Human induced pluripotent stem cell (hiPSC)-derived osteoclasts offer a potentially scalable platform for studying osteoclast biology and anti-resorptive pharmacology. However, differentiation efficiency and inter-line reproducibility remain limited. To address this limitation, we investigated whether cytokine priming during the monocyte-to-macrophage transition is an important determinant of efficient and reproducible osteoclast differentiation from hiPSCs across diverse genetic backgrounds. Monocytes derived from three independent hiPSC lines were primed briefly with M-CSF, GM-CSF, or no cytokine, followed by osteoclast differentiation with M-CSF plus RANKL. Among these conditions, M-CSF priming reproducibly increased osteoclast differentiation efficiency compared with GM-CSF priming or no priming, yielding > 70% CD51/CD61[+] cells across hiPSC lines derived from distinct tissue sources. Osteoclast maturation and function were then assessed by multinucleation, TRAP staining, and bone resorption assays, which further showed that M-CSF priming enhanced osteoclast maturation and resorptive function across the tested hiPSC lines. We also performed extracellular flux-based measurements of OCR/ECAR and ATP production rates to examine metabolic changes associated with osteoclast maturation. This analysis showed that an early rise in OXPHOS-derived ATP production preceded peak induction of mature osteoclast genes, suggesting that early oxidative metabolism may support subsequent fusion and functional maturation. Finally, we assessed the pharmacological responsiveness of hiPSC-derived osteoclasts using two clinically used bisphosphonates, alendronate and zoledronate, both of which elicited dose-dependent anti-osteoclastic effects. Collectively, these results indicate that brief M-CSF priming enhances the efficiency and inter-line reproducibility of hiPSC-derived osteoclast differentiation, while early oxidative metabolism temporally precedes osteoclast gene induction. This improved reproducibility and functional maturation may allow hiPSC-derived osteoclasts to support mechanistic studies of osteoclastogenesis and the evaluation of pharmacological responses to osteoclast-targeted agents.

Bisphosphonates (BPs; alendronate and risedronate) have been approved as a therapy for osteoporosis. While the efficacy of these drugs appears similar, differences in safety and gastrointestinal tract (GIT) adverse effects are lacking. Consequently, a systematic review and a meta-analysis were carried out reporting on the incidence rate of GIT adverse effects and the safety of alendronate versus risedronate. In this study, the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines were used. This study collected articles from Embase, PubMed, Cochrane library, and Web of Science. Only RCTs that focused on the comparison between alendronate and risedronate in treating patients with osteoporosis were included. Risk ratios (RRs), 95% confidence intervals (CIs), and random-effects model were calculated. This meta-analysis identified 2202 studies including 4151 participants. There were six eligible RCTs. The incidence rate of GIT adverse effects in patients who received alendronate was 20.19%, while the incidence rate in patients who received risedronate was 18.30%. The rate of GIT adverse effects was not significantly different with alendronate compared to risedronate (RR: 1.11; 95% CI: 0.86-1.42; P < 0.44). In this review, the incidence rate of alendronate-associated gastrointestinal (GI) adverse effects was slightly higher than the incidence rate of risedronate-associated GI adverse effects. Although the finding is not statistically significant, switching the patient from alendronate to risedronate may nevertheless lower the incidence rate of GI side effects.

A case is discussed of a postmenopausal woman started on alendronate treatment based on a low T-score in the total hip. 3 yr later, her total hip score remains below -2.5, the target set by the ASBMR task force on goal-directed therapy.(1) The important question at this juncture is whether she should change to a different treatment that might result in better improvement in total hip BMD or if she should continue treatment with alendronate.

