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The aim of this study was to observe levels of oxidative DNA damage in patients with multiple injuries in correlation with the nutritional support that the patients have received during their hospital stay. Oxidative DNA damage was evaluated in two periods of time, first evaluation was performed during standard nutritional support according to the ESPEN guidelines. Second evaluation was performed after a change in nutrition according to individual parameters of metabolism and utilization of nutritional components based on indirect calorimetric measurements. This study included 6 patients with multiple injuries hospitalized in the Intensive Care Unit 1 at the Department of Surgery, University Hospital in Hradec Kralove. In this experiment DNA isolated from peripheral lymphocytes was used to evaluate oxidative DNA damage. This DNA was analyzed using the Comet Assay method. The enzymatic version of the Comet Assay was used to determine the oxidative damage of purines and pyrimidines, and the alkaline version was used for detection of single strand breaks. Mann-Whitney test was used for statistic evaluation the difference between both measuremetns, correlation analysis for relations between Comet Assay results and clinical parameters. Significant correlations between a total amount of nutrients given...

Irradiamos amostras de DNA com luz UVA em diferentes condições para estudar os possíveis mecanismos envolvidos na indução de lesões de DNA por essa radiação. As lesões de DNA formadas após as irradiações foram quantificadas com enzimas de reparo de DNA que reconhecem e clivam os sítios contendo bases oxidadas e dímeros de pirimidina (CPDs). Complementando essas análises, foram realizados ensaios com anticorpos e HPLC-ED. NaCl e uma maior concentração de DNA são capazes de diminuir a indução de CPDs. Danos gerados por estresse oxidativo são inibidos na presença de azida de sódio e quelantes de metais, indicando o envolvimento de oxigênio singlete e reações de Fenton, na geração dessas lesões. Água deuterada e DNA mais concentrado aumentaram a indução de bases oxidadas. Quanto maior a quantidade de DNA irradiado, mais oxigênio singlete é formado, o que indica um possível mecanismo de fotossensibilização endógeno. / DNA samples were irradiated with UVA light in different conditions for studying the possible mechanisms involved in the induction of DNA lesions by this radiation. DNA lesions formed after irradiation were quantified with DNA repair enzymes, which recognize and cleave the sites containing oxidized bases and pyrimidine dimers (CPDs). Complementing these analyses, tests were performed with antibodies and HPLC-ED. NaCl and more concentrated DNA are capable of reducing the induction of CPDs. Damage caused by oxidative stress is inhibited in the presence of sodium azide and metal chelators, indicating the involvement of singlet oxygen and Fenton reactions, in the generation of these lesions. Deuterated water and more concentrated DNA increased the induction of oxidized bases. The bigger the amount of irradiated DNA, the more singlet oxygen is formed, which indicates a possible endogenous photosensitization mechanism.

Ma, Yongzheng. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references. / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

No description available.

