Introduction: Collagen and glycosaminoglycans (GAGs) such as hyaluronan (HA) and chondroitin sulfate (CS) are basic elements of bone structure and collagen-GAG composites are currently developed for a wide range of applications. Here, we report on the molecular and cellular effects of GAGs and their sulfated derivatives on osteocytes, which are fundamental orchestrators of bone remodeling.
Materials and methods: The effects of native and sulfate-modified GAGs on viability, necrosis, apoptosis, and gene expression were studied in the murine MLO-Y4 and the rat UMR 106-01 cell lines, which both display properties of primary osteocytes. Necrosis and apoptosis were determined using ELISA photometric immunoassays of DNA fragmentation, and viability was evaluated with a fluometric assay. The gene expression profile was examined by real-time PCR.
Results: Native and sulfated GAGs were stable and non-cytotoxic. At a concentration of 200 μg/ml, unsulfated HA did not reduce apoptosis compared to control, whereas highly sulfated HA led to a significant reduction of apoptosis both in comparison to control and unsulfated HA (P<0.05). Moreover, highly sulfated CS decreased apoptosis by 30% compared to control and to its native form (P<0.05). Similar results were observed for cell necrosis. Both forms of HA significantly increased cell viability when compared to control (P<0.05), whereas CS did not affect cell viability. At concentrations ranging from 10 to 200 μg/ml, unsulfated HA dose-dependently increased the RANKL:OPG ratio compared to control, whereas highly sulfated HA significantly downregulated the RANKL:OPG ratio when compared to its native form (both P<0.05). The expression of SOST, the gene encoding sclerostin was also reduced by 38% by highly sulfated HA when compared to control (P<0.05). Native HA and CS did not alter SOST expression.
Conclusion: Highly sulfated HA may maintain the phenotype of healthy and functional osteocytes but the clinical significance of these findings needs to be validated in vivo.
18 - 21 May 2013
European Calcified Tissue Society