Date of Award

12-2024

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biomedical Engineering and Sciences

First Advisor

Christopher A. Bashur

Second Advisor

Venkat Keshav Chivukula

Third Advisor

Siddhartha Bhattacharyya

Fourth Advisor

Vipuil Kishore

Abstract

Vascular cognitive impairment and dementia (VCID) is one of the main causes of dementia. VCID is caused by age and other genetic factors that lead to disruptions in the blood brain barrier (BBB) and allow blood components to enter the brain. The currently available treatments of VCID include opening the BBB with high-frequency-ultrasound to deliver drugs to the brain. However, this only treats the already developed disease. Consequently, we developed a new drug delivery system whose goal is to restore the brain capillary endothelial cells (BCEC) lining the blood vessel. The objective of this study was to demonstrate the feasibility of a drug delivery system using microbubbles (MBs) as a carrier for incorporating carbon monoxide (CO) and providing enough CO delivery that could induce BCECs growth to accomplish our goal which is to restore the BBB. To achieve our objective, we completed two studies. First, we investigated in-silico the impact of CO released from fully-ruptured-MBs on BCECs. Second, we investigated in-vitro the impact of ultrasound on the MBs and in-silico the trajectory of unruptured-MBs under ultrasound and blood flow forces, and the amount available to be endocytosed by BCECs.

In the first study, we considered ultrasound-rupture of all of the MBs away from BBB to avoid causing any BBB damage, and we computationally modeled the concentration reaching the BBB with pulsatile blood flow, gravity, lift, drag forces, and binding kinetics of CO with hemoglobin (Hb). While this study did not involve experimental analysis, the simulations considered perfluoropentane-based MBs loaded with CO that we have synthesized previously. The CO and Hb binding kinetic without blood flow showed that they do not bind very quickly and the percentage of released CO that reaches the BBB is high enough to impact the BCECs. With pulsatile blood flow, the binding kinetics is even slower. Furthermore, we determined that potentially beneficial doses are well within the safety threshold. Finally, the concentration of available CO at the BBB was investigated with pulsatile blood flow for both a simple tube and a patient specific geometry. Overall, this study showed feasibility of CO gas delivery from MBs ruptured outside the BBB, including concentrations at the BBB wall that are both non-toxic and above levels that are required to impact BCECs.

In the second study, we considered any CO-loaded MBs that are not ruptured by the ultrasound and their ability to travel near the wall of the BBB where they can be endocytosed by BCECs. Experimentally, we designed a hydrogel-based phantom with an embedded 3 mm diameter straw, and the results showed increased rupture, displacement, and speed as we increase ultrasound intensities from 10 to 100%. We also verified that MBs can be endocytosis by culturing GFP+ human umbilical ECs (HUVECs) and imaging with time-lapse, confocal microscopy. Our computational results showed that the MBs primarily moved in the direction of blood flow with limited impact of the direction of gravity with a simple tube geometry, but the direction of gravity had more of an effect for a patient specific geometry. Interestingly, ultrasound application caused the MBs to move in the direction of the sound waves, and the speed and distance of movement depended on both the frequency of the ultrasound and the size of the particles. When both ultrasound and blood flow are considered together, the blood flow component was most important. However, movement of MBs towards the vessel wall due to ultrasound did result in a lower number of MBs available at the BBB wall. Overall, this study showed that the interaction of ultrasound with MBs is also an important consideration when determining the dose of CO that would be available to the BCECs at the BBB.

Overall, this study showed that the novel delivery strategy of CO to repair the BBB via ultrasound sensitive microbubbles is feasible and should be further investigated in vivo. The CO will be delivered to the BCECs both from CO released by ruptured microbubbles and unruptured microbubbles endocytosed by BCECs at the BBB at doses that can be beneficial.

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