Modulation of endothelin-1-induced cytosolic free calcium mobilization and mitogen-activated protein kinase activation by erythropoietin in vascular smooth muscle cells.
BACKGROUND
It has been reported that human recombinant erythropoietin (rHuEPO) modulates the sensitivity of the cardiovascular system to vasoconstrictors. We investigated whether rHuEPO has modulative effects on the endothelin-1 (ET-1)-induced elevation of cytosolic free calcium concentration ([Ca2+]i) and mitogen-activated protein (MAP) kinase activation in vascular smooth muscle cells (VSMC).


METHODS
[Ca2+]i was measured by fura-2/AM, and MAP kinase activation was analyzed by Western blotting.


RESULTS
Exposure of VSMC to rHuEPO prior to stimulation with ET-1 enhanced both basal and ET-1-induced elevation of [Ca2+]i in a dose-dependent manner in the presence of extracellular Ca2+. The synergistic effect was also retained in the absence of extracellular Ca2+ after exposure to rHuEPO. However, the effect was diminished in the presence of extracellular Ca2+ combined with the intracellular Ca2+ release inhibitor TMB-8, PKC inhibitor, or PKC depletion. Exposure to rHuEPO also had a synergistic effect on the activation of MAP kinase induced by ET-1; however, this effect was diminished in the presence of the Ca2+ chelator BAPTA-AM.


CONCLUSIONS
The results suggest that rHuEPO has synergistic effects on ET-1-induced [Ca2+]i mobilization, particularly on intracellular Ca2+ release, and MAP kinase activation in VSMC.
Kusano
E
E
Department of Nephrology, Jichi Medical School, Tochigi, Japan. eishun@jichi.ac.jp
Akimoto
T
T
Umino
T
T
Yanagiba
S
S
Inoue
M
M
Ito
C
C
Ando
Y
Y
Asano
Y
Y
D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism*; D019332:Endothelin-1 / Q000494:pharmacology*; D004789:Enzyme Activation; D004921:Erythropoietin / Q000494:pharmacology*; D005110:Extracellular Space / Q000378:metabolism; D006801:Humans; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D008297:Male; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D009994:Osmolar Concentration; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000172:deficiency / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D019332:Endothelin-1; D007476:Ionophores; D011994:Recombinant Proteins; D004921:Erythropoietin; D015759:Ionomycin; D011493:Protein Kinase C; D020928:Mitogen-Activated Protein Kinases; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium