fevereiro de 2013. As prevalências de deficiência de ferro, anemia por deficiência de ferro e anemia foram avaliadas por meio da concentração plasmática de ferritina e receptor de transferrina, hemoglobina e proteína C-reativa. Foi utilizada regressão linear múltipla para estimar o efeito de variáveis independentes sobre o log das concentrações plasmáticas de ferritina. Estas variáveis foram condições socioeconômicas, demográficas, maternas, gestacionais, antropomêtricas, amamentação, uso de suplemento, e parâmetros bioquímicos. RESULTADOS: Com relação ao estado nutricional de ferro, as prevalências de deficiência de ferro e anemia por deficiência de ferro foram de 14,1% e 1,5% respectivamente. Além disso, foi encontrada prevalência de 5,6% de anemia nos lactentes estudados. Os fatores associados a ferritina foram o folato, a vitamina B12 e o uso de suplemento de ferro no momento da coleta, os quais cada unidade elevaram o log da concentração plasmática de ferritina em 0,009 μg/L, 0,001 μg/L e 0,315 μg/L, respectivamente. CONCLUSÃO: Os dados do presente estudo evidenciaram baixas prevalências de deficiência de ferro e anemia nas crianças estudadas. O uso de suplemento de ferro e as concentrações séricas das vitaminas B12 e folato correlacionaram-se as concentrações de ferritina e consequentemente, o estado nutricional de ferro nesta população.
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Thomas, Carla. "The validation and use of the rat intestinal epithelial cell line 6 (IEC-6) to study the role of ferroportin1 and divalent metal transporter 1 in the uptake of iron from Fe(II) and Fe(III)." University of Western Australia. Physiology Discipline Group, 2003. http://theses.library.uwa.edu.au/adt-WU2004.0019.
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[Formulae and special characters can only be approximated here. Please see the pdf version of the abstract for an accurate reproduction.] Iron is vital for almost all living organisms by participating in a wide variety of metabolic processes, including oxygen transport, DNA synthesis, and electron transport. However, iron concentrations in body tissues must be tightly regulated because excessive iron leads to tissue damage, as a result of formation of free radicals. In mammals since no controlled means of eliminating unwanted iron has evolved, body iron balance is maintained by alterations in dietary iron intake. This occurs in the duodenum where most dietary iron is absorbed. Absorption involves at least two steps, uptake of iron from the intestinal lumen and then its transport into the body, processes that occur at the apical and basal membranes of enterocytes, respectively. In chapter one of this thesis the background information relevant to iron absorption is described. Despite numerous studies, the role of these proteins in iron absorption remains unclear, partly because many studies have reported them in non-enterocyte cell lines where the expression of the proteins involved in iron absorption is unlikely and therefore the physiological significance of the findings uncertain. Therefore, the study of iron absorption would value from additional cell lines of intestinal origin being used, preferably derived from a species used to comprehensively study this process in vivo, namely the rat. Validation of such a model would enable comparisons to be made from a molecular level to its relevance in the whole organism. In chapter 3 of this thesis, the rat intestinal cell line 6 (IEC-6) was examined as a model of intestinal iron transport. IEC-6 cells expressed many of the proteins involved in iron absorption, but not the ferrireductase Dcytb, sucrase or αvβ3 integrin. In addition, in IEC-6 cells the expression of the apical transporter divalent metal transporter 1 (DMT1), the iron storage protein ferritin, the uptake of Fe(II) and Fe(III) were regulated by cellular iron stores as is seen in vivo. This suggests that IEC-6 cells are of a lower villus enterocyte phenotype. Presented in chapter 4 is the study of the uptake of iron from Fe(II):ascorbate and Fe(III):citrate by IEC-6 cells in the presence of a blocking antibody to the putative basolateral transporter ferroportin1 and of colchicine and vinblastine, different pHs, and over-expression of DMT1. It was shown that optimal Fe(II) uptake required a low extracellular pH and was dependent on DMT1. Uptake of Fe(III) functioned optimally at a neutral pH, did not require surface ferrireduction, and was increased during over-expression of DMT1. These observations suggest that intravesicular ferrireduction takes place before transport of Fe(II) to the cytoplasm by DMT1. This pathway was not blocked by a functional antibody against αvβ3 integrin but was inhibited by competition with unlabeled iron citrate or citrate alone. Surprisingly, a functional antibody against ferroportin1 had no effect on efflux but significantly reduced (p<0.05) uptake of Fe(II) by 40-50% and Fe(III) by 90%, indicating two separate pathways for the uptake of iron from Fe(II)-ascorbate and from Fe(III)-citrate in IEC-6 cells. Presented in chapter 5 is the development and validation of a technique for the removal of freshly isolated enterocytes from the rat duodenum and their use to study iron transport processes that enabled comparisons to be made between these cells, IEC-6 cells and the human enterocyte cell line Caco-2 cells. In chapter 6 a blocking antibody to ferroportin1 was shown to inhibit uptake of Fe(II) but not release of iron in freshly isolated duodenal enterocytes from rats and Caco-2 cells supporting the findings obtained with IEC-6 cells described in chapter 4. Fe(II) uptake was reduced only when the antibody was in contact with the apical membrane indicating its expression at the microvillus membrane. Confirming this, ferroportin1 was shown along the microvillus membrane of Caco-2 cells, in enriched microvillus membrane preparations and in enterocytes of duodenum tissue of rats where it co-localised with lactase. The significant findings to emerge from this thesis are that the IEC-6 cell is a valid model to study iron absorption producing results consistent with those found in freshly isolated enterocytes and in human enterocyte-like cells. In particular, ferroportin1 functions in the uptake of iron at the apical membrane possibly by modulating surface binding of Fe(II) to DMT1 or the activity of DMT1. In addition to this in Fe(II) uptake from Fe(III) ferroportin1 may also affect the number of Fe(III): citrate binding sites. Preliminary studies further characterizing the function of ferroportin1 at the apical membrane and at intracellular sites of IEC-6 cells along with integration of these data are discussed in chapter 7.
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Navaroli, Deanna M. "Molecular Mechanisms of Endocytosis: Trafficking and Functional Requirements for the Transferrin Receptor, Small Interfering RNAs and Dopamine Transporter: A Dissertation." eScholarship@UMMS, 2004. http://escholarship.umassmed.edu/gsbs_diss/592.
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Endocytosis is an essential function of eukaryotic cells, providing crucial nutrients and playing key roles in interactions of the plasma membrane with the environment. The classical view of the endocytic pathway, where vesicles from the plasma membrane fuse with a homogenous population of early endosomes from which cargo is sorted, has recently been challenged by the finding of multiple subpopulations of endosomes. These subpopulations vary in their content of phosphatidylinositol 3- phosphate (PI3P) and Rab binding proteins. The role of these endosomal subpopulations is unclear, as is the role of multiple PI3P effectors, which are ubiquitously expressed and highly conserved. One possibility is that the different subpopulations represent stages in the maturation of the endocytic pathway. Alternatively, endosome subpopulations may be specialized for different functions, such as preferential trafficking of specific endocytosed cargo. To determine whether specific receptors are targeted to distinct populations of endosomes, we have built a platform for total internal reflection fluorescence (TIRF) microscopy coupled with structured illumination capabilities named TESM (TIRF Epifluorescence Structured light Microscope.) In this study, TESM, along with standard biochemical and molecular biological tools, was used to analyze the dynamic distribution of two highly conserved Rab5 and PI3P effectors, EEA1 and Rabenosyn-5, and systematically study the trafficking of transferrin. Rabenosyn-5 is necessary for proper expression of the transferrin receptor as well as internalization and recycling of transferrin-transferrin receptor complexes. Results of combining TIRF with structured light Epifluorescence (SLE) indicate that the endogenous populations of EEA1 and Rabenoysn-5 are both distinct and partially overlapping.
The application of antisense oligonucleotides as potential therapeutic agents requires effective methods for their delivery to the cytoplasm of target cells. In collaboration with RXi Pharmaceuticals we show the efficient cellular uptake of the antisense oligonucleotide sd-rxRNA® in the absence of delivery vehicle or protein carrier. In this study TIRF, SLE, and biochemical approaches were utilized to determine whether sd-rxRNA traffics and functions along specific endosomal pathways. Sd-rxRNA was found to traffic along the degradative pathway and require EEA1 to functionally silence its target. These new findings will help define the cellular pathways involved in RNA silencing.
Neurotransmitter reuptake and reuse by neurotransmitter transport proteins is fundamental to transmitter homeostasis and synaptic signaling. In order to understand how trafficking regulates transporters in the brain and how this system may be disregulated in monoamine-related pathologies, the transporter internalization signals and their molecular partners must be defined. We utilized a yeast two-hybrid system to identify proteins that interact with the dopamine transporter (DAT) endocytic signal. The small, membrane associated, GTPase Rin was determined to specifically and functionally interact with the DAT endocytic signal, regulating constitutive and protein kinase C (PKC) – stimulated DAT endocytosis. The results presented in this study provide new insights into functions and components of endocytosis and enhance the understanding of endocytic organization.
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Navaroli, Deanna M. "Molecular Mechanisms of Endocytosis: Trafficking and Functional Requirements for the Transferrin Receptor, Small Interfering RNAs and Dopamine Transporter: A Dissertation." eScholarship@UMMS, 2012. https://escholarship.umassmed.edu/gsbs_diss/592.
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Endocytosis is an essential function of eukaryotic cells, providing crucial nutrients and playing key roles in interactions of the plasma membrane with the environment. The classical view of the endocytic pathway, where vesicles from the plasma membrane fuse with a homogenous population of early endosomes from which cargo is sorted, has recently been challenged by the finding of multiple subpopulations of endosomes. These subpopulations vary in their content of phosphatidylinositol 3- phosphate (PI3P) and Rab binding proteins. The role of these endosomal subpopulations is unclear, as is the role of multiple PI3P effectors, which are ubiquitously expressed and highly conserved. One possibility is that the different subpopulations represent stages in the maturation of the endocytic pathway. Alternatively, endosome subpopulations may be specialized for different functions, such as preferential trafficking of specific endocytosed cargo. To determine whether specific receptors are targeted to distinct populations of endosomes, we have built a platform for total internal reflection fluorescence (TIRF) microscopy coupled with structured illumination capabilities named TESM (TIRF Epifluorescence Structured light Microscope.) In this study, TESM, along with standard biochemical and molecular biological tools, was used to analyze the dynamic distribution of two highly conserved Rab5 and PI3P effectors, EEA1 and Rabenosyn-5, and systematically study the trafficking of transferrin. Rabenosyn-5 is necessary for proper expression of the transferrin receptor as well as internalization and recycling of transferrin-transferrin receptor complexes. Results of combining TIRF with structured light Epifluorescence (SLE) indicate that the endogenous populations of EEA1 and Rabenoysn-5 are both distinct and partially overlapping.
The application of antisense oligonucleotides as potential therapeutic agents requires effective methods for their delivery to the cytoplasm of target cells. In collaboration with RXi Pharmaceuticals we show the efficient cellular uptake of the antisense oligonucleotide sd-rxRNA® in the absence of delivery vehicle or protein carrier. In this study TIRF, SLE, and biochemical approaches were utilized to determine whether sd-rxRNA traffics and functions along specific endosomal pathways. Sd-rxRNA was found to traffic along the degradative pathway and require EEA1 to functionally silence its target. These new findings will help define the cellular pathways involved in RNA silencing.
Neurotransmitter reuptake and reuse by neurotransmitter transport proteins is fundamental to transmitter homeostasis and synaptic signaling. In order to understand how trafficking regulates transporters in the brain and how this system may be disregulated in monoamine-related pathologies, the transporter internalization signals and their molecular partners must be defined. We utilized a yeast two-hybrid system to identify proteins that interact with the dopamine transporter (DAT) endocytic signal. The small, membrane associated, GTPase Rin was determined to specifically and functionally interact with the DAT endocytic signal, regulating constitutive and protein kinase C (PKC) – stimulated DAT endocytosis. The results presented in this study provide new insights into functions and components of endocytosis and enhance the understanding of endocytic organization.
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Fouquet, Guillemette. "Régulation de l’érythropoïèse : rôle des récepteurs à la transferrine et d’un phytoestrogène." Thesis, Université Paris-Saclay (ComUE), 2019. http://www.theses.fr/2019SACLS293.
