Transcriptional regulation of iNOS by IL-1 beta in cultured rat pulmonary artery smooth muscle cells.
Transcriptional regulation of iNOS by IL-1 beta in cultured rat pulmonary artery smooth muscle cells. Am. J. Physiol. 271 (Lung Cell. Mol. Physiol. 15): L166-L171, 1996.-Interleukin-1 beta (IL-1 beta) is the critical cytokine affecting peripheral vascular expression of inducible nitric oxide synthase (iNOS). Accordingly, we sought to determine a role for IL-1 beta in stimulating iNOS transcription in cultured rat pulmonary artery smooth muscle cells (RPASMC). Treatment of RPASMC with IL-1 beta caused a concentration-dependent increase in iNOS gene expression by Northern and Western blotting. To demonstrate IL-1 beta-mediated transcriptional activation, we used transient liposome-mediated transfection of RPASMC with promoter-luciferase constructs containing deletional mutations of the murine macrophage iNOS 5' flanking promoter region. IL-1 beta increased promoter activity approximately two- to threefold over baseline in fragments ranging from -1592 (full-length) to -242 bp. Activity was lost, however, when the promoter fragment was shorter than -242 bp. IL-1 beta-mediated increases in steady-state iNOS mRNA were sensitive to pyrrolidine dithiocarbamate (PDTC), an inhibitor of NF-kappa B activation. Nuclear proteins from IL-1 beta-stimulated cells demonstrated PDTC-sensitive binding to an oligonucleotide containing the sequence for the NF-kappa B binding element present in the region between -242 and -42 bp. These data document that IL-1 beta, by itself, increases iNOS expression in RPASMC by transcriptional activation, mediated in part by NF-kappa B.
Wong
H R
HR
Department of Pharmacology, University of Pittsburgh School of Medicine, Pennsylvania 15261, USA.
Finder
J D
JD
Wasserloos
K
K
Lowenstein
C J
CJ
Geller
D A
DA
Billiar
T R
TR
Pitt
B R
BR
Davies
P
P
D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D004586:Electrophoresis; D004790:Enzyme Induction / Q000187:drug effects; D015870:Gene Expression / Q000187:drug effects; D007375:Interleukin-1 / Q000494:pharmacology*; D008297:Male; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000201:enzymology*; D016328:NF-kappa B / Q000037:antagonists & inhibitors; D019001:Nitric Oxide Synthase / Q000235:genetics* / Q000378:metabolism*; D015345:Oligonucleotide Probes; D011651:Pulmonary Artery / Q000166:cytology / Q000201:enzymology*; D011759:Pyrrolidines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D013859:Thiocarbamates / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects*
D007375:Interleukin-1; D016328:NF-kappa B; D015345:Oligonucleotide Probes; D011759:Pyrrolidines; D013859:Thiocarbamates; C020972:pyrrolidine dithiocarbamic acid; D019001:Nitric Oxide Synthase