Liu Bonnie Pui-ling. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 134-163). / Abstracts in English and Chinese. / Acknowledgments --- p.i / Abbreviations --- p.ii / Abstract / English --- p.1 / Chinese --- p.3 / Chapter Chapter 1 --- General Introduction / Chapter 1.1 --- Colorectal Cancer Statistics --- p.5 / Chapter 1.2 --- Colon Cancer --- p.5 / Chapter 1.3 --- Treatment --- p.6 / Chapter 1.4 --- Effects of Cytotoxic Drug Treatment --- p.7 / Chapter 1.5 --- Cell Cycle --- p.8 / Chapter 1.6 --- Oxidases --- p.9 / Chapter 1.7 --- Chemistry of Novel β-carboline: Tangutorine --- p.11 / Chapter 1.8 --- Aim of Study --- p.14 / Chapter Chapter 2 --- Cytotoxicity / Chapter 2.1 --- Introduction --- p.15 / Chapter 2.2 --- Materials and Methods --- p.18 / Chapter 2.3 --- Results --- p.23 / Chapter 2.4 --- Discussion --- p.44 / Chapter Chapter 3 --- Oxidase Activity and Protein Oxidation / Chapter 3.1 --- Introduction --- p.48 / Chapter 3.2 --- Materials and Methods --- p.54 / Chapter 3.3 --- Results --- p.60 / Chapter 3.4 --- Discussion --- p.80 / Chapter Chapter 4 --- Cell Cycle / Chapter 4.1 --- Introduction --- p.89 / Chapter 4.2 --- Materials and Methods --- p.93 / Chapter 4.3 --- Results --- p.96 / Chapter 4.4 --- Discussion --- p.118 / Chapter Chapter 5 --- General Discussion --- p.126 / References --- p.134

O presente estudo objetivou avaliar os efeitos da inflamação e das expressões de metalotioneínas (MTs) e ciclooxigenase-2 (COX-2) na desnervação associada à carcinogênese experimental colônica, através da avaliação de imunoexpressão de COX-2, mieloperoxidase (MPO), neutrophil elastase (NE) e cluster de diferenciação 68 (CD68). Ainda, para analisar a presença de mutações e de danos ao material genético, realizou-se a técnica de imunoistoquímica para identificação de células imunomarcadas para MTs e histonas gamma-H2AX (H2AX). 52 ratos machos Wistar foram divididos em seus respectivos grupos: C (controle), B (controle dos animais desnervados), D (aplicação de DMH 30 mg/Kg ou 125 mg/Kg), BD (Cloreto de benzalcônio 0,3% na serosa intestinal, com posterior aplicação de DMH) e DB (aplicação de DMH, com posterior administração de BAC), sendo subdivididos de acordo com o tempo em que foram mortos, sendo: 24 horas, 72 horas (para avaliação da fase de iniciação) e 20 dias (para avaliação da fase de promoção). Como resultado, a desnervação foi capaz de proteger as células colônicas contra os efeitos desse carcinógeno, o que pode ser observado através da diminuição significativa da imunomarcação de MTs e de H2AX nos grupos desnervados. Ainda, através do aumento do número de células positivamente marcadas para MPO, NE e CD68, neutrófilos, macrófagos e/ou monócitos podem estar estritamente relacionados ao papel protetor da desnervação contra o câncer em um período de 20 dias após administração do DMH, uma vez que os animais desnervados apresentaram uma significativa diminuição desses marcadores. A desnervação aumentou o número de células imunomarcadas para MTs em animais que receberam apenas a aplicação do DMH e foram mortos após 20 dias - o que possivelmente diminuiu a expressão de COX-2 encontrada nesse estudo. Assim, sugere-se que os possíveis mecanismos protetores desempenhados pela desnervação contra o câncer sejam: (i) o aumento de MTs - conhecidamente protetoras à célula, e (ii) atuação da desnervação sobre o processo inflamatório, mais especificamente sobre neutrófilos e macrófagos e/ou monócitos. / This study evaluated the effects of inflammation and expression of metallothionein (MTs) and cyclooxygenase-2 (COX-2) on denervation associated to colonic experimental carcinogenesis by immunohistochemical evaluation of COX-2, myeloperoxidase (MPO), neutrophil elastase (NE) and cluster of differentiation 68 (CD68). Also, to analyze the presence of mutations and damage to DNA, was performed the immunohistochemical technique to identify positive immunostained cells for MTs and histone gamma-H2AX (H2AX). 52 male Wistar rats were divided into their respective groups: C (control), CB (control related to denervated animals), D (application of DMH 30 mg/Kg or 125 mg/kg), BD (0.3% benzalkhonium chloride administration in the intestinal serosa, and subsequent application of DMH) and DB (application of DMH with subsequent administration of BAC, in serous). The animals were subdivided according to euthanasia time: 24 hours, 72 hours (for evaluation of the initiation phase of the carcinogenesis process) and 20 days (for assessment of promoting phase). As result, the denervation process was able to protect colonic cells against the carcinogen effects, which can be seen through the significant decrease in immunostained cells for MTs and H2AX. Moreover, through the increased expression of MPO, NE and CD68, we found that neutrophils, macrophages and/or monocytes are closely related to inflammation on development of early colonic experimental carcinogenesis (20 days groups) - which was reverted by denervation. The denervation increased the number of immunostained cells for MTs in denervated animals that received application of DMH - what could possibly decreased COX-2 expression found in this study. Thus, it is suggested that the possible protective mechanisms performed by denervation against cancer are: (i) increased MTs - known to be protective to the cell, and (ii) actuation on inflammatory process, more specifically on neutrophils and macrophages and/or monocytes.

