Bone Formation Function Research Articles

The multifaceted roles of mitochondria in osteoblasts: from energy production to mitochondrial-derived vesicle secretion.

Mitochondria in osteoblasts have been demonstrated to play multiple crucial functions in bone formation from intracellular adenosine triphosphate production to extracellular secretion of mitochondrial components. The present review explores the current knowledge about mitochondrial biology in osteoblasts, including mitochondrial biogenesis, bioenergetics, oxidative stress generation, and dynamic changes in morphology. Special attention is given to recent findings, including mitochondrial donut formation in osteoblasts, which actively generates mitochondrial-derived vesicles (MDVs), followed by extracellular secretion of small mitochondria and MDVs. We also discuss the therapeutic effects of targeting osteoblast mitochondria, highlighting their potential applications in improving bone health.

Fibrous periosteum repairs bone fracture and maintains the healed bone throughout mouse adulthood

Bone is regarded as one of few tissues that heals without fibrous scar. The outer layer of the periosteum is covered with fibrous tissue, whose function in bone formation is unknown. We herein developed a system to distinguish the fate of fibrous-layer periosteal cells (FL-PCs) from the skeletal stem/progenitor cells (SSPCs) in the cambium-layer periosteum and bone marrow in mice. We showed that FL-PCs did not participate in steady-state osteogenesis, but formed the main body of fibrocartilaginous callus during fracture healing. Moreover, FL-PCs invaded the cambium-layer periosteum and bone marrow after fracture, forming neo-SSPCs that continued to maintain the healed bones throughout adulthood. The FL-PC-derived neo-SSPCs expressed lower levels of osteogenic signature genes and displayed lower osteogenic differentiation activity than the preexisting SSPCs. Consistent with this, healed bones were thinner and formed more slowly than normal bones. Thus, the fibrous periosteum becomes the cellular origin of bones after fracture and alters bone properties permanently.

The genetic basis of the leafy seadragon's unique camouflage morphology and avenues for its efficient conservation derived from habitat modeling.

The leafy seadragon certainly is among evolution's most "beautiful and wonderful" species aptly named for its extraordinary camouflage mimicking its coastal seaweed habitat. However, limited information is known about the genetic basis of its phenotypes and conspicuous camouflage. Here, we revealed genomic signatures of rapid evolution and positive selection in core genes related to its camouflage, which allowed us to predict population dynamics for this species. Comparative genomic analysis revealed that seadragons have the smallest olfactory repertoires among all ray-finned fishes, suggesting adaptations to the highly specialized habitat. Other positively selected and rapidly evolving genes that serve in bone development and coloration are highly expressed in the leaf-like appendages, supporting a recent adaptive shift in camouflage appendage formation. Knock-out of bmp6 results in dysplastic intermuscular bones with a significantly reduced number in zebrafish, implying its important function in bone formation. Global climate change-induced loss of seagrass beds now severely threatens the continued existence of this enigmatic species. The leafy seadragon has a historically small population size likely due to its specific habitat requirements that further exacerbate its vulnerability to climate change. Therefore, taking climate change-induced range shifts into account while developing future protection strategies.

Taohong Siwu Decoction enhances human bone marrow mesenchymal stem cells proliferation, migration and osteogenic differentiation via VEGF-FAK signaling in vitro.

