Resumo: As aderências pulmonares em cães são uma sequela comum, após intervenção cirúrgica, prejudicando eventuais reintervenções. Neste estudo, pretendeu-se verificar, em cães, a eficiência da solução de cloreto de sódio 0,9%, adicionada ou não com dexametasona, na prevenção de aderências após toracotomia intercostal, Para isso, foram utilizados 15 cães, separados em três grupos de cinco, denominados A, B e C e submetidos a toracotomia no quinto espaço intercostal esquerdo. Nos cães do grupo A, foi feita somente a toracotomia e a toracorrafia; no grupo B foram realizadas a toracotomia, a toracorrafia e a injeção na cavidade pleural de 10ml de solução de cloreto de sódio a 0,9% associada com 1mg kg-1 de dexametasona. No grupo C, além da toracotomia e toracorrafia foram injetados na cavidade pleural 10ml kg-1 de solução de cloreto de sódio a 0,9%. Após 15 dias da toracotomia com os três tratamentos foi efetuada a toracoscopia transdiafragmática para se determinar a presença e o escore de aderências entre o pulmão e a parede costal. Os resultados demonstraram presença de aderências na maioria dos animais do grupo A e reduzida a nenhuma aderência nos demais grupos. Na avaliação estatística, foi aplicado o teste quiquadrado, com nível de significância de 5% (P≤0,05). O uso de solução de cloreto de sódio a 0,9% adicionada ou não com dexametasona no espaço pleural reduz ou evita aderências pulmonares após a toracotomia intercostal / Abstract: Pulmonary adhesions in dogs are a common sequela after surgical intervention, undermining any interventions. This study aimed at determining, in dogs, the efficacy of sodium chloride solution 0.9% with or without dexamethasone in order to prevent adhesions after intercostal thoracotomy. Fifteen dogs were separated into three groups of five animals, A, B and C and underwent thoracotomy in the fifth left intercostal space. In the dogs of Group A were performed only a thoracotomy and thoracorraphy; in the dogs of group B, were performed a thoracotomy, thoracorraphy and injection into the pleural cavity of 10ml of isotonic sodium chloride and 1mg kg-1 of dexamethasone; In the dogs of group C, were performed the thoracotomy thoracorraphy and injected into the pleural cavity 10ml kg-1 of isotonic sodium chloride. After 15 days of thoracotomy, was performed transdiaphragmatic thoracocospy to determine the presence and score of adhesions between the lung and chest wall. The results demonstrated the presence of adhesions in the majority of group A and reduced or no adhesions in the other groups. For statistical evaluation, we applied the chi-square test with significance level of 5% (P≤0.05). The sodium chloride solution 0.9% with or without dexamethasone into the pleural space prevented or reduced lung adhesions after intercostal thoracotomy / Orientador: José Antônio Marques / Coorientador: Alceu Gaspar Raiser / Banca: Silvana Martinez Baraldi Artoni / Banca: Richard da Rocha Filgueiras / Mestre

A montagem e desmontagem das adesões focais (AF) desempenham um papel fundamental em diversos processos celulares, incluindo migração celular e sobrevivência. Resultados prévios do nosso laboratório mostram que fibroblastos nulos ou silenciados para miosina-Va sofrem um atraso na desmontagem das adesões, sugerindo um papel para a miosinaVa neste processo. Neste trabalho, visamos analisar a dinâmica de montagem das AF em fibroblastos murinos imortalizados NIH3T3, utilizando sondas fluorescentes para visualização de componentes de adesão focal. A formação das AF foi analisada após estímulo por soro de células quiescentes, o que leva a intensa polimerização de actina, reorganização do citoesqueleto e montagem das AF. A cinética de montagem das AF foi observada em ensaios ao longo do tempo, de células fixadas em 0, 5, 15, 30, 120 minutos após estímulo, e marcadas para miosina-Va fosforilada (p-miosina-Va, S1650), FAK fosforilada (p-FAK, Y397), vinculina, dinamina-2, integrina-?1, faloidina, Ki67 e DAPI. Os nossos resultados mostraram um aumento de fluorescência de p-miosina-Va por todo o citoplasma após a estímulo com soro, e revelaram que a p-miosina-Va co-localiza com pFAK nas AF logo após o estímulo, essa localização da p-miosina-Va nas AF diminui ao passar do tempo e retorna após 120 minutos. Isto é consistente com os resultados anteriores de um papel da miosina-Va na dinâmica das AF. Também é possível perceber uma maior concentração de p-miosina-Va e dinamina-2 na região perinuclear, 5 minutos após estímulo, e o espalhamento de ambas as proteínas pelo