JBMR Plus

Three-dimensional dual-energy X-ray absorptiometry (3D-DXA) reconstructs proximal femur models from standard scans to estimate cortical and trabecular bone parameters. The aim of this study was to evaluate 3D-DXA against quantitative computed tomography (QCT) across independent international cohorts. The study included 537 subjects from four cohorts: an adult population from Spain, a postmenopausal female population from the United States, an osteoarthrosis population and a young population, both from Japan. Subjects underwent both 3D-DXA and QCT imaging. Accuracy was assessed using linear regression and Bland-Altman analysis to evaluate systematic and random errors. 3D-DXA parameters strongly correlated with QCT across all datasets, with correlation coefficients between 0.82 and 0.97. Random errors were consistent across cohorts and ranged between 16.55 and 19.91 mg/cm3 for integral volumetric bone mineral density (vBMD), between 13.52 and 18.47 mg/cm3 for trabecular vBMD, and between 9.13 and 11.37 mg/cm2 for cortical surface bone mineral density (sBMD). Systematic errors ranged between -14.84 and 4.50 mg/cm3 for integral vBMD, between -8.31 and 14.41 mg/cm3 for trabecular vBMD, and between -5.58 and 3.21 mg/cm2 for cortical sBMD. The variations in systematic errors were likely attributable to differences in QCT acquisition protocols. Overall, these results demonstrate consistent agreement between 3D-DXA and QCT across sex, age, ethnicity, geographic regions, and clinical profiles. Taken together, these findings support the use of 3D-DXA as an accurate, non-invasive, and clinically accessible technology for advanced assessment of the cortical and trabecular compartments of the proximal femur.

PurposeOsteoporosis and associated hip fractures are a major public health concern. Dualenergy X-ray Absorptiometry (DXA) remains the diagnostic gold standard, but its areal (a) bone mineral density (BMD) measurements have limited sensitivity, as many fractures occur at T-scores above -2.5. Three-dimensional (3D) DXA provides compartment-specific volumetric parameters of the hip, potentially improving osteoporosis management. This study aimed to establish reference data for 3D-DXA parameters to improve osteoporosis management and investigate potential compartmental imbalances at the hip. MethodThis multicenter, cross-sectional, population-based observational study (SEIOMM-3D-DXA project), supported by the Spanish Society for Bone and Mineral Metabolism (SEIOMM), analyzed hip DXA scans from 1366 Spanish men and women across six centers. 3D-DXA analyses were conducted using the 3D-Shaper software (3D-Shaper Medical, Barcelona, Spain), producing estimates of trabecular volumetric (v) BMD and cortical surface (s) BMD. Age- and sex-specific reference curves were generated using the LMS method, and thresholds were established based on regression with T-score values. Moreover, trabecular vBMD and cortical sBMD Z-scores were calculated to evaluate potential compartmental imbalances. ResultsThe derived aBMD curves closely aligned with the NHANES III Caucasian reference. Sex-specific thresholds for trabecular vBMD and cortical sBMD were established for patient stratification. Z-score comparisons revealed significant discrepancies between trabecular and cortical compartments in 52.0% of females and 48.7% of males, underlining the importance of compartment-specific bone assessment. ConclusionsThis study establishes reference curves for clinical interpretation of 3D-DXA parameters and demonstrates the potential of 3D-DXA to capture compartmental imbalances at the hip. Mini AbstractIn this study, hip scans from over a thousand men and women in Spain were analyzed to create normative reference values for 3D-DXA parameters. These results can help doctors better stratify people based on the status of each part of the bone and improve the management of osteoporosis.

