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Analyzing and modeling data is a well established research area and a vast variety of different methods have been developed over the last decades. Most of these methods assume fixed positions of data points; only recently uncertainty in data has caught attention as potentially useful source of information. In order to provide a deeper insight into this subject, this thesis concerns itself with the following essential question: Can information on uncertainty of feature values be exploited to improve in silico modeling? For this reason a state-of-art random forest algorithm is developed using Matlab R. In addition, three techniques of handling uncertain numeric features are presented and incorporated in different modified versions of random forests. To test the hypothesis six realworld data sets were provided by AstraZeneca. The data describe biochemical features of chemical compounds, including the results of an Ames test; a widely used technique to determine the mutagenicity of chemical substances. Each of the datasets contains a single uncertain numeric feature, represented as an expected value and an error estimate. Themodified algorithms are then applied on the six data sets in order to obtain classifiers, able to predict the outcome of an Ames test. The hypothesis is tested using a paired t-test and the results reveal that information on uncertainty can indeed improve the performance of in silico models.

<p>Analyzing and modeling data is a well established research area and a vast variety of different methods have been developed over the last decades. Most of these methods assume fixed positions of data points; only recently uncertainty in data has caught attention as potentially useful source of information. In order to provide a deeper insight into this subject, this thesis concerns itself with the following essential question: Can information on uncertainty of feature values be exploited to improve in silico modeling? For this reason a state-of-art random forest algorithm is developed using Matlab R. In addition, three techniques of handling uncertain numeric features are presented and incorporated in different modified versions of random forests. To test the hypothesis six realworld data sets were provided by AstraZeneca. The data describe biochemical features of chemical compounds, including the results of an Ames test; a widely used technique to determine the mutagenicity of chemical substances. Each of the datasets contains a single uncertain numeric feature, represented as an expected value and an error estimate. Themodified algorithms are then applied on the six data sets in order to obtain classifiers, able to predict the outcome of an Ames test. The hypothesis is tested using a paired t-test and the results reveal that information on uncertainty can indeed improve the performance of in silico models.</p>

Thesis (D. Min.)--Midwestern Baptist Theological Seminary, 2008. / Abstract. Includes bibliographical references (leaves 120-125)

Diss.-- Franeker (H. Calsbeek, defendant).

