Bone Development Research Articles

Importance of adequate and timely diagnosis in non-neoplasic bone lesions: case report

Non-ossifying fibroma is defined as a bone lesion of non-neoplasic origin, characterized by the presence of fibrous tissue in eddies, it differs from the fibrous cortical defect due to its size, being the result of the periosteal cortical bone development defect that leads to failure of ossification. We present a case of a 12-year-old female adolescent who comes to this center presenting knee pain, claudication while walking and reduced range of motion of the left knee and that in the CT scan shows a hyper-enhanced lesion, hypointense on T1, hyperintense on T2, in addition to cortical lesion at the level of both distal femurs. Planning is carried out for taking a percutaneous biopsy whose report reports a non-ossifying fibroma of the bilateral distal femur, and it is decided to perform surgery on the right distal femur, with good results, in the case the clinical and radiographic characteristics of the patient and the resolution of this are described.

The “toxic window” of amoxicillin exposure during pregnancy on long bone development in fetal mice

AimsAmoxicillin is a broad-spectrum beta-lactam antibiotic used to treat infectious diseases in pregnant women. Studies have shown that prenatal amoxicillin exposure (PAmE) has developmental toxicity on fetal development. However, the effect of PAmE on long bone development has not been reported. This study aimed to investigate the “toxic window” of PAmE on long bone development and explore its possible mechanism in fetal mice. Materials and methodsPregnant mice were administered amoxicillin by gavage at different stages (gestational day (GD)10–12 and GD16–18), different doses (150 and 300 mg/kg·d) and different courses (single and multiple courses). Fetal femurs were collected at GD18 and bone development related indicators were detected. Key findingsThe results showed that PAmE significantly reduced the length of the femur and primary ossification center of fetal mice, and inhibited the development of fetal growth plate. Meanwhile, PAmE inhibited the development of bone marrow mesenchymal stem cells, osteoclasts and endothelial cells in fetal long bone. Further, we found the fetal long bone developmental toxicity induced by PAmE was most significant at late-pregnancy (GD16–18), high dose (300 mg/kg·d) and multiple-course group. Besides, PAmE inhibited the expression of Wnt/β-catenin signaling pathway in fetal long bone. The β-catenin mRNA expression was significantly positively correlated with the development indexes of fetal long bone. SignificancePAmE has toxic effects on long bone development, and there was an obvious “toxic window” of PAmE on the long bone development in fetal mice. The Wnt/β-catenin signaling pathway may mediate PAmE-induced fetal long bone development inhibition.

Time of exercise differentially impacts bone growth in mice.

Although physical training has been shown to improve bone mass, the time of day to exercise for optimal bone growth remains uncertain. Here we show that engaging in physical activity during the early active phase, as opposed to the subsequent active or rest phase, results in a more substantial increase in bone length of male and female mice. Transcriptomic and metabolomic methodologies identify that exercise during the early active phase significantly upregulates genes associated with bone development and metabolism. Notably, oxidative phosphorylation-related genes show a rhythmic expression in the chondrification centre, with a peak at the early active phase, when more rhythmic genes in bone metabolism are expressed and bone growth is synergistically promoted by affecting oxidative phosphorylation, which is confirmed by subsequent pharmacological investigations. Finally, we construct a signalling network to predict the impact of exercise on bone growth. Collectively, our research sheds light on the intricacies of human exercise physiology, offering valuable implications for interventions.

Biomimetic Hydrogels as the Inductive Endochondral Ossification Template for Promoting Bone Regeneration.

Repairing critical size bone defects (CSBD) is a major clinical challenge and requires effective intervention by biomaterial scaffolds. Inspired by the fact that the cartilaginous template-based endochondral ossification (ECO) process is crucial to bone healing and development, developing biomimetic biomaterials to promote ECO is recognized as a promising approach for repairing CSBD. With the unique highly hydrated 3D polymeric network, hydrogels can be designed to closely emulate the physiochemical properties of cartilage matrix to facilitate ECO. In this review, the various preparation methods of hydrogels possessing the specific physiochemical properties required for promoting ECO are introduced. The materiobiological impacts of the physicochemical properties of hydrogels, such as mechanical properties, topographical structures and chemical compositions on ECO, and the associated molecular mechanisms related to the BMP, Wnt, TGF-β, HIF-1α, FGF, and RhoA signaling pathways are further summarized. This review provides a detailed coverage on the materiobiological insights required for the design and preparation of hydrogel-based biomaterials to facilitate bone regeneration.