BackgroundBone formation during development and repair is divergently modulated by osteoblast (OB)-derived vascular endothelial growth factor (VEGF) which drives the skeletal sexual dimorphism of the bone vasculature. While the extracellular matrix (ECM) provides both structural and instructive cues to developing vasculature, the contributions of the bone matrix to this skeletal vascular dimorphism in bone remains undefined at the cellular level. MethodsPrimary OBs were isolated from neonatal female and male C57BL/6 long bones and cultured under basal or osteogenic conditions. ECM composition was quantified by Raman spectroscopy. Primary murine bone marrow endothelial cells (BMECs) were seeded directly onto established OB layers and maintained in heterotypic cocultures to assess contact-mediated effects of OB ECM on BMEC survival and expansion. OB-conditioned media (CM) was used to evaluate soluble-factor contributions, with VEGF-A concentration quantified by ELISA. ResultsRaman spectroscopy, on individual OBs from monotypic cultures, revealed sexually dimorphic ECM signatures that were independent of cellular growth profiles. Female OB matrices were enriched with type I collagen-specific proline and hydroxyproline and octacalcium phosphate with enhanced collagen intra-strand stability consistent with a matrix-dominant signature. Male OB matrices exhibited relatively lower type I collagen content and higher carbonated apatite resulting in an elevated mineral-to-matrix ratio indicative of advanced mineral maturation. After 24-hours of heterotypic culture of BMECs with OBs, BMEC numbers were 1.39-fold higher when in contact with male OBs. CM treatment of BMECs did not recapitulate these effects despite higher VEGF-A release from male OBs. ConclusionsSex differences in OB ECM are linked to divergent, contact-dependent regulation of BMEC behaviour. These findings suggest that differences in matrix maturation stat contribute to the sex-specific regulation of the skeletal vascular niche. Elucidating the mechanisms that regulate sex-specific OB-ECM production may reveal new therapeutic targets for selectively modulating pathological skeletal angiogenesis in men and women. SummaryBone is a sexually dimorphic organ, with men and women differing in bone size, strength and risk of fracture. The skeletal vasculature is essential for bone growth and repair, with bone forming osteoblast (OB) cells influencing blood vessel development through the skeletal extracellular matrix (ECM). Although the interactions between OB and vascular cells are crucial for lifelong skeletal health, it is not yet known whether sex differences in bone structure between men and women arise from differences in OB activity, or whether this divergence is driven by sex differences in blood vessel growth. Here, we show that male and female mouse OBs deposit distinct ECMs that differentially influence vascular endothelial cell behaviour. Female OBs produce a collagen-rich matrix with low mineral content. In contrast, male OB matrices contain less collagen and more mineral while releasing elevated levels of blood vessel promoting VEGF-A than females. When placed directly onto these OBs, vascular cell growth was greater when in contact with male than female OBs. Notably, this sex-dependent effect requires direct contact between both cell types and was not reproduced by exposure to OB-derived substances alone. These findings identify a cellular mechanism by which sex differences in OB matrix composition influences vascular cell behaviour in bone. Understanding how OB-vascular interactions differ by sex may help explain variation in bone health, healing capacity and disease risk between men and women. Further, our approach may support the discovery of new therapeutic targets that support bone growth and repair while targeting abnormal blood vessel growth in a sex-specific manner. HighlightsO_LIPrimary OBs from male and female C57BL/6 mouse long bones synthesise compositionally distinct ECMs. C_LIO_LIFemale OB matrices are type I collagen-rich and enriched with octacalcium phosphate, whereas male OB matrices contain less type I collagen and higher levels of carbonated apatite. C_LIO_LIBone marrow-derived endothelial cell (BMEC) growth is enhanced in heterotypic cocultures with male, but not female, OBs after 24 hours. C_LIO_LIMale OBs release higher levels of the pro-angiogenic factor VEGF-A than female OBs. C_LIO_LIThe sex-specific effects of the OB ECM on BMECs is contact-dependent and are not reproduced by treatment with OB-derived conditioned media. C_LI

Osteoblasts generate bone by secreting collagen and mineralizing it in response to various signaling cues. We have previously shown that the majority of ATP generated by differentiated osteoblasts in response to glucose is through glycolysis in contrast to undifferentiated cells that are more dependent on oxidative phosphorylation. To confirm our previous findings, metabolomics was performed for unlabeled polar metabolites, revealing elevated glycolytic metabolites at the later stages of differentiation. Krebs cycle (TCA cycle) metabolites were also changed confirming metabolic rerouting with differentiation. We hypothesized that an increase in mitophagy shifts ATP generation towards glycolysis resulting in the observed bioenergetic and metabolic changes. Utilizing calvarial osteoblasts isolated from a mitophagy reporter mouse model (MitoQC), an increase in mitophagy and the mitophagy receptor, Bnip3, was observed with osteoblast differentiation. KD of Bnip3 in osteoblasts inhibited differentiation and mineralization arising from impaired mitochondrial function. In vivo, male Bnip3 null mice exhibited a significant decrease in osteoblast numbers resulting in lower bone mass. Mechanistically we identified decreased fusion and increased fission factors, impaired stress signaling and increased proapoptotic factors in the absence of Bnip3. These data demonstrate for the first time that BNIP3 expression and mitophagy during osteoblast differentiation are necessary for relieving mitochondrial stress to maintain optimal bone mass.

