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Marsh, D. J. "The αvβ6 integrin in cancer." Thesis, University College London (University of London), 2011. http://discovery.ucl.ac.uk/1331896/.
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The epithelial restricted αvβ6 integrin is known to have minimal expression in healthy tissue and to be upregulated in cancer and healing wounds. This thesis explores the role of αvβ6 in cancer and tests the hypothesis that αvβ6 has a prognostic and therapeutic utility in cancer. Using immunohistochemistry, increased αvβ6 expression was found in nonmelanoma skin cancers (NMSC), particularly in morphoeic type basal cell carcinoma. In cell culture experiments, αvβ6 was found to activate TGF-β and promote myofibroblast differentiation, producing a tumour stroma rich in smooth muscle actin (SMA). These findings prompted a study of αvβ6 and SMA as prognostic indicators in oral squamous cell carcinoma (OSCC). A study of 282 cases of OSCC found that although αvβ6 was not a prognostic marker, patients with high SMA levels had a highly significant increased risk of disease specific mortality (HR 3.06 [CI 1.65-5.66], p<0.001). Next, the utility of αvβ6 as a target was explored through the development of a single chain antibody fragment (scFv) specific for αvβ6. The scFv was tested for the delivery of targeted magnetic fluid hyperthermia (MFH), an experimental cancer treatment based on the generation of heat by magnetic nanoparticles when placed within an alternating magnetic field. The αvβ6-specific scFv (B6.3) was manufactured and high ligand specificity confirmed on ELISA and FACS analysis. B6.3 was successfully conjugated to two alternative iron nanoparticles. In-vitro studies demonstrated increased cellular uptake of scFv-nanoparticle complexes and greater cellular toxicity on exposure to MFH compared to nanoparticles alone. In conclusion, αvβ6 is a potential target for therapy in NMSC and OSCC. SMA is found to be an independent prognostic marker in OSCC and αvβ6 identified as a proinvasive factor in morphoeic BCC. Finally, the production αvβ6 specific scFvs and use for in-vitro MFH potentiates the development of αvβ6 targeted MFH cancer therapy.
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Vallath, Sabarinath S. "Studying the role of integrin αVβ6 in pancreatic cancer." Thesis, Queen Mary, University of London, 2013. http://qmro.qmul.ac.uk/xmlui/handle/123456789/8663.
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Pancreatic cancer is often referred to as the “silent killer“ due to the asymptomatic nature of the disease in the early stages and the extremely poor prognosis overall. The average one-year survival rate for PDAC patients is 24% (American Cancer Society, facts and figures, 2010), decreasing to 5%-6% over 5 years (WHO report, Pancreatic cancer, 2010). Only 20% of patients are suitable for surgical resection at the time of diagnosis and treatment options available to PDAC patients have not improved significantly over the past few decades. Thus novel therapeutic approaches are essential to treat this disease. Our experimental, clinical and pre-clinical data suggest integrin αvβ6 may be a suitable target. Bioinformatics studies using the Pancreatic Expression Database revealed that the β6 gene (ITGB6) was highly up regulated in pancreatic ductal carcinoma (PDAC) compared with normal pancreas. Further analysis carried out showed that there was a significant correlation between ITGB6 expression at the mRNA level and survival in a cohort of 292 PDAC patients. Immunohistochemistry analysis on two separate patient cohorts (n=118 and n=147) showed that normal pancreas lacked αvβ6 expression whereas 91% of PDAC tissues expressed αvβ6 at the protein level. There was no significant correlation between αvβ6 expression and survival at the protein level in both cohorts of patients tested. Flow cytometry and Western blotting analyses on a panel of PDAC cell lines confirmed expression of αvβ6 in PDAC cell lines. This study investigated the functional role of αvβ6 in PDAC cell lines. Antibody mediated function blockade of αvβ6 significantly inhibited proliferation in a dose dependent manner, specifically in αvβ6 positive PDAC cell lines. A significant reduction in migration and invasion was also observed in a panel of αvβ6 positive PDAC cell lines when treated with an αvβ6 function-blocking antibody. αvβ6 targeted antibody mediated therapy in combination with gemcitabine significantly inhibited tumour growth in a physiologically relevant pre-clinical subcutaneous xenograft model of PDAC. These data reaffirms that αvβ6 is a potential novel therapeutic target and an αvβ6 specific function-blocking antibody can be used as a novel agent to treat pancreatic adenocarcinoma patients.
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Hayward, Mary-Kate. "Mechanostimulation of integrin αvβ6 and fibronectin in DCIS myoepithelial cells." Thesis, Queen Mary, University of London, 2018. http://qmro.qmul.ac.uk/xmlui/handle/123456789/54057.
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Alterations to the tumour microenvironment is a common feature of many cancers, including breast cancer, and there is increasing evidence that alterations to the microenvironment, including; increased integrin expression, ECM deposition and protease activity, promote cancer progression. Most invasive breast cancers arise from a preinvasive stage, ductal carcinoma in situ (DCIS). Previous work in our laboratory has shown the microenvironment of DCIS is altered, such that myoepithelial cells (MECs) switch to a tumour-promoting phenotype, associated with upregulation of integrin αvβ6 and fibronectin (FN) expression. Mechanisms by which integrin αvβ6 and FN expression are regulated is unclear. We show DCIS progression into invasion is accompanied by an increase in MEC expression of integrin αvβ6 and periductal FN deposition, and their expression were associated in DCIS. These findings were modelled in isolated primary DCIS-MECs, primary normal MECs and MEC lines, with and without integrin αvβ6 expression, where integrin αvβ6-positive MECs upregulating FN expression. We identified integrin αvβ6-positive DCIS ducts were larger than integrin αvβ6-negative DCIS ducts, and mechanical stretching of primary normal MECs and a normal MEC line led to upregulation of integrin αvβ6 expression and FN deposition in a TGFβ-dependent manner. We further show upregulation of integrin αvβ6 and FN by MECs mediate TGFβ-dependent upregulation of MMP13 which promotes breast cancer cell invasion in vitro. These data show altered tissue mechanics in DCIS and MEC expression of integrin αvβ6 and FN deposition are linked, and implicate TGFβ in their activation. These findings suggest integrin αvβ6 and FN may be used as markers to stratify DCIS patients.
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Ylipalosaari, M. (Merja). "Matrix metalloproteinases (MMPs) in oral carcinomas." Doctoral thesis, University of Oulu, 2005. http://urn.fi/urn:isbn:9514277309.
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Abstract
Matrix metalloproteinases, MMPs, are a family of enzymes capable of modulating connective tissue components. The expression of several MMPs is increased in oral squamous cell carcinomas (OSCCs). They are assumed to have an important role in the development and progression of OSCCs. However, the exact role and mechanism of the regulation of MMPs in malignant transformation are still largely unknown.
In this study, tumour-associated trypsin-2 (TAT-2) was detected in OSCC tissue sections, and its role in MMP-2 and -9 regulation in carcinoma cells was evaluated. The TAT-2 gene was transfected into two different OSCC cell lines and one immortalized oral epithelial cell line. In TAT-2-transfected cells, MMP-9 activation increased OSCC cell invasion in chicken chorionallantoic membrane assay. Increased intravasation was prevented by tumour-associated trypsin inhibitor or specific gelatinase-inhibiting CTT-peptide. TAT-2 also converted MMP-1, -8, -13 and -3 into smaller molecular weight forms in vitro. However, TAT-2-transfected OSCC cells showed no conversion. TAT-2 was demonstrated to degrade powerfully type I collagen into small fragments in vitro. The cell surface receptor αvβ6 integrin is strongly up-regulated in OSCCs. By using β6-transfected OSCC cells, it was demonstrated that αvβ6 integrin down-regulates MMP-13 expression. However, this integrin did not regulate other collagenases or TIMP-1. β6-transfected cells invaded more efficiently through the basement membrane matrix, but their migration through type I collagen remained unchanged. MMP-8 expression was detected for the first time in head and neck squamous cell carcinoma (HNSCC) cell lines and corresponding cultured dermal and tumour fibroblasts. The localization of MMP-8 in HNSCC was determined by immunohistochemical stainings and in situ hybridization. MMP-8 production levels in carcinoma cells were faint and sporadic in HNSCCs sections. Ninety-two primary mobile tongue SCCs were subjected to MMP-8 immunohistochemical staining, and the staining results were compared to survival rates. MMP-8 was associated with improved disease-free survival in females but not in males.
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Dirheimer, Luca. "Ciblage de modèles cellulaires 3D par des agents de contrastes fluoresçant dans le proche infrarouge : application à la chirurgie guidée par la fluorescence des cancers de la tête et du cou." Electronic Thesis or Diss., Université de Lorraine, 2024. http://www.theses.fr/2024LORR0159.
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La résection chirurgicale constitue le traitement de première intention des cancers de la tête et du cou (HNSCC). La berge peropératoire est un facteur de pronostic majeur pour la survie globale des patients. Actuellement, il existe peu d’outils pour discriminer de manière fiable et en temps réel le tissu tumoral du tissu sain. La chirurgie guidée par la fluorescence (FGS) proche infrarouge (NIR) est une méthode d’imagerie utilisant le marquage fluorescent des tissus tumoraux afin de fournir une image de contraste augmentée. Le but de ces travaux est l’étude de la distribution de Quantum Dots (QDs) et d’IRDye-680, deux agents de contrastes fluorescents, couplés au peptide A20FMDV2 afin de cibler les cellules tumorales ORL au travers de l’intégrine αVβ6, surexprimée dans ces cancers. L’étude de l’accumulation et de la localisation de ces agents est réalisée sur modèle sphéroïde en monoculture et coculture (cellules cancéreuses de la langue/fibroblastes) afin de mieux représenter l’impact du microenvironnement tumoral sur la délivrance des agents de contraste. L’étude se poursuivra par le développement de nouveaux modèles sphéroïdes, mimant davantage le microenvironnement tumoral de la sphère ORLSurgical resection is the first-line treatment for head and neck cancer (HNSCC). Theintraoperative margin is a major prognostic factor for the overall survival of patients. Currently, there are few tools to reliably discriminate tumor tissue from healthy tissue in real time. Near Infrared (NIR) Fluorescence Guided Surgery (FGS) is an imaging method using fluorescent labeling of tumor tissue to provide an enhanced contrast image. The aim of this work is to study the distribution of Quantum Dots (QDs) and IRDye-680, two fluorescent contrast agents, coupled to the A20FMDV2 peptide to target ENT tumor cells through the αVβ6 integrin, which is overexpressed in these cancers. The accumulation and localization of these agents was studied using spheroid models in monoculture and coculture (tongue cancer cells/fibroblasts) to better represent the impact of the tumor microenvironment on the delivery of these contrast agents. The study is continuing with the development of new spheroid models that better represent the tumor microenvironment found in the ENT sphere
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Nieberler, Markus [Verfasser], Klaus-Dietrich Akademischer Betreuer] Wolff, Andreas [Akademischer Betreuer] Kolk, and Henning [Akademischer Betreuer] [Bier. "Entwicklung und klinische Etablierung einer intraoperativen zytologischen Diagnostik der Knocheninfiltration bei Kopf-Hals-Karzinomen mit Charakterisierung von αvβ6 Integrin als Biomarker invasiver Karzinomzellen / Markus Peter Nieberler. Gutachter: Klaus-Dietrich Wolff ; Andreas Kolk ; Henning August Bier. Betreuer: Klaus-Dietrich Wolff." München : Universitätsbibliothek der TU München, 2014. http://d-nb.info/1058214454/34.
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Mathias, Lucas Solla. "Ativação da via MAPK/ERK e Integrina αvβ3 pela ação da triiodotironina (T3) na modulação da expressão gênica de adipocinas e modificação do perfil lipídico em adipócitos, 3T3-L1." Botucatu, 2019. http://hdl.handle.net/11449/181721.
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Orientador: Miriane de OliveiraResumo: Introdução: O hormônio triiodotironina (T3) influencia o metabolismo e desenvolvimento do tecido adiposo (TA), modulando a proliferação e diferenciação de adipócitos, podendo agir sobre os reguladores do processo de adipogênese, como o receptor ativado por proliferador de peroxissomo (PPARy). O TA está envolvido na regulação da energia corporal, sintetizando e secretando substâncias denominadas adipocinas, dentre elas a adiponectina e leptina. A adiponectina está relacionada ao aumento da sensibilidade à insulina, enquanto a leptina está envolvida com o gasto energético. O T3 pode desencadear ações por ativação de vias extranucleares, dentre elas a via MAPK/ERK e integrina αVβ3. Objetivo: Verificar a ação do T3, com participação das vias extranucleares MAPK/ERK e integrina αVβ3, na modulação de adiponectina e leptina, além de avaliar os parâmetros relacionados ao perfil adipogênico e dano de DNA. Métodos: Adipócitos, 3T3-L1, foram tratados com T3 (10nM) por uma hora, na ausência ou presença dos inibidores de MAPK/ERK – PD98059 (PD, 50uM) e da integrina αvβ3 – ácido tetraiodotiroácetico (Tetrac, 10-4M). A ausência de qualquer tratamento foi considerada grupo controle (C). Após o período de tratamento foi realizado PCRq-RT para analisar a expressão de mRNA de adiponectina e leptina, e Western Blot para expressão proteica de adiponectina, leptina, PPARy, pAKT e pERK; a viabilidade celular foi realizada pelo ensaio de MTT; a quantificação do acúmulo lipídico pelos ens... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: Introduction: The hormone triiodothyronine (T3) influences the metabolism and development of adipose tissue (TA), modulating the proliferation and differentiation of adipocytes, and can act on regulators of the adipogenic differentiation process, such as the peroxisome proliferator activated receptor). TA is involved in the regulation of body energy, synthesizing and secreting substances called adipokines, among them adiponectin and leptin. Adiponectin is related to increased insulin synaptic, since leptin is involved in energy expenditure. T3 can trigger actions by activation of extranuclear pathways, including MAPK / ERK and integrin α Vβ3. Objective: Given the role of T3 in TA and the importance of adipokines, the objective of this study is to verify the action of T3 with the participation of extranuclear pathways in the modulation of adiponectin and leptin and the parameters related to the adipogenic profile. Methods: Adipocytes, 3T3-L1, were treated with a physiological dose of T3 (10nM) for one hour, in the absence or presence of MAPK / ERK-PD98059 (PD) and integrin αvβ3 - tetraiodothyrocetic (Tetrac) integrin inhibitors. The absence of any treatment was considered as a control group (C). After the treatment period PCRqRT was performed to analyze the expression of leptin and adiponectin mRNA, and Western Blot for protein expression of adiponectin, leptin, PPARγ, pAKT and pERK; cell viability was performed by the MTT assay; the quantification of lipid accumulation by the... (Complete abstract click electronic access below)
Mestre
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Elsharif, Amal A. M. "Functional Investigation of Dual αvβ3 and αllbβ3 Integrin Inhibition in Haematological and Solid Tumour Models." Thesis, University of Bradford, 2018. http://hdl.handle.net/10454/16883.
