Ectodermal inhibition of cartilage differentiation in micromass culture of chick limb bud mesenchyme in relation to gene expression and cell shape

Development ◽  
1989 ◽  
Vol 105 (4) ◽  
pp. 769-777
Author(s):  
B.C. Gregg ◽  
A. Rowe ◽  
P.M. Brickell ◽  
L. Wolpert
Keyword(s):  
Cell Differentiation ◽  
Cell Shape ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Limb Bud ◽  
Type I ◽  
Micromass Culture ◽  
Cartilage Cell ◽  
Cartilage Differentiation ◽  
Mesenchyme Cells

Ectoderm inhibits the formation of cartilage by chick wing bud mesenchyme in micromass culture. This suggests that the pattern of cartilage formation in the limb bud may result from a restriction of cartilage cell differentiation to the limb bud core as cells leave the progress zone. We have used in situ hybridization to investigate whether ectodermal inhibition in micromass culture occurs at the level of gene transcription. We found that ectoderm completely inhibited the accumulation of cartilage-specific type II collagen transcripts in the mesenchyme cells, whilst the level of type I collagen transcripts was unaffected. Morphometric analysis of electron micrographs revealed that inhibition of chondrogenesis in micromass culture was not preceded by cell flattening. In fact, a rounded cell shape was found not to be a prerequisite for cartilage cell differentiation in micromass.

Changes in the patterns of collagens and fibronectin during limb-bud chondrogenesis

Development ◽  
1980 ◽  
Vol 57 (1) ◽  
pp. 51-60
Author(s):  
W. Dessau ◽  
H. Von Der Mark ◽  
K. Von Der Mark ◽  
S. Fischer
Keyword(s):  
Type I Collagen ◽  
Type Ii Collagen ◽  
Limb Bud ◽  
Condensation Process ◽  
Type I ◽  
Cellular Interactions ◽  
Type Ii ◽  
Cell Matrix ◽  
Cartilage Differentiation ◽  
Mesenchyme Cell

The distribution and sequence of appearance of fibronectin and of type-I and type-II collagen in the developing cartilage models of embryonic chick hind-limb buds was studied by immunofluorescence, using specific antibodies directed against these proteins. Fibronectin and type-I collagen are evenly distributed throughout the intercellular space ofthe mesenchyme prior to condensation of core mesenchyme of the limb anlage and formationof the cartilage blastema. With the onset of the condensation process fibronectin and type-I collagen appear to increase in the cartilage blastema compared to the surrounding loose mesenchyme, reaching a maximal density at the time of cartilage differentiation. The latter process is marked by the appearance of type-II collagen in the cartilage blastema. As cartilage differentiation progresses, type-I collagen is gradually replaced by type-II collagen; fibronectin disappears and is completely absent from mature cartilage. The transient appearance of type-I collagen and fibronectin suggests a temporal role in cell-matrix or cell-cell interactions in chondrogenesis, since it had been shown that(a) type-I collagen substrates stimulate cell proliferation and cartilage differentiation in limb-bud mesenchyme cell cultures; (b) fibronectin mediates attachment of cells to collagen substrates; and (c) fibronectin is directly involved in cellular interactions in chondrocyte cultures.

In situ hybridization reveals differential spatial distribution of mRNAs for type I and type II collagen in the chick limb bud

Development ◽  
1988 ◽  
Vol 103 (1) ◽  
pp. 111-118 ◽  
Author(s):  
C.J. Devlin ◽  
P.M. Brickell ◽  
E.R. Taylor ◽  
A. Hornbruch ◽  
R.K. Craig ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Limb Development ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Limb Bud ◽  
Type I ◽  
Type Ii ◽  
Mrna Accumulation ◽  
Rna Probes

During limb development, type I collagen disappears from the region where cartilage develops and synthesis of type II collagen, which is characteristic of cartilage, begins. In situ hybridization using antisense RNA probes was used to investigate the spatial localization of type I and type II collagen mRNAs. The distribution of the mRNA for type II collagen corresponded well with the pattern of type II collagen synthesis, suggesting control at the level of transcription and mRNA accumulation. In contrast, the pattern of mRNA for type I collagen remained more or less uniform and did not correspond with the synthesis of the protein, suggesting control primarily at the level of translation or of RNA processing.