3D-DXA reconstructs DXA hip scans to 3-dimensional images allowing measurement of trabecular and cortical bone parameters. Given the higher image quality of GE Healthcare iDXA than GE Healthcare Prodigy, it could be hypothesized that the reconstruction might differ, thereby affecting 3D-DXA results. The aim of the study was to assess agreement and precision of 3D-DXA cortical and trabecular femur parameters between Prodigy and iDXA densitometers in adult subjects. The study cohort was composed of 391 men and women recruited from 3 clinical centers (USA and Brazil). All subjects were scanned on either Prodigy or iDXA scanners. Short-term precision was assessed on two Prodigy and two iDXA densitometers. 3D-DXA analyses were performed using 3D-Shaper software version 2.14. Agreement between densitometers was assessed by regression and Bland-Altman analyses. Short-term precision was determined following International Society for Clinical Densitometry recommendations. Strong agreements for 3D-DXA parameters were obtained between devices regardless of the center or the DXA device model (all R2 > 0.96). Bland-Altman analyses demonstrated statistically (p < 0.05), but not clinically, significant difference between both aBMD and 3D-DXA measurements obtained using Prodigy and iDXA scanners. Short-term precision of areal BMD and 3D-DXA parameters was similar between densitometers. This study demonstrated excellent 3D-DXA measurement agreement and similar precision between iDXA and Prodigy densitometers. These data provide evidence that no adjustments are required when using 3D-Shaper software on iDXA or Prodigy instruments. Mini AbstractWe assessed agreement and precision of 3D-DXA parameters between GE Healthcare Prodigy and iDXA densitometers in adults. Strong agreement was observed between devices, and short-term precision was comparable. Findings indicate that no adjustment is needed when using 3D-DXA with GE Healthcare densitometers.

Bone is a dynamic tissue that continuously adapts its structure in response to mechanical loading, an essential process for maintaining skeletal health. However, this adaptive capacity declines with aging, contributing to increased fragility and fracture risk. Developing therapeutic strategies that preserve or restore bone mechanoadaptation in patients with increased bone fragility requires identifying key molecular regulators of this process. We applied spatial transcriptomics (GeoMx, NanoString) to characterize gene expression changes induced by mechanical loading in the murine tibia, focusing on periosteal and bone compartments in regions under tension and compression. Spatial data were validated and cross-compared with previously published bulk RNA-seq and laser-capture microdissection datasets, identifying a set of 12 genes consistently regulated by loading across independent platforms and laboratories. As part of a functional analysis, we selected Slc13a5, a citrate transporter implicated in bone mineralization and metabolism. Conditional deletion of Slc13a5 in osteolineage cells using Osteocalcin-Cre significantly increased the loading-induced mineralizing surface in tensile regions compared with Cre- Slc13a5fl/fl littermates. In addition, Slc13a5 cKO mice exhibited lower resorption around the neutral axis after loading compared to controls. Together, these findings identify Slc13a5 as a regulator of bone adaptation in regions experiencing low mechanical stimulation and suggest it as a potential therapeutic target for conditions characterized by impaired mechanoadaptive responses. This study highlights spatial transcriptomics as a powerful gene discovery framework for bone, enabling identification of novel targets to understand mechanisms and develop therapies. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=181 SRC="FIGDIR/small/711126v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@5bf180org.highwire.dtl.DTLVardef@4c33b7org.highwire.dtl.DTLVardef@d75668org.highwire.dtl.DTLVardef@169fa97_HPS_FORMAT_FIGEXP M_FIG C_FIG

Protein kinase A (PKA) is involved in bone biology and is a key mediator of parathyroid hormone signaling in the osteoblast. However, the consequences of sustained PKA activation in bone are unclear. In this study, we inducibly activated PKA in osteoblasts by deleting its major regulatory subunit, Prkar1a, using a Col11-driven Cre system. Prkar1aob-/-mice demonstrated rapid and profound bone pathologies in their femurs, lumbar and caudal vertebrae with cortical bone breakdown and cortical trabecularization. This phenotype was characterized by increased bone turnover and elevated osteoblastic and osteoclastic activities. Transcriptomic and qPCR analyses showed an impairment of osteoblast differentiation with a defect in ossification, expansion of stromal cells, and numbers of both osteoblastic and osteoclastic precursors. Moreover, there were alterations in gene expression of chemokines and Wnt members with enhanced osteoclastogenesis. Altogether, activation of PKA in osteoblasts by inducible deletion of Prkar1a causes a profound high bone turnover phenotype resembling several human bone diseases.

Social isolation is a known modifiable risk factor for many chronic diseases including cardiovascular, metabolic, and brain disorders. Recent research has demonstrated that social isolation is similarly detrimental to skeletal health, but these effects may be sexually dimorphic. In rodents, isolation negatively affects bone in adult male mice, but not in females. However, these sex differences have not been systematically investigated, and it is unknown if they persist with long-term social isolation. The goal of our study was to investigate if isolation-induced bone loss may occur on different timescales between female and male mice, as well as investigate the potential roles of estrogen and testosterone. We examined bone changes in grouped (4 mice/cage) or isolated (1 mouse/cage) female and male 16-week-old C57BL/6J mice after 2, 4, or 8 weeks of treatment. We found that social isolation through single housing significantly reduced bone parameters across treatment lengths in male mice (20% reduction in Tb.BV/TV; 8% reduction in Ct.Th.) but not in females even with prolonged isolation. Isolation also decreased biomechanical properties in the femur of male but not female mice. While the females overall bone phenotype was unaffected, isolated females did show an increase in bone turnover markers with as little as 2 weeks of isolation. Isolation also altered estrogen-related gene expression in male mice isolated for 4 or 8 weeks. Overall, our results demonstrate that short- and long-term social isolation has sexually dimorphic effects on murine bone. These findings have important clinical implications for individuals at risk for social isolation, as well as for pre-clinical rodent models utilizing single housing.

