Understanding Mechanisms Driving Cranial Mesenchyme Morphogenesis and Neural Fold Elevation During Normal and Abnormal Neural Tube Closure
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Neural tube closure allows for the development of the central nervous system—the brain and spinal cord. Disruption in this process leads to neural tube defects (NTDs). NTDs can be fatal or lead to long-term disability and are one of the most common structural birth defects. Morphogenesis within the neural plate, which rolls up into the neural tube, is well-studied. Yet, understanding the role of the underlying cranial mesenchyme (CM) and the cranial mesenchymal surrounding extracellular matrix (CM-ECM) is limited. Thus, the overarching goal of this dissertation is to elucidate mechanisms involving the CM morphogenesis and the role of the surrounding CM-ECM during neural tube fold elevation and disrupted closure leading to NTDs. The CM is derived from paraxial mesoderm (PM-CM) and neural crest (NC-CM) derived cells, and both are implicated in NTDs. In Chapter 2, using bulk-RNA sequencing, changes in the expression of genes encoding the CM-ECM within the CM were analyzed in a time-series experiment in wild-type embryos. The analysis demonstrated changes in CM-ECM composition as the neural folds elevated. Specifically, the expression of fibronectin and other ECM components change as the neural folds elevate, presumably altering the properties of the CM needed to support neural fold elevation. In Chapter 3, the Hectd1 mouse model of NTDs with failure of neural fold elevation and CM morphogenesis was used to elucidate how disruption of CM morphogenesis disrupts neural fold elevation. Using RNA sequencing, functional assays, and epistasis analysis, experiments tested how the secretion of extracellular heat shock protein 90 (eHSP90) and ECM changes altered the interaction of PM-CM and the NC-CM cells, resulting in abnormal CM behavior disrupting CM expansion and neural fold elevation. The results demonstrate that increased eHSP90 from NC-CM cells causes excess migration in the Hectd1 mouse model, partly by stabilizing excess fibronectin surrounding both NC-CM and PM-CM cells. Interestingly, a loss of HSP90 or reduced Fibronectin 1 (Fn1) gene dosage in Fn1-/+ embryo partially rescued NTDs in the Hectd1 model. Finally, Chapter 4 explores the novel insights gained through time-lapse live imaging experiments of neural tube closure that observed cell behavior in real time during neural tube formation. Together, the research presented here provides a better understanding of the critical role of normal cranial mesenchyme morphogenesis and extracellular matrix changes for proper neural fold elevation. Moreover, the research explains how disruption in these processes caused by the activation of aberrant mechanisms involving eHSP90 and fibronectin can lead to NTDs in the Hectd1 mouse model.
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