A Low-fat Diet Containing Heated Soybean Oil Promotes Hyperglycemia in C57BL/6J Mice Introduction: The metabolic effects of consuming mildly oxidized lipids as compared to highly oxidized lipids are not well documented. Consumption of highly oxidized polyunsaturated fatty acids, even in a low-fat diet, may be a threat to one’s health. Previous studies in our lab have shown that mice consuming soybean oil heated for 3 hours, compared to unheated, gain less body weight, but more fat pad mass, and 3T3-L1 adipocytes treated with soybean oil heated for ≥ 6 hours have abrogated triglyceride accumulation. Another study showed that rats fed highly oxidized oil (heated > 24 hrs), as compared to fresh soybean oil and fish oil, had lower fat pad mass and weight gain but developed glucose intolerance. This suggested that the extent of lipid oxidation determined the subsequent metabolic risk. Objective: Our aim, in the current study, is to investigate if a low-fat diet with soybean oil heated for increasing time points (3h, 6h and 9h) alter fat mass and glucose tolerance. Methods: Six week old, male, C57BL/6J mice were randomly divided into six groups (n=8/group). Three groups were fed a low fat diet with soybean oil heated for 3 (3hr-HO), 6 (6hr-HO), or 9 (9hr-HO) hours for 16 weeks. Another three groups were pair-fed to each of the 3hr-HO, 6hr-HO and 9hr-HO groups with a low-fat diet containing unheated oil (PF-UHO). Food consumption was recorded every 3-4 days, and body weights were recorded weekly. Soy oil in the diets was analyzed for products of oxidation. At 16 weeks, blood glucose levels were measured after a 6 hour fast; fat pad and liver weights were recorded, and blood was collected by cardiac puncture for serum insulin analysis. Results: Final weight gain was not significantly different between all HO groups as a percent of their respective PF groups (p >0.05). The feeding efficiency for 3hr, 6hr, and 9hr-HO groups as a percent of PF was 92.75, 113.02, and 111.28, respectively. Mean weights of all fat pads for HO groups decreased with heating time as a percent of PF, although these differences were not statistically significant. Blood glucose was lowest in the 3hr-HO group and significantly increased from 3hr-HO group to 6hr-HO group (p=0.021) as a percent of PF. Serum insulin levels decreased for the HO groups as heating time increased, although these differences were not statistically significant. Conclusion: Consuming a diet with increasing amounts of oxidized lipids decreased fat pad mass and insulin levels, while increasing fasting glucose levels. This paradoxical relationship between increased glucose in the presence of decreased insulin in c57BL/6J mice could be due to either reduced insulin secretion or increased insulin resistance. Further research in our lab will aim to analyze triglyceride accumulation in the liver and muscle cells of these mice to determine if oxidized lipids promote ectopic fat deposition.

The purpose of the study was to investigate how different types of filtrates (oxidized and unoxidized) would effect on oxygen delignification for softwood pulp, as well as study its impact on yield final-pH, viscosity and kappa number.

The CE-QUAL-W2 model has been selected as a tool for use in water quality management studies of the Occoquan Reservoir. In order for the model to achieve its best possible predictive capability, additional quantitative information on denitrification rates in the reservoir was required. A microcosm operating protocol was developed to obtain such information and also to enhance the understanding of complex nitrate-sediment-phosphorus interactions. The microcosm system developed was a biphasic system, consisting of a single continuous stirred tank reactor (CSTR), or a series of CSTRs containing representative sediment and water samples from the reservoir. The system was configured to simulate the bottom waters in the upper reaches of the reservoir during anoxic conditions. Nitrate concentrations in the microcosm system were monitored, and first-order denitrification-rate constants calculated to be used as an input to the reservoir water quality model CE-QUAL-W2. Other water chemistries were also monitored to investigate the nitrate effects on water quality. From the investigation results, it appears that the first-order denitrification-rate constant of the model should be set at 0.22 day-1 instead of the model default value. Nitrate was also observed to be removed by chemical and/or biologically mediated reduction by reduced forms of manganese. Once the nitrate was depleted, soluble manganese was released from the sediment first, followed by soluble iron. The release of phosphorus was not observed in this study after the depletion of nitrate, but nevertheless, was believed to occur. The absence of the release was attributed to phosphorus adsorption to the Plexiglas reactor walls. / Master of Science

