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The role of endothelial cells during lung organogenesis

Havrilak, Jamie Ann
http://rave.ohiolink.edu/etdc/view?acc_num=ucin1428065611

Abstract Details

2015, PhD, University of Cincinnati, Medicine: Molecular and Developmental Biology.
Early respiratory development begins with the specification of the lung cell fate, followed by the emergence of lung buds from the ventral foregut endoderm. The respiratory tree is then elaborated through branching morphogenesis, and proliferation and differentiation of specialized cell types along the proximal-distal axis produces airways and alveoli, respectively. Lung development is orchestrated through crosstalk between the epithelium and mesenchyme; morphogenesis and differentiation of the lung relies on diffusible signaling molecules that cross tissue layers to activate a complex network of transcription factors to drive development. Significant progress has been made in understanding the molecular regulation of lung morphogenesis through paracrine signaling pathways and transcription factors essential for lung development. The lung mesenchyme is comprised of multiple cell types, however, and which of the mesenchymal cell types are producing these molecules that drive epithelial morphogenesis remains unknown. To determine if endothelial cells are required for epithelial branching morphogenesis we cultured E12.5 lung explants in the presence of 3 different VEGFR inhibitors, which and found epithelial branching was not repressed. We additionally recombined isolated E12.5 lung epithelium (LgE) with mesenchyme from which the endothelial cells had been removed, and found that LgE still branched when cultured with the endothelial cell-depleted mesenchyme. Additionally, we found that LgE did not branch when cultured with human lung microvascular endothelial cells or mouse primary endothelial cells. These data demonstrate that endothelial cells are neither necessary nor sufficient to induce epithelial branching morphogenesis in vitro. To determine if endothelial cells are required for respiratory specification and lung bud initiation, we examined lung formation in E8.5 wild-type embryos cultured in the presence of VEGFR inhibitors and found that specification still occurred when endothelial cell signaling was hindered. However, several caveats to these experiments led us to further investigate this process by utilizing a genetic mouse model in which embryos do not develop a vascular system due to a mutation in the VEGF receptor Flk-1. Due to the decreased size and ultimate embryonic lethality as a result of the Flk-1 mutation, we conducted in vitro foregut culture experiments at E8.5, when wild-type and Flk-1-/- mutant embryos were indistinguishable. As visualized by whole mount confocal microscopy, we found that cultured E8.5 Flk-1-/- mutants expressed Nkx2.1 (a marker for respiratory progenitor cells) in the prospective lung field endoderm to the same extent as wild type littermates, and initiated lung bud formation (measured by SP-C expression), demonstrating that endothelial cells are not necessary for lung specification and bud initiation in vitro. These data collectively suggest that early events in lung organogenesis, such as specification, bud initiation, and terminal branching are not dependent on endothelial cells.
John Shannon, Ph.D. (Committee Chair)
James Lessard, Ph.D. (Committee Member)
Robert Brackenbury, Ph.D. (Committee Member)
Timothy Lecras, Ph.D. (Committee Member)
James Wells, Ph.D. (Committee Member)
159 p.
Developmental Biology
Lung; Tissue interactions; Branching morphogenesis; Endothelial cells; Specification

Recommended Citations

Citations

  • Havrilak, J. A. (2015). The role of endothelial cells during lung organogenesis [Doctoral dissertation, University of Cincinnati]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1428065611

    APA Style (7th edition)

  • Havrilak, Jamie. The role of endothelial cells during lung organogenesis. 2015. University of Cincinnati, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=ucin1428065611.

    MLA Style (8th edition)

  • Havrilak, Jamie. "The role of endothelial cells during lung organogenesis." Doctoral dissertation, University of Cincinnati, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1428065611

    Chicago Manual of Style (17th edition)

ucin1428065611
241
© 2015, some rights reserved.Creative Commons License The role of endothelial cells during lung organogenesis by Jamie Ann Havrilak is licensed under a Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License. Based on a work at etd.ohiolink.edu.
This open access ETD is published by University of Cincinnati and OhioLINK.