scholarly journals Hypertrophic chondrocytes undergo further differentiation in culture

1992 ◽  
Vol 117 (2) ◽  
pp. 427-435 ◽  
Author(s):  
F Descalzi Cancedda ◽  
C Gentili ◽  
P Manduca ◽  
R Cancedda
Keyword(s):  
Collagen Synthesis ◽  
Type I Collagen ◽  
Bone Cells ◽  
Single Cells ◽  
Protein A ◽  
Culture Conditions ◽  
Hypertrophic Chondrocytes ◽  
Calcium Deposition ◽  
Type I ◽  
Collagen Secretion

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 collagenous domains, and is glycosylated. The Ch21 protein, a marker of hypertrophic chondrocytes and bone cells, was synthesized throughout the culture. We have defined this additional differentiation stage as an osteoblast-like stage. Calcium deposition in the extracellular matrix occurred regardless of the addition of beta glycerophosphate to the culture medium. Comparable results were obtained both when the cells were plated at low density and when they were already at confluence and maintained in culture without passaging up to 50 d. When retinoic acid was added to the hypertrophic chondrocyte culture between day 1 and day 5 the maturation of the cells to the osteoblast-like stage was highly accelerated. The switch in the collagen secretion was already observed after 2 d and the production of the 63-kD protein after 3 d. Mineralization was observed after 15-20 d.

scholarly journals Thyroid hormone, insulin, and glucocorticoids are sufficient to support chondrocyte differentiation to hypertrophy: a serum-free analysis.

1992 ◽  
Vol 119 (4) ◽  
pp. 989-995 ◽  
Author(s):  
R Quarto ◽  
G Campanile ◽  
R Cancedda ◽  
B Dozin
Keyword(s):  
Collagen Synthesis ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Hypertrophic Chondrocytes ◽  
Type I ◽  
Type Ii ◽  
Type X Collagen ◽  
Serum Free ◽  
Low Levels ◽  
Complete Cell

Chondrocytes from chicken embryo tibia can be maintained in culture as adherent cells in Coon's modified Ham's F-12 medium supplemented with 10% FCS. In this condition, they dedifferentiate, losing type II collagen expression in favor of type I collagen synthesis. Their differentiation to hypertrophy can be obtained by transferring them to suspension culture. Differentiation is evidenced by the shift from type I to type II and type IX collagen synthesis and the following predominant expression of type X collagen, all markers of specific stages of the differentiation process. To identify the factors required for differentiation, we developed a serum-free culture system where only the addition of triiodothyronine (T3; 10(-11) M), insulin (60 ng/ml), and dexamethasone (10(-9) M) to the F-12 medium was sufficient to obtain hypertrophic chondrocytes. In this hormonal context, chondrocytes display the same changes in the pattern of protein synthesis as described above. For proper and complete cell maturation, T3 and insulin concentrations cannot be modified. Insulin cannot be substituted by insulin-like growth factor-I, but dexamethasone concentration can be decreased to 10(-12) M without chondrogenesis being impaired. In the latter case, the expression of type X collagen and its mRNA are inversely proportional to dexamethasone concentration. When ascorbic acid is added to the hormone-supplemented medium, differentiating chondrocytes organize their matrix leading to a cartilage-like structure with hypertrophic chondrocytes embedded in lacunae. However, this structure does not present detectable calcification, at variance with control cultures maintained in FCS. Accordingly, in the presence of the hormone mixture, the differentiating chondrocytes have low levels of alkaline phosphatase activity. This report indicates that T3 and insulin are primary factors involved in the onset and progression of chondrogenesis, while dexamethasone supports cell viability and modulates some differentiated functions.

Melatonin stimulates proliferation and type I collagen synthesis in human bone cells in vitro

1999 ◽  
Vol 27 (2) ◽  
pp. 106-110 ◽  
Author(s):  
Osamu Nakade ◽  
Hiroki Koyama ◽  
Hirohiko Ariji ◽  
Akihiro Yajima ◽  
Tohru Kaku
Keyword(s):  
Collagen Synthesis ◽  
Type I Collagen ◽  
Bone Cells ◽  
Human Bone ◽  
Type I