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L’érythropoïèse est un processus extrêmement prolifératif, et qui doit donc être très étroitement régulé. L’érythropoïétine (EPO) est l’un des facteurs absolument nécessaires à l’érythropoïèse. Cependant, dans la moelle osseuse, la quantité d'EPO circulante est sous-optimale et la capacité des érythroblastes à survivre dépend donc de leur sensibilité à l'EPO. Les facteurs modulant la réponse à l'EPO au cours de l'érythropoïèse sont encore largement inconnus.Nous avons donc voulu explorer plusieurs facteurs pouvant potentiellement être impliqués dans la régulation de l’érythropoïèse et plus précisément dans la réponse à l’EPO : tout d’abord, la transferrine ainsi que ses récepteurs (TfR), la transferrine et le TfR1 étant également essentiels à l’érythropoïèse, ainsi qu’un phytoestrogène provenant d’une plante nommée Curcuma comosa, les oestrogènes étant eux aussi connus pour favoriser l’érythropoïèse.Concernant la transferrine, nous avons voulu principalement explorer son rôle sur la signalisation, ayant récemment montré au laboratoire que le TfR1, essentiellement connu pour son rôle dans l’endocytose du fer, est également capable d’entraîner une signalisation.Nous avons montré que la transferrine potentialise la stimulation induite par l’EPO des voies ERK, AKT et STAT5. Cet effet est conservé même en l’absence d’endocytose du TfR1. Aucune coopération n’a été trouvée entre la transferrine et le stem cell factor (SCF).Nous avons également observé qu’en l’absence du TfR2, il existe une augmentation de l’expression de l’EPO-R et de la signalisation induite par l’EPO, sans impact de la transferrine dans ce contexte. Par ailleurs, nous avons montré que le Curcuma comosa améliore la prolifération et la différenciation des progéniteurs érythroïdes précoces, par un mécanisme de potentialisation de la signalisation induite par l’EPO impliquant le récepteur aux oestrogènes ER-α.En conclusion, la transferrine et ses récepteurs, ainsi qu’un phytoestrogène et l’ER-α, sont impliqués dans la régulation de l’érythropoïèse via leur action sur la signalisation induite par l’EPO. L’approfondissement de ces données pourrait ouvrir de nouvelles pistes thérapeutiques dans le traitement de l’anémieErythropoiesis is an extremely proliferative process and must be very closely regulated. Erythropoietin (EPO) is one of the major factors necessary for erythropoiesis. However, in the bone marrow, the amount of circulating EPO is suboptimal and the ability of erythroblasts to survive therefore depends on their sensitivity to EPO. The factors modulating the response to EPO during erythropoiesis are still largely unknown. We therefore wanted to explore several factors that could potentially be involved in the regulation of erythropoiesis and more specifically in the response to EPO: first, transferrin and its receptors (TfR), transferrin and TfR1 being also essential for erythropoiesis, as well as a phytoestrogen from a plant called Curcuma comosa, as estrogens are also known to promote erythropoiesis. Regarding transferrin, we mainly wanted to explore its role on signaling, having recently shown in the laboratory that TfR1, essentially known for its role in iron endocytosis, is a signaling-competent receptor. We have shown that transferrin potentiates EPO-induced stimulation of the ERK, AKT and STAT5 pathways. This effect is maintained even in the absence of TfR1 endocytosis. No cooperation was found between transferrin and stem cell factor (SCF). We also observed that in the absence of TfR2, there is an increase in EPO-R expression and EPO-induced signaling, without any impact of transferrin in this context.In addition, we have shown that Curcuma comosa improves the proliferation and differentiation of early erythroid progenitors through a mechanism involving the ER-α estrogen receptor, able to potentiate EPO-induced signaling. In conclusion, transferrin and its receptors, as well as a phytoestrogen and ER-α, are involved in the regulation of erythropoiesis through their action on EPO-induced signaling. Further investigation of these data could provide new therapeutic strategies in the treatment of anemia
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CABRAL, FILHO Paulo Euzébio. "Desenvolvimento de sondas multimodais baseadas em pontos quânticos para aplicações biomédicas." Universidade Federal de Pernambuco, 2016. https://repositorio.ufpe.br/handle/123456789/18443.
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Os pontos quânticos ou quantum dots (QDs) são nanocristais fluorescentes de semicondutores com propriedades ópticas únicas, tendo como principais vantagens: (1) alta resistência à fotodegradação, possibilitando o acompanhamento de eventos biológicos em tempo real e, (2) superfície ativa, permitindo a conjugação a biomoléculas que vão propiciar especificidade às marcações, além de possibilitar também sua ligação a outras nanopartículas. Com isso, é possível quantificar uma variedade de biomoléculas em células e tecidos e desenvolver nanossondas bimodais (magnético-fluorescentes) baseadas em QDs. O desenvolvimento de nanopartículas bimodais pode aliar as vantagens das técnicas baseadas em fluorescência com as de imagem por ressonância magnética (IRM). Entretanto, a obtenção de sondas bimodais é ainda um desafio, pois durante a conjugação devem ser mantidas as propriedades fluorescentes e magnéticas das nanopartículas, e com isso ainda há poucos trabalhos que façam aplicações em sistemas biológicos. O objetivo desta tese se caracteriza pelo desenvolvimento de sondas com propriedades multimodais baseadas em QDs de Telureto de Cádmio (CdTe) associadas a nanopartículas magnéticas de óxido de ferro como marcadores sítio-específicos em células cancerígenas. Inicialmente os QDs foram conjugados covalentemente à transferrina (Tf) [QDs-Tf] para a quantificação específica de seus receptores (TfRs) em células HeLa e em duas linhagens de glioblastoma (U87 e DBTRG). Através de ensaios de saturação do TfR, foi possível inferir sobre a taxa de renovação deste receptor nessas células. Os resultados mostraram que as células HeLa e as DBTRG possuem uma maior quantidade do TfR quando comparadas às U87. As DBTRG apresentaram maior taxa de renovação do TfR, quando comparadas aos outros dois tipos, demonstrando que os conjugados QDs-Tf são potenciais ferramentas para o estudo da biologia celular do câncer. Posteriormente, nanossondas bimodais (QDsMNPs), baseadas em QDs associados a nanopartículas magnéticas de óxido de ferro, foram obtidas por conjugação covalente. De acordo com as caracterizações, QDs-MNPs mantiveram suas propriedades ópticas e magnéticas e apresentaram-se como potenciais sondas inespecíficas para fluorescência e para aquisição de imagens por RM ponderadas em T2 (tempo de relaxação nuclear transversal). A conjugação prévia dos QDs a Tf, além de fornecer informações sobre a biologia do câncer, auxiliou também na padronização da marcação específica do TfR em células cancerígenas e no estabelecimento de protocolos de conjugação das sondas bimodais a Tf. Por fim, as QDs-MNPs foram conjugadas covalentemente a Tf e essa nova sonda multimodal [(QDs-MNPs)-Tf] reconheceu especificamente os TfR em células HeLa. As caracterizações indicaram que o sistema multimodal não apresentou alteração significativa nas propriedades ópticas e exibiu uma maior relaxividade transversal (r2), se mostrando igualmente potencial sonda para análise por fluorescência e IRM ponderada em T2. Neste trabalho foram obtidas nanossondas promissoras para serem aplicadas na compreensão da biologia celular do câncer, além de auxiliar em métodos diagnósticos e terapêuticos para essa doença.
Quantum dots (QDs) are fluorescent semiconductor nanocrystals with unique optical properties, which have as major advantages: (1) the high resistance to photobleaching, making possible to monitor biological events in real-time and, (2) active surface, allowing the conjugation not only with biomolecules for specific labeling, but also to other nanoparticles. Thus, it would be possible to quantify a variety of biomolecules in cells and tissues, as well as to develop bimodal nanoprobes (fluorescent-magnetic) [BNPs] based on QDs. The development of BNPs can help to combine the advantages of the fluorescence with the resonance magnetic imaging techniques. However, the preparation of bimodal probes can still be considered a challenge, since the fluorescent and magnetic nanoparticles’ properties need to be preserved after conjugation. Therefore, there are still few works applying BNPs in biological studies. The aim of this thesis was to develop nanoprobes, with multimodal properties, based on cadmium telluride (CdTe) QDs conjugated with iron oxide magnetic nanoparticles (MNPs), for site-specific labeling in cancer cells. For this, initially, QDs were covalently coupling to transferrin (Tf) [QDs-Tf] and used to quantify the transferrin receptor (TfRs) in HeLa cells as well as in two glioblastoma lines (U87 and DBTRG). Furthermore, by a TfR saturation assay, it was possible to study the recycling rate of this receptor in cells studied. The results showed that HeLa and DBTRG cells present a higher amount of TfRs when compared to U87. DBTGR showed a higher TfR recycling rate, when compared to the other two lineages, demonstrating that QDs-Tf conjugates are potential tools to study the cancer cell biology. BNPs, based on the conjugation of QDs with MNPs (QDs-MNPs), were obtained by covalent coupling. According to characterizations, the BNPs remained with their optical and magnetic properties preserved and showed to be potential unspecific probes for fluorescence analysis and for T2-weighted magnetic resonance imaging (MRI) acquisition. The conjugation of QDs to Tf, performed previously, was a valuable step not only to provide us information about the biology of cancer cells, but also for the standardization of TfR specific labeling and the establishment of protocol to conjugate the BNPs with Tf. Therefore, QDs-MNPs were also covalently coupling to Tf and this new multimodal nanotool [(QDs-MNPs)Tf] was also able to recognize specifically TfRs in HeLa cells. The multimodal nanosystems presented their fluorescent properties practically unchanged and also exhibited a higher transversal relaxivity (r2), when compared to bare BNPs, showing likewise potential to be used for fluorescence and T2-weighted MRI analyses. In this work, it was developed promising nanoprobes, able to be applied for the cancer cell biology comprehension, and with potential for helping in the improvement of diagnostic and therapeutic methods for this disease.
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Araujo, Felipe Saldanha de. "Avaliação fenotípica dos linfócitos T em um modelo animal de deficiência de ferro." Universidade de São Paulo, 2006. http://www.teses.usp.br/teses/disponiveis/60/60135/tde-17052007-094115/.
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O ferro é um elemento chave em muitos processos metabólicos, como transporte de oxigênio, síntese de hormônios esteróides, respiração celular, transporte de elétrons, síntese de DNA, proliferação e diferenciação celular e regulação gênica. A deficiência de ferro é a desordem nutricional mais comum afetando aproximadamente um terço da população mundial. Pequenos déficits no compartimento funcional de ferro têm sérias conseqüências sobre o sistema imune, principalmente na imunidade mediada por células. A abordagem dos pais ou responsáveis, as exigências éticas e a aderência de crianças da mesma faixa etária e sem outros problemas que afetem o metabolismo do ferro e o sistema imune são as principais dificuldades enfrentadas no desenvolvimento de pesquisas com seres humanos, sendo necessário o estabelecimento de modelos experimentais. Este trabalho teve como objetivo estabelecer um modelo de indução e recuperação de deficiência de ferro em camundongos, visando a sua utilização em estudos sobre alterações do sistema imune induzidas por esta deficiência. A deficiência de ferro foi induzida por ingestão de uma ração com baixo teor de ferro (5 mg /kg de ração) por 4 e 8 semanas. No termino deste período foram determinados: concentração de hemoglobina (colorimetrico), hematócrito (microhematócrito), estoques de ferro hepático (espectrometria de absorção atômica) e fenotipagem (citometria de fluxo) dos linfócitos presentes no sangue periférico e em suspensão de células do baço dos animais dos grupos controle (C) e deficiente em ferro (DF), sendo avaliado a porcentagem de células T CD4+ e CD8+, bem como a expressão do receptor de transferrina (CD71+) nessas subpopulações. Não houve diferenças na concentração de hemoglobina e no valor do hematócrito entre os animais dos grupos DF e C, porém os estoques de ferro estavam significantemente reduzidos nos animais do grupo DF de quatro (p<0,05) e oito (p<0,01) semanas. Não houve diferenças na porcentagem de linfócitos T CD4+ e T CD8+ entre os animais dos grupos DF e C, porém os animais deficientes em ferro apresentaram maior porcentagem de linfócitos T CD8+ do baço expressando CD71+ (p< 0,001). Este trabalho sugere que a depleção nos estoques de ferro não altera a proporção dos subtipos de linfócitos, porem as células T CD8 + do baço são mais sensíveis à deficiência de ferro.Iron have a crucial role in several metabolic pathways, such oxygen transport, steroid hormone synthesis, cellular respiration, electron transport, DNA synthesis, cellular proliferation and differentiation and genic regulation. The iron deficiency is most common disorder nutrition, affecting about 30% world population. Deficits in iron functional compartment have serious delays about immunity systems, especially in the cellular immunity. Because of environmental problems, age, deficiency of nutrients other than iron, prevalence of infection, which may make human studies difficult, we used an animal model. This work aimed established iron deficiency induction and recuperation in mouse, for study about immune systems alteration. Iron deficiency was induced by feeding mice a diet that contained only 5 mg Fe/Kg for 4 and 8 weeks. After this period were determined: hemoglobin (colorimetry), hematocrit (microhematocrit), liver iron stores (atomic absorption spectrophotometer) and we performed a flow cytometry analyses in peripheral blood and spleen lymphocytes in control (C) and iron deficient (ID) mouse. We defined the effects of iron deficiency on T-cell subset and expression of cell-surface transferrin receptor (CD71+) in these cells. Hemoglobin concentration and hematocrit of ID mice were not difference those of C mice, but iron stores of ID mice (4 and 8 weeks) were reduced (p< 0,05 and p< 0,01; respectively). Although T-cells subsets in peripheral blood and spleen were not altered, iron deficiency significantly increased the number of spleen T CD8+ cells that express CD 71+ (p< 0,001). Data suggest that depletion of iron storage not alter T-cells subsets and spleen T CD8+ is the most sensible subset in iron deficiency.
