Development of computational and experimental methods for biomass composition and evaluation of its impact in genome-scale models prediction

The use of genome-scale metabolic models has been rapidly increasing in fields such as metabolic engineering. An important part of a metabolic model is the biomass equation since this reaction will ultimately determine the predictive capacity of the model in terms of essentiality and flux distributions. Thus, in order to obtain a reliable metabolic model the biomass precursors and their coefficients must be as precise as possible. Ideally, determination of the biomass composition would be performed experimentally, but when no experimental data are available this is established by approximation to closely related organisms. Computational methods however, can extract some information from the genome such as amino acid and nucleotide compositions. The main objectives of this study were to compare the biomass composition of several organisms and to evaluate how biomass precursor coefficients affected the predictability of several genome-scale metabolic models by comparing predictions with experimental data in literature. For that, the biomass macromolecular composition was experimentally determined and the amino acid composition was both experimentally and computationally estimated for several organisms. Sensitivity analysis studies were also performed with the Escherichia coli iAF1260 metabolic model concerning specific growth rates and flux distributions. The results obtained suggest that the macromolecular composition is conserved among related organisms. Contrasting, experimental data for amino acid composition seem to have no similarities for related organisms. It was also observed that the impact of macromolecular composition on specific growth rates and flux distributions is larger than the impact of amino acid composition, even when data from closely related organisms are used.

Avaliação experimental da utilização de conectores indentados em elementos mistos de aço e betão leve

Dissertação de mestrado integrado em Engenharia Civil

Fluência no betão desde as primeiras idades: um estudo experimental

Dissertação de mestrado integrado em Engenharia Civil

Efeito da berinjela sobre os lípides plasmáticos, a peroxidação lipídica e a reversão da disfunção endotelial na hipercolesterolemia experimental

OBJETIVO: Estudar o efeito do suco da berinjela sobre os lípides plasmáticos, o colesterol tecidual, a peroxidação lipídica das LDL nativas, oxidadas e da parede arterial e o relaxamento dependente do endotélio, em coelhos hipercolesterolêmicos. MÉTODOS: Coelhos foram separados em grupos controle (GC), hipercolesterolêmico (GH) e berinjela (GB), (n=10). Os animais do GC foram alimentados com ração normal, o GH e o GB com ração acrescentada de colesterol (0,5%) e gordura de babaçu (10%) durante 30 dias. Ao GB acrescentou-se suco de berinjela, nos últimos 15 dias do experimento. Os lípides plasmáticos foram medidos através de kits enzimáticos, a peroxidação lipídica pela dosagem do malondialdeído (MDA) e o relaxamento dependente do endotélio, por curvas de concentração efeito pela acetilcolina e nitroprussiato. RESULTADOS: O peso dos animais foi menor no GB em relação ao GC e GH (p<0,05). O colesterol total plasmático, as LDL e os triglicérides que se elevaram no GH, reduziram-se em 19%, 29% e 38%, respectivamente no GB (p<0,05). O colesterol tecidual foi menor no GB (50%) em relação ao GH (p<0,05). Nos animais do GB ocorreu redução significante do teor de MDA nas LDL nativas (56%) e oxidadas (22%), assim como na parede arterial (30%) (p<0,05). O relaxamento máximo dependente do endotélio, , elevou-se significantemente em 28% no GB em relação ao GH (p<0,05). CONCLUSÃO: O suco de berinjela administrado a coelhos hipercolesterolêmicos reduziu significantemente o peso corpóreo, o colesterol total, as LDL plasmáticas, os triglicérides, o colesterol tecidual, a peroxidação lipídica das LDL nativas, oxidadas e da parede arterial, assim como aumentou o relaxamento dependente do endotélio.