Taohong Siwu Decoction (THSWD) is a conventional traditional Chinese prescription aiming at promoting blood circulation and alleviating blood stasis. It is widely prescribed in instances of ischemic strokes, cardiovascular diseases, osteoporosis and bone fracture. However, its molecular functions in bone formation remain uncharacterized. This study aims to explore the potential effects of THSWD treatment on human bone marrow mesenchymal stem cells (BMSCs) proliferation, osteogenic differentiation, and migration. BMSCs undergo osteogenic, adipogenic, and chondrogenic differentiation to determine cell stemness. BMSCs were treated with low dose (200μg/ml), medium dose (400μg/ml) and high dose (600μg/ml) THSWD. The cell viability was determined by CCK-8 assays, the osteogenic differentiation ability was determined by alizarin red staining and ALP staining, and cell migration was determined by wound healing and transwell assays. The effect of THSWD on the vascular endothelial growth factor (VEGF)/focal adhesion kinase (FAK) pathway was determined by immunoblotting. THSWD time-dependently and dose-dependently promoted BMSC viability. Moreover, THSWD also promoted BMSC osteogenic differentiation and migration. As opposed to THSWD, VEGF receptor inhibitor Bevacizumab suppressed BMSC osteogenic differentiation and migration. In BMSCs that have been co-treated with THSWD and Bevacizumab, THSWD effects on BMSC functions were partially eliminated by Bevacizumab. Moreover, THSWD treatment boosted VEGF content in the supernatant and was conducive to the phosphorylation of FAK and Src, whereas Bevacizumab exerted opposite effects; similarly, Bevacizumab partially abolished THSWD effects on VEGF and FAK (Tyr397) and Src (Tyr418) phosphorylation. THSWD enhances the capacities of BMSCs to proliferate, differentiate, and migrate, possibly through VEGF and the FAK-Src, thereby improving fracture healing.

Physicochemical properties of bone marrow mesenchymal stem cells encapsulated in microcapsules combined with calcium phosphate cement and their ectopic bone formation.

Calcium phosphate bone cement (CPC) serves as an excellent scaffold material for bone tissue engineering owing to its good biocompatibility, injectability, self-setting property and three-dimensional porous structure. However, its clinical use is limited due to the cytotoxic effect of its setting reaction on cells and difficulties in degradation into bone. In this study, bone marrow mesenchymal stem cells (BMSCs) were encapsulated in alginate chitosan alginate (ACA) microcapsules and compounded with calcium phosphate bone cement. Changes in the compressive strength, porosity, injectability and collapsibility of CPC at different volume ratios of microcapsules were evaluated. At a 40% volume ratio of microcapsules, the composite scaffold displayed high porosity and injectability with good collapsibility and compressive strength. Cell live/dead double staining, Cell Counting Kit-8 (CCK-8) assays and scanning electron microscopy were used to detect the viability, proliferation and adhesion of cells after cell microcapsules were combined with CPC. The results revealed that cells protected by microcapsules proliferated and adhered better than those that were directly combined with CPC paste, and cell microcapsules could effectively form macropores in scaffold material. The composite was subsequently implanted subcutaneously on the backs of nude mice, and ectopic osteogenesis of the scaffold was detected via haematoxylin-eosin (H&E), Masson’s trichrome and Goldner’s trichrome staining. CPC clearly displayed better new bone formation function and degradability after addition of pure microcapsules and cell microcapsules. Furthermore, the cell microcapsule treatment group showed greater osteogenesis than the pure microcapsule group. Collectively, these results indicate that BMSCs encapsulated in ACA microcapsules combined with CPC composite scaffolds have good application prospects as bone tissue engineering materials.

Effects of Mechanical Stress Stimulation on Function and Expression Mechanism of Osteoblasts.

Osteoclasts and osteoblasts play a major role in bone tissue homeostasis. The homeostasis and integrity of bone tissue are maintained by ensuring a balance between osteoclastic and osteogenic activities. The remodeling of bone tissue is a continuous ongoing process. Osteoclasts mainly play a role in bone resorption, whereas osteoblasts are mainly involved in bone remodeling processes, such as bone cell formation, mineralization, and secretion. These cell types balance and restrict each other to maintain bone tissue metabolism. Bone tissue is very sensitive to mechanical stress stimulation. Unloading and loading of mechanical stress are closely related to the differentiation and formation of osteoclasts and bone resorption function as well as the differentiation and formation of osteoblasts and bone formation function. Consequently, mechanical stress exerts an important influence on the bone microenvironment and bone metabolism. This review focuses on the effects of different forms of mechanical stress stimulation (including gravity, continuously compressive pressure, tensile strain, and fluid shear stress) on osteoclast and osteoblast function and expression mechanism. This article highlights the involvement of osteoclasts and osteoblasts in activating different mechanical transduction pathways and reports changings in their differentiation, formation, and functional mechanism induced by the application of different types of mechanical stress to bone tissue. This review could provide new ideas for further microscopic studies of bone health, disease, and tissue damage reconstruction.