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Thornley, Thomas B. "IFN-α/β Induction by dsRNA and Toll-Like Receptors Shortens Allograft Survival Induced by Costimulation Blockade: A Dissertation." eScholarship@UMMS, 2006. https://escholarship.umassmed.edu/gsbs_diss/254.
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Costimulation blockade protocols are promising alternatives to the use of chronic immunosuppression for promoting long-term allograft survival. However, the efficacy of costimulation blockade-based protocols is decreased by environmental insults such as viral infections. For example, lymphocytic choriomeningitis virus (LCMV) infection at the time of costimulation blockade treatment abrogates skin allograft survival in mice. In this dissertation, we test the hypothesis that viruses shorten allograft survival by activating the innate immune system through pattern-recognition receptors (PRRs), such as toll-like receptors (TLRs).
To investigate the role of innate immunity in shortening allograft survival, costimulation blockade-treated mice were co-injected with TLR2 (Pam3Cys), TLR3 (polyinosinic:polycytidylic acid, poly(I:C)), TLR4 (lipopolysaccharide, LPS), or TLR9 (CpG DNA) agonists, followed by transplantation with skin allografts 7 days later. Costimulation blockade prolonged skin allograft survival that was shortened in mice coinjected with TLR agonists. To investigate the underlying mechanisms of this observation, we used synchimeric mice, which circulate trace populations of anti-H2b transgenic alloreactive CD8+ T cells. In synchimeric mice treated with costimulation blockade, co-administration of all four TLR agonists prevented deletion of alloreactive CD8+ T cells. These alloreactive CD8+ T cells 1) expressed the proliferation marker Ki-67, 2) upregulated CD44, and 3) failed to undergo apoptosis. We also demonstrate that costimulation blockade-treated CD8α-deficient mice exhibit prolonged allograft survival when co-injected with LPS. These data suggest that TLR agonists shorten allograft survival by impairing the apoptosis of alloreactive CD8+T cells.
We further delineate the mechanism by which TLR agonists shorten allograft survival by demonstrating that LPS and poly(I:C) fail to shorten allograft survival in IFNRI- deficient mice. Interestingly, the ability of poly(I:C) to more potently induce IFN-α/β than LPS correlates with its superior abilities to shorten islet allograft survival and induce allo-specific CTL activity as measured by an in vivo cytotoxicity assay. The ability to shorten allograft survival and induce IFN-α/β is a TLR-dependent process for LPS, but is a TLR-independent process for poly(I:C). Strikingly, the injection of IFN-β impairs alloreactive CD8+T cell deletion and shortens allograft survival, similar to LPS and poly(I:C). These data suggest that LPS and poly(I:C) shorten allograft survival by inducing IFN-α/β through two different mechanisms.
Finally, we present data showing that viruses (LCMV, Pichinde virus, murine cytomegalovirus and vaccinia virus) impair alloreactive CD8+T cell deletion and shorten allograft survival, in a manner comparable to LPS and poly(I:C). Similar to LPS, LCMV and MCMV exhibit an impaired ability to shorten allograft survival in MyD88-deficient mice. These data suggest that the MyD88 pathway is required for certain viruses and TLR-agonists to shorten allograft survival.
In this dissertation, we present data supporting an important role for TLRs and IFN- α/β in shortening allograft induced by costimulation blockade. Our findings suggest that targeting these pathways during the peri-transplant period may enhance the efficacy of costimulation blockade protocols in the clinic.
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Ahmad, Shakil [Verfasser], Gregor [Akademischer Betreuer] Eichele, and Lutz [Akademischer Betreuer] Walter. "Effect of acute phase cytokines on iron uptake in hepatocytes and differential localization of Lipocalin-2 and Transferrin receptors in rat hepatic and extra hepatic organs / Shakil Ahmad. Gutachter: Gregor Eichele ; Lutz Walter. Betreuer: Gregor Eichele." Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2014. http://d-nb.info/1057776289/34.
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Szulczewski, Vívian. "Estudo in vitro sobre a interação celular e vias endocíticas de papilomavírus humano (HPV) em leucócitos do sangue periférico." Universidade de São Paulo, 2009. http://www.teses.usp.br/teses/disponiveis/87/87131/tde-19082009-134506/.
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O papilomavírus humano (HPV) é o principal agente etiológico do câncer cervical e anogenital, sendo o HPV16 e o HPV18 os vírus de alto risco. Estudos recentes evidenciaram que além da transmissão sexual do HPV, há outras formas de contágio. Entretanto, a dificuldade na obtenção de quantidades viáveis do tipo selvagem ou mutante do HPV tem limitado em muito os estudos de diversos aspectos da biologia do papilomavírus. Este estudo investigou a possibilidade de o HPV infectar células leucocitárias do sangue periférico humano. Concluímos que as VLPs L1L2 do HPV16 podem utilizar a via endocítica do ferro mediada por clatrina, através do complexo VLPs-Transferrina-Receptor de Transferrina, permanecendo de forma latente em leucócitos. Esta porta de entrada oportunista poderia explicar a propagação crescente e alarmante deste agravo à saúde humana, motivo de preocupação nos sistemas mundiais de saúde pública. Este trabalho demonstrou pela primeira vez a internalização de VLPs L1L2 do HPV16 em leucócitos do sangue periférico humano.Human papillomavirus (HPV) is the primary etiologic agent of anogenital and cervical cancer, caused mainly by the high-risk HPV16 and HPV18 viruses. Recent studies revealed that besides the sexual transmission of HPV, there are other forms of contagion. However, the difficulty in obtaining quantities of viable wild-type or mutant of HPV constitutes a limiting factor in the studies of various aspects of the biology of human papillomavirus. This study investigated the possibility of HPV infect the cells of human peripheral blood leukocytes. We conclude that the VLPs L1L2 of HPV16 may use the iron endocytic pathway clathrin-mediated through the complex VLPs-Transferrin-Transferrin Receptor and remained so latent in leukocytes. This port of entry opportunist could explain the growing and alarming spread of this disease to human health, cause for concern in the global public health. This study showed for the first time the internalization of VLPs L1L2 of HPV16 in human peripheral blood leukocytes.
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Sharma, Nita Devi. "Molecular definition of the interaction of transferrin with the meningococcal and human transferrin receptor." Thesis, Birkbeck (University of London), 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.338641.
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Stokes, Russell Hayden. "Meningococcal transferrin binding proteins A and B form a functional human serum transferrin receptor." Thesis, King's College London (University of London), 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.313503.
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Melhem, Rana. "Ciblage du récepteur de la transferrine de type 1 (TfR1) et du métabolisme du Fer dans le cancer du pancréas." Thesis, Montpellier, 2017. http://www.theses.fr/2017MONTT011.
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L'adénocarcinome canalaire pancréatique (PDAC) est une maladie agressive à pronostic sombre et à forte mortalité. Il est donc crucial de rechercher de nouvelles cibles thérapeutiques et de nouveaux traitements. Une option intéressante pourrait être le ciblage du métabolisme du Fer. En effet, la transformation cellulaire s'accompagne généralement d'un accroissement des besoins en fer et de l'augmentation du récepteur de la transferrine de type 1, TfR1, le récepteur majeur impliqué dans l'approvisionnement des cellules en Fer par l'internalisation de la transferrine plasmatique chargée en fer. Nous avons utilisé un anticorps monoclonal humain IgG1 anti-TfR1 (H7) pour cibler le TfR1 dans le PDAC. Le traitement in vitro de 3 lignées de PDAC, établies à partir de tumeurs primaires de patients (BxPC3 et HPAC) ou d'une métastase hépatique (CFPAC) par H7 inhibe la viabilité cellulaire en réduisant la prolifération et induisant l'apoptose. H7 bloque efficacement l'internalisation de la transferrine chargée en Fer avec pour conséquence une baisse du Fer libre intracellulaire, une augmentation du TfR1 et une diminution de la ferritine, protéine de stockage du fer. Le traitement par H7 induit également l'expression du suppresseur de tumeur NDRG1 (N-myc downstream regulated gene 1), une cible prometteuse dans le cancer du pancréas, et la formation de sphères par la lignée HPAC in vitro, montrant que la déprivation en fer inhibe aussi les cellules initiatrices de tumeur dans ce modèle. Enfin, H7 recrute les cellules Natural Killer in vitro et induit efficacement l’ADCC (cytotoxicité cellulaire dépendante des anticorps). In vivo, dans 2 modèles de PDAC chez la souris (greffe de la lignée BxPC3 ou d’une tumeur dérivée d’un patient (PDX)), H7 diminue la croissance tumorale et augmente l'activité anti-tumorale du traitement chimiothérapeutique standard (gemcitabine). Ces résultats suggèrent que le ciblage du TfR1 par l'anticorps H7, seul ou en combinaison avec le traitement chimiothérapeutique standard est une stratégie prometteuse pour le traitement du PDACPancreatic ductal adenocarcinoma (PDAC) is a highly aggressive disease associated with poor diagnosis and high mortality. It is therefore necessary to search for new therapeutic targets and treatments. One of the interesting options would be targeting iron metabolism. Indeed, cell transformation is generally accompanied with increased needs for iron together with increased expression of the transferrin receptor 1, TfR1, the major receptor involved in cellular iron supply via the internalization of plasma transferrin loaded with iron.We have used a fully human internalizing anti-TfR1 antibody (IgG1), namely H7, to target TfR1 in PDAC. On three PDAC cell lines, BxPC3, HPAC (established from primary tumor), and CFPAC (established from hepatic metastasis), H7 treatment decreased cellular viability in vitro as a result of combined proliferation inhibition and apoptosis induction. H7 blocked efficiently transferrin internalization, and, likely due to a decrease in the labile iron pool, induced the upregulation of TfR1 and the downregulation of the iron storage protein ferritin. Interestingly, H7 treatment also induced the expression of the metastasis suppressor N-myc downstream regulated gene 1 (NDRG1), a promising therapeutic target in pancreatic cancer. H7 also decreased the ability of HPAC cell line to form tumor sphere in vitro indicating its inhibitory effect tumor initiating cells. Finally, H7 was able to recruit Natural killer cells and mediate antibody-dependent cell cytotoxicity on PDAC cell lines in vitro. In vivo, both in a PDAC cell line (BxPC3) and a patient derived xenograft (PDX) mouse model, H7 treatment decreases tumor growth and increases the anti-tumor activity of the Gemcitabine standard treatment. These data provide evidence that targeting pancreatic cancer with the iron depriving anti-TfR1 antibody, alone or in combination with gemcitabine might be a promising strategy in PDAC
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Adam, Mohammed A. "Fate of the transferrin receptor during in vitro maturation of sheep reticulocytes and post-translational modifications of the transferrin receptor." Thesis, McGill University, 1986. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=70354.