Ho Cheuk-nang. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 89-94). / Abstracts in English and Chinese. / Acknowledge --- p.ii / Table of Contents --- p.iii / Lists of Tables --- p.vi / Lists of Figures --- p.vii / Abstract (English Version) --- p.x / Abstract (Chinese Version) --- p.xii / Chapter 1. --- Literature Survey of Phosphorus Chemical Shift Studies of DNA --- p.1 / Chapter 1.1 --- Introduction --- p.1 / Chapter 1.2 --- General Review of DNA Structures --- p.2 / Chapter 1.2.1 --- "Nomenclature, Symbols and Numbering Scheme" --- p.2 / Chapter 1.2.2 --- Conformations of DNA --- p.7 / Chapter 1.2.3 --- Random Coil State --- p.8 / Chapter 1.3 --- Phosphorus Chemical Shift Studies of DNA --- p.9 / Chapter 1.4 --- Purpose of This Work --- p.10 / Chapter 2. --- Methodology for Studying the Sequence Effect on Random Coil Phosphorus Chemical Shift --- p.12 / Chapter 2.1 --- Introduction --- p.12 / Chapter 2.2 --- Determination of Sequence Effect --- p.12 / Chapter 2.3 --- Design of Sequence --- p.13 / Chapter 2.4 --- Sample Preparation --- p.14 / Chapter 2.5 --- Measurement of Phosphorus Chemical Shift --- p.16 / Chapter 2.5.1 --- Proton Resonance Assignments --- p.16 / Chapter 2.5.2 --- Phosphorus Resonance Assignments --- p.20 / Chapter 2.6 --- Determination of Sugar Conformation --- p.23 / Chapter 2.7 --- Determination of Backbone Conformation --- p.25 / Chapter 3. --- Results and Discussion of Random Coil Phosphorus Chemical Shift of DNA --- p.28 / Chapter 3.1 --- Introduction --- p.28 / Chapter 3.2 --- Resonance Assignments --- p.28 / Chapter 3.3.1 --- Proton Resonance Assignments --- p.31 / Chapter 3.3.2 --- Phosphorus Resonance Assignments --- p.31 / Chapter 3.3 --- Verification of Random Coil State --- p.31 / Chapter 3.2.1 --- Variable Temperature Proton Chemical Shift --- p.31 / Chapter 3.2.2 --- Sugar Conformation --- p.32 / Chapter 3.2.3 --- Backbone Conformation --- p.33 / Chapter 3.4 --- Random Coil Phosphorus Chemical Shift of DNA --- p.33 / Chapter 3.4.1 --- Temperature Effect --- p.33 / Chapter 3.4.2 --- Neighbor Effect --- p.36 / Chapter 3.5 --- Random Coil Phosphorus Chemical Shift Prediction --- p.42 / Chapter 3.5.1 --- Dimer Model Prediction Protocol --- p.42 / Chapter 3.5.2 --- Trimer Model Prediction Protocol --- p.45 / Chapter 4. --- Conclusions and Future Work --- p.48 / Appendix I H6/H8-H1' regions of 2D NOESY spectra of SS1-SS16 --- p.49 / "Appendix II 'H and 31P resonance assignments and 3Jhi1,h2,3Jh1，h2´ح and 3Jh3，p coupling constants of SS1-SS16" --- p.57 / Appendix III H3' regions 1H-31P HSQC spectra of SS1-SS16 --- p.65 / Appendix IV H1，-H2，/H2´حregions of DQF-COSY spectra of SS1- SS16 --- p.73 / Appendix V H3' regions of 1H-31P selective heteronuclear COSY spectra of SS1-SS16 --- p.81 / References --- p.89

For the successful detection of selected DNA sequences or mutated genes associated with human disease, there are several challenges that the current research aims to overcome - higher sensitivity, greater selectivity and rapid assaying time. An electrochemical device using redox-active intercalators to transduce DNA hybridization via long-range electron transfer is presented in this thesis which aims to address the above challenges. The DNA recognition interface is composed of thiolated single-stranded DNA (ss-DNA) and a diluent component both of which are self-assembled onto a gold electrode. This project seeks to advance fundamental insight into issues that impact the structure and behavior of surface-immobilized DNA towards hybridization with target complementary ss-DNA. After the optimal conditions have been identified for the construction of a reproducible DNA recognition layer, a stepwise detection scheme using an anionic intercalator, as the redox molecule is introduced for the DNA transduction. The stepwise detection relies on the absence of any electrochemistry prior to DNA hybridization. Upon hybridization, the perfectly stacked DNA is capable of mediating the electrochemical oxidation and reduction of intercalated species and hence voltammetric peaks become evident. Although excellent selectivity towards single-base mismatch detection is achieved, this detection scheme has a high detection limit and slow assaying time. However, this is overcome by a novel in situ approach where the electrochemistry is performed in the presence of both complementary target DNA and intercalator. The effect of different DNA recognition interfaces on hybridization is also investigated using electrochemical and gravimetric techniques where the hybridization efficiency, kinetics and affinity constant of hybridization were assessed. These measurements showed that the length of the diluent layer has a large impact on the time taken to form a perfect duplex but no impact on the initial recognition of the target DNA by the immobilized probe DNA. Fundamental aspects of the DNA technology towards assaying small molecules which have binding affinity to DNA are also investigated. The probe ss-DNA sensing interface was found to be highly sensitive towards detection of Cd2+. The long-range electron transfer approach was also utilized in gaining more insight knowledge of the interaction of cisplatin, an anti-cancer drug with the DNA.

Graduation date: 1997