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Matrix-bound Nanovesicles as an Extracellular Source of Lysyl Oxidase

Lee, Yoojin (2021) Matrix-bound Nanovesicles as an Extracellular Source of Lysyl Oxidase. Doctoral Dissertation, University of Pittsburgh. (Unpublished)

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Biologic scaffolds produced from extracellular matrix (ECM) have been commonly implemented as inductive templates for constructive tissue remodeling in multiple musculoskeletal sites. The discovery of matrix-bound nanovesicles (MBV), a unique subpopulation of extracellular vesicles embedded within the ECM, has prompted the further study of MBV structure, internal cargo, and overall function. MBV are more specifically defined as lipid-bound extracellular vesicles that possess the ability to modulate cell behavior as a function of their internal cargo of biologically active signaling molecules (e.g., microRNAs, proteins, lipids, cytokines). The lipid membrane of MBV is decorated with a variety of surface proteins that are unique to the local function and anatomic location from which MBV are harvested.
Lysyl oxidase (LOX) is an enzyme responsible for catalyzing collagen cross-linking and serves an essential role in tissue stabilization. In this dissertation, LOX is investigated because of its function of regulating cross-linking of collagen in tissues and its potential to be implemented as a therapy in damaged or healing tissues that lack sufficient cross-linking. There are substantial challenges associated with isolating and purifying LOX in high quantities, which limits full characterization and understanding of the protein. Herein, MBV is isolated from multiple tissue types in abundance and is presented as a promising method of isolating LOX.
The objectives of the present dissertation were to isolate MBV-associated LOX from ECM bioscaffolds and to determine its effects on the development of collagen fibrils in vitro. ECM bioscaffolds were solubilized using enzymatic digestion with collagenase and elastase, and incubation in a high salt solution. Results show that MBV-associated LOX in the ECM is present
in its 52 kDa pro-peptide form and that it is enzymatically active when isolated with elastase digestion. MBV-associated LOX activity significantly decreases when treated with LOX inhibitor β-aminopropionitrile (BAPN) or when proteinase K is utilized to remove surface proteins. Furthermore, MBV-associated LOX enhances the formation of cross-linked collagen and the strength collagen constructs in vitro. MBV offer an attractive method to deliver surface-associated LOX, which through its direct role in collagen cross-linking and matrix regulation for enhancing the biomechanics of remodeled tissue, specifically its strength.


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Item Type: University of Pittsburgh ETD
Status: Unpublished
CreatorsEmailPitt UsernameORCID
Lee, Yoojinlee.yooj.c@gmail.comyol400000-0002-1197-1588
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Thesis AdvisorBadylak,
Committee MemberDavidson,
Committee MemberWoo,
Committee MemberHussey,
Committee MemberZdanowicz,
Date: 26 January 2021
Date Type: Publication
Defense Date: 14 July 2020
Approval Date: 26 January 2021
Submission Date: 30 June 2020
Access Restriction: No restriction; Release the ETD for access worldwide immediately.
Number of Pages: 125
Institution: University of Pittsburgh
Schools and Programs: Swanson School of Engineering > Bioengineering
Degree: PhD - Doctor of Philosophy
Thesis Type: Doctoral Dissertation
Refereed: Yes
Uncontrolled Keywords: Matrix-bound nanovesicles, lysyl oxidase, extracellular matrix
Date Deposited: 26 Jan 2021 14:17
Last Modified: 26 Jan 2021 14:17


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