Date of Award
2021
Embargo Period
2-26-2026
Document Type
Dissertation - MUSC Only
Degree Name
Doctor of Philosophy (PhD)
Department
Pathology and Laboratory Medicine
College
College of Graduate Studies
First Advisor
William D. Hill
Second Advisor
Robin C. Muise-Helmericks
Third Advisor
Joe B. Blumer
Fourth Advisor
Amanda C. LaRue
Fifth Advisor
Louis M. Luttrell
Sixth Advisor
Meghan E. McGee-Lawrence
Abstract
Osteoporosis is a disease that leads to high morbidity and mortality. It represents a reduced bone formation to bone resorption ratio. With most current treatments for osteoporosis targeting bone resorption, there is a need for new interventions that can increase osteogenesis. CXCL12 is a pro-osteogenic chemokine that can recruit bone marrow mesenchymal stem cells (BMSCs) to injury sites and increase their proliferation and osteogenic differentiation. This work aims to study mechanisms for translational and post-translational regulation of CXCL12 via the aryl hydrocarbon receptor (AhR), miRNAs, and the enzyme dipeptidyl peptidase 4 (DPP4). We utilized a wild type and mutated 3’-UTR luciferase assay to demonstrate direct regulation of CXCL12 by miR-29b-1-5p. We also showed that this inhibition of CXCL12 translation is downstream of AhR activation by kynurenine (KYN), a tryptophan metabolite that accumulates in bone marrow with aging. Using ELISA, we showed that total CXCL12 levels declined in the murine bone marrow. We also showed that cell culture media from aged BMSCs contained higher levels of KYN compared to BMSCs isolated from younger mice. Treating BMSCs with KYN lead to inhibition of osteogenic differentiation and decreased CXCL12 levels in cell culture media, while adding a miR-29b-1-5p inhibitor blocked KYN’s effect on CXCL12 levels. Post-translationally, CXCL12 is cleaved by DPP4 within minutes of its release. Using proteomics and phospho-proteomics approaches, we demonstrated that DPP4-cleaved CXCL12 is a functional antagonist to “intact” CXCL12. We showed that cleaved CXCL12 inhibits osteogenesis and osteoclastogenesis, and induces senescence in BMSCs. We also utilized BRET assays to show that cleaved CXCL12 can still signal via the ACKR3 β-arrestin pathway, but cannot activate G-protein signaling downstream of CXCR4 activation. Interestingly, using immunoprecipitation mass spectrometry, we found that while cleaved CXCL12 makes up almost 54% of total CXCL12 in human plasma, it made up almost 78% of total CXCL12 in bone marrow interstitial fluid. In conclusion, we demonstrated how CXCL12 is important for BMSCs osteogenic differentiation and explored two mechanisms that inhibit pro-osteogenic functions of CXCL12 with age; KYN-driven epigenetic inhibition and a DPP4 generated CXCL12 metabolite with cryptic CXCR4 and ACKR3 receptor based signaling. These two pathways could provide two potential clinical targets for prevention and/or treatment of osteoporosis.
Recommended Citation
Elmansi, Ahmed M., "Age-Related Translational and Post-translational Regulation of the Pro-Osteogenic Chemokine CXCL12 in the Bone Marrow Microenvironment" (2021). MUSC Theses and Dissertations. 747.
https://medica-musc.researchcommons.org/theses/747
Rights
All rights reserved. Copyright is held by the author.