Effects of lisinopril on experimental ischemia in rats. Influence of infarct size

OBJECTIVE - Angiotensin-converting enzyme inhibitors (ACEIs) have gained importance in preventing or attenuating the process of ventricular remodeling after myocardial infarction. The significance of infarct size in regard to the response to ACEIs, however, is controversial. This study aimed to analyze the effects of lisinopril on mortality rate, cardiac function, degree of cardiac hypertrophy and fibrosis in rats with different infarct sizes. METHODS - Lisinopril (20 mg/kg/day) dissolved in drinking water was administered to rats immediately after coronary artery occlusion. After being sacrificed, the infarcted animals were divided into two groups: one group of animals with small infarcts (< 40% of the left ventricle) and another group of animals with large infarcts (> 40% of the left ventricle). RESULTS - The mortality rate was 31.7% in treated rats and 47% in the untreated rats. There was no statistical difference between the groups with small and large infarcts in regard to myocardial concentration of hydroxyproline. In small infarcts, the treatment attenuated the heart dysfunction characterized by lower levels of blood pressure and lower values of the first derivative of pressure and of the negative derivative of pressure. The degree of hypertrophy was also attenuated in small infarcts. In regard to large infarcts, no differences between the groups were observed. CONCLUSION - Treatment with the ACEIs had no effect on mortality rate and on the amount of fibrosis. The protective effect of lisinopril on heart function and on the degree of hypertrophy could only be detected in small infarcts

Experimental analysis of Maritime pine and Iroko single shear dowel-type connections

"Available online 21 March 2016"

Effects of losartan on ventricular remodeling in experimental infarction in rats

OBJECTIVE: To evaluate the effects of losartan on ventricular remodeling and on survival after myocardial infarction in rats. METHODS: After surgical occlusion of left coronary artery, 84 surviving male Wistar rats were divided into two groups: LO treated with losartan (20mg/kg/day, n=33) and NT (n=51), without medication. After 3 months, we analyzed mortality; ventricular to body mass ratio (VM /BM); myocardial hydroxyproline concentration (HOP); isovolumetric pressure, +dp/dt, -dp/dt, and diastolic volume/left ventricle mass ratio (VO/LV). RESULTS: Mortality was: LO = 22%, and NT = 47% (p<0.05). Ventricular mass,(VM/BM, mg/g) was 4.14 ± 0.76 and 3.54±0.48, in the NT and LO groups, respectively (p<0.05). HOP (median) was 4.92 upsilong/mg in the LO and 5.54 upsilong/g in the NT group (p>0.05). The V0/LV values (median) were 0.24 mL/g in group LO and 0.31 mL/g in group NT (p<0.05) compared to NT group. There were no differences between the groups for +dp/dt and -dp/dt parameters. CONCLUSION: 1- The use of losartan myocardial infarction causes an attenuation of ventricular remodeling, bringing about an increased survival, an attenuation of ventricular hypertrophy and dilation, and an improvement of the isovolumetric pressure; 2- the treatment does not modify the myocardial collagen concentration.

Effect of Propafenone on the Contractile Activity of Latissimus Dorsi Muscle Isolated in an Organ Chamber: Experimental Study in Rats

OBJECTIVE: To study the effect of propafenone on the contractile function of latissimus dorsi muscle isolated from rats in an organ chamber. METHODS: We studied 20 latissimus dorsi muscles of Wistar rats and divided them into 2 groups: group I (n=10), or control group - we studied the feasibility of muscle contractility; group II (n=10), in which the contralateral muscles were grouped - we analyzed the effect of propafenone on muscle contractility. After building a muscle ring, 8 periods of sequential 2-minute baths were performed, with intervals of preprogrammed electrical stimulation using a pacemaker of 50 stimuli/min. In group II, propafenone, at the concentration of 9.8 µg/mL, was added to the bath in period 2 and withdrawn in period 4. RESULTS: In group I, no significant depression in muscle contraction occurred up to period 5 (p>0.05). In group II, a significant depression occurred in all periods, except between the last 2 periods (p<0.05). Comparing groups I and II only in period 1, which was a standard period for both groups, we found no significant difference (p>0.05). CONCLUSION: Propafenone had a depressing effect on the contractile function of latissimus dorsi muscle isolated from rats and studied in an organ chamber.

