Bone Research最新文献

Proteoglycans, key components of non-collagenous proteins in the bone matrix, attract water through their negatively charged glycosaminoglycan chains. Among these proteoglycans, biglycan (Bgn) and decorin (Dcn) are major subtypes, yet their distinct roles in bone remain largely elusive. In this study, we utilized single knockout (KO) mouse models and successfully generated double KO (dKO) models despite challenges with low yield. Bgn deficiency, but not Dcn deficiency, decreased trabecular bone mass, with more pronounced bone loss in dKO mice. Low-field nuclear magnetic resonance measurements showed a marked decrease in bound water among all KO groups, especially in Bgn KO and dKO mice. Moreover, both Bgn KO and dKO mice exhibited reduced fracture toughness compared to Dcn KO mice. Dcn was significantly upregulated in Bgn KO mice, while a modest upregulation of Bgn was observed in Dcn KO mice, indicating Bgn’s predominant role in bone. High resolution atomic force microscopy showed decreased in situ permanent energy dissipation and increased elastic modulus in the extrafibrillar matrix of Bgn/Dcn deficient mice, which were diminished upon dehydration. Furthermore, we found that both Bgn and Dcn are indispensable for the activation of ERK and p38 MAPK signaling pathways. Collectively, our results highlight the distinct and indispensable roles of Bgn and Dcn in maintaining bone structure, water retention, and bulk/in situ tissue properties in the bone matrix, with Bgn exerting a predominant influence.

Degenerative spine and joint diseases, including intervertebral disc degeneration (IDD), ossification of the spinal ligaments (OSL), and osteoarthritis (OA), are common musculoskeletal diseases that cause pain or disability to the patients. However, the pathogenesis of these musculoskeletal disorders is complex and has not been elucidated clearly to date. As a matter of fact, the spine and joints are not independent of other organs and tissues. Recently, accumulating evidence demonstrates the association between obesity and degenerative musculoskeletal diseases. Obesity is a common metabolic disease characterized by excessive adipose tissue or abnormal adipose distribution in the body. Excessive mechanical stress is regarded as a critical risk factor for obesity-related pathology. Additionally, obesity-related factors, mainly including lipid metabolism disorder, dysregulated pro-inflammatory adipokines and cytokines, are reported as plausible links between obesity and various human diseases. Importantly, these obesity-related factors are deeply involved in the regulation of cell phenotypes and cell fates, extracellular matrix (ECM) metabolism, and inflammation in the pathophysiological processes of degenerative spine and joint diseases. In this study, we systematically discuss the potential cellular and molecular mechanisms underlying obesity in these degenerative musculoskeletal diseases, and hope to provide novel insights for developing targeted therapeutic strategies.

Recent genome-wide association studies (GWAS) identified 518 significant loci associated with bone mineral density (BMD), including variants at the RUNX1 locus (rs13046645, rs2834676, and rs2834694). However, their regulatory impact on RUNX1 expression and bone formation remained unclear. This study utilized human induced pluripotent stem cells (iPSCs) differentiated into osteoblasts to investigate these variants’ regulatory roles. CRISPR/Cas9 was employed to generate mutant (Δ) iPSC lines lacking these loci at the RUNX1 locus. Deletion lines (Δ1 and Δ2) were created in iPSCs to assess the effects of removing regions containing these loci. Deletion lines exhibited enhanced osteogenic potential, with increased expression of osteogenic marker genes and Alizarin Red staining. Circularized chromosome conformation capture (4C-Seq) was utilized to analyze interactions between BMD-associated loci and the RUNX1 promoter during osteogenesis. Analysis revealed altered chromatin interactions with multiple gene promoters including RUNX1 isoform, as well as SETD4, a histone methyltransferase, indicating their regulatory influence. Interestingly, both deletion lines notably stimulated the expression of the long isoform of RUNX1, with more modest effects on the shorter isoform. Consistent upregulation of SETD4 and other predicted targets within the Δ2 deletion suggested its removal removed a regulatory hub constraining expression of multiple genes at this locus. In vivo experiments using a bone defect model in mice demonstrated increased bone regeneration with homozygous deletion of the Δ2 region. These findings indicate that BMD-associated variants within the RUNX1 locus regulate multiple effector genes involved in osteoblast commitment, providing valuable insights into genetic regulation of bone density and potential therapeutic targets.

Osteoarthritis (OA) poses a significant challenge in orthopedics. Inflammatory pathways are regarded as central mechanisms in the onset and progression of OA. Growing evidence suggests that senescence acts as a mediator in inflammation-induced OA. Given the lack of effective treatments for OA, there is an urgent need for a clearer understanding of its pathogenesis. In this review, we systematically summarize the cross-talk between cellular senescence and inflammation in OA. We begin by focusing on the mechanisms and hallmarks of cellular senescence, summarizing evidence that supports the relationship between cellular senescence and inflammation. We then discuss the mechanisms of interaction between cellular senescence and inflammation, including senescence-associated secretory phenotypes (SASP) and the effects of pro- and anti-inflammatory interventions on cellular senescence. Additionally, we focus on various types of cellular senescence in OA, including senescence in cartilage, subchondral bone, synovium, infrapatellar fat pad, stem cells, and immune cells, elucidating their mechanisms and impacts on OA. Finally, we highlight the potential of therapies targeting senescent cells in OA as a strategy for promoting cartilage regeneration.

