Neural Crest Cell Development in the Nervous System of Normal Gut and in Hirschsprung's Disease

This dissertation, "Neural Crest Cell Development in the Nervous System of Normal Gut and in Hirschsprung's Disease" by Ming, Fu, 付明, was obtained from The University of Hong Kong (Pokfulam, Hong Kong) and is being sold pursuant to Creative Commons: Attribution 3.0 Hong Kong License. The content of this dissertation has not been altered in any way. We have altered the formatting in order to facilitate the ease of printing and reading of the dissertation. All rights not granted by the above license are retained by the author. Abstract: Abstract of thesis entitled Neural Crest Cell Development in the Nervous System of Normal Gut and in Hirschsprung's Disease Submitted by Ming FU For the degree of Doctor of Philosophy at the University of Hong Kong in November 2003 The enteric nervous system (ENS) controls the peristalsis and endocrine functions of the gut. At early embryonic stage neural crest cells (NCCs) from the neural tube migrate and colonize the entire gut, proliferating and differentiating into neurons and glia that coalesce into neural plexuses at specific regions as the smooth muscle of the gut differentiates. Perturbations of these developmental processes can result in Hirschsprung's disease (HSCR) in newborn children. Studies in animal models have identified signaling pathways for ENS development, but parallel studies in humans are lacking. The aim of this study was to understand the molecular and cellular basis of ENS development and to define the molecular defects characteristic of HSCR. Specific markers for NCCs, neurons, glia and smooth muscle were used, and it was found that the differentiation of both the smooth muscle of the gut and the neural plexuses followed a rostro-caudal manner. The neural plexuses also differentiated centripetally: those between the outer longitudinal and inner circular smooth muscles differentiated before those on the lumenal side. This developmental pattern in humans is similar to that in mice, but different from that in chicks. The temporal and spatial expression patterns of HOXB5 in human embryos were found to correlate with the migration of NCCs, and the differentiation of enteric neuroblasts and gut mesenchyme. Gut mesenchyme and NCCs express HOXB5, and HOXB5 expression was switched off in the mesenchyme and NCCs once these cells differentiated. These findings suggest that HOXB5 plays a role in ENS development and also contributes to the topographic patterning of the gut mesenchyme. Gene expression study showed that NCCs, glia and neurons in human embryonic and normal infant guts expressed SOX10. In HSCR gut SOX10 transcripts were restricted to the hypertrophic nerve trunk. To analyze the effect of Sox10 mutation on the migration of the NCCs, ex vivo gut explants were set up on a natural Sox10 mutant mouse (Dom). The results suggested that Sox10 may modulate the responsiveness of the NCCs to mesenchyme-derived factor GDNF (glia cell-line derived neurotrophic factor), and that "defective" responsiveness of NCCs to GDNF inductions may contribute to the HSCR phenotype in the Dom mice. Using in vitro NCC culture and ex vivo gut explants, it was shown that the endoderm-derived morphogen sonic hedgehog (Shh) promotes the proliferation and aggregation of enteric NCCs, restricts the migration of NCCs, and overrides the neuronal induction effect of GDNF and BMP2 (bone morphogenetic protein 2). Present study suggests that Shh regulates the topographic patterning of the ENS by modulating the responsiveness of NCCs to mesenchyme- derived factors, and that in its presence the pool of NCCs can expand and be maintained in long-term neurospheres without loosing their stem cell properties. The potential for applying Shh in the expansion of NCCs and in stem cell therapy for HSCR and other NCC-associated gut anomalies deserves further investigation. In conclusion, the findings of this study suggest that genes that encode components affecting the cell-autonomo