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Invasion and metastasis of cancer is the leading cause of increased mortality. In addition, haematological malignancies (leukaemia and lymphoma) are a significant cause of morbidity and mortality in both children and adults. Therefore, new treatments which will inhibit cancer progression are required. Integrin adhesion receptors, particularly the RGD-binding integrin subfamily comprising αvβ3, αvβ5, αvβ6, αvβ8, αllbβ3, α5β1, α8β1 and αvβ1 are related to progress and spread of cancer and poor prognosis. Because of the importance of integrin biology in the regulation of cancer dissemination, the integrin receptors are being utilised as targets to regulate cancer progression. The goal of this study was to develop a dual αvβ3/ αIIbβ3 expressing model for testing integrin antagonists. Expression of αv, αIIb, and β3 integrin subunits was characterised using immunofluorescence and flow cytometry in a panel of cell lines. After characterising the expression of αv, αIIb and β3 integrin subunits in inducible and natural expression models (K562 and MCF-7 cells respectively), functional tests for cellular adhesion, detachment and migration were determined. Phorbol 12-myristate 13-acetate (PMA)-treated K562 cells showed increased adhesion on fibrinogen compared to untreated cells. Adhesion of cancer cells (K562 ± PMA and MCF-7) to fibrinogen was inhibited and detachment was induced by the known β3 antagonists, cRGDfV and GR104453.
Migration of cancer cells (K562 without PMA and MCF-7) was inhibited by combination of the known β3 antagonists. A panel of 12 novel small molecules developed in the ICT was investigated for cytotoxicity and activity in the validated function assays. ICT9055 was the most potent antagonist in inhibition of cell adhesion, migration, and inducing cell detachment. The data presented in this thesis had selected models and assays for evaluating small molecule integrin antagonists and identified ICT9055 as a promising molecule to develop for further preclinical evaluation.The Libyan Embassy; Omer Al Mukhtar University, Faculty of Medical Technology, Derna, Libya.
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Ahmedah, Hanadi Talal A. "Correlation between the expression of integrins and their role in cancer progression : expression pattern of integrins αvβ3, αvβ5 and α5β1 in clinical and experimental tumour samples." Thesis, University of Bradford, 2015. http://hdl.handle.net/10454/14284.
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The integrins play a crucial role in cancer cell proliferation, migration, differentiation, survival and angiogenesis. It has been shown that integrin expression is positively correlated to cancer dissemination, this suggests targeting selected integrins as an anti-metastatic strategy. The aim of this study is to investigate the effect of novel antagonists of α5β1, αvβ3 and αvβ5 integrins on cancer cell migration, a key process in tumour cell dissemination. Immunohistochemistry was used to evaluate the expression of α5, αv, β3 and β5 integrin subunits in prostate cancer tissues. Furthermore the expression of these integrin subunits in tumour and normal human head and neck tissues was compared. The expression profile of these integrin subunits in established human cancer cell lines was subsequently evaluated using immunodetection methods in cells and xenograft tumour samples. The effect of integrin inhibition on cell migration was then assessed using neutralizing antibodies against αvβ3, αvβ5, and α5β1 integrins in the scratch-wound healing assay. This assay was then used to evaluate the potential of novel small molecule integrin antagonists in preventing tumour cell migration. In H & N tissues, αvβ3, αvβ5 and α5β1 integrins are extensively expressed in tumour tissues but weakly expressed in normal tissue from the same patient. Further, prostate cancer tissues expressed variable levels of αvβ3, αvβ5 and α5β1 integrins. αvβ3 and αvβ5 integrins were expressed in variable levels in OSC-19, PC-3, DU145, DLD-1, HT-29, HUVEC, MCF-7, MCF-7ADR and M14 human tumour cell lines and in OSC-19, PC-3, HT-29 and MCF-7 xenografts. α5β1 integrin was expressed in all cell lines and xenografts except in MCF-7 cell line and HT-29 cell line and xenograft. Overall, the expression was elevated in xenografts compared to the corresponding cultured cells. Based on the expression profile and ability of cells to migrate, three cell lines (DLD-1 colon, DU145 prostate and OSC-19 HNSCC) were selected as models to further evaluate the potential of novel small molecule integrin antagonists to inhibit cell migration. The cell lines were characterized by using neutralizing antibodies against αvβ3, αvβ5, and α5β1 integrins to determine which of these three integrins were primarily involved in tumour cell migration. In DLD-1 and DU145, blocking αvβ5 and αvβ3 significantly inhibited migration, whilst the migration of OSC-19 was 50% inhibited by a multi-integrin inhibitor combination. Among the antagonists, ICT9055 and ICT9072 significantly decreased DLD-1 cell migration by 70% and 60% respectively while ICT9023, ICT9024, and ICT9026 significantly decreased DU145 cell migration by 60%, 60% and 50% respectively. The findings suggest that single integrin inhibition is not sufficient to prevent cell migration whereas dual or multiple inhibition is more effective. Two novel anti-migratory agents were identified in colon cancer and three in prostate cancer which would warrant further investigation.
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Ahmedah, Hanadi T. A. "Correlation between the expression of integrins and their role in cancer progression. Expression pattern of integrins αvβ3, αvβ5 and α5β1 in clinical and experimental tumour samples." Thesis, University of Bradford, 2015. http://hdl.handle.net/10454/14284.
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The integrins play a crucial role in cancer cell proliferation, migration,
differentiation, survival and angiogenesis. It has been shown that integrin
expression is positively correlated to cancer dissemination, this suggests
targeting selected integrins as an anti-metastatic strategy. The aim of this study
is to investigate the effect of novel antagonists of α5β1, αvβ3 and αvβ5 integrins
on cancer cell migration, a key process in tumour cell dissemination.
Immunohistochemistry was used to evaluate the expression of α5, αv, β3 and
β5 integrin subunits in prostate cancer tissues. Furthermore the expression of
these integrin subunits in tumour and normal human head and neck tissues was
compared. The expression profile of these integrin subunits in established
human cancer cell lines was subsequently evaluated using immunodetection
methods in cells and xenograft tumour samples. The effect of integrin inhibition
on cell migration was then assessed using neutralizing antibodies against αvβ3,
αvβ5, and α5β1 integrins in the scratch-wound healing assay. This assay was
then used to evaluate the potential of novel small molecule integrin antagonists in preventing tumour cell migration. In H & N tissues, αvβ3, αvβ5 and α5β1
integrins are extensively expressed in tumour tissues but weakly expressed in normal tissue from the same patient. Further, prostate cancer tissues expressed
variable levels of αvβ3, αvβ5 and α5β1 integrins. αvβ3 and αvβ5 integrins were
expressed in variable levels in OSC-19, PC-3, DU145, DLD-1, HT-29, HUVEC,
MCF-7, MCF-7ADR and M14 human tumour cell lines and in OSC-19, PC-3,
HT-29 and MCF-7 xenografts. α5β1 integrin was expressed in all cell lines and
xenografts except in MCF-7 cell line and HT-29 cell line and xenograft. Overall,
the expression was elevated in xenografts compared to the corresponding
cultured cells. Based on the expression profile and ability of cells to migrate,
three cell lines (DLD-1 colon, DU145 prostate and OSC-19 HNSCC) were
selected as models to further evaluate the potential of novel small molecule
integrin antagonists to inhibit cell migration. The cell lines were characterized by
using neutralizing antibodies against αvβ3, αvβ5, and α5β1 integrins to
determine which of these three integrins were primarily involved in tumour cell
migration. In DLD-1 and DU145, blocking αvβ5 and αvβ3 significantly inhibited migration, whilst the migration of OSC-19 was 50% inhibited by a multi-integrin
inhibitor combination. Among the antagonists, ICT9055 and ICT9072
significantly decreased DLD-1 cell migration by 70% and 60% respectively while
ICT9023, ICT9024, and ICT9026 significantly decreased DU145 cell migration
by 60%, 60% and 50% respectively. The findings suggest that single integrin
inhibition is not sufficient to prevent cell migration whereas dual or multiple
inhibition is more effective. Two novel anti-migratory agents were identified in
colon cancer and three in prostate cancer which would warrant further
investigation.Princess Nora Bint Abdul Rahman University
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Alshammari, Fatemah O. F. O. "An immunohistopathological and functional investigation of β3 integrin antagonism as a therapeutic strategy in cancer : characterisation, development, and utilisation of preclinical cancer models to investigate novel β3 integrin anatgonists." Thesis, University of Bradford, 2013. http://hdl.handle.net/10454/6327.
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Tumour cell dissemination is a major issue with the treatment of cancer, thus new therapeutic strategies which can control this process are needed. Antagonism of integrins highly expressed in tumours is one potential strategy. The integrins are transmembrane glycoprotein adhesive receptors. Two of the integrins, αVβ3 and αIIbβ3, are highly expressed in a number of tumours and induce bi-directional signalling through their interaction with extracellular matrix proteins, and growth factor receptors. Through this signalling they play an important role in a number of cellular processes that are involved in tumour dissemination such as tumour growth, migration, invasion, metastasis and angiogenesis. Dual αIIbβ3 and αVβ3 integrin antagonism will have a direct effect on β3-expressing tumour cells that leads to the inhibition of cell migration and dissemination. Furthermore, through targeting tumour cell interaction with endothelial cells and platelets, this will also lead to inhibition of angiogenesis and metastasis. The aim of this project was to characterise the expression of αVβ3 and αIIbβ3 integrin in a panel of tumour cell lines and in human tumour xenograft samples, and to develop and utilise cell-based models to investigate potential novel β3 antagonists. The expression of αV and β3 subunits was detected in xenograft tissue using immunoblotting techniques. A panel of cell lines of different tumour types including melanoma, prostate, breast, colon and non small cell lung carcinoma was then characterised for αVβ3 and αIIbβ3 integrin expression using immunoblotting and immunocytochemistry. Melanoma cell lines demonstrated the strongest αVβ3 expression. No αIIbβ3 integrin expression was seen in any of the cell lines evaluated. A selection of cell lines with varying αVβ3 expression were then used to develop a functional test for cell migration, the scratch wound healing assay. Migration of tumour cells that expressed αVβ3 integrin was inhibited by the known β3 antagonists, cRGDfV peptide and LM609 antibody. A panel of 12 potential novel β3 integrin antagonists was screened for cytotoxicity and activity in the validated scratch assay. ICT9055 was the most effective antagonist in inhibition of M14 cell migration as determined by the scratch assay, with an IC₅₀ of < 0.1 μM. Therefore the work presented in this thesis has established models and tools for evaluating potential novel β3 integrin antagonists, and identified a promising molecule to progress for further preclinical evaluation.
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Antonow, Michelli Barcelos. "Desenvolvimento e caracterização físico-química de nanocápsulas multiparedes complexadas com zinco e funcionalizadas com RGD para reconhecimento por integrinas ανβ3 presentes em células tumorais." reponame:Biblioteca Digital de Teses e Dissertações da UFRGS, 2016. http://hdl.handle.net/10183/149502.
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A funcionalização de superfície nas nanocápsulas contendo doxorrubicina com o peptídeo RGD é uma estratégia promissora devido a ligação preferencial na integrina αvβ3 expressa em células tumorais. Este estudo objetivou o desenvolvimento, caracterização e estudos biológicos de nanocápsulas multiparedes com doxorrubicina e funcionalizadas com RGD. Para isso, na primeira etapa do trabalho foi realizada a síntese do peptídeo RGD. Os produtos obtidos foram caracterizados por análises de infravermelho e RMN de 1H. Na segunda etapa foram desenvolvidas formulações de nanocápsulas com doxorrubicina ou cloridrato de doxorrubicina, e, nanocápsulas multiparedes revestidas com quitosana, íons zinco, RGD ou fenilalanina. Essas suspensões foram caracterizadas através da determinação do pH, diâmetro de partícula por diferentes técnicas, potencial zeta, eficiência de encapsulação e eficiência de associação do RGD na superfície da nanopartícula. Na terceira etapa, foram realizados ensaios de viabilidade celular por MTT após 24 e 72h com as formulações desenvolvidas em células de câncer de mama (MCF7) e glioblastoma humano (U87MG). As formulações apresentaram diferentes valores de citotoxicidade e, utilizando o Gráfico de Pareto foi possível determinar os fatores que exercem maior influencia. Em células MCF7 foi a concentração de fármaco e tempo de tratamento e, nas células U87MG além desses fatores, a funcionalização mostrou-se determinante. Além disso, foi avaliada a captação das nanocápsulas funcionalizadas com RGD e fenilalanina após 24h nas células tumorais e células de queratinócitos humanos (HaCat), com diferentes níveis de expressão da integrina αvβ3. O estudo mostrou menores valores de captação nas células HaCat (sem expressão de integrina αvβ3) para as duas formulações testadas. Finalmente as nanocápsulas funcionalizadas com RGD apresentaram maior captação em células U87MG com maior expressão da integrina αvβ3.The surface functionalization in nanocapsules containing doxorubicin with RGD peptide is a promising strategy due to preferential binding in the αvβ3 integrin expressed on tumor cells. This study aimed the development, characterization, and biological studies of multiwall nanocapsules containing doxorubicin and functionalized with RGD. For this reason, in the first stage of this study the synthesis of RGD peptide was performed and the products characterized by infrared analysis and 1H NMR. Besides, nanocapsules formulations were developed containing doxorubicin or doxorubicin hydrochloride, and multiwall nanocapsules coated with chitosan, zinc ions, RGD or phenylalanine. These suspensions were characterized by pH determination, particle diameter by different techniques, zeta potential, encapsulation efficiency, and association efficiency of RGD on the surface of the nanoparticle. Additionally, it was performed cell viability assays by MTT after 24 and 72 hours with formulations developed in breast cancer (MCF7) and human glioblastoma cells (U87MG). Formulations showed different cytotoxicity values. The Pareto chart was possible to determine factors that have more influence. In MCF7 cells was drug concentration and treatment time, and U87MG cells, besides these factors, the functionalization was decisive. Furthermore, it was performed the cellular uptake of nanocapsules functionalized with RGD or phenylalanine after 24 hours in tumor cells and human keratinocyte cells (HaCaT), with different levels of expression αvβ3 integrin. The study showed less uptake in HaCaT cells (without expression αvβ3 integrin) for the two formulations applied, and the nanocapsules functionalized with RGD showed more uptake in U87MG cells, with higher expression of integrin αvβ3.
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Acharya, Mridu. "Expression and function of the αVβ5 integrin during human B lymphopoiesis." Thesis, University of Glasgow, 2008. http://theses.gla.ac.uk/172/.
