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Bone Development (bone + development)
Selected AbstractsLocalization of Indian hedgehog and PTH/PTHrP receptor expression in relation to chondrocyte proliferation during mouse bone developmentDEVELOPMENT GROWTH & DIFFERENTIATION, Issue 2 2005Helen E. MacLean We have developed a useful approach to examine the pattern of gene expression in comparison to cell proliferation, using double in situ hybridization and immunofluorescence. Using this system, we examined the expression of Indian hedgehog (Ihh) and PTH/PTHrP receptor (PPR) mRNA in relation to chondrocyte proliferation during embryonic mouse bone development. Both genes are expressed strongly in prehypertrophic and early hypertrophic chondrocytes, and there is a strong correlation between upregulation of both Ihh and PPR expression and chondrocyte cell cycle arrest. At embryonic day (E14.5), PPR mRNA upregulation begins in the columnar chondrocytes just prior to cell cycle exit, but at later time points expression is only observed in the postproliferative region. In contrast, Ihh mRNA expression overlaps slightly with the region of columnar proliferating chondrocytes at all stages. This study provides further evidence that in the developing growth plate, cell cycle exit and upregulation of Ihh and PPR mRNA expression are coupled. [source] Studies on the appearance of skeletal anomalies in red porgy: effect of culture intensiveness, feeding habits and nutritional quality of live preysJOURNAL OF APPLIED ICHTHYOLOGY, Issue 2 2010M. S. Izquierdo Summary Despite the great interest of red porgy as a new species for Mediterranean aquaculture, its commercial production is constrained by the high incidence of skeletal deformities occurring in this species under culture conditions. Several studies have been conducted to better understand the origin of these anomalies in this species, using different system intensiveness, rotifers enrichment products or rotifers docosahexaenoic acid content. The first study showed that culture intensification increased the number of fish with an extra vertebrae, what was probably related to the different nutritional quality of live preys employed in each treatment, since water temperature, salinity and genetic background were identical for the different batches of fish studied. Total incidence of skeletal abnormalities was higher in the intensive system, particularly cranial abnormalities and kyphosis in the cephalic vertebrae. In both rearing systems the most common skeletal anomalies were vertebral column disorders, lordosis and fused vertebrae, their localization along the column being affected by the culture intensiveness. Rotifer enrichment, predominantly its docosahexaenoic acid content significantly affected deformities occurrence. A marked positive effect of rotifer docosahexaenoic acid content was found on larval survival. X-ray studies denoted elevated levels of bone abnormalities associated, in both trials, to low docosahexaenoic acid content in live preys. Among different anomalies, the presence of fused vertebrae was the most frequent deformity for both rearing trials. A 50% reduction in the number of deformed fish for each type of deformity was obtained when the larvae were fed higher docosahexaenoic acid levels, denoting the important role of this fatty acid in bone development. Further studies are needed to elucidate the importance of essential fatty acids on the development of bone deformities in fish, since the functions of these fatty acids differ among them and can lead to very different effects in fish metabolism, including bone formation. [source] Regulation of Osteogenesis-Angiogenesis Coupling by HIFs and VEGF,,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 8 2009Ernestina Schipani Abstract Bone is a highly vascularized tissue, but the function of angiogenesis in bone modeling and remodeling is still poorly defined, and the molecular mechanisms that regulate angiogenesis in bone are only partially elucidated. Genetic manipulations in mice have recently highlighted the critical role of the hypoxia-inducible-factor/vascular endothelial growth factor pathway in coupling angiogenesis and osteogenesis. In this brief perspective, we review the current understanding of the mechanisms responsible for this coupling. Elucidation of such mechanisms will expand our knowledge of bone development and homeostasis, and it may aid in the design of new therapies for accelerating bone regeneration and repair. [source] Sustained BMP Signaling in Osteoblasts Stimulates Bone Formation by Promoting Angiogenesis and Osteoblast Differentiation,,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 7 2009Fengjie Zhang Abstract Angiogenesis and bone formation are tightly coupled during the formation of the skeleton. Bone morphogenetic protein (BMP) signaling is required for both bone development and angiogenesis. We recently identified endosome-associated FYVE-domain protein (endofin) as a Smad anchor for BMP receptor