Collagen expression in embryonic chicken chondrocytes treated with phorbol myristate acetate

1985 ◽  
Vol 5 (6) ◽  
pp. 1415-1424
Author(s):  
M H Finer ◽  
L C Gerstenfeld ◽  
D Young ◽  
P Doty ◽  
H Boedtker
Keyword(s):  
Morphological Change ◽  
Type I Collagen ◽  
Late Phase ◽  
Tumor Promoter ◽  
Type I ◽  
Mrna Levels ◽  
Type Ii ◽  
Collagen Mrna ◽  
Type I Procollagen ◽  
Embryonic Chicken

Growth of embryonic chicken sternal chondrocytes in the presence of phorbol-12-myristate-13-acetate (PMA), a potent tumor promoter, resulted in a dramatic morphological change from spherical floating cells to adherent fibroblastic cells. This morphological change was accompanied by a quantitative switch from synthesis of cartilage-specific type II procollagen to type I procollagen. Type II procollagen mRNA levels decreased 10-fold in PMA-treated cells. Activation of type I collagen genes led to the accumulation of type I procollagen mRNA levels comparable to those of type II mRNA in these cells. However, only type I procollagen mRNA was translated. In addition to gene activation, unprocessed pro alpha 1(I) transcripts present at low levels in control chondrocytes were processed to mature mRNA species. Redifferentiation of PMA-treated chondrocytes was possible if cells were removed from PMA after the morphological change and cessation of type II procollagen synthesis but before detectable amounts of type I procollagen were synthesized. Production of type I collagen thus marks a late phase of chondrocyte "dedifferentiation" from which reversion is no longer possible. Redifferentiated cell populations contained 24-fold more pro alpha 1(II) collagen mRNA than pro alpha 1(I) collagen mRNA, but the rates of procollagen synthesis were comparable. This suggests that the PMA-mediated dedifferentiation of chondrocytes as well as their redifferentiation is under both transcriptional and posttranscriptional regulation.

scholarly journals Collagen expression in embryonic chicken chondrocytes treated with phorbol myristate acetate.

1985 ◽  
Vol 5 (6) ◽  
pp. 1415-1424 ◽  
Author(s):  
M H Finer ◽  
L C Gerstenfeld ◽  
D Young ◽  
P Doty ◽  
H Boedtker
Keyword(s):  
Morphological Change ◽  
Type I Collagen ◽  
Late Phase ◽  
Tumor Promoter ◽  
Type I ◽  
Mrna Levels ◽  
Type Ii ◽  
Collagen Mrna ◽  
Type I Procollagen ◽  
Embryonic Chicken

Growth of embryonic chicken sternal chondrocytes in the presence of phorbol-12-myristate-13-acetate (PMA), a potent tumor promoter, resulted in a dramatic morphological change from spherical floating cells to adherent fibroblastic cells. This morphological change was accompanied by a quantitative switch from synthesis of cartilage-specific type II procollagen to type I procollagen. Type II procollagen mRNA levels decreased 10-fold in PMA-treated cells. Activation of type I collagen genes led to the accumulation of type I procollagen mRNA levels comparable to those of type II mRNA in these cells. However, only type I procollagen mRNA was translated. In addition to gene activation, unprocessed pro alpha 1(I) transcripts present at low levels in control chondrocytes were processed to mature mRNA species. Redifferentiation of PMA-treated chondrocytes was possible if cells were removed from PMA after the morphological change and cessation of type II procollagen synthesis but before detectable amounts of type I procollagen were synthesized. Production of type I collagen thus marks a late phase of chondrocyte "dedifferentiation" from which reversion is no longer possible. Redifferentiated cell populations contained 24-fold more pro alpha 1(II) collagen mRNA than pro alpha 1(I) collagen mRNA, but the rates of procollagen synthesis were comparable. This suggests that the PMA-mediated dedifferentiation of chondrocytes as well as their redifferentiation is under both transcriptional and posttranscriptional regulation.

scholarly journals Altered beta-actin gene expression in phorbol myristate acetate-treated chondrocytes and fibroblasts.