A realização do sujeito em complementos infinitivos de verbos causativos e percetivos no português língua de herança

Dissertação de mestrado em Ciências da Linguagem

Development of a water-in-oil-in-water multiple emulsion system integrating biomimetic aqueous-core lipid nanodroplets for protein entity stabilization. Part I: experimental factorial design

Lipid nanoballoons integrating multiple emulsions of the type water-in-oil-in-water enclose, at least in theory, a biomimetic aqueous-core suitable for housing hydrophilic biomolecules such as proteins, peptides and bacteriophage particles. The research effort entertained in this paper reports a full statistical 23x31 factorial design study (three variables at two levels and one variable at three levels) to optimize biomimetic aqueous-core lipid nanoballoons for housing hydrophilic protein entities. The concentrations of protein, lipophilic and hydrophilic emulsifiers, and homogenization speed were set as the four independent variables, whereas the mean particle hydrodynamic size (HS), zeta potential (ZP) and polydispersity index (PI) were set as the dependent variables. The V23x31 factorial design constructed led to optimization of the higher (+1) and lower (-1) levels, with triplicate testing for the central (0) level, thus producing thirty three experiments and leading to selection of the optimized processing parameters as 0.015% (w/w) protein entity, 0.75% (w/w) lipophilic emulsifier (soybean lecithin) and 0.50% (w/w) hydrophilic emulsifier (poloxamer 188). In the present research effort, statistical optimization and production of protein derivatives encompassing full stabilization of their three-dimensional structure, has been attempted via housing said molecular entities within biomimetic aqueous-core lipid nanoballoons integrating a multiple (W/O/W) emulsion.

Experimental validation of a proposed single-phase five-level active rectifier operating with model predictive current control

This paper presents a model predictive current control applied to a proposed single-phase five-level active rectifier (FLAR). This current control strategy uses the discrete-time nature of the active rectifier to define its state in each sampling interval. Although the switching frequency is not constant, this current control strategy allows to follow the reference with low total harmonic distortion (THDF). The implementation of the active rectifier that was used to obtain the experimental results is described in detail along the paper, presenting the circuit topology, the principle of operation, the power theory, and the current control strategy. The experimental results confirm the robustness and good performance (with low current THDF and controlled output voltage) of the proposed single-phase FLAR operating with model predictive current control.

Experimental procedures for the identification of material properties of intervertebal disc - a contribution for the development of biomimetic substitutes for the nucleus pulposus

Tese de Doutoramento em Engenharia Mecânica.

Experimental Model of Gene Transfection in Healthy Canine Myocardium: Perspectives of Gene Therapy for Ischemic Heart Disease

OBJECTIVE: To assess the transfection of the gene that encodes green fluorescent protein (GFP) through direct intramyocardial injection. METHODS: The pREGFP plasmid vector was used. The EGFP gene was inserted downstream from the constitutive promoter of the Rous sarcoma virus. Five male dogs were used (mean weight 13.5 kg), in which 0.5 mL of saline solution (n=1) or 0.5 mL of plasmid solution containing 0.5 µg of pREGFP/dog (n=4) were injected into the myocardium of the left ventricular lateral wall. The dogs were euthanized 1 week later, and cardiac biopsies were obtained. RESULTS: Fluorescence microscopy showed differences between the cells transfected and not transfected with pREGFP plasmid. Mild fluorescence was observed in the cardiac fibers that received saline solution; however, the myocardial cells transfected with pREGFP had overt EGFP expression. CONCLUSION: Transfection with the EGFP gene in healthy canine myocardium was effective. The reproduction of this efficacy using vascular endothelial growth factor (VEGF) instead of EGFP aims at developing gene therapy for ischemic heart disease.