METTL7A-mediated m6A modification of corin reverses bisphosphonates-impaired osteogenic differentiation of orofacial BMSCs

Bisphosphonate-related osteonecrosis of jaw (BRONJ) is characterized by impaired osteogenic differentiation of orofacial bone marrow stromal cells (BMSCs). Corin has recently been demonstrated to act as a key regulator in bone development and orthopedic disorders. However, the role of corin in BRONJ-related BMSCs dysfunction remains unclarified. A m6A epitranscriptomic microarray study from our group shows that the CORIN gene is significantly upregulated and m6A hypermethylated during orofacial BMSCs osteogenic differentiation. Corin knockdown inhibits BMSCs osteogenic differentiation, whereas corin overexpression or soluble corin (sCorin) exerts a promotion effect. Furthermore, corin expression is negatively regulated by bisphosphonates (BPs). Corin overexpression or sCorin reverses BPs-impaired BMSCs differentiation ability. Mechanistically, we find altered expression of phos-ERK in corin knockdown/overexpression BMSCs and BMSCs under sCorin stimulation. PD98059 (a selective ERK inhibitor) blocks the corin-mediated promotion effect. With regard to the high methylation level of corin during osteogenic differentiation, we apply a non-selective m6A methylase inhibitor, Cycloleucine, which also blocks the corin-mediated promotion effect. Furthermore, we demonstrate that METTL7A modulates corin m6A modification and reverses BPs-impaired BMSCs function, indicating that METTL7A regulates corin expression and thus contributes to orofacial BMSCs differentiation ability. To conclude, our study reveals that corin reverses BPs-induced BMSCs dysfunction, and METTL7A-mediated corin m6A modification underlies corin promotion of osteogenic differentiation via the ERK pathway. We hope this brings new insights into future clinical treatments for BRONJ.

Serum sclerostin as a biomarker of disease activity in ankylosing spondylitis in correlation with radiographic imaging

BackgroundThe wingless signaling pathway of bone development is inhibited by sclerostin, which may contribute to the etiology of ankylosing spondylitis.AimThe study aimed to evaluate serum sclerostin levels in ankylosing spondylitis patients and investigate how it correlated with radiographic damage using the Spondylo-arthritis Research Consortium of Canada index (SPARCC), disease activity, and functional impairment.ResultsThis cross-sectional case–control study revealed a significantly lower mean serum sclerostin (11.28 ng/ml) in AS patients compared with controls (101.25 ng/ml). Serum sclerostin levels showed a significant negative correlation with each of Bath Ankylosing Spondylitis Metrology Index (BASMI) (p = 0.043), sacroiliac joints SPARCC, spine SPARCC, and overall SPARCC scores (p = 0.012, p = 0.036, and p = 0.007). The detection of AS, serum sclerostin levels ≤ 20 ng/ml showed 100% sensitivity and specificity.ConclusionSerum sclerostin had good discriminating power between ankylosing spondylitis cases and healthy control individuals and was correlated with subclinical activity status on magnetic resonance imaging.

#49 Uremic toxins and inflammation: Metabolic pathways affected in stage 5 kidney disease

