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Access Type
WSU Access
Date of Award
January 2022
Degree Type
Dissertation
Degree Name
Ph.D.
Department
Biomedical Engineering
First Advisor
Howard W. Matthew
Abstract
Knee osteoarthritis (OA) involves complex and heterogeneous changes to articular cartilage (AC), with morphologically- and molecularly-distinct regions presenting simultaneously. The stromal cell-derived factor-1 (SDF-1)/CXCR4 axis is a key influencer of chondrocyte phenotype during OA progression, but chondrocyte CXCR4 expression is not well-characterized. Molecular imaging presents an opportunity to study CXCR4 expression patterns across the articular surface. The purpose of this study was to develop and apply a small molecule-based, near infrared fluorescent (NIR) probe to map CXCR4 expression within AC, and correlate it to AC morphology in a preclinical model of anterior cruciate ligament (ACL) injury. A novel CXCR4 probe was developed and robustly validated. In vitro NIR imaging confirmed that probe binding to CXCR4+ cells (HUVECs) was significantly ablated via competitive binding. Signal correlated strongly to CXCR4+ cell density. Flow cytometry confirmed strong correlation between probe- and antibody-based CXCR4 labeling. Ex vivo NIR imaging of rat femoral explants confirmed that probe-based AC signal was significantly higher than un-conjugated fluorophore, could be ablated via competitive inhibition, and was sensitive to upregulation of CXCR4. Fluorescent histology of probe labeling confirmed full penetration through uncalcified AC, with signal localized to chondrocytes. A custom sample holder compatible with both NIR and micro-computed tomography (µCT) systems was fabricated and combined with advanced image processing strategies to generate paired maps probe-based NIR signal and AC thickness to be generated. Female Lewis rats were subjected to closed ACL rupture (n=8/endpoint) or sham (n=4/endpoint), incubated with probe, and imaged via NIR and contrast-enhanced µCT at 1- and 4-weeks post-injury. AC of injured femora and tibiae displayed characteristic morphological hallmarks of OA, with increased AC thickness (AC.Th) and surface deviation at 1- and 4-weeks. Injured medial femoral condyles exhibited regions of hypertrophic, pathologically thick AC adjacent to thin, degenerate regions, and corresponded to distinct regions of high and low probe-based NIR signal. NIR signal deviation was significantly higher in injured femora compared to sham at 1-week post-injury, while no significant differences in mean NIR signal were observed. Understanding chondrocyte CXCR4 expression patterns enhances overall understanding of disease progression and may inform future treatment strategies.
Recommended Citation
Newton, Michael Daniel, "Molecular Mapping Of Articular Cartilage Cxcr4 Expression After Acl Injury Via A Novel Small Molecule-Based Probe" (2022). Wayne State University Dissertations. 3806.
https://digitalcommons.wayne.edu/oa_dissertations/3806