Efeitos da estimulação com corrente elétrica contínua pulsátil sobre as propriedades eletrofisiológicas atriais: estudo experimental da fibrilação atrial em cães

OBJETIVO: Avaliar se a estimulação atrial com corrente elétrica contínua pulsátil induz fibrilação atrial e os seus efeitos sobre as propriedades eletrofisiológicas atriais e as alterações histológicas atriais. MÉTODOS: Foram submetidos à toracotomia lateral direita 22 cães e implantados eletrodos de marcapasso no sulcus terminalis (ST), apêndice atrial direito (ADb) e na região póstero-inferior do átrio esquerdo (AE); um par de eletrodos foi suturado na auriculeta direita para estimulação com bateria alcalina de 9 Volts conectada a uma sistema (LM 555) que transforma a energia contínua linear da bateria em corrente contínua pulsátil, durante 60 min. A biópsia epicárdica atrial foi realizada antes e após a estimulação atrial. RESULTADOS: Não foram observadas diferenças nas durações dos períodos refratários efetivos atriais. Os tempos de condução intra-atrial, interatrial, bem como dos extra-estímulos atriais também prolongaram-se. A duração dos eletrogramas atriais prolongou-se durante ritmo sinusal e estimulação atrial programada; em 68% dos cães a fibrilação atrial foi induzida e sustentou-se. Foram observados edema intersticial e bandas de contração celular no subepicárdio à microscopia óptica, e intensa desorganização miofibrilar e aumento do tamanho das mitocôndrias à microscopia eletrônica. CONCLUSÃO: Esta técnica de estimulação atrial induz fibrilação atrial e provoca modificações atriais que aumentam sua vulnerabilidade para o surgimento de fibrilação atrial.

Remodelação miocárdica após infarto agudo do miocárdio experimental em ratos: efeito do bloqueio do sistema renina angiotensina aldosterona

OBJETIVO: Verificar a ação do lisinopril e do losartan sobre a remodelação miocárdica no infarto experimental em ratos. MÉTODOS: Ratos machos Wistar foram submetidos a infarto e tratados com lisinopril 20 mg/kg/dia (LIS, n=13) ou losartan 20 mg/kg/dia (LOS, n=11), ou mantidos sem tratamento (NT, n=11), por três meses e os resultados comparados com grupo controle (CONT, n=11) de ratos sem infarto. Após a eutanásia, o ventrículo esquerdo foi separado e pesado. Foram medidas a área seccional dos miócitos (AC), fração de colágeno intersticial (CVF) e a hidroxiprolina (HOP) miocárdica. As variáveis foram comparadas pela ANOVA de uma via, para nível de significância de p<0,05. RESULTADOS: O infarto agudo promoveu a hipertrofia do ventrículo esquerdo e os tratamentos com lisinopril e losartam preveniram a hipertrofia quantificada pelo peso do ventrículo esquerdo (LOS=1,06± 0,12g, LIS=0,97±0,18g, NT=1,26±0,17g, CONT=1,02± 0,09g; p<0,05), pelo peso de ventrículo esquerdo corrigido pelo peso corporal VE/PC (LOS=2,37±0,21mg/g, LIS=2,41± 0,38mg/g,NT=2,82±0,37mg/g, CONT=2,27± 0,15mg/g) e pela medida da AC do ventrículo esquerdo (LOS=210±39µ², LIS=217±35µ², NT=256±35µ², CONT= 158±06 µ²; p<0,05). O CVF foi significantemente maior no ventrículo esquerdo do grupo infartado e houve prevenção do aumento com os tratamentos (LOS=1,16±0,4%, LIS=1,27± 0,5%, NT=1,8± 0,4%, CONT=0,7±0,5%). A HOP foi maior no grupo infartado (NT=6,91±2,98mg/g vs. CONT=2,81±1,21mg/g) e não alterou com o tratamento. CONCLUSÃO: A remodelação miocárdica pós-infarto é caracterizada por aumento da massa ventricular remanescente e aumento de colágeno intersticial. O bloqueador da enzima conversora da angiotensina e o antagonista seletivo AT1 da angiotensina II previnem a hipertrofia do miócito e a fibrose intersticial.