activation. Endofin contains a protein-phosphatase pp1c binding domain, which negatively modulates BMP signals through dephosphorylation of the BMP type I receptor. A single point mutation of endofin (F872A) disrupts interaction between the catalytic subunit pp1c and sensitizes BMP signaling in vitro. To study the functional impact of this mutation in vivo, we targeted expression of an endofin (F872A) transgene to osteoblasts. Mice expressing this mutant transgene had increased levels of phosphorylated Smad1 in osteoblasts and showed increased bone formation. Trabecular bone volume was significantly increased in the transgenic mice compared with the wildtype littermates with corresponding increases in trabecular bone thickness and number. Interestingly, the transgenic mice also had a pronounced increase in the density of the bone vasculature measured using contrast-enhanced ,CT imaging of Microfil-perfused bones. The vessel surface and volume were both increased in association with elevated levels of vascular endothelial growth factor (VEGF) in osteoblasts. Endothelial sprouting from the endofin (F872A) mutant embryonic metatarsals cultured ex vivo was increased compared with controls and was abolished by an addition of a VEGF neutralizing antibody. In conclusion, osteoblast targeted expression of a mutant endofin protein lacking the pp1c binding activity results in sustained signaling of the BMP type I receptor, which increases bone formation and skeletal angiogenesis. [source] Murine and Chicken Chondrocytes Regulate Osteoclastogenesis by Producing RANKL in Response to BMP2,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 3 2008Michihiko Usui Abstract Chondrocytes express RANKL, but their role in osteoclastogenesis is not clear. We report that hypertrophic chondrocytes induce osteoclast formation through RANKL production stimulated by BMP2 and Runx2/Smad1 and thus they may regulate resorption of calcified matrix by osteoclasts at growth plates. Introduction: Bone morphogenetic protein (BMP) signaling and Runx2 regulate chondrogenesis during bone development and fracture repair and RANKL expression by osteoblast/stromal cells. Chondrocytes express RANKL, and this expression is stimulated by vitamin D3, but it is not known if chondrocytes directly support osteoclast formation or if BMPs or Runx2 is involved in this potential regulation of osteoclastogenesis. Material and Methods: The chondrocyte cell line, ATDC5, primary mouse sternal chondrocytes, and chick sternal chondrocytes were used. Cells were treated with BMP2, and expression of RANKL and chondrocyte marker genes was determined by real-time RT-PCR and Western blot. Chondrocytes and spleen-derived osteoclast precursors ± BMP2 were co-cultured to examine the effect of chondrocyte-produced RANKL on osteoclast formation. A reporter assay was used to determine whether BMP2-induced RANKL production is through transcriptional regulation of the RANKL promoter and whether it is mediated by Runx2. Results: BMP2 significantly increased expression of RANKL mRNA and protein in all three types of chondrocytes, particularly by Col X-expressing and upper sternal chondrocytes. Chondrocytes constitutively induced osteoclast formation. This effect was increased significantly by BMP2 and prevented by RANK:Fc. BMP2 significantly increased luciferase activity of the RANKL-luc reporter, and Smad1 increased this effect. Deletion or mutation of Runx2 binding sites within the RANKL promoter or overexpression of a dominant negative Runx2 abolished BMP2- and Smad1-mediated activation of RANKL promoter activity. Conclusions: Hypertrophic chondrocytes may regulate osteoclastogenesis at growth plates to remove calcified matrix through BMP-induced RANKL expression. [source] Growth and Bone Mineral Accretion During Puberty in Chinese Girls: A Five-Year Longitudinal Study,,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 2 2008Kun Zhu Abstract There are few longitudinal data on bone development during puberty in children with low calcium intake. This 5-yr longitudinal study showed that, in Chinese girls, the mean apparent calcium retention efficiency during puberty was 40.9%, PHV occurred at 3,0 yr before menarche, and peak bone mineral accretion occurred 1 yr later than PHV. Chinese girls have high calcium retention efficiency during puberty. Introduction: There are few longitudinal data on bone development during puberty in children with low dietary calcium intake. The aim of this study was to examine the rate of growth and bone mineral accretion and study the predictors of total body BMC during puberty in a 5-yr longitudinal study with Chinese girls. Materials and Methods: Ninety-two girls, 9.5,10.5 yr of age at baseline, from the unsupplemented control group of a school milk intervention trial were included in this analysis. Data on anthropometric measurements, total body BMC as assessed by DXA, and calcium intake as assessed by a 3-day food record were obtained at baseline and 1, 2, 4, and 5 yr. Results: The mean age of