Author ORCID Identifier

https://orcid.org/0000-0002-0481-5464

Defense Date

2021

Document Type

Dissertation

Degree Name

Doctor of Philosophy

Department

Integrative Life Sciences

First Advisor

Barbara D. Boyan

Second Advisor

Henry J. Donahue

Third Advisor

Preetam Ghosh

Fourth Advisor

Zvi Schwartz

Fifth Advisor

Tarynn Witten

Abstract

Growth plate chondrocytes are the driving force of long bone elongation and they accomplish this vital task, in part, by producing, remodeling, and finally mineralizing the cartilage tissue that makes up the growth plate. This mineralized tissue is turned over again as bone forming cells invade and began the process of turning mineralized cartilage into bone. Chondrocytes undergo a distinct series of observable phases as they move through this process transitioning from a stage of proliferation to one of hypertrophy and finally capillary invasion. Chondrocytes produce matrix vesicles (MVs) that they release into the growth plate and that attach to collagen in the extracellular matrix (ECM). These MVs are involved in chondrocyte regulation, tissue remodeling, and matrix mineralization.

This entire process is highly regulated by various factors and hormones. 1α,25-dihydroxyvitamin D3 [1α,25(OH)2D3] is a well-studied hormone with known regulation of both chondrocytes and MVs. With the recent discovery of selectively exported microRNA in growth plate MVs there is the possibility of an additional regulator of growth plate maturation. The central hypothesis of this work was that MV microRNA are able to regulate growth plate chondrocytes and that the production and packaging of MV microRNA is regulated by 1α,25(OH)2D3.

Specific aim 1 determined whether microRNA were selectively exported in MVs, if those microRNA remain protected within the MVs, if certain microRNA are found in ex-vivo tissue, and the ability of specific microRNA to regulate growth plate chondrocytes. Specific aim 2 determined the regulatory potential of 1α,25(OH)2D3 on microRNA production and packaging into MVs and the known mRNA and pathways targeted by the exported microRNA that were differentially expressed following 1α,25(OH)2D3 treatment as well as the impact of 1α,25(OH)2D3 on MV endocytosis and the release of microRNA from MVs. Specific aim 3 examined the transcriptome following specific microRNA transfections and adapted an approach to target the interactions between specific tagged microRNA and the corresponding mRNA followed by RNAseq of the mRNA for the purpose of pathway analysis.

Rights

© The Author

Is Part Of

VCU University Archives

Is Part Of

VCU Theses and Dissertations

Date of Submission

5-13-2022

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