The role of platelet-derived growth factor-BB-induced increase in cytosolic free Ca2+ in activation of mitogen-activated protein kinase and DNA synthesis in vascular smooth muscle cells.
BACKGROUND
Platelet derived growth factor (PDGF)-BB is an important vascular smooth muscle cell (VSMC) mitogen. PDGF-BB induces an increase in intracellular free calcium concentration ([Ca2+]i), an activation of mitogen-activated protein (MAP) kinase and an increase in DNA synthesis. The increase in [Ca2+]i is thought to be an important second messenger in the intracellular signalling cascade, leading to growth of VSMC.


OBJECTIVE
The aim of the present study was to elucidate the role of the PDGF-BB-induced increase in [Ca2+]i in the activation of MAP kinase and increase in DNA synthesis. Binding of [Ca2+]i was performed by the intracellular chelator bis-(2-amino-5-methylphenoxy) ethane-N,N,N',N'-tetraacetic acid tetraacetoxymethyl ester (MAPTAM).


METHODS
Ca2+ levels were measured by the Fura-2 method. MAP kinase activation was determined by Western blotting. DNA synthesis was determined by measurement of incorporation of [3H]-thymidine into the cell DNA.


RESULTS
Administration of 50 ng/ml PDGF-BB induced an increase in [Ca2+]i, an activation of MAP kinase and an increase in DNA synthesis. In bis-(2-amino-5-methylphenoxy) ethane-N,N,N'N'-tetraacetic acid tetraacetoxymethyl ester (MAPTAM)-treated cells the PDGF-BB-induced effect on [Ca2+]i was totally blunted, whereas no effect on MAP kinase activation and DNA synthesis could be observed.


CONCLUSIONS
These findings show that the effect of PDGF-BB on MAP kinase activation is independent of calcium level. [Ca2+]i might be implicated in the PDGF-BB-induced mitogenic process only in conjugation with other signalling components.
Seewald
S
S
Medizinische Universitäts-Poliklinik, Bonn, Germany.
Sachinidis
A
A
Seul
C
C
Kettenhofen
R
R
Ko
Y
Y
Vetter
H
H
D000818:Animals; D000925:Anticoagulants / Q000494:pharmacology*; D001013:Aorta, Thoracic / Q000166:cytology; D000077214:Becaplermin; D002118:Calcium / Q000378:metabolism*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000187:drug effects / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis*; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D007424:Intracellular Fluid / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology*; D020574:Proto-Oncogene Proteins c-sis; D051381:Rats; D011921:Rats, Inbred WKY; D011994:Recombinant Proteins
D000925:Anticoagulants; D002614:Chelating Agents; D010982:Platelet-Derived Growth Factor; D020574:Proto-Oncogene Proteins c-sis; D011994:Recombinant Proteins; D000077214:Becaplermin; D004533:Egtazic Acid; D004247:DNA; C058180:5,5'-dimethyl-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetate; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002118:Calcium