citoplasma com o passar do tempo. Demonstramos, por Western blotting, que o estímulo por soro não causa alteração na quantidade total de miosina-Va em nenhum dos tempos analisados em relação à condição de quiescência, mas induz, após 5 e 15 minutos, um aumento apreciável de p-miosina-Va, que sofre queda e variações nos tempos posteriores. Para nosso conhecimento, esta é a primeira demonstração de que a fosforilação da miosina-Va aumenta em resposta ao soro e estamos investigando se este evento está ligado à dinâmica das adesões focais em fibroblastos / The assembly and disassembly of focal adhesions (FA) play a critical role in several cellular process, including cell migration and survival. Previous work from our laboratory showed that fibroblasts without myosin-Va show a delay in focal adhesion disassembly, suggesting a role for myosin-Va in this process. In this work, we aim at imaging the dynamics of focal adhesion disassembly and reassembly in cells, with fluorescent probes for visualization of focal adhesion components. Here, we used murine NIH3T3 fibroblasts to analyze FA formation after serum stimulation of quiescent cells, which leads to intense polymerization of actin and reorganization of the cytoskeleton and FA assembly. The kinetics of FA assembly was observed in a time-course assay of cells fixed at 0, 5, 15, 30 and 120 min after serum stimulation, and stained for phosphorylated myosin-Va (p-myosin-Va, S1650), phosphorylated FAK (p-FAK, Y397), vinculin, phalloidin and DAPI. Our results showed an increase of pmyosin-Va staining throughout the cytoplasm upon serum stimulation, and revealed that pmyosin-Va does not colocalize with FAK in FA at early time points. However, colocalization is observed after 30 to 120 min. This is consistent with previous results of a role for myosin-Va in FA disassembly. It is also possible to observe a higher concentration of p-myosin-Va and dynamin-2 in the perinuclear region 5 minutes after stimulation, and the spreading of both proteins in the cytoplasm over time. We demonstrate by Western blotting that serum stimulation does not cause change in total amount of myosin-Va, in any of the times analyzed in relation to the quiescent condition, but induces, after 5 and 15 minutes, an appreciable increase of pmyosin-Va suffering drop and variations in the later times. To our knowledge, this is the first demonstration that phosphorylation of myosin-Va increases in response to serum and we are investigating whether this event is connected to the dynamics of focal adhesions in fibroblasts

Orientador: Anselmo Eduardo Diniz / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia Mecanica / Made available in DSpace on 2018-08-13T03:28:51Z (GMT). No. of bitstreams: 1 ArroyoOsorio_JoseManuel_D.pdf: 25100736 bytes, checksum: 163b1aa900bc2b44578e310b2fccbe2b (MD5) Previous issue date: 2009 / Resumo: A usinagem é um dos processos de fabricação fundamentais da indústria contemporânea sendo o metal duro recoberto o material mais utilizado na fabricação das ferramentas de corte. A manufatura deste tipo de ferramentas inclui o processamento da superfície do substrato, previamente à deposição do recobrimento, com o objetivo de obter uma elevada resistência de aderência na interface substrato-recobrimento. O método mais difundido com este propósito e o jateamento do substrato com micropartículas de alumina que, embora amplamente utilizado e efetivo, gera poluição, risco à saúde do operador, consome muitos recursos de tempo e mão de obra e apresenta dificuldade no controle da rugosidade. Por outro lado a texturização a laser é uma tecnologia de engenharia de superfície baseada num laser pulsado que modifica a superfície do material através de fusão, evaporação, sublimação e subsequente solidificação. Nesta tese foi explorada a efetividade da texturização a laser como método alternativo ao jateamento na preparação pré-recobrimento da superfície do substrato. Este processo não apresenta poluição, permite o processamento seletivo de áreas especificas da ferramenta e, sendo automatizável, potencialmente utiliza menos recursos de tempo e mão de obra. As ferramentas selecionadas para experimentar foram duas classes de metal duro com recobrimento composto de TiCN+Al2O3+TiN depositado pelo processo MT-CVD, e, na preparação pré-recobrimento da superfície dos substratos experimentais foi utilizado um laser CuHBr pulsado. De acordo com a dinâmica do processo laser utilizado, é possível controlar a rugosidade do substrato e produzir uma grande diversidade de