Runt-related transcription factor 1 is required for murine osteoblast differentiation and bone formation

Despite years of research investigating osteoblast differentiation, the mechanisms by which transcription factors regulate osteoblast maturation, bone formation, and bone homeostasis is still unclear. It has been reported that runt-related transcription factor 1 (Runx1) is expressed in osteoblast progenitors, pre-osteoblasts, and mature osteoblasts; yet, surprisingly, the exact function of RUNX1 in osteoblast maturation and bone formation remains unknown. Here, we generated and characterized a pre-osteoblast and differentiating chondrocyte-specific Runx1 conditional knockout mouse model to study RUNX1's function in bone formation. Runx1 ablation in osteoblast precursors and differentiating chondrocytes via osterix-Cre (Osx-Cre) resulted in an osteoporotic phenotype and decreased bone density in the long bones and skulls of Runx1f/fOsx-Cre mice compared with Runx1f/f and Osx-Cre mice. RUNX1 deficiency reduced the expression of SRY-box transcription factor 9 (SOX9), Indian hedgehog signaling molecule (IHH), Patched (PTC), and cyclin D1 in the growth plate, and also reduced the expression of osteocalcin (OCN), OSX, activating transcription factor 4 (ATF4), and RUNX2 in osteoblasts. ChIP assays and promoter activity mapping revealed that RUNX1 directly associates with the Runx2 gene promoter and up-regulates Runx2 expression. Furthermore, the ChIP data also showed that RUNX1 associates with the Ocn promoter. In conclusion, RUNX1 up-regulates the expression of Runx2 and multiple bone-specific genes, and plays an indispensable role in bone formation and homeostasis in both trabecular and cortical bone. We propose that stimulating Runx1 activity may be useful in therapeutic approaches for managing some bone diseases such as osteoporosis.

Hedgehog Inhibitors Suppress Osteoclastogenesis in In Vitro Cultures, and Deletion of Smo in Macrophage/Osteoclast Lineage Prevents Age-Related Bone Loss.

The functional role of the Hedgehog (Hh)-signaling pathway has been widely investigated in bone physiology/development. Previous studies have, however, focused primarily on Hh functions in bone formation, while its roles in bone resorption have not been fully elucidated. Here, we found that cyclopamine (smoothened (Smo) inhibitor), GANT-58 (GLI1 inhibitor), or GANT-61 (GLI1/2 inhibitor) significantly inhibited RANKL-induced osteoclast differentiation of bone marrow-derived macrophages. Although the inhibitory effects were exerted by cyclopamine or GANT-61 treatment during 0–48 h (early stage of osteoclast differentiation) or 48–96 h (late stage of osteoclast differentiation) after RANKL stimulation, GANT-58 suppressed osteoclast formation only during the early stage. These results suggest that the Smo-GLI1/2 axis mediates the whole process of osteoclastogenesis and that GLI1 activation is requisite only during early cellular events of osteoclastogenesis. Additionally, macrophage/osteoclast-specific deletion of Smo in mice was found to attenuate the aging phenotype characterized by trabecular low bone mass, suggesting that blockage of the Hh-signaling pathway in the osteoclast lineage plays a protective role against age-related bone loss. Our findings reveal a specific role of the Hh-signaling pathway in bone resorption and highlight that its inhibitors show potential as therapeutic agents that block osteoclast formation in the treatment of senile osteoporosis.

Berberine Promotes Osteogenic Differentiation of Human Dental Pulp Stem Cells Through Activating EGFR-MAPK-Runx2 Pathways.

