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Manisha C Yadav Ana Maria Sper Simão Sonoko Narisawa Carmen Huesa Marc D McKee Colin Farquharson José Luis Millán 《Journal of bone and mineral research》2011,26(2):286-297
Endochondral ossification is a carefully orchestrated process mediated by promoters and inhibitors of mineralization. Phosphatases are implicated, but their identities and functions remain unclear. Alkaline phosphatase (TNAP) plays a crucial role promoting mineralization of the extracellular matrix by restricting the concentration of the calcification inhibitor inorganic pyrophosphate (PPi). Mutations in the TNAP gene cause hypophosphatasia, a heritable form of rickets and osteomalacia. Here we show that PHOSPHO1, a phosphatase with specificity for phosphoethanolamine and phosphocholine, plays a functional role in the initiation of calcification and that ablation of PHOSPHO1 and TNAP function prevents skeletal mineralization. Phospho1?/? mice display growth plate abnormalities, spontaneous fractures, bowed long bones, osteomalacia, and scoliosis in early life. Primary cultures of Phospho1?/? tibial growth plate chondrocytes and chondrocyte‐derived matrix vesicles (MVs) show reduced mineralizing ability, and plasma samples from Phospho1?/? mice show reduced levels of TNAP and elevated plasma PPi concentrations. However, transgenic overexpression of TNAP does not correct the bone phenotype in Phospho1?/? mice despite normalization of their plasma PPi levels. In contrast, double ablation of PHOSPHO1 and TNAP function leads to the complete absence of skeletal mineralization and perinatal lethality. We conclude that PHOSPHO1 has a nonredundant functional role during endochondral ossification, and based on these data and a review of the current literature, we propose an inclusive model of skeletal calcification that involves intravesicular PHOSPHO1 function and Pi influx into MVs in the initiation of mineralization and the functions of TNAP, nucleotide pyrophosphatase phosphodiesterase‐1, and collagen in the extravesicular progression of mineralization. © 2011 American Society for Bone and Mineral Research. 相似文献
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Epiphyseal Chondrocyte Secondary Ossification Centers Require Thyroid Hormone Activation of Indian Hedgehog and Osterix Signaling 下载免费PDF全文
Weirong Xing Shaohong Cheng Jon Wergedal Subburaman Mohan 《Journal of bone and mineral research》2014,29(10):2262-2275
Thyroid hormones (THs) are known to regulate endochondral ossification during skeletal development via acting directly in chondrocytes and osteoblasts. In this study, we focused on TH effects on the secondary ossification center (SOC) because the time of appearance of SOCs in several species coincides with the time when peak levels of TH are attained. Accordingly, micro–computed tomography (µCT) evaluation of femurs and tibias at day 21 in TH‐deficient and control mice revealed that endochondral ossification of SOCs is severely compromised owing to TH deficiency and that TH treatment for 10 days completely rescued this phenotype. Staining of cartilage and bone in the epiphysis revealed that whereas all of the cartilage is converted into bone in the prepubertal control mice, this conversion failed to occur in the TH‐deficient mice. Immunohistochemistry studies revealed that TH treatment of thyroid stimulating hormone receptor mutant (Tshr?/?) mice induced expression of Indian hedgehog (Ihh) and Osx in type 2 collagen (Col2)‐expressing chondrocytes in the SOC at day 7, which subsequently differentiate into type 10 collagen (Col10)/osteocalcin‐expressing chondro/osteoblasts at day 10. Consistent with these data, treatment of tibia cultures from 3‐day‐old mice with 10 ng/mL TH increased expression of Osx, Col10, alkaline phosphatase (ALP), and osteocalcin in the epiphysis by sixfold to 60‐fold. Furthermore, knockdown of the TH‐induced increase in Osx expression using lentiviral small hairpin RNA (shRNA) significantly blocked TH‐induced ALP and osteocalcin expression in chondrocytes. Treatment of chondrogenic cells with an Ihh inhibitor abolished chondro/osteoblast differentiation and SOC formation. Our findings indicate that TH regulates the SOC initiation and progression via differentiating chondrocytes into bone matrix–producing osteoblasts by stimulating Ihh and Osx expression in chondrocytes. © 2014 American Society for Bone and Mineral Research. 相似文献
