Sclerostin, the protein product of the Sost gene, is a potent inhibitor of bone formation. Among bone cells, sclerostin is found nearly exclusively in the osteocytes, the cell type that historically has been implicated in sensing and initiating mechanical signaling. The recent discovery of the antagonistic effects of sclerostin on Lrp5 receptor signaling, a crucial mediator of skeletal mechanotransduction, provides a potential mechanism for the osteocytes to control mechanotransduction, by adjusting their sclerostin (Wnt inhibitory) signal output to modulate Wnt signaling in the effector cell population. We investigated the mechanoregulation of Sost and sclerostin under enhanced (ulnar loading) and reduced (hindlimb unloading) loading conditions. Sost transcripts and sclerostin protein levels were dramatically reduced by ulnar loading. Portions of the ulnar cortex receiving a greater strain stimulus were associated with a greater reduction in Sost staining intensity and sclerostin-positive osteocytes (revealed via in situ hybridization and immunohistochemistry, respectively) than were lower strain portions of the tissue. Hindlimb unloading yielded a significant increase in Sost expression in the tibia. Modulation of sclerostin levels appears to be a finely tuned mechanism by which osteocytes coordinate regional and local osteogenesis in response to increased mechanical stimulation, perhaps via releasing the local inhibition of Wnt/Lrp5 signaling.

Osteocalcin (OCN), the most abundant noncollagenous protein in the bone matrix, is reported to be a bone-derived endocrine hormone with wide-ranging effects on many aspects of physiology, including glucose metabolism and male fertility. Many of these observations were made using an OCN-deficient mouse allele (Osc^–) in which the 2 OCN-encoding genes in mice, Bglap and Bglap2, were deleted in ES cells by homologous recombination. Here we describe mice with a new Bglap and Bglap2 double-knockout (dko) allele (Bglap/2^{p.Pro25fs17Ter}) that was generated by CRISPR/Cas9-mediated gene editing. Mice homozygous for this new allele do not express full-length Bglap or Bglap2 mRNA and have no immunodetectable OCN in their serum. FTIR imaging of cortical bone in these homozygous knockout animals finds alterations in the collagen maturity and carbonate to phosphate ratio in the cortical bone, compared with wild-type littermates. However, μCT and 3-point bending tests do not find differences from wild-type littermates with respect to bone mass and strength. In contrast to the previously reported OCN-deficient mice with the Osc⁻allele, serum glucose levels and male fertility in the OCN-deficient mice with the Bglap/2^{pPro25fs17Ter} allele did not have significant differences from wild-type littermates. We cannot explain the absence of endocrine effects in mice with this new knockout allele. Possible explanations include the effects of each mutated allele on the transcription of neighboring genes, or differences in genetic background and environment. So that our findings can be confirmed and extended by other interested investigators, we are donating this new Bglap and Bglap2 double-knockout strain to the Jackson Laboratories for academic distribution. Cells that make and maintain bone express proteins that function either locally or systemically. The former proteins, such as type 1 collagen, affect the material properties of the skeleton, while the latter, such as fibroblast growth factor 23, enable the skeleton to communicate with other organ systems. Mutations that affect the functions of most bone-cell-expressed proteins cause diseases that have similar features in humans and other mammals such as mice, for example, brittle bone diseases for type 1 collagen mutations and hypophosphatemic rickets for mutations in fibroblast growth factor 23. Our study focuses on another bone-cell-expressed protein, osteocalcin, which has been suggested to function locally to affect bone strength and systemically as a hormone. Studies using osteocalcin knockout mice led other investigators to suggest endocrine roles for osteocalcin in regulating blood glucose, male fertility, muscle mass, brain development, behavior, and cognition. We therefore decided to generate a new strain of osteocalcin knockout mice that could also be used to investigate these nonskeletal effects. To our surprise, the osteocalcin knockout mice we created did not significantly differ from wild-type mice for the three phenotypes we examined: bone strength, blood glucose, and male fertility. Our data are consistent with findings from osteocalcin knockout rats but are inconsistent with data from the original osteocalcin knockout mice. Because we do not know why our new strain fails to recapitulate the phenotypes previously reported for another knockout mouse stain, we have donated our mice to a public repository so that they can be easily obtained and studied in other academic laboratories.

