Application of 2,3-Naphthalenediamine in Labeling Natural Carbohydrates for Capillary Electrophoresis

Neutral and acidic monosaccharide components in <em>Ganoderma lucidum</em> polysaccharide are readily labeled with 2,3-naphthalenediamine, and the resulting saccharide-naphthimidazole (NAIM) derivatives are quantified by capillary electrophoresis (CE) in borate buffer. Using sulfated-α-cyclodextrin as the chiral selector, enantiomers of monosaccharide-NAIMs are resolved on CE in phosphate buffer, allowing a simultaneous determination of the absolute configuration and sugar composition in the mucilage polysaccharide of a medicinal herb<em> Dendrobium</em> <em>huoshanense</em>. Together with the specific enzymatic reactions of various glycoside hydrolases on the NAIM derivatives of glycans, the structures of natural glycans can be deduced from the digestion products identified by CE analysis. Though heparin dissachrides could be successfully derived with the NAIM-labeling method, the heparin derivatives with the same degree of sulfation could not be separated by CE

Structural Characterization and Antioxidative Activity of Low-Molecular-Weights Beta-1,3-Glucan from the Residue of Extracted Ganoderma lucidum Fruiting Bodies

The major cell wall constituent of Ganoderma lucidum (G. lucidum) is β-1,3-glucan. This study examined the polysaccharide from the residues of alkaline-extracted fruiting bodies using high-performance anion-exchange chromatography (HPAEC), and it employed nuclear magnetic resonance (NMR) and mass spectrometry (MS) to confirm the structures. We have successfully isolated low-molecular-weight β-1,3-glucan (LMG), in high yields, from the waste residue of extracted fruiting bodies of G. lucidum. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay evaluated the capability of LMG to suppress H2O2-induced cell death in RAW264.7 cells, identifying that LMG protected cells from H2O2-induced damage. LMG treatment decreased H2O2-induced intracellular reactive oxygen species (ROS) production. LMG also influenced sphingomyelinase (SMase) activity, stimulated by cell death to induce ceramide formation, and then increase cell ROS production. Estimation of the activities of neutral and acid SMases in vitro showed that LMG suppressed the activities of both neutral and acid SMases in a concentration-dependent manner. These results suggest that LMG, a water-soluble β-1,3-glucan recycled from extracted residue of G. lucidum, possesses antioxidant capability against H2O2-induced cell death by attenuating intracellular ROS and inhibiting SMase activity

比較邏輯迴歸模式與類神經網路模 式對內科加護病房存活率之預測

在醫療費用支出緊縮的政策下，隨著醫療技術的發展與人口老化的雙重壓力下，將可 預見重症醫療照護對有限醫療資源將造成更大的壓力。因此本研究的目的係比較邏輯 迴歸與類神經網路二種模式，對內科加護病房病人存活率之預測能力，提供一更倫理 與客觀的存活率預測系統，以進一步促使內科加護病房資源能更有效率之營運。此二 個模式使用於2002年1月至2004年1月期間住進台灣某醫學中心內科加護病房1,496位 病人的APACHE-II(Acute Physiology and Chronic Health Evaluation-II)及GCS( Glasgow Coma Scale)分數來進行存活率之預測。研究結果顯示類神經網路模式相較 於邏輯迴歸模式在存活者(86. 7%,n=361)與整體病患(74.7%,n=498)之預測能力均較 佳。 Under the policy of restraint in medical expenditure and with the dual pressures of medical technology development and population aging, the critical care services will exert even greater pressure on the limited medical resources. Therefore, the objective of this study is to compare the abilities of two models, the Logistic Regression Model and the Neural Network Model, to predict the survival of critical care patients, in order to provide a more ethical and objective survival prediction system, as well as to promote more effective management of the resources of the medical intensive care unit (MICU). The two models use the Acute Physiology and Chronic Health Evaluation-II (APACHE-II ) and Glasgow Coma Scale (GCS) scores of 1,496 patients stayed who in the MICU of a Taiwan medical center during January 2002-January 2004 to conduct the survival prediction . The study results show that the Neural Network Model has a better predictive ability than the Logistic Regression Model both with regard to the survivors (86.7%, n=361) and with regard to the entire population of patients studied (74 .7%, n=498)

Inhibition of Thymidylate Synthase Activity by Antisense Oligodeoxynucleotide and Possible Role in Thymineless Treatment

