Amiodarone and N-desethylamiodarone enhance endothelial nitric oxide production in human endothelial cells.
Amiodarone (AM) is a potent vasodilator and exhibits diverse cardiovascular protective effects in vivo, but their underlying mechanisms remain unsettled. We investigated the effects of AM and N-desethylamiodarone (DEA), the major metabolite of AM, on endothelial nitric oxide (NO) production using cultured human umbilical vein endothelial cells (HUVECs). The release of NO was evaluated as measured by nitrite, a stable metabolite of NO, using the Griess reaction and also measured directly by a NO-selective electrode. The expression of each nitric oxide synthase (NOS) mRNA was examined by reverse transcriptase-polymerase chain reaction (RT-PCR), and the effects of AM on eNOS mRNA expression were studied by quantitative real-time RT-PCR. AM and DEA (1-30 microM) enhanced NO production in a concentration-dependent manner. DEA was capable of producing more NO than AM. L-NAME, a nonselective NOS inhibitor, EGTA, a Ca(2+)-chelating agent, and nickel, a nonspecific Ca(2+) blocker, all inhibited AM-induced NO production. However, LY294002, an Akt pathway inhibitor and SB202190, a MAP kinase inhibitor, did not significantly suppress the production. In RT-PCR analysis, only eNOS mRNA was detected. Treatment with AM for 4 hours did not show a significant increase in the expression of eNOS mRNA. AM lower than 30 microM did not induce apoptosis, net cell loss, or LDH release from cells. The present study provides the first evidence that therapeutic concentrations of AM and DEA enhance eNOS-mediated NO production without any toxic or apoptotic effects. This mechanism may underlie the cardiovascular protective effects of AM and its metabolite observed in a clinical setting.
Department of Cardiovascular Medicine, University of Tokyo, Tokyo, Japan.
Shinya
S
Kishida
Toshiaki
T
Nakajima
Ji
J
Ma
Taisuke
T
Jo
Hiroyuki
H
Imuta
Hitoshi
H
Oonuma
Haruko
H
Iida
Haruhito
H
Takano
Toshihiro
T
Morita
Ryozo
R
Nagai
D000638:Amiodarone / Q000031:analogs & derivatives / Q000494:pharmacology*; D018929:Cell Culture Techniques; D004730:Endothelium, Vascular / Q000187:drug effects*; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D009569:Nitric Oxide / Q000096:biosynthesis*; D019001:Nitric Oxide Synthase / Q000096:biosynthesis*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014665:Vasodilator Agents / Q000494:pharmacology*
D014665:Vasodilator Agents; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; C036116:desethylamiodarone; D000638:Amiodarone