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The transferrin receptor is a membrane protein which is involved in the binding and endocytosis of transferrin, the main Fe$ sp{3+}$ carrying protein. The receptor and transferrin function to provide cells with iron, an essential nutrient. As red cells mature and lose their ability to make hemoglobin, they also lose their capacity to bind transferrin and take up iron. The transferrin receptor in sheep reticulocytes has been shown to undergo externalization in vesicular form during maturation of sheep reticulocytes in vitro. Microscopic studies show that the receptor is internalized into endosomes and externalization, in vesicular form, occurs via formation of multivesicular elements (or bodies), which ultimately fuse with plasma membranes releasing the inclusion vesicles bearing the transferrin receptor. The released receptor seems intact, having the same molecular weight and $ sp{125}$I-iodo-tyrosyl tryptic peptide map as the native receptor. The externalized receptor can also bind transferrin and the anti-receptor antibody.The transferrin receptor can be fatty acylated and phosphorylated in intact cells and in isolated plasma membranes. The phosphorylation of the receptor is not affected by the binding of transferrin or anti-transferrin receptor antibody. However, $ beta$-phorbol esters can stimulate the phosphorylation of the receptor suggesting that protein kinase C may be responsible for receptor phosphorylation. During reticulocyte maturation, the externalized transferrin receptor is not phosphorylated. Furthermore, the ability of this externalized receptor to undergo phosphorylation by protein kinase C is also lost, suggesting a maturation induced change in the receptor compared to the cell associated receptor. This change may be a signal which determines whether the transferrin receptor is to be segregated for externalization during red cell maturation or recycled for iron delivery.
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Boulton, Ian Charles. "In vitro characterisation of the meningococcal transferrin receptor." Thesis, Birkbeck (University of London), 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.266085.
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Stangland, Jenna Emily. "Biochemical Markers of Iron Status in Recreational Female Runners." The Ohio State University, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=osu1374168831.
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Luz, Henrique Luís Lopes Ferreira Reguengo da. "Ferro, Firritina, transferrina e receptores solúveis de transferrina em doentes com esclerose múltipla." Master's thesis, Faculdade de Farmácia da Universidade do Porto, 2008. http://hdl.handle.net/10216/20747.
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Luz, Henrique Luís Lopes Ferreira Reguengo da. "Ferro, Firritina, transferrina e receptores solúveis de transferrina em doentes com esclerose múltipla." Dissertação, Faculdade de Farmácia da Universidade do Porto, 2008. http://hdl.handle.net/10216/20747.
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Lok, Chun-nam, and 陸振南. "Regulation of transferrin receptor expression in human leukemic HL-60 cells: gene expression and cellular signaling." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 1996. http://hub.hku.hk/bib/B31235141.
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Lok, Chun-nam. "Regulation of transferrin receptor expression in human leukemic HL-60 cells : gene expression and cellular signaling /." Hong Kong : University of Hong Kong, 1996. http://sunzi.lib.hku.hk/hkuto/record.jsp?B17310659.
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Vieillevoye, Maud. "Role and expression of transferrin receptor 2 in erythropoiesis." Thesis, Paris 5, 2013. http://www.theses.fr/2013PA05S020.
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L’érythropoïèse est le processus de différentiation d’un progéniteur érythroïde multipotent en globules rouges. La différentiation érythroïde est essentiellement contrôlée par le récepteur à l’érythropoïétine (EPOR). Nous avons montré que le récepteur à la transferrine de type 2 (TFR2) est un membre important du complexe formé par l’EPOR. Le TFR2 présente, comme l’EPOR une expression restreinte qui dépend du type cellulaire. Ainsi son expression n’a pu être détectée que dans le foie, l’érythron et l’intestin grêle. Le rôle du TFR2 a été exploré dans les hépatocytes et il a été montré qu’il joue le rôle d’un senseur de fer dans cette lignée et de ce fait contribue à l’homéostasie du fer. Nous avons déterminé le rôle du TFR2 dans les érythroblastes et montré que TFR2 est une protéine escorte de l’EPOR qui contribue à l’érythropoïèse in vitro et in vivo. De plus, nos travaux montrent que le TFR2 est requis pour la production de GDF15 (Growth Differentiation Factor 15) dans les érythroblastes. D’autre part nous avons démontré que la production de GDF15 est augmentée par l’EPO, la déplétion intracellulaire en fer et l’activité transactivatrice de P53. L’inhibition de l’expression de P53, réalisée au cours de l’étude de son rôle dans la production de GDF15, a révélé son implication dans l’érythropoïèse normale. Nous avons mis en évidence l’existence de plusieurs formes du TFR2. Deux d’entre elles résultent de l’utilisation de sites distincts d’initiation de la traduction. Ces deux isoformes sont régulée différemment au cours de la maturation des érythroblastes. La troisième isoforme, appelée TFR2 soluble (sTFR2), est relargée dans le plasma suite au clivage du TFR2. Nous avons montré que la production du sTFR2 est inhibée en présence du ligand de TFR2, la transferrine saturée en fer (holoTF) alors que le TFR2 est stabilisé dans ces mêmes conditions. Les rôles spécifiques des trois formes du TFR2 doivent encore être élucidésErythropoiesis is the differentiation process of a multipotent erythroid progenitor into red blood cells. Erythroid differentiation is primarily controlled by the erythropoietin receptor (EPOR). We showed that the Transferrin receptor 2 (TFR2) is an important member of the EPOR complex. TFR2 has like EPOR a lineage-restricted expression and can solely be detected in the liver, erythron and small intestine. TFR2 function has been explored in hepatocytes where it plays the role of an iron sensor and contributes to iron homeostasis. We determined the role of TFR2 in erythroblasts and showed that TFR2 is an escort protein for EPOR that contributes to optimal erythropoiesis in vitro and in vivo. Moreover we evidenced that TFR2 is absolutely required for the production of Growth differentiation factor 15 (GDF15) in erythroblasts. We further demonstrated that GDF15 production is increased by EPO levels, by intracellular iron depletion as well as by P53 trans-activation activity. The inhibition of P53 expression, realized for the study of its role in GDF15 production, revealed its implication in normal erythropoiesis. We evidenced that TFR2 is expressed under several forms, two of which result from the utilization of distinct translational initiation sites. These two isoforms are differently regulated during erythroid maturation. The third form called soluble TFR2 (sTFR2) is released in the plasma after TFR2 cleavage. We showed that sTFR2 production is inhibited in the presence of TFR2 ligand, iron loaded transferrin (holoTF) whereas cell surface TFR2 expression is stabilized by holoTF. The specific roles of the three forms of TFR2 expressed by erythroblasts remain to be elucidated
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Yoon, Hye-Sun Melissa 1977. "Molecular cloning and characterization of mouse transferrin receptor 2." Thesis, McGill University, 2002. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=33861.
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Iron (Fe) plays an essential role in numerous metabolic processes. The transferrin receptor (TfR) is a cell membrane-associated protein that serves as a gatekeeper in regulating cellular uptake of Fe from transferrin (Tf) by TfR-mediated endocytosis. TfRs are expressed ubiquitously, but their highest levels of expression are on immature erythroid cells (which are the most avid consumers of Fe in the organism) and on rapidly dividing cells, both normal and malignant. In proliferating non-erythroid cells TfR expression is feedback inhibited by Fe through the interaction of specific binding proteins, iron regulatory proteins (IRPs), with iron responsive elements (IREs) in the 3 ' untranslated region (UTR) of TfR mRNA. However, results from our lab indicate that in differentiating murine erythroleukemia (MEL) cells, TfR expression responds only slightly to an increase in intracellular Fe. An explanation for this is lacking. Importantly, a second TfR (TfR2) has recently been cloned in humans. TfR2 is highly homologous with the classical receptor, TfR1 (except for the lack of IREs in the 3' UTR), and seems to have a similar function, but shows different tissue distribution. It has been speculated that TfR2 is expressed in erythroid cells since its message is abundant in K562 cells, a human cell line that can differentiate along an erythroid lineage, and can be induced to synthesize hemoglobin. Thus, it is important to determine whether TfR2 is expressed in erythroid cells, since this could possibly explain differences in TfR regulation in erythroid vs. non-erythroid cells. In this study, I have cloned mouse TfR2 (mTfR2) by RACE (rapid amplification of cDNA ends) and sequenced it. The sequence analysis shows approximately 50% and 82% homology to mouse TfR1 and human TfR2, respectively. Tissue distribution of mTfR2, based on Northern blot analysis indicated that, in the mouse, TfR2 mRNA was predominantly expressed in the liver and its level increased about 2 fold after a
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Kaur, Ishwinder. "Nuclear translocation and transferrin-transferrin receptor interaction of IPSE/[alpha}-1, a secretory glycoprotein from Schistosoma mansoni." Thesis, University of Nottingham, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.508222.
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Helminthic parasites have evolved with immune modulating machinery to manipulate their host's immune response and thus survive unscathed for years and in some cases even decades. However, the underlying molecular mechanisms governing the host-parasite relationship are still largely unknown. Therefore detailed investigation and evaluation of parasitic molecules is desirable. One such molecule worthy of attention is IPSE/a-1 (lnterleukin-4 inducing principle from schistosome eggs). IPSE/a-1 is a secretory glycoprotein produced exclusively by the egg stage of Schistosoma mansoni, which activates human basophils in non-antigen specific IgE dependent mechanisn;t. Sequence analyses of IPSE/a-1 using bioinformatic subcellular localisation prediction tools revealed a putative nuclear localisation signal (NLS) at the C-terminus. The present work was conducted to confirm whether this sequence ('125-PKRRRTY-131') was both necessary and sufficient for nuclear localisation of IPSE/a-1 and other heterologous GFP proteins. A plasmid encoding EGFP-IPSE/a-1, as well as a truncated mutant lacking amino acids 125-134, was transfected into Huh7 and U2-0S cell lines, and fluorescence of the fusion protein was determined by confocal laser scanning microscopy. EGFP-IPSE/a-1 was found to be exclusively nuclear, whereas the mutant displayed both nuclear and cytoplasmic staining. Furthermore, insertion of the IPSE/a-1 NLS into a tetra-EGFP construct showed nuclear localisation, and alanine scanning mutagenesis revealed a requirement for the KRRR residues. IPSE/a-l also binds to transferrin, which lead to downstream effect on cellular proliferation. Besides, fluorescence microscopy revealed that recombinant IPSE/a-l protein added exogenously to culture medium was internalized by variant Chinese hamster ovary (CRO) cells expressing the human transferrin receptor and was found in the nuclei of these cells Western blotting further confirmed this temporal relocalisation of IPSE/a-l from cytosolic to nuclear fractions. In addition, IPSE/a-l exhibited a DNA-binding activity that appeared to be dependent on the C-terminal NLS sequence. In summary, the main achievement of this work is the identification and characterization of a NLS in IPSE/a-l that is functional in mammalian cells, which will form the basis for further investigations into the biological significance of this nuclear targeting and DNA interaction e.g. IPSE/a-l may function as transcription factor in the nucleus. The properties of IPSE/a-l described here also make it an interesting potential vehicle for intracellular and nuclear drug delivery.
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Parsons, Tina. "Receptor-mediated iron and haem transport in Haemophilus." Thesis, University of Nottingham, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.282581.
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Sakamoto, Souichiro. "H-Ferritin Is Preferentially Incorporated by Human Erythroid Cells through Transferrin Receptor 1 in a Threshold-Dependent Manner." Kyoto University, 2016. http://hdl.handle.net/2433/215433.
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Giannetti, Anthony Michael Rees Douglas C. "Biochemical, biophysical, and cellular investigations of the interactions of transferrin receptor with transferrin and the hereditary hemochromatosis protein, HFE /." Diss., Pasadena, Calif. : California Institute of Technology, 2004. http://resolver.caltech.edu/CaltechETD:etd-05262004-173612.
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Dussiot-Abraham, Michaël. "Nouveaux acteurs contribuant à la régulation de l’érythropoïèse normale et inefficace : le récepteur à la transferrine et le récepteur à l'activine IIA." Thesis, Paris 11, 2013. http://www.theses.fr/2013PA11T030.
