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Dissertations / Theses on the topic 'Desmosine'
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1
Hopkins, Gemma V. "The mechanism of desmosome internalisation." Thesis, University of Manchester, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.493936.
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Desmosomes are intercellular adhesive junctions providing architectural integrity to tissues that are subject to mechanical stress. During wound healing downregulation of desmosomal adhesion is necessary to facilitate tissue remodeling. Ultrastructural observations suggest that downregulation may occur by internalisation of whole desmosomes but the mechanism of downregulation is not understood. Protein kinase C alpha (PKCα) has been shown to colocalise with desmosomes at the wound edge and to modulate their adhesion. This thesis investigates the internalisation of desmosomes with particular reference to the roles of the cytoskeleton and PKC.
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Tan, Celia I. C. "A radiological and biochemical perspective on ageing and degeneration of the human thoracic intervertebral disc." University of Western Australia. School of Surgery and Pathology, 2004. http://theses.library.uwa.edu.au/adt-WU2004.0059.
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Disc degenerative changes are directly or indirectly associated with spinal pain and disability. Literature revealed a high prevalence of disc degeneration in the thoracic region, however thoracic MRI degeneration trends and information on disc biochemical matrix constituents are limited for thoracic discs compared to lumbar and cervical discs. The objective of this thesis was to use MRI to investigate the prevalence of disc degenerative changes affecting the human thoracic spine, and to determine the factors affecting spinal disc biochemical matrix. A 3-point subjective MRI grading scale was used to grade the films. The feasibility of using archived formalin-fixed cadaver material was investigated to analyse collagen and elastin crosslinks. The prevalence of degenerative changes in human thoracic discs and vertebrae (T1 to T12) was determined retrospectively from an audit of 216 MRI cases, using sagittal T1- and T2-weighted MR images. In a subsequent series of ex-vivo studies, human thoracic discs and LF from 26 formalin-fixed and two fresh spines, involving all thoracic levels, were examined macroscopically to determine the degeneration status. Subsequently, disc and ligament tissues were analysed biochemically for collagen (pyridinoline and deoxypyridinoline) and elastin (desmosine and isodesmosine) crosslinks. These crosslinks were extracted from hydrolysed samples by cellulose partition chromatography, and analysed by reverse-phase HPLC. Collagen content was determined using its hydroxyproline content, and proteoglycan content was assayed using a modified DMB assay for chondroitin sulphate. Finally the MRI and macroscopic assessments of thoracic discs, were compared with the biochemical data from two fresh cadaver thoracic spines. The 3-point MRI grading scale had a high inter- (k = 0.57 to 0.78) and intra-rater (k = 0.71 to 0.87) reliability. There were no significant differences in the collagen and elastin content and extent of collagen crosslinks between formalin fixed and unfixed ligament and disc tissues, after 25 weeks of formalin fixation. From the in-vivo MRI series of investigations (n = 216 MRI films), the prevalence of thoracic disc degenerative and vertebral morphological changes revealed significant age, gender and spinal level trends (p < 0.05).Generally, males had a higher propensity for disc degeneration in contrast to females, especially older females, where the trend showed a higher prevalence of osteophytes and vertebral body changes. In particular, the mid and lower thoracic levels have a higher prevalence of degenerative changes, except for osteophytes and anterior vertebral wedging. With increased age, there was a concomitant increase in anterior wedging and bi-concavity and disc degenerative changes except for end-plates. The biochemical investigations on the ex-vivo series of formalin-fixed thoracic discs (n = 303) also revealed significant changes in the disc matrix due to degeneration status, age, gender and spinal regional factors. With increased age, normal disc matrices have significantly lower collagen content and extent of pyridinoline (p < 0.001). In contrast, the degenerated disc matrix revealed significantly higher collagen content and extent of deoxypyridinoline (p < 0.05). These findings suggest that an altered matrix existed in normal ageing discs, which render the disc prone to injury and degeneration over the life span. The higher collagen and deoxypyridinoline in degenerated disc matrices reflects an increase in chondrocyte synthesis, and is also a novel finding, suggesting that they may be used as markers of ageing and degeneration processes. The biochemical investigations on another series of ex-vivo spinal LF tissues (n = 364), revealed that this had a lower collagen and pyridinoline, but significantly higher elastin and deoxypyridinoline compared to spinal discs (p < 0.05). Elastin crosslinks however were difficult to detect in spinal discs, being present in negligible amounts in a few lumbar discs. The elastin crosslinks in the LF were not significantly affected by age, but were significantly higher in calcified, and female ligamentum tissues, and also in the lumbar region (p < 0.05). These MRI prevalence findings enhanced our knowledge of vertebral body and disc degeneration trends in the thoracic region and contributed to the interpretation of MR images for pathology in the human thoracic spine. Information on the associated collagenous and elastic changes in the disc and ligamentum matrices provide original data and insight on the pathogenesis of degeneration in the disc matrix from a biochemical perspective, highlighting gender, age and spinal level influences on the matrix tensile strength and cellular synthetic activities.
3
Winfield, Kaye R. "Extraction of desmosines from urine : an indicator for inflammatory lung damage." University of Western Australia. School of Paediatrics and Child Health, 2007. http://theses.library.uwa.edu.au/adt-WU2007.0059.
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[Truncated abstract] Urinary desmosines have been proposed as a biomarker for inflammatory lung damage. Desmosine, a breakdown product of elastin, is an effective marker of the degradation of elastin and has been studied in many disease scenarios where there is acute and chronic lung inflammation. Lung matrix degradation has been proven in vitro and in vivo with many experiments showing that the excess proteases degrades lung matrix. The secretion of proteases by neutrophils is an innate response of the body to the invasion by micro organisms and when secreted in excess, the protective anti-protease mechanism is swamped. Chronic inflammation and persistent infection eventually leads to bronchiectasis and respiratory failure. Urinary desmosine has been shown to be elevated in respiratory conditions with acute and chronic inflammation . . . Urinary desmosine levels in a large cohort of healthy children have been established using this method and predictive Z-score formulae have been developed to use in children with lung disease. Exploration of these scores in children with CF have shown that the levels of urinary desmosine appear to be sensitive to the clinical setting, where high urinary desmosine levels were present during exacerbation and significantly reduced when treated for infection with antibiotic therapy and physiotherapy. The study of young children under the age of seven was undertaken to determine if the urinary desmosine levels could indicate when lung damage was occurring and to determine what mechanisms might be involved. Since there appeared to be no apparent relationship between elevated desmosines and proteases in the lung in young children with CF, further studies are required to define the mechanisms behind increased elastin metabolism in those children.
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Winfield, Kaye R. "Extraction of desmosines from urine : an indicator for inflammatory lung damage /." Connect to this title, 2006. http://theses.library.uwa.edu.au/adt-WU2007.0059.
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Lloyd, Susan. "Calcium-independent desmosome adhesion : aquisition, maintenance and modulation." Thesis, University of Southampton, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.295856.
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Measures, H. R. "A study of desmosome formation in kidney epithelial cells." Thesis, University of Southampton, 1988. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.234435.
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Cabral, Rita Meira e. Cruz de Vasconcelos. "The role of desmoplakin and plakoglobin in desmosome associated disease." Thesis, Queen Mary, University of London, 2010. http://qmro.qmul.ac.uk/xmlui/handle/123456789/2413.
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Desmosomes are intercellular junctions that connect intermediate filaments (IFs) of adjacent cells within tissues, generating a large and mechanically resilient network. Desmosomes are prominent in epithelia and the myocardium. Their importance for maintenance of the strength and flexibility of these tissues is underlined by mutations in desmosomal genes which compromise skin or heart and in some instances both. This thesis is focused on two obligate plaque proteins of the desmosome, desmoplakin (DP) and plakoglobin (PG), and their role in disease of skin and heart. A novel homozygous nonsense mutation, p.S24X, in the gene encoding PG, JUP, was identified in three non-consanguineous Argentinean patients, resulting in skin fragility, palmoplantar keratoderma (PPK) and woolly hair with no symptoms of cardiomyopathy. p.S24X mRNA was readily detected and an alternative AUG codon translates an N-terminally truncated PG, which is expressed in patient and parental skin samples at barely detectable levels. At the same time another novel homozygous mutation, c.468G>A, was identified in JUP causing a very similar phenotype in two Kuwaiti siblings and resulting also in very low levels of PG in the skin. These results suggest that PG is essential for maintenance of skin integrity, but not for normal heart development in children. The gene encoding DP, DSP, produces two alternative splice variants, DPI and DPII. A novel DP alternative splice isoform, DPIb, is described. As shown by RT-PCR, DSPIb is expressed in several epithelial and cardiovascular tissues and is the only DSP isoform detected in the aorta. Western blotting proteins from HaCaT and K1 cells identified this novel isoform, which has been previously missed due to its similarity in size to DPII. DPIb may help compensate defects in patients with DSPI-specific mutations. The molecular mechanisms of three different DSP mutations leading to skin disease or skin disease associated with cardiomyopathy were investigated. Mutations causing DPI/DPII haploinsufficiency and DPI knock-out lead to decreased expression levels of the desmosomal proteins DSC2, DSC3 and PKP1 in an experimental system using HaCaT keratinocytes. Expression of a mutant DSPI construct harbouring a dominant 30 bp insertion results in the formation of large DPI aggregates at the cell membrane of HaCaT cells. The spectrum of DSP and JUP mutations which cause different clinical phenotypes is discussed.
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ZORILA, FLORESCU SILVANA MARIA. "Alteration du profil d'expression des composants desmosomaux et des jonctions adherentes dans les carcinomes epidermoides de l'oropharynx et leur valeur predictive : etude in vivo et in vitro." Paris 7, 2001. http://www.theses.fr/2001PA07GA02.
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DEPONDT, JOEL. "Evaluation prospective des cytokeratines et des composants desmosomaux comme marqueurs diagnostiques et pronostiques des carcinomees epidermoides de l'oropharynx." Paris 7, 2000. http://www.theses.fr/2000PA07GA01.
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Asimaki, Angeliki. "Arrhythmogenic right ventricular cardiomyopathy, a disease of the desmosome : genetic and functional studies." Thesis, University College London (University of London), 2008. http://discovery.ucl.ac.uk/1443947/.