menarche was 12.1 ± 1.0 yr. The mean annual rate of bone mineral accretion was 197.4 g/yr during the follow-up period, representing a calcium accretion rate of 162.3 mg/d. This calcium retention rate and the average dietary calcium intake of 444.1 mg/d gave an apparent calcium retention efficiency of 40.9%. Peak height velocity (PHV) occurred at 3,0 yr before menarche. Peak bone mineral accretion occurred 1 yr later than PHV. There was a decrease in size-corrected BMD in the year before menarche. In the linear mixed-effects model analysis containing body size and lifestyle factors, we found that height, body weight, and calcium intake were significant independent predictors of total body BMC. Conclusions: Chinese girls with low habitual dietary calcium intake have high calcium retention efficiency during puberty. Because calcium intake is a significant predictor of total body BMC, increasing dietary calcium intake may have beneficial effects on bone mineral accretion in these girls. [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] Effect of Osteoblast-Targeted Expression of Bcl-2 in Bone: Differential Response in Male and Female Mice,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 8 2005Alexander G Pantschenko Abstract Transgenic mice (Col2.3Bcl-2) with osteoblast-targeted human Bcl-2 expression were established. Phenotypically, these mice were smaller than their wildtype littermates and showed differential effects of the transgene on bone parameters and osteoblast activity dependent on sex. The net effect was an abrogation of sex differences normally observed in wildtype mice and an inhibition of bone loss with age. Ex vivo osteoblast cultures showed that the transgene had no effect on osteoblast proliferation, but decreased bone formation. Estrogen was shown to stimulate endogenous Bcl-2 message levels. These studies suggest a link between Bcl-2 and sex regulation of bone development and age-related bone loss. Introduction: Whereas Bcl-2 has been shown to be an important regulator of apoptosis in development, differentiation, and disease, its role in bone homeostasis and development is not well understood. We have previously showed that the induction of glucocorticoid-induced apoptosis occurred through a dose-dependent decrease in Bcl-2. Estrogen prevented glucocorticoid-induced osteoblast apoptosis in vivo and in vitro by preventing the decrease in Bcl-2 in osteoblasts. Therefore, Bcl-2 may be an important regulator of bone growth through mechanisms that control osteoblast longevity and function. Materials and Methods: Col2.3Bcl-2 mice were developed carrying a 2.3-kb region of the type I collagen promoter driving 1.8 kb of human Bcl-2 (hBcl-2). Tissue specific expression of hBcl-2 in immunoassays validated the transgenic animal model. Histomorphometry and DXA were performed. Proliferation, mineralization, and glucocorticoid-induced apoptosis were examined in ex vivo cultures of osteoblasts. The effect of estrogen on mouse Bcl-2 in ex vivo osteoblast cultures was assayed by RT-PCR and Q-PCR. Results and Conclusions: Two Col2.3Bcl-2 (tg/+) founder lines were established and appeared normal except that they were smaller than their nontransgenic wildtype (+/+) littermates at 1, 2, and 6 months of age, with the greatest differences at 2 months. Immunohistochemistry showed hBcl-2 in osteoblasts at the growth plate and cortical surfaces. Nontransgenic littermates were negative. Western blots revealed hBcl-2 only in type I collagen-expressing tissues. Histomorphometry of 2-month-old mice showed a significant decrease in tg/+ calvaria width with no significant differences in femoral trabecular area or cortical width compared with +/+. However, tg/+ males had significantly more trabecular bone than tg/+ females. Female +/+ mice showed increased bone turnover with elevated osteoblast and osteoclast parameters compared with +/+ males. Col2.3Bcl-2 mice did not show such significant differences between sexes. Male tg/+ mice had a 76.5 ± 1.5% increase in ObS/BS with no significant differences in bone formation rate (BFR) or mineral apposition rate (MAR) compared with male +/+ mice. Transgenic females had a significant 48.4 ± 0.1% and 20.1 ± 5.8% decrease in BFR and MAR, respectively, compared with +/+ females. Osteoclast and osteocyte parameters were unchanged. By 6 months, femurs from female and male +/+ mice had lost a significant amount of their percent of trabecular bone compared with 2-month-old mice. There was little to no change in femoral bone in the tg/+ mice with age. Ex vivo cultures of osteoblasts from +/+ and Col2.3Bcl-2 mice showed a decrease in mineralization, no effect on proliferation, and an inhibition of glucocorticoid-induced apoptosis in Col2.3Bcl-2 cultures. Estrogen was shown to increase mouse Bcl-2 transcript levels in osteoblast cultures of wildtype mice, supporting a role for Bcl-2 in the sex-related differences in bone phenotype regulated by estrogen. Therefore, Bcl-2 differentially affected bone phenotype in male and female