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The integrin αVβ5 is a receptor for sCD23 molecule and the αVβ5-CD23 interaction sustains proliferation of the pre-B cell line SMS-SB. This thesis describes further investigation into the role of αVβ5 integrin during human B cell development. B cell development in the bone marrow involves stepwise maturation of progenitor cells through different defined stages. A tight regulation of proliferation and differentiation mediates the progression of progenitor cells through this developmental pathway. A variety of signals from soluble molecules and adhesive interactions regulate this balance of proliferation and differentiation. The main aim of this work was to assess the importance of the integrin αVβ5 during B cell development by defining its expression and function during specific stages of B cell development in the bone marrow. The αVβ5 integrin was expressed by B cell precursors in the bone marrow, by different pre-B cell lines and the αVβ5-CD23 interaction sustained proliferation of some pre-B cell lines. The pre-B cell lines SMS-SB, RS4;11 and 697 showed a significant proliferative response to αVβ5 stimulation by sCD23, a sCD23-derived long peptide and anti-αVβ5 MAb 15F11. Transitional and more mature B cell lines down-regulated αVβ5 expression and did not show a proliferative response. Both αVβ5 and αVβ3 integrin could be detected on normal bone marrow B cell precursor populations, though αVβ5 was the more highly expressed integrin. In preliminary functional experiments, stimulation of CD19+/κ- cells with sCD23 induced cell proliferation whereas equivalent treatment of CD19+/κ+ cells did not. The data are consistent with the interpretation that αVβ5 integrin is expressed in B cell precursors but that its ability to sustain growth of these cells wanes as the cells mature towards a membrane immunoglobulin-positive state. The αVβ5-mediated proliferation was enhanced by the chemokine SDF-1 and PDGF but not by the cytokines IL-3, IL-4, IL-7 or IL-11. This effect was apparently restricted to earlier B cell precursors and was independent of levels of expression of both αVβ5 and CXCR4. Stimulation of SMS-SB cells by αVβ5 ligands provoked ERK phosphorylation and co-stimulation with SDF-1 promoted a more rapid and sustained ERK activation. PDGF induced a similar effect on αVβ5-mediated activation of ERKphosphorylation. These data suggest that ligation of αVβ5 by soluble, adhesion-independent stimuli activates ERK phosphorylation and this pathway can be modulated by inputs from G-protein-coupled and tyrosine kinase receptors. The murine pro-B cell line BAF03 also displayed αVβ5-mediated proliferation in response to human CD23. Preliminary experiments showed that human CD23 sustained growth of murine bone marrow B cell precursors. Therefore, these data suggest that murine B cells can also use αVβ5 integrin to sustain their growth and the studies described here in human cell lines could be translated into in vivo murine models. Further work is needed to confirm the proliferative response due to the αVβ5-CD23 interaction in normal B cell precursors in the bone marrow and to define the exact stage of development where this interaction is critical. In addition to its expression in the bone marrow B cell precursors, previous work has also demonstrated the expression of αVβ5 integrin in B cells from patients with ALL. Therefore, the αVβ5-CD23 interaction could have important implications not only in proliferation of normal B cell precursors but also in proliferation of neoplastic B cells. These data identify the αVβ5-CD23 interaction as a potentially important interaction during early B cell development, as αVβ5 expression and function is stage-specific, regulated by other molecules and can be demonstrated in both human and murine cell lines.
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Seelke, Sandra [Verfasser], Nina [Akademischer Betreuer] Kollmar, Sigrid [Akademischer Betreuer] Hoyer-Fender, and Wilfried [Akademischer Betreuer] Kramer. "Untersuchung bispezifischer Intradiabodies gegen die Integrine αvβ3, αvβ5 und α5β1 auf ihre anti-angiogenen Eigenschaften / Sandra Seelke. Gutachter: Sigrid Hoyer-Fender ; Wilfried Kramer. Betreuer: Nina Kollmar." Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2013. http://d-nb.info/1044173068/34.
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Alkhedaiade, Adel Qlayel Hamdan. "Studies of αvβ integrin functions in human B cell precursors." Thesis, University of Glasgow, 2011. http://theses.gla.ac.uk/3044/.
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The αVβ5 integrin is a member of integrin family that binds to different ligands such as vitronectin, fibronectin and soluble CD23 in order to mediate different biological responses such as cell growth, adhesion and metastasis. It is expressed by B cell precursors and by different acute lymphocytic leukaemia cell lines such as SMS-SB cells. This thesis is an attempt to explain how the sCD23- αVβ5 integrin interaction stimulates SMS-SB cell growth and to study the role of the αVβ5 integrin and other receptors such as PDGF receptor and CXCR4 in B cell development in the bone marrow. The maturation and differentiation of B-cells occur due to several factors that impact on gene expression in its development program. This program is divided into two main phases, the antigen-independent B-cell development phase and antigen-dependent B-cell development phase, respectively. The antigen - independent phase of B cell development starts from the pluripotent haemopoietic stem cell (PHSC) and progresses through several successive stages which are identified by somatic recombination and rearrangement of both heavy and light chain genes. Soluble CD23 and LP (a synthetic peptide derived from soluble CD23) significantly stimulate SMS-SB growth while a smaller growth stimulation is caused by either SDF1-α or PDGFAB. There are different signalling targets involved in the αVβ5 integrin-mediated proliferation due to its binding to either sCD23 or LP. These ligands enhance the association between the αVβ5 integrin and the PDGF receptor which promote the phosphorylation of both Jak2 and STAT5. Moreover, cell growth was reduced and the phosphorylation of Jak2 and STAT5 was also knocked down with using either PDGF receptor inhibitor (AG1295) or Jak2 inhibitor (AG490). Both soluble CD23 and LP activate the STAT5-DNA binding and strongly increase its transcriptional activity. In addition, both ligands induce the phosphorylation of other different substrates such as STAT2, c-Src, c-yes and AMPKα2 which might be related to cell growth stimulation. The αVβ5 integrin ligands also promote the phosphorylation of ERK1/2, p90RSK and activate a SRF transfected reporter gene. However, ERK1/2 and p90RSK phosphorylation was completely blocked by the specific MEK inhibitor (U0126). In similar context, SDF1-α stimulates the transcriptional activity of SRF but not STAT5 while PDGFAB does the opposite. Finally, soluble CD23 induces the proliferation of 697 and BAF03 which are other pre-B cell line models. These data suggest that the αVβ5 integrin-ligated ligands stimulate SMS-SB cell growth by promoting different signalling pathways, mainly Jak2/STAT5 and MEK/ERK1/2 pathway. Further work is required to determine the role of STAT5, p90RSK, c-Src and SRF in stimulating either the proliferation or apoptosis that promoted by the αVβ5 integrin-sCD23 interaction and to investigate the relationship between the activation of these targets.
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Fenton, Thomas. "Integrin αvβ8 on human dendritic cells : a role in intestinal immune homeostasis." Thesis, University of Manchester, 2015. https://www.research.manchester.ac.uk/portal/en/theses/integrin-alphav8-on-human-dendritic-cells-a-role-in-intestinal-immune-homeostasis(3820bc76-2c42-4db6-a344-b66e5b77769d).html.
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Intestinal inflammatory disorders such as Crohn’s disease contribute significantly to human mortality and morbidity. Although the cells and molecules involved in suppression of intestinal inflammation have been extensively documented in mouse models, a full understanding of how these work together in the healthy and diseased gut remains elusive. It is known, however, that tight regulation over TH17 cells and regulatory T cells (Treg) is required to maintain immune homeostasis in the intestine. Activation of the cytokine transforming growth factor-β (TGFβ), which is secreted by immune cells as an inactive complex, plays a crucial role in the induction of both Treg and TH17 cells. Recent work has shown that the αvβ8 integrin is required for activation of TGFβ by murine dendritic cells (DC). Murine integrin αvβ8 is thus of fundamental importance for Treg and TH17 induction and, subsequently, for intestinal immune homeostasis. However, little is known about the signals controlling the expression of integrin αvβ8 on intestinal DC. Furthermore, whether this system is also important for regulation of the human system is entirely unknown. Here, expression of integrin αvβ8 is shown on human intestinal CD1c+CD103+SIRPα+ DC and CD1c+CD103-SIRPα+ DC, but not on CD141+CD103+SIRPα- DC. Expression of integrin αvβ8 is increased on DC from Crohn’s disease patients, and on DC from non-IBD donors treated with LPS. Both LPS and a number of probiotic bacteria were also able to induce integrin αvβ8 expression on human monocyte-derived DC (moDC), which suggested that the microbiota may play a role in the regulation of human integrin αvβ8. Importantly, we have also shown that TGFβ is activated by integrin αvβ8 on human moDC, and that integrin αvβ8 promotes expression of forkhead box P3 (FOXP3) in naïve human T cells in vitro. Together, these data suggest that integrin αvβ8-mediated activation of TGFβ by DC may play an important role in the regulation of human T cell responses in the human intestine, and that this pathway may be perturbed during intestinal inflammation.
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Steri, Veronica. "Elucidating the role of endothelial αvβ3-integrin in tumour growth and angiogenesis." Thesis, University of East Anglia, 2015. https://ueaeprints.uea.ac.uk/56762/.
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Angiogenesis, the formation of new vessels from pre-existing ones, is essential for primary tumour growth as well as for metastasis, and endothelial cells play a central role in this process: they drive blood vessel formation in response to signals from the local environment by a mechanism that is integrin-dependent. αvβ3-integrin seemingly poses an ideal anti-angiogenic target. Its expression is vastly up-regulated in neo-angiogenic vessels, while its expression in quiescent vasculature is minimal. However, anti-angiogenic therapy targeting αvβ3-integrin has proven somewhat disappointing. In part, this may relate to the fact that αvβ3-integrin is not expressed solely by endothelial cells, but across a wide range of cell types that each contribute to angiogenesis. In this thesis, I describe my studies on understanding the role of αvβ3-integrin as expressed specifically by endothelial cells in tumour growth and angiogenesis using endothelial specific β3-integrin deficient mice. I have shown that inducible deletion of endothelial β3-integrin inhibits tumour growth and angiogenesis preventatively, while its constitutive deletion is ineffective; furthermore, I have found that even the inducible deletion does not alter angiogenesis in already established tumours. The findings described in this thesis re-establish αvβ3-integrin as good antiangiogenic target, but imply that timing and length of inhibition are critical factors to be considered when targeting endothelial β3-integrin-expression.
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Schäfer, Markus [Verfasser], and M. [Akademischer Betreuer] Bastmeyer. "Mechanische Kräfte regulieren die Bindungspromiskuität von αVβ3-Integrin / Markus Schäfer ; Betreuer: M. Bastmeyer." Karlsruhe : KIT-Bibliothek, 2019. http://d-nb.info/1200471148/34.
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Andriu, Alexandra. "Evaluating the utility of αvβ3 integrin antagonists to detect and treat angiogenic tumour cells." Thesis, University of Aberdeen, 2018. http://digitool.abdn.ac.uk:80/webclient/DeliveryManager?pid=240026.
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Tumour angiogenesis, the formation of new blood vessels within a tumour, is a hallmark of cancers, that allows them to grow beyond a critical size and to metastasize to other organs. As the key driver of tumour angiogenesis, αvβ3 integrin is both an established therapeutic target for anti-angiogenic drugs and a biomarker for imaging agents. Accurate detection of αvβ3 integrin in angiogenic tumours impacts patient prognosis and could report on therapy response. Over the last twenty years, novel αvβ3 integrin-targeted radiotracers have been developed for PET imaging of tumour angiogenesis. While most radiotracers have shown their utility as diagnostic tools, only a few focussed on evaluating response to treatment, partly due to complex biology of the integrin receptors. The aim of this project was the in-vitro biological testing of a novel αvβ3-targeted radiotracer, [3H]ZMPZAT71, and the corresponding unlabelled compound for targeted delivery of a cytotoxic drug, paclitaxel (PTX). Firstly, this piece of work involved validation of this radiotracer to assess expression of αvβ3 integrin. Secondly, [3H]ZMPZAT71 was investigated as a biomarker for assessing response to pharmacological inhibitors of cell signalling pathways. Thirdly, the targeting moiety (ZMPZAT71) conjugated with paclitaxel was studied for integrinmediated drug delivery. The results presented herein demonstrate that radiotracer binding to αvβ3 integrin is dependent not only on the expression levels, but also on the activation status of αvβ3 integrin. Furthermore, this piece of information was used to explain radiotracer binding in response to various pharmacological inhibitors of key cell signalling pathways. Additionally, the integrin-targeted chemotherapeutic exhibited selective cytotoxic effect, explained by enhanced apoptosis of cancer cells compared to PTX alone, together with anti-migratory and anti-invasive effects from the targeting moiety. This study provides valuable information about the molecular mechanisms regulated by αvβ3 integrin, supporting the development of integrin-targeted therapeutics and imaging.
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van, Wieringen Tijs. "Intra- and Extracellular Modulation of Integrin-directed Connective Tissue Cell Contraction." Doctoral thesis, Uppsala universitet, Institutionen för medicinsk biokemi och mikrobiologi, 2009. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-102349.
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All blood vessels in the microvasculature are embedded in loose connective tissue, which regulates the transport of fluid to and from tissues. The intersti-tial fluid pressure (IFP) is one of the forces that control this transport. A lowering of IFP in vivo results in an increased transport of fluid from the circulation into the underhydrated connective tissues, resulting in edema formation. During homeostasis, contractile connective tissue cells exert a tension on the connective tissue fibrous network by binding with β1 in-tegrins, thereby actively controlling IFP. During inflammation, the IFP is lowered but platelet-derived growth factor (PDGF)-BB induces an IFP nor-malization dependent on integrin αVβ3. We demonstrate that extracellular proteins from Streptococcus equi subspecies equi modulated cell-mediated and integrin αVβ3-directed collagen gel contraction in vitro. One of these proteins, the collagen- and fibronectin binding FNE, stimulated contraction by a process dependent on fibronectin synthesis. This study identified a pos-sible novel virulence mechanism for bacteria based on the ability of bacteria to modulate the edema response. Another protein, the collagen-binding pro-tein CNE, inhibited contraction and this led to the identification of sites in collagen monomers that potentially are involved in connecting αVβ3 to the collagen network. PDGF-BB and prostaglandin E1 (PGE1) stimulate and inhibit collagen gel contraction in vitro and normalize and lower IFP, respec-tively. We showed that these agents affected both similar and different sets of actin-binding proteins. PDGF-BB stimulated actin cytoskeleton dynamics whereas PGE1 inhibited processes dependent on cytoskeletal motor and adhesive functions, suggesting that these different activities may partly ex-plain the contrasting effects of PGE1 and PDGF-BB on contraction and IFP. Mutation of the phosphatidylinositol 3’-kinase (PI3K), but not phospholipase C (PLC)γ activation site, rendered cells unable to respond to PDGF-BB in contraction and in activation of the actin binding and severing protein cofilin. Ability to activate cofilin after PDGF-BB stimulation correlated with ability to respond to PDGF-BB in contraction, suggesting a role for cofilin in this process downstream of PDGF receptor-activated PI3K. Many proteins can modulate contraction either by affecting the extracellular matrix and cell adhesions or by altering cytoskeletal dynamics. Knowledge on how these proteins might influence IFP is likely to be of clinical importance for treat-ment of inflammatory conditions including anaphylaxis, septic shock and also carcinoma growth.
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Casals, Soler Gemma. "Investigación de la osteopontina y la integrina αvβ3 como marcadores de receptividad endometrial para la implantación embrionaria." Doctoral thesis, Universitat de Barcelona, 2014. http://hdl.handle.net/10803/301774.
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El estudio del endometrio y de su estado de receptividad para la implantación embrionaria es crucial en la evaluación de la fertilidad de la mujer. No obstante, se ha demostrado que el estudio histológico clásico carece de utilidad clínica práctica para valorar la adecuada capacidad del endometrio para implantar embriones. Por tanto, la búsqueda de nuevos marcadores de receptividad endometrial constituye una línea de investigación de gran interés. Se ha descrito que tanto la integrina αvβ3 como la osteopontina presentan unos patrones de expresión temporales y espaciales específicos en los diferentes tipos celulares del endometrio, que definirían la fase de receptividad endometrial para la implantación embrionaria o "ventana de implantación".
