Down-regulation of ERK but not MEK phosphorylation in cultured endothelial cells by repeated changes in cyclic stretch.
OBJECTIVE
Effects of cyclic stretch on endothelial cells are studied usually by exposing cells cultured under stretch-free conditions to some levels of cyclic stretch, but in vivo these cells experience both increase and decrease in stretch. Experiments were designed to study how endothelial cells maintained under certain levels of cyclic stretch responded to shifts in stretch frequencies and amplitudes.


METHODS
Confluent endothelial cells cultured on flexible silicone membranes with or without pre-stretching for 2-12 h were exposed to various levels of stretch amplitude or frequency and assayed for extracellular signal-regulated kinase 1/2 (ERK) phosphorylation.


RESULTS
When endothelial cells without pre-stretching were cyclically stretched, ERK phosphorylation increased, peaking approximately 15 min and slowly decreased. In contrast, when pre-stretched cells were exposed to either higher or lower stretch condition, ERK phosphorylation transiently decreased within 5 min, indicating that some mechanism which down-regulated ERK phosphorylation was activated. Because phosphorylation of ERK kinase (MEK) was not inhibited in these cells, this mechanism targeted ERK directly, not the upstream kinases of the Ras-Raf-MEK-ERK cascade. Furthermore, this ERK down-regulation in pre-stretched cells was not induced by agonists, was inhibited by Na(3)VO(4) but not okadaic acid, and was detected in the cytosolic fraction. Repeated shifts in stretch conditions induced continuous down-regulation of ERK but not MEK phosphorylation.


CONCLUSIONS
Endothelial cells are capable of down-regulating ERK phosphorylation in a cyclic stretch- and tyrosine phosphatase-dependent manner. Frequent changes in stretch conditions constitutively activated this ability, which could play some role in regulating ERK activity in endothelial cells in vivo.
Shi
Feng
F
Cardiovascular Research Institute, University of Rochester, 601 Elmwood Avenue, Box 679, Rochester, NY 14642, USA.
Chiu
Yi-Jen
YJ
Cho
Youngsun
Y
Bullard
Tara A
TA
Sokabe
Masahiro
M
Fujiwara
Keigi
K
D000818:Animals; D001011:Aorta; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D002417:Cattle; D048429:Cell Size; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D015536:Down-Regulation; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D005419:Flavonoids / Q000494:pharmacology; D007106:Immune Sera / Q000494:pharmacology; D040542:Mechanotransduction, Cellular / Q000187:drug effects / Q000502:physiology*; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism*; D019319:Okadaic Acid / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors; D010766:Phosphorylation / Q000187:drug effects; D014635:Valproic Acid / Q000494:pharmacology
D004791:Enzyme Inhibitors; D005419:Flavonoids; D007106:Immune Sera; D019319:Okadaic Acid; D014635:Valproic Acid; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D020929:Mitogen-Activated Protein Kinase Kinases; D010749:Phosphoprotein Phosphatases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one