Bone Research最新文献

Gain-of-function mutations in fibroblast growth factor receptor (FGFR) genes lead to chondrodysplasia and craniosynostoses. FGFR signaling has a key role in the formation and repair of the craniofacial skeleton. Here, we analyzed the impact of Fgfr2- and Fgfr3-activating mutations on mandibular bone formation and endochondral bone repair after non-stabilized mandibular fractures in mouse models of Crouzon syndrome (Crz) and hypochondroplasia (Hch). Bone mineralization of the calluses was abnormally high in Crz mice and abnormally low in Hch mice. The latter model presented pseudarthrosis and impaired chondrocyte differentiation. Spatial transcriptomic analyses of the Hch callus revealed abnormally low expression of Col11, Col1a, Dmp1 genes in mature chondrocytes. We found that the expression of genes involved in autophagy and apoptosis (Smad1, Comp, Birc2) was significantly perturbed and that the Dusp3, Dusp9, and Socs3 genes controlling the mitogen-activated protein kinase pathway were overexpressed. Lastly, we found that treatment with a tyrosine kinase inhibitor (BGJ398, infigratinib) or a C-type natriuretic peptide (BMN111, vosoritide) fully rescued the defective endochondral bone repair observed in Hch mice. Taken as a whole, our findings show that FGFR3 is a critical orchestrator of bone repair and provide a rationale for the development of potential treatments for patients with FGFR3-osteochondrodysplasia.

Reproductive hormones associated with the hypothalamic-pituitary-gonadal (HPG) axis are closely linked to bone homeostasis. In this study, we demonstrate that Gonadotropin inhibitory hormone (GnIH, one of the key reproductive hormones upstream of the HPG axis) plays an indispensable role in regulating bone homeostasis and maintaining bone mass. We find that deficiency of GnIH or its receptor Gpr147 leads to a significant reduction in bone mineral density (BMD) in mice primarily by enhancement of osteoclast activation in vivo and in vitro. Mechanistically, GnIH/Gpr147 inhibits osteoclastogenesis by the PI3K/AKT, MAPK, NF-κB and Nfatc1 signaling pathways. Furthermore, GnIH treatment was able to alleviate bone loss in aging, ovariectomy (OVX) or LPS-induced mice. Moreover, the therapy using green light promotes the release of GnIH and rescues OVX-induced bone loss. In humans, serum GnIH increases and bone resorption markers decrease after green light exposure. Therefore, our study elucidates that GnIH plays an important role in maintaining bone homeostasis via modulating osteoclast differentiation and demonstrates the potential of GnIH therapy or green light therapy in preventing osteoporosis.

Mechanical stress modulates bone formation and organization of the extracellular matrix (ECM), the interaction of which affects heterotopic ossification (HO). However, the mechanically sensitive cell populations in HO and the underlying mechanism remain elusive. Here, we show that the mechanical protein Polysyctin-1 (PC1, Pkd1) regulates CTSK lineage tendon-derived mesenchymal stem cell (TDMSC) fate and ECM organization, thus affecting HO progression. First, we revealed that CTSK lineage TDMSCs are the major source of osteoblasts and fibroblasts in HO and are responsive to mechanical cues via single-cell RNA sequencing analysis and experiments with a lineage tracing mouse model. Moreover, we showed that PC1 mediates the mechanosignal transduction of CTSK lineage TDMSCs to regulate osteogenic and fibrogenic differentiation and alters the ECM architecture by facilitating TAZ nuclear translocation. Conditional gene depletion of Pkd1 or Taz in CTSK lineage cells and pharmaceutical intervention in the PC1-TAZ axis disrupt osteogenesis, fibrogenesis and ECM organization, and consequently attenuate HO progression. These findings suggest that mechanically sensitive CTSK-lineage TDMSCs contribute to heterotopic ossification through PC1-TAZ signaling axis mediated cell fate determination and ECM organization.

