Neural crest cells form during early vertebrate development and migrate throughout the embryo, laying the framework for a diverse array of structures. For well over a century, embryologists have investigated these cells. Researchers have tracked neural crest migration patterns, defined neural crest derivatives, and – with the advent of molecular biology techniques and mutational assays – have begun to determine which genes are involved in neural crest formation. From these studies, we have recognized that components of the peripheral nervous system, craniofacial structures, and pigment cells can all be counted among the derivatives of the neural crest. With the realization that the neural crest can go on to form such a myriad of structures, scientists have come to understand that defects in neural crest development are responsible for several human diseases. Cancers of the peripheral nervous system, craniofacial malformations, and melanoma have all been characterized as neural crest-associated pathologies. As we gain insight into what genes are involved in neural crest development, we are beginning to understand how neural crest-associated diseases manifest and progress.
Despite over a century of study on neural crest development, early stages of mammalian neural crest formation – especially that of the human neural crest – remain largely unexplored. Human neural crest development is remarkably difficult to assess, given the limited availability of tissue and underlying ethical concerns. To date, no study has comprehensively assessed the in vivo expression patterns of multiple neural crest markers in human embryos when neural crest migration is underway. Research in model organisms can help to supplement the gaps in our knowledge of human neural crest formation; however, extrapolating from model organisms to humans carries with it a particular set of challenges. Insights into neural crest development gained from the frog, chick and mouse may not always translate directly to the human. Furthermore, early neural crest formation is difficult to investigate in the mouse (the standard mammalian model system) given its small embryonic size and unique morphology. Thus, mammalian neural crest development – specifically at its earliest phases – is still not well understood.
The over-arching goal of this work is to expand our knowledge of the mammalian, and ultimately human, neural crest. In Chapter 2, we describe the expression patterns of several neural crest markers in human embryos between Carnegie Stages 12 and 18. The expression of many of these markers has never before been reported in early human embryos. Importantly, we note the in vivo expression of the neural crest-associated transcription factors Pax3, Sox9, Sox10, p75NTR, Pax7 and AP-2α in either the premigratory or migratory neural crest at Carnegie Stage 12 and/or 13. We also comment on marker expression in neural crest derivatives at these and older stages, postulating on the roles these factors play in human pathologies. Finally, we note that markers that have classically been used to identify human neural crest cells in vitro are actually widely expressed throughout the human embryo (p75NTR), or do not appear to be specific for the human neural crest (HNK-1).
In Chapter 3, we investigate the early stages of mammalian neural crest development by establishing the rabbit as a new model system for this cell population. Given the difficulties in assessing murine neural crest formation, paired with the limited availability of human embryonic tissue, we turned to the rabbit in the hopes of better understanding all stages of mammalian neural crest formation. Here, we provide the first analysis of neural crest marker expression in rabbit embryos, and further compare expression patterns between mouse, chick, rabbit and human embryos. Similar to other model systems, Pax3, Pax7, Sox9, Sox10, AP-2α, and Msx1/2 are all expressed in presumptive rabbit neural crest cells at some stage. Importantly, we provide the first evidence of neural crest specification in a mammalian system. Explants derived from specific regions of the rabbit gastrula give rise to presumptive neural crest cells, as evidenced by the expression of Pax7 and/or Sox10. Furthermore, the expression of these markers appears largely independent of the mesodermal marker Brachyury. Additionally, we also demonstrate that rabbit whole mount embryos cultured in vitro display normal neural crest development, and touch on some of our preliminary success with rabbit embryo electroporations. Collectively, this work provides new insights into the development of the mammalian neural crest, and establishes the rabbit as a fantastic model system for future work evaluating these cells.
|Commitee:||Holley, Scott, Sakkas, Denny, Slack, Frank|
|Department:||Molecular, Cellular, and Developmental Biology|
|School Location:||United States -- Connecticut|
|Source:||DAI-B 74/05(E), Dissertation Abstracts International|
|Subjects:||Neurosciences, Cellular biology, Developmental biology|
|Keywords:||Embryo, Neural crest, Pigment cells, Rabbit, Specification|
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