Orientadores: Ângela Maria Moraes, José Alberto Fracassi da Silva / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia Química / Made available in DSpace on 2018-08-19T03:16:00Z (GMT). No. of bitstreams: 1 RammazzinaFilho_WalterAnibal_M.pdf: 10957404 bytes, checksum: d23a9aea438343039da493db09603556 (MD5) Previous issue date: 2011 / Resumo: O uso de implantes em cirurgias de correção da parede abdominal é freqüentemente necessário. Telas de poli(propileno) podem ser empregadas com sucesso para essa finalidade, possuindo boa aceitação tecidual e baixo custo. O uso deste tipo de biomaterial pode, porém, ocasionar a aderência indesejada entre tecidos e/ou entre órgãos como o fígado e os intestinos e a tela, resultando em dores abdominais, obstrução intestinal e infertilidade. Neste contexto, o objetivo deste trabalho foi o de desenvolver uma estratégia de recobrimento de telas de poli(propileno) enfocando a deposição de soluções de quitosana de massa molar baixa e média e de polietilenoglicol (PEG) de massa molar igual a 1000 Da por electrospinning. Para fins de comparação, telas de poli(propileno) foram alternativamente recobertas por imersão com diferentes soluções combinando quitosana e PEG. Nos estudos de recobrimento via electrospinning, foram avaliadas as variáveis tipo de solvente para a dissolução da quitosana e do PEG, vazão de solução de recobrimento, proporção entre quitosana e PEG, diferença de potencial usada durante a deposição e distância entre a tela e o jato de injeção. As telas recobertas através da imersão em soluções de quitosana a 1% (tanto com a de baixa massa molar quanto a com massa molar média) e em soluções de quitosana misturada ao PEG também com concentração de sólidos total de 1% apresentaram-se satisfatórias para fins de barreira física em cirurgias de hérnia, considerando-se o aspecto, a uniformidade, os valores médios de espessura (de 1140 e 990 mm, respectivamente), a perda de massa em fluido corpóreo simulado (5,1 e 8,9%, respectivamente) e a capacidade de absorção do mesmo fluido (0,76 e 0,59 g/g, respectivamente). Já o recobrimento por electrospinning resultou no depósito de fibras emaranhadas na superfície das telas, que apresentaram, então espessuras finais variando entre 581 e 612 mm, perdas de massa entre 4,9 e 9,2% em fluido corpóreo simulado, capacidade de absorção de 0,17 a 0,36 g/g em fluido corpóreo simulado e diâmetros de fibras de 20,9 a 92,2 mm. O recobrimento via electrospinning resultou em biomateriais menos espessos e com menos massa associada, com bom potencial de uso na aplicação pretendida / Abstract: The use of implants in surgical correction of the abdominal wall is frequently required. Polypropylene meshes can be successfully employed for this purpose, having good tissue acceptance and low cost. The use of this biomaterial, however, can lead to undesired adhesion between tissues and/or between organs such as the liver and intestines and the mesh, resulting in abdominal pain, bowel obstruction and infertility. In this context, the objective of this work was to develop different coatings of polypropylene meshes, focusing on the deposition of solutions of chitosan of low and medium molecular weight and polyethylene glycol (PEG) of molecular weight equal to 1000 Da by electrospinning. For comparison, polypropylene meshes were alternatively coated by immersion in different solutions combining chitosan and PEG. In the studies of coating via electrospinning, the effect of the variables type of solvent for the dissolution of chitosan and PEG, coating solution flow rate, chitosan to PEG mass ratio, the potential difference used during the deposition and the distance between the mesh and the injection jet were evaluated. Meshes coated by immersion in chitosan (both with low and medium molar mass) and PEG solutions with total solids concentration of 1% were satisfactory for purposes of physical barrier in hernia surgery, considering aspect, uniformity, mean thickness (1140 and 990 mm, respectively), mass loss (5.1 and 8.9%, respectively) and the capacity of fluid absorption (0.76 and 0.59 g/g, respectively). Coating by electrospinning resulted in the deposition of entangled fibers on the surface of the meshes, which had final thickness ranging between 581 and 612 mm, losses of mass between 4.9 and 9.2% in simulated body fluid, absorption capacity of 0.17 to 0.36 g/g in the same fluid and fiber diameters from 20.9 to 92.2 mm. Coating the meshes via electrospinning resulted in thinner biomaterials and with less associated mass, with good potential for use in the intended application / Mestrado / Desenvolvimento de Processos Biotecnologicos / Mestre em Engenharia Química