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Sonoko Narisawa Manisha C Yadav José Luis Millán 《Journal of bone and mineral research》2013,28(7):1587-1598
Functional ablation of tissue‐nonspecific alkaline phosphatase (TNAP) (Alpl?/? mice) leads to hypophosphatasia, characterized by rickets/osteomalacia attributable to elevated levels of extracellular inorganic pyrophosphate, a potent mineralization inhibitor. Osteopontin (OPN) is also elevated in the plasma and skeleton of Alpl?/? mice. Phosphorylated OPN is known to inhibit mineralization, however, the phosphorylation status of the increased OPN found in Alpl?/? mice is unknown. Here, we generated a transgenic mouse line expressing human TNAP under control of an osteoblast‐specific Col1a1 promoter (Col1a1‐Tnap). The transgene is expressed in osteoblasts, periosteum, and cortical bones, and plasma levels of TNAP in mice expressing Col1a1‐Tnap are 10 to 20 times higher than those of wild‐type mice. The Col1a1‐Tnap animals are healthy and exhibit increased bone mineralization by micro–computed tomography (µCT) analysis. Crossbreeding of Col1a1‐Tnap transgenic mice to Alpl?/? mice rescues the lethal hypophosphatasia phenotype characteristic of this disease model. Osteoblasts from [Col1a1‐Tnap] mice mineralize better than nontransgenic controls and osteoblasts from [Col1a1‐Tnap+/?; Alpl?/?] mice are able to mineralize to the level of Alpl+/? heterozygous osteoblasts, whereas Alpl?/? osteoblasts show no mineralization. We found that the increased levels of OPN in bone tissue of Alpl?/? mice are comprised of phosphorylated forms of OPN whereas wild‐type (WT) and [Col1a1‐Tnap+/?; Alpl?/?] mice had both phosphorylated and dephosphorylated forms of OPN. OPN from [Col1a1‐Tnap] osteoblasts were more dephosphorylated than nontransgenic control cells. Titanium dioxide‐liquid chromatography and tandem mass spectrometry analysis revealed that OPN peptides derived from Alpl?/? bone and osteoblasts yielded a higher proportion of phosphorylated peptides than samples from WT mice, and at least two phosphopeptides, p(S174FQVS178DEQY182PDAT186DEDLT191)SHMK and FRIp(S299HELES304S305S306S307)EVN, with one nonlocalized site each, appear to be preferred sites of TNAP action on OPN. Our data suggest that the promineralization role of TNAP may be related not only to its accepted pyrophosphatase activity but also to its ability to modify the phosphorylation status of OPN. 相似文献
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Daniel Toben Ireen Schroeder Thaqif El Khassawna Manav Mehta Jan‐Erik Hoffmann Jan‐Tilmann Frisch Hanna Schell Jasmin Lienau Alessandro Serra Andreas Radbruch Georg N Duda 《Journal of bone and mineral research》2011,26(1):113-124
Fracture healing is a unique biologic process starting with an initial inflammatory response. As in other regenerative processes, bone and the immune system interact closely during fracture healing. This project was aimed at further elucidating how the host immune system participates in fracture healing. A standard closed femoral fracture was created in wild‐type (WT) and recombination activating gene 1 knockout (RAG1?/?) mice lacking the adaptive immune system. Healing was investigated using micro–computed tomography (µCT), biomechanical testing, and histologic and mRNA expression analyses. Biomechanical testing demonstrated a significantly higher torsional moment on days 14 and 21 in the RAG1?/? mice compared to the WT group. µCT evaluation of RAG1?/? specimens showed earlier mineralization and remodeling. Histologically, endochondral ossification and remodeling were accelerated in the RAG1?/? compared with the WT mice. Histomorphometric analysis on day 7 showed a significantly higher fraction of bone and a significantly lower fraction of cartilage in the callus of the RAG1?