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Osteogenic Differentiation (osteogenic + differentiation)
Terms modified by Osteogenic Differentiation Selected AbstractsDysregulated BMP Signaling and Enhanced Osteogenic Differentiation of Connective Tissue Progenitor Cells From Patients With Fibrodysplasia Ossificans Progressiva (FOP),JOURNAL OF BONE AND MINERAL RESEARCH, Issue 3 2008Paul C Billings Abstract The study of FOP, a disabling genetic disorder of progressive heterotopic ossification, is hampered by the lack of readily available connective tissue progenitor cells. We isolated such cells from discarded primary teeth of patients with FOP and controls and discovered dysregulation of BMP signaling and rapid osteoblast differentiation in FOP cells compared with control cells. Introduction: Fibrodysplasia ossificans progressiva (FOP), the most disabling condition of progressive heterotopic ossification in humans, is caused by a recurrent heterozygous missense mutation in activin receptor IA (ACVR1), a bone morphogenetic protein (BMP) type I receptor, in all classically affected individuals. A comprehensive understanding of FOP has been limited, in part, by a lack of readily available connective tissue progenitor cells in which to study the molecular pathology of this disorder. Materials and Methods: We derived connective tissue progenitor cells from discarded primary teeth (SHED cells) of patients with FOP and controls and examined BMP signaling and osteogenic differentiation in these cells. Results: SHED cells transmitted BMP signals through both the SMAD and p38 mitogen-activated protein kinase (MAPK) pathways and responded to BMP4 treatment by inducing BMP responsive genes. FOP cells showed ligand-independent BMP signaling and ligand-dependent hyper-responsiveness to BMP stimulation. Furthermore, FOP cells showed more rapid differentiation to an osteogenic phenotype than control cells. Conclusions: This is the first study of BMP signaling and osteogenic differentiation in connective tissue progenitor cells from patients with FOP. Our data strongly support both basal and ligand-stimulated dysregulation of BMP signaling consistent with in silico studies of the mutant ACVR1 receptor in this condition. This study substantially extends our understanding of dysregulated BMP signaling in a progenitor cell population relevant to the pathogenesis of this catastrophic disorder of progressive ectopic ossification. [source] Ethanol Alters the Osteogenic Differentiation of Amniotic Fluid-Derived Stem CellsALCOHOLISM, Issue 10 2010Jennifer A. Hipp Background:, Fetal alcohol spectrum disorder (FASD) is a set of developmental defects caused by prenatal alcohol exposure. Clinical manifestations of FASD are highly variable and include mental retardation and developmental defects of the heart, kidney, muscle, skeleton, and craniofacial structures. Specific effects of ethanol on fetal cells include induction of apoptosis as well as inhibition of proliferation, differentiation, and migration. This complex set of responses suggests that a bioinformatics approach could clarify some of the pathways involved in these responses. Methods:, In this study, the responses of fetal stem cells derived from the amniotic fluid (AFSCs) to treatment with ethanol have been examined. Large-scale transcriptome analysis of ethanol-treated AFSCs indicates that genes involved in skeletal development and ossification are up-regulated in these cells. Therefore, the effect of ethanol on osteogenic differentiation of AFSCs was studied. Results:, Exposure to ethanol during the first 48 hours of an osteogenic differentiation protocol increased in vitro calcium deposition by AFSCs and increased alkaline phosphatase activity. In contrast, ethanol treatment later in the differentiation protocol (day 8) had no significant effect on the activity of alkaline phosphatase. Conclusions:, These results suggest that transient exposure of AFSCs to ethanol during early differentiation enhances osteogenic differentiation of the cells. [source] Polypyrrole Thin Films Formed by Admicellar Polymerization Support the Osteogenic Differentiation of Mesenchymal Stem CellsMACROMOLECULAR BIOSCIENCE, Issue 8 2004Harold Castano Abstract Summary: The objective of this study was to evaluate the attachment, proliferation, and differentiation of rat mesenchymal stem cells (MSC) toward the osteoblastic phenotype seeded on polypyrrole (PPy) thin films made by admicellar polymerization. Three different concentrations of pyrrole (Py) monomer (20, 35, and 50,×,10,3M) were used with the PPy films deposited on tissue culture polystyrene dishes (TCP). Regular TCP dishes and PPy polymerized on TCP by chemical polymerization without surfactant using 5,×,10,3M Py, were used as controls. Rat MSC were seeded on these surfaces and cultured for up to 20 d in osteogenic media. Surface topography was characterized by atomic force microscopy, X-ray photoelectron spectroscopy, and static contact angle. Cell attachment, proliferation, alkaline phosphatase (ALP) activity, and calcium content were measured to evaluate the ability of MSC to adhere and differentiate on PPy-coated TCP. Increased monomer concentrations resulted in PPy films of increased thickness and surface roughness. PPy films generated by different monomer concentrations induced drastically different cellular events. A wide spectrum of cell attachment characteristics (from excellent cell attachment to the complete inability to adhere) were obtained by varying the monomer concentration from 20 m to 50,×,10,3M. In particular the 20,×,10,3M PPy thin films demonstrated superior induction of MSC osteogenicity, which was comparable to standard TCP dishes, unlike PPy films of similar thickness prepared by chemical polymerization without surfactant. Adhesion of mesenchymal stem cells on tissue culture plates (TCP) coated with polypyrrole thin films made by admicellar polymerization. [source] Periodically Discontinuous Induction of Bone Marrow Stem Cells toward Osteogenic Differentiation in VitroBIOTECHNOLOGY PROGRESS, Issue 3 2008Zhen Wang This paper reveals that a discontinuous in vitro induction, namely, the periodic presence and absence of foreign induction factors, might be, under a certain condition, more effective to stimulate stem cells' differentiation than a continuous induction. Bone marrow stem cells (BMSCs) derived from Sprague Dawley rats were employed to examine the effects of discontinuous additions of osteogenic supplements with a series of alternate frequency in contrast to those with continuous induction or no induction. The results demonstrated that a suitable discontinuous induction was more able to achieve osteogenesis than not only no induction but also the associated continuous induction. Additionally, the osteogenic supplements were confirmed to enhance cell differentiation but suppress cell proliferation. So, the combination of differentiation extent per cell and cell number accounts for the "unexpected" good osteogenic effect of the discontinuous induction. The induction effect was found to be dependent upon alternate frequency, and the optimum alternate period in our experimental systems was determined to be around 4 