Date of Award

1-1-2016

Language

English

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

College/School/Department

Department of Biological Sciences

Content Description

1 online resource (ii, viii, 201 pages) : illustrations (some color)

Dissertation/Thesis Chair

Melinda Larsen

Committee Members

Guohao Dai, Prashanth Rangan, Paolo Forni

Keywords

Angiocrine factor, Endothelial precursors, Epithelial cell differentiation, Epithelial patterning, Ex vivo tissue reconstitution, Salivary gland development, Endothelial cells, Cell differentiation, Vascular endothelial growth factors, Submandibular gland, Salivary glands

Subject Categories

Bioinformatics | Developmental Biology

Abstract

Functional vasculature regulates organ formation and regeneration. Perfusion-independent and nutrition-independent endothelial regulation of epithelial patterning is of considerable interest for application in restoring parenchymal function. During murine submandibular salivary gland development, the vasculature co-develops with the epithelium; however, it is still unknown if the vasculature has instructive effects on the epithelium. A systems biological study revealed that endothelial genes are enriched in epithelial clefts, suggesting a potential involvement of endothelial cells in early morphogenesis. Using immunodepletion of the endothelial cells from the mesenchyme of reconstituted glands, a requirement for endothelial cells in epithelial patterning was demonstrated. Specifically, lobule formation was disrupted with enlargement of the endbuds. Pharmacological inhibition and siRNA genetic knockdown phenocopied the lobule disruption identified with endothelial deficiecy. In addition, it suggested that VEGFR2/Kdr-dependent signaling is required for epithelial patterning. Addition of exogenous endothelial cells to reconstituted glands restored defective epithelial patterning restulting from endothelial depletion. I further asked what endothelial factors are involved in regulation of epithelial progenitors since VEGFR2-inhibition promoted ductalization, and endothelial cell-depletion increased cleft widening, suggesting that acinar and ductal differentiation are regulated by endothelial cells. Several endothelial soluble factors were identified from cultures with embryonic mesenchyme. Interestingly, IGFBP2 was identified as a putative endothelial-dependent angiocrine effector that regulates salivary gland epithelial patterning, showing partial restoration of epithelial defects with endothelial depletion and VEGFR2 inhibition. Our results indicate that endothelial cells are required for epithelial patterning in the salivary gland. Taken together, these observations indicate that VEGFR2+ endothelial cells in early developing salivary glands produce angiocrine factors that regulate epithelial organ pattern formation and differentiation. The results provided in this dissertation highly suggest that endothelial cells and signaling are necessary for submandibular salivary gland organ development and epithelial differentiation. Furthermore, submandibular salivary gland endothelial cell isolation, in vitro culture, and lentiviral manipulation procedures that have been established in this study will provide a platform to further elucidate mechansms of endothelial regulation of the epithelium.

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