Similar to the mesenchymal stem cells (MSCs), dental pulp stem cells (DPSCs) also have pluripotent differentiation characteristic and may be more ideal for tissue regeneration, especially in tooth regeneration engineering. However, bacterial infection may be a powerful obstacle. Berberine (BBR), known with antibacterial effects, was recently found to play functions in bone formation through promoting osteogenic differentiation from pluripotent stem cells. However, whether BBR also function in DPSCs osteogenic differentiation has not yet been reported. Primary DPSCs were isolated from dental pulp tissues extracted from human impacted mandibular third molars, and identified by flow cytometry for cell surface antigen molecules. A dexamethasone osteogenic medium was used to induce DPSCs osteogenic differentiation. BBR (1μM and 5μM) was pre-added to into medium, and then cell proliferation, spheroid formation and osteogenic differentiation capacities of DPSCs were analyzed, as well as the underlying molecules modulation mechanism. Flow cytometry identified that CD44, CD90, CD81 and CD105 positively expressed in the isolated hDPSCs, with CD34 and CD45 negetively expressed. BBR enhanced the cell proliferation of hDPSCs in a dose-dependent pattern, and promoted dexamethasone-induced osteogenic differentiation via enhancing Runx2 transcription factor activity followed by upregulating osteogenesis markers expression, whereas the adipogenic differentiation of hDPSCs was suppressed dramatically by BBR. The EGFR and MAPK pathways were activated by BBR, and inhibitors for these pathways significantly suppressed the osteogenic differentiation promotion of BBR. These results have revealed a novel mechanism that berberine might promote hDPSCs osteogenic differentiation through activating EGFR-MAPK-Runx2 signaling pathways.

Micrornas at the Interface between Osteogenesis and Angiogenesis as Targets for Bone Regeneration.

Bone formation and regeneration is a multistep complex process crucially determined by the formation of blood vessels in the growth plate region. This is preceded by the expression of growth factors, notably the vascular endothelial growth factor (VEGF), secreted by osteogenic cells, as well as the corresponding response of endothelial cells, although the exact mechanisms remain to be clarified. Thereby, coordinated coupling between osteogenesis and angiogenesis is initiated and sustained. The precise interplay of these two fundamental processes is crucial during times of rapid bone growth or fracture repair in adults. Deviations in this balance might lead to pathologic conditions such as osteoarthritis and ectopic bone formation. Besides VEGF, the recently discovered important regulatory and modifying functions of microRNAs also support this key mechanism. These comprise two principal categories of microRNAs that were identified with specific functions in bone formation (osteomiRs) and/or angiogenesis (angiomiRs). However, as hypoxia is a major driving force behind bone angiogenesis, a third group involved in this process is represented by hypoxia-inducible microRNAs (hypoxamiRs). This review was focused on the identification of microRNAs that were found to have an active role in osteogenesis as well as angiogenesis to date that were termed “CouplingmiRs (CPLGmiRs)”. Outlined representatives therefore represent microRNAs that already have been associated with an active role in osteogenic-angiogenic coupling or are presumed to have its potential. Elucidation of the molecular mechanisms governing bone angiogenesis are of great relevance for improving therapeutic options in bone regeneration, tissue-engineering, and the treatment of bone-related diseases.

A novel mutation in CDH11, encoding cadherin-11, cause Branchioskeletogenital (Elsahy-Waters) syndrome.

Cadherins are cell-adhesion molecules that control morphogenesis, cell migration, and cell shape changes during multiple developmental processes. Until now four distinct cadherins have been implicated in human Mendelian disorders, mainly featuring skin, retinal and hearing manifestations. Branchio-skeleto-genital (or Elsahy-Waters) syndrome (BSGS) is an ultra-rare condition featuring a characteristic face, premature loss of teeth, vertebral and genital anomalies, and intellectual disability. We have studied two sibs with BSGS originally described by Castori et al. in 2010. Exome sequencing led to the identification of a novel homozygous nonsense variant in the first exon of the cadherin-11 gene (CDH11), which results in a prematurely truncated form of the protein. Recessive variants in CDH11 have been recently demonstrated in two other sporadic patients and a pair of sisters affected by BSGS. Although the function of this cadherin (also termed Osteoblast-Cadherin) is not completely understood, its prevalent expression in osteoblastic cell lines and up-regulation during differentiation suggest a specific function in bone formation and development. This study identifies a novel loss-of-function variant in CDH11 as a cause of BSGS and supports the role of cadherin-11 as a key player in axial and craniofacial malformations.

