The aim of this thesis was to develop a molecular imaging probe for the urokinase plasminogen activator (uPA) system, which has been shown to play a critical role in cancer metastasis, tumour aggressiveness and likelihood of progression. Two classes of small molecule inhibitors carrying isotopes of iodine were synthesized and evaluated using in vitro assays and in vivo studies. Lead compounds showed high affinity for the target with Ki values in the low nanomolar range (1b = 1.4 nM, 1e = 6.1 nM, 1g = 2.6 nM and 2a = 2.1 nM). Biodistribution studies of the reversible compounds (1b, 1e, 1g) showed rapid clearance, accumulation in the gall bladder and intestines and little to no tumour uptake (<1 %ID/g). The irreversible inhibitor (2a) showed specificity for the target through SDS-PAGE and biodistribution studies. Analysis of the biodistribution pattern showed retention in the tumour over time reaching a maximum at 24 h post-injection of 1.95 %ID/g with tumour-to-blood ratio being 0.65 at 24 h, 1.13 at 48 h and 1.09 at 96 h post-injection. A parallel strategy reported involved targeting the uPA receptor (uPAR) through the use of antibodies and bioorthogonal chemisty based on radiolabeled tetrazines and transcyclooctene (TCO) functionalized biomolecules. A new tetrazine synthon was developed that can be readily labeled with both 99mTc and 18F where the products were produced in 75 and 31 % radiochemical yields. Stability studies showed the compounds are suitable for use in vivo. Biodistribution studies were carried out in CD1 mice and results showed that both probes had sufficient distribution patterns to warrant use in pre-targeting strategies. Their reactivity with TCO, including functionalized derivatives such as TCO-anti-uPAR, was also demonstrated creating the means to develop PET and SPECT probes for imaging the urokinase system using a single prosthetic group. / Thesis / Master of Science (MSc)

La distròfia muscular de Duchenne (DMD) és una de les malalaties recessives lligades al cromosoma X més comuns essent la seva causa primària el dèficit de distrofina, proteïna localitzada en la cara interna del sarcolema. La soca de ratolí mdx també presenta un dèficit total de distrofina i és el model animal més àmpliament utilitzat per aquesta malaltia. Malgrat el seu dèficit de distrofina, el ratolí mdx no presenta un fenotip distròfic tan sever com l'observat en la DMD humana. S'han apuntat dues hipòtesis per tractar d'explicar aquestes diferències entre la malaltia humana i el model murí. La primera atribuirïa al ratolí capacitat regenerativa més gran, mentre que la segona li atribuirïa un descens del seu ritme de necrosi a partir de cert moment de la seva vida. En aquest treball intentem clarificar aquest aspecte mitjançant diferents aproximacions: i) caracterizació morfològica de la patologia muscular durant tota la vida del ratolí mdx i estudi de marcadors de necrosi-regeneració, ii) estudi de la expressió de utrofina (proteïna que presenta elevada homologia amb la distrofina) i b-distroglicà (el lligand natural de la distrofina), iii) caracterització del paper de la proteasa uPA en la regeneració muscular, iv) el doble mutant mdx/uPA-/- com model de necrosi espontània i capacitat regenerativa disminuïda i finalment v) identificació i caracterització de noves proteïnes implicades en el procés de regeneració muscular i/o protecció enfront de la necrosi. Els resultats derivats de la caracterització morfològica apunten que el ratolí mdx no presenta necrosi durant les seves dues primers setmanes de vida, moment a partir del qual comença un increment d'aquest fenòmen arribant a un màxim al voltant dels dos mesos per a, posteriorment experimentar un descens gradual però important de l'activitat necròtica. L'anàlisi evolutiu de marcadors de necrosi-regeneració també comfirma aquesta aparició i posterior decrement de la necrosi muscular. L'estudi evolutiu de l'expressió de utrofina, ens ha permès observar com aquesta preoteïna despareix a les dues setmanes de vida, coincidint amb l'inici del descens de la necrosi muscular en els ratolins mdx, mentre que el seu posterior increment en aquests ratolins coincideix amb l'inici de la fase en que la necrosis es mostra molt més lleu. A més, els nivells de utrofina presenten una bona correlació amb els nivells b-distroglicà, comfirmant la capacitat de la utrofina per unir-se amb el que és el lligand natural de la distrofina. Aquests resultats atribueixen a la utrofina un paper com a substituta de al distrofina en el ratolí carent de distrofina.Per altre banda, el ratolí carent de uPA (uPA-/-) presenta una important reducció de la capacitat regenerativa muscular, presentant un retard temporal important en el procés regeneratiu, una acumulació de fibrina en el teixit necròtic i una disminució del reclutament de macròfags en la zona lesionada respecte els ratolins control. El tractament amb Ancrod (una toxina fibrinolítica) restaura sensiblement la seva su capacitat de regeneració. Per la seva banda, el múscul del ratolí doble mutant mdx/uPA-/- presenta un aspecte més distròfic, destacant importants acumulacions de fibres necròtiques calcificades, un augment de grups necròtics deguts a una disminució de la seva capacitat regenerativa, un pes corporal menor que les soques mdx i uPA-/- així com una mortalitat propera al 80% tot i que aquesta es concentra exclusivament en el periode de necrosi màxima. Aquests resultats confirmen la importància que té la proteasa uPA en el procés de la regeneració muscular i ens mostren com la capacitat regenerativa només només és limitant en el ratolí mdx durant el periode de màxima necrosi.Finalment, mitjançant un estudi d'expressió diferencial de gens entre ratolins controls i mdx hem pogut identificar 9 seqüències clarament sobreexpressades en ratolins mdx, essent les més interessants la tetraspanina CD63 i la seqüència anomenada B2. Experiments destinats a analitzar l'expressió d'aquestes dues seqüències ens indiquen que CD63 és també expressada en els ratolins control tot i que a nivells lleugerament inferiors que els mdx i que la seva expressió és elevada però constant durant la diferenciació de la línia mioblàstica murina C2C12 in vitro. La B2 es clarament diferencial, essent expressada per ratolins mdx i no expressada o a nivells molt baixos en controls, presentant un pic d'expressió als 3 i 6 dies de diferenciació en la línia mioblàstica murina C2C12 in vitro. / Duchenne Muscular Dystrophy (DMD) is one of the most common X-linked diseases. DMD is due to mutations in the DMD gene, which results in lack or dysfunction of dystrophin, a 427 KDa protein localised at the inner face of the sarcolemma. The mdx mice, the most useful animal model for DMD, carry a nonsense mutation in exon 23 of the dystrophin gene that causes lack of dystrophin. Despite some genetic, biochemical and histological similarities between DMD and mdx mice, clinical manifestations are less severe in mdx mice than humans. To explain these differences, some authors proposed that mdx mice have greater regenerative capacity than humans, thus allowing them to repair the continuous necrotic events of their skeletal muscle fibres. On the other hand, it has been suggested that a decreased necrosis in mid-age and old mdx mice may underlie the clinical differences between mdx mice and DMD patients. In this work we clarify these questions using different approximations: i) morphologic characterisation of the muscular pathology during mdx mice lifespan and study of the evolution of necrosis-regeneration markers, ii) evolutive utrophin (dystrophin high-homology protein) expression study and b-dystroglycan (the natural ligand of dystrophin), iii) characterisation of the role of urokinase type plasminogen activator (uPA) during muscular regeneration, iv) the double mutant mdx/uPA-/- as both spontaneous necrosis and impaired regeneration model and finally v) identification and characterisation of new sequences implicated in the muscular regeneration process and or in the protection against necrosis. The results obtained in the morphologic characterisation indicate that necrosis in mdx mice is scarce during first two weeks, become very active with a peak at two months and then progressively fade, remaining at very low levels. The evolutive analysis of the necrosis-regeneration markers also confirms its apparition and posterior decrement. The evolutive study of utrophin expression shows that this protein practically disappear at two weeks of age, coinciding with the onset of necrosis and its posterior increase in mdx mice is concomitant with the decrease of the muscular necrosis. Furthermore, utrophin levels show a consistent correlation with b-dystroglycan levels, confirming that utrophin is able to bind and stabilise b-dystroglycan in the sarcolemma. These results confirm a role of utrophin as a substitute of dystrophin in the mdx mice sarcolemma.The uPA deficient mice (uPA-/-) suffer an important muscular regenerative capacity reduction, showing a severe delay in the regeneration process, an abnormal fibrin accumulation in the necrotic tissue and a reduction of macrophage recruitment in the zone of injury. Ancrod treatment (fibrinolitic toxin) restores significantly its regenerative capacity. Interestingly, the double mutant mdx/uPA-/- muscle shows a more severe phenotype than mdx, with important accumulation of necrotic fibbers with calcium depositions, an increment of degenerative-regenerative groups due to the reduction of its regenerative capacity, a reductions on its corporal weight compared to mdx and uPA -/- strains and the mortality of the strain is near the 80% but concentrated exclusively in the maximal necrosis period. These results demonstrate the critical importance of uPA in the muscular regeneration process and show that, in a mdx background, the regenerative capacity is only limiting during the maximal necrosis period.Finally, using a differential expression study comparing mdx and control mice we have identified 9 sequences overexpressed in mdx mice, being the more interesting ones the tetraspanin CD63 and an unknown sequence named B2. The characterisation of its expression shows that CD63 is expressed in control mice but in the mdx mice its expression is slightly higher and that its expression is high and constant during the differentiation of the murine mioblastic line C2C12 in vitro. B2 expression is clearly differential, induced in mdx mice and not expressed or expressed at a very low levels in controls, and shows a peak of expression at days 3 and 6 days of differentiation in the murine mioblastic line C2C12 in vitro.

O presente trabalho utiliza marcadores moleculares para auxiliar na caracterização e filogenia das espécies de macroalgas vermelhas do gênero Hypnea na costa do Brasil. O gênero Hypnea Lamouroux (1813) apresenta cerca de 67 espécies descritas e possui distribuição geográfica em águas quentes ao redor do mundo. Além disso, o gênero possui grande importância econômica e ecológica, como fonte de alimento e produção industrial de carragenana. Porém, a identificação das espécies de Hypnea com base exclusivamente em dados morfológicos é dificultada em virtude da plasticidade fenotípica presente no grupo, de sua morfologia relativamente simples e da ampla distribuição geográfica de suas espécies. Em vista disso, utilizamos a técnica de \"DNA barcoding\" que permite a análise de um grande número de amostras. Neste trabalho foram utilizados dois \"DNA barcodes\" (a região 5\' do gene mitocondrial cox1 e o \"universal plastid amplicon\" UPA), e para as análises filogenéticas foi utilizado o gene plastidial rbcL. Além disso, estudos morfológicos foram feitos a fim de delimitar o real valor dos caracteres morfo-anatômicos citados na literatura para a separação de espécies do gênero Hypnea. Ao todo, foram obtidas 230 amostras brasileiras de Hypnea, provenientes de 11 estados brasileiros, e 10 amostras de outros países. Um total de 367 sequências de marcadores moleculares foi obtido neste trabalho. Confirmamos a ocorrência de nove espécies para o gênero: H. cervicornis, \"H. flexicaulis\", \"H. musciformis\", H. spinella, \"H. stellulifera\", Hypnea sp. 1, Hypnea sp. 2, Hypnea sp. 3 e Hypnea sp. 4. As amostras coletadas e previamente identificadas em campo como H. cornuta revelaram-se, pelos estudos da biologia molecular, serem \"H. stellulifera\". As espécies H. musciformis, H. nigrescens, H. valentiae foram consideradas variações morfológicas de uma mesma espécie, denominada de \"H. musciformis\". A identificação das espécies com base apenas em características morfológicas mostrou-se insatisfatória, devido principalmente a plasticidade fenotípica presente no grupo, além da existência de espécies com morfologias convergentes. A técnica de \"DNA barcode\", principalmente com relação ao marcador cox1, mostrou-se essencial na identificação e delimitação das espécies, revelando cenários que passariam despercebidos com o uso apenas da morfologia / This study uses molecular markers to aid in the characterization and phylogeny of species of the genus Hypnea, a red macroalgae, on the coast of Brazil. The genus Hypnea Lamouroux (1813) presents about 67 described species and has geographical distribution in warm waters around the world. Furthermore, the genus has great economic and ecological importance as a source of food and industrial production of carrageenan. However, the identification of the species of Hypnea based solely on morphological data is difficult due to phenotypic plasticity present in this group, its relatively simple morphology and broad geographical distribution of its species. In view of this, we use the technique of \"DNA barcoding\" that allows the analysis of a large number of samples. In this work we used two \"DNA barcodes\" (the 5 \'region of the mitochondrial gene cox1 and universal plastid amplicon UPA), and for phylogenetic analysis the plastid gene rbcL was used. In addition, morphological studies were made in order to delimit the actual value of morpho-anatomical caracters cited in the literature for the separation of species of Hypnea. Altogether, 230 Hypnea samples were obtained from 11 Brazilian states, and 10 samples from other countries. A total of 367 sequences of molecular markers were obtained in this study. We confirm the occurrence of nine species of the genus: H. cervicornis, \"H. flexicaulis\", \"H. musciformis\", \"H. spinella, \"H. stellulifera\", Hypnea sp. 1, Hypnea sp. 2, Hypnea sp. 3 and Hypnea sp. 4. Samples collected in the field and previously identified as H. cornuta based on molecular data, proved to be \"H. stellulifera\". The species H. musciformis, H. nigrescens and H. valentiae were considered morphological variations of the same species, named \"H. musciformis\". The identification of species based on morphological characteristics proved unsatisfactory, mainly due to phenotypic plasticity in this group and the existence of species with convergent morphologies. The technique of \"DNA barcode\", especially with respect to cox1 marker, was essential for the identification and definition of species, revealing scenarios that would go unnoticed by using only morphology

Duchenne muscular dystrophy (DMD) is a fatal degenerative disorder of locomotor and respiratory muscles, in which myofibers are progressively replaced by non-muscular fibrotic tissue. Here, we show that fibrin/ogen accumulates in dystrophic muscles of DMD patients and of the mdx mouse model of DMD. Genetic loss or pharmacological depletion of fibrin/ogen in mdx mice attenuated muscular dystrophy progression and improved locomotor capacity. More importantly, fibrin/ogen depletion reduced fibrosis in mdx mouse diaphragm. Our data indicate that fibrin/ogen, through induction of IL-1 Ò, drives the synthesis of TGF Ò by mdx macrophages, which in turn, induces collagen production in mdx fibroblasts. Fibrin/ogen-produced TGF Ò further amplifies collagen accumulation through recruitment and activation of pro-fibrotic alternatively activated macrophages. Fibrin/ogen also stimulated collagen synthesis directly in mdx fibroblasts, via Ñv Ò3 integrin engagement. In addition, when analyzing a group of 39 DMD patients, fibrin/ogen accumulation in locomotor muscles was found associated with fibrosis and disease severity. These data unveil a novel role of fibrin/ogen in muscular dystrophy and, importantly, in the replacement of muscle by fibrotic tissue.

O presente trabalho utiliza marcadores moleculares para auxiliar na caracterização e filogenia das espécies de macroalgas vermelhas do gênero Hypnea na costa do Brasil. O gênero Hypnea Lamouroux (1813) apresenta cerca de 67 espécies descritas e possui distribuição geográfica em águas quentes ao redor do mundo. Além disso, o gênero possui grande importância econômica e ecológica, como fonte de alimento e produção industrial de carragenana. Porém, a identificação das espécies de Hypnea com base exclusivamente em dados morfológicos é dificultada em virtude da plasticidade fenotípica presente no grupo, de sua morfologia relativamente simples e da ampla distribuição geográfica de suas espécies. Em vista disso, utilizamos a técnica de \"DNA barcoding\" que permite a análise de um grande número de amostras. Neste trabalho foram utilizados dois \"DNA barcodes\" (a região 5\' do gene mitocondrial cox1 e o \"universal plastid amplicon\" UPA), e para as análises filogenéticas foi utilizado o gene plastidial rbcL. Além disso, estudos morfológicos foram feitos a fim de delimitar o real valor dos caracteres morfo-anatômicos citados na literatura para a separação de espécies do gênero Hypnea. Ao todo, foram obtidas 230 amostras brasileiras de Hypnea, provenientes de 11 estados brasileiros, e 10 amostras de outros países. Um total de 367 sequências de marcadores moleculares foi obtido neste trabalho. Confirmamos a ocorrência de nove espécies para o gênero: H. cervicornis, \"H. flexicaulis\", \"H. musciformis\", H. spinella, \"H. stellulifera\", Hypnea sp. 1, Hypnea sp. 2, Hypnea sp. 3 e Hypnea sp. 4. As amostras coletadas e previamente identificadas em campo como H. cornuta revelaram-se, pelos estudos da biologia molecular, serem \"H. stellulifera\". As espécies H. musciformis, H. nigrescens, H. valentiae foram consideradas variações morfológicas de uma mesma espécie, denominada de \"H. musciformis\". A identificação das espécies com base apenas em características morfológicas mostrou-se insatisfatória, devido principalmente a plasticidade fenotípica presente no grupo, além da existência de espécies com morfologias convergentes. A técnica de \"DNA barcode\", principalmente com relação ao marcador cox1, mostrou-se essencial na identificação e delimitação das espécies, revelando cenários que passariam despercebidos com o uso apenas da morfologia / This study uses molecular markers to aid in the characterization and phylogeny of species of the genus Hypnea, a red macroalgae, on the coast of Brazil. The genus Hypnea Lamouroux (1813) presents about 67 described species and has geographical distribution in warm waters around the world. Furthermore, the genus has great economic and ecological importance as a source of food and industrial production of carrageenan. However, the identification of the species of Hypnea based solely on morphological data is difficult due to phenotypic plasticity present in this group, its relatively simple morphology and broad geographical distribution of its species. In view of this, we use the technique of \"DNA barcoding\" that allows the analysis of a large number of samples. In this work we used two \"DNA barcodes\" (the 5 \'region of the mitochondrial gene cox1 and universal plastid amplicon UPA), and for phylogenetic analysis the plastid gene rbcL was used. In addition, morphological studies were made in order to delimit the actual value of morpho-anatomical caracters cited in the literature for the separation of species of Hypnea. Altogether, 230 Hypnea samples were obtained from 11 Brazilian states, and 10 samples from other countries. A total of 367 sequences of molecular markers were obtained in this study. We confirm the occurrence of nine species of the genus: H. cervicornis, \"H. flexicaulis\", \"H. musciformis\", \"H. spinella, \"H. stellulifera\", Hypnea sp. 1, Hypnea sp. 2, Hypnea sp. 3 and Hypnea sp. 4. Samples collected in the field and previously identified as H. cornuta based on molecular data, proved to be \"H. stellulifera\". The species H. musciformis, H. nigrescens and H. valentiae were considered morphological variations of the same species, named \"H. musciformis\". The identification of species based on morphological characteristics proved unsatisfactory, mainly due to phenotypic plasticity in this group and the existence of species with convergent morphologies. The technique of \"DNA barcode\", especially with respect to cox1 marker, was essential for the identification and definition of species, revealing scenarios that would go unnoticed by using only morphology

Doctor of Philosophy / Department of Chemistry / Stefan H. Bossmann / This paper describes two methods for utilizing cancer associated proteases for targeting cancer therapy to the tumor. The first method is designing a drug delivery system based on liposomes that are sensitive to cancer associated proteases. Upon contact with the protease, the liposome releases its contents. The second method is designing a prodrug that is based on a porin isolated from Mycobacterium smegmatis. The porin is modified with protease consensus sequences, inhibiting its toxicity. Upon contact with the protease, the drug is activated. Protease sensitive liposomes were synthesized that were sensitive to urokinase plasminogen activator. This was done by synthesizing a cholesterol-anchored, uPA consensus – sequence-containing, acrylic acid block copolymer and using it to form a covalently bound polymer cage around the outside of a hypertonic liposome. Liposomes were synthesized that had a diameter of 136 nm. Upon addition of the polymer the diameter increased by 2.69 nm, indicating it had successfully embedded into the liposome membrane. After crosslinking with either a short peptide containing a lysine (so that it is a diamine) or ethylenediamine, the diameter increased between 5.33 nm and 14.1 nm (depending on the type and amount of the crosslinked). Fluorescence release assays showed that the polymer cage could add in excess of thirty atmospheres of osmotic pressure resistance, and, under isobaric conditions, would prevent release of much of the liposomal contents. Upon treatment with uPA, the polymer caged liposomes released a significantly larger amount of their contents making the liposomes protease sensitive. MspA was shown to be a very stable protein able to be imaged by AFM. AFM imaging demonstrated that MspA is able to form native pore structures in membranes making it a good imitator of the membrane attack complex. MspA was demonstrated to be highly cytotoxic, but poor at distinguishing between cells. Pro-MspA was synthesized by adding a hydrophilic peptide to MspA that prevents insertion. A uPA cleavage sequence embedded causes the MspA to become activated at the cancer site. This was demonstrated in tests against uPA and non-uPA producing cell lines.

