Resistance training presents beneficial effects on bone development of adolescents engaged in swimming but not in impact sports: ABCD Growth Study.

BACKGROUND: Sports practice during adolescence is important to enhance bone development, although it may provide different effects depending on the mechanical impact present in the sport. Besides, resistance training (RT) may also induce bone changes directly (via muscle contractions) and indirectly (via myokines). However, there have been no studies analyzing the longitudinal influence of engaging in sport with and without added mechanical load. Thus, this study aims to analyze the combined effects of sports participation and resistance training on areal bone mineral density (aBMD) accrual in adolescent athletes participating in swimming and impact sports for 12-months. METHODS: This was a 12-month longitudinal study. The sample comprised 91 adolescents (21 females) aged 10 to 18 years, engaged in impact sports (basketball, tennis, track & field, baseball and gymnastics, n = 66) and non-impact sport (swimming, n = 25). The sample was divided according to resistance training participation: impact sports only (n = 45), impact sports + resistance training (n = 21), swimming-only (n = 17) and swimming + resistance training (n = 8). aBMD and soft tissues were measured using dual-energy X-ray absorptiometry. Generalized linear models analysis was used for the resistance training (RT) x type of sport interaction in predicting aBMD changes overtime, adjusting for maturation, sex and baseline aBMD. RESULTS: After 12-months, all groups showed a significant increase in aBMD, except for the swimming groups (regardless of resistant training), which showed a significant loss in spine aBMD (-0.045 [-0.085 to -0.004] g/cm2 in swimming-only and - 0.047 [-0.073 to -0.021] g/cm2 in swimming + RT). In comparisons between groups, only swimming + RT group, compared with swimming-only group presented higher upper limbs aBMD (0.096 g/cm2 [0.074 to 0.118] in swimming + RT vs. 0.046 [0.032 to 0.060] g/cm2 in swimming only; p < 0.05) and whole body less head (WBLH) aBMD (0.039 [0.024 to 0.054] g/cm2 in swimming + RT vs. 0.017 [0.007 to 0.027] g/cm2 swimming-only; p < 0.05). CONCLUSION: Despite the significant gain in aBMD in all groups and body sites after 12-months, except for the spine site of swimmers, the results indicate that participation in RT seems to improve aBMD accrual in swimmers at the upper limbs and WBLH.

Noncoding RNAs in skeletal development and disorders.

Protein-encoding genes only constitute less than 2% of total human genomic sequences, and 98% of genetic information was previously referred to as "junk DNA". Meanwhile, non-coding RNAs (ncRNAs) consist of approximately 60% of the transcriptional output of human cells. Thousands of ncRNAs have been identified in recent decades, and their essential roles in the regulation of gene expression in diverse cellular pathways associated with fundamental cell processes, including proliferation, differentiation, apoptosis, and metabolism, have been extensively investigated. Furthermore, the gene regulation networks they form modulate gene expression in normal development and under pathological conditions. In this review, we integrate current information about the classification, biogenesis, and function of ncRNAs and how these ncRNAs support skeletal development through their regulation of critical genes and signaling pathways in vivo. We also summarize the updated knowledge of ncRNAs involved in common skeletal diseases and disorders, including but not limited to osteoporosis, osteoarthritis, rheumatoid arthritis, scoliosis, and intervertebral disc degeneration, by highlighting their roles established from in vivo, in vitro, and ex vivo studies.

