Local Adaptations of Osteocyte Proteins to Increased and Decreased Mechanical Forces Correlate with Osteoblast Levels

Abstract

Osteocytes, cells embedded in the mineralized matrix of bone, are believed to be the primary mechanosensors of bone tissue. They signal to both osteoblasts (bone forming cells) and osteoclasts (bone resorbing cells) by releasing certain proteins. Sclerostin, interleukin-6 (IL-6), and insulin-like growth factor-I (IGF-I) are three such proteins that signal to osteoblasts to increase (via IGF-I and IL-6) or decrease (via sclerostin) osteoblast activity. PURPOSE: To determine if the osteocyte protein response to mechanical unloading is restricted to the unloaded bone or is a systemic signal. Using a hindlimb unloading (HU) rodent model, we hypothesized the unloaded hindlimb would have altered prevalence of osteocyte proteins while the weight-bearing forelimb would have no differences. METHODS: Male Sprague Dawley rats (6-mo old) were subjected to HU (n=7) for 28 days. Age-matched controls (CON; n=7) had normal weight-bearing activity on all four limbs for 28 days. The unloaded distal femur (DF) and the weight-bearing proximal humerus (PH) were compared in HU vs CON. RESULTS: Immunohistochemical staining of the cancellous region to quantify %positive osteocytes revealed 19% higher %sclerostin+ osteocytes in the DF in HU, but 30% lower %sclerostin+ at the PH. Both %IGF-I+ and %IL-6+ osteocytes were lower at the DF (by 29% and 25%, respectively), but higher at the PH by 94% and 48%. Staining for osterix, a marker of osteoblasts, showed 60% lower %osterix+ cancellous bone surface in HU in the DF; however, the PH had 48% more %osterix+ surface in HU. All comparisons were statistically significant at p<0.05. CONCLUSION: After 28 days of HU, the unloaded DF had higher sclerostin osteocyte prevalence and lower IL-6 and IGF-I osteocyte prevalence as well as lower osteoblast surface as expected with unloading. Our results indicate that the osteocytes in the PH are signaling osteoblasts to increase formation, which is an unexpected finding based on the conventional notion that the forelimbs of HU animals are normally loaded and not overloaded. The opposite response of osteocyte proteins and osteoblast surface in bones within the same animal that are experiencing both unloading and loading indicates a precise, localized mechanism by which osteocytes sense mechanical strain and signal to local cells to adapt to those changes.