Fibrotic remodeling of nucleus pulposus (NP) leads to structural and mechanical anomalies of intervertebral discs that prone to degeneration, leading to low back pain incidence and disability. Emergence of fibroblastic cells in disc degeneration has been reported, yet their nature and origin remain elusive. In this study, we performed an integrative analysis of multiple single-cell RNA sequencing datasets to interrogate the cellular heterogeneity and fibroblast-like entities in degenerative human NP specimens. We found that disc degeneration severity is associated with an enrichment of fibrocyte phenotype, characterized by CD45 and collagen I dual positivity, and expression of myofibroblast marker α-smooth muscle actin. Refined clustering and classification distinguished the fibrocyte-like populations as subtypes in the NP cells - and immunocytes-clusters, expressing disc degeneration markers HTRA1 and ANGPTL4 and genes related to response to TGF-β. In injury-induced mouse disc degeneration model, fibrocytes were found recruited into the NP undergoing fibrosis and adopted a myofibroblast phenotype. Depleting the fibrocytes in CD11b-DTR mice in which myeloid-derived lineages were ablated by diphtheria toxin could markedly attenuate fibrous modeling and myofibroblast formation in the NP of the degenerative discs, and prevent disc height loss and histomorphological abnormalities. Marker analysis supports that disc degeneration progression is dependent on a function of CD45⁺COL1A1⁺ and αSMA⁺ cells. Our findings reveal that myeloid-derived fibrocytes play a pivotal role in NP fibrosis and may therefore be a target for modifying disc degeneration and promoting its repair.

Osteocytes are the main cells in mineralized bone tissue. Elevated osteocyte apoptosis has been observed in lytic bone lesions of patients with multiple myeloma. However, their precise contribution to bone metastasis remains unclear. Here, we investigated the pathogenic mechanisms driving melanoma-induced osteocyte death. Both in vivo models and in vitro assays were combined with untargeted RNA sequencing approaches to explore the pathways governing melanoma-induced osteocyte death. We could show that ferroptosis is the primary mechanism behind osteocyte death in the context of melanoma bone metastasis. HMOX1 was identified as a crucial regulatory factor in this process, directly involved in inducing ferroptosis and affecting osteocyte viability. We uncover a non-canonical pathway that involves excessive autophagy-mediated ferritin degradation, highlighting the complex relationship between autophagy and ferroptosis in melanoma-induced osteocyte death. In addition, HIF1α pathway was shown as an upstream regulator, providing a potential target for modulating HMOX1 expression and influencing autophagy-dependent ferroptosis. In conclusion, our study provides insight into the pathogenic mechanisms of osteocyte death induced by melanoma bone metastasis, with a specific focus on ferroptosis and its regulation. This would enhance our comprehension of melanoma-induced osteocyte death.

Tissue clearing combined with high-resolution confocal imaging is a cutting-edge approach for dissecting the three-dimensional (3D) architecture of tissues and deciphering cellular spatial interactions under physiological and pathological conditions. Deciphering the spatial interaction of leptin receptor-expressing (LepR⁺) stromal cells with other compartments in the bone marrow is crucial for a deeper understanding of the stem cell niche and the skeletal tissue. In this study, we introduce an optimized protocol for the 3D analysis of skeletal tissues, enabling the visualization of hematopoietic and stromal cells, especially LepR⁺ stromal cells, within optically cleared bone hemisections. Our method preserves the 3D tissue architecture and is extendable to other hematopoietic sites such as calvaria and vertebrae. The protocol entails tissue fixation, decalcification, and cryosectioning to reveal the marrow cavity. Completed within approximately 12 days, this process yields highly transparent tissues that maintain genetically encoded or antibody-stained fluorescent signals. The bone hemisections are compatible with diverse antibody labeling strategies. Confocal microscopy of these transparent samples allows for qualitative and quantitative image analysis using Aivia or Bitplane Imaris software, assessing a spectrum of parameters. With proper storage, the fluorescent signal in the stained and cleared bone hemisections remains intact for at least 2–3 months. This protocol is robust, straightforward to implement, and highly reproducible, offering a valuable tool for tissue architecture and cellular interaction studies.

The cranial mesenchyme, originating from both neural crest and mesoderm, imparts remarkable regional specificity and complexity to postnatal calvarial tissue. While the distinct embryonic origins of the superior and dura periosteum of the cranial parietal bone have been described, the extent of their respective contributions to bone and vessel formation during adult bone defect repair remains superficially explored. Utilizing transgenic mouse models in conjunction with high-resolution multiphoton laser scanning microscopy (MPLSM), we have separately evaluated bone and vessel formation in the superior and dura periosteum before and after injury, as well as following intermittent treatment of recombinant peptide of human parathyroid hormone (rhPTH), Teriparatide. Our results show that new bone formation along the dura surface is three times greater than that along the superior periosteal surface following injury, regardless of Teriparatide treatment. Targeted deletion of PTH receptor PTH1R via SMA-CreER and Col 1a (2.3)-CreER results in selective reduction of bone formation, suggesting different progenitor cell pools in the adult superior and dura periosteum. Consistently, analyses of microvasculature show higher vessel density and better organized arterial-venous vessel network associated with a 10-fold more osteoblast clusters at dura periosteum as compared to superior periosteum. Intermittent rhPTH treatment further enhances the arterial vessel ratio at dura periosteum and type H vessel formation in cortical bone marrow space. Taken together, our study demonstrates a site-dependent coordinated osteogenic and angiogenic response, which is determined by regional osteogenic progenitor pool as well as the coupling blood vessel network at the site of cranial defect repair.