O Orlistat é um membro de uma classe de drogas usadas como tratamento para obesidade. No entanto, a segurança de seu uso em longo prazo ainda não é conhecida. O Orlistat exerce sua atividade no lúmen do trato gastrintestinal inibindo a enzima lipase pancreática, responsável pela hidrólise dos triacilglicerídeos normalmente ingeridos com a dieta. Esse medicamento, quimicamente sintetizado, é um derivado da lipstatina, inibidora natural da lipase produzida pelo Streptomyces toxytricini. Assim, a excreção de gordura fecal fica significativamente aumentada com o uso do Orlistat. Estudos epidemiológicos e em modelos experimentais, sugerem que o aumento das dietas hipergordurosas tem efeito promotor para o câncer colorretal. Tal efeito deve ser relacionado, pelo menos parcialmente, às mudanças intra-colônicas causadas pela ação direta da gordura nas células da mucosa do cólon, os colonócitos. O estudo atual tem como objetivo verificar os efeitos do Orlistat na formação colônica de focos de criptas aberrantes (FCA) e na proliferação celular epitelial da mucosa gastrintestinal. Ratos Wistar machos receberam dieta padrão ou dieta com aumento de gordura, suplementada ou não com Orlistat (200 mg/kg), e duas doses semanais do carcinógeno químico Dimetilhidrazina (DMH) (25 mg/kg). Após 30 dias, nos animais tratados com DMH, o Orlistat foi associado a um aumento significativo no número de FCAs colônicos e na proliferação celular epitelial da mucosa do cólon, independentemente da dieta. Os achados obtidos neste trabalho permitem concluir que o aumento do teor de gordura na luz do cólon distal, em decorrência da ação do Orlistat, pode potencializar a ação da DMH na formação de FCAs e no aumento da proliferação celular epitelial da mucosa colônica. / Orlistat is a member of a drug class used as obesity treatment. However, the security of its use in a long period of time is not known yet. Orlistat has its activity in the lumen of the gastrointestinal tract inhibiting the pancreatic lipase enzyme responsible for the hydrolyze of the triglycerides that usually are swallowed with the diet. This drug chemically synthesized, is a derived from lipstatina, that inhibit naturally the lipase produced by Streptomyces Toxytricini. So, the excretion of fat excrement stays significantly increased with the use of Orlistat. Epidemiologic studies and in experimental pattern, suggest that the increase of the high-fat diets facilitate the appearance of the colorectal cancer. Such effect must be related, at least partially to the changes intracolonic caused by the direct action of the fat in the cells of the colon mucous, the colonocytes. The current study has as objective to check the effects of the Orlistat on the formation of rat colonic aberrant crypt foci (ACF) and in the epithelial cell proliferation of the gastrointestinal mucous. Male Wistar rats received either a standard diet or a high fat diet (HFD), supplemented or not with Orlistat (200 mg/kg chow) and two doses of the carcinogen dimethyl-hydrazine (25 mg/Kg). After 30 days, Orlistat was associated to a significant increase in the number of colonic ACFs and cell proliferation in DMH-treated animals, independently of the HFD. The find got in this study permit to conclude that the increase in the level of adiposity inside the distal colon, due to Orlistat action, can potencializar the DMH action in the formation of ACFs and in the increase of epithelial cell proliferation of the colônica mucous.

