Erythropoietin modulates angiotensin II- or noradrenaline-induced Ca(2+) mobilization in cultured rat vascular smooth-muscle cells.
BACKGROUND
It has been reported that human recombinant erythropoietin (rHuEpo) modulates the sensitivity of the cardiovascular system to angiotensin II (Ang II) or noradrenaline (NA). In the present study, we explored the effect of rHuEpo on the responsiveness of Ang II- or NA-induced cytosolic free calcium ([Ca(2+)]i) mobilization in cultured rat vascular smooth-muscle cells (VSMC).


METHODS
[Ca(2+)]i concentrations in VSMC were measured by using the calcium-sensitive fluorescent dye fura-2.


RESULTS
The addition of rHuEpo (250 U/ml) alone induced elevation in [Ca(2+)]i, which remained significantly elevated above basal level for at least 60 min in the presence of extracellular Ca(2+). Pre-incubation with specific protein kinase C (PKC) inhibitor calphostin C (1 micromol/l) significantly reduced the peak and the sustained elevations of [Ca(2+)]i. Pre-treatment with rHuEpo for 60 min increased both basal [Ca(2+)]i and the changes in [Ca(2+)]i by Ang II or NA in a dose-dependent manner in the presence of extracellular Ca(2+). The synergistic effects of rHuEpo with Ang II or NA were also retained when VSMC were bathed in the Ca(2+)-free medium after the pre-incubation of rHuEpo. Conversely, they were diminished in the presence of extracellular Ca(2+) combined with intracellular Ca(2+) release inhibitor 8-(NN-diethylamino)octyl-1,3,4,5-trimethoxybenzoate (TMB-8). The synergistic effects of rHuEpo were also diminished by PKC depletion or by PKC inhibitor.


CONCLUSIONS
These observations suggest that rHuEpo has synergistic effects on Ang II- or NA-induced [Ca(2+)]i mobilization, particularly on intracellular Ca(2+) release, in VSMC. This may be a potential mechanism contributing to hypertension associated with rHuEpo therapy.
Department of Nephrology, Jichi Medical School, Tochigi, Japan.
T
T
Akimoto
E
E
Kusano
N
N
Fujita
K
K
Okada
O
O
Saito
S
S
Ono
Y
Y
Ando
S
S
Homma
T
T
Saito
Y
Y
Asano
D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D004357:Drug Synergism; D004789:Enzyme Activation; D004921:Erythropoietin / Q000494:pharmacology*; D005110:Extracellular Space / Q000378:metabolism; D005707:Gallic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D009638:Norepinephrine / Q000494:pharmacology*; D009994:Osmolar Concentration; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D002121:Calcium Channel Blockers; D007476:Ionophores; D011994:Recombinant Proteins; D004921:Erythropoietin; D000804:Angiotensin II; D015759:Ionomycin; C006014:8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate; D005707:Gallic Acid; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; D009638:Norepinephrine