Introduction: The human menstrual cycle is regulated by sex-steroid hormones, including oestrogen (E), progesterone (P4) and androgens which act by ligand binding to their cognate receptors. Perturbation of the complex series of events governing the menstrual cycle may lead to heavy menstrual bleeding (HMB). This is a common debilitating condition and often associated with uterine fibroids. There remains an unmet need for effective, long-term medical treatment so women avoid surgery and preserve their fertility. Selective progesterone receptor modulators (SPRMs, e.g. ulipristal acetate, UPA) are synthetic ligands that bind the progesterone receptor (PR). Many SPRMs have been developed but only mifepristone (for the management of unwanted pregnancy) and UPA are in current clinical use. UPA is licensed for the intermittent treatment of symptomatic fibroids. SPRMs have potential utility for treatment of HMB as administration rapidly induces amenorrhoea but the mechanisms by which this is achieved are unknown. SPRM administration results in unique endometrial morphological changes (progesterone receptor modulator-associated endometrial changes; PAEC). Despite endometrial unopposed estradiol exposure these morphological changes do not appear to be associated with malignancy or pre-malignancy risk. Indeed endometrial cell proliferation appears reduced despite relative progesterone-antagonism. Based upon findings with other SPRMs it was hypothesised that: (i) administration of UPA would have an endometrial specific effect upon the reproductive tract, with regard to alteration in morphology, localisation of sex steroid receptors (SSR) and cell proliferation.; (ii) administration of UPA would impact upon progesterone-regulated (Pregulated) genes in the endometrium. Methods: The data presented within this thesis are derived from biopsies obtained at hysterectomy from the endometrium, fallopian tubes and cervices of women with symptomatic fibroids administered UPA for 8-15 weeks. Samples were obtained for histological assessment, immunohistochemistry and RNA extraction for subsequent quantitative RT-qPCR of sex-steroid receptors (SSR) and proliferation markers. In addition key P-regulated genes within the endometrium were investigated by RT-qPCR and selected protein expression. To further interrogate the anti-proliferative effect, RNA was extracted from “paired” endometrial biopsies from the same woman in the proliferative phase of the menstrual cycle and following subsequent treatment with UPA for at least eight weeks and microarray gene analyses undertaken. Results: Morphological alteration of the endometrium with UPA administration was consistent with previously published data, but with a higher prevalence than previously described. There was a striking alteration in expression and localization of SSRs, particularly PR and androgen receptor (AR), and alteration of many P-regulated genes, consistent with UPA acting with low progesteroneagonism within the endometrium. There was no alteration of SSR expression within the cervix and proliferation was unchanged. Fallopian tube morphology and SSR expression was consistent with proliferative phase but cell proliferation was reduced following UPA administration, consistent with secretory phase levels. Microarray analyses identified multiple transcripts altered relative to proliferative phase, with GREM2 the most significantly down-regulated gene and MUC1 one of the most significantly upregulated genes. Consistent with low levels of mitotic figures and cell proliferation, the most down regulated KEGG pathway was the cell cycle. Multiple elements within this were subsequently validated (RT-qPCR) and included key regulators of all elements of the mitotic cell cycle, many of which were novel to those previously described following administration of another SPRM, mifepristone. In summary the novel data presented in this thesis considerably extend the data available to date concerning the actions of the SPRM, UPA, on the female reproductive tract, and increases knowledge regarding a compound with promising utility for the management of the debilitating complaint of HMB.

Dissertação (mestrado)—Universidade de Brasília, Faculdade de Ciências da Saúde, Programa de Pós-Graduação em Enfermagem, 2017. / Submitted by Albânia Cézar de Melo (albania@bce.unb.br) on 2017-09-28T16:35:43Z No. of bitstreams: 1 2017_SimoneSouzaNascimento.pdf: 3875863 bytes, checksum: a504c8969b6cadd611b14f833737610d (MD5) / Approved for entry into archive by Raquel Viana (raquelviana@bce.unb.br) on 2017-10-09T21:28:57Z (GMT) No. of bitstreams: 1 2017_SimoneSouzaNascimento.pdf: 3875863 bytes, checksum: a504c8969b6cadd611b14f833737610d (MD5) / Made available in DSpace on 2017-10-09T21:28:57Z (GMT). No. of bitstreams: 1 2017_SimoneSouzaNascimento.pdf: 3875863 bytes, checksum: a504c8969b6cadd611b14f833737610d (MD5) Previous issue date: 2017-10-09 / Introdução: Os serviços de emergência tem funcionado como porta de entrada para o sistema de saúde, mesmo em casos sem gravidade, o que causa filas de espera, superlotação, demora para atendimento, insatisfação do usuário e sobrecarga de serviço aos profissionais. É neste contexto que a qualidade da assistência e a implantação do processo de enfermagem como ferramenta de trabalho, apesar de importante, tem sido deixado em segundo plano. O objetivo deste estudo foi avaliar os diagnósticos de enfermagem dos clientes atendidos em uma Unidade de Pronto Atendimento, admitidos nas Salas Amarela e Vermelha, fundamentado no referencial teórico de Imogene King e na Taxonomia da NANDA-I. Método: Tratou-se de pesquisa de abordagem quantitativa do tipo descritiva, com delineamento transversal. Foi realizado em Unidade de Pronto Atendimento do Distrito Federal, com os pacientes classificados como laranja e/ou vermelho admitidos nas Salas Amarela e Vermelha. Os dados foram coletados no período de novembro de 2016 a fevereiro de 2017, a partir da aplicação do Processo de Enfermagem e estruturados no modelo de Imogene King. A coleta foi realizada pela própria autora, com 50 clientes, que assinaram o Termo de Consentimento Livre e Esclarecido. Resultados: Quanto ao perfil observou-se discreta predominância de indivíduos do sexo feminino (54%). A