transgenic mice, altered bone cell activity associated with sex-related differences, and decreased bone formation, suggesting that apoptosis is necessary for mineralization. In addition, Bcl-2 targeted to mature osteoblasts seemed to delay bone development, producing a smaller transgenic mouse compared with wildtype littermates. These studies suggest that expression of Bcl-2 in osteoblasts is important in regulating bone mass in development and in the normal aging process of bone. [source] Possible Roles of Runx1 and Sox9 in Incipient Intramembranous Ossification,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 10 2004Takashi Yamashiro DDS Abstract We evaluated the detailed expression patterns of Runx1 and Sox9 in various types of bone formation, and determined whether Runx1 expression was affected by Runx2 deficiency and Runx2 expression by Runx1 deficiency. Our results indicate that both Runx1 and Sox9 are intensely expressed in the future osteogenic cell compartment and in cartilage. The pattern of Runx1 and Sox9 expression suggests that both genes could potentially be involved in incipient intramembranous bone formation during craniofacial development. Introduction:Runx1, a gene essential for hematopoiesis, contains RUNX binding sites in its promoter region, suggesting possible cross-regulation with Runx2 and potential regulatory roles in bone development. On the other hand, Sox9 is essential for chondrogenesis, and haploinsufficiency of Sox9 leads to premature ossification of the skeletal system. In this study, we studied the possible roles of Runx1 and Sox9 in bone development. Materials and Methods:Runx1, Runx2/Osf2, and Sox9 expression was evaluated by in situ hybridization in the growing craniofacial bones of embryonic day (E)12,16 mice and in the endochondral bone-forming regions of embryonic and postnatal long bones. In addition, we evaluated Runx2/Osf2 expression in the growing face of Runx1 knockout mice at E12.5 and Runx1 expression in Runx2 knockout mice at E14.5. Results:Runx1 and Sox9 were expressed in cartilage, and the regions of expression expanded to the neighboring Runx2 -expressing osteogenic regions. Expression of both Runx1 and Sox9 was markedly downregulated on ossification. Runx1 and Sox9 expression was absent in the regions of endochondral bone formation and in actively modeling or remodeling bone tissues in the long bones as well as in ossified craniofacial bones. Runx2 expression was not affected by gene disruption of Runx1, whereas the expression domains of Runx1 were extended in Runx2,/, mice compared with wildtype mice. Conclusions:Runx1 and Sox9 are specifically expressed in the osteogenic cell compartments in the craniofacial bones and the bone collar of long bones, and this expression is downregulated on terminal differentiation of osteoblasts. Our results suggest that Runx1 may play a role in incipient intramembranous bone formation. [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] Genetically Modified Animal Models as Tools for Studying Bone and Mineral Metabolism,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 6 2004Rachel A Davey Abstract Genetic modification of mice is a powerful tool for the study of bone development and metabolism. This review discusses the advantages and disadvantages of various approaches used in bone-related research and the contributions these studies have made to bone biology. Genetic modification of mice is a powerful tool for the study of bone development and metabolism. This review discusses the advantages and disadvantages of various approaches used in bone-related research and the contributions these studies have made to bone biology. The approaches to genetic modification included in this review are (1) overexpression of genes, (2) global gene knockouts, (3) tissue-specific gene deletion, and (4) gene knock-in models. This review also highlights issues that should be considered when using genetically modified animal models, including the rigorous control of genetic background, use of appropriate control lines, and confirmation of tissue specificity of gene expression where appropriate. This technology provides a unique and powerful way to probe the function of genes and is already revolutionizing our approach to understanding the physiology of bone development and metabolism. [source] Cyclo-Oxygenase 2 Function Is Essential for Bone Fracture Healing,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 6 2002Ann Marie Simon Abstract Despite the molecular and histological similarities between fetal bone development and fracture healing, inflammation is an early phase of fracture healing that does not occur during development. Cyclo-oxygenase 2 (COX-2) is induced at inflammation sites and produces proinflammatory prostaglandins. To determine if COX-2 functions in fracture healing, rats were treated with COX-2-selective nonsteroidal anti-inflammatory drugs (NSAIDs) to stop COX-2-dependent prostaglandin production. Radiographic, histological, and mechanical testing determined