Chan, Ka Ki Alex. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (leaves 245-263). / Abstracts in English and Chinese. / Abstract --- p.i / Chinese abstract --- p.iii / Acknowledgements --- p.v / Table of contents --- p.vii / Chapter Chapter One --- General introduction --- p.1 / Chapter 1.1 --- Structure and function of the enteric nervous system --- p.1 / Chapter 1.2 --- Neural crest cells (NCC) --- p.5 / Chapter 1.2.1 --- Vagal neural crest cells --- p.7 / Chapter 1.2.2 --- Sacral neural crest cells --- p.10 / Chapter 1.3 --- Prespecialization of the neural crest cells to form ENS --- p.15 / Chapter 1.4 --- Signaling pathways involved in ENS development --- p.19 / Chapter 1.4.1 --- Endothelin signaling pathway --- p.20 / Chapter 1.4.2 --- Ret signaling pathway: GDNF/Ret/GFRa1 --- p.22 / Chapter 1.4.3 --- Ret signaling pathway: NRTN/Ret/GFRa2 --- p.26 / Chapter 1.4.4 --- Phox2b --- p.28 / Chapter 1.4.5 --- Sox10 --- p.29 / Chapter 1.5 --- Hirschsprung's Disease (HSCR) --- p.31 / Chapter 1.6 --- Objective of studies --- p.32 / Figures and legends --- p.35 / Chapter Chapter Two --- The early migratory pathways of mouse sacral neural crest cells --- p.39 / Chapter 2.1 --- Introduction --- p.39 / Chapter 2.2 --- Materials and Methods --- p.46 / Chapter 2.2.1 --- Animals --- p.46 / Chapter 2.2.2 --- Isolation of the mouse embryos at E95 --- p.46 / Chapter 2.2.3 --- Preparation ofWGA-Au --- p.47 / Chapter 2.2.4 --- Preparation of Dil --- p.48 / Chapter 2.2.5 --- Microinjection ofWGA-Au or Dil --- p.48 / Chapter 2.2.6 --- Preparation of rat serum --- p.49 / Chapter 2.2.7 --- Preparation of culture medium --- p.50 / Chapter 2.2.8 --- in vitro whole embryo culture system --- p.50 / Chapter 2.2.9 --- Examination of embryo after culture --- p.51 / Chapter 2.2.10 --- Histological preparation of WGA-Au labelled embryos --- p.51 / Chapter 2.2.11 --- Silver enhancement staining on sections of WGA-Au labelled embryo --- p.52 / Chapter 2.2.12 --- Histological preparation of Dil labelled embryos --- p.53 / Chapter 2.2.13 --- Reconstruction of the mouse embryos --- p.53 / Chapter 2.2.14 --- Cell counting on labelled sacral NCC between the anterior and posterior halves of the somite --- p.54 / Chapter 2.2.15 --- Cell counting on migrating labelled sacral NCC for each somite at different developmental stages --- p.55 / Chapter 2.3 --- Results --- p.57 / Chapter 2.3.1 --- Development of E9.5 mouse embryo in vitro and in vivo --- p.57 / Chapter 2.3.2 --- Labelling of sacral neural crest cells by means of different cell markers --- p.58 / Chapter 2.3.3 --- Migration of sacral neural crest cells at different developmental stages --- p.59 / Chapter 2.3.3.1 --- Distribution of sacral NCC at the 26th somite stage --- p.60 / Chapter 2.3.3.2 --- Distribution of sacral NCC at the 28th somite stage --- p.61 / Chapter 2.3.3.3 --- Distribution of sacral NCC at the 30th somite stage --- p.61 / Chapter 2.3.3.4 --- Distribution of sacral NCC at the 32nd somite stage --- p.63 / Chapter 2.3.3.5 --- Distribution of sacral NCC at the 34th somite stage --- p.64 / Chapter 2.3.4 --- Defined migration pathways of the sacral neural crest cells --- p.65 / Chapter 2.3.5 --- Quantification of migrating sacral NCC at different somite axial levels at different developmental stages --- p.66 / Chapter 2.4 --- Discussion --- p.68 / Chapter 2.4.1 --- E9.5 mouse embryo grew normally in vitro using whole embryo culture --- p.69 / Chapter 2.4.2 --- Migration of sacral neural crest cells at 26th somite stage --- p.70 / Chapter 2.4.3 --- Migration of sacral neural crest cells at 28th somite stage --- p.72 / Chapter 2.4.4 --- Migration or sacral neural crest cells at 30th somite stage --- p.73 / Chapter 2.4.5 --- Migration of sacral neural crest cells at 32nd somite --- p.75 / Chapter 2.4.6 --- Migration of sacral neural crest cells at 34th somite stage --- p.77 / Chapter 2.4.7 --- Majority of sacral neural crest cells migrate along the dorsomedial pathway --- p.80 / Figures and Legends --- p.82 / Tables --- p.136 / Chapter Chapter Three --- The early migratory pathways of Dom mouse sacral neural crest cells --- p.139 / Chapter 3.1 --- Introduction --- p.139 / Chapter 3.2 --- Materials and Methods --- p.145 / Chapter 3.2.1 --- Animals --- p.145 / Chapter 3.2.2 --- In vitro culture of Dom mouse embryos --- p.145 / Chapter 3.2.3 --- Genotyping by polymerase chain reaction (PCR) --- p.146 / Chapter 3.2.4 --- Treatment of the harvested Dom mouse embryos --- p.147 / Chapter 3.2.5 --- Reconstruction of images and cell counting --- p.148 / Chapter 3.2.6 --- Percentage of migrating sacral neural crest cells reduction in Dom mouse embryo --- p.148 / Chapter 3.3 --- Results --- p.150 / Chapter 3.3.1 --- Migration of sacral neural crest cells in Dom mouse embryos at different developmental stages --- p.150 / Chapter 3.3.1.1 --- Distribution of sacral neural crest cells of Dom mouse embryos at the 26th somite stage --- p.150 / Chapter 3.3.1.2 --- Distribution