topografias e estruturas de superfície através da variação da intensidade e/ou da quantidade de pulsos do laser. Também foi verificado que o processamento com laser produz sempre transformações do WC, presente no substrato do metal duro, em novas fases não estequiométricas com menor teor de C. Por tentativa e erro, foram determinados dois conjuntos de parâmetros laser que produziram a melhor resistência de aderência avaliada através de ensaios de indentação: 02 pulsos, 410 MW·cm-2 e 64 pulsos, 239 MW. cm-2. Em ensaios de corte, tipo fresamento de topo a seco de aço molde P20 com ferramentas toroidais, foi comparado o desempenho de arestas microjateadas (comerciais) contra arestas texturizadas a laser (experimentais) encontrando uma vida média igual para ambos os tipos de arestas. Na análise detalhada dos mecanismos de desgaste foram encontradas em todas as arestas evidências de difusão, attrition, abrasão, deformação, delaminação, trincas e um mecanismo secundário, chamado desgaste por microfusão do material da aresta. Não houve diferenças notáveis entre as arestas comerciais e as experimentais que fossem atribuíveis ao processo de preparação da superfície do substrato para a deposição do recobrimento / Abstract: Machining is one of the fundamental manufacturing processes of contemporary industry and coated cemented carbide is the most used cutting tool material. Manufacturing of this kind of tools includes the substrate surface processing previously to coating deposition in order to achieve high adhesion resistance in the substrate-coating interface. The most spread out method with this intention is the substrate blasting with alumina micro-particles, which, although so widely used and effective, generates pollution, operator health risks, consumes considerable time and labor resources and presents roughness control difficulty. In the other hand, laser texturing is a surface engineering technology based on a pulsed laser which modifies the material surface through melting, evaporation, sublimation and subsequent solidification. This thesis aims to explore the laser texturing effectiveness as an alternative method for substrate surface pre-coating preparation. This process does not generate pollution, allows selective tool areas for processing and, being automated, it potentially requires less time and labor resources. The tools selected for the experiments were two kinds of coated cemented carbide with a composed TiCN+Al2O3+TiN MT-CVD coating and for the experimental substrate surface pre-coating preparation, a pulsed CuHBr laser was used. With the laser process dynamics used, it is possible to control the substrate roughness and to produce a great diversity of surface topographies and structures by adjusting the intensity and/or the amount of laser pulses. Also, it was verified that the laser processing always produces WC (present in the substrate) transformations in new non- stoichiometric phases with less C content. By trial and error method, two laser parameters sets were determined which produced the best adhesion resistance as evaluated through indentation tests: 02 pulses, 410 MW·cm-2 and 64 pulses, 239 MW·cm-2. In dry face milling cutting tests of mould P20 steel with toroidal tools, it was compared the performance of micro-blasted tools (commercial) against laser textured (experimental) ones finding an equal average life for both tool types. Detailed wear mechanisms analysis showed evidences in all the cutting edges of diffusion, attrition, abrasion, deformation, delamination, cracks and a secondary mechanism, called micro-melting wear of the material of the edge. There were no remarkable wear differences between the commercial edges and the experimental ones which could be attributable to the substrate surface pre-coating preparation process / Doutorado / Materiais e Processos de Fabricação / Doutor em Engenharia Mecânica