Purpose To determine whether clinically significant weight loss (>5% of body weight) is associated with slower 2-year knee cartilage degeneration in individuals with and without radiographic osteoarthritis. This study used a cartilage structural assessment score derived from the spatial distribution of cartilage thickness, referred to as the Cartilage Thickness Score (CTh-Score). It is based on cartilage thickness patterns and scores the cartilage between 0 and 100, with higher scores indicating greater severity. Methods We conducted a retrospective matched cohort study within the Osteoarthritis Initiative. High-resolution cartilage thickness maps (CTh-Maps), along with their corresponding CTh-Score, were extracted from a public repository. Participants with complete radiographic and MRI data at baseline and 24 months were stratified by baseline Kellgren-Lawrence (KL) grade into non-radiographic OA (non-ROA; KL<2) and radiographic OA (ROA; KL>=2). Within strata, cases (>5% 2-year weight loss) were propensity score-matched 1:2 to weight-stable controls on age, sex, height, weight, KL grade, joint space width (JSW), KOOS Pain, baseline CTh-Score, and mean cartilage thickness in the medial and lateral femoral and tibial compartments. The primary outcome was 2-year change (delta) in CTh-Score, where higher values indicate worsening. Secondary outcomes were delta JSW, delta regional mean cartilage thickness, and delta KOOS Pain. Non-parametric tests were used. Results We included 164 cases and 328 controls in non-ROA, and 266 cases and 532 controls in ROA. Median (interquartile range) weight loss was -6.10 kg (-8.90, -4.70) versus +0.30 kg (-1.30, 2.20) in non-ROA and -6.80 kg (-9.10, -5.02) versus +0.40 kg (-1.40, 2.82) in ROA (both p<0.001). Weight loss was associated with significantly smaller 2-year increases in CTh-Score: in non-ROA, median 1.58 (0.61, 6.53) vs 3.14 (0.44, 7.12) (p=0.005); in ROA, median 1.69 (0.97, 6.71) vs 2.90 (0.19, 7.38) (p=0.004). No between-group differences were detected for delta JSW or delta regional mean cartilage thickness in any of the 4 ROIs. A trend toward greater KOOS Pain improvement with weight loss was observed in ROA: 2.75 (-3.35, 13.40) vs 0.00 (-5.60, 8.40) (p=0.06). Conclusions Achieving >5% weight loss over 2 years is associated with approximately 50% lower progression in median cartilage degeneration, as assessed by CTh-Score, in both non-ROA and ROA. No change was observed with conventional structural metrics. These findings support weight management as a structural disease-modifying strategy and highlight CTh-Score as a sensitive endpoint.

Individuals with chronic kidney disease (CKD) have higher rates of hip fracture and post-fracture mortality. Although they may develop age-related osteoporosis similar to those without CKD, they may also exhibit CKD-related metabolic bone disease (MBD), characterized by low, high, or mixed turnover at similar levels of bone mineral density (BMD). Because BMD does not provide information about turnover status, clinical decision-making is challenging. This study evaluated the associations between circulating bone-turnover biomarkers and static histomorphometry in patients undergoing hip-fracture surgery. In this cross-sectional study, we enrolled adults with and without CKD, defined as estimated glomerular filtration rate (eGFR) [&le;]60 ml/min/1.73m{superscript 2} (CKD-EPI 2021), undergoing hip-fracture surgery. Blood samples, bone specimens from the femoral head or greater trochanter, and demographic and clinical data were collected at the time of surgery. Plasma biomarkers included -Klotho, bone alkaline phosphatase (BAP), dickkopf-related protein 1 (DKK-1), fibroblast growth factor 23 (FGF23), tartrate-resistant acid phosphatase 5b (TRAP5b), parathyroid hormone (PTH), and sclerostin. Logistic regression models, adjusted for age, gender, eGFR, and osteoporosis, assessed associations with CKD status. Tertiles of osteoblast surface (Ob.S/BS) and eroded surface (ES/BS) were defined in participants without CKD and applied to the full cohort. Multinomial and multivariable linear regression evaluated associations of biomarkers with these histomorphometry parameters. Among 97 enrolled participants (mean age 80 {+/-} 11 years; 67% female), 68% had CKD. Of 75 with complete biomarker and histomorphometry data, 96% demonstrated low bone turnover. CKD was associated with lower trabecular thickness (Tb.Th) and higher osteoid thickness (O.Th), osteoid volume (OV/BV), and osteoid surface (OS/BS), suggesting thinner, largely unmineralized trabeculae. Higher BAP (222.2% difference per doubling; 95% CI 77.2-485.8) and TRAP5b (319.3%; 95% CI 128.3-669.5) were directly associated with Ob.S/BS and ES/BS, whereas sclerostin was inversely associated with ES/BS (-28.9%; 95% CI -44.8 to -7.1). PTH was not associated with bone-turnover measures. These findings suggest that BAP, TRAP5b, and sclerostin may provide useful adjunct information alongside PTH for assessing bone turnover and guiding therapy in patients with and without CKD.

