Previous research has provided evidence in support of apoptotic osteocytes playing a role in the bone remodeling process. In this study, we examined the regional and quadrantal variations of apoptotic and viable osteocytes in cyclically loaded and unloaded samples. Left tibias of C57 Black 6 Taconic mice (C57Bl/6) were cyclically loaded for either 2 weeks or 5 weeks, with the right tibias being used as controls. After loading, tibias were resected, processed, and then stained using either a TUNEL stain, to show apoptotic osteocytes, or a 2.0% methyl green solution, to reveal viable cells. Cross-sectional images from each tibia were then captured and analyzed in each region (distal, midshaft and proximal) and quadrant (cranial, lateral, caudal, and medial) by counting the number of osteocytes, both apoptotic and viable, and subsequently calculating the percentages and densities of those osteocytes. Individual analysis of each sample group showed that the 5 week loaded bones, with the most statistically significant p-values, had the most regional variations within the samples, specifically showing decreased apoptotic and viable osteocytes in the lateral quadrants. Comparative analysis revealed a statistically significant higher percentage and density of apoptotic osteocytes in 5 week loaded samples compared to all other samples. This provides further quantitative evidence in support of apoptotic osteocytes playing a role in bone remodeling.

ABSTRACT A Study of Osteocyte Apoptosis by Region and Quadrant in Murine Cortical Bone Jessica Kristen Chan Osteocytes undergo apoptosis to spatially and temporally initiate bone remodeling. This study investigates the distribution of apoptotic osteocytes within different quadrants and regions of cortical bone and compares the frequency of osteocyte apoptosis to regional factors associated with bone remodeling. Specifically, the quantity of apoptosis was compared to levels of the bone morphagenic protein antagonists noggin and gremlin. Samples of unloaded right tibial bone obtained from C57/Bl/6 mice underwent TUNEL staining for apoptotic osteocytes and were counterstained with methyl green to detect osteocyte viability. Cross sectional areas of bone were divided into four quadrants (cranial, caudal, medial, and lateral) and three regions (proximal, midshaft, and distal) for analysis. Densities and percentages of osteocytes were measured within each area. While the results show that there were no differences among quadrants, regional variations were found in osteocyte apoptosis. A significantly higher density of apoptotic osteocytes was found in the midshaft region which also displayed higher levels of BMP antagonists. Using regression analysis, a positive linear relationship between apoptotic osteocytes and gremlin was established while noggin showed a negative linear correlation for the percentage of apoptotic osteocytes. Further studies are needed to observe the distribution of apoptotic osteocytes within loaded bone to confirm the exact relationship between osteocyte apoptosis and bone remodeling. Keywords: osteocyte apoptosis, bone remodeling