Abstract
The effects of 2-naphthylethyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline (THI 53), on nitric oxide (NO) production and inducible nitric-oxide synthase (iNOS) protein induction by lipopolysaccharide (LPS) were investigated in RAW 264.7 cells and mice. In cells, THI 53 concentration dependently reduced NO production and iNOS protein induction by LPS. In addition, THI 53 inhibited NO production and iNOS protein induction in LPS-treated mice. LPS-mediated iNOS protein induction was inhibited significantly by the specific tyrosine kinase inhibitor α-cyano-(3-hydroxy-4-nitro)cinnamonitrile (AG126) as well as by THI 53. In addition, a c-Jun NH2-terminal kinase (JNK) inhibitor anthra[1,9-cd]pyrazole-6 (2H)-one) (SP600125) but not an extracellular regulated kinase inhibitor [2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one (PD98029)] or a p38 inhibitor [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole (SB230580)] reduced the iNOS protein level induced by LPS. Moreover, a Janus kinase 2 (JAK2) inhibitor α-cyano-(3,4-dihydroxy)-N-benzylcinnamide (AG490) dose-dependently prevented LPS-mediated iNOS protein induction. LPS activated phosphorylations of tyrosine kinases, especially tyrosine kinase 2 (Tyk2) and signal transducer and activator of transcription-1 (STAT-1); these were reduced by THI 53. LPS also phosphorylated the JNK pathway; however, this phosphorylation was unaffected by THI 53. Interestingly, a JNK inhibitor (SP600125) and another tyrosine kinase inhibitor (genistein) significantly inhibited STAT-1 phosphorylation, suggesting that the LPS-activated JNK pathway and a tyrosine kinase pathway (especially Tyk2) may link to the STAT-1 pathway, which is involved in iNOS induction. However, THI 53 regulates LPS-mediated iNOS protein induction by affecting the Tyk2/JAK2-STAT-1 pathway, not the JNK pathway. The inhibition by THI 53 of LPS-induced NO production was recovered by a tyrosine phosphatase inhibitor (Na3VO4), which supports the possibility that THI 53 inhibits the LPS-induced inflammatory response through regulation of tyrosine kinase pathways. THI 53 also inhibited LPS-mediated interferon (IFN)-β production and nuclear factor-κB (NF-κB) activation. Thus, THI 53 may regulate LPS-mediated inflammatory response through both the NF-κB and IFN-β/Tyk2/JAK2-STAT-1 pathways.
Footnotes
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This study was supported by the Korea Science and Engineering Foundation (R13-2005-005-01003-0).
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H.J.K. and K.T. contributed equally to this work.
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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.106.112052.
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ABBREVIATIONS: iNOS, inducible nitric-oxide synthase; NO, nitric oxide; LPS, lipopolysaccharide; NF-κB, nuclear factor-κB; MAPK, mitogen-activated protein kinase; ERK, extracellular regulated kinase; JNK, c-Jun N-terminal kinase; JAK, Janus protein tyrosine kinase; IκB, inhibitor of NF-κB; STAT, signal transducer and activator of transcription; Tyk2, tyrosine kinase 2; THI 53, 2-naphtylethyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline; AG490, α-cyano-(3,4-dihydroxy)-N-benzylcinnamide; AG126, α-cyano-(3-hydroxy-4-nitro)cinnamonitrile; SB203580, [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole]; PD98029, ·; ECL, enhanced chemoluminescence; SP600125 (anthra[1,9-cd]pyrazole-6 (2H)-one); MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium; NOx, nitrite; PVDF, polyvinylidene difluoride; BSA, bovine serum albumin; IFN, interferon; MAP, mitogen-activated protein; AP-1, activator protein-1; THI 52, 1-naphthylethyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline; TLR4, Toll-like receptor 4; PD98029, 2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one.
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↵1 Current affiliation: Department of Pharmacology, College of Medicine, Yeungnam University, Daegu, Korea.
- Received August 4, 2006.
- Accepted November 13, 2006.
- The American Society for Pharmacology and Experimental Therapeutics
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