The bone matrix is precisely maintained and optimized to resist fractures. However, aging and disease deteriorate the bone matrix and increase fragility. Individuals with type 2 diabetes (T2D) have an elevated risk of bone fracture despite apparently normal bone mass. The chronic hyperglycemia in T2D promotes the formation of advanced glycation end-products (AGEs) in the bone tissue and modify the matrix mechanics. AGEs also bind to its receptor, RAGE, to activate inflammation and alter homeostasis. Using a leptin-receptor deficient mouse model of diabetes, we used a combination of high-resolution methods across multiple scales to evaluate the microarchitectural-, material- and cellular- level changes affected by the modulation of RAGE. To demonstrate the relevance of RAGE, we genetically ablated RAGE (RAGE-null) before the onset of diabetes; and to demonstrate the potency of RAGE as a disease modifying therapy, a RAGE antagonist (FPS-ZM1) was administered after prolonged diabetes. Diabetes impaired bone microstructure, the homeostatic actions of bone cells, the bone matrix nanomechanics, and whole- bone strength. The constitutive ablation of RAGE in diabetic animals prevented AGEs accumulation and the decline of trabecular connectivity; protected against the loss of osteocyte lacunae density and morphology; and maintained the matrix nanomechanics and bone strength. The inhibition of RAGE after the onset of diabetes reversed AGE accumulation and loss of bone volume; rescued osteocyte lacunae density and osteoclast activity; and restored matrix nanomechanics and bone strength. These results suggest that RAGE is a viable therapeutic target for diabetes-mediated impairments of bone quality. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=129 SRC="FIGDIR/small/716153v1_ufig1.gif" ALT="Figure 1000"> View larger version (26K): org.highwire.dtl.DTLVardef@6d8a5borg.highwire.dtl.DTLVardef@1967b5borg.highwire.dtl.DTLVardef@7cb1edorg.highwire.dtl.DTLVardef@85491d_HPS_FORMAT_FIGEXP M_FIG C_FIG

Osteocytes play a central role in bone remodeling, mineral metabolism, and skeletal homeostasis, but direct molecular analysis of human osteocytes remains technically challenging because they are embedded within the mineralized bone matrix. Surgically obtained human bone specimens provide valuable material for studying human bone biology; however, surface-associated cells, marrow-derived cells, and adherent soft tissues can confound downstream transcript analysis. Here, we describe a bone fragment-based protocol for preparing surgically obtained human bone specimens for molecular analysis of osteocyte-associated transcripts. The protocol consists of mechanical trimming, mincing into small bone fragments, repeated washing, and five sequential rounds of collagenase digestion to reduce non-osteocytic cellular components associated with the bone surface and marrow spaces. The remaining mineralized bone fragments are then frozen in liquid nitrogen, cryogenically pulverized, and lysed in TRIzol reagent for total RNA extraction. Histological validation using residual maxillary bone specimens showed that sequential collagenase digestion markedly reduced adherent soft tissue and extra-matrix nuclei while preserving osteocyte lacunar occupancy. This protocol provides a practical workflow for bone fragment-based RNA analysis focused on osteocyte-associated transcripts in human bone specimens. Specifications table O_TBL View this table: org.highwire.dtl.DTLVardef@1cec618org.highwire.dtl.DTLVardef@2f746forg.highwire.dtl.DTLVardef@1854247org.highwire.dtl.DTLVardef@1c26c1aorg.highwire.dtl.DTLVardef@1473a88_HPS_FORMAT_FIGEXP M_TBL C_TBL

