Objectives: To establish a mouse orthotopic bladder cancer model with consistent tumor-take rate. This orthotopic model was subsequently used to evaluate small animal imaging techniques and investigate new therapeutic agents for bladder cancer treatment. / Materials and Methods: Different orthotopic implantation techniques have been tested. MBT-2 cells and syngeneic C3H/He mice were used in all experiments. Chemical bladder pre-treatment with different agents (saline, hydrochloric acid, trypsin and poly-L-lysine) and different concentration of instilled tumor cells (1 x 10⁶ or 2 x 10⁶) were investigated. In the second part of the experiment, trans-abdominal micro-ultrasound imaging (MUI) technique was investigated and validated. Bladder tumor growths were monitored with longitudinal measurement. Mice were killed at every MUI session. Bladder tumor volumes were measured and correlated with gross stereomicroscopy. Using the optimized orthotopic bladder cancer model, targeted contrast enhanced micro-ultrasound imaging has been investigated. VEGFR2 targeted contrast agent was prepared and injected intravenously before imaging sessions. The intra-tumoral perfusion, VEGFR2 expression and blood volume in real time were quantified. Contrast enhanced MUI was performed on Days 14 and 21. The feasibility of targeted contrast enhanced micro-ultrasound imaging was confirmed. After the establishment of orthotopic model and in vivo molecular imaging techniques, this robust platform was used for investigating new treatment agent in localized bladder cancer. Tumor-bearing mice were randomized into control and sunitinibtreated (40 mg/kg) groups. Tumor volume, intra-tumoral perfusion, and in vivo VEGFR2 expression were measured using a targeted contrast-enhanced micro-ultrasound imaging system. The effects of sunitinib malate on angiogenesis and cellular proliferation were measured by CD31 and Ki-67 immunohistochemistry. The clinical outcomes including total bladder weight, tumor stage, and survival were evaluated. / Results: A consistent tumor take-rate of over 90% was achieved by using poly-L-lysine pretreatment with 2 x 10⁶ MBT-2 cells in all of the experiments. MUI identified all tumors that were present on final histology. Measurements of tumor size by MUI and gross microscopy had a high correlation coefficient (r = 0.97). Measurements of intra-tumoral perfusion and in vivo VEGFR2 expression were also proved to be feasible. After the technical refinement and modification, complete measurements could be performed in all mice (n = 10) at 2 consecutive imaging sessions. No adverse effects occurred due to anesthesia or the ultrasound contrast agent. This is the first report of applying targeted contrast enhanced MUI in orthotopic bladder cancer model. Finally, sunitinib was found to have significant tumor growth inhibition in both in vitro and in vivo experiments. In the orthotopic model, tumors in sunitinib-treated mice had reduced tumor volume and stage, lower proliferation index and micro-vessel density. Sunitinib prolonged survival in tumor-bearing mice as compared to control group. / Conclusions: The development of reliable orthotopic animal models assists in the discovery of novel therapeutic agents. The establishment in the methods of implantation with improved tumor-take rate and the advances in imaging technology form the important foundation of basic research in bladder cancer. Trans-abdominal MUI is proven to be a valuable tool for translational studies involving orthotopic mouse bladder cancer models. Furthermore, the first report of the application of targeted contrast enhanced MUI in deep-seated tumor in bladder has been published. It enables investigators to monitor tumor angiogenesis and vascular changes after treatment. It will be useful for direct, noninvasive, in vivo evaluation of anti-angiogenesis therapeutic agents. The preclinical study has demonstrated the activities of a new class of targeted therapy against localized bladder cancer in an orthotopic mouse model. Sunitinib inhibits tumor growth and thus decreases the tumor burden and prolongs survival compared with placebo. These results provide a rationale for future clinical trials using VEGFR-targeted treatments of localized bladder cancer in the neo-adjuvant and adjuvant settings. / Chan, Shu Yin Eddie. / Thesis (M.D) Chinese University of Hong Kong, 2014. / Includes bibliographical references (leaves 189-212).

