The role of protein kinase cascades and proto-oncogene expression and activation in the ihibition by camp of PDGF and pmainduced human fibroblast growth

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Abstract

Fibroblast proliferation is a key event in several normal processes like wound and tissue repair and the control of abnormal cell proliferation leading to fibrosis is a primary problem in numerous diseases. In this study, cAMP level-raising agents were shown to inhibit human lung fibroblast proliferation induced by PDGF (Platelet-derived Growth Factor) and PMA (phosbol 12-myristate 13-acetate). We wanted to know at which level of the PDGF and PMA-signalling pathways cAMP interferes. The effects of PDGF are mediated via the activation of protein kinase cascades leading to the activation of transcription factors and gene expression. We first investigated the role ot phosphatidylinositol 3-kinase (PI-3K), protein kinase C (PKC) and mitogen-activated protein kinases (MAPK) in PDGF-induced cell proliferation. Wortmannin, a specific inhibitor of PI-3K, had no effect on PDGF-induced cell growth, suggesting that PI-3K activation is not essential to this fibroblastic response. We then tested the effect of PDGF on PKC activation using a colorimetric assay kit. Contrary to PMA, PDGF didn't affect PKC activity i n these cells. Finally, PDGF was shown to induce MAPK phosphorylation (detected by gels shifts on immunoblotting) and activation (estimated by immune complex kinase assay). PMA also activated MAPK but to a less extend than PDGF. DibutyrylcAMP, an analogue of cAMP, had no effect on PDGF and PMAinduced Mapk activation suggesting that cAMP interferes either at a step downstream of the MAPK cascade or at an other level of PDGF and PMA-signalling pathways. We are therefore presently investigating the possible effect of cAMP on the binding interaction between transcription factors (AP-1 i.e. Fos/Jun or Jun /Jun dimers and Myc-Max) and consensus DNA binding elements (by E.M.S.A.) and also on the expression of the protooncogenes c-myc, c-jun and c-fos (by Northern-Blot).
Original languageEnglish
JournalBiochemical Society transactions
Volume24
Issue number4
Publication statusPublished - 1 Jan 1996

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Proto-Oncogenes
Platelet-Derived Growth Factor
Fibroblasts
Protein Kinases
Chemical activation
Growth
Myristic Acid
Phosphatidylinositol 3-Kinase
Mitogen-Activated Protein Kinases
Acetates
Protein Kinase C
Cell proliferation
Assays
Cell Proliferation
Phosphorylation
Transcription Factor AP-1
Cell growth
Antigen-Antibody Complex
Immunoblotting
Gene expression

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@article{9d2a8fd914bf431b8daafe159f9a1198,
title = "The role of protein kinase cascades and proto-oncogene expression and activation in the ihibition by camp of PDGF and pmainduced human fibroblast growth",
abstract = "Fibroblast proliferation is a key event in several normal processes like wound and tissue repair and the control of abnormal cell proliferation leading to fibrosis is a primary problem in numerous diseases. In this study, cAMP level-raising agents were shown to inhibit human lung fibroblast proliferation induced by PDGF (Platelet-derived Growth Factor) and PMA (phosbol 12-myristate 13-acetate). We wanted to know at which level of the PDGF and PMA-signalling pathways cAMP interferes. The effects of PDGF are mediated via the activation of protein kinase cascades leading to the activation of transcription factors and gene expression. We first investigated the role ot phosphatidylinositol 3-kinase (PI-3K), protein kinase C (PKC) and mitogen-activated protein kinases (MAPK) in PDGF-induced cell proliferation. Wortmannin, a specific inhibitor of PI-3K, had no effect on PDGF-induced cell growth, suggesting that PI-3K activation is not essential to this fibroblastic response. We then tested the effect of PDGF on PKC activation using a colorimetric assay kit. Contrary to PMA, PDGF didn't affect PKC activity i n these cells. Finally, PDGF was shown to induce MAPK phosphorylation (detected by gels shifts on immunoblotting) and activation (estimated by immune complex kinase assay). PMA also activated MAPK but to a less extend than PDGF. DibutyrylcAMP, an analogue of cAMP, had no effect on PDGF and PMAinduced Mapk activation suggesting that cAMP interferes either at a step downstream of the MAPK cascade or at an other level of PDGF and PMA-signalling pathways. We are therefore presently investigating the possible effect of cAMP on the binding interaction between transcription factors (AP-1 i.e. Fos/Jun or Jun /Jun dimers and Myc-Max) and consensus DNA binding elements (by E.M.S.A.) and also on the expression of the protooncogenes c-myc, c-jun and c-fos (by Northern-Blot).",
author = "M. Burton and {Van Steenbrugge}, M. and Martine Raes",
note = "Copyright 2006 Elsevier B.V., All rights reserved.",
year = "1996",
month = "1",
day = "1",
language = "English",
volume = "24",
journal = "Biochemical Society transactions",
issn = "0300-5127",
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T1 - The role of protein kinase cascades and proto-oncogene expression and activation in the ihibition by camp of PDGF and pmainduced human fibroblast growth

AU - Burton, M.