Background Automated pelvic CT segmentation has advanced to reliable coarse bone extraction. Yet the structured anatomical hierarchy required for morphometry, fixation planning, bone quality mapping, and arthroplasty workflows remains unachieved. This study developed and validated a fully automated anatomy-informed pipeline that converts standard pelvic CT into a comprehensive, surgeon-readable subsegmentation of the pelvis and proximal femur. Methods Pelvic CT datasets were retrospectively collected from anonymized archives of hospitals affiliated with the Directorate of Health Affairs, Sharqia, Egypt. After eligibility screening, 757 normal adult cases were processed using a custom one-click 3D Slicer pipeline integrating TotalSegmentator for coarse extraction, followed by deterministic anatomy-based subsegmentation into 81 segments. One hundred randomly selected cases were validated against expert-corrected reference segmentations using Dice similarity coefficient, volume difference, surface distance metrics, and bilateral symmetry analysis. Results Of 1,316 screened cases, 757 met eligibility criteria. Across 8,100 case-segment observations, the pipeline achieved a mean Dice of 0.9926 +/- 0.0465. Complete agreement was observed for the sacrum, ilium, acetabulum, anterior and posterior columns, sciatic buttress, and all landmarks. Relative decreases were confined to boundary-dependent regions. Bilateral symmetry analysis confirmed a median surface agreement of 99.85% within 5 mm. Conclusion The pipeline demonstrated high accuracy and reproducibility across a large normal adult dataset, establishing a structured anatomical foundation for quantitative pelvic analysis and surgical planning workflows. Clinical feasibility across abnormal anatomy and decision-level applications awaits dedicated validation.

ObjectiveTo evaluate the Cartilage Thickness Score (CTh-Score) as a quantitative measure of cartilage damage severity by assessing its association with three osteoarthritis (OA) milestones and comparing its performance with conventional morphometric measures: radiographic minimum joint space width (JSW) and regional average cartilage thickness. MethodsData were obtained from the Osteoarthritis Initiative (OAI) and the publicly available OAI CTh-Maps and CTh-Score dataset. Three matched case-control designs were used to represent major OA milestones: (i) incident radiographic OA onset, (ii) combined pain and structural progression, and (iii) knee replacement (KR) in the coming 2 years. Progression subjects were extracted from the FNIH Biomarkers Consortium cohort. Cases and controls were compared at 4 years (T-4Y), 2 years (T-2Y), and 0 years (T0) before the milestone. MRI-based CTh-Score and regional average cartilage thickness, as well as JSW, were analyzed cross-sectionally and longitudinally. Associations with case status were assessed using adjusted logistic regression models, and responsiveness was evaluated using longitudinal change and standardized response means. ResultsThe onset cohort included 307 matched case-control pairs, the progression cohort 164 cases and 369 controls, and the KR cohort 81 cases and 324 controls. Across all three study designs, the CTh-Score significantly differentiated cases from controls at all timepoints. In the onset cohort, the CTh-Score was higher in future cases than controls at T-4Y (16.2 vs 12.6, p=0.007), T-2Y (23.5 vs 16.7, p<0.001), and T0 (39.8 vs 18.6, p<0.001), whereas JSW and regional thickness measures showed limited or later discrimination. Similar findings were observed for progression (43.2 vs 33.0 at T-4Y; p<0.001) and KR (55.4 vs 46.1 at T-4Y; p=0.02) cohorts. Longitudinally, CTh-Score changes differentiated cases from controls earlier and more consistently than JSW or regional average thickness, and its responsiveness was consistently the highest across OA milestones and time intervals. In adjusted models, the CTh-Score was independently associated with all outcomes at T-4Y and T-2Y, with odds ratios per standard deviation increase ranging from 1.3 to 2.2. ConclusionThe CTh-Score captures high-resolution cartilage thickness patterns associated with OA onset, progression, and future knee replacement, outperforming conventional morphometric measures in early discrimination, responsiveness, and predictive association. These findings support CTh-Score as a sensitive quantitative marker of cartilage damage severity across the OA continuum.

BackgroundCranioplasty following decompressive craniectomy can be performed using various implant materials, with titanium and polyetheretherketone (PEEK) being the most commonly used synthetic options. However, their comparative safety and clinical performance remain debated. This systematic review and meta-analysis aimed to compare titanium-based cranioplasty with PEEK and other synthetic or autologous materials regarding implant survival, complications, functional outcomes, cosmetic results, and operative metrics. MethodsThis systematic review was conducted in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) 2020 guidelines and registered in PROSPERO (CRD). A comprehensive search was performed in PubMed, Embase, Scopus, Web of Science, and the Cochrane Database of Systematic Reviews (CDSR) without language or date restrictions. A total of 1,026 records were identified (Embase n = 263, Web of Science n = 272, Scopus n = 293, PubMed n = 193). After removal of 550 duplicates, 78 articles underwent full-text review, and 38 comparative studies met the eligibility criteria for qualitative synthesis. Three studies directly comparing titanium and PEEK with extractable infection data were included in the meta-analysis. Risk of bias was assessed using Joanna Briggs Institute (JBI) tools. ResultsForty-one studies encompassing heterogeneous patient populations and study designs were included, predominantly retrospective cohort studies. Titanium demonstrated shorter operative times and lower intraoperative blood loss compared with autologous bone and, in most studies, compared with PEEK and PMMA. Implant survival outcomes were heterogeneous: PEEK frequently showed lower exposure rates but higher rates of subgaleal fluid collection. Compared with autologous bone, titanium had higher exposure rates but avoided resorption-related failures. Infection outcomes varied across materials; however, pooled meta-analysis demonstrated a significantly lower odds of postoperative infection with titanium compared with PEEK (random-effects model), with moderate heterogeneity. Functional and neurological outcomes were largely comparable across materials, and cosmetic satisfaction was generally high regardless of implant type. ConclusionsTitanium cranioplasty provides favorable operative efficiency and competitive complication rates compared with alternative materials. While exposure risk may be higher than PEEK, pooled evidence suggests a lower infection risk with titanium. Overall, implant material selection should consider patient-specific risk factors, defect characteristics, and surgeon expertise. Further high-quality prospective studies are warranted to strengthen comparative evidence.