Abstract Background and Aims Chronic kidney disease (CKD) is a global health threat, affecting around 12% of the population. The uremic phenotype associated with CKD is considered a predictor for the high cardiovascular and overall mortality in these patients and inflammation is regarded as an essential contributor to it. We seek to discover new inflammation-related metabolites in the CKD population through the use of semitargeted metabolomics, in order to gain insight into the pathways involved and identify potential biomarkers. Method The study employed an observational prospective design to investigate metabolite impact on inflammation in 50 non-dialysis-dependent stage 5 CKD patients. Blood samples were collected before the start of the first haemodialysis session. A semitargeted metabolomic approach was followed, utilising liquid chromatography and high-resolution mass spectrometry with an in-house library of polar compounds. Univariate (Mann-Whitney test) and multivariate (logistic regression with LASSO regularization) analyses identified metabolomic variables associated with inflammation. Results Univariate analysis revealed that adenosine-5′-phosphosulfate (APS) (p-value (p) 0.0211), dimethyglycine (p 0.0015), pyruvate (p 0.0066), lactate (p 0.0156) and 2-ketobutyric acid (p 0.0458) were significantly increased with inflammation, with differences over 1,5-fold changes (inflammation = yes vs inflammation = no). Cholic acid (p 0.01655), homogentisic acid (p 0.0007), 2-phenylpropionic acid (p 0.0351) and 3-indoxylsulfate (p 0.0087) show similar patterns but in the opposite direction, as observed in the volcano plot in Fig. 1. Multivariate analysis revealed significant associations between metabolites and inflammation in CKD stage 5 patients. Specifically, odds ratios for N-Butyrylglycine (OR: 1.87), dimethylglycine (OR: 1.81), 2-Oxoisopentanoic acid (OR: 1.70), and pyruvate (OR: 1.71) were related to a higher probability of experiencing inflammation. Conversely, homogentisic acid (OR: 072), 2-Phenylpropionic acid (OR: 0.77), and 2-Methylglutaric acid (OR: 0.78) exhibited an OR<1, suggestive of decreased probability. This is graphically represented in Fig. 2. Conclusion In our semi-targeted metabolomics study on stage 5 CKD patients, we pinpointed various metabolites linked to inflammation. Notably, dimethylglycine, a uremic toxin, may contribute to vascular damage, while pyruvate's abnormal metabolism is a known feature in heart failure and chronic kidney disease. N-butyrilglycine and 2-oxoisopentanoic acid were also associated with inflammation, potentially tied to mitochondrial fatty acid beta oxidation and incomplete breakdown of branched-chain amino acids. APS, impacting bone development and intestinal permeability, showed a positive correlation with inflammation. Conversely, homogentisic acid and phenylpropionic acid were negatively correlated. While homogentisic acid's anti-inflammatory role isn't clear, phenylpropionic acid exhibits protective effects against hepatotoxicity by reducing immune response. 2-methylglutaric acid and cholic acid were inversely correlated with inflammation, suggesting a protective effect related to fatty acid metabolism and bile acid regulation. These identified metabolites enhance our understanding of CKD-associated inflammation and present strong candidates for future biomarker validation with potential for clinical translation.

Single Nucleotide Polymorphisms in RUNX2 and BMP2 contributes to different vertical facial profile.

The vertical facial profile is a crucial factor for facial harmony with significant implications for both aesthetic satisfaction and orthodontic treatment planning. However, the role of single nucleotide polymorphisms (SNPs) in the development of vertical facial proportions is still poorly understood. This study aimed to investigate the potential impact of some SNPs in genes associated with craniofacial bone development on the establishment of different vertical facial profiles. Vertical facial profiles were assessed by two senior orthodontists through pre-treatment digital lateral cephalograms. The vertical facial profile type was determined by recommended measurement according to the American Board of Orthodontics. Healthy orthodontic patients were divided into the following groups: "Normodivergent" (control group), "Hyperdivergent" and "Hypodivergent". Patients with a history of orthodontic or facial surgical intervention were excluded. Genomic DNA extracted from saliva samples was used for the genotyping of 7 SNPs in RUNX2, BMP2, BMP4 and SMAD6 genes using real-time polymerase chain reactions (PCR). The genotype distribution between groups was evaluated by uni- and multivariate analysis adjusted by age (alpha = 5%). A total of 272 patients were included, 158 (58.1%) were "Normodivergent", 68 (25.0%) were "Hyperdivergent", and 46 (16.9%) were "Hypodivergent". The SNPs rs1200425 (RUNX2) and rs1005464 (BMP2) were associated with a hyperdivergent vertical profile in uni- and multivariate analysis (p-value < 0.05). Synergistic effect was observed when evaluating both SNPs rs1200425- rs1005464 simultaneously (Prevalence Ratio = 4.0; 95% Confidence Interval = 1.2-13.4; p-value = 0.022). In conclusion, this study supports a link between genetic factors and the establishment of vertical facial profiles. SNPs in RUNX2 and BMP2 genes were identified as potential contributors to hyperdivergent facial profiles.

Bone and Joint-on-Chip Platforms: Construction Strategies and Applications.

