scholarly journals Nature of collagens synthesized by monkey periodontal-ligament fibroblasts in vitro

1978 ◽  
Vol 170 (1) ◽  
pp. 63-71 ◽  
Author(s):  
H F Limeback ◽  
J Sodek ◽  
D M Brunette
Keyword(s):  
Periodontal Ligament ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Deae Cellulose ◽  
Type Iii Collagen ◽  
Type I ◽  
Periodontal Ligament Fibroblasts ◽  
Type Iii ◽  
Type I Procollagen

1. First subcultures of fibroblast-like cells from adult monkey periodontal ligament were incubated in the presence of 14C-labelled amino acids and produced significant amounts of type-I and type-III collagens. 2. The proportion of type-III collagen produced was calculated on the basis of the recovery of procollagens from DEAE-cellulose chromatography to be approx. 20%, and at least 10% when analysed as collagens on CM-cellulose chromatography. 3. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of the procollagens, the collagens and their CNBr peptides was used to confirm the identity of the collagen types. 4. In serum-free media extensive conversion of type-I procollagen, but not of type-III procollagen, into collagen was observed, suggesting that a specific type-I procollagen peptidase was produced. 5. The pattern of collagen synthesis was not significantly different from that obtained with fibroblasts derived from skin corium of the same animals.

scholarly journals Metabolism of rabbit skin collagen. Differences in the apparent turnover rates of type-I- and type-III-collagen precursors determined by constant intravenous infusion of labelled amino acids

1979 ◽  
Vol 181 (1) ◽  
pp. 75-82 ◽  
Author(s):  
S P Robins
Keyword(s):  
Amino Acid ◽  
Collagen Synthesis ◽  
Specific Radioactivity ◽  
Deae Cellulose ◽  
Type Iii Collagen ◽  
Type I ◽  
Turnover Rates ◽  
Type Iii ◽  
Type I Procollagen ◽  
Skin Collagen

Growing rabbits were infused for up to 10 h with labelled proline, tyrosine and leucine to achieve plateau conditions within body free pools, for [3H]proline infusion, blood free-proline specific radioactivity remained constant after about 1 h. For individual animals, type-I- and type-III-collagen precursors were isolated by precipitation with (NH4)2SO4 and DEAE-cellulose chromatography. Experiments where 3H- and 14C-labelled proline and tyrosine were infused concurrently for different periods of time showed that type I procollagen reached plateau specific radioactivity within 3 h and 90% of the plateau value after 2 h infusion, corresponding to a calculated apparent t 1/2 of less than 26 min. Plateau values for type I procollagen were taken as precursor amino acid pool specific radioactivities. The type-III-collagen-precursor fractions consistently showed lower rates of label incorporation and, by assuming that both type I and type III collagens are synthesized from the same amino acid pools, kinetic analysis revealed an apparent t 1/2 for the isolated type-III-collagen precursors of 3.9 h. For proline, there were large variations between animals in the ratio between the precursor pool for collagen synthesis and the skin homogenate free pool (0.31 +/- 0.13, mean +/- S.D.), so that collagen-synthesis rates based solely on total tissue free-pool values for proline are subject to large and inconsistent errors.

scholarly journals Characterization of pepsin-solubilized bovine heart-valve collagen

1978 ◽  
Vol 173 (3) ◽  
pp. 885-894 ◽  
Author(s):  
R I Bashey ◽  
H M Bashey ◽  
S A Jimenez
Keyword(s):  
Heart Valve ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Heart Valves ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
Type Iii Collagen ◽  
Type I ◽  
Type Iii

Collagens extracted from heart valves by using limited pepsin digestion were fractionated by differential salt precipitation. Collagen types were identified by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, amino acid analysis and cleavage with CNBr. Heart-valve collagen was heterogeneous in nature, consisting of a mixture of type-I and type-III collagens. The identity of type-III collagen was established on the basis of (a) insolubility in 1.7 M-NaC1 at neutral pH, (b) behaviour of this collagen fraction on gel electrophoresis under reducing and non-reducing conditions, (c) amino acid analysis showing a hydroxyproline/proline ratio greater than 1, and (d) profile of CNBr peptides on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis showing a peak characteristic for type-III collagen containing peptides alpha1(III)CB8 and alpha1(III)CB3. In addition to types-I and -III collagen, a collagen polypeptide not previously described in heart valves was identified. This polypeptide represented approx. 30% of the collagen fraction precipitated at 4.0 M-NaCl, it migrated between beta- and alpha1-collagen chains on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and its electrophoretic behaviour was not affected by disulphide-bond reduction. All collagen fractions from the heart valves contained increased amounts of hydroxylysine when compared with type-I and -III collagens from other tissues. The presence of beta- and gamma-chains and higher aggregates in pepsin-solubilized collagen indicated that these collagens were highly cross-linked and suggested that some of these cross-links involved the triple-helical regions of the molecule. It is likely that the higher hydroxylysine content of heart-valve collagen is responsible for the high degree of intermolecular cross-linking and may be the result of an adaptive mechanism for the specialized function of these tissues.

