TY - JOUR
T1 - H2O2 injury causes Ca2+-dependent and independent hydrolysis of phosphatidylcholine in alveolar epithelial cells
AU - Rice, K. L.
AU - Duane, P. G.
AU - Archer, S. L.
AU - Gilboe, D. P.
AU - Niewoehner, D. E.
PY - 1992
Y1 - 1992
N2 - Oxidants may play a central role in the pathogenesis of adult respiratory distress syndrome, and phospholipase activation is a potential mechanism of oxidant-induced injury of alveolar epithelial cells. Studies were performed in rat alveolar type II epithelial cells (RAEC) after 3 days in culture. As measured by 51Cr and lactate dehydrogenase release, H2O2 caused time- and dose-dependent cytotoxicity to RAEC. RAEC phospholipids labeled with [14C]- stearic acid ([14C]SA) and [3H]arachidonic acid ([3H]AA) released free fatty acids in response to H2O2 in a manner that closely paralleled the cytotoxicity indexes. Analysis of phospholipid subclasses indicated that phosphatidylcholine was preferentially affected. Analysis for putative products of phospholipase activity revealed significant increases in diacylglycerol and phosphorylcholine, expected products of phospholipase C, as well as significant increases in L-α-lysophosphatidylcholine and L-α- glycerophosphocholine, expected products of phospholipase A2. Increases in phospholipase D activity were not detected. To determine whether H2O2- stimulated phospholipase activity might be Ca2+ stimulated, RAEC were loaded with fura-2/AM, and changes in intracellular Ca2+ concentrations ([Ca2+](i)) were monitored by epifluorescent microscopy. Exposure to H2O2 caused elevations in [Ca2+](i), and the time and dose relationships were consistent with the hypothesis that the release of [14C]SA and [3H]AA is related to changes in cellular Ca2+ concentrations. Additionally, pretreatment with MAPTAM, an intracellular chelator of calcium, partially blocked H2O2-mediated [3H]AA liberation. However, experiments in saponin- permeabilized RAEC, in which [Ca2+](i) was strongly buffered by ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'-tetraacetic acid, indicate that H2O2-induced phospholipase activity also has a Ca2+-independent component. Both H2O2-mediated increases in [Ca2+](i) and liberation of free fatty acids may be triggered by a loss of energy stores, as both events are preceded by profound depletion of cellular ATP levels.
AB - Oxidants may play a central role in the pathogenesis of adult respiratory distress syndrome, and phospholipase activation is a potential mechanism of oxidant-induced injury of alveolar epithelial cells. Studies were performed in rat alveolar type II epithelial cells (RAEC) after 3 days in culture. As measured by 51Cr and lactate dehydrogenase release, H2O2 caused time- and dose-dependent cytotoxicity to RAEC. RAEC phospholipids labeled with [14C]- stearic acid ([14C]SA) and [3H]arachidonic acid ([3H]AA) released free fatty acids in response to H2O2 in a manner that closely paralleled the cytotoxicity indexes. Analysis of phospholipid subclasses indicated that phosphatidylcholine was preferentially affected. Analysis for putative products of phospholipase activity revealed significant increases in diacylglycerol and phosphorylcholine, expected products of phospholipase C, as well as significant increases in L-α-lysophosphatidylcholine and L-α- glycerophosphocholine, expected products of phospholipase A2. Increases in phospholipase D activity were not detected. To determine whether H2O2- stimulated phospholipase activity might be Ca2+ stimulated, RAEC were loaded with fura-2/AM, and changes in intracellular Ca2+ concentrations ([Ca2+](i)) were monitored by epifluorescent microscopy. Exposure to H2O2 caused elevations in [Ca2+](i), and the time and dose relationships were consistent with the hypothesis that the release of [14C]SA and [3H]AA is related to changes in cellular Ca2+ concentrations. Additionally, pretreatment with MAPTAM, an intracellular chelator of calcium, partially blocked H2O2-mediated [3H]AA liberation. However, experiments in saponin- permeabilized RAEC, in which [Ca2+](i) was strongly buffered by ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'-tetraacetic acid, indicate that H2O2-induced phospholipase activity also has a Ca2+-independent component. Both H2O2-mediated increases in [Ca2+](i) and liberation of free fatty acids may be triggered by a loss of energy stores, as both events are preceded by profound depletion of cellular ATP levels.
KW - adenosine 5'- triphosphate
KW - calcium ions
KW - hydrogen peroxide
KW - lung epithelial cells
KW - oxidants
KW - phospholipases
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U2 - 10.1152/ajplung.1992.263.4.l430
DO - 10.1152/ajplung.1992.263.4.l430
M3 - Article
C2 - 1415720
AN - SCOPUS:0026940030
SN - 0002-9513
VL - 263
SP - L430-L438
JO - American Journal of Physiology - Lung Cellular and Molecular Physiology
JF - American Journal of Physiology - Lung Cellular and Molecular Physiology
IS - 4 7-4
ER -