The efficient delivery and intracellular release of genes remain a huge challenge for gene therapy. Calcium phosphate (CaP) is a commonly used gene delivery vector, but traditional CaP nanoparticles have poor stability. They are too easy to aggregate to be taken up by cells. Herein mannitol-disulfide-alendronate (Man-SS-Aln) composed of a mannitol group, a disulfide bond, and a diphosphate group was synthesized and further crosslinked with calcium ions to form the hybrid calcium phosphate nanovector named CaP-MSSA. It was shown that CaP-MSSA had excellent stability, gene loading and protecting ability, redox-responsive release profiles and good biocompatibility. The hybrid CaP-MSSA nanovector was found to enhance the cellular uptake through caveolae-mediated endocytosis and realize the rapid and controlled release of loaded genes in cells through the redox-responsive property of disulfide bonds, so as to exert good gene transfection efficiency in vitro and in vivo even at a single administration, which is of great significance for the gene therapy in the clinic. This strategy highlights a promising approach to produce a stable, CaP-based gene delivery system with improved cellular uptake and redox-responsive gene release profiles for enhanced transgene expression.

Mechanical unloading (disuse) leads to reductions in bone and muscle mass. Bone and muscle adaptation are often studied together in terms of mechanical stimulus, but non-mechanical (biological) crosstalk during periods of disuse and how this is affected by age and bone-preserving pharmaceuticals has not been assessed. This study aimed to determine how mechanical unloading and concurrent bisphosphonate treatment affect bone and muscle structure and function in young, middle-aged, and old mice. We hypothesized that unloading would cause bone loss in untreated mice, but bisphosphonate treatment would prevent this loss. Additionally, we expected that unloading would result in muscle atrophy and reduced contraction force, but we hypothesized that these reductions would be partially mitigated by bisphosphonate treatment due to decreased release of osteokines, and that this mitigation would decrease with age. To investigate these hypotheses, young (3-mo, n = 40), middle-aged (12-mo, n = 40), and old (20-m, n = 40) male C57BL/6 J mice received biweekly subcutaneous bisphosphonate injections (0.03 mg alendronate/mouse) or vehicle injections starting one week before unloading. Mice underwent hindlimb unloading (HLU) via tail suspension for 14 days. Maximum force production of the hind limb was measured after 14 days of unloading. Femurs were imaged with micro-computed tomography (μCT 35, SCANCO Medical AG); cortical bone was analyzed at the mid-diaphysis, and trabecular bone was analyzed at the distal femur to determine bone microstructural outcomes. Muscle fiber cross-sectional area (CSA) and fiber type were analyzed via IHC. Muscle myostatin and serum TGF-β1 levels were measured via ELISA. Achilles tendon mechanical properties were also assessed using tensile testing. We found that HLU decreased the mass of the triceps surae muscles, and this loss was not recovered during bisphosphonate treatment. Muscle mass in old mice decreased during HLU with bisphosphate treatment. Maximum hindlimb force production and respective force to muscle mass ratio differed between all age groups and did not correlate with bone or muscle changes. Muscle myostatin concentrations increased with age (p = 0.040), bisphosphonate treatment (p = 0.003), and unloading (p = 0.002), as well as due to the interaction of treatment and unloading (p = 0.0239). Serum levels of TGF-β1 increased with age (p < 0.0001) and unloading (p = 0.012), as well as with interactions between both age and treatment (p = 0.006) and age and unloading (p = 0.013). Age impacted muscle, tendon, and bone responses to unloading and/or bisphosphonate treatment. Bone and muscle adaptation to unloading are different across the lifespan, as are the effects of bisphosphonate treatment. Characterizing these changes is essential for understanding clinical outcomes related to periods of disuse and clinical bone and muscle preserving treatments during bedrest, immobilization, or even spaceflight across different age groups.