BackgroundLongitudinal bone growth occurs via the process of endochondral ossification, involving a complex interplay of chondrocyte proliferation, differentiation, and matrix remodelling. As with all mammalian cells, chondrocytes require dynamin for mitochondrial fission, to shuttle vesicles from the Golgi apparatus, and for both clathrin- and caveolin-mediated endocytosis. Here, we aimed to test the functions of dynamin on bone growth. To do so, we applied dynasore - a small molecule that is a reversible dynamin inhibitor - to mouse metatarsal bones cultured ex vivo. We assessed gross changes using bone length measurements and histomorphometry, and combined this with EdU detection, immunostaining, super-resolution microscopy and transmission electron microscopy. ResultsDynasore induced a dose-dependent hormetic effect on bone elongation: while high concentrations (220 {micro}M) impaired growth and abolished chondrocyte proliferation, low-dose treatment (40 {micro}M) significantly increased longitudinal bone growth. Histological analysis demonstrated that low dose dynasore augmented epiphyseal cartilage expansion and matrix accumulation, particularly within the resting and proliferative zones, while reducing chondrocyte proliferation. Immunostaining indicated that 40 {micro}M dynasore preserved collagen type X synthesis, activated mTORC1 signalling, and blocked autophagy, based on SQSTM1 accumulation. Low dose dynasore treatment expanded the thickness of the filamentous actin layer at the plasma membrane and deepened collagen fiber-containing endocytic pits, indicating that impaired cartilage remodelling was associated with growth-associated matrix accumulation. ConclusionsThis study reveals that dynasore exerts hormetic effects on growth plate chondrocytes, wherein low doses stimulate bone elongation, and high doses impair chondrocyte function.

ObjectiveX-linked Hypophosphatemia is associated with dental complications, including spontaneous endodontic infections (abscesses) in non-carious teeth and severe periodontal loss. Previous studies have mainly focused on dentin Hypomineralization; however, the structural basis underlying periodontal tissue failure remains unclear. We aimed to investigate histoanatomical abnormalities in the dentin and periodontium of Hyp mice to clarify structural consequences of Phex deficiency in adult molars. MethodsWe performed detailed histological and scanning electron microscopy analyses on the molar regions of untreated adult Hyp mice and wild-type littermates, with particular attention to the structural integrity of the root and periodontal ligament. Additionally, odontoblast process morphology and periodontal attachment abnormalities were evaluated. ResultsHyp molars exhibited marked root abnormalities, including radicular shunt-like defects and disorganized odontoblast processes, particularly in furcation and radicular dentin. Periodontal attachment showed characteristic asymmetry: detachment from the cementum surface was frequently observed, whereas attachment to the alveolar bone surface was relatively preserved. These changes were accompanied by thinning and discontinuity of Sharpeys fibers and increased vascularity in the periodontal ligament. ConclusionsThese findings provide a histoanatomical framework for understanding refractory dental complications in X-linked hypophosphatemia and support the importance of intervention during root development.

Macrophages/osteoclasts are highly fusogenic cells that interact closely with bone-metastatic breast cancer cells. These cancer cells adapt to bone microenvironments by undergoing osteomimicry, acquiring bone-like phenotypes. Exploration using human breast cancer-bone metastases dataset revealed that a small population of epithelial breast cancer cells express osteoclast-like and osteomimicry genes at the single-cell level. Cell fusion and cell-in-cell (CIC) processes are two uncommon yet prognostically significant mechanisms in cancer. We showed that co-culture between murine breast cancer cells and osteoclasts yielded a unique osteoclast phenotype through dynamic cell-in-cell (CIC) interactions and fusion-like behaviours between pre-osteoclasts/mature osteoclasts and breast tumor cells, resulting in osteoclast-tumor hybrid-like cells. These tumor cell interactions characterized by membrane retention and nuclear adjacency to host nuclei were consistently observed throughout osteoclast differentiation. Single-cell sequencing analysis and interpretative assays on hybrid-like cells revealed altered extracellular matrix (ECM) modification processes, immunoregulatory, and cancer-associated pathways compared to unfused osteoclasts. Tumor cells co-cultured with osteoclasts expressed hematopoietic and osteoclast-lineage factors more strongly than tumor cells cultured alone with their effects amplified under direct cell-cell contact. The presence of these hybrid-like cells was validated in human breast cancer-bone metastases. We propose that disseminated bone-tropic breast cancer cells were stimulated by osteoclasts to undergo a non-canonical, dynamic osteoclast differentiation and CIC formation to form hybrid-like cells that may facilitate bone metastatic lesions.