AN ABSTRACT OF THE DISSERTATION OF Cristal M. Hill, for the Doctor of Philosophy degree in Molecular Biology, Microbiology, and Biochemistry, presented on January 22nd 2016, at Southern Illinois University Carbondale. THE METABOLIC EFFECTS OF DIET-INDUCED OBESITY AND GROWTH HORMONE TREATMENT IN LONG-LIVED MICE WITH ALTERED INSULIN AND INSULIN-LIKE GROWTH FACTOR -1 SIGNALING MAJOR PROFESSOR: Dr. Andrzej Bartke It is well established that high calorie diets providing mostly fat and simple carbohydrates as nutrients promote obesity and are associated with metabolic syndromes such as type 2 diabetes and cardiovascular disease. However, the effects of these types of diets in genetically long lived mice remain to be fully elucidated. The effects of high calorie diets have been reported to induce inflammation and alter longevity. However, when viewed in the context of the growth hormone (GH) pathway, these types of diets that have negative impact on IGF-1 and insulin signaling. To examine high calorie diet and GH-treatment effects in long-lived mice, we designed a three part study using hypopituitary Ames dwarf mice that have primary altered endocrine signaling and Pregnancy Associated Plasma Protein-A knockout mice that have normal endocrine signaling. Most importantly, together these studies investigate the detrimental effects of high energy feeding promoting obesity and influencing adipokine profiles that regulate or alter insulin/ IGF-1 signaling that may possibly impair glucose homeostasis in the context of the GH-axis. Longevity and aging are influenced by common intracellular signals of the insulin/insulin-like growth factor (IGF)-1 (IIS) pathway. Abnormally high levels of bioactive IGF-1 increase the development of various cancers and may contribute to metabolic diseases such as insulin resistance. Enhanced availability of IGF-1 is promoted by cleavage of IGF binding proteins (IGFBPs) by proteases, including the pregnancy associated plasma protein-A (PAPPA). In vitro, PAPPA is regulated by pro-inflammatory cytokines (PICs) such as interleukin (IL)-6 and tumor necrosis factor -a (TNF-a). Mice born with deficiency of the Papp-a gene [PAPP-A knockout (KO) mice] live ∼30–40 % longer than their normal littermates and have decreased bioactive IGF-1 on standard diets. In the first study, our objective was to elucidate how the effects of high-fat, high-sucrose diet (HFHS) promote obesity, induce metabolic dysfunction, and alter systemic cytokine levels in PAPP-A KO and normal mice. We show that PAPP-A KO mice fed HFHS diet for 10 weeks were more glucose tolerant and had enhanced insulin sensitivity compared to normal mice fed identical diet. PAPP-A KO mice fed HFHS diet had lower levels of pro-inflammatory cytokines (IL-2, IL-6, and TNF-α) compared to normal mice fed the same diet. Moreover, anti-inflammatory cytokine (IL-4 and adiponectin) levels were higher in PAPP-A KO mice fed HFHS diet compared to normal mice fed HFHS. Circulating PAPP-A levels were elevated in normal mice fed an HFHS diet compared to normal mice fed a standard, low-fat, low-sucrose (LFLS) diet. Indirect calorimetry, at 10 weeks of feeding HFHS diet, showed significantly increased oxygen consumption (VO2) in PAPP-A KO mice fed HFHS diet compared to normal mice fed the same diet. Furthermore, respiratory quotient (RQ) was significantly lower in PAPP-A KO mice fed HFHS diet compared to normal (N) mice fed HFHS diet indicating PAPP-A KO mice fed HFHS diet are able to rely on fat as their primary source of energy more so than normal controls. We conclude that PAPP-A KO mice are resistant to the HFHS diet induction of metabolic dysfunction associated with higher levels of anti-inflammatory cytokines and have a remarkably metabolically flexible phenotype and that some of the effects of HFHS diet in normal animals may be due to increased levels of PAPP-A. We continued our investigations of high calorie diet effects in long-lived endocrine disrupted Ames dwarf mice. Ames dwarf mice are hypopituitary, thus lacking the production of GH. GH stimulates the production of IGF-1; induces insulin resistance, alters inflammatory cytokine levels and can reduce life expectancy in both humans and mice. Disruption of GH signaling by reducing plasma GH levels significantly or deleting GH receptors extends health span and life span in mice as observed in Ames dwarfs. Metabolic stressors such as high-fat diet (HFD) may alter longevity through the GH signaling pathway. Our objective was to investigate the effects of HFD in Ames dwarf and control mice to elucidate the interactions on environmental (diet) and genetic mechanisms that regulate metabolism in aging processes. We show that Ames dwarf mice fed HFD for 12 weeks are sensitive to weight gain and increase in subcutaneous and visceral adiposity, yet are more insulin sensitive and have higher levels of adiponectin compared to control mice fed either standard diet (STD) or HFD. Interleukin 6 levels were lower in Ames dwarf mice fed HFD than control mice fed either STD or HFD. Energy expenditure was higher in Ames dwarf mice fed HFD than control mice fed STD or HFD. Moreover, we show that transplant of epididymal white adipose tissue (eWAT) from Ames dwarf mice fed HFD is able to improve insulin sensitivity in control mice fed the same diet. We conclude that Ames dwarf mice are resistant to the detrimental metabolic effects of HFD and the visceral adipose tissue of Ames dwarf mice can recuse metabolic dysfunction in control mice. In the third study, we investigated the effects of early-life GH-treatment in Ames dwarf mice starting at 1week of age. The focus of this study was to examine the metabolic effects of GH- treatment and HFD feeding during young age, which is the most critical time for biological maturation. In this study, one week old Ames dwarf and control mice were injected with either GH or saline for 6 weeks and fed STD. At 7 weeks of age, test for insulin sensitivity and calorimetric measurements were performed and the animals were subjected to diet switch from STD to HFD for 12 weeks post GH-treatment. With these preliminary data, we focus on the detrimental effects of GH-treatment during development and on the interaction of the effects of GH and diet. We first show that early-life-GH treatment in hypopituitary Ames dwarf mice induces a slight reduction of insulin sensitivity and decreased use of fatty acids as indicated by indirect calorimetry, thus promoting metabolic dysfunction. In addition, we show that the effects of early-life GH-treatment and high fat diet in Ames dwarf mice worsen insulin sensitivity and impair substrate utilization. We will continue to investigate the expression of genes that are associated with metabolism and longevity in these animals. Inhibition of proteases, such as PAPP-A, may be a therapeutic treatment to decrease the activity of biologically active IGF- to induce protection from metabolic dysfunction, including insulin resistance, in humans. Furthermore, it is not likely to inhibit GH/insulin/ IGF-1 signaling in healthy humans at young age, decreasing the activity of the insulin/ IGF-1 pathway at middle age may be beneficial in human therapies in the aims of protecting against metabolic dysfunction. Combined, these studies provide novel information on the interaction of the GH pathway and diets that induce obesity and metabolic dysfunction. Thus, mice with either primarily altered endocrine signaling or deletion of proteases that increase local IGF-1 signaling are protected from the detrimental effects of high calorie diets on metabolic function and energy expenditure.