Organ-on-a-chip, also known as "tissue chip," is an advanced platform based on microfluidic systems for constructing miniature organ models in vitro. They can replicate the complex physiological and pathological responses of human organs. In recent years, the development of bone and joint-on-chip platforms aims to simulate the complex physiological and pathological processes occurring in human bones and joints, including cell-cell interactions, the interplay of various biochemical factors, the effects of mechanical stimuli, and the intricate connections between multiple organs. In the future, bone and joint-on-chip platforms will integrate the advantages of multiple disciplines, bringing more possibilities for exploring disease mechanisms, drug screening, and personalized medicine. This review explores the construction and application of Organ-on-a-chip technology in bone and joint disease research, proposes a modular construction concept, and discusses the new opportunities and future challenges in the construction and application of bone and joint-on-chip platforms.

Cohort profile of an early life observational cohort in China: Bone and MicroBiOme onset (BAMBOO) study

PurposeThe Bone And MicroBiOme Onset (BAMBOO) study is an ongoing prospective observational cohort study conducted in Tianjin, China, aiming to determine age-appropriate trajectories for microbiome maturation and bone development and...

Fufang Zhenshu Tiaozhi capsule enhances bone formation and safeguards against glucocorticoid-induced osteoporosis through innovative Mekk2-mediated β-catenin deubiquitination.

Bone homeostasis depends on the regulation of β-catenin in osteoblasts. Glucocorticoids (GCs) are known to diminish β-catenin activity via Wnt pathway signaling, leading to osteoporosis. Conversely, activating β-catenin in osteoblasts through mitogen-activated protein kinase kinase kinase 2 (Mekk2) offers an innovative approach to combat GC-induced osteoporosis (GIOP). Fufang Zhenshu Tiaozhi (FTZ) capsules have shown effectiveness in treating GIOP, but the mechanisms behind this are still unclear. In this study, Mekk2 knockout mice (Mekk2-/-) was generated by CRISPR/Cas9. These mice were then subjected to Alcian Blue-Alizarin Red staining and immunofluorescence to assess their bone and cartilage development. To establish models of GIOP, both Mekk2-/- and wild-type (WT) mice were treated with dexamethasone (DXMS) and subsequently given FTZ capsules. We analyzed the resulting phenotypic changes in these mice using Micro-CT scans and histomorphological studies. Primary osteoblasts, isolated from both Mekk2-/- and WT mice, underwent qRT-PCR to measure key osteogenesis markers, including Runx2, Sp7, Bgalp, Col1a1 and Alp. Cells were then exposed to treatments with either FTZ or Wnt3a and the phosphorylation levels of β-catenin and Mekk2, along with the protein expression of Runx2, were evaluated using Western blotting and immunoprecipitation. Additionally, C3H10T1/2 cells transfected with TOPflash-luciferase and Renilla luciferase reporters were treated with FTZ and Wnt3a to measure β-catenin activity. In our study, administering FTZ in vivo effectively prevented bone loss typically induced by GCs. However, it's important to note that this protective effect was substantially reduced in mice lacking Mekk2. Additionally, FTZ showed a significant ability to enhance osteogenic differentiation in primary osteoblasts, doing so by altering the expression of Mekk2. Intriguingly, the impact of FTZ on Mekk2 appears to function through a pathway separate from the traditional Wnt signaling route. Furthermore, our findings indicate that FTZ also promotes the deubiquitination of β-catenin, contributing further to its positive effects on bone health. This study suggests that FTZ plays a significant role in protecting bone mass in cases of GIOP. The mechanism through which FTZ confers this benefit involves the activation of Mekk2/β-catenin signaling pathways, which represents a promising alternative strategy to counteract the deleterious effects of GIOP by augmenting osteoblastogenesis.

METTL3 Modulates Ctsk+ Lineage Supporting Cranial Osteogenesis via Hedgehog

N6-methyladenosine (m6A) modification, a eukaryotic messenger RNA modification catalyzed by methyltransferase-like 3 (METTL3), plays a pivotal role in stem cell fate determination. Calvarial bone development and maintenance are orchestrated by the cranial sutures. Cathepsin K (CTSK)–positive calvarial stem cells (CSCs) contribute to mice calvarial ossification. However, the role of m6A modification in regulating Ctsk+ lineage cells during calvarial development remains elusive. Here, we showed that METTL3 was colocalized with cranial nonosteoclastic Ctsk+ lineage cells, which were also associated with GLI1 expression. During neonatal development, depletion of Mettl3 in the Ctsk+ lineage cells delayed suture formation and decreased mineralization. During adulthood maintenance, loss of Mettl3 in the Ctsk+ lineage cells impaired calvarial bone formation, which was featured by the increased bone porosity, enhanced bone marrow cavity, and decreased number of osteocytes with the less-developed cellular outline. The analysis of methylated RNA immunoprecipitation sequencing and RNA sequencing data indicated that loss of METTL3 reduced Hedgehog (Hh) signaling pathway. Restoration of Hh signaling pathway by crossing Sufufl/+ alleles or by local administration of SAG21 partially rescued the abnormity. Our data indicate that METTL3 modulates Ctsk+ lineage cells supporting calvarial bone formation by regulating the Hh signaling pathway, providing new insights for clinical treatment of skull vault osseous diseases.

Deletion of Asb15b gene can lead to a significant decrease in zebrafish intermuscular bone

Intermuscular bones, which are present in numerous economically significant fish species, have a negative impact on the development of aquaculture. The Asb15b gene, primarily expressed in skeletal muscle, plays a crucial role in regulating protein turnover and the development of muscle fibers. It stimulates protein synthesis and controls the differentiation of muscle fibers. In this study, we employed CRISPR/Cas9 technology to generate homozygous zebrafish strains with 7 bp and 49 bp deletions in the Asb15b gene. Subsequent analyses using skeleton staining demonstrated a substantial reduction in the number of intermuscular bones in adult Asb15b−/− −7 bp and Asb15b−/− −49 bp mutants compared to the wild-type zebrafish, with decreases of 30 % (P < 0.001) and 40 % (P < 0.0001), respectively. Histological experiments further revealed that the diameter and number of muscle fibers in adult Asb15b−/− mutants did not exhibit significant changes when compared to wild-type zebrafish. Moreover, qRT-PCR experiments demonstrated significant differences in the expression of bmp6 and runx2b genes, which are key regulators of intermuscular bone development, during different stages of intermuscular bone development in Asb15b−/− mutants. This study strongly suggests that the Asb15b gene plays a crucial role in regulating intermuscular bone development in fish and lays the groundwork for further exploration of the role of the Asb15b gene in zebrafish intermuscular bone development.

Transcriptome Analysis Elucidates the Potential Key Genes Involved in Rib Development in bmp6-Deficient Silver Carp (Hypophthalmichthys molitrix).

Bone morphogenetic protein 6 (BMP-6) is a constituent of the TGF-β superfamily, known for its ability to stimulate bone and cartilage formation. The investigation of bmp6's involvement in the formation of intermuscular bones in fish has garnered significant attention in recent years. The rib cage is an important skeletal structure that plays a protective function for internal organs in fish. However, there has been limited research conducted on the effects of the bmp6 gene on rib development. Silver carp is one of four major fish in China, favoured for its affordability and tender muscle. Nevertheless, the presence of numerous intermuscular bones in silver carp significantly hinders the advancement of its palatability and suitability for processing. This study showcases the effective utilisation of CRISPR/Cas9 technology for the purpose of disrupting the bmp6 gene in silver carp, leading to the creation of chimeras in the P0 generation, marking the first instance of such an achievement. The chimeras exhibited complete viability, normal appearance, and partial intermuscular bones loss, with approximately 30% of them displaying rib bifurcation or bending. Subsequently, a transcriptome analysis on ribs of P0 chimeras and wild-type silver carp was conducted, leading to the identification of 934 genes exhibiting differential expression, of which 483 were found to be up-regulated and 451 were found to be down-regulated. The results of the KEGG analysis revealed that the "NF-kappa B signalling pathway", "Hippo signalling pathway", "osteoclast differentiation", and "haematopoietic cell lineage" exhibited enrichment and displayed a significant correlation with bone development. The up-regulated genes such as tnfα, fos, and ctgf in pathways may facilitate the proliferation and differentiation of osteoclasts, whereas the down-regulation of genes such as tgfb2 and tgfbr1 in pathways may hinder the formation and specialisation of osteoblasts, ultimately resulting in rib abnormalities. This study presents novel findings on the impact of bmp6 gene deletion on the rib development of silver carp, while simultaneously investigating the previously unexplored molecular mechanisms underlying rib defects in fish.

IRE1α pathway: A potential bone metabolism mediator.

Osteoblasts and osteoclasts collaborate in bone metabolism, facilitating bone development, maintaining normal bone density and strength, and aiding in the repair of pathological damage. Endoplasmic reticulum stress (ERS) can disrupt the intracellular equilibrium between osteoclast and osteoblast, resulting in dysfunctional bone metabolism. The inositol-requiring enzyme-1α (IRE1α) pathway-the most conservative unfolded protein response pathway activated by ERS-is crucial in regulating cell metabolism. This involvement encompasses functions such as inflammation, autophagy, and apoptosis. Many studies have highlighted the potential roles of the IRE1α pathway in osteoblasts, chondrocytes, and osteoclasts and its implication in certain bone-related diseases. These findings suggest that it may serve as a mediator for bone metabolism. However, relevant reviews on the role of the IRE1α pathway in bone metabolism remain unavailable. Therefore, this review aims to explore recent research that elucidated the intricate roles of the IRE1α pathway in bone metabolism, specifically in osteogenesis, chondrogenesis, osteoclastogenesis, and osteo-immunology. The findings may provide novel insights into regulating bone metabolism and treating bone-related diseases.

Hereditary human diseases with skeletal pathology – molecular pathogenesis and clinical characteristics

Differential diagnosis of hereditary diseases is challenging due to similar clinical manifestations, diversity of nosologies, wide clinical variability and genetic heterogeneity, severity, rare frequency and complex molecular etiology. In some of them, connective tissue as a whole is involved in the pathological process, affecting almost all organs and systems of the human body, and in particular bone tissue, disrupting its remodeling and microarchitecture. The defect can occur at any of the stages of remodeling: during the initiation process, during bone resorption, osteoblast differentiation, osteoid mineralization, etc. Violation of the microarchitecture of bone tissue is accompanied by the development of low-traumatic fractures and deformities, early osteoarthritis, the formation of contractures and chronic pain syndrome. Genetic studies of patients have expanded knowledge about the molecular signaling pathways that coordinate bone development and metabolism, the nature of disease inheritance, clinical features, and specific bone biomarkers. This article provides an overview of key cellular mechanisms, features of diagnosis and treatment of hereditary human diseases that affect the condition of bone tissue and skeleton: osteogenesis imperfecta, EhlersDanlos syndrome, Marfan syndrome, juvenile osteoporosis, hypophosphatasia, osteopetrosis, progressive diaphyseal dysplasia, mucopolysaccharidoses, achondroplasia, multiple hereditary exostoses. Doctors of different specialties can encounter these diseases, and making a correct diagnosis will make it possible to determine the correct algorithm for patient management and begin timely treatment, the ability to prevent the development of severe complications, improve the quality of life of patients, restore maximum working capacity and reduce the percentage of disability.

Exosomes: A New Hope for Angiogenesis-Mediated Bone Regeneration.

Bone is a metabolically dynamic structure that is generally remodeled throughout the lifetime of an individual but often causes problems with increasing age. A key player for bone development and homeostasis, but also under pathological conditions, is the bone vasculature. This complex system of arteries, veins, and capillaries forms distinct structures where each subset of endothelial cells has important functions. Starting with the basic process of angiogenesis and bone-specific blood vessel formation, coupled with initial bone formation, the importance of different vascular structures is highlighted with respect to how these structures are maintained or changed during homeostasis, aging, and pathological conditions. After exemplifying the current knowledge on bone vasculature, this review will move on to exosomes, a novel hotspot of scientific research. Exosomes will be introduced starting from their discovery via current isolation procedures and state-of-the-art characterization to their role in bone vascular development, homeostasis, and bone regeneration and repair while summarizing the underlying signal transduction pathways. With respect to their role in these processes, especially mesenchymal stem cell-derived extracellular vesicles are of interest, which leads to a discussion on patented applications and an update on ongoing clinical trials. Taken together, this review provides an overview of bone vasculature and bone regeneration, with a major focus on how exosomes influence this intricate system, as they might be useful for therapeutic purposes in the near future.

Exploring the Regulators of Keratinization: Role of BMP-2 in Oral Mucosa.