Made available in DSpace on 2015-07-22T20:50:43Z (GMT). No. of bitstreams: 0 Previous issue date: 2008-11-26 / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Objetivos: As lipoproteínas oxidadas e os anticorpos anti-LDL oxidada (anti-LDLox) têm sido detectados no plasma e em lesões ateroscleróticas em humanos. No entanto, o papel destes autoanticorpos na proteção vascular ou na patogênese das síndromes coronarianas agudas (SCA) permanece não elucidado. Nós examinamos a relação entre os títulos de IgG humana anti-LDLox com marcadores de risco para a doença cardiovascular. Métodos: Títulos de autoanticorpos anti-LDLox foram mensurados em indivíduos portadores de hipertensão arterial em estágio 1 (n=94), sem outros fatores de risco, e em indivíduos com síndrome metabólica após recente síndrome coronariana aguda (n=116). Os autoanticorpos contra a LDL oxidada pelo cobre foram avaliados por ELISA. Resultados: pacientes com hipertensão arterial apresentaram menor índice de massa corpórea e circunferência abdominal, maiores níveis de pressão arterial sistólica e diastólica quando comparados aos portadores de SCA (p<0,001). O HDL-C e a Apo A1 foram maiores, enquanto os triglicérides e a Apo B foram menores nos pacientes do grupo hipertensão em estágio 1 (p<0,0001). Os títulos de anticorpos anti-LDLox foram maiores no grupo hipertensão comparados aos do grupo SCA, e os hipertensos do primeiro grupo apresentaram níveis de PCR menores do que indivíduos com SCA (p<0,0001). A análise conjunta de ambos os grupos mostrou, em análise univariada, significante correlação inversa para a PCR (r=-0,284), IMC (r=-0,256), circumferência abdominal (r=-0,368), apo B (r=-0,191) e glicemia (r=-0,303) e correlações positivas entre pressão arterial sistólica e diastólica (r=0,319 e r=0,167, respectivamente), HDL-C e Apo A1 (r=0,224 e r=0,257, respectivamente), com os títulos de anticorpos anti-LDLox (p<0,02). Regressão linear múltipla mostrou que a PCRas, glicemia e circunferência abdominal permaneceram independente e negativamente associados com os títulos de anticorpos anti-LDLox. Conclusões: nossos resultados sugerem que os títulos baixos de anticorpos circulantes anti-LDLox possam estar associados com maior risco cardiovascular. / Objectives: Oxidized lipoproteins and antibodies anti-oxidized LDL (anti-oxLDL) have been detected in human plasma and in atherosclerotic lesions. However, the role of these autoantibodies in the maintenance of health or in the pathogenesis of acute coronary syndromes (ACS) remains unclear. We examined the relationship of human IgG antibodies anti- ox LDL with cardiovascular disease risk markers. Methods: Titers of human anti-oxLDL were measured in hypertensive subjects in stage 1 (n=94) without other risk factors, and in individuals with metabolic syndrome after recent acute coronary syndrome (n=116). Autoantibodies against copper ion oxidized LDL were measured by ELISA. Results: Hipertensive patients presented lower BMI, waist circunference, higher blood pressure levels than those with ACS (p<0.001). HDL-C and Apo A1 were higher, whereas triglycerides and Apo B were lower in those with hypertension stage 1 (p<0.0001). Anti-oxLDL titers were higher in hypertensive patients compared to those with acute coronary syndromes, and hypertensive patients presented lower hs-CRP than those with ACS (p<0.0001). Taken into account both populations, univariate analysis showed small, but significant inverse correlations between the hs-CRP (r=-0.284), BMI (r=-0.256), waist circunference (r=-0.368), apo B (r= -0.191), and blood glucose (r= - 0.303) and positive correlations between systolic and diastolic blood pressure (r=0.319 and r=0.167, respectively), HDL-C and Apo A1 (r=0.224 and r=0.257, respectively), with anti-ox LDL titers (p<0.02). After multiple linear regression, hs-CRP, fasting glycemia and waist circunference remained independently associated with anti-oxLDL. Conclusions: Our results suggest that low titers of circulating anti-oxLDL antibodies may be associated with increased cardiovascular risk. / TEDE / BV UNIFESP: Teses e dissertações

