Date of Award

Spring 5-2021

Document Type

Dissertation

Degree Name

Doctor of Engineering (DEng)

Department

Biomedical Engineering

First Advisor

Mark A. DeCoster

Abstract

The brain accounts for 20% of overall energy metabolism in the body though it just comprises 2% of the total body mass but has a limited capacity of storing energy unlike other critical organs in the body such as the heart and liver. This energy along with oxygen and nutrients is supplied by cerebral blood flow (CBF), any interruption of which can cease the brain function within seconds with a potential irreversible neuronal injury, within minutes. Vascular cells along with astrocytes and neurons are a part of a recently developed concept known as the Neurovascular Unit responsible for Neurovascular coupling (NVC), a phenomenon whereby an increase in neuronal activity leads to elevation of local CBF. The blood-brain barrier(BBB) chiefly composed of the vascular cells and astrocytes comprises the major role in NVC and thus is central to understanding brain functions and disorders.

Nitric oxide (NO) is a calcium-dependent vasoactive mediator that heavily influences NVC while intracellular calcium (Ca++) is a second messenger that serves for complex signaling roles in the brain. Impairment in the homeostasis of these signals can lead to extreme functional alterations in the brain. Furthermore, both NO and Ca++ changes have been shown to affect BBB permeability which is a common feature observed in many neurological disorders. Therefore, this research investigates the alteration of NO and Ca++ signals in the rat brain microvascular endothelial cells (BMVECs) and astrocytes, the two major cell types of BBB during inflammation, and further evaluates the effect of cellular interaction between them in modulating these signals by developing their co-culture model. It focuses on phenotypic and biochemical changes in BBB due to inflammation, as inflammation related failure of BBB is implicated in the initiation or progression of a wide variety of neurological disorders. Both NO and Ca++ were found to increase excessively in BMVECs unlike the astrocytes during inflammation, and in the co-culture model, the presence of astrocytes was found to provide negative feedback to these elevated NO and Ca++signals from BMVECs.

Considering the dominant role of NO and endothelial cells in NVC, this research investigates a potential application of a novel material, copper -cystine biohybrid known as Copper High Aspect Ratio Structures (CuHARS) in catalyzing NO in BMVECs in normal as well as in induced inflammatory conditions. CuHARS was able to increase the NO concentration in normal BMVECs by stimulating endothelial nitric oxide synthase (eNOS) activity, known to have neuroprotective roles besides inhibiting the usually harmful NO synthesis in an inflammatory condition. Thus, CuHARS displays potential in therapeutic applications for NO related NVC disorders in addition to the antimicrobial and wound healing applications studied earlier.

The tight junctions, unique to the endothelial lining of the brain, limit the entry of drugs against neurological and mental disorders from reaching into the brain. This poses an immense challenge in developing treatments for brain disorders. This research explores the naturally available Halloysite nanotubes (HNTs) for drug loading and delivery across the BBB. HNTs loaded with a fluorescent dye (RITC) and ionomycin separately were tested on BMVECs to study periodic and real-time attributes of HNTs in drug delivery. HNTs were found to deliver the payload gradually over an extended period to the BMVECs cells.

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