days. Since it is very common to change culture medium every 2,4 days, such a strategy of discontinuous induction does not bring any extra manual work but reduces the consumption of foreign induction factors and significantly enhances the global differentiation efficacy. Our work thus affords a convenient and practical approach to achieve differentiation of BMSCs, which might be useful for potential large-scale culture and differentiation of stem cells. Meanwhile, the existence of optimum frequency implies some unknown inherent rhythms of cell proliferation and differentiation. [source] Multilineage mesenchymal differentiation potential of human trabecular bone-derived cellsJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 5 2002Ulrich Nöth Abstract Explant cultures of adult human trabecular bone fragments give rise to osteoblastic cells, that are known to express osteoblast-related genes and mineralize extracellular matrix. These osteoblastic cells have also been shown to undergo adipogenesis in vitro and chondrogenesis in vivo. Here we report the in vitro developmental potential of adult human osteoblastic cells (hOB) derived from explant cultures of collagenase-pretreated trabecular bone fragments. In addition to osteogenic and adipogenic differentiation, these cells are capable of chondrogenic differentiation in vitro in a manner similar to adult human bone marrow-derived mesenchymal progenitor cells. High-density pellet cultures of hOB maintained in chemically defined serum-free medium, supplemented with transforming growth factor-,1, were composed of morphologically distinct, chondrocyte-like cells expressing mRNA transcripts of collagen types II, IX and X, and aggrecan. The cells within the high-density pellet cultures were surrounded by a sulfated prote-oglycan-rich extracellular matrix that immunostained for collagen type II and proteoglycan link protein. Osteogenic differentiation of hOB was verified by an increased number of alkaline phosphatase-positive cells, that expressed osteoblast-related transcripts such as alkaline phosphatase, collagen type I, osteopontin and osteocalcin, and formed mineralized matrix in monolayer cultures treated with ascorbate, ,-glycerophosphate, and bone morphogenetic protein-2. Adipogenic differentiation of hOB was determined by the appearance of intracellular lipid droplets, and expression of adipocyte-specific genes, such as lipoprotein lipase and peroxisome proliferator-activated receptor ,2, in monolayer cultures treated with dexamethasone, indomethacin, insulin and 3-isobutyl-l-methylxanthine. Taken together, these results show that cells derived from collagenase-treated adult human trabecular bone fragments have the potential to differentiate into multiple mesenchymal lineages in vitro, indicating their developmental plasticity and suggesting their mesenchymal progenitor nature. © 2002 Orthopaedic Research Society. Published by Elsevier Science Ltd. All rights reserved. [source] Proteomic analysis of osteogenic differentiation of dental follicle precursor cellsELECTROPHORESIS, Issue 7 2009Christian Morsczeck Abstract Recently, there has been an increased interest in unravelling the molecular mechanisms and cellular pathways controlling the differentiation and proliferation of human stem cell lines. Proteome analysis has proven to be an effective approach to comprehensive analysis of the regulatory network of differentiation. In the present study we applied 2-DE combined with capillary-LC-MS/MS analysis to profile differentially regulated proteins upon differentiation of dental follicle precursor cells (DFPCs). Out of 115 differentially regulated proteins, glutamine synthetase, lysosomal proteinase cathepsin B proteins, plastin 3 T-isoform, beta-actin, superoxide dismutases, and transgelin were found to be highly up-regulated, whereas cofilin-1, pro-alpha 1 collagen, destrin, prolyl 4-hydrolase and dihydrolipoamide dehydrogenase were found to be highly down-regulated. The group of up-regulated proteins is associated with actin-bundling and defence against oxidative cellular stress, whereas down-regulated proteins were associated with collagen biosynthesis. Bioinformatic analyses of the entire data set confirmed these findings that represent significant steps towards the understanding of DFPC differentiation. The bioinformatic analyses suggest that proteins associated with cell cycle progression and protein metabolism were down-regulated and proteins involved in catabolism, cell motility and biological quality were up-regulated. These results display the general physiological state of DFPCs before and after osteogenic differentiation. We also identified regulatory proteins, such as the transcription factors TP53 and Sp-1, associated with the differentiation process. Further studies will investigate the impact of identified regulatory proteins for cell proliferation and osteogenic differentiation in DFPCs. [source] The proteasome inhibitor bortezomib inhibits FGF-2-induced reduction of TAZ levels in osteoblast-like cellsEUROPEAN JOURNAL OF HAEMATOLOGY, Issue 1 2010Homare Eda Abstract Objectives:,Bortezomib (PS-341; VelcadeÔ), a proteasome inhibitor, is used as a therapeutic agent for multiple myeloma. Bortezomib has been shown to strongly induce osteoblast differentiation and elevate the levels of osteoblast-related differentiation markers in the serum of patients with myeloma. Bortezomib also reportedly increases the activity of the transcription factor, Runx2. However, the mechanism of action by which bortezomib-elevated Runx2 activity mediates osteoblast differentiation remains unclear. On the other hand, fibroblast growth factor 2 (FGF-2) is found at high levels in patients with multiple myeloma. We previously reported that FGF-2 reduces the levels of the transcriptional coactivator with PDZ-binding motif (TAZ). We therefore investigated the effects of bortezomib on TAZ protein levels in the presence of FGF-2. Methods: Osteoblastic MC3T3-E1 cells were treated with different concentrations of bortezomib in the presence or absence of FGF-2 and various biologic responses were investigated by immunoblotting, RT-PCR, quantitative PCR, and alizarin red staining. Results: We found that bortezomib inhibited FGF-2-induced reduction of TAZ levels through a pathway other than that used for proteasome inhibition, while maintaining TAZ function, which in turn, enhanced the expression of Runx2-transcribed osteogenic differentiation markers. Bortezomib also suppressed the antimineralization effect of FGF-2. Conclusions: These findings suggest that bortezomib inhibited FGF-2-induced reduction of TAZ and consequently stimulated osteogenic differentiation independently of proteasome inhibition. These findings may contribute to elucidate the osteolytic mechanism in multiple myeloma, and to the development of new drugs for multiple myeloma and other osteolytic diseases. [source] Expression of Osterix in mechanical stress-induced osteogenic differentiation of periodontal ligament cells in vitroEUROPEAN JOURNAL OF ORAL SCIENCES, Issue 3 2008Yanhong Zhao Osterix (Osx) is an osteoblast-specific transcription factor required for the differentiation of pre-osteoblasts into functional osteoblasts. This