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Runt-related transcription factor (RUNX1) is a transcription factor closely involved in hematopoiesis. RUNX1 gene mutation plays an essential pathogenic role in the initiation and development of hematological tumors, especially in acute myeloid leukemia. Recent studies have shown that RUNX1 is also involved in the regulation of bone development and the pathological progression of bone-related diseases. RUNX1 promotes the differentiation of mesenchymal stem cells into chondrocytes and osteoblasts and modulates the maturation and extracellular matrix formation of chondrocytes. The expression of RUNX1 in mesenchymal stem cells, chondrocytes, and osteoblasts is of great significance for maintaining normal bone development and the mass and quality of bones. RUNX1 also inhibits the differentiation and bone resorptive activities of osteoclasts, which may be influenced by sexual dimorphism. In addition, RUNX1 deficiency contributes to the pathogenesis of osteoarthritis, delayed fracture healing, and osteoporosis, which was revealed by the RUNX1 conditional knockout modeling in mice. However, the roles of RUNX1 in regulating the hypertrophic differentiation of chondrocytes, the sexual dimorphism of activities of osteoclasts, as well as bone loss in diabetes mellitus, senescence, infection, chronic inflammation, etc, are still not fully understood. This review provides a systematic summary of the research progress concerning RUNX1 in the field of bone biology, offering new ideas for using RUNX1 as a potential target for bone related diseases, especially osteoarthritis, delayed fracture healing, and osteoporosis.

Chondrocyte hypertrophy in the growth plate promotes stress anisotropy affecting long bone development through chondrocyte column formation.

The length of long bones is determined by column formation of proliferative chondrocytes and subsequent chondrocyte hypertrophy in the growth plate during bone development. Despite the importance of mechanical loading in long bone development, the mechanical conditions of the cells within the growth plate, such as the stress field, remain unclear owing to the difficulty in investigating spatiotemporal changes within dynamically growing tissues. In this study, the mechanisms of longitudinal bone growth were investigated from a mechanical perspective through column formation of proliferative chondrocytes within the growth plate before secondary ossification center formation using continuum-based particle models (CbPMs). A one-factor model, which simply describes essential aspects of a biological signaling cascade regulating cell activities within the growth plate, was developed and incorporated into CbPM. Subsequently, the developmental process and maintenance of the growth plate structure and resulting bone morphogenesis were simulated. Thus, stress anisotropy in the proliferative zone that affects bone elongation through chondrocyte column formation was identified and found to be promoted by chondrocyte hypertrophy. These results provide further insights into the mechanical regulation of multicellular dynamics during bone development.

The Origin and Fate of Chondrocytes: Cell Plasticity in Physiological Setting.

PURPOSE OF REVIEW: Here, we discuss the origin of chondrocytes, their destiny, and their plasticity in relationship to bone growth, articulation, and formation of the trabeculae. We also consider these processes from a biological, clinical, and evolutionary perspective. RECENT FINDINGS: Chondrocytes, which provide the template for the formation of most bones, are responsible for skeletal growth and articulation during postnatal life. In recent years our understanding of the fate of these cells has changed dramatically. Current evidence indicates a paradoxical situation during skeletogenesis, with some cells of mesenchymal condensation differentiating directly into osteoblasts, whereas others of the same kind give rise to highly similar osteoblasts via a complex process of differentiation involving several chondrocyte intermediates. The situation becomes even more paradoxical during postnatal growth when stem cells in the growth plate produce differentiated, functional progenies, which thereafter presumably dedifferentiate into another type of stem cell. Such a remarkable transition from one cell type to another under postnatal physiological conditions provides a fascinating example of cellular plasticity that may have valuable clinical implications.

Impact loading intensifies cortical bone (re)modeling and alters longitudinal bone growth of pubertal rats.

Physical exercise is important for musculoskeletal development during puberty, which builds bone mass foundation for later in life. However, strenuous levels of training might bring adverse effects to bone health, reducing longitudinal bone growth. Animal models with various levels of physical exercise were largely used to provide knowledge to clinical settings. Experiments from our previous studies applied different levels of mechanical loading on rat tibia during puberty accompanied by weekly in vivo micro-CT scans. In the present article, we apply 3D image registration-based methods to retrospectively analyze part of the previously acquired micro-CT data. Longitudinal bone growth, growth plate thickness, and cortical bone (re)modeling were evaluated from rats' age of 28-77 days. Our results show that impact loading inhibited proximal bone growth throughout puberty. We hypothesize that impact loading might bring different growth alterations to the distal and proximal growth plates. High impact loading might lead to pathological consequence of osteochondrosis and catch-up growth due to growth inhibition. Impact loading also increased cortical bone (re)modeling before and after the peak proximal bone growth period of young rats, of which the latter case might be caused by the shift from modeling to remodeling as the dominant activity toward the end of rat puberty. We confirm that the tibial endosteum is more mechano-sensitive than the periosteum in response to mechanical loading. To our knowledge, this is the first study to follow up bone growth and bone (re)modeling of young rats throughout the entire puberty with a weekly time interval.