/? mice than in the WT mice. Endochondral ossification was accelerated in the RAG1?/? mice. Lymphocytes were present during the physiologic repair process, with high numbers in the hematoma on day 3 and during formation of the hard callus on day 14 in the WT mice. Expression of inflammatory cytokines was reduced in the RAG1?/? mice. In contrast, expression of anti‐inflammatory interleukin 10 (IL‐10) was strongly upregulated in RAG1?/? mice, indicating protective effects. This study revealed an unexpected phenotype of enhanced fracture healing in RAG1?/? mice, suggesting detrimental functions of lymphocytes on fracture healing. The shift from proinflammatory to anti‐inflammatory cytokines suggests that immunomodulatory intervention strategies that maximise the regenerative and minimize the destructive effects of inflammation may lead to enhanced fracture repair. © 2011 American Society for Bone and Mineral Research. 相似文献
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Christian Richard Rujuan Huo Rana Samadfam Isabel Bolivar Dengshun Miao Edward M Brown Geoffrey N Hendy David Goltzman 《Journal of bone and mineral research》2010,25(7):1627-1636
We examined parathyroid and skeletal function in 3‐month‐old mice expressing the null mutation for 25‐hydroxyvitamin D–1α‐hydroxylase [1α(OH)ase?/?] and in mice expressing the null mutation for both the 1α(OH)ase and the calcium‐sensing receptor [Casr?/?1α(OH)ase?/?] genes. On a normal diet, all mice were hypocalcemic, with markedly increased parathyroid hormone (PTH), increased trabecular bone volume, increased osteoblast activity, poorly mineralized bone, enlarged and distorted cartilaginous growth plates, and marked growth retardation, especially in the compound mutants. Osteoclast numbers were reduced in the Casr?/?1α(OH)ase?/? mice. On a high‐lactose, high‐calcium, high‐phosphorus “rescue” diet, serum calcium and PTH were normal in the 1α(OH)ase?/? mice but increased in the Casr?/?1α(OH)ase?/? mice with reduced serum phosphorus. Growth plate architecture and mineralization were improved in both mutants, but linear growth of the double mutants remained abnormal. Mineralization of bone improved in all mice, but osteoblast activity and trabecular bone volume remained elevated in the Casr?/?1α(OH)ase?/? mice. These studies support a role for calcium‐stimulated maturation of the cartilaginous growth plate and mineralization of the growth plate and bone and calcium‐stimulated CaSR‐mediated effects on bone resorption. PTH‐mediated bone resorption may require calcium‐stimulated CaSR‐mediated enhancement of osteoclastic activity. © 2010 American Society for Bone and Mineral Research. © 2010 American Society for Bone and Mineral Research 相似文献
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Rebecca Bell Jun Li Elizabeth F. Shewman Jorge O. Galante Brian J. Cole Bernard R. Bach Jr. Karen L. Troy Katalin Mikecz John D. Sandy Anna H. Plaas Vincent M. Wang 《Journal of orthopaedic research》2013,31(10):1540-1548
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Qiuqian Wu MD PhD Di Chen Michael J Zuscik Regis J O'Keefe Randy N Rosier 《Journal of bone and mineral research》2008,23(4):552-563
Ectopic expression of Smurf2 in chondrocytes and perichondrial cells accelerated endochondral ossification by stimulating chondrocyte maturation and osteoblast development through upregulation of β‐catenin in Col2a1‐Smurf2 embryos. The mechanism underlying Smurf2‐mediated morphological changes during embryonic development may provide new mechanistic insights and potential targets for prevention and treatment of human osteoarthritis. Introduction : Our recent finding that adult Col2a1‐Smurf2 mice have an osteoarthritis‐like phenotype in knee joints prompted us to examine the role of Smurf2 in the regulation of chondrocyte maturation and osteoblast differentiation during embryonic endochondral ossification. Materials and Methods : We analyzed gene expression and morphological changes in developing limbs by immunofluorescence, immunohistochemistry, Western blot, skeletal preparation, and histology. A series of markers for chondrocyte maturation and osteoblast differentiation in