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Mutation analysis of the recognized ARVC genes and of further candidate genes was performed on a large cohort of ARVC patients. Several novel mutations were identified and three further desmosomal genes were linked to the disease: plakophilin2, desmocollin2 and desmoglein2. Heart and skin samples from ARVC patients were subjected to microscopic examination and immunohistochemistry to study the effect of the newly-identified mutations on the structure of cell adhesion complexes.;The functional effects of a particular novel mutation were thoroughly examined in vitro. S39_K40insS is the first dominant ARVC-causing plakoglobin mutation to be reported. Yeast-two hybrid analysis was used to investigate the effect of S39_K40insS on the proteins interactions established by plakoglobin. A HEK293 cell line stably expressing the mutant protein was generated and used to study the effects of S39_K40insS on desmosomal structure, cell proliferation, cell death, subcellular localization and expression levels of proteins involved in adhesion and signalling and cellular responses to defined mechanical load. A recombinant adenovirus expressing the mutant protein was generated and used to transfect neonatal rat ventricular cardiomyocytes, whose behaviour and responses were subsequently analysed. The functional consequences of S39_K40insS were compared with those of PK215del2, a previously reported recessive plakoglobin mutation known to underlie Naxos disease, a syndromic form of ARVC.;These results point towards novel mechanisms of disease pathogenesis, that apart from weakened cell-cell adhesion involve altered protein turnover kinetics and defects in signalling pathways. Similar studies should improve our understanding of ARVC and provide a more accurate diagnostic algorithm.
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Parker, Andrew E. "A molecular analysis of desmosomal glycoproteins II and III : constituents of the desmosome." Thesis, Open University, 1991. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.303573.
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Bharathan, Navaneetha Krishnan. "USING THE FROG EPIDERMIS TO UNCOVER DESMOSOME FUNCTION AND REGULATION IN THE DEVELOPING EMBRYO." VCU Scholars Compass, 2018. https://scholarscompass.vcu.edu/etd/5313.
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The desmosome is one of the major cell adhesion junctions found in the epithelia, heart, and hair follicle. Described as a “rivet” that hold cells together, it provides these tissues with the integrity to withstand the tremendous forces they face in everyday life. Defects in this junction can lead to devastating diseases where patients are susceptible to skin infections and cardiovascular defects. Limited treatments exist for diseases of the desmosome, and strategies do not target all symptoms. Therefore, delineating the function and regulation of desmosomes is of paramount importance for the development of prevention and treatment strategies. The Xenopus laevis has been utilized for the study of embryonic development and tissue movements. This study takes advantage of the frog model to study a key desmosomal protein, desmoplakin (Dsp), in the epidermal development of the embryo. First, Xenopus embryonic epidermis has junctional desmosomes as early as the blastula stages. Desmosomes numbers per junction increase as the embryo develops. Dsp is present in many epidermally-derived structures in the embryo at varying levels. Xenopus embryos deficient in desmoplakin have phenotypic defects in epidermal structures and the heart, mimicking mammalian models. Embryos with reduced Dsp exhibit an increased susceptibility to epidermal damage under applied mechanical forces. Assays also reveal a potential role for desmosomes in radial intercalation, a process through which cells move from the inner to the outer epidermal layers. Embryos with reduced Dsp exhibit a slight reduction in intercalation and defects in intercalating cell types, including multiciliated cells and small secretory cells. Finally, c-Jun N-terminal kinase (JNK) may have a potential role in the regulation of desmosome assembly and adhesion. Embryos with deficient Dsp display a partial recovery of mechanical integrity when treated with a JNK inhibitor.
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Völlner, Frauke [Verfasser], Anna [Gutachter] Starzinski-Powitz, and Ritva [Gutachter] Tikkanen. "Flotillins as novel regulators of desmosome dynamics / Frauke Völlner ; Gutachter: Anna Starzinski-Powitz, Ritva Tikkanen." Frankfurt am Main : Universitätsbibliothek Johann Christian Senckenberg, 2017. http://d-nb.info/1138276944/34.
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Cooper, Nichola. "The role of Protein Kinase Cα in the skin and cutaneous wound healing." Thesis, University of Manchester, 2014. https://www.research.manchester.ac.uk/portal/en/theses/the-role-of-protein-kinase-c-alpha-in-the-skin-and-cutaneous-wound-healing(b0dc19c1-6bd6-4662-adca-010c9f19d785).html.