O presente trabalho teve como objetivo avaliar a emissão de metano por cordeiros alimentados em diferentes pastagens tropicais: Panicum maximum cv. IZ-5 (capim aruana) e Cajanus cajan cv. anão (feijão guandu), e suas relações com a composição química destas pastagens. Os trabalhos foram conduzidos na Estação Experimental Agronômica da UFRGS, nos anos de 2015 e 2016. O delineamento experimental foi de blocos completos casualizados com três repetições. O capítulo II relata os resultados da emissão de metano por cordeiros pastejando nos seguintes tratamentos: 1) ARU – somente capim aruana; 2) FG - somente feijão Guandu; 3) AFG – meio piquete com capim aruana, e a outra metade com feijão Guandu. Para avaliação de metano foi utilizado a técnica com marcador SF6 . No experimento do capítulo II, utilizou-se parcela subdividida, sendo a subparcela dois animais testes que receberam polietileno glicol (PEG) para quelar o tanino presente no feijão guandu, e dois que receberam água como controle. As emissões de metano por consumo de matéria seca (EMICONMSDIA) não apresentaram diferença entre os tratamentos, mesmo considerando as dosificações com polietileno glicol (PEG) ou água (P=0,9235). O tratamento FG (P=0,036) apresentou mais altos níveis de taninos condensados do que os demais tratamentos Apesar de diferenças nas suas características químicas, a emissão de metano de cordeiros é semelhante entre pastagens tropicais de capim aruana (Panicum maximum cv. IZ-5), feijão Guandu (Cajanus cajan cv. Anão. Os taninos condensados presentes na forrageira feijão guandu, nas condições experimentais deste trabalho, não demonstraram potencial para contribuir na redução da emissão de metano por cordeiros. No capítulo III, os tratamentos, manejo dos animais e delineamento experimental foram semelhantes ao capítulo II. Os resultados demonstraram que não há uma relação direta entre as características bioquímicas do alimento e a emissão de metano. Essa desassociação entre a emissão de metano e a composição química da pastagem foi constatada tanto quando relacionou-se a emissão por animal (EMI), quanto quando relacionou-se com a emissão por quilograma de matéria seca ingerida (EMICONMSDIA). Conclui-se, então, que não é possível relacionar a emissão de metano de cordeiros com as características químicas das plantas forrageiras tropicais perenes, capim aruana e feijão Guandu. / The researches carried out in the present work had the objective of evaluating the emission of methane by lambs fed on different tropical pastures: Panicum maximum cv. IZ-5 (Aruana grass) and Cajanus cajan cv. dwarf (pigeon pea), and their relationships with the chemical composition of these pastures. The works were conducted at the Experimental Agronomic Station of UFRGS, between January and April 2016, Chapter II; and from February to April 2015, Chapter III. The experimental design was of randomized complete blocks with three replicates. The blocking was carried out with the objective of eliminating variations of slope of the terrain (flat area, sloping at the top and sloping slope). Chapter II reports the results of methane emission by grazing lambs in the following treatments: 1) ARU - only aruana grass; 2) FG - only Guandu beans; 3) AFG - half picket with aruana grass, and the other half with Guandu beans. For methane evaluation, the SF6 marker technique was used. In the experiment of chapter II, a subdivided plot was used, the subplot being two test animals that received polyethylene glycol (PEG) to chelate the tannin present in the pigeon pea, and two that received water as control. Emissions of methane by dry matter intake (EMICONMSDIA) did not show any difference between treatments, even considering the dosages with polyethylene glycol (PEG) or water (P = 0.9235) FG treatment (P = 0.036) presented higher levels of condensed tannins than the other treatments. Despite differences in their chemical characteristics, the methane emission of lambs is similar between tropical pastures of Aruana grass (Panicum maximum cv. IZ-5), Guandu beans (Cajanus cajan cv. Dwarf), and when these forages are offered at will , in two lanes in the picket. The condensed tannins present in the forage pigeon pea, under the experimental conditions of this work, did not demonstrate the potential to contribute in the reduction of the methane emission by lambs. In Chapter III, treatments, animal management and experimental design were similar to Chapter II. The relationships between the chemical composition of pastures of nutritional influence showed to be quite variable in the methane emission response. The results showed that there is no direct relationship between the biochemical characteristics of the food and the emission of methane. This dissociation between the methane emission and the chemical composition of the pasture was verified both when the emission per animal (EMI) was related and when it was related to the emission per kilogram of ingested dry matter (EMICONMSDIA). It is concluded that it is not possible to relate the emission of methane with the chemical characteristics of perennial tropical forage plants Panicum maximum cv. IZ-5, and Cajanus cajan cv. Dwarf.

