title: TGF-β2 reduces nitric oxide synthase mRNA through a ROCK-dependent pathway in airway epithelial cells.
abstract: Exhaled NO (eNO) is a potential noninvasive biomarker of inflammation in asthma. The significant intersubject variability of eNO within clinically similar patients has contributed to its limited clinical application. Arginase and NO synthase (NOS) utilize the same substrate (l-arginine) and contribute to the fibrotic and inflammatory features of asthma, respectively. Interestingly, TGF-β(2) can increase the expression of arginase, stimulates fibrosis, and is overexpressed in asthma. We hypothesized that TGF-β(2)-enhanced arginase activity would decrease gas phase NO release from lung epithelial cells by limiting l-arginine availability for NOS. Our results show that TGF-β(2) (5 ng/ml) significantly enhances total arginase activity up to two- to threefold in both primary small airway epithelial cells (SAECs) and the A549 cell line. Preincubation with TGF-β(2) prior to cytokine (IL-1β, TNF-α, and IFN-γ, 10 ng/ml each) stimulation decreases gas phase NO release to baseline levels (from 1.66 ± 0.52 to 0.30 ± 0.12 pl·s(-1)·cm(-2) and from 0.27 ± 0.03 pl·s(-1)·cm(-2) to near zero in SAEC and A549 cells, respectively). Addition of arginase inhibitor (N(ω)-hydroxy-nor-l-arginine) or small interfering RNA only partly reverses the reduction. In contrast, Rho-kinase (ROCK) pathway inhibitor (Y-27632) completely recovers the cytokine-induced NO flux in the present of TGF-β(2). Inducible NO synthase (iNOS) mRNA and protein levels change in a similar trend as NO release from the cells. We conclude that TGF-β(2) impacts cytokine-induced NO production in airway epithelial cells by reducing iNOS mRNA and protein levels through a ROCK-dependent pathway.
authors.affiliation: Dept. of Biomedical Engineering, 2420 Engineering Hall, Univ. of California, Irvine, Irvine, CA 92697-2730, USA.
authors.forename: Jingjing
authors.initials: J
authors.lastname: Jiang
authors.forename: Steven C
authors.initials: SC
authors.lastname: George
mesh_terms: D000577:Amides / Q000494:pharmacology; D001119:Arginase / Q000037:antagonists & inhibitors / Q000378:metabolism; D001120:Arginine / Q000031:analogs & derivatives / Q000494:pharmacology; D001249:Asthma / Q000378:metabolism; D002460:Cell Line; D004847:Epithelial Cells / Q000378:metabolism; D006801:Humans; D008168:Lung / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000096:biosynthesis / Q000378:metabolism; D011725:Pyridines / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D053781:Transforming Growth Factor beta2 / Q000378:metabolism*; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors
chemical_list: D000577:Amides; C404735:N(omega)-hydroxynorarginine; D011725:Pyridines; D012333:RNA, Messenger; D053781:Transforming Growth Factor beta2; C108830:Y 27632; D001120:Arginine; D019001:Nitric Oxide Synthase; D054460:rho-Associated Kinases; D001119:Arginase