The work of the Cal Poly Tissue Engineering Lab is primarily focused on the fabrication, characterization, and improvement of “Blood Vessel Mimics” (BVMs), tissue engineered constructs used to evaluate cellular response to vascular medical devices. Currently, cells are grown onto fibrous, porous tubes made using an in-house electrospinning process from PLGA, a biocompatible co-polymer. The adhesion and proliferation of cells in a BVM is reliant on the micro-scale structure of the PLGA scaffold, and as such it is of great importance for the electrospinning process to consistently produce scaffolds of similar morphologies. Additionally, it has been shown that cell proliferation increases with scaffolds of smaller fibers and pores than the current electrospinning protocol can produce. Finally, the Tissue Engineering Lab has interest in testing devices in more tortuous BVM bioreactor designs, however the use of relatively rigid PLGA scaffolds has severely limited the ability to construct more complicated vessel geometries.
The overall goal of this thesis was to improve fabrication and characterization of electrospun polymer scaffolds for BVM use. The specific aims of this thesis were to: 1) Improve scaffold characterization by comparing two techniques for fiber diameter measurement and implementing a technique for pore area measurement. 2) Reduce scaffold fiber diameter and pore area by investigating humidity and solvent composition electrospinning parameters. 3) Reduce process variability by developing a more specific electrospinning protocol. 4) Improve scaffold consistency and use by understanding and reducing PLGA scaffold shrinkage. 5) Identify and evaluate more flexible polymers as potential alternatives for electrospun BVM scaffolds.
In order to accomplish these aims, first, several BVM and outside literature images were taken and evaluated with current and prospective fiber diameter techniques, and with 2 prospective pore area techniques to characterize accuracy and consistency of each method. It was found that the prospective fiber diameter measurement technique was not superior to the current method. The techniques developed for pore area measurement were found to produce results that differed significantly from each other and from the published value for a given image. Next, changes to environmental and solution composition parameters were made with the hopes of reducing fiber diameter and pore area of electrospun PLGA scaffolds. Changes in relative humidity did not appear to significantly affect scaffold fiber diameter while changes to solvent composition, specifically the use of acetone, resulted in fibers significantly smaller than those regularly achieved in the BVM lab. Next, several sources of variability in the electrospinning protocol were identified and subsequently altered to improve consistency and usability. Specifically, this included redefining the precision with which PLGA mass was measured, repositioning electrical equipment to reduce the effect of stray electrostatic forces on the polymer solution jet, attempting to control the temperature and humidity inside the electrospinning enclosure, and improving the ease with which scaffolds are removed from their mandrels through alternative mandrel surface treatments. In addition to overall process variability, the issue of scaffold shrinkage during BVM use was investigated and two possible treatments, exposure to either ethanol or elevated temperatures, were proposed based on previous electrospinning literature results. Each was tested for their effectiveness in mitigating shrinkage through exposure to BVM setup-mimicking conditions. It was found that both treatments reduced scaffold shrinkage compared to control samples when exposed to BVM setup-mimicking conditions. Finally, 3 flexible polymers were selected and electrospun to compare against typical PLGA results and to conduct a kink radius test as a metric for measuring flexibility as it pertains to the proposed BVM lab application. It was concluded that two types of thermoplastic polyurethane (tPU) were not acceptable electrospinning materials for use in the BVM lab. Additionally, while polycaprolactone (PCL) could be successfully electrospun it could not undergo the amount bending required for more tortuous BVM bioreactor designs without kinking.
Overall, the work in this thesis provided insight into multiple scaffold characterization techniques, reduced overall electrospinning variability in the fabrication and use of PLGA scaffolds, and defined processing parameters that have been shown to yield scaffolds with smaller morphological features than all prior Tissue Engineering Lab work. By creating better, more effective scaffolds, researchers in the Tissue Engineering Lab can more accurately mimic the structure and properties of native blood vessels; this, in turn, will result in BVM cell responses that more closely resemble that of native tissue. Creating consistent and appropriate BVMs will then lead to impactful contributions to the existing body of tissue engineering research and to better preclinical device testing.
| Identifer | oai:union.ndltd.org:CALPOLY/oai:digitalcommons.calpoly.edu:theses-3029 |
| Date | 01 September 2017 |
| Creators | Dowey, Evan M |
| Publisher | DigitalCommons@CalPoly |
| Source Sets | California Polytechnic State University |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Master's Theses |
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