John G. Logan

The genetic landscape of diseases associated with changes in bone mineral density (BMD), such as osteoporosis, is only partially understood. Here, we explored data from 3,823 mutant mouse strains for BMD, a measure that is frequently altered in a range of bone pathologies, including osteoporosis. A total of 200 genes were found to significantly affect BMD. This pool of BMD genes comprised 141 genes with previously unknown functions in bone biology and was complementary to pools derived from recent human studies. Nineteen of the 141 genes also caused skeletal abnormalities. Examination of the BMD genes in osteoclasts and osteoblasts underscored BMD pathways, including vesicle transport, in these cells and together with in silico bone turnover studies resulted in the prioritization of candidate genes for further investigation. Overall, the results add novel pathophysiological and molecular insight into bone health and disease.

Functional characterisation of cell-type-specific regulatory networks is key to establish a causal link between genetic variation and phenotype. The osteoclast offers a unique model for interrogating the contribution of co-regulated genes to in vivo phenotype as its multinucleation and resorption activities determine quantifiable skeletal traits. Here we took advantage of a trans-regulated gene network (MMnet, macrophage multinucleation network) which we found to be significantly enriched for GWAS variants associated with bone-related phenotypes. We found that the network hub gene Bcat1 and seven other co-regulated MMnet genes out of 13, regulate bone function. Specifically, global (Pik3cb^-/-, Atp8b2^+/-, Igsf8^-/-, Eml1^-/-, Appl2^-/-, Deptor^-/-) and myeloid-specific Slc40a1 knockout mice displayed abnormal bone phenotypes. We report opposing effects of MMnet genes on bone mass in mice and osteoclast multinucleation/resorption in humans with strong correlation between the two. These results identify MMnet as a functionally conserved network that regulates osteoclast multinucleation and bone mass.

Osteocytes are master regulators of the skeleton. We mapped the transcriptome of osteocytes from different skeletal sites, across age and sexes in mice to reveal genes and molecular programs that control this complex cellular-network. We define an osteocyte transcriptome signature of 1239 genes that distinguishes osteocytes from other cells. 77% have no previously known role in the skeleton and are enriched for genes regulating neuronal network formation, suggesting this program is important in osteocyte communication. We evaluated 19 skeletal parameters in 733 knockout mouse lines and reveal 26 osteocyte transcriptome signature genes that control bone structure and function. We showed osteocyte transcriptome signature genes are enriched for human orthologs that cause monogenic skeletal disorders (P=2.4×10 ^-22 ) and are associated with the polygenic diseases osteoporosis (P=1.8×10 ^-13 ) and osteoarthritis (P=1.6×10 ^-7 ). Thus, we reveal the molecular landscape that regulates osteocyte network formation and function and establish the importance of osteocytes in human skeletal disease.

We thank Dr Faucz and colleagues for sharing their hypothesis that hypothyroidism, rather than growth hormone (GH) excess, may be the main driver for the acrome

Background: Thyroid hormones act in bone and cartilage via thyroid hormone receptor alpha (TRα). In the absence of triiodothyronine (T3), TRα interacts with co-repressors, including nuclear receptor co-repressor-1 (NCoR1), which recruit histone deacetylases (HDACs) and mediate transcriptional repression. Dominant-negative mutations of TRα cause resistance to thyroid hormone alpha (RTHα; OMIM 614450), characterized by excessive repression of T3 target genes leading to delayed skeletal development, growth retardation, and bone dysplasia. Treatment with thyroxine has been of limited benefit, even in mildly affected individuals, and there is a need for new therapeutic strategies. It was hypothesized that (i) the skeletal manifestations of RTHα are mediated by the persistent TRα/NCoR1/HDAC repressor complex containing mutant TRα, and (ii) treatment with the HDAC inhibitor suberoylanilide hydroxamic acid (SAHA) would ameliorate these manifestations. Methods: The skeletal phenotypes of (i) Thra1 ^PV/+ mice, a well characterized model of RTHα; (ii) Ncor1 ^ΔID/ΔID mice, which express an NCoR1 mutant that fails to interact with TRα; and (iii) Thra1 ^PV/+ Ncor1 ^ΔID/ΔID double-mutant adult mice were determined. Wild-type, Thra1 ^PV/+ , Ncor1 ^ΔID/ΔID , and Thra1 ^PV/+ Ncor1 ^ΔID/ΔID double-mutant mice were also treated with SAHA to determine whether HDAC inhibition results in amelioration of skeletal abnormalities. Results: Thra1 ^PV/+: mice had a severe skeletal dysplasia, characterized by short stature, abnormal bone morphology, and increased bone mineral content. Despite normal bone length, Ncor1 ^ΔID/ΔID mice displayed increased cortical bone mass, mineralization, and strength. Thra1 ^PV/+ Ncor1 ^ΔID/ΔID double-mutant mice displayed only a small improvement of skeletal abnormalities compared to Thra1 ^PV/+ mice. Treatment with SAHA to inhibit histone deacetylation had no beneficial or detrimental effects on bone structure, mineralization, or strength in wild-type or mutant mice. Conclusions: These studies indicate treatment with SAHA is unlikely to improve the skeletal manifestations of RTHα. Nevertheless, the findings (i) confirm that TRα1 has a critical role in the regulation of skeletal development and adult bone mass, (ii) suggest a physiological role for alternative co-repressors that interact with TR in skeletal cells, and (iii) demonstrate a novel role for NCoR1 in the regulation of adult bone mass and strength.