An in silico model for woven bone adaptation to heavy loading conditions in murine tibia.

Existing in silico models for lamellar bone adaptation to mechanical loading are unsuitable for predicting woven bone growth. This anomaly is due to the difference in mechanobiology of the woven bone with respect to that of the lamellar bone. The present study is aimed at developing an in silico bone-adaptation model for woven bone at cellular and tissue levels. The diffusion of Ca2+ ions reaching lining cells from the osteocytic network and the bone cortex in response to a mechanical loading on the cortical bone has been considered as a stimulus. The diffusion of ions within osteocytic network has been computed with a lacunar-canalicular network (LCN) in which bone cells are uniformly arranged. Strain energy density is assumed to regulate ion flow within the network when the induced normal strain is above a threshold level. If the induced strain exceeds another higher threshold level, then the strain with a power constant is additionally assumed to regulate the stimulus. The intracellular flow of Ca2+ ions within the LCN has been simulated using Fick's laws of diffusion, using a finite element method. The ion diffusion from bone cortex to vesicles has been formulated as a normal strain with a power constant. The stimuli reaching the surface cells are assumed to form the new bone. The mathematical model closely predicts woven bone growth in mouse and rat tibia for various in vivo loading conditions. This model is the first to predict woven bone growth at tissue and cellular levels in response to heavy mechanical loading.

Nox4 expression in osteo-progenitors controls bone development in mice during early life.

Tightly regulated and cell-specific NADPH-oxidases (Nox) represent one of the major sources of reactive oxygen species (ROS) signaling molecules that are involved in tissue development and stem cell self-renewal. We have characterized the role of Nox4 in osteo-progenitors during postnatal bone development. Nox4 expression in bone and ROS generation were increased during early osteoblast differentiation and bone development. Stromal osteoblastic cell self-renewal, proliferation and ROS production were significantly lower in samples from whole-body Nox4 knockout mice (Nox4-/-) and conditional knockout (CKO) mice with depletion of Nox4 in the limb bud mesenchyme compared with those from control mice (Nox4fl/fl), but they were reversed after 9 passages. In both sexes, bone volume, trabecular number and bone mineral density were significantly lower in 3-week old CKO and Nox4-/- mice compared with Nox4fl/fl controls. This was reflected in serum levels of bone formation markers alkaline phosphatase (ALP) and procollagen 1 intact N-terminal propeptide (P1NP). However, under-developed bone formation in 3-week old CKO and Nox4-/- mice quickly caught up to levels of control mice by 6-week of age, remained no different at 13-week of age, and was reversed in 32-week old male mice. Osteoclastogenesis showed no differences among groups, however, CTX1 reflecting osteoclast activity was significantly higher in 3-week old male CKO and Nox4-/- mice compared with control mice, and significantly lower in 32-week old Nox4-/- mice compared with control mice. These data suggest that Nox4 expression and ROS signaling in bone and osteoblastic cells coordinately play an important role in osteoblast differentiation, proliferation and maturation.

Klotho in Osx+-mesenchymal progenitors exerts pro-osteogenic and anti-inflammatory effects during mandibular alveolar bone formation and repair.