The oral mucosa functions as a physico-chemical and immune barrier to external stimuli, and an adequate width of the keratinized mucosa around the teeth or implants is crucial to maintaining them in a healthy and stable condition. In this study, for the first time, bulk RNA-seq analysis was performed to explore the gene expression of laser microdissected epithelium and lamina propria from mice, aiming to investigate the differences between keratinized and non-keratinized oral mucosa. Based on the differentially expressed genes (DEGs) and Gene Ontology (GO) Enrichment Analysis, bone morphogenetic protein 2 (BMP-2) was identified to be a potential regulator of oral mucosal keratinization. Monoculture and epithelial-mesenchymal cell co-culture models in the air-liquid interface (ALI) indicated that BMP-2 has direct and positive effects on epithelial keratinization and proliferation. We further performed bulk RNA-seq of the ALI monoculture stimulated with BMP-2 in an attempt to identify the downstream factors promoting epithelial keratinization and proliferation. Analysis of the DEGs identified, among others, IGF2, ID1, LTBP1, LOX, SERPINE1, IL24, and MMP1 as key factors. In summary, these results revealed the involvement of a well-known growth factor responsible for bone development, BMP-2, in the mechanism of oral mucosal keratinization and proliferation, and pointed out the possible downstream genes involved in this mechanism.

RanGAP1 maintains chromosome stability in limb bud mesenchymal cells during bone development

BackgroundBone development involves the rapid proliferation and differentiation of osteogenic lineage cells, which makes accurate chromosomal segregation crucial for ensuring cell proliferation and maintaining chromosomal stability. However, the mechanism underlying the maintenance of chromosome stability during the rapid proliferation and differentiation of Prx1-expressing limb bud mesenchymal cells into osteoblastic precursor cells remains unexplored. MethodsA transgenic mouse model of RanGAP1 knockout of limb and head mesenchymal progenitor cells was constructed to explore the impact of RanGAP1 deletion on bone development by histomorphology and immunostaining. Subsequently, G-banding karyotyping analysis and immunofluorescence staining were used to examine the effects of RanGAP1 deficiency on chromosome instability. Finally, the effects of RanGAP1 deficiency on chromothripsis and bone development signaling pathways were elucidated by whole-genome sequencing, RNA-sequencing, and qPCR. ResultsThe ablation of RanGAP1 in limb and head mesenchymal progenitor cells expressing Prx1 in mice resulted in embryonic lethality, severe cartilage and bone dysplasia, and complete loss of cranial vault formation. Moreover, RanGAP1 loss inhibited chondrogenic or osteogenic differentiation of mesenchymal stem cells (MSCs). Most importantly, we found that RanGAP1 loss in limb bud mesenchymal cells triggered missegregation of chromosomes, resulting in chromothripsis of chromosomes 1q and 14q, further inhibiting the expression of key genes involved in multiple bone development signaling pathways such as WNT, Hedgehog, TGF-β/BMP, and PI3K/AKT in the chromothripsis regions, ultimately disrupting skeletal development. ConclusionsOur results establish RanGAP1 as a critical regulator of bone development, as it supports this process by preserving chromosome stability in Prx1-expressing limb bud mesenchymal cells.

Diatomite-incorporated hierarchical scaffolds for osteochondral regeneration

Osteochondral regeneration involves the highly challenging and complex reconstruction of cartilage and subchondral bone. Silicon (Si) ions play a crucial role in bone development. Current research on Si ions mainly focuses on bone repair, by using silicate bioceramics with complex ion compositions. However, it is unclear whether the Si ions have important effect on cartilage regeneration. Developing a scaffold that solely releases Si ions to simultaneously promote subchondral bone repair and stimulate cartilage regeneration is critically important. Diatomite (DE) is a natural diatomaceous sediment that can stably release Si ions, known for its abundant availability, low cost, and environmental friendliness. Herein, a hierarchical osteochondral repair scaffold is uniquely designed by incorporating gradient DE into GelMA hydrogel. The adding DE microparticles provides a specific Si source for controlled Si ions release, which not only promotes osteogenic differentiation of rBMSCs (rabbit bone marrow mesenchymal stem cells) but also enhances proliferation and maturation of chondrocytes. Moreover, DE-incorporated hierarchical scaffolds significantly promoted the regeneration of cartilage and subchondral bone. The study suggests the significant role of Si ions in promoting cartilage regeneration and solidifies their foundational role in enhancing bone repair. Furthermore, it offers an economic and eco-friendly strategy for developing high value-added osteochondral regenerative bioscaffolds from low-value ocean natural materials.