Este trabalho apresenta as propriedades ópticas dos complexos Európio Tetraciclinas (EuTcs) na presença de LDL e de LDL oxidada com potenciais aplicações em análises clínicas. Foram escolhidos quatro elementos da família das Tetraciclinas: Tetraciclina (Tc), Clorotetraciclina (CTc), Metatetraciclina (MTc) e Oxitetraciclina (OTc) para fazerem parte dos complexos com o íon európio. As melhores condições para se formar os complexos eficientemente foram determinadas, através das medidas dos parâmetros ópticos como: absorção, emissão e de tempo de vida. As melhores concentrações de európio nos complexos EuTcs e possíveis influências de íons inorgânicos normalmente presentes no plasma sanguíneo também foram analisadas. As amostras foram preparadas em pH neutro e a luminescência visível do lantanídeo foi detectada após tempo de repouso das amostras de 15 minutos. Os resultados deste trabalho mostraram que as moléculas de LDL e de LDL oxidada apresentaram um importante papel no aumento da intensidade de emissão dos complexos das Tcs. As medidas realizadas com os complexos EuTcs não apresentaram deslocamentos nos comprimentos de onda dos espectros de absorção e de emissão na presença de LDL, o que demonstra a ausência de interação direta entre as moléculas de Tcs e as moléculas de LDL e LDL oxidada. No entanto, o íon európio pode interagir em diferentes sítios das moléculas de tetraciclinas o que diferenciou a intensidade de emissão de cada complexo. Comparando os resultados obtidos entre os complexos de EuTcs, o complexo EuTc foi o que apresentou perspectivas promissoras na quantificação de LDL e LDL oxidada. / This work presents the optical properties of europium complexes - Tetracyclines (EuTcs) in the presence of LDL and oxidized LDL with potential applications in clinical analysis. Four elements were chosen from the Tetracyclines family: Tetracycline (Tc), Chlortetracycline (CTc), Metatetraciclina (MTc) and Oxytetracycline (OTc) to be part of complexes with europium ion. The best conditions to form the complex efficiently were determined through measurements of optical parameters such as absorption, emission and lifetime. The best concentrations of europium complexes in EuTcs and possible influences of inorganic ions normally present in blood plasma were also analyzed. The samples were prepared at neutral pH and the visible luminescence of lanthanide was detected after resting time of the samples of 15 minutes. These results showed that the molecules of LDL and oxidized LDL have an important role in increase of the emission intensity for Tcs complexes . The measurements performed with the complex EuTcs showed no shifts in the wavelengths of the absorption and emission spectra in the presence of LDL, which demonstrates the absence of direct interaction between the molecules of Tcs and the molecules of LDL and oxidized LDL. However, the europium ion can interact at different sites of the tetracyclines molecules which differed the emission intensity of each complex. Comparing the results obtained between the complexes EuTcs, the complex EuTc is the one that presented the promising prospects in the quantification of LDL and oxidized LDL.

Cancer cell esterases are often overexpressed and some have chiral specificities different from those of corresponding normal cells. Carboxylesterases in particular are known to be overexpressed in several cancers. Additionally, cancer cells often exhibit high levels of intrinsic oxidative stress that is required for survival and an aggressive phenotype. We hypothesized that these 2 characteristics of cancer cells could be exploited to aid in the detection and treatment of prostate cancer. We have developed a fluorogenic ester probe that is activated by carboxylesterase to help distinguish tumorigenic cells from nontumorigenic prostate cells. Ester prodrugs have the same activation mechanism and have been thought to be a promising approach in cancer therapy. Prodrugs are inactive drugs that can be selectively activated by a specific enzyme. We have developed a chiral ester prodrug strategy using native polyacrylamide gel electrophoresis (n-PAGE) and proteomic methods to compare and identify the esterase profiles of several tumorigenic and nontumorigenic prostate cell lines. Our results showed that cell lysates from LNCaP, DU 145, and PC3 prostate cancer cell lines exhibit differential esterase activity compared with non-tumorigenic RWPE-1 prostate cell lysates when incubated with α- naphthyl acetate or α-naphthyl N-acetyl-alaninate ester substrates and a diazonium salt. We have identified oxidized protein hydrolase (OPH), a serine esterase/protease that catalyzes the removal of N-acylated residues from proteins, to be differentially expressed between some tumorigenic and nontumorigenic prostate cell lines. OPH was found to have high hydrolytic activity towards the S-isomer of α-naphthyl N-acetylalaninate (S-ANAA) chiral ester. LNCaP lysates incubated with N-acetyl-alanyl-p-nitroanilide, a known OPH substrate, had twofold higher OPH activity compared with RWPE-1 lysates. We have also developed and tested novel glutathione depleting prodrugs modeled after S-ANAA that increase oxidative stress and induced apoptosis in tumorigenic prostate cells with little effect on nontumorigenic RWPE-1 cells. These results suggest that ester molecular beacon probes and ester prodrugs may be effective in identifying and treating prostate cancer tumors that overexpress esterases with little effect on normal prostate cells.

