Journal of Bone and Mineral Research最新文献

Osteoporosis is a major public health problem among older adults. Forty percent of older U.S. adults take multivitamin/multimineral (MVM) supplementation. The effects of MVM supplementation on fractures is unclear. Preclinical and observational studies suggest that MVM and flavanols may have beneficial effects on bone. We conducted an ancillary study to COcoa Supplement and Multivitamin Outcomes Study (COSMOS; NCT05232669) designed to investigate incident fracture and injurious falls in 21 442 COSMOS participants (12 666 females aged ≥65 years and 8776 males aged ≥60 years) randomized in a 2x2 factorial design to 1 of 4 intervention groups: cocoa extract + MVM, cocoa extract + MVM placebo, cocoa extract placebo + MVM, or double placebo. The daily cocoa extract supplement contained 500 mg/d flavanols and 80 mg/d (-)-epicatechin (Mars Edge); the daily MVM supplement was Centrum Silver® (Haleon). The median (interquartile range) duration of the intervention was 3.6 (3.2-4.2) years. Annually, participants self-reported incident fractures. In intention-to-treat analyses, we examined the effects of cocoa extract and MVM on the primary outcomes of total clinical fracture (hip, upper leg, forearm/wrist, pelvis, upper arm/shoulder, spine, knee, or other), hip fracture, and nonvertebral fracture, and secondary outcomes of clinical spine, forearm/wrist, major osteoporotic, and pelvic fracture using Cox proportional hazards models. During the intervention period, 2083 incident clinical fractures occurred. Compared with placebo, cocoa extract was not significantly associated with lower risk of incident clinical fracture (adjusted hazard ratio [aHR] 1.03, 95% CI 0.95-1.12) or nonvertebral fracture (aHR 1.05, 95% CI 0.96-1.14). MVM supplementation was not associated with lower risk of total clinical fracture (aHR 1.09, 95% CI 1.00-1.19), hip fracture (aHR 1.06, 95% CI 0.80-1.42), or nonvertebral fracture (aHR 1.10, 95% CI 1.00-1.20). These findings do not support the use of cocoa extract or MVM to decrease fracture risk in older individuals not selected for pre-existing osteoporosis.

Lysine demethylase 6B (KDM6B) plays a role in regulating osteoblast differentiation and fetal bone ossification. Nevertheless, its involvement in regulating postnatal bone homeostasis and bone mass accrual remains unclear. In this study, we generated mice lacking Kdm6b gene specifically in mesenchyme and osteoprogenitor cells using a conditional strategy. The adult mice of both mutant strains had decreased cancellous bone mass. The absence of Kdm6b in mesenchyme led to decreased numbers of osteoblasts and osteoclasts, increased marrow adipocytes, as well as repressed bone formation and resorption. Additionally, Kdm6b-deficient bone marrow stromal cells (BMSCs) displayed impaired osteogenic differentiation and exerted an inhibitory effect on osteoclastogenesis. RNA-seq combined with gene expression analysis uncovered downregulation of collagen triple helix repeat containing 1 (CTHRC1) and a lower receptor activator of nuclear factor κB ligand (RANKL)/osteoprotegerin (OPG) ratio in BMSCs of the mutant mice. Further mechanistic explorations demonstrated that KDM6B epigenetically upregulated CTHRC1 expression by removing the repressive H3K27me3 mark from its promoter, thereby triggering PKCδ/MAPKs signaling to facilitate osteoblast differentiation. CTHRC1 was able to mitigate the dysregulated osteogenic and adipogenic differentiation induced by Kdm6b deficiency. This study provides evidence that KDM6B regulates postnatal bone homeostasis through balancing osteoblast and osteoclast differentiation. Given its predominant promotion of osteoblastic bone formation over osteoclastic bone resorption, KDM6B tends to promote postnatal bone mass accrual.