Thymidylate synthase (TS) is an important target for chemotherapeutic treatment of cancer. However, efficacy of TS-targeted anticancer drugs is limited by the development of drug resistance as a result of TS gene amplification. In this work, a phosphorothioated antisense oligonucleotide ( ODN), designated ATS-2, was used to suppress cellular synthesis of TS. ATS-2 at 0.2 muM concentration was mixed with lipofectin in a charge ratio of 1:1 and was used to treat the human embryonic kidney (HEK) cell line. A reduction of TS mRNA and protein was achieved. Furthermore, a dose- dependent reduction of cumulative viable cells of up to 98% was observed. Flow cytometer analysis of cell cycle progression indicates that ATS-2- treated cells were arrested and went into apoptosis at the S phase, possibly because of thymidine shortage, suggesting that ATS-2 is specifically effective for dividing cells. When used in combination with the anticancer drug FdUrd, ATS-2 exerted a additive inhibitory effect on cellular proliferation. To elucidate the possible role of cellular thymidine kinase (TdR kinase) in ATS-2 treatment, a second cell line, HeLa , was used. Both HEK and HeLa have similar rates of cell division and ODN uptake. In contrast to HEK, which was shown to have very low levels of TdR kinase activity in [H-3]thymidine incorporation experiments, [H-3] thymidine incorporation in HeLa. was 15-fold greater than that of HEK. We found that HeLa cells were sensitive to FdUrd but were rather resistant to ATS-2. On the contrary, HEK cells were sensitive to ATS- 2 but insensitive to FdUrd. Effects of ATS-2 and FdUrd are, therefore, complementary in thymineless treatment too

Correction/Mutation of Acid Alpha-D-Glucosidase Gene by Modified Single- Stranded Oligonucleotides: In Vitro and in Vivo Studies

Deficiency in acid alpha-D-glucosidase results in Pompe's disease. Modified single-stranded oligonucleotide (ODN) was designed to correct the acid alpha-D-glucosidase gene with a C1935-->A (Asp-->Glu) point mutation which causes a complete loss of enzymatic activity for glycogen digestion in the lysosome. The ODN vectors contained a stretch of normal oligonucleotide flanked by phosphorothioated sequences. The 25mer and 35 mer ODNs were homologous to the target sequence, except for a mismatched base in the middle. The ODNs caused permanent and inheritable restoration of acid alpha-D-glucosidase activity in skin fibroblast cells carrying this mutation derived from a Pompe's disease patient Gene correction was confirmed by amplification refractory mutation system-PCR (ARMS-PCR), restriction fragment length polymorphism (RFLP) and direct DNA cloning and sequencing. The increased acid alpha-D-glucosidase activity was detected using 4-MUG as the artificial substrate. The correction efficiency, ranging from 0.5 to 4% , was dependent on the length and polarity of the MSSOV used , the optimal design being a sense-strand 35mer ODNs. Repeated treatment of the mutant fibroblast cells with the ODNs substantially increased correction. We also constructed ODN vectors to trigger specific and in vivo nonsense mutation in the mouse acid alpha-D-glucosidase gene. The ODNs were in complex with YEEE-K-18, an asialoglycoprotein- receptor ligand tagged with polylysine and targeted to hepatocytes and renal cells in vivo through intravenous injection. The mutated genotype was detected in the liver and the kidney by ARMS-PCR and glycogen accumulation in the lysosome of the liver cells. The studies demonstrate the utility of single -stranded ODN to direct targeted gene correction or mutation in a human hereditary disease and in an animal model. Our data open the possibility of developing ODN vector as a therapeutic approach for treatment of human hereditary diseases caused by point mutation

Use of EDTA Derivatization to Characterize Interactions Between Oligodeoxyribonucleoside Methylphosphonates and Nucleic Acids