that fracture healing failed in rats treated with COX-2-selective NSAIDs (celecoxib and rofecoxib). Normal fracture healing also failed in mice homozygous for a null mutation in the COX-2 gene. This shows that COX-2 activity is necessary for normal fracture healing and confirms that the effects of COX-2-selective NSAIDs on fracture healing is caused by inhibition of COX-2 activity and not from a drug side effect. Histological observations suggest that COX-2 is required for normal endochondral ossification during fracture healing. Because mice lacking Cox2 form normal skeletons, our observations indicate that fetal bone development and fracture healing are different and that COX-2 function is specifically essential for fracture healing. [source] Changes in Bone Density During Childhood and Adolescence: An Approach Based on Bone's Biological OrganizationJOURNAL OF BONE AND MINERAL RESEARCH, Issue 4 2001Frank Rauch Abstract Bone densitometry has great potential to improve our understanding of bone development. However, densitometric data in children rarely are interpreted in light of the biological processes they reflect. To strengthen the link between bone densitometry and the physiology of bone development, we review the literature on physiological mechanisms and structural changes determining bone mineral density (BMD). BMD (defined as mass of mineral per unit volume) is analyzed in three levels: in bone material (BMDmaterial), in a bone's trabecular and cortical tissue compartments (BMDcompartment), and in the entire bone (BMDtotal). BMDmaterial of the femoral midshaft cortex decreases after birth to a nadir in the first year of life and thereafter increases. In iliac trabecular bone, BMDmaterial also increases from infancy to adulthood, reflecting the decrease in bone turnover. BMDmaterial cannot be determined with current noninvasive techniques because of insufficient spatial resolution. BMDcompartment of the femoral midshaft cortex decreases in the first months after birth followed by a rapid increase during the next 2 years and slower changes thereafter, reflecting changes in both relative bone volume and BMDmaterial. Trabecular BMDcompartment increases in vertebral bodies but not at the distal radius. Quantitative computed tomography (QCT) allows for the determination of both trabecular and cortical BMDcompartment, whereas projectional techniques such as dual-energy X-ray absorptiometry (DXA) can be used only to assess cortical BMDcompartment of long bone diaphyses. BMDtotal of long bones decreases by about 30% in the first months after birth, reflecting a redistribution of bone tissue from the endocortical to the periosteal surface. In children of school age and in adolescents, changes in BMDtotal are site-specific. There is a marked rise in BMDtotal at locations where relative cortical area increases (metacarpal bones, phalanges, and forearm), but little change at the femoral neck and midshaft. BMDtotal can be measured by QCT at any site of the skeleton, regardless of bone shape. DXA allows the estimation of BMDtotal at skeletal sites, which have an approximately circular cross-section. The system presented here may help to interpret densitometric results in growing subjects on a physiological basis. [source] Phosphate regulates embryonic endochondral bone developmentJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 3 2009Alena A. Zalutskaya Abstract Phosphate is required for terminal differentiation of hypertrophic chondrocytes during postnatal growth plate maturation. In vitro models of chondrocyte differentiation demonstrate that 7,mM phosphate, a concentration analogous to that of the late gestational fetus, activates the mitochondrial apoptotic pathway in hypertrophic chondrocytes. This raises the question as to whether extracellular phosphate modulates chondrocyte differentiation and apoptosis during embryonic endochondral bone formation. To address this question, we performed investigations in the mouse metatarsal culture model that recapitulates in vivo bone development. Metatarsals were cultured for 4, 8, and 12 days with 1.25 and 7,mM phosphate. Metatarsals cultured with 7,mM phosphate showed a decrease in proliferation compared to those cultured in 1.25,mM phosphate. This decrease in proliferation was accompanied by an early enhancement in hypertrophic chondrocyte differentiation, associated with an increase in FGF18 expression. By 8 days in culture, an increase caspase-9 activation and apoptosis of hypertrophic chondrocytes was observed in the metatarsals cultured in 7,mM phosphate. Immunohistochemical analyses of embryonic bones demonstrated activation of caspase-9 in hypertrophic chondrocytes, associated with vascular invasion. Thus, these investigations demonstrate that phosphate promotes chondrocyte differentiation during embryonic development and implicate a physiological role for phosphate activation of the mitochondrial apoptotic pathway during embryonic endochondral bone formation. J. Cell. Biochem. 