study sought to examine the changes of Osx expression in periodontal ligament cells (PDLC) subjected to mechanical force, and to investigate whether Osx is involved in the mechanical stress-induced differentiation of PDLC. Human PDLC were exposed to centrifugal force for 1,12 h. Real-time polymerase chain reaction (PCR), western blot, and immunofluorescence assays were used to examine the mRNA and protein expression of Osx and its subcellular localization. Furthermore, PDLC were transfected with the expression vector pcDNA3.1 flag-Osx and subjected to mechanical force for 6 h. The changes in alkaline phosphatase (ALP) activity and in the expression of core-binding factor alpha1 (Cbfa1), ALP, osteopontin, bone sialoprotein, osteocalcin, and collagen I were measured. After the application of mechanical force, Osx was upregulated in a time-dependent manner at both mRNA and protein levels, and Osx protein was translocated from the cytosol into the cell nuclei. Overexpression of Osx did not affect the expression of Cbfa1, but it significantly enhanced the ALP activity and the mRNA expression of all the aforementioned osteogenic marker genes, all of which increased further under mechanical stress. These results suggest that Osx might play an important role in the mechanical stress-induced osteogenic differentiation of PDLC and therefore be involved in alveolar bone remodeling during orthodontic therapy. [source] Labeling of Adipose-Derived Stem Cells by Oleic-Acid-Modified Magnetic NanoparticlesADVANCED FUNCTIONAL MATERIALS, Issue 8 2009Lian Cen Abstract The in vivo tracking of adipose derived stem cells (ASCs) is of essential concern when they are used as seed cells in tissue engineering. This study explores the feasibility of using magnetic nanoparticles (MNs), a type of contrast agents in magnetic resonance imaging (MRI), to label ASCs such that the labeled ASCs could be tracked in vivo by MRI non-invasively and repeatedly. To do this, MNs of <10,nm surface-coated with oleic acid are synthesized via a high-temperature solution-phase reaction. Cytotoxicity of the as-synthesized MNs at concentrations up to 0.1,mg,mL,1 on 104,cells,mL,1 ASCs is evaluated by LDH release. Since only minor cytotoxicity is detected, the effects of the labeling technique on cellular behaviors and uptake by labeled cells are investigated. Cell proliferation and differentiation with and without MNs are compared. The results show that proliferation of ASCs (104,cells,mL,1) labeled by MNs (0.05,mg,mL,1) is significantly enhanced and dependent on the labeling time. The MNs are located in the vesicles within cytoplasm of ASCs. The cellular uptake reaches as high as ,180,pg/cell. Nevertheless, the labeled ASCs still maintained adipogenic and osteogenic differentiation. Hence, the feasibility of labeling ASCs by oleic acid coated MNs is ascertained and it was better to label the cells during their quiescent stage. The labeled ASCs can also be in vivo detected by MRI in a subcutaneous model in vivo. Further MRI tracking of the labeled ASCs in long-term follow-up would thus follow this current study. [source] Cell proliferation and osteogenic differentiation of growing pig cranial suturesJOURNAL OF ANATOMY, Issue 3 2007Zongyang Sun Abstract Bone growth at the cranial sutures relies on proliferation of osteogenic progenitor cells and/or differentiation of osteoblasts. The current study was undertaken to assess these events in relation to suture growth and fusion. A total of 21 pigs, divided into three age groups (0.5,1.5 months, 3,4 months and 5,7 months), were used for immunohistochemical evaluation of cell proliferation (BrdU) and osteogenic differentiation (Cbfa1/Runx2) in the interfrontal and interparietal sutures. Proliferation and osteogenic differentiation were both more prominent near the bone fronts than in the central zone. With age, both proliferation and osteogenic differentiation diminished. Proliferation ceased on the endocranial (dura mater) side by the age of 3,4 months. Proliferation on the pericranial side was accompanied by active bone formation and initiation of suture fusion from this side. In conclusion, (1) decreased suture bone growth with age reflects decreased cell proliferation and probably also osteogenic differentiation, and (2) suture fusion occurs from the pericranial side where activity remains relatively high. [source] Origin matters: Differences in embryonic tissue origin and Wnt signaling determine the osteogenic potential and healing capacity of frontal and parietal calvarial bonesJOURNAL OF BONE AND MINERAL RESEARCH, Issue 7 2010Natalina Quarto Abstract Calvarial bones arise from two embryonic tissues, namely, the neural crest and the mesoderm. In this study we have addressed the important question of whether disparate embryonic tissue origins impart variable osteogenic potential and regenerative capacity to calvarial bones, as well as what the underlying molecular mechanism(s). Thus, by performing in vitro and in vivo studies, we have investigated whether differences exist between neural crest,derived frontal and paraxial mesodermal,derived parietal bone. Of interest, our data indicate that calvarial bone osteoblasts of neural crest origin have superior potential for osteogenic differentiation. Furthermore, neural crest,derived frontal bone displays a superior capacity to undergo osseous healing compared with calvarial bone of paraxial mesoderm origin. Our study identified both in vitro and in vivo enhanced endogenous canonical Wnt signaling in frontal bone compared with parietal bone. In addition, we demonstrate that constitutive activation of canonical Wnt signaling in paraxial mesodermal,derived parietal osteoblasts mimics the osteogenic potential of frontal osteoblasts, whereas knockdown of canonical Wnt signaling dramatically impairs the greater osteogenic potential of neural crest,derived frontal osteoblasts. Moreover, fibroblast growth factor 2 (FGF-2) treatment induces phosphorylation of GSK-3, and increases the nuclear levels of ,-catenin in osteoblasts, suggesting that enhanced activation of Wnt signaling might be mediated by FGF. Taken together, our data provide compelling evidence that indeed embryonic tissue origin makes a difference and that active canonical Wnt signaling plays a major role in contributing to the superior intrinsic osteogenic potential and tissue regeneration observed in neural crest,derived frontal bone. © 2010 American Society for Bone and Mineral Research [source] Mechanical stretching induces osteoprotegerin in differentiating C2C12 precursor cells through noncanonical Wnt Pathways,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 5 2010Hsiao-Chi Yu Abstract Mechanical loading is known to be important for maintaining the formation and resorption rates of bone. To study the mechanisms by which mechanical loading regulates osteogenesis, we investigated the role of the Wnt pathway in C2C12 cells committed to osteogenic differentiation in response to cyclic mechanical stretching. Osteoprotegerin (OPG) acts as a decoy receptor for RANKL to inhibit osteoclastogenesis and resorption of bone. Our results demonstrate that stretching leads to a sustained increase in OPG expression in