Exploration of Decellularized Vegetal Scaffolds as New Biomaterials for Tissue Engineering

In tissue engineering (TE), biomaterials are critical to guiding cell growth and tissue regeneration. With the shortage of available human donors to fill the growing need, alternative options from both synthetic and natural sources have been explored. However, a gold standard approach has yet to emerge and new sources are sought. Recently, scaffolds resulting from the decellularization of vegetal tissue have been investigated to provide a biocompatible, easy to handle and sustainable alternative to current TE constructs. However, the exploration of these biomaterials is still in the nascent stage and the goal of this research project is to advance their characterization. Specifically, we first investigated a new decellularization method to replace the time-consuming and damaging conventional serial chemical treatment. We thus showed that the use of supercritical carbon dioxide in combination with 2% peracetic acid is more efficient timewise while maintaining the scaffold’s structural, mechanical and biochemical properties and cell-supporting capabilities. Second, we explored the mechanical properties of these scaffolds. We demonstrated that the stiffness of decellularized vegetal tissues matches the range of stiffness of human tissues while their elasticity under cyclic strain behaves as heterogeneously as mammalian tissue. Interestingly, we also showed that cells seeded on the scaffolds can sense these mechanical cues and respond through mechanotransduction pathways such as YAP/TAZ signaling leading to altered cellular proliferation, morphology and metabolism. Third, we extended the portfolio of plant-based scaffolds by investigating their ability to be used as a drug delivery system. Using cannabidiol in an exploratory study, we showed that scaffolds could adsorb 70% of the drug within 24h and release up to 12% within the next 24h. Based on these results, we will evaluate the capacity of scaffolds with different biochemical (polymer composition) and physical (hydrophilicity, porosity) properties for the retention/release of several classes of drugs. Fourth, we developed a new approach to overcome the current challenge of repopulating the natural vein network of leaves with human endothelial cells in order to vascularize the future constructs. By controlling hydration level and capillarity effect, we were able to move fluid into the entire length and ramification of the network and advance cells into secondary, micron-sized capillaries located at long distances from the injection site and the main stem. Fifth, to demonstrate the clinical potential of these findings, we showed that decellularized plant-based materials could be assembled to generate a multi-layered “skin-on-leaf” system with similar mechanical properties and cellular composition to human skin or be used as a substitute scaffolding for peripheral nerve growth and replaced current animal templates. Finally, in the last stage of this work, we initiated a new investigation using plant-derived extracellular vesicles (pEV) isolated from fresh vegetal sources to serve as nanosystems for drug delivery to work in complement with the decellularized scaffolds. We showed that pEV from spinach leaves, turmeric roots and olives could be reproducibly isolated by rounds of increasing centrifugation speed and purified by either size-exclusion chromatography or ultracentrifugation with a sucrose gradient. These pEVs also showed biocompatibility and bioactivity, through their radiation modulation effect, when incubated with human cells exposed to radiation, thus suggesting their potential as therapeutic target. Altogether, this data advanced our knowledge on decellularized plant-based scaffolds by establishing new techniques for their fabrication and maturation, and by showing their physiological relevance. This data showed the large diversity of potential translational applications of these biomaterials (and the plant kingdom at large) and confirmed their promising interest for biomedical and clinical research.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

doctoral

Discipline thesis:degree_discipline

Graduate College

Grantor dc:publisher

The University of Arizona.

Year dc:date.issued

2022

Author dc:creator

• Harris, Ashlee Frances

Advisor dc:contributor.advisor

• Zenhausern, Frederic

Committee members dc:contributor.committeemember

• Lacombe, Jerome
• Yoon, Jeong-Yeol
• Kim, Minkyu

Statement dc:rights

• Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction, presentation (such as public display or performance) of protected items is prohibited except with permission of the author.

Licence dc:rights.uri

• http://rightsstatements.org/vocab/InC/1.0/

Language dc:language.iso

en

Handle dc:identifier.uri

http://hdl.handle.net/10150/668067

OAI identifier oai:identifier

oai:repository.arizona.edu:10150/668067