Sulfatides trigger increase of cytosolic free calcium and enhanced expression of tumor necrosis factor-α and interleukin-8 mRNA in human neutrophils. Evidence for a role of L-selectin as a signaling molecule

C. Laudanna, G. Constantin, P. Baron, E. Scarpini, G. Scarlato, G. Cabrini, C. Dechecchi, F. Rossi, M. A. Cassatella, G. Berton

Research output: Contribution to journalArticle

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Abstract

Sulfatides have been established recently as ligands for L-selectin, and we investigated whether they trigger transmembrane signals through ligation of L-selectin. We found that sulfatides trigger the increase of cytosolic free calcium in neutrophils and that this effect was strictly dependent on sulfation of the galactose ring, as non-sulfated galactocerebrosides were not stimulatory. Chymotrypsin and phorbol 12-myristate 13-acetate treatment of neutrophils caused shedding of L-selectin, but not of class I major histocompatibility complex antigens or β2 integrins, and blunted the capability of neutrophils to respond to sulfatides with an increase of cytosolic free calcium. Four different anti-L-selectin antibodies (DREG-200, LAM1/3, LAM1/6, and LAM1/10), but not four control antibodies directed against different surface molecules of neutrophils, also triggered an increase of cytosolic free calcium. The anti-L-selectin antibodies were stimulatory both if used in a soluble form, after cross-linking with anti- mouse F(ab') 2 fragments, and immobilized to protein A of Staphylococcus aureus through the Fc fragment. With immobilized antibodies, an increase of cytosolic free calcium was found also by plating neutrophils on antibodies bound to protein A-coated coverslips and monitoring the increase of cytosolic free calcium by fluorescence microscopy. Both sulfatides and anti-L-selectin antibody effects were not inhibited by pertussis toxin, thus indicating that a pertussis toxin-sensitive GTP-binding protein was not involved in signal transduction. Sulfatides also triggered an increase of tumor necrosis factor- α and interleukin-8 mRNAs in neutrophils. Also to act as stimuli of cytokine mRNA expression, sulfatides required sulfation of the galactose ring, as non- sulfated galactocerebrosides were not stimulatory, and depended on expression of L-selectin, as shedding of this molecules induced by chymotrypsin blunted their effects. These findings suggest that L-selectin can transduce signals activating selective cell function.

Original languageEnglish
Pages (from-to)4021-4026
Number of pages6
JournalJournal of Biological Chemistry
Volume269
Issue number6
Publication statusPublished - 1994

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Sulfoglycosphingolipids
L-Selectin
Interleukin-8
Neutrophils
Tumor Necrosis Factor-alpha
Calcium
Messenger RNA
Molecules
Antibodies
Galactosylceramides
Pertussis Toxin
Staphylococcal Protein A
Chymotrypsin
Galactose
Immobilized Proteins
Immobilized Antibodies
Immunoglobulin Fc Fragments
Signal transduction
Histocompatibility Antigens Class I
Fluorescence microscopy

ASJC Scopus subject areas

  • Biochemistry

Cite this

Sulfatides trigger increase of cytosolic free calcium and enhanced expression of tumor necrosis factor-α and interleukin-8 mRNA in human neutrophils. Evidence for a role of L-selectin as a signaling molecule. / Laudanna, C.; Constantin, G.; Baron, P.; Scarpini, E.; Scarlato, G.; Cabrini, G.; Dechecchi, C.; Rossi, F.; Cassatella, M. A.; Berton, G.

In: Journal of Biological Chemistry, Vol. 269, No. 6, 1994, p. 4021-4026.

Research output: Contribution to journalArticle

Laudanna, C. ; Constantin, G. ; Baron, P. ; Scarpini, E. ; Scarlato, G. ; Cabrini, G. ; Dechecchi, C. ; Rossi, F. ; Cassatella, M. A. ; Berton, G. / Sulfatides trigger increase of cytosolic free calcium and enhanced expression of tumor necrosis factor-α and interleukin-8 mRNA in human neutrophils. Evidence for a role of L-selectin as a signaling molecule. In: Journal of Biological Chemistry. 1994 ; Vol. 269, No. 6. pp. 4021-4026.
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abstract = "Sulfatides have been established recently as ligands for L-selectin, and we investigated whether they trigger transmembrane signals through ligation of L-selectin. We found that sulfatides trigger the increase of cytosolic free calcium in neutrophils and that this effect was strictly dependent on sulfation of the galactose ring, as non-sulfated galactocerebrosides were not stimulatory. Chymotrypsin and phorbol 12-myristate 13-acetate treatment of neutrophils caused shedding of L-selectin, but not of class I major histocompatibility complex antigens or β2 integrins, and blunted the capability of neutrophils to respond to sulfatides with an increase of cytosolic free calcium. Four different anti-L-selectin antibodies (DREG-200, LAM1/3, LAM1/6, and LAM1/10), but not four control antibodies directed against different surface molecules of neutrophils, also triggered an increase of cytosolic free calcium. The anti-L-selectin antibodies were stimulatory both if used in a soluble form, after cross-linking with anti- mouse F(ab') 2 fragments, and immobilized to protein A of Staphylococcus aureus through the Fc fragment. With immobilized antibodies, an increase of cytosolic free calcium was found also by plating neutrophils on antibodies bound to protein A-coated coverslips and monitoring the increase of cytosolic free calcium by fluorescence microscopy. Both sulfatides and anti-L-selectin antibody effects were not inhibited by pertussis toxin, thus indicating that a pertussis toxin-sensitive GTP-binding protein was not involved in signal transduction. Sulfatides also triggered an increase of tumor necrosis factor- α and interleukin-8 mRNAs in neutrophils. Also to act as stimuli of cytokine mRNA expression, sulfatides required sulfation of the galactose ring, as non- sulfated galactocerebrosides were not stimulatory, and depended on expression of L-selectin, as shedding of this molecules induced by chymotrypsin blunted their effects. These findings suggest that L-selectin can transduce signals activating selective cell function.",
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AU - Constantin, G.

AU - Baron, P.

AU - Scarpini, E.

AU - Scarlato, G.

AU - Cabrini, G.

AU - Dechecchi, C.

AU - Rossi, F.

AU - Cassatella, M. A.

AU - Berton, G.

PY - 1994

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