Oxidized phospholipids containing phosphatidylcholine (PC-OxPLs), present on oxidized low-density lipoproteins (OxLDL) and apoptotic cells, are pathogenic in osteoporosis. PC-OxPLs are bound by the IgM E06 antibody, which blocks their deleterious effects. Overexpression of the antigen-binding domain of E06 (E06-scFv) increases bone mass in 6-month-old mice and attenuates high fat diet- and age-induced bone loss by increasing osteoblast number and function. The mechanisms mediating the anabolic effect of oxidized phospholipids blockade are unclear. In previous work, we showed that E06-scFv upregulates the expression of Wnt10b and Wnt signaling in vivo. We report herein that the bulk of Wnt10b in the bone microenvironment originates from osteoblast lineage cells. OxLDL decreases the expression of Wnt10b and Wnt target genes in osteoblastic cells, and single cell RNA-sequencing (scRNA-seq) of mesenchymal cells reveals an increase in Wnt10b and Wnt target genes in pre-osteoblasts and osteoblasts of E06-scFv transgenic mice compared to WT. Loss of Wnt10b prevents the increases in BMD by DXA and in vertebral and femoral cancellous bone in 6-month-old E06-scFv transgenic male mice, and attenuates these increases in female mice. Collectively, these findings indicate that oxidized phospholipids decrease bone mass, at least in part, by suppressing Wnt10b.

Juvenile female Klotho-deficient mice have severe cortical abnormalities, but how the remaining bone matrix partitions deformation across length scales is unclear. We combined in situ tensile testing with synchrotron small-angle X-ray diffraction to quantify tissue and mineralized-collagen-fibril deformation in femoral cortical bone from 5-week-old female wild-type, heterozygous, and homozygous Klotho-deficient mice (n = 5 per group). Quantitative backscattered electron imaging (qBEI) assessed tibial cortical thickness, mineralization and the dark-area fraction of the same sections; phase-contrast microtomography of femur provided qualitative structural context. Effective fibril modulus was higher in heterozygous (14.6 ± 3.5 GPa, p = 0.012) and homozygous mice (13.4 ± 5.5 GPa, p = 0.010) than in wild-type mice (4.3 ± 0.8 GPa). The fibril-to-tissue strain ratio was lower in both Klotho-deficient groups (p = 0.018 and p = 0.028), whereas fibril-orientation dispersion did not differ (p = 0.823). Tissue modulus, failure strain, and failure stress showed no statistically significant genotype differences. Tibial cortices were thinner in both Klotho-deficient groups; the dark-area fraction was higher in homozygous mice (p < 0.001), and mean calcium content was lower in homozygous than wild-type mice (p = 0.011). Mineralization heterogeneity did not differ (p = 0.811). These exploratory findings are consistent with altered tissue-to-fibril strain partitioning associated with Klotho deficiency, together with marked cortical structural deficits, but do not establish the molecular origin of the response. The local X-ray dose was estimated at approximately 0.6 kGy per measurement location, well below the absorbed dose at which X-ray irradiation alters the deformation behaviour of cortical bone. Only female mice at a single juvenile age were studied, so the findings cannot be extrapolated to males or to older animals.

Neuroblastoma amplified sequence gene (NBAS) variants are associated with short stature, optic atrophy, and Pelger-Huët anomaly (SOPH) syndrome. We previously identified compound heterozygous variants in NBAS to cause atypical Osteogenesis Imperfecta (OI), with these patients presenting with short stature, developmental delay, and recurrent long-bone fractures. However, skeletal disease progression due to these variants and the disease mechanisms underlying NBAS-associated OI remain poorly understood. Here, we provide a clinical update on previously identified patients and investigate the role of NBAS during skeletal development using zebrafish knockout and patient-specific missense variant zebrafish models. Homozygous knockout larvae exhibited delayed operculum development, reduced bone ossification, and defects in Meckel's cartilage morphology and its underlying cellular structure. Homozygous missense larvae displayed milder cartilage defects without any major defects to early skeletal structures. Seemingly opposing phenotypes were observed in compound heterozygous zebrafish carrying the knockout and missense alleles in trans, with no obvious phenotypes seen in the Meckel's cartilage and accelerated operculum development observed. Together, our results demonstrate that different nbas variants differentially affect skeletal development, suggesting complex spectrums of phenotypic and pathogenic mechanisms in NBAS-associated atypical OI.