En base a lo anterior, nuestra hipótesis de trabajo es que la identificación y estudio de la osteopontina y la integrina αvβ3 como marcadores endometriales de implantación embrionaria en diferentes situaciones clínicas, debería permitir ir más allá de los simples criterios clásicos morfo-histológicos valorados por microscopia óptica y de esta forma poder definir de una manera más precisa y objetiva el grado de receptividad endometrial para la implantación embrionaria.
Para ello, se han realizado 3 estudios, publicados en revistas del primer cuartil de la especialidad:
I. “Osteopontin and αvβ3 integrin expression in the endometrium of infertile and fertile women”
Casals G, Ordi J, Creus M, Fábregues F, Casamitjana R, Quinto L, Campo E, Balasch J.
Reprod Biomed Online, 2008;16(6):808-816
II. “Osteopontin and αvβ3 integrin as markers of endometrial receptivity: the effect of different hormone therapies”
Casals G, Ordi J, Creus M, Fábregues F, Carmona F, Casamitjana R, Balasch J.
Reprod Biomed Online, 2010;21(3):349-359
III. “Expression pattern of osteopontin and αvβ3 integrin during the implantation window in infertile patients with early stages of endometriosis”
Casals G, Ordi J, Creus M, Fábregues F, Carmona F, Casamitjana R, Balasch J.
Hum Reprod, 2012;27(3):805-813
Según los datos obtenidos en estos estudios, podemos afirmar que no existe una relación causa-efecto entre la presencia o ausencia de estos dos marcadores endometriales (osteopontina e integrina αvβ3) durante la denominada "ventana de implantación" y la esterilidad de la mujer asociada o no a la presencia de endometriosis. Por otra parte, la administración de diferentes hormonas con efectos probados sobre el endometrio modifica la expresión de osteopontina e integrina αvβ3 sólo en tanto en cuanto son capaces de influir sobre el estado de maduración histológica de la mucosa endometrial. Por tanto, el interés de su uso rutinario en la valoración de la paciente estéril no puede confirmarse de acuerdo con los resultados de esta Tesis Doctoral.The study of the human endometrium as a fertility-determining factor and understanding the factors that contribute to a receptive endometrium are, at present, important areas of research. Investigation of endometrial function has traditionally been assessed by morphological criteria. However, the relationship between histological changes and endometrial receptivity has been seriously questioned in recent years and, consequently, a number of new biomarkers of endometrial receptivity have been described. The study of integrin αvβ3 and osteopontin (OPN) has been proposed as a means of distinguishing receptive from non-receptive endometrium in clinical practice and as a new method to investigate the impaired endometrial receptivity in certain groups of infertile patients. Both glycoproteins have been found to be coordinately expressed in the human endometrium across the menstrual cycle and maximally expressed at the time of the implantation window. According to these findings, we have developed and subsequently published the following studies: I. “Osteopontin and αvβ3 integrin expression in the endometrium of infertile and fertile women” Casals G, Ordi J, Creus M, Fábregues F, Casamitjana R, Quinto L, Campo E, Balasch J. Reprod Biomed Online, 2008;16(6):808-816 II. “Osteopontin and αvβ3 integrin as markers of endometrial receptivity: the effect of different hormone therapies” Casals G, Ordi J, Creus M, Fábregues F, Carmona F, Casamitjana R, Balasch J. Reprod Biomed Online, 2010;21(3):349-359 III. “Expression pattern of osteopontin and αvβ3 integrin during the implantation window in infertile patients with early stages of endometriosis” Casals G, Ordi J, Creus M, Fábregues F, Carmona F, Casamitjana R, Balasch J. Hum Reprod, 2012;27(3):805-813 The results of these studies indicate that although the expression of the OPN:αvβ3 integrin complex is closely correlated with histological maturation of endometrium evaluated by histological dating, neither OPN nor αvβ3 alone or in combination are useful markers of endometrial functional receptivity: there were no differences in expression or coexpression of these two markers between fertile controls and infertile patients. On the other hand, endometrial OPN and αvβ3 integrin expression or co-expression during the window of implantation are not impaired in patients with stage I–II endometriosis. Finally, OPN and alphavbeta3 integrin expression was closely related to endometrial maturation and this was irrespective of the hormonal treatment received. In conclusion, the results of our studies do not support the routine use of these markers in the clinical practice.
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Shuttleworth, Elinor. "The role of integrin αvβ8 on human monocytes and macrophages in intestinal immune homeostasis." Thesis, University of Manchester, 2018. https://www.research.manchester.ac.uk/portal/en/theses/the-role-of-integrin-alpha-v-beta-8-on-human-monocytes-and-macrophages-in-intestinal-immune-homeostasis(b1f62522-19ba-49d7-8b09-b9d8e1bbbf6b).html.
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Intestinal immune cells remain tolerant to the trillions of commensal bacteria present in the gut, with perturbations of this process implicated in development of inflammatory bowel disease (IBD). The cytokine TGF-β is a key factor promoting intestinal immune tolerance, but is secreted in a latent state that requires activation to function. Binding of TGF-β to the integrin αvβ8 is a principal mechanism of TGF-β activation, with mouse models demonstrating a crucial role for αvβ8 expression by dendritic cells and regulatory T cells in intestinal immune regulation. Despite this evidence, very little is known regarding the importance of this activating integrin in human intestinal homeostasis. Utilising flow cytometry here we find that integrin αvβ8 is highly expressed on peripheral blood monocytes with highest levels on intermediate CD14++CD16+ monocytes. Upon monocyte to macrophage differentiation high β8 expression is observed on anti-inflammatory M-CSF differentiated macrophages versus pro-inflammatory GM-CSF macrophages. In monocytes, expression of β8 is upregulated by specific bacterial TLR ligands. Utilising a TGF-β reporter cell line both monocytes and M-CSF MDM display an enhanced ability to activate TGF-β in an αvβ8-dependent manner. Data presented here indicate that macrophage αvβ8-dependent TGF-β activation does not alter expression of surface markers associated with a tolerogenic macrophage phenotype, phagocytosis, or production of the anti-inflammatory cytokine IL-10; nor does TGF-β appear to influence the metabolic profile of macrophages, key differences of which are associated with pro- or anti-inflammatory phenotype. However, the previously undescribed finding of integrin αvβ8 expression on human monocytes and macrophages, which was subsequently confirmed in intestinal populations and found to be downregulated in inflamed IBD mucosa, may highlight an important functional pathway in intestinal immune homeostasis and represent a potential future therapeutic target in IBD.
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Burgett, Monica E. "Direct contact with perivascular tumor cells enhances integrin αvβ3 signaling and migration of endothelial cells." Kent State University / OhioLINK, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=kent1466174564.
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FIRMO, MORAIS EDUARDO MANUEL. "Role of αVβ3 integrin in cortical synaptic transmission: relevance for Autism Spectrum Disorder and Epilepsy." Doctoral thesis, Università degli studi di Genova, 2019. http://hdl.handle.net/11567/942181.
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Autism Spectrum Disorder (ASD) is a group of neurodevelopmental disorders characterized by social communication impairments and restricted interests. Epilepsy is a brain disorder diagnosed after the occurrence of at least one epileptic seizure. ASD and epilepsy are often comorbid.
The integrin αVβ3 is a synaptic cell adhesion molecule that binds to counter-receptors and extracellular matrix proteins. Itgb3, the gene encoding the β3 subunit of integrin αVβ3, has been associated to ASD in humans, and constitutive knockout (KO) mice for this gene exhibit ASD-like behaviors, while brain-specific conditional KO mice for the other subunit of integrin αVβ3 (αV) have higher propensity for seizures.
Integrin αVβ3 may, therefore, play important neuronal roles in brain regions relevant for ASD and epilepsy. To test this hypothesis, I expanded the behavioral characterization of Itgb3 constitutive KO mice and characterized the behavioral phenotype of an Itgav conditional KO mouse specific for excitatory neurons of the forebrain. In these mouse models, I also investigated the synaptic dysfunctions of excitatory synapses onto layer V pyramidal neurons of the medial prefrontal cortex.
I find that loss of integrin αVβ3 leads to deficits in social novelty preference, hyper-reactivity to acoustic stimuli of high intensity, and reduces the threshold for pharmacologically induced seizures. This correlates with a reduction in the content of GluA2-lacking AMPA receptors in excitatory inputs to layer V pyramidal neurons.
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Raj, April. "Mechanistic studies to evaluate the targeting specificity of novel RGD Micelles to the αVβ3 integrin receptor." Scholarly Commons, 2012. https://scholarlycommons.pacific.edu/uop_etds/830.
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Current chemotherapeutics pose many di sadvantages due to their lack of specificity and low therapeutic index. To overcome these challenges, research has focused its attention on the development of nano-based delivery systems that can penetrate the leaky vasculature of tumor endothelium, use site-directed ligands that can bind with high affinity and specific ity to tumor cells, physically entrap poorly soluble drugs, and deliver these cytotoxic agents directly to the tumor site. One approach to nanosystem drug delivery is with the use of peptide amphiphiles (PAs) that are conjugated with the Arginine-Glycine-Aspartic Acid (RGD) motif to actively target a αVβ3 integrin receptors on cancer cells or tumor endothelium. The current work is focused on mechanistic studies to evaluate the uptake of novel RGD amphiphi les with varying alkyl chain lengths (palmitic acid : Cl 6 and stearic acid: C 18) and hydrophilic linkers, 8-amino- 3,6-dioxaoctonoic acid (ADA) or glucose, as micellar delivery systems of hydrophobic anticancer agents. PAs were confirmed for their self-assembling properties and further evaluated for their RGD-mediated binding specificity to purified αVβ3 integrin through a competitive binding fluorescence polarization assay (with novel RGD micelles displacing an integrin-bound fluorescent RGD probe by as much as 63.03%). Ultimately, these nanocarriers were assessed for their ability to deliver phys ically entrapped fluorescein isoth iocyanate (FITC) to A2058 cells overexpressing αVβ3 integrin receptors. Results from confocal microscopy indicate that uptake of RGD micelles was driven by an energy-dependent mechanism, as statistically significant levels of FITC internalization was seen at 37°C versus 4°C (p-value<0.05 for all treatment groups); moreover, intracellular fluorescence was notably higher (as much as 4-fold) when delivered through novel RGD conjugates as opposed to its free form. Regardless of chain length and the number of hydrophilic linkers, all RGD PAs showed promising results as micellar carriers that can effectively deliver their payload to the target tumor site via receptor mediated endocytosis.
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Massaro, Raffaele <1988>. "Interaction of Glycoproteins H/L from human α-herpesviruses with αvβ integrins." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2017. http://amsdottorato.unibo.it/7967/1/raffaele_massaro_tesi.pdf.
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Herpes Simplex Virus (HSV) and Varicella-Zoster Virus (VZV) belong to the viral subfamily Alphaherpesvirinae, and are important neurotropic human pathogens. HSV and VZV fuse their envelope with the membrane of the host cell, a process termed cell entry, accomplished by a conserved trio of viral glycoproteins (gB, gH and gL). Host cell receptors mediate viral entry, i.e. nectin1 is recognized by the viral tropism factor gD in HSV, while the host receptor is still elusive in VZV. Integrins are widely distributed adhesion membrane proteins, and give critical contribution to the entry of most human Herpesviruses. αvβ6 or αvβ8 integrins are interchangeable receptors for HSV gH/gL, and they could synchronize fusion exerted by gB with the virion endocytosis they mediate. αvβ6 or αvβ8 integrins modify the glycoprotein machinery of HSV, but the modification they induce on gH/gL heterodimer was never investigated. The results presented here argue in favor of an integrin dependent mechanism which involves the conserved glycoprotein apparatus of HSV and seems to be essential to HSV entry. This mechanism, which also depends on the presence of gB and nectin1-bound gD, entails the dissociation of gL from gH/gL heterodimer, and its release in the media of culture cells upon binding of αvβ6 or αvβ8 integrins receptors. Recent findings disclose both the resemblance between HSV and VZV gH/gL heterodimer structures and the role of integrins of the alpha V family as receptors for VZV entry, a role probably shared with Myelin Associated Glycoprotein (MAG). However, while MAG function in fusion was associated with VZV gB, it was not clarified which glycoprotein(s) was(were) involved in integrin-mediated VZV entry. Data presented here argue in favor of integrin direct binding with a part of the conserved VZV entry machinery, while VZV fusion seems to be the result of more complex interactions.
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Ellison, Timothy. "Elucidating how αvβ3-integrin regulates neuropilin-1's role in tumour angiogenesis in order to improve anti-angiogenic therapy." Thesis, University of East Anglia, 2015. https://ueaeprints.uea.ac.uk/57412/.
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αvβ3-integrin expression is vastly upregulated in tumour vasculature, and has long been considered a key molecule in promoting tumour angiogenesis. However, its initial promise as a target in anti-angiogenic therapy has wavered since the αvβ3-integrin antagonist cilengitide did not meet its end point in Phase III clinical trials for the treatment of glioblastoma. This failure corresponds with the enhanced tumour growth and angiogenesis observed in β3-integrin-knockout mice, which potentially occurs via a compensatory upregulation of VEGFR2 and enhancement in VEGFR2-neuropilin-1 interactions. Here, I show that tumour growth and angiogenesis are sensitive to neuropilin-1 perturbation even with only a 50% reduction in β3-integrin expression. β3-integrin-heterozygous, but not wild-type, mice show an increased dependence on neuropilin-1 that is not related to changes in neuropilin-1-mediated VEGFR2 function. Rather, the suppression of β3-integrin leads to the activation of a neuropilin-1-dependent endothelial cell migration pathway via a mechanism in which NRP1 is mobilised away from mature focal adhesions following VEGF-stimulation. Concordantly, the simultaneous genetic targeting of both molecules significantly impairs paxillin activation and focal adhesion turnover in endothelial cells, and thus inhibits endothelial cell migration, and tumour growth and angiogenesis, even in established tumours. These findings therefore provide important pre-clinical evidence that pharmacologically targeting both molecules in unison might be an effective anti-angiogenic therapy for patients with advanced cancers.
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Lidén, Åsa. "Integrin αVβ3-Directed Contraction by Connective Tissue Cells : Role in Control of Interstitial Fluid Pressure and Modulation by Bacterial Proteins." Doctoral thesis, Uppsala University, Department of Medical Biochemistry and Microbiology, 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-6601.
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This thesis aimed at studying mechanisms involved in control of tissue fluid homeostasis during inflammation.
The interstitial fluid pressure (PIF) is of importance for control of tissue fluid balance. A lowering of PIF in vivo will result in a transport of fluid from the circulation into the tissue, leading to edema. Loose connective tissues that surround blood vessels have an intrinsic ability to take up fluid and swell. The connective tissue cells exert a tension on the fibrous network of the tissues, thereby preventing the tissues from swelling. Under normal homeostasis, the interactions between the cells and the fibrous network are mediated by β1 integrins. Connective tissue cells are in this way actively controlling PIF.
