scholarly journals Collagen gene expression during limb cartilage differentiation.

1986 ◽  
Vol 102 (4) ◽  
pp. 1151-1156 ◽  
Author(s):  
R A Kosher ◽  
W M Kulyk ◽  
S W Gay
Keyword(s):  
Gene Expression ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Mesenchymal Cells ◽  
Type I ◽  
Collagen Gene ◽  
Type Ii ◽  
Collagen Mrna ◽  
Cytoplasmic Type

As limb mesenchymal cells differentiate into chondrocytes, they initiate the synthesis of type II collagen and cease synthesizing type I collagen. Changes in the cytoplasmic levels of type I and type II collagen mRNAs during the course of limb chondrogenesis in vivo and in vitro were examined using cloned cDNA probes. A striking increase in cytoplasmic type II collagen mRNA occurs coincident with the crucial condensation stage of chondrogenesis in vitro, in which prechondrogenic mesenchymal cells become closely juxtaposed before depositing a cartilage matrix. Thereafter, a continuous and progressive increase in the accumulation of cytoplasmic type II collagen mRNA occurs which parallels the progressive accumulation of cartilage matrix by cells. The onset of overt chondrogenesis, however, does not involve activation of the transcription of the type II collagen gene. Low levels of type II collagen mRNA are present in the cytoplasm of prechondrogenic mesenchymal cells at the earliest stages of limb development, well before the accumulation of detectable levels of type II collagen. Type I collagen gene expression during chondrogenesis is regulated, at least in part, at the translational level. Type I collagen mRNAs are present in the cytoplasm of differentiated chondrocytes, which have ceased synthesizing detectable amounts of type I collagen.

scholarly journals Immunological and biochemical studies of collagen type transition during in vitro chondrogenesis of chick limb mesodermal cells

1977 ◽  
Vol 73 (3) ◽  
pp. 736-747 ◽  
Author(s):  
K Von Der Mark ◽  
H Von Der Mark
Keyword(s):  
Collagen Synthesis ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Mesenchymal Cells ◽  
Gradual Transition ◽  
Type I ◽  
Type Ii ◽  
Total Collagen ◽  
Collagen Antibodies

This work describes an approach to monitor chondrogenesis of stage-24 chick limb mesodermal cells in vitro by analyzing the onset of type II collagen synthesis with carboxymethyl-cellulose chromatography, immunofluorescence, and radioimmunoassay. This procedure allowed specific and quantitative determination of chondrocytes in the presence of fibroblasts and myoblasts, both of which synthesize type I collagen. Chondrogenesis was studied in high-density cell preparations on tissue culture plastic dishes and on agar base. It was found that stage-24 limb mesenchymal cells initially synthesized only type I collagen. With the onset of chondrogenesis, a gradual transition to type II collagen synthesis was observed. In cell aggregates formed over agar, type II collagen synthesis started after 1 day in culture and reached levels of 80-90 percent of the total collagen synthesis at 6-8 days. At that time, the cells in the center of the aggregates had acquired the typical chondrocyte phenotype and stained only with type II collagen antibodies, whereas the peripheral cells had developed into a "perichondrium" and stained with type I and type II collagen antibodies. On plastic dishes plated with 5 X 10(6) cells per 35mm dish, cartilage nodules developed after 4-6 days, but the type II collagen synthesis only reached levels of 10-20 percent of the total collagen. The majority of the cells differentiated into fibroblasts and myoblasts and synthesized type I collagen. These studies demonstrate that analysis of cell specific types of collagen provides a useful method for detailing the specific events in the differentiation of mesenchymal cells in vitro.