Maxillofacial bone defects are commonly seen in clinical practice. A clearer understanding of the regulatory network directing maxillofacial bone formation will promote the development of novel therapeutic approaches for bone regeneration. The fibroblast growth factor (FGF) signalling pathway is critical for the development of maxillofacial bone. Klotho, a type I transmembrane protein, is an important components of FGF receptor complexes. Recent studies have reported the presence of Klotho expression in bone. However, the role of Klotho in cranioskeletal development and repair remains unknown. Here, we use a genetic strategy to report that deletion of Klotho in Osx-positive mesenchymal progenitors leads to a significant reduction in osteogenesis under physiological and pathological conditions. Klotho-deficient mensenchymal progenitors also suppress osteoclastogenesis in vitro and in vivo. Under conditions of inflammation and trauma-induced bone loss, we find that Klotho exerts an inhibitory function on inflammation-induced TNFR signaling by attenuating Rankl expression. More importantly, we show for the first time that Klotho is present in human alveolar bone, with a distinct expression pattern under both normal and pathological conditions. In summary, our results identify the mechanism whereby Klotho expressed in Osx+-mensenchymal progenitors controls osteoblast differentiation and osteoclastogenesis during mandibular alveolar bone formation and repair. Klotho-mediated signaling is an important component of alveolar bone remodeling and regeneration. It may also be a target for future therapeutics.

Characterisation of Growth Plate Dynamics in Murine Models of Osteoarthritis.

The purpose of this study was to investigate growth plate dynamics in surgical and loading murine models of osteoarthritis, to understand whether abnormalities in these dynamics are associated with osteoarthritis development. 8-week-old C57BL/6 male mice underwent destabilisation of medial meniscus (DMM) (n = 8) surgery in right knee joints. Contralateral left knee joints had no intervention (controls). In 16-week-old C57BL/6 male mice (n = 6), osteoarthritis was induced using non-invasive mechanical loading of right knee joints with peak force of 11N. Non-loaded left knee joints were internal controls. Chondrocyte transiency in tibial articular cartilage and growth plate was confirmed by histology and immunohistochemistry. Tibial subchondral bone parameters were measured using microCT and correlated to 3-dimensional (3D) growth plate bridging analysis. Higher expression of chondrocyte hypertrophy markers; Col10a1 and MMP13 were observed in tibial articular cartilage chondrocytes of DMM and loaded mice. In tibial growth plate, Col10a1 and MMP13 expressions were widely expressed in a significantly enlarged zone of proliferative and hypertrophic chondrocytes in DMM (p=0.002 and p<0.0001, respectively) and loaded (both p<0.0001) tibiae of mice compared to their controls. 3D quantification revealed enriched growth plate bridging and higher bridge densities in medial compared to lateral tibiae of DMM and loaded knee joints of the mice. Growth plate dynamics were associated with increased subchondral bone volume fraction (BV/TV; %) in medial tibiae of DMM and loaded knee joints and epiphyseal trabecular bone volume fraction in medial tibiae of loaded knee joints. The results confirm articular cartilage chondrocyte transiency in a surgical and loaded murine models of osteoarthritis. Herein, we reveal spatial variation of growth plate bridging in surgical and loaded osteoarthritis models and how these may contribute to anatomical variation in vulnerability of osteoarthritis development.

Evolution and Future of Growth Plate Therapeutics.

BACKGROUND: Longitudinal bone growth is regulated by multiple endocrine signals (e.g., growth hormone, insulin-like growth factor I, estrogen, and androgen) and local factors (e.g., fibroblast growth factors and their receptors and the C-natriuretic peptide/natriuretic peptide receptor-B pathway). SUMMARY: Abnormalities in both endocrine and local regulation of growth plate physiology cause many disorders of human skeletal growth. Knowledge of these pathways creates therapeutic potential for sustaining or even augmenting linear growth. Key Message: During the past 4 decades, advances in understanding growth plate physiology have been accompanied by development and implementation of growth-promoting treatments that have progressed in both efficacy and specificity of action. This paper reviews the history and continuing evolution of growth plate therapeutics.

PiRNA-63049 inhibits bone formation through Wnt/ß-catenin signaling pathway.