Here we show a previously unrecognized function for the integrin αVβ3, namely in the control of PIF. During inflammation the β1 integrin function is disturbed and the connective tissue cells release their tension on the fibrous network resulting in a lowering of PIF. Such a lowering can be restored by platelet-derived growth factor (PDGF) -BB. We demonstrated that PDGF-BB restored PIF through a mechanism that was dependent on integrin αVβ3. This was shown by the inability of PDGF-BB to restore a lowered PIF in the presence of anti-integrin β3 IgG or a peptide inhibitor of integrin αVβ3. PDGF-BB was in addition unable to normalize a lowered PIF in β3 null mice. Furthermore, we demonstrated that extracellular proteins from Streptococcus equi modulated αVβ3-mediated collagen gel contraction. Because of the established concordance between collagen gel contraction in vitro and control of PIF in vivo, a potential role for these proteins in control of tissue fluid homeostasis during inflammation could be assumed. Sepsis and septic shock are severe, and sometimes lethal, conditions. Knowledge of how bacterial components influence PIF and the mechanisms for tissue fluid control during inflammatory reactions is likely to be of clinical importance in treating sepsis and septic shock.
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Broich, Kerstin [Verfasser]. "Importance of αvβ3 [alpha-v-beta-3] integrin in arteriogenesis in the peripheral circulation of the rabbit / eingereicht von Kerstin Broich." Gießen : DVG-Service, 2004. http://d-nb.info/972754466/34.
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Ribeiro, Lívia Carolina de Abreu. "Efeito da desintegrina recombinante DisBa-01 incorporada em micelas ou livre em células portando ou não a integrina αvβ3 /." Araraquara, 2013. http://hdl.handle.net/11449/108420.
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Orientador :Iracilda Zeppone CarlosBanca: Danielle Cardoso Geraldo Maia
Banca: Márcia Antoniazi Michelin
Banca: Dagmar Ruth Stach-Machado
Banca: Leila Aparecida Chiavacci
Resumo: Introdução: a integrina αvβ3 causa adesão celular à vitronectina e está expressa em diversos tumores humanos, mas está em níveis muito reduzidos nos tecidos normais, sendo mais notável em células derivadas da medula óssea. A desintegrina recombinante DisBa-01 se liga à integrina αvβ3, inibindo a adesão celular à vitronectina. Apresenta também uma capacidade antimetastática in vivo e antitrombótica in vitro. As micelas são sistemas de transporte que podem direcionar o fármaco ao tecido alvo de forma passiva, se acumulando onde a microvasculatura esteja mais permeável, como no caso do câncer. Objetivos: verificar a perda de adesão, a liberação de mediadores, a citotoxicidade e a anoikis gerados pela desintegrina DisBa-01, tanto em sua forma livre como incorporada em micelas, em linhagens celulares que expressem ou não a integrina αvβ3. Métodos: a desintegrina foi expressa em bactérias Escherichia coli a partir de um cDNA fusionado ao vetor pET28a, e então purificada por cromatografias e diálises. As micelas controle (M-C) ou contendo a desintegrina (M-DB) foram estruturadas com polissorbato 80 e fosfatidilcolina de soja, e foi observado o diâmetro médio e índice de polidispersidade por espalhamento dinâmico de luz, o potencial zeta por microeletroforese, a eficiência de encapsulação por dosagem protéica de Lowry, a avaliação estrutural da desintegrina encapsulada por espectroscopias de dicroísmo circular e de fluorescência e a estrutura das micelas pela curva de SAXS. As micelas e a proteína livre foram testadas em linhagens HUVEC e SC, contendo a integrina αvβ3, e K562, não contendo esta integrina. Foram avaliados a citotoxicidade pelo teste de MTT, a inibição de adesão celular ou descolamento a vitronectina e fibronectina por quantificação das células aderidas com cristal violeta, a anoikis por observação das células viáveis e em apoptose se aderidas ou não por MTT, Apo-Direct ...
Abstract: Introduction: integrin αvβ3 sustain cellular adhesion to vitronectin and is expressed on a diversity of human tumors but is present at very low levels on normal tissues, with notable expression on bone marrow-derived cells. The recombinant disintegrin DisBa-01 binds to αvβ3 integrin, inhibiting cell adhesion to vitronectin. It also features an in vivo anti-metastatic ability and in vitro anti-thrombotic function. Micelles are a transport system that can direct a drug to the target tissue passively, accumulating where microvasculature is more permeable, which happens on cancer. Objectives: to verify adhesion loss, mediators production, citotoxicity and anoikis generated by disintegrin DisBa-01, both in its free form or incorporated into micelles, in cell lines expressing or not αvβ3 integrin. Methods: the disintegrin was expressed in Escherichia coli using a cDNA fused to vector pET28a, and then purified by chromatography and dialyses. Control micelles (M-C) or containing disintegrin (M-DB) were assembled using polysorbate 80 and soy phosphatidylcholine, and it was observed the average diameter and polydispersity index by dynamic light scattering, zeta potential by microelectrophoresis, the efficiency of encapsulation by protein determination using Lowry reagent, structural assessment of disintegrin encapsulated by circular dichroism spectroscopy and fluorescence spectroscopy and micelles' structure by SAXS curve. The protein free or incorporated into micelles were tested in HUVEC and SC, containing integrin αvβ3, and in K562, not containing this integrin. Cytotoxicity was evaluated by MTT assay, inhibition of cell adhesion and detachment to vitronectin or fibronectin by quantifying the adherent cells with crystal violet, the anoikis by observation of viable cells and apoptosis, whether attached or not, by MTT, Apo-Direct and annexin V, and production of mediators VEGF-A, IL-8, TGF-β, TNF-α, IL-12 and ...
Doutor
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Ribeiro, Lívia Carolina de Abreu [UNESP]. "Efeito da desintegrina recombinante DisBa-01 incorporada em micelas ou livre em células portando ou não a integrina αvβ3." Universidade Estadual Paulista (UNESP), 2013. http://hdl.handle.net/11449/108420.
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Bolsa de estudos
Introdução: a integrina αvβ3 causa adesão celular à vitronectina e está expressa em diversos tumores humanos, mas está em níveis muito reduzidos nos tecidos normais, sendo mais notável em células derivadas da medula óssea. A desintegrina recombinante DisBa-01 se liga à integrina αvβ3, inibindo a adesão celular à vitronectina. Apresenta também uma capacidade antimetastática in vivo e antitrombótica in vitro. As micelas são sistemas de transporte que podem direcionar o fármaco ao tecido alvo de forma passiva, se acumulando onde a microvasculatura esteja mais permeável, como no caso do câncer. Objetivos: verificar a perda de adesão, a liberação de mediadores, a citotoxicidade e a anoikis gerados pela desintegrina DisBa-01, tanto em sua forma livre como incorporada em micelas, em linhagens celulares que expressem ou não a integrina αvβ3. Métodos: a desintegrina foi expressa em bactérias Escherichia coli a partir de um cDNA fusionado ao vetor pET28a, e então purificada por cromatografias e diálises. As micelas controle (M-C) ou contendo a desintegrina (M-DB) foram estruturadas com polissorbato 80 e fosfatidilcolina de soja, e foi observado o diâmetro médio e índice de polidispersidade por espalhamento dinâmico de luz, o potencial zeta por microeletroforese, a eficiência de encapsulação por dosagem protéica de Lowry, a avaliação estrutural da desintegrina encapsulada por espectroscopias de dicroísmo circular e de fluorescência e a estrutura das micelas pela curva de SAXS. As micelas e a proteína livre foram testadas em linhagens HUVEC e SC, contendo a integrina αvβ3, e K562, não contendo esta integrina. Foram avaliados a citotoxicidade pelo teste de MTT, a inibição de adesão celular ou descolamento a vitronectina e fibronectina por quantificação das células aderidas com cristal violeta, a anoikis por observação das células viáveis e em apoptose se aderidas ou não por MTT, Apo-Direct ...
Introduction: integrin αvβ3 sustain cellular adhesion to vitronectin and is expressed on a diversity of human tumors but is present at very low levels on normal tissues, with notable expression on bone marrow-derived cells. The recombinant disintegrin DisBa-01 binds to αvβ3 integrin, inhibiting cell adhesion to vitronectin. It also features an in vivo anti-metastatic ability and in vitro anti-thrombotic function. Micelles are a transport system that can direct a drug to the target tissue passively, accumulating where microvasculature is more permeable, which happens on cancer. Objectives: to verify adhesion loss, mediators production, citotoxicity and anoikis generated by disintegrin DisBa-01, both in its free form or incorporated into micelles, in cell lines expressing or not αvβ3 integrin. Methods: the disintegrin was expressed in Escherichia coli using a cDNA fused to vector pET28a, and then purified by chromatography and dialyses. Control micelles (M-C) or containing disintegrin (M-DB) were assembled using polysorbate 80 and soy phosphatidylcholine, and it was observed the average diameter and polydispersity index by dynamic light scattering, zeta potential by microelectrophoresis, the efficiency of encapsulation by protein determination using Lowry reagent, structural assessment of disintegrin encapsulated by circular dichroism spectroscopy and fluorescence spectroscopy and micelles’ structure by SAXS curve. The protein free or incorporated into micelles were tested in HUVEC and SC, containing integrin αvβ3, and in K562, not containing this integrin. Cytotoxicity was evaluated by MTT assay, inhibition of cell adhesion and detachment to vitronectin or fibronectin by quantifying the adherent cells with crystal violet, the anoikis by observation of viable cells and apoptosis, whether attached or not, by MTT, Apo-Direct and annexin V, and production of mediators VEGF-A, IL-8, TGF-β, TNF-α, IL-12 and ...
FAPESP: 10/05428-0
FAPESP: 10/01568-2
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Müller, Martina Verfasser], Horst [Akademischer Betreuer] [Kessler, Ute [Akademischer Betreuer] Reuning, and Steffen Johannes [Akademischer Betreuer] Glaser. "Impact of the Integrin αvβ3 Transmembrane Domain Sequence and the Cytoplasmic Contacts on Cell/Matrix Adhesiveness and Integrin-mediated Cell Signalling / Martina Müller. Gutachter: Ute Reuning ; Steffen Johannes Glaser. Betreuer: Horst Kessler." München : Universitätsbibliothek der TU München, 2012. http://d-nb.info/1019853611/34.
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Brunie, Leonora Verfasser], Ute [Akademischer Betreuer] Reuning, Ernst J. [Akademischer Betreuer] Rummeny, and Barbara [Akademischer Betreuer] [Schmalfeldt. "Generation of integrin αvβ3 cytoplasmic and transmembrane domain mutants: characterisation of the impact of integrin conformation on human ovarian cancer cell proliferation / Leonora Brunie. Gutachter: Ute Reuning ; Ernst J. Rummeny ; Barbara Schmalfeldt. Betreuer: Ute Reuning." München : Universitätsbibliothek der TU München, 2012. http://d-nb.info/1031513906/34.
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Leoni, Valerio <1985>. "Role of αvβ3 – Integrin and TLR2 in the innate response to Herpes Simplex Virus infection and Delivery of retargeted oncolytic Herpes Simplex via carrier cells." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2014. http://amsdottorato.unibo.it/6464/1/Leoni_Valerio_tesi.pdf.
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In the first part of my thesis I studied the mechanism of initiation of the innate response to HSV-1. Innate immune response is the first line of defense set up by the cell to counteract pathogens infection and it is elicited by the activation of a number of membrane or intracellular receptors and sensors, collectively indicated as PRRs, Patter Recognition Receptors. We reported that the HSV pathogen-associated molecular patterns (PAMP) that activate Toll-like receptor 2 (TLR2) and lead to the initiation of innate response are the virion glycoproteins gH/gL and gB, which constitute the conserved fusion core apparatus across the Herpesvirus. Specifically gH/gL is sufficient to initiate a signaling cascade which leads to NF-κB activation. Then, by gain and loss-of-function approaches, we found that αvβ3-integrin is a sensor of and plays a crucial role in the innate defense against HSV-1. We showed that αvβ3-integrin signals through a pathway that concurs with TLR2, affects activation/induction of interferons type 1, NF-κB, and a polarized set of cytokines and receptors. Thus, we demonstrated that gH/gL is sufficient to induce IFN1 and NF-κB via this pathway. From these data, we proposed that αvβ3-integrin is considered a class of non-TLR pattern recognition receptors. In the second part of my thesis I studied the capacity of human mesenchymal stromal cells isolated by fetal membranes (FM-hMSCs) to be used as carrier cells for the delivery of retargeted R-LM249 virus. The use of systemically administrated carrier cells to deliver oncolytic viruses to tumoral targets is a promising strategy in oncolytic virotherapy. We observed that FM-hMSCs can be infected by R-LM249 and we optimized the infection condition; then we demonstrate that stromal cells sustain the replication of retargeted R-LM249 and spread it to target tumoral cells. From these preliminary data FM-hMSCs resulted suitable to be used as carrier cells
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Leoni, Valerio <1985>. "Role of αvβ3 – Integrin and TLR2 in the innate response to Herpes Simplex Virus infection and Delivery of retargeted oncolytic Herpes Simplex via carrier cells." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2014. http://amsdottorato.unibo.it/6464/.
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In the first part of my thesis I studied the mechanism of initiation of the innate response to HSV-1. Innate immune response is the first line of defense set up by the cell to counteract pathogens infection and it is elicited by the activation of a number of membrane or intracellular receptors and sensors, collectively indicated as PRRs, Patter Recognition Receptors. We reported that the HSV pathogen-associated molecular patterns (PAMP) that activate Toll-like receptor 2 (TLR2) and lead to the initiation of innate response are the virion glycoproteins gH/gL and gB, which constitute the conserved fusion core apparatus across the Herpesvirus. Specifically gH/gL is sufficient to initiate a signaling cascade which leads to NF-κB activation. Then, by gain and loss-of-function approaches, we found that αvβ3-integrin is a sensor of and plays a crucial role in the innate defense against HSV-1. We showed that αvβ3-integrin signals through a pathway that concurs with TLR2, affects activation/induction of interferons type 1, NF-κB, and a polarized set of cytokines and receptors. Thus, we demonstrated that gH/gL is sufficient to induce IFN1 and NF-κB via this pathway. From these data, we proposed that αvβ3-integrin is considered a class of non-TLR pattern recognition receptors. In the second part of my thesis I studied the capacity of human mesenchymal stromal cells isolated by fetal membranes (FM-hMSCs) to be used as carrier cells for the delivery of retargeted R-LM249 virus. The use of systemically administrated carrier cells to deliver oncolytic viruses to tumoral targets is a promising strategy in oncolytic virotherapy. We observed that FM-hMSCs can be infected by R-LM249 and we optimized the infection condition; then we demonstrate that stromal cells sustain the replication of retargeted R-LM249 and spread it to target tumoral cells. From these preliminary data FM-hMSCs resulted suitable to be used as carrier cells
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This, Sébastien. "Régulation des réponses immunitaires adaptives par l'intégrine αvβ8 - Implications pour l'immunité des muqueuses et la réponse humorale." Thesis, Lyon, 2020. http://www.theses.fr/2020LYSEN011.