Low-intensity pulsed ultrasound promotes spinal fusion and enhances migration and proliferation of MG63s through sonic hedgehog signaling pathway.

Low-intensity pulsed ultrasound (LIPUS) has been found to accelerate the healing process of spinal fusion via a process closely related to osteoblast differentiation and migration. Sonic hedgehog (Shh) signaling plays an important role in development and homeostasis, including a critical function in bone formation. However, its role in spinal fusion during LIPUS treatment is still unknown. This study showed that LIPUS treatment after spinal fusion surgery increased bone formation. The increased bone mass under LIPUS treatment appeared to result from the increased migration and proliferation of osteoblasts, resulting from upregulation of the Shh signaling pathway. In contrast, inhibition of Shh reduced the migratory and proliferative ability of osteoblast-like MG63 cells and blocked the efficacy of LIPUS treatment.

Analysis of Ca2+ response of osteocyte network by three-dimensional time-lapse imaging in living bone.

Osteocytes form a three-dimensional (3D) cellular network within the mineralized bone matrix. The cellular network has important roles in mechanosensation and mechanotransduction related to bone homeostasis. We visualized the embedded osteocyte network in chick calvariae and observed the flow-induced Ca2+ signaling in osteocytes using 3D time-lapse imaging. In response to the flow, intracellular Ca2+ ([Ca2+]i) significantly increased in developmentally mature osteocytes in comparison with young osteocytes in the bone matrix. To investigate the differences in response between young and developmentally mature osteocytes in detail, we evaluated the expression of osteocyte-related genes using the osteocyte-like cell line MLO-Y4, which was 3D-cultured within type I collagen gels. We found that the c-Fos, Cx43, Panx3, Col1a1, and OCN mRNA levels significantly increased on day 15 in comparison with day 7. These findings indicate that developmentally mature osteocytes are more responsive to mechanical stress than young osteocytes and have important functions in bone formation and remodeling.

P2X7Rs are involved in cell death, growth and cellular signaling in primary human osteoblasts

The ionotropic ATP-gated P2X7 receptor (P2X7R) is involved in the regulation of many physiological functions including bone metabolism. Several studies on osteoblasts from rodents and human osteoblast-like cell lines have addressed the expression and function of P2X7R on these bone-forming cells however; its role in human primary osteoblasts has not yet been reported. The aim of this study was to assess the expression of the P2X7R in bone marrow-derived stromal cells and in primary human trabecular osteoblasts and to determine the function in bone formation and cell signaling. We report that osteoblasts derived from human trabecular explants express a functional P2X7R capable of agonist-induced increase in intracellular calcium concentration and a positive permeability to fluorescent dyes. These osteoblasts are fully differentiated cells with alkaline phosphatase activity and the ability to form mineralized nodules. We show that the transcriptional regulation of osteoblastic markers can be modulated by P2X7R activity or blockade thereby influencing the differentiation, proliferation and bone matrix formation by these primary human osteoblasts. Finally, we demonstrate that the P2X7R is involved in propagation of mechanically-induced intercellular signaling in addition to the known mechanisms involving calcium signaling via P2Y2 receptors and gap junction.

MiR-106b-5p and miR-17-5p suppress osteogenic differentiation by targeting Smad5 and inhibit bone formation