1985 ◽  
Vol 5 (6) ◽  
pp. 1425-1433 ◽  
Author(s):  
L C Gerstenfeld ◽  
M H Finer ◽  
H Boedtker
Keyword(s):  
Steady State ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Tumor Promoter ◽  
Type I ◽  
Mrna Levels ◽  
Collagen Mrna ◽  
Beta Actin ◽  
Steady State Levels ◽  
Actin Mrna

Phorbol-12-myristate-13-acetate (PMA), a potent tumor promoter, was shown to have opposite effects on the cellular morphology and steady-state levels of beta-actin mRNA in embryonic chicken muscle fibroblasts and sternal chondrocytes. When fibroblasts were treated with PMA, they formed foci of densely packed cells, ceased to adhere to culture plates, and had significantly reduced levels of beta-actin mRNA and protein. Conversely, when treated with PMA, floating chondrocytes attached to culture dishes, spread out, and began to accumulate high levels of beta-actin mRNA and proteins. In the sternal chondrocytes the stimulation of the beta-actin mRNA production was accompanied by increased steady-state levels of fibronectin mRNAs and protein. These alterations were concomitant with a fivefold reduction in type II collagen mRNA and a cessation in its protein production. After fibronectin and actin mRNAs and proteins reached their maximal levels, type I collagen mRNA and protein synthesis were turned on. Removal of PMA resulted in reduced beta-actin mRNA levels in chondrocytes and in a further alteration in the cell morphology. These observed correlations between changes in cell adhesion and morphology and beta-actin expression suggest that the effect of PMA on cell shape and adhesion may result in changes in the microfilament organization of the cytoskeleton which ultimately lead to changes in the extracellular matrix produced by the cells.

scholarly journals Lack of a phenotype in transgenic mice aberrantly expressing COL2A1 mRNA because of highly selective post-transcriptional down-regulation

2000 ◽  
Vol 345 (2) ◽  
pp. 377-384 ◽  
Author(s):  
Constance M. YUAN ◽  
Leena ALA-KOKKO ◽  
Dominique LE GUELLEC ◽  
Suzanne FRANC ◽  
Andrzej FERTALA ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Type I ◽  
Mrna Levels ◽  
Type Ii ◽  
Col2a1 Gene ◽  
Type I Procollagen ◽  
Steady State Levels ◽  
Hybridization In Situ

We reported previously that a 1.9-kb 5ʹ-fragment from the human COL1A1 gene drove transcription of a promoterless human COL2A1 gene in tissues of transgenic mice that normally express the COL1A1 but not the COL2A1 gene. In the present study, we have established that the aberrant transcription of the COL2A1 gene did not produce any gross or microscopic phenotype, because the transcripts were not efficiently translated in cells that do not normally express the COL2A1 gene. In two lines of transgenic mice, the mRNA levels from the transgene were 30% to 45% of the mRNA for the proα1(I) chain of type I procollagen, the most abundant mRNA in the same tissues. Analysis of collagens extracted from skin of the transgenic mice indicated that triple-helical type II collagen, with the normal pattern of cyanogen bromide peptides, was synthesized from the transgene. However, the level of type II collagen in skin was less than 2% of the level of type I collagen. Hybridization in situ indicated the presence of mRNA for both COL2A1 and COL1A1 in the same cells. Immunofluorescence staining for type II collagen, however, was negative in the same tissues. The results, therefore, indicated that many mesenchymal cells in the transgenic mice had high steady-state levels of the homologous mRNAs for type I and type II procollagen, but only the mRNAs for type I procollagen were efficiently translated.