Multiple sclerosis (MS) is a chronic autoimmune disorder of the central nervous system (CNS) and a common cause for neurological disabilities in young adults. Although it is known that the peripheral immune landscape is altered in people with MS (pwMS), the impact on other organ systems than the CNS is frequently overlooked. In addition to neurological deficits, pwMS suffer from impaired bone health and increased fracture risk. However, the mechanisms underlying bone loss in pwMS are poorly understood. Here, we investigated the compartment-specific bone microarchitecture as well as cellular and molecular mechanisms of altered bone remodeling in pwMS and the corresponding mouse model of experimental autoimmune encephalomyelitis (EAE). We show that pwMS and EAE mice have reduced bone mineral density characterized by a combined loss of trabecular and cortical bone. Intriguingly, bone loss in EAE followed a biphasic dynamic defined by increased osteocyte apoptosis associated with decreased bone formation in acute EAE and increased bone resorption in the chronic phase, which could be explained by increased CXCL13/CXCR5 signaling. In conclusion, the identified stage-dependent mechanism for bone loss in EAE may help to develop improved strategies for osteoporosis treatment in pwMS.

Cerebral palsy (CP) is a non-progressive neurological syndrome resulting in abnormal muscle tone, movement, and posture. It is unclear whether ambulatory children with CP have deficits in bone quantity or quality. Furthermore, the relationship between abnormal muscle tone, altered function, and bone health remains largely unexplored. This observational study investigated bone mineral density (BMD) and microarchitecture in ambulatory children with spastic CP and associations of BMD with function, muscle spasticity, and gait. Children with spasticity in both lower limbs (n = 12) aged 3-8 years were recruited. Areal BMD was measured with dual-energy x-ray absorptiometry (DXA) at the proximal femur and lateral distal femur and compared to normative data. High-resolution peripheral quantitative computed tomography (HR-pQCT) was performed at the metaphyseal tibia and radius in a subset of participants (n = 5) and compared to healthy children (n = 7). Gait pathology and cardiopulmonary function were investigated with the Gait Deviation Index, Edinburgh Visual Gait Score, and energy expenditure index. DXA aBMD Z-scores at the lateral distal femur were within a normal range. Instead, the CP group's median aBMD Z-score at the proximal femur was -1.8 (interquartile range: -2.2, -1.2, p=.03) indicating potential skeletal fragility. Strong correlations were found between gait pathology and DXA-based bone outcomes (correlation coefficient 0.62 (p=.04) to 0.73 (p=.01)) as well as energy expenditure index and DXA-based bone outcomes (correlation coefficient -0.63 (p=.03) to -0.98 (p = <0.001)). At the metaphyseal tibia, children with spastic CP had significant deficits in HR-pQCT-measured bone geometry and trabecular microarchitecture: 35% lower total area, 42% lower trabecular area, and 48% lower trabecular number than controls. HR-pQCT parameters were similar between groups at the metaphyseal radius. These differences in tibial metaphysis size and trabecular microarchitecture are similar to those observed in disuse and thus could be a result of abnormal biomechanics or low levels of physical activity.

Osteogenesis imperfecta (OI) Type V is typically characterized by radial head dislocation, calcification of interosseous membrane and hyperplastic callus. It is caused by the c.-14C > T mutation in the 5' UTR of IFITM5 gene, adding five amino acids (MALEP) to the N-terminal of IFITM5 protein. Previous studies have suggested a neomorphic function of the MALEP-IFITM5 protein. However, the underlying mechanisms remain unclear due to embryonic lethality in previous mouse models. Therefore, we developed an inducible mouse model (Ifitm5flox c.-14C > T) that could be induced by Cre expressed at different developmental stages to explore the pathogenic effects of the neomorphic MALEP-IFITM5. The mutant Ifitm5 allele could be regulated by the endogenous regulatory elements after Cre recombination, maintaining its spatiotemporal expression pattern and physiological level. Specifically, Prx1-Cre; Ifitm5flox c.-14C > T mutant mice were born with fractures in all limbs, showing impaired ossification and enhanced chondrogenesis associated with increased SOX9 abundance. Analyses of single-cell RNA sequencing data revealed arrested osteogenesis in Prx1-Cre; Ifitm5flox c.-14C > T mouse. A major population of cells expressing both osteogenic and chondrogenic signature genes was identified in the mutant mouse. Reduced expression of SP7 and SOST in the cortical regions of mutant mice confirmed delayed osteocyte maturation and compromised osteogenesis. Elevated bone marrow adipocytes were found in the adult mutant mice. Ectopic chondrogenesis and SOX9 expression were also observed in the perichondrium regions of Col1a1-Cre; Ifitm5flox c.-14C > T and Ocn-Cre; Ifitm5flox c.-14C > T mutant mice. The inducible Ifitm5flox c.-14C > T mouse model and integrated single-cell transcriptomic analyses elucidated that ectopic expression of SOX9 and disrupted homeostatic balance among osteogenesis, chondrogenesis and adipogenesis may contribute to the pathogenesis caused by MALEP-IFITM5, helping to gain deeper insights into the molecular mechanisms of type V OI.