Osteogenesis is a complex process which relies on the coordination of signals and transcription factors. BMP-2/Smad5 signal transduction pathway plays an important role in the process. Recent evidence indicates that microRNAs (miRNAs) act as important post-transcriptional regulators in a large number of biological processes including osteoblast differentiation. In this study, we investigated the effect of miR-106b-5p and miR-17-5p on osteogenic differentiation. We observed an obvious decreasement in miR-106b-5p and miR-17-5p levels during osteogenic differentiation. By using gain- and loss-of function experiments, we noticed that miR-106b-5p and miR-17-5p could suppress the osteogenic differentiation of C2C12 and MC3T3-E1 cells. In addition, we performed bioinformatic analysis, western blot and luciferase reporter assay to confirm that miR-106b-5p and miR-17-5p could regulate Smad5 expression negatively. When we inhibited Smad5 expression by siRNA, the effects of miR-106b-5p and miR-17-5p inhibition on osteogenesis of C2C12 cells could be significantly reversed by Smad5 RNA interference. Furthermore, silencing of miR-106b-5p and miR-17-5p in sham and ovariectomized (OVX) mice increased bone formation and bone mass, resulting in an improvement of trabecular microarchitecture. Taken together, these data showed that miR-106b-5p and miR-17-5p are novel Smad5 regulators, and they have a crucially physiological function in bone formation and osteoblast differentiation.

Osteomodulin regulates diameter and alters shape of collagen fibrils

Osteomodulin (OMD) is a member of the small leucine-rich repeat proteoglycan family, which is involved in the organization of the extracellular matrix. OMD is located in bone tissue and is reportedly important for bone mineralization. However, the details of OMD function in bone formation are poorly understood. Using the baculovirus expression system, we produced recombinant human OMD and analyzed its interaction with type I collagen, which is abundant in bone. In this result, OMD directly interacted with purified immobilized collagen and OMD suppressed collagen fibril formation in a turbidity assay. Morphological analysis of collagen in the presence or absence of OMD demonstrated that OMD reduces the diameter and changes the shape of collagen fibrils. We conclude that OMD regulates the extracellular matrix during bone formation.

Proteomics Analysis of the Zebrafish Skeletal Extracellular Matrix

The extracellular matrix of the immature and mature skeleton is key to the development and function of the skeletal system. Notwithstanding its importance, it has been technically challenging to obtain a comprehensive picture of the changes in skeletal composition throughout the development of bone and cartilage. In this study, we analyzed the extracellular protein composition of the zebrafish skeleton using a mass spectrometry-based approach, resulting in the identification of 262 extracellular proteins, including most of the bone and cartilage specific proteins previously reported in mammalian species. By comparing these extracellular proteins at larval, juvenile, and adult developmental stages, 123 proteins were found that differed significantly in abundance during development. Proteins with a reported function in bone formation increased in abundance during zebrafish development, while analysis of the cartilage matrix revealed major compositional changes during development. The protein list includes ligands and inhibitors of various signaling pathways implicated in skeletogenesis such as the Int/Wingless as well as the insulin-like growth factor signaling pathways. This first proteomic analysis of zebrafish skeletal development reveals that the zebrafish skeleton is comparable with the skeleton of other vertebrate species including mammals. In addition, our study reveals 6 novel proteins that have never been related to vertebrate skeletogenesis and shows a surprisingly large number of differences in the cartilage and bone proteome between the head, axis and caudal fin regions. Our study provides the first systematic assessment of bone and cartilage protein composition in an entire vertebrate at different stages of development.

Genotype–phenotype study in type V osteogenesis imperfecta

Type V osteogenesis imperfecta (OI) presents with moderate-to-severe skeletal deformity and is characterized by hyperplastic callus formation at fracture sites and calcification of the interosseous membranes of the forearm and lower leg. The facial dysmorphism is not well characterized and has not been described in previous reports. Inheritance is autosomal dominant, although the genetic aetiology remained unknown until very recently. The aims of this study were to establish the genetic aetiology in patients with type V OI and further characterize patients with this condition, and to ascertain whether they have a similar clinical phenotype and facial dysmorphism. Three families (one mother-daughter pair and two singletons) were identified with the above features and further investigations (molecular genetic analysis and skin biopsy including electron microscopy, histology and collagen species analysis) were performed. Accurate phenotyping of patients with type V OI was performed. PCR amplification was performed using the Sheffield Diagnostic Genetics Service pyrosequencing assay for the interferon-induced transmembrane protein-5 (IFITM5) gene. All the patients had been confirmed to have a heterozygous variant, c.[-14C>T];[=], in the 5'-UTR of the IFITM5 gene, which is located in the transcribed region of this gene. This recurrent mutation, in IFITM5, also known as bone-restricted interferon-induced transmembrane protein-like protein or BRIL, encodes a protein with a function in bone formation and plays an important role in osteoblast formation. All four patients in this study appear to have similar clinical features and facial dysmorphism, including a short, up-turned nose, a small mouth, a prominent chin and greyish-blue sclerae. Skin biopsy in one patient showed clumping of elastic fibres and normal biochemical analysis of collagen. We have been able to characterize patients with type V OI further and confirm the genetic aetiology in this distinct form of OI. Accurate phenotyping (including describing the common facial features) before investigations is important to enable the useful interpretation of genetic results and/or target-specific gene testing.