京都大学 / 0048 / 新制・論文博士 / 博士(農学) / 乙第12821号 / 論農博第2794号 / 新制||農||1025(附属図書館) / 学位論文||H26||N4816(農学部図書室) / 31308 / (主査)教授 河田 照雄, 教授 保川 清, 准教授 橋本 渉 / 学位規則第4条第2項該当 / Doctor of Agricultural Science / Kyoto University / DGAM

The oncogene c-MYC has guanine-rich and complementary cytosine-rich sequences in its P1 promoter region. The P1 promoter is responsible for over 90% of the c-MYC expression. Downregulation of c-MYC expression represents a novel therapeutic approach to more than 50% of all cancers. A stable i-motif formed by the c-MYC C-rich sequence would be an attractive target for cancer treatment. We have previously shown that c-MYC promoter sequences can form stable i-motifs in acidic solution (pH 4.5-5.5). The question is whether c-MYC promoter sequence i-motif will be stable at physiological pH. In this work, we have investigated the stability of mutant c-MYC i-motif in solutions having pH values from 4 to 7 and containing co-solutes or molecular crowding agents. The crowded nuclear environment was modeled by the addition of polyethylene glycol (PEG, having molecular weights from 200 to 12000 g/mol) at concentrations of 10% to 40% w/w. Circular dichroism spectroscopy (CD) and differential scanning calorimetry (DSC) were used to establish the presence and stability of c-MYC i-motifs in buffer solutions having pH values of 4 to 7. The results of these studies are: 1) the addition of up to 20% w/w glycerol does not increase i-motif stability, 2) the addition of 30% PEG results in an increase in i-motif stability to pH values as high as 6.7, 3) i-motif stability is increased with increased PEG concentration and increased PEG molecular weight, and 4) the effects of PEG size and concentration are not linear, with larger PEGs forming DNA/PEG complexes, which destabilize the i-motif. In summary, we have shown that the c-MYC i-motif can exist as a stable structure at pH as high as 6.7 in a crowded environment. Molecular crowding, largely an excluded volume effect, drives the formation of the more compact i-motif, even at higher pH values where the cytosine imino-nitrogen is deprotonated and neutral C-C pairs can form only two H-bonds. Based on this research, it seems possible that a stable c-MYC promoter sequence i-motif could form at physiological pH and would be a reasonable drug target for new cancer therapies.

In this thesis, the successful fabrication of 3D-printed microfluidic devices will be discussed. Fabrication is performed with a low-cost commercially available stereolithographic 3D printer utilizing a custom PEGDA resin formulation tailored for low non-specific protein adsorption based on my colleagues' work [Rogers et al., Anal. Chem. 83, 6418 (2011)]. Horizontal microfluidic channels with designed rectangular cross sectional dimensions as small as 300 um wide and 150 um tall are printed with 100% yield, as are cylindrical vertical microfluidic channels with 300 um designed (334 um actual) diameters. Moreover, two different resins developed by our group are utilized in the process of 3D-printing which is the novel aspect about this thesis since other groups have not done research on this aspect of 3D-printing.