scholarly journals Chondrogenic Potential of Mouse Calvarial Mesenchyme

2005 ◽  
Vol 53 (5) ◽  
pp. 653-663 ◽  
Author(s):  
Thomas Åberg ◽  
Ritva Rice ◽  
David Rice ◽  
Irma Thesleff ◽  
Janna Waltimo-Sirén
Keyword(s):  
Type I Collagen ◽  
Type Ii Collagen ◽  
Type I ◽  
Embryonic Mouse ◽  
Chondrogenic Potential ◽  
Intramembranous Bone ◽  
Bone And Cartilage Development ◽  
Mesenchyme Cells ◽  
Normal Differentiation

Facial and calvarial bones form intramembranously without a cartilagenous model; however, cultured chick calvarial mesenchyme cells may differentiate into both osteoblasts and chondroblasts and, in rodents, small cartilages occasionally form at the sutures in vivo. Therefore, we wanted to investigate what factors regulate normal differentiation of calvarial mesenchymal cells directly into osteoblasts. In embryonic mouse heads and in cultured tissue explants, we analyzed the expression of selected transcription factors and extracellular matrix molecules associated with bone and cartilage development. Cartilage markers Sox9 and type II collagen were expressed in all craniofacial cartilages. In addition, Msx2 and type I collagen were expressed in sense capsule cartilages. We also observed that the undifferentiated calvarial mesenchyme and the osteogenic fronts in the jaw expressed Co∗∗∗l2A1. Moreover, we found that cultured mouse calvarial mesenchyme could develop into cartilage. Of the 49 explants that contained mesenchyme, intramembranous ossification occurred in 35%. Only cartilage formed in 4%, and both cartilage and bone formed in 4%. Our study confirms that calvarial mesenchyme, which normally gives rise to intramembranous bone, also has chondrogenic potential.

scholarly journals Versican contributes to ligament formation of knee joints

PLoS ONE ◽  
2021 ◽  
Vol 16 (4) ◽  
pp. e0250366
Author(s):  
Tomoko Higuchi ◽  
Daisuke Suzuki ◽  
Takafumi Watanabe ◽  
Kanda Fanhchaksai ◽  
Keiko Ota ◽  
...  
Keyword(s):  
Knee Joint ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Perinatal Period ◽  
Conditional Knockout ◽  
Limb Bud ◽  
Knee Joints ◽  
Type I ◽  
Cruciate Ligaments ◽  
Joint Function

Versican is a large proteoglycan in the extracellular matrix. During embryonic stages, it plays a crucial role in the development of cartilage, heart, and dermis. Previously, we reported that Prx1-Vcan conditional knockout mice, lacking Vcan expression in mesenchymal condensation areas of the limb bud, show the impaired joint formation and delayed cartilage development. Here, we investigated their phenotype in adults and found that they develop swelling of the knee joint. Histologically, their newborn joint exhibited impaired formation of both anterior and posterior cruciate ligaments. Immunostaining revealed a decrease in scleraxis-positive cells in both articular cartilage and ligament of Prx1-Vcan knee joint, spotty patterns of type I collagen, and the presence of type II collagen concomitant with the absence of versican expression. These results suggest that versican expression during the perinatal period is required for cruciate ligaments’ formation and that its depletion affects joint function in later ages.

scholarly journals Transcriptomics Study to Determine the Molecular Mechanism by which sIL-13Rα2-Fc Inhibits Caudal Intervertebral Disc Degeneration in Rats

2020 ◽  
Vol 2020 ◽  
pp. 1-13
Author(s):  
Xin Wang ◽  
Jianshi Tan ◽  
Junhao Sun ◽  
Pengzhong Fang ◽  
Jinlei Chen ◽  
...  
Keyword(s):  
Intervertebral Disc ◽  
Disc Degeneration ◽  
Intervertebral Disc Degeneration ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Type I ◽  
Model Group ◽  
Expression Levels ◽  
Protein Levels ◽  
Collagen Protein