Type VI collagen expression is upregulated in the early events of chondrocyte differentiation

Development ◽  
1993 ◽  
Vol 117 (1) ◽  
pp. 245-251
Author(s):  
R. Quarto ◽  
B. Dozin ◽  
P. Bonaldo ◽  
R. Cancedda ◽  
A. Colombatti
Keyword(s):  
Suspension Culture ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Type I ◽  
Type Ii ◽  
Type Vi Collagen ◽  
Full Spectrum ◽  
Collagen Expression ◽  
Hypertrophic Chondrocyte

Dedifferentiated chondrocytes cultured adherent to the substratum proliferate and synthesize large amounts of type I collagen but when transferred to suspension culture they decrease proliferation, resume the chondrogenic phenotype and the synthesis of type II collagen, and continue their maturation to hypertrophic chondrocyte (Castagnola et al., 1986, J. Cell Biol. 102, 2310–2317). In this report, we describe the developmentally regulated expression of type VI collagen in vitro in differentiating avian chondrocytes. Type VI collagen mRNA is barely detectable in dedifferentiated chondrocytes as long as the attachment to the substratum is maintained, but increases very rapidly upon passage of the cells into suspension culture reaching a peak after 48 hours and declining after 5–6 days of suspension culture. The first evidence of a rise in the mRNA steady-state levels is obtained already at 6 hours for the alpha 3(VI) chain. Immunoprecipitation of metabolically labeled cells with type VI collagen antibodies reveals that the early mRNA rise is paralleled by an increased secretion of type VI collagen in cell media. Induction of type VI collagen is not the consequence of trypsin treatment of dedifferentiated cells since exposure to the actin-disrupting drug cytochalasin or detachment of the cells by mechanical procedures has similar effects. In 13-day-old chicken embryo tibiae, where the full spectrum of the chondrogenic differentiation process is represented, expression of type VI collagen is restricted to the articular cartilage where chondrocytes developmental stage is comparable to stage I (high levels of type II collagen expression).(ABSTRACT TRUNCATED AT 250 WORDS)

Tissue specificity of type I collagen gene expression is determined at both transcriptional and post-transcriptional levels

1984 ◽  
Vol 4 (9) ◽  
pp. 1843-1852
Author(s):  
R J Focht ◽  
S L Adams
Keyword(s):  
Gene Expression ◽  
Rna Structure ◽  
Type I Collagen ◽  
Translational Efficiency ◽  
Type I ◽  
Collagen Gene ◽  
Differentiated Cells ◽  
Collagen Gene Expression

We analyzed the control of type I collagen synthesis in four kinds of differentiated cells from chicken embryos which synthesize very different amounts of the protein. Tendon, skin, and smooth muscle cells were found to have identical amounts of type I collagen RNAs; however, the RNAs had inherently different translatabilities, which were observed both in vivo and in vitro. Chondrocytes also had substantial amounts of type I collagen RNAs, even though they directed no detectable synthesis of the protein either in vivo or in vitro. Type I collagen RNAs in chondrocytes display altered electrophoretic mobilities, suggesting that in these cells the reduction in translational efficiency may be mediated in part by changes in the RNA structure. These data indicate that control of type I collagen gene expression is a complex process which is exerted at both transcriptional and post-transcriptional levels.

Distribution of type I and type II collagen gene expression during the development of human long bones

Bone ◽  
1990 ◽  
Vol 11 (4) ◽  
pp. 275-279 ◽  
Author(s):  
S. Mundlos ◽  
H. Engel ◽  
I. Michel-Behnke ◽  
B. Zabel
Keyword(s):  
Gene Expression ◽  
Type Ii Collagen ◽  
Type I ◽  
Collagen Gene ◽  
Long Bones ◽  
Type Ii ◽  
Collagen Gene Expression

scholarly journals Tissue specificity of type I collagen gene expression is determined at both transcriptional and post-transcriptional levels.

1984 ◽  
Vol 4 (9) ◽  
pp. 1843-1852 ◽  
Author(s):  
R J Focht ◽  
S L Adams
Keyword(s):  
Gene Expression ◽  
Rna Structure ◽  
Type I Collagen ◽  
Translational Efficiency ◽  
Type I ◽  
Collagen Gene ◽  
Differentiated Cells ◽  
Collagen Gene Expression

We analyzed the control of type I collagen synthesis in four kinds of differentiated cells from chicken embryos which synthesize very different amounts of the protein. Tendon, skin, and smooth muscle cells were found to have identical amounts of type I collagen RNAs; however, the RNAs had inherently different translatabilities, which were observed both in vivo and in vitro. Chondrocytes also had substantial amounts of type I collagen RNAs, even though they directed no detectable synthesis of the protein either in vivo or in vitro. Type I collagen RNAs in chondrocytes display altered electrophoretic mobilities, suggesting that in these cells the reduction in translational efficiency may be mediated in part by changes in the RNA structure. These data indicate that control of type I collagen gene expression is a complex process which is exerted at both transcriptional and post-transcriptional levels.