Cementocytes are terminally differentiated cells embedded in cellular cementum, an important hard tissue covering the apical regions of tooth roots. However, the roles of cementocytes in cellular cementum remain enigmatic. Here, we show that Murine Double Minute 2 (Mdm2), an E3 ubiquitin ligase that plays vital roles in regulating cell proliferation, apoptosis, and differentiation to influence tissue or organ development, is highly expressed in the cementocytes of mice. To investigate the role of cementocyte-expressed Mdm2, Dmp1-Cre;Mdm2flox/flox (Mdm2 cKO)mice were obtained to inactivate Mdm2 in cementocytes. The results showed that Mdm2 was successfully ablated and Mdm2 cKO mice display increased cementocyte apoptosis and reduced cellular cementum volume. p53, the canonical substrate of Mdm2, was accumulated and hyperactivated in the cementocytes of Mdm2 cKO mice and in cultured IDG-CM6 cells (a cementocyte cell line) treated with Nutlin3a, an inhibitor of Mdm2. Further experiments showed that inactivation of one allele of p53 significantly rescued the increased cementocyte apoptosis and the decreased cellular cementum volume in Mdm2 cKO mice. Therefore, p53 is targeted by Mdm2 for degradation and mediates the role of Mdm2 in cementocyte survival and cellular cementum volume. Notably, Mdm2 cKO mice exhibited decreased differentiation of cementoblasts (the cell type primarily responsible for cementum deposition) and reduced rate of cellular cementum deposition. Meanwhile, OCCM-30 cells (a cementoblast cell line) showed diminished migration, proliferation, differentiation, and mineralization ability after culture with conditioned medium (CM) from Nutlin3a-pretreated IDG-CM6 cells. Intriguingly, Mdm2 cKO mice displayed significantly increased osteoclast formation and cementum resorption. Meanwhile, in vitro experiments verified that CM from Nutlin3a-pretreated IDG-CM6 cells induced osteoclast differentiation of bone marrow macrophages. Collectively, these results demonstrate that Mdm2-mediated degradation of p53 promotes cementocyte survival, and that cementocytes affect the cell behaviors of cementoblasts and osteoclasts through a paracrine mode to modulate cellular cementum volume.