Objetivo: Avaliar a eficiência da Triancinolona Acetonida como agente modulador da resposta inflamatória e cicatricial em coelhos submetidos a cirurgia de músculo extra-ocular. Método: Foi realizado estudo prospectivo, mascarado, em dois estádios. No primeiro estádio, dez coelhos foram submetidos a retrocesso do músculo reto superior em ambos os olhos. Aplicouse, em um deles, 0,15 cc de Triancinolona Acetonida (40mg/cc) nos tecidos circunjacentes ao local de reinserção muscular e, como controle, 0,15cc de solução de cloreto de sódio a 0,9% no local equivalente no olho contra-lateral. Quinze dias após, cinco coelhos foram submetidos a exenteração das órbitas e os restantes dos animais tiveram o mesmo procedimento realizado após trinta dias. O material do sítio de reinserção muscular foi avaliado por meio de análise histopatológica qualitativa e quantitativa (morfometria). No segundo estádio, com incrementação da agressão cirúrgica, dezesseis coelhos foram submetidos aos mesmos procedimentos com exenteração das órbitas após quinze dias, e posterior análise histopatológica dos tecidos. Resultados: Houve efeito inibitório sobre a intensidade da resposta inflamatória nos olhos tratados em comparação com os olhos controle. Conclusão: Nas condições de realização do presente estudo o uso per-operatório da triancinolona acetonida foi efetivo no controle da resposta inflamatória em olhos de coelhos submetidos a cirurgia de músculo extra-ocular. / Purpose: To evaluate the efficiency of triamcinolone acetonide (TRI) in limiting the postoperative inflammatory response and scarring following strabismus surgery. Methods: A prospective, two-stage, masked, controlled trial was conducted. In the first stage, the inflammatory response at the extraocular reattachment site was analyzed following superior rectus recession in ten rabbits. One eye had 0,15 cc of triamcinolone acetonide (40mg/cc) applied around the new insertion site and, similarly, 0,15 cc of isotonic saline solution (0,9%) was applied to the fellow eye following the same procedure, thus serving as a control. Fifteen days later, orbital exenteration was performed in five rabbits and the remaining five were exenterated thirty days later. The reattachment site tissues were submitted to qualitative and quantitative histological examinations. In the second stage 16 rabbits were submitted to amplified surgical trauma, after which the aforementioned steps were also carried out. Granuloma total area at the extraocular muscle reattachment sites of control and treated eyes were compared. Results: There was an inhibitory effect of TRI on the inflammatory response of treated eyes as compared to control eyes. Conclusions: TRI was effective in controlling the postoperative inflammatory response in rabbit eyes submitted to traumatic recession of the superior rectus muscle.

Orientador: Orlando Petrucci Junior / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-11T21:17:28Z (GMT). No. of bitstreams: 1 Oliveira_PedroPauloMartinsde_D.pdf: 3207291 bytes, checksum: 643cd7870725684beed7998d60351274 (MD5) Previous issue date: 2008 / Resumo: Introdução: Reoperações representam cerca de 20% das cirurgias cardíacas, quase na totalidade com esternotomia, onde há a formação de aderências entre o coração, esterno e estruturas adjacentes. Lesões das câmaras cardíacas e grandes vasos pela serra esternal ou na dissecção dessas estruturas resultam em aumento da morbimortalidade e do sangramento perioperatório. Vários autores propuseram o uso de substitutos pericárdicos biológicos e sintéticos na tentativa de diminuir o risco de acidentes nas reoperações, porém sem resultados consistentes em longo prazo. A membrana de poli (álcool vinílico) - PVAl reticulado formam um hidrogel bastante estudado como biomaterial, com boa biocompatibilidade e características favoráveis ao seu emprego como substituto pericárdico. Objetivo: Caracterizar a membrana de PVAl quanto à capacidade de absorção de água, calcificação e citotoxidade e estudar o comportamento biológico da mesma como substituto pericárdico. Metodologia: Foram utilizadas amostras da membrana de PVAl reticulada por irradiação e realizados ensaios de citotoxicidade em culturas de células VERO, da capacidade de absorção de água e de calcificação após o implante in vivo. Ratos da Raça Wistar foram divididos em quatro grupos: Grupo controle - pericardiotomia. Grupo Talco - colocação de talco sobre o epicárdio. Grupo PVAl - colocada membrana de PVAl circundando o coração. Grupo PVAl + Talco - colocado talco sobre o epicárdio e a membrana de PVAl circundando o coração. Após oito semanas foi realizada análise macroscópica e histológica dos corações. Avaliação estatística foi realizada com análise de variância (ANOVA) e teste de Dunnett com significância p<0,05. Resultados: A membrana de PVAl não apresentou citotoxicidade, sua capacidade de absorção de água foi de 42,4 ± 0,89% e mostrou valor médio de 0,00422± 0,00256% de cálcio da massa total do material analisado. Na análise macroscópica observou-se maior aderência no grupo Talco. Na análise histológica o grupo PVAl + Talco apresentou maior espessura epicárdica. Os grupos T e PVAl + Talco apresentaram maior