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Aging Cell Feb 2021Impaired osteoblast function is involved in osteoporosis, and microRNA (miRNA) dysregulation may cause abnormal osteoblast osteogenic activity. However, the influence of...
Impaired osteoblast function is involved in osteoporosis, and microRNA (miRNA) dysregulation may cause abnormal osteoblast osteogenic activity. However, the influence of miRNA on osteoblast activity and the underlying mechanisms remain elusive. In this study, miR-103-3p was found to be negatively correlated with bone formation in bone specimens from elderly women with fractures and ovariectomized (OVX) mice. Additionally, miR-103-3p directly targeted Mettl14 to inhibit osteoblast activity, and METTL14-dependent N -methyladenosine (m A) methylation inhibited miR-103-3p processing by the microprocessor protein DGCR8 and promoted osteoblast activity. Moreover, miR-103-3p inhibited bone formation in vivo, and therapeutic inhibition of miR-103-3p counteracted the decreased bone formation in OVX mice. Further, METTL14 was negatively correlated with miR-103-3p but positively correlated with bone formation in bone specimens from elderly women with fractures and OVX mice. Collectively, our results highlight the critical roles of the miR-103-3p/METTL14/m A signaling axis in osteoblast activity, identifying this axis as a potential target for ameliorating osteoporosis.
Topics: Animals; Bone Resorption; Methyltransferases; Mice; MicroRNAs; Osteoblasts
PubMed: 33440070
DOI: 10.1111/acel.13298 -
Laboratory Investigation; a Journal of... Feb 2019Osteoblast differentiation plays a critical role in bone formation and maintaining balance in bone remodeling. Runt-related transcription factor 2 (Runx2) is a central...
Osteoblast differentiation plays a critical role in bone formation and maintaining balance in bone remodeling. Runt-related transcription factor 2 (Runx2) is a central transcription factor regulating osteoblast differentiation and promoting bone mineralization. Until now, the molecular regulatory basis and especially the gene regulatory network of osteogenic differentiation have been unclear. Krüppel-like factor 2 (KLF2) is a zinc finger structure and DNA-binding transcription factor. The current study aimed to investigate the physiological function of KLF2 in osteoblast differentiation. Our results indicate that KLF2 is expressed in pre-osteoblast MC3T3-E1 cells and primary osteoblasts. Interestingly, KLF2 expression is increased in osteoblasts during the osteoblastic differentiation process. Overexpression of KLF2 in MC3T3-E1 cells promoted the expression of the osteoblastic differentiation marker genes Alp, Osx, and Ocn, and stimulated mineralization by increasing Runx2 expression at both the mRNA and protein levels. In contrast, knockdown of KLF2 produced the opposite effects. Importantly, we found that KLF2 could physically interact with Runx2. KLF2 promoted osteoblast differentiation by regulating Runx2 and physically interacting with Runx2. Taken together, the findings of this study identify KLF2 as a novel regulator of osteoblast differentiation. Our findings suggest that KLF2 might be a new therapeutic target for bone disease.
Topics: Animals; Cell Differentiation; Cell Line; Core Binding Factor Alpha 1 Subunit; Gene Knockdown Techniques; Human Umbilical Vein Endothelial Cells; Humans; Kruppel-Like Transcription Factors; Mice; Osteoblasts
PubMed: 30429507
DOI: 10.1038/s41374-018-0149-x -
Arbutin ameliorates glucocorticoid-induced osteoporosis through activating autophagy in osteoblasts.Experimental Biology and Medicine... Jul 2021Chronic long-term glucocorticoid use causes osteoporosis partly by interrupting osteoblast homeostasis and exacerbating bone loss. Arbutin, a natural hydroquinone...
Chronic long-term glucocorticoid use causes osteoporosis partly by interrupting osteoblast homeostasis and exacerbating bone loss. Arbutin, a natural hydroquinone glycoside, has been reported to have biological activities related to the differentiation of osteoblasts and osteoclasts. However, the role and underlying mechanism of arbutin in glucocorticoid-induced osteoporosis are elusive. In this study, we demonstrated that arbutin administration ameliorated osteoporotic disorders in glucocorticoid dexamethasone (Dex)-induced mouse model, including attenuating the loss of bone mass and trabecular microstructure, promoting bone formation, suppressing bone resorption, and activating autophagy in bone tissues. Furthermore, Dex-stimulated mouse osteoblastic MC3T3-E1 cells were treated with arbutin. Arbutin treatment rescued Dex-induced repression of osteoblast differentiation and mineralization, the downregulation of osteogenic gene expression, reduced autophagic marker expression, and decreased autophagic puncta formation. The application of autophagy inhibitor 3-MA decreased autophagy, differentiation, and mineralization of MC3T3-E1 cells triggered by arbutin. Taken together, our findings suggest that arbutin treatment fends off glucocorticoid-induced osteoporosis, partly through promoting differentiation and mineralization of osteoblasts by autophagy activation.
Topics: Animals; Arbutin; Autophagy; Cell Line; Dexamethasone; Glucocorticoids; Male; Mice; Mice, Inbred C57BL; Osteoblasts; Osteoporosis
PubMed: 33757338
DOI: 10.1177/15353702211002136 -
Biological Trace Element Research Feb 2022Little attention has been paid to the tolerance of osteoblasts to fluoride in distinct differentiation stages, and the role of TGF-β1 in fluoride-treated osteoblast...
Little attention has been paid to the tolerance of osteoblasts to fluoride in distinct differentiation stages, and the role of TGF-β1 in fluoride-treated osteoblast differentiation of progenitors and precursors was rarely mentioned in previous studies. The present study aimed to clarify how fluoride affected different differentiation stages of osteoblasts, and to elucidate the role of TGF-β1 in this process. We assessed cell migration, proliferation, DNA damage, and apoptosis of early-differentiated osteoblasts derived from bone marrow stem cells (BMSCs) exposed to fluoride with or without TGF-β1. Subsequently, MC3T3-E1 cells cultured with mineral induction medium were treated with fluoride to test fluoride's effect on late-differentiated osteoblasts. The specific fluoride concentrations and treatment times were chosen to evaluate the role of TGF-β1 in fluoride-induced osteoblastic differentiation and function. Results showed early-differentiated osteoblasts treated with a low dose of fluoride grew and moved more rapidly. TGF-β1 promoted cell proliferation and inhibited cell apoptosis in early-differentiated osteoblasts exposed to a low fluoride dose, but enhanced apoptosis at higher fluoride conditions. In the late-differentiated osteoblasts, the fluorine dose range with anabolic effects was narrowed, and the fluoride range with catabolic effects was widened. Treatment with a low fluoride dose stimulated the alkaline phosphatase (ALP) expression. TGF-β1 treatment inhibited Runx2 expression but increased RANKL expression in late-differentiated osteoblasts exposed to fluoride. Meanwhile, TGF-β1 treatments activated Smad3 phosphorylation but blocked Wnt10b expression in osteoblasts. We conclude that TGF-β1 plays an essential role in fluoride-induced differentiation and osteoblast function via activation of Smad3 instead of Wnt10 signaling.
Topics: 3T3 Cells; Animals; Cell Differentiation; Fluorides; Mice; Osteoblasts; Osteogenesis; Signal Transduction; Transforming Growth Factor beta1
PubMed: 34031801
DOI: 10.1007/s12011-021-02686-2 -
The FEBS Journal Oct 2020Gene expression in extant animals might reveal how skeletal cells have evolved over the past 500 million years. The cells that make up cartilage (chondrocytes) and bone... (Review)
Review
Gene expression in extant animals might reveal how skeletal cells have evolved over the past 500 million years. The cells that make up cartilage (chondrocytes) and bone (osteoblasts) express many of the same genes, but they also have important molecular differences that allow us to distinguish them as separate cell types. For example, traditional studies of later-diverged vertebrates, such as mouse and chick, defined the genes Col2a1 and sex-determining region Y-box 9 as cartilage-specific. However, recent studies have shown that osteoblasts of earlier-diverged vertebrates, such as frog, gar, and zebrafish, express these 'chondrogenic' markers. In this review, we examine the resulting hypothesis that chondrogenic gene expression became repressed in osteoblasts over evolutionary time. The amphibian is an underexplored skeletal model that is uniquely positioned to address this hypothesis, especially given that it diverged when life transitioned from water to land. Given the relationship between phylogeny and ontogeny, a novel discovery for skeletal cell evolution might bolster our understanding of skeletal cell development.
