Evidence for altered osteoclastogenesis in splenocyte cultures from Cyp27b1 knockout mice

• Published in: The Journal of steroid biochemistry and molecular biology 2016-11, Vol.164, p.353-360
• Main Authors: Reinke, Daniel C., Kogawa, Masakazu, Barratt, Kate R., Morris, Howard A., Anderson, Paul H., Atkins, Gerald J.
• Format: Article
• Language: English
• Subjects: 25-Hydroxyvitamin D; 25-Hydroxyvitamin D3 1-alpha-Hydroxylase - deficiency; 25-Hydroxyvitamin D3 1-alpha-Hydroxylase - genetics; Acid phosphatase (tartrate-resistant); Animals; Apoptosis; Bcl-2 protein; bcl-2-Associated X Protein - genetics; bcl-2-Associated X Protein - metabolism; Bone resorption; Bone Resorption - genetics; Bone Resorption - metabolism; Bone Resorption - pathology; Calcitriol - metabolism; Cell cycle; Cell Differentiation; Clonal deletion; CYP27B1; Gene deletion; Gene Expression Regulation; Metabolism; Mice; Mice, Knockout; Osteoclast; Osteoclastogenesis; Osteoclasts; Osteoclasts - metabolism; Osteoclasts - pathology; Osteogenesis - genetics; Phosphatase; Primary Cell Culture; Proto-Oncogene Proteins c-bcl-2 - genetics; Proto-Oncogene Proteins c-bcl-2 - metabolism; Spleen - cytology; Spleen - metabolism; Splenocytes; Tartrate-Resistant Acid Phosphatase - genetics; Tartrate-Resistant Acid Phosphatase - metabolism; Vitamin D; Vitamin D - analogs & derivatives; Vitamin D - metabolism
• ISSN: 0960-0760; 1879-1220
• DOI: 10.1016/j.jsbmb.2015.11.015

•Osteoclast differentiation in mice with global deletion of Cyp27b1 (Cyp27b1 KO) was studied.•Cyp27b1 KO splenocytes gave rise to fewer osteoclasts in response to recombinant RANKL/M-CSF.•Cyp27b1 KO derived osteoclasts had enhanced resorptive capacity. The association between increased serum 25-hydroxyvitamin D (25D) and reduced osteoclastic bone resorption is well known. Previously, we have demonstrated that mechanism by which this occurs, may include the conversion of 25D to 1,25-dihydroxyvitamin D (1,25D) by osteoclasts, catalysed by the CYP27B1 enzyme. Local 1,25D synthesis in osteoclasts was shown to regulate osteoclastogenesis and moderating resorptive activity. Thus, we hypothesised that osteoclasts differentiated from mice with global deletion of the Cyp27b1 gene (Cyp27b1 KO) would display enhanced resorptive capacity due to the lack of an ameliorating effect of 1,25D. Splenocytes isolated from Cyp27b1 KO mice or their wild-type (WT) littermates between 6 and 8 weeks of age were cultured under osteoclast-forming conditions for up to 14 days. Osteoclast formation was measured by staining for the osteoclast marker tartrate resistant acid phosphatase (TRAP). Bone resorption activity was measured by plating the cells on a bone-like substrate. In Cyp27b1 KO cultures, osteoclastogenesis was reduced, as indicated by fewer TRAP-positive multinucleated cells at all time points measured (p