número de células inflamatórias. Conclusão: A membrana não é citotóxica, apresentou boa capacidade de hidratação, a absorção de cálcio foi desprezível, não induziu formação de aderências pericárdicas, não provocou aumento da espessura epicárdica e não induziu aumento de migração de células de resposta inflamatória para o epicárdio, mostrando-se interessante para a aplicação desejada. / Abstract: Background: Cardiac surgery reoperations represent around 20% of all surgical procedures. The main incision is sternotomy and after the first operation there are adherences joining the heart, sternum and neighboring structures. Cardiac chambers and great vessels lesions caused by sternal saw increase morbidity and mortality as well as perioperatory bleeding. Several authors had tried pericardial replacement with biological or synthetic materials in order to decrease risks at reoperations, however with no significant results on long term. Polyvinyl alcohol (PVAl) is a well-known hydrogel, with good biocompatibility and favorable properties as a pericardium replacement. Objective: Describe the biological PVAl behavior as a pericardial replacement. Methodology: PVAl samples were reticulated by radiation. Cytotoxicity direct and indirect tests with VERO cells were performed. We tested absorption water capability and in vivo calcification. Wistar rats were divided in four groups: Control - pericardium abrasion; Talc - talc insertion surrounding the heart; PVAl membrane - PVAl surrounding the heart; PVAl + talc - talc and PVAl membrane insertion surrounding the heart. All animals were kept for 8 weeks and euthanized for study. Macroscopic and microscopic analyses were performed. Statistical analyses were performed with ANOVA and Dunnett post test. Results: The PVAl membrane showed no cytotoxicity. The water absorption capability was 42,4 ± 0,89%. The calcification test showed only 0.00422± 0.00256% of calcium in the total mass of analyzed material. Macroscopic analysis showed higher adherences in the talc group. Histological analysis showed higher epicardium thickness in the PVAl + talc group, higher inflammatory cells in the talcum and PVAl + talc groups. Conclusion: The PVAl membrane hasn't cytotoxicity. It has good water absorption capability and calcification was insubstantial. The membrane showed neither adherences formation nor inflammatory response ...Note: The complete abstract is available with the full electronic digital thesis or dissertations. / Doutorado / Pesquisa Experimental / Doutor em Cirurgia

Post surgical adhesions (PSAs) are an inevitable outcome of surgery and their presence leads to pathogeneses and significant economic impact. The studies within this thesis utilised standard and reproducible abrasion models, in rabbits, pigs and rats, to investigate the formation and maturation of PSAs with strict quantitative analyses. These studies have shown that the development of PSAs is a series of complex, multi-factorial processes. PSA development can be classified into two stages: (i) PSA modelling occurring up to/including 16 hours post injury characterised by the inflammatory response and fibrin deposition and maturation; and (ii) PSA remodelling occurring from 16 hours onwards and characterised by tissue repair, collagen deposition and maturation and chemical mediation by TGF-P. Treatment with VivostatTM System Derived (novel) Fibrin Sealant significantly reduced the formation of PSAs with mean PSA reduction of 80% for the rabbit uterine horn abrasion model, from 3 separate studies; 83% for the pig stomach/colon/caecum abrasion model, from 2 separate studies; 80% for the rat caecum abrasion model. This is significantly better than other fibrin sealants investigated in this thesis. PSA prevention with novel fibrin sealant demonstrated a similar pattern to PSA development, with two stages of development evident: (i) tissue generation modelling occurring up to/including 16 hours post injury characterised by the inflammatory response and fibrin deposition and maturation; and (ii) tissue generation remodelling occurring from 16 hours onwards and characterised by tissue repair, collagen deposition and maturation and chemical mediation by TGF-P. However the extent and subsequent time taken for these changes to occur was significantly reduced. The prevention of PSAs and alterations of wound healing by novel fibrin sealant is most probably due to the sealant acting as a haemostat, as well as a physical barrier. Thus preventing fibrinous and subsequent fibrous PSA formation.