idade dos clientes entrevistados variou entre 18 e 91 anos, com média de 50,48 anos. Em relação à escolaridade, a amostra caracterizou-se com baixa escolaridade. Quanto a raça/cor houve predominância de indivíduos que se autodeclararam pardos (44%). Em relação ao estado conjugal, 60% eram casados. Sobre os dados referentes à moradia, 76% estavam na zona urbana e 62% moravam em imóvel próprio. Saneamento básico completo foi identificado em 64% da amostra. Sobre a fonte de renda, 44% exerciam alguma atividade laboral e 36% estavam desempregados. Sobre as condições patológicas pré-existentes as mais prevalentes foram a hipertensão arterial (40,0%) e o diabete melito (30,0%). A amostra também caracterizou-se pela polimedicação. Dos indivíduos 20% eram tabagistas, 22,0% eram etilistas e apenas 18,0% praticavam algum exercício físico regularmente. Apenas 34,0% dos clientes já haviam passado por pelo menos um atendimento, em outro momento, na unidade de estudo. Sobre a classificação de risco, 94,0% foram classificados como Laranja e 68% foram entrevistados na Sala Amarela. As queixas predominantes foram dor no peito e falta de ar com 28,0% cada. Foram identificados 13 fluxogramas diferentes nos clientes pertencentes ao estudo, e os de maior prevalência foram Dor torácica (18,0%), Mal-estar em adulto (14,0%), Dispneia em adulto e Dor abdominal em adulto (12,0% cada). Utilizando a taxonomia da NANDA-I foram arrolados ao todo 722 diagnósticos de enfermagem com média de 14,44 diagnósticos por cliente. Foram identificados 94 diagnósticos, sendo 54 diagnósticos com foco no problema (reais), dois diagnósticos de promoção da saúde e 38 de risco (ou vulnerabilidade). Analisando sob a perspectiva dos três sistemas mencionados por King observou-se que os diagnósticos encontrados contemplaram todos os sistemas propostos, e estão divididos de forma percentual em sistema pessoal com 88,3%, sistema interpessoal com 9,6% e sistema social com 2,1%. Apenas nove diagnósticos apresentaram frequências superiores a 40%, sendo eles: risco de infecção (90,0%), conhecimento deficiente (80,0%), risco de quedas (64,0%), estilo de vida sedentário (62,0%), dentição prejudicada (60,0%), risco de perfusão renal ineficaz (48,0%), risco de sangramento (46,0%), fadiga (46,0%) e dor aguda (42,0%). Entre alguns fatores causais presentes nos diagnósticos prioritários destacaram-se: conhecimento deficiente por insuficiência de informações sobre o tratamento ou o problema de saúde; sedentarismo por interesse insuficientes pela atividade física; dentição prejudicada por dificuldades de acesso a cuidados dentários profissionais; fadiga decorrente de condições fisiológicas limitantes; e dor devido a agente lesivo biológico. Conclusão: Os resultados obtidos neste estudo, servirão de apoio aos profissionais atuantes em unidades de pronto atendimento, tanto no que se refere a realização de uma classificação de risco detalhada e na aplicação do processo de enfermagem. O estudo permitiu elaborar reflexões sobre a importância de se conhecer o perfil da população que procura o serviço, bem como conhecer a essência do raciocínio clínico e as diferentes habilidades, que envolvem a aplicação do processo de enfermagem. / Introduction: The emergency services have been working as entrance door for the system of health, even in cases without gravity, what causes wait lines, over crowed, is long for attendance, the user's dissatisfaction and service overload to the professionals. It is in this context that the quality of the attendance and the implantation of the nursing process as work tool, in spite of important, it has been left in second plan. The objective of this study was to evaluate the nursing diagnosis of the clients assisted in an Unit of Ready Attendance, admitted in the Yellow and Red Rooms, based in Imogene King theory and in Taxonomy of the NANDA-I. Method: It was treated of research of quantitative approach of the descriptive type, with traverse design. It was accomplished in Unit of Ready Attendance of Distrito Federal, with the patients classified as orange red and/or admitted in the Yellow and Red Rooms. The data were collected in the period of November from 2016 to February of 2017, starting from the application of the Nursing Process and structured in Imogene King model. The collection was accomplished by the own author, with 50 clients, that signed the Term of Consent Free and Explained. Results: With relationship to the profile individuals' of the feminine sex discreet predominance was observed (54%). the interviewed customers' age varied between 18 and 91 years, with 50,48 year-old average. In relation to the education, the sample was characterized with low education. As the breed/color had individuals self-declared predominance that brown (44%). In relation to the matrimonial state, 60% were married. On the referring data to the home, 76% were in the urban zone and 62% lived in immobile own. Complete basic sanitation was identified in 64% of the sample. On the source of income, 44% worked and 36% were unemployed. About the pré-existent pathological conditions the more prevalents were the hypertension (40,0%) and the diabetes (30,0%). The sample was also characterized by the polypharmacy. Of the individuals 20% were smoker, 22,0% were drinker and 18,0% only practiced regularly some physical exercise. Only 34,0% of the customers had already passed for at least an attendance, in another moment, in the unit of study. About the risk classification, 94,0% were classified as Orange and 68% were interviewed in the Yellow Room. The predominant complaints were pain in the chest and lack of air with 28,0% each. It were identified 13 differents flow chart in the customers belonging to the study, and the one of larger prevalence were Thoracic pain (18,0%), Indisposition in adult (14,0%), Dyspnea in adult and Abdominal pain in adult (12,0% each). Using the Taxonomy of the NANDA-I was inventoried to the whole 722 nursing diagnosis with average of 14,44 diagnosis for client. It were identified 94 diagnosis, being 54 diagnosis with focus in the problem (real), two diagnosis of promotion of the health and 38 of risk (or vulnerability). Analyzing under the perspective of the three systems mentioned by King it was observed that the