The culture of chick embryo chondrocytes and the control of their differentiated functions in vitro *1I. Characterization of the chondrocyte-specific phenotypes

Capasso, Olga

doi:10.1016/0014-4827(82)90423-2

Cited by 82 publications

(36 citation statements)

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“…As expected, the hypertrophic chondrocytes from subgroup I clones, when replated on plastic dishes after mild digestion of extracellular matrix (Fig. 1), presented the characteristic epithelial-like morphology (5). The cells from the subgroup 2 clones, when replated, presented the same morphology of the starting dedifferentiated cell population (Fig.…”

Section: Based On Their Properties Clones Can Be Divided In Two Subgsupporting

confidence: 74%

“…We observed that most of the colonies had a characteristic chondrocyte phenotype (5) and that the dedifferentiated phenotype was shown only by slightly more than one third of the colonies. The absolute number of the colonies formed in the presence of medium conditioned by hypertrophic chondrocytes was too low to have a statistically significant distribu- tion according to the different phenotype.…”

Section: The Cloning Ej~ciency Of Primary Cultures Is Dramatically Inmentioning

confidence: 77%

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In vitro development of hypertrophic chondrocytes starting from selected clones of dedifferentiated cells.

Quarto

Dozin

Tacchetti

et al. 1990

The Journal of Cell Biology

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Abstract. Single cells from enzymatically dissociated chick embryo tibiae have been cloned and expanded in fresh or conditioned culture media. A cloning efficiency of ,,o13% was obtained using medium conditioned by dedifferentiated chondrocytes. A cloning efficiency of only 1.4% was obtained when conditioned medium from hypertrophic chondrocytes was used, and efficiencies of essentially 0 were found with fresh medium or medium conditioned by J2-3T3 mouse fibroblasts. Cell clones were selected by morphological criteria and clones showing a dedifferentiated phenotype (fibroblast-like) were further characterized. Out of 38 clones analyzed, 17 were able to differentiate to the hypertrophic chondrocyte stage and reconstitute hypertrophic cartilage when placed in the appropriate culture conditions. Cells from these clones expressed the typical markers of chondrocyte differentiation, i.e., type II and type X collagens. Clones not undergoing differentiation continued to express only type I collagen. Hypertrophic chondrocytes from differentiating clones were analyzed at the single cell level by immunofluorescence; all the cells were positive for type X collagen, while ,x,50% of them showed positivity for type II collagen. (6,19,24). Chondrocytes from Hamburger and Hamilton stage 28-30 (17) chick embryo tibiae dedifferentiate when cultured on plastic dishes. These cells, as well as dedifferentiated chondrocytes from another source (4), maintain the dedifferentiated phenotype as long as the conditions for the cell attachment to the plastic are kept. When the same cells are transferred into suspension culture, they resume the chondrocytic phenotype and mature to hypertrophic chondrocytes through a transient stage of cell aggregation (7). This process is accompanied by an increase in the length of the cell cycle and in the number of quiescent and degenerating cells (16), as well as by a modulation in the transcriptional activity of the collagen genes (9). We have recently reported the identification, purification, and characterization of a novel, low molecular weight protein, named Ch21, expressed and secreted by the in vitro differentiating chondrocytes at a late stage of development (11). When dedifferentiated cells are cultured in suspension in the constant presence of ascorbic acid, they organize their own extracellular matrix and develop into a tissue closely resembling calcifying hypertrophic cartilage (28,29).The heterogeneity of the dedifferentiated cell population did not allow us to definitely state whether the cells followed successive stages of differentiation or the culture conditions made a selection of cell subpopulations. To answer this important question we selected clones of dedifferentiated cells to follow the respective behavior of each clone in terms of morphology and collagen phenotype in the different culture conditions. We show that up to 45 % of the cell populations obtained from single cloned dedifferentiated cells are still able to differentiate to hypertrophic chondrocytes and to reconstitute hyp...

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Section: Based On Their Properties Clones Can Be Divided In Two Subgsupporting

confidence: 74%

Section: The Cloning Ej~ciency Of Primary Cultures Is Dramatically Inmentioning

confidence: 77%

In vitro development of hypertrophic chondrocytes starting from selected clones of dedifferentiated cells.

Quarto

Dozin

Tacchetti

et al. 1990

The Journal of Cell Biology

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“…Type II collagen, which represents approximately 85-90% of the total tissue collagen of hyaline cartilage (Burgeson et al 1982), is also present in different amounts in elastic cartilage and fibrocartilage. The presence of Type X collagen is associated mainly with hypertrophic cartilage, either in vitro (Capasso et al 1982;Gibson et al 1982) or in vivo in growth plate (Nerlich et al 1992) or sternal cartilage destined to become mineralized (Gibson et al 1984). Type X collagen has also been identified in the fibrocartilage at the bone insertion site of the Achilles tendon (Fukuta et al 1998).…”

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confidence: 99%

Immunochemical and Mechanical Characterization of Cartilage Subtypes in Rabbit

Naumann

Dennis

Awadallah

et al. 2002

J Histochem Cytochem.

143

124

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S U M M A R YCartilage is categorized into three general subgroups, hyaline, elastic, and fibrocartilage, based primarily on morphologic criteria and secondarily on collagen (Types I and II) and elastin content. To more precisely define the different cartilage subtypes, rabbit cartilage isolated from joint, nose, auricle, epiglottis, and meniscus was characterized by immunohistochemical (IHC) localization of elastin and of collagen Types I, II, V, VI, and X, by biochemical analysis of total glycosaminoglycan (GAG) content, and by biomechanical indentation assay. Toluidine blue staining and safranin-O staining were used for morphological assessment of the cartilage subtypes. IHC staining of the cartilage samples showed a characteristic pattern of staining for the collagen antibodies that varied in both location and intensity. Auricular cartilage is discriminated from other subtypes by interterritorial elastin staining and no staining for Type VI collagen. Epiglottal cartilage is characterized by positive elastin staining and intense staining for Type VI collagen. The unique pattern for nasal cartilage is intense staining for Type V collagen and collagen X, whereas articular cartilage is negative for elastin (interterritorially) and only weakly positive for collagen Types V and VI. Meniscal cartilage shows the greatest intensity of staining for Type I collagen, weak staining for collagens V and VI, and no staining with antibody to collagen Type X. Matching cartilage samples were categorized by total GAG content, which showed increasing total GAG content from elastic cartilage (auricle, epiglottis) to fibrocartilage (meniscus) to hyaline cartilage (nose, knee joint). Analysis of aggregate modulus showed nasal and auricular cartilage to have the greatest stiffness, epiglottal and meniscal tissue the lowest, and articular cartilage intermediate. This study illustrates the differences and identifies unique characteristics of the different cartilage subtypes in rabbits. The results provide a baseline of data for generating and evaluating engineered repair cartilage tissue synthesized in vitro or for post-implantation analysis.

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“…The process is characterized by the differentiation of prechondrogenic mesenchymal cells to resting, proliferating, and hypertrophic chondrocytes (1)(2)(3)(4)(5)(6)(7)(8) and then by the replacement of hypertrophic cartilage by bone, i.e. cartilage calcification, erosion, invasion by blood vessels, and onset of osteogenesis.…”

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confidence: 99%

The Developmentally Regulated Avian Ch21 Lipocalin Is an Extracellular Fatty Acid-binding Protein

Cancedda

Malpeli

Gentili

et al. 1996

Journal of Biological Chemistry

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Ch21, a developmentally regulated extracellular protein expressed in chick embryos and in cultured chondrocytes, was expressed in the baculovirus system, and the recombinant protein was purified to homogeneity by gel-filtration chromatography. Separation of two isoforms was achieved on an ion-exchange column. Previous work had shown that Ch21 belongs to the superfamily of lipocalins, which are transport proteins for small hydrophobic molecules. Studies were performed to identify the Ch21 ligand.By analysis of recombinant Ch21 on native polyacrylamide gel electrophoresis and by Lipidex assay, the binding of fatty acid to the protein was shown and a preferential binding of long-chain unsaturated fatty acids was observed. Both isoforms had the same behavior. Ch21, also present in chicken serum, represents the first extracellular protein able to selectively bind and transport fatty acid in extracellular fluids and serum. We propose to rename the Ch21 protein as extracellular fatty acid-binding protein (Ex-FABP).In the last several years, our laboratory has focused on the cellular and molecular mechanisms that control endochondral bone formation. The process is characterized by the differentiation of prechondrogenic mesenchymal cells to resting, proliferating, and hypertrophic chondrocytes (1-8) and then by the replacement of hypertrophic cartilage by bone, i.e. cartilage calcification, erosion, invasion by blood vessels, and onset of osteogenesis. Hypertrophic chondrocytes play a critical role in the last part of the process. In fact they can undergo further differentiation to osteoblast-like cells both in vitro (9) and, in selected bone regions, in vivo (10 -14) and are essential to the deposition of the first bone on newly synthesized bone sialoprotein-rich matrix ("bone priming").

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The culture of chick embryo chondrocytes and the control of their differentiated functions in vitro *1I. Characterization of the chondrocyte-specific phenotypes

Cited by 82 publications

References 16 publications

In vitro development of hypertrophic chondrocytes starting from selected clones of dedifferentiated cells.

In vitro development of hypertrophic chondrocytes starting from selected clones of dedifferentiated cells.

Immunochemical and Mechanical Characterization of Cartilage Subtypes in Rabbit

The Developmentally Regulated Avian Ch21 Lipocalin Is an Extracellular Fatty Acid-binding Protein

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