Bone remodeling is a dynamic process between bone formation mediated by osteoblasts and bone resorption mediated by osteoclasts. Disrupted bone remodeling is a key factor in postmenopausal osteoporosis, a metabolic disorder characterized by deteriorated bone microarchitecture and increased risk of fracture. Recent studies have shown that piwi-binding RNA (piRNA) is involved in the pathogenesis of certain diseases at the post-transcriptional level. Here, we analyzed piRNA-63049 (piR-63049), which may play an essential role in bone remodeling. The expression of piR-63049 significantly increased in both bone tissues and plasma of osteoporotic rats and postmenopausal osteoporotic patients. Overexpressing piR-63049 could inhibit the osteoblastogenesis of bone marrow stromal cells (BMSCs) while knocking down piR-63049 could promote the osteoblastogenesis of BMSCs through the Wnt2b/ß-catenin signaling pathway. Moreover, knocking-down piR-63049 (piR-63049-antagonist) in vivo could attenuate the bone loss in ovariectomized rats by promoting bone formation. Taken together, the current study shows that piR-63049 inhibits bone formation through the Wnt2b/ß-catenin signaling pathway. This novel piRNA may be a potential target to increase bone formation in bone loss disorders such as postmenopausal osteoporosis.

Congenital Metabolic Bone Disorders as a Cause of Bone Fragility.

Bone fragility is a pathological condition caused by altered homeostasis of the mineralized bone mass with deterioration of the microarchitecture of the bone tissue, which results in a reduction of bone strength and an increased risk of fracture, even in the absence of high-impact trauma. The most common cause of bone fragility is primary osteoporosis in the elderly. However, bone fragility can manifest at any age, within the context of a wide spectrum of congenital rare bone metabolic diseases in which the inherited genetic defect alters correct bone modeling and remodeling at different points and aspects of bone synthesis and/or bone resorption, leading to defective bone tissue highly prone to long bone bowing, stress fractures and pseudofractures, and/or fragility fractures. To date, over 100 different Mendelian-inherited metabolic bone disorders have been identified and included in the OMIM database, associated with germinal heterozygote, compound heterozygote, or homozygote mutations, affecting over 80 different genes involved in the regulation of bone and mineral metabolism. This manuscript reviews clinical bone phenotypes, and the associated bone fragility in rare congenital metabolic bone disorders, following a disease taxonomic classification based on deranged bone metabolic activity.

Pancreas deficiency modifies bone development in the ovine fetus near term.

Hormones have an important role in the regulation of fetal growth and development, especially in response to nutrient availability in utero. Using micro-CT and an electromagnetic three-point bend test, this study examined the effect of pancreas removal at 0.8 fraction of gestation on the developing bone structure and mechanical strength in fetal sheep. When fetuses were studied at 10 and 25 days after surgery, pancreatectomy caused hypoinsulinaemia, hyperglycaemia and growth retardation which was associated with low plasma concentrations of leptin and a marker of osteoclast activity and collagen degradation. In pancreatectomized fetuses compared to control fetuses, limb lengths were shorter, and trabecular (Tb) bone in the metatarsi showed greater bone volume fraction, Tb thickness, degree of anisotropy and porosity, and lower fractional bone surface area and Tb spacing. Mechanical strength testing showed that pancreas deficiency was associated with increased stiffness and a greater maximal weight load at fracture in a subset of fetuses studied near term. Overall, pancreas deficiency in utero slowed the growth of the fetal skeleton and adapted the developing bone to generate a more compact and connected structure. Maintenance of bone strength in growth-retarded limbs is especially important in a precocial species in preparation for skeletal loading and locomotion at birth.

Reconstitution of the host holobiont in germ-free born male rats acutely increases bone growth and affects marrow cellular content.

Integration of microbiota in a host begins at birth and progresses during adolescence, forming a multidirectional system of physiological interactions. Here, we present an instantaneous effect of natural, bacterial gut colonization on the acceleration of longitudinal and radial bone growth in germ-free born, 7-wk-old male rats. Changes in bone mass and structure were analyzed after 10 days following the onset of colonization through cohousing with conventional rats and revealed unprecedented acceleration of bone accrual in cortical and trabecular compartments, increased bone tissue mineral density, improved proliferation and hypertrophy of growth plate chondrocytes, bone lengthening, and preferential deposition of periosteal bone in the tibia diaphysis. In addition, the number of small in size adipocytes increased, whereas the number of megakaryocytes decreased, in the bone marrow of conventionalized germ-free rats indicating that not only bone mass but also bone marrow environment is under control of gut microbiota signaling. The changes in bone status paralleled with a positive shift in microbiota composition toward short-chain fatty acids (SCFA)-producing microbes and a considerable increase in cecal SCFA concentrations, specifically butyrate. Furthermore, reconstitution of the host holobiont increased hepatic expression of IGF-1 and its circulating levels. Elevated serum levels of 25-hydroxy vitamin D and alkaline phosphatase pointed toward an active process of bone formation. The acute stimulatory effect on bone growth occurred independently of body mass increase. Overall, the presented model of conventionalized germ-free rats could be used to study microbiota-based therapeutics for combatting dysbiosis-related bone disorders.