Topics: Animals; Chondrogenesis; Osteoblasts
PubMed: 31994313
DOI: 10.1111/febs.15228 -
Cells Sep 2021The complex multidimensional skeletal organization can adapt its structure in accordance with external contexts, demonstrating excellent self-renewal capacity. Thus,... (Review)
Review
The complex multidimensional skeletal organization can adapt its structure in accordance with external contexts, demonstrating excellent self-renewal capacity. Thus, optimal extracellular environmental properties are critical for bone regeneration and inextricably linked to the mechanical and biological states of bone. It is interesting to note that the microstructure of bone depends not only on genetic determinants (which control the bone remodeling loop through autocrine and paracrine signals) but also, more importantly, on the continuous response of cells to external mechanical cues. In particular, bone cells sense mechanical signals such as shear, tensile, loading and vibration, and once activated, they react by regulating bone anabolism. Although several specific surrounding conditions needed for osteoblast cells to specifically augment bone formation have been empirically discovered, most of the underlying biomechanical cellular processes underneath remain largely unknown. Nevertheless, exogenous stimuli of endogenous osteogenesis can be applied to promote the mineral apposition rate, bone formation, bone mass and bone strength, as well as expediting fracture repair and bone regeneration. The following review summarizes the latest studies related to the proliferation and differentiation of osteoblastic cells, enhanced by mechanical forces or supplemental signaling factors (such as trace metals, nutraceuticals, vitamins and exosomes), providing a thorough overview of the exogenous osteogenic agents which can be exploited to modulate and influence the mechanically induced anabolism of bone. Furthermore, this review aims to discuss the emerging role of extracellular stimuli in skeletal metabolism as well as their potential roles and provide new perspectives for the treatment of bone disorders.
Topics: Anabolic Agents; Animals; Bone Regeneration; Cell Differentiation; Humans; Mechanotransduction, Cellular; Osteoblasts; Signal Transduction
PubMed: 34572032
DOI: 10.3390/cells10092383 -
Molecular and Cellular Biology Apr 2015Congenital osteopenia is a bone demineralization condition that is associated with elevated fracture risk in human infants. Here we show that Runx3, like Runx2, is...
Congenital osteopenia is a bone demineralization condition that is associated with elevated fracture risk in human infants. Here we show that Runx3, like Runx2, is expressed in precommitted embryonic osteoblasts and that Runx3-deficient mice develop severe congenital osteopenia. Runx3-deficient osteoblast-specific (Runx3(fl/fl)/Col1α1-cre), but not chondrocyte-specific (Runx3(fl/fl)/Col1α2-cre), mice are osteopenic. This demonstrates that an osteoblastic cell-autonomous function of Runx3 is required for proper osteogenesis. Bone histomorphometry revealed that decreased osteoblast numbers and reduced mineral deposition capacity in Runx3-deficient mice cause this bone formation deficiency. Neonatal bone and cultured primary osteoblast analyses revealed a Runx3-deficiency-associated decrease in the number of active osteoblasts resulting from diminished proliferation and not from enhanced osteoblast apoptosis. These findings are supported by Runx3-null culture transcriptome analyses showing significant decreases in the levels of osteoblastic markers and increases in the levels of Notch signaling components. Thus, while Runx2 is mandatory for the osteoblastic lineage commitment, Runx3 is nonredundantly required for the proliferation of these precommitted cells, to generate adequate numbers of active osteoblasts. Human RUNX3 resides on chromosome 1p36, a region that is associated with osteoporosis. Therefore, RUNX3 might also be involved in human bone mineralization.
Topics: Animals; Apoptosis; Bone Development; Bone Diseases, Metabolic; Bone and Bones; Cells, Cultured; Core Binding Factor Alpha 3 Subunit; Gene Deletion; Gene Expression Regulation, Developmental; Humans; Mice; Mice, Knockout; Osteoblasts; Osteogenesis; Transcriptome
PubMed: 25605327
DOI: 10.1128/MCB.01106-14 -
Journal of Bone and Mineral Metabolism May 2018Ankylosing spondylitis (AS) is characterized by excessive bone formation with syndesmophytes, leading to bony ankylosis. The contribution of osteoblasts to the...