Los quad trees propuestos por Bentley en el 1974 son una estructura de datos diseñada para resolver búsquedas asociativas. Esta estructura es una generalización de los árboles binarios de búsqueda y es interesante porque es clásica y es utilizada. Además, es una estructura de datos jerárquica de propósito general que puede ser generalizada fácilmente a múltiples dimensiones. El problema de los quad trees es que es difícil borrar elementos o, dicho de otra manera, que sean dinámicos. Pese a que el borrado en dos dimensiones está definido por H. Samet, el algoritmo es complicado y difícil de implementar. Es más, este algoritmo también es difícil de generalizar para dimensiones mayores a 2. Además, en el algoritmo clásico de inserción, la forma del árbol depende del orden en el que se insertan las llaves. Si las llaves son generadas de manera independiente por una distribución de probabilidad continua, se obtiene un árbol aleatorio cuya altura esperada es logarítmica con respecto al número de llaves del árbol, sin embargo, si las llaves se insertan en orden, la altura del árbol es lineal, lo que tiene un impacto en la eficiencia de las búsquedas exactas en dicho árbol (que en el caso peor son de coste proporcional a la altura del árbol) y también en el de las búsquedas asociativas. En el artículo, Randomized insertion and deletion in point quad trees, se propone, utilizando algoritmos aleatorios, un algoritmo de borrado simple y escalable a más dimensiones. El algoritmo es fácil de describir y de generalizar y es bastante más sencillo de implementar que el de Samet. Además, los algoritmos de inserción y borrado en randomized quad trees garantizan que los quad trees resultantes sean siempre aleatorios. En este trabajo final de grado se estudian los algoritmos randomizados propuestos en el artículo citado anteriormente, se implementan y se propone una implementación alternativa y más eficiente que la original. Finalmente, se analiza experimental y exhaustivamente la eficiencia de los algoritmos de inserción y borrado propuestos. Posteriormente se comparan los resultados con algoritmos alternativos; una implementación con cola de prioridad y una implementación sin randomizar. Los resultados experimentales muestran que los algoritmos randomizados propuestos: 1) funcionan correctamente para cualquier dimensión (de hecho se han probado hasta dimensión 6, pero los algoritmos son válidos para cualquier dimensión, especialmente el borrado), 2) producen árboles aleatorios que cumplen con los costes esperados dados en la literatura, 3) compiten en eficiencia con la implementación con cola de prioridad propuesta especialmente para dimensiones menores de 5, y 4) compiten en eficiencia (son mucho mejores) que el algoritmo de inserción clásico en el caso en el que las llaves a insertar no sean aleatorias o estén dadas en orden. De hecho, este último algoritmo no produce árboles aleatorios cuando las llaves no son generadas de manera aleatoria.The quad trees proposed by Bentley in 1974 are a data structure designed to solve associative queries. This structure is a generalization of binary search trees and is interesting because it is classical and it is used. In addition, it is a general-purpose hierarchical data structure that can easily be generalized to multiple dimensions. The problem with quad trees is that it is difficult to delete elements, or in other words, to be dynamic. Although the two-dimensional deletion is defined by H. Samet, the algorithm is complicated and difficult to implement. Moreover, this algorithm is also difficult to generalize for dimensions greater than 2. In addition, in the classic algorithm of insertion, the shape of the tree depends on the order in which the keys are inserted, if the keys are generated independently by a continuous probability distribution, a random tree is obtained, whose expected height is logarithmic in regard to the number of keys of the tree, however, if the keys are inserted in order, the height of the tree is linear, which has an impact on the efficiency of the exact searches in said tree ( which in worst case their cost is proportional to the height of the tree) and also in the associative queries. In the article, Randomized insertion and deletion in point quad trees, it is proposed, using random algorithms, a simple and scalable deletion algorithm for more dimensions. The algorithm is easy to describe and generalize and is much simpler to implement than the Samet algorithm. In addition, the insertion and deletion algorithms in randomized quad trees guarantee that the resulting quad trees are always random. In this final degree project, the randomized algorithms proposed in the aforementioned article are studied and implemented, and an alternative and more efficient implementation than the original one is proposed. Finally, the efficiency of the proposed insertion and deletion algorithms is experimentally and exhaustively analysed. Subsequently, the results are compared with alternative algorithms; an implementation with priority queue and an implementation without randomization. The experimental results show that the proposed randomized algorithms: 1) work correctly for any dimension (in fact they have been tested up to dimension 6, but the algorithms are valid for any dimension, especially the deletion), 2) produce random trees that meet the costs expected in the literature, 3) compete in efficiency with the implementation with priority queue proposed especially for dimensions less than 5, and 4) compete in efficiency (are much better) than the classical insertion algorithm in the case where the keys to be inserted are not random or are given in order. In fact, this last algorithm does not produce random trees when the keys are not generated randomly

Diagnostic Value of Urine Deoxypyridinoline for Detecting Bone Metastases in Breast Cancer Patients