Atypical femoral fracture (AFF) is a rare fracture associated with long-term bisphosphonate treatment that occurs in the subtrochanteric and midshaft femur. AFF is not fully explained by bone mineral density motivating identification of biomarkers of AFF risk that reflect bone quality. Cortical bone water has been linked to mechanical properties of bone. Ultra-short echo time magnetic resonance imaging (UTE MRI) can quantify water compartments in bone, including matrix-associated bound water (BW) and porosity-related pore water (PW). Here, we investigated the influence of BW and PW on fracture formation in the AFF region using MRI-based extended finite element modeling and applied fuzzy approximation to capture nonlinear effects between BW and PW on mechanical outcomes. Simulation results showed that individually, mean BW and PW were not correlated with FE-predicted whole femur fracture load or stiffness. Mean porosity index, which is the ratio of PW to total water, was negatively associated with strain at the crack site (R = -0.54, p = 0.026). Stiffness values from BW and PW-based FE simulations were significantly correlated (R = 0.92, p < 0.01), whereas no correlation was observed for fracture load. Fuzzy sensitivity analysis showed that PW and BW are interrelated variables, and their relative influence on stiffness and strain at crack location depends on their levels within the AFF region. These results provide new insights into mechanisms of AFF using MRI including the compartment-specific roles of bone water.

Romosozumab is an antisclerostin antibody that increases bone formation and decreases bone resorption. This study aimed to identify predictors of the response to romosozumab in women with postmenopausal osteoporosis previously treated with denosumab. The study had an observational pre-post design and included 167 women with primary osteoporosis. Romosozumab 210 mg was administered subcutaneously every 4 weeks for 12 months in 87 patients who had been treated with denosumab (DMAb group) and 80 who had not (naïve group). The safety, incidence of new fractures, and changes in bone mineral density (BMD) and bone turnover markers were recorded. At 12 months, a response was defined as a ≥ 3.0% increase in BMD and a non-response as a ≥ 3.0% decrease in BMD. Non-serious adverse events occurred in six patients (3.6%) in each group. New fractures occurred in three patients (4.0%) in the DMAb group and one (1.6%) in the naïve group. At 12 months, the changes in BMD at the lumbar spine and total hip were + 12.6% and + 4.7%, respectively, in the naïve group and + 3.9% and + 0.5% in the DMAb group. Bone turnover markers increased gradually after discontinuation of denosumab. A higher P1NP at baseline was associated with a response to romosozumab and a higher TRACP-5b at 3 months and longer duration of treatment with denosumab were associated with no BMD response at the total hip (TH). Longer denosumab use (≥18 months) followed by romosozumab may be associated with reduced TH BMD response and requires validation in larger prospective studies.

Early life experiences of physical activity have been shown in humans, rodents, and chickens to have long-lasting effects on bone density, structure, and fracture susceptibility. Nevertheless, commercial egg-laying chickens reared with opportunities for exercise still exhibit high fracture incidence. Thus, this study compared two genetic strains of female egg-laying chickens raised in four housing systems differing in structural complexity and the amount and type of exercise possible. At 6, 11, and 16 weeks of age, long bones of the wings and legs were analyzed for length, diameter, breaking strength via 3-point bending, and DEXA-based bone mineral content. Cortical bone density and microstructure was assessed with micro-computed tomography images in tibiae at 16 weeks of age. We found that both rearing environment (i.e. loading history) and genetic strain influenced bone structural and mechanical properties. Compared to conventionally cage-housed chickens, aviary-housed birds consistently exhibited stronger, longer, and wider bones normalized to body weight, as well as tibial microstructural changes associated with increased bone strength. Surprisingly, most improvements compared to caged birds were observed in the moderate-complexity aviary group. The white-feathered genetic strain tended to have longer, stronger bones than the brown (normalized to body weight). Overall, restricted activity was detrimental to bone strength and structure, and bone structure and strength differed between genetic strains of chickens. These data allow for a broader, cross-species understanding of the relationship between bone form and function, and also inform egg farmers when selecting optimal housing environments to improve the welfare and bone health of their flocks.

Patients with Alzheimer's disease (AD) often exhibit abnormal bone loss alongside cognitive impairment. However, the mechanisms linking these two conditions remain insufficiently understood. We first employed amyloid precursor protein/presenilin 1 (APP/PS1) mice as the experimental model and demonstrated that they exhibit significant bone loss concurrent with cognitive impairment. Subsequently, we analyzed public datasets for AD and osteoporosis (OP) and identified CXC chemokine ligand 16 (CXCL16) as a shared hub gene between the two diseases. Transcriptomic sequencing further showed increased CXCL16 expression in the femoral tissue of APP/PS1 mice. Immunoblot analysis of brain and bone tissues, together with serum enzyme-linked immunosorbent assay (ELISA), verified the systemic elevation of CXCL16. These findings suggest that CXCL16 may link cognitive impairment with bone loss in APP/PS1 mice. In vitro experiments showed that CXCL16 did not directly affect the differentiation of bone marrow mesenchymal stem cells or bone marrow-derived macrophages. Instead, CXCL16 acted through neutrophils by promoting their migration and potentiating PMA-stimulated neutrophil extracellular trap (NET) formation. Conditioned medium from NET-producing neutrophils inhibited osteogenic differentiation and promoted osteoclastogenesis. In APP/PS1 mice, antagonism of CXCR6, the cognate receptor for CXCL16, with ML339 improved trabecular bone microarchitecture and osteogenic activity and alleviated cognitive impairment. Degradation of NETs with deoxyribonuclease I (DNase I) produced similar effects. Collectively, these results implicate CXCL16-associated enhancement of NET formation in AD-related skeletal deterioration and highlight the CXCL16-NETs pathway as a possible intervention target for maintaining bone homeostasis.