108: 668,674, 2009. © 2009 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] Skeletal tissue engineering using embryonic stem cellsJOURNAL OF TISSUE ENGINEERING AND REGENERATIVE MEDICINE, Issue 3 2010Jojanneke M. Jukes Abstract Various cell types have been investigated as candidate cell sources for cartilage and bone tissue engineering. In this review, we focused on chondrogenic and osteogenic differentiation of mouse and human embryonic stem cells (ESCs) and their potential in cartilage and bone tissue engineering. A decade ago, mouse ESCs were first used as a model to study cartilage and bone development and essential genes, factors and conditions for chondrogenesis and osteogenesis were unravelled. This knowledge, combined with data from the differentiation of adult stem cells, led to successful chondrogenic and osteogenic differentiation of mouse ESCs and later also human ESCs. Next, researchers focused on the use of ESCs for skeletal tissue engineering. Cartilage and bone tissue was formed in vivo using ESCs. However, the amount, homogeneity and stability of the cartilage and bone formed were still insufficient for clinical application. The current protocols require improvement not only in differentiation efficiency but also in ESC-specific hurdles, such as tumourigenicity and immunorejection. In addition, some of the general tissue engineering challenges, such as cell seeding and nutrient limitation in larger constructs, will also apply for ESCs. In conclusion, there are still many challenges, but there is potential for ESCs in skeletal tissue engineering. Copyright © 2009 John Wiley & Sons, Ltd. [source] Recent development of small molecular specific inhibitor of protein tyrosine phosphatase 1BMEDICINAL RESEARCH REVIEWS, Issue 4 2007Seokjoon Lee Abstract Protein tyrosine phosphatases (PTPs), a large family of signaling enzymes, play essential roles in intracellular signal transduction by regulating the cellular level of tyrosine phosphorylation to control cell growth and differentiation, metabolism, cell migration, gene transcription, ion-channel activity, immune response, cell apoptosis, and bone development. Among all PTPs, protein tyrosine phosphatase 1B (PTP1B) plays a seminal role in cellular signaling and in many human diseases, including cancer, diabetes, and obesity. Therefore, small molecular inhibitors of PTP1B can be promising drug candidates. Because of the structural homologies in many families of PTPs, it is a challenging task to find inhibitors specific to each PTP. Recent studies suggested that secondary binding pockets or peripheral binding sites around the conserved active site should be exploited to design novel potent and selective PTP1B inhibitors. In this review, we discuss the structural and biological features of small molecular PTP1B-specific inhibitors, with particular emphasis on small molecular inhibitors targeting PTP1B over the other PTPs that have been synthesized in the past 4 years. © 2006 Wiley Periodicals, Inc. Med Res Rev, 27, No. 4, 553,573, 2007 [source] A new bone vascular perfusion compound for the simultaneous analysis of bone and vasculatureMICROSCOPY RESEARCH AND TECHNIQUE, Issue 7 2010Krista L. Sider Abstract Bone is a highly vascular tissue, which plays an important role in bone development and healing. The ability to analyze both the bone and vasculature simultaneously can enhance the understanding of wound healing, development, and disease in bone. At present, analysis methods are limited in their ability to allow for this simultaneous analysis of bone and bone vasculature in three dimensions, without using the most recent dual-energy computed tomography (CT) techniques. In this study, we present a new barium sulfate (BaSO4) radiopaque vascular perfusion compound for performing postmortem microangiography with single-beam microcomputed tomography (microCT), which allows for such simultaneous analysis. This compound differs from currently available contrast mediums due to (1) the high weight-to-volume ratio of BaSO4 achieved, (2) small BaSO4 aggregate size (<5 ,m), (3) minimal additives, and (4) its miscibility with blood and saline. Most notably, it achieves a radiodensity of 2.4× that of cortical bone, with high perfusion of both the arterial and venous systems and the intervening capillary bed, resulting in an in vivo radiodensity that ranges from that of bone to titanium. Our results, verified using a rat femoral gap-healing model, show that the compound is uniquely suited to high-contrast imaging of the vasculature in the presence of undecalcified bone, with a versatility to be used in other tissues. Microsc. Res. Tech., 2010. © 2009 Wiley-Liss, Inc. [source] Body composition and its components in preterm and term newborns: A cross-sectional, multimodal investigationAMERICAN JOURNAL OF HUMAN BIOLOGY, Issue 1 2010Irfan Ahmad A prospective, cross-sectional, observational study in preterm and term infants was performed to compare multimodal measurements of body composition, namely, limb ultrasound, bone quantitative