The endothelium-bone axis in development, homeostasis and bone and joint disease.

Blood vessels form a versatile transport network that is best known for its critical roles in processes such as tissue oxygenation, metabolism and immune surveillance. The vasculature also provides local, often organ-specific, molecular signals that control the behaviour of other cell types in their vicinity during development, homeostasis and regeneration, and also in disease processes. In the skeletal system, the local vasculature is actively involved in both bone formation and resorption. In addition, blood vessels participate in inflammatory processes and contribute to the pathogenesis of diseases that affect the joints, such as rheumatoid arthritis and osteoarthritis. This Review summarizes the current understanding of the architecture, angiogenic growth and functional properties of the bone vasculature. The effects of ageing and pathological conditions, including arthritis and osteoporosis, are also discussed.

Endogenous Glucocorticoid Metabolism in Bone: Friend or Foe.

The role of tissue specific metabolism of endogenous glucocorticoids (GCs) in the pathogenesis of human disease has been a field of intense interest over the last 20 years, fuelling clinical trials of metabolism inhibitors in the treatment of an array of metabolic diseases. Localised pre-receptor metabolism of endogenous and therapeutic GCs by the 11ß-hydroxysteroid dehydrogenase (11ß-HSD) enzymes (which interconvert endogenous GCs between their inactive and active forms) are increasingly recognised as being critical in mediating both their positive and negative actions on bone homeostasis. In this review we explore the roles of endogenous and therapeutic GC metabolism by the 11ß-HSD enzymes in the context of bone metabolism and bone cell function, and consider future strategies aimed at modulating this system in order to manage and treat various bone diseases.

Novel polychrome staining distinguishing osteochondral tissue and bone cells in decalcified paraffin sections.

The bone is a dynamic and metabolically active organ in which growth and resorption of the osteochondral matrix is orchestrated by osteoblasts and osteoclasts. For decalcified paraffin-embedded specimens, decalcifying agents alter the staining intensity, and excess decalcification interferes with bone staining. Robust bone staining methods independent of the decalcification conditions and animal species are lacking. In this study, we have developed a novel polychrome staining method, named JFRL staining, which stains the components of osteochondral tissue in different colors. With this staining we could visualize the hyaline cartilage as blue by alcian blue, osteoid as red by picrosirius red, and mineralized bone as green by picro-light green SF or picro-naphthol green B and easily distinguished osteoblasts, osteocytes, and osteoclasts. In mineralized bone, this staining revealed the obvious lamellar structures and woven bone. Notably, this staining was independent of the decalcification conditions and experimental animal species examined. To verify the usefulness of JFRL staining, we observed cotton rat tail which has shorter length and shows a false autotomy. The caudal vertebrae were normally developed via endochondral ossification without a fracture plane. At 6 months of age, the number of chondrocytes declined and the hypertrophic zone was absent at the epiphyseal plate, which might reflect the shorter tail. In conclusion, JFRL staining is the first method to simultaneously distinguish osteochondral matrix and bone cells in one section regardless of decalcifying conditions. This robust staining will provide new information for a wide number of biomedical fields, including bone development, physiology, and pathology.

GATA4-driven miR-206-3p signatures control orofacial bone development by regulating osteogenic and osteoclastic activity.