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L’érythropoïèse est le processus de formation des globules rouges. L’anémie demeure à l’heure actuelle un problème de santé publique majeur. Par conséquent, une meilleure compréhension des mécanismes impliqués dans le contrôle de ce processus dans des conditions physiologiques et pathologiques, ainsi que l’établissement de stratégies thérapeutiques ciblées constituent un enjeu de recherche majeur. Le récepteur de la transferrine 1 (CD71/RTf1) est un élément essentiel de l'érythropoïèse, la majorité des travaux de recherche étant focalisés sur son rôle indéniable dans le métabolisme du fer. Cependant, de nouveaux ligands du RTf1 ont été découverts ouvrant de nouvelles perspectives relatives aux fonctionnalités de ce récepteur. Ayant démontré que le RTf1 fixait les immunoglobulines A1 (IgA1), nous nous sommes intéressés au rôle des IgA1 dans l’érythropoïèse. Nous montrons que le RTf1 lié aux polymères d'IgA1 (pIgA1) induit la croissance et une augmentation de la prolifération des érythroblastes en concentration sous-optimale d'érythropoïétine (Epo). De même, l'expression transgénique d’IgA1 humaine (souris alpha1-KI), ou le traitement de souris de type sauvage avec les pIgA1 permettent une récupération accélérée de l’anémie aiguë. L’engagement du RTf1 module la sensibilité à l'Epo, en diminuant le seuil d'activation cellulaire, et en induisant les voies de signalisation MAPK/ERK et phosphatidylinositol-3-kinase (PI3K/AKT). Ces données mettent en évidence un nouveau rôle du RTf1 en tant que régulateur positif de l'érythropoïèse. Parallèlement au RTf1, nous avons identifié un autre récepteur pouvant constituer une cible thérapeutique pour corriger une érythropoïèse inefficace : le récepteur de l’activine de type IIA (ActRIIA). Dans un modèle murin de Beta-thalassémie intermédiaire (Hbbth1/th1), résultant d'une déficience génétique de la chaîne Beta de la globine, nous montrons que l'administration d'une protéine de fusion constituée du domaine extracellulaire de l’ActRIIA lié à un fragment Fc d’IgG de souris (RAP-011), corrige l'anémie, augmente le taux d'hémoglobine et diminue la splénomégalie. Ce traitement favorise l’érythropoïèse splénique et diminue la saturation de la transferrine et l’hémolyse. Fait intéressant, des niveaux élevés de GDF11 (Growth Differentiation Factor 11) sont observés sur des coupes spléniques de souris thalassémiques ainsi que dans le sérum de patients thalassémiques. In vivo, l’inhibition de l’interaction GDF11/ActRIIa par le RAP-011 favorise l’apoptose des érythroblastes précoces par la voie Fas/FasLigand. Ces résultats suggèrent que l’activation constitutive des signaux GDF11/ActRIIA contribue à l’établissement d’une érythropoïèse inefficace caractéristique de la Beta-thalassémie. La neutralisation de cette signalisation inverse ce processus. En conclusion, nos travaux ouvrent de nouvelles perspectives dans la compréhension de l'hématopoïèse normale et pathologique, et pourraient conduire à envisager des traitements innovants pour l'anémieAnemia produced by a variety of underlying causes is the most common disorder of the blood, and remains a major global public health problem associated with a poor quality of life for many patients. Thus, better understanding the erythroid process in physiological and pathological conditions, and developing new strategies to boost erythropoiesis appear of great interest. Transferrin receptor 1 (CD71/TfR1) plays an essential role in erythropoiesis, and investigations of TfR1 functions have been focused on their undeniable role in iron metabolism. However, recent data demonstrate that TfR1 is a multi-ligand receptor that participates in a wide array of cellular functions. We have identified TfR1 as a receptor for A1 isotype immunoglobulins (IgA1). In this work, we show that pIgA1s are able through their interaction with the TfR1, to stimulate erythropoiesis by sensitizing erythroblasts to Epo. Likewise, transgenic expression of human IgA1 (Alpha1-KI mice) or treatment of wild-type mice with pIgA1 accelerated recovery from acute anemia. TfR1 engagement by pIgA1 increased cell sensitivity to Epo by inducing activation of mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K) signaling pathways. These findings unveiled a new role of TfR1 as a signaling competent molecule positively regulating erythropoiesis. In addition to TfR1, our work identifies another receptor as a putative target for correcting ineffective erythropoiesis: the activin receptor IIA (ActRIIA). Indeed, using a mouse model of Beta-thalassemia intermedia (Hbbth1/th1) resulting from a genetic deficiency of Beta-globin chain, we show that administration of a ligand trap (named RAP-011), consisting in a fusion protein between the extracellular domain of ActRIIA and the Fc fragment of a mouse IgG, improves anemia, increases total hemoglobin levels and decreases splenomegaly. In addition, targeting ActRIIa signaling corrects ineffective erythropoiesis in the spleen, reduces hemolysis and transferrin saturation. Interestingly, high levels of Growth Differentiation Factor 11 (GDF11) are detected in spleen sections from Beta-thalassemic mice, as well as in sera from thalassemic patients. In addition, the inactivation of GDF11 promotes terminal erythroblast differentiation. Finally, blockade of the GDF11/ActRIIa signaling, promotes premature apoptosis of early erythroblasts through induction of Fas/FasLigand pathway. Therefore, these results first suggest that constitutive GDF11/ActRIIa signaling pathway may promote ineffective erythropoiesis in Beta-thalassemia intermedia, and secondly, support the use of ActRIIa traps for the treatment of chronic anemia and ineffective erythropoiesis. Altogether, these results open new perspectives in the understanding of normal and pathological hematopoiesis and lead to propose innovative treatments for anemia
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Chiu, Shih-Jiuan. "Receptor-mediated DNA-based therapeutics delivery." Columbus, Ohio : Ohio State University, 2005. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1127403022.
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Chan, Yuen-yee Roxanne. "Studies on receptor-mediated uptake of transferrin and iron acquisition by rabbit reticulocytes and a rat hepatoma cell line /." Hong Kong : University of Hong Kong, 1986. http://sunzi.lib.hku.hk/hkuto/record.jsp?B12325193.
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Rosendale, Morgane. "Visualisation et perturbation de la dynamique spatio-temporelle de l’endocytose." Thesis, Bordeaux, 2015. http://www.theses.fr/2015BORD0071/document.
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L’endocytose dépendante de la clathrine (EDC) est un processus fondamental des cellules eucaryotes. Elle se caractérise par la formation d’invaginations à la membrane plasmique aboutissant à la création de petites vésicules par l’action de la dynamine. Dans le cerveau, elle est impliquée dans la dépression synaptique à long terme, un corrélat cellulaire de la mémoire. La morphologie complexe des neurones et le contrôle précis du code neuronal suggèrent qu’elle puisse être régulée spatialement et temporellement dans ces cellules. Le but de mon travail a été de développer de nouveaux outils pour visualiser et perturber l’EDC afin d’étudier ce type de régulation. Le premier de ces outils est pHuji, un senseur de pH rouge génétiquement encodable. Je l’ai utilisé avec un senseur de pH vert existant pour montrer que dans les cellules NIH- 3T3, le récepteur β2-adrénergique est internalisé dans une sous-population de vésicules contenant le récepteur à la transferrine constitutivement endocyté. Le deuxième est une nouvelle méthode d’imagerie permettant de visualiser l’activité d’endocytose de structures recouvertes de clathrine optiquement stables dans des neurones d’hippocampe. J’ai ainsi pu suivre pour la première fois la cinétique d’internalisation de récepteurs au glutamate de type AMPA dans des conditions de plasticité. Enfin, j'ai élaboré un test combinant imagerie et patch-clamp afin de développer un bloqueur peptidique spécifique de l'EDC. En utilisant des peptides dimériques, j’ai montré que la dynamine se lie à ses partenaires via des interactions multimériques. En conclusion, ce travail propose une boite à outils permettant d’élucider les mécanismes de l’EDC avec une grande résolution spatiale et temporelleClathrin mediated endocytosis (CME) is a fundamental process of all eukaryotic cells. At the level of the plasma membrane, it is characterized by the formation of deep invaginations resulting in the creation of small vesicles after membrane scission by dynamin. In the central nervous system, it is involved in the expression of synaptic long term depression, a proposed cellular correlate of learning and memory. The complex morphology of neurons and the precise timing of neuronal firing suggest that endocytosis may be spatially and temporally regulated in those cells. The aim of the work presented here was to develop new tools to visualize and perturb CME in order to study such regulation. The first tool to be characterized was pHuji, a genetically encoded red pH-sensor. I used it in combination with an existing green pHsensor to demonstrate that in NIH-3T3 cells, the β2-adrenergic receptor was internalized in a subset of vesicles containing the constitutively endocytosed transferrin receptor. The second tool is a new imaging method that allowed me to monitor the endocytic activity of optically stable clathrin coated structures in hippocampal neurons. I was thus able to visualize for the first time the kinetics of internalization of AMPA-type glutamate receptors under plasticity inducing conditions. Finally, I set up an assay combining imaging and cell dialysis in order to develop a specific peptide-based inhibitor of CME. Using dimeric peptides, I found that the interplay between dynamin and its binding partners relies on multimeric interactions. Altogether, this work provides a toolbox to decipher the mechanisms of vesicle formation with high spatial and temporal resolution
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Nada, Dina. "Gallium-68 and fluorine-18 labeling of a peptide binding to the human transferrin receptor and determination of its uptake into transferrin receptor-expressing human cell lines." Thesis, McGill University, 2010. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=87014.
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An often proposed mechanism to overcome the system of efflux transporters and tight junctions present at the blood brain barrier (BBB) is the receptor-mediated transport via endocytosis. Among the proposed transport vehicles, the transferrin receptor (TfR) has been considered to be one of the most attractive targets. Besides transferrin itself and the antibody OX26 (anti-rat TfR-antibody), it has recently been shown that peptides are also able to bind to the transferrin receptor and are internalized into TfR expressing cells. In this study we evaluated the ability of the 68Ga-labeled peptide THRPPMWSPVWP to become internalized into human TfR bearing cells to pre-evaluate its potential to act as a carrier system for small molecules across the BBB utilizing the TfR receptor located on the endothelial cells. To prove the validity of this concept we first synthesized the peptide THRPPMWSPVWP, and then a conjugate consisting of the D2 receptor ligand fallypride and the peptide which was synthesized by our collaborators and evaluated as to its binding affinity towards the D2 receptor. The conjugate still showed a high receptor affinity (Ki = 27 nM) despite the strong structural alteration exerted by the peptide.For radioactive-labeling, the reported TfR-binding peptide THRPPMWSPVWP was derivatized with p-SCN-Bn-NOTA. It was then labelled with 68Ga by our collaborators. The uptake of the 68Ga-labelled peptide into TfR-bearing cells was investigated using two human TfR-expressing cell lines, the human glioblastoma cell line U87MG and the human colon adenocarcinoma cell line HT-29 to assess the peptides' potential as a carrier molecule targeting the TfR receptor . The binding was found to be less than 1% after 60 min incubation at 37°C.
These results suggest that the concept of using peptide THRPPMWSPVWP to act as a carrier system for short lived PET radio-pharmaceuticals across the BBB within a reasonable time span is doubtful. However, further experiments have to be carried out to determine the potential of this peptide as transferrin receptor targeting carrier molecule for therapeutic agents.
Besides, and owing to the need for optimization of the synthetic procedures for 18F labeling of biologically-active peptides for clinical PET, we also investigated the possibility to synthesize a glucose based labeling synthon 2,3,4-tri-O-acetyl-1-azido-6-[18F]fluoro-β-D-glucopyranoside, to radiolabel alkyne derivatized peptides with 18F using click chemistry. Our strategy would combine the advantages of carbohydration of peptides with the fast and efficient click reaction. We synthesized 2,3,4-tri-O-acetyl-1-azido-6-O-p-toluene sulfonyl-β-D- glucopyranoside in an overall chemical yield of 80%, which served as a labeling precursor bearing an azide group ready for coupling to an alkyne derivative of a biologically active peptide by click chemistry and a p-toluene sulfonyl group ready for radio-labeling with 18F by a nucleophilic substitution reaction. The second compound was 2,3,4-tri-O-acetyl-1-azido-6-fluoro-β-D-glucopyranoside in an overall chemical yield of 77%, which served as a reference compound to establish HPLC and TLC conditions for the radio-labeling experiments. Initial labeling experiments with nucleophilic 18F did not yield the desired 18F-labeled glucose derivative so far. We are currently taking efforts to radiolabel the precursor with 18F buy re-crystallizing the precursor several times as it was proven by HPLC that it still contains some impurities. These impurities can compromise the intended labeling reaction with 18F.