C2C12 cells. The expression of osteogenic marker genes, such as osteocalcin and alkaline phosphatase, was transiently decreased by stretching at 24 hours and returned to control levels at 48 hours. The addition of inhibitors of the canonical Wnt/,-catenin pathways, such as the secreted FZD-related peptide sRFP2, as well as siRNA-mediated knockdown, did not inhibit the effect of stretching on OPG expression. In contrast, treatment with inhibitors of noncanonical Wnt signaling, including KN93, and siRNA for Nemo-like kinase (NLK) blocked most of the mechanical inductive effect on OPG. Furthermore, stretching-induced OPG production in the culture medium was able to inhibit the osteoclast formation of bone marrow macrophages. These results suggest that mechanical stretching may play an important role in bone remodeling through the upregulation of OPG and that the mechanical signaling leading to OPG induction involves the noncanonical Wnt pathway. © 2010 American Society for Bone and Mineral Research [source] Age-Related Changes in the Osteogenic Differentiation Potential of Mouse Bone Marrow Stromal Cells,,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 7 2008Weixi Zhang Abstract Age-dependent bone loss has been well documented in both human and animal models. Although the underlying causal mechanisms are probably multifactorial, it has been hypothesized that alterations in progenitor cell number or function are important. Little is known regarding the properties of bone marrow stromal cells (BMSCs) or bone progenitor cells during the aging process, so the question of whether aging alters BMSC/progenitor osteogenic differentiation remains unanswered. In this study, we examined age-dependent changes in bone marrow progenitor cell number and differentiation potential between mature (3 and 6 mo old), middle-aged (12 and 18 mo old), and aged (24 mo old) C57BL/6 mice. BMSCs or progenitors were isolated from five age groups of C57BL/6 mice using negative immunodepletion and positive immunoselection approaches. The osteogenic differentiation potential of multipotent BMSCs was determined using standard osteogenic differentiation procedures. Our results show that both BMSC/progenitor number and differentiation potential increase between the ages of 3 and 18 mo and decrease rapidly thereafter with advancing age. These results are consistent with the changes of the mRNA levels of osteoblast lineage-associated genes. Our data suggest that the decline in BMSC number and osteogenic differentiation capacity are important factors contributing to age-related bone loss. [source] Dysregulated BMP Signaling and Enhanced Osteogenic Differentiation of Connective Tissue Progenitor Cells From Patients With Fibrodysplasia Ossificans Progressiva (FOP),JOURNAL OF BONE AND MINERAL RESEARCH, Issue 3 2008Paul C Billings Abstract The study of FOP, a disabling genetic disorder of progressive heterotopic ossification, is hampered by the lack of readily available connective tissue progenitor cells. We isolated such cells from discarded primary teeth of patients with FOP and controls and discovered dysregulation of BMP signaling and rapid osteoblast differentiation in FOP cells compared with control cells. Introduction: Fibrodysplasia ossificans progressiva (FOP), the most disabling condition of progressive heterotopic ossification in humans, is caused by a recurrent heterozygous missense mutation in activin receptor IA (ACVR1), a bone morphogenetic protein (BMP) type I receptor, in all classically affected individuals. A comprehensive understanding of FOP has been limited, in part, by a lack of readily available connective tissue progenitor cells in which to study the molecular pathology of this disorder. Materials and Methods: We derived connective tissue progenitor cells from discarded primary teeth (SHED cells) of patients with FOP and controls and examined BMP signaling and osteogenic differentiation in these cells. Results: SHED cells transmitted BMP signals through both the SMAD and p38 mitogen-activated protein kinase (MAPK) pathways and responded to BMP4 treatment by inducing BMP responsive genes. FOP cells showed ligand-independent BMP signaling and ligand-dependent hyper-responsiveness to BMP stimulation. Furthermore, FOP cells showed more rapid differentiation to an osteogenic phenotype than control cells. Conclusions: This is the first study of BMP signaling and osteogenic differentiation in connective tissue progenitor cells from patients with FOP. Our data strongly support both basal and ligand-stimulated dysregulation of BMP signaling consistent with in silico studies of the mutant ACVR1 receptor in this condition. This study substantially extends our understanding of dysregulated BMP signaling in a progenitor cell population relevant to the pathogenesis of this catastrophic disorder of progressive ectopic ossification. [source] An In Vivo Model to Study Osteogenic Gene Regulation: Targeting an Avian Retroviral Receptor (TVA) to Bone With the Bone Sialoprotein (BSP) Promoter,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 8 2005Ling Li Abstract To study bone development in vivo, a transgenic mouse model was established in which an avian retroviral receptor (TVA) gene driven by the BSP promoter was selectively expressed in skeletal tissues. The model was validated by showing suppressed BSP expression and delayed bone and tooth formation after infection with a virus expressing a mutated Cbfa1/Runx2 gene. Introduction: Tissue-specific expression of the avian retroviral (TVA) receptor can be used to efficiently target ectopic expression of genes in vivo. To determine the use of this approach for studies of osteogenic differentiation and bone formation at specific developmental stages, transgenic mice expressing the TVA receptor under the control of a 5-kb bone sialoprotein (BSP) promoter were generated. The mice were first analyzed for tissue-specific expression of the TVA gene and then, after infection with a viral construct, for the effects of a dominant-negative form of the Cbfa1/Runx2 transcription factor on bone formation. Materials and Methods: We first generated transgenic mice (BSP/TVA) in which the TVA gene was expressed under the control of a 4.9-kb mouse BSP promoter. The tissue-specific expression of the TVA gene was analyzed by RT-PCR, in situ hybridization, and immunohistochemistry and compared with the expression of the endogenous BSP gene. A 396-bp fragment of mutated Cbfa1/Runx2 (Cbfa1mu) encoding the DNA-binding domain was cloned into a RCASBP (A) viral vector, which was used to infect neonatal BSP/TVA mice. Results and Conclusion: Expression of the TVA receptor mRNA and protein in the transgenic mice was consistent with the expression of endogenous BSP. Four days after systemic infection with the Cbfa1mu-RCASBP (A) vector, RT-PCR analyses revealed that the expression of BSP mRNA in tibia and mandibles was virtually abolished, whereas a 30% reduction was seen in calvarial bone. After 9 days, BSP expression in the tibia and mandible was reduced by 45% in comparison with control animals infected with an empty RCASBP vector, whereas BSP expression in the membranous bone of calvariae was decreased ,15%. However, after 4 