Ate impact. Several clinical and basic science reports indicate that muscle forces play a dominant role in generating the mechanical stimulus in exercise-induced bone gain. Although it is, in most cases, difficult to separate the effects of gravitational forces acting on body mass from muscle contractions, several well-conceived experiments offer considerable insight into the propensity of muscle-derived forces per se to drive the adaptive response in bone. Load-induced osteogenesis requires that mechanical signals come packaged with particular characteristics, all of which can be generated from either gravitational or muscle forces. Neither of these two sources have been demonstrated empirically to be the source of bone's adaptive response, but a convincing body of data suggests that muscle contractions are present, significant, and capable of accounting for most of the adaptive responses....

Bone fragility fractures are caused by low bone mass or impaired bone quality. Osteoblast/osteoclast coordination determines bone mass, but the factors that control bone quality are poorly understood. Osteocytes regulate osteoblast and osteoclast activity on bone surfaces but can also directly reorganize the bone matrix to improve bone quality through perilacunar/canalicular remodeling; however, the molecular mechanisms remain unclear. We previously found that deleting the transcriptional regulators Yes-associated protein (YAP) and transcriptional co-activator with PDZ-motif (TAZ) from osteoblast-lineage cells caused lethality in mice due to skeletal fragility. Here, we tested the hypothesis that YAP and TAZ regulate osteocyte-mediated bone remodeling by conditional ablation of both YAP and TAZ from mouse osteocytes using 8 kb-DMP1-Cre. Osteocyte-conditional YAP/TAZ deletion reduced bone mass and dysregulated matrix collagen content and organization, which together decreased bone mechanical properties. Further, YAP/TAZ deletion impaired osteocyte perilacunar/canalicular remodeling by reducing canalicular network density, length, and branching, as well as perilacunar flourochrome-labeled mineral deposition. Consistent with recent studies identifying TGF-β as a key inducer of osteocyte expression of matrix-remodeling enzymes, YAP/TAZ deletion in vivo decreased osteocyte expression of matrix proteases MMP13, MMP14, and CTSK. In vitro, pharmacologic inhibition of YAP/TAZ transcriptional activity in osteocyte-like cells abrogated TGF-β-induced matrix protease gene expression. Together, these data show that YAP and TAZ control bone matrix accrual, organization, and mechanical properties by regulating osteocyte-mediated bone remodeling. Elucidating the signaling pathways that control perilacunar/canalicular remodeling may enable future therapeutic targeting of bone quality to reverse skeletal fragility. © 2019 American Society for Bone and Mineral Research.

The WNT pathway has become an attractive target for skeletal therapies. High-bone-mass phenotypes in patients with loss-of-function mutations in the LRP5/6 inhibitor Sost (sclerosteosis), or in its downstream enhancer region (van Buchem disease), highlight the utility of targeting Sost/sclerostin to improve bone properties. Sclerostin-neutralizing antibody is highly osteoanabolic in animal models and in human clinical trials, but antibody-based inhibition of another potent LRP5/6 antagonist, Dkk1, is largely inefficacious for building bone in the unperturbed adult skeleton. Here, we show that conditional deletion of Dkk1 from bone also has negligible effects on bone mass. Dkk1 inhibition increases Sost expression, suggesting a potential compensatory mechanism that might explain why Dkk1 suppression lacks anabolic action. To test this concept, we deleted Sost from osteocytes in, or administered sclerostin neutralizing antibody to, mice with a Dkk1-deficient skeleton. A robust anabolic response to Dkk1 deletion was manifest only when Sost/sclerostin was impaired. Whole-body DXA scans, μCT measurements of the femur and spine, histomorphometric measures of femoral bone formation rates, and biomechanical properties of whole bones confirmed the anabolic potential of Dkk1 inhibition in the absence of sclerostin. Further, combined administration of sclerostin and Dkk1 antibody in WT mice produced a synergistic effect on bone gain that greatly exceeded individual or additive effects of the therapies, confirming the therapeutic potential of inhibiting multiple WNT antagonists for skeletal health. In conclusion, the osteoanabolic effects of Dkk1 inhibition can be realized if sclerostin upregulation is prevented. Anabolic therapies for patients with low bone mass might benefit from a strategy that accounts for the compensatory milieu of WNT inhibitors in bone tissue.