ultrasound, and dual X-ray absorptiometry (DXA). One hundred and two preterm and term infants appropriate for gestational age were enrolled from the newborn nursery and neonatal intensive care unit. Infants were included when they were medically stable, in an open crib, on full enteral feeds and within 1 week of anticipated discharge. Correlations among the various measurements of body composition were performed using standard techniques. A comparison between preterm infant (born at 28,32 weeks) reaching term to term-born infants was performed. Limb ultrasound estimates of cross-sectional areas of lean and fat tissue in a region of tissue (i.e., the leg) were remarkably correlated with regional and whole-body estimates of fat-free mass and fat obtained from DXA suggesting the potential usefulness of muscle ultrasound as an investigative tool for studying aspects of body composition in this fragile population. There was a weak but significant correlation between quantitative ultrasound measurements of bone strength and DXA-derived bone mineral density (BMD). Preterm infants reaching term had significantly lower body weight, length, head circumference, muscle and fat cross-sectional area, bone speed of sound, whole-body and regional lean body mass, fat mass, and BMD compared to term-born infants. Current postnatal care and nutritional support in preterm infants is still unable to match the in-utero environment for optimal growth and bone development. The use of relatively simple bedside, noninvasive body composition measurements may assist in understanding how changes in different components of body composition early in life affect later growth and development. Am. J. Hum. Biol. 2010. © 2009 Wiley-Liss, Inc. [source] Spatiotemporal Localization of VEGF-A Isoforms in the Mouse Postnatal Growth PlateTHE ANATOMICAL RECORD : ADVANCES IN INTEGRATIVE ANATOMY AND EVOLUTIONARY BIOLOGY, Issue 1 2008Kristin D. Evans Abstract Vascular endothelial growth factor (VEGF) is implicated as a key angiogenic factor in the development of endochondral long bone. Several studies have evaluated the role of VEGF in prenatal endochondral bone development, but few have evaluated VEGF postnatally. Growth plates from mice at postnatal ages 14 (P14), 35 (P35), 49 (P49), and 77 (P77) days were examined for differential expression of the primary VEGF-A mRNA isoforms: VEGF 120, VEGF 164, and VEGF 188. VEGF 120 isoform expression was stable across all ages, whereas VEGF 164 had significantly less expression at P35 and P49 and VEGF l88 expression increased with increasing age. The proportion of transcript isoforms expressed at a given age also changed with VEGF 120 being expressed more highly at P35 and P49 than the other two isoforms. Changes in VEGF mRNA isoforms across cell types within the growth plate were assessed by Percoll fractionation of growth plate cells at age P28. Cells of the proliferative and early hypertrophic regions had significantly higher total VEGF mRNA expression relative to the resting and late hypertrophic regions. VEGF protein expression assessed by immunohistochemistry showed variable expression patterns with increasing postnatal age. In contrast, FLK-1 (VEGF Receptor-2) expression was restricted to the hypertrophic region. These results indicate that VEGF continues to play a significant role in endochondral bone development throughout the entire growth phase of postnatal bone development. Anat Rec, 291:6,13, 2007. © 2007 Wiley-Liss, Inc. [source] Study of the Structure of Canine Mesenchymal Stem Cell Osteogenic CultureANATOMIA, HISTOLOGIA, EMBRYOLOGIA, Issue 5 2010M. B. Eslaminejad With 6 figures and 1 table Summary This study was designed to investigate the morphological features of osteogenic cultures that were established from canine marrow derived-mesenchymal stem cells (MSCs). Tripotent canine MSCs were plated in osteogenic conditions for 3 weeks, at the end of which the cultures were observed by light and transmission electron microscopy. Alkaline phosphatase (ALP) activity of the culture was determined during the differentiation period. To assess whether endochondral or intramembranous ossification was involved in MSC bone differentiation, the cultures were explored for cartilage-related gene expression. Multiple nodule-like cell aggregates appeared to form in the osteogenic cultures. These nodules were covered by a periosteum-like layer and osteocyte-like cells of varying morphology were located in lacuna-like cavities within the nodule mass. Furthermore, the bone nodules possessed an abundant matrix in which clearly striated collagen I fibres were arranged in perpendicular bundles. Matrix vesicles involving in matrix mineralization were evident in the nodules. This was in accordance with increased ALP activity in the culture. No expression of cartilage-related genes was observed, which suggested that osteogenesis might occur by intramembranous ossification. In conclusion, canine MSCs could be an appropriate model for studying in vitro bone development. [source] Roles