Diagnostiquées à partir de douleurs pelviennes, de dysfonctionnements menstruels ou dans le cadre d’un bilan d’infertilité, les adhérences intra-utérines sont des accolements fibreux des faces internes de la cavité utérine, qui peuvent obstruer partiellement ou totalement la cavité utérine. Tout acte traumatique dans la cavité utérine est considéré comme pourvoyeurs d’adhérences post-opératoires. Pour prévenir l’apparition de ces adhérences intra-utérines, nous avons développé un dispositif médical anti-adhérentiel, résorbable et autodéployable. Pour cela, nous avons synthétisé une famille de copolymères innovants constitués de poly (acide lactique) – poly (éthylène oxyde) – poly (acide lactique) avec des propriétés de gonflement adaptés à l’application. L’évaluation biologique in vitro des copolymères a permis de démontrer l’absence de cytotoxicité et le caractère anti-adhérentiel des biomatériaux en contact avec des cellules endométriales humaines. Les études in vivo ont permis de démontrer l’efficacité du copolymère sélectionné comme barrière physique résorbable permettant la non-adhérence de tissus lésés. L’implantation du copolymère n’entraîne pas de réactions biologiques majeures dans les cornes utérines et n’altère pas la fonction de reproduction de l’animal. De plus, l’étude du déploiement du dispositif dans des utérus issus d’hystérectomies chez des patientes nous a permis de confirmer que le dispositif médical développé est adapté à la morphologie utérine. / The symptoms of intrauterine adhesions are pelvic pain, menstrual abnormalities and infertility. Intrauterine adhesions result in the fibrous adherence of opposing uterine walls, which produce partial or complete obliteration in the uterine cavity. Trauma to a gravid uterine cavity is known to be the main cause of adhesions formation. In order to prevent postsurgical adhesion formation, a new anti-adhesive and resorbable medical device was developed to maintain separated uterine walls after surgical trauma. Anti-adhesive barrier was obtained from polylactic acid – polyethylene oxyde – polylactic acid triblock copolymers presenting specific swelling properties. In vitro biocompatibility and anti-adhesive effects of triblock copolymers were demonstrated using human endometrial cells. In vivo adhesion models have allowed us to confirm the anti-adhesive efficiency and the accuracy of the degradation time of a copolymer. This copolymer does not cause major biological responses after implantation in uterine horns and does not alter reproductive functions. A deployment study of the medical device in uterus of patients from hysterectomy has allowed us to demonstrate that our medical device is adapted to the uterine morphology.

Vascular smooth muscle cells (VSMC) constitute an important component of blood vessels and are primarily responsible for vessel contraction. In vascular disorders such as hypertension and atherosclerosis as well as pregnancy and exercise, VSMC demonstrate increased capacity to proliferate and migrate, resulting in vascular remodelling. The actin cytoskeleton is an important component of vascular contractility and is also essential for proliferation and migration of VSMC. Vasoactive agonists such as Endothelin-1 (ET-1) and Noradrenaline (NA), have been shown to mediate VSMC contraction through changes in actin cytoskeleton and focal adhesion (FA) remodelling, and have also been reported to cause VSMC migration in the appropriate setting. The aim of this study was to investigate the signalling mechanisms responsible for FA dependent actin cytoskeleton remodelling of VSMC in response to ET-1 and NA, with a special focus on Hydrogen peroxide-inducible clone 5 (Hic-5). The latter is a FA protein shown to regulate actin cytoskeleton dynamics in small arteries in response to Noradrenaline (NA) and the response of VSMC to arterial injury and abdominal aortic aneurysm. We have shown that Src-dependent tyrosine phosphorylation of Hic-5 regulated its subcellular localisation in mouse embryonic fibroblasts and VSMC, but was not responsible for the effects of ET-1 and NA on actin filament remodelling or Hic-5 redistribution in VSMC. ET-1 stimulation caused an increase in Hic-5 localisation at FAs concurrent with an increase in the density of actin filaments, whereas NA stimulation caused a decrease in Hic-5 localisation at FAs in VSMC concurrent with actin filament redistribution at the cell cortex. Hic-5 was the FA protein that demonstrated the most dramatic changes in subcellular localisation in response to ET-1 and NA, when compared to paxillin (Hic-5 homologue) or vinculin (classical FA marker). NA-mediated changes in Hic-5 localisation and actin filament distribution were more pronounced compared to ET-1-mediated changes. Further investigation into the NA-induced changes suggested that actin filament disassembly preceded Hic-5 relocalisation from FAs to the cytosol. These results show that vasoactive peptides cause Hic-5 relocalisation and actin filament rearrangement in VSMCs in an agonist-dependent manner. Given that VSMC FA remodelling and actin cytoskeleton reorganisation occur during contraction and arterial remodelling, our data identify Hic-5 as a key regulator of these processes in response to NA and ET-1. Furthermore, these data have implications in agonist- specific VSM function such as migration and contraction.

