DOI
https://doi.org/10.25772/XWW7-6A25
Defense Date
2023
Document Type
Dissertation
Degree Name
Doctor of Philosophy
Department
Biomedical Engineering
First Advisor
Dr. Jennifer Puetzer
Second Advisor
Dr. Henry Donahue
Third Advisor
Dr. Rene Olivares-Navarrete
Fourth Advisor
Dr. Lesley Chow
Fifth Advisor
Dr. Seth Cheatham
Abstract
The anterior cruciate ligament (ACL) connects to bone via structurally complex attachments known as entheses. Entheses are composed of gradients in organization, composition, mineralization, and cell phenotype and are critical for proper load transfer between mechanically dissimilar elastic ligaments and stiff bone. Currently, these gradients are not restored in natural healing, repair, or in engineered replacements, yielding limited repair options and high failure rates, demonstrating the need to develop native-like entheses for improved functional and lasting repair. The collective focus of this work was to drive development of native-like entheses in engineered ligaments by mimicking developmentally inspired mechanical cues. We found that compressive boundary clamps, which restrict cellular contraction and produce a zonal tensile-compressive environment at the clamping interface, guide ligament fibroblasts to produce multizonal, early postnatal-like enthesis gradients in organization and composition. Further, the addition of βTCP under these clamps enhanced maturation of these engineered entheses mirroring development via endochondral ossification (Chapter 2). Next, the application of developmental mechanical cues mimicking growth rate and cyclic muscle loading were found to differentially drive maturation of entheses in ACL fibroblast seeded engineered ligament constructs (Chapter 3). Based on the exciting findings we found of this culture system driving ACL fibroblasts to form immature entheses, we were interested in evaluating whether these mechanical cues were enough to drive human mesenchymal stem cells (hMSCs) to differentiate and produce similar zonal entheses. Static mechanical cues were found to drive zonal stem cell differentiation and accelerated early postnatal-like enthesis gradients in hMSC seeded constructs (Chapter 4). Further, application of dynamic mechanical cues drove further maturation and differentiation of hMSC constructs beyond early postnatal-like development towards later postnatal enthesis development with more mature native-like gradients in collagen organization, matrix composition, and cell phenotype (Chapter 5). Collectively, developmental mechanical cues were able to drive the development of some of the most physiologically appropriate engineered entheses to date. Further, this work established and characterized a promising in vitro culture system that closely mirrors native enthesis development. This is a promising platform for use in future studies to investigate and understand how mechanical cues can further improve enthesis regeneration efforts moving forward.
Rights
© Michael Ethan Brown
Is Part Of
VCU University Archives
Is Part Of
VCU Theses and Dissertations
Date of Submission
5-10-2023