Autosomal dominant hypocalcemia (ADH) is due to enhanced calcium-dependent signaling caused by heterozygous gain-of-function (GOF) variants in the CASR gene (ADH1) or in the GNA11 gene, encoding Gα11 (ADH2). Both ADH1 and ADH2 are associated with hypocalcemia and normal or inappropriately low levels of circulating PTH. ADH1 patients typically manifest hypercalciuria, while ADH2 is associated with short stature in approximately 42% of cases. We evaluated a 10-yr-old boy with hypoparathyroidism and short stature. Biochemical analyses revealed hypocalcemia, hyperphosphatemia, and inconsistent hypercalciuria. Genetic analyses revealed a de novo heterozygous p.Leu723Arg variant in CASR. We characterized the expression of recombinant WT and Leu723Arg calcium-sensing receptor (CaSR) proteins in HEK293 cells and assessed G protein activation in vitro by CaSR using bioluminescence resonance energy transfer. Transient expression studies showed the Leu723Arg variant was normally expressed but resulted in a significantly lower EC50 for extracellular calcium activation of G11 but not other G proteins (ie, Gi, Gq, Gs). The Leu723Arg substitution has a novel GOF phenotype that leads to biased CaSR activation of G11 signaling, suggesting that residue 723 specifies activation of G11 but not other G proteins. Similar studies of a previously described CaSR variant associated with hypoparathyroidism and short stature, Leu616Val, showed no changes in any G protein pathways, indicating it is likely to be a benign variant. Given the preferential activation of G11 by the Leu723Arg CaSR variant, we propose that the patient's short stature shares a similar basis to that in patients with ADH2 due to GOF variants in GNA11.

Bone homeostasis within the skeletal system is predominantly maintained by bone formation and resorption, where formation of new bone involves maturation of stromal cells to mineral and matrix secreting mature osteoblasts, which requires cellular energy or adenosine triphosphate. Alterations in systemic metabolism can influence osteoblast function. In line with this, type 2 diabetes mellitus (T2DM), a common metabolic disorder is also associated with reduced bone formation and increased risk of fracture. Impairment in lipid metabolism is one of the key features associated with T2DM-related pathologies in multiple tissues. Therefore, we tested the hypothesis that the reduced bone formation reported in obese murine models of impaired glucose tolerance is a function of disrupted lipid metabolism in osteoblasts. We first confirmed that mice fed a high-fat diet (HFD) have reduced bone microarchitecture along with lower bone formation rates. Interestingly, osteoblasts from obese mice harbor higher numbers of cytosolic lipid droplets along with decreased bioenergetic profiles compared to control cells. Further supporting this observation, bone cortex demonstrated higher total lipid content in HFD fed mice compared to control-fed mice. As a further proof of principle, we generated a novel murine model to conditionally delete Plin2 in osteoblast-progenitor cells using Prrx1-Cre, to enhance lipid droplet breakdown. Our data demonstrate that knocking down Plin2 in an osteoprogenitor specific manner protects from HFD induced osteoblast dysfunction. Furthermore, the mechanism of action involves enhanced osteoblast fatty acid oxidation. In conclusion, the current studies establish that HFD induced glucose intolerance leads to perturbations in osteoblast lipid metabolism, thus causing lower bone formation, which can be protected against by increasing fatty acid oxidation.

Motivated by studies showing an association between beta blocker (BB) use and positive bone outcomes, a pilot randomized control trial was performed at the Mayo Clinic which randomized postmenopausal women to placebo, propranolol (40 or 80 mg twice daily), atenolol (50 mg/d), or nebivolol (5 mg/d) to determine changes in bone turnover markers (BTMs) and in BMD over 20 wk. Pharmacogenetic effects and microRNA-mediated mechanisms involving beta adrenergic receptor and related genes have previously been found. We sought to validate these effects and discover new candidates in an ancillary study to the pilot clinical trial. We genotyped all participants and performed microRNA (miRNA) sequencing at baseline and at 20 wk for 24 participants from the atenolol or placebo groups. We discovered several variants in ADRB1, ADRB2, and HDAC4 which showed significant pharmacogenetic effects with BMD at multiple sites and with BTMs. Our miRNA results showed a significant treatment effect for miR-19a-3p over time with atenolol use in the low-responder group compared to placebo. Overall, the longitudinal miRNA analysis showed a large number of miRNAs which were up-regulated over the trial in the low responders but not the high responders compared to placebo, of which miR-19a-3p was one example. Finally, we compared the response to atenolol treatment for cardiovascular traits (pulse and blood pressure) with the response for the bone resorption marker, C-terminal telopeptide, and found a largely independent effect. Our results have implications for personalized therapy and for understanding mechanisms of BB treatment effect on bone.