R Castro1, X H Sun, X-B Liu, J R Martinez, G H Zhang. 1. Department of Pediatrics, University of Texas Health Science Center, San Antonio, TX 78229-3900, USA. castror@uthscsa.edu
Abstract
OBJECTIVE: Pulmonary surfactants reduce alveolar surface tension and alter inflammatory cell function. We studied the effects of surfactant preparations on Ca2+ influx regulated by protein kinase C (PKC) and mitogen-activated protein kinases (MAPK) and cytokine secretion in the alveolar macrophage (AM) cell line NR8383. METHODS: Fura-2-loaded AMs were stimulated with zymosan (200 microg/ml), 1,2-dioctanoyl-sn-glycerol (DOG, 20 microM) or C6-ceramide (C6C, 10 microM) in the presence of exogenous surfactants (beractant, calfactant or colfosceril) or surfactant phospholipid (dipalmitoyl phosphatidylcholine, DPPC), at 250 microg/ml phospholipid and changes in cytosolic free Ca2+ (Delta[Ca2+]i) and cytokines were measured. RESULTS: Zymosan-induced Delta[Ca2+]i (117 +/- 5 nM) at 3 min was reduced (p <0.001) by beractant (50 +/- 6 nM), colfosceril (61 +/- 2 nM), calfactant (46 +/- 5 nM), and DPPC (52 +/- 5 nM). Beractant inhibited the Delta[Ca2+]i by PKC stimulation with DOG and all preparations reduced the MAPK-induced Ca2+ influx by C6C. Beractant and Ca2+ channel blocker SKF 96365 (10 microM) together abolished the zymosan-stimulated Delta[Ca2+]i. Zymosan-stimulated TNF-alpha and IL-1beta secretion was also inhibited by surfactant pretreatment. CONCLUSIONS: These results indicate that exogenous surfactant inhibits Ca2+ influx and cytokine secretion in zymosan-stimulated AMs. This anti-inflammatory activity may be through an interaction with downstream signaling elements or Ca2+ channels.
OBJECTIVE: Pulmonary surfactants reduce alveolar surface tension and alter inflammatory cell function. We studied the effects of surfactant preparations on Ca2+ influx regulated by protein kinase C (PKC) and mitogen-activated protein kinases (MAPK) and cytokine secretion in the alveolar macrophage (AM) cell line NR8383. METHODS: Fura-2-loaded AMs were stimulated with zymosan (200 microg/ml), 1,2-dioctanoyl-sn-glycerol (DOG, 20 microM) or C6-ceramide (C6C, 10 microM) in the presence of exogenous surfactants (beractant, calfactant or colfosceril) or surfactant phospholipid (dipalmitoyl phosphatidylcholine, DPPC), at 250 microg/ml phospholipid and changes in cytosolic free Ca2+ (Delta[Ca2+]i) and cytokines were measured. RESULTS:Zymosan-induced Delta[Ca2+]i (117 +/- 5 nM) at 3 min was reduced (p <0.001) by beractant (50 +/- 6 nM), colfosceril (61 +/- 2 nM), calfactant (46 +/- 5 nM), and DPPC (52 +/- 5 nM). Beractant inhibited the Delta[Ca2+]i by PKC stimulation with DOG and all preparations reduced the MAPK-induced Ca2+ influx by C6C. Beractant and Ca2+ channel blocker SKF 96365 (10 microM) together abolished the zymosan-stimulated Delta[Ca2+]i. Zymosan-stimulated TNF-alpha and IL-1beta secretion was also inhibited by surfactant pretreatment. CONCLUSIONS: These results indicate that exogenous surfactant inhibits Ca2+ influx and cytokine secretion in zymosan-stimulated AMs. This anti-inflammatory activity may be through an interaction with downstream signaling elements or Ca2+ channels.