Date Approved

8-19-2022

Embargo Period

8-22-2022

Document Type

Dissertation

Degree Name

Ph.D. Doctor of Philosophy

Department

Biomedical Engineering

College

Henry M. Rowan College of Engineering

Advisor

Peter A. Galie, Ph.D.

Committee Member 1

Mary Staehle, Ph.D.

Committee Member 2

Erik Brewer, Ph.D.

Committee Member 3

Sebastian Vega, Ph.D.

Committee Member 4

Allison Andrews, Ph.D.

Keywords

Blood-Brain Barrier, Mechanotransduction, Shear Stress, Small GTPases, Ischemic stroke

Subject(s)

Cerebrovascular disease

Disciplines

Biomedical Engineering and Bioengineering

Abstract

Fluid shear stress is an important mediator of vascular permeability, yet the molecular mechanisms underlying the effect of shear on the blood-brain barrier (BBB) have yet to be clarified in cerebral vasculature despite its importance for brain homeostasis.. Neurological symptoms including the formation of microclots, stroke, and other neurological pathologies associated with changes in cerebral blood flow are hallmarks of BBB dysfunction. The in vitro model used in this dissertation is compatible with real-time measurement of barrier function using a transendothelial electrical resistance as well as immunocytochemistry and dextran permeability assays. These experiments reveal that there is a threshold level of shear stress required for barrier formation and that the composition of the extracellular matrix, specifically the presence of high molecular weight hyaluronan, dictates the flow response. Gene editing to modulate the expression of CD44, a mechanosensitive receptor for hyaluronan, demonstrates that the receptor is required for the endothelial response to shear stress. Manipulation of small GTPase activity reveals CD44 activates Rac1 while inhibiting RhoA activation. Additionally, adducin-gamma localizes to tight junctions in response to shear stress and RhoA inhibition and is required to maintain the barrier. This dissertation identifies specific components of the mechanosensing complex associated with the BBB response to fluid shear stress and, therefore, illuminates potential targets for barrier manipulation in vivo.

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