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Chronic wounds represent a severe socio-economic burden and a key area of unmet clinical need. PKCα is ubiquitous in the skin, particularly the epidermis and functions in numerous pathways that are fundamental to wound repair. By utilising a global PKCα-/- mouse we have identified PKCα-regulated processes both in unwounded skin and during wound healing. PKCα-/- mice display considerably delayed wound healing with a dramatic reduction in re-epithelialisation. By analysing the ultrastructure of the epidermis, I have shown that this delay directly correlates with a failure of wound edge desmosomes to switch to a their adhesive properties. A major risk factor for the development of chronic wounds is age. Crucially, this delay in modulating cell adhesion is conserved in human chronic wounds and aged murine skin. Furthermore, manipulation of PKCα using an inducible bitransgenic mouse containing epidermal specific constitutively active PKCα can accelerate the modulation of desmosome adhesion and subsequently improve re-epithelialisation. Global gene expression analysis of PKCα-/- skin and wounds revealed further defects. Upon wounding, we observed a failure to correctly regulate expression of key collagen and Wnt signalling genes that are essential for correct and timely wound healing. Finally, intrinsic gene expression changes were identified in the skin of PKCα-/- mice, specifically a downregulation of multiple extracellular matrix genes. Of note was the downregulation of small leucine-rich proteoglycans which led to alterations to dermal collagen structure and skin tensile strength. These changes render the PKCα-/- skin susceptible to breaking and wound development. To conclude, we have identified multiple roles for PKCα intrinsically in the skin and also during cutaneous wound healing. Importantly, these intrinsic changes appear to predispose PKCα-/- skin to the development of cutaneous wounds and altered wound-specific processes that manifest in a delayed healing phenotype.
15
AMAR, LAURENCE. "Role de la proteine kinase c dans la regulation des desmosomes au niveau des cellules epitheliales. Etude in vitro sur les keratinocytes de rat foetal et la lignee hela, derivant d'un adenocarcinome cervical humain." Paris 7, 1999. http://www.theses.fr/1999PA07GA03.
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Cozzani, Emanuele. "Immunopathologie des pemphigus auto-immuns : caractérisation immunochimiques des antigènes desmosomiaux et implication dans le diagnostic sérologique." Lyon 1, 1997. http://www.theses.fr/1997LYO1T329.
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Andersen, Nicholas John. "Characterization of mammalian exocyst subunit Sec3." Diss., University of Iowa, 2009. https://ir.uiowa.edu/etd/327.
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The Exocyst is a hetero-octameric complex involved in tethering of post-Golgi vesicle transport to sites of membrane expansion. In budding yeast, the Exocyst targets vesicles to bud site resulting in bud emergence and abscission of the daughter cell. Mammalian Exocyst is recruited to developing lateral membranes after cadherin mediated adhesion and then is segregated to adherens junctional complexes (AJC). In polarized epithelia, the Exocyst is required for basal-lateral transport of LDL receptor. Additional Exocyst subunit localizations and functions have been identified. It is not known whether these supplementary roles can be attributed to the Exocyst or other unidentified Exocyst subcomplexes. Sec3, an Exocyst subunit, is hypothesized to be a landmark of polarization in yeast. In polarized epithelia, GFP tagged Sec3 remained cytosolic in polarized epithelia unlike Sec6/8. Sec3-GFP was recruited to lateral membranes only after dual over expression of heterologous GLYT1. Little is known about endogenous mammalian Sec3. Our work suggests Sec3 defines an Exocyst subcomplex that is required for desmosome integrity. Sec3 and additional subunits (Sec6, Sec8, Sec15, Exo70, and Exo84) were present at desmosomes, but Sec3 failed to localize to AJC. Only antibodies to Sec6 and Sec8 labeled AJC. Reduction of Sec3 protein expression resulted in the impairment of desmosome morphology and function with no detrimental effect on adherens junctions. These data suggest the existence of functionally different Exocyst subcomplexes. Sec3-exocyst recruited minus-end directed microtubule motor KifC3 to desmosomes. KifC3 was previously shown to be recruited with a microtubule anchoring complex to basal-lateral membrane. This suggests Sec3 may recruit KifC3 to organize microtubules at desmosomes. This would establish a pathway to efficiently transport newly synthesized basal-lateral cargo. These results suggest a novel mechanism of the Exocyst to regulate post-Golgi vesicular transport and intercellular adhesion.
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GRIBBINS, KEVIN MICHAEL. "THE CYTOLOGY OF SPERMATOGENESIS AND ULTRASTRUCTURE OF THE SEMINIFEROUS EPITHELIUM IN REPTILES." University of Cincinnati / OhioLINK, 2003. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1053715753.
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Mouquet, Hugo. "LE ROLE DE L'AUTOANTIGENE DANS LES MALADIES AUTO-IMMUNES : ETUDE DE LA DESMOGLEINE 1 AU COURS DES PEMPHIGUS." Phd thesis, Université de Rouen, 2006. http://tel.archives-ouvertes.fr/tel-00130209.