developing limbs were examined by in situ hybridization. Results : Ectopic overexpression of Smurf2 driven by the Col2a1 promoter was detected in chondrocytes and in the perichondrium/periosteum of 16.5 dpc transgenic limbs. Ectopic Smurf2 expression in cells of the chondrogenic lineage inhibited chondrocyte differentiation and stimulated maturation; ectopic Smurf2 in cells of the osteoblastic lineage stimulated osteoblast differentiation. Mechanistically, this could be caused by a dramatic increase in the expression of β‐catenin protein levels in the chondrocytes and perichondrial/periosteal cells of the Col2a1‐Smurf2 limbs. Conclusions : Ectopic expression of Smurf2 driven by the Col2a1 promoter accelerated the process of endochondral ossification including chondrocyte maturation and osteoblast differentiation through upregulation of β‐catenin, suggesting a possible mechanism for development of osteoarthritis seen in these mice. 相似文献
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Yamasaki A Itabashi M Sakai Y Ito H Ishiwari Y Nagatsuka H Nagai N 《Calcified tissue international》2001,68(1):53-60
The osteosclerotic (oc/oc) mouse, a genetically distinct murine mutation that has a functional defect in its osteoclasts, also has rickets and shows an altered endochondral ossification in the epiphyseal growth plate. The disorder is morphologically characterized by an abnormal extension of hypertrophic cartilage at 10 days after birth, which is later (21 days after birth) incorporated into the metaphyseal woven bone without breakdown of the cartilage matrix following vascular invasion of chondrocyte lacunae. In situ hybridization revealed that the extending hypertrophic chondrocytes expressed type I and type II collagen mRNA, as well as that of type X collagen and that the osteoblasts in the metaphysis expressed type II and type X collagen mRNA, in addition to type I collagen mRNA. The topographic distribution of the signals suggests a possible co-expression of each collagen gene in the individual cells. Immunohistochemically, an overlapping deposition of type I, type II, and type X collagen was observed in both the extending cartilage and metaphyseal bony trabeculae. Such aberrant gene expression and synthesis of collagen indicate that pathologic ossification takes place in the epiphyseal/metaphyseal junction of oc/oc mouse femur in different way than in normal endochondral ossification. This abnormality is probably not due to a developmental disorder in the epiphyseal plate but to the failure in conversion of cartilage into bone, since the epiphyseal plate otherwise appeared normal, showing orderly stratified zones with a proper expression of cartilage-specific genes. 相似文献
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Leptin regulates chondrocyte differentiation and matrix maturation during endochondral ossification 总被引:3,自引:0,他引:3
Kishida Y Hirao M Tamai N Nampei A Fujimoto T Nakase T Shimizu N Yoshikawa H Myoui A 《BONE》2005,37(5):607-621
Leptin has been suggested to mediate a variety of actions, including bone development, via its ubiquitously expressed receptor (Ob-Rb). In this study, we investigated the role of leptin in endochondral ossification at the growth plate. The growth plates of wild-type and ob/ob mice were analyzed. Effects of leptin on chondrocyte gene expression, cell cycle, apoptosis and matrix mineralization were assessed using primary chondrocyte culture and the ATDC5 cell differentiation culture system. Immunohistochemistry and in situ hybridization showed that leptin was localized in prehypertrophic chondrocytes in normal mice and that Ob-Rb was localized in hypertrophic chondrocytes in normal and ob/ob mice. Growth plates of ob/ob mice were more fragile than those of wild-type mice in a mechanical test and were broken easily at the chondro-osseous junction. The growth plates of ob/ob mice showed disturbed columnar structure, decreased type X collagen expression, less organized collagen fibril arrangement, increased apoptosis and premature mineralization. Leptin administration in ob/ob mice led to an increase in femoral and humeral lengths and decrease in