Un mécanisme proposé expliquant le transport par efflux présent dans la 'blood brain barrier' consiste en un système de transport membranaire assisté faisant intervenir des récepteurs spécifiques, par d'endocytose. Parmi les vésicules de transport proposées, les récepteurs de transferrine (TfR) sont considérés comme les plus attractives. A part la transferrine et l'anticorps pour clone MRC OX-26 (anticorps TfR anti-rat), il a été montré que des peptides sont aussi capables de se lier spécifiquement au récepteur de la transferrine, puis d'y être internalisé dans les cellules possédant les récepteurs à transferrine. Dans cette étude, nous avons évalué la possibilité du peptide 'THRPPMWSPVWP' marqué avec du 68G de s'internaliser dans des cellules humaines présentant les récepteurs TfR. Nous avons aussi évalué son potentiel de se comporter comme vecteur de transport de petites molécules à travers la 'blood brain barrier'. Nous avons utilisé des cellules endothéliales avec des récepteurs TfR. Pour prouver la validité du concept, nous avons synthétisé le peptide THRPPMWSPVWP, puis conjugué au fallypride de type D2, qui est un ligand avec une grande affinité pour les récepteurs du TfR. L'ensemble a été évalué par nos collaborateurs afin de vérifier l'affinité de liaison vis-à-vis des récepteurs du TfR. Ce conjugué à montré une grande affinité précités (Ki = 27 nM), malgré de fortes altérations au niveau de sa structure causées par la présence du peptide. Pour le marquage radioactif, le peptide THRPPMWSPVWP bioconjugué ci-dessus a été dérivé avec le p-SCN-Bn-NOTA et radio-marqué au 68Ga par nos collaborateurs. Les propriétés de ce peptide ont été étudiées en utilisant deux lignées cellulaires différentes présentant les récepteurs TfR, soit des cellules de type tumoral glial (U-87 MG) et des cellules carcinomes de côlon humain (HT-29), afin d'étudier le potentiel du peptide à se comporter comm
D'autre part, du fait de l'optimisation des procédures de synthèse pour le radio marquage au 18F de peptides biologiques actifs, dans le cadre d'études cliniques de 'PET', nous avons envisagé de synthétiser un nouveau synthon radio-marqué dérivant du glucose, le 2,3,4-tri-O-acetyl-1-azido-6-[18F]fluoro-β-D-glucopyranoside, pour radio-marquer des dérivés de type 'alkyne', par la technique de 'click chemistry'. Notre stratégie combine les avantages du processus de 'carbohydratation' des peptides et de la réaction de 'click chemistry'. Nous avons synthétisé le 2,3,4-tri-O-acetyl-1-azido-6-O- p-toluene sulfonyl β-D-glucopyranoside avec un rendement de 80%. Ce dernier possède un groupe p-toluène sulfonyle qui sert de précurseur pour radio-marquer la molécule au 18F par une réaction de substitution nucléophile, et présente également un groupement 'azide', permettant le couplage rapide avec un peptide actif possédant une fonction 'alkyne'. Le 2eme composé préparé est le 2,3,4-tri-O-acetyl-1-azido-6-fluoro-β-D-glucopyranoside, avec un rendement de 77%. Ce dernier servant de référence pour établir les conditions HPLC et par CCM nécessaires au radio-marquage. Les études préliminaires avec le 18F comme nucléophile n'ont pas conduit au composé de type 'glucose' radio-marqué au 18F. Nos efforts actuels se portent sur le radio-marquage du précurseur au 18F en recristallisant ce dernier. Il a été montré par HPLC que ce dernier contient des impuretés, qui empêcheraient le radio-marquage au 18F.
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Jullié, Damien. "Les endosomes de recyclage fusionnent transitoirement avec la membrane plasmique des dendrites neuronales." Thesis, Bordeaux 2, 2012. http://www.theses.fr/2012BOR21942/document.
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Le trafic membranaire est essentiel dans les neurones pour la morphogénèse, le recyclage des vésicules synaptiques et des récepteurs. L’exocytose des récepteurs AMPA contenus dans les endosomes de recyclage (ER) est nécessaire pour l’expression de la plasticité synaptique à long terme (PLT). Pour étudier ce mécanisme, nous avons visualisé l’exocytose par microscopie de fluorescence sur des neurones en culture transfectés avec le récepteur de la transferrine (TfR), un marqueur des ER, fusionné à la phluorine. Un examen systématique des événements d'exocytose a révélé des différences de comportement. Dans la plupart des cas, les récepteurs diffusent rapidement dans la membrane plasmique après exocytose (discharge), mais dans environ 25% des cas, les récepteurs restent concentrés (display). L’utilisation de changements rapides de pH extracellulaire autour de la cellule montre que l’exocytose est transitoire : après quelques secondes (médiane 2.6s) les récepteurs sont réinternalisés. Ce mécanisme a pu être étendu aux récepteurs AMPA et β2-adrenergique, pour lesquels l’exocytose de type display avait déjà été décrite. L’imagerie deux couleurs montre que Rab11, un marqueur des ER, est enrichie au site d’exocytose. L’expression d’un dominant négatif de Rab11 connu pour inhiber la PLT provoque une diminution spécifique de la fréquence des évènements discharge. Dans nos recherches sur le mécanisme de l’exocytose, nous avons testé l’implication des protéines SNARE dans la fusion membranaire. Ainsi VAMP4 est enrichie avec le TfR dans les ER qui sont exocytés à une fréquence équivalente. De plus, elle est requise pour le recyclage du TfRMembrane trafficking is essential for neuronal function: from growth of neurons and synapse formation to recycling of synaptic vesicles and receptors, questions concerning exocytosis and endocytosis are stimulating neurobiology research. In particular, trafficking of glutamate receptors present in recycling endosomes (REs) is necessary for the expression of long term potentiation (LTP). To investigate the mechanism of exocytosis in dendrites, we have imaged cultured rat hippocampal neurons transfected with transferrin receptor, a classical marker of REs, tagged with phluorin. As for AMPA receptors or β2-adrenegric receptors, single exocytic events has revealed two main behaviors: in most cases, receptors diffuse quickly in the plasma membrane after exocytosis (discharge events), but receptors can also remain clustered (display events). Using fast extracellular pH changes around the recorded cell, we show that for display events exocytosis is transient: after a few seconds (median 2.6 s) receptors are internalized. Moreover, using two color imaging of single exocytosis events with markers of neuronal compartments, we found that Rab11 is enriched at the exocytosis site, confirming the endosomal origin of the vesicles. Overexpression of a dominant negative form of Rab11 known to impair LTP decreases selectively the frequency of discharge events. As SNARE proteins are involved in virtually all membrane fusion processes, we investigated the role of Vamp proteins in somatodendritic exocytosis events. We found that Vamp4, unlike Vamp2 or Vamp7, is enriched in TfR containing compartments and can undergo exocytosis at high frequency and is required for TfR exocytosis
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Byrne, Shaina. "Investigation of the Molecular Basis of Receptor Mediated Iron Release from Transferrin." ScholarWorks @ UVM, 2009. http://scholarworks.uvm.edu/graddis/38.
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Human serum transferrin (hTF) is a bilobal glycoprotein that plays a central role in iron metabolism. Each lobe of hTF (N- and C-lobe) can reversibly bind a single ferric iron. Iron binds to hTF at neutral pH in the plasma; diferric hTF binds to specific hTF receptors (TFR) on the cell surface and the complex undergoes receptor mediated endocytosis. The pH within the endosome is lowered to ~5.6 and iron is released from hTF. Apo hTF remains bound to the TFR and recycles back to the cell surface. Upon fusion with the plasma membrane, apo hTF dissociates from the TFR and is free to bind more iron and continue the cycle. The iron release process is complicated by various factors which include pH, anions, a chelator, lobe-lobe cooperativity and interaction with the TFR. All of these influence iron release in a complex manner. Because they are intricately linked, it is difficult to determine the effect of any single parameter. We have utilized stopped-flow and steady-state fluorescence and urea gel electrophoresis to dissect the iron release process as a function of lobe-lobe interactions, the presence of the TFR, and changes in pH and salt concentration. Application of recombinant protein production and site-directed mutagenesis has allowed us to generate a variety of hTF constructs in which the iron status of each lobe is completely controlled. Thus, we have created authentic monoferric hTFs unable to bind iron in one lobe, diferric hTFs with iron locked in one lobe and diferric hTF in which iron can be removed from both lobes. Importantly, we have produced the soluble portion of the TFR (sTFR) to analyze interactions between hTF and the sTFR and to monitor iron release from hTF/sTFR complexes. Together, we are able to provide a more precise picture of iron release from the two lobes of hTF in the presence and absence of the TFR. Steady-state fluorescence emission scans and urea gel electrophoresis provide a qualitative evaluation of the iron status of each construct after a predetermined incubation in iron removal buffer (i.e. an endpoint). However, these techniques do not provide information regarding the kinetic pathway to reach that endpoint. Combined with stopped-flow fluorescence time-based kinetics, a more precise assessment of the iron release process has been obtained. We have determined that changes in pH and salt affect endpoint iron release from the C-lobe, but not the N-lobe, however, the kinetics of iron release from both lobes are highly sensitive to pH and salt. Kinetic analysis in the absence and presence of the sTFR reveals the complexity of the iron release process. In the absence of the sTFR, the kinetics of iron release are insensitive to the iron status of the opposite lobe. However, in the presence of the sTFR, the kinetics of iron release from both lobes are affected by the iron status of the opposite lobe. Determination of conformational changes induced by anion binding, lobe-lobe communication and sTFR interactions have now been confidently assigned. We have created kinetic models of iron release from diferric hTF ± the sTFR and incorporated specific events pertaining to anion binding, lobe-lobe communication and conformational changes associated with sTFR interactions. We provide irrefutable evidence that a critical role of the sTFR is to accelerate the rate of iron release from the C-lobe, while decreasing the rate of iron release from the N-lobe such that the two lobes effectively release iron on a time scale relevant to one cycle of endocytosis.
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Petrie, Robert G. "Receptor-mediated endocytosis of testicular transferrin by germinal cells of the rat testis." Thesis, McGill University, 1991. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=60453.
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The present study examines events of the Sertoli cell iron delivery pathway following the secretion of diferric testicular transferrin (tTf) in to the adluminal compartment of the rat seminiferous epithelium. The unidirectional secretion of tTf by Sertoli cells was verified, in vivo, and it was shown that this protein is internalized by adluminal germ cells. It was further determined by Scatchard analysis that this internalization was mediated by high affinity transferrin binding sites on the surface of round spermatids, numbering 1453/cell and displaying a K$ sb{ rm d}$ = 0.6 $ times$ 10$ sp{-9}$ M. Northern blot analysis of RNA isolated from adluminal germ cells, namely spermatocytes, round spermatids and elongating spermatids, indicated that these cells expressed Tf receptor mRNA and ferritin mRNA in levels inversely related to their stage of maturation. Finally it was determined that following binding and internalization in round spermatids, Tf became associated with the endosomal compartment and was recycled back to the cell surface. Diferric tTf binds to Tf receptor on the surface of adluminal germ cells, is internalized by receptor-mediated endocytosis and the apotTf-Tf receptor complex is recycled back to the cell surface where apotTf is released into the adlumenal fluid.
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Ahn, Jinhi. "Transferrin receptor expression in the sheep reticulocyte : biosynthesis and fate during reticulocyte maturation." Thesis, McGill University, 1992. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=39437.
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The transferrin receptor (TfR) is a membrane protein whose function is to deliver transferrin-bound iron into the cell. Erythroid precursor cells and reticulocytes require huge amounts of iron for hemoglobin synthesis and express large numbers of TfRs. However, as the reticulocyte matures into an erythrocyte, TfRs are lost from the cell, as are a number of other membrane proteins. Nearly a decade ago, it was shown that TfRs are released from the sheep reticulocyte in vesicular form (in exosomes) during culture. Exosome formation may be the mechanism for remodelling of the plasma membrane during reticulocyte maturation. In this thesis, we determine whether the sheep reticulocyte has the capacity to sythesize TfRs and whether TfR biosynthesis or externalization is controlled directly by iron. Although the sheep reticulocyte incorporates ($ sp{35}$S) methionine into the TfR, the labelled TfRs are incompletely glycosylated and are not transported to the cell surface. TfR synthesis is regulated by heme, rather than directly by iron. Newly synthesized TfRs are not externalized; however, pulse-chase experiments showed that they are lost slowly by an iron-dependent mechanism. Externalization of preexisting TfRs is stimulated by hemin but not iron. We propose that hemin catalyzes an oxidative change in the TfR which may facilitate the removal of the TfR from the recycling pathway and allow it to be targeted to multivesicular bodies and ultimately released in exosomes. We suggest that the TfR ectodomain can be cleaved from the released vesicles and propose that exosomal membranes are the source of the soluble truncated TfRs found in the circulation.
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Reutens, Gail. "Effect of a mutation in transferrin receptor 2 on the uptake of iron." Thesis, Reutens, Gail (2005) Effect of a mutation in transferrin receptor 2 on the uptake of iron. Honours thesis, Murdoch University, 2005. https://researchrepository.murdoch.edu.au/id/eprint/52266/.
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Hereditary haemochromatosis is an iron overload disorder associated with misregulation of iron metabolism. Subtype III arises from mutations in the transferrin receptor 2 {TfR2) gene. TfR2 is thought to mediate transferrin bound iron (TBI) uptake by a lowaffinity receptor-mediated endocytic process. The V22F mutation in TfR2 is situated adjacent to the putative TfR2 endocytosis signal and results in a valine to phenylalanine substitution and has been shown to cause haemochromatosis.
The aims of this study were to investigate the functional effects of the V22F mutation on the uptake of iron. To study the functional effects of TfR2V22F on cellular iron uptake, the V22F mutation (TfR2inF) was generated by site-directed mutagenesis and transfected into the Chinese Hamster Ovary (CHO-TRVb) cell line that is deficient in endogenous transferrin receptors. To confirm the subcellular localisation of TfR2, cells containing each construct were subjected to immunocytochemistry using a specific antibody, before being visualised by fluorescence microscopy. The distribution of TfR2 V22F and TfR2WT was predominantly on the cellular surface, consistent with being a surface receptor.
Cells were incubated with l25I-Tf-59Fe and treated with Pronase to separate into membrane-bound and internalised fractions. Higher levels of surface and internalised transferrin and iron were observed in cells containing TfR2WT compared with TfR2VEC, signifying that TfR2 mediates TBI uptake. Mutant cells exhibited increased TBI uptake. suggesting that the mutation causes an up-regulation in TBI uptake compared with TfR2WT. To investigate the effects of the mutation on TfR2 cycling, the endocytic and exocytic rates were determined. Cells were incubated with different concentrations of transferrin (0.1 to 10|iM) to measure endocytosis. On increasing concentrations, transferrin internalisation in mutants and wild-type was saturable, consistent with a receptor-mediated process. Similar Kms were obtained for CHO-TRVbV22F. and CHO-TRVwt , however, the mutants exhibited a 1.7-fold higher Kmax, suggestive of an increased rate of endocytosis. Similarly, after loading cells with 125l-Tf-59Fe, followed by incubation in efflux medium, CHO-TRVbV22F demonstrated a 1.6-fold higher rate of transferrin release when compared to CHO-TRYbVVT. Rates of iron release were similar between cells, showing the mutation does not affect iron release but increases the exocytic rate of transferrin
When cells were incubated with non-transferrin bound iron (NTBI) in the form of ^Feaccumulated low levels of NTBI in contrast to cells expressing and TfR2WT, suggesting that TfR2 mediates NTBI uptake. The rate of uptake was markedly higher in the mutants than wild-type, which implies that the V22F citrate, CHO-TRVbVEC TfR2V22F mutation induces an increased rate of uptake of NTBI over and above that apparent in CHO-TRVbWT.
The results suggest that cells containing TfR2V22F, in comparison to cells containing ^ WT TfR2 , have an enhanced ability to internalise TBI and NTBI in vitro. These findings are consistent with the notion that the V22F mutation affects TfR2-mediated endocytosis, leading to type III haemochromatosis in vivo.
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Chiu, Shihjiuan. "Receptor-mediated DNA-based therapeutics delivery." The Ohio State University, 2005. http://rave.ohiolink.edu/etdc/view?acc_num=osu1127403022.
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Li, Jing. "Studies on iron-dependent regulation of transferrin receptor gene expression in chicken HD3 cells." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp04/mq29742.pdf.
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Li, Jing 1957. "Studies on iron-dependent regulation of transferrin receptor gene expression in chicken HD3 cells." Thesis, McGill University, 1996. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=27368.
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Expression of the transferrin receptor (TfR) gene is developmentally regulated in mammalian cells (1), an iron-dependent post-transcription regulatory mechanism has been demonstrated in non-erythroid cell (2,3). In order to understand the molecular mechanism underlying the iron-dependent regulation of the TfR gene expression in the differentiating erythroid cell, a virus-transformed chicken erythroblast cell line, HD3 was employed in this study. We have investigated the possible effects of iron on the TfR gene expression that might involve regulatory mechanisms at the transcriptional and/or the post-transcriptional level during the HD3 cell differentiation.We found that treatment of the differentiating HD3 cell with Pyridoxal Isonicotinoyl Hydrazone (PIH) causes an increase in TfR mRNA accumulation as a result of increased TfR gene transcription. No evidence was obtained for stabilization of TfR mRNA. These data indicate that there may be another distinct iron-dependent regulatory mechanism for the TfR gene expression during erythroid cell differentiation.
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陳遠儀 and Yuen-yee Roxanne Chan. "Studies on receptor-mediated uptake of transferrin and iron acquisition by rabbit reticulocytes and a rat hepatoma cell line." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 1986. http://hub.hku.hk/bib/B31207571.
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Suarez, Ortegon Milton Fabian. "Markers of iron status and cardiometabolic disease risk : an exploration of the association based on cross-sectional and prospective studies in multiple populations." Thesis, University of Edinburgh, 2017. http://hdl.handle.net/1842/31470.
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The aim of this thesis is to contribute to the understanding of iron metabolism, as a factor associated with cardiometabolic risk, by undertaking secondary data analyses. The objectives were to identify gaps in existing knowledge in terms of populations studied and alternative iron markers, and to attempt to fill the gaps with additional analyses and interpretation. Serum ferritin was the most widely available measure of iron status but the role of serum transferrin and soluble transferrin receptor (sTfR) levels was considered where available. I have taken a life-course approach with analyses in childhood and adulthood, and have included both intermediate factors such as the metabolic syndrome (MetS), and disease diagnoses of diabetes and cardiovascular disease as outcomes. Chapter one presents a review of empirical research literature on the relationship between iron metabolism and cardiometabolic risk, concepts surrounding iron markers and the study outcomes. This chapter also describes the gaps in understanding the iron-cardiometabolic risk relationship, which are subsequently explored in chapters two to six. Chapter two explores the link between serum ferritin and transferrin and MetS in cross-sectional and prospective studies of 725 Spanish children and 567 Chilean adolescents. I found associations between both ends of the ferritin distribution and MetS or glucose metabolism markers in different paediatric populations. For instance, whereas in the Spanish children there was a decrease of 0.02 SD units in the change of MetS score over time for every SD unit increase in ferritin, in the Chilean male adolescents being in the highest tertile of ferritin (v. the lowest) was associated with an increase of 0.25 SD units of MetS score. Furthermore, sustained high ferritin levels at various time points and gradual increase of ferritin during childhood were associated with higher MetS score in adolescence. The third chapter describes the association between serum ferritin status and MetS in adults in two cross-sectional studies of Scottish populations (2,047 individuals from Shetland Islands and 8,563 subjects from the Scottish Health Surveys (SHeS) 1995- 1998). I also examined the overall association between ferritin, MetS and each MetS component in adults, by conducting a meta-analysis and investigating potential relevant sources of heterogeneity for the association. Interestingly, ferritin levels were positively associated with MetS in the Scottish populations, but the association was not independent of the effect of covariates, mainly body mass index (BMI) and transaminase levels [Men Odds ratio (OR) 95% confidence interval (CI) 1.43(0.83- 2.46); Postmenopausal women OR (95%CI) 1.09(0.62-1.90); Premenopausal women OR (95%CI) 1.02(0.42-2.46), P > 0.05]. The meta-analysis supported this finding by describing hepatic injury markers and BMI as the major attenuating factors of the ferritin-MetS association. Chapter four investigates the association between sTfR or ferritin, and MetS in 725 Croatian adults in a cross-sectional study. There was no evidence of an association between sTfR and MetS [Men OR (95%CI) 1.35(0.90-2.02); Postmenopausal women OR (95%CI) 0.73(0.47-1.15); Premenopausal women OR (95%CI) 0.87(0.66-1.17), P > 0.05]. In contrast serum ferritin, was positively and independently associated with MetS in men and postmenopausal women (P < 0.05) [Men OR (95%CI) 1.78(1.31- 2.42); Postmenopausal women OR (95%CI) 1.71(1.12-2.62); Premenopausal women OR (95%CI) 1.24(0.85-1.80)]. These contrasting results suggest that different iron markers reflect different physiological processes other than iron metabolism. Chapter five evaluates the longitudinal association between serum ferritin and several cardiometabolic disease outcomes (CMDs) in the nationally representative SHeS 1995 and 1998 (n = 6,497). I found an independent positive longitudinal association between ferritin and cerebrovascular disease (CEVD), which was strengthened by using higher cut-points for increased ferritin [higher v. lowest sextile fully adjusted Hazard ratio(HR) 95%CI 2.08 (1.09-3.94), P=0.024], and a not significant association with coronary heart disease (CHD) after adjustment for covariates. My analyses confirmed the widely established association with type 2 diabetes (T2D) [whole sample fully adjusted HR 95% CI 1.59(1.10-2.34), P=0.006], even with serum ferritin within the normal range. The above set of observations confirm ferritin as biomarker mainly related to the development of T2D and identifies the need to investigate the association between ferritin and CEVD in other populations. Chapter six investigates whether ferritin is associated with risk for cardiovascular complications among people with T2D using cross-sectional study designs in two populations with differing baseline cardiovascular risk (Spanish study SIDIAP n=38,617) and (Edinburgh Type 2 Diabetes Study (ET2DS) n= 821) with additional analysis of follow-up data for ET2DS. Interestingly, ferritin levels were negatively associated with prevalence of cardiovascular disease, mainly CHD, in people with T2D in both studies [ET2DS OR (95%CI): 0.80(0.67-0.96), P=0.020; SIDIAP study: 0.85(0.83-0.88), P < 0.001). Ferritin was also negatively associated with incident cardiovascular disease in ET2DS: HR 95% CI: 0.39(0.16-0.93), P=0.035. Therefore, the association between iron status and CMD risk in people with T2D appears to differ from that in general populations in which a positive association has been more commonly described. In conclusion, serum ferritin is associated with cardiometabolic risk in different ways in a variety of populations. Inconsistent associations for other iron markers suggest that iron biomarkers reflect factors other than iron homeostasis that influence cardiometabolic risk. The association between iron markers and MetS appears to differ between populations. This thesis illustrates the complex relationship between iron metabolism markers, MetS and CMD, and identifies the need for further research on the topic in order to extend knowledge about pathophysiology and the potential for measures of iron status as biomarkers for CMD.
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Yoshinaga, Masanori. "Regnase-1 Maintains Iron Homeostasis via the Degradation of Transferrin Receptor 1 and Prolyl-Hydroxylase-Domain-Containing Protein 3 mRNAs." Kyoto University, 2020. http://hdl.handle.net/2433/253191.
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Papista, Christina. "Study of IgA receptors and transglutaminase 2 in IgA nephropathy and coeliac disease." Paris 7, 2013. http://www.theses.fr/2013PA077014.