of sacral neural crest cells of Dom mouse embryos at the 28th somite stage --- p.151 / Chapter 3.3.1.3 --- Distribution of sacral neural crest cells of Dom mouse embryos at the 30th somite stage --- p.152 / Chapter 3.3.1.4 --- Distribution of sacral neural crest cells of Dom mouse embryos at the 32nd somite stage --- p.154 / Chapter 3.3.1.5 --- Distribution of sacral neural crest cells of Dom mouse embryos at the 34th somite stage --- p.156 / Chapter 3.3.2 --- Number of migrating sacral NCC of different genotypes of Dom mouse embryos at different developmental stage --- p.158 / Chapter 3.4 --- Discussion --- p.160 / Chapter 3.4.1 --- The use of Dom mouse model to study the etiology of Hirschsprung's disease (HSCR) --- p.161 / Chapter 3.4.2 --- Migration of sacral NCC in Dom mouse embryos --- p.164 / Figures and legends --- p.169 / Tables --- p.230 / Chapter Chapter Four --- General discussion and conclusions --- p.236 / Appendix --- p.241 / References --- p.245

Introduction L’évolution à long terme de la maladie de Hirschsprung (HSCR) est souvent associée à des complications et à un dysfonctionnement de l’intestin, source de conséquences importantes sur la qualité de vie (QdV). L’objectif principal de cette étude est, dans une cohorte québécoise de HSCR, d’étudier la QdV en utilisant un outil spécifique et d’en déterminer les facteurs prédictifs. Méthodes Étude prospective de cohorte et cas-témoins. Les questionnaires 'HAQL' (QdV spécifique de HSCR), 'PedsQL’ (QdV générale), sur le stress et sur la situation sociale ont été administrés, ainsi qu’un journal des selles. Le contenu en cortisol des cheveux (CCC) (mesure du stress chronique) a été quantifié par dosage immuno-enzymatique (ELISA). Résultats 72 patients (72% garçons) et 117 contrôles (65% garçons) ont été analysés. L'âge médian [IQR1-IQR3] à l'inclusion était de 12,1 ans [8-17,5] et 12,6 ans [10,2-15,1] respectivement. La QdV liée à la santé globale était comparable entre les patients et les contrôles. Parmi les patients, la QdV spécifique à la HSCR mesurée par le HAQL était inférieure chez les enfants de 8 à11 ans par rapport aux adolescents de 12 à 16 ans (valeurs moyenne 539,3±66,5/700 vs. 622,6± 54,6/700; p=0,002). La dimension ‘Continence fécale pendant la journée’ était la plus affectée chez les patients âgés de 8 à 11 ans (valeur moyenne 52,6±25,3 sur 100). Chez les patients 12 à 16 ans, la dimension la plus affectée était le ‘Fonctionnement physique’. La prévalence de l'incontinence fécale mesurée chez les patients était de 85% chez les enfants, de 40% chez les adolescents et de 12% chez les adultes. Une association significative entre incontinence fécale et jeune âge était vu (p=<0,0001). Aucune association n'a été démontrée entre incontinence fécale et stress chronique (HCC, événements stressants) ou situation sociale chez les patients. Conclusion L’étude de la QdV par un outil spécifique de la HSCR est nécessaire pour une évaluation adéquate de l’état psychosocial dans cette population qui est à risque d’un dysfonctionnement de l’intestin à long terme. / Introduction Multimorbidity and bowel dysfunction are affecting patients with Hirschsprung disease (HSCR) on long-term follow-up, having an important impact on quality of life (QoL). The primary aim of this study is to evaluate the disease-specific QoL with determination of its predictive factors in a French-Canadian cohort of HSCR patients. Methods Prospective cohort and case-control study. The questionnaires ‘HAQL’ (disease-specific QoL questionnaire), 'PedsQL’ (global health related QoL), ‘Stressful life events’, a questionnaire about the socio-economic state and a stool diary were requested to fill in. Hair cortisol concentration (HCC) (measure of chronic stress) was measured using an enzyme-linked immunosorbent assay kit (ELISA). Results 72 patients (72% males) and 117 controls (65% males) were analyzed. Median [IQR1, IQR3] age at study inclusion was 12.1 years [8,17.5] in patients and 12.6 years [10.2,15.1] in controls. General health related QoL was comparable between patients and controls. In the patient’s group, children 8 to 11 years reported lower disease-specific QoL than adolescents (12 to 16 years) (mean scores 539.3±66.5/700 vs. 622.6±54.6/700; p=0.002). The dimension ‘Fecal continence during daytime’ was the most affected one in children (mean score 52.6± 25.3/100) and the dimension ‘Physical functioning’ was the most affected one in adolescents. Prevalence of fecal incontinence/soiling in the patient’s group was 85% in children, 40% in adolescents and 12% in adults. Younger age was associated with a higher prevalence of fecal incontinence (p=<0,0001). No association was seen between presence of fecal soiling/incontinence and chronic stress (HCC, Stressful live events) or social situation. Conclusion Disease-specific QoL investigation is mandatory in HSCR patients, to encounter and evaluate adequately psychosocial problems related to long term bowel dysfunction.