the proportional length of the calcified hypertrophic zone to the whole hypertrophic zone. In primary chondrocyte culture, the matrix mineralization in ob/ob chondrocytes was stronger than that of wild-type mice; this mineralization in both types of mice was abolished by the addition of exogenous leptin (10 ng/ml). During ATDC5 cell differentiation culture, exogenous leptin at a concentration of 1-10 ng/ml (equivalent to the normal serum concentration of leptin) altered type X collagen mRNA expression and suppressed apoptosis, cell growth and matrix calcification. In conclusion, we demonstrated that leptin modulates several events associated with terminal differentiation of chondrocytes. Our finding that the growth plates of ob/ob mice were fragile implies a disturbance in the differentiation/maturation process of growth plates due to depletion of leptin signaling in ob/ob mice. These findings suggest that peripheral leptin signaling plays an essential role in endochondral ossification at the growth plate. 相似文献
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Susan W. Volk Shalin R. Shah Arthur J. Cohen Yanjian Wang Becky K. Brisson Laurie K. Vogel Kurt D. Hankenson Sherrill L. Adams 《Calcified tissue international》2014,94(6):621-631
Type III collagen (Col3), a fibril-forming collagen, is a major extracellular matrix component in a variety of internal organs and skin. It is also expressed at high levels during embryonic skeletal development and is expressed by osteoblasts in mature bone. Loss of function mutations in the gene encoding Col3 (Col3a1) are associated with vascular Ehlers–Danlos syndrome (EDS). Although the most significant clinical consequences of this syndrome are associated with catastrophic failure and impaired healing of soft tissues, several studies have documented skeletal abnormalities in vascular EDS patients. However, there are no reports of the role of Col3 deficiency on the murine skeleton. We compared craniofacial and skeletal phenotypes in young (6–8 weeks) and middle-aged (>1 year) control (Col3+/+) and haploinsufficient (Col3+/?) mice, as well as young null (Col3?/?) mice by microcomputed tomography (μCT). Although Col3+/? mice did not have significant craniofacial abnormalities based upon cranial morphometrics, μCT analysis of distal femur trabecular bone demonstrated significant reductions in bone volume (BV), bone volume fraction (BV/TV), connectivity density, structure model index and trabecular thickness in young adult female Col3+/? mice relative to wild-type littermates. The reduction in BV/TV persisted in female mice at 1 year of age. Next, we evaluated the role of Col3 in vitro. Osteogenesis assays revealed that cultures of mesenchymal progenitors collected from Col3?/? embryos display decreased alkaline phosphatase activity and reduced capacity to undergo mineralization. Consistent with this data, a reduction in expression of osteogenic markers (type I collagen, osteocalcin and bone sialoprotein) correlates with reduced bone Col3 expression in Col3+/? mice and with age in vivo. A small but significant reduction in osteoclast numbers was found in Col3+/? compared to Col3+/+ bones. Taken together, these findings indicate that Col3 plays a role in development of trabecular bone through its effects on osteoblast differentiation. 相似文献
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Summary Type X collagen is a significant component of the extracellular matrix of the hypertrophic zone of physeal cartilage, but
its precise role in endochondral ossification has not been determined. The concentration of type X collagen increases in physeal
cartilage in chicks with vitamin D deficiency. The purpose of our study was to determine whether defective endochondral ossification
due to vitamin D deficiency was associated with abnormalities in the distribution of type X collagen in the proximal tibiotarsus
of chicks. To accomplish this, we induced vitamin D deficiency in broiler chicks and sequentially evaluated the pattern of
type X collagen immunoreactivity in the proximal tibiotarsus using a monoclonal antibody specific for chicken type X collagen.
Type X collagen immunoreactivity was present in the matrix of the prehypertrophic zone, hypertrophic zone, cartilage cores
of the primary spongiosa, and within the chondrocytes of the prehypertrophic and early hypertrophic zones in vitamin D-deficient
and D-replete chicks. However, rachitic chicks exhibited two consistent differences in type X collagen immunoreactivity: hypertrophic
chondrocytes in the late hypertrophic zone and primary spongiosa contained intracellular type X collagen; and type X collagen
was concentrated into laminated aggregates in the pericellular and territorial matrices in the late hypertrophic zone and
primary spongiosa. We conclude from these findings that (1) normal serum concentrations of 1,25(OHD)2D3 and 25OHD3 are not required for type X collagen production; (2) type X collagen production does not decrease in the most mature zones
of the physes in chicks with vitamin D deficiency; and (3) newly secreted type X collagen accumulates in the pericellular
and territorial matrices of the late hypertrophic zone and primary spongiosa of rachitic chicks, perhaps because it is not
readily incorporated into the interterritorial matrix. 相似文献
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Zachary M. Working Elizabeth R. Morris Jiun Chiun Chang Ryan F. Coghlan Brian Johnstone Theodore Miclau William A. Horton Chelsea S. Bahney 《Journal of orthopaedic research》2021,39(1):53-62
Currently, there are no standardized methods for quantitatively measuring fracture repair. Physicians rely on subjective physical examinations and qualitative evaluation of radiographs to detect mineralized tissue. Since most fractures heal indirectly through a cartilage intermediate, these tools are limited in their diagnostic utility of early repair. Prior to converting to the bone, cartilage undergoes hypertrophic maturation, characterized by the deposition of a provisional collagen X matrix. The objective of this study was to characterize the kinetics of a novel collagen X biomarker relative to other biological measurements of fracture healing using a murine model of endochondral fracture repair in which a closed, mid‐shaft tibia fracture was created using the classic drop‐weight technique. Serum was collected 5 to 42 days post‐fracture in male and female mice and compared to uninjured controls (n = 8‐12). Collagen X in the serum was quantified using a recently validated ELISA‐based bioassay (“Cxm”)1 and compared to genetic and histological markers of fracture healing and inflammation. We found the Cxm biomarker reliably increased from baseline to a statistically unique peak 14 days post‐fracture that then resolved to pre‐fracture levels by 3 weeks following injury. The shape and timing of the Cxm curve followed the genetic and histological expression of collagen X but did not show a strong correlation with local inflammatory states. Assessment of fracture healing progress is crucial to making correct and timely clinical decisions for patients. This Cxm bioassay represents a minimally invasive, inexpensive technique that could provide reliable information on the biology of the fracture to significantly improve clinical care. 相似文献
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Frieda Chen Ruolin Guo Luisa Moreno Esther Rosenthal Tanya Zappitelli Ralph A Zirngibl Ann Flenniken Marc Grynpas Lucy R Osborne Lee Adamson Janet Rossant Jane E Aubin 《Journal of bone and mineral research》2014,29(6):1412-1423
By using a genome‐wide N‐ethyl‐N‐nitrosourea (ENU)‐induced dominant mutagenesis screen in mice, a founder with low bone mineral density (BMD) was identified. Mapping and sequencing revealed a T to C transition in a splice donor of the collagen alpha1 type I (Col1a1) gene, resulting in the skipping of exon 9 and a predicted 18‐amino acid deletion within the N‐terminal region of the triple helical domain of Col1a1. Col1a1Jrt/+ mice were smaller in size, had lower BMD associated with decreased bone volume/tissue volume (BV/TV) and reduced trabecular number, and furthermore exhibited mechanically weak, brittle, fracture‐prone bones, a hallmark of osteogenesis imperfecta (OI). Several markers of osteoblast differentiation were upregulated in mutant bone, and histomorphometry showed that the proportion of trabecular bone surfaces covered by activated osteoblasts (Ob.S/BS and N.Ob/BS) was elevated, but bone surfaces undergoing resorption (Oc.S/BS and N.Oc/BS) were not. The number of bone marrow stromal osteoprogenitors (CFU‐ALP) was unaffected, but mineralization was decreased in cultures from young Col1a1Jrt/+ versus +/+ mice. Total collagen and type I collagen content of matrices deposited by Col1a1Jrt/+ dermal fibroblasts in culture was ~40% and 30%, respectively, that of +/+ cells, suggesting that mutant collagen chains exerted a dominant negative effect on type I collagen biosynthesis. Mutant collagen fibrils were also markedly smaller in diameter than +/+ fibrils in bone, tendon, and extracellular matrices deposited by dermal fibroblasts in vitro. Col1a1Jrt/+ mice also exhibited traits associated with Ehlers‐Danlos syndrome (EDS): Their skin had reduced tensile properties, tail tendon appeared more frayed, and a third of the young adult mice had noticeable curvature of the spine. Col1a1Jrt/+ is the first reported model of combined OI/EDS and will be useful for exploring aspects of OI and EDS pathophysiology and treatment. © 2014 American Society for Bone and Mineral Research. 相似文献
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Annina Sipola Lotta Seppinen Taina Pihlajaniemi Juha Tuukkanen 《Calcified tissue international》2009,85(5):412-420
Endostatin, a fragment of collagen XVIII, can inhibit vascular endothelial growth factor (VEGF) signaling. VEGF is known to
be crucial for bone development. The aims of this study were to investigate the influences of endostatin on osteoblast behavior
in vitro and the roles of collagen XVIII/endostatin on bone development in vivo. For the in vitro experiments, MC3T3-E1 osteoblasts
were treated with VEGF-A, 2 μg/ml endostatin, 20 μg/ml endostatin, VEGF-A + 2 μg/ml endostatin, or VEGF-A + 20 μg/ml endostatin.