ABSTRACT Background: Polyunsaturated fatty acids (PUFA) are found in nuts and seeds, salad dressings and vegetable oil and are prone to oxidation during storage and food preparation. Evidence supports that consumption of oxidized lipids promotes atherosclerosis and glucose intolerance in animal models. However there is a dearth of evidence with regard to the amount of oxidized lipids consumed and its association with parameters of adiposity and glucose homeostasis in humans. Objective: The objective of this study is to estimate the amount of oxidized lipids in common foods and the oxidized lipid consumption in the US population using the data from National Health and Nutrition Examination Survey (NHANES) 2005-06. The second objective of this study is to investigate if there is an association between consumption of oxidized lipids with markers of adiposity and glucose tolerance. Methods- Foods with possible high oxidized lipid content were selected from the NHANES food frequency questionnaire. Oxidized lipid content /Peroxide Values (PV) of these foods were determined from published values in the literature. Oxidized lipid consumption was stratified into tertiles to determine the relationship between consumption of oxidized lipids and markers of adiposity. Regression analysis was used to explore to the extent to which body fat % and HOMA- IR scores could be attributed to oxidized lipid intake. Results- The estimated mean daily consumption of oxidized lipids was 0.625 meq/kg of fat for the US population. Estimated mean consumption of oxidized lipids was significantly greater in men compared to women, in children compared to adults and among African Americans compared to other races. In both men and women it was observed that the markers of adiposity like body fat%, waist circumference, triceps skinfold decreased significantly with increased consumption of oxidized lipids. However in women (below 18 years) there was a significant increase in HOMA-IR with increased consumption of oxidized lipids. Conclusion- Increased consumption of oxidized lipids is associated with decreased fat mass but increased glucose intolerance in women, but not in men.

Background: Foods rich in polyunsaturated fatty acids (PUFA) are susceptible to oxidation through heating or storage. Oxidized lipids are known to act as ligands for a transcription factor (PPAR-gamma) that affects adipocyte differentiation and insulin sensitivity. Objective: The purpose of this study was to determine the amounts of oxidation products of a variety of PUFA containing foods over time, and to determine whether extracted fats from these foods act as ligands for PPAR-gamma. Method: To study the effect of room-temperature storage on oxidation, 5 foods (walnuts, sunflower seeds, ground flax, fish oil capsules, and infant formula) were purchased and stored at room temperature for 1, 2, and 3 months. To determine oxidation levels in fried foods, French fries and chicken nuggets were used. Fat was extracted from each food and the levels of oxidation products were analyzed by spectrophotometry and kits designed to measure oxidation products. Using a fluorescence polarization-based ligand screening assay kit, fat extracted from foods was analyzed for its binding affinity for PPAR-gamma. Results: Among foods stored at room temperature, the levels of oxidation products did not change significantly with time. Most foods exhibited the highest levels of oxidation at the purchase date. Infant formula and ground flax demonstrated higher levels of oxidation products than did other foods. In preliminary ligand binding assays, extracted fat from French fries showed the greatest binding affinity for PPAR-gamma; a select few other oils showed slight affinity. Discussion: Surprisingly, storage time did not affect oxidation levels. The greatest amount of oxidation may occur during pre-purchase storage conditions. The processing of formula and ground flax may be the cause of the relatively higher oxidation levels in those foods. The binding affinity for PPAR-gamma demonstrated by French fries needs further investigation. Conclusion: Certain oxidized lipids from foods may act as ligands for PPAR-gamma. Further research is required not only to determine which component of these PUFA-containing foods activates PPAR-gamma but also to determine whether that component acts as an agonist or antagonist for PPAR-gamma.