Wearable technology offers the promise of decentralized and personalized healthcare, which can both alleviate current burdens on medical resources, and also help individuals to be more informed about their health. The heterogeneity of disease phenotypes necessitates adaptations to both diagnosing and surveilling disease, but to ensure user adoption and behavioral change, there needs to be a convenient way to amass such health information continuously. This can be in part accomplished by the development of continuously monitoring, compact wearable medical sensors and analytics technology that provide updates on analyte and biosignal measurements at regular intervals in situ. This dissertation investigates methods for collecting and analyzing information from wearable devices with these principles in mind. In Aim 1, we developed new methods for analysis of cardiovascular biosignals. Current methods of estimating left ventricular mass index (LVMI, a strong risk factor for cardiac outcomes), rely on the analysis of echocardiographic signals. Though still the gold standard, echocardiography can typically only be performed in the clinic, making it inconvenient to obtain frequent measurements of LVMI. Frequent measurements can be useful for monitoring cardiac risk, particularly for high-risk individuals, so we investigated the feasibility of predicting LVMI using a deep learning-based approach through ambulatory blood pressure readings, a one-time laboratory test and demographic information. We find that adding blood pressure waveform information in conjunction with multitask learning improved prediction errors (compared to baseline linear regression and neural network models), pointing to its potential as a clinical tool. Using transfer learning, we developed a model that does not require waveform data, but achieved similar prediction accuracies as methods that do require such data – opening the door to use cases that eliminate the need for wearing a blood pressure cuff continuously during the measurement period. Overall, such a technique has the potential to provide information to individuals who are at high risk of cardiac outcomes both inside and outside the clinic. In Aims 2 and 3, we developed a minimally invasive hydrogel patch for continuous monitoring of calcium, as proof-of-concept for wearable measurement of a wide variety of analytes typically assayed in the lab – a technology that can facilitate treatment and management of many prevalent diseases. Specifically, in Aim 2, we engineered a DNA polyacrylamide hydrogel microneedle array that sensed physiologically relevant calcium levels, for potential use by individuals who have hypoparathyroidism, a condition in which blood calcium levels are low and calcium supplements are needed. A negative mold was made using a CNC mill, the positive mold was cast in silicone, and the aptamer along with acrylamide and bis-acrylamide was seeded into the silicone mold. The DNA hydrogel was then fabricated using a simple UV curing protocol. The optimized DNA hydrogel was specific to calcium, used simple fabrication methods and had a fast, reversible signal response. Finally, in Aim 3, we developed the DNA hydrogel sensor into a wearable, integrated system with real-time fluorescence monitoring for testing in vivo. The microneedle array needed to be hydrated for the DNA aptamer to function, but polyacrylamide was too weak in its hydrated state to effectively pierce through skin epidermis. We demonstrated a method for strengthening our hydrogel system with polyethylene glycol diacrylate (PEGDA), while maintaining an optically translucent gel for detection purposes. We conducted piercing studies with a skin phantom on different microneedle array sizes and shapes, and determined that a 3x3 array of beveled microneedles required the least amount of force to pierce through a skin phantom. A custom complementary metal-oxide semiconductor (CMOS) system was developed to capture real-time fluorescence signals from the microneedle array, which correlated to calcium levels in vitro. This setup was then validated in a rat study. In this dissertation, we demonstrated methods for monitoring human biosignals using signal processing techniques, material innovations and integrated sensing platforms. While a work in progress, this dissertation is a step towards realizing the goal of decentralized, connected health for earlier detection and better management of disease.