Intervertebral disc degeneration (IDD), osteoarthritis (OA), and osteoporosis (OP) are common musculoskeletal disorders (MSDs) with similar age-related risk factors, representing the leading causes of disability. However, successful therapeutic development and translation have been hampered by the lack of clinically-relevant animal models. In this study, we investigated the potential suitability of the tree shrew, a small mammal with a close genetic relationship to primates, as a new animal model for MSDs. Age-related spontaneous IDD in parallel with a gradual disappearance of notochordal cells were commonly observed in tree shrews upon skeletal maturity with no sex differences, while age-related osteoporotic changes including bone loss in the metaphyses were primarily presented in aged females, similar to observations in humans. Moreover, in the osteochondral defect model, tree shrew cartilage exhibited behavior similar to that of humans, characterized by a more restricted self-healing capacity compared to the rapid spontaneous healing of joint surfaces observed in rats. The induced OA model in tree shrews was highly efficient and reproducible, characterized by gradual deterioration of articular cartilage, recapitulating the human OA phenotype to some degree. Surgery-induced IDD models were successfully established in tree shrews, in which the lumbar spine instability model developed slow progressive disc degeneration with more similarity to the clinical state, whereas the needle puncture model led to the rapid development of IDD with more severe symptoms. Taken together, our findings pave the way for the development of the tree shrew as a new animal model for the study of MSDs and aging.

Accumulating research has shed light on the significance of skeletal interoception, in maintaining physiological and metabolic homeostasis related to bone health. This review provides a comprehensive analysis of how skeletal interoception influences bone homeostasis, delving into the complex interplay between the nervous system and skeletal system. One key focus of the review is the role of various factors such as prostaglandin E2 (PGE2) in skeletal health via skeletal interoception. It explores how nerves innervating the bone tissue communicate with the central nervous system to regulate bone remodeling, a process critical for maintaining bone strength and integrity. Additionally, the review highlights the advancements in biomaterials designed to utilize skeletal interoception for enhancing bone regeneration and treatment of bone disorders. These biomaterials, tailored to interact with the body’s interoceptive pathways, are positioned at the forefront of innovative treatments for conditions like osteoporosis and fractures. They represent a convergence of bioengineering, neuroscience, and orthopedics, aiming to create more efficient and targeted therapies for bone-related disorders. In conclusion, the review underscores the importance of skeletal interoception in physiological regulation and its potential in developing more effective therapies for bone regeneration. It emphasizes the need for further research to fully understand the mechanisms of skeletal interoception and to harness its therapeutic potential fully.

Bone morphogenetic proteins are essential for bone regeneration/fracture healing but can also induce heterotopic ossification (HO). Understanding accessory factors modulating BMP signaling would provide both a means of enhancing BMP-dependent regeneration while preventing HO. This study focuses on the ability of the collagen receptor, discoidin domain receptor 2 (DDR2), to regulate BMP activity. As will be shown, induction of bone formation by subcutaneous BMP2 implants is severely compromised in Ddr2-deficient mice. In addition, Ddr2 deficiency attenuates HO in mice expressing the ACVR1 mutation associated with human fibrodysplasia ossificans progressiva. In cells migrating into BMP2 implants, DDR2 is co-expressed with GLI1, a skeletal stem cell marker, and DDR2/GLI1-positive cells participate in BMP2-induced bone formation where they contribute to chondrogenic and osteogenic lineages. Consistent with this distribution, conditional knockout of Ddr2 in Gli1-expressing cells inhibited bone formation to the same extent seen in globally Ddr2-deficient animals. This response was explained by selective inhibition of Gli1⁺ cell proliferation without changes in apoptosis. The basis for this DDR2 requirement was explored further using bone marrow stromal cells. Although Ddr2 deficiency inhibited BMP2-dependent chondrocyte and osteoblast differentiation and in vivo, bone formation, early BMP responses including SMAD phosphorylation remained largely intact. Instead, Ddr2 deficiency reduced the nuclear/cytoplasmic ratio of the Hippo pathway intermediates, YAP and TAZ. This suggests that DDR2 regulates Hippo pathway-mediated responses to the collagen matrix, which subsequently affect BMP responsiveness. In summary, DDR2 is an important modulator of BMP signaling and a potential therapeutic target both for enhancing regeneration and treating HO.