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Intracellular calcium mobilization and phospholipid degradation in sphingosylphosphorylcholine-stimulated human airway epithelial cells.

ABSTRACT: Extracellular sphingosylphosphorylcholine (SPC) caused a remarkable elevation in the intracellular Ca2+ concentration ([Ca2+]i) in immortalized human airway epithelial cells (CFNP9o-). An increase in total inositol phosphates formation was determined; however, the dose responses for [Ca2+]i elevation and inositol phosphates production were slightly different and, furthermore, PMA and pertussis toxin almost completely inhibited [Ca2+]i mobilization by SPC, whereas inositol phosphates production was only partially reduced. The possible direct interaction of SPC with Ca2+ channels of intracellular stores was determined by experiments with permeabilized cells, where SPC failed to evoke Ca2+ release, whereas lysophosphatidic acid was shown to be effective. The level of phosphatidic acid was increased by SPC only in the presence of AACOCF3, a specific inhibitor of phospholipase A2 (PLA2) and blocked by both pertussis toxin and R59022, an inhibitor of diacylglycerol kinase. R59022 enhanced diacylglycerol production by SPC and also significantly reduced [Ca2+]i mobilization. Only polyunsaturated diacylglycerol and phosphatidic acid were generated by SPC. Lastly, SPC caused stimulation of arachidonic acid release, indicating the involvement of PLA2. Taken together, these data suggest that, after SPC stimulation, phospholipase C-derived diacylglycerol is phosphorylated by a diacylglycerol kinase to phosphatidic acid, which is further hydrolysed by PLA2 activity to arachidonic and lysophosphatidic acids. We propose that lysophosphatidic acid might be the intracellular messenger able to release Ca2+ from internal stores.

SUBMITTER: Orlati S

PROVIDER: S-EPMC1219734 | biostudies-other | 1998 Sep

REPOSITORIES: biostudies-other

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Project description:The pleiotrophic cytokine interleukin (IL)-13 features prominently in allergic and inflammatory diseases. In allergic asthma, IL-13 is well established as an inducer of airway inflammation and tissue remodeling. We demonstrated previously that IL-13 induces release of transforming growth factor-alpha (TGFalpha) from human bronchial epithelial cells, with proliferation of these cells mediated by the autocrine/paracrine action of this growth factor. TGFalpha exists as an integral membrane protein and requires proteolytic processing to its mature form, with a disintegrin and metalloproteinase (ADAM)17 responsible for this processing in a variety of tissues.In this study, normal human bronchial epithelial (NHBE) cells grown in air/liquid interface (ALI) culture were used to examine the mechanisms whereby IL-13 induces release of TGFalpha and cellular proliferation. Inhibitors and antisense RNA were used to examine the role of ADAM17 in these processes, while IL-13-induced changes in the intracellular expression of TGFalpha and ADAM17 were visualized by confocal microscopy.IL-13 was found to induce proliferation of NHBE cells, and release of TGFalpha, in an ADAM17-dependent manner; however, this IL-13-induced proliferation did not appear to result solely from ADAM17 activation. Rather, IL-13 induced a change in the location of TGFalpha expression from intracellular to apical regions of the NHBE cells. The apical region was also found to be a site of significant ADAM17 expression, even prior to IL-13 stimulation.Results from this study indicate that ADAM17 mediates IL-13-induced proliferation and TGFalpha shedding in NHBE cells. Furthermore, they provide the first example wherein a cytokine (IL-13) induces a change in the intracellular expression pattern of a growth factor, apparently inducing redistribution of intracellular stores of TGFalpha to the apical region of NHBE cells where expression of ADAM17 is prominent. Thus, IL-13-induced, ADAM17-mediated release of TGFalpha, and subsequent epithelial cell proliferation, could contribute to the epithelial hypertrophy, as well as other features, associated with airway remodeling in allergic asthma.

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Intracellular calcium mobilization and phospholipid degradation in sphingosylphosphorylcholine-stimulated human airway epithelial cells.

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