Osteoporosis (OP) is a common and fracture-prone skeletal disease characterized by deteriorated trabecular microstructure and pathologically involving various forms of regulated bone cell death. However, the exact role, cellular nature and regulatory mechanisms of ferroptosis in OP are not fully understood. Here, we reported that OP femurs from ovariectomized (Ovx) mice exhibited pronounced iron deposition, ferroptosis, and transcriptional suppression of a key anti-ferroptotic factor GPX4 (glutathione peroxidase 4). GPX4 suppression was accompanied by hypermethylation of the Gpx4 promoter and an increase in DNA methyltransferases DNMT1/3a/3b and was transcriptionally promoted by repressive KLF5 and the transcriptional corepressors NCoR and SnoN. Conversely, DNMT inhibition with SGI-1027 reversed promoter hypermethylation, GPX4 suppression and ferroptotic osteoporosis. In cultured primary bone cells, ferric ammonium citrate (FAC) mimicking iron loading similarly induced GPX4 suppression and ferroptosis in osteoblasts but not in osteoclasts, which were rescued by siRNA-mediated individual knockdown of DNMT 1/3a/3b. Intriguingly, SGI-1027 alleviated the ferroptotic changes caused by FAC, but not by a GPX4 inactivator RSL3. More importantly, we generated a strain of osteoblast-specific Gpx4 haplo-deficient mice Gpx4^Ob+/− that developed spontaneous and more severe ferroptotic OP alterations after Ovx operation, and showed that GPX4 inactivation by RSL3 or semi-knockout in osteoblasts largely abolished the anti-ferroptotic and osteoprotective effects of SGI-1027. Taken together, our data suggest that GPX4 epigenetic suppression caused by DNMT aberration and the resulting osteoblastic ferroptosis contribute significantly to OP pathogenesis, and that the strategies preserving GPX4 by DNMT intervention are potentially effective to treat OP and related bone disorders.

Intervertebral disc degeneration (IVDD), a disease associated with ageing, is characterised by a notable increase in senescent nucleus pulposus cells (NPCs) as IVDD progresses. However, the specific mechanisms that regulate the senescence of NPCs remain unknown. In this study, we observed impaired autophagy in IVDD-NPCs, which contributed to the upregulation of NPCs senescence and the senescence-associated secretory phenotype (SASP). The dysregulated SASP disrupted NPCs viability and initiated extracellular matrix degradation. Conversely, the restoration of autophagy reversed the senescence phenotype by inhibiting GATA binding protein 4 (GATA4). Moreover, we made the novel observation that a cross-talk between histone H3 lysine 4 trimethylation (H3K4me3) modification and N6-methyladenosine(m⁶A)-methylated modification regulates autophagy in IVDD-NPCs. Mechanistically, lysine methyltransferase 2A (KMT2A) promoted the expression of methyltransferase-like 3 (METTL3) through H3K4me3 modification, whereas METTL3-mediated m⁶A modification reduced the expression of autophagy-associated 4a (ATG4a) by attenuating its RNA stability, leading to autophagy damage in NPCs. Silencing KMT2A and METTL3 enhanced autophagic flux and suppressed SASP expression in IVDD-NPCs. Therefore, targeting the H3K4me3-regulated METTL3/ATG4a/GATA4 axis may represent a promising new therapeutic strategy for IVDD.

The concept and development of bone/cartilage organoids are rapidly gaining momentum, providing opportunities for both fundamental and translational research in bone biology. Bone/cartilage organoids, essentially miniature bone/cartilage tissues grown in vitro, enable the study of complex cellular interactions, biological processes, and disease pathology in a representative and controlled environment. This review provides a comprehensive and up-to-date overview of the field, focusing on the strategies for bone/cartilage organoid construction strategies, progresses in the research, and potential applications. We delve into the significance of selecting appropriate cells, matrix gels, cytokines/inducers, and construction techniques. Moreover, we explore the role of bone/cartilage organoids in advancing our understanding of bone/cartilage reconstruction, disease modeling, drug screening, disease prevention, and treatment strategies. While acknowledging the potential of these organoids, we discuss the inherent challenges and limitations in the field and propose potential solutions, including the use of bioprinting for organoid induction, AI for improved screening processes, and the exploration of assembloids for more complex, multicellular bone/cartilage organoids models. We believe that with continuous refinement and standardization, bone/cartilage organoids can profoundly impact patient-specific therapeutic interventions and lead the way in regenerative medicine.