The role of extracellular modulators of canonical Wnt signaling in bone metabolism and diseases

ObjectivesThe Wnt signaling pathway is a key pathway in various processes, including bone metabolism. In this review, current knowledge of all extracellular modulators of the canonical Wnt signaling in bone metabolism is summarized and discussed. MethodsThe PubMed database was searched using the following keywords: canonical Wnt signaling, β-catenin bone metabolism, BMD, osteoblast, osteoporosis, Wnt, LRPs, Frizzleds, sFRPs, sclerostin or SOST, dickkopfs, Wif1, R-spondins, glypicans, SOST-dc1 and kremen, all separately as well as in different combinations. ResultsCanonical Wnt signaling is considered to be one of the major pathways regulating bone formation. Consequently, a large number of studies were performed to elucidate the role of numerous proteins in canonical Wnt signaling and bone metabolism. These studies led to the identification of novel modulators of the pathway like the R-spondin and glypican protein families. Furthermore novel insights are gained in the regulatory role of the different Wnt proteins. Finally, due to its function in bone formation, the pathway is an interesting target for the development of therapeutics for osteoporosis and other bone diseases. In this review, we discuss the promising results of the Wnt modulators sclerostin, Dkk1 and sFRP1 as targets for osteoporosis treatment. ConclusionThe increasing number of studies into the exact function of all proteins in the canonical Wnt pathway in general and in bone metabolism already led to novel insights in the regulation of the canonical Wnt pathway. In this review we covered the current knowledge of all extracellular modulators of canonical Wnt signaling.

Polymorphisms in bone morphogenetic protein 3 and the risk of papillary thyroid cancer

Bone morphogenetic proteins (BMPs) are members of the transforming growth factor beta (TGFβ) superfamily with well-described functions in bone formation. Although disrupted BMP signaling in tumor development has been investigated, a genetic association for BMP3 in papillary thyroid cancer (PTC) has remained largely unexplored. In this study, we investigated whether BMP3 single nucleotide polymorphisms (SNPs) are associated with the development of PTC and its clinicopathological features. A total of 103 PTC patients and 324 control subjects were enrolled. One promoter SNP (rs13138132; -1919C/A) and one missense mutation (rs3733549; Arg192Gln) in BMP3 were genotyped by direct sequencing. SNPStats, SNPAnalyzer, Helixtree and Haploview version 4.2 were used to evaluate the genetic data. Multiple logistic regression models were used to calculate odds ratios (ORs), 95% confidence intervals (CIs) and P-values. The missense SNP (rs3733549) was weakly associated with the development of PTC in a codominant model (AA vs. GG; P=0.017) and a recessive model (AA vs. GG/GA; P=0.023). Additionally, in an analysis according to clinicopathological features, rs13138132 was significantly associated with extra-thyroidal invasion in a codominant model (CA vs. CC; P=0.006) and a dominant model (CA/AA vs. CC; P=0.0023). We also identified that the frequency of the A allele in the promoter SNP (rs13138132) was increased in PTC patients with extrathyroidal invasion (P=0.004). Our data suggest that rs3733549 in BMP3 is associated with the development of PTC and that the A allele of rs13138932 in BMP3 is a risk factor for extrathyroidal invasion.