AU - Van Steenbrugge, M.

AU - Raes, Martine

N1 - Copyright 2006 Elsevier B.V., All rights reserved.

PY - 1996/1/1

Y1 - 1996/1/1

N2 - Fibroblast proliferation is a key event in several normal processes like wound and tissue repair and the control of abnormal cell proliferation leading to fibrosis is a primary problem in numerous diseases. In this study, cAMP level-raising agents were shown to inhibit human lung fibroblast proliferation induced by PDGF (Platelet-derived Growth Factor) and PMA (phosbol 12-myristate 13-acetate). We wanted to know at which level of the PDGF and PMA-signalling pathways cAMP interferes. The effects of PDGF are mediated via the activation of protein kinase cascades leading to the activation of transcription factors and gene expression. We first investigated the role ot phosphatidylinositol 3-kinase (PI-3K), protein kinase C (PKC) and mitogen-activated protein kinases (MAPK) in PDGF-induced cell proliferation. Wortmannin, a specific inhibitor of PI-3K, had no effect on PDGF-induced cell growth, suggesting that PI-3K activation is not essential to this fibroblastic response. We then tested the effect of PDGF on PKC activation using a colorimetric assay kit. Contrary to PMA, PDGF didn't affect PKC activity i n these cells. Finally, PDGF was shown to induce MAPK phosphorylation (detected by gels shifts on immunoblotting) and activation (estimated by immune complex kinase assay). PMA also activated MAPK but to a less extend than PDGF. DibutyrylcAMP, an analogue of cAMP, had no effect on PDGF and PMAinduced Mapk activation suggesting that cAMP interferes either at a step downstream of the MAPK cascade or at an other level of PDGF and PMA-signalling pathways. We are therefore presently investigating the possible effect of cAMP on the binding interaction between transcription factors (AP-1 i.e. Fos/Jun or Jun /Jun dimers and Myc-Max) and consensus DNA binding elements (by E.M.S.A.) and also on the expression of the protooncogenes c-myc, c-jun and c-fos (by Northern-Blot).

AB - Fibroblast proliferation is a key event in several normal processes like wound and tissue repair and the control of abnormal cell proliferation leading to fibrosis is a primary problem in numerous diseases. In this study, cAMP level-raising agents were shown to inhibit human lung fibroblast proliferation induced by PDGF (Platelet-derived Growth Factor) and PMA (phosbol 12-myristate 13-acetate). We wanted to know at which level of the PDGF and PMA-signalling pathways cAMP interferes. The effects of PDGF are mediated via the activation of protein kinase cascades leading to the activation of transcription factors and gene expression. We first investigated the role ot phosphatidylinositol 3-kinase (PI-3K), protein kinase C (PKC) and mitogen-activated protein kinases (MAPK) in PDGF-induced cell proliferation. Wortmannin, a specific inhibitor of PI-3K, had no effect on PDGF-induced cell growth, suggesting that PI-3K activation is not essential to this fibroblastic response. We then tested the effect of PDGF on PKC activation using a colorimetric assay kit. Contrary to PMA, PDGF didn't affect PKC activity i n these cells. Finally, PDGF was shown to induce MAPK phosphorylation (detected by gels shifts on immunoblotting) and activation (estimated by immune complex kinase assay). PMA also activated MAPK but to a less extend than PDGF. DibutyrylcAMP, an analogue of cAMP, had no effect on PDGF and PMAinduced Mapk activation suggesting that cAMP interferes either at a step downstream of the MAPK cascade or at an other level of PDGF and PMA-signalling pathways. We are therefore presently investigating the possible effect of cAMP on the binding interaction between transcription factors (AP-1 i.e. Fos/Jun or Jun /Jun dimers and Myc-Max) and consensus DNA binding elements (by E.M.S.A.) and also on the expression of the protooncogenes c-myc, c-jun and c-fos (by Northern-Blot).

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