Background. Intervertebral disc degeneration is related to tissue fibrosis. ADAMTS can degrade the important components of the ECM during the process of intervertebral disc degeneration, ultimately resulting in the loss of intervertebral disc function. sIL-13Rα2-Fc can inhibit fibrosis and slow down the degeneration process, but the mechanism involved remains unclear. Objective. To determine the mechanism by which sIL-13Rα2-Fc inhibits ECM degradation and reduces intervertebral disc tissue fibrosis using a transcriptomics analysis. Methods. A rat model of caudal intervertebral disc degeneration was established, and Sirius red staining was used to observe the pathological changes in the caudal intervertebral disc. Transcriptome sequencing was employed to assess the gene expression profiles of the intervertebral disc tissues in the model group and the sIL-13Rα2-Fc-treated group. Differentially expressed genes were identified and analyzed using GO annotation and KEGG pathway analyses. Real-time fluorescence quantitative PCR was used to verify the expression levels of candidate genes. The levels of GAG and HA were quantitatively assessed by ELISA, and the levels of collagen I and collagen II were analyzed by western blotting. Results. Sirius red staining showed that in the model group, the annulus fibrosus was disordered, the number of breaks increased, and the type I collagen protein levels increased, whereas in the sIL-13Rα2-Fc group, the annulus fibrosus was ordered, the number of breaks decreased, and the type II collagen protein levels increased. In comparison with the model group, we identified 58 differentially expressed genes in the sIL-13Rα2-Fc group, and these were involved in 35 signaling pathways. Compared with those in the model group, the mRNA expression levels of Rnux1, Sod2, and Tnfaip6 in the IL-13Rα2-Fc group were upregulated, and the mRNA expression levels of Aldh3a1, Galnt3, Fgf1, Celsr1, and Adamts8 were downregulated; these results were verified by real-time fluorescence quantitative PCR. TIMP-1 (an ADAMTS inhibitor) and TIMP-1 combined with the sIL-13Rα2-Fc intervention increased the levels of GAG and HA, inhibited the expression of type I collagen, and promoted the expression of type II collagen. Conclusion. Adamts8 may participate in the degradation of ECM components such as GAG and HA and lead to an imbalance in the ECM of the intervertebral disc, resulting in intervertebral disc degeneration. sIL-13Rα2-Fc promoted anabolism of the ECM and increased the levels of ECM components by inhibiting the expression of Adamts8, thus maintaining the dynamic equilibrium of the ECM and ultimately delaying intervertebral disc degeneration.

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)

Type I collagen induces mesenchymal cell differentiation into myofibroblasts through YAP-induced TGF-β1 activation

Biochimie ◽  
2018 ◽  
Vol 150 ◽  
pp. 110-130 ◽  
Author(s):  
Xiaoling Liu ◽  
Xinyu Long ◽  
Weiwei Liu ◽  
Yeli Zhao ◽  
Toshihiko Hayashi ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
Type I Collagen ◽  
Mesenchymal Cell ◽  
Type I ◽  
Tgf Β1

Construction of Biomimetic Environments with A Synthetic Peptide Analogue of Collagen.

1998 ◽  
Vol 530 ◽  
Author(s):  
Rajendra S. Bhatnagar ◽  
Jing Jing Qian ◽  
Anna Wedrychowska ◽  
Nancy Smith
Keyword(s):  
Cell Differentiation ◽  
Synthetic Peptide ◽  
Type I Collagen ◽  
Peptide Ligand ◽  
Type I ◽  
Surrounding Matrix ◽  
Collagen Receptors ◽  
Specific Receptors ◽  
Tractional Forces ◽  
And Migration

AbstractThe flow of chemical and mechanical signals among cells, and between cells and their environment plays a crucial role in cell differentiation and morphogenesis. In tissues, type I collagen serves as the template for cell anchorage and migration, and it mediates the flux of regulatory signals via highly specific receptors. Cells respond to mechanical cues by secreting growth factors and remodeling their surrounding matrix in an exquisitely orchestrated spatial and temporal program of matrix turnover and organization. Cellular tractional forces contribute to the organization and orientation of the newly synthesized matrix, establishing the template for subsequent morphogenesis. The junction between cells and collagen plays a key role in cell differentiation, morphogenesis and tissue remodeling. An optimal biomimetic environment would emulate this pathway for the exchange of stimuli. To achieve this goal, we have constructed templates which place cells in apposition to P-15, a synthetic peptide ligand for collagen receptors. These environments prompted 3-D colony formation, induced increased osteogenic differentiation, and the deposition of highly oriented and organized matrix by human dermal and gingival fibroblasts and by osteoblast like HOS cells. These observations support our concept for biomimetic environments for tissue engineering.