Osteoblast proliferation and matrix mineralization were analyzed. Faxitron, pQCT, and histological analyses were performed
on hindleg bones of transgenic mice overexpressing endostatin (ES-tg) and mice lacking collagen XVIII (Col18a1
−/−) to study bone development in vivo. Treatment of cells with endostatin decreased osteoblast proliferation. Moreover, VEGF-A
together with endostatin (2 μg/ml) decreased osteoblast proliferation and matrix mineralization. In vivo, Col18a1
−/− and ES-tg mice displayed no differences in bone density or mineral content during bone development, but ES-tg bones grew
in length more slowly compared to the controls. The formation of secondary ossification centers was delayed in Col18a1
−/− mice. Immunohistochemistry revealed collagen XVIII in basement membranes of periosteal and bone marrow vessels and at muscle
attachment sites. In conclusion, endostatin affects osteoblast behavior in vitro, the effects being boosted by simultaneous
treatment with VEGF. In vivo, Col18a1
−/− and ES-tg mice show mild delays in bone development. These changes are transitory and suggest that collagen XVIII/endostatin
does not play an indispensable role in skeletal development. 相似文献
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Endochondral bone formation occurs through a series of developmentally regulated cellular stages, from initial formation
of cartilage tissue to calcified cartilage, resorption, and replacement by bone tissue. Nasal cartilage cells isolated by
enzymatic digestion from rat fetuses were seeded at a final density of 105 cell/cm2 and cultured in Dulbecco's modified Eagle medium (DMEM) supplemented with 10% fetal calf serum in the presence of ascorbic
acid and β-glycerophosphate. First, cells lost their phenotype but in this condition they rapidly reexpressed the chondrocyte
phenotype and were able to form calcified cartilaginous nodules with the morphological appearance of cartilage mineralization
that occurs in vivo during endochondral ossification. In this mineralizing chondrocyte culture system, we investigated, between day 3 and day
15, the pattern expression of types II and X collagen, proteoglycan core protein, characteristic markers of chondrocyte differentiation,
as well as alkaline phosphatase and osteocalcin associated with the mineralization process. Analysis of labeled collagen and
immunoblotting revealed type I collagen synthesis associated with the loss of chondrocyte phenotype at the beginning of the
culture. However, our culture conditions promoted extracellular matrix mineralization and cell differentiation towards the
hypertrophic phenotype. This differentiation process was characterized by the induction of type X collagen mRNA, alkaline
phosphatase, and diminished expression of type II collagen and core protein of large proteoglycan after an increase in their
mRNA levels before the mineralizing process. These results revealed distinct switches of the specific molecular markers and
indicated a similar temporal expression to that observed in vivo recapitulating all stages of the differentiation program in vitro.
Received: 12 December 1996 / Accepted: 26 June 1997 相似文献
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Gretl Hendrickx Verena Fischer Astrid Liedert Simon von Kroge Melanie Haffner-Luntzer Laura Brylka Eva Pawlus Michaela Schweizer Timur Yorgan Anke Baranowsky Tim Rolvien Mona Neven Udo Schumacher David J Beech Michael Amling Anita Ignatius Thorsten Schinke 《Journal of bone and mineral research》2021,36(2):369-384