Regulation of Lipopolysaccharide-Induced Inducible Nitric-Oxide Synthase Expression through the Nuclear Factor-κB Pathway and Interferon-β/Tyrosine Kinase 2/Janus Tyrosine Kinase 2-Signal Transducer and Activator of Transcription-1 Signaling Cascades by 2-Naphthylethyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline (THI 53), a New Synthetic Isoquinoline Alkaloid

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 alpha-cyano-(3-hydroxy-4-nitro)cinnamonitrile (AG126) as well as by THI 53. In addition, a c-Jun NH(2)-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 alpha-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 (Na(3)VO(4)), 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)-beta production and nuclear factor-kappaB (NF-kappaB) activation. Thus, THI 53 may regulate LPS-mediated inflammatory response through both the NF-kappaB and IFN-beta/Tyk2/JAK2-STAT-1 pathways.