Implant-associated bone infection involves a complex interplay between pathogenic stimuli and host cell responses, yet analysis in preclinical models has typically relied on qualitative or semi-quantitative measures. We aimed to establish a quantified evaluation framework to define host-pathogen relationships in a preclinical implant infection model. Staphylococcus aureus-coated stainless-steel implants were inserted trans-cortically in mouse tibiae and bone changes recorded longitudinally by in vivo micro-CT. An automated segmentation task list was developed to independently isolate and quantify cortical, periosteal-reactive, and trabecular bone compartments. RGB trichrome histomorphometry was used to quantify bone matrix integrity, osteocyte lacunar geometry, and osteoclastic activity. Droplet digital PCR was used to determine absolute bacterial and host genome copy number. Infected implants produced marked reductions in trabecular bone volume fraction, number, and bone mineral density (BMD), together with decreased cortical bone volume fraction and increased cortical porosity, accompanied by significant elevations in periosteal bone volume fraction. Histologically, infected bone exhibited increased eroded surface indicative of osteoclastic resorption, extensive degraded bone matrix and pathological remodelling of osteocyte lacunae toward circularity, consistent with an osteocytic osteolysis response. Infection-induced changes to cortical bone structure correlated mostly with host cell rather than bacterial load; however, cortical BMD negatively correlated with the bacterial:host genome ratio. This multifaceted, quantified framework reveals distinct pathobiological effects of implant-associated infection on trabecular, cortical, and periosteal bone compartments, bone matrix and osteocyte and osteoclast populations, consistent with reports in human patients, suggesting that major pathological changes are driven by the host bone cell response to infection.

Staphylococcus aureus osteomyelitis drives progressive bone destruction through dysregulated osteoblast-osteoclast crosstalk, yet the role of exosome-mediated circular RNA (circRNA) communication in this process remains poorly understood. This study aimed to determine whether exosomal circRNAs derived from S. aureus-infected osteoblasts orchestrate osteoclast-mediated bone resorption. Exosomes were isolated from hFOB1.19 osteoblasts with or without S. aureus infection, characterized by transmission electron microscopy, nanoparticle tracking analysis, and Western blotting, and their circRNA profiles were screened by RNA sequencing. A murine femoral osteomyelitis model was established via intramedullary S. aureus inoculation followed by tail-vein exosome injection, with bone microarchitecture and osteoclastogenesis assessed by micro-CT, histology, TRAP/ALP staining, qPCR, and Western blotting. The circ_0001843/miR-369-5p/TAFA5 axis was validated by dual-luciferase reporter assays. S. aureus infection markedly upregulated exosomal circ_0001843. In vivo, exosomes from infected osteoblasts aggravated bone destruction, evidenced by decreased BV/TV, increased trabecular separation, enhanced osteoclast activity, and suppressed osteogenic markers. In vitro, circ_0001843 knockdown attenuated RANKL-induced osteoclast differentiation while promoting osteoblast proliferation and mineralization; these protective effects were reversed by exosomes from infected osteoblasts or miR-369-5p inhibition. Mechanistically, circ_0001843 sponged miR-369-5p to derepress TAFA5, thereby disrupting bone remodeling homeostasis. These findings identify exosomal circ_0001843 as a novel mediator of infection-induced bone loss via the miR-369-5p/TAFA5 axis, highlighting a potential therapeutic target for osteomyelitis-associated bone destruction.

Teriparatide (PTH 1-34) is an anabolic agent used to treat osteoporosis, yet clinical response varies widely among patients. To investigate genetic and sex-specific determinants of skeletal response, we administered intermittent PTH to male and female mice from eight genetically diverse inbred strains. Mice were treated for four weeks, and bone phenotypes were assessed via DXA, microCT, and mechanical testing. Response to PTH was highly strain- and sex-dependent, with some strains responding at the femur but not the spine, and vice versa. Heritability estimates for PTH-induced changes in areal bone mineral density (aBMD), cortical bone area at the femoral mid-diaphysis, femur strength, and trabecular bone volume fraction (BV/TV) ranged from moderate to high, with BV/TV showing the strongest genetic influence. Cortical bone response mechanisms differed by sex: males exhibited periosteal expansion, while females showed endosteal remodeling. These findings mirror clinical observations where non-response to PTH can be anatomic site specific. Further, our study suggests that genetic background and sex significantly influence bone response to PTH. Our data support the future use of genetically diverse mouse models to elucidate the genetic architecture of skeletal response to PTH.