Author ORCID Identifier

https://orcid.org/0000-0003-0945-3631

Semester

Fall

Date of Graduation

2023

Document Type

Dissertation

Degree Type

PhD

College

School of Medicine

Department

Biochemistry

Committee Chair

Mark Tseytlin

Committee Co-Chair

Timothy Eubank

Committee Member

Matthew Dietz

Committee Member

Michael Gunther

Committee Member

Ming Pei

Committee Member

Elena Pugacheva

Abstract

Electron paramagnetic resonance (EPR), and its molecular imaging modality, is a powerful tool to noninvasively map various biological and chemical markers within objects of interest. Reliable data acquisition is a major impeding factor for longitudinal hands-off measurements. Measurements are especially challenging in biomedical applications, as live objects are not static. Frequent changes occur that require constant fine recalibration of the EPR detection system, called the resonator. To enable longitudinal imaging, a technology permitting automatic digital control of resonator coupling, tuning, and EPR data acquisition was developed. Automation was achieved through the utilization of a microcontroller and digital peripheral components such as digitally controlled capacitors, a digital frequency source, and a printed circuit board resonator. Several applications of this technology have been suggested and tested, including in vivo EPR imaging. The first was to develop a tool for the optimization of light-based 3D printing, for which oxygen plays a major role. Towards this goal, an EPR oxygen-sensitive probe was incorporated into 3D printing resin. Oxygen depletion was measured during the 3D printing process as the polymerization front progressed. After printing, oxygen depletion was again measured during the post-curing process, proposed as a method to optimize post-curing light intensity, temperature, and duration in order to produce quality 3D printed constructs. The second application of longitudinal EPR imaging was directed toward resolving an important problem of oxygen delivery to thick (>1 cm) bioprinted models. Oxygen-sensitive EPR probes (water-soluble trityls or crystalline lithium octa-n-butoxynaphthalocyanine) were introduced into bioinks (liquid hydrogels containing cells, nutrients, and other biological factors) before printing. Bioinks become solid structures after printing due to crosslinking. EPR imaging was demonstrated to measure oxygen consumption by the cells embedded in the bioprints. As expected, an increase of oxygen depletion was observed by introducing a nutrient (pyruvate) to bioink. A numerical MATLAB simulation program was developed to predict rates of oxygen consumption by the cells in the bioprint. The input parameters for the mathematical model include the size and number of cells, the diffusion coefficient of the media, and rates of oxygen transfer through the cell membrane. The software is being further refined and optimized for computational speed. Future efforts will be aimed at improving the speed and scope of EPR automatic digital control, imaging the oxygen depletion process in commercial 3D printers, and applying EPR mapping of oxygen consumption rate to quantify the delivery of oxygen to cells deep inside bioprinted tissue models. Optimizing the delivery of oxygen to cells would overcome the challenge of the limit of diffusion, facilitating the development of larger and more complex bioprinted tissues and organs. These complex tissue and organ models are envisioned for use in drug testing, biomedical research, and, in the distant future, implants in humans.

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