scholarly journals The protein phosphatase inhibitor okadaic acid suppresses type I collagen gene expression in cultured fibroblasts at the transcriptional level

1995 ◽  
Vol 308 (3) ◽  
pp. 995-999 ◽  
Author(s):  
J Westermarck ◽  
E Ilvonen ◽  
V M Kähäri
Keyword(s):  
Gene Expression ◽  
Type I Collagen ◽  
Protein Phosphatases ◽  
Type I ◽  
Collagen Gene ◽  
Mrna Levels ◽  
Collagen Mrna ◽  
Collagen Gene Expression ◽  
Nih 3T3 ◽  
Collagen Promoter

Type I collagen is the most abundant component of the extracellular matrix of human connective tissues. We have examined the effect of okadaic acid (OA), an inhibitor of phosphoserine- and-phosphothreonine-specific protein phosphatases 1 and 2A, on type I collagen gene expression by fibroblasts in culture. Treatment of human skin fibroblasts with OA potently reduced type I and type III collagen mRNA levels, maximally by over 90%. The inhibitory effect of OA on type I and III collagen mRNA abundance was not prevented by cycloheximide, and was not affected by simultaneous treatment with dexamethasone or retinoic acid. OA also abrogated the enhancing effect of transforming growth factor-beta (TGF-beta) on type I and III collagen mRNA levels. Treatment of transiently transfected NIH-3T3 fibroblasts with OA suppressed the activity of a 3.5 kb human pro alpha 2(I) collagen promoter/chloramphenicol acetyltransferase construct maximally, by 70%. In addition, OA treatment of NIH-3T3 cells abrogated enhancement of pro alpha 2(I) collagen promoter activity by TGF-beta. These results indicate that protein phosphatases 1 and 2A have an important role as positive regulators of type I and III collagen gene expression. The results also suggest that selective inhibition of activity of protein phosphatases 1 and 2A may offer a novel approach for preventing excessive collagen accumulation in fibrotic disorders.

scholarly journals Synthesis of cartilage matrix by mammalian chondrocytes in vitro. III. Effects of ascorbate.

1984 ◽  
Vol 99 (6) ◽  
pp. 1960-1969 ◽  
Author(s):  
J C Daniel ◽  
B U Pauli ◽  
K E Kuettner
Keyword(s):  
Extracellular Matrix ◽  
Type I Collagen ◽  
Phenotypic Expression ◽  
Type Ii Collagen ◽  
Ruthenium Red ◽  
Type I ◽  
Type Ii ◽  
Morphological Examination ◽  
Associated Matrix

Chondrocytes isolated from bovine articular cartilage were plated at high density and grown in the presence or absence of ascorbate. Collagen and proteoglycans, the major matrix macromolecules synthesized by these cells, were isolated at times during the course of the culture period and characterized. In both control and ascorbate-treated cultures, type II collagen and cartilage proteoglycans accumulated in the cell-associated matrix. Control cells secreted proteoglycans and type II collagen into the medium, whereas with time in culture, ascorbate-treated cells secreted an increasing proportion of types I and III collagens into the medium. The ascorbate-treated cells did not incorporate type I collagen into the cell-associated matrix, but continued to accumulate type II collagen in this compartment. Upon removal of ascorbate, the cells ceased to synthesize type I collagen. Morphological examination of ascorbate-treated and control chondrocyte culture revealed that both collagen and proteoglycans were deposited into the extracellular matrix. The ascorbate-treated cells accumulated a more extensive matrix that was rich in collagen fibrils and ruthenium red-positive proteoglycans. This study demonstrated that although ascorbate facilitates the formation of an extracellular matrix in chondrocyte cultures, it can also cause a reversible alteration in the phenotypic expression of those cells in vitro.

scholarly journals Nuclear factor binding to an AP-1 site is associated with the activation of pro-α 1(I)-collagen gene in dedifferentiating chondrocytes