Growth disorders in the orofacial bone development process may lead to orofacial deformities. The balance between bone matrix formation by mesenchymal lineage osteoblasts and bone resorption by osteoclasts is vital for orofacial bone development. Although the mechanisms of orofacial mesenchymal stem cells (OMSCs) in orofacial bone development have been studied intensively, the communication between OMSCs and osteoclasts remains largely unclear. Methods: We used a neural crest cell-specific knockout mouse model to investigate orofacial bone development in GATA-binding protein 4 (GATA4) morphants. We investigated the underlying mechanisms of OMSCs-derived exosomes (OMExos) on osteoclastogenesis and bone resorption activity in vitro. miRNAs were extracted from OMExos, and differences in miRNA abundances were determined using an Affymetrix miRNA array. Luciferase reporter assays were used to validate the binding between GATA4 and miR-206-3p in OMSCs and to confirm the putative binding of miR-206-3p and its target genes in OMSCs and osteoclasts. The regulatory mechanism of the GATA4-miR-206-3p axis in OMSC osteogenic differentiation and osteoclastogenesis was examined in vitro and in vivo. Results: Wnt1-Cre;Gata4fl/fl mice (cKO) not only presented inhibited bone formation but also showed active bone resorption. Osteoclasts cocultured in vitro with cKO OMSCs presented an increased capacity for osteoclastogenesis, which was exosome-dependent. Affymetrix miRNA array analysis showed that miR-206-3p was downregulated in exosomes from shGATA4 OMSCs. Moreover, the transcriptional activity of miR-206-3p was directly regulated by GATA4 in OMSCs. We further demonstrated that miR-206-3p played a key role in the regulation of orofacial bone development by directly targeting bone morphogenetic protein-3 (Bmp3) and nuclear factor of activated T -cells, cytoplasmic 1 (NFATc1). OMExos and agomiR-206-3p enhanced bone mass in Wnt1-cre;Gata4fl/fl mice by augmenting trabecular bone structure and decreasing osteoclast numbers. Conclusion: Our findings confirm that miR-206-3p is an important downstream factor of GATA4 that regulates the functions of OMSCs and osteoclasts. These results demonstrate the efficiency of OMExos and microRNA agomirs in promoting bone regeneration, which provide an ideal therapeutic tool for orofacial bone deformities in the future.

Probiotics Stimulate Bone Formation in Obese Mice via Histone Methylations.

Rationale: Manipulation of the gut microbiome can prevent pathologic bone loss. However, the effects of probiotics on mitochondrial epigenetic remodeling and skeletal homeostasis in the high-fat diet (HFD)-linked obesity remains to be explored. Here, we examined the impact of probiotics supplementation on mitochondrial biogenesis and bone homeostasis through the histone methylation mechanism in HFD fed obese mice. Methods: 16S rRNA gene sequencing was performed to study the microbiota composition in the gut and microbial dysbiosis in obese mouse model. High resolution (microPET/CT) imaging was performed to demonstrate the obese associated colonic inflammation. Obese-associated upregulation of target miRNA in osteoblast was investigated using a microRNA qPCR array. Osteoblastic mitochondrial mass was evaluated using confocal imaging. Overexpression of mitochondrial transcription factor (Tfam) was used to investigate the glycolysis and mitochondrial bioenergetic metabolism using Tfam-transgenic (Tg) mice fed on HFD. The bone formation and mechanical strength was evaluated by microCT analysis and three-point bending analysis. Results: High-resolution imaging (µ-CT) and mechanical testing revealed that probiotics induced a significant increase of trabecular bone volume and bone mechanical strength respectively in obese mice. Probiotics or Indole-3-propionic acid (IPA) treatment directly to obese mice, prevents gut inflammation, and improved osteoblast mineralization. Mechanistically, probiotics treatment increases mitochondrial transcription factor A (Tfam) expression in osteoblasts by promoting Kdm6b/Jmjd3 histone demethylase, which inhibits H3K27me3 epigenetic methylation at the Tfam promoter. Furthermore, Tfam-transgenic (Tg) mice, fed with HFD, did not experience obesity-linked reduction of glucose uptake, mitochondrial biogenesis and mineralization in osteoblasts. Conclusions: These results suggest that the probiotics mediated changes in the gut microbiome and its derived metabolite, IPA are potentially be a novel agent for regulating bone anabolism via the gut-bone axis.