Inflammatory bowel diseases, (IBD), are a group of chronic disorders of the gastro-intestinal tract, and include Crohn’s disease (CD) and Ulcerative Colitis (UC). The pathogenesis is not known, but involves at least in part a loss of tolerance towards the commensal colonic microbiota. In this thesis, we show in animal models of CD and UC that the colonic mucosal blood flow increased compared to healthy animals. This blood flow increase is due to an up regulation of endothelial nitric oxide synthase (NOS). Further, we show in the UC model that the thickness of the firmly adherent colonic mucus layer increased compared to healthy animals. This increase is due to an up regulation of inducible NOS in the epithelium. Both the blood flow and mucus thickness increase appear to be protective mechanisms.  We demonstrate that the firmly adherent colonic mucus layer acts as a partial barrier towards luminal bacteria. In the UC model, this barrier is destroyed, causing increased bacterial translocation. The adhesion molecule P-selectin was up regulated in the UC model, leading to increased interactions between leukocytes and the endothelium, but also increased interactions between platelets and the endothelium. This indicates that not only leukocytes, but also platelets are involved in colonic inflammation. The addition of the probiotic bacterial strain Lactobacillus reuteri prevented disease by normalizing P-selectin levels and endothelial interactions with leukocytes and platelets. Lactobacillus reuteri also decreased bacterial translocation over the epithelium. In summary, this thesis highlights the importance of colonic barrier functions, and investigates the role of the microbiota in experimental IBD.

Thesis (Ph. D.)--University of Nevada, Reno, 1999. / Includes bibliographical references. Online version available on the World Wide Web.

Thesis (Ph. D.)--University of Missouri--Columbia, 2001. / Typescript. Vita. Includes bibliographical references (leaves 95-113). Also available on the Internet.

No description available.

In questo studio abbiamo analizzato l'impatto dellethermophiluds Streptococcus sull'epitelio del colon di ratto. Dopo la generazione del modello di ratto axenico e inoculato con S. thermophilus abbiamo investigato l'interazione tra il batterio e epithelo del colon di ratto. Dopo questo studio integrativo abbiamo analizzato l’espressione genica del colon usando due diversi approcci: Ibridazione sottrattiva I trascritti ottenuti dopo il sequenziamento dei cloni ottenuti con la SSH sono stati raggruppati in diversi categorie funzionali: arresto del ciclo cellulare e induzione del differenziamento, comunicazione cellulare e binding . due geni candidati sono stati privilegiati, krupel like fattore 4 e 14-3-3σ. Questi geni candidati sono stati analizzati mediante RT-PCR, qRT-PCR e Western Blot in animali mono-associati e in animali germ-free. I test hanno confermato che l’espressione dei geni candidati aumenta in presenza di S. thermophilus. Microarray Analysis. L’spressione genica è stata misurata per due gruppi di animali: i) ratti privi di germi, ii) ratti mono-associati con il ceppo LMD9 di Streptococcus thermophilus. I risultati delle analisi dei dati di microarray indicano che Streptococcus thermophilus influenza notevolmente l'espressione genica nelle cellule epiteliali del colon. / In this study we have investigated the impact of Streptococcus thermophiluds on the rat colonic epithelium. After generation of the model axenic rat and inoculated with S. thermophilus we have investigated the interplay between bacteria and host colon. Colonic epithelium gene expression was investigated also, with two different approaches: Suppressive Subtractive Hybridization. The subtraction library was prepared subtracting mRNA between epithelial cells from colonic mono-associated rats and germ-free rats. The transcripts generated by SSH were grouped into divers Functional groups: cell-cycle arrest and induction of differentiation; cell-communication and binding. Tow candidates genes were privileged, krupel like factor 4 and 14-3-3σ. These candidate genes were tested by RT-PCR, qRT-PCR and Western blot in mono-associated animals and in germ-free animals. The tests confirmed that candidate genes increase their expression in the presence of S. thermophilus. Microarray analysis. Gene expression was measured in tow groups of animals: i) germ-free rats; ii) mono-associated rats inoculated with LMD9 strain of Streptococcus thermophilus. The results of microarray analysis data show that Streptococcus thermophilus remarkably affected gene expression in the colonic epithelial cells. Streptococcus thermophilus enhanced the expression of genes involved in different pathways in the host, compared to the gem free group.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 4 uppsatser.

Thesis (Ph. D.) -- University of Texas Southwestern Medical Center at Dallas, 2005. / Vita. Bibliography: 49-53.