and 8 weeks, there was almost no change in BSP expression in any of the bone tissues. In comparison, a reduction in osteopontin expression was only observed 9 days after viral transfection in the three bones. Histomorphological examination revealed that bone formation and tooth development were delayed in some of the mice infected with mutated Cbfa1. These studies show that BSP/TVA transgenic mice can be used to target genes to sites of osteogenesis, providing a unique system for studying molecular events associated with bone formation in vivo. [source] Strategies for Directing the Differentiation of Stem Cells Into the Osteogenic Lineage In Vitro,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 9 2004Boon Chin Heng Abstract A major area in regenerative medicine is the application of stem cells in bone reconstruction and bone tissue engineering. This will require well-defined and efficient protocols for directing the differentiation of stem cells into the osteogenic lineage, followed by their selective purification and proliferation in vitro. The development of such protocols would reduce the likelihood of spontaneous differentiation of stem cells into divergent lineages on transplantation, as well as reduce the risk of teratoma formation in the case of embryonic stem cells. Additionally, such protocols could provide useful in vitro models for studying osteogenesis and bone development, and facilitate the genetic manipulation of stem cells for therapeutic applications. The development of pharmokinetic and cytotoxicity/genotoxicity screening tests for bone-related biomaterials and drugs could also use protocols developed for the osteogenic differentiation of stem cells. This review critically examines the various strategies that could be used to direct the differentiation of stem cells into the osteogenic lineage in vitro. [source] Shock Wave Application Enhances Pertussis Toxin Protein-Sensitive Bone Formation of Segmental Femoral Defect in Rats,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 12 2003Yeung-Jen Chen Abstract Extracorporeal shock waves (ESWs) elicit a dose-dependent effect on the healing of segmental femoral defects in rats. After ESW treatment, the segmental defect underwent progressive mesenchymal aggregation, endochondral ossification, and hard callus formation. Along with the intensive bone formation, there was a persistent increase in TGF-,1 and BMP-2 expression. Pretreatment with pertussis toxin reduced ESW-promoted callus formation and gap healing, which presumably suggests that Gi proteins mediate osteogenic signaling. Introduction: Extracorporeal shock waves (ESWs) have previously been used to promote bone repair. In our previous report, we found that ESWs promoted osteogenic differentiation of mesenchymal cells through membrane perturbation and activation of Ras protein. In this report, we show that ESWs elicit a dose-dependent effect on the healing of segmental defects and that Gi proteins play an important role in mediating ESW stimulation. Materials and Methods: Rats with segmental femoral defects were subjected to ESW treatment at different energy flux densities (EFD) and impulses. Bone mass (mineral density and calcium content), osteogenic activities (bone alkaline phosphatase activity and osteocalcin content), and immunohistochemistry were assessed. Results: An optimal ESW energy (500 impulses at 0.16 mJ/mm2 EFD) stimulated complete bone healing without complications. ESW-augmented healing was characterized by significant increases (p < 0.01) in callus size, bone mineral density, and bone tissue formation. With exposure to ESW, alkaline phosphatase activity and osteocalcin production in calluses were found to be significantly enhanced (p < 0.05). After ESW treatment, the histological changes we noted included progressive mesenchymal aggregation, endochondral ossification, and hard callus formation. Intensive bone formation was associated with a persistent increase in transforming growth factor-beta 1 (TGF-,1) and bone morphogenetic protein-2 (BMP-2) expression, suggesting both growth factors were active in ESW-promoted bone formation. We also found that pertussis toxin, an inhibitor of membrane-bound Gi proteins, significantly reduced (p < 0.01) ESW promotion of callus formation and fracture healing. Conclusion: ESW treatments enhanced bone formation and the healing of segmental femoral defects in rats. It also seems likely that TGF-,1 and BMP-2 are important osteogenic factors for ESW promotion of fracture healing, presumably through Gi protein-mediated osteogenic signaling. [source] Foetal and adult cardiomyocyte progenitor cells have different developmental potentialJOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 4 2010Patrick Van Vliet Abstract In the past years, cardiovascular progenitor cells have been isolated from the human heart and characterized. Up to date, no studies have been reported in which the developmental potential of foetal and adult cardiovascular progenitors was tested simultaneously. However, intrinsic differences will likely affect interpretations regarding progenitor cell potential and application for regenerative medicine. Here we report a direct comparison between human foetal and adult heart-derived cardiomyocyte progenitor cells (CMPCs). We show that foetal and adult CMPCs have distinct preferences to differentiate into mesodermal lineages. Under pro-angiogenic conditions, foetal CMPCs form more endothelial but less smooth muscle cells than adult CMPCs. Foetal CMPCs can also develop towards adipocytes, whereas neither foetal nor adult CMPCs show significant osteogenic differentiation. Interestingly, although both cell types differentiate into heart muscle cells, adult CMPCs give rise to electrophysiologically more mature cardiomyocytes than foetal CMPCs. Taken together, foetal CMPCs are suitable for molecular cell biology and developmental studies. The potential of adult CMPCs to form mature cardiomyocytes and smooth muscle cells may be essential for cardiac repair after transplantation into the injured heart. [source] BMP-9-induced osteogenic differentiation of mesenchymal progenitors requires functional canonical Wnt/,-catenin signallingJOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 8b 2009Ni Tang Abstract Bone morphogenetic protein 9 (BMP-9) is a member of the transforming growth factor (TGF)-,/BMP superfamily, and we have demonstrated that it is one of the most potent BMPs to induce osteoblast differentiation of mesenchymal stem cells (MSCs). Here, we sought to investigate if canonical Wnt/,-catenin signalling plays an important role in BMP-9-induced osteogenic differentiation of MSCs. Wnt3A and BMP-9 enhanced each other's ability to induce alkaline phosphatase (ALP) in MSCs and mouse embryonic fibroblasts (MEFs). Wnt antagonist FrzB was shown to inhibit BMP-9-induced ALP activity more effectively than Dkk1, whereas a secreted form of LPR-5 or low-density lipoprotein receptor-related protein (LRP)-6 exerted no inhibitory effect on BMP-9-induced ALP activity. ,-Catenin knockdown in MSCs and MEFs diminished BMP-9-induced ALP activity, and led to a decrease in BMP-9-induced osteocalcin reporter activity and BMP-9-induced expression