Sustained contraction and loss of NO production in TGFβ 1- treated endothelial cells

M. Watanabe, M. Oike, Y. Ohta, H. Nawata, Y. Ito

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Abstract

Background and purpose: Transforming growth factor β 1 (TGFβ 1) is generated in atherosclerotic and injured vessel walls. We examined whether the endothelial-to-mesenchymal transdifferentiation induced by TGFβ 1 affects endothelial functions. Experimental approach: Bovine aortic endothelial cells (BAECs) were treated with 3 ng ml -1 TGFβ 1 for 7 days. Contraction of TGFβ 1-treated BAECs was assessed by collagen gel contraction assay. Protein expression and phosphorylation were assessed by Western blotting. Intracellular Ca 2+ concentration and NO production were measured using fura2 and DAF-2, respectively. Key results: TGFβ 1-treated BAECs showed dense actin fibers and expressed smooth muscle marker proteins; they also changed into smooth muscle-like, spindle-shaped cells in collagen gel cultures. ATP (10 μM) induced a gradual contraction of collagen gels containing TGFβ 1-treated BAECs but not of gels containing control BAECs. ATP-induced contraction of TGFβ 1-treated BAECs was not reversed by the removal of ATP but was partially suppressed by a high concentration of sodium nitroprusside (1μM). TGFβ 1-treated BAECs showed sustained phosphorylation of myosin light chain in response to ATP and low levels of basal MYPT1 expression. ATP-induced Ca 2+ transients as well as eNOS protein expression were not affected by TGFβ 1 in BAECs. However, ATP-induced NO production was significantly reduced in TGFβ 1-treated BAECs. Anti-TGFβ 1 antibody abolished all of these TGFβ 1-induced changes in BAECs.Conclusions and Implications:Mesenchymal transdifferentiation induced by TGFβ 1 leads to sustained contraction and reduced NO production in endothelial cells. Such effects, therefore, would not be beneficial for vascular integrity.

Original languageEnglish
Pages (from-to)355-364
Number of pages10
JournalBritish Journal of Pharmacology
Volume149
Issue number4
DOIs
Publication statusPublished - Oct 11 2006

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Transforming Growth Factors
Endothelial Cells
Adenosine Triphosphate
Gels
Collagen
Smooth Muscle
Phosphorylation
Muscle Spindles
Myosin Light Chains
Muscle Proteins
Nitroprusside
Blood Vessels
Actins
Proteins
Western Blotting

All Science Journal Classification (ASJC) codes

  • Pharmacology

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Sustained contraction and loss of NO production in TGFβ 1- treated endothelial cells. / Watanabe, M.; Oike, M.; Ohta, Y.; Nawata, H.; Ito, Y.

In: British Journal of Pharmacology, Vol. 149, No. 4, 11.10.2006, p. 355-364.

Research output: Contribution to journalArticle

Watanabe, M. ; Oike, M. ; Ohta, Y. ; Nawata, H. ; Ito, Y. / Sustained contraction and loss of NO production in TGFβ 1- treated endothelial cells. In: British Journal of Pharmacology. 2006 ; Vol. 149, No. 4. pp. 355-364.
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AB - Background and purpose: Transforming growth factor β 1 (TGFβ 1) is generated in atherosclerotic and injured vessel walls. We examined whether the endothelial-to-mesenchymal transdifferentiation induced by TGFβ 1 affects endothelial functions. Experimental approach: Bovine aortic endothelial cells (BAECs) were treated with 3 ng ml -1 TGFβ 1 for 7 days. Contraction of TGFβ 1-treated BAECs was assessed by collagen gel contraction assay. Protein expression and phosphorylation were assessed by Western blotting. Intracellular Ca 2+ concentration and NO production were measured using fura2 and DAF-2, respectively. Key results: TGFβ 1-treated BAECs showed dense actin fibers and expressed smooth muscle marker proteins; they also changed into smooth muscle-like, spindle-shaped cells in collagen gel cultures. ATP (10 μM) induced a gradual contraction of collagen gels containing TGFβ 1-treated BAECs but not of gels containing control BAECs. ATP-induced contraction of TGFβ 1-treated BAECs was not reversed by the removal of ATP but was partially suppressed by a high concentration of sodium nitroprusside (1μM). TGFβ 1-treated BAECs showed sustained phosphorylation of myosin light chain in response to ATP and low levels of basal MYPT1 expression. ATP-induced Ca 2+ transients as well as eNOS protein expression were not affected by TGFβ 1 in BAECs. However, ATP-induced NO production was significantly reduced in TGFβ 1-treated BAECs. Anti-TGFβ 1 antibody abolished all of these TGFβ 1-induced changes in BAECs.Conclusions and Implications:Mesenchymal transdifferentiation induced by TGFβ 1 leads to sustained contraction and reduced NO production in endothelial cells. Such effects, therefore, would not be beneficial for vascular integrity.

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