found diagnosis contemplated all the proposed systems, and it are divided in a percentile way in personal system with 88,3%, interpersonal system with 9,6% and social system with 2,1%. only nine diagnosis presented superior frequencies to 40%, being them: infection risk (90,0%), deficient knowledge (80,0%), risk of falls (64,0%), sedentary lifestyle (62,0%), prejudiced teething (60,0%), risk of ineffective renal perfusion (48,0%), bleeding risk (46,0%), it fatigues (46,0%) and acute pain (42,0%). Among some factors causal presents in the priority diagnosis stood out: deficient knowledge for inadequacy of information on the treatment or the problem of health; sedentarism for insufficient interest for the physical activity; prejudiced teething for access difficulties the professional dental cares; fatigues due to physiologic limiting conditions; and pain due to biological harmful agent. Conclusion: The results obtained in this study, it will serve as support to the active professionals in units of ready attendance, so much in what it refers the accomplishment of a risk classification detailed and in the application of the nursing process. The study allowed to elaborate reflections on the importance of knowing the profile of the population that seeks the service, as well as to know the essence of the clinical reasoning and the different abilities, that involve the application of the nursing process. / Introducción: Los servicios de la emergencia han estado trabajando como la puerta de la entrada para el sistema de salud, incluso en los casos sin la gravedad, que qué causas esperan las líneas, encima de braveado, es largo para la asistencia, el descontento del usuario y carga excesiva de servicio a los profesionales. Está en este contexto que la calidad de la asistencia y la implantación del proceso de enfermería como la herramienta de trabajo, a pesar de importante, se ha salido en segundo plan. El objetivo de este estudio era evaluar los diagnósticos de enfermería de los clientes ayudados en una Unidad de Asistencia Lista, admitida en los Cuartos Amarillo y Rojo, basado en la de teoría Imogene King y en Taxonomía del NANDA-I. El método: Se trató de investigación de acercamiento cuantitativo del tipo descriptivo, con el plan atravesado. Era cumplido en la Unidad de Asistencia Lista de Distrito Federal, con los pacientes clasificados como la naranja rojo y/o admitió en los Cuartos Amarillo y Rojo. Los datos eran reunido en el periodo de noviembre de 2016 a febrero de 2017, empezando de la aplicación del Proceso de Enfermería y estructuró en modelo de Imogene King. La colección era cumplida por el propio autor, con 50 clientes que firmaron el Término de Consentimiento Libre y Explicó. Los resultados: Con la relación a los individuos del perfil del sexo femenino el predominio discreto se observó (54%). la edad de los clientes entrevistados varió entre 18 y 91 años, con 50,48 promedio año-viejo. Respecto a la educación, la muestra se caracterizó con la educación baja. Cuando los raza/color tenían el ego de los individuos declarar el predominio que el atezado (44%). Respecto al estado matrimonial, 60% estaban casados. En los datos refiriéndose a la casa, 76% estaban en la zona urbana y 62% vivieron en inmóvil propio. Se identificó la higienización básica completa en 64% de la muestra. En la fuente de ingreso, 44% trabajaron y 36% eran desempleado. Sobre las condiciones patológicas pré-existentes el más los prevalentes eran la hipertensión (40,0%) y la diabetes (30,0%). La muestra también se caracterizó por el polifarmacia. De los individuos 20% eran fumadores, 22,0% eran bebedores y sólo 18,0% practicaron algún ejercicio físico regularmente. Sólo 34,0% de los clientes ya habían pasado para por lo menos una asistencia, en otro momento, en la unidad de estudio. Sobre la clasificación de riesgo, 94,0% eran clasificados como la Naranja y 68% se entrevistó en el Cuarto Amarillo. Las quejas predominantes eran el dolor en el pecho y falta de aire con 28,0% cada uno. Se identificaron 13 mapa de flujo diferente en los clientes que pertenecen al estudio, y el uno de predominio más grande era el Dolor torácico (18,0%), Indisposición en el adulto (14,0%), Disnea en el adulto y el Dolor abdominal en el adulto (12,0% cada uno). Usando el Taxonomy del NANDA-I me inventarié a los 722 diagnósticos de la lactancia enteros con el promedio de 14,44 diagnósticos para cliente. Se identificaron 94 diagnósticos, mientras siendo 54 diagnósticos con el enfoque en el problema (real), dos diagnósticos de promoción de la salud y 38 de riesgo (o vulnerabilidad). Analizando bajo la perspectiva de los tres sistemas mencionaron por King que fue observado que los diagnósticos encontrados contemplaron todos los sistemas propuestos, y son dividido de una manera del percentil en el sistema personal con 88,3%, sistema interpersonal con 9,6% y el sistema social con 2,1%. sólo nueve diagnósticos presentaron las frecuencias superiores a 40%, mientras siendo: el riesgo de infección (90,0%), el conocimiento deficiente (80,0%), riesgo de caídas (64,0%), el estilo de vida sedentario (62,0%), el echando los dientes prejuiciado (60,0%), riesgo de perfusión renal ineficaz (48,0%), el riesgo sangrante (46,0%), fatiga (46,0%) y el dolor agudo (42,0%). Entre algunos factores los regalos causales en los diagnósticos de prioridad destacados: el conocimiento deficiente para la insuficiencia de información en el tratamiento o el problema de salud; el sedentarios para el interés insuficiente para la actividad física; perjudicado echando los dientes para las dificultades de acceso los cuidados dentales profesionales; las fatigas debido a condiciones fisiologicas que las limita; y dolor debido al agente dañoso biológico. Conclusión: Los resultados obtuvieron en este estudio, servirán como el apoyo a los profesionales activos en las unidades de asistencia lista, tanto en lo que se refiere al logro de una clasificación de riesgo detallado y en la aplicación del proceso de enfermería. El estudio permitió elaborar las reflexiones en la importancia de saber el perfil de la población que busca el servicio, así como para saber el ser del razonamiento clínico y las habilidades diferentes que involucran la aplicación del proceso de enfermería.