Altered beta-actin gene expression in phorbol myristate acetate-treated chondrocytes and fibroblasts

1985 ◽  
Vol 5 (6) ◽  
pp. 1425-1433
Author(s):  
L C Gerstenfeld ◽  
M H Finer ◽  
H Boedtker
Keyword(s):  
Steady State ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Tumor Promoter ◽  
Type I ◽  
Mrna Levels ◽  
Collagen Mrna ◽  
Beta Actin ◽  
Steady State Levels ◽  
Actin Mrna

Phorbol-12-myristate-13-acetate (PMA), a potent tumor promoter, was shown to have opposite effects on the cellular morphology and steady-state levels of beta-actin mRNA in embryonic chicken muscle fibroblasts and sternal chondrocytes. When fibroblasts were treated with PMA, they formed foci of densely packed cells, ceased to adhere to culture plates, and had significantly reduced levels of beta-actin mRNA and protein. Conversely, when treated with PMA, floating chondrocytes attached to culture dishes, spread out, and began to accumulate high levels of beta-actin mRNA and proteins. In the sternal chondrocytes the stimulation of the beta-actin mRNA production was accompanied by increased steady-state levels of fibronectin mRNAs and protein. These alterations were concomitant with a fivefold reduction in type II collagen mRNA and a cessation in its protein production. After fibronectin and actin mRNAs and proteins reached their maximal levels, type I collagen mRNA and protein synthesis were turned on. Removal of PMA resulted in reduced beta-actin mRNA levels in chondrocytes and in a further alteration in the cell morphology. These observed correlations between changes in cell adhesion and morphology and beta-actin expression suggest that the effect of PMA on cell shape and adhesion may result in changes in the microfilament organization of the cytoskeleton which ultimately lead to changes in the extracellular matrix produced by the cells.

scholarly journals Regulation of type I collagen mRNA in lung fibroblasts by cystine availability

1998 ◽  
Vol 331 (2) ◽  
pp. 417-422 ◽  
Author(s):  
David C. RISHIKOF ◽  
Ping-Ping KUANG ◽  
Christine POLIKS ◽  
Ronald H. GOLDSTEIN
Keyword(s):  
Amino Acids ◽  
Steady State ◽  
Amino Acid ◽  
Type I Collagen ◽  
State Level ◽  
Lung Fibroblasts ◽  
Type I ◽  
Mrna Levels ◽  
Collagen Mrna ◽  
Amino Acid Availability

The steady-state level of α1(I) collagen mRNA is regulated by amino acid availability in human lung fibroblasts. Depletion of amino acids decreases α1(I) collagen mRNA levels and repletion of amino acids induces rapid re-expression of α1(I) mRNA. In these studies, we examined the requirements for individual amino acids on the regulation of α1(I) collagen mRNA. We found that re-expression of α1(I) collagen mRNA was critically dependent on cystine but not on other amino acids. However, the addition of cystine alone did not result in re-expression of α1(I) collagen mRNA. Following amino acid depletion, the addition of cystine with selective amino acids increased α1(I) collagen mRNA levels. The combination of glutamine and cystine increased α1(I) collagen mRNA levels 6.3-fold. Methionine or a branch-chain amino acid (leucine, isoleucine or valine) also acted in combination with cystine to increase α1(I) collagen mRNA expression, whereas other amino acids were not effective. The prolonged absence of cystine lowered steady-state levels of α1(I) collagen mRNA through a mechanism involving decreases in both the rate of gene transcription as assessed by nuclear run-on experiments and mRNA stability as assessed by half-life determination in the presence of actinomycin D. The effect of cystine was not mediated via alterations in the level of glutathione, the major redox buffer in cells, as determined by the addition of buthionine sulphoximine, an inhibitor of γ-glutamylcysteine synthetase. These data suggest that cystine directly affects the regulation of α1(I) collagen mRNA.

scholarly journals Regulation of type I collagen mRNA levels in fibroblasts

1986 ◽  
Vol 157 (2) ◽  
pp. 433-439 ◽  
Author(s):  
Tilman VOSS ◽  
Paul BORNSTEIN
Keyword(s):  
Type I Collagen ◽  
Type I ◽  
Mrna Levels ◽  
Collagen Mrna