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A la suite d’une infection, la capacité de l’hôte à produire une réponse immunitaire efficace et adaptée est cruciale pour se défendre et générer une réponse mémoire permettant de se protéger contre une future réinfection. En revanche, il est absolument nécessaire que cette réponse soit correctement régulée afin d’éviter sa persistante ou son activation incontrôlée pouvant mener au développement de maladies immunitaires. Le système immunitaire comprend un ensemble complexe de cellules et de molécules immunitaires. En particulier, la capacité des cellules immunitaires à produire et à répondre aux cytokines est absolument critique pour l’orchestration des réponses immunitaires. Mon travail de thèse s’est intéressé à une molécule particulière, la cytokine Transforming Growth Factor β ou TGFβ. Contrairement à la plupart des autres cytokines, le TGFβ est produit sous une forme latente et doit être activé afin de pouvoir se lier à son récepteur et induire une réponse sur la cellule cible. Nous avons montré par le passé que l’intégrine αvβ8 joue un rôle clé dans le processus d’activation du TGFβ et ainsi dans la régulation des réponses immunitaires dépendantes du TGFβ.Plus précisément, j’ai cherché au cours de ma thèse à comprendre le rôle de l’intégrine αvβ8 dans la régulation des réponses immunitaires intestinales et la régulation des réponses humorales.Mon travail s’est plus particulièrement concentré sur la compréhension de trois processus : 1/ l’induction de lymphocytes TREG et TH17 dans l’intestin nécessaires au maintien de l’homéostasie immunitaire intestinale, 2/ la régulation des réponses humorales de classe A (IgA) intestinales et 3/ la régulation des réponses humorales T-dépendantes au cours de la réaction du Centre GerminatifThe ability of a host to generate an appropriate immune response is critical to provide protection against a particular pathogen and to provide long-lasting memory against future reinfection. However, this immune response must be tightly regulated to prevent its persistence or inadequate activation which can lead to the development of immune pathologies. Mammalian immune system comprises a wide array of immune cells and molecules. In particular, the ability ofimmune cells to secrete and respond to cytokines is central to the orchestration of immune responses. My PhD project has focused on the role of a particular cytokine named Transforming Growth Factor β (TGFβ). Unlike most other cytokines, TGFβ is secreted in a latent form and must be activated to bind its receptor and induce response on target cell. Our team and others have shown that αvβ8 integrin plays a critical role in TGFβ activation and thus the regulation of TGFβ-dependent immune responses. More precisely, I investigated the role of αvβ8 integrin in the regulation of intestinal immunityand humoral B cell responses. In particular, my work focused on three immune processes: 1/ the induction of TREG and TH17 in Mucosal Associated Lymphoid Tissues and 2/ the regulation ofintestinal IgA humoral responses and 3/ the regulation of T-dependent B cell responses during the germinal center reaction
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Schmitz, Carla Regina. "Avaliação da Milk Fat Globule Epidermal Growth Factor 8 (MFG-E8), da integrina αvβ3 e da Leukemia Inhibitory Factor (LIF) na implantação embrionária humana : estudo em modelo in vitro e no endométrio de mulheres com e sem endometriose." reponame:Biblioteca Digital de Teses e Dissertações da UFRGS, 2015. http://hdl.handle.net/10183/131165.
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Base teórica: O processo de implantação do embrião no ser humano é extremamente complexo e, ao mesmo tempo, essencial para que a mulher possa engravidar. Neste processo, em que o endométrio precisa sofrer uma série de mudanças para tornar-se receptivo, a adequada expressão de MFG-E8 (milk fat globule epidermal growth factor 8), seu receptor a integrina αvβ3 e LIF (leukemia inhibitory factor) parecem ter um papel importante. Além do mais, mulheres com infertilidade e endometriose podem apresentar a falha de implantação como uma grande barreira para obter seu sucesso terapêutico. Objetivos: Avaliar o papel de MFG-E8 e do seu receptor integrina αvβ3 em um modelo de implantação in vitro com uma linhagem celular trofoblástica e outra de epitélio endometrial. Comparar a expressão de MFG-E8, de integrina αvβ3 e de LIF no endométrio de pacientes férteis e inférteis com endometriose durante a janela de implantação. Métodos: No primeiro ensaio, utilizando-se uma linhagem celular bem diferenciada de adenocarcinoma de endométrio (células Ishikawa) e uma linhagem de coriocarcinoma de trofoblasto, o modelo in vitro de implantação humana foi estabelecido. Para investigação do impacto do bloqueio de MFG-E8 e integrina αvβ3, ambas linhagens celulares foram pré-tratadas com anticorpos contra estas proteínas em diferente concentrações antes do ensaio de adesão. No ensaio subsequente, para comparar a expressão de MFG-E8, de integrina αvβ3 e de LIF no endométrio humano, foram realizadas biópsias no período da janela de implantação (LH+7 a LH+10) com cateter de Pipelle. As amostras foram submetidas a imunohistoquímica, e analisadas através do HSCORE. Resultados: Na avaliação in vitro observamos que as células Ishikawa pré-tratadas com anticorpo anti-MFG-E8 causaram diminuição da adesão das esferas Jar dose-dependente. Por outro lado, o pré-tratamento das esferas Jar não resultou em diminuição significativa da adesão. Pré-tratamento com anticorpos anti-integrina αvβ3, tanto de células Ishikawa como de esferas Jar, causaram inibição significativa, dose-dependente, da adesão das esferas. A análise imunohistoquímica das biópsias realizadas durante a janela de implantação mostrou uma expressão aumentada de MFG-E8 em pacientes com endometriose e infertilidade. Além do mais, houve expressão diminuída de LIF no grupo em estudo. Contudo, não houve diferença estatisticamente significativa na expressão de integrina αvβ3 entre os grupos em estudo. Conclusão: Este estudo demonstrou que, quando se bloqueia MFG-E8 ou seu receptor integrina αvβ3 em células Ishikawa em um modelo in vitro, ocorre uma diminuição de adesão das células Jar. Além do mais, bloqueando-se a integrina αvβ3 nas esferas Jar, também ocorre uma diminuição da adesão destas nas células Ishikawa. No entanto, quando estudamos o endométrio in vivo de pacientes com endometriose e infertilidade, encontramos a expressão aumentada de MFG-E8 e diminuída de LIF durante a janela de implantação no endométrio.Background: The human implantation process is very complex and, at the same time, it is essential for women to achieve pregnancy. In this process, where the human endometrium must go through a lot of changes in order to become receptive, an adequate expression of MFG-E8 (milk fat globule epidermal growth factor 8), integrin αvβ3 and LIF (leukemia inhibitory factor) appear to play an important role. Furthermore, women with endometriosis and infertility may have in their implantation process the key to achieve pregnancy. Objectives: To investigate the role of MFG-E8 and its receptor integrin αvβ3 in the attachment of trophoblast cells to the endometrial epithelium, in an in vitro model. To compare endometrial expression of MFG-E8, integrin αvβ3 and LIF between fertile patients and patients with endometriosis and infertility during the window of implantation. Methods: In our first assay, by using a well-differentiated endometrial adenocarcinoma cell line (Ishikawa cells) and choriocarcinoma human trophoblast cells (Jar cells), an in vitro model mimicking human implantation was established. To investigate the impact of blocking MFG-E8 and integrin αvβ3, the cell lines were pretreated with antibodies against those proteins at different concentrations before the attachment assay. Moreover, to compare endometrial expression of MFG-E8, integrin αvβ3 and LIF, endometrial biopsies were performed during the window of implantation (LH+7 to LH+10) with the Pipelle catheter. The samples were submitted immunochemistry, and analyzed with HSCORE. Results: Pretreatment of Ishikawa cells with anti-MFG-E8 antibody caused a dosedependent and significant inhibition of attachment is our in vitro assay. On the other hand, pretreatment of Jar spheroids did not result in a significant effect on the attachment rate. Pretreatment of Ishikawa cells as well as Jar spheroids with anti-integrin avb3 antibodies resulted in a dose-dependent, significant inhibition of attachment. The immunochemistry analysis of the endometrial biopsies performed during the window of implantation showed increased MFG-E8 expression in patients with endometriosis and infertility. Moreover, there was lower LIF expression in the study group. Conclusion: This study showed that blocking MFG-E8 and its receptor integrin αvβ3 in Ishikawa cells diminishes Jar spheroid attachment in an in vitro model. Moreover, blocking integrin αvβ3 in the trophoblastic cells also diminished their attachment to the Ishikawa monolayer. Nevertheless, when we studied the endometrium of patients with endometriosis and infertility, we saw an increased expression of MFG-E8 and decreased expression of LIF during the window of implantation.
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Braeuer, Miriam [Verfasser], and Heidrun [Akademischer Betreuer] Potschka. "Komparative Evaluation der Tracer Avebetrin und Aquibeprin zur Detektion der Integrine α5β1 und αvβ3 als Prädiktionsmarker der Revaskularisierung nach akutem Herzinfarkt im Rattenmodell / Miriam Braeuer ; Betreuer: Heidrun Potschka." München : Universitätsbibliothek der Ludwig-Maximilians-Universität, 2018. http://d-nb.info/116957212X/34.
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Geiger, Pamina Xenia Charlotte [Verfasser], Ute [Akademischer Betreuer] Reuning, Manfred [Akademischer Betreuer] Schmitt, and Ernst J. [Akademischer Betreuer] Rummeny. "Einfluss des Metastasierungssuppressors KAI1 CD 82 auf das Integrin αvβ3-vermittelte Migrations- und spreading-Verhalten humaner Ovarialkarzinomzellen / Pamina Xenia Charlotte Geiger. Gutachter: Ute Reuning ; Manfred Schmitt ; Ernst J. Rummeny. Betreuer: Ute Reuning." München : Universitätsbibliothek der TU München, 2012. http://d-nb.info/1031512500/34.
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Upheber, Sina [Verfasser], Ute [Akademischer Betreuer] Reuning, Manfred [Akademischer Betreuer] Schmitt, and Barbara [Akademischer Betreuer] Schmalfeldt. "Die Integrin αvβ3-vermittelte Migration humaner Ovarialkarzinomzellen als Funktion des Metastasierungssuppressors KAI1 (CD82) und seiner Splice-Variante sowie deren Interaktion mit dem Zell/Zell-Adhäsionsmolekül E-Cadherin / Sina Upheber. Gutachter: Barbara Schmalfeldt ; Manfred Schmitt. Betreuer: Ute Reuning ; Manfred Schmitt." München : Universitätsbibliothek der TU München, 2015. http://d-nb.info/1075595924/34.
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Héroux, Julie. "Des lapins watanabe au syndrome hyper IgE humain : caractérisation précoce de l'athérosclérose utilisant une probe optique ciblant l'integrin aVb3." Thesis, Grenoble, 2012. http://www.theses.fr/2012GRENS041/document.
Full textAbstract:
La détection précoce de l’athérosclérose, avant le développement de ses séquellespathologiques, comme l’infarctus du myocarde, l’angine ou l’accident cérébrauxvasculaire(ACV), représente un important défi au niveau de la médecine diagnostiqueactuelle. Malgré les récentes avances technologiques, les maladies cardiovasculairesdemeurent la principale cause de décès dans les pays occidentaux et la détection à unstage plus précoce s’avère nécessaire pour permettre une intervention thérapeutiqueadéquate. Notre étude se concentre sur la détection de l’athérosclérose, plusspécifiquement la vulnérabilité de la plaque, grâce à l’imagerie moléculaire combinée àl’observation pathologique. Afin de prédire la rupture de la plaque, l’imageriemoléculaire a émergé comme outil diagnostique puissant suite au développementcroissant de sondes ayant de l’affinité pour les molécules cibles du processusd’athérosclérose. Comme résultantes, ces molécules sélectives possédant une forteaffinité pour des cibles surexprimées durant le processus de formation de la plaque,comme l’αvβ3 par exemple, devrait représentées des sondes prometteuses pour ladétection de l’athérosclérose.Objectif L’objectif global de notre étude était d’évaluer et de prédire lavulnérabilité de la plaque d’athérome à l’aide de différents marqueurs moléculaires. Leprincipal objectif de notre recherche était d’évaluer la possibilité de détecter précocementla plaque en utilisant une ITOP (integrin targeted optical probe). Cette sonde synthétiquenouvellement développée et ciblant l’intégrine αvβ3 avait déjà démontré une affinité etspécificité particulièrement élevée pour le récepteur de l’αvβ3 dans le cancer. Nousavons également exploré la relation entre cette sonde et l’observation pathologique desplaques d’athéromes sur le modèle animale WHHL et sur des plaques humainesprovenant de différents patients.Procédure et Résultats Les expériences ont été réalisées sur un total de 12 lapinsWatanabe hyperlipidémiques de souche WHHL (Watanabe heritable hyperlipidemic) et 1lapin contrôle NZW (New Zealand White). Premièrement, notre ITOP, marquée avec lafluorescéine isothiocyanate (FITC), a été utilisée pour détecter in vitro et ex vivo laprésence du récepteur de l’αvβ3. La microscopie à fluorescence a révélé un importantmarquage de la plaque d’athérome, lequel était absent dans les tissus provenant des lapinscontrôles NZW. Le marquage a été détecté au niveau de segments de plaques provenantde deux régions distinctes de l’aorte ascendante et descendante dans chaque lapin. Lesignal a été détecté principalement au niveau de l’adventitia et de l’intima proximale desvaisseaux aortiques, correspondant directement à l’expression de l’intégrine αvβ3,déterminée par essai immunochimique avec un anticorps contre l’αvβ3. De plus, uneforte association s’est révélée entre le niveau de marquage de la sonde ciblant l’αvβ3 etl’épaisseur de l’adventitia. Deuxièmement, nous avons évalué notre sonde sur deséchantillons humains affectés par l’athérosclérose et comparé les résultats avec uneévaluation morphologique. Nous avons remarqué la même tendance que chez le lapin, soiun marquage plus important lorsque l’adventitia s’épaissi. Finalement, nous avons testé lasonde sur des artères coronaires provenant d’une autopsie d’un patient affecté par le ADHIESet comparé les résultats avec l’évaluation morphologique de leurs artèrescoronaires. Nous avons trouvé un lien entre la morphologie de la plaque et la prévalenced’anévrysmes coronaires chez ces patients.Conclusion L’expression de l’αvβ3 est reliée à la foi aux processus inflammatoires età la sténose. Notre ITOP à marqué efficacement in vitro le premier type de plaqued’athérome classé comme avancé (type IV) et pouvant produire des manifestationscliniques. En combinaison avec l’imagerie noninvasive détectant la sténose, il pourraits’avéré utile dans la détection de la plaque vulnérablePurpose The detection of early atherosclerosis, before the development of its later sequelae of myocardial infarction, angina or stroke, constitutes an important challenge in current diagnostic medicine. Despite all the recent technological advances, cardiovascular disease remains the leading cause of death in the Western World and needs to be detected at an earlier stage to allow for more timely therapeutic intervention. This study is focusing on the detection of atherosclerosis or more specifically plaque vulnerability with the help of molecular imaging and pathological observation. Effectively, to predict plaque rupture, molecular imaging has emerged as a powerful diagnostic tool, consequent to the development of a growing number of new probes with affinity for key molecular targets. As a result, such selective molecule with high affinity for overexpressed target in plaque formation, as αvβ3 integrin, should have promise as a probe for imaging atherosclerosis. With the help of molecular imaging combined with pathological observations, we can better comprehend, predict, and detect plaque vulnerability and rupture. Objectives The overall objective of this study is to evaluate different molecular tools to predict the vulnerability of the atheromatous plaque. The major objective of the research was to investigate the possibility of detecting atherosclerotic plaque by using a newly developed synthetic αvβ3 integrin targeted optical probe (ITOP) showing particularly high affinity and specificity for the αvβ3 receptor. We also investigate the relation between this probe and pathological observation of atherosclerotic plaques from WHHL animal model and different human samples. Procedures and Results For this study, experiments were performed on 12 Watanabe heritable hyperlipidemic (WHHL) rabbits and 1 New Zealand White (NZW) rabbits for control. First, our ITOP labeled with fluorescein isothiocyanate was used for detecting the presence of αvβ3 receptors in vitro and ex vivo on a Watanabe rabbit model. Fluorescence microscopy demonstrated a strong labeling of atherosclerotic plaques, which was absent in tissue from normal NZW rabbits. Segments of plaque accumulation from two distinct regions of ascending and descending aortas were labeled in each rabbit. The signal was found principally in the adventitia and proximal intima of the aortic vessel, corresponding directly to the expression of integrin αvβ3 as determined by antibody assay. Moreover, there was a close association between the level of labeling with the αvβ3 targeted probe and the thickness of the adventitia. Secondly, the ITOP was evaluated on human atherosclerotic samples, and was found to efficiently labeled atherosclerotic plaques. Moreover, we observed the same tendency as in the Watanabe rabbit: the ITOP intensity correlated with the degree of adventitial thickening. Finally, we tested the ITOP on Job's Syndrome coronary arteries, and have been able to detect a plaque corresponding to the first type of advanced atherosclerosis (type IV). We also found a relationship between plaque morphology and predisposition to aneurysms in Job's syndrome. Conclusions αvβ3 expression is related to inflammatory and stenotic processes. Our ITOP can efficiently label in vitro the first type of advanced atherosclerotic plaque. In combination with noninvasive imaging techniques that evaluate stenosis, it has great potential for the detection of vulnerable plaque
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Garcia, Fulle Maria Isabel. "Expression of αvβ6 integrin in the junctional epithelium." Thesis, 2005. http://hdl.handle.net/2429/16745.