Osteocytes and adipocytes represent cells with disparate functions. Osteocytes regulate bone metabolism (remodeling) and bone homeostasis, while adipocytes regulate energy metabolism and energy storage. Here, we demonstrate that osteocyte phenotype consists of adipocytic features which are under control of peroxisome proliferator-activated receptor gamma (PPARG), a master regulator of adipocyte differentiation and function. Using a mouse model with osteocyte-specific deletion of PPARG (OT{gamma}KO) and osteocyte cellular model of MLO-Y4 cells edited with CRISPR/Cas9 for PPARG deficiency, we are demonstrating that under PPARG control osteocytes produce and secrete adiponectin (ADIPOQ), and they are equipped in adipocyte-specific mechanisms for lipid-storage and their metabolism. Under PPARG, osteocytes accumulate lipid droplets which correlate with their capability to cover up to 20% of energy requirements from fatty acids metabolism. Although osteocytes like osteoblasts mainly express perilipin 2 (Plin2), however similarly to adipocytes, lipid droplets accumulation is associated with expression of perilipin 1 (Plin1) under PPARG control. Similarly, lipids accumulation and metabolism involve adipocyte-specific activities including fatty acids binding protein 4 (Fabp4), hormone-specific lipase (Hsl) and adipocyte-specific triglyceride lipase (Atgl), which expression are under PPARG control. These studies provide a new understanding of osteocyte biology which include adipocyte-like endocrine and lipid metabolism features probably reflecting an adaptation to their unique localization and a need for a maintenance of functional fitness in these conditions. They deepen our comprehension of the crossroads of osteocyte and adipocyte function and underscore the therapeutic potential of targeting common molecular pathways in both cell types for managing metabolic disorders and skeletal diseases.

Keratin-positive giant cell-rich tumor (KPGCT) is a newly described bone and soft tissue tumor. The tumor is characterized by scattered keratin-positive cells and the presence of HMGA2::NCOR2 fusions. It is not known if the HMGA2::NCOR2 fusion is located in the keratin-positive cells, and little is known about how KPGCT develops. KPGCT shares some histologic features with tenosynovial giant cell tumor (TGCT), a soft tissue tumor with CSF1 rearrangements. Single-nuclei RNA sequencing (snRNA-seq) and Xenium spatial transcriptomics were used to elucidate the mechanisms driving KPGCT and compare KPGCT to TGCT. We show that the neoplastic cells in KPGCT constitute only a minority of cells in the tumor, and that they co-express keratin, HMGA2 and CSF1. The neoplastic cells in KPGCT express no synovial markers, confirming KPGCT as a distinct entity, separate from TGCT. The bulk of the tumor consists of CSF1R-expressing macrophages and osteoclast-like giant cells, suggesting an important role for CSF1-CSF1R signaling. In addition, we find that the cells with the HMGA2 translocation show activation of the hippo signaling pathway, which is known to regulate CSF1 expression. We show that the CSF1-CSF1R axis, possibly regulated through the hippo signaling pathway, plays an important role in KPGCT. This axis likely stimulates the migration and proliferation of macrophages, which form the majority of cells in the tumor, as well as their differentiation into osteoclasts-like giant cells. These results provide a rationale for the use of CSF1R inhibitors, which have already shown efficacy in TGCT, as a therapy for KPGCT. SignificanceKeratin-positive giant cell-rich tumor (KPGCT) is a rare, newly described soft tissue and bone tumor. By examining this tumour on a single-cell level, we confirm the identity of the neoplastic cells on a molecular level, showing these form a minority of cells in the tumor. We show that activation of the hippo pathway in the neoplastic cells is a likely driver of tumorigenesis. Additionally, we show the neoplastic cells produce large amounts of CSF1, attracting the macrophages that form the majority of cells in the tumor. This finding gives supporting evidence for anecdotal reports of response to CSF1 inhibitor therapy. Finally, we identify key differences between KPGCT and tenosynovial giant cell tumor, a tumor that shares histological features with KPGCT.