Ankylosing spondylitis (AS) is characterized by excessive bone formation with syndesmophytes, leading to bony ankylosis. The contribution of osteoblasts to the pathogenesis of ankylosis is poorly understood. The aim of this study was to determine molecular differences between disease controls (Ct) and AS bone-derived cells (BdCs) during osteogenic differentiation with or without inflammation using AS patient serum. We confirmed osteoblastic differentiation of Ct and AS BdCs under osteogenic medium by observing morphological changes and measuring osteoblastic differentiation markers. Osteoblast differentiation was detected by alkaline phosphatase (ALP) staining and activity, and alizarin red and hydroxyapatite staining. Osteoblast-specific markers were analyzed by quantitative reverse-transcriptase-polymerase chain reaction, immunoblotting, and immunostaining. To examine the effects of inflammation, we added AS and healthy control serum to Ct and AS BdCs, and then analyzed osteoblast-specific markers. AS BdCs showed elevated basal intercellular and extracellular ALP activity compared to Ct. When osteoblast differentiation was induced, AS BdCs exhibited higher expression of osteoblast-specific marker genes and faster mineralization than Ct, indicating that these cells differentiated more rapidly into osteoblasts. ALP activity and mineralization accelerated when serum from AS patients was added to Ct and AS BdCs. Our results revealed that AS BdCs showed significantly increased osteoblastic activity and differentiation capacity by regulating osteoblast-specific transcription factors and proteins compared to Ct BdCs. Active inflammation of AS serum accelerated osteoblastic activity. Our study could provide useful basic data for understanding the molecular mechanism of ankylosis in AS.
Topics: Adult; Bone and Bones; Cell Differentiation; Cells, Cultured; Humans; Male; Osteoblasts; Osteogenesis; Spondylitis, Ankylosing
PubMed: 28589411
DOI: 10.1007/s00774-017-0846-3 -
Biochemical and Biophysical Research... Jan 2018This study was designed to identify and characterize primary bone-derived cells (BdCs) and investigate the potential role of osteoblast differentiation. Primary BdCs...
This study was designed to identify and characterize primary bone-derived cells (BdCs) and investigate the potential role of osteoblast differentiation. Primary BdCs were isolated from surgical bone for comparative analysis with mesenchymal stem cells (MSCs) and fetal osteoblasts (FOBs) and for potential differentiation to mature osteoblasts. Using three different cells, we successfully cultivated human osteoblast differentiation and activity which were evaluated using microarray and biochemical methods. BdCs are more correlated to MSCs in bioinformatics result and similar with FOBs in gene expression. In particular, Osterix, osteoprogenitor marker, was high expressed in BdCs, while the expression in MSCs and FOBs were very low. Furthermore, BdCs exhibited a marked alkaline phosphatase (ALP) expression, early stage of osteogenic marker, and retained osteogenic properties and physiological changes into maturation as in FOBs. BdCs also showed an increase in bone morphogenic protein 2 (BMP2), osteopontin (OPN), and osteocalcin (OCN) mRNA expressions during differentiation. This study suggests that BdCs may be osteoprogenitor cells or undifferentiated preosteoblasts with strong capacity to differentiate toward mature osteoblasts.
Topics: Cell Differentiation; Cells, Cultured; Humans; Mesenchymal Stem Cells; Osteoblasts; Proteome
PubMed: 29180008
DOI: 10.1016/j.bbrc.2017.11.155 -
Frontiers in Bioscience (Landmark... Oct 2022Recently, single-cell RNA sequencing (scRNA-seq) technology was increasingly used to study transcriptomics at a single-cell resolution, scRNA-seq analysis was...
BACKGROUND
Recently, single-cell RNA sequencing (scRNA-seq) technology was increasingly used to study transcriptomics at a single-cell resolution, scRNA-seq analysis was complicated by the "dropout", where the data only captures a small fraction of the transcriptome. This phenomenon can lead to the fact that the actual expressed transcript may not be detected. We previously performed osteoblast subtypes classification and dissection on freshly isolated human osteoblasts.
MATERIALS AND METHODS
Here, we used the scImpute method to impute the missing values of dropout genes from a scRNA-seq dataset generated on freshly isolated human osteoblasts.
RESULTS
Based on the imputed gene expression patterns, we discovered three new osteoblast subtypes. Specifically, these newfound osteoblast subtypes are osteoblast progenitors, and two undetermined osteoblasts. Osteoblast progenitors showed significantly high expression of proliferation related genes ( and ). Analysis of each subtype showed that in addition to bone formation, these undetermined osteoblasts may involve osteoclast and adipocyte differentiation and have the potential function of regulate immune activation.
CONCLUSIONS
Our findings provided a new perspective for studying the osteoblast heterogeneity and potential biological functions of these freshly isolated human osteoblasts at the single-cell level, which provides further insight into osteoblasts subtypes under various (pathological) physiological conditions.
Topics: Humans; RNA-Seq; Osteoblasts; Transcriptome; Cell Differentiation; Osteogenesis; Gene Expression Profiling
PubMed: 36336853
DOI: 10.31083/j.fbl2710295