Title

A Novel Role for Reelin in Coronary Vasculogenesis

Date of Award

7-2019

Degree Type

Thesis

Degree Name

Master of Science in Biomedical Sciences

First Advisor

Cathy Hatcher, PhD

Second Advisor

Ruth Borghaei, PhD

Third Advisor

Heather Montie, PhD

Abstract

Congenital coronary artery anomalies occur in up to 5% of the human population. Such anomalies can lead to myocardial ischemia, myocardial infarction, and sudden death. Vascular repair processes often recapitulate developmental mechanisms. Therefore, elucidation of signaling mechanisms that govern formation of nascent coronary vessels is important to the development of therapeutic strategies to regrow congenitally malformed or damaged coronary vessels. Mice with an epicardial-specific deletion of Tbx5, or Tbx5epi-/- mice, display impaired formation of the coronary vascular plexus, punctate cardiac hemorrhaging, and reduced Reln mRNA expression. The Reln gene encodes the extracellular matrix glycoprotein Reelin. Reelin has been most extensively studied in the brain, where it orchestrates neuronal migration to the cortex and the hippocampus. However, immunostaining of wild type mouse hearts revealed that Reelin is expressed in endothelial cells (ECs) of some nascent coronary vessels in the embryonic mouse heart. In addition, Reelin has also been linked to numerous other biological processes, including formation of lymphatic vessels.

Study Objective: The aim of this study was to clarify the contributions of Reelin to the formation of the coronary vasculature with particular focus on the contributions of Reelin to the endothelial cell barrier.

Methods: First, an in vitro cell membrane permeability assay protocol was established with human dermal microvascular endothelial cells (HDMECs) to model formation of the endothelial cell barrier in coronary vessels. Second, HDMECs transfected with RELN siRNA or a non-targeting control siRNA or were utilized in the newly established cell membrane permeability assay protocol to explore the functional contributions of Reelin to the permeability of the coronary vasculature.

Results: A protocol for the in vitro examination of vascular endothelial cell monolayer permeability was established. Permeability assays were performed using RELN siRNAtransfected cells and non-targeting control siRNA-transfected cells. The results indicated a role for Reelin in increasing vascular endothelial cell membrane permeability.

Conclusions: An in vitro cell membrane permeability assay was established and will likely be a valuable tool for future laboratory applications. The results of this research project revealed possible novel roles for Reelin in increasing vascular endothelial cell membrane permeability and maintaining typical endothelial cell morphology.

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