Elevated plasma concentrations of lipoprotein(a) (Lp(a)) are associated with increased risk of atherothrombotic disease. Lp(a) is a unique lipoprotein consisting of a low density lipoprotein-like moiety covalently linked to apolipoprotein(a) (apo(a)), a homologue of the fibrinolytic proenzyme plasminogen. Apo(a) is extremely heterogeneous in size with small isoforms being independently associated with increased cardiovascular risk. Several in vitro and in vivo studies have shown that Lp(a)/apo(a) can inhibit tissue-type plasminogen activator (tPA)-mediated plasminogen activation on fibrin surfaces, although the mechanism of inhibition by apo(a) remains controversial. Essential to fibrin clot lysis are a number of plasmin-dependent positive feedback reactions that enhance the efficiency of plasminogen activation, including the plasmin-mediated conversion of Glu1-plasminogen to Lys78-plasminogen. Additionally, abnormal fibrin clot structures have been associated with both an increased risk of cardiovascular disease and elevated Lp(a) levels. Similarly, oxidized phospholipids have been implicated in the development of cardiovascular disease, and are not only preferentially carried by Lp(a) in the plasma but have also been shown to covalently-modify both apo(a) and plasminogen. In this thesis, we built upon the understanding of the role of apo(a) in plasminogen activation on the fibrin/degraded fibrin surface by determining that: (i) apo(a) inhibits plasmin-mediated Glu1-plasminogen to Lys78-plasminogen conversion and identifying the critical domains in apo(a) responsible for this effect, (ii) apo(a) isoform size does not affect either the inhibition of tPA-mediated plasminogen activation or the inhibition of plasmin-mediated Glu1-plasminogen to Lys78-plasminogen conversion, (iii) apo(a) modifies fibrin clot structure to form more dense clots with thinner fibers and reduced permeability, modifications that enhance the ability of apo(a) to inhibit tPA-mediated plasminogen activation and (iv) the phosphorus content of apo(a) affects its ability to inhibit tPA-mediated plasminogen activation and the phosphorus content of plasminogen affects its ability to be activated by tPA. By understanding these individual reactions, each of which has the potential to affect the broader fibrin clot lysis process, we have expanded our understanding of the overall effect of Lp(a)/apo(a) in the inhibition of plasminogen activation on the fibrin/degraded fibrin surface and thus broadened our understanding of how Lp(a)/apo(a) may mediate the inhibition of thrombolysis in vivo. / Thesis (Ph.D, Biochemistry) -- Queen's University, 2011-12-14 08:26:54.99

Atherosclerotic cardiovascular disease is the leading cause of death of adults in North America. Diabetes is a classical risk factor for atherosclerotic cardiovascular disease. Plasminogen activator inhibitor-1 (PAI-1) is the major physiological inhibitor of fibrinolysis. Elevated levels of PAI-1, oxidized low-density lipoprotein (oxLDL) and glycated LDL (glyLDL) were detected in patients with diabetes. Increased oxidative stress is associated with diabetic cardiovascular complications. Previous studies in our laboratory demonstrated that oxLDL or glyLDL increased the production of PAI-1 or reactive oxygen species (ROS) in vascular endothelial cells (EC). This study was undertaken to investigate transmembrane signaling mechanisms involved in oxLDL or glyLDL-induced upregulation of PAI-1 in cultured vascular EC. Further, we examined the mechanism for oxLDL or glyLDL-induced oxidative stress in EC. The results of the present studies demonstrated novel transmembrane signaling pathway for oxLDL-induced PAI-1 production in vascular EC. We demonstrated that lectin-like oxLDL receptor-1, H-Ras, a small G-protein and Raf-1/ERK-1/2 mediate oxLDL-induced PAI-1 expression in cultured EC. GlyLDL may activate EC via a distinct transmembrane signaling pathway. The results of the present study demonstrated that receptor for advanced glycation end products, NADPH oxidase and H-Ras/Raf-1 are implicated in the upregulation of heat shock factor-1 or PAI-1 in vascular EC under diabetes-associated metabolic stress. We investigated the effects of oxLDL or glyLDL on mitochondrial function in EC. Treatment with oxLDL or glyLDL significantly impaired the activities of electron transport chain (ETC) enzymes and also increased mitochondria-associated ROS in EC. The findings suggest that oxLDL or glyLDL attenuated activity of ETC and increased ROS generation in EC, which potentially contributes to oxidative stress in vasculature. In conclusion, diabetes-associated lipoproteins may upregulate stress response mediators and PAI-1 production via distinct transmembrane signaling pathways. OxLDL or glyLDL may increase ROS production via NOX activation and the impairment of mitochondrial ETC enzyme activity in EC. The understanding and identification of the regulatory mechanisms involved in diabetes-associated lipoprotein-induced signaling may help pharmacological design for the management of diabetic cardiovascular complications.