Emerging evidence suggests a significant role of gut microbiome in bone health. Aging is well recognized as a crucial factor influencing the gut microbiome. In this study, we investigated whether age-dependent microbial change contributes to age-related bone loss in CB6F1 mice. The bone phenotype of 24-month-old germ-free (GF) mice was indistinguishable compared to their littermates colonized by fecal transplant at 1-month-old. Moreover, bone loss from 3 to 24-month-old was comparable between GF and specific pathogen-free (SPF) mice. Thus, GF mice were not protected from age-related bone loss. 16S rRNA gene sequencing of fecal samples from 3-month and 24-month-old SPF males indicated an age-dependent microbial shift with an alteration in energy and nutrient metabolism potential. An integrative analysis of 16S predicted metagenome function and LC-MS fecal metabolome revealed an enrichment of protein and amino acid biosynthesis pathways in aged mice. Microbial S-adenosyl methionine metabolism was increased in the aged mice, which has previously been associated with the host aging process. Collectively, aging caused microbial taxonomic and functional alteration in mice. To demonstrate the functional importance of young and old microbiome to bone, we colonized GF mice with fecal microbiome from 3-month or 24-month-old SPF donor mice for 1 and 8 months. The effect of microbial colonization on bone phenotypes was independent of the microbiome donors’ age. In conclusion, our study indicates age-related bone loss occurs independent of gut microbiome.

Osteonecrosis of the femoral head (ONFH) is a common complication of glucocorticoid (GC) therapy. Recent advances demonstrate that sympathetic nerves regulate bone homeostasis, and GCs lower the sympathetic tone. Here, we show that the dramatically decreased sympathetic tone is closely associated with the pathogenesis of GC-induced ONFH. GCs activate the glucocorticoid receptor (GR) but hinder the activation of the mineralocorticoid receptor (MR) on neurons in the hypothalamic paraventricular nucleus (PVN). This disrupts the balance of corticosteroid receptors (GR/MR) and subsequently reduces the sympathetic outflow in the PVN. Vascular endothelial cells rapidly react to inhibition of sympathetic tone by provoking endothelial apoptosis in adult male mice treated with methylprednisolone (MPS) daily for 3 days, and we find substantially reduced H-type vessels in the femoral heads of MPS-treated ONFH mice. Importantly, treatment with a GR inhibitor (RU486) in the PVN promotes the activation of MR and rebalances the ratio of GR and MR, thus effectively boosting sympathetic outflow, as shown by an increase in tyrosine hydroxylase expression in both the PVN and the sympathetic postganglionic neurons and an increase in norepinephrine levels in both the serum and bone marrow of the femoral head of MPS-treated mice. Rebalancing the corticosteroid receptors mitigates GC-induced endothelial impairment and ONFH and promotes angiogenesis coupled with osteogenesis in the femoral head, while these effects are abolished by chemical sympathectomy with 6-OHDA or adrenergic receptor-β2 (Adrb2) knockout. Furthermore, activating Adrb2 signaling in vivo is sufficient to rescue the GC-induced ONFH phenotype. Mechanistically, norepinephrine increases the expression of the key glycolytic gene 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3) via Adrb2-cyclic AMP response element-binding protein (CREB) signaling. Endothelial-specific overexpression of PFKFB3 attenuates endothelial impairment and prevents severe osteonecrosis in MPS-treated Adrb2 knockout mice. Thus, GC inhibits sympathetic tone via the hypothalamic descending pathway, which, in turn, acts as a mediator of GC-induced ONFH.

Osteoarthritis (OA) is an age-related cartilage-degenerating joint disease. Mitochondrial dysfunction has been reported to promote the development of OA. Poly (ADP-ribose) polymerase family member 12 (PARP12) is a key regulator of mitochondrial function, protein translation, and inflammation. However, the role of PARP12 in OA-based cartilage degradation and the underlying mechanisms are relatively unknown. Here, we first demonstrated that PARP12 inhibits mitophagy and promotes OA progression in human OA cartilage and a monosodium iodoacetate-induced rat OA model. Using mass spectrometry and co-immunoprecipitation assay, PARP12 was shown to interact with ISG15, upregulate mitofusin 1 and 2 (MFN1/2) ISGylation, which downregulated MFN1/2 ubiquitination and SUMOylation, thereby inhibiting PINK1/Parkin-dependent chondrocyte mitophagy and promoting cartilage degradation. Moreover, inflammatory cytokine-induced interferon regulatory factor 1 (IRF1) activation was required for the upregulation of PARP12 expression, and it directly bound to the PARP12 promoter to activate transcription. XAV-939 inhibited PARP12 expression and suppressed OA pathogenesis in vitro and in vivo. Clinically, PARP12 can be used to predict the severity of OA; thus, it represents a new target for the study of mitophagy and OA progression. In brief, the IRF1-mediated upregulation of PARP12 promoted cartilage degradation by inhibiting PINK1/Parkin-dependent mitophagy via ISG15-based attenuation of MFN1/2 ubiquitylation and SUMOylation. Our data provide new insights into the molecular mechanisms underlying PARP12-based regulation of mitophagy and can facilitate the development of therapeutic strategies for the treatment of OA.