Orientador: Edson Luis Maistro / Resumo: Cerca de 11% dos medicamentos considerados essenciais pela OMS são derivados de plantas medicinais, daí a importância do desenvolvimento de testes científicos sobre essas plantas, avaliando, além do seu real potencial farmacológico, a genotoxicidade, mutagenicidade, antigenotoxicidade, citotoxicidade, dentre outros, para esclarecer se o uso de tais plantas por seres humanos é seguro. Crataegus oxyacantha é uma planta originalmente encontrada na Europa e, devido à suas potencialidades medicinais, já há muito tempo foi introduzida no continente sulamericano. Pertencente à família Rosaceae, a árvore forma, na primavera, cachos grandes de flores brancas ou rosas de fragrância agradável, que no outono se transformam em pequenos frutos vermelhos. A importância da C. oxyacantha se dá pela presença comprovada de flavonóides, que são conhecidos por sua ação antioxidante. Devido a inexistência na literatura de estudos investigando a toxicidade genética de C. oxyacantha para os seres humanos, o presente estudo foi elaborado visando avaliar se o extrato de frutos desta planta apresenta efeitos citotóxico, genotóxico e clastogênico/aneugênico em leucócitos humanos e células HepG2 em cultura, e mutagênico em cepas de Salmonella typhimurium (teste de Ames). Os resultados da análise de genotoxicidade mostraram que o extrato não apresentou efeitos genotóxicos nas concentrações de 2,5 e 5,0 µg/ml em ambos os tipos celulares analisados, no entanto, em concentrações acima de 10 µg/ml verificou-s... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: About 11% of medicines considered essential by WHO are derived from medicinal plants, being important to develop scientific tests on these plants, evaluating several aspects as its pharmacological potential, genotoxicity, mutagenicity, antigenotoxicity, cytotoxicity, among others, to clarify if the use of such plants is safe for humans. Crataegus oxyacantha is a plant originally found in Europe and due to its medicinal potential, was at long time introduced in the South American continent. Belonging to the Rosaceae family, the tree form in the spring, large bunches of white flowers and pleasant fragrance of roses, which in autumn turn into small red fruits. The importance of C. oxyacantha is attributed to the presence of flavonoids as constituents, which are known for their antioxidant activity. Considering the absence in the literature of studies investigating the genetic toxicity of C. oxyacantha to humans, this study was designed to evaluate if the fruits extract of this plant present a cytotoxic, genotoxic and clastogenic/aneugenic effects in cultured human HepG2 and leukocytes cells, and mutagenicity in Salmonella typhimurium strains (Ames test). The results of genotoxicity analysis evidenced that the extract showed no genotoxic effects at 2.5 and 5.0 ug/ml concentrations, in both cell types tested, however, at concentrations above of 10 ug/ml significant DNA damage was observed. The micronucleus test results showed that the concentrations above of 10 ug/ml also produced... (Complete abstract click electronic access below) / Mestre

Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Martina Klaučová Supervisor: prof. PharmDr. Petr Pávek, Ph.D. Consultant: PharmDr. Ivona Pávková, Ph.D. Diploma thesis title: Ames test in the drug development Background: Thesis objective is the determination of potential genotoxicity of newly developed drugs within primary testing and the introduction of the Ames microfluctuation test which can be used in common laboratory conditions. Methods: I used commercially supplied kit based on the principles of Ames test which detects reverse mutation through colour changes of the samples using bacterial strains S. typhimurium. At first I had to study literary sources and then I could design the procedures of the Ames microfluctuation test, preparation of the chemicals and storage of the strains which are optimal for all laboratories. Results: The drug samples T6445 and T6447 with 30 µM concentration tested by metabolic activation S9 on bacterial strain ST TA 98 show genotoxicity. The sample UOCHB1 with 30 µM concentration tested without activation shows possible genotoxicity on both strains ST TA 98 and ST TA 100. Other samples do not show any toxicity. I used 3 different procedures during the designation of assay. The most suitable version of the...

Ames dwarf (df/df) mice are homozygous for a spontaneous mutation in the prop1 gene due to which there is no development of anterior pituitary cells – somatotrophs, lactotrophs and thyrotrophs, leading to a deficiency of growth hormone (GH), prolactin (PRL) and thyrotropin (TSH). They tend to become obese as they age, but still live longer and healthier lives compared to their wild-type littermates, being very insulin sensitive, showing no signs of diabetes and cancer. These mutant mice also have high circulating levels of anti-inflammatory and antidiabetic adiponectin. Plasma levels of this adipokine usually decrease with an increase in accumulation of visceral fat (VF). We thus believe that VF in df/df mice, developed in the absence of GH signaling, may be functionally different from the same fat depots in normal (N) mice and may be beneficial, rather than detrimental, to the overall health of the animal. We performed surgeries involving removal of VF depots (epididymal and perirenal fat) in both groups of mice and hypothesize that the beneficial effects of visceral fat removal (VFR) will be present exclusively in N mice as VF in df/df mice contributes to enhanced insulin sensitivity by producing decreased levels of pro-inflammatory adipokines like TNF and IL-6. We found that VFR improved insulin sensitivity only in N mice but not in the df/df mice. This intervention led to an upregulation of certain players of the insulin signaling pathway in the skeletal muscle of N mice only, with no alteration in df/df mice. The subcutaneous fat of df/df mice showed a downregulation of these insulin signaling genes upon VFR. Compared to N mice, epididymal fat of df/df mice (sham-operated) had increased gene expression of some of the players involved in insulin signaling and a decrease in transcript levels of TNFa. Ames dwarf mice had decreased levels of IL-6 protein in EF and in circulation. High circulating levels of adiponectin and iv decreased levels of IL-6 in circulation could contribute to the high insulin sensitivity observed in the Ames dwarf mice. Understanding the mechanisms responsible for VF having positive effects on insulin signaling in df/df mice would be important for future treatment of obese diabetic patients.

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