Full textAbstract:
Cell-cell binding and cell-extracellular matrix adhesion is mediated by cell surface receptors
known as integrins, which play important roles during wound healing. Junctional epithelium (JE)
that links gingiva to tooth enamel mimics wound epithelium. The αvβ6 integrin is an epithelial
integrin that is not normally expressed by oral gingival epithelium but is induced during wound
healing, and has also been shown to be expressed in squamous cell carcinomas and in
leukoplakia specimens. Furthermore, αvβ6 integrin can bind and activate transforming growth
factor-β (TGF-β), suggesting a central role for the integrin in this unique epithelium. Using
human gingival specimens from extracted teeth and human gingival specimens attached to
decalcified teeth, the presence of JE was confirmed using simple epithelia tissue markers
(cytokeratin-19 and laminin-5). The presence of αvβ6 integrin in the JE and some of its putative
ligands (such as tenascin, fibronectin and TGF-β) was confirmed using immunohistochemical
labelling. Paraffin sections of wild-type (FVB), β6integrin-overexpressing (B6F1) and
P6integrin-knock-out mice (β6-/-) were stained with hematoxylin and eosin and observed under
the light microscope to analyze the morphology of the JE. Interestingly, the analysis of the β6-/-
sections showed striking alterations in the morphology and cellular organization of the JE. The
findings of this study suggest that αvβ6 integrin is constitutively expressed in the JE, in which it
could function as an immunoregulator via activation of TGF-β1.Dentistry, Faculty of
Graduate
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Al-Dahlawi, Salwa. "The role of αvβ6 integrin in epidermal wound healing." Thesis, 2004. http://hdl.handle.net/2429/15613.
Full textAbstract:
The αvβ6 integrin is an exclusively epithelial integrin that is highly expressed during
fetal development. In adult tissue αvβ6 integrin is re-expressed during inflammation,
carcinogenesis and in wound healing. The objective of this study was to investigate
whether compromised wound healing by hydrocortisone treatment is altered in mice
deficient or overexpressing the β6 subunit in comparison to wild type littermates.
Three groups of age and sex matched mice were used; wild type (WT), β6 integrin
deficient (β6√-), and human β6 integrin overexpressing transgenic mice (hβ6Fl). Each
group was subdivided into untreated (control) and treatment groups. Treatment
groups received a daily intraperitoneal injection of 1 mg of hydrocortisone for 13
days. Excisional 4 mm wounds were made on the dorsal surface of the mice and
wound healing was evaluated daily. Comparisons between different groups were
made from the histological analysis of three and ten days wound healing. Aged (22-
month-old) β6 integrin deficient (β6√-) animals showed a significant delay in wound
healing when compared to their aged matched wild type animals. The most significant
delay was observed at the stages where significant granulation tissue formation is
occurring (four to seven days post wounding). Hydrocortisone treatment significantly
delayed wound healing in wild type and β6 integrin deficient mice in comparison to
the untreated controls. However, hydrocortisone treatment in human P6 integrin
overexpressing (hβ6Fl) animals did not cause a significant delay in wound healing.
The results of this study indicated that the presence of αvβ6 integrin plays an
important role in wound healing in animals compromised by either age or stress.
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Ching-HaoTsai and 蔡景皓. "Design of Integrin αvβ6 or αIIbβ3-specific Antagonists using Disintegrin Scaffolds." Thesis, 2018. http://ndltd.ncl.edu.tw/handle/t5nny9.
Full textAbstract:
碩士國立成功大學
生物化學暨分子生物學研究所
106
Integrins are a family of heterodimeric receptors that are expressed on the surface of most cells, where they mediate cell-cell and cell-extracellular matrix interactions. Integrin αvβ6 is epithelial-specific and strongly induced during wound healing, inflammation and carcinogenesis. Integrin αIIbβ3 plays a critical role in platelet aggregation that is essential for hemostasis and thrombosis. It was reported that proteins with ARGDLXXL and KGD motifs can selectively bind to integrins αvβ6 and αIIbβ3, respectively. In this study, we propose to use rhodostomin (Rho) and trimucrin (Tmu), snake venom disintegrins, as scaffolds to develop integrins αvβ6 or αIIbβ3-specific antagonists. In the study on the design of integrin αvβ6-specific antagonists, we found that the 48ARGDRP and 48ARGDKP mutants exhibited high affinity with the IC50 values of 24.0 and 36.4 nM. In contrast, the 48ARGD(D/E)P and 48ARGDLP mutants exhibited low affinity with the IC50 values of 〉 5000 and 357.3 nM. These results suggest that the binding of integrin αvβ6 prefers the C-terminal residue adjacent to the RGD motif with positively charged residues but not with negatively charged and hydrophobic residues. In summary, we found that the relative affinity of 48ARGDXP mutants to integrin αvβ6 were 48ARGDRP 〉 48ARGDKP 〉 48ARGDWP 〉 48ARGDMP 〉 48ARGDAP 〉 48ARGDLP〉 48ARGDDP. We also incorporated the 48ARGDLAAL amino acid sequence of CagL in Helicobacter pylori, the integrin αvβ6-binding loop, into Rho, and it exhibited low affinity with the IC50 value of about 2000 nM. These results suggested that the corporation of the RGD motif into different scaffolds may exhibit different conformations. In addition, 52RP derived mutants exhibited better activity than mutants derived from 52LA, suggesting that binding of integrins αvβ6 prefers 52RP in Rho. From the docking results of 48ARGDRP binding to integrin αvβ6, we can see the difference interactions that LAP binding to integrins αvβ6 processes, which may provide us a new perspective for future design in selectivity. In the study on the design of integrin αIIbβ3-specific antagonist, we found that 41KKKRT-50AKGDRR, 41MKKGT-50AKGDRR, and 41IEEGT-50AKGDRP mutants had the IC50 values of 127.1, 115.6, and 511.2 nM. These results indicate that the binding of integrin αIIbβ3 prefers positively charged residues in the linker region of Tmu. The results of this study will serve as the basis for the design of integrin αvβ6- or αIIbβ3-specific drugs for the treatments of fibrosis and myocardial infarction.
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Turaga, Ravi C. "PROAGIO (A PROTEIN DESIGNED TO TARGET INTEGRIN αVβ3)." 2017. http://scholarworks.gsu.edu/biology_diss/189.
Full textAbstract:
Large efforts have been made to target integrin αVβ3 of endothelial cells. We have successfully developed a new class of protein (Ref to as ProAgio) by rational protein design using a stable host protein, domain 1 of cell adhesion protein CD2. ProAgio is designed to target integrin αVβ3 at a novel site and induces angiogenic endothelial cell apoptosis by recruiting and activating caspase 8 to the cytoplasmic domain of the targeted integrins. Tests with tumor xenograft models show that ProAgio strongly inhibits tumor growth. Histology analyses indicate that tumor vessels are reduced, while the established vasculatures are not affected. Toxicity analyses demonstrate that ProAgio is not toxic to mouse. Our study develops an effective anti-angiogenesis agent and provides a new platform for development of therapeutics by targeting integrins. We have successfully developed an anti-angiogenesis protein targeting integrin αVβ3 at a novel site by rational protein design. The developed agent is not toxic to non-cancerous blood vessels and other tissue/organs, providing an excellent candidate for future potential clinical development. Our developed protein is one of the very few examples that do not act through targeting VEGF/VEGFR or any other RTK pathways. The βA groove is present in almost all other β integrins. This approach may be applicable to develop agents targeting the similar βA groove of other integrin pairs, which can address wide array of pathological conditions such as AMD, Rheumatoid Arthritis, Osteoporosis etc.
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Lössner, Daniela [Verfasser]. "Tumorbiologische Rolle des Integrins αvβ3 [Alpha-v-Beta-3] beim humanen Ovarialkarzinom : differentielle Genexpression des Epidermal-growth-factor-Receptors (EGF-R) und der Integrin-linked-Kinase (ILK) als Funktion des Integrins αvβ3 ; Evaluierung der Bindung spezifischer Integrin-Antagonisten mittels Oberflächenplasmonenspektroskopie / Daniela Lössner." 2007. http://d-nb.info/988098326/34.
Full text
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Hsieh, I.-Shan, and 謝宜珊. "Role of αvβ3 integrin in the action of osteopontin in cancer cells." Thesis, 2015. http://ndltd.ncl.edu.tw/handle/c3z7nn.
Full textAbstract:
博士國立臺灣大學
藥理學研究所
103
Osteopontin, known as a secreted phosphoprotein, has a functional RGD domain and acts as a regulator of cytoskeleton dynamics and gene expression. Extracellular osteopontin functions through its interaction with cell surface receptors, including various integrins (αvβ1, αvβ3, αvβ5, α4β1, and α9β1) and CD44. Binding of osteopontin to these receptors can elicit a wild range of functions, such as cell adhesion, survival, and migration. Osteopontin is overexpressed in various cancers and has a crucial role in all stages of cancer. Elevated ostoepontin has been correlated with poor survival of patients with cancer in different tumor types. Accordingly, clinical studies have shown high osteopontin plasma concentration in patients with metastatic tumors compared with normal samples. Here we investigated the role of osteopontin in drug resistance, cancer cell survival and cancer therapy. It was found that osteopontin was upregulated in hypoxic human prostate cancer cells and osteosarcoma cells. Glucose transporters were also regulated in the hypoxia condition. Osteopontin upregulated drug transporter-p-glycoprotein expression in prostate cancer cells. P-glycoprotein is a subfamily of ATP-binding cassette transporter (ABC transporter). Cancer cells overexpressing ABC transporters actively pump out a variety of compounds from cells and decrease the therapeutic effects of chemotherapeutic agents. Using daunomycin, a chemotherapeutic agent with autofluorescence, to evaluate the ABC transporter pump activity, and osteopontin was found to increase the drug pumping-out activity. Long-term treatment with low-dose of daunomyain contributed to a drug resistance condition and further enhanced the overexpression of osteopontin. Released osteopontin inhibited daunomycin-induced cell death, which was antagonized by αvβ3 antibody. Knockdown of endogenous osteopontin potentiated the daunomycin-induced apoptosis. Furthermore, knockdown of osteopontin also enhanced the cell death caused by other chemotherapeutic drugs, including paclitaxel, doxorubicin, actinomycin-D and rapamycin, which are also the p-glycoprotein substrate. The animal studies showed that osteopontin knockdown enhanced the cytotoxic action of daunomycin. These results indicate that osteopontin is a potential therapeutic target for cancer therapy to reduce the drug resistance in sensitive tumors. On the other hand, endogenously released osteopontin regulated the expression of glucose transporter 1 and glucose transporter 3 in osteosarcoma and enhanced glucose uptake into cells via the αvβ3 integrin. Knockdown of osteopontin induced cell death in 20% of osteosarcoma cells. Phloretin, a glucose transporter inhibitor, also caused cell death by treatment alone. The phloretin-induced cell death was significantly enhanced in osteopontin knockdown osteosarcoma cells. Combination of a low dose of phloretin and chemotherapeutic drugs, such as daunomycin, 5-Fu, etoposide, and methotrexate, exhibited synergistic cytotoxic effects in three osteosarcoma cell lines. Inhibition of glucose transporters markedly potentiated the apoptotic sensitivity of chemotherapeutic drugs in osteosarcoma. These results indicate that the combination of a low dose of a glucose transporter inhibitor with cytotoxic drugs may be beneficial for treating osteosarcoma patients. According to the results shown above studies, we know that osteopontin plays an important role in drug resistance and influences cancer cells survival during cancer therapy via integrin αvβ3. We thus investigated the effect of integrin antagonist, Rhodostomin mutant-RGD-related proteins, HSA (C34S)-ARLDDL, PEG-ARLDDL, ARLDDL and KKKRT-ARGDNP, in cancer therapy. During treatment of these four RGD-related proteins, they can inhibit prostate cancer, melanoma, osteosarcoma tumor growth, effectively. The inhibition of tumor growth effect is more significant when combination RGD-protein combined with chemotherapy drug of daunomycin. The RGD-proteins also can inhibit cancer cell adhesion and angiogenesis. αvβ3 integrin is highly expressed in some tumor cells and involved in tumor progression. Osteopontin and RGD-related proteins influence tumor growth, cell adhesion, survival via αvβ3 integrins. αvβ3 integrin can be a good target for cancer therapy. Our studies demonstrate that inhibition of αvβ3 integrin with RGD-related proteins or in combination with chemotherapy improve the anticancer efficacy. This may be a new strategy for cancer therapy.
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chen, Tsai-kun, and 陳蔡昆. "The Role of the RXDDL Motif of Rhodostomin in Recognizing Integrin αVβ3." Thesis, 2007. http://ndltd.ncl.edu.tw/handle/98544917370609495610.