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

1990 ◽  
Vol 110 (4) ◽  
pp. 1379-1386 ◽  
Author(s):  
R Quarto ◽  
B Dozin ◽  
C Tacchetti ◽  
G Campanile ◽  
C Malfatto ◽  
...  
Keyword(s):  
Type I Collagen ◽  
Culture Media ◽  
Single Cells ◽  
Type Ii Collagen ◽  
Hypertrophic Chondrocytes ◽  
Type I ◽  
Type Ii ◽  
Cloning Efficiency ◽  
Morphological Criteria

Single cells from enzymatically dissociated chick embryo tibiae have been cloned and expanded in fresh or conditioned culture media. A cloning efficiency of approximately 13% 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 approximately 50% of them showed positivity for type II collagen.

scholarly journals Autophagy facilitates type I collagen synthesis in periodontal ligament cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Tomomi Nakamura ◽  
Motozo Yamashita ◽  
Kuniko Ikegami ◽  
Mio Suzuki ◽  
Manabu Yanagita ◽  
...  
Keyword(s):  
Periodontal Ligament ◽  
Collagen Synthesis ◽  
Type I Collagen ◽  
Periodontal Diseases ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Physiological Role ◽  
Type I ◽  
Periodontal Tissue ◽  
Pdl Cells

AbstractAutophagy is a lysosomal protein degradation system in which the cell self-digests its intracellular protein components and organelles. Defects in autophagy contribute to the pathogenesis of age-related chronic diseases, such as myocardial infarction and rheumatoid arthritis, through defects in the extracellular matrix (ECM). However, little is known about autophagy in periodontal diseases characterised by the breakdown of periodontal tissue. Tooth-supportive periodontal ligament (PDL) tissue contains PDL cells that produce various ECM proteins such as collagen to maintain homeostasis in periodontal tissue. In this study, we aimed to clarify the physiological role of autophagy in periodontal tissue. We found that autophagy regulated type I collagen synthesis by elimination of misfolded proteins in human PDL (HPDL) cells. Inhibition of autophagy by E-64d and pepstatin A (PSA) or siATG5 treatment suppressed collagen production in HPDL cells at mRNA and protein levels. Immunoelectron microscopy revealed collagen fragments in autolysosomes. Accumulation of misfolded collagen in HPDL cells was confirmed by sodium dodecyl sulfate–polyacrylamide gel electrophoresis. E-64d and PSA treatment suppressed and rapamycin treatment accelerated the hard tissue-forming ability of HPDL cells. Our findings suggest that autophagy is a crucial regulatory process that facilitates type I collagen synthesis and partly regulates osteoblastic differentiation of PDL cells.

scholarly journals Combining Sandblasting, Alkaline Etching, and Collagen Immobilization to Promote Cell Growth on Biomedical Titanium Implants

Polymers ◽  
2021 ◽  
Vol 13 (15) ◽  
pp. 2550
Author(s):  
Chia-Fei Liu ◽  
Kai-Chun Chang ◽  
Ying-Sui Sun ◽  
Diem Thuy Nguyen ◽  
Her-Hsiung Huang
Keyword(s):  
Cell Growth ◽  
Type I Collagen ◽  
Bone Cells ◽  
Surface Characteristics ◽  
Attenuated Total Reflection ◽  
Titanium Implants ◽  
Type I ◽  
Alkaline Etching ◽  
Marrow Mesenchymal Stem Cells ◽  
Collagen Immobilization

Our objective in this study was to promote the growth of bone cells on biomedical titanium (Ti) implant surfaces via surface modification involving sandblasting, alkaline etching, and type I collagen immobilization using the natural cross-linker genipin. The resulting surface was characterized in terms topography, roughness, wettability, and functional groups, respectively using field emission scanning electron microscopy, 3D profilometry, and attenuated total reflection-Fourier transform infrared spectroscopy. We then evaluated the adhesion, proliferation, initial differentiation, and mineralization of human bone marrow mesenchymal stem cells (hMSCs). Results show that sandblasting treatment greatly enhanced surface roughness to promote cell adhesion and proliferation and that the immobilization of type I collagen using genipin enhanced initial cell differentiation as well as mineralization in the extracellular matrix of hMSCs. Interestingly, the nano/submicro-scale pore network and/or hydrophilic features on sandblasted rough Ti surfaces were insufficient to promote cell growth. However, the combination of all proposed surface treatments produced ideal surface characteristics suited to Ti implant applications.