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Les pemphigus sont des maladies auto-immunes spécifiques d'organe qui affectent la peau et les muqueuses. Ils sont caractérisés par la production d'autoanticorps pathogènes dirigés contre des protéines du desmosome, plus particulièrement, les desmogléines qui permettent l'adhésion entre eux des kératinocytes de l'épiderme. Au cours des pemphigus superficiels (PS), la réponse auto-immune est dirigée contre la desmogléine 1 (Dsg1). Chez les malades atteints de PS, des lymphocytes T autoréactifs vis à vis de la Dsg1 sont présents et interviennent dans la production d'autoanticorps anti-Dsg1. Cet autoantigène est aussi la cible de la réponse auto-immune au cours d'autres formes de pemphigus tels que les pemphigus vulgaire et paranéoplasique et par conséquent, semble jouer un rôle clé dans ces maladies. Depuis plus d'une décennie, le rôle de l'autoantigène lui-même dans l'initiation, la propagation et la pérennisation de la réponse auto-immune a été conforté par de nombreux arguments expérimentaux. En nous appuyant sur ce concept, nous avons entrepris d'étudier l'intervention de la Dsg1 dans les mécanismes physiopathologiques qui concourent au développement des pemphigus. Dans un premier temps, nous avons démontré l'expression dans l'épiderme humain, d'une isoforme tronquée de la Dsg1 générée par épissage alternatif des transcrits DSG1. Cette Dsg1 soluble est porteuse d'une séquence peptidique spécifique qui se fixe avec une forte affinité à certaines molécules HLA de classe II de susceptibilité au PS en particulier, à la molécule DRB1*0102. Ce peptide est par ailleurs capable d'induire la prolifération de cellules mononuclées du sang périphérique chez 50% des malades atteints de la forme sporadique de PS. Ces patients expriment des allèles HLA de classe II associés à la maladie et deux d'entre eux sont porteurs de la molécule DRB1*0102. Ainsi, la modification de la Dsg1 par épissage alternatif pourrait-elle intervenir dans la rupture de la tolérance au niveau du compartiment T chez les individus prédisposés génétiquement par l'expression de certains allèles HLA de classe II et in fine, conduire à l'initiation d'une réponse auto-immune B dirigée contre la Dsg1. En second lieu, nous avons observé une diversification de la réponse anticorps chez des souris normales immunisées avec la région extracellulaire recombinante de la Dsg1. Les animaux immunisés produisent non seulement des IgG dirigées contre la Dsg1 mais aussi, contre d'autres protéines de l'épiderme. Nous avons dérivé cinq anticorps monoclonaux à partir des splénocytes isolés de ces souris et montré que trois d'entre eux sont dirigés spécifiquement contre la Dsg1. Les deux autres, 10A1 et CK1, ne réagissent pas avec la Dsg1 mais reconnaissent des protéines épidermiques de plus haut poids moléculaire, compatible avec ceux des autoantigènes de la famille des plakines spécifiquement reconnus par les anticorps au cours du pemphigus paranéoplasique, e.g. l'envoplakine et la périplakine. Grâce à une analyse protéomique ciblée combinant l'immunocriblage d'une carte protéique 2D d'épiderme humain et la spectrométrie de masse MALDI-ToF, nous avons montré que la protéine cible du 10A1 est l'envoplakine, et que le CK1 reconnaît à la fois l'envoplakine et la périplakine. Ainsi, en accord ce modèle expérimental murin, la réponse B anti-Dsg1 pourrait-elle gouverner la réponse vis à vis d'autres protéines du desmosome en particulier, les plakines, mimant de ce fait la diversité de la réponse auto-immune B observée au cours du pemphigus paranéoplasique. Enfin, nous démontrons par des analyses en PCR que l'ARNm de la Dsg1 est exprimé dans le thymus humain normal, que son expression augmente avec l'âge et de ce fait, que l'absence de l'expression thymique de cette autoantigène ne constitue pas l'origine de la rupture de la tolérance qui concoure au développement des PS. Nos résultats mettent en exergue le rôle la Dsg1 dans le processus auto-immun au cours des pemphigus, d'une part, à la phase d'initiation, avec l'intervention de cet autoantigène modifié par épissage alternatif dans la réponse lymphocytaire T et d'autre part, à la phase de propagation, avec la diversification de la réponse anticorps vis à vis d'autres antigènes desmosomiaux induite chez des souris normales immunisées avec cet autoantigène.
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Bush, David Roy. "The LC/MS/MS Analysis of Pyridinoline and Desmosines in Hypertensive Mouse Aorta, Elucidation of Arginine and Proline Fragmentation for the Analysis of Arginine Metabolism in Mosquitoes by LC/MS/MS, and Mudpit Identification of B. Pseudomallei Proteins in Urine." Diss., The University of Arizona, 2013. http://hdl.handle.net/10150/293615.
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This dissertation presents an investigation into the application of mass spectrometry to the detection and quantitation of proteins and amino acids in complex biological samples. This is accomplished by two dimensional chromatography (strong cation exchange / reverse phase) coupled to tandem mass spectrometry followed by peptide spectrum matching for the detection of Burkholderia pseudomallei proteins in infected patient urine samples and reverse phase liquid chromatography coupled to a triple quadrupole and quadrupole time of flight mass spectrometers for the quantitation of cross-linked or free amino acids in mouse aorta or mosquito excreta, respectively. B. pseudomallei is a pathogenic gram negative bacillus that is endemic to the populations of Southeast Asia and is the causative agent of the disease melioidosis. LC/LC/MS/MS was used to identify candidate B. pseudomallei proteins for the development of a lateral flow immunoassay feasible for use in the impoverished communities that melioidosis affects. Three proteins (GroEL, FliC, and BipC) were identified, and have been detected in western blots and ELISAs of patient urine. Angiotensin II is known to increase both hypertension and vascular and cellular extracellular matrix remodeling, but little is known about the underlying mechanism of angiotensin II action on ECM remodeling. It has been hypothesized that the cross-linking of collagen by the enzyme lysyl oxidase increases the stiffness of the vasculature as increased levels of lysyl oxidase expression and activity have been observed in angiotensin II models of hypertension. LC/MS/MS was used to show that the cross-linking of ECM proteins increased over time in mice treated with angiotensin II over 4 weeks using a novel method to account for tissue heterogeneity in mouse aorta samples. Mosquitoes are missing a key enzyme in the urea cycle which makes arginine both an essential amino acid for mosquitoes but also makes mosquitoes unable catabolize arginine into non-toxic metabolites. MS/MS was used to show that mosquitoes excrete high levels of arginine after feeding. In addition, a previously undescribed fragmentation of arginine was elucidated using ¹⁸O labeling for future metabolism studies that would require the determination of individual arginine carbons based on fragmentation spectra.
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Dedeić, Zinaida. "Investigating the role of iASPP in cutaneous disorders." Thesis, University of Oxford, 2014. http://ora.ox.ac.uk/objects/uuid:9d393f2d-1e85-46fe-a751-427a0faa23f4.
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Desmosomes are intercellular junctions that anchor intermediate filaments to the sites of intercellular contacts. They are critical for maintaining the integrity of tissues that experience constant mechanical and structural stresses, like the skin and heart. Perturbation of desmosomal adhesion can lead to devastating epidermal and myocardial diseases. However, little is known about the regulators of desmosomes and the role of desmosomes in cell signalling events. Recent work has suggested that iASPP, an inhibitor of the p53 family of proteins, localises at the intercalated discs where desmosomes reside. However, its role at the desmosomes has remained elusive. Thus, in this thesis, it was investigated whether iASPP is a dual function protein that links desmosome adhesion to gene expression and if desmosome-related diseases develop in the absence of iASPP. iASPP was found to be a novel regulator of desmosomes, co-localising with them by physically interacting with the desmosomal components desmoplakin and K5 intermediate filaments. Loss of iASPP resulted in increased phosphorylation and solubilisation of desmoplakin, leading to the formation of K5 aggregates. This culminated in disrupted intercellular adhesion and enhanced cellular migration. Consistent with the role of iASPP in the maintenance of desmosomal adhesion integrity, focal palmoplantar keratoderma was observed in iASPP-deficient mice — a disorder often associated with desmosome dysfunction. This was accompanied by disrupted intracellular signalling, as exemplified by the disrupted expression of differentiation markers; an increase in the thickness of cell layers expressing differentiation marker K1 was noted, and K5 and K6 cells were ectopically expressed throughout the diseased palmoplantar epidermis. Impaired intercellular adhesion and migration had consequences for wound healing, as iASPP-deficient mice exhibited delayed wound closure. Furthermore, defects in eyelid closure in iASPP-deficient mice were found to be due to increased apoptosis. The localisation of apoptotic cells at the leading edge of the eyelid epidermis implied that apoptosis might have occurred due to a loss of cell-matrix or cell-cell contact, i.e. anoikis. Taken together, these results suggest that iASPP is involved in pathological (palmoplantar keratoderma), physiological (wound healing) and developmental processes (embryonic eyelid closure) through its regulation of desmosomes and their dynamics. Therefore, iASPP represents a new candidate gene in cutaneous disorders and could be implicated in a variety of epidermal and myocardial diseases.
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Yu, Chen-Han, and 余承翰. "Investigations on Desmosome Related Genes in Taiwanese Patients with Arrhythmogenic Right Ventricular Dysplasia." Thesis, 2007. http://ndltd.ncl.edu.tw/handle/58214392455622938019.
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碩士臺灣大學
藥理學研究所
95
Background: Arrhythmogenic right ventricular dysplasia (ARVD) is an inherited disorder of cardiac tissue that often occurs in people during the prime of life before 40 years old. There are four desmosome-related genes: DSP, PKP2, DSG2, and DSC2, which encode for four uniform desmosomal proteins: desmoplakin, plakophilin-2, desmoglein-2, and desmocollin-2. If a mutation of the above genes occurs, the normal function of desmosome may be impaired and results in the occurrence of arrhythmogenic right ventricular dysplasia. To date, a few mutations have been described. In this study, we enrolled 6 Taiwanese patients with ARVD and used polymerase chain reaction technique to screen the mutation in DSP, PKP2, DSG2, DSC2 genes. Methods and Results: We enrolled 6 Taiwanese patients with ARVD and 50 healthy volunteers. PCR protocol is set up to amplify 24exons of DSP, 14 exons of PKP2, 15 exons of DSG2, and 16 exons of DSC2. After agarose gel electrophoresis of products, we cut the correct band and purified it. The variations in nucleotides was identified using of DNA sequencing reaction. Five mutations were identified in 6 patients with ARVD, one in DSP and the others in DSG2. These mutations were Arg2339Gln (DSP), Asn330Asp (DSG2)、Phe531Cys (DSG2)、Glu713Lys (DSG2) and one mutant splice product. All of these mutations were novel mutations and have never been reported before. Conclusions: We used polymerase chain reaction and DNA sequencing reaction to screen the exons of DSP, PKP2, DSG2, DSC2, and identified 5 mutations in 6 Taiwanese patients with ARVD. These five novel mutations in desmosome-related genes further prove the relationship between ARVD and desmosome.
