title: Chitinase activates protease-activated receptor-2 in human airway epithelial cells.
abstract: Mammalian chitinase released by airway epithelia is thought to be an important mediator of disease manifestation in an experimental model of asthma. However, the intracellular signaling mechanisms engaged by exogenous chitinase in human airway epithelial cells are unknown. Here, we investigated the direct effects of exogenous chitinase from Streptomyces griseus on Ca(2+) signaling in human airway epithelial cells. Spectrofluorometry was used to measure intracellular Ca(2+) concentration ([Ca(2+)](i)) in fura-2-AM-loaded cells. S. griseus chitinase induced dose-dependent [Ca(2+)](i) increases in normal human bronchial epithelial cells and promoted [Ca(2+)](i) oscillations in H292 cells. Chitinase-induced [Ca(2+)](i) oscillations were independent of extracellular Ca(2+), suggesting that the observed [Ca(2+)](i) increases were due to Ca(2+) release from intracellular stores. Accordingly, after depleting endoplasmic reticulum (ER) Ca(2+) with the ER Ca(2+) ATPase inhibitor, thapsigargin, chitinase-mediated [Ca(2+)](i) increases were abolished. Treatment with the phospholipase C (PLC) inhibitor U73122 or the 1, 4, 5-trisinositolphosphate (IP(3)) receptor inhibitor 2-APB attenuated chitinase-induced [Ca(2+)](i) increases. Desensitization of protease-activated receptor-2 (PAR-2) by repetitive agonist stimulation or siRNA-mediated PAR-2 knock-down revealed that chitinase-mediated [Ca(2+)](i) increases were exclusively mediated by PAR-2 activation. Finally, chitinase was found to cleave a model peptide representing the cleavage site of PAR-2 and enhanced IL-8 production. These results indicate that exogenous chitinase is a potent proteolytic activator of PAR-2 that can directly induce PLC/IP(3)-dependent Ca(2+) signaling in human airway epithelial cells.
authors.affiliation: Department of Oral Biology, Brain Korea 21 Project, Center for Natural Defense System, Yonsei University College of Dentistry, Seoul, Korea.
authors.forename: Jeong Hee
authors.initials: JH
authors.lastname: Hong
authors.forename: Jung Yeon
authors.initials: JY
authors.lastname: Hong
authors.forename: Boryung
authors.initials: B
authors.lastname: Park
authors.forename: Syng-Ill
authors.initials: SI
authors.lastname: Lee
authors.forename: Jeong Taeg
authors.initials: JT
authors.lastname: Seo
authors.forename: Kyu-Earn
authors.initials: KE
authors.lastname: Kim
authors.forename: Myung Hyun
authors.initials: MH
authors.lastname: Sohn
authors.forename: Dong Min
authors.initials: DM
authors.lastname: Shin
mesh_terms: D001980:Bronchi / Q000166:cytology* / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling; D002478:Cells, Cultured; D002688:Chitinases / Q000378:metabolism*; D004847:Epithelial Cells / Q000378:metabolism*; D006801:Humans; D016209:Interleukin-8 / Q000096:biosynthesis; D007700:Kinetics; D044464:Receptor, PAR-2 / Q000235:genetics / Q000378:metabolism*; D013305:Streptomyces griseus / Q000201:enzymology*; D013379:Substrate Specificity
chemical_list: D016209:Interleukin-8; D044464:Receptor, PAR-2; D002688:Chitinases; D002118:Calcium