In the version of this article initially published, in Fig. 5a, the data in the right column of ‘DAAM2 gRNA1’ were incorrectly plotted as circles indicating ‘untreated’ rather than as squares indicating ‘treated’. The error has been corrected in the HTML and PDF versions of the article.

NFκB is implicated in cancer and bone remodelling, and we have recently reported that the verified NFκB inhibitor Parthenolide (PTN) reduced osteolysis and skeletal tumour growth in models of metastatic breast cancer. Here, we took advantage of in vitro and ex vivo bone cell and organ cultures to study the effects of PTN on the ability of prostate cancer cells and their derived factors to regulate bone cell activity and osteolysis. PTN inhibited the in vitro growth of a panel of human, mouse and rat prostate cancer cells in a concentration-dependent manner with a varying degree of potency. In prostate cancer cell—osteoclast co-cultures, the rat Mat-Ly-Lu, but not human PC3 or mouse RM1-BT, enhanced RANKL stimulated osteoclast formation and PTN reduced these effects without affecting prostate cancer cell viability. In the absence of cancer cells, PTN reduced the support of Mat-Ly-Lu conditioned medium for the adhesion and spreading of osteoclast precursors, and survival of mature osteoclasts. Pre-exposure of osteoblasts to PTN prior to the addition of conditioned medium from Mat-Ly-Lu cells suppressed their ability to support the formation of osteoclasts by inhibition of RANKL/OPG ratio. PTN enhanced the ability of Mat-Ly-Lu derived factors to increase calvarial osteoblast differentiation and growth. Ex vivo, PTN enhanced bone volume in calvaria organ—Mat-Ly-Lu cell co-culture, without affecting Mat-Ly-Lu viability or apoptosis. Mechanistic studies in osteoclasts and osteoblasts confirmed that PTN inhibit NFκB activation related to derived factors from Mat-Ly-Lu cells. Collectively, these findings suggest that pharmacological inhibition of the skeletal NFκB signalling pathway reduces prostate cancer related osteolysis, but further studies in the therapeutic implications of NFκB inhibition in cells of the osteoblastic lineage are needed.

This chapter describes the use of point projection digital microradiography for rapid imaging and quantitation of bone mineral content in mice.

Osteoporosis is characterized by low bone mineral density (BMD) and fragility fracture and affects over 200 million people worldwide. Bone quality describes the material properties that contribute to strength independently of bone mineral density, and its quantitative analysis is a major priority in osteoporosis research. Tissue mineralization is a fundamental process requiring calcium and phosphate transporters. Here we identify impaired bone quality and strength in Slc20a2^‐/‐ mice lacking the phosphate transporter SLC20A2. Juveniles had abnormal endochondral and intramembranous ossification, decreased mineral accrual and short stature. Adults exhibited only small reductions in bone mass and mineralization but a profound impairment of bone strength. Bone quality was severely impaired in Slc20a2^‐/‐ mice: yield load (‐2.3 SD), maximum load (‐1.7 SD), and stiffness (‐2.7 SD) were all below values predicted from their bone mineral content as determined in a cohort of 320 wild‐type controls. These studies identify Slc20a2 as a physiological regulator of tissue mineralization and highlight its critical role in the determination of bone quality and strength. This article is protected by copyright. All rights reserved