The effect of sphingosine on the release of fibronectin from human lung fibroblasts.
Previous work from our laboratory (Scheidl et al. (1992) Biochim. Biophys. Acta 1135, 295-300) has shown that sphingosine (Sph), a known inhibitor of protein kinase C (PKC) inhibited the release of fibronectin (FN) into the media of fibroblast cells in culture and that this effect was independent of PKC. We now report that the action of Sph was time dependent (maximum inhibition in 2 h), concentration dependent (0.5-10 microM, at which concentration Sph was not toxic), and rapidly reversible after removal of Sph. Incubating with [35S]methionine, we found Sph reduced FN release either by inhibition of FN synthesis or FN secretion. To distinguish between these two possibilities, cells were treated with monensin, an inhibitor of FN secretion, which together with Sph showed no additive effect on FN release. Cycloheximide also inhibited FN release, but this inhibition was additive to that with monensin. We concluded that Sph inhibited secretion of FN, not synthesis. Furthermore, intracellular FN declined less in Sph-treated than in untreated control cells. By cell-surface iodination with Na125I and lactoperoxidase, followed by immunoprecipitation with antibodies to FN and alpha 5 beta 1-integrin, we showed that labeled multimeric and dimeric cell-surface FN declined, whereas integrins remained unchanged upon Sph treatment. This result suggested that though the number of cell-surface receptors for FN was not affected by Sph, their affinity may be reduced. In addition, we found that either pre- or co-treatment for 30 min with Sph caused a robust inhibition of cell adhesion to FN-coated plastic. We conclude that Sph rapidly inhibits FN secretion, lowers cell-surface FN, and greatly reduces cell adhesion to a FN substratum.
Scita
G
G
Department of Nutritional Sciences, University of California at Berkeley 94720.
Wolf
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G
D002448:Cell Adhesion / Q000187:drug effects; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D005353:Fibronectins / Q000378:metabolism*; D006801:Humans; D008168:Lung / Q000187:drug effects / Q000378:metabolism*; D013110:Sphingosine / Q000494:pharmacology*
D005353:Fibronectins; D013110:Sphingosine