Deoxypyridinoline (Dpd), a crosslink product of collagen molecules found in bone and excreted in urine during bone degradation, has been described as a marker of bone turnover in metastatic breast cancer. In this study, the urine deoxypyridinoline/creatinine (Dpd/Cre) ratio was determined by enzyme immunoassay in urine samples from 116 women with breast cancer. Bone metastases were confirmed by x-ray or CT scan, with follow-up >6 mo. The urine Dpd/Cre ratio was significantly higher in patients with bone metastasis, compared to those without bone metastasis (p <0.05). In patients with bone metastasis, ratios of urine Dpd/Cre were higher in those with multiple lesions, compared to those with a solitary lesion, and the values also reflected therapeutic response (p <0.05). Serial monitoring of urine Dpd/Cre revealed that an elevation was correlated with disease progression. Patients with stable bone disease under effective therapy had significant diminution of the urine Dpd/Cre ratios, compared to those with progression of bone disease (p <0.05). In conclusion, the urine Dpd/Cre ratio may be a useful marker for detecting bone metastases and evaluating their response to therapy

Antimutagenicity of Supercritical Co2 Extracts of Terminalia Catappa Leaves and Cytotoxicity of the Extracts to Human Hepatoma Cells

Natural antimutagens may prevent cancer and are therefore of great interest to oncologists and the public at large. Phytochemicals are potent antimutagen candidates. When the Ames test was applied to examine the antimutagenic potency of supercritical carbon dioxide (SC-CO2) extracts of Terminalia catappa leaves at a dose of 0.5 mg/plate, toxicity and mutagenicity were not detected. The antimutagenic activity of SC-CO2 extracts increased with decreases of temperature (60, 50, and 40 degreesC) and pressure (4000, 3000, and 2000 psi) used for extraction. The most potent antimutagenicity was observed in extracts obtained at 40 degreesC and 2000 psi. At a dose of 0.5 mg of extract/plate, approximately 80% of the mutagenicity of benzo[a]pyrene (B[a]P, with S-9) and 46% of the mutagenicity of N-methyl-N'-nitroguanidine (MNNG, without S-9) were inhibited. Media supplemented with SC-CO2 extracts at a range of 0-500 mug /mL were used to cultivate human hepatoma (Huh 7) and normal liver (Chang liver) cells. The viability of the cells was assayed by measuring cellular acid phosphatase activity. A dose-dependent growth inhibition of both types of cells was observed. The SC-CO2 extracts were more cytotoxic to Huh 7 cells than to Chang liver cells. The observation that SC-CO2 extracts of T. catappa leaves did not induce mutagenicity at the doses tested while exhibiting potent antimutagenicity and were more cytotoxic to human hepatoma cells than to normal liver cells is of merit and warrants further investigation

Bdellovibrio in Methylotropic Bacteria

The aim of this thesis is to develop from scratch a Python algorithm able to analyse some input data regarding a real coverage area of different technologies and provide an output of the distribution of all their elements. This algorithm has been implemented through the QGIS platform, able to read information tables, plot them in a distribution layer and provide the information in Python vectors. The purpose of the algorithm is to understand the distribution of the QGIS layer, directly related to a coverage distribution near Prague city, and provide a list with the most important neighbours for each polygon in this layer. With all of this mentioned, the code has been implemented taking into account that the work will be continued and the main objective is to analyse huge amount of data and provide a simplified output. This document will introduce the general concepts needed to understand how the data has to be managed, how the algorithm has been developed and how can we thread the results that we obtain from this code.El objetivo de esta tesis es desarrollar desde cero un algoritmo de Python capaz de analizar algunos datos de entrada con respecto a un área de cobertura real de diferentes tecnologías y proporcionar una salida de la distribución de todos sus elementos. Este algoritmo se ha implementado a través de la plataforma QGIS, capaz de leer tablas de información, trazarlas en una capa de distribución y proporcionar la información en vectores Python.L'objectiu d'aquesta tesi és desenvolupar des de zero un algoritme de Python capaç d'analitzar algunes dades d'entrada sobre una àrea de cobertura real de diferents tecnologies i proporcionar una sortida de la distribució de tots els seus elements. Aquest algorisme s'ha implementat a través de la plataforma QGIS, capaç de llegir taules d'informació, dibuixar-les en una capa de distribució i proporcionar la informació en vectors Python