Developmental control of chondrogenesis and osteogenesis (original) (raw)
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Endochondral ossification: How cartilage is converted into bone in the developing skeleton
The International Journal of Biochemistry & Cell Biology, 2008
Endochondral ossification is the process by which the embryonic cartilaginous model of most bones contributes to longitudinal growth and is gradually replaced by bone. During endochondral ossification, chondrocytes proliferate, undergo hypertrophy and die; the cartilage extracellular matrix they construct is then invaded by blood vessels, osteoclasts, bone marrow cells and osteoblasts, the last of which deposit bone on remnants of cartilage matrix. The sequential changes in chondrocyte behaviour are tightly regulated by both systemic factors and locally secreted factors, which act on receptors to effect intracellular signalling and activation of chondrocyte-selective transcription factors. Systemic factors that regulate the behaviour of chondrocytes in growth cartilage include growth hormone and thyroid hormone, and the local secreted factors include Indian hedgehog, parathyroid hormone-related peptide, fibroblast growth factors and components of the cartilage extracellular matrix. Transcription factors that play critical roles in regulation of chondrocyte gene expression under the control of these extracellular factors include Runx2, Sox9 and MEF2C. The invasion of cartilage matrix by the ossification front is dependent on its resorption by members of the matrix metalloproteinase family, as well as the presence of blood vessels and bone-resorbing osteoclasts. This review, which places an emphasis on recent advances and current areas of debate, discusses the complex interactions between cell types and signalling pathways that govern endochondral ossification.
The development of embryonic bone and cartilage in tissue culture
PubMed, 1983
Embryonic chick long bone develops in a series of temporally controlled, cellular events and involves the integration of at least three distinctly different sets of cells: collar osteoblasts, core osteoblasts, and resorptive or osteoclastic cells. The morphology of the long bones is established by the developing cartilage rudiment or model. All of these events seem to be influenced by positional cues. The cultivation of all of these cells and their presumptive progenitor cells potentially allows a detailed analysis of their individual and collective phenotypic traits. Future studies can include how long bones form, how bone-forming and bone-resorbing cells interact, and how osteogenic cells influence each other throughout each stage of their respective developmental lineages.
Journal of Bone and Mineral Research, 2009
Differentiation of hypertrophic chondrocytes to an osteoblast-like phenotype occurs in vivo in the hypertrophic cartilage of chick embryo tibiae underneath early or prospective periosteum and in cartilage around vascular canals. Synthesis of type I collagen by hypertrophic chondrocytes was shown by immunolocalization of the C propeptide. By enzyme cytochemistry it was instead shown that, in vivo, further differentiating hypertrophic chondrocytes express alkaline phosphatase at the time of initial mineral deposition. Evidence that hypertrophic chondrocytes may resume proliferation was obtained by BrdU labeling. A monoclonal antibody (LA5) was isolated and characterized that recognizes a hypertrophic chondrocyte membrane protein. In addition to staining hypertrophic chondrocytes surrounded by a type I1 and type X collagen-stainable matrix, the LA5 antibodies also stained elongated chondrocytes at the cartilagehone collar interface and cells incorporated in the first layer of bone and osteoid matrix.
The Anatomical Record, 1983
Mandibular condyles of fetal mice 19 to 20 days in utero were kept in an organ culture system for up to 10 days. After 2 days in culture the cartilage of the mandibular condyle appeared to have maintained all its inherent structural characteristics, including its various cell layers: chondroprogenitor, chondroblastic, and hypertrophic. After 5 days in culture no chondroblasts could be seen and, instead, the entire cartilage was occupied by hypertrophic chondrocytes. At the same time, the mesenchymal cells at the chondroprogenitor zone differentiated into osteoblasts which produced osteoid. Light microscopic examinations showed that the newly formed osteoid did not stain with acidic toluidine blue or with alcian blue, but stained intensively with the van Gieson stain and with Periodic acid-Schiff (PAS). The osteoid reacted with antibodies against type I collagen but not with antibodies against type I1 collagen, Electron microscopic examinations showed that the mineralization appeared to be associated with collagen fibers in bone rather than with matrix vesicles in the cartilage. The process of bone formation progressed with time and by the 10th day new bone replaced almost the entire cartilage, thus forming an expanded layer of membrane bone. This in vitro system represents an experimental model whereby undifferentiated precursor cells transform into osteoblasts with the subsequent formation of a typical membrane bone.
Tissue Engineering Part B: Reviews, 2009
Cartilage is the first skeletal tissue to be formed during embryogenesis leading to the creation of all mature cartilages and bones, with the exception of the flat bones in the skull. Therefore, errors occurring during the process of chondrogenesis, the formation of cartilage, often lead to severe skeletal malformations such as dysplasias. There are hundreds of skeletal dysplasias, and the molecular genetic etiology of some remains more elusive than of others. Many efforts have aimed at understanding the morphogenetic event of chondrogenesis in normal individuals, of which the main morphogenetic and regulatory mechanisms will be reviewed here. For instance, many signaling molecules that guide chondrogenesis-for example, transforming growth factor-b, bone morphogenetic proteins, fibroblast growth factors, and Wnts, as well as transcriptional regulators such as the Sox family-have already been identified. Moreover, extracellular matrix components also play an important role in this developmental event, as evidenced by the promotion of the chondrogenic potential of chondroprogenitor cells caused by collagen II and proteoglycans like versican. The growing evidence of the elements that control chondrogenesis and the increasing number of different sources of progenitor cells will, hopefully, help to create tissue engineering platforms that could overcome many developmental or degenerative diseases associated with cartilage defects.
Utilization of Bone Morphogenetic Protein Receptors During Chondrocyte Maturation
Journal of Bone and Mineral Research, 2000
USCs more effectively than DN-BMPR-IA. These results imply that the major BMPR required for BMP induction of chondrocyte hypertrophy is BMPR-IB, and that difference between permanent and prehypertrophic chondrocytes is not caused by absence of receptors required for BMP signaling. (J Bone Miner Res 2000;15:1630 -1639) Key words: hypertrophic, bone morphogenetic protein, receptors, chondrocytes, cartilage Recently, we have shown that BMP-2, -4, and -7 are all capable of inducing chondrocyte maturation in chondrocytes from chick embryos that are destined for hypertrophy. (15) Cultures of prehypertrophic upper (cephalic) ster-
Stem Cells, 2007
The study of human embryonic stem cells (hESCs) can provide invaluable insights into the development of numerous human cell and tissue types in vitro. In this study, we addressed the potential of hESCs to undergo chondrogenesis and demonstrated the potential of hESC-derived embryoid bodies (EBs) to undergo a well-defined full-span chondrogenesis from chondrogenic induction to hypertrophic maturation. We compared chondrogenic differentiation of hESCs through EB direct-plating outgrowth system and EB-derived high-density micromass systems under defined serumfree chondrogenic conditions and demonstrated that cell-tocell contact and bone morphogenetic protein 2 (BMP2) treatment enhanced chondrocyte differentiation, resulting in the formation of cartilaginous matrix rich in collagens and proteoglycans. Provision of a high-density three-dimensional (3D) microenvironment at the beginning of differentiation is critical in driving chondrogenesis because increasing EB seeding numbers in the EB-outgrowth system was unable to enhance chondrogenesis. Temporal order of chondrogenic differentiation and hypertrophic maturation indicated by the gene expression profiles of Col 1, Col 2, and Col 10, and the deposition of extracellular matrix (ECM) proteins, proteoglycans, and collagen II and X demonstrated that the in vivo progression of chondrocyte maturation is recapitulated in the hESC-derived EB model system established in this study. Furthermore, we also showed that BMP2 can influence EB differentiation to multiple cell fates, including that of extraembryonic endodermal and mesenchymal lineages in the EB-outgrowth system, but was more committed to driving the chondrogenic cell fate in the EB micromass system. Overall, our findings provide a potential 3D model system using hESCs to delineate gene function in lineage commitment and restriction of chondrogenesis during embryonic cartilage development.
Specification of chondrocytes and cartilage tissues from embryonic stem cells
Osteoarthritis primarily affects the articular cartilage of synovial joints. Cell and/or cartilage replacement is a promising therapy, provided there is access to appropriate tissue and sufficient numbers of articular chondrocytes. Embryonic stem cells (ESCs) represent a potentially unlimited source of chondrocytes and tissues as they can generate a broad spectrum of cell types under appropriate conditions in vitro. Here, we demonstrate that mouse ESC-derived chondrogenic mesoderm arises from a Flk-1 -/Pdgfrα + (F -P + ) population that emerges in a defined temporal pattern following the development of an early cardiogenic F -P + population. Specification of the late-arising F -P + population with BMP4 generated a highly enriched population of chondrocytes expressing genes associated with growth plate hypertrophic chondrocytes. By contrast, specification with Gdf5, together with inhibition of hedgehog and BMP signaling pathways, generated a population of non-hypertrophic chondrocytes that displayed properties of articular chondrocytes. The two chondrocyte populations retained their hypertrophic and non-hypertrophic properties when induced to generate spatially organized proteoglycan-rich cartilage-like tissue in vitro. Transplantation of either type of chondrocyte, or tissue generated from them, into immunodeficient recipients resulted in the development of cartilage tissue and bone within an 8-week period. Significant ossification was not observed when the tissue was transplanted into osteoblast-depleted mice or into diffusion chambers that prevent vascularization. Thus, through stage-specific manipulation of appropriate signaling pathways it is possible to efficiently and reproducibly derive hypertrophic and non-hypertrophic chondrocyte populations from mouse ESCs that are able to generate distinct cartilage-like tissue in vitro and maintain a cartilage tissue phenotype within an avascular and/or osteoblast-free niche in vivo.
Altered cartilage phenotype expressed during intramembranous bone formation
Journal of Bone and Mineral Research, 2009
The sequential phenotypic expression occurring during intramembranous bone formation was investigated using the tooth extraction socket created in rat alveolar bone in vivo. The differential expression of bone extracellular matrix genes, such as collagen I and osteocalcin, was confirmed by RNA transfer blot analysis and in situ hybridization during the active healing period of the bony socket. To clarify the possible involvement of the chondrogenic phenotype during the process of intramembranous bone formation, the expression of cartilage collagen I1 and IX was further examined in this model. It was found that both a,(II) and al(IX) mRNAs were present, but the al(IX) mRNA was a transcript from the downstream start site/promoter, which is a different site in the al(IX) gene from that used in hyaline cartilage. In situ hybridization indicated that the a,(IX) message was expressed by cells associated with bone matrix in the early formation stage. This finding led to the investigation of type IX collagen expression by osteogenic cells isolated from newborn rat calvariae, in which only the truncated form of al(IX) mRNA was indicated by RNA transfer analysis. The expression of collagen I1 and a truncated form of collagen I X may represent an early phenotypic feature of osteoblast differentiation.
Hypertrophic chondrocytes undergo further differentiation in culture
Journal of Cell Biology, 1992
Conditions have been defined for promoting growth and differentiation of hypertrophic chondrocytes obtained in culture starting from chick embryo tibiae. Hypertrophic chondrocytes, grown in suspension culture as described (Castagnola P., G. Moro, F. Descalzi Cancedda, and R. Cancedda. 1986. J. Cell Biol. 102 :2310-2317), when they reached the stage of single cells, were transferred to substrate-dependent culture conditions in the presence of ascorbic acid. Cells showed a change in morphology, became more elongated and flattened, expressed alkaline phosphatase, and eventually mineralized. Type II and X collagen synthesis was halted and replaced by type I collagen synthesis. In addition the cells started to produce and to secrete in large amount a protein with an apparent molecular mass of 82 KD in reducing conditions and 63 KD in unreducing conditions. This protein is soluble in acidic solutions, does not contain oNG bone organogenesis occurs in the embryo by endochondral ossification from undifferentiated mesenchyme. During the early stages of development, mesenchymal cells in the limb buds condense to form a core of differentiated chondrocytes ; osteogenesis starts at the periphery ofthe cartilage core, which is subsequently invaded by blood vessels and replaced by bone marrow and trabecular bone. After birth, similar events take place in the long bone growth plate and at the bone fracture sites. Bone formation and remodeling have been extensively investigated, starting from pioneering work describing the morphological and biochemical changes occurring during early bone formation to more recent studies aimed at the elucidations of the cellular and molecular mechanisms involved (7, 22, 34). It is widely agreed that cells present in a continuous collar surrounding, but separated from the cartilage rudiment, give rise to osteoblasts, i.e., cells responsible for the synthesis and mineralization of the osteoid extracellular matrix. In the past, occasionally and recently more frequently, it has been postulated thatgrowth platehypertrophic chondrocytes might also contribute to the formation of a bone matrix, since in some organ culturesthese cells start to express bone markers. During culture of mouse mandibular condyles, the expression of type I collagen, osteonectin, alkaline phosphatase, osteopontin, and osteocalcin by mature chondrocytes was detected by in situ hybridization (38). A morphological study