of lipid-soluble vitamins during ontogeny of marine fish larvaeAQUACULTURE RESEARCH, Issue 5 2010Kristin Hamre Abstract The roles of lipid-soluble vitamins during ontogeny of marine fish larvae are a subject topic where only fragments of the whole picture are known. Most of the research has been focussed on the larval requirements and the availability of these vitamins in the live feed organisms used for early-stage larvae, while the function of the vitamins in the larvae themselves is largely unknown. Our knowledge is mostly extrapolated from research on other vertebrates and also in part from juvenile and adult fish. Vitamin A is known to be essential for establishing body and organ axes in vertebrate embryos and interacts with other nutrients such as vitamin D and fatty acids through the steroid/thyroid nuclear hormone receptor family. In marine fish larvae, excess vitamin A stimulates pigmentation, but at the same time induces vertebral deformities. Live feed organisms contain very little vitamin A but marine fish larvae appear to convert carotenoids in Artemia and copepods to vitamin A, while rotifers, which contain little carotenoids, should be enriched with vitamin A. Vitamin E acts as an antioxidant and is important for the protection of marine fish larvae against the oxidation pressure probably present in intensive rearing systems. Vitamin E may also have other roles connected to its modulation of cell and tissue red-ox balance. In marine fish larvae and juveniles, vitamin E has been shown to enhance the symptoms of vitamin C deficiency, while protecting against the oxidative effect of n-3 fatty acids. Vitamin D is important for the modulation of calcium and phosphorus homeostasis and for the development of the vertebrate skeleton. Vitamin K influences bone development and coagulation of the blood. There is little information on vitamins D and K connected to the ontogeny of marine fish larvae. [source] Effect of dietary protein level and source on bone mineralization in ratsBIOFACTORS, Issue 1-4 2004M. A. Gralak Abstract Bone mineralization was studied in rats. Animals were divided into three feeding groups: LCP - diet with 13.5% crude protein in DM (5% of gluten, 10% of casein), HCP - diet with 21.2% CP in DM (8% of gluten, 10% of casein), and LSM - diet based on grain meals and meat-bone meal (21% CP in DM). After 28 days feeding, animals were euthanased by cervical dislocation and femur bones were collected, weighed and kept frozen until analyses. Diets with 21% protein (HCP, LSM) significantly increased weight of femur bones. Despite of the substantially higher ash level (7.1%) in the LSM diet than in the LCP diet (3.4%), rats of both groups had the similar bone concentration of Ca (15.7 ± 1.1 vs. 17.4 ± 1.1 g/kg) and Zn (178.7 ± 7.9 vs. 173.0 ± 8.5 mg/kg). However bone density in LSM rats was significantly higher than in LCP ones. Although rats fed HCP diet had intermediate bone density, the bone concentration of Ca (11.4 ± 0.5 g/kg) and Zn (145.1 ± 2.9 mg/kg) was significantly lower, than in animals fed LCP and LSM diets. This was related to the very wide protein/calcium (37:1 g/g) and protein/zinc (5.3:1 g/mg) ratios in HCP diet. Those ratios were narrowest in the LSM diet: 16.2:1 (CP/Ca) and 2.6:1 (CP/Zn). It can be conluded that protein/mineral ratio in a diet is a very important factor in bone development, besides dietary protein and ash contents itselves. [source] Role of annexin 1 gene expression in mouse craniofacial bone developmentBIRTH DEFECTS RESEARCH, Issue 7 2007Amilcar Sabino Damazo Abstract BACKGROUND: Annexin 1 is a 37-kDa protein that has complex intra- and extracellular effects. To discover whether the absence of this protein alters bone development, we monitored this event in the annexin-A1 null mice in comparison with littermate wild-type controls. METHODS: Radiographic and densitometry methods were used for the assessment of bone in annexin-A1 null mice at a gross level. We used whole-skeleton staining, histological analysis, and Western blotting techniques to monitor changes at the tissue and cellular levels. RESULTS: There were no gross differences in the appendicular skeleton between the genotypes, but an anomalous development of the skull was observed in the annexin-A1 null mice. This was characterized in the newborn annexin-A1 null animals by a delayed intramembranous ossification of the skull, incomplete fusion of the interfrontal suture and palatine bone, and the presence of an abnormal suture structure. The annexin-A1 gene was shown to be active in osteocytes during this phase and COX-2 was abundantly expressed in cartilage and bone taken from annexin-A1 null mice. CONCLUSIONS: Expression of the annexin-A1 gene is important for the normal development of the skull in mice, possibly through the regulation of osteoblast differentiation and a secondary effect on the expression of components of the cPLA2-COX-2 system. Birth Defects Research (Part A), 2007. © 2007 Wiley-Liss, Inc. [source] | ||||||||||||||||