Osteocyte Apoptosis Caused by Hindlimb Unloading is Required to Trigger Osteocyte RANKL Production and Subsequent Resorption of Cortical and Trabecular Bone in Mice Femurs

Abstract

Osteocyte apoptosis is essential to activate bone remodeling in response to fatigue microdamage and estrogen withdrawal, such that apoptosis inhibition in vivo prevents the onset of osteoclastic resorption. Osteocyte apoptosis has also been spatially linked to bone resorption owing to disuse, but whether apoptosis plays a similar controlling role is unclear. We, therefore, 1) evaluated the spatial and temporal effects of disuse from hindlimb unloading (HLU) on osteocyte apoptosis, receptor activator of NF-κB ligand (RANKL) expression, bone resorption, and loss in mouse femora, and 2) tested whether osteocyte apoptosis was required to activate osteoclastic activity in cortical and trabecular bone by treating animals subjected to HLU with the pan-caspase apoptosis inhibitor, QVD (quinolyl-valyl-O-methylaspartyl-[-2,6-difluorophenoxy]-methylketone). Immunohistochemistry was used to identify apoptotic and RANKL-producing osteocytes in femoral diaphysis and distal trabecular bone, and μCT was used to determine the extent of trabecular bone loss owing to HLU. In both cortical and trabecular bone, 5 days of HLU increased osteocyte apoptosis significantly (3- and 4-fold, respectively, p < 0.05 versus Ctrl). At day 14, the apoptotic osteocyte number in femoral cortices declined to near control levels but remained elevated in trabeculae (3-fold versus Ctrl, p < 0.05). The number of osteocytes producing RANKL in both bone compartments was also significantly increased at day 5 of HLU (>1.5-fold versus Ctrl, p <0.05) and further increased by day 14. Increases in osteocyte apoptosis and RANKL production preceded increases in bone resorption at both endocortical and trabecular surfaces. QVD completely inhibited not only the HLU-triggered increases in osteocyte apoptosis but also RANKL production and activation of bone resorption at both sites. Finally, μCT studies revealed that apoptosis inhibition completely prevented the trabecular bone loss caused by HLU. Together these data indicate that osteocyte apoptosis plays a central and controlling role in triggering osteocyte RANKL production and the activation of new resorption leading to bone loss in disuse.

Role of endoplasmic reticulum stress in disuse osteoporosis

“…The tail-suspended animal model has been widely accepted as an effective disuse osteoporosis model for simulating bone loss [15,22,41]. In the current tail suspension experiment, bone formation was mildly inhibited and bone resorption was markedly increased in unloaded mice, these 20 changes are in accord with previous study [14,[42][43][44]. Our correlation analysis and linear regression showed that the expression level of p-eIF2α was positively associated with the number of bone-forming osteoblasts, and negatively associated with that of bone-resorbing osteoclasts.…”

β-aminoisobutyric Acid, l-BAIBA, Is a Muscle-Derived Osteocyte Survival Factor

“…Blocking osteocyte apoptosis using an apoptosis inhibitor has been shown to prevent bone loss induced by hindlimb unloading (Cabahug-Zuckerman et al, 2016). As BAIBA production is increased with exercise (Roberts et al, 2014), the hindlimb unloading assay, a model for disuse or lack of exercise, was performed to determine if BAIBA could prevent bone loss.…”

“…The detrimental effects of unloading on bone mass through osteocytes are well described (Basso and Heersche, 2006), and it has previously been shown that blocking osteocyte apoptosis prevents bone loss due to hindlimb unloading (Cabahug-Zuckerman et al, 2016). We found that providing L-BAIBA in drinking water for 2 weeks of hindlimb unloading retained bone mass, more so in males than females, most likely due to the greater amount of trabecular bone in males compared to females even with using 3-month-old females as compared to 5-month-old males.…”

Cellular and Molecular Pathways Leading to External Root Resorption

“…A possible explanation for this different appetite and resorption behavior is that dentin contains more matrix proteins of noncollagenous origin, such as osteopontin, when compared with bone (Azari et al 2011). The absence of regulatory osteocytes and osteocyte proteins in dentin may also account for the quantitative differences (Cabahug-Zuckerman et al 2016). Pretreatment with osteopontin or dentin sialoprotein has been reported to induce an increase in IL-1 (Yao et al 2008).…”