Cardiovascular disease is the leading cause of human death worldwide. Currently, the prevalence of cardiovascular disease and health care costs associated with its onset continue to increase in both developed and developing societies. Concordant with the need to improve preventative measures is the imperative to develop more effective and efficient remedies for incident cardiovascular pathologies. Increased aortic stiffness with aging has recently emerged as an early, independent, and consistent physiological predictor of cardiovascular disease and represents an attractive target for possible therapeutic options. The success of any biomedical strategy in this regard is incumbent upon comprehension of biological processes and mechanical properties attributable to constituent components within the aortic wall. This dissertation tested the hypothesis that aging-induced changes to smooth muscle maintenance of biomechanical homeostasis within the aorta lead to undesirable increases in stiffness, correlative with increased risk of negative cardiovascular outcomes. Conventionally, mechanical studies and models have identified extracellular matrix as the primary determinant of changes in stiffness, but new research presented here shows that this may not be true. In viable ex vivo preparations of aortic tissue, roughly half of the maximal elastic modulus results from alpha-agonist activation of smooth muscle cells. Investigation of the biochemical interactions that characterize this effect revealed a link between aging and decreased expression of Src, a kinase involved in numerous signaling pathways governing cellular growth and survival, as well as defective regulation of focal adhesions between the smooth muscle cells and extracellular matrix. These findings were integrated into a model of aortic contractility and stiffness that establishes an aging-impaired regulatory complex comprising focal adhesions and non-muscle actin cytoskeleton in vascular smooth muscle cells. A better understanding of the mechanisms underlying this model may motivate the design of potential therapeutics, deliverable to previously overlooked target sites within aortic smooth muscle, and ultimately novel treatments for aging-induced cardiovascular disease. / 2017-10-27T00:00:00Z

One of the characteristics that distinguishes living things from non-living things is motility. On the cellular level, the motility or non-motility of different types of cells can be life building, life-saving or life-threatening. A thorough study of cell motion is needed to help understand the underlying mechanisms that enhance or prohibit cell motion. We introduce a discrete centroid model of cell motion in the context of a generalized random walk. We find an approximation for the theoretical mean square displacement (MSD) that uses a subset of the state space to estimate the MSD for the entire space. We give some intuition as to why this is an unexpectedly good estimate. A lower and upper bound for the MSD is also given. We extend the centroid model to an ODE model and use it to analyze the distribution of focal adhesion (FA) lifetimes gathered from experimental data. We found that in all but one case a unimodal, non-symmetric gamma distribution is a good match for the experimental data. We use a detach-rate function in the ODE model to determine how long a FA will persist before it detaches. A detach-rate function that is dependent on both force and time produces distributions with a best fit gamma curve that closely matches the data. Using the data gathered from the matching simulations, we calculate both the cell speed and mean FA lifetime and compare them. Where available, we also compare this relationship to that of the experimental data and find that the simulation reasonably matches it in most cases. In both the simulations and experimental data, the cell speed and mean FA lifetime are related, with longer mean lifetimes being indicative of slower speeds. We suspect that one of the main predictors of cell speed for migrating cells is the distribution of the FA lifetimes.