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Kuo, Yu-Ting, and 郭育廷. "Functional roles of KRT6-KRT14 fusion variant 9 in desmosome remodeling and cell spreading/migration." Thesis, 2018. http://ndltd.ncl.edu.tw/handle/49yg2x.
Full textAbstract:
碩士國立中山大學
生物醫學研究所
106
Keratins are cytoskeleton proteins with major functions in controlling/maintaining cell morphology and stiffness. Recent studies indicated possible involvement of keratin filaments in cancer developments. In our previous study, we have identified the existence of keratin fusions between KRT6 and KRT14 genes in oral squamous cell carcinoma (OSCC), and patients with the history of chewing betel nut showed higher frequency to harbor such fusions in their tumor tissues. In this study, we studied bio-functional roles of a major keratin fusion isotype K6-K14/V9, which correlated with bigger tumor size, cell migration and invasion. We established a tet-off inducible gene expression system in OSCC cells. After gene being turned on, K6-K14/V9 formed a peripheral-dominant network with the loss of peri-nuclear arrays. Interestingly, the desmosome junction was found to be dramatically reduced, leading to loss of cell-cell contact, that subsequently up-regulated genes related actin filament activation via mechano-transduction. Actin-associated protrusion at cell leading edge was also increased by K6-K14/V9 to enhance cell migration and invasion. This study uncover new functions/definitions for keratin filament in cancer development, Mechanisms revealed in this study can help us to design new strategies for new drug development and clinical treatments.
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Barahona-Dussault, Catherine. "Rôle du test génétique dans la cardiomyopathie arythmogène du ventricule droit. Étude sur une cohorte prospective unicentrique." Thèse, 2008. http://hdl.handle.net/1866/2789.
Full textAbstract:
La cardiomyopathie/dysplasie arythmogène du ventricule droit (ARVC/D) est un
désordre d’origine génétique caractérisé par le remplacement du myocarde par du tissus
fibro-adipeux dans le ventricule droit. Ce désordre est responsable d’un grand
pourcentage de mort subite, spécialement chez les plus jeunes. ARVC/D est difficile à
diagnostiquer avec les outils cliniques actuels. Elle est causée en grande majorité par
des mutations dans les protéines desmosomales. ARVC/D a donc des implications
d’une grande importance chez les membres de la famille, qui peuvent sans le savoir,
être aussi à risque de mort subite. Dans le but d’améliorer le diagnostique, un nouvel
outil, le test génétique, est de plus en plus utilisé.
Hypothèses: Dans le but d’évaluer la valeur du test génétique en complément du test
clinique classique chez ARVC/D nous avons effectué une investigation clinique et
génétique chez 23 cas-index atteints.
Méthodes: Les cas-index sont diagnostiqué après une mort subite dans la famille ou
après un examen clinique poussé pour arythmies. Le diagnostique d’ARVC/D a été fait
avec les outils cliniques selon les critères. L’analyse génétique des protéines
desmosomales associées à la maladie a été effectuée en séquençant leurs exons ainsi que
les régions introniques nécessaires à l’épissage alternatif.
Résultats: Le diagnostique clinique était clair dans 18/23 et incertain dans 5/23 des
individus. Nous avons identifié 15 différentes mutations chez 10 cas-index. 64% des
mutations n’avaient jamais été décrites. De plus, nous avons observé la présence de
double ou triple mutant dans 40% des cas-index positifs. Les individus avec mutations
sont plus jeunes et ont plus de symptômes que les individus sans mutation.
Conclusion: Les tests génétiques sont positifs dans 43% des patients avec ARVC/D.
L’utilisation de la technologie génétique basée sur l’identification de mutations connues
a une valeur limitée vu le haut pourcentage des mutations nouvelles dans la maladie. La
présence de double, même de triple mutant n’est pas associé avec un phénotype plus
sévère, mais renforce l’idée de la nécessité d’un test génétique pour tous les gènes. Le
test génétique est un outil fort utile à ajouter aux tests cliniques pour le diagnostique des
patients qui ne remplissent pas tous les critères cliniques de la maladie.
Mots clés: génétique, ARVC/D, mort subite, desmosomeArrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVC/D) is a genetic disorder characterized by the presence of fibro-fatty replacement of the myocardium in the right ventricle. The disease is thought responsible for an important percentage of sudden cardiac death in the young. Hence the disease is usually difficult to diagnose with present clinical tools. ARVC/D is it caused in greater part by mutations in desmosomal proteins. The diagnosis of the genetic carriers bears important implications in family members, who unknowingly may be at risk for sudden death. In order to improve the diagnosis, a new tool, genetic testing, is increasingly being used. Hypothesis: In order to assess the value of genetic testing in complementing clinical testing in ARVC/D, we undertook the project to collect and perform clinical and genetic investigation in 23 probands with the disease. Methods: The probands were usually identified either after the death of a family member or after their clinical investigation for arrhythmias. The diagnosis of ARVC was made with clinical tools according to accepted criteria. Genetic analysis of desmosomal proteins previously associated with the disease was performed by sequencing the exons and intron-exon boundaries. Results: The clinical diagnosis was clear in 18/23 and suspicious in 5/23 individuals. We identified 15 different mutations in 10 probands. 64% of the mutations were not previously described. Interestingly we also observed the presence of double or triple mutants in 40% of the positive individuals. Individuals with mutations were younger and had more symptoms than individuals with no mutation. Conclusion: Genetic testing is useful in 43% of patients with ARVC. The use of mutation-based genetic technology has a very limited value due to the high percentage of previously unknown mutations in this disease. The presence of double and even triple mutants is not associated with a more severe phenotype but it indicates the need to have genetic testing performed for all genes for familial screening. Genetic testing is a useful tool to add to the clinical testing for the diagnosis of patients who do not completely fulfill the clinical criteria for the disease. Key words: genetic, ARVC/D, sudden death, desmosome
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Sumigray, Kaelyn D. "Novel Roles for Desmosomes in Cytoskeletal Organization." Diss., 2011. http://hdl.handle.net/10161/5009.