<p>I put department up there as Biomedical Engineering. The full title should be: School of Biomedical Engineering.</p> / <p>The work presented in this thesis was focused on the surface modification of biomaterials with combinations of polyethylene glycol (PEG) and bioactive molecules (protein anticoagulants) for improved blood compatibility. Since the fate of biomaterials in contact with blood depends significantly on plasma protein-surface interactions, the objective of this work was to reduce non-specific protein adsorption using PEG and to promote specific protein interactions that could inhibit clot formation using protein anticoagulants as modifiers.</p> <p>Two anticoagulant molecules were used in this work: hirudin, a specific inhibitor of thrombin and corn trypsin inhibitor (CTI), a specific inhibitor of clotting factor XIIa. Gold, used as a model substrate, was modified with PEG and anticoagulant molecules using two methods referred to as sequential and direct. In the sequential method PEG was first immobilized on the surface and then the bioactive molecule was attached (conjugated) to the PEG. In the direct method, a PEG-bioactive molecule conjugate was first formed and then immobilized on the surface. Surfaces were characterized by contact angle, ellipsometry and x-ray photoelectron spectroscopy (XPS). Uptake of the bioactive molecules was measured by radiolabeling. Biointeraction studies included plasma protein adsorption, bioactivity assays using chromogenic substrates and clotting time assays. For PEG-hirudin and PEG-CTI surfaces (both direct and sequential) the protein resistance was similar to that of the PEG-alone surfaces. Despite having a lower density of bioactive molecule (both hirudin and CTI), the sequential surfaces showed superior bioactivity compared to the direct ones.</p> <p>To determine the optimal ratio of free PEG and bioactive molecule-PEG conjugate on the surface (best combination of protein resistance and bioactivity), PEG-CTI was immobilized on gold substrate with varying ratio of conjugated to free PEG using both direct and sequential methods. As the ratio increased, protein resistance was maintained while specific interactions (bioactivity) increased. The optimal composition appeared to be where all PEG molecules are conjugated to a CTI molecule.</p> <p>In the final part of this project, PEG and CTI were immobilized on polyurethane as a material with applicability to medical device construction. A sequential method was developed for this substrate. Comparison of the PEG-CTI surface with PEG only or CTI only surfaces indicated that the combination of PEG-CTI was effective both in reducing non-specific protein adsorption and promoting the specific interactions of CTI with its target plasma protein, factor XIIa. In fact, the presence of PEG improved CTI interactions with FXIIa compared with CTI only surfaces. Thus, sequential attachment of PEG and CTI may be effective for modifying polyurethane surfaces used in blood-contacting medical devices.</p> / Doctor of Philosophy (PhD)

Fouling properties of materials such as polyethylene glycol (PEG) have been extensively studied over the past decades. Traditionally, the factors believed to result in protein adsorption resistance have included i) steric exclusion arising from the compression of longer chains and ii) grafting density contribution which may provide shielding from the underlying material. Recent studies have suggested that PEG interaction with water may also play a role in its ability to resist protein adsorption suggesting that steric exclusion may not be the only mechanism occurring during PEG/protein interactions. Star-shaped PEG polymers have been utilized in protein adsorption studies due to their high PEG segment concentration, which allows to increase the PEG chain grafting density compared to that achieved with linear PEG chains. Most studies that have investigated the interactions of tethered linear and star-shaped PEG layers with proteins have considered linear PEG molecules with molecular weights several orders of magnitude smaller than those considered for star-shaped PEG molecules (i.e. 10 000 g/mol vs. 200 000 g/mol, respectively). Additionally, the star-shaped PEG molecules which have been considered in the literature had up to ~70 arms and were therefore modeled by hard-sphere like structures and low chain densities near the surface due to steric hindrance. This resulted in some difficulties to achieve grafted PEG chain overlap for star molecules. Here, triethoxysilane end-functionalized linear PEG molecules have been synthesized and utilized to form star-shaped PEG derivatives based on ethoxy hydrolysis and condensation reactions. This resulted in PEG stars with up to ~4 arms, which were found to result in grafted star-shaped PEG chains with significant chain overlap. Linear PEG derivatives were synthesized so that their molecular weight would match the overall molecular weight of the star-shaped PEG molecules. These 2 PEG molecular architectures were covalently self-assembled to hydroxylated silicon wafers and the thickness, grafting density, and conformation of these films were studied. The adsorption of human albumin (serum protein) on linear and star-shaped PEG films was compared to that obtained on control samples, i.e. uncoated silicon wafers. Both film architectures were found to significantly lower albumin adsorption. / Ph. D.