Detection of enzyme activity has gained popularity in molecular imaging because increased activity of enzymes such as urokinase plasminogen activator (uPA) can serve as biomarkers and assist in cancer diagnosis. Chemical exchange saturation transfer (CEST) Magnetic Resonance Imaging (MRI) is a non-invasive technique that can be utilized to detect enzyme activity; however, CEST MRI is not the only technique that can assess enzyme activity. Chapter 1 provides an overview of various imaging modalities that have been used to detect enzyme activity in vivo. Advances made in probe-design are discussed, in addition to advantages and disadvantages of each technique. Chapter 2 focuses on detection of uPA activity in a pancreatic cancer tumor model using a catalyCEST MRI contrast agent. Chapter 2 also discusses the importance of uPA in tumor biology, addresses the synthesis of the contrast agent, and evaluates the results of in vivo detection and ex vivo validation of uPA activity in response to therapy of pancreatic tumor models of Capan-2. The in vivo and ex vivo results showed no significant difference in uPA activity between chemotherapy-treated and non-treated mice. Additionally, no significant difference was observed between before and after chemotherapy-treated groups. Chapter 3 addresses some of the limitations of the study detailed in Chapter 2 and proposes improvements.

Despite recent advances in early detection, in 2017 prostate cancer is estimated to claim over 26,000 lives in the United States alone. Prostate cancer related morbidity and mortality is a result of secondary skeletal metastasis. Therefore, better understanding of the underlying molecular mechanisms of prostate tumor cell migration and subsequent metastasis is vital for improved clinical outcomes. Interestingly, integrin α6, a laminin receptor, is highly expressed in a number of aggressive tumor types including prostate and is associated with increased metastasis and reduced patient survival. Preliminary studies by our group found that α6 integrin undergoes a post-translational modification mediated by the serine protease, uPA and its receptor, uPAR, leading to the cleavage of α6 integrin and production of the tumor specific structural variant integrin α6p. Cleavage of this laminin receptor and production of α6p variant gives rise to an aggressive phenotype, markedly increasing tumor cell migration and invasion. Thus, the work conducted here sought to identify the function and efficacy of these prominent proteins in various aspects of tumor cell migration as well as the factors regulating α6 integrin cleavage. Interestingly, utilization of a co-culture system of prostate tumor cells and macrophages found that a direct and indirect interaction between the two cell populations influenced α6 integrin cleavage. Specifically, prostate tumor cell interactions with macrophages, a known immune cell population that is highly observed in a number of primary tumors, resulted in increased protein levels of uPAR on the surface of prostate tumor cells that led to a significant production of α6p and subsequently increased invasion. Additionally, key downstream effectors of integrin signaling, including FAK, ILK, and actin, were found to regulate production of the tumor specific variant integrin α6p. Depletion of FAK, ILK, or actin, resulted in a significant increase in uPAR protein expression and subsequent α6 integrin cleavage, a regulatory event previously not known of these integrin signaling effector molecules. In addition, silencing of another prominently expressed laminin receptor, integrin α3β1, led to a significant increase in the cohesive collective phenotype exhibited by aggressive prostate tumor cells that was found to be facilitated by α6 integrin cleavage. Depletion of integrin α3β1 resulted in increased surface uPAR expression and increased lateral association with α6 integrin, which resulted in a striking increase in α6p production, a novel finding showing the regulation of one laminin receptor is dependent on the expression of another. Furthermore, the expression of α6 integrin as well as uPAR, was found to be highly expressed in aggressive pancreatic tumor cells. This expression pattern was found to significantly increase in response to the development of drug resistance and increased invasive potential. This finding showed a never before seen efficacy of integrin and uPAR expression in dictating acquired drug resistance in pancreatic tumor cells and demonstrates their potential use as prognostic biomarkers for acquired chemotherapy resistance. Taken together, the work conducted here illustrates the utility in further understanding the role of integrins and their regulation in mediating tumor cell migration and subsequent metastasis in the progression of aggressive epithelial cancers.