Cytokine-activated Jak-2 is involved in inducible nitric oxide synthase expression independent from NF-kappaB activation in vascular smooth muscle cells.
Inflammatory cytokines, such as interleukin (IL)-1beta and tumor necrosis factor (TNF)-alpha, activate nuclear factor-kappa B (NF-kappaB) which transactivates inducible nitric oxide synthase (iNOS) gene in vascular smooth muscle cells (VSMCs). However, it remains obscure whether cytokine-mediated iNOS expression in VSMCs requires signaling pathway(s) other than NF-kappaB activation. The present study was designed to elucidate whether protein tyrosine kinases (PTKs) are involved in the cytokine-induced NF-kappaB activation and iNOS expression in cultured rat VSMCs. Both IL-1beta and TNF-alpha stimulated NF-kappaB activity, iNOS mRNA and protein expression with massive nitrite/nitrate (NOx) production in rat VSMCs. PTK inhibitors (genistein, herbimycin A) dose-dependently inhibited the cytokine-stimulated NOx production and iNOS mRNA expression. However, neither genistein nor herbimycin A affected the cytokine-stimulated phosphorylation and degradation of IkappaB-alpha, or NF-kappaB activation, whereas they completely blocked the cytokine-stimulated iNOS transcriptional activity. Tyrphostin B42 (AG490), a Jak-2 tyrosine kinase inhibitor, similarly blocked the cytokine-induced NOx production, iNOS expression and its promoter activity without affecting NF-kappaB-dependent transcription. Transfection of a dominant-negative Jak-2 mutant antagonized the cytokine-induced NOx production and iNOS expression, while wild-type Jak-2 expressing construct was without effect. These data indicate that the cytokine-induced iNOS expression involves activation of Jak-2 signaling pathway independent from NF-kappaB activation in rat VSMCs.
Doi
Masaru
M
Department of Clinical and Molecular Endocrinology, Tokyo Medical and Dental University Graduate School, 1-5-45, Yushima, Bunkyo-ku 113-8519, Tokyo, Japan.
Shichiri
Masayoshi
M
Katsuyama
Koichi
K
Ishimaru
Shinya
S
Hirata
Yukio
Y
D000818:Animals; D001665:Binding Sites / Q000187:drug effects; D016207:Cytokines / Q000378:metabolism* / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007375:Interleukin-1 / Q000378:metabolism; D053614:Janus Kinase 2; D008297:Male; D023421:Models, Animal; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000201:enzymology; D016328:NF-kappa B / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000096:biosynthesis*; D052247:Nitric Oxide Synthase Type II; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D011518:Proto-Oncogene Proteins; D051381:Rats; D017208:Rats, Wistar; D015533:Transcriptional Activation / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D016207:Cytokines; D004791:Enzyme Inhibitors; D007375:Interleukin-1; D016328:NF-kappa B; D011518:Proto-Oncogene Proteins; D014409:Tumor Necrosis Factor-alpha; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat; D011505:Protein-Tyrosine Kinases; C507927:Jak2 protein, rat; D053614:Janus Kinase 2