Relationships between mesangial cell proliferation and types I and IV collagen mRNA levels in vitro

1995 ◽  
Vol 269 (3) ◽  
pp. C554-C562 ◽  
Author(s):  
C. J. He ◽  
L. J. Striker ◽  
M. Tsokos ◽  
C. W. Yang ◽  
E. P. Peten ◽  
...  
Keyword(s):  
Type I Collagen ◽  
Mesangial Cell ◽  
Mesangial Cells ◽  
Methyl Cellulose ◽  
Collagen Gel ◽  
Type I ◽  
Mrna Levels ◽  
Collagen Mrna ◽  
Type Iv

Changes in the composition of the mesangial extracellular matrix (ECM) and cell turnover are present in glomerular disease. To determine if ECM changes play a role in perpetuating mesangial cell dysfunction, we examined a line of mouse mesangial cells cultured on films or gels of several ECM components and also on methyl cellulose, an inert substrate that prevents attachment. Cells on films of fibronectin or type IV or I collagen had persistently high growth rates and high levels of alpha 1-I and alpha 1-IV collagen mRNAs. In contrast, on gels of type IV or I collagen or matrigel, the growth rate was low. The alpha 1-IV collagen mRNA levels were low on type IV collagen gel or matrigel, whereas the alpha 1-I collagen mRNA levels remained high. In contrast, the alpha 1-I collagen mRNA levels were low on type I collagen gel, and the alpha 1-IV collagen mRNA levels were high. Cells on methyl cellulose formed floating aggregates, did not proliferate, and had a 5- to 10-fold decrease in both alpha 1-I and alpha 1-IV collagen mRNA levels. These phenotypic changes were largely reversible. Finally, when matrigel was layered over cells on fibronectin films, alpha 1-IV collagen mRNA levels decreased, but alpha 1-I collagen mRNA levels and proliferation remained high. Thus proliferation and alpha 1-I and alpha 1-IV collagen mRNA levels in mesangial cells were independently regulated and depended on attachment and the nature of the adjacent matrix.

Nonsynchronous changes in myocardial collagen mRNA and protein during aging: effect of DOCA-salt hypertension

1994 ◽  
Vol 267 (6) ◽  
pp. H2237-H2244 ◽  
Author(s):  
S. Besse ◽  
V. Robert ◽  
P. Assayag ◽  
C. Delcayre ◽  
B. Swynghedauw
Keyword(s):  
Aging Effect ◽  
Type I ◽  
Mrna Levels ◽  
Salt Treatment ◽  
Collagen Mrna ◽  
Type I Procollagen ◽  
Salt Hypertension ◽  
Old Rats ◽  
Modify Collagen ◽  
Myocardial Collagen

Myocardial fibrosis has been investigated in 3-, 16-, and 24-mo-old normal rats and also in 24-mo-old rats subjected to deoxycorticosterone acetate (DOCA)-salt treatment-induced-hypertension. Collagen content was assessed both histologically and by hydroxyproline assay. Type I and III procollagen mRNA levels were quantitated by Slot Blot analyses. Aging is associated with fibrosis as shown both biochemically (hydroxyproline concentration in 3-, 16-, and 24-mo-old rats was 0.70 +/- 0.05, 0.92 +/- 0.07, and 1.57 +/- 0.13 mg/g of left ventricle, respectively, P < 0.05 and P < 0.0001 vs. 3 mo) and histologically. By contrast, type I procollagen mRNA levels decreased during aging (from -63%, P < 0.001 in 16-mo-old rats and -51%, P < 0.01 in 24-mo-old rats vs. 3-mo-old rats) as well as type III procollagen mRNA levels. DOCA-salt treatment in 24-mo-old rats had no effect on either the degree of fibrosis or the mRNA levels. We conclude that nonsynchronous changes in myocardial collagen mRNA and protein occur during aging, indicating translational and/or posttranslational mechanisms in collagen regulation. Hypertension during senescence did not modify collagen deposition at either the protein or mRNA 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.

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