Lesões na inervação do trato urinário inferior ocasionado por traumatismo raquimedular afetam geralmente o músculo detrusor e o esfíncteres uretrais. Estas alterações acarretam problemas basicamente de incontinência urinária e aumento da pressão intravesical, decorrente deste traumatismo, trazendo consequências para o funcionamento do sistema urinário superior. Quantificar os elementos fibrosos da matriz extracelular e fibras musculares das bexigas neurogênicas hiper-reflexas comparando-as com bexigas normais. Foram utilizadas 6 amostras de bexigas neurogênicas de indivíduos que foram submetidos a cirurgia de reparação por cistoenteroplastia realizados pelo serviço de urologia do Hospital Municipal Souza Aguiar, estas amostras foram fixadas imediatamente em solução tamponada de formalina a 10%. O controle com amostras iguais as do estudo extraída de cadáveres cuja causa morte não relacionava-se ao sistema urogenital macroscópicamente. O material foi submetido as seguintes técnicas histoquímicas: H&E, van Gieson e Resorcina Fucsina resorcina de Weigert com prévia oxidação pela oxona. Imunohistoquímica: anti-elastina. A observação dos cortes corados pelo van Gieson demonstrou uma diminuição significativa do músculo liso de 13% e aumento do colágeno em 72% e as fibras do sistema elástico um aumento de 101%. Conclusão. Nas bexigas neurogênicas hiper-reflexas o músculo detrusor e os elementos fibrosos da matriz foram profundamente modificados. As fibras do sistema elástico foram as mais afetadas. / Lesions on lower urinary tract innervations caused by spinal cord injuries usually affect the detrusor muscle and urethral sphincter. Beside the smooth muscle fibers, the collagen fibers and elastic system fibers, fibrous components of the extracellular matrix of the bladder wall, are strongly related to vesicle bladder compliance. For this reason the aim of this work is to quantify the fibrous elements of the extracellular matrix and muscle fibers of the neurogenic bladder hyperreflexia. Samples of neurogenic bladder were obtained from six men who had previously undergone surgical repair. The control group samples (n=6) were similarly obtained from patients whose deaths were not related to the urogenital system. The samples were stained using the following histochemical techniques: H&E, Van Gieson, Weigert and Sirius Red. Sections stained with Sirius Red were observed under polarization light microscopy to characterize possible different kinds of collagen. Immunohistochemical technique was used to characterize and quantify the elastic system fibers. Quantification analysis was performed by stereological methods. An increase of 72% of the collagen was observed. Nevertheless, the most significant difference observed was the raising of 101% of the elastic system fibers. Contrary the smooth muscle fibers showed a decrease of 13%. In the neurogenic bladder with detrusor hyperreflexia the fibrous elements of the extracellular matrix and smooth muscle fibers were greatly modified. The elastic system fibers seem to be the most affected in this disease.