Absence of collagen deficiency in familial cerebral aneurysms

1989 ◽  
Vol 70 (6) ◽  
pp. 837-840 ◽  
Author(s):  
Richard Leblanc ◽  
Andrés M. Lozano ◽  
Michel van der Rest ◽  
Ronald David Guttmann
Keyword(s):  
Cell Lines ◽  
Polyacrylamide Gel Electrophoresis ◽  
Control Cell ◽  
Sodium Dodecyl ◽  
Cerebral Aneurysms ◽  
Type Iii Collagen ◽  
Type I ◽  
Content Type ◽  
Type Iii ◽  
Fine Print

✓ It has been suggested that a deficiency in the expression of type III collagen may play a role in the pathogenesis of cerebral aneurysms. To test this hypothesis in cases of familial cerebral aneurysms, fibroblast cell cultures were established and the expression of collagen types I and III was studied in a patient with three cerebral aneurysms whose mother and sister also had cerebral aneurysms. Cultured skin fibroblasts were labeled with tritiated proline. The collagens and procollagens were precipitated and run on sodium dodecyl sulfate-polyacrylamide gel electrophoresis after reduction to analyze procollagen chains. Control cell lines were analyzed simultaneously. Quantitation of the ratios of type III to type I procollagen synthesis was achieved by integration of the intensities of the pro-α1 (III), pro-α1(I), and pro-α2(I) bands on fluorograms of electrophoretic gels of medium proteins. There was no difference in type I and III procollagen levels observed between the cells from the aneurysm patient and those from the control cell lines. These data do not support the hypothesis that familial cerebral aneurysms are caused by a deficiency of type III collagen.

scholarly journals Changes in synthesis of types-I and -III collagen during matrix-induced endochondral bone differentiation in rat

1980 ◽  
Vol 186 (3) ◽  
pp. 919-924 ◽  
Author(s):  
B U Steinmann ◽  
A H Reddi
Keyword(s):  
Type I Collagen ◽  
Bone Matrix ◽  
Sodium Dodecyl ◽  
Mesenchymal Cell ◽  
Type Iii Collagen ◽  
Type I ◽  
Endochondral Bone ◽  
Type Iii

The changes in rates of hydroxyproline formation and biosynthesis of types-I and -III collagen during bone matrix-induced sequential differentiation of cartilage, bone and bone marrow in rat were investigated. Biosynthesis of types-I and -III collagen at different stages of this sequence was studied by labelling in vivo and in vitro with [2,3-3H]proline. Pepsin-solubilized collagens were separated by sodium dodecyl sulphate/polyacrylamide-slab-gel electrophoresis. The results revealed that maximal amounts of type-III collagen were synthesized on day 3 during mesenchymal-cell proliferation. Thereafter, there was a gradual decline in type-III collagen synthesis. On days 9-20 during bone formation predominantly type-I collagen was synthesized. Similar results were obtained by the use of labelling techniques both in vivo and in vitro.

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 Separation of the Binding Step from the Collagen Platelet Reaction

1977 ◽  
Author(s):  
P.L. Kronick ◽  
S.A. Jimenez
Keyword(s):  
Type I Collagen ◽  
Specific Activity ◽  
Adenine Nucleotides ◽  
High Specific Activity ◽  
Type Iii Collagen ◽  
Type I ◽  
Type Iii ◽  
Valid Comparison ◽  
Platelet Reaction

Determination of activity of most agents in stimulating platelets to aggregate or release adenine nucleotides is conveniently done by titrating the platelet reaction with the agent. Platelets have previously been titrated with different types of collagen (types I, II, and III) in this way to compare the activities of the collagens. It has been concluded that the order of activity is type III>I>II. Whether this order is due to differences in binding was not obvious from these experiments because the binding was not determined directly. We have developed a method of comparing activities by measuring the targeted dose for each point in the titration - the amount of collagen which actually binds to platelets. The collagens used in these experiments were prepared in vitro from embryonic chick tissue to give labelled products of extremely high specific activity without structural alteration. We find that type I collagen is at least 20 times as active as previously reported, and that the activity of Type III collagen is not significantly higher when the amounts bound are taken into account. The fraction of the labelled tendon collagen which was bound to platelets was identified as type I by its hydroxyproline/proline ratio. Direct measurement of the bound fraction in dose-response studies is required for valid comparison of collagen activities.