HSP47, a collagen-specific chaperone protein, is a limiting factor for type I collagen secretion in aged skin

2017 ◽  
Vol 86 (2) ◽  
pp. e92
Author(s):  
MinJu Pyo ◽  
Jun Sang Park ◽  
Young Hun Lee ◽  
Dong Hun Lee ◽  
Jin Ho Chung ◽  
...  
Keyword(s):  
Type I Collagen ◽  
Limiting Factor ◽  
Type I ◽  
Chaperone Protein ◽  
Collagen Secretion ◽  
Aged Skin

scholarly journals Endothelial cell growth factor and heparin regulate collagen gene expression in keloid fibroblasts

1991 ◽  
Vol 278 (3) ◽  
pp. 863-869 ◽  
Author(s):  
E M L Tan ◽  
J Peltonen
Keyword(s):  
Gene Expression ◽  
Endothelial Cell ◽  
Collagen Synthesis ◽  
Type I Collagen ◽  
Collagen Type ◽  
Collagen Type I ◽  
Type I ◽  
Collagen Gene ◽  
Endothelial Cell Growth Factor ◽  
Endothelial Cell Growth

Keloids are benign cutaneous tumours characterized by excess deposition of collagen, specifically type I collagen. We report here that collagen biosynthesis, as measured by hydroxyproline synthesis, was markedly inhibited by 65-80% by the combination of endothelial cell growth factor (ECGF) supplement and heparin in keloid fibroblast cultures. Fibroblast cultures that were incubated with ECGF alone also demonstrated a measurable decrease of approx. 50% in collagen synthesis compared with control cultures. The inhibition of collagen synthesis was related to the down-regulation of collagen gene expression. Quantitative measurements of mRNA-cDNA hybrids revealed that the gene expression of collagen type I was decreased by more than 80% by heparin and ECGF. Markedly diminished levels of mRNA encoding collagen type I were also observed in cultures incubated with ECGF alone. The results show that ECGF and heparin elicit a negative regulatory effect on collagen production, and that this inhibition is due largely to the down-regulation of the pro-alpha 1(I) of type I collagen gene. Furthermore, ECGF has a potent suppressive effect, and heparin provides an additive effect to this inhibitory phenomenon.

Surfactant components modulate fibroblast apoptosis and type I collagen and collagenase-1 expression

2000 ◽  
Vol 279 (5) ◽  
pp. L950-L957 ◽  
Author(s):  
Luis Vázquez De Lara ◽  
Carina Becerril ◽  
Martha Montaño ◽  
Carlos Ramos ◽  
Vilma Maldonado ◽  
...  
Keyword(s):  
Growth Rate ◽  
Type I Collagen ◽  
Protein A ◽  
Surfactant Protein ◽  
Northern Blotting ◽  
Tissue Inhibitor Of Metalloproteinase ◽  
Lung Fibroblasts ◽  
Type I ◽  
Chromogenic Assay ◽  
Collagen Accumulation

During lung injury, fibroblasts migrate into the alveolar spaces where they can be exposed to pulmonary surfactant. We examined the effects of Survanta and surfactant protein A (SP-A) on fibroblast growth and apoptosis and on type I collagen, collagenase-1, and tissue inhibitor of metalloproteinase (TIMP)-1 expression. Lung fibroblasts were treated with 100, 500, and 1,000 μg/ml of Survanta; 10, 50, and 100 μg/ml of SP-A; and 500 μg/ml of Survanta plus 50 μg/ml of SP-A. Growth rate was evaluated by a formazan-based chromogenic assay, apoptosis was evaluated by DNA end labeling and ELISA, and collagen, collagenase-1, and TIMP-1 were evaluated by Northern blotting. Survanta provoked fibroblast apoptosis, induced collagenase-1 expression, and decreased type I collagen affecting mRNA stability ∼10-fold as assessed with the use of actinomycin D. Collagen synthesis and collagenase activity paralleled the gene expression results. SP-A increased collagen expression ∼2-fold and had no effect on collagenase-1, TIMP-1, or growth rate. When fibroblasts were exposed to a combination of Survanta plus SP-A, the effects of Survanta were partially reversed. These findings suggest that surfactant lipids may protect against intraluminal fibrogenesis by inducing fibroblast apoptosis and decreasing collagen accumulation.

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