Chitinase activates protease-activated receptor-2 in human airway epithelial cells.
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.
Hong
Jeong Hee
JH
Department of Oral Biology, Brain Korea 21 Project, Center for Natural Defense System, Yonsei University College of Dentistry, Seoul, Korea.
Hong
Jung Yeon
JY
Park
Boryung
B
Lee
Syng-Ill
SI
Seo
Jeong Taeg
JT
Kim
Kyu-Earn
KE
Sohn
Myung Hyun
MH
Shin
Dong Min
DM
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
D016209:Interleukin-8; D044464:Receptor, PAR-2; D002688:Chitinases; D002118:Calcium