1993 ◽  
Vol 294 (2) ◽  
pp. 365-371 ◽  
Author(s):  
A Määttä ◽  
V Glumoff ◽  
P Paakkonen ◽  
D Liska ◽  
R P K Penttinen ◽  
...  
Keyword(s):  
Gene Expression ◽  
Type I Collagen ◽  
Specific Interaction ◽  
Binding Activity ◽  
Type I ◽  
Collagen Gene ◽  
Type Ii ◽  
Mobility Shift ◽  
First Intron ◽  
Fetal Fibroblasts

Isolated chondrocytes grown on plastic gradually lose their differentiated phenotype upon subculturing. This dedifferentiation is manifested by an altered production of extracellular-matrix molecules (ECM): e.g., the cartilage specific type II collagen is replaced by types I and III. We have studied the regulation of ECM gene expression in dedifferentiating human and murine fetal chondrocytes. Nuclear extracts from dedifferentiated cells, human fetal fibroblasts and 3T3 cells contained a protein that bound in an electrophoretic mobility shift assay to an AP-1 site in the first intron of the human alpha 1(I) collagen gene. This binding activity was not present in freshly isolated human or murine chondrocytes, which produced type II, but not type I, collagen mRNA in culture. Thus the binding activity was induced simultaneously with alpha 1(I)-collagen-gene expression during dedifferentiation. The specific interaction was sensitive to dephosphorylation of the nuclear extract and to chemical modification of reduced cysteine residues. The AP-1 site we studied had previously been shown to be a positive transcriptional contributor in the first intron to the expression of the alpha 1(I) collagen gene. In transient transfections into dedifferentiating chondrocytes, an alpha 1(I) collagen expression plasmid carrying a mutated AP-1 site in the first intron resulted in three-times-lower reporter gene RNA levels than a plasmid carrying the respective functional AP-1 site. These data suggest that the AP-1 sequence and its respective trans-acting factors may play a role in the transcriptional regulation of the alpha 1(I) collagen gene during dedifferentiation of chondrocytes.

scholarly journals Efficacy of collagen and alginate hydrogels for the prevention of rat chondrocyte dedifferentiation

2018 ◽  
Vol 9 ◽  
pp. 204173141880243 ◽  
Author(s):  
Guang-Zhen Jin ◽  
Hae-Won Kim
Keyword(s):  
Tissue Engineering ◽  
Type I Collagen ◽  
Cartilage Tissue Engineering ◽  
Collagen Gel ◽  
Type Ii Collagen ◽  
Cartilage Tissue ◽  
Type I ◽  
Type Ii ◽  
Chondrogenic Phenotype

Dedifferentiation of chondrocytes remains a major problem in cartilage tissue engineering. The development of hydrogels that can preserve chondrogenic phenotype and prevent chondrocyte dedifferentiation is a meaningful strategy to solve dedifferentiation problem of chondrocytes. In the present study, three gels were prepared (alginate gel (Alg gel), type I collagen gel (Col gel), and their combination gel (Alg/Col gel)), and the in vitro efficacy of chondrocytes culture while preserving their phenotypes was investigated. While Col gel became substantially contracted with time, the cells encapsulated in Alg gel preserved the shape over the culture period of 14 days. The mechanical and cell-associated contraction behaviors of Alg/Col gel were similar to those of Alg. The cells in Alg and Alg/Col gels exhibited round morphology, whereas those in Col gel became elongated (i.e. fibroblast-like) during cultures. The cells proliferated with time in all gels with the highest proliferation being attained in Col gel. The expression of chondrogenic genes, including SOX9, type II collagen, and aggrecan, was significantly up-regulated in Alg/Col gel and Col gel, particularly in Col gel. However, the chondrocyte dedifferentiation markers, type I collagen and alkaline phosphatase ( ALP), were also expressed at significant levels in Col gel, which being contrasted with the events in Alg and Alg/Col gels. The current results suggest the cells cultured in hydrogels can express chondrocyte dedifferentiation markers as well as chondrocyte markers, which draws attention to choose proper hydrogels for chondrocyte-based cartilage tissue engineering.

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