A radioterapia é frequentemente utilizada no tratamento de tumores da próstata, porém durante esse procedimento a bexiga sadia usualmente sofre efeitos colaterais. Através do uso de um modelo animal para irradiação pélvica, avaliamos se a suplementação nutricional com L-glutamina poderia prevenir possíveis danos na parede da bexiga, especialmente em suas camadas mais superficiais. Ratos Wistar adultos machos com idade entre 3 e 4 meses foram separados em grupos de 8 animais: grupo controle que não recebeu a irradiação; grupos somente irradiados que foram mortos 7 (R7) e 15 dias (R15) após a irradiação (dose única de 10 Gy na região pélvico-abdominal); grupos irradiados e suplementados com L-glutamina (0,65g/kg de peso por dia), que foram mortos 7 (RG7) ou 15 após a irradiação. Células e vasos sanguíneos da lâmina própria, bem como o urotélio, foram avaliados com métodos histológicos. No urotélio foram feitas análises da altura e densidade nuclear e na lâmina própria densidade celular, densidade vascular e o número de mastócitos. Os resultados mostraram que em R7, a altura e densidade nuclear do urotélio e a densidade celular da lâmina própria não foram alterados significativamente. Entretanto a densidade dos vasos sanguíneos foi reduzida em 48% (p<0,05) e essa alteração foi evitada pela glutamina (p <0,02). No grupo R15, a densidade celular do epitélio aumentou em 35% (p<0,02). A densidade celular da lâmina própria não apresentou diferença estatística entre os grupos. Os mastócitos na lâmina própria foram reduzidos em R7 e R15. Apesar de ainda reduzidos em RG7 em RG15 houve aumento no número desse tipo celular o que sugere uma ação positiva da glutamina. Células &#945;-actina positivas na lâmina própria formam uma camada suburotelial e foram identificadas como miofibroblastos. A espessura dessa camada aumentou em R7, mas foi semelhante ao controle em RG7, enquanto alterações em R15 e RG15 foram menos evidentes. Esses resultados mostraram que a utilização da L-glutamina antes e após a radioterapia deve ser considerada para uso humano na proteção da bexiga contra os efeitos da radiação. / Radiotherapy is often used to treat prostate tumors, but the normal bladder is usually adversely affected. Using an animal model of pelvic radiation, we investigated whether glutamine nutritional supplementation can prevent radiation-induced damage to the bladder, especially in its more superficial layers. Male rats aged 3 to 4 months were divided into groups of 8 animals each: controls, which consisted intact animals; radiated-only rats, which were sacrificed 7 (R7) or 15 (R15) days after a radiation session (10 Gy aimed at the pelvico-abdominal region); and radiated rats receiving L-glutamine supplementation (0.65 g/kg body weight/day), which were sacrificed 7 (RG7) or 15 (RG15) days after the radiation session. Morphological and morphometric analysis of the urothelium were made. Nuclear density, lamina propria cell density and mast cells numbers per area were counted. The results showed that, in R7, epithelial thickness, epithelial cell density, and cell density in the lamina propria were not significantly affected. However, density of blood vessels in R7 was reduced by 48% (p < 0.05) and this alteration was mostly prevented by glutamine (p < 0.02). In R15, density of blood vessels in the lamina propria was not significantly modified. However, epithelial thickness was reduced by 25% (p < 0.05) in R15, and this effect was prevented by glutamine (p < 0.01). In R15, epithelial cell density was increased by 35% (p < 0.02), but glutamine did not protect against this radiation-induced increase. Cell density in the lamina propria was likewise unaffected in R15. Density of mast cells in the lamina propria was markedly reduced in R7 and R15. The density was still reduced in RG7, but a higher density in RG15 suggested a glutamine-mediated recovery. Alpha-actin positive cells in the lamina propria formed a suburothelial layer and were identified as myofibroblasts. Thickness of this layer was increased in R7, but was similar to controls in RG7, while changes in R15 and RG15 were less evident. In conclusion, pelvic radiation leads to significant acute and post-acute alterations in the composition and structural features of the vesical lamina propria and epithelium. Most of these changes, however, can be prevented by glutamine nutritional supplementation. These results emphasize, therefore, the potential use of this aminoacid as a radioprotective drug.

Group B Streptococcus, or GBS, is a gram positive bacteria commonly found in the gastrointestinal, genital and urinary tract of healthy adults. Between 10% and 30% of all pregnant women are colonized with GBS in the vagina or rectum. While GBS colonized mothers typically show no symptoms or adverse health effects, the bacteria can be passed to their child during labor and delivery. Although significant progress has been made in the identification and treatment of GBS, it remains the leading infectious cause of Page 14 2014 Appalachian Student Research Forum morbidity and mortality among newborns in the United States. The current guidelines recommended by the Center for Disease Control and Prevention (CDC) and endorsed by the American College of Obstetrics and Gynecology (ACOG) is to test pregnant women for GBS colonization between 34-37 weeks of gestation. The current gold standard for identification of GBS colonization is the use of selective enrichment broth (SEB) followed by culture and biochemical testing. Identified concerns with the culture procedure are: 1) the length of time it takes to get the results, 2) the lower sensitivity if the SEB step is left out to improve turn-around-time (TAT) and 3) the limited number of qualified technicians available to perform the complex test. Recently, several semi-automated molecular assays have been developed for identification of GBS which are marketed as having equivalent sensitivity and specificity to SEB followed by culture. The goals of this study were to: 1) validate the sensitivity and specificity of an FDA approved GBS molecular assay (Illumigene, Meridian Bioscience) and 2) evaluate a new testing strategy utilizing SEB followed by the Illumigene GBS assay to see if it offers an improvement in TAT when compared to SEB culture in our in-house microbiology lab and to those sent out to a national reference lab for GBS DNA assay. During the validation process, 20 consecutive samples were submitted to SEB followed by simultaneous in-house culture and Illumigene assay for GBS. The method validation experiments were analyzed using the EP Evaluator version 11 statistical software (Data Innovations). Comparison of TAT was evaluated utilizing a blinded report generated from our Laboratory Information System (Harvest, Orchard Software) for a 2 month period for the GBS tests performed using the SEB followed by Illumigene molecular assay (n=73), in-house SEB followed by culture (n=50) and send-out reference lab GBS DNA assay (n=43) procedures. The TAT (hrs) for each method (Mean±SEM) were determined from the time of collection until result approval. The Illumigene assay had a high sensitivity (100%) and specificity (100%) when compared to SEB followed by culture for identification of GBS. Utilization of the Illumigene assay following SEB significantly (p