of late osteogenic markers. Furthermore, ,-catenin knockdown or FrzB overexpression inhibited BMP-9-induced mineralization in vitro and ectopic bone formation in vivo, resulting in immature osteogenesis and the formation of chondrogenic matrix. Chromatin immunoprecipitation (ChIP) analysis indicated that BMP-9 induced recruitment of both Runx2 and ,-catenin to the osteocalcin promoter. Thus, we have demonstrated that canonical Wnt signalling, possibly through interactions between ,-catenin and Runx2, plays an important role in BMP-9-induced osteogenic differentiation of MSCs. [source] The polymine spermine regulates osteogenic differentiation in adipose stem cells,JOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 5a 2008G.S. Tjabringa Abstract For bone tissue engineering, it is important that mesenchymal stem cells (MSCs) differentiate into osteoblasts. To develop a method for differentiation of adipose tissue-derived mesenchymal stem cells (AT-MSCs) along the osteogenic lineage, we studied the effect of polyamines, which are organic cations implicated in bone growth and development, on differentiation of AT-MSCs. Treatment of goat-derived AT-MSCs with 1,25-dihydroxyvitamin-D3 (1,25(OH)2D3), which stimulates osteogenic differentiation, for 7 days induced gene expression of the polyamine-modulated transcription factor-1 (PMF-1) and spermidine/spermine N (1)-acetyltransferase (SSAT), which are both involved in polyamine metabolism, suggesting that polyamines are involved in osteogenic differentiation of AT-MSCs. Furthermore, treatment of AT-MSCs with the polyamine spermine-regulated gene expression of runx-2, a transcription factor involved in early stages of osteogenic differentiation, and that of osteopontin, a bone matrix protein expressed in later stages of osteogenic differentiation. Runx-2 gene expression was increased 4 and 14 days after a short 30 min. treatment with spermine, while osteopontin gene expression was only increased 4 days after spermine treatment. Finally, alkaline phosphatase activity, which is intimately involved in the formation of extracellular matrix of bone, was increased 4 weeks after the 30 min.-spermine treatment of AT-MSCs. In conclusion, this study shows for the first time that the polyamine spermine regulates differentiation of AT-MSCs along the osteogenic lineage, which can be used as a new method for differentiation of AT-MSCs along the osteogenic lineage. Therefore, polyamines may constitute a promising tool for bone tissue engineering approaches using AT-MSCs, such as a one-step surgical procedure for spinal interbody fusion. [source] Oxysterol-induced osteogenic differentiation of marrow stromal cells is regulated by Dkk-1 inhibitable and PI3-kinase mediated signalingJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 2 2008Christopher M. Amantea Abstract Osteoporosis and its complications cause morbidity and mortality in the aging population, and result from increased bone resorption by osteoclasts in parallel with decreased bone formation by osteoblasts. A widely accepted strategy for improving bone health is targeting osteoprogenitor cells in order to stimulate their osteogenic differentiation and bone forming properties through the use of osteoinductive/anabolic factors. We previously reported that specific naturally occurring oxysterols have potent osteoinductive properties, mediated in part through activation of hedgehog signaling in osteoprogenitor cells. In the present report, we further demonstrate the molecular mechanism(s) by which oxysterols induce osteogenesis. In addition to activating the hedgehog signaling pathway, oxysterol-induced osteogenic differentiation is mediated through a Wnt signaling-related, Dkk-1-inhibitable mechanism. Bone marrow stromal cells (MSC) treated with oxysterols demonstrated increased expression of osteogenic differentiation markers, along with selective induced expression of Wnt target genes. These oxysterol effects, which occurred in the absence of ,-catenin accumulation or TCF/Lef activation, were inhibited by the hedgehog pathway inhibitor, cyclopamine, and/or by the Wnt pathway inhibitor, Dkk-1. Furthermore, the inhibitors of PI3-Kinase signaling, LY 294002 and wortmanin, inhibited oxysterol-induced osteogenic differentiation and induction of Wnt signaling target genes. Finally, activators of canonical Wnt signaling, Wnt3a and Wnt1, inhibited spontaneous, oxysterol-, and Shh-induced osteogenic differentiation of bone marrow stromal cells, suggesting the involvement of a non-canonical Wnt pathway in pro-osteogenic differentiation events. Osteogenic oxysterols are, therefore, important small molecule modulators of critical signaling pathways in pluripotent mesenchymal cells that regulate numerous developmental and post-developmental processes. J. Cell. Biochem. 105: 424,436, 2008. © 2008 Wiley-Liss, Inc. [source] Wnt 3a promotes proliferation and suppresses osteogenic differentiation of adult human mesenchymal stem cells,JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 6 2004Genevieve M. Boland Abstract Multipotential adult mesenchymal stem cells (MSCs) are able to differentiate along several known lineages, and lineage commitment is tightly regulated through specific cellular mediators and interactions. Recent observations of a low/high bone-mass phenotype in patients expressing a loss-/gain-of-function mutation in LRP5, a coreceptor of the Wnt family of signaling molecules, suggest the importance of Wnt signaling in bone formation, possibly involving MSCs. To analyze the role of Wnt signaling in mesenchymal osteogenesis, we have profiled the expression of WNTs and their receptors, FRIZZLEDs (FZDs), and several secreted Wnt inhibitors, such as SFRPs, and examined the effect of Wnt 3a, as a representative canonical Wnt member, during MSC osteogenesis in vitro. WNT11, FZD6, SFRP2, and SFRP3 are upregulated during MSC osteogenesis, while WNT9A and FZD7 are downregulated. MSCs also respond to exogenous Wnt 3a, based on increased ,-catenin nuclearization and activation of a Wnt-responsive promoter, and the magnitude of this response depends on the MSC differentiation state. Wnt 3a exposure inhibits MSC osteogenic differentiation, with decreased matrix mineralization and reduced alkaline phosphatase mRNA and activity. Wnt 3a treatment of fully osteogenically differentiated MSCs also suppresses osteoblastic marker gene expression. The Wnt 3a effect is accompanied by increased cell number, resulting from both increased proliferation and decreased apoptosis, particularly during expansion of undifferentiated MSCs. The osteo-suppressive effects of Wnt 3a are fully reversible, i.e., treatment prior to osteogenic induction does not compromise subsequent MSC osteogenesis. The results also showed that sFRP3 treatment attenuates some of the observed Wnt 3a effects on MSCs, and that inhibition of canonical Wnt signaling using a dominant negative TCF1 enhances MSC osteogenesis. Interestingly, expression of Wnt 5a, a non-canonical Wnt member, appeared to promote osteogenesis. Taken together, these findings suggest that canonical Wnt signaling functions in maintaining an undifferentiated, proliferating progenitor MSC population, whereas