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La néphropathie à IgA (NIgA), une cause majeure d'insuffisance rénale dans le monde, est caractérisée par une pathogenèse complexe, incluant des facteurs inconnus favorisant la formation des complexes d'IgAl dans la circulation et des dépôts de ces complexes dans le mésangium. Des anomalies des récepteurs aux IgA sont impliquées: de complexes circulants d'IgAl avec la forme soluble (s) du récepteur myéloïde CD89 et la surexpression du récepteur mesangial aux IgAl le TfRl (récepteur de la transferrine 1). Dans la première partie de cette thèse, en utilisant un nouveau model murin de NIgA exprimant à la fois PIgAl et le CD89 humains, nous avons démontré que les complexe IgAl-CD89s déclenchent une procédure pathogénique incluant la surexpression de TfRl et de l'enzyme transglutaminase2 (TGase2), permettant des dépôts mésangiaux de complexes d'IgAl et l'activation cellulaire. Des antigènes alimentaires, notamment le gluten, ont été associés avec la NIgA sans que le mécanisme soit connu. Avec u modèle murin innovant: des souris sensibilisées au gluten, développé dans la deuxième partie de cette étude, démontrant le rôle des IgA, TGase2, TfRl et de microbiotes intestinaux dans la maladie cœliaque induits par gluten, j'ai conclu cette thèse par une troisième partie axée sur l'effet du gluten dans la NIgA. J'ai montré que le gluten associé au CD89, est impliqué dans la formation des complexes IgAl-CD89s et dans la réponse mucosale IgAl exacerbée, induisant le développement de la NIgA. Des marqueurs cruciaux: CD89, TfRl, TGase2 et gluten, émergent de cette thèse, corrèlant la NIgA avec la maladie cœliaque et offrant de nouvelles cibles thérapeutiques aux deux pathologiesIgA nephropathy (IgAN), a major cause of renal failure Worldwide, is characterized by a complex pathogenesis, which consists of unknown factors favoring formation of macromolecular IgAl complexes in the circulation and complex deposition in the mesangium. IgA receptor abnormalities are implicated, including circulating complexes of IgAl with the soluble (s) form of the myeloid receptor CD89 and over-expression of the mesangial IgAl receptor TfRl (transferrin receptor 1). In the first part of this thesis, using a new IgAN mouse model expressing both human IgAl and CD89, w demonstrated that IgAl-sCD89 complexes initiated a process of auto-amplification involving over-expression of TfR and the cross-linking enzyme transglutaminase2 (TGase2), allowing increased mesangial deposition of pathogenic IgA complexes and chronic mesangial cell activation. Food antigens, notably gluten, are also associated with IgAN onset, bi the mechanism is still unknown. Inspired by a novel model of gluten-sensitized mice developed in the second part of this study showing the role of IgA, TGase2, TfRl and the intestinal microbiota in the induction of immune responses an coeliac-like disease induced by gluten, I concluded this thesis with a third part focused on the effect of gluten in IgA1 using the same sensitization method. I showed that gluten in association with CD89, was implicated in IgAl-sCD8 complex formation and exacerbating IgAl mucosal response, resulting in a breakdown of oral tolerance and IgA1 development. Crucial players: CD89, TfRl, TGase2 and gluten, emerge from this thesis, affecting both IgAN and coeliac disease and proposing new therapeutic targets for both pathologies
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Guimarães, Jacqueline da Silva. "Alterações do metabolismo do ferro nas talassemias." Universidade de São Paulo, 2014. http://www.teses.usp.br/teses/disponiveis/60/60135/tde-17042015-113612/.
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As síndromes talassêmicas (?- e ?-talassemia) são as desordens mais comuns e frequentes associadas com eritropoese ineficaz. O desbalanço na produção das cadeias ?- e ?-globinas resulta no comprometimento da produção de eritrócitos, em anemia e aumento de progenitores eritroides no sangue periférico. Enquanto os pacientes homozigóticos afetados por essas desordens demonstram alterações características dos parâmetros relacionados a eritropoese, a relação entre grau de anemia, eritropoese alterada e disfunção do metabolismo de ferro ainda não foram investigados nos indivíduos com ?+-talassemia heterozigótica ou ?+-talassêmia. Duzentos e vinte seis indivíduos (75 do gênero feminino e 151 do gênero masculino) foram recrutados e divididos em 5 grupos: Controle (n=28), doadores de sangue regulares (DSR, n=23), ?+-talassemia heterozigótica (TAT, n=14), ?+-thalassemia (traço ?-talassêmico, TBT, n=20) e ?0-talassemia, (?-talassemia maior, BTM, n=27). As amostras foram analisadas para parâmetros hematológicos (Micros ABX 60); ferro sérico, capacidade total de ligação ao ferro e saturação de transferrina por método colorimétrico (Pointe Scientific, Inc., Canton, MI, USA), ferritina e proteína C-reativa ultra sensível por imunoensaio (Immulite 1000); receptor solúvel de transferrina, eritropoetina, fator de diferenciação do crescimento 15 (R&D Systems) e hepcidina (Intrinsic LifeSciences, La Jolla, CA) por ELISA. As razões sTfR/log ferritina e (hepcidina/ferritina)/sTfR foram calculadas para avaliar o metabolismo do ferro. sTfR/log ferritina pode distinguir depleção dos estoques de ferro de eritropoese deficiente de ferro, enquanto (hepcidina/ferritina)/sTfR pode avaliar os estímulos contrários (disponibilidade de ferro e atividade eritropoética) que controlam a síntese de hepcidina e a absorção de ferro, na ausência de estímulos inflamatórios. Foi demonstrado que TAT teve significativa redução da hepcidina e aumento do receptor solúvel de transferrina, com parâmetros hematológicos relativamente normais. Em contraste, todos os parâmetros hematológicos de TBT foram significativamente diferentes do Controle, incluindo aumento dos níveis do receptor solúvel de transferrina, ferritina, eritropoetina e fator de diferenciação do crescimento 15. Essas alterações em ambos os grupos sugerem um balanço alterado entre eritropoese e metabolismo de ferro. Os índices sTfR/log ferritina e (hepcidina/ferritina)/sTfR estão, respectivamente, aumentado e reduzido comparados ao Controle, proporcional a severidade de cada grupo talassêmico. Em conclusão, destacamos que, pela primeira vez, foram descritas alterações no metabolismo de ferro em indivíduos com ?+-talassemia heterozigótica. Esses dados demonstram que, no contexto da saúde pública, são necessários identificação e acompanhamento dos portadores de ?+-talassemia.The thalassemia syndromes (?- and ?-thalassemia) are the most common and frequent disorders associated with ineffective erythropoiesis. Imbalance of ?- or ?-globin chain production results in impaired red blood cell synthesis, anemia and more erythroid progenitors in the blood stream. While patients affected by these disorders show definitive altered parameters related to erythropoiesis, the relationship between the degree of anemia, altered erythropoiesis and dysfunctional iron metabolism have not been investigated in both carriers of ?-thalassemia and ?-thalassemia. 226 subjects (75 females and 151 males) were recruited to this study and divided in 5 groups: Control (n=28), repeat blood donors (DSR, n=23), ?+-thalassemia heterozygous carriers (TAT, n=14), ?+-thalassemia (?-thalassemia trait, TBT, n=20) and ?0-thalassemia, (?-thalassemia major, BTM, n=27). Samples were tested for hematological parameters (Micros ABX 60); serum iron, total iron binding capacity, and transferrin saturation by the colorimetric method (Pointe Scientific, Inc., Canton, MI, USA), ferritin and high sensitive C-reactive protein by immunoassay (Immulite 1000); soluble transferrin receptor, erythropoietin and growth differentiation factor 15 (R&D Systems) and hepcidin (Intrinsic LifeSciences, La Jolla, CA) by ELISA. Were calculated the ratios sTfR/log ferritin and (hepcidin/ferritin)/sTfR to evaluate iron metabolism. sTfR/log ferritin can distinguish storage iron depletion from iron-deficient erythropoiesis, while (hepcidin/ferritin)/sTfR can be utilized to explore and quantify the opposing forces (i.e. iron availability and erythropoietic activity) regulating hepcidin synthesis and iron absorption in absence of inflammatory stimuli. We demonstrate that TAT have a significantly reduced hepcidin and increased soluble transferrin receptor levels but relatively normal hematological findings. In contrast, TBT have all hematological parameters significantly different from controls, including increased soluble transferrin receptor, ferritin, erythropoietin and growth differentiation factor 15 levels. These changings in both groups suggest an altered balance between erythropoiesis and iron metabolism. The indexes sTfR/log ferritin and (hepcidin/ferritin)/sTfR are respectively increased and reduced relative to controls, proportional to the severity of each thalassemia group. In conclusion, we emphasize that, for the first time in the literature, subjects with heterozygous ?+-thalassemia have altered iron metabolism. Our data demonstrate that within the context of public health, identification and monitoring of patients with ?+-thalassemia are needed.
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Krugner, Fernando. "Niveis dos receptores soluveis de transferrina e graus de maturidade dos reticulocitos na talassemia alfa+." [s.n.], 2004. http://repositorio.unicamp.br/jspui/handle/REPOSIP/313387.
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Orientadores: Maria de Fatima Sonati, Helena Z. W. GrottoDissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas
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Resumo: Objetivo. Foi sugerido que, o aumento do número de células vermelhas (RBC) mais jovens na circulação em crianças a+-talassêmicas poderiam estar correlacionados com as altas freqüências do alelo a+-talassêmico em regiões endêmicas de malária. A avaliação dos reticulócitos (RET) nessa condição, contudo, não tem sido realizada até o presente momento. Nosso objetivo foi determinar a contagem de RET circulantes e suas frações de maturidade, além do Receptor solúvel de Transferrina (sTfR) e dos níveis de eritropoietina sérica (s-Epo), em heterozigotos da talassemia a+ da região Sudeste do Brasil, área não sujeita à ação da malária. Material e Métodos. Foram estudados 121 portadores de talassemia a+ (-a3.7/aa) (T) e 249 controles normais (aa/aa) (C), sub-classificados de acordo com as seguintes faixas etárias:1-5 anos (T=27;C=41), 6-10 (T=18;C=42), 11-15 (T=16;C=44), 16-20 (T=20;C=42) e maior de 20 (T=40;C=80), todos com níveis normais de Ferritina (FER). A análise dos RET foi feita utilizando citometria de fluxo, a determinação dos níveis de sTfR e s-Epo por imunonefelometria e quimioluminescência, respectivamente. Resultados. Não houve nenhuma diferença estatística entre T e C na avaliação dos RET [porcentagens e valores absolutos, p=0,2643 e 0,5421; nas frações de maturidade (alta, média e baixa), (p=0,2579, 0,2196 e 0,4192); e no Índice de Maturidade RET (RMI), p=0,2471, respectivamente], tão quanto nos níveis de s-Epo (p=0,5711). Os níveis de sTfR foram significativamente mais altos em T (p=0,0001) nos sub-grupos de 1-5 anos e maior de 20 (p=0,0082 e 0,0436, respectivamente). Conclusões. Embora os níveis de sTfR tenham sido mais altos, não foi observado qualquer alteração no número e maturação dos RET, nesses a+-talassêmicos aqui analisados, uma região livre de malária. Os resultados são compatíveis com uma eritropoiese compensatória
Abstract: Background and Objective. It has been suggested that an increased number of young circulating red blood cells (RBC) in a+-thalassemic children could be related to the high frequencies of the a+-thalassemic allele in malaria endemic areas. Reticulocyte (RET) evaluation in this condition, however, has not been performed so far. Our objective was to determine the RET number and maturation degree, in addition to the soluble transferrin receptor (sTfR) and serum erythropoietin (s-Epo) levels, in a+-thalassemia heterozygotes from Southeastern Brazil, an area not subjected to malaria. Design and Methods. We studied 121 a+-thalassemia carriers (-a3.7/aa) (T) and 249 normal controls (aa/aa) (C), sub-classified according to age [1-5 years (T=27;C=41), 6-10 (T=18;C=42), 11-15 (T=16;C=44), 16-20 (T=20;C=42) and over 20 (T=40;C=80)], all of them with normal ferritin levels. RET analyzes were performed by flow cytometry and the sTfR and s-Epo levels determined by immunonephelometry and chemiluminescence, respectively. Results. There was no statistical difference between T and C regarding the RET evaluation [percentages and absolute values, p=0.2643 and 0.5421; high, medium and low maturation degree, p=0.2579, 0.2196 and 0.4192; RET Maturity Index (RMI), p=0.2471, respectively], as well as the s-Epo levels (p=0.5711). The sTfR concentrations were higher in T (p=0.0001), reaching statistical significance in the 1-5 and over 20 subgroups (p=0.0082 and 0.0436, respectively). Interpretation and Conclusions. Although the higher sTfR levels, we could not observe any alteration in RET number and maturation in the a+-thalassemic population analyzed here, a region free from malaria. These results are compatible with a compensatory erythopoiesis
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Ciencias Biomedicas
Mestre em Ciências Médicas