Full textAbstract:
Microtubules often adopt non-centrosomal arrays in differentiated tissues, where they are important for providing structure to the cell and maintaining polarity. Although the formation and organization of centrosomal arrays has been well-characterized, little is known about how microtubules form non-centrosomal arrays.
In the mouse epidermis, centrosomes in differentiated cells lose their microtubule-anchoring ability through the loss of proteins from the centrosome. Instead, microtubules are organized around the cell cortex. The cell-cell adhesion protein desmoplakin is required for this organization. Our model is that desmoplakin recruits microtubule-anchoring proteins like ninein to the desmosome, where they subsequently recruit and organize microtubules.
To test this model, we confirmed that the microtubule-binding proteins Lis1, Ndel1, and CLIP170 are recruited by desmoplakin to the cell cortex. Furthermore, by creating an epidermis-specific conditional Lis1 knockout mouse, I found that Lis1 is required for cortical microtubule organization. Surprisingly, however, Lis1 is also required for desmosome stability. This work reveals essential desmosome-associated components that control cortical microtubule organization and unexpected roles for centrosomal proteins in epidermal function.
Although Lis1 is required for microtubule organization, it is not sufficient. I created a culture-based system to determine what other factors may be required for cortical organization for microtubules. My work reveals that stabilization of the microtubules is sufficient to induce their cortical organization. Functionally, cortical microtubules are important for increasing the mechanical integrity of cell sheets by engaging adherens junctions. In turn, tight junction activity is increased. Therefore, I propose that cortical microtubules in the epidermis are important in forming a robust barrier by cooperatively strengthening each cell-cell junction.
To determine whether desmosomes play similar roles in simple epithelia as stratified epithelia, I examined intestinal epithelial-specific conditional desmoplakin conditional knockout mice. Unexpectedly, I found that desmoplakin is not required for cell-cell adhesion and tissue integrity in the small intestine. Furthermore, it does not organize intermediate filaments. Desmoplakin is required, however, for proper microvillus architecture.
Overall, my studies highlight novel tissue-specific roles for desmosomes, in particular desmoplakin, in organizing and integrating different cytoskeletal networks. How desmoplakin's function is regulated in each tissue will be a new interesting area of research.
Dissertation
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Zhang, Kehan. "Mechanotransduction through cytoskeleton and junctions in cardiomyopathies." Thesis, 2020. https://hdl.handle.net/2144/41029.
Full textAbstract:
Cardiomyopathies represent a heterogeneous group of diseases of the heart muscle that often lead to progressive heart failure with high morbidity and mortality. In a significant and increasing percentage of the patient population, cardiomyopathies have been associated with hereditary mutations in genes encoding critical cellular components that make up the cytoarchitecture of cardiac muscle cells, or cardiomyocytes. While specific mutations have been linked to different classes of cardiomyopathies, it is however not well understood how these mutations cause cytostructural abnormalities that ultimately lead to dysfunction of cardiomyocytes. To gain insights into the pathogenesis of inherited cardiomyopathies, we focus in this thesis on a particular set of mutations in the cardiac cytoskeleton and desmosomes that are associated with dilated and arrhythmogenic cardiomyopathies, and probe their pathogenic mechanisms using cardiomyocytes derived from human induced pluripotent stem cells and bioengineered culture platforms. In part one, we describe the mechanical and molecular basis for the assembly of sarcomeres, the fundamental contractile units within cardiomyocytes, and reveal how mutations in titin (TTN) abolish this process by disrupting cell-matrix interaction and impairing diastolic force generation, a hallmark of dilated cardiomyopathy. In the second part of this thesis, we reveal that plakophilin-2 (PKP2) mutations that are associated with arrhythmogenic cardiomyopathy lead to impaired systolic function by destabilizing cell-cell junctions and in turn disrupting sarcomere stability and organization. Together, our studies establish a deeper understanding of how cell-matrix and cell-cell interactions contribute to the organization and function of cardiomyocytes and how disruption of these interactions by pathogenic mutations lead to cardiac dysfunction.2022-05-18T00:00:00Z
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Lee, DonChing, and 李東慶. "Identification and Characterization of Proteins Interacting with Pinin, A Moonlighting Protein Located Both at the Desmosome and within the Nucleus." Thesis, 2000. http://ndltd.ncl.edu.tw/handle/23211914414592074843.
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