Abstract
Sphingosine 1-phosphate (S1P) is a lipid mediator that exerts potent and diverse biological effects on several cardiovascular cells. We investigated the effect of S1P on interleukin (IL)-1β-induced nitric oxide (NO) production and inducible NO synthase (iNOS) expression in rat vascular smooth muscle cells (VSMCs). S1P inhibited NO production at concentrations higher than 0.1 μM; this was associated with the inhibition of iNOS protein and mRNA expression. S1P also inhibited IL-1β-induced GTP cyclohydrolase I (GTPCH) mRNA expression. Pertussis toxin (PTX) partially attenuated the inhibitory effects of S1P on NO production and iNOS protein induction, whereas it completely blocked the inhibitory effects on iNOS and GTPCH mRNA expression. S1P inhibited iNOS expression in Ca2+-depleted conditions; PTX did not modify this effect. The Rho kinase inhibitor Y 27632 partially but significantly attenuated the inhibitory effect of S1P on iNOS expression in Ca2+-depleted condition but did not affect it in the presence of Ca2+. S1P significantly inhibited IL-1β-induced persistent activation of extracellular signal-regulated kinase (ERK) but had no effect in Ca2+-depleted conditions. Thus, S1P inhibits IL-1β induction of NO production and iNOS expression in rat VSMCs through multiple mechanisms involving both PTX-sensitive and -insensitive G proteins coupled to S1P receptors. Furthermore, Ca2+-dependent ERK inhibition and Ca2+-independent Rho kinase activation might be involved in the inhibitory mechanism of iNOS expression. Through its action on NO production by VSMCs, S1P may play an important role in the progression of local vascular injury associated with thrombosis, atherosclerosis, and hypertension.
Footnotes
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This study was supported in part by The High Technology Research Program from the Ministry of Education, Science, Sports and Culture of Japan and by The Akiyama Foundation. R.L. was supported by National Institutes of Health Grants HL34215, HL73400, HL47073, and HL46403.
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.107.127290.
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ABBREVIATIONS: NO, nitric oxide; NOS, nitric-oxide synthase; VSMC, vascular smooth muscle cell; iNOS, inducible nitric-oxide synthase; IL, interleukin; ERK, extracellular signal-regulated kinase; S1P, sphingosine 1-phosphate; PTX, pertussis toxin; MAPK, mitogen-activated protein kinase; eNOS, endothelial nitric-oxide synthase; nNOS, neuronal nitric-oxide synthase; GTPCH, GTP cyclohydrolase I; RT, reverse transcription; PCR, polymerase chain reaction; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; BH4, tetrahydrobiopterin; BAPTA-AM, 1,2-bis-(o-aminophenoxy)-ethane-N,N,N′,N′-tetraacetic acid tetraacetoxy-methyl ester; PD 98059, 2′-amino-3′-methoxyflavone; SB 203580, 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)5-(4-pyridyl)1H-imidazole; GF 109203X, 2-[1-(3-dimethylaminopropyl)-1H-indol-3-yl]-3-(1H-indol-3-yl)-maleimide; Y 27632, (R)-(+)-trans-N-(4-pyridyl)-4-(1-aminoethyl)-cyclohexanecarboxamide, 2HCl; rottlerin, 1-[6-[(3-acetyl-2,4,6-trihydroxy-5-methylphenyl)methyl]-5,7-dihydroxy-2,2-dimethyl-2H-1-benzopyran-8-yl]-3-phenyl-2-propen-1-one; 1400W, N-([3-(aminomethyl)phenyl]methyl)ethanimidamide dihydrochloride; PKC, protein kinase C; p-ERK, phosphorylated ERK; t-ERK, total ERK.
- Received June 14, 2007.
- Accepted December 28, 2007.
- The American Society for Pharmacology and Experimental Therapeutics
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