non-canonical Wnts facilitate osteogenic differentiation. Release from canonical Wnt regulation is a prerequisite for MSC differentiation. Thus, loss-/gain-of-function mutations of LRP5 would perturb Wnt signaling and depress/promote bone formation by affecting the progenitor cell pool. Elucidating Wnt regulation of MSC differentiation is important for their potential application in tissue regeneration. Published 2004 Wiley-Liss, Inc. [source] To go or not to go: Migration of human mesenchymal progenitor cells stimulated by isoforms of PDGFJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 5 2004Jörg Fiedler Abstract The recruitment of mesenchymal progenitor cells (MPCs) and their subsequent differentiation to osteoblasts is mandatory for bone development, remodeling, and repair. To study the possible involvement of platelet-derived growth factor (PDGF) isoforms, primary human MPCs and osteogenic differentiated progenitor cells (dOB) were examined for chemotaxic response to homodimeric human platelet-derived growth factor AA, -BB, and heterodimeric PDGF-AB. The role of PDGF receptors was addressed by preincubation with PDGF receptor alpha and beta chain specific antibodies. Migration of MPCs, dOB, and primary osteoblasts (OB) was stimulated by the addition of rhPDGF-AA, rhPDGF-BB, and rhPDGF-AB. The effect was highest in MPCs and for rhPDGF-BB, and declining with osteogenic differentiation. Preincubation with the receptor alpha specific antibody decreased the CI to borderline values while pretreatment with the receptor beta specific antibody led to a complete loss of chemotactic response to PDGF isoforms. In control experiments, basal migration values and rhBMP-2 as well as rxBMP-4 induced chemotaxis of MPC were not influenced by the addition of receptor alpha or beta antibodies. Interestingly, without preincubation the parallel exposure of MPC to rhTGF-,1 instantaneously leads to a selective loss of migratory stimulation by rhPDGF-AA. The chemotactic effect of PDGF isoforms for primary human MPCs and the influence of osteogenic differentiation suggest a functional role for recruitment of MPCs during bone development and remodeling. Moreover, these observations may be useful for novel approaches towards guided tissue regeneration or tissue engineering of bone. © 2004 Wiley-Liss, Inc. [source] Estrogen modulates estrogen receptor , and , expression, osteogenic activity, and apoptosis in mesenchymal stem cells (MSCs) of osteoporotic miceJOURNAL OF CELLULAR BIOCHEMISTRY, Issue S36 2001Shuanhu Zhou Abstract In the mouse, ovariectomy (OVX) leads to significant reductions in cancellous bone volume while estrogen (17,-estradiol, E2) replacement not only prevents bone loss but can increase bone formation. As the E2-dependent increase in bone formation would require the proliferation and differentiation of osteoblast precursors, we hypothesized that E2 regulates mesenchymal stem cells (MSCs) activity in mouse bone marrow. We therefore investigated proliferation, differentiation, apoptosis, and estrogen receptor (ER) , and , expression of primary culture MSCs isolated from OVX and sham-operated mice. MSCs, treated in vitro with 10,7 M E2, displayed a significant increase in ER, mRNA and protein expression as well as alkaline phosphatase (ALP) activity and proliferation rate. In contrast, E2 treatment resulted in a decrease in ER, mRNA and protein expression as well as apoptosis in both OVX and sham mice. E2 up-regulated the mRNA expression of osteogenic genes for ALP, collagen I, TGF-,1, BMP-2, and cbfa1 in MSCs. In a comparison of the relative mRNA expression and protein levels for two ER isoforms, ER, was the predominant form expressed in MSCs obtained from both OVX and sham-operated mice. Cumulatively, these results indicate that estrogen in vitro directly augments the proliferation and differentiation, ER, expression, osteogenic gene expression and, inhibits apoptosis and ER, expression in MSCs obtained from OVX and sham-operated mice. Co-expression of ER,, but not ER,, and osteogenic differentiation markers might indicate that ER, function as an activator and ER, function as a repressor in the osteogenic differentiation in MSCs. These results suggest that mouse MSCs are anabolic targets of estrogen action, via ER, activation. J. Cell. Biochem. Suppl. 36: 144,155, 2001. © 2001 Wiley-Liss, Inc. [source] Osterix is a key target for mechanical signals in human thoracic ligament flavum cellsJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2007Dongwei Fan Mechanical stress is considered to be an important factor in the progression of thoracic ossification of the ligament flavum (TOLF). To elucidate the mechanism underlying mechanical stress-induced TOLF, we investigated the effect of stretching on cultured flavum ligament cells derived from TOLF and non-TOLF patients. We found that the mRNA expression of alkaline phosphatase (ALP), osteocalcin, Runx2, and osterix, but not that of Dlx5 and Msx2, was significantly increased by stretching in TOLF cells. In addition, the effect seems to be finely tuned by stretching-triggered activation of distinct mitogen-activated protein kinase cascades. Specifically, a p38 specific inhibitor, SB203580, significantly inhibited stretching-induced osterix expression as well as ALP activity, whereas a specific inhibitor of ERK1/2, U0126, prevented stretching-induced Runx2 expression. We showed that overexpression of osterix resulted in a significant increase of ALP activity in TOLF cells, and osterix-specific RNAi completely abrogated the stretching-induced ALP activity, indicating that osterix plays a key role in stretching-stimulated osteogenic effect in TOLF cells. These results suggest that mechanical stress plays important roles in the progression of TOLF through induction of osteogenic differentiation of TOLF cells, and our findings support that osterix functions as a molecular link between mechanostressing and osteogenic differentiation. J. Cell. Physiol. 211: 577,584, 2007. © 2007 Wiley-Liss, Inc. [source] The regulation of osteogenesis by ECM rigidity in MC3T3-E1 cells requires MAPK activationJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2007Chirag B. Khatiwala Once thought to provide only structural support to tissues by acting as a scaffold to which cells bind, it is now widely recognized that the extracellular matrix (ECM) provides instructive signals that dictate cell behavior. Recently we demonstrated that mechanical cues intrinsic to the ECM directly regulate the behavior of pre-osteoblastic MC3T3-E1 cells. We hypothesized that one possible mechanism by which ECM compliance exerts its influence on osteogenesis is by modulating the mitogen-activated protein kinase (MAPK) pathway. To address this hypothesis, the differentiation of MC3T3-E1 cells cultured on poly(ethylene glycol) (PEG)-based model substrates with tunable mechanical properties was assessed. Alkaline phosphatase (ALP) levels at days 7 and 14 were found to be significantly higher in cells grown on stiffer substrates (423.9 kPa hydrogels and rigid tissue culture polystyrene (TCPS) control) than on a soft hydrogel (13.7 kPa). Osteocalcin (OCN) and bone sialoprotein (BSP) gene expression levels followed a similar trend. In parallel, MAPK activity was significantly higher in cells cultured on stiffer substrates at both time points. Inhibiting this activation pharmacologically, using PD98059, resulted in significantly lower ALP levels, OCN, and BSP gene expression levels on the hydrogels. Interestingly, the effectiveness of PD98059 was itself dependent on substrate stiffness, with marked inhibition of MAPK phosphorylation in cells grown on compliant hydrogels but insignificant reduction in cells grown on TCPS. Together, these data confirm a role for MAPK in the regulation of osteogenic differentiation by ECM compliance. J. Cell. Physiol. 211: 661,672, 2007. © 2007 Wiley-Liss, Inc. [source] Alterations in intranuclear localization of Runx2 affect biological activity,,JOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2006Sayyed K. Zaidi The transcription factor Runx2 controls osteoblast proliferation and differentiation. Runx2 organizes and assembles gene-regulatory complexes in nuclear microenvironments where target genes are activated or suppressed in a context-dependent manner. Intranuclear localization of Runx2 is mediated by the nuclear matrix-targeting signal (NMTS), an autonomous motif with a loop (L1)-turn-loop (L2) structure that forms predicted protein,protein interaction surfaces. Here we examined the functional consequences of introducing mutations in the L1 and L2 loops of the NMTS. These mutant proteins enter the nucleus, interact with the hetero-dimeric partner Cbf,, and bind to DNA in vitro and in vivo. In addition, these mutants retain interaction with the carboxy-terminus interacting co-regulatory proteins that include TLE, YAP, and Smads. However, two critical mutations in the L2 domain of the NMTS decrease association of Runx2 with the nuclear matrix. These subnuclear targeting defective (STD) mutants of Runx2 compromise target gene activation or repression. The biological significance of these findings is reflected by decreased osteogenic differentiation of mesenchymal progenitors, concomitant with major changes in gene expression profiles, upon expression of the STD Runx2 mutant. Our results demonstrate that fidelity of temporal and spatial localization of Runx2 within the nucleus is functionally linked with biological activity. J. Cell. Physiol. 209: 935,942, 2006. © 2006 Wiley-Liss, Inc. [source] In vivo anabolic effect of strontium on trabecular bone was associated with increased osteoblastogenesis of bone marrow stromal cells,JOURNAL OF ORTHOPAEDIC RESEARCH, Issue 9 2010Songlin Peng Abstract In vitro studies have demonstrated that strontium (Sr) could increase osteogenic differentiation of bone marrow stromal cells (BMSCs). We investigated the in vivo effect of Sr on BMSCs. Thirty-six female rats were randomly divided into the following groups: sham operated and treated with either vehicle (Sham,+,Veh) or Sr compound (Sham,+,Sr) and ovariectomized and treated with either vehicle (OVX,+,Veh) or Sr compound (OVX,+,Sr). Vehicle and Sr were orally administrated daily starting immediately after the surgery and continuing for 12 weeks. The anabolic effect of Sr on trabecular bone was determined at the structural and tissue level by microCT and histomorphometry, respectively. Colony formation assays demonstrated that BMSCs exhibited higher osteogenic colony but lower adipogenic colony in Sr-treated versus Veh-treated OVX rats. The mRNA level of osteogenic genes was higher, while the mRNA level of adipogenic genes was lower in BMSCs from Sr-treated versus Veh-treated Sham and OVX rats. The effect of Sr on rat BMSCs was reproducible in human BMSCs. Taken together, this study suggests that the anabolic effect of Sr on normal or osteoporotic bones is associated with increased osteoblastic differentiation of BMSCs. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:1208,1214, 2010 [source] Putative heterotopic ossification progenitor cells derived from traumatized muscle,JOURNAL OF ORTHOPAEDIC RESEARCH, Issue 12 2009Wesley M. Jackson Abstract Heterotopic ossification (HO) is a frequent complication following combat-related trauma, but the pathogenesis of traumatic HO is poorly understood. Building on our recent identification of mesenchymal progenitor cells (MPCs) in traumatically injured muscle, the goal of this study was to evaluate the osteogenic potential of the MPCs in order to assess the role of these cells in HO formation. Compared to bone marrow-derived mesenchymal stem cells (MSCs), a well-characterized population of osteoprogenitor cells, the MPCs exhibited several significant differences during osteogenic differentiation and in the expression of genes related to osteogenesis. Upon osteogenic induction, MPCs showed increased alkaline phosphatase activity, production of a mineralized matrix, and up-regulated expression of the osteoblast-associated genes CBFA1 and alkaline phosphatase. However, MPCs did not appear to reach terminal differentiation as the expression of osteocalcin was not substantially up-regulated. With the exception of a few genes, the osteogenic gene expression profile of traumatized muscle-derived MPCs was comparable to that of the MSCs after osteogenic induction. These findings indicate that traumatized muscle-derived MPCs have the potential to function as osteoprogenitor cells when exposed to the appropriate biochemical environment and are the putative osteoprogenitor cells that initiate ectopic bone formation in HO. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27:1645,1651, 2009 [source] Effect of hyaluronan on osteogenic differentiation of porcine bone marrow stromal cells in vitroJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 5 2008Lijin Zou Abstract Hyaluronan (HA) plays a predominant role in tissue morphogenesis, cell migration, proliferation, and cell differentiation. The aims of the present study were to investigate whether (i) prolonged presence of high concentration (4.0 mg/mL) 800 KDa HA and (ii) pretreatment with HA can modify osteogenic differentiation of pig bone marrow stromal cells (pBMSC). Cell proliferation and mineralization were measured. Expression of differentiation-related genes was evaluated by means of real-time reverse transcription polymerase chain reaction (RT-PCR). HA increased cell proliferation on day 7. HA decreased the basal level of bone-related gene expression and increased the basal level of sox9 marginally during 7-day pretreatment with HA. HA increased calcium deposit on day 21. cbfa1, ALP, and type 1, collagen (Col1) expression was increased when pBMSC were cultivated in osteogenic medium, whereas their expression was decreased in the presence of HA on day 7. On day 14, the addition of HA upregulated cbfa1 and ALP expression compared to osteogenic medium group; there was no significant difference in Col1 expression. At day 21, osteocalcin (OC) expression showed 2.5-fold upregulation over osteogenic medium. These results suggest that exogenous HA stimulates endogenous HA, which together may play a synergetic role in osteogenic differentiation under osteoinducing conditions although gene expression was inhibited at the early stage. © 2007 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 26:713,720, 2008 [source] | ||||||||||||||||