The purpose of this study is to investigate whether a prolonged delay in diagnosis of bladder cancer will result in worse outcomes for those patients, compared to those patients with a shorter diagnostic time interval. Data was collected on 247 patients newly diagnosed with transitional cell carcinoma of the bladder from January 1996 to December 2006 (10 years). The medical records of these patients were reviewed for demographics, pathological stage, date of consultation to the genitourinary (GU) service, and date of diagnosis by transurethral resection of bladder tumor (TURBT). The specialty delay was calculated as the time between the date of consultation to the GU service to the establishment of a diagnosis by TURBT. Univariate analyses were performed to test the association of specialty delay with clinical features and all-cause mortality. The median specialty delay in this study was 100 days. There was a trend towards a longer specialty delay for muscle-invasive disease (T2-T4) in comparison to superficial disease (Ta and T1). There was a significant correlation between all-cause mortality and increasing clinical stage (p=0.01). There was a paradoxical finding that patients with a specialty delay greater than 100 days had a significant reduction in all-cause death in comparison to patients with a specialty delay of 100 days or less (relative risk=0.59; 95% CI 0.36-0.90; p=0.01). In conclusion, this study did not confirm the hypothesis that a prolonged specialty delay in patients diagnosed with bladder cancer would result in a worse prognosis. In fact, there was a paradoxical finding that patients with a specialty delay greater than the median delay of 100 days had a better prognosis.

This study was prompted by the inconsistent reports and apparent controversies that exist in the biomedical literature on the responses of diabetic bladder strips to cholinergic nerve stimulation or exogenous administration of muscarinic agonists, especially acetylcholine (ACh), in vitro. In the present study, acetylcholine-induced contractions of urinary bladders isolated from normoglycaemic (normal) and streptozotocin-treated, diabetic Wistar rats were examined under physiological conditions. Mechanical contractile changes of the isolated urinary bladders of STZ-treated, diabetic rats in response to bath-applied acetylcholine were compared with those obtained from isolated urinary bladders of normal, age-matched, control rats. Results obtained show that urinary bladders from diabetic rats consistently weighed more, and were always more spontaneously active after mounting, than those of the age-matched normal, control rats. ft A Acetylcholine (ACh, 10" -10" M) provoked concentration-related, atropine-sensitive contractions of the isolated urinary bladders of both diabetic and age-matched normal, control rats. However, acetylcholine always induced more powerful and greater contractions of the diabetic bladders compared with bladders from the age-matched normal, control rats. The enhanced contractile responses of the diabetic bladder strips to bath-applied ACh were detected soon after induction of diabetes, and the magnitude and/or intensity of the enhanced contractile responses to ACh continued to increase as the diabetic state of the animals progressed. Although this preliminary study could not establish the mechanism of the increased contractile responsiveness of the diabetic bladders to the muscarinic agonist (ACh) used, the results tend to suggest that alterations in diabetic urinary bladder synaptosomal, vesicle-bound neurotransmitter (ACh) concentrations and the compensatory increase in the density of muscarinic M3-receptor population in diabetic bladders are two of the most attractive plausible mechanisms of the increased diabetic bladder responsiveness to bath-applied acetylcholine. / Thesis (M.Sc.Pharm.)-University of KwaZulu-Natal, Westville, 2006.

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

Thesis (Ph. D.)--University of Virginia, 2008. / Title from title page. Includes bibliographical references. Also available online through Digital Dissertations.

Thesis (Ph. D.)--University of Missouri-Columbia, 2006. / The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Vita. Includes bibliographical references.

Thesis (Ph.D.). / Written for the Dept. of Pharmacology & Therapeutics. Title from title page of PDF (viewed 2009/06/10). Includes bibliographical references.