Affinity biosensors are crucial in clinical settings for their ability to provide rapid, accurate and sensitive detection of analytes, enabling early diagnosis and monitoring of diseases. These biosensors operate on the principle of high-affinity binding between recognition receptors and the target analyte to cause a physicochemical change which is converted by a transducer into a measured signals to obtain the analyte concentration. Current commercial affinity biosensors, utilizing optical, electrochemical and mass-based transduction methods, offer high sensitivity and specificity of analyte measurements for clinical applications, but face limitations such as high costs and bulkiness for optical biosensors, difficulty in detecting small molecules for mass-based biosensors, and complexity and label degradation issues for electrochemical biosensors. 2D nanomaterial-based aptameric nanosensors, which utilize 2D nanomaterials as transduction materials and aptamers as receptors to specifically recognize target analytes, hold the potential to address these limitations of traditional commercial biosensors. However, the nanosenors still face challenges for practical applications in clinical settings, such as nonspecific binding in physiological media, the necessity for individual sensor calibration, and limitations in current 2D nanomaterial properties and performance degradation over time. This thesis focuses on developing 2D-nanomaterial based aptameric FET nanosensors to enhance their adaptability toward practical clinical applications. We first present the optimization of surface modification of aptameric graphene nanosensors for measurement of biomarkers in undiluted physiological media. In these sensors, graphene surfaces are coated with a polyethylene glycol (PEG) nanolayer to minimize nonspecific adsorption of matrix molecules. We perform a systematic study of the aptamer and PEG attachment schemes and parameters, including the impact of the serial versus parallel PEG and aptamer attachment scheme, PEG molecular weight and surface density, and aptamer surface density on the sensor behavior, and then use the understanding from this parametric study to identify optimal surface modification of the nanosensor to enable sensitive and specific biomarker measurements in undiluted physiological media. We then present a calibration-free method that exploits kinetic measurements to enable the calibration-free operation of aptameric graphene nanosensors. In our kinetically based calibration-free method, a time constant estimated based on the aptamer-analyte kinetic binding process is used to determine the analyte concentration. The time constant depends on the to-be-measured analyte concentration and reaction rate constants 𝑘_on and 𝑘_off, respectively, which is unaffected by variations in measurement conditions and sensor properties, thereby allowing devices functionalized with the same aptamer to be used without individual calibration once one particular device is calibrated, and enabling accurate analyte concentration determination independent of variations in parameters. Next, we present an affinity nanosensor that uses Ni₃(HITP)₂ metal-organic framework (MOF) as the conducting channel which is functionalized with an aptamer for specific biomarker recognition. Binding between the aptamer and the target biomarker induces a change in the carrier density in the MOF and resulting in measurable changes in FET characteristics for determination of the biomarker concentration. 𝘐𝘯-𝘴𝘪𝘵𝘶 synthesis of the MOF enhances the adhesion between the MOF film and electrodes to reduce noise during sensor operations, while also allowing devices to be potentially mass-produceable at reduced cost. Thanks to the porous structure of the MOF, the aptamer surface density on the MOF is optimized to enhance sensitivity for biomarker measurements. We finally present the preliminary development of aptameric graphene nanosensors for accurate and stable extended-time measurement of analyte. We perform a systematic study of the stability of the nanosensor including the graphene, aptamers and aptamer attachment, to identify the cause of performance degradation, and use the understanding to develop a differential nanosensor functionalized the aptamer through covalent reaction and implement graphene compensation method to achieve the accurate and stable extended-time analyte measurement.

The problem was to determine the effectiveness of using polyethylene glycol 1000 in the treatment of green wood for the purpose of forming projects made of wood veneer and of simple design for use in junior high or high school woodworking classes. The purpose of this study was to seek answers to the following questions. 1. Is polyethylene glycol 1000 an effective stabilizing agent for green wood veneer that can be used in school woodworking classes? 2. Can green wood veneer treated with polyethylene glycol 1000 be bent to form simple woodwork projects? 3. Can green wood veneer treated with polyethylene glycol 1000 be successfully used in junior high and high school woodworking classes? 4. What length of treatment time is best for green wood veneer that is to be used to form simple bent wood projects? 5. Is one-fourth inch thickness suitable for green wood veneer that is to be treated with polyethylene glycol 1000 and used to form simple bent wood projects?