Type I and III procollagen propeptides in growth hormone-deficient patients: effects of increasing doses of GH

1991 ◽  
Vol 124 (3) ◽  
pp. 278-282 ◽  
Author(s):  
Lars T. Jensen ◽  
Jens O.L. Jørgensen ◽  
Juha Risteli ◽  
Jens S. Christiansen ◽  
Ib Lorenzen
Keyword(s):  
Type Iii Collagen ◽  
Type I ◽  
Dose Dependency ◽  
Loose Connective Tissue ◽  
Gh Therapy ◽  
Type Iii ◽  
Type I Procollagen ◽  
Type Iii Procollagen ◽  
Procollagen Propeptides ◽  
Serum Type

Abstract. The effect of increasing doses of growth hormone on collagen synthesis in GH-treated GH-deficient patients was determined in a short-term study. The synthesis of type I and III collagen was estimated by measurements of the carboxyterminal propeptide of type I procollagen and the aminoterminal propeptide of type III procollagen. Type I collagen is mainly found in bone and type III collagen in loose connective tissue. We observed a GH dose dependency of both procollagen propeptides. Serum type I procollagen propeptide was significantly higher following GH doses of 4 and 6 IU/day for 14 days compared with 2 IU/day (normal replacement dose) (p=0.04). Withdrawal of GH therapy for 14 days resulted in wider variation, but not significantly different from the levels at 2, 4 and 6 IU/day. A dose dependency was found regarding type III procollagen propeptide, showing significantly higher serum concentrations at a GH dose of 4 IU/day compared with 2 IU/day (p=0.001), and of 6 IU/day compared with 4 IU/day (p=0.001). Withdrawal of GH therapy resulted in significantly lower type III procollagen propeptide concentrations compared with those at a GH dose of 4 and 6 IU/day (p=0.03). Serum type III procollagen propeptide increased twice as much as type I procollagen propeptide, by 47 vs 25%, at a GH dose of 6 IU/day compared with 2 IU/day. The differences between the effects on type I and type III collagen may reflect differences in secretion or turn-over rate of collagen in bone and loose connective tissue. Serum type I and type III procollagen propeptides may prove useful as monitors of GH therapy, especially regarding the GH dose levels in the individual patients.

An osteonectinlike protein in porcine periodontal ligament and its synthesis by periodontal ligament fibroblasts

1984 ◽  
Vol 62 (6) ◽  
pp. 470-478 ◽  
Author(s):  
Safia Wasi ◽  
Kichibee Otsuka ◽  
Kam-Ling Yao ◽  
Pierre S. Tung ◽  
Jane E. Aubin ◽  
...  
Keyword(s):  
Periodontal Ligament ◽  
Culture Medium ◽  
Alveolar Bone ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Relative Mass ◽  
Bovine Bone ◽  
Binding Studies ◽  
Periodontal Ligament Fibroblasts ◽  
Sds Page

Periodontal ligament, a soft connective tissue that lies between cementum and alveolar bone in the periodontium, has been shown to contain an osteonectinlike protein. The similarity between porcine ligament osteonectin and bovine bone osteonectin was evident from immunochemical studies, from migration characteristics on sodium dodecyl sulfate – polyacrylamide gel electrophoresis (SDS–PAGE) and from binding studies on hydroxyapatite. Using immunotransfer and immunodot analyses, ligament osteonectin was found to be extractable from tissues with 4 M guanidine–HCl (GuHCl) and 4 M GuHCl − 0.5 M EDTA and to comigrate with authentic bovine osteonectin on SDS–PAGE with a relative mass ~ 38 000. Furthermore, osteonectin from guanidine extracts of ligament was bound to hydroxyapatite in the presence of 4 M GuHCl. Immunofluorescence studies showed the osteonectin to be distributed throughout the extracellular matrix of the ligament and to be present within the ligament fibroblasts in a perinuclear, punctate distribution. Biosynthesis of osteonectin by ligament fibroblasts was studied following pulse-chase labelling with [35S]methionine and immunoprecipitation. The labelled osteonectin in the chased culture medium represented ~0.5% of the total labelled proteins secreted. It comigrated on SDS–PAGE with the corresponding labelled protein from pulsed cells and with the protein extracted from the tissue.

Phagocytosis of Fibronectin and Collagens Type I, III, and V by Human Gingival and Periodontal Ligament Fibroblasts In Vitro

2001 ◽  
Vol 72 (10) ◽  
pp. 1340-1347 ◽  
Author(s):  
Monique T. van der Pauw ◽  
Theo Van den Bos ◽  
Vincent Everts ◽  
Wouter Beertsen
Keyword(s):  
Periodontal Ligament ◽  
Type I ◽  
Periodontal Ligament Fibroblasts
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