Full textAbstract:
碩士國立成功大學
生物化學研究所
95
Integrins are heterodimeric cell surface receptors required for cell trafficking and for adhesion to other cell types and to constituents of the extracellular matrix. The ligands of integrins utilize RGD, ILDV, or short sequences as a key structural component for their integrin-binding site. Disintegrins are a family of RGD-containing and cysteine-rich proteins isolated from snake venoms. Many reports show that the RGD motif and the amino acid residues flanking the RGD sequence of disintgrins play an important role in recognizing integrins. They mainly interact with the β1 and β3 families of integrins. In this study we used rhodostomin (Rho) as the scaffold to study the roles of the RGD motif and the amino acid residues flanking the RGD sequence of disintegrins in recognizing disintegrins. Rho is a disintegrin that contains 68 amino acids including 6 disulfide bonds and a PRGDMP sequence at the positions of 48-53. Our previous study showed that Rho containing either the residue flanking the R and D residues or the DL residues C-terminally adjacent to RGD motif had high potency and specificity to integrin αVβ3. Therefore, we incorporated the 48PRXDDL53 motif into Rho to study the effect of the residue flanking the R and D residues in recognizing integrins. In addition to PRGDDL, PRLDDL, and PRIDDL mutants, I have expressed seventeen mutants PRHDDL, PRYDDL, PRADDL, PRCDDL, PRDDDL, PREDDL, PRFDDL, PRKDDL, PRMDDL, PRNDDL, PRPDDL, PRQDDL, PRRDDL, PRSDDL, PRTDDL, PRVDDL and PRWDDL in Pichia pastoris and purified them to homogeneity. The mutant proteins produced in P. pastoris were 4.3-16.5mg/L. The experimental molecular weights of the mutants deviate <1 Da when compared with calculated values, indicating the formation of six disulfide bond in these mutants. The analysis of platelet aggregation and cell adhesion assays showed that Rho mutants-containing the PRXDDL motif has low potency to integrin α5β1 and αIIbβ3. The mutants PRMDDL, PRPDDL, PRWDDL, and PRLDDL can selectively inhibit integrin αvβ3 with the IC50 value of 175, 188, 387, and 342 nM, respectively. This study may serve as the basis for exploring the structure and function relationships of integrins and disintegrins and for designing integrin αvβ3-specific disintegrins.
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Τσιουπινάκη, Κωνσταντία. "Molecular imaging of spatio-temporal distribution of angiogenesis in hindlimb ischemia model and diabetic milieu." Thesis, 2013. http://hdl.handle.net/10889/7944.
Full textAbstract:
In the current thesis, spatio-temporal evaluation of the endogenous angiogenic response to ischemia was perfomed. After vascular occlusion, ischemic angiogenesis is an important reparative mechanism and can ameliorate the outcome of ischemic disease. Diabetic foot ulcers affect almost 15% of diabetic patients and are the leading cause of amputations worldwide (Yoon et al., 2005). Diminished blood flow because of atherosclerotic occlusive disease of the peripheral arteries of diabetic patients, in conjunction with anatomic and functional microcirculatory impairments contribute to development of trophic ulcerations, infections and gangrene of the lower extremities, frequently requiring amputation of the leg (Sasso et al., 2005). Numerous studies have confirmed the impaired post-ischemic angiogenesis in diabetes (Yoon et al., 2005; Sasso et al., 2005). Consequently, wound healing patterns are disturbed in diabetes mainly due to decreased ischemia-driven angiogenesis (Yoon et al., 2005). Integrin ανβ3 is a promising imaging target of angiogenic activity which is up-regulated on activated endothelial cells (ECs) but not on quiescent ones. Molecular imaging (MI) of ανβ3 integrin expression with the aid of a dedicated high resolution gamma camera, is a very sensitive imaging approach for the evaluation of angiogenesis in the rabbit hindlimb ischemia model. Furthermore, diabetes mellitus (DM) was induced, to study the effects of this pathology on the spatio-temporal distribution of angiogenesis. In order to evaluate the whole spectrum of endogenous process of collateralization after occlusion of an artery, Digital Subtraction Angiography (DSA) was also used for the visualization of larger collaterals.
During the first part of the study DM experimental protocol was investigated in order to find the appropriate protocol for the induction of long-term diabetic animal model, as it is a methodology that has not yet been standardized. At the same time a cohort of animals underwent endovascular embolization for the establishment of hindlimb ischemia and were imaged with the aid of a MI radiotracer technique in order to deal with unresolved issues and establish the imaging protocol. The study included seven New Zealand White (NZW) rabbits that underwent unilateral percutaneous endovascular embolization of the femoral artery, for the establishment of hindlimb ischemia that triggers the endogenous process of collateralization. The contralateral limb was not embolized and served as a control. The employed radiotracer for angiogenesis imaging, was a 99mTc labeled cyclic RGD peptide [c RGDfk-His]-99mTc that binds specifically to ανβ3 integrin via the three amino acid sequence Arginine-Glycine-Aspartic acid or RGD. Image acquisition was performed with a high resolution gamma camera and all animals underwent molecular imaging on the 3rd and the 9th day post-embolization. In all animals DSA was performed on the 9th day post-embolization.
The acquired images demonstrated that retention of the radiotracer at the ischemic tissue is remarkably increased compared to the non-ischemic hindlimb (normal limb) (16020 ± 2309 vs. 13139 ± 2493 on day 3; p=0.0014 and 21616 ± 2528 vs. 13362 ± 2529 on day 9; p<0.0001, respectively). In addition, radiotracer retention in normal limbs seems to be increased at day 9 in normal limbs compared to day 3 (p=0.0112). DSA at day 9, demonstrated that the mean vessel length detected was significantly superior in the normal compared to the ischemic limb (mean value 3680 ± 369.8 vs.2772 ± 267.7; p< 0.0001, respectively).
Angiogenesis was successfully detected using a 99mTc labeled cyclic RGD peptide MI technique and was significantly more pronounced in the ischemic compared to normal limbs, both at day 3 and day 9 after embolization. The peak of the phenomenon was detected at day 9. Increased mean vessel length in the normal compared to the ischemic limb demonstrates that although angiogenesis is pronounced in day 9, arteriogenesis is not sufficiently pronounced and that the phenomenon of arteriogenesis has just initiated.
The study of the angiogenic response to ischemia, has not yet been completed as MI of diabetic animals with hindlimb ischemia is underway and not completed due to many difficulties and delay in different phases of the experiment. With the conclusion of the MI of diabetic animals with hindlimb ischemia, the study will be completed and we expect to demonstrate the effect of DM on the spatio-temporal pattern of angiogenesis, providing a valuable tool in clinical practice for the precise and early diagnosis and therapy assessment of the ‘diabetic foot’.Σκοπός της παρούσας μελέτης ήταν η χωρο-χρονική εκτίμηση της ενδογενούς αγγειογενετικής διαδικασίας ως απόκριση στο ερέθισμα της προκλητής ισχαιμίας. Μετά από απόφραξη αρτηρίας, η επαγόμενη αγγειογένεση είναι ένα σημαντικός μηχανισμός αποκατάστασης που μπορεί να περιορίσει το αποτέλεσμα της ισχαιμίας. Το έλκος του ‘διαβητικού ποδιού’ εμφανίζεται στο 15% περίπου των ασθενών που πάσχουν από διαβήτη και αποτελεί την κύρια αιτία ακρωτηριασμού του κάτω άκρου, μετά τα ατυχήματα (Yoon et al., 2005). Η μειωμένη αιματική ροή λόγω της αποφρακτικής αρτηριοπάθειας που οφείλεται στην αθηροσκληρωτική προσβολή των περιφερικών αρτηριών των διαβητικών, σε συνδυασμό με ανατομική και λειτουργική φθορά του αγγειακού δικτύου της μικροκυκλοφορίας, οδηγούν στην ανάπτυξη ελκών, μολύνσεων και γάγγραινας των κάτω άκρων που πολύ συχνά οδηγεί στον ακρωτηριασμό του ποδιού (Sasso et al., 2005). Πολυάριθμες μελέτες όμως έχουν δείξει ότι η αγγειογένεση που επάγεται από κρίσιμη ισχαιμία, στην παθολογία του διαβήτη δεν είναι φυσιολογική (Yoon et al., 2005; Sasso et al., 2005). Συνεπώς στον διαβήτη και ο μηχανισμός επούλωσης πληγών δεν συμβαίνει φυσιολογικά κυρίως λόγω ελαττωματικής ενδογενούς αγγειογένεσης ως απόκριση στην ισχαιμία (Yoon et al., 2005). Το μόριο της ιντεγκρίνης ανβ3 υπερεκφράζεται στα ενεργοποιημένα ενδοθηλιακά κύτταρα που συμμετέχουν στην αγγειογενετική διαδικασία αλλά όχι στο ανενεργό ή αλλιώς «σιωπηλό» ενδοθήλιο. Συνεπώς αποτελεί έναν πολύ καλό μοριακό στόχο για απεικόνιση και δείκτη της αγγειογενετικής δραστηριότητας. Η Μοριακή Απεικόνιση (ΜΑ) του επιπέδου έκφρασης του μορίου ιντεγκρίνης ανβ3 με τη χρήση ραδιοϊσοτοπικών τεχνικών έχει το πλεονέκτημα υψηλής ευαισθησίας ανίχνευσης πολύ χαμηλών συγκεντρώσεων του ραδιοϊχνηθέτη σε σχέση με τις συμβατικές τεχνικές (x-ray computed tomography (CT) angiography, contrast-enhanced ultrasound και high-resolution magnetic resonance angiography). Ο σκοπός της μελέτης μας ήταν να ερευνήσουμε τις δυνατότητες της ΜΑ με την βοήθεια γ-κάμερας υψηλής διακριτικής ικανότητας και πειραματικού μοντέλου κονίκλου με ίσχαιμο οπίσθιο άκρο. Επιπλέον ένα σημαντικό μέρος της μελέτης αφορούσε την εφαρμογή του πρωτοκόλλου για τη πρόκληση διαβήτη ώστε να γίνει μελέτη της επίδρασης της συγκεκριμένης παθολογίας στην χωρο-χρονική κατανομή της αγγειογένεσης. Προκειμένου να αποδοθεί μία συνολική εκτίμηση του ενδογενούς μηχανισμού αποκατάστασης του αγγειακού δικτύου, εφαρμόστηκε Ψηφιακή Αφαιρετική Αγγειογραφία για την εκτίμηση της δημιουργίας παράπλευρου δικτύου. Στη πρώτη φάση της μελέτης, έγινε διερεύνηση του πρωτοκόλλου πρόκλησης διαβήτη δεδομένου ότι η μεθοδολογία παρουσιάζει έλλειψη προτυποποίησης. Παράλληλα εφαρμόστηκε το πρωτόκολλο ισχαιμίας σε μια ομάδα λευκών κονίκλων Νέας Ζηλανδίας για την διερεύνηση του πρωτοκόλλου ΜΑ. Στην μελέτη, χρησιμοποιήθηκαν συνολικά επτά κόνικλοι Νέας Ζηλανδίας οι οποίοι υπεβλήθησαν σε εμβολισμό της μηριαίας αρτηρίας ενός από τα δύο οπίσθια άκρα για την πρόκληση οξείας ισχαιμίας. Στους κόνικλους έγινε ενδοφλέβια έγχυση του κυκλικού επισημασμένου πεπτιδίου [c RGDfk-His]-99mTc που περιέχει την αλληλουχία τριών αμινοξέων Αργινίνης-Γλυκίνης-Ασπαρτικού οξέος (Arginine-Glycine-Aspartic acid or RGD), μέσω της οποίας δεσμεύεται το πεπτίδιο στο μόριο της ιντεγκρίνης ανβ3. Η απεικόνιση του επιπέδου έκφρασης του μορίου ιντεγκρίνης ανβ3 πραγματοποιήθηκε με γ-κάμερα υψηλής διακριτικής ικανότητας την 3η και την 9η ημέρα μετά την απόφραξη της μηριαίας αρτηρίας. Ψηφιακή Αφαιρετική Αγγειογραφία πραγματοποιήθηκε την 9η ημέρα μετά την απόφραξη της μηριαίας αρτηρίας. Τα δεδομένα από την ποσοτικοποίηση των απεικονιστικών δεδομένων, έδειξαν ότι υπάρχει αυξημένη πρόσληψη του ραδιοϊχνηθέτη στη περιοχή ισχαιμίας σε σχέση με τη περιοχή φυσιολογικής αιμάτωσης του ετερόπλευρου άκρου (16020 ± 2309 έναντι 13139 ± 2493 την 3η ημέρα, p=0.0014 και 21616 ± 2528 έναντι 13362 ± 2529 την 9η ημέρα, p<0.0001, αντίστοιχα). Επιπλέον η πρόσληψη του ραδιοϊχνηθέτη στα φυσιολογικά άκρα φαίνεται να είναι αυξημένη την 9η ημέρα σε σχέση με την 3η ημέρα (p=0.0112), γεγονός που μπορεί να αποδοθεί στην σταδιακή συγκέντρωση ενεργοποιημένου ενδοθηλίου και στους φυσιολογικούς ιστούς. Η Ψηφιακή Αφαιρετική Αγγειογραφία έδειξε ότι την 9η ημέρα που έγινε η λήψη δεδομένων, το μέσο μήκος αγγείων στα φυσιολογικά άκρα ήταν αρκετά μεγαλύτερο σε σχέση με τα ίσχαιμα άκρα (μέση τιμή 3680 ± 369.8 έναντι 2772 ± 267.7, p< 0.0001, αντίστοιχα). Η ραδιοϊσοτοπική τεχνική που εφαρμόστηκε για την απεικόνιση του επιπέδου έκφρασης του μορίου ιντεγκρίνης ανβ3 στη παρούσα μελέτη, έδειξε ότι υπάρχει αυξημένη πρόσληψη του ραδιοϊχνηθέτη στη περιοχή ισχαιμίας σε σχέση με τη περιοχή φυσιολογικής αιμάτωσης του ετερόπλευρου άκρου, την 3η ημέρα και την 9η ημέρα μετά την απόφραξη της μηριαίας αρτηρίας. Τα πειραματικά δεδομένα δείχνουν επίσης ότι το φαινόμενο της αγγειογένεσης κορυφώνεται την 9η ημέρα μετά την πρόκληση ισχαιμίας. Επιπλέον τα δεδομένα από τη Ψηφιακή Αφαιρετική Αγγειογραφία την 9η ημέρα, δείχνουν ότι ο ενδογενής μηχανισμός σχηματισμού παράπλευρου δικτύου αν και έχει πυροδοτηθεί δεν έχουν σχηματιστεί ακόμα μεγαλύτερα αγγεία και γι αυτό το λόγο η αρτηριογένεση υπολείπεται της αγγειογένεσης σε αυτή τη φάση. Για την ολοκλήρωση της μελέτης, εκκρεμεί η απεικόνιση των διαβητικών κονίκλων με ίσχαιμο οπίσθιο άκρο η οποία καθυστέρησε λόγω επιμέρους δυσκολιών στη διαδικασία των πειραμάτων. Η ΜΑ δεικτών της αγγειογένεσης σε άκρο που πάσχει από ισχαιμία παρουσία διαβήτη, δύναται να έχει τεράστια εφαρμογή στην κλινική πράξη για την ιατρική παρακολούθηση του ‘διαβητικού ποδιού’.
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Kym, Eugene Yongshik (Gene). "Engineered Discoidin Domain from Factor VIII Binds αvβ3 Integrin with Antibody-like Affinity." Thesis, 2014. https://thesis.library.caltech.edu/8449/1/Gene_Kym_Thesis.pdf.
Full textAbstract:
Alternative scaffolds are non-antibody proteins that can be engineered to bind new targets. They have found useful niches in the therapeutic space due to their smaller size and the ease with which they can be engineered to be bispecific. We sought a new scaffold that could be used for therapeutic ends and chose the C2 discoidin domain of factor VIII, which is well studied and of human origin. Using yeast surface display, we engineered the C2 domain to bind to αvβ3 integrin with a 16 nM affinity while retaining its thermal stability and monomeric nature. We obtained a crystal structure of the engineered domain at 2.1 Å resolution. We have christened this discoidin domain alternative scaffold the “discobody.”