Glucocorticoid-induced osteoporosis (GIOP) poses a critical long-term complication for lymphoma survivors, with cumulative incidence of fractures following R-CHOP-like therapy. Existing GIOP guidelines, typically based on chronic low-dose steroid use, are insufficient for managing this acute, high-risk toxicity of lymphoma therapy-related GIOP (LTR-GIOP). This prospective, multi-center, phase 3 randomized controlled trial (n=100; median age 74-years) compared the efficacy and safety of denosumab versus alendronate in newly diagnosed lymphoma patients receiving R-CHOP-like therapy. Patients were randomized into two groups: one received a total of two subcutaneous injections of denosumab every 6 months, and the other received oral alendronate once a week for 12 months. This study was named DENOSULY and the cases were collected by the Hokuriku Hematology Oncology Study Group. The primary endpoint was the percentage change in lumbar spine (LS) bone mineral density (BMD) at 12 months. Consequently, denosumab demonstrated superiority over alendronate in LS(L1-L4) BMD change (denosumab: +2.8%±4.4% vs. alendronate: -1.3%±5.6%; p=0.0010). Crucially, denosumab also showed superiority at the femoral neck (denosumab: +2.8%±5.8% vs. alendronate: -3.6%±10.3%; p=0.0020), a site where superiority is infrequently demonstrated in non-LTR-GIOP comparisons. Denosumab achieved stronger suppression of the bone resorption marker TRACP-5b (p=0.0003). Since denosumab showed significant superiority over alendronate at both the lumbar spine and femoral neck, denosumab may be the preferred agent in LTR-GIOP. Recognizing R-CHOP recipients as a very high-risk population, our findings underscore the need for immediate, enhanced prophylaxis with denosumab to prevent LTR-GIOP and improve long-term survivorship and quality of life. This trial was registered at www.umin.ac.jp as UMIN000038881.

The majority of clinical research continues to focus on evaluating the efficacy of teriparatide and bisphosphonates in the treatment of osteoporosis. This systematic review gathered data from extensive research assessing the efficacy, safety, and clinical outcomes of various medications. The objective was to determine the effectiveness and clinical implications of teriparatide and bisphosphonates in the treatment of postmenopausal osteoporosis. We analyzed 15 comprehensive studies, encompassing randomized controlled trials (RCTs), systematic reviews, and meta-analyses. The primary studies included large trials and several medium-sized trials that examined changes in bone mineral density (BMD), fracture risk, and adverse outcomes. Teriparatide significantly lowered vertebral fracture risk compared with risedronate in patients with severe osteoporosis and increased lumbar spine BMD to a comparable extent as alendronate, though via a distinct anabolic mechanism. Combination therapy with zoledronic acid resulted in greater BMD gains than either agent alone. Most adverse events were mild and transient, including injection-site reactions, nausea, and dizziness, with no significant difference in serious adverse event rates compared with bisphosphonates. Network meta-analyses indicate that romosozumab may achieve greater early spine BMD gains than teriparatide. Teriparatide is effective in lowering vertebral fracture risk and enhancing BMD in postmenopausal osteoporosis. Anabolic agents, including teriparatide, are recommended as first-line therapy for patients at very high risk of fractures (e.g., very low BMD with prevalent fractures or fractures occurring during glucocorticoid therapy), followed by a transition to antiresorptive therapy. Treatment selection should be guided by guideline-based risk stratification rather than applying a uniform stepwise approach.

Background/Objectives: Bone-targeted drug delivery systems hold great promise for treating skeletal diseases, yet the optimal strategy for functionalizing lipid nanoparticles (LNPs) with bone-homing ligands remains insufficiently explored. Herein, we compared two alendronate sodium (Alen) modification approaches (pre-conjugation and post-conjugation) for constructing bone-targeted LNPs capable of delivering mRNA to skeletal tissues. Methods: LNPs were fabricated via microfluidic mixing, and the 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-polyethylene glycol-alendronate conjugate (DSPE-PEG-Alen) required for the pre-conjugation method was synthesized. The bone-targeting ability of LNPs prepared by the two Alen modification strategies was evaluated using an in vitro hydroxyapatite (HAP) binding assay. Furthermore, the physicochemical properties, bone-targeting performance, mRNA delivery efficiency, and biosafety of the LNPs prepared by the post-conjugation method were assessed through cellular uptake, in vivo imaging, and other methods. Results: Hydroxyapatite binding assays revealed that the post-conjugation strategy afforded significantly superior bone affinity compared to the pre-conjugation approach. In addition, ex vivo bone fragment binding experiments further confirmed that the bone-targeting LNPs prepared by the post-conjugation method exhibited stronger bone-binding capability compared to unmodified LNPs. The optimized Alen-LNPs demonstrated efficient cellular uptake and functional mRNA translation in bone marrow mesenchymal stem cells with negligible cytotoxicity. In vivo studies in mice confirmed the preferential accumulation of Alen-LNPs in bone tissues, with successful green fluorescent protein (GFP) mRNA translation detected in bone tissue sections. Histopathological analysis confirmed the biosafety of the formulation. Conclusions: This study establishes the post-conjugation strategy as the superior approach for Alen functionalization of LNPs, providing a robust and reproducible platform for bone-targeted mRNA therapeutics.

Critical-sized bone defects represent a prevalent and intractable challenge in orthopedic due to poor self-healing ability and prolonged treatment cycles. Biomaterials are recognized as promising strategies for effective bone defect repair, for the capability of regulating microenvironment and facilitating osteogenic differentiation. Herein, an inorganic-organic multifunctional composite hydrogel was developed with methacrylated gelatin (GelMA) as the matrix, functionalized with alendronate modified ceria quantum dots (AHA@CQDs) and nano-hydroxyapatite (nHAP). AHA@CQDs regulated local oxidative stress to promote cell proliferation and differentiation, while nHAP enhanced osteogenic differentiation and mechanical stability of the hydrogel. In vitro experiments confirmed that the composite hydrogel exhibited excellent biocompatibility, antioxidant property and cell migration ability, while also exhibiting remarkable potential in promoting the proliferation and osteogenic differentiation of rat bone marrow mesenchymal stem cells (BMSCs). Furthermore, after being implanted into a rat cranial defect model, the composite hydrogel significantly promoted new bone formation and upregulated the expression of osteogenic related biomarkers. Collectively, this multifunctional composite hydrogel provides an effective approach for repairing critical-sized bone defects.

Osteogenesis imperfecta (OI) is a group of genetic connective tissue disorders characterized by poor bone matrix and increased bone fragility. Bisphosphonates, a group of antiresorptive drugs FDA-approved for the treatment of osteoporosis, are commonly used to treat OI, but their effects on fracture risk and long-term efficacy in growing children remain unclear. Here, we use the oim mouse model of severe OI to investigate the long-term impact of alendronate, a nitrogen-containing bisphosphonate, on bone fragility and examine sex-specific responses through (i) traditional fracture mechanics testing paired with full-field digital image correlation strain maps around the propagating crack, (ii) porosity measurements using synchrotron microtomography, (iii) collagen organization assessment via second harmonic generation microscopy, and (iv) tissue composition and crosslinking content by Raman spectroscopy and molecular fluorescence. Male and female oim/oim mice exhibit distinct compositional and structural abnormalities that influence disease severity and response to therapy, ultimately affecting fracture resistance. Alendronate treatment does not improve fracture resistance of oim/oim bones, although it increases cortical thickness and cortical porosity, and is associated with the appearance of trabecular-like structures spanning the medullary cavity at mid-shaft, resembling the radiographic zebra lines observed in pediatric OI population. Changes are more pronounced in female oim/oim mice, where load-bearing capacity increases together with canal porosity. These results highlight the continuous need for treatments that rescue OI bone fragility by improving the poor bone matrix and emphasize the importance of considering sexual dimorphism when designing therapies for OI. STATEMENT OF SIGNIFICANCE: Bisphosphonates are widely used anti-resorptive drugs approved for osteoporosis and administered also to children with bone fragility disorders, like osteogenesis imperfecta (OI). Their benefits in young patients, however, have been debated: while some studies report fewer fractures and improved pain, others find limited effects. The impact of long-term bisphosphonate treatment on fracture resistance in growing OI bones remains unclear. Using an interdisciplinary multiscale approach combining advanced imaging and fracture mechanics on OI mouse (oim/oim) bones, this study solves the controversy. We demonstrate that bisphosphonates do not improve bone fragility. However, they enhance bone structural stability, allowing higher loads to be sustained by increasing cortical thickness and generating mid-shaft trabecular bands, resembling the "zebra lines" observed in the bones of children with OI. These changes were particularly evident in female mice.