Characterizing the effect of substrate stiffness on neural stem cell differentiation

2012 ◽  
Vol 1498 ◽  
pp. 47-52
Author(s):  
Colleen T. Curley ◽  
Kristen Fanale ◽  
Sabrina S. Jedlicka
Keyword(s):  
Stem Cells ◽  
Cell Differentiation ◽  
Neural Stem Cells ◽  
Cortical Neurons ◽  
Type I Collagen ◽  
Stem Cell Differentiation ◽  
Synaptic Proteins ◽  
Substrate Stiffness ◽  
Type I ◽  
Polyacrylamide Gels

ABSTRACTDifferentiated neurons (dorsal root ganglia and cortical neurons) have been shown to develop longer neurite extensions on softer materials than stiffer ones, but previous studies do not address the ability of neural stem cells to undergo differentiation as a result of material elasticity. In this study, we investigate neuronal differentiation of C17.2 neural stem cells due to growth on polyacrylamide gels of variable elastic moduli. Neurite growth, synapse formation, and mode of division (asymmetric vs. symmetric) were all assessed to characterize differentiation. C17.2 neural stem cells were seeded onto polyacrylamide gels coated with Type I collagen. The cells were then serum starved over a 14 day period, fixed, and analyzed for biochemical markers of differentiation. For division studies, time-lapse imaging of cells on various substrates was performed during serum withdrawal using the Nikon Biostation. Division events were analyzed using ImageJ to quantify sizes of resulting daughter. Data shows that C17.2 cell differentiation (as dictated by number and type of division events) is dependent upon substrate stiffness, with softer polyacrylamide surfaces (140 Pa) leading to increased populations of neurons and increased neurite length. Our data also indicates that the ability of neural stem cells to express synaptic proteins and develop synapses is dependent upon material elasticity.

scholarly journals Thrombospondin gene expression by endothelial cells in culture is modulated by cell proliferation, cell shape and the substratum

1990 ◽  
Vol 268 (1) ◽  
pp. 225-230 ◽  
Author(s):  
A E Canfield ◽  
R P Boot-Handford ◽  
A M Schor
Keyword(s):  
Gene Expression ◽  
Cell Proliferation ◽  
Endothelial Cells ◽  
Cell Shape ◽  
Type I Collagen ◽  
Three Dimensional ◽  
Type I ◽  
Mrna Levels ◽  
Collagen Gels ◽  
Cells Cultured

Endothelial cells plated on the surface of a two-dimensional substratum (gelatin-coated dishes, dishes coated with native type I collagen or collagen gels) form a cobblestone monolayer at confluence, whereas cells plated within a three-dimensional gel matrix elongate into a sprouting morphology and self-associate into tube-like structures. In this study, we have compared the synthesis of thrombospondin by quiescent endothelial cells displaying (a) the same morphological phenotype (cobblestone) on different substrata (gelatin and collagen) and (b) different morphological phenotypes (cobblestone and sprouting) on the same substratum (collagen). We demonstrate that thrombospondin is a major biosynthetic product of confluent, quiescent cells cultured on dishes coated with either gelatin or collagen, and that the synthesis of this protein is markedly decreased when cells are plated on or in three-dimensional collagen gels. Moreover, we demonstrate that cells plated in gel (sprouting) secrete less thrombospondin than do cells plated on the gel surface (cobblestone). The regulation of thrombospondin synthesis is reversible and occurs at the level of transcription, as steady-state mRNA levels for thrombospondin decrease in a manner comparable with the levels of protein secreted by these cells. We also show that mRNA levels for laminin B2 chains are increased when cells are cultured on and in collagen gels compared with on gelatin-coated dishes, suggesting that the syntheses of thrombospondin and laminin are regulated by different mechanisms. When cells are cultured on gelatin- or collagen-coated dishes, thrombospondin gene expression is directly proportional to the proliferative state of the cultures. By contrast, the synthesis of thrombospondin by cells cultured on collagen gels remains at equally low levels whether they are labelled when they are sparse and rapidly proliferating or when they are confluent and quiescent. Fibronectin synthesis was found to increase with increasing confluency of the cells plated on all three substrata. These results demonstrate that thrombospondin gene expression is modulated by cell shape, cell proliferation and the nature of the substratum used for cell culture.

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