Date of Award

Summer 2025

Rights

Access is available to all users

Document Type

Thesis

Degree Name

Master of Science (MS) in Biology

Department

Biology

Abstract

Bone remodeling is a dynamic process maintained by the balance between osteoblast-mediated bone formation and osteoclast-driven resorption. When this balance is disrupted, pathological bone loss occurs, as seen in osteoporosis, rheumatoid arthritis, and cancer-induced bone disease (Teitelbaum, 2007; Korol & Baumeister, 2023; Ensrud & Crandall, 2017). Osteoclasts, derived from hematopoietic stem cells through a macrophage lineage, play a central role in bone resorption. However, the identity of all proteins of osteoclast precursors remains unclear. CD68, a lysosomal glycoprotein widely expressed in macrophages and monocyte-lineage cells (Holness & Simmons, 1993), has been proposed as a marker of osteoclast precursors. Some studies have demonstrated that CD68- positive bone marrow cells differentiate into osteoclasts under the influence of RANKL and M-CSF (Pettit et al., 2008), while others suggest CD68 expression may not be exclusive to osteoclast lineage cells (Witwicka et al., 2015; Jackson et al., 2017). This study aim was to determine whether CD68-positive bone marrow cells serve as more effective osteoclast precursors with greater osteoclastogenic potential than CD68-negative populations. Bone marrow cells were harvested from C57Bl/6 mice and subjected to fluorescence-activated cell sorting (FACS) based on CD68 and RANK expression. The cell population data that was collected was then analyzed for correlation between CD68 and RANK in bone marrow populations. Sorted populations also underwent osteoclast differentiation in vitro using Macrophage Colony-Stimulating Factor and Receptor Activator of Nuclear factor kappa-B stimulation. Tartrate-resistant acid phosphatase (TRAP) staining and image analysis will assess osteoclast formation. We hypothesized that bone marrow osteoclast populations would have a high correlation between CD68 and Receptor Activator of Nuclear factor kappa-B Ligand. Also that they would form larger, multinucleated TRAP-positive osteoclasts and exhibit elevated osteoclast gene expression compared to CD68- negative cells. Even if CD68 proves to be correlated with RANK in bone marrow populations more testing would be required before it could be considered a reliable marker of osteoclast precursors. However, if further research confirms this it could serve as a therapeutic target for modulating osteoclast activity in diseases involving excessive bone resorption (Boyle et al., 2003; Kong et al., 1999; Batoon et al., 2021). By